Published in the United States _ : VOLUME 14 - NUMBER 1—
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Amphibian & Reptile Conservation 14(1) [General Section]: 1-9 (e218).
The ecology, distribution, status, threats, and conservation of the Common Water Monitor (Varanus salvator) in the Dhaleswari River of Assam, India
‘*Muhammed Khairujjaman Mazumder, ?Amir Sohail Choudhury, 7Rofik Ahmed Barbhuiya, 3Himabrata Chakravarty, and *Badruzzaman Barbhuiya
'Department of Zoology, Dhemaji College, Dhemaji, 787057, Assam, INDIA *Department of Ecology and Environmental Science, Assam University, Silchar 788011, Assam, INDIA *Department of Zoology, Srikishan Sarda College, Hailakandi 788151, Assam, INDIA *District Level Laboratory, Public Health Engineering Department, Hailakandi 788155, Assam, INDIA
Abstract.—The Common Water Monitor, Varanus salvator (Laurenti, 1768), is a large monitor lizard distributed in southern and south-east Asia, including India which remains closely associated with water bodies, such as rivers and lakes. Although IUCN considers it to be ‘Least Concern,’ the Common Monitor Lizard faces several threats throughout its global distribution range, and the status of the species is decreasing rapidly. The Dhaleswari River of Assam (India) is one of the most important abodes of this species, where it is locally known by the names ‘/rong’ and ‘Shanda.’ Geographically, the Dhaleswari River is located in southern Assam (India), which falls within the Indo-Burma Biodiversity hotspot area. Unfortunately, most of the wildlife of southern Assam (India) are poorly studied, and this varanid is one of the most ignored species of the region. The present study was conducted along the Dhaleswari River, Assam, India, to elucidate the distribution, status, ecology, threats, and conservation of the Common Water Monitor, and is the first report on this species from this river. The results show that the Dhaleswari River still serves as a habitat of the species, with a viable population. Further, the species was found to prefer smaller rivers with clayed soil and bushes, and it faces major threats from habitat destruction, hunting for flesh and oil, and conflicts with humans. Based on our observations, we discuss recommendations for the conservation of this large varanid.
Keywords. Asia, Barak Valley, Indo-Burma Biodiversity hotspot, Reptilia, Sauria, Varanidae
Citation: Mazumder MK, Choudhury AS, Barbhuiya RA, Chakravarty H, Barbhuiya B. 2020. The ecology, distribution, status, threats, and conservation of the Common Water Monitor (Varanus salvator) in the Dhaleswari River of Assam, India. Amphibian & Reptile Conservation 14(1) [General Section]: 1-9 (e218).
Copyright: © 2020 Mazumder et al. This is an open access article distributed under the terms of the Creative Commons Attribution License [Attribu- tion 4.0 International (CC BY 4.0): https://creativecommons.org/licenses/by/4.0/], which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. The official and authorized publication credit sources, which will be duly enforced, are
as follows: official journal title Amphibian & Reptile Conservation; official journal website: amphibian-reptile-conservation.org. Received: 3 March 2018; Accepted: 23 May 2019; Published: 30 January 2020
Introduction
The Common Water Monitor (Varanus salvator) is the largest monitor lizard of India, and second largest of the world, after the Komodo Dragon (V. komodoensis). It has the widest global distribution among all varanids (Traeholt 1994). The distribution of the species in the Indian subcontinent was described by Das (1994), Daniel (1983), and Koch et al. (2013), while Smith (1935) reported its occurence and distribution in Northeast India, of which the state of Assam 1s a part. Anderson (1982) and Auffenberg (1986) reported the occurence of this species from Assam (India). Among the four recognized subspecies of the Common Water Monitor, V. s. macromaculatus occurs in Assam (Traeholt 1994; Auffenberg 1986). The Common Water Monitor is generally found associated with water systems including
Correspondence. *khairujjaman1987@gmail.com
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rivers and wetlands (Ahmed et al. 2009; Cota et al. 2009). It is seldom found far from water (Smith 1935), and only rarely beyond 200 m from water bodies (Cota et al. 2009). The Common Water Monitor has a wide trophic niche, and it consumes a variety of prey species including crabs, fishes, other lizards, snakes, domestic fowl, and eggs of other animals, in addition to scavenging (Ahmed et al. 2009).
Some of the important publications on the reptiles of Northeast India include Ahmed et al. (2009), Das (2008), and Choudhury (1989, 1992, 1993a,b, 1995, 1996a,b, 1998, 2011). However, none of these articles provide special treatment of either the Common Water Monitor, or southern Assam. Southern Assam (India), is also known as the Barak Valley, and comprises the districts of Cachar, Hailakandi, and Karimganj. It is a part of the Indo-Burma Biodiversity hotspot, and
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92°27' E 92°40°E
Barak river
24°20'N te]
Legends
P: Panchgram K: Kanchanpur B: Bowerghat M: Monacherra L: Lala Town
24°38' N
8: Sahabad
Fig. 1. (A) Map of India, highlighting Assam. (B) Map of Assam, highlighting Hailakandi district. (C) Map of Hailakandi district showing the two rivers, Dhaleswari River and Katakhal River, with distribution of the Common Water Monitor (Varanus salvator). Dots represent sighting locations during the present survey; current distribution in the Dhaleswari River is shown in green. Further downstream, despite no present records, occurrence in the past (1970s—1980s) was reported by several interviewees (shown in yellow). No reports on present occurrence in the Katakhal River (shown in red) could be found. Map by A.S. Choudhury.
harbors a myriad wildlife assemblage (Myers et al. 2000; Choudhury 1997, 2013; Mazumder 2014), including a diverse herpetofauna (Ahmed et al. 2009). Two species of monitor lizard are reported to inhabit the region, the Bengal Monitor (Varanus bengalensis) [Das 2008] and the Common Water Monitor (Varanus salvator) [Ahmed et al. 2009; Whitaker and Whitaker 1980]. Once common in most of the rivers of Assam, the habitat of the Common Water Monitor is decreasing rapidly (Ahmed et al. 2009), although its IUCN status currently remains Least Concern (Bennet et al. 2010).
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The present study was conducted in the Dhaleswari River of southern Assam (India) to elucidate the distribution (past and current), status, ecology (behavior and habitat), threats, and conservation of the Common Water Monitor. Based on the findings of this study, we provide recommendations for the conservation of this species. The present study is particularly important since some of the majestic mega-fauna of southern Assam, including Gaur (Bos gaurus), Asiatic Wild Water Buffalo (Bubalus arnee), Indian One-horned Rhino (Rhinoceros unicornis), Sumatran Rhino (Dicerorhinus sumatrensis), Javan Rhino (R. sondaicus), Royal Bengal Tiger (Panthera tigris), and notably two mega-reptiles: the Gharial (Gavialis gangeticus) and Marsh Crocodile (Crocodylus palustris), have been extirpated in the last century (Choudhury 1997, 1998, 2013, 2016; Singha 2009), while the Ganges River Dolphin (Platanista gangetica gangetica) is on the brink of extirpation (Mazumder et al. 2014).
Materials and Methods
Study site. The present study was conducted in the Dhaleswari River, located in the district of Hailakandi, in southern Assam of India. The climate is of the tropical monsoon type, with average elevation of 21 m asl, average annual precipitation of 2,400—2,800 mm, and the temperature varies from 37 °C in the summer to 7 °C in the winter (Choudhury and Choudhury 2017). The Dhaleswari River originates in the Mizo Hills in the state Mizoram (India), flows through hilly terrains towards north, and enters the Hailakandi district. Further downstream, the river bifurcates at Shahabad (Gainja- Khauri) of Hailakandi (24°28°51.8°N, 92°34738.3”E), whereby its water is diverted to an artificial channel now called Katakhal River with a sluice gate, thereby eliminating the immediate downstream _ section. However, further downstream, smaller streams join the river, thereby enhancing its flow discharge, and it finally drains into the Barak River at Panchgram, Hailakandi (24°51°54’”N, 92°36°32”E; Fig. 1). The length of the river from Shahabad to Panchgram is approximately 110 km, with an elevation ranging from 20-33 m. The river is almost dry with no flow in the winter season, especially in the upstream half of its length, while significant flow is attained in the monsoon season (April to October).
The present study was conducted in a stretch of the river from Shahabad (24°28751.8"N, 92°34’38.3”E) to Bowerghat (24°38°37.4"N, 92°32713”E), covering an approximate length of 56 km. The area is inhabited by Bengali community people who cultivate rice paddies (between June and November) in the plain areas, and vegetables (between November and March) in the plains as well as on the river banks. The river bank has an abundance of bushes and bamboo groves, as well as large trees. Brief surveys were also made in the Katakhal River (the other river of Hailakandi district) from Sahabad to
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Te eek
Fig. 2. Photographs relevant to the habitat and threats of the Common
a Ee Nee , aS Water Monitor (Varanus salvator) in the Dhaleswari River,
PE? i =o =
Assam, India. (A) Research team interacting with the locals at Rongpur 5, Hailakandi. (B) Habitat of the Common Water Monitor in the Dhaleswari River, showing bushes and other features on the banks. (C) The sandy bank of the Katakhal River, prone to erosion and landslides, is the habitat not preferred by the Common Water Monitor. (D) The sluice gate at the mouth of Dhaleswari River at Shahabad, which prevents water flow into it and diverts the water to the Katakhal River. (E) Encroachment and conversion of the
Dhaleswari River into fisheries by the locals building dikes at Rongpur 2. Photos by A.S. Choudhury.
its confluence with the Barak River at Katakhalmukh (24°51°21”N, 92°37°27°E; Fig. 1). The Katakhal River has greater depth, width, flow rate, and flow discharge, and it has abrupt banks (with sandy soil) which experience frequent and severe erosion and landslides.
Survey techniques. Preliminary discussions with the riparian local people and forest officials were conducted to locate extant populations in the Dhaleswari and Katakhal rivers of Hailakandi, southern Assam. The discussions revealed that the Common Water Monitor
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does not occur in the Katakhal River, while it continues to inhabit the Dhaleswari River between Shahabad and Bowerghat (Fig. 1). Two researchers independently carried out an on-foot survey simultaneously along both banks of the Dhaleswari River, covering a length of 56 km (of the total river length of ~110 km), from Shahabad to Bowerghat, to conduct focal counts of the species and the number of nests available in the river bank. The burrows made by the lizard were considered to be nests, while others, including crevices and burrows of other animals, were considered temporary refuges. The
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Table 1. Geo-coordinates of locations along the Dhaleswari River, Assam, India, where live specimens of the Common Water Monitor (Varanus salvator) were sighted during the survey, including the number of individuals sited at each location.
Geo-coordinates No. of individuals
Site No. Location : Latitude Longitude sighted 1 Shahbad 24°28'°51.8°N 92°34’38.3"E 1 2 Shahabad 1 24°29°31"N 92°34’33”E 1 3 Rongpur 2 24°30°18.4"N 92°34’39.8"E 1 4 Rongpur 5 24°31°31”"N 92°34’ 36"E 1 5 Rongpur 5 24°31°39.7°N 92°35’°00.7"E 2 6 Rongpur 4 24°31°42.6"N 92°34’°57.7°E 2 ei Abdullahpur 1 24°32’08”"N 92°35°23”E 2 8 Tantoo Road Bridge 24°32°22.1"N 92°35°33.2”E 1 9 Dhanipur 24°33’29"N 92°35’48”"E 2 10 Lala Rural College 24°33°32”N 92°36 12°E 2 11 Lalaghat Nala confluence 24°33°36”N 02°35 21°E 3 12 Kaya Khal confluence 24°34’00”"N 92°34’?24”E 2 13 Bhabanipur 24°34°19"N 92°34’54”°E 2 14 Sarbanandapur 24°34°19"N 92°34’51°E 1 15 Behula 24°34’44"N 92°34’03”E 1 16 Aenakhal Tea Factory 24°35’18"N 92°33’04”E 1 17 Aenakhal Market 24°35°55”N 92°32°352°R 1 18 Monacherra (Lakhinagar) 24°36’45"N 92°32’54”E 3 19 Kukinagar 24°37°26"N 92°32’22”E 2 20 Bowerghat 24°38°37.4"N 92°32713”"E 1
diameter of each nest hole was measured and topology was identified whenever possible. Different types of nests were recorded, which were verified by local guides. Upon sighting of a lizard, the Global Positioning System (GPS) coordinates were recorded using a digital GPS machine (eTrex 20X, Garmin, China). The surveyors made keen observations in the trees for possible occurrences of the varanid, since this species has been reported to climb trees for basking (Ahmed et al. 2009). Since no specimens were found in the Dhaleswari River further downstream of Bowerghat, or in the Katakhal River, those negative data are not incorporated into the further analysis. The total time spent in the field survey, for direct observations and counting of the lizards, was 118 h for each observer. In addition, potential threats to the monitor lizard and its habitat were observed during the survey.
Interviews of local people. Interviews of the locals (n = 30), generally the older folks, were conducted regarding the occurrence of the lizards, their nesting places, food, incidences of hunting, and potential conflicts (Fig. 2A). In addition to the local people of the area, many others hailing from different localities in southern Assam were also interviewed to get an idea of the past and present occurrences of the monitor lizard in the Hailakandi district, and in the Dhaleswari River and Katakhal River in particular. The interviews used colored photographs, pictorial guides and sketches, and
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a closed-ended questionnaire. The total time spent on the interviews was 55 h.
Results Distribution and Population Status
Past and current distribution. Interactions with local people, forest personnel, and elderly people of the villages revealed that in the 1970s, the whole length of the Dhaleswari River (~110 km) had viable and abundant populations of the Common Water Monitor. However, the population declined gradually, and the species currently inhabits only the section of the river between Shahabad and Bowerghat (Fig. 1). The species also occurs in the tributaries or smaller water channels (mu//ahs) joining the Dhaleswari River in this river section, most importantly the Lalaghat Nala, Kaya Khal, and others. In addition, stray individuals have been reported from Kanchanpur (~10 km downstream of the current range). It 1s often found near human habitations, crop fields, ponds, and lakes near the current main distribution range. Neither interviews nor surveys revealed the occurrence of this species in the Katakhal River, and we therefore argue that the species has been extirpated from this river.
Population status. During the study, 32 individuals were directly sighted at 20 locations in the 56 km of the river
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section surveyed, for a linear density of 0.57 individuals/ km of the river course (Table 1). The total number of active nests found during the survey was 215, for a linear density of 3.84 nests/km of the river section. In addition, the survey found 63 burrows and crevices which were used by the lizards as refuges.
Ecology
Habitat and nesting ecology. The current habitat of the monitor in the Dhaleswari River includes areas with river channel widths of 1—5 m in the winter months, with no water present. However, in the monsoon season, the river width may reach up to 30 m, with significant flow and discharge. The depth of the river in the study area varies from 0-10 m. The width of the river bank is 10—30 m, and the banks become flooded with the high waters of the monsoons. The river banks are inclined, and not vertically abrupt. The approximate distances of human habitations from the river bank range from 5—50 m. The river bank is characterized by the presence of bushes, secondary tree growth, planted woodlands, bamboo groves, and occasional crop fields (mainly vegetables), with or without bamboo fencing (Fig. 2B). However, the lizard is absent in the Katakhal River, which indicates that it does not prefer a river where flow and discharge are high, and banks are abrupt and steep with sandy soil which are prone to erosion (Fig. 2C).
The lizard made nests in the inclined river banks in areas where anthropogenic pressure is less intensive. The diameters of the nest entrance holes were 20-25 cm (23.59 + 1.14 cm; mean + SD; n = 40). The vicinity of the nests was very clean, due to movement of the lizards, while the entries of some nests were half-sealed with soil. The entries of the nests were horizontal, or inclined downward or upward; however, the tunnel rose upwards immediately at an approximate distance of ~0.3—0.6 m from the entrance hole. Several other micro-habitats were found, which were natural crevices or burrows of other animals that were used by the monitors, probably as refuges. In these larger crevices, the lizards generally occurred in groups of several individuals.
Behavioral ecology. During winter months, the Common Water Monitor basks in the sun, and has been found to climb trees for basking, especially in the morning. During monsoons, the river water level increased and the nests were flooded. This factor drove the monitors to hide in the bushes, forage near human settlements, and shelter in trees. According to the locals (n = 30), the monitor has been found to feed on fishes, snakes, small lizards, crabs, and carcasses of other larger animals which were thrown into the river. There are several poultry farms near the river banks, as well as slaughter houses near populated areas, and people often dispose of carcasses and remnants of slaughtered animals in the rivers. In such places, the Common Water Monitor was frequently encountered.
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Monitors with newborns were observed in the month of April, suggesting that eggs hatched in the early monsoon showers which coincided with the nests becoming submerged by flood waters. Also, the abundance of fish and other prey organisms were greater in this season, thereby making it more suitable for the newborns.
Threats
Like any other aquatic or water body associated species, and the monitors of other habitats, there are several threats to the Common Water Monitors in the Dhaleswar1 River, which threaten the survival of the species.
Habitat loss and destruction. Most of the rivers of Assam (India) are under immense human pressure due to pollution, overfishing, extraction of water for agricultural and domestic uses, encroachment, and other uses; and the Dhaleswari River is no exception. The diversion of the river water to the Katakhal River by the sluice gate at Shahabad (Fig. 2D) has led to a decline in available water in the parent stream, and a loss of biodiversity. This water diversion also invites cultivation on the river banks. In several places on the banks, people made fences of bamboo or steel wire, thereby reducing the available space for the lizards on the banks. However, due to decreased river flow, erosion, and land-slides, many sections of the river could not be studied.
The Dhaleswari River is highly polluted due to dumping of domestic and agricultural wastes, sewage, carcasses of animals, and other refuse. All the wastes which are dumped continue to accumulate until the next monsoon. This leads to deterioration of water quality, and thereby causes depletion of aquatic diversity and prey species, although it does not appear to harm the lizards directly.
Prey species depletion. The Dhaleswari River does not maintain water flow in the winter months, and it fully dries up gradually upstream. In addition, people make dikes in the banks to catch any remaining fish left in the river, and these dikes are used for human movement as well. Unfortunately, the upstream section of the river has been encroached and converted into fisheries, and flow in certain areas has been diverted resulting in extremely narrow channels (Fig. 2E). In this situation, prey species, mainly fishes and those dependent on the fishes, barely remain in the river. The severe prey depletion is thus among the most important threats to the Common Water Monitor in this river.
Hunting. Unfortunately, there were several instances of hunting of the Common Water Monitor throughout its range in the river. It is poached for meat, and also for oil. The tribal community people of the region, mainly the Manipuri, Khasi, Mizo, Naga, Tea tribes, and others, hunt the monitor as a protein source. The oil extracted
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Fig. 3. A Common Water Monitor (Varanus salvator) killed for venturing into a human habitation at Rongpur 6, Hailakandi. It was subsequently buried. Photo by R.A. Barbhuiya.
from the base of the tail of the lizard, called Shandar tel (‘Oil of the Monitor lizard’ in the local language), 1s used as a sexual lubricant by man. Although the Bengali community people, who are the majority in the present study area, do not consume the lizard themselves, they catch any individuals that venture into the settlements and sell them to the tribes. A moderate sized individual may cost up to 1,000-1,500 INR (~14-21 USD), while the oil is sold at the rate of 500 INR/L (~7 USD).
Human-monitor lizard conflicts. During the monsoon months, the preferred sites of nesting or refuge are submerged, and thus the lizards venture into human habitations for foraging, mainly in the afternoon hours. They often prey on domestic poultry or venture into artificial ponds and prey on fishes. Thus, the local people who do not consume the meat of the lizard consider them to be pests and often kill them. Such killing is an age- old practice. In addition, many people (mainly children) believe that the lizard may physically harm them, or even kill them, and that the bite of the lizard 1s poisonous. Thus, the local people often harm the Common Water Monitor whenever it occurs within their reach. Despite occasional attacks on domestic animals and venturing into ponds, the lizard was not reported by any of the respondents to actually attack man. In 2011, one individual was caught from village Kanchanpur, and killed (Saifur Rahman Laskar, Kanchanpur, pers. comm.). According
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to the interviewees, there were about 25 (range 20-30) incidences of hunting Common Water Monitors in 2014 in the study area. In the month of August 2015, as many as six cases of such hunting were recorded from the river section between Bowerghat and Monacherra, a stretch of ~5 km. On 5 September 2015, one lizard ~1.8 m long (Fig. 3) was killed from Rongpur 6 village for attacking poultry, and subsequently buried. However, since the responses of only 30 interviewees were taken, the exact number of incidences is estimated to be much higher than is indicated here.
