Zoosyst. Evol. 97 (2) 2021, 315-343 | DOI 10.3897/zse.97.63936 ee Ee BERLIN Uncovering the herpetological diversity of small forest fragments in south-eastern Madagascar (Haute Matsiatra) Francesco Belluardo', Darwin Diaz Quirés!, Javier Lobon-Rovira!, Goncalo M. Rosa*®?, Malalatiana Rasoazanany*, Franco Andreone®, Angelica Crottini' © 1 CIBIO Centro de Investigagdo em Biodiversidade e Recursos Genéticos, Universidade do Porto, Rua Padre Armando Quintas, Campus de Vairdo, 4485—661, Vairdo, Portugal 2 Institute of Zoology, Zoological Society of London, Outer Circle, Regent's Park, NW1, 4RY, London, United Kingdom 3 Centre for Ecology, Evolution and Environmental Changes (CE3C), Faculdade de Ciéncias da Universidade de Lisboa, Campo Grande, Edificio C2, 5° Piso, Sala 2.5.46, 1749-016, Lisboa, Portugal 4 Mention Zoologie et Biodiversité Animal, Domaine des Sciences et Technologies, Université d’Antananarivo, B.P. 906, 101, Antananarivo, Madagascar 5 Museo Regionale di Scienze Naturali, Via G. Giolitti, 36, 10123, Torino, Italy 6 Departamento de Biologia, Faculdade de Ciéncias da Universidade do Porto, Rua Campo Alegre, 4169-007, Porto, Portugal http://zoobank. org/5381781A-2801-4240-9A CE-F53CE3264B70 Corresponding author: Francesco Belluardo (france89belluardo@gmail.com) Academic editor: Johannes Penner # Received 3 February 2021 # Accepted 20 May 2021 Published | July 2021 Abstract Madagascar has historically suffered from high fragmentation of forested habitats, often leading to biodiversity loss. Neverthless, forest fragments still retain high levels of biological diversity. The Haute Matsiatra Region (south-eastern Madagascar) hosts the renowned Andringitra National Park and several surrounding isolated forest fragments embedded in a matrix of human-dominated landscape. During a herpetological survey conducted in the Region, we visited a total of 25 sites. We applied a molecular taxonomic approach to identify the collected material and generate new reference sequences to improve the molecular identification of Malagasy herpetofauna. We identified a total of 28 amphibian and 38 squamate taxa and provided a systematic account for each one of them. Nine of the identified taxa are candidate species, amongst which one was newly identified. We extended the known distributional range of 21 taxa (nine amphibians and 12 squamates). Although the largest forest fragments hold a higher number of species, we also detected a relatively high herpetological diversity in small patches. Our results highlight the importance of investigating small forest fragments to contribute to a better understanding of the patterns of diversity and distribution of the amphibians and reptiles of Madagascar. Key Words herpetofauna, forest patches, Andringitra, barcoding, 16S, COI, microendemic, rapid assessment Introduction genera being entirely limited to Madagascar. At the spe- cies level, 92% of native non-marine “reptiles” (intended Ranked as one of the top megadiversity hotspots on Earth, Madagascar hosts exceptional and highly threat- ened fauna and flora (Myers et al. 2000; Wilmé et al. 2006). The proportion of native endemic vertebrate fau- na is remarkable, with families, subfamilies and several as all Sauropsida, excluding birds) and 100% of native amphibians are endemic to the Island (Glaw and Vences 2007; AmphibiaWeb 2021; Uetz et al. 2021). Yet, some recent anthropogenic introductions are currently known (e.g. Duttaphrynus melanostictus, Licata et al. 2020; Copyright Francesco Belluardo et al. This is an open access article distributed under the terms of the Creative Commons Attribution License (CC BY 4.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. 316 Hoplobatrachus tigerinus, Mohanty et al. 2021; Agama agama, Wagner et al. 2009; Indotyphlops braminus, Uetz et al. 2021). The great environmental and bioclimatic heterogeneity of the Island has played a major role in the diversification of the rich Malagasy biota (Vences et al. 2009; Ganzhorn et al. 2014). Following Cornet (1974) and Schatz (2000), five major biomes are recognised in Madagascar: the eastern rainforest, the western dry de- ciduous forest, the sub-humid forest of the central high- lands, the southern sub-arid spiny forest and the montane thickets. The main biodiversity distributional patterns largely follow this bioclimatic subdivision, with the highest abundance of both amphibian and reptile species found along the eastern rainforest belt (Goodman and Benstead 2003; Wollenberg et al. 2008; Crottini et al. 2012a; Brown et al. 2014, 2016). Amphibians and reptiles are particularly diverse, with ca. 369 and 440 currently recognised species, respective- ly (Glaw and Vences 2007; AmphibiaWeb 2021; Uetz et al. 2021). Nevertheless, the level of undescribed diversi- ty is high, as reported by Nagy et al. (2012) for reptiles and Perl et al. (2014) for amphibians. Field research ef- forts over the last three decades, the widespread use of large-scale species inventories and the application of an integrative taxonomic approach employing molecular, morphological and bioacoustic (for amphibians) identi- fication, coupled with voucher collection (Yoder et al. 2005; Padial et al. 2010), enabled impressive progress in uncovering this hidden diversity (e.g. Rosa et al. 2012; Cocca et al. 2018). In times of major biodiversity loss, field research 1s fundamental to catalogue world biodi- versity and represents a fundamental step for its conser- vation (Dijkstra 2016). As in several other places of the world (Bohm et al. 2013), a large portion of Malagasy herpetofauna is at high risk of extinction (e.g. Irwin et al. 2010; Jenkins et al. 2014), with 46.2% of the assessed amphibians and 37.7% of the assessed reptiles currently listed as threatened according to the IUCN Red List cat- egories (Vulnerable, Endangered and Critically Endan- gered) (IUCN 2020). With most species being forest dwellers, the major threat to Malagasy amphibians and reptiles is forest loss, degradation and fragmentation (e.g. Irwin et al. 2010; Bohm et al. 2013; Jenkins et al. 2014; Riemann et al. 2017). The extent of deforestation that Malagasy biota has historically undergone is dramatic (Hornac 1943; Ja- rosz 1993; Vieilledent et al. 2018). In the first 30 years of French colonisation (1895-1925), the amount of pri- mary forest that was destroyed is estimated to be 70% of the vegetation present in the pre-colonial period (Hornac 1943; Jarosz 1993), while a further 44% of the remaining forest was estimated to be lost in the period of 1953-2014 (Vieilledent et al. 2018). Deforestation not only results in an overall decrease in the surface of forested areas, but is also responsible for the heavy fragmentation of remaining forests. In fact, Vieilledent et al. (2018) estimated that, in 2014, 46% of Malagasy remaining forests were less than 100 m distant from forest edges. Species richness tends zse.pensoft.net Belluardo, F. et al.: Herpetological survey in Haute Matsiatra to decrease with fragment size and species composition is also affected by fragmentation, with forest specialists disappearing rapidly with increasing degradation and fragmentation (e.g. Vallan 2002; Lehtinen and Ramana- manjato 2006; Riemann et al. 2015; Nopper et al. 2018). Nevertheless, the diversity and endemism in small forest fragments remain notable (e.g. Rosa et al. 2012). These small patches, embedded in a mosaic landscape, can act as refugia to local herpetofauna (Crottini et al. 2011a; Durkin et al. 2011; Riemann et al. 2015). The description of several new species, microendemic to these tiny forest fragments, has further increased the awareness on their conservation value (e.g. Gehring et al. 2010; Rosa et al. 2014; Jenkins et al. 2014; Crottini et al. 2015; Protzel et al. 2018). The Region of Andringitra is located in south-eastern Madagascar (Haute Matsiatra Region). This area is dom- inated by the Andringitra Massif, protected by Andringi- tra National Park (Fig. 1). Most of the herpetological re- search conducted in this Region has been focused on the protected area. The Andringitra Massif was first surveyed over 90 years ago and the first species inventories took place during the 1970s (see Paulian et al. 1971; Nicoll and Langrand 1989; Blommers-Schlésser and Blanc 1993; Goodman 1996; Raselimanana 1999; Goodman and Razafindratsita 2001). In 1993, Goodman (1996) car- ried out a thorough herpetological assessment within the framework of a multidisciplinary inventory of the east- ern slopes of the massif. Several amphibian and reptile species were collected, providing a fundamental contri- bution to the knowledge of the local herpetofauna. The most updated species list of Andringitra National Park includes 50 amphibians and 40 reptiles (of which five are locally endemic to the Andringitra Massif ), resulting from several biological inventories (both published and unpublished) and observations from the area (Goodman et al. 2018). In the area surrounding the Andringitra Massif (espe- cially the western side), several small patches of forest are the remains of a much more extensive forest cover (Fig. 1). This area is dominated by a human-modified landscape made of pastures, villages and rice fields (Crottini et al. 2012b; Gould and Andrianomena 2015). Knowledge of the herpetofauna that inhabits these small forest patches is scarce (see Glaw and Vences 2007) and most of the information comes from Anja Community Reserve (e.g. Crottini et al. 2011b, 2012b, 2015), a tiny forest fragment managed by the local community (Fig. 1A). Three reptile species were recently described from Anja and, to date, they are reliably known only from this Reserve (or from a few scattered isolated rock boulders close by): Brookesia brunoi Crottini, Miralles, Glaw, Harris, Lima & Vences, 2012; Paragehyra felicitae Crot- tini, Harris, Miralles, Glaw, Jenkins, Randrianantoandro, Bauer & Vences, 2014 and Phelsuma gouldi Crottini, Gehring, Glaw, Harris, Lima & Vences, 2011. These ex- ceptional findings stress the conservation value of this small forest fragment and point to the importance and Zoosyst. Evol. 97 (2) 2021, 315-343 Figure 1. Map of the study area and sampling sites. The borders of Andringitra National Park (red) and Paysage Harmonieux Protégé du Corridor Forestier Ambositra—Vondrozo (Ambositra—Vondrozo Forest Corridor) (blue) are shown. See Suppl. material 1: Table S1 for more details on the sampled localities. Map data ©2015 Google (QGIS Development Team 2020). ANP ES — An- dringitra National Park Eastern Slopes, ANP WS — Andringitra National Park Western Slopes. A. Anja; B. Sakaviro; C. Ambato- mainty; D. Western slopes of the Andringitra Massif from Iantaranomby (ANP WS); E. Imaitso (ANP ES); F. Belambo (ANP ES); G. Fivahona—Velotsoa; H. Map with sampling sites. Photographs by Javier Lobon-Rovira. potential herpetological interest of all other nearby frag- ments of the Region. We surveyed the Region of Andringitra, collected tis- sue samples and specimens, and performed a species-lev- el identification of the sampled amphibians and reptiles. Despite visiting some localities within the borders of Andringitra National Park, we focused our efforts on the several forest fragments surrounding the Massif, to fill the knowledge gap of the herpetological diversity of these poorly explored areas. Here, we provide a first barcoding reference database for the surveyed areas. Methods Study area The study area is in the administrative region of Haute Matsiatra, encompassing a portion of the Andringi- tra Massif and the areas in the immediate surroundings (Fig. 1). This mountain chain dominates the area and is composed of several granitic peaks, amongst which Pic Boby soars as the highest of the region (2,658 m a.s.1.) and as the second highest in the whole country (Nicoll and Langrand 1989; Goodman 1996). The Massif is located at the south-eastern limit between the eastern escarpment and the central high plateau, which determines a sharp bioclimatic gradient with humid conditions in the eastern part and drier weather in the western (Goodman 1996). The regional climate can be defined as cold and humid with marked seasonality (Vidal Romani et al. 2002). Be- tween May and October, the weather 1s cold and dry, with extreme temperatures that can drop below 0 °C at night, whereas the following season, from November to April, it is warm and wet with heavy rainfalls that represent 80% of the yearly precipitations (Vidal Romani et al. 2002). The strong elevational and climatic variability is respon- sible for the great diversity of habitats, amongst which there are lowland rainforest and dry forest in the east- ern and western parts of the Region, respectively, and, at higher elevations, montane meadows, heathlands and rocky outcrops (Goodman 1996; Goodman et al. 2018). Most of the Massif is included in Andringitra National Park, which protects an overall area of 31,160 ha (Nicoll zse.pensoft.net 318 and Langrand 1989). Paysage Harmonieux Protégé du Corridor Forestier Ambositra—Vondrozo (thereafter “Ambositra—Vondrozo Forest Corridor’) is eastwards of Andringitra National Park and protects the ca. 200 km of low-elevation forest corridor connecting Vondro- zo, Andringitra and Pic d’Ivohibe Special Reserve (ca. 10 km further south than Andringitra National Park) with Ranomafana National Park and Ambositra further north (Fig. 1) (Goodman et al. 2018). Besides these protected areas, three private Reserves are present in the surround- ings of the Andringitra Massif: Anja Community Reserve, Sakaviro Community Reserve and Tsaranoro Valley For- est (Fig. 1A and B). They are characterised by fragments of semi-arid deciduous forest located at ca. 950 m a.s.l. at the base of low-elevation granitic mountains rising a few hundred metres relative to the ground level. These for- est fragments are particularly small: 36 ha (Anja), 14 ha (Sakaviro) and 46 ha (Tsaranoro) (Crottini et al. 2012b; Gould and Andrianomena 2015). Several other small for- est fragments are scattered throughout the area, especially in the western part of the Region, but no legal protection is known for these sites. We sampled in 25 localities, eight of which are within Andringitra National Park, one locality within Ambositra—Vondrozo Forest Corridor and 16 sites are located in the surroundings of the Andringitra Massif (Fig. 1; Suppl. material 1: Table S1). In the present study, we refer to Andringitra as the overall study area comprising the eponymous Massif and the forest frag- ments in the surrounding areas that were investigated, irrespective of the limits of the protected areas present in the region (Fig. 1). Sampling The samples included in this study were collected be- tween 2009 and 2018, although most of the sampling ef- fort was deployed between the 13th of November and the 18th of December 2018. This period matches the onset of the rainy season and the peak of activity of reptiles and amphibians. We spent a minimum of two days at each sampling locality (defined as where campsites were es- tablished). Continuous opportunistic searches took place while moving amongst different sites (Suppl. material 1: Table S1). Animals were opportunistically sought in all microhabitats during both day and night searches, visual- ly detected and caught by hand. Each individual was pho- tographed and the geographic location was recorded with a GPS receiver. One tissue sample was taken from each collected individual. Samples were stored in 96% etha- nol and the caught animals were released upon sampling. We identified each individual, based on the morpholog- ical descriptions provided in Glaw and Vences (2007) and subsequent descriptions and, in the case of putative candidate species, a limited number of specimens across different localities were collected as vouchers. These in- dividuals were anesthetised and subsequently euthanised with an overdose of MS222 either by immersion in a zse.pensoft.net Belluardo, F. et al.: Herpetological survey in Haute Matsiatra saturated solution (amphibians) or through intracoelom- ic injection (reptiles) of the same solution. All voucher specimens were fixed in 96% ethanol and placed in 70% ethanol for long-term storage. Laboratory procedures Total genomic DNA was extracted from tissue samples following the protocol described in Bruford et al. (1992), consisting of a high-salt extraction using proteinase K digestion (10 mg/ml concentration). We amplified a frag- ment of ca. 550 bp of the 3’ terminus of the mitochon- drial 16S rRNA gene (16S) for all amphibian samples (Palumbi et al. 1991) and a fragment of ca. 650 bp of the cytochrome oxidase I gene (COI) for reptiles (Nagy et al. 2012), both of which have been widely used for mo- lecular taxonomic identification of Malagasy herpetofau- na (Vences et al. 