Lack of research and awareness. Unfortunately, the Common Water Monitors are the least studied of the large animals in the region. To the best of our knowledge, there are no reports on the Common Water Monitors from this region, save for the few records simply reporting its occurrence, and no mentions in the literature of its occurrence in Dhaleswari River. Moreover, no specific awareness campaign has yet been undertaken to educate the locals, and the lizard remains among the most ignored wildlife of the region.
Discussion
The freshwater bodies of the world, both lotic and lentic, are habitats with highly diverse wildlife assemblages as well as immense anthropogenic pressures, which threaten the survival of these species (Dudgeon 2000). Amidst a myriad of threats, the Common Water Monitor still survives in habitats that are highly disturbed. In the Dhaleswari River of Hailakandi (Assam, India), there is still a good population of the species, inhabiting the upstream stretch of the river which represents about one- half of the ~110 km total length of the river course. This varanid used to be very common throughout the region in the 1970s. However, today the species 1s encountered only in the 56 km stretch between Bowerghat and Shahabad (Fig. 1). During the survey, 32 individual Common Water Monitors were directly sighted in this river section, for a linear density of 0.57 individuals/km of the river course (Table 1). However, the linear density of the active nests was found to be 3.84/km, which suggests that the actual number of lizards inhabiting the river section is quite a bit higher than the number of individuals seen.
No present record of the species in the Katakhal River could be found. Since this river is much larger, with greater width, depth, flow rate, and flow discharge compared to the Dhaleswari, it further supports our assumption that the Common Water Monitor prefers smaller river habitats (Fig. 2B). In the Katakhal River, due to high flow velocity and discharge, the banks are frequently eroded in the monsoons, and thus monitor nests would be damaged. Since the species appears to use its nests repeatedly (for more than one year), larger rivers with sandy banks and prevalent erosion and landslides (Fig. 2C) would not serve as good habitats, and thus the
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monitors avoid these rivers. This survey reveals that the nests are made on high grounds, which is consistent with earlier reports (Biswas and Kar 1980). The direction of the nests was found to be turned upwards in the present study area. Biswas and Kar (1980) also mentioned that the monitor seals the nest hole after laying eggs by scraping up soil, and we speculated that the half- sealed nests found in the present study area are probably similarly sealed and contain eggs that were laid.
During the present field work, some other fauna from the river section were recorded, which are prey of either the lizard or its competitors, or which prey on the eggs and hatchlings of the Common Water Monitor. Other lizards recorded from the present survey were Indian Garden Lizard (Calotes versicolor), Tokay Gecko (Gekko gecko), Bronze Grass Skink (Eutropis macularia), and Many-lined Grass Skink (Eutropis multidasciata), and the snakes included Checkered Keelback (Xenochrophis piscator), Red-necked Keelback (Rhabdophis subminiatus), Indian Rat Snake (Pitvas mucosa), Banded Krait (Bungarus fasciatus), Greater Black Krait (Bungarus niger), King Cobra (Ophiophagus hannah), Monocled Cobra (Naja kaouthia), Common Water Snake (Enhydris enhydris), and Indo-Chinese Rat Snake (Ptvas korros). The amphibians included Common Asian Toad (Duttaphrynus melanostictus), Indian Skipping Frog (Euphlyctis cyanophlyctis), Indian Bull Frog (Hoplobatrachus tigerinus), and Common Tree Frog (Polypedates teraiensis). The fishes recorded from the river were Sperata aor, Sperata seenghala, Channa punctatus, Notopterus notopterus, Clarias batrachus, Heteropneustes fossilis, Pethia ticto, and Anabas_ testudineus. The common fishes cultured by the local people in the ponds and fisheries in the adjoining areas (which are often prey of the Common Water Monitor) were Labeo rohita, Clarias gariepinus, Ctenopharyngodon idella, Cyprinus carpio, Gibelion catla, Hypophthalmychthys molitrix, and Labeo calbasu.
The water monitor of Dhaleswari River faces immense threats, which include anthropogenic pressures in terms of over-fishing, construction of dikes, conversion of the river into fisheries (Fig. 2E), cultivation of crops, and fencing of potential cultivable areas on the bank. Further, the species is frequently hunted as well as persecuted as a pest (Fig. 3). Thus, habitat destruction, hunting, and retaliatory killings are the major conservation issues in the present study area. These threats, in addition to others, have extirpated the other majestic aquatic mega- fauna of the river, including the Ganges River Dolphin Platanista gangetica gangetica (Mazumder et al. 2014). In spite of all these potential pressures, the Common Water Monitor could survive well in the river thanks to certain attributes, most notably its higher adaptability to human-modified habitats and wider food niche. In fact, the Common Water Monitor of Dhaleswari River is more common in town areas, where the dumping of municipal wastes provides better provisions for this lizard.
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Moreover, the many poultry farms on the river banks deliberately dispose of dead birds near the nests of the lizards. In some areas where the monitor 1s deliberately or incidentally provisioned, it is often concentrated.
The Indian Wildlife (Protection) Act (1972) listed the species under Schedule I, thereby conferring maximum legal protection. Although the IUCN regards this species to be stable and considers it “Least Concerned’ (LC), the status of the population in southern Assam (India) is decreasing. Thus, the river section between Shahabad and Bowerghat requires special care, by limiting anthropogenic pressures. The Assam Fishery Rules (1953) should be enforced, as they restrict fishing of brood fishes in the spawning season, and prohibit fishing using specific methods in specific seasons, including dewatering, which will enhance fish stock. The construction of dikes, encroachment, and pollution of the river should be checked, and sewage should be treated before discharge. Hunting should be strictly dealt with and the relevant legislation should be strictly enforced. The people who consume the meat and use the oil of the lizards, should be motivated to stop doing so through education and outreach programs. The ecology, population status, distribution, and threats of the Common Water Monitor need to be properly evaluated and a specific long-term conservation action plan should be devised. Government and non-government organizations should come forward to work together for the conservation of this lizard. Above all, we as human beings and the dwellers of the Indo-Burma Biodiversity hotspot region should endeavor to save our pristine wildlife assemblage, or else the Common Water Monitor will be the next large reptile, after the Gharial and Marsh Crocodile, to become extirpated from southern Assam, India.
Acknowledgements.—We sincerely acknowledge some of our local guides and resource people for sharing their valuable experiences. To name a few, Abdur Rahim Mazumder (Ujankupa), Saifur Rahman Laskar (Kanchanpur), Lukan Uddin Laskar (Monacherra), Hanna Momtaz Begum Mazumder (Lakshminagar), Ali Akbar Barbhutya (Krishnapur), Faruk Ahmed Barbhuiya (Shahabad), Zinna Mohd. Badruzzaman Mazumder (Ujankupa), Riyazul Azhar Laskar (Bowerghat), Imrana Begam Choudhury (Ujankupa), Badrun Nehar Laskar (Ujankupa), Yasmin Choudhury (Hailakandi town), and Rojob Uddin Laskar (Monacherra) deserve special thanks. Ruhul Amin Ahmed (Mohanpur, Hailakandji), Jabed Ahmed (Chiporsangan, Hailakandi), Hanif Mohd. Choudhury (Shahabad), Arif Ahmed Barbhuiya (Krishnapur), and Mizazur Rohaman Mazumder (Hailakandi town W/N-IV, Hailakandi) accompanied us during field visits. We are especially thankful to Dr. Anwaruddin Choudhury, an eminent Naturalist and Conservation Biologist, for his valuable suggestions. We acknowledge the sincere efforts of the three anonymous reviewers and the handling editor Halli Boman for suggesting improvements and corrections.
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Choudhury A. 2013. The Mammals of North East India. 1*' Edition. Gibbon Books and the Rhino Foundation for Nature in North East India, Guwahati, Assam, India. 431 p.
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Derantyagala PEP. 1944. Four new races of the “Kabaragoya” lizard, Varanus salvator. Spolia Zeylanica 24: 59-62.
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Koch A, Ziegler T, Bohme W, Arida E, Auliya M. 2013. Pressing problems: distribution, threats, and conservation status of the monitor lizards (Varanidae: Varanus spp.) of Southeast Asia and the Indo- Australian Archipelago. Herpetological Conservation and Biology 8(3): 1-62.
Mazumder MK, Boro F, Barbhuiya B, Singha U. 2014. A study of the winter congregation sites of the Gangetic River Dolphin in southern Assam, India, with reference to conservation. Global Ecology and Conservation 2: 359-366.
Mazumder MK. 2014. Diversity, habitat preferences, and conservation of the primates of Southern Assam, India: the story of a primate paradise. Journal of Asia- Pacific Biodiversity 7: 347-354.
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Singha TP. 2009. Habitat ecology and conservation strategy of the fresh water dolphin, Platanista gangetica Roxburgh, in the river Barak in southern Assam. Ph.D. Dissertation, Assam University, Silchar, India. 297 p.
Smith MA. 1935. The Fauna of British India, including Ceylon and Burma. Reptilia and Amphibia, Volume II. Sauria. Taylor and Francis, London, United Kingdom. 440 p.
Traeholt C. 1994. Notes on the Water Monitor Varanus salvator as a scavenger. Malaysian Nature Journal 47: 345-353.
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Mazumder et al.
Muhammed Khairujjaman Mazumder is currently working as Assistant Professor in the Department of Zoology, Dhemaji College, Assam, India. Muhammed obtained his Ph.D. from the Department of Life Science and Bioinformatics, Assam University, Silchar, India, in the subject of Neurobiology. He has a keen interest in the natural history of Northeast India, particularly Assam, and has authored several articles in journals of international repute and book chapters on the natural history of Assam, as well as Northeast India. Muhammed is particularly interested in mammals, and both aquatic and water-dependent species, their habitats, ecology, and conservation.
Amir Sohail Choudhury developed a keen interest in wildlife in childhood, being motivated by Dr. Anwaruddin Choudhury (one of the eminent naturalists of Northeast India). Amir completed post-graduate work in Ecology and Environmental Sciences with a specialization in Wildlife. His research interests include studying the population dynamics and habitat ecology of the birds and mammals of Assam, India. He has authored several articles and book chapters on the natural history of Northeast India, particularly Assam. Further, Amir has documented reptiles from the southern part of Assam, with several publications to his credit. He is a freelancer, and is very interested in wildlife photography.
Rofik Ahmed Barbhuiya is a young naturalist who has been involved in the study and conservation of wildlife, especially primates, elephants, and birds, since his school years. Rofik is a post-graduate in Ecology and Environmental Science at Assam University, Silchar, India, and is currently pursuing his Ph.D. from the same department on the behavioral ecology of the Capped Langur of Assam (India). He has conducted several awareness programs for the conservation of Hoolock Gibbon, Phayre’s Langur, Capped Langur, and elephants, and has several articles published in international journals.
Himabrata Chakravarty is currently serving as Associate Professor and Head, Department of Zoology, Srikishan Sarda College, Hailakandi, Assam, India. Himabrata was awarded a Ph.D. by Assam University, Silchar, India, and his research interests include the ecology of birds, mammals, and reptiles. He has authored several articles and book chapters on the wildlife of Assam (India), and has been engaged in rescue, rehabilitation, and conservation programs for a long time. Himabrata also has a keen interest in wildlife photography.
Badruzzaman Barbhuiya is currently working as an Assistant Chemist in the District Level Laboratory, Public Health Engineering Department, Government of Assam, Hailakandi, Assam, India. Badruzzaman has a Chemistry background, and a keen interest in studying water quality and habitat ecology of aquatic species in southern Assam. He has experience in the analysis of water parameters in the region, and the mitigation of issues related to drinking water quality. With immense concern for the environment and climatic change, he remains involved in various awareness activities with NGOs.
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Official journal website: amphibian-reptile-conservation.org
Amphibian & Reptile Conservation 14(1) [General Section]: 10—21 (e219).
Climatic niche, natural history, and conservation status of the Porthole Treefrog, Charadrahyla taeniopus (Gunther, 1901) (Anura: Hylidae) in Mexico
‘Raciel Cruz-Elizalde, *Itzel Magno-Benitez, *Christian Berriozabal-lslas, 7Raul Ortiz-Pulido, 2*Aurelio Ramirez-Bautista, and “Raquel Hernandez-Austria
'Museo de Zoologia “Alfonso L. Herrera”, Departamento de Biologia Evolutiva, Facultad de Ciencias, Universidad Nacional Autonoma de México (UNAM). A.P. 70-399, Ciudad de México CP 04510, MEXICO *Laboratorio de Ecologia de Poblaciones, Centro de Investigaciones Biologicas, Instituto de Ciencias Bdsicas e Ingenieria, Universidad Autonoma del Estado de Hidalgo, Km 4.5 carretera Pachuca-Tulancingo, 42184, Mineral de La Reforma, Hidalgo, MEXICO +Programa Educativo de Ingenieria en Biotecnologia. Universidad Politécnica de Quintana Roo. Av. Arco Bicentenario, M 11, Lote 1119-33, Sm 255, 77500 Cancun, Quintana Roo, MEXICO ‘Departamento de Zoologia, Instituto de Biologia, Universidad Nacional Autonoma de México, Apartado Postal 70-153, 04510 Ciudad de México, MEXICO
Abstract.—Amphibian species of the family Hylidae exhibit a high degree of endemism in Mexico. To better understand ongoing declines of many amphibian populations, especially for endemic species that are particularly vulnerable to extinction, information on diverse aspects of their biological makeup is required, including their ecology. This study provides an analysis of the distribution, natural history, feeding habits, reproduction, morphology, and conservation status of Charadrahyla taeniopus, a species endemic to central Mexico. The distribution of this species extends along the Sierra Madre Oriental, primarily in cloud forest. Based on changes in climatic niche, decreases of 14.14% and 37% of its distributional range are predicted to occur by the years 2050 and 2070, respectively. An examination of the stomach contents from 31 adults and two juveniles revealed plant materials and arthropods as major parts of their diet. Charadrahyla taeniopus is sexually dimorphic in size. Females were larger than males, and after correcting for body size, females had larger jaws than males. Based on guidelines proposed by national legislation (NOM-059), we propose that this species should continue to be classified as Threatened. Further studies are necessary to classify it in a high conservation category by international legislation (IUCN) guidelines, due to the high vulnerability indicated by the Environmental Vulnerability Score, which is caused by an accelerated loss of habitat. Charadrahyla taeniopus is a good model for analyzing the conservation status of hylid frogs from temperate areas and in highly transformed environments, as this species exemplifies the conservation status of endemic amphibians in central Mexico.
Keywords. Amphibians, Central America, cloud forest, diet, morphology, reproduction
Citation: Cruz-Elizalde R, Magno-Benitez |, Berriozabal-Islas C, Ortiz-Pulido R, Ramirez-Bautista A, Hernandez-Austria R. 2020. Climatic niche, natural history, and conservation status of the Porthole Treefrog, Charadrahyla taeniopus (Gunther, 1901) (Anura: Hylidae) in Mexico. Amphibian & Reptile Conservation 14(1) [General Section]: 10-21 (e219).
Copyright: © 2020 Cruz-Elizalde et al. This is an open access article distributed under the terms of the Creative Commons Attribution License [At- tribution 4.0 International (CC BY 4.0): https://creativecommons.org/licenses/by/4.0/], which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. The official and authorized publication credit sources, which will be duly enforced, are as follows: official journal title Amphibian & Reptile Conservation; official journal website: amphibian-reptile-conservation.org.
Received: 24 October 2018; Accepted: 16 March 2019; Published: 11 February 2020
Introduction
Scientists are concerned about the worldwide decline of amphibians associated with habitat degradation (Delia et al. 2013; Wilson et al. 2013), which is caused by a variety of factors, including shifts in land use, increased pollution, and the splitting and fragmentation of habitat (Becker et al. 2007; Ochoa-Ochoa et al. 2009; Cruz- Elizalde et al. 2015). As a result, many species are globally threatened (Lips et al. 2004; Ochoa-Ochoa et al. 2009) by human activities that are destroying habitat, leading to population decline, extirpation, or even species extinction (Delia et al. 2013). The creation of
Correspondence. *ramibautistaa@gmail.com Amphib. Reptile Conserv.
regulations and laws at national and international levels, and resources such as the Red List of the International Union for the Conservation of Nature (IUCN), have been important tools for the conservation and management of biodiversity (Wilson et al. 2013).
In Mexico, flora and fauna are protected by the NOM- 059, a regulation that provides a way to evaluate the threat level or conservation status of species through a nationally-recognized method: the Método de Evaluacion del Riesgo de Extincion de las Especies Silvestres en Mexico (method for evaluation of the extinction risk of wild species, MER; DOF 2010). MER has been used to evaluate the conservation status of diverse plant taxa,
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om, ae re
Fig. 1. Female individual of Charadrahyla taeniopus (A) in a cloud forest and (B) at Tenango de Doria, Hidalgo, Mexico. Photos by Uriel Hernandez-Salinas (A) and Raciel Cruz- Elizalde (B). but major groups of animals, including many species of amphibians, have not yet been evaluated (DOF 2010). Moreover, the lack of knowledge of the distribution and natural history of amphibian species populations impedes the proper application and evaluation of the MER. For example, Lithobates johni (Blair, 1965) is an endemic species that is considered endangered, and was reported as extirpated at the type locality in Palictla, San Luis Potosi, Mexico (DOF 2010; Campos-Rodriguez et al. 2012). However, Hernandez-Austria et al. (2015) found several robust populations of the species in the state of Hidalgo. According to MER, conservation assessments should be updated to include current data and to focus on such critical factors as population density, reproductive period, and natural history (Wilson et al. 2013). One method for assessing conservation status that takes these factors into consideration is the Environmental Vulnerability Score (EVS). It is based on an algorithm proposed by Wilson et al. (2013) for amphibian species inhabiting Mexico, and has been accepted for biodiversity and conservation studies of this group at a variety of spatial scales (Johnson et al. 2015; Mata-Silva et al. 2015; Lemos-Espinal et al. 2018a,b). Use of this index has enabled better estimates of the conservation status of amphibian species at regional (Cruz-Elizalde et al. 2015, 2016) and state levels
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(Johnson et al. 2015; Mata-Silva et al. 2015) by utilizing information on ecological distribution and reproductive modes (Wilson et al. 2013).
Approximately 99 species of the family Hylidae occur in Mexico (Wilson et al. 2013; Parra-Olea et al. 2014; Canseco-Marquez et al. 2017; Johnson et al. 2017; Jiménez-Arcos et al. 2019), and 68 of these are endemic to the country (Parra-Olea et al. 2014; Caviedes-Solis et al. 2015; Canseco-Marquez et al. 2017; Johnson et al. 2017). Many of the endemic species are distributed in temperate and tropical environments (Flores- Villela et al. 2010; Delia et al. 2013). The family Hylidae exhibits a remarkable species richness and diversity in montane regions (Duellman 2001; Flores-Villela et al. 2010), which are at high risk because of a shift in land use from forests to agroecosystems such as shade coffee plantations and grazing areas (Ochoa- Ochoa et al. 2009; Santos-Barrera and Urbina-Cardona 2011; Murrieta-Galindo et al. 2013). Furthermore, the temperate environments of Mexico are expected to be affected by climate change, leading to a decrease in species richness and diversity (Lips et al. 2004; Urbina- Cardona and Flores-Villela 2010).
The genus Charadrahyla (Faivovich et al. 2005) is composed of ten species (Frost 2019) which inhabit the highlands of the Sierra Madre Oriental, Sierra Madre Occidental, and the sierras of Oaxaca and Chiapas (Duellman 2001; Campbell et al. 2009; Frost 2019). Little 1s known about their ecology and natural history, so most of the species are assigned to categories of high extinction risk by the IUCN. Four of the species, C. juanitae (Synder, 1972), C. nephila (Mendelson and Campbell, 1999), C. pinorum (Taylor, 1937), and C. taeniopus (Gunther, 1901) are in the Vulnerable category; C. chaneque (Duellman, 1961) is Endangered; C. altipotens (Duellman, 1968) and C. trux (Adler and Dennis, 1972) are Critically Endangered; and C. sakbah Jiménez-Arcos, Calzada-Arciniega, Alfaro- Juantorena, Vazquez-Reyes, Blair, and Parra-Olea, 2019, C. esperancensis Canseco-Marquez, Ramirez- Gonzalez, and Gonzalez-Bernal, 2017, and C. tecuani Campbell, Blancas-Hernandez, and Smith, 2009 have not been evaluated (IUCN 2019). According to NOM- 059-SEMARNAT-2010, C. altipotens, C. chaneque, and C. pinorum are in the Subject to Special Protection (Pr) category; C. juanitae, C. taeniopus, and C. trux are Threatened (A); and the remaining four have not been evaluated (DOF 2010).
Charadrahyla taeniopus (Fig. 1A) inhabits un- disturbed cloud forests (Fig. 1B) and pine-oak forests in the Sierra Madre Oriental (Duellman 2001; Kaplan and Heimes 2015), at elevations from 1,100 to 1,200 m in the states of Hidalgo, Puebla, and Veracruz (Duellman 2001). To date, knowledge about the potential distribution, ecology, reproduction, and natural history of this species is limited (Duellman 2001; Kaplan and Heimes 2015). The primary goal of this study is to assess changes in
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the current and future (2050 and 2070) climatic niche of this species in Mexico. Secondary goals include characterizing the feeding habits, basic reproductive parameters, morphological variation, and conservation status of this species. This information can be used to develop future conservation strategies in environments with high species numbers and endemism, such as the cloud forests of the Sierra Madre Oriental (Ponce-Reyes et al. 2012).
Materials and Methods Data Collection
Occurrence data for C. taeniopus were obtained from (i) the databases of the Global Biodiversity Information Facility (GBIF), the Comision Nacional para el Conocimiento y Uso de la Biodiversidad (CONABIO), and HerpNet; (11) records from publications by Duellman (2001) and Campbell et al. (2009); and (ii1) specimens from field work (sporadic collecting in the state of Hidalgo from 2008 to 2016) deposited in the Coleccion Herpetologica, Centro de Investigaciones Bioldgicas at the Universidad Autonoma del Estado de Hidalgo (Appendix 1).
Climatic Niche Modelling
The 37 unique occurrence records of C. taeniopus were used to generate climatic niche models. For this study, climate information was obtained from the 19 current climate layers available in the WorldClim database version 1.4 (Hijmans et al. 2005). These climate layers contain the averages of meteorological conditions recorded from North America, with a spatial resolution of 2.5 arc-min. To avoid overrepresentation of environmental variables, a bivariate correlation analysis was carried out with the aim of reducing multicollinearity among the input variables (Merow et al. 2013; Varela et al. 2014). For variables that were highly correlated (r > 0.7), the variable was chosen that exhibited the greatest variation or that represented the greatest biological meaning for the actual distribution of the species (e.g., temperature or precipitation). After this procedure, eight climate variables were retained: annual mean temperature (BIO 1), temperature seasonality (standard deviation x 100, BIO 4), maximum temperature of the warmest month (BIO 5), minimum temperature of the coldest month (BIO 6), annual temperature range (BIO 5—BIO 6, BIO 7), mean temperature of the wettest quarter (BIO 8), mean temperature of the coldest quarter (BIO 11), and precipitation in the driest month (BIO 14; Hijmans et al. 2005). Subsequently, MAXENT was used, the selected climate variables were projected onto a map of Mexico and climate change scenarios were estimated (Phillips and Dudik 2008). Each model was replicated 100 times, the maximum number of repetitions allowed by available
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computing power (Dambach and Rodder 2011). Average models for the present and for the future years 2050 and 2070 were then obtained. Current and future projections were estimated using the CCSM-GCM model under the greenhouse concentration scenario RCP8.5, which represents a pessimistic scenario (RCP8.5 = +8.5 W/m”).
Finally, to assess the impact of climate change on habitat suitability for the species, the percent change between current and future potential distribution areas was estimated. To determine habitat suitability, the formula % change = [(S,-S,)/S,]* 100% was used, where S, 1s the total area that the species occupies in the country according to the current distribution scenario, and S, is the total area that the species could occupy in the country under future climate change conditions (Gutiérrez and Trejo 2014).
Diet
Stomach contents were removed from 31 adults and two juveniles (Appendix 1), and a stereomicroscope was used to classify all identifiable organisms to order, including plant material. The number of prey items in each stomach was tallied (n), the number of stomachs with each prey category (i) was determined (F;,), and the percentage of stomachs with prey category 1 (YF) was calculated. Also, the number of prey 1ttems belonging to each prey category (N) and the numerical percentage of total abundance (%N) represented by each prey category were determined. Then the volume of each prey item (mm*) was calculated using the formula for an ellipsoid (Selby 1965; Duré and Kehr 2004; Duré et al. 2009): V = 4/3 n (length/2) (width/2)’. The food importance index (1) was calculated using the formula of Biavati et al. (2004), which is I = (F%+N%+V%)/3, where F% is the percentage of occurrence, N% is numerical percentage, and V% is volumetric percentage. The trophic niche amplitude was measured with Levin’s standardized index using the formula B, = ((1/Xp,7)-1)/n-1, where p, is the proportion of each prey category with respect to the total number of prey found in each sex, and 7 is the number of prey categories in the diet of individuals (Hurlbert 1978). The overlap in dietary composition between sexes was analyzed with Pianka’s index (1986): O,, = P,P. a »P’,,. where p, and P, are the numerical proportions of prey belonging to the 7" category that was used by organisms (sexes) j and & (Pianka 1986; Gadsden and Palacios-Orona 1997). Analyses of amplitude and overlap for males, females, and juveniles were carried out in the Ecological Methodology v. 6.1.1. program (http://www. exetersoftware.com [Accessed: 26 January 2016]).
Reproduction Thirty-one preserved adults (11 females and 20 males; Appendix 1) were examined to assess reproductive
parameters. Fat bodies and livers from all individuals,
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Fig. 2. Current (A), and future 2050 (B) and 2070 (C) scenarios of the climatic niche of Charadrahyla taeniopus in Sierra Madre Oriental province (degraded area). The color scale indicates the probability of occupancy for the species.
and egg mass in females or testes mass in males were weighed to the nearest 0.0001 g using an analytical balance. For males, the length and width of testes were measured to the nearest 0.01 mm with a digital caliper, and testicular volume was calculated using the formula for an ellipsoid (Selby 1965; Duré and Kehr 2004). In females, 10% of the total egg mass was excised from the oviducts, and the eggs contained therein were counted. The eggs were counted in a Petri dish with water using a stereomicroscope. Since the data were not normally distributed, Spearman’s correlation test was used to evaluate correlations between body size and either egg mass or testicular mass. A Mann-Whitney U test was performed to evaluate the differences in mass, volume, length, and width of the testes.
Morphology
For all adult specimens (11 females and 20 males), digital calipers were used to measure the following (each + 0.1 mm): snout-vent length (SVL), internarial distance (IND), eye diameter (ED), interorbital diameter (OD), tympanum diameter (TD), head length (HL), head width (HW), head height (HH), jaw length (JL), jaw width (JW), length from forearm to the fourth finger (LFFT), forearm length (FOL), length of the humerus (LHU), thigh length (THL), tibia length (TL), and foot length (FL; Watters et al. 2016). The normality of the data was checked with a Kolmogorov-Smirnov test. Since the data were normal, an analysis of covariance (ANCOVA) was conducted to analyze sexual dimorphism. The function of the ANCOVA was to eliminate, through linear regressions, the effect of SVL (covariate) on the dependent variables (IND, ED, IOD, TD, HL, HW, HH, JL, JW, LFFT, FOL, LHU, THL, TL, and FL), and to check whether the regression slopes were different between the sexes (factor; Zar 2009). The data are presented as mean + | SE.
Amphib. Reptile Conserv.
Conservation Status
Conservation status was summarized by consulting Mexican regulations (NOM-059-SEMARNAT-2010; DOF 2010) and the IUCN (2019) Red List, and an Environmental Vulnerability Score (EVS; Wilson et al. 2013) was generated. The EVS recognizes three categories of risk: low (3-9 points), medium (10-13), and high (14-19). The score is the result of adding points assigned to features of a species based on three criteria: (i) extent of the geographic distribution, (11) extent of ecological distribution (vegetation types used), and (111) type of reproductive mode (Wilson et al. 2013).
Results
Areas of Occupation and Exchange Rates of Climate Availability
The distribution of C. taeniopus was restricted to cloud forests in the central region of the Sierra Madre Oriental (Fig. 2A—C) in the states of Hidalgo, Puebla, Veracruz, and the northern portion of Oaxaca, Mexico. The analysis of habitat occupancy under current conditions and in the future (to 2050 and 2070), showed a general loss of climatic niche in much of the range of C. taeniopus. This loss occurred in temperate areas, where the current area of occupation (18,262.23 km’) will decrease to 15,678.45 km? by 2050, which represents a habitat availability decrease of 14.14% (Fig. 2B); and to an area of 11,032.93 km? by the year 2070, which represents a habitat availability decrease of 37.08% (Fig. 2C).
Diet
Fourteen taxa were identified in the stomachs, with 11 taxa present in both males and females (Table 1). The
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Table 2. Mean values + 1 SE, and range in parentheses, of morphological characteristics of adult Charadrahyla taeniopus (SVL = snout-vent length, IND = internarial distance, ED = eye diameter, IOD = interorbital diameter, TD = tympanum diameter, HL = head length, HW = head width, HH = head height, JL = jaw length, JW = jaw width, LFFT = length from forearm to the fourth finger, FOL = forearm length, LHU = length of the humerus, THL = thigh length, TL = tibia length, FL = foot length). Comparisons were
made with ANCOVA, with SVL as the covariate.
Characteristics Female (” = 11) Male (n = 20) ain F df P
IND 5.58 + 0.24 (3.87-7.08) 4.94 + 0.18 (2.91-6.72) 1.133 1, 28 0.296 ED 6.22 + 0.23 (5.06-7.91) 5.86 + 0.21 (4.86-8.80) 0.065 1, 28 0.800 IOD 7.02 + 0.36 (4.24-8.74) 5.21 + 0.39 (1.57-8.18) 5.396 1, 28 0.027 TD 3.75 + 0.17 (2.64-4.62) 3.29 + 0.26 (1.99-6.85) 0.457 1, 28 0.504 Ai 19.82 + 0.97 (12.60—24.09) 18.46 + 0.38 (15.81—22.91) 0.002 1, 28 0.966 HW 19.99 + 0.70 (13.94—22.51) 16.45 + 0.32 (14.01-19.41) 24.521 1, 28 <0.001 HH 8.14 + 0.36 (5.56-9.19) 6.83 + 0.32 (4.95-9.08) 3.553 1, 28 0.069 JL 17.28 + 0.66 (12.33—20.68) 15.33 + 0.26 (13.57-17.45) 6.433 1, 28 0.017 JW 19.91 + 0.80 (13.05—22.99) 17.22 + 0.31 (15.20—20.87) 19.166 1, 28 <0.001 LFFT 16.83 + 1.00 (12.03—21.05) 15.60 + 0.53 (11.58-20.37) 0.003 1, 28 0.959 FOL 13.78 + 0.65 (9.44-16.91) 13.12 + 0.29 (11.08-15.58) 1.387 1, 28 0.249 LHU 16.38 + 0.84 (10.77—20.80) 16.07 + 0.51 (10.48-19.26) 2.365 1, 28 0.135 THL 31.32 + 1.44 (20.21-38.05) 30.17 + 0.65 (25.19-37.96) 0.927 1, 28 0.344 su, 32.68 + 1.24 (22.02—37.08) 30.68 + 0.54 (27.84—37.26) 0.016 28 0.900 FL 43.06 + 1.77 (30.21—51.14) 37.92 + 2.17 (34.38-49.26) 0.035 1,27 0.853
most important prey categories, according to the values of food importance for the species and for each sex, were orthopterans, plant material (leaves), and ants (Table 1). The overlap in diet between the sexes was high (O,, = 0.822; 63.21—-100%), with males presenting a slightly higher value (B = 0.526) of diet niche breadth than females (B = 0.504).
Reproduction
The mean number of eggs was 722 + 277.53 (range 426— 1,138, n= 11). There was no correlation between female SVL and either number of eggs (7, = 0.09, P = 0.79, n = 11) or egg mass in females (r, = 0.09, P = 0.79, n = 11). There were no differences among weights, lengths, widths, or volumes of the testes (P > 0.05 in all cases). The average weight, length, width, and volume of the right testis was 0.198 g, 14.78 mm, 6.05 mm, and 305.57 mm*°, respectively; and for the left testis the averages were 0.204 g, 14.63 mm, 6.0 mm, and 293.25 mm’, respectively. There was no correlation between SVL and testicular volume (7, = 0.31, P = 0.17, n = 20), but there was a positive correlation between SVL and testicular mass in males (7, = 0.49, P = 0.02, n = 20).
Morphology
Five of the 15 characteristics measured exhibited sexual dimorphism, with females higher than males in SVL,
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IOD, HW, JL, and JW (Table 2). Females (mean SVL = 63.94 + 2.35 mm; range 45.27-74.31, n = 11) were larger than males (mean SVL = 59.70 + 1.09 mm, range 52.90-71.05, n= 20; U= 60, P = 0.04).
Conservation Status
Charadrahyla taeniopus is listed in conservation standards (DOF 2010; IUCN 2019) as being in high risk categories. According to the Mexican Standard NOM-059-SEMARNAT-2010 (DOF 2010), the species is considered to be Threatened. The IUCN Red List of Threatened Species places the species in the Vulnerable category, with a distributional area less than 20,000 km? in fragmented environments and with declining populations (status Blab[ii]; IUCN 2019). In the EVS, it was classified as a medium environmental vulnerability species, with a value of 13 points. This EVS category was calculated from: (1) its distribution in Mexico, but not exclusive to the type locality (5 points), (1) its occurrence in two vegetation types (pine-oak and cloud forest, 7 points), and (111) a reproductive mode with egg laying in lentic or lotic water bodies (1 point; Wilson et al. 2013).
Discussion
Conserving native populations of tree frogs at a local scale requires information on their ecological distribution,
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feeding habits, reproduction, and morphology (Delia et al. 2013; Toledo et al. 2014). The analyses reported here suggest that the distribution of C. taeniopus will potentially decrease during the next 50 years. Microhabitats in currently occupied habitats (montane environments) are subject to change because of temperature and moisture shifts, and also because of changes in vegetation cover associated with high deforestation rates (Kaplan and Heimes 2015) and potential climate change, including shifts in temperature and moisture (Ponce-Reyes et al. 2012). For example, several authors including Pineda and Halffter (2004), Pineda et al. (2005), and Murrieta- Galindo et al. (2013), have suggested that the existence of abundant vegetation and native shrub cover provide appropriate humidity and temperature conditions for the permanence of hylid frogs in temperate environments such as cloud forests. If the abiotic and biotic conditions change in the forests inhabited by C. taeniopus, this species could be negatively affected. Loss of climatic niche in our models is consistent with that reported by Roxburgh et al. (2004). These authors mentioned that the expected changes could generate ecological scenarios that will delimit the overall distribution of arboreal species from cloud forests (Roxburgh et al. 2004; Pineda et al. 2005), and therefore could affect their associations with their environment (Urbina-Cardona and Flores- Villela 2010; Ponce-Reyes et al. 2012).
In addition to the above considerations, the thermal tolerances of anurans in high elevation or low temperature environments can determine the presence and distribution of their populations (Wells 2007). The hylid frogs are an example of this, as their limits of distribution are in high latitude regions such as the arid and semi-arid climates of northern Mexico (Wiens et al. 2006). This may be the result of the thermal tolerances that hylid species show in temperate environments, which are different from those of species that occur in tropical environments (Navas 2006; Wells 2007). To date, there are no studies of thermal tolerances or maximum/minimum temperature limits for C. taeniopus; therefore, it is very difficult to know the behavior of individuals and/or populations of this species in their distribution area. Future field studies, and in situ and laboratory experiments on thermal preferences are therefore necessary for this species. They could complement the results obtained in the potential distribution model of the species, enabling the analysis of variables that could be interacting to a greater degree with the biology of the organism, and improving determinations of the distribution range of the species (Gross and Price 2000; Wiens et al. 2006).
Ochoa-Ochoa et al. (2009), stated that in addition to the loss of vegetation, climate change is a determining factor in the loss of amphibian species in conserved environments, mainly in sites outside of natural protected areas (NPAs). The Sierra Madre Oriental Corridor occupies large areas of cloud forest, a type of environment that is highly threatened by the effects of
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climate change (Ponce-Reyes et al. 2012), and in which the known distribution of the species is not included in any NPA (IUCN 2019). This shows the importance of evaluating the distribution of highly vulnerable hylid frogs throughout the potential distribution range based on climatic niche models and climate change scenarios. The results are worrisome, because despite the fact that amphibian richness in Mexico is high (Johnson et al. 2015, 2017), more than 50% of the species are listed in high vulnerability categories by the IUCN (Delia et al. 2013; Caviedes-Solis et al. 2015; IUCN 2019; Johnson et al. 2017). For example, recent studies have found that some mountain hylid species have not been recorded over prolonged periods of time (Delia et al. 2013; Caviedes-Solis et al. 2015). Due to multiple factors, such as vegetation loss, pollution, and in particular climate change, populations of these species tend to occur in highly vulnerable sites (Lips et al. 2004; Stuart et al. 2004). Therefore, the species that inhabit this type of environment (cloud forest, pine-oak) are highly threatened (Ochoa-Ochoa et al. 2009; Caviedes-Solis et al. 2015).
Inaddition to habitat fragmentation and climate change, the presence of the pathogenic fungus Batrachochytrium dendrobatidis Longcore, Pessier, and Nichols, 1999 (Bd) has contributed to amphibian population and species losses in Mexico (Mendoza-Almeralla et al. 2015, 2016) and other regions of the world (Lips et al. 2003; Fisher et al. 2009). However, Bd has not been detected thus far in C. taeniopus (Murrieta-Galindo et al. 2014; Hernandez- Austria 2017). Therefore, further studies are needed to examine the potential presence of Bd in C. taeniopus populations through their distribution area (Hernandez- Austria 2017).
The lack of information on the natural history of this species inhibits the development of strategies for its conservation (Toledo et al. 2014). The data presented here on diet provide valuable information on the basic ecology of C. taeniopus. The diet of this species consists of orthopterans, plant material, and ants, and there is a high degree of overlap in diet between the sexes. In C. taeniopus, plant material is the second most important food item. This is particularly notable since the diet of most anuran species in Mexico consists primarily of arthropods (Ramirez-Bautista and Lemos-Espinal 2004; Suazo-Ortufio et al. 2007), and the ingestion of plant material, such as leaves and flowers, is usually considered to be accidental (Evans and Lampo 1996). In the case of C. taeniopus, further studies are necessary to determine if consumption of plant material (leaves) is accidental or part of their diet, which would be unusual, but not unprecedented. For example, some species of tree frogs, such as Ptychohyla zophodes Campbell and Duellman, 2000 (Luria-Manzano 2012) and Xenohyla truncata (Izecksohn, 1959) do consume large quantities of plant material, and the latter (XY. truncata) has been reported as entirely omnivorous, consuming fruits, seeds,
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and flowers (da Silva and Britto-Pereira 2006).
Egg number and the relative sizes of eggs vary greatly in amphibians (Vitt and Caldwell 2009), and they are often related to female body size (Jorgensen 1992; Hartmann et al. 2010). The data presented here show that egg number is not related to female body size in C. taeniopus. This may be due to the fact that its reproductive period may have a longer duration, and the sample size obtained from the collections only reflects the behavior of the females in the first part of the year (March-April), not in the entire reproductive period. Females with eggs were found throughout the year, and aggregations of individuals of both sexes and amplexus were observed in the field in August. This seasonal variation in the correlation between egg size and size of females has been reported for other anuran species such as Leptodactylus fuscus (Schneider, 1799), L. podicipinus (Cope, 1862), and Dendropsophus nanus (Boulenger, 1889) [Prado and Haddad 2005]. Furthermore, testicular mass, but not testicular volume, increases with larger SVL. These data suggest that larger males invest more energy in sperm production to have greater reproductive success (Byrne et al. 2002).
Most species of frogs (nearly 90%) are sexually dimorphic, with females being larger than males (Wells 2007), and C. taeniopus is no exception. The larger size of females compared to males is presumably associated with the potential to produce more eggs. However, no correlation was found between SVL of females and egg number. Another explanation for sexual size dimorphism could be differences in growth rates (Kupfer 2007), in which the growth rate of males is faster than that of females in order to reach sexual maturity at a smaller size and compete with other males for access to calling sites, thereby maximizing the number of matings (Kupfer 2007; Wells 2007). Also, considering the ecological hypothesis to explain the sexual dimorphism, the larger jaw size in females compared to the males might indicate a larger gape in females, which could allow for partitioning of food resources in terms of prey size (Luria-Manzano 2012). However, additional studies on microhabitat use, behavior, and reproduction are required before the ecological significance of the sexual dimorphism in C. taeniopus can be determined.