2005); Nagy et al. 2012. Whenever the amplification of the two markers was either unsuccessful or not informative to provide an accurate species identi- fication, we amplified additional markers, namely the 5’ terminus of the 16S fragment (16S 5’) for amphibians and 16S, NADH dehydrogenase subunit 1 (ND1), NADH de- hydrogenase subunit 2 (ND2) or Cytochrome b (Cytb) for reptiles (see Suppl. material 2: Table S2 for primers and PCR conditions). Successfully amplified samples were sequenced with an ABI 3730XL automated sequencer at Macrogen Inc. (Spain). Chromatograms were manually checked and edited, when necessary, with BIOEDIT 7.2.6 (Hall 1999). Molecular species identification Newly-generated sequences were aligned with the Clust- al W algorithm implemented in BIOEDIT 7.2.6 (Thomp- son et al. 1994; Hall 1999). Neighbor joining trees, based on Kimura 2—parameter model distances, were computed for each investigated gene with MEGA X 10.0.5 (Ku- mar et al. 2018) to roughly divide the samples into ge- netically uniform groups (16S for amphibians and COI for reptiles). Each group was compared to the molecular database available in GenBank through the BLAST algo- rithm (https://blast.ncbi.nlm.nih.gov/Blast.cgi) (Altschul et al. 1997), using nblast default parameters, to retrieve the most similar homologous sequences available in the online database. Species identification was based on the inter-specific thresholds suggested by Vieites et al. (2009) for the 16S fragment in amphibians (3%) and the different thresholds for the different Malagasy reptile groups sug- gested by Nagy et al. (2012) for the COI fragment. Mo- lecular identification was confirmed by the analysis of the photographic records. In a few instances where molecular data were not available, sample identification was based on the photographic records or the morphological exam- ination of the collected specimens. We used MEGA X 10.0.5 (Kumar et al. 2018) to compute the average uncor- Zoosyst. Evol. 97 (2) 2021, 315-343 rected pairwise genetic distances (uncorrected p—distanc- es) for each identified taxon. Whenever it was not possi- ble to assign a sample to a formally described species, we used the definitions of different categories of candidate species proposed in Vieites et al. (2009). The working names for the candidate species follow the nomenclature proposed in Vieites et al. (2009) and following publica- tions (e.g. Zimkus et al. 2017) or, in case of first identifi- cation, we used the species epithet of the morphologically most similar species, which was prefixed with “sp. aff.” and followed by the locality name in quotation marks. To verify if each taxon was already reported from the Region, we retrieved information from Glaw and Vences (2007), Brown et al. (2014), the IUCN Red List of Threat- ened Species (IUCN 2020), Goodman et al. (2018) and tax- on-specific publications. Species records that were found to be outside the known species distribution were considered range extensions. Information on species identification, distribution, microhabitat preferences and intra-specific ge- netic variability of each identified taxon are reported in the section Species accounts. Results We generated a total of 520 sequences (308 of amphibians and 212 of reptiles; see Suppl. material 3: Table S3 and Suppl. material 4: Table S4 for GenBank accession num- bers (MZ285088—MZ285597) and sampling information and identification of each analysed sample; see Suppl. ma- terial 9: Table S7 for locality records of all identified spe- cies). Thirteen records (four for amphibians and nine for reptiles) were based on the inspection of photographic ma- terial or the morphological examination of the preserved specimens. Neighbor joining trees of the 16S (amphib- ians) and COI (reptiles) gene fragments are provided in Suppl. material 7: Fig. S1 and Suppl. material 8: Fig. S2. We sampled a total of 28 amphibian taxa. Most of them belong to the Mantellidae family, of which were 11 Boophinae, 10 Mantellinae and two Laliostominae. We also recorded two hyperoliids and Ptychadena mas- careniensis (Ptychadenidae). Only two microhylids, both belonging to the subfamily Scaphiophryninae, were sampled. We collected four candidate species (Boophis sp. Ca33, Mantidactylus sp. Cal4, Mantidactylus sp. Ca48 and Ptychadena sp. aff. mascareniensis “OTU1”). We contributed to extending the current species range for nine taxa (Heterixalus luteostriatus, Scaphiophryne (Pseudohemisus) calcarata, Boophis (Sahona) doulioti, Boophis (Boophis) boppa, Boophis (Boophis) occidenta- lis, Boophis (Boophis) rhodoscelis, Gephyromantis (Phy- lacomantis) corvus, Mantella betsileo and Mantidactylus (Brygoomantis) sp. Cal4). Amongst the collected materi- al, two taxa are microendemic to the area: Boophis lau- renti and Mantidactylus bourgati. We identified a total of 38 squamate reptiles. Geckos and chameleons are the most represented groups in our sampling, with 10 taxa each. They are followed by pseu- 319 doxyrhophiid snakes (six), skinks (four), gerrhosaurids (three), oplurids (two), sanziniid snakes (two) and one psammophiid snake. We collected five candidate species (Lygodactylus sp. aff. pictus Ca01 “Isalo”, Paragehyra sp. aff. felicitae ““Tsaranoro”, Paroedura sp. aff. bastardi Lineage D, Trachylepis sp. aff. vato and Pseudoxyrho- pus sp. Ca2). Amongst these, Paragehyra sp. aff. felic- itae “Tsaranoro” was unknown to science and identified in the present study for the first time. We contributed to extending the known distribution area for 12 taxa (Furci- fer nicosiai, Furcifer willsti, Lygodactylus sp. aff. pictus Ca01 “Isalo”, Paragehyra felicitae, Paroedura rennerae, Paroedura sp. aff. bastardi Lineage D, Phelsuma gouldi, Phelsuma lineata elanthana, Zonosaurus laticaudatus, Trachylepis sp. aft. vato, Leioheterodon modestus and Pseudoxyrhopus sp. Ca2). Amongst the collected mate- rial, five taxa are microendemic to the surveyed region (Brookesia brunoi, Paragehyra felicitae, Paragehyra sp. aff. felicitae “Tsaranoro”, Paroedura sp. aff. bastardi Lin- eage D and Phelsuma gouldi). Species accounts Amphibians Family Hyperoliidae Subfamily Hyperoliinae Heterixalus betsileo (Grandidier, 1872) Fig. 2L Heterixalus betsileo occurs in Madagascar’s central high- lands, where it can be found in swamps and rice fields (Glaw and Vences 2007). Reported within Andringitra Na- tional Park, which is close to the southern limit of its dis- tributional range (Glaw and Vences 2007; Goodman et al. 2018), the species was sampled at night within the forests of Anja and Sakaviro, in the western part of the surveyed region, between ca. 930 and 1,020 m a.s.l. (Suppl. material 3: Table S3). The species resulted in being common in both fragments. The two sampled individuals are genetically al- most identical to each other (Suppl. material 5: Table S5) and to published sequences of this taxon sampled in An- dringitra (16S: EF646668, JQ346497; 16S 5’: EF646633) and Andasibe (e.g. 16S: EF646661). Heterixalus luteostriatus (Andersson, 1910) Fig. 2M Heterixalus luteostriatus has a wide and discontinuous distribution across north-western and western Mad- agascar, including the Isalo Massif. This species can commonly be found in swamps and rice fields (Glaw and Vences 2007; Mercurio et al. 2008) and the record reported here constitutes an extension of the known species distributional range by ca. 180 km to the east from the Isalo Massif (Glaw and Vences 2007). Our study found this taxon at Anja, Anja—Ambilo, Iarintsena, zse.pensoft.net 320 : 2 Belluardo, F. et al.: Herpetological survey in Haute Matsiatra Figure 2. Hyperoliid, microhylid, ptychadenid and mantellid (subfamilies Boophinae and Laliostominae) species identified in this study. Sampling localities for each photographed individual are provided. ANP ES — Andringitra National Park Eastern Slopes; ANP WS - Andringitra National Park Western Slopes (Fig. 1; Suppl. material 1: Table $1). A. Boophis (Boophis) ankaratra from Imaitso (ANP ES); B. Boophis (Boophis) boppa from Iantaranomby (ANP WS); C. Boophis (Sahona) doulioti from Ambalavao; D. Boophis (Boophis) laurenti from Iantaranomby (ANP WS); E. Boophis (Boophis) luteus from Fivahona—Velotsoa; F. Boophis (Boophis) majori from Asaramanitra (ANP ES); G. Boophis (Boophis) obscurus from Imaitso (ANP ES); H. Boophis (Boophis) occidentalis from Andramena (ANP WS); I. Boophis (Boophis) popi from Imaitso (ANP ES); J. Boophis (Boophis) sp. Ca33 from Asaramanitra (ANP ES); K. Boophis (Boophis) rhodoscelis from Fivahona—Ambavanala; L. Heterixalus betsileo from Sakaviro; M. Heterixalus luteostriatus from Anja; N. Scaphiophryne (Scaphiophryne) madagascariensis from Andramena (ANP WS); O. Scaphiophryne (Pseudohemisus) calcarata from Ambalavao; P. Laliostoma labrosum from Anja; Q. Aglyptodactylus madagas- cariensis from Namoly; R. Ptychadena sp. aff. mascareniensis “OTU1” from Iantaranomby (ANP WS). Photographs by Javier Lobon-Rovira (A—J, L—-R) and Francesco Belluardo (K). Sakaviro and Tsaranoro (between ca. 940 and 1,020 m a.s.1.) (Suppl. material 3: Table S3), sometimes in synto- py with H. betsileo. This species seemed abundant and was found at both day (sleeping on leaves) and night on small shrubs in areas with degraded and open vegeta- tion outside forests. The 16S sequences of the individual sampled during our survey are identical to each other (Suppl. material 5: Table S5) and individuals from Isalo (e.g. KX066672). The genetic distance between the in- dividuals of H. /uteostriatus and H. betsileo sampled in this study is 3.33%. zse.pensoft.net Family Microhylidae Subfamily Scaphiophryninae Scaphiophryne (Pseudohemisus) calcarata (Mocquard, 1895) Fig. 20 Scaphiophryne calcarata belongs to the subgenus Pseudohemisus. A recent study assigned the lectotype of five nomina (including Calophrynus_ calcaratus Mocquard, 1895) (Scherz et al. 2021). This taxonomic Zoosyst. Evol. 97 (2) 2021, 315-343 proposal, together with the analysis of newly-collect- ed material, enabled the restriction of the lineage of Scaphiophryne calcarata to the southern and south-west- ern portion of the Island. The samples analysed in this study, collected at night in Ambalavao and Sakaviro (at ca. 1,000-1,010 m a.s.l.), are assigned to this lineage. They were included in Scherz et al. (2021) contribut- ing to extend the known distributional range by ca. 180 km in a northern direction. This species was rare and only two individuals were found active at night: one in the garden of a hotel and the second on the ground in an open environment next to a small forest patch sur- rounded by pastures (Suppl. material 3: Table S3). The two analysed samples are identical to each other (Sup- pl. material 5: Table S5) and to individuals from Isalo (e.g. MH063283), and are almost identical to the oth- er published sequences of this species (e.g. Berenty, AY 834192; Tolagnaro AY834193). The genetic distance from individuals of S. obscura and the north-west lineage is ca. 3.3% (e.g. Kirindy, KU937802; Isalo, KX066692) and ca. 4% (Ampijoroa, KU937797), respectively. Scaphiophryne (Scaphiophryne) madagascariensis (Boulenger, 1882) Fig. 2N This species is distributed in the central highlands, be- tween the Ankaratra Massif in the north and the Andringi- tra Massif, in the south. This frog generally mhabits high elevation sites, both above and below the tree line (Glaw and Vences 2007; Goodman et al. 2018). Scaphiophryne madagascariensis was sampled at Andramena, Asarama- nitra and Belambo, on both the eastern and western slopes of the Massif and between ca. 1,580 and 1,740 ma.s.l. The species was frequently encountered during our surveys. The individuals were often spotted in clearings next to a forest (Suppl. material 3: Table S3), both during day and night. We observed no genetic difference amongst the anal- ysed samples (Suppl. material 5: Table S5). When compar- ing the samples analysed here with the sequences available in Genbank, we found no difference between our samples and individuals collected in the same area (Andohariana Plateau, DQ787110) and they are less than 1% divergent from individuals from Ankaratra (KC1 80053). Family Ptychadenidae Ptychadena sp. aff. mascareniensis “OTU1” UCS Fig. 2R Ptychadena mascareniensis (Duméril & Bibron, 1841) is the most widespread amphibian in Madagascar. It is usu- ally found next to any pond, swamp and rice field, often outside the forest. Until a few years ago, Malagasy pop- ulations were considered conspecific to the populations from mainland Africa and it was thought that the species had been recently introduced to the Island (Glaw and Vences 2007). A recent study showed that P. mascare- 321 niensis 1S a species complex. Malagasy populations are not conspecific with the populations of the African con- tinent and the populations assigned to P. mascareniensis from Madagascar belong to three operational taxonomic units (OTUs) in need of taxonomic revision (Zimkus et al. 2017). All the samples analysed in this study belong to P. sp. aff. mascareniensis “OTU1” (sensu Zimkus et al. 2017). This species was amongst the most frequent and abundant frogs we recorded, found both during day and night, despite being more commonly active during the day. It was normally found in rice fields or close to temporary water bodies, between ca. 870 and 1,650 m a.s.l. (Suppl. material 3: Table S3). We sampled this taxon at Andra- mena, Asaramanitra, Iantaranomby, Imaitso, Ambalavao, Ambatomainty, Antanifotsy 1, 3, 5, Anja, Fivahona—Am- bavanala, Fivahona—Velotsoa, Namoly and Tsaranoro. The individuals, analysed for this study, are genetically uniform amongst each other (Suppl. material 5: Table S5) and are almost identical to other individuals collected in the area (Andohariana: AY517587 and AY517588) and across Madagascar (e.g. Toliara, KX836419; Ranomafa- na, KX836413; Bemaraha, KX836392; Andohahela, KX836390; Isalo, KX066671). Family Mantellidae Subfamily Boophinae Boophis (Sahona) doulioti (Angel, 1934) Fig. 2C Distributed in western and southern Madagascar, this tax- on is mostly found in open areas and secondary vegetation (Glaw and Vences 2007). Boophis doulioti had not been reported to this area before and our finding represents an extension of the species distributional range by ca. 150 km towards the east. Individuals were spotted active at night in Anja and Jarintsena (on the western side of the Andrin- ditra Massif), perching on trees and shrubs within semi-ar- id deciduous forest, in open environments next to villages, close to rice fields and within the town of Ambalavao at an elevation of ca. 950—1,030 m a.s.l (Suppl. material 3: Table S3). The analysed samples are all identical to each other (Suppl. material 5: Table S5) and samples collect- ed at Isalo (KX066561), Tranomaro (MK132751) and Ranomafana (AY848515; this record being reported as Boophis tephraeomystax (Duméril, 1853)). Boophis (Boophis) ankaratra Andreone, 1993 Fig. 2A Boophis ankaratra is commonly found in the central highlands at high-elevation locations, both in rainfor- est and degraded gallery vegetation (Glaw and Vences 2007; IUCN SSC Amphibian Specialist Group 2016a). We collected a single individual during a night search at Imaitso, within an area of dense rainforest (ca. 1,670 m a.s.l.), perching on riverine vegetation surrounding a small stream (Suppl. material 3: Table S3). This sample zse.pensoft.net 322 is identical to other available sequences from the An- dringitra Region (Andringitra: AF411611; Imaitso For- est: DQ068396, DQ068397, DQ068398). The individu- al analysed in this study is molecularly almost identical to the population from Ranomafana (difference: 0.30%; GU974475). Boophis (Boophis) boppa Hutter, Lambert, Cobb, An- driampenomanana & Vences, 2015 Fig. 2B Boophis boppa has been recently described from Ranomafana National Park and Antoetra (Andreone et al. 2007; Hutter et al. 2015). This record represents a range extension for the species by ca. 80 km towards the south. Individuals were locally abundant and were collected at Asaramanitra (along the eastern slope of the Massif) and Iantaranomby (on the western slope), at an elevation of ca. 1,580—1,600 m a.s.l. Sampled individuals were perch- ing at night on riverine vegetation along large streams in both forested areas and open environments with scattered trees (Suppl. material 3: Table S3). The analysed samples collected in this study are identical (Suppl. material 5: Table S5). They are also identical to individuals from the type locality (Ranomafana: KT588038) and very similar (99.80% similarity) to individuals from Antoetra (e.g. AY 848438). Boophis (Boophis) laurenti (Guibé, 1947) Fig. 2D This species is currently known only from Andringitra National Park, where it can be found in montane heath- lands above 1,500 m a.s.l. (UCN SSC