Based on the information about climatic niche, feeding habits, reproduction, and morphology, C. taeniopus is highly threatened because it is distributed in environments (i.e., cloud, oak, and pine-oak forests) that are currently being dismantled by fragmentation and climate change (Ponce-Reyes et al. 2012). As with other hylid frogs (Caviedes-Solis et al. 2015), C. taeniopus could face a rapid rate of population decline, as has occurred in other species inhabiting the temperate areas of cloud forest in Oaxaca (Delia et al. 2013; Mata-Silva et al. 2015), Chiapas (Johnson et al. 2015), and areas of the Sierra Madre Oriental (Flores-Villela et al. 2010). To add to the information presented in this study, additional
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studies on demography, ecology, physiological tolerances to temperature, length of the reproductive period, effect of fragmentation on populations, and population dynamics of this species should be conducted in order to devise efficient conservation strategies for C. taeniopus, and other species of anurans that inhabit the temperate and tropical montane environments of central and southern Mexico (Delia et al. 2013; Caviedes-Solis et al. 2015).
Acknowledgments —We thank Abraham Lozano, J. Daniel Lara Tufifio, Diego Juarez Escamilla, Concepcion Puga, Uriel Hernandez Salinas, and Luis M. Badillo Saldafia for their help in the field and laboratory. We thank three anonymous reviewers for comments that greatly improved the manuscript. We thank Margaret Schroeder for her revision of the English. Thanks to Irene Goyenechea Mayer Goyenechea for providing the numbers of specimens from Colecci6n Herpetologica CIB-UAEH. This study was supported by the CONABIO JMO001, Fomix-CONACyT-191908 Biodiversidad del Estado de Hidalgo-3a, UAEH-DI-ICBI-BI-SF-008 and ICBIPAI-29 projects. Specimens were collected under SEMARNAT-SGPA/DGVS/02726/10 and SGPA/ DGVS/11746/13 permits issued by SEMARNAT and assessed by Mexican environmental protection laws.
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Appendix 1. Voucher numbers of Charadrahyla taeniopus specimens analyzed in this study from Coleccion Herpetologica del Centro de Investigaciones Biologicas, Universidad Autonoma del Estado de Hidalgo, México. Year 2008, September: CIB 5422-5425 (males), December: CIB 5426 (female); 2009, April: CIB 5427 (female), June: CIB 5428 (male), CIB 5429-5430 (females), July: CIB 5430 (female), August CIB 5431 (male), CIB 5432 (female), October: CIB 5433-5435 (males), CIB 5436-5437 (juveniles, females), November CIB 5438 (female), CIB 5439 (juvenile, female); 2010, June: CIB 5440 (male); 2011, April CIB 5441 (female), CIB 5442-5443 (males), CIB 5444 (female), CIB 5445 (juvenile, female), CIB 5446 (male), May CIB 5447-5449 (males), June CIB 5450 (male); 2012, March CIB 5451-5452 (females), CIB 5453 (male); 2015, April CIB 5454-5545 (males), CIB 5456 (female).
Raciel Cruz-Elizalde is a Mexican herpetologist who received his B.Sc. in Biology, M.Sc. in Biodiversity and Conservation, and Ph.D. in Biodiversity and Conservation from the Autonomous University of Hidalgo State, Mexico. He is currently conducting a postdoctoral stay at the Department of Evolutionary Biology, Faculty of Science, National Autonomous University of Mexico (UNAM). Raciel is interested in the ecology, life history evolution, diversity, and conservation of amphibians and reptiles in Mexico. His current research includes life history evolution of diverse lizard species of the genus Sceloporus, conservation issues in natural protected areas, and analysis of ecological and morphological traits in the composition of amphibian and reptile species assemblages.
Itzel Magno-Benitez is a Mexican herpetologist. Itzel recetved her B.Sc. in Biology from the Autonomous University of Hidalgo State, Mexico, and she is interested in the ecology and conservation of amphibians and reptiles in Mexico. Her current research interests include the anthropic effects on conservation and ecology (feeding habits, reproduction, and morphological variation) of amphibians and reptiles in the temperate and arid environments of Mexico.
Christian Berriozabal Islas received a Ph.D. from the Autonomous University of Hidalgo State, Mexico. His research focuses on the diversity of amphibians and reptiles, conservation, and the effects of climate change on tropical lizards and kinosternid turtles.
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Raul Ortiz-Pulido is a Mexican scientist, amateur astronomer, scientific communicator, and artist (landscape photographer). Ratil received a B.Sc. degree from the Universidad Veracruzana (1994) and a doctorate in sciences from the Instituto de Ecologia, A.C. (2000). He has been working with animal ecology since 1991, particularly the relationship between food abundance and individuals of different species. In 1997, Raul started the Huitzil Mexican Journal of Ornithology and in 2002 the astronomical society of his current university. In 2005, he was elected president of CIPAMEX, a Mexican association of scientists. Raul has written more than 200 scientific papers and given more than 250 talks, many of them popularizing science. He is a reviewer for 27 indexed scientific journals and associate editor of four of them. Currently, Raul is associated with the Population Ecology Laboratory of the Universidad Autonoma del Estado de Hidalgo, where he has worked since 2001.
Aurelio Ramirez-Bautista began his herpetological career conducting research as an undergraduate student at the Los Tuxtlas Biological Field Station, Veracruz, Mexico. Aurelio received his B.Sc. in Biology from Universidad Veracruzana in Veracruz, Mexico, his M.Sc. and Doctorate degrees from the Universidad Nacional Autonoma de México (UNAM), and a postdoctoral appointment at the University of Oklahoma, Norman, Oklahoma, USA. Aurelio’s main research involves studies on ecology, demography, reproduction, conservation, and life history evolution, using amphibians and reptiles of Mexico as models. He was president of the Sociedad Herpetologica Mexicana and is currently associate editor of Mesoamerican Herpetology. Aurelio is a professor at Universidad Autonoma del Estado de Hidalgo (UAEH), teaching population ecology, herpetology, and natural history of amphibians and reptiles.
Raquel Hernandez-Austria is a biologist with an M.Sc. in Biodiversity and Conservation Sciences from the Universidad Autonoma del Estado de Hidalgo, Mexico. Raquel’s interest is the study of amphibian and reptile diversity and conservation. For her undergraduate degree, she compared the food habits of two syntopic species of Lithobates, and for her Master’s degree she evaluated the presence of Batrachochytrium dendrobatidis in anuran species of Hidalgo State. Raquel has participated in various Mexican congresses and is an author and co-author of many scientific papers related to amphibians and reptiles.
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“eptile-conse
Detection of Ophidiomyces ophiodiicola at two mid-Atlantic natural areas in Anne Arundel County, Maryland and Fairfax County, Virginia, USA
‘*Lauren D. Fuchs, ?Todd A. Tupper, *Robert Aguilar, ‘Eva B. Lorentz, ?Christine A. Bozarth, 2David J. Fernandez, and *David M. Lawlor
'George Mason University, Department of Systems Biology, 10900 University Blvd, Manassas, Virginia 20110 USA *Northern Virginia Community College, Division of Math, Science, Technologies and Business, 5000 Dawes Avenue Alexandria, Virginia 2231 USA °*Smithsonian Environmental Research Center, Fish and Invertebrate Ecology Lab, 647 Contees Wharf Road, Edgewater, Maryland 21037 USA *Huntley Meadows Park, Natural Resource Management Division, 3701 Lockheed Blvd Alexandria, Virginia 22306 USA
Abstract.—Since the early 2000s, ophidiomycosis has been reported with increasing frequency and associated with widespread morbidity in numerous North American snake species. Ophidiomyces ophiodiicola (Oo), the etiologic agent of ophidiomycosis, has been detected in over 30 species throughout most of the eastern United States, as well as in Europe and Australia; however, it is suspected that the distribution of this pathogen may be underestimated due to a lack of standardized inventories. To contribute to the existing but limited data on ophidiomycosis in the mid-Atlantic United States, snakes were sampled for Oo at two natural areas in this region—one in Anne Arundel County, Maryland and one in Fairfax County, Virginia. Ophidiomyces ophiodiicola was detected at both study sites. Thirty-four of 61 (55.7%) samples across eight species tested positive for the pathogen, with the highest detection rates occurring in Nerodia sipedon (73.1%) and Pantherophis alleghaniensis (70%). Ophidiomyces ophiodiicola was detected in snakes with (71.4%) and without (34.6%) Clinical signs of ophidiomycosis. These results support the need for both increased Oo monitoring throughout
the region, and implementation of more standardized and unbiased sampling protocols.
Keywords. Colubridae, ophidiomycosis, population decline, Reptilia, Serpentes, snake fungal disease
Citation: Fuchs LD, Tupper TA, Aguilar R, Lorentz EJ, Bozarth CA, Fernandez DJ, Lawlor DM. 2020. Detection of Ophidiomyces ophiodiicola at two mid-Atlantic natural areas in Anne Arundel County, Maryland and Fairfax County, Virginia, USA. Amphibian & Reptile Conservation 14(1) [General
Section]: 22—28 (e220).
Copyright: © 2020 Fuchs et al. This is an open access article distributed under the terms of the Creative Commons Attribution License [Attribution 4.0 International (CC BY 4.0): https://creativecommons.org/licenses/by/4.0/], which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. The official and authorized publication credit sources, which will be duly enforced, are as follows: official journal title Amphibian & Reptile Conservation; official journal website: amphibian-reptile-conservation.org.
Received: 18 April 2019; Accepted: 5 January 2020; Published: 12 February 2020
Introduction
Ophidiomycosis has emerged as a growing threat to snakes throughout much of North America (Dolinski et al. 2014; Allender et al. 2015; Lorch et al. 2016; Paré and Sigler 2016) and has been associated with widespread morbidity in numerous species (Guthrie et al. 2016: Lorch et al. 2016; Stengle 2018). The disease is attributed to Ophidiomyces ophiodiicola (Oo), a mycotic pathogen that is only known to infect snakes (Allender et al. 2015; Lorch et al. 2016; Paré and Sigler 2016). Clinical manifestations of infection (see Fig. 1) typically include scabs, crusty scales, superficial pustules, subcutaneous nodules, and dysecdysis (Dolinski et al. 2014; McBride et al. 2015; Tetzlaff et al. 2015). Ophidiomycosis infections are generally chronic, but mild; however, severe infections with high mortality have been reported in several viperid species (Allender et al. 2013; Sigler et
Correspondence. */fuchs@masonlive.gmu.edu
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al. 2013; Sleeman 2013; Lorch et al. 2015, 2016; Stengle et al. 2018). The precise mechanisms that influence lethal outcomes of the disease are still unclear, but are likely multifaceted (Lorch et al. 2015; Guthrie et al. 2016). Since 2006, ophidiomycosis has been increasingly documented, with cases of infection reported in at least 20 states, including Maryland and Virginia (Allender et al. 2015; Guthrie et al. 2016; Tupper et al. 2015, 2018, 2019). Despite growing reports of ophidiomycosis throughout the mid-Atlantic, systematic studies designed to assess its prevalence using non-incidental sampling methods are limited. The rising incidence of ophidiomycosis, coupled with habitat loss, pollution, and other anthropogenic stressors, poses an added challenge for snake conservation, underscoring the significance of ongoing disease monitoring (Franklinos et al. 2017; Kucherenko et al. 2018). The objective of this study was to assess the presence and prevalence of Oo at two mid-
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Fig. 1. The ventral scales of a symptomatic Northern Black Racer (Coluber constrictor) infected with Ophidiomyces ophiodiicola. This 41 g male was captured and swabbed on 20 May 2018 at Huntley Meadows Park. Its total length was 123.2 cm and snout-to-vent length was 94.6 cm. Photo by Eva Lorentz.
Atlantic natural areas located in Maryland and Virginia, USA. The results obtained contribute toward an improved understanding of the distribution and prevalence of ophidiomycosis in the region.
Materials and Methods
Area-constrained visual encounter searches (Crump and Scott 1994) were used to sample for Oo in snakes from Huntley Meadows Park (HMP; 38°45’36.57” N, 77°05’44.13” W; Fig. 2) in Fairfax County, Virginia, and at the Smithsonian Environmental Research Center (SERC; 38°53717.41”N, 76°33’ 15.52” W; Fig. 3) in Anne Arundel County, Maryland, between 22 April 2018 and 9 October 2018. Snakes were hand-captured (wearing sterile nitrile gloves) and visually inspected for clinical signs of ophidiomycosis (Allender et al. 2011; Clark et al. 2011). Then, using a modified protocol developed by Allender et al. (2016), snake skins were sampled with sterile dry swabs (no. MW113, Medical Wire and Equipment Company, Durham, North Carolina, USA) from all craniofacial scales and along the entire ventral length of the body separately, swabbing each region five times, taking care to swab any lesions, pustules, nodules, or displaced scales on snakes which showed signs of infection (Allender et al. 2011, 2016). Swabs were stored in sterile 1.5 mL microcentrifuge tubes and kept frozen until molecular analysis. Prior to release, each snake was measured, weighed, and photographed to help in differentiating conspecifics. Aseptic techniques were employed and appropriate biosecurity protocols were followed (see Rzadkowska et al. 2016; VHS 2016) to limit the transmission of Oo.
For the Oo assay, DNA was eluted from the swabs using the Purification of Total DNA from Animal Tissues Protocol (Qiagen®, Valencia, California, USA). To ensure samples were not contaminated during the
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Oe 05.0% | 2 a es Kilometers
Fig. 2. Location of Huntley Meadows Park (HMP) snake capture locations. Black markers = all samples positive; white markers = all samples negative; gray markers = samples either positive or negative.
extraction process, a negative control was used, which included all elements of the extraction mixture other than DNA. Following methods described by Allender et al. (2015), 2.5 uL of eluted DNA was combined with 12.5 uL Sso Advanced™ universal probes supermix (Bio- Rad, Hercules, California, USA), 1.25 uL of a combined target-specific primer (OphioITS-F and OphiolTS-R)- probe, and water, creating a 25 uL reaction mixture. The DNA was amplified via GPCR using a CFX96 Touch™ Real-Time PCR Detection System (Bio-Rad, Hercules, California, USA), with the following cycling parameters: 1 cycle at 50 °C for 2 min, 1 cycle at 95 °C for 10 min, 40 cycles of 95 °C for 15 sec and 60 °C for 60 sec, followed by a final cycle at 72 °C for 10 min.
For each round of qPCR, a positive control was included by adding 2.5 uL of a plasmid containing Oo (obtained from the Wildlife Epidemiology Laboratory at Illinois University at Urbana-Champaign, Illinois, USA) to a designated well containing the 22.5 uL mixture of primer-probe, and water (as described above). A well was also included for the negative control, which contained only the 22.5 uL mixture, but no DNA. These controls were used to determine whether the reaction mixture was prepared accurately, and to ensure that samples were not contaminated during qPCR preparation. Up to five rounds of qPCR were performed for each sample. A sample was considered positive if at least three rounds (per sample) had a lower cycle threshold (C,) than the
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: 2 es Kilometers
Fig. 3. Location of Smithsonian Environmental Research Center (SERC) snake capture locations. Black markers = all samples positive; white markers = all samples negative; gray markers = samples either positive or negative.
lowest detected standard dilution for the positive control (Allender et al. 2016).
Sampling sites within the study areas (indicating locations of positive and negative samples) were plotted with ESRI ArcMap (version 10.6). Snake nomenclature corresponds with Crother et al. (2017). Tables and descriptive statistics were completed with Microsoft Excel for Office 365 (Microsoft Corporation, Redmond, Washington, USA).
Results
Sixty snakes (35 from HMP and 25 from SERC) across nine species were captured and swabbed (Table 1). Northern Watersnake (Nerodia sipedon) comprised the largest proportion (n = 26; 43.3%) of the captures. Eastern Ratsnake (Pantherophis alleghaniensis, n= 10), Common Ribbonsnake (Thamnophis sauritus, n = 9), and Eastern Wormsnake (Carphophis amoenus, n = 7) were also well- represented, comprising 16.7%, 15%, and 11% of the total snake sample, respectively. Northern Black Racer (Coluber constrictor, n = 2), Eastern Kingsnake (Lampropeltis getula; n = 1), Northern Ring-necked Snake (Diadophis punctatus, n = 1), Eastern Gartersnake (Thamnophis sirtalis, n= 2), and Dekay’s Brownsnake (Storeria dekayi; n = 2) were all sparsely represented. Ophidiomyces ophiodiicola was detected in 33 snakes and in a shed skin of a Northern Black Racer, yielding an overall detection rate of 55.7%. More than half of the positive samples (55.9%) were from a single species—Northern Watersnake. Of the nine species sampled, Northern Watersnake had the highest detection rate (73.1%), followed closely by Eastern Ratsnake (70%). Northern Black Racer, Eastern Wormsnake, and Common Ribbonsnake were positive in 66.7%, 28.6%, and 11.1% of samples, respectively. Only one Eastern Kingsnake and one Northern Ring-necked snake were sampled, and both were positive. Dekay’s Brownsnake was positive in one of two samples and Eastern Gartersnake was the only species that did not test positive for Oo. Twenty-five of the 35 (71.4%) snakes showing clinical signs tested positive for Oo, and nine of the 26 (34.6%) without clinical signs were Oo positive (Table 1). Prevalence varied between study locations, with 34.6% of snakes testing positive at HMP and 84.6% at SERC. Of the 34 snakes testing positive, Oo was detected in both swabs in 18 snakes (52.9%) and in only one of two swabs (nine from the craniofacial swab only, seven from the body swab only) in 16 snakes (47.1%).
Table 1. Prevalence by species. S/+ = positive with clinical signs, A/+ = positive without clinical signs.
Species Eastern Wormsnake (Carphophis amoenus amoenus) Northern Black Racer (Coluber constrictor constrictor)
Northern Ring-Necked Snake (Diadophis punctatus edwardsii)
Eastern Kingsnake (Lampropeltis getula)
Northern Watersnake (Nerodia sipedon sipedon) Eastern Ratsnake (Pantherophis alleghaniensis) Dekay's Brownsnake (Storeria dekayi)
Common Ribbonsnake (Thamnophis saurita saurita) Eastern Gartersnake (Thamnophis sirtalis sirtalis) Total or overall prevalence
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24
Prevalence (%)
N_ Positive Forspecies Overall S/+ A/+ 7 2 28.6 59 0 es 3 2 66.7 5.9 2 0 1 1 100 2.9 0 1 1 1 100 2.9 1 0 26 19 73.1 559 15 4 10 7 70 20.6 > 2 2 1 50 2.9 1 0 9 1 11.1 2.9 1 0 2 0 0 0 0 0 61 34 - 55.7 25 9
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Discussion
Although Oo has previously been documented in Maryland and Virginia (Guthrie et al. 2016; Tupper et al. 2018), this work is one of only two studies (see Guthrie et al. 2016) to investigate Oo in these states. In Maryland, observations of fungal dermatitis have been reported from the Smithsonian Environmental Research Center (SERC) since 2014 (Tupper et al. 2015), with Oo recently being confirmed as the etiological agent of a dermal infection in Northern Watersnake (Tupper et al. 2018). These results add four new species (Eastern Wormsnake, Northern Black Racer, Northern Ring-necked Snake, and Eastern Ratsnake) to the documented host range of this pathogen in Maryland, which previously included only Northern Watersnake (Tupper et al. 2018) and Timber Rattlesnake (Crotalus horridus;, Tupper et al. 2019). In eastern Virginia, Guthrie et al. (2016) documented Oo in four species (all with clinical signs): Northern Watersnake (n= 3), Rainbow Snake (Farancia erytrogramma; n= 1), Northern Black Racer (n = 2), and Brown Watersnake (Nerodia taxispilota, n = 2). This study adds four new hosts to the list of Oo positive species occurring in Virginia: Eastern Kingsnake, Eastern Ratsnake, Dekay’s Brownsnake, and Common Ribbonsnake.
The overall detection rate of 57.4% is among the highest reported (except see McKenzie et al. 2018) across the eastern and midwestern United States (Smeenk et al. 2016; Allender et al. 2016). The prevalence of Oo throughout these regions appears to be highly variable, with detection rates as low as 0% and 4.9% in Ohio and Michigan, respectively (Smeenk et al. 2016; Allender et al. 2016), and up to nearly 62% in eastern Kentucky (McKenzie et al. 2018). We interpret these rates cautiously, however, taking into consideration the variation in species sampled between studies. It is still unclear how susceptibility and severity of infection differ between species (Grisnik et al. 2018), but the composition of species sampled in this study may partly explain the overall prevalence and the relatively high proportion of Oo positive snakes that did not show clinical signs of the disease.
In this study, Oo was detected in eight of the nine species sampled, which was not surprising given that each of these species has previously tested positive for the pathogen in the eastern and mid-western United States (Lorch et al. 2016; Persons et al. 2017; Grisnik et al. 2018; McKenzie et al. 2018). However, the small sample sizes in certain species made it impossible to assess how each of these species actually influence the overall detection rate. Ophidiomyces ophiodiicola was found to be most prevalent in Northern Watersnake, with a detection rate of 73%. This species represented nearly 43% of the total sample and thus had a strong influence on overall prevalence (55.7%). Prior studies with similar proportions of aquatic species have also demonstrated relatively high Oo detection rates among Northern
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Watersnakes and other species with aquatic affiliations. However, this trend in detection may partially reflect the habitat preferences of the pathogen (Lorch et al. 2016; McKenzie et al. 2018), rather than an inherent biological susceptibility to the pathogen. Additional work is needed to better understand susceptibility to the disease.
Variability in sampling methods between studies should also be considered when interpreting results (McCoy et al. 2017; Grisnik et al. 2018; Hileman et al. 2018; McKenzie et al. 2018). For instance, the number of sterile dry swab applicators used per snake has been shown to influence detectability of Oo, with the use of only one applicator greatly increasing the probability of obtaining false-negatives (Hileman et al. 2018). The results obtained here support this concept, with 47.1% of snakes testing positive for Oo in only one of two swabs.
Underestimation of the prevalence of Oo may also occur when diagnostic tests are limited only to snakes that present clinical manifestation of infection (see Guthrie et al. 2016). While clinical signs have been associated with a higher probability of PCR-positive results (Allender et al. 2016), studies have also demonstrated that anywhere from 6% (Bohuski et al. 2015) to 38% (Hileman et al. 2018) of snakes without clinical signs test positive for Oo. The data reported here support these studies, with 26.5% of Oo positive snakes in this sample showing no signs of infection. One possible explanation is that clinical signs may be subtle and overlooked during inspection, because a snake is either in the early stages of infection or effectively clearing the infection through repeated sheds (Lorch et al. 2016; Grisnik et al. 2018; Hileman et al. 2018). Detection without clinical signs may also reflect the absence of infection in a specimen altogether. Ophidiomyces ophiodiicola can persist as a saprobe in the soil, which can facilitate transmission and increase the likelihood of a snake encountering Oo (Allender et al. 2015; Lorch et al. 2016). The presence of Oo on the skin, however, does not necessarily indicate infection. Therefore, while swabbing can be an effective, low-cost, and minimally invasive method for detecting the pathogen, it cannot be used to infer or imply infection status.
Results from this study confirm that Oo is present and relatively prevalent in both Maryland and Virginia, and that the presence of Oo is more often accompanied by clinical manifestations consistent with ophidiomycosis than not. The geographic distribution and host range of the pathogen are still largely unknown (Burbrink et al. 2017), and ophidiomycosis may be more widely distributed than documented cases suggest (USGS 2018). Some have proposed that biased sampling methods may result in underestimations of prevalence within a population (Grisnik et al. 2018; Hileman et al. 2018). This potential for inaccurate assessments highlights the need for more standardized sampling efforts and diagnostic protocols. Based on the increasing number of reports of ophidiomycosis throughout the eastern United States, we suggest increased efforts to identify and monitor
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Oo throughout the mid-Atlantic region. Additionally, enhanced biosecurity protocols should be implemented to limit disease transmission throughout the region.
Acknowledgements.—This work was funded by a NOVA Foundation Innovation Grant. Valerie Czach, Amanda Lee, and Ashely Davis assisted with field and lab work. Maggie Emblom-Callahan, Susan Williams, Tatiana Stantcheva, and the NOVA Police Department provided logistical support. This work was approved by the Institutional Animal Care and Use Committee at George Mason University (reference #0396), the Virginia Department of Game and Inland Fisheries (permit # 062364), and the Maryland Department of Natural Resources (permit #57025).
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Sigler L, Hambleton S, Pare JA. 2013. Molecular characterization of reptile pathogens currently known as members of the Chrysosporium anamorph of Nanniziiopsis vriesii complex and relationship with some human-associated isolates. Journal of Clinical Microbiology 51: 3,338-3,357.
Sleeman J. 2013. Snake fungal disease in the United States. National Wildlife Health Center Wildlife Health Bulletin 2013(2): 1-3.
Smeenk NA, Lipps Jr. GJ, Allender MC, Backus K, Freeman M, Hribar D, Parsley M. 2016. Snake fungal disease, Ophidiomyces ophiodiicola, in Ohio, USA.
Herpetological Review 47: 592-594.
Stengle A. 2018. Habitat selection, connectivity, and population genetics of a Timber Rattlesnake (Crotalus horridus) metapopulation in southwestern Massachusetts and New England. Ph.D. Dissertation, University of Massachusetts, Amherst, Department of Organismic and Evolutionary Biology. 213 p.
Tetzlaff S, Allender MC, Ravesi M, Smith J, Kingsbury B. 2015. First report of snake fungal disease from Michigan, USA involving Massasaugas, Sistrurus catenatus (Rafinesque 1818). Herpetology Notes 8: 31-33.
Tupper TA, Aguilar R, Ferrara A, Hay C. 2015. Field notes: Snake fungal disease. Catesbeiana 35: 36-37. Tupper TA, Aguilar R, Fuchs LD. 2018. Field notes: snake fungal disease: MD. Catesbeiana 38: 78-79. Tupper TA, Fuchs LD, Aguilar R. 2019. Molecular confirmation of Ophidiomyces ophiodiicola in Crotalus horridus and Agkistrodon contortrix (Allegany County, Maryland and Rappahannock County, Virginia).
Catesbeiana 39: 35-37.
USGS (United States Geological Survey). 2019. Snake fungal disease. Available: https:/Awww.usgs.gov/ centers/nwhc/science/snake-fungal-disease [Accessed 20 January 2020].
VHS (Virginia Herpetological Society). 2016. Disinfection protocol. Available: http://www. virginiaherpetologicalsociety.com/research/disease/ index.htm [Accessed 1 March 2019].
WVDNR (West Virginia Division of Natural Resources). 2017. Snake fungal disease detected in West Virginia’s snakes. Available: —http://www.wvdnr. gov/2017news/17news098.shtm [Accessed | February 2019].
Amphib. Reptile Conserv.
Lauren Fuchs is a Ph.D. student in Environmental Science and Policy at George Mason University (Fairfax, Virginia, USA). Lauren is also a graduate teaching assistant, and volunteer at the Smithsonian Environmental Research Center (Edgewater, Maryland, USA). Her main area of interest is in the diseases of herpetofauna.
Todd Tupper is a Professor of Biology at Northern Virginia Community College (Alexandria, Virginia, USA), an affiliate faculty member at George Mason University (Fairfax, Virginia, USA), and a Visiting Researcher at the Smithsonian Environmental Research Center (Edgewater, Maryland, USA). Todd teaches general biology, zoology, and biostatistics. His areas of interest are biology education, and amphibian and reptile monitoring and conservation.
Robert Aguilar is a biologist at the Smithsonian Environmental Research Center (Edgewater, Maryland, USA). His areas of interest are quite varied and include fish and invertebrate ecology and phylogenetics, herpetological diseases, wildlife inventory and monitoring, and science education. Robert helps with the Maryland Biodiversity Project and supervises undergraduate researchers at the Smithsonian Environmental Research Center.
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Ophidiomycosis in the mid-Atlantic USA
Eva Lorentz is an undergraduate senior in Biology at George Mason University (Fairfax, Virginia, USA) and a volunteer for the Smithsonian Environmental Research Center (Edgewater, Maryland, USA).
Christine Bozarth is an Associate Professor of Environmental Science at Northern Virginia Community College (Alexandria, Virginia, USA). Her areas of interest are in population genetics, wildlife inventory and monitoring, and science education.
David Fernandez is a Professor of Biology at Northern Virginia Community College (Alexandria, Virginia, USA). David teaches cell biology, general biology, and anatomy and physiology. His areas of interest are in hepatic cancers and wildlife disease. His areas of interest are in hepatocyte biology, tumor biology, and herpetological diseases.
David Lawlor is a biologist at Huntley Meadows Park in Alexandria, Virginia, USA. His areas of interest are in natural history, natural resource management, and science education. David also coordinates and oversees the wildlife research that occurs at Huntley Meadows.
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Official journal website: amphibian-reptile-conservation.org
Amphibian & Reptile Conservation 14(1) [General Section]: 29-42 (e221).
A
AEs ae
le-conse*
Insights into the natural history of the endemic Harlequin
Toad, Atelopus laetissimus Ruiz-Carranza, Ardila-Robayo,
and Hernandez-Camacho, 1994 (Anura: Bufonidae), in the Sierra Nevada de Santa Marta, Colombia
'*Hernan D. Granda-Rodriguez, 7Andrés Camilo Montes-Correa, *Juan David Jiménez-Bolafo, ‘Alberto J. Alaniz, Pedro E. Cattan, and °Patricio Hernaez
'Programa de Ingenieria Ambiental, Facultad de Ciencias Agropecuarias, Universidad de Cundinamarca, Facatativa, COLOMBIA *%Grupo de Investigacion en Manejo y Conservacion de Fauna, Flora y Ecosistemas Estratégicos Neotropicales (MIKU), Universidad del Magdalena, Santa Marta, COLOMBIA ‘Centro de Estudios en Ecologia Espacial y Medio Ambiente, Ecogeografia, Santiago, CHILE °Facultad de Ciencias Veterinarias y Pecuarias, Universidad de Chile, Santiago, CHILE °Centro de Estudios Marinos y Limnologicos, Facultad de Ciencias, Universidad de Tarapaca, Arica, CHILE
Abstract.—Atelopus laetissimus is a bufonid toad that inhabits the mountainous areas of the Sierra Nevada de Santa Marta (SNSM), Colombia. This species is endemic and endangered, so information about its ecology and distribution are crucial for the conservation of this toad. Here, the relative abundance, habitat and microhabitat uses, and vocalization of A. /laetissimus are described from the San Lorenzo creek in the SNSM, as well as its potential distribution in the SNSM. To this end, 447 individuals were analyzed during several sampling trips from 2010 to 2012. Against expectations, population density was significantly higher in the stream than in the riparian forest. Overall, A. laetissimus used seven different diurnal microhabitats, with a high preference for leaf litter substrates and rocks. The rate of recaptures decreased linearly across the survey nights. Two types of vocalizations related to the advertisement call of A. latissimus were recorded: a series of pulsed calls like a buzz and another short call, lacking pulses or partially pulsed. According to this analysis, the areas with higher habitat suitability for A. laetissimus were located principally in the northern and northwestern regions of the SNSM, in agreement with literature. Moreover, the data modeling indicated a significant increase in habitat loss from 2013 to 2017. The information presented here should be considered as a starting point for the conservation of this species.
Keywords. Advertisement call, amphibian decline, conservation, ecology, habitat loss, habitat suitability, home range, microhabitat selection, nocturnal site fidelity
Resumen.—Atelopus laetissimus es un sapo de la familia Bufonidae que habita las zonas montanosas de la Sierra Nevada de Santa Marta (SNSM), Colombia. Para esta especie endemica y en peligro, la informacion ecologica y de distribucion es crucial para su conservacion. En el presente trabajo describimos la abundancia relativa, usos de habitat y microhabitat, y las vocalizaciones de A. laetissimus, asi como su distribucion potencial en la SNSM. Para esto, analizamos 477 individuos durante varios muestreos entre 2010 y 2012. Contra las expectativas, la densidad poblacional fue significativamente mayor en el lecho de la quebrada que en el bosque ribereno adyacente. En general, A. laetissimus utilizo siete microhabitats diurnos, con una alta preferencia por los sustratos de hojarasca y rocosos. La tasa de capturas decrecio linealmente a lo largo de los muestreos nocturnos. Registramos dos tipos de vocalizaciones relacionadas con el llamado de anuncio de A. laetissimus. Una serie de Ilamados pulsados como zumbidos y otros mas cortos, con pulsos ausentes oO parcialmente pulsados. De acuerdo con nuestros analisis, las areas con mayor idoneidad de habitat se localizan en los sectores septentrionales y noroccidentales de la SNSM, lo que es concordante con la literatura. Ademas, el modelo construido indica un incremento significativo de la pérdida de habitat entre 2013-2017. Esta informacion debe ser considerada como punto de partida para la conservacion de esta especie.
Palabras clave. Ambito doméstico (home range), conservacion, declive de los anfibios, ecologia, fidelidad de percha nocturna, idoneidad de habitat, llamado de anuncio, pérdida de habitat, seleccion de microhabitat
Citation: Granda-Rodriguez HD, Montes-Correa AC, Jiménez-Bolafio JD, Alaniz AJ, Cattan PE, Hernaez P. 2020. Insights into the natural history of the endemic Harlequin Toad, Atelopus /aetissimus Ruiz-Carranza, Ardila-Robayo, and Hernandez-Camacho, 1994 (Anura: Bufonidae), in the Sierra Nevada de Santa Marta, Colombia. Amphibian & Reptile Conservation 14(1) [General Section]: 29-42 (e221).
Copyright: © 2020 Granda-Rodriguez et al. This is an open access article distributed under the terms of the Creative Commons Attribution License [Attribution 4.0 International (CC BY 4.0): https://creativecommons.org/licenses/by/4.0/], which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. The official and authorized publication credit sources, which will be duly enforced, are as follows: official journal title Amphibian & Reptile Conservation; official journal website: amphibian-reptile-conservation.org.
Received: 3 April 2019; Accepted: 31 October 2019; Published: 17 February 2020
Correspondence. !*hernangrandar@gmail.com;, *andresc.montes@gmail.com, * herpetos4@gmail.com; * alberto.alaniz@ug.uchile.cl, ° pahernaez@gmail.com
Amphib. Reptile Conserv. 29 February 2020 | Volume 14 | Number 1 | e221
Natural history of Harlequin Toad, Atelopus /aetissimus in Colombia
Introduction
The harlequin toads (Bufonidae: Ate/opus) are small amphibians (<10 cm) which have aposematic coloration and predominantly diurnal activity periods (Lotters 1996). With a worldwide diversity of 96 described species (Frost 2019), the conservation of these amphibians has been seriously affected during the last decades due to the population decreases detected in a majority of species and the extinction of others, and currently 97.92% of the species of this genus are included in the IUCN Red List of Threatened Species, with one extinct species (Young et al. 2001; La Marca et al. 2005; Gascon et al. 2007; Tapia et al. 2017; IUCN 2019). Among the main risk factors for the decline of these populations are the loss of habitat, the introduction of exotic species which are potential predators and competitors, as well as deaths by pathogens (mainly by the chytrid fungus Batrachochytrium dendrobatidis [Bd]|) and _ climate change (Lotters 2007; Catenazzi 2015; Barrio-Amoros and Abarca 2016; Valenzuela-Sanchez et al. 2017).
In general, information about the population biology and ecology of the different Ate/opus species 1s relatively scarce. For example, the males of the species Ate/opus cruciger (Lichtenstein and Martens, 1856) are known to remain longer in the streams than the females (Sexton 1958). But, on the other hand, studies conducted on Atelopus carbonerensis (Rivero, 1974 “1972”) in Venezuela, found that they remain almost all year in their habitat, except in the dry season when individuals migrate towards the streams for reproduction (Dole and Durant 1974). In the Variable Harlequin Toad of Costa Rica, Atelopus varius (Lichtenstein and Martens, 1856), both males and females are territorial and have fidelity for their reproduction sites. This indicates that even temporary alterations of the aggregation patterns of individuals between dry and rainy seasons, when they are more dispersed due to the increase in humidity (Crump 1988; Pounds and Crump 1989), results in an apparent decrease in detection (Gonzalez-Maya et al. 2013). In Panama, the Golden Toad, Ate/opus zeteki (Dunn, 1933) 1s concentrated in streams at the beginning of the breeding season, which occurs only during the transition between the rainy season and the dry season (Karraker et al. 2006). Another study determined site fidelity, habitat utilization, and range of households in the Suriname Toad, Atelopus hoogmoedi (Lescure, 1974) during the rainy season (Luger et al. 2009).
The Sierra Nevada of Santa Marta (SNSM) is a mountain massif located in the Caribbean region of Colombia. The particular conditions of isolation and vegetation of this mountainous system, which is not connected to the Andes, have led to a series of speciation processes in several groups of vertebrates, such as amphibians and reptiles (Ruthven 1922; Bernal- Carlo 1991; Sanchez-Pacheco et al. 2017), mammals (Alberico et al. 2000), and birds (Strewe and Navarro
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2004). This applies to the harlequin toads (Bufonidae: Atelopus Dumeéril and Bibron, 1841), whose current diversity includes five endemic species for this region: Atelopus arsyecue Rueda-Almonacid, 1994, Atelopus carrikeri Ruthven, 1916, Atelopus laetissimus Ruiz- Carranza, Ardila-Robayo, and Hernandez-Camacho, 1994, Atelopus nahumae Ruiz-Carranza, Ardila-Robayo, and Hernandez-Camacho, 1994, and Atelopus walkeri Rivero, 1963. These species can be found between 800 and 4,500 m asl, and from the tropical moist forests to the paramos of the SNSM (Ruthven 1916; Rueda-Almonacid 1994; Ruiz-Carranza et al. 1994; Rueda-Almonacid et al. 2005).
This study examines some aspects of the natural history and ecology of the Harlequin Toad, A. /aetissimus. Previous studies have shown that A. /aetissimus inhabits streams and rivers in the mountainous areas of the northwestern sector of the SNSM (Granda-Rodriguez et al. 2008, 2012; Rueda-Solano et al. 2016a), and it is classified as Endangered (EN) by the IUCN (Granda et al. 2008). A recent study reported individuals infected by the chytrid fungus Bd (Flechas et al. 2017), the pathogen that has led to the decline and disappearance of many amphibian populations globally (Young et al. 2001; Catenazzi 2015). This study also includes an estimation of relative abundance, population density, microhabitat preference, spatial dynamics, and vocalization, in a population of A. /aetissimus in the sector of San Lorenzo, Santa Marta, Colombia. Additionally, the potential distribution and patterns of habitat loss for this species during the 21" century were modeled, recognizing the great influence of habitat loss in recent decades on the extinction of many Neotropical species (Young et al. 2001; Marca et al. 2005; Lotters 2007).
Materials and Methods Study Area
The study area corresponds to the San Lorenzo creek (11°6’56.21” N, 74°3’0.18” W, 2,100 m asl), an affluent of the upper basin of Gaira river, northwestern sector of the SNSM, Santa Marta district, Magdalena department (= state), Colombian Caribbean. The principal vegetation unit of this area corresponds to lower mountain humid forest (sensu Espinal and Montenegro 1963). According to Granda-Rodriguez et al. (2012), the annual averages of rainfall and temperature are 2,622 mm and 13.6 °C, respectively, and the climatic regime is unimodal bi- seasonal, with a dry period from December to March, and a rainy period from April to November.
Estimations of Relative Abundance and Population Density
Relative abundance (RA) was estimated from data col- lected in seven field campaigns conducted from 2010 to
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2012 (October 2010; April, June, and December 2011; January and February 2012). Each fieldtrip had a duration of 12 days, with seven hours of daily work (0800-1200 h, 1800-2100 h). Individuals were detected through Visual Encounter Survey (Crump and Scott 1994), where two observers performed random walks. The sampling effort was 84 h per observer for each field campaign, reach- ing 558 h per observer in total. Relative abundance was calculated as the number of individuals/(h < observers), or ind/[h x obs] (Lips 1999). Sex was assigned by the size of individuals, assuming that females had a snout- vent length (SVL) > 40 mm and juveniles < 35 mm, and specimens within this range were considered as potential males. Sexual determination also considered the pres- ence of eggs in the corporeal cavity noted through skin, calling behavior, and amplectant couples. Sex was deter- mined in this way because the sexually dimorphic char- acters typically useful for population studies (La Marca et al. 1990 “1989;” Lampo et al. 2017) have not been established for A. /aetissimus.
Population density was estimated through 40 perma- nent transects of 20 x 4 m (Jaeger 1994), with 20 located in the riparian forest and 20 in the stream. These tran- sects were positioned parallel to the stream, separated by at least 20 m. Two observers walked along the tran- sect counting the individuals only once. The medians of density obtained in each riparian forest and stream were compared using the Wilcoxon test for independent sam-
ples (W). Habitat Selection and Dispersal Patterns
Testing the microhabitat preferences followed the con- cept of the third and fourth levels of habitat selection according to Johnson (1980), that indicate which com- ponents of the habitat are used and their proportions of use. For diurnal microhabitat, the substrate occupied by each individual was recorded according to the seven cat- egories proposed by Granda-Rodriguez et al. (2008b): (1) rocks, (II) leaf litter, (III) fallen trunks, (IV) ferns, (V) leaves, (VI) bare floor, and (VII) others. The proportion of area for each microhabitat category was measured in 15 random plots of 4 m? located at the side of the stream. A Chi-squared test (y) was performed, where the expect- ed frequency was the number of individuals by substrate (N), while the observed frequency was calculated as the total number of individual (N) per area proportion of the substrate (%). Microhabitat selection was assumed when the proportion of substrate used was different from its availability, following the method of Molina-Zuluaga and Gutiérrez-Cardenas (2007). To determine which substrates were selected by individuals, this analysis was repeated after deleting the categories most used or those that seemed to be used disproportionally to their avail- ability (Molina-Zuluaga and Gutiérrez-Cardenas 2007). In cases where significant differences in the second anal-
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ysis were not found, the deleted category was considered to be preferred by the species.