Amphibian Spe- cialist Group 2016b; Goodman et al. 2018). We sampled Boophis laurenti either on mossy rocks in streams or perching on dense vegetation and scattered trees close to running water. The species was detected during day and night (although more frequently active at night) and seemed common along both the western (Andramena, Iantaranomby) and the eastern (Diavolana) slopes of the Massif (between ca. 1,580 and 1,740 m a.s.l.) (Suppl. material 3: Table S3). Our samples are genetically uni- form to each other (Suppl. material 5: Tables S5) and in comparison with previously available sequences (Ando- hariana: AY848599, AY659964; Andringitra: AY659963; Cuvette Boby: AY848575). Boophis (Boophis) luteus (Boulenger, 1882) Fig. 2E This species can be found along streams in rainforest and secondary vegetation in several localities of eastern Madagascar (including Andringitra National Park), but also in Isalo in the south-west and Ambohitantely in the central highlands (Glaw and Vences 2007; Cocca et al. 2018; Goodman et al. 2018). The species was rare and only two individuals were spotted at Fivahona—Velot- zse.pensoft.net Belluardo, F. et al.: Herpetological survey in Haute Matsiatra soa, in the eastern part of the surveyed region, at ca. 1,270 m a.s.l. (Suppl. material 3: Table S3). Both indi- viduals were found in roosting positions on trees near a stream at night. They show limited genetic differen- tiation and are almost identical to individuals collected at Ranomafana (e.g. FJ559330) and Isalo (FJ559354) (Suppl. material 5: Table S5). Boophis (Boophis) majori (Boulenger, 1896) Fig. 2F Boophis majori is distributed in eastern Madagascar with- in a restricted region comprised of Antoetra, Ranomafa- na, Andringitra National Park (Imaitso Forest) (Brown et al. 2014) and Ivohibe, where it can be found on trees along rainforest streams (Glaw and Vences 2007; IUCN SSC Amphibian Specialist Group 2016c; Goodman et al. 2018). Boophis majori was a common species. During a night search, we collected three samples perching on trees along a large stream within rainforest habitat in Asaramanitra, on the eastern slope of the Massif, at ca. 1,590 m as.l. (Suppl. material 3: Table $3). These sam- ples are identical to each other (Suppl. material 5: Table S5) and have 0.50% genetic distance with samples from Ranomafana (e.g. AY 848586). Boophis (Boophis) obscurus (Boettger, 1913) Fig. 2G Boophis obscurus was recently resurrected from the synonymy with Boophis goudotii Tschudi, 1838 (Glaw et al. 2010). The species is distributed in south-eastern Madagascar between Ranomafana, Isalo and Andringitra National Park (Andohariana Plateau) (Glaw et al. 2010; IUCN SSC Amphibian Specialist Group 2016d; Good- man et al. 2018). Boophis obsurus was common and abundant across the Region and was sampled in Andra- mena, Asaramanitra, Belambo, Iantaranomby, Imaitso, Riandahy, Siranandambo, Fivahona—Ambavanala, Fiva- hona—Velotsoa, Namoly and Tsaranoro (Suppl. material 3: Table S3). Boophis obscurus was collected between ca. 950 and 1,740 m as.l., normally found on rocks and boulders along water streams. It was sampled during both diurnal and nocturnal searches, despite being more frequent at night. The samples analysed in this study are molecularly similar to each other (Suppl. material 5: Table S5) and another sample collected from the same area (AY 848568). They show only limited genetic differ- ence to samples from other localities (e.g. Ranomafana: GU975058; Isalo: KX066565). Boophis (Boophis) occidentalis Glaw & Vences, 1994 Fig. 2H Boophis occidentalis has a discontinuous distribution in western (Tsingy de Bemaraha) and central Madagas- car (between Isalo, Zazafotsy and Antoetra), where it is found in dry deciduous forest (Vences et al. 2003; Glaw Zoosyst. Evol. 97 (2) 2021, 315-343 and Vences 2007). This species was previously not re- ported from our study area and this record extends the species distributional range by ca. 50 km towards the east. This species was collected in the eastern and west- ern part of the Region (Andramena, Iantaranomby, Am- batomainty, Fivahona—Velotsoa and Tsaranoro; Suppl. material 3: Table S3) between ca. 920 and 1,740 ma.s.l., where it was often found on high trees close to streams. Boophis occidentalis was particularly common in Tsara- noro, whereas, in the other sampling sites, it was rarer. The individuals were all spotted at night. The collected samples are identical (Suppl. material 5: Table S5) and show high genetic affinity with individuals from Isalo (e.g. KX066570) and Antoetra (AY341720), with less than 1% genetic distance. Boophis (Boophis) popi Kéhler, Glaw, Rosa, Gehring, Pabijan, Andreone & Vences, 2011 Fig-2] Boophis popi is distributed in central-eastern Madagascar between Tsinjoarivo and Andringitra. This species can be found along streams in montane rainforest in a narrow elevational range of 1,000—1,500 ma.s.l. (Andreone et al. 2007; Kohler et al. 2011; Goodman et al. 2018). The sam- ples included in the present study were collected on the eastern slopes of the Massif at Imaitso (where other indi- viduals were previously sampled) at higher elevation (ca. 1,520—1,690 m a.s.1.), slightly above the currently known range for the species (Kohler et al. 2011) (Suppl. mate- rial 3: Table S3). The species was particularly abundant in this site. The collected individuals were found along small slow-flowing streams within rainforest, perching on the riverine vegetation. They were mostly spotted at night, despite a single individual being found during the day. The analysed samples are almost identical to each other (Suppl. material 5: Table S5) and to previously available sequences (Andringitra: e.g. JN679879; An- toetra: e.g. AY848551). Boophis (Boophis) rhodoscelis (Boulenger, 1882) Fig. 2K This species is distributed between Ranomafana and An- toetra, where it can be found in swampy areas. Previous records from Ambohitantely have recently been assigned to Boophis andrangoloaka (Ahl, 1928) (Glaw et al. 2010). Boophis rhodoscelis was not yet reported from the study area and this record represents a range extension by ca. 70 km to the south. The species was rare. Two call- ing males were sampled at Fivahona—Ambavanala, in the eastern part of the surveyed region. The individuals were found during the night at ca. 1,480 m a.s.l. perching on trees next to a fast-flowing stream within rainforest (Sup- pl. material 3: Table S3). These two samples are identical to each other (Suppl. material 5: Table S5) and to samples collected at Antoetra (AY848616) and Ranomafana (e.g. AY 848619). 323 Boophis (Boophis) sp. Ca33 UCS Fig. 2J This unconfirmed candidate species is part of the spe- cies group comprising Boophis microtympanum (Boett- ger, 1881) and B. /aurenti. Boophis sp. Ca33 (following the terminology as in Vieites et al. (2009) and reported in Glaw and Vences (2007) as B. sp. aff. microtympanum) is morphologically similar to B. microtympanum from which it is distinguished by a more uniform dorsal colouration rather than the vermiculated pattern described in the nom- inal species. Vocalisations of the two taxa are very similar and present only some quantitative difference in note du- ration (Glaw and Vences 2007; Vieites et al. 2009). Glaw and Vences (2007) report the presence of Boophis sp. Ca33 in the surveyed area at Cirque Namoly and Vieites et al. (2009) states that this taxon was found to live in sympatry with B. microtympanum, an observation that might support the distinction of these taxa (sensu Miralles et al. 2021), although the genetic differentiation at the 16S marker is below the standard threshold of 3% (Vieites et al. 2009). The species was not abundant. We sampled three individ- uals of Boophis sp. Ca33 at Asaramanitra, on the eastern slopes of the Massif. These individuals were found at night perching on branches next to a large fast-flowing stream at ca. 1,590 m a.s.l. of elevation (Suppl. material 3: Table S3). The samples analysed here are identical to each other (Suppl. material 5: Table S5), to other available sequenc- es from the area (AY848597) and Mahahira (AY 848604; in the Ranomafana area). They are also almost identical to samples from Itremo (JF903885). Individuals from Ankaratra (HM769929), Col des Tapias (AJ315918), Ambohitantely (HM769928) and Andasibe (AY848598) attributed to B. microtympanum are ca. 1.60% different. Subfamily