To determine the nocturnal site fidelity, 60 speci- mens were marked with sub-epidermal alphanumeric tags (Visible Implant Alpha Tags, Northwestern Marine Technology Inc., 1.5 x 2.5. mm), and detected at night with a fluorescence lantern (Courtois et al. 2013). Dur- ing 13 continuous nights (1900-2200 h, 36 h x obs), two observers looked for marked individuals to determine if they stayed in the same sites. The date, hour, location, and distance from the previous capture site were record- ed for each recapture. The potential relationship between the number of individuals recaptured and the number of nights of survey was explored using a linear regression.
To describe the patterns of diurnal horizontal move- ment, each marked and recaptured specimen was spa- tially located in a Cartesian diagram consisting of a 50 m transect along the stream delimited by a reference point every 5 m (y axis), and the perpendicular distance of the specimen to the transect (x axis). Then, the distance from diurnal to nocturnal microhabitat was measured when possible. This was carried out in two sampling sessions of three days, with a three-day interval between them. The sampling times on the first day were 0800-1100 h and 1400-1600 h, on the second at 0600-0900 h and 1300-— 1500 h, and on the third at 0900-1200 h and 1500-1700 h. This sequence was repeated successively. Aggressive behavior observed during the survey was described fol- lowing the terminology of Crump (1988).
Advertisement Call
On 30 January 2012, 270 seconds (s) of the advertisement call (sensu Wells 2007) of a male of A. /aetissimus was recorded in San Lorenzo creek, at 1732 h, using a Sony (ICD-PX312) digital recorder. Although the advertisement call is not easy to define, we consider that the recorded calls belong to this functional category because they were emit- ted regularly by a solitary male in situ (without manipula- tion), who had no interactions with individuals of the same sex (which might indicate aggressive calls) or the opposite sex (which might indicate courtship calls). Air tempera- ture and relative humidity at the recording moment were 12.3 °C and 75%, respectively. The traits of advertisement calls were quantified using the software PRAAT 6.0.13 for Windows (Broesma and Weenik 2007). The parameters of the advertisement call measured were: call duration (in seconds: s), number of pulses per call, pulse duration (s), interpulse interval (s), rate of pulses per second (pulses/s), frequency range (Hz), dominant frequency (Hz), and vis- ible harmonics (Hz). Means and standard deviations (SD) were calculated for each call parameter. The terminology proposed and revised by Cocroft et al. (1990) was used for call types and by Kohler et al. (2017) for call parameters. Spectrograms and oscillograms were generated using the Seewave package (Sueur et al. 2008) in R environment (R Core Team 2018).
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>
So ip
Relative abundance (ind/[h x obs])
0.0-
2010-VIl 2014-IIl 2041-VI 2011-XI 2011-XII 2012-1 2012-II
Oy Pe
4
0.00-
Forest Stream
Fig. 1. (A) Temporal variation of relative abundance (ind/[h x obs]). (B) Population density (m7) per habitat of Atelopus laetissimus.
Roman numerals represent the months of the surveys. Potential Distribution and Habitat Loss
A species distribution model was performed, which pre- dicts the habitat suitability with predictive algorithms integrated from environmental data and museum re- cords (Phillips et al. 2017). The available records of A. laetissimus were compiled from Global Biodiversity Information Facility (GBIF), and the authors’ own data, considering all known records from 1969 to 2017. Nine- teen bioclimatic layers from Worldclim 2.0. (Fick and Hijymans 2017) plus altitude, human footprint (Venter et al. 2016), and solar radiation were used as predictor variables. First, data were explored through a prelimi- nary model including all variables, with the technique of maximum entropy (MaxEnt software, 3.4.1, Phillips et al. 2017), considering that this algorithm is not hin- dered by a minimum number of occurrences. Variables with correlation indexes > 0.7 and lower contributions to the exploratory model were removed. To reduce the over- fitting of the model, collinearity was determined with a Spearman correlation test. According to van Proosdjj et al. (2016), the size of the background was considered in relation to the prevalence of the species to be modeled (< 25 localities), since this criterion generates acceptable results for species with restricted distributions.
A model of seven-fold bootstrap technique was per- formed, using 65% of data for training and 35% of data for testing (Puschendorf et al. 2008), considering the small number of locations that could be used (Elith et al. 2011). The average and standard deviation of the pre- dicted suitability were used as a final model, and as a spatially explicit measure of the reliability of the predic- tion, respectively. The accuracy of the model was esti- mated using the metric of the area under the curve of the receiver operating characteristic (AUC, Elith et al. 2011).
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Additionally, the distribution extent was calculated us- ing the IUCN methodology (2019), based on the area of occupation (AOO) and extent of occurrence (EOO). The AOO was calculated as the intersection of the species oc- currence with a square grid of 2 x 2 km, while EOO cor- responded to the minimum convex polygon drawn on the peripheral localities of the distribution area. Both AOO and EOO were calculated using only records after 2010. To determine habitat loss, the resulting map from the dis- tribution model was overlapped with forest cover loss maps from 2000 to 2017. These forest cover loss maps were gen- erated by Hansen et al. (2013), who monitored the changes of forest cover annually with a spatial resolution of 30 m. This product has shown important benefits in terms of its feasibility for evaluating the loss and fragmentation of habi- tat for forest specialist species (Alaniz et al. 2018; Carva- jal et al. 2018). The cumulative and annual habitat losses were calculated for the potential distribution, AOO, and EOO. For the annual habitat loss, fourth order polynomial regressions were performed to test the trends of the multian- nual habitat loss. The AOO and EOO estimations, and their respective trends, allowed a suggested threat classification based on criterion B of IUCN (2019) Red List of Species.
Results Relative Abundance and Population Density
A total of 447 individuals of A. /aetissimus were record- ed, distributed potentially among 428 males, 16 females, and three juveniles. The numbers of individuals per survey fluctuated from 32 to 90 (median + interquartile range, 78 + 44.5 individuals), with a general RA of 0.38 ind/(h x obs), and 0.21 to 0.54 ind/(h x obs) for each sur- vey (Fig. la). Atelopus laetissimus showed a population
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30-
ha o
Recaptures (n)
5 10 Nights of survey
Transect along the stream (m)
= o 1
= oO
e 5 tae 30- 4 Individuals #| A01 S| A3B6 4 sily a| Aa 0 — 20; [a | aaa
[a | ast
2 3 2 lel 5 0- 479 0.4 0.8 1.2 1.6 Distance to stream (m)
Fig. 2. Temporal variation of the number of recaptures (A) and movement patterns (B) of Ate/opus laetissimus.
density from 0 to 0.13 ind/m? (0.04 + 0.06 ind/m7). Popu- lation density was significantly higher (Wilcoxon test, W = 277.5, p = 0.036, Fig. 1b) in the stream (O—0.12 ind/ m’, 0.05 + 0.07 ind/m7) than in the riparian forest (O-0.11 ind/m?, 0.03 + 0.05 ind/m?). The corresponding author will provide tables of raw data for individual specimens on request.
Habitat Selection and Dispersal Patterns
Atelopus laetissimus used seven different diurnal mi- crohabitats, which were also used differentially regard- ing their availability (y?7 = 120.121, df = 6, p < 0.001). Although the leaf litter and rocks were the most used substrates (Table 1), significant differences in the use of microhabitat were still evident when these were removed from the analysis (7 = 471.991, df = 4, p < 0.001), sug- gesting that there is no preference for these substrates. From 60 marked specimens, three (5%) to 31 (52%) were recaptured per night of survey. The recapture rate was higher than 30% until the fifth night, while after the ninth night it was reduced to less than 7%, showing a lin- ear decrease across the sampling nights (7° = 0.86, F_,, = 69.99, p < 0.001, Fig. 2a). The height of the noctur- nal microhabitat ranged between 10 and 168 cm (mean + SD, 73.03 + 48.41 cm). The nocturnal site fidelity of A. laetissimus did not appear to be related to its height or to the SVL of specimens. Only six of the 60 tagged individuals were recaptured more than four times. These specimens showed an average home range of 0.35 + 0.21 m? (0.1-0.59 m’, Table 2, Fig. 2b), with an aver- age horizontal displacement of 1.92 + 0.82 m (0.8-3 m) relative to the nocturnal site. Most of the specimens were separated from each other by at least 5 m, but specimens Al and A4 were very close to each other, so aggressive behavior between them could be observed. Specimen
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A4 pounced and squashed specimen A1, then they be- gan actively “wrestling.” This situation lasted about 120 seconds, until Al fled. The “winner” male (A4) did not chase the male who left. The males did not emit vocaliza- tions during the event.
Advertisement Call
Two types of vocalizations were recorded in a male A. laetissimus. The first call type corresponded to a short series of pulses like a buzz (mean + SD, range, N; 27 + 5.63 pulses, 7-33 pulses, 26 calls, Fig. 3a), with a dura- tion of 0.41 + 0.112 s (0.111—0.805 s, 26 calls). These pulsed calls showed modulated amplitude, where the amplitude increased along the call and decreased again at the last pulse. The pulse duration was 0.009 + 0.006 s (0.001— 0.099 s, 705 pulses), emitted at a rate of 67.55 + 9.428 pulses/s (32.298—-76.167 pulses/s, 26 calls). In most of these calls, the last pulse had a longer duration. The interpulse interval duration was 0.006 + 0.004 s (0.0001—0.097 s, 652 interpulse intervals). These calls showed an ascending modulated frequency, although in some cases the frequency decreased notably at the last pulse. The frequency range was 1,287—8,558 Hz, while the dominant frequency was 1,921.433 + 114.391 Hz (1,480.95—2,155.55 Hz, 631 pulses). In addition, the pulsed call of A. /aetissimus showed three harmonics: first at 2,640.54-4,923 Hz, second at 3,661.56—6,644 Hz, and third at 5,771.95-8,558 Hz. The second type was a short call (Fig. 3b), with a duration of 0.06 + 0.006 s (0.05—0.07, 12 calls). The short calls showed a vari- able structure, either unpulsed (Fig. 3b), partially pulsed (Fig. 3c), or pulsed (Fig. 3d). The short calls showed a low dominant frequency (1,649.59 + 32.715, 1,584.2- 1,705.99, 12 calls) and were produced irregularly, from a variable series, and alternated among vocalizations of
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Table 1. Microhabitat selection by Ate/opus laetissimus and y? values for each substrate.
Substrate Proportion of area Individuals () ee Ia WRG REA x? Rocks 0.59 49 66.22 4.48 Leaf litter 0.28 28 31319 0.33 Others 0.03 2 355 0.59 Ferns 0.01 3 0.75 6.81 Leaves 0.05 9 5.65 1.99 Bare floor 0.01 12 13 104.56 Fallen trunks 0.04 ih 4.52 1.36 N 1 113 h3 120.12
the first type (Fig. 3e), or within vocalizations of the first type (Fig. 3f).
Potential Distribution and Habitat Loss
The areas with higher habitat suitability for A. /aetissi- mus are located mainly in the northern and northwestern sectors of the SNSM, which agrees with the occurrence localities. The model reached an AUC of 0.971 (+ 0.011, Fig. 4). The explanatory variables with the highest con- tributions to the suitability prediction were the average temperature of the coldest trimester (Bio 11), range of an- nual temperature (Bio 7), elevation, and human footprint. Atelopus laetissimus shows a maximum of suitability at 120 mm of rainfall at the coldest trimester; for annual temperature, it shows high suitability at middle ranges, rapidly decreasing at under 12 °C. The suitability shows a Gaussian trend regarding elevation, with a maximum at 2,000-—3,000 m asl. Habitat suitability of A. /aetissimus is inversely related with human footprint (Fig. 4).
The potential distribution area is 1,740.95 km’, which corresponds to a continuous area in the northwestern sec-
tor of the SNSM, and a smaller and fragmented area in the northern sector. The AOO calculated by 13 plots was 54 km’, while the EOO was 1,074.47 km?. The habitat loss from 2000 to 2017 was larger in the southern sector of the SNSM, but was smaller in the northern and northwest- ern sectors of the massif. Ate/opus laetissimus lost 1.48% of its habitat based on the potential distribution, 1.16% of its AOO, and 2.51% of its EOO (Fig. 5). A significant increasing trend in habitat loss was detected from 2013 to 2017, where the last year showed the greatest loss of potential habitat for the species. As A. /aetissimus shows an EOO smaller than 5,000 km? (criteria B1b[i], B1 b[ii], and B1bfiti]) and an AOO smaller than 500 km? (criteria B2b[i], B2b[11], and B2b/i11]), these data reinforce its clas- sification in the Endangered (EN) category.
Discussion Relative Abundance and Density
The results of these surveys showed that Ate/opus laetis- simus 1S an easily detectable species in the northwestern
Frequency (kHz) N T
Cc
Amplitude (dB)
Amplitude
0.0
0.06 0.08
Time (s)
Frequency (kHz)
Amplitude
Time (s)
Time (s)
Time (s)
Figure 3. Acoustic repertoire of the advertisement call of Ate/opus laetissimus. Conventional pulsed call (A), unpulsed short call (B), partially pulsed short call (C), pulsed short call (D), partially pulsed short call before pulsed call (E), and partially pulsed short call within pulsed call (F). The corresponding author will provide tables of raw data for individual specimens on request.
Amphib. Reptile Conserv.
34
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Granda-Rodriguez et al.
Table 2. Movement patterns of six Ate/opus laetissimus individuals.
Specimen Home range (m7?)
A4 Onn A44 0.47 A49 0.33 Al 0.54 A6l 0.1
A36 0.59 Mean 0.36 SE 0.21 Minimum 0.1
Maximum 0.59
sector of the SNSM. Several studies in this zone have reported more than 100 specimens in less than 150 h of survey effort (Granda-Rodriguez et al. 2012; Rocha-Usu- ga et al. 2017; Rueda-Solano et al. 2016a). Nevertheless, at less than 2,100 m of elevation, the relative abundances recorded for the species were significantly lower (Car-
| ces f ar
Occurrences
a Peet eh, att Piep den oT Ce eee a kt Fy :
Recaptures (7) Distance to nocturnal sites (m)
i ae 5 2 8 3 > 0.8 10 1.8 b) Pia 6.67 FOZ 207 0.82 5 0.8 10 3
vajalino-Fernandez et al. 2008, 2013; Granda-Rodriguez et al. 2012). In the model performed in this study, the most suitable habitat was in the altitudinal range be- tween 2,000 and 3,000 m. It is possible that changes in the physical and structural characteristics of the habitat at lower altitudes contribute to the decrease in either the
Habitat loss
© = After 2000
@ Before 2000 Suitability
al Deforestation 2000-2017 IUCN criteria (B)
MM 01-02 MM 02-03 |] 03-04 [ _]04-05 | |05-06
Colombia
i o9-1
Ps
Fig. 4. Habitat suitability estimate (upper left panel), minimum convex polygon of extent of occurrence (EEO, upper right panel),
and area of occupation (AOO, lower left panel) of Ate/opus /aetissimus. The total deforested area for the analyzed period and species
occurrence locations are provided in red.
Amphib. Reptile Conserv.
35 February 2020 | Volume 14 | Number 1 | e221
Natural history of Harlequin Toad, Atelopus /aetissimus in Colombia
on '
Habitat loss (Ha) @
2005 2010 2015
Fig. 5. Estimated annual habitat loss for Ate/opus laetissimus in the last decade in the potential distribution (A), area of occurrence (AOO, B), and extent of occurrence (EOO, C).
occupation or detection of A. /aetissimus.
Populations of A. /aetissimus appear to be highly dis- proportionate in males. The first publication that men- tioned this observation (Rocha-Usuga et al. 2017) did not describe how the males were differentiated from the fe- males. A sexual proportion that is biased to males can be related to differences in the mortality rate by sex. How- ever, it is also possible that some individuals categorized as males corresponded to small females, especially since the sexuality was assumed based on size (e.g., GOmez- Hoyos et al. 2017). This is an important detail that has been addressed only rarely in population studies of At- elopus (Gomez-Hoyos et al. 2014; Gonzalez-Maya et al. 2018), but it is important due to the conservation interest in the species of this genus.
Regarding population density, there is no previously published information for A. /aetissimus. Since this study provides the first estimation of this population attribute, it is not possible to estimate variations among different populations of this species. However, comparing the population density of A. /aetissimus obtained here with lowland species, the densities obtained with distance- based models for Atelopus spurrelli Boulenger, 1914 and Atelopus elegans (Boulenger, 1882) were slightly lower (0.03 and 0.01 ind/m’, respectively, GOmez-Hoyos et al. 2014, 2017). On the other hand, the observed density of Atelopus hoogmoedi (0.47 ind/m’, Luger et al. 2009) was higher than the mean density of A. /aetissimus found in this study.
Some species, such as Atelopus ignescens (Cornalia, 1849) and A. varius, had dense populations before severe population declines, with reports of 0.025—0.75 ind/m?
Amphib. Reptile Conserv.
and 0.065—0.755 ind/m?, respectively (Ron et al. 2003; La Marca et al. 2005). Atelopus cruciger 1s one of the few species with information on population density after a decline, which was 0.005—0.057 ind/m* (Lampo et al. 2012). Populations of A. cruciger had a high prevalence of chytrid fungus, but remained stable because of the high recruitment rate of healthy individuals in the popu- lation (Lampo et al. 2017).
Habitat Selection and Movement Patterns
The differential use of several substrates by A. /aetissi- mus has been previously reported, where leaf-litter and rocks were mainly used (Granda-Rodriguez et al. 2008b). The data reported here reinforce these findings, suggest- ing that this species selects the most available substrates. The structural complexity of the riparian forest occupied by A. /aetissimus can influence its differential pattern of habitat use, as has been described for some anurans from southeastern Asia (Gillespie et al. 2004). Habitat selection allows organisms to avoid adverse environmen- tal conditions, like extremely low temperatures (Navas 1996). Recently, A. /aetissimus has been described as a thermoconforming species, showing a direct relation- ship between the temperature of substrate and the activ- ity temperature (Rueda-Solano et al. 2016b). Therefore, the differential selection of substrates could be associ- ated with some thermoregulatory strategy. The results of these surveys show that A. /aetissimus exhibits relatively high nocturnal site fidelity, although the recapture rate was decreasing gradually, probably due to the manipula- tion of the specimens in each recapture. Recently, Rueda- Solano and Warketin (2016) reported that A. /aetissimus use the nocturnal sites for predatory activities, guided by the vibration of the substrate (leaves and ferns), suggest- ing that the use of a nocturnal perch is not exclusively for rest.
Regarding the home range of Afe/opus, some species such as A. carbonerensis and A. hoogmoedi possess a mean home range much larger than A. /aetissimus (41 m? and 38.1 + 17.7 m?, respectively; Dole and Durant 1974; Luger et al. 2009), which could be the result of seasonal variation. The results here indicate that individuals of A. laetissimus can remain, at least for a short period, near to the stream defending their territories.
Advertisement Call
The pulsed call is the most commonly known vocaliza- tion in harlequin toads, being present in at least 17 spe- cies (Asquith and Altig 1989; Cocroft et al. 1990; Ibafiez et al. 1995; Jaslow 1979; Lescure 1981; Lotters et al. 1999, 2002; this study). The pulsed call of A. /aetissimus consists of a short series of pulses (7-33 pulses) emitted rapidly, which is remarkably different from the pulsed calls of A. barbotini Lescure, 1981 (41-53 pulses per call, 30.35-—33.97 pulses/s, 2,000—3,000 Hz; Lescure 1981),
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A. flavescens Dumeril and Bibron, 1841 (45-58 pulses per call, 29.76—34.78 pulses/s, 2,500—3,000 Hz; Lescure 1981), A. franciscus Lescure, 1974 (31-39 pulses per call, 22.97—23.78 pulses/s, 2,300-3,000 Hz; Lescure 1981), A. hoogmoedi (40-42 pulses per call, 33.61—35 pulses/s, 2,300—3,000 Hz; Lescure 1981), A. spumarius Cope, 1871 (20-37 pulses per call, 38.55—45.96 pulses/s, 3,600—-4,400 Hz; Asquith and Altig 1987; Lescure 1981) or A. reticulatus Lotters, Haas, Schick, and Bohme, 2002 (27-32 pulses per call, 75—76 pulses/s, 3,282 Hz; Lotters et al. 2002) by having a higher number of pulses repli- cated more quickly at a lower dominant frequency. Like- wise, it differs from the pulsed call of A. zeteki Dunn, 1933 (42-52 pulses per call, 115-146 pulses/s, 1,381- 1,510 Hz; Cocroft et al. 1990), by having fewer pulses replicated more quickly at a higher dominant frequency.
Other species such as A. cruciger (84—99 pulses per call, 2,400—2,870 Hz; Cocroft et al. 1990), A. limosus Ibafiez, Jaramillo, and Solis, 1995 (31-45 pulses per call, 146.4-156.3 pulses/s, 2,600—2,800 Hz; Ibafiez et al. 1995), and A. varius (43-56 pulses per call, 119-123 pulses/s, 1,750—1,965 Hz; Cocroft et al. 1990) exhibit pulsed calls with higher numbers of pulses emitted at considerably faster rates than A. /aetissimus. On the oth- er hand, the structure of pulsed calls of A. /aetissimus 1s very similar to the calls of A. chiriquiensis Shreve, 1936 (18—33 pulses per call, 59.5—82.3 pulses/s, 2,000—2,700 Hz, Jaslow 1979), A. exiguus (Boettger, 1892) [19-21 pulses per call, 2,150—2,700 Hz, Coloma et al. 2000], A. minutulus Ruiz-Carranza, Hernandez-Camacho, and Ardila-Robayo, 1988, (14—21 pulses per call, 59.5—67.9 pulses/s, 2,700—3,150 Hz, Cocroft et al. 1990), A. nice- fori Rivero, 1963 (21-24 pulses per call, 53.9-65.7 pulses/s, 2,630—2,871 Hz, Cocroft et al. 1990), A. senex Taylor, 1952 (30-34 pulses per call, Cocroft et al. 1990), and A. tricolor Boulenger, 1902 (16-19 pulses per call, 2,970-3,450 Hz, Lotters et al. 1999). Pulsed calls of these species also consist of shorter calls emitted at faster rates, but with higher frequencies than A. /aetissimus in all cases.
The second type of vocalization (short calls) has been described for 12 species (Carvajalino-Fernandez et al. 2017; Ibafiez et al. 1995; Jaslow 1979; Lotters et al. 1999, 2002; this study). This call is the more variable of the two in terms of structure, by the definitions pro- posed by Cocroft et al. (1990), which includes several vocalizations emitted in different social contexts. Both pulsed and short calls of A. /aetissimus corresponded to the advertisement call context (sensu Wells 2007). This is probably the same situation for A. varius, whose short calls were obtained in the field, without apparent interac- tion among individuals (Cocroft et al. 1990). Short calls recorded in captivity for A. cruciger and A. spumarius also can be related to advertisement calls. This call is also reported in male-female interactions, in an amplectant couple of A. zeteki. Wells (2007) described this type of interaction as courtship calls. Encounter calls were re-
Amphib. Reptile Conserv.
corded in a male-male aggressive interaction in A. chiri- quiensis (Jaslow 1979). Nevertheless, most of the short calls described were release calls obtained at the moment of specimen manipulation (A. chiriquiensis, Jaslow 1979; A. limosus, Ibafiez et al. 1995; A. nahumae, Carvajalino- Fernandez et al. 2017; A. peruensis and_A. tricolor, Lot- ters et al. 1999). In addition, short calls of A. tricolor cannot be included in any of these categories, since the context of the recording was not described clearly (Lot- ters et al. 2002). Previously, the role of vocalizations in the communication of the genus Ate/opus has been dis- torted by the absence of several elements of the auditory apparatus (McDiarmid 1971) and the conspicuousness of its visual communication (Jaslow 1979; Crump 1988). Nevertheless, the complexity and diversity of vocaliza- tions described and reviewed here suggests that their roles in communication may be underestimated.
Potential Distribution and Habitat Loss
According to IUCN SSC Amphibian Specialist Group (2014), A. laetissimus is a species restricted to the moist low montane forest life zone, at altitudes between 1,500-— 2,880 and an area of 797 km?. In this study, localities are reported between 900 and 2,880 m asl and an EOO of 1,074 km/?, higher than that reported by IUCN. De- spite this increase in the distribution of the species which could be associated with new samplings, we recommend its status of Endangered (EN) be maintained. The niche model of A. /aetissimus suggests that the potential distri- bution is restricted to forests in humid zones at the north- western and northern flanks of the SNSM. The results show that the AOO of the species may not exceed 52 km”. The analysis by the model suggested the environmental layers that most influenced the distribution of the species are the average temperature of the coldest quarter, the elevation, and the human footprint. In the case of am- phibians, important influences of the climate on their dis- tribution have been described, therefore, factors such as climate change could significantly alter populations of A. laetissimus. Among the potential changes are alterations of the precipitation and temperature regimes, resulting in an increase in the annual temperature ranges and thus affecting habitat quality and the availability of specific resources for A. /aetissimus (Zhang and Yan 2014). On the other hand, human influence generates a very marked negative effect on this type of species in terms of degra- dation and loss of habitat (Grant et al. 2016).
The model performed here represents the first empiri- cal estimation of the distribution of this species based on distribution modeling, and it also uses the largest compi- lation of localities. Additionally, recent samples in areas with high suitability predicted by the model support the reliability of the prediction (Rueda and Warkentin 2016). In any case, the predictions of these models should be in- terpreted with caution and they should be considered as a first approximation to the real distribution of the species,
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helping to focus the sampling efforts in order to further adjust and refine the predictions of the distribution of this species in the future.
Although the habitat loss identified does not represent a high percentage in relation to the total habitat of the species, the trends showed a significant increase in loss in recent years (Ribeiro et al. 2018). However, considering that Hansen et al. (2013) does not differentiate between types of vegetation (natural and exotic plantations), the results could vary. This is important since A. /aetissimus has scarcely been associated with exotic plantations of Pinus spp., or in streams associated with this type of cov- erage in the locality of San Lorenzo (Granda-Rodriguez et al. 2012). In addition, it is also not known how coffee and avocado plantations can affect this species, since in the middle- and upper-part of the distribution large areas of these crops are present (Fundacion Pro-Sierra Nevada de Santa Marta 2000). It is necessary to carry out studies at smaller scales in the distribution area of A. /aetissimus, allowing the identification of the landscape dynamics of forest patch isolations, connectivity, and the different elements that may have negative consequences for this species (Palmeirim et al. 2018). In this sense, remaining remnants of forest become important for the maintenance of Atelopus laetissimus, as well as other endemic spe- cies of the SNSM. However, in the southern sector of the SNSM there is high fragmentation and habitat degrada- tion; and this site is (was?) inhabited by A. arsyecue and A. walker, species that have not been seen in the field for more than 20 years, providing possible evidence for the risk of disappearance of this type of toad.
Acknowledgements.—We are grateful to the people from Minca village and Unidad de Parques Naturales. We thank Vanesa Pacheco, Adolfo del Portillo, Miguel De Luque, and Liliana Saboya for their assistance during the field work. We thank Cristian Estades, Jaime Hernandez, and the anonymous reviewers for their valuable com- ments on the manuscript.
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Ecography 39: 542-552. Young BE, Lips KR, Reaser JK, Ibafiez R, Salas AW, Venter O, Sanderson EW, Magrach A, Allan JR, Beher Cedefio JR, Coloma LA, Ron S, La Marca E, Meyer J, Jones KR, Watson JEM. 2016. Sixteen years of JR, et al. 2001. Population declines and priorities for change in the global terrestrial human footprint and amphibian conservation in Latin America. Conserva- implications for biodiversity conservation. Nature tion Biology 15: 1,213—1,223. Communications 7: 12,558. Zhang X, Yan X. 2014. Spatiotemporal change in geo- Wells K. 2007. The Behavior and Ecology of Amphib- graphical distribution of global climate types in the ians. University Chicago Press, Chicago, Illinois, context of climate warming. Climate Dynamics 43: USA. 1,161 p. 595-605.
Hernan Granda Rodriguez is a Biologist from the University of Magdalena, Santa Marta, Colombia, who obtained his Master’s degree in Wild Areas and Nature Conservation from the University of Chile, Santiago, Chile. Hernan is currently a professor in the environmental engineering program at the University of Cundinamarca, Cundinamarca, Colombia. His research interests focus on the conservation of amphibians and reptiles, from the perspectives of both biological issues and social issues, such as the public's perception of herpetofauna.
. Andrés Camilo Montes-Correa is a Colombian young biologist and herpetologist (in training) at the Uni- versidad del Magdalena (Santa Marta, Colombia). His research interests include the taxonomy, systematics, and ecology of the Colombian herpetofauna, especially lizards and freshwater turtles. Andrés has conducted investigations on habitat use by freshwater turtles of the Colombian Llanos (for his B.Sc. thesis); ecology, taxonomy, and distribution of small geckos and cryptic dendrobatid frogs; and synecological studies of herpetofauna. His current project is on the taxonomy, ecology, distribution, and conservation of Lepido- blepharis miyatai, a small and endangered gecko endemic to the coastal mountains (Sierra Nevada de Santa “ Marta) of the Colombian Caribbean.
Juan David Jiménez-Bolaiio is a wildlife photographer and has a B.Sc. in Biology from the University of Magdalena (Santa Marta, Magdalena, Colombia). Juan’s work has focused mainly on the ecological and diversity patterns of Neotropical herpetofauna. Currently, Juan is interested in studying diversity patterns in high Andean environments.
Alberto J. Alaniz, Geographer, B.Sc., M.Sc. His work is focused on ecological modelling applied to biological conservation and epidemiology, and his recent research has been on the development of spatially explicit infection risk models for Zika virus at global and local scales. Currently, Alberto is a researcher in the Ecology and Conservation Lab in the Technological Faculty at the University of Chile, and he is a founding member of the Center for Spatial Ecology and Environment (Ecogeografia), located in the city of . Santiago, Chile.
Patricio Hernaez is a Marine Biologist by training at the Universidad Arturo Prat (Chile), with a Master’s degree in Biology from the Universidad de Costa Rica (Costa Rica), and a Ph.D. in Biological Sciences from the Universidade de Sao Paulo (Brazil). His research focus is divided into three main lines: (1) Taxonomy of recent groups with an emphasis on Crustacea, (11) Population dynamics, and (ili) Fisheries. Patricio is currently a researcher associated with the Centro de Investigaciones Marinas y Limnologicas of the Universidad de Tarapaca, Chile, and the Grupo de Pesquisa em Biologia de Crustaceos of the Universidade Estadual Paulista ‘Julio de Mesquita,’ Brazil.
Pedro E. Cattan has a D.V.M. and a Ph.D. in Biological Sciences from the University of Chile, and he was the Chairman of the Department of Biological Sciences of the University of Chile for 10 years. Pedro has developed two main lines of research, one on the study of populations in wild vertebrates and the other on disease ecology at the wildlife level. He currently directs projects on the vectors and wild reservoirs of Chagas disease.
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Official journal website: amphibian-reptile-conservation.org
Amphibian & Reptile Conservation 14(1) [General Section]: 43-54 (e222).
Do growth rate and survival differ between undisturbed and disturbed environments for Sceloporus spinosus Wiegmann, 1828 (Squamata: Phrynosomatidae) from Oaxaca, Mexico?
‘Carlos A. Torres Barragan, ?Uriel Hernandez Salinas, and **Aurelio Ramirez-Bautista
‘Instituto Politécnico Nacional, Centro Interdisciplinario de Investigacion para el Desarrollo Integral Regional (CIIDIR) Unidad Oaxaca, Hornos No. 1003, Col. Noche Buena, Santa Cruz Xoxocotlan, Oaxaca, Oaxaca 71230, MEXICO ?Instituto Politécnico Nacional, Centro Interdisciplinario de Investigacion para el Desarrollo Integral Regional (CIIDIR) Unidad Durango, Calle Sigma 119 Fraccionamiento 20 de Noviembre II, Durango, Durango 34220, MEXICO ?Laboratorio de Ecologia de Poblaciones, Centro de Investigaciones Bioldgicas, Instituto de Ciencias Bdsicas e Ingenieria, Universidad Autonoma del Estado de Hidalgo, Km 4.5 carretera Pachuca-Tulancingo, 42184, Mineral de La Reforma, Hidalgo, MEXICO
Abstract.—Demography is intimately related to the evolution of the life history of a species, since it describes the patterns of variation in the growth, maturation, reproduction, and survival of an organism through populations, species, and environments. In this study the growth, survivorship, and population structure were evaluated for an oviparous lizard, Sceloporus spinosus from two sites, a relatively undisturbed area (UA) and a disturbed area (DA; zone of land-use change) within the Natural Protected Area Yagul of southern Oaxaca, Mexico. The results showed different relative densities between seasons (higher during the wet season than the dry season), but not between populations. Males and females from the UA and DA showed similar growth rate patterns, and both sexes reached sexual maturity at a similar body size. The highest survival rates and recapture probabilities were found in the UA; however, males from both populations showed higher survival rates than females. Overall, this study suggests that land-use changes do not seem to cause wide variation in the analyzed demographic characteristics of this species. This work describes and quantifies demographic effects on some life history characteristics of aspecies endemic to Mexico. We argue for the need to analyze and compare many capture-recapture data for a species between locations in order to obtain a better assessment of the variation in the life history characteristics analyzed.
Keywords. Age class, Cormack-Jolly-Seber model, demography, density, natural protected area, toe-clipping method
Citation: Torres Barragan CA, Hernandez Salinas U, Ramirez-Bautista A. 2020. Do growth rate and survival differ between undisturbed and disturbed environments for Sceloporus spinosus Wiegmann, 1828 (Squamata: Phrynosomatidae) from Oaxaca, Mexico? Amphibian & Reptile Conservation 14(1) [General Section]: 43-54 (e222).
Copyright: © 2020 Torres Barragan et al. This is an open access article distributed under the terms of the Creative Commons Attribution License [At- tribution 4.0 International (CC BY 4.0): https://creativecommons.org/licenses/by/4.0/], which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. The official and authorized publication credit sources, which will be duly enforced, are as follows: official journal title Amphibian & Reptile Conservation; official journal website: amphibian-reptile-conservation.org.
Received: 7 August 2018; Accepted: 21 October 2019; Published: 18 February 2020
Introduction characteristics of the environments that they inhabit (e.g., pristine or fragmented). Therefore, the conservation
The variation in life history characteristics of lizards is _ priority of populations of a widely distributed species, is
considered to be an outcome of phenotypic plasticity driven by changing environmental conditions (Stearns 1992; Adolph and Porter 1996). Studies investigating demographic parameters (e.g., density, sex ratio, natality, growth rate, age classes) and life history (e.g., SVL at sexual maturity, survival, reproduction, fecundity) in lizards have shown that different life strategies (e.g., growth rate, survival) in these vertebrates have evolved due to environmental changes caused by habitat loss and by changes in land use (Dunham 1982; Stearns 1992). This implies that the populations of any species may evolve different life history strategies according to the
to assess the tolerance of their life history characteristics to certain environmental factors (precipitation, humidity, radiation, pollution, deforestation, and others) that occur throughout the distribution of the species; therefore, this is a feasible method to test for changes in their fitness (Walkup et al. 2017). Evolution within and across species that inhabit fluctuating environments has resulted in changes of their life history strategies, such as size and age at sexual maturity, fecundity (clutch size), growth rate, and survival (Stearns 1992); and these changes have been found within different populations of a single species that is widely distributed (Dunham 1982; Cruz
Correspondence. ! augusto.torres007@gmail.com, ? uhernndez3@gmail.com, **ramibautistaa@gmail.com
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Sceloporus spinosus in Oaxaca, Mexico
et al. 2014; Pérez-Mendoza et al. 2014; Cruz-Elizalde and Ramirez-Bautista 2016). These variations in life history characteristics have been documented in several species that inhabit environments with different degrees of disturbance (Cruz et al. 2014; Cruz-Elizalde and Ramirez-Bautista 2016; Walkup et al. 2017). However, strong fragmentation of habitats by land use change, pollution, and global warming have adverse effects on these strategies, that consequently lead to population decline at the local level (Sinervo et al. 2010).
Due to concerns regarding the effects of environmental disturbances, some _ researchers have generated conservation models for various biological groups (e.g., birds, Escalante et al. 1998; mammals, Ceballos and Oliva 2005) that include significant amounts of information on the natural history of individual species. For example, Sinervo et al. (2010) described patterns of species decline and extinction in a diverse assemblage of lizard species of genus Sce/oporus under thermoregulatory stress induced by global warming. Chavez (2011) and Calder6n-Mandujano (2011) noted that land use change is another factor that has resulted in high levels of population decimation and extinction among lizards and amphibians. However, each species responds in different ways according to the pressures of their local environment (Tews et al. 2004; Suazo-Ortufio et al. 2007). This pattern has been documented in several wide-ranging species that occur in relatively pristine habitats as well as sites that are subject to varying degrees of disturbance (e.g., Sceloporus grammicus, Pérez-Mendoza et al. 2014; S. minor, Garcia-Rosales et al. 2017; and S. variabilis, Cruz-Elizalde and Ramirez- Bautista 2016). Therefore, considering these factors, herein, the effects of a pristine and a disturbed habitat on some life history characteristics were evaluated in two populations of Sce/oporus spinosus (Eastern Spiny Lizard) in southeastern Mexico.
Sceloporus spinosus 1s a species endemic to Mexico, and adults are of medium body size for the genus (120 mm snout-vent length, SVL; Ramirez-Bautista et al. 2014). The scales of the body are strongly keeled and mucronate. This species feeds on insects and other invertebrates, and it 1s oviparous with a clutch size of eight to 31 eggs and a mean of 18.5 (Valdéz-Gonzalez and Ramirez-Bautista 2002). This lizard is found from Durango to Oaxaca, and inhabits arboreal and saxicolous landscapes (Torres-Barragan 2015) in both temperate and semiarid regions, at an elevation range from 1,900 to 2,700 m (Canseco-Marquez and Gutiérrez-Mayén 2010). In Yagul Natural Protected Area (NPA), this species is distributed in a mountain range with elevations from 1,600 to 2,000 m. So far, there is limited information regarding demographic aspects of this species in pristine and disturbed areas such as Yagul NPA. In this framework, the goal of this study was to compare and assess key demographic characteristics, such as density, growth, survival, and population structure of S. spinosus in two
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contrasting environments, in Oaxaca, Mexico. Therefore, considering that land use change promotes variation in demographic characteristics and life histories in various vertebrate groups (Adolph and Porter 1996; Flatt and Heyland 2011), these demographic characteristics were expected to differ as a function of the environments where each population of Sceloporus spinosus occurred.
Materials and Methods Study Area
This study was carried out at two sites in Yagul Natural Protected Area in the municipality of Tlacolula de Matamoros, Oaxaca, Mexico (Fig. 1). The municipality encompasses 1,076 ha, ranging in elevation from 600 to 2,500 m. The vegetation at the site is represented by tropical dry forest; however, much of it has been replaced by crops and grazing areas. The climate is semi-warm, with temperatures ranging from 16 °C to 26 °C, and mean annual precipitation from 400 to 800 mm (INEGI 2005). Two sites of 1 ha each were chosen for this study. The first site was considered the undisturbed area (UA; 16.957922 N, -96.429953 W; 1,800 m), with a vegetation cover of 83% and an arboreal density of 697 individual trees/ha. This cover includes 80% tropical dry forest, 10% flood zone, 3% reedbed (Arundo donax), and 2% surface without vegetation (Torres-Barragan 2015). The second site was a disturbed area (DA; 16.959617 N, -96.450633 W; 1,652 m; Fig. 1). This site is an open area with agave plant cultivation and extensive grazing areas; canopy cover 1s 1% with an arboreal density of eight individual trees/ha; 50% of the land is used for cultivation of agave plants (Agave angustifolia), 20% for induced pasture, 10% for living fences (Prosopis, Yucca, Celtis, Acacia, Opuntia, and Schinus), and 20% of the surface has no vegetation (exposed floor; Torres-Barragan 2015).
Data Collection
Twelve sampling events were conducted at each site (UA and DA) from January 2014 to January 2015. Each sampling event was carried out over a single three-day period in each month at each of the two sites (for a total of six sampling days per month), with a sampling effort of three people from 0900-1800 h.
The method of mark-recapture of Lemos-Espinal and Ballinger (1995) and Ramirez-Bautista (1995) was used in this study. This method consisted of ectomization of phalanges (e.g., toe-clipping), a permanent marking technique that makes it possible to recognize every previously marked individual during each subsequent sampling event. Toe-clipping is commonly used to follow cohorts of lizard populations (Dunham 1978; Tinkle 1961, 1969). More recently, Guimaraes et al. (2014) and Olivera-Tlahuel et al. (2017) expressed some concern when using this method due to observed effects on the
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1877000
Tlacolula de Matamoros
1875000 1876000
1874000
) Salado River 770000
771000
772000
SYMBOLOGY
MUA [DA
—— Watercourses
[]} Polygon MNY
1876000
Level curve
1875000
~—-—= Road
LC] Urban area
ee Archaeological Area Yagul
1874000
773000
Scale 1:27804
Fig. 1. Map of the study area. The green polygon depicts Yagul Natural Protected Area, including the two sampling sites (UA =
undisturbed area; DA = disturbed area, land use change).
behavior and health of some lizard species, and therefore, on survival. Although toe-clipping could affect survival, lizards at both sites were toe-clipped similarly, so any negative bias in survival estimates should apply equally to both sites. The SVL of each lizard was measured with a digital caliper (to the nearest 0.01 mm), and body mass with a balance (+ 0.01 g).