Laliostominae Aglyptodactylus madagascariensis (Duméril, 1853) Fig. 2Q This species inhabits rainforest in northern and east- ern Madagascar. The Andringitra Massif represents the southernmost limit of its distributional range and the highest elevational record (Imaitso forest, 1,509 m a.s.1.) for the species (Kohler et al. 2015; Goodman et al. 2018). It is an explosive breeder reproducing in tempo- rary ponds, but it can often be found on the forest floor outside the breeding season (Glaw and Vences 2007). The four analysed individuals were found on the floor of the rainforest at Imaitso and Namoly, in the eastern part of the Region at an elevation range between ca. 1,550 and 1,650 m a.s.l. (Suppl. material 3: Table S3). They were recorded during both night and day, although more frequently spotted at night. The collected samples are genetically identical to each other (Suppl. material 5: Table S5) and to other samples from the Region (e.g. KT159884). They are slightly different (ca. 0.40% ge- zse.pensoft.net 324 netic distance) from individuals collected in Ranomafa- na (e.g. AY847991). Laliostoma labrosum (Cope, 1868) Fig. 2P This species 1s widely distributed in dry habitats of west- ern, northern and southern Madagascar (Glaw and Venc- es 2007). Within the Region of Andringitra, this species was already reported from the town of Ambalavao (e.g. AY 848009). Laliostoma labrosum was not frequently en- countered during our surveys. Individuals were spotted at night in the western portion of the Region in Iantaranomby and Anja. The encounters took place on the floor of semi-ar- id deciduous forest, close to large streams in open environ- ments and within the town of Ambalavao, in a hotel garden, at an elevation between ca. 980 and 1,560 m a.s.l. (Suppl. Material 3: Table S3). The analysed samples are genetically identical to each other (Suppl. material 5: Table S5) and to other populations from different parts of Madagascar (e.g. Toliara: KR337974; Ankarafantsika: KR337954; Kirindy: KR337951:; Isalo: KX066667; Tsaratanana: KR337858). Subfamily Mantellinae Gephyromantis (Phylacomantis) corvus (Glaw & Venc- es, 1994) Fig. 3H The samples of this taxon, collected at Anja, Sakaviro and Tsaranoro (in the western portion of the surveyed area; Suppl. material 3: Table S3), have been included in a re- cent taxonomic study (Cocca et al. 2020) where the au- thors synonymised Gephyromantis azzurrae Mercurio & Andreone, 2007 with G. corvus and described the second Phylacomantis lineage inhabiting Isalo as Gephyromantis kintana Cocca, Andreone, Belluardo, Rosa, Randrianirina, Glaw & Crottini, 2020. Previously known only from the Isalo Massif, where it inhabits large and deep canyons with fast-flowing water and gallery forests (Glaw and Vences 2007; Mercurio and Andreone 2007), these records rep- resent an important range extension for the species by ca. 180 km towards east (Cocca et al. 2020). This species was rare. The individuals were found during night searches at an elevational range of ca. 950—1,020 m a.s.l. along small canyon-like streams in banks, ravines and crevices both within semi-arid deciduous forest and in open habitats. The analysed samples are identical (Suppl. material 5: Table S5) and have a limited genetic differentiation (ca. 0.60%) from the individuals of Isalo (K-X066651). Gephyromantis (Gephyromantis) blanci Guibé, 1974 Fig. 3G Males of Gephyromantis blanci are easily found in rainforest and secondary vegetation while calling on the forest floor or low branches. Ambalamarovandana, zse.pensoft.net Belluardo, F. et al.: Herpetological survey in Haute Matsiatra located in the eastern slopes of the Andringitra Mas- sif, is the type locality of the species (Glaw and Vences 2007; IUCN SSC Amphibian Specialist Group 2016¢e; Goodman et al. 2018). Gephyromantis blanci was \ocal- ly abundant and was sampled at Imaitso and Namoly, in the eastern part of the Region, at an elevation range of ca. 1,540—-1,690 m a.s.l. (Suppl. material 3: Table S3). All collected individuals were males calling from the forest floor within dense rainforest, spotted both during day and night-time. The analysed samples are genet- ically identical to each other and to sequences previ- ously available from the area (e.g. AY848324) (Suppl. material 5: Table S5). Mantella betsileo (Grandidier, 1872) Fig. 3] This species is discontinuously reported from multiple localities in western Madagascar between the Isalo Mas- sif, at the south, and Bemaraha, at the north. Within Isalo National Park, this species can be observed around both temporary and permanent water bodies, generally outside of the canyons (Glaw and Vences 2007; Mercurio et al. 2008). Although not yet confirmed from the surveyed area, the species was described from individuals likely collected in the Betsileo Region, which includes the Re- gion of Andringitra. Sightings of Mantella betsileo were rare in our survey. We found this taxon during the night in the western portion of the Region at Ambatomainty, where two individuals were spotted active within a humid pit in the ground at ca. 930 m a.s.l. of elevation. Another individual was found in Anja, in an open area close to a granitic boulder. These records extend the known distri- butional range of the species by ca. 180 km towards the east (Suppl. material 3: Table S3). The analysed samples are identical (Suppl. material 5: Table S5) and very simi- lar to individuals from Isalo (e.g. EF674841) and Moron- dava (AF215288). Mantidactylus (Brygoomantis) betsileanus (Boulenger, 1882) Fig. 3B This species is distributed in central-eastern Madagascar where it is often found along slow-flowing streams within rainforest, but it can also be found in degraded vegetation and human-dominated areas (e.g. rice fields) (Glaw and Vences 2007). We sampled M. betsileanus at Fivahona— Ambavanala, Fivahona—Velotsoa, Namoly and Tsaranoro, both in the eastern and western sides of the surveyed area (between ca. 930 and 1,650 m a.s.l.) (Suppl. material 3: Table S3). The species seemed abundant, especially in Fivahona—Velotsoa and Tsaranoro. The individuals were spotted both during day and night-time along the banks of slow-flowing streams within forest. Molecularly, the individuals analysed here are uniform (Suppl. material 5: Table S5) and are almost identical to individuals collected at Itremo (JF903887), Ranomafana (AY 848275), Andasibe Zoosyst. Evol. 97 (2) 2021, 315-343 S25 Figure 3. Amphibians of the mantellid subfamily Mantellinae identified in this study. Sampling localities for each photographed individual are provided. ANP ES — Andringitra National Park Eastern Slopes; ANP WS — Andringitra National Park Western Slopes (Fig. 1; Suppl. material 1: Table S1). A. Mantidactylus (Bryogoomantis) bourgati from Namoly; B. Mantidactylus (Bryogoomantis) betsileanus from Namoly; C. Mantidactylus (Chonomantis) delormei from Imaitso (ANP ES); D. Mantidactylus (Ochthomantis) femoralis from Asaramanitra (ANP ES); E. Mantidactylus (Bryogoomantis) sp. Cal4 from Tsaranoro; F. Mantidactylus (Hyloba- trachus) sp. Ca48 from Fivahona—Velotsoa; G. Gephyromantis (Gephyromantis) blanci from Imaitso (ANP ES); H. Gephyromantis (Phylacomantis) corvus from Sakaviro; I. Mantella betsileo from Ambatomainty; J. Subadult and tadpoles of Spinomantis elegans from Imaitso (ANP ES). Photographs by Javier Lobon-Rovira. (e.g. FJ559234), Fierenana (e.g. EF606877) and Mandraka (AY 848238). Mantidactylus (Brygoomantis) bourgati Guibé, 1974 Fig. 3A Endemic to the Andringitra Massif (already reported from Imaitso Forest and Andohariana Plateau), the species is known to live along streams within forest above the tree line (Glaw and Vences 2007). Ambalamarovandana, lo- cated in the eastern slopes of the Massif, is the type lo- cality of the species. We sampled Mantidactylus bourgati across a large portion of the surveyed sites (Andramena, Asaramanitra, Belambo, Iantaranomby, Imaitso, Rianda- hy, Siranandambo, Fivahona—-Ambavanala, Fivahona— Velotsoa, Namoly and Tsaranoro; Suppl. material 3: Table S3), both on the western and eastern portion of the Region at a wide elevational range (between ca. 930 and 1,740 m a.s.l.). In all sites, M. bourgati was abundant and the indi- viduals were spotted during both day and night along the banks of streams within forest. Analysed individuals are genetically almost identical (Suppl. material 5: Table S5). zse.pensoft.net 