Relative Density and Population Structure
Based on the number of captured and recaptured individuals from both populations during the study, the relative density of each population was determined using the equation, N = M/R, where N = number of unknown individuals in the population; M = the number of marked individuals; and R = the number of recaptured individuals/surface area. To determine the population structure for each site, size classes (SVL) were determined based on those used by Leyte- Manrique et al. (2017) with Sceloporus grammicus. These authors related the SVL of each age category based on anatomical traits, yielding classification categories of: offspring (SVL < 48 mm), juveniles (49-69 mm), and adults (females and males > 70 mm). Females were considered to be adults if they contained eggs in the oviduct, which were identified by palpation of the ventral region (Galan 1997). Whereas males were considered to be adults when they showed the bulky tail base indicative of sperm production (Lozano et al. 2014). Relative densities of lizards were compared
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between locations and seasons by means of a Student’s t-test (Zar 2014).
Growth Rate
Growth rates were assessed for males and females from each population by considering only those lizards with recapture intervals greater than 30 and less than 100 days. Therefore, growth rate was estimated with the formula: GR = (SVL, — SVL,)/days, where growth rate (GR) is the difference in recorded SVL between the last recapture (SVL,) and first capture (SVL,) divided by the number of days that had elapsed (Dunham 1978; Zamora-A brego et al. 2012). Then, nonlinear regression models of Von Bertalanffy, logistic by body size (SVL), and logistic by body mass were used (Dunham 1978); and growth rates for both sexes and populations were compared. The first model (Von Bertalanffy) describes a pattern in which smaller individuals (in SVL) show faster growth rates than larger ones (Dunham 1978; Zamora-Abrego et al. 2012). In contrast, the logistic models predict that individuals smaller in SVL will grow moderately faster to reach intermediate sizes, and after reaching their maximum growth rate, that rate will decrease in a non-linear direction as size increases (Dunham 1978; Schoener and Schoener 1978; Zamora-Abrego et al. 2012). The difference between the two models 1s that the maximum growth rate attained under a logistic by length (SVL) model is observed at early ages, while maximum growth rate under a logistic by body mass model will
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be observed at later ages (Dunham 1978). Detailed descriptions of each model can be reviewed in Dunham (1978) and Schoener and Schoener (1978). Selection of the best model was based on the best fit to the observed growth rates for both sexes, chosen by the lowest value of the residual mean square (RMS) and highest values of coefficients of determination or correlation (R*?; Dunham 1978; Schoener and Schoener 1978).
Once selected, the models were developed with confidence intervals following Schoener and Schoener (1978) for the growth parameter (r), and the asymptotic (A,) was calculated by the formula:
@,— JRF, ani S} = Bj = 9, + JRF, en S}
where B, is the adjusted parameter j, 0, is the parameter estimated B., S, 1s the asymptotic standard deviation of bees _ {9 is the value F _, of a tail with & and N —k degrees of freedom, N is sample size, and k 1s the number of adjusted parameters. These confidence intervals provide maximum reliability for each parameter (r and A,) and are considered significantly different between sexes if they do not overlap (Schoener and Schoener 1978). On the other hand, the residuals of the growth rate (removal of effect size) were used to determine by two- way ANOVA if there were differences in the patterns of growth rates between factors (sexes and populations). The residuals are the result of the relationship between the SVL and the growth rate under the model with the best fit (Schoener and Schoener 1978). Finally, based on the values of the growth parameter (r) and asymptotic (A,) obtained from the best-fit model, together with the average values of SVL of offsprings at hatching (LO = 42 mm in SVL), the ages in days were determined for both males and females at which they reach sexual maturity (Dunham 1978; Schoener and Schoener 1978; Zamora- Abrego et al. 2012). The growth models for both sexes and populations were developed with Statistica program, version 7.0.
Estimates of the Survival Models
Captures and recaptures of marked individuals allowed estimates of demographic parameters, such as survival (@) and recapture (p); and both parameters were estimated from different models that represent distinct biological hypotheses of survival (Lebreton et al. 1992). Both and p can be constant (c) over time or vary as a function of time (¢) and between sex, and for their assessment a general model was considered which allowed the determination of whether survival rates and recaptures were different between sexes and populations. This model is: @ (groups [males and females in two populations = four groups]*time) p (groups*time), and it calculates the probability that survival and rate of recapture are different between groups over different periods of time.
Amphib. Reptile Conserv.
For developing the different models based on encounter histories of each individual (e.g., 100101), zero represents sampling when a lizard was not seen in the area, while 1 represents those sampling events when lizards were marked and recaptured (Lebreton et al. 1992). Encounter histories were analyzed and modeled with software Mark 6.0 (White and Burnham 1999) using the subprogram “only recaptures” with the goal of obtaining estimates of survival and recapture rates grounded in the techniques of maximum likelihood under the model developed by Cormack-Jolly-Seber (Lebreton et al. 1992). The model that showed the best fit to the capture-recapture data was the one with the lower Akaike information criterion (AIC) value; however, when there was a difference of 2 between A/C values of the two models, both models were assumed to have approximately the same fit to the data (Burnham and Anderson 2002). Survival and recapture results are represented with confidence intervals of 95%.
Results Relative Density and Population Structure
In the surveys, 271 individuals were marked across both sites (UA = 149 and DA = 122): of these, 113 were recaptured (73 in UA and 40 in DA; Table 1). In some cases, several individuals were captured as juveniles and then recaptured as adults; but most of the recaptures were adult males and females (Table 1). The relative density between seasons was different (oN = -2.023, P = 0.05; wet: 6.95 + 0.41 [4.92-10.07]; dry: 9.70 + 1.29 [2.68— 17.21]), but not between populations (¢,,, = -0.116, P = 0.98; UA: 8.33 + 0.97 [5.68-16.11]; DA: 8.33 + 1.11 [3.28-17.21]). The relative density for UA (both sexes and all age classes) was 149 individuals/ha, whereas for DA it was 122 individuals/ha. In UA, offspring emerged from July to December, but the peak hatching period was in September; juveniles were recorded from October to April, but the highest population of this age class was in December; adults were present throughout the year, but the highest numbers of captures were in April and May (Table 1). Offspring from DA were found from August to November, with peak density in August; juveniles appeared from October to March, with density peaking in December; and adults were seen from September to July, with the greatest densities from April to May (Table 1).
Growth Rate
The length logistic model showed the best fit to the growth rate data for males and females from UA; in contrast, the Von Bertalanffy model showed the best fit to the growth rate data for both males and females of DA (Fig. 2 and Table 2). The logistic model by length showed that growth rates for males (r + EE: 0.007 + 0.0005) and females (r + EE: 0.008 + 0.0008) from UA were similar, whereas the asymptotic growth curve of females (A, =
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Table 1. Numbers of Sce/oporus spinosus individuals in each age class during each study month at Yagul Natural Protected Area. UA = Undisturbed area, DA = Disturbed area. * Indicates the rainy months. A = adult, J = juvenile.
UA Offspring Juveniles Adults
Samples Males Females Males Females Males Females Recaptures January 0 0 1 2 1 0 0 February 0 0) is) 4 2 1 19A March 0 0 2 4 7 2 3GA, 19, 295 April 0 0) 3 3 9 9 ASA, 52,28) May 0) 0 0) 0) 7 10 AGA, SQA June* 0) 0) 0) 0) 2 6 IGA, SOA July* 0) 1 0) 0 7 4 SSA, 29A August* 3 1 0 0 2 3 29,14) September* i) 2 0) 0 5 1 30'A, 129A, 23) October a 1 0) 1 4 3 3GA, 29A, 28, 19) November 4 0) 3 2 6 0) SSA, 28, 19) December 1 0) 7 2 2 0) SSA, 3Q9A Density (#/area) 0.0019 0.0037 0.0093
Total 14 5 19 18 54 39 73
DA Offspring Juveniles Adults
Samples Males Females Males Females Males Females
January 0 l 2 2 2 2 0 February 0) 0 1 0 2 1 0 March 0 0 2 2 5 3 IGA April 0) 0) 0) 0 10 6 6GA, 32A May 0) 0 0) 0 11 10 AGA, SQA June* 0) 0) 0) 0 1 5 29A,1dA July* 0) 0 0) 0) 4 5 SSA, 29A August* 3 4 0) 0) 0 0 0 September* 1 2 0 0 1 3 0 October 1 0 2 4 0) 2 19J,29A November 1 0 4 2 0 2 19A, 3d) December 0) 0 2) 6 1 1 IGA, 19, 295 Density (#/area) 0.0013 0.0032 0.0077
Total 6 yi 16 16 37 40 40
EE: 105.359 + 3.225 mm) was slightly higher than that of males (A, + EE: 101.706 + 1.587 mm; Table 2). On the other hand, for males and females from DA, the Von Bertalanffy model showed that male (r + EE: 0.005 + 0.0008) and female (r+ EE: 0.005 + 0.0011) growth rates were similar (Table 2); however, the females reached an asymptotic size (maximum size) that was slightly larger (A, + EE: 108.058 + 4.139 mm) than the males (A, = EE: 103.253 + 3.723 mm; Table 2). Average values of the residuals of growth rates for males and females from UA, obtained with both the logistic by length model and a two-way ANOVA, did not show significant differences (males: -0.001 + 0.009; females: 0.003 + 0.019) between dry (F’,, = 0.047, P = 0.8300) and wet seasons (0.001 +
1,29
Amphib. Reptile Conserv.
0.010, -0.004 + 0.011, respectively), between sexes (F, ,, = 0.003, P = 0.9577), or interactions between factors (season*sex; F, ,, = 0.116, P = 0.7358). A similar pattern occurred for DA, where no differences were found in males (-0.003 + 0.013) and females (-0.007 + 0.015) between dry and wet seasons (Pag = 0.975, P = 0.3333; males = 0.021 + 0.009 and females = 0.015 + 0.033), between sexes (F, ,, = 0.044, P = 0.8358), or interactions between factors (season*sex; Ff’ ,, = 0.004, P = 0.9472).
On the other hand, there were no differences in the overall growth rates of males from the two populations (fF ,, = 0.234, P = 0.6317), in neither dry (UA: -0.001+ 0.009; DA: -0.003+0.013) nor wet (UA: 0.001 + 0.010; DA: 0.021 + 0.009) seasons (F, , = 0.535, P = 0.4699);
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Sceloporus spinosus in Oaxaca, Mexico
Growth rate (day/mm)
SVL (mm)
= ho wn
_ ho
Growth rate (day/mm) Growth rate (day/mm)
bd oS nm
40 50 60 70 380 90 100 110 50 60 70 80 90 100 110 SVL (mm) SVL (mm) Fig. 2. Growth rate of Sce/oporus spinosus. (A) Undisturbed area (UA) males, (B) Disturbed area (DA) males, (C) UA females, and
(D) DA females. Black circles represent data points for individual lizards. Modeled relationships between growth and body sizes of males and females: solid lines = Von Bertalanffy, dashed lines = logistic by length, and dotted lines = logistic by mass.
— = 240 alice © 200 - ri] -| = 160 - : 120 o | = 80 | oO | om 40 £ = 0. x DA UA € 0.0120 0.0120 - O — 0.0100 J 0.0100 - i % M—_ oD — o rT 0.0080 0.0080 - =o 2 = 0.0060 0.0060 - a o © 0.0040 0.0040 - © Oo Pa g 0.0020 0.0020 - 0 0.0000 0.0000 = DA UA DA UA
Fig. 3. Means and 95% confidence intervals of the Asymptotic growth (A,) and Characteristic growth (r) parameters obtained by the Von Bertalanffy and logistic by length models for males and females of Sce/oporus spinosus in both Disturbed area (DA) and Undisturbed area (UA) populations.
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therefore, the interaction term (locality*season) was not significant (F' ,, = 0.401, P = 0.5312). The same pattern was found in females, as there were no differences between localities (F’ tog OOS 0.8130), or seasons (F fap ee Ee 0.7284); and, therefore, the interaction term (locality*season) was also not significant (F,,, = 0.587, P=0.4519).
Growth parameters (r and A,) were similar for males and females in UA and DA populations (Fig. 3); confirming that there is no difference between the sexes. In UA, the growth rate showed that males reached sexual maturity at SVL 85 mm at age 210 d (7 months), whereas females attained sexual maturity at SVL 89 mm at age 280 d (9 months). In DA, males reached sexual maturity at SVL 81 mm at age 210 d, and females at SVL 85 mm at age 280 d.
Survival Model Estimation
To analyze survival rate (@) and recapture (p) of S. spinosus in UA and DA populations, a set of models was developed (Table 3). The single model that described survival rate as varying between groups (sexes) and where the recapture rate was constant [® (sex) p(c)] was chosen as the best fit for both populations (Table 3A). Based on this model, the survival rate for males from
UA (0.82) was higher than that of females (0.70), while the recapture rate was similar for both sexes (0.40; Table 4A). In addition, the survival rate for males from DA (0.75) was higher than that of females (0.65), and the probability of recapture was higher for males (0.40) than for females (0.35; Table 4A). These values are lower than those found for UA, which suggests a higher probability of survival and recapture in UA than DA. On the other hand, survival and recapture rates by season (wet and dry) in UA and DA populations showed that the model with the best fit was ® (c) p(season), indicating that survival rate is constant, and the probability of recapture varies between seasons (Table 3B). According to this model, the survival rates in both populations were higher in the dry (UA: 0.76, DA: 0.92) than the wet (UA: 0.54, DA: 0.49; Table 4B) season.
Discussion
More lizards were marked at the UA site than the DA site during this study. However, at both UA and DA the greatest numbers of adult recaptures were in April and May, due to the peak in reproductive activity during these months (Valdéz-Gonzalez and Ramirez-Bautista 2002). According to the recapture data, lizards born in August-September reached the minimum SVL at sexual
Table 2. Growth parameters for Sce/oporus spinosus males and females from UA and DA populations obtained from each growth model. RMS = residual mean square, R* = coefficient of determination, A, = asymptotic of growth, r = parameter of growth, + =
standard error. UA Model RMS Males (n = 20)
Von Bertalanffy 0.029
Logistic by length 0.019
Logistic by weight 0.025 Females (n = 13)
Von Bertalanffy 0.029
Logistic by length 0.019
Logistic by weight 0.023 DA Model RMS Males (” = 15)
Von Bertalanffy 0.027
Logistic by length 0.035
Logistic by weight 0.048 Females (n = 13)
Von Bertalanffy 0.030
Logistic by length 0.032
Logistic by weight 0.429
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49
R? A, r 0.730 109.480 + 5.223 0.003 + 0.0003 0.849 101.706 + 1.587 0.007 + 0.0005 0.797 100.081 + 1.114 0.012 + 0.0009 0.595 122.243 + 12.423 0.003 + 0.0007 0.736 105.359 + 3.225 0.008 + 0.0008 0.687 102 482° 2°29 1 0.012 +0.0011
R? A, r 0.809 103.253 + 3.723 0.005 + 0.0008 0.756 99.762 + 2.410 0.0099 + 0.0010 0.664 98.523 + 1.995 0.0141 + 0.0016 0.625 108.058 + 4.139 0.005 + 0.0011 0.601 100.178 + 3.225 0.008 + 0.0012 0.046 99 340 + 3.7378 0.012 + 0.0017
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Sceloporus spinosus in Oaxaca, Mexico
Table 3. Models describing survival rate (@) and recapture (p) of Sceloporus spinosus males and females in UA and DA with the Jolly-Saber model using the Mark program. The models are fitted with the Mark program considering that ® y p (probability of survival and recapture) can be either constant (c) or varying between sex (s), season (dry and wet), and population. AIC = Measurement of the level of adjustment and parsimony of each model, A = difference of AIC, W, = weight of AIC, K = number of parameters. A. Models for estimation of sex and populations.
Model AIC A, W, K Dee) 231.98 0 0.69 12 ® (c) p(c) 234.09 2.11 0.24 13 ® (population) p(c) 238.22 “625. =:0,03" 412: ® (population) p(sex) 240.54 857 0.01 13 ® (sex) p(c) 241.68 9.71 0.01 3 ® (c) p(sex) 242.11 10.13 0 3 ® (population) p(population) 243.39 11.42 0 21 ® (sex) p(sex) 243.84 11.86 0 4
B. Models for estimation of season (dry and wet) and populations.
Model AIC A. Ww. K
Ui I
® (c) p(season) 224,02 0 0.88 3
® (season) p(c) 231.98 4.96 0.07 12
® (season) p(population) 237.17 JOA6. .0,01 3
® (c) p(c) 240.11 13.1 0 2 O (ft) p(t) 243.39 16.38 0 21 ® (population) p(season) 247.87 20.86 0 2
maturity (> 80 mm) in April-May of the following year, a similar pattern seen in another population of the same species (Valdéz-Gonzalez and Ramirez-Bautista 2002). Recruitment of offspring, together with adult males and females, and a few subadults, results in population growth with respect to these age structures. These events are synchronized with an increase in ambient temperatures (from 20.9 °C in April to 21.2 °C in May) and precipitation (from 113.3 mm in June to 114.4 mm in May) in the region, which also coincide with high production of food in the environment (Dunham 1982; Ramirez-Bautista and Vitt 1997).
In general, these results revealed that lizard density was significantly similar between populations, but not between seasons (higher in the wet season than in the dry season). The higher density of lizards found in the wet season could be explained by a high supply of the food consumed by this species. However, this assumption brings up additional questions regarding the feeding habits of other species living in sympatry with S. spinosus in disturbed and undisturbed environments. Therefore, it is necessary to investigate whether those sympatric species have the same possibilities of acquiring available resources (food and microhabitat), or if they
Amphib. Reptile Conserv.
display different activity schedules that allow them to obtain resources more efficiently. Such studies would certainly expand our knowledge on the natural history of this species, and consequently enable the development of more effective conservation strategies.
The growth rate models used here showed that males and females in both populations grow at the same rate and reach maximum SVL at similar sizes. These results were found by the logistic model by length (UA) and Von Bertalanffy (DA), models that are known to fit most growth analyses for at least some lizard species of the family Phrynosomatidae (Lemos-Espinal and Ballinger 1995; Zufiga-Vega et al. 2008; Pérez-Mendoza et al. 2014; Ramirez-Bautista et al. 2016). The pattern of low growth rate variation in both populations observed in this study could suggest that populations of this species are able to inhabit areas with certain degrees of disturbance, as has been shown in other species of lizards analyzed by D’Cruze and Kumar (2011) in both disturbed and undisturbed environments.
On the other hand, a homologous pattern in growth rates for males and females in both UA and DA populations may be due to the similarities in SVL at birth and the SVL at sexual maturity. For the former (SVL at birth), growth rates are likely to be regulated by predation intensity, acting mainly on offspring and juveniles of both populations (Schoener 1979; Andrews 1982). This interpretation could be different 1f both demographic parameters (survivorship and recaptures) evaluated for each site had changed based on an increased number of recaptures. This pattern has also been observed in populations of S. grammicus from Central Mexico (Pérez-Mendoza et al. 2013, 2014).
The life history characteristics studied here for this species could have significant plasticity among its populations, and therefore, small differences in temperature, precipitation, and food between UA and DA would not have apparent effects (Valdez-Gonzalez and Ramirez-Bautista 2002; Valencia-Limon et al. 2014). A similar pattern in growth rates also occurs between sexes and age classes in other lizard genera (e.g., Xenosaurus spp.; Molina-Zuluaga et al. 2013).
The low variation in SVL at sexual maturity observed in the growth curve within and between populations is partially explained by the absence of sexual dimorphism with respect to SVL (Valdéz-Gonzalez and Ramirez- Bautista 2002; Ramirez-Bautista et al. 2013). Walkup et al. (2017) pointed out that Uta stansburiana, Aspidoscelis marmorata, A. sexlineata, and Sceloporus consobrinus present generalist habits in microhabitat choice, being able to inhabit sites with different degrees of disturbance. Consequently, these species tend to present reduced variation in some of their demographic characteristics, as a measure of phenotypic plasticity towards different degrees of environmental disturbance.
In this study, lizards from UA were numerically more abundant and showed a slightly higher survival rate
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