326 Mantidactylus (Brygoomantis) sp. Ca14 UCS Fig. 3E Mantidactylus sp. Cal4 is closely related to Mantidacty- lus alutus (Peracca, 1893) and morphologically similar to Mantidactylus ulcerosus (Boettger, 1880), relative to which it exhibits significant bioacoustic differences, slight mor- phological dissimilarities and substantial mitochondrial di- vergence (Vieites et al. 2009). This taxon is already known from Isalo and Tsingy de Bemaraha where it is typically encountered along slow-flowing streams in forested areas (Glaw and Vences 2007; Cocca et al. 2018). This record ex- tends the known distributional range of this taxon by ca. 180 km towards the east from the Isalo Massif. The species was rare, with only two individuals sampled at night tn Tsara- noro (at ca. 910 m a.s.l.), in the western part of the Region (Suppl. material 3: Table S3). They were spotted in a small pond below a large boulder. These two samples show no ge- netic differentiation to each other (Suppl. material 5: Table S5) and with samples from Isalo (e.g. KX066586). Mantidactylus (Chonomantis) delormei Angel, 1938 Fig. 3C This species is typically found along streams in montane forests between Ranomafana and the Andringitra Massif, which is also its type locality (Glaw and Vences 2007; Goodman et al. 2018). We sampled Mantidactylus delo- rmei at Asaramanitra, Belambo and Imaitso, along the eastern slopes of the Massif between ca. 1,570 and 1,710 m_a.s.l. Although a leaf litter-dwelling species, invididu- als were always in proximity to streams within rainforest (Suppl. material 3: Table S3). The animals were sampled during both day and night searches, although they were more frequent at night. The analysed individuals are al- most identical to each other (Suppl. material 5: Table S5) and to a previously available sequence from Andringi- tra (AY 848148) and 1.1% distant from individuals from Ranomafana (e.g. GU975171). Mantidactylus (Ochthomantis) femoralis (Boulenger, 1882) Fig. 3D The type locality of this taxon is “East Betsileo”, which roughly corresponds to the surveyed area and the lineage currently assigned to this name is known from Andringitra and the Isalo Massifs (Glaw and Vences 2007; Cocca et al. 2018; Goodman et al. 2018). This locally abundant species was recorded at Asaramanitra, Iantaranomby, Imaitso, Ri- andahy, Siranandambo, Anya, Fivahona—Velotsoa and Tsa- ranoro, at an elevation range between ca. 930 and 1,730 m a.s.l., both in the eastern and western portions of the study area, showing a similar distribution to 1 bourgati (Suppl. material 3: Table S3). The sampled individuals were ob- served along streams banks within forest and were collect- ed during both day and night-time, despite being more fre- quent at night. They are genetically identical to each other (Suppl. material 5: Table S5) and almost identical with pre- zse.pensoft.net Belluardo, F. et al.: Herpetological survey in Haute Matsiatra viously available sequences (e.g. Andringitra: HQ610918; Isalo: AY324813). Mantidactylus (Hylobatrachus) sp. Ca48 UCS Pig-3F Mantidactylus sp. Ca48 is morphologically similar to both Mantidactylus lugubris (Duméril, 1853) and Mantidacty- lus cowanii (Boulenger, 1882). This undescribed lineage is widely distributed amongst Isalo, Itremo, Antoetra (in sympatry with M. cowanii), Ranomafana, Ambohitsara, Vondrozo and Manombo (Cocca et al. 2018; Scherz et al. 2019). Goodman et al. (2018) report the possible presence of M. /ugubris in the species list of Andringitra National Park although the record is considered as doubt- ful. We found this taxon at Asaramanitra, Iantaranomby and Fivahona—Velotsoa, in both the eastern and western parts of the Region (between ca. 930 and 1,650 m a.s.l.) (Suppl. material 3: Table S3). The species seemed com- mon in Fivahona—Velotsoa, whereas, in the other sites, it was rarer. Individuals were found at night on rocks along streams. The analysed samples are genetically identical (Suppl. material 5: Table S5) and ca. 100% identical to individuals from Ranomafana (e.g. MK447667), Am- bohitsara (e.g. MK447637), Ambatolahy (MK447645), Valohoaka (MK447661), Miranony (MK447658) and Manombo (AY 848186). Spinomantis elegans (Guibé, 1974) Figes) Spinomantis elegans 1s distributed in south-eastern Mada- gascar between Ranomafana to the north and Andohahela to the south, including the area of the Andringitra Massif, which 1s close to the type locality (Ivohibe). This species lives at high elevations and is often found within small caves, between outcrops or hiding below rocks, both within forest and above the tree line (Glaw and Vences 2007). Spinomantis elegans was rarely encountered in our surveyes. We sampled a tadpole in a small pond at Imaitso and a subadult next to a large permanent stream at Asaramanitra, on the eastern slopes of the Massif at ca. 1,540-1,600 m a.s.l. of elevation (Suppl. material 3: Table S3). Both individuals were found at night. The two analysed samples are identical to each other (Suppl. mate- rial 5: Table S5) and previously available sequences (e.g. Cuvette Boby: AY659960; Ranomafana: AY 848405). Reptiles Family Chamaeleonidae Subfamily Brookesiinae Brookesia brunoi Crottini, Miralles, Glaw, Harris, Lima & Vences, 2012 Fig. 4A This species is currently known only from Anja Com- munity Reserve where it is typically encountered on Zoosyst. Evol. 97 (2) 2021, 315-343 327 Figure 4. Chameleonid species identified in this study. The picture of the individual identified as Furcifer willsii from Fivahona— Velotsoa, found within the stomach content of a Mimophis mahfalensis, is not shown. Sampling localities for each photographed individual are provided. ANP ES — Andringitra National Park Eastern Slopes; ANP WS — Andringitra National Park Western Slopes (Fig. 1; Suppl. material 1: Table S1). A. Brookesia brunoi from Anja; B. Calumma andringitraense from Imaitso (ANP ES); C. Calumma crypticum from Imaitso (ANP ES); D. Calumma fallax from Asaramanitra (ANP ES); E. Palleon nasus from Namoly; FE. Furcifer lateralis from lantaranomby (ANP WS); G. Furcifer major from Anja; H. Furcifer nicosiai from Tsaranoro; I. Furcifer oustaleti from Anja. Photographs by Javier Lobon-Rovira. the leaf litter during the day or roosting at a few cen- timetres height after the sunset, within the semi-arid deciduous forest patches of the Reserve (Crottini et al. 2012b). Brookesia brunoi was found only at Anja, at an elevation of ca. 970-980 m a.s.l. (Suppl. mate- rial 4: Table S4). The species seemed common in this forest fragment. All sampled individuals were found during the day moving on the leaf litter. Their sequenc- es are molecularly uniform (Suppl. material 6: Table S6, COI) and show a maximum genetic distance of ca. 1.50% from previously available sequences (e.g. ND2: JX101752). Palleon nasus (Boulenger, 1887) Fig. 4E Palleon nasus has originally been described from “Ekongo”, which probably refers to Ikongo, ca. 55 km north-east of the Andringitra Massif. This species is distributed in south-eastern Madagascar between Ranomafana and Andringitra to the north and Tolagna- ro to the south-east. Palleon nasus has been subdivided into two subspecies, based on morphological differ- ences: P. n. nasus (Boulenger, 1887) and P. n. pauliani (Brygoo, Blanc & Domergue, 1972), the latter being de- scribed from Manjarivolo (in the Andringitra Massif), where it was collected at an elevation of 1,620—1,650 m a.s.1. Based on morphological examination, Goodman (1996) identified P. n. nasus (specimens collected at an elevation of 720—1,630 m a.s.I.) during a herpetological assessment on the eastern slopes of the Massif. We col- lected six individuals of P. nasus in the eastern parts of the Region (Imaitso and Namoly), at an elevation range between ca. 1,580 and 1,640 m a.s.l. (Suppl. material 4: Table S4). The species was common. The animals were all found at night within rainforest while roosting on branches (one individual close to the floor and the other at ca. 2 m from the ground) and they were not active. The analysed samples are genetically uniform to each other (Suppl. material 6: Table S6) and they show 10.5% genetic distance to sequences obtained from individuals of Andohahela (COI: JQ909283) and are ca. 4% distant to an individual from Ranomafana (16S: HQ130509). Morphological examination of the collected individuals suggests a closer affinity with Pon. nasus. zse.pensoft.net 328 Subfamily Chamaeleoninae Calumma andringitraense (Brygoo, Blanc & Domer- gue, 1972) Fig. 4B This species 1s known from Andringitra, which represents the type locality, and the Andohahela Massif (Glaw and Vences 2007), where it inhabits montane rainforest at an elevation range of 1,550—1,680 m a.s.l. (Goodman 1996; Goodman et al. 2018). We collected samples of this spe- cies at Imaitso and Fivahona—Ambavanala, in the eastern part of the Region, while roosting on branches in dense rainforest at an elevation between ca. 1,480 and 1,560 m a.s.l. (Suppl. material 4: Table S4). They were all spotted at night in a sleeping position. The species was abundant in Imaitso, while rarer in Fivahona—Ambavanala. The an- alysed individuals are genetically uniform to each oth- er (Suppl. material 6: Table S6) and they have ca. 5.5% genetic distance to individuals from Andohahela (COI: JQ909303). Calumma crypticum Raxworthy & Nussbaum, 2006 Fig. 4C Calumma crypticum has a scattered distribution in- cluding the Tsaratanana Massif in the north and sever- al localities in the central highlands, Ranomafana and Andohahela. It inhabits montane forests at an elevation between ca. 1,050 and 1,850 m a.s.l. (Boumans et al. 2007; Glaw and Vences 2007; Randrianantoandro et al. 2010). Boumans et al. (2007) showed that the species is composed of several intra-specific lineages charac- terised by a certain degree of mitochondrial differenti- ation at the 16S gene. This species was the most com- mon chameleon we encountered during our sampling. We collected the species at Asaramanitra, Belambo, Imaitso, Fivahona—Velotsoa and Namoly, in the eastern portion of the Region (Suppl. material 4: Table S4). The sampled individuals were found both during the day and at night (sleeping) while roosting on branches in dense rainforest. They are genetically quite similar to each other (Suppl. material 6: Table S6) and show limited genetic differentiation compared to other samples col- lected in Ranomafana (COI: ca. 3%, JQ909308; 16S: ca. 0.50%, EF210643). Calumma fallax (Mocquard, 1900) Fig. 4D Following its new definition, Calumma fallax 1s distribut- ed in eastern Madagascar from Andohahela, to the south, to Mandraka to the north, where it can be found in rain- forest at low and middle elevations (Glaw and Vences 2007; Gehring et al. 2011, 2012; Prétzel et al. 2020). The type locality of this taxon is Ikongo, which is located ca. 55 km north-east of the Andringitra Massif. We sampled this species at Asaramanitra, Fivahona—Ambavanala, zse.pensoft.net Belluardo, F. et al.: Herpetological survey in Haute Matsiatra Fivahona—Velotsoa and Namoly in the rainforest of the eastern part of the surveyed area at an elevation range between ca. 1,490 and 1,670 m a.s.l. (Suppl. material 4: Table S4). The species was common in Namoly, while in the other sites, it was rarer. The animals were spotted at night on tree branches while sleeping. The analysed indi- viduals are molecularly uniform across the different sam- pling localities (Suppl. material 6: Table S6) and show a 4% genetic distance from individuals of the same species from Ranomafana (ND2: JQ734064). Furcifer lateralis (Gray, 1831) Fig. 4F Florio et al. (2012) revised the taxonomy of the F: /at- eralis complex, assigning the populations of southern and north-western Madagascar to Furcifer major (Bry- goo, 1971) and Furcifer viridis Florio, Ingram, Rako- tondravony, Louis Jr. & Raxworthy, 2012, respectively. The distribution of F: /ateralis was restricted to eastern Madagascar, where it can be found within rainforest, at forest edges, in shrubby grasslands and more degraded vegetational formations (Raselimanana and Rakotomala- la 2003; Glaw and Vences 2007). The species is reported from Andringitra (Goodman et al. 2018) and was sam- pled on the western slopes of the Massif (lakanga and Iantaranomby) where it was encountered with low fre- quency. Only two individuals were collected during the day, actively perching in open grassland with scattered trees at an elevation between ca. 900 and 1,560 m a.s.l. (Suppl. material 4: Table S4). The analysed samples are molecularly identical to each other (Suppl. material 6: Table S6) and show limited genetic differentiation (16S) from samples collected at Cirque Namoly (EF210582; within the study area), Vondrozo (EF210589) and Tampi- na forest (EF210593). Furcifer major (Brygoo, 1971) Fig. 4G Distributed in southern Madagascar, F? major inhab- its almost any arid habitat including human-disturbed environments (Raselimanana and Rakotomalala 2003; Florio et al. 2012). We sampled this species at Ambato- mainty, Anja, Anja—Ambilo and Sakaviro (Suppl. mate- rial 4: Table S4), on the western part of the Region at only ca. 10 km from the localities where we collected F. lateralis. Individuals were spotted during both day and night (sleeping), perching on tree branches. De- spite present in multiple sites, the species was local- ly rare to encounter. Relative to F Jateralis, F? major was found at a lower elevation (between ca. 930 and 1,030 m a.s.l.) and in semi-arid deciduous forest. The analysed samples are genetically identical to each other (Suppl. material 6: Table S6) and show 2.6% genetic distance (COI) from conspecific populations from Isalo (e.g. MH063344). Zoosyst. Evol. 97 (2) 2021, 315-343 Furcifer nicosiai Jesu, Mattioli & Schimmenti, 1999 Fig. 4H This chameleon was first described from Tsingy de Be- maraha, in western Madagascar, where it is mostly found within dense sub-humid and dry forests. This species has been later reported from Paysage Harmonieux Protégé de Beanka, Paysage Harmonieux Protégé du Complexe Tsi- membo Manambolomaty, Paysage Harmonieux Protégé du Complexe Lac—Forét Ambondrombe, Paysage Harmo- nieux Protégé de Menabe Antimena and Réserve Spéciale d’ Andranomena (Goodman et al. 2018). We sampled Fur- cifer nicosiai at Tsaranoro, in the western part of the re- gion, at ca. 960-970 maz.s.l. (Suppl. material 4: Table S4). This record was included in Belluardo et al. (2021), along with other new records from central and western Mada- gascar, significantly expanding the known distribution of this species by ca. 300 km towards the south-east. F’ nico- siai did not seem abundant. We sampled three individuals both during night (sleeping) and day while roosting on branches within semi-arid deciduous forest. The analysed individuals are genetically identical to each other (Suppl. material 6: Table S6) and they show 4% (COI: JQ909373) and 3% (16S: HF57045) genetic distance with individuals from the type locality (Tsingy de Bemaraha). Furcifer oustaleti (Mocquard, 1894) Clade D Fig. 41 This species 1s widely distributed in Madagascar. It can inhabit a wide variety of habitats, including degraded vegetation and human-dominated environments (Glaw and Vences 2007; Florio and Raxworthy 2016). We iden- tified this species at Ambalavao, Anja, Anja—Ambilo and Sakaviro (in the western part of the Region) at an eleva- tion range of ca. 980—1,010 m a.s.l. (Suppl. material 4: Table S4). The species seemed quite common. The indi- viduals were found during the day, both within semi-arid deciduous forest and in human-associated environments, actively perching on tree braches. The analysed animals are molecularly identical to each other (Suppl. material 6: Table S6) and to individuals from Isalo (COI: MH063345; 16S: MH063288) attributed to Clade D (sensu Florio and Raxworthy 2016), which is distributed in southern, cen- tral and north-western Madagascar. Furcifer willsii (Giinther, 1890) This chameleon is distributed in central-eastern Mada- gascar and in the north, in the area of Tsaratanana, while records from western Madagascar require verification (Glaw and Vences 2007). Furcifer willsii can be found in rainforest where it is usually spotted roosting high above the ground. The only recorded individual was sampled in Fivahona—Velotsoa, in the eastern part of the study area (ca. 1,270 maz.s.I.) and it was recovered in the stomach of a specimen of Mimophis mahfalensis (Grandidier, 1867) (ACZC11133) (Suppl. material 4: Table S4) (Lob6n-Ro- 329 vira et al. 2020). The previous southernmost record of this chameleon (Ikongo) is at ca. 55 km north-east from Fiva- hona—Velotsoa. The sample was included in Lobon-Ro- vira et al. (2020), contributing to extend the species dis- tributional range further south. The sample shows 7.2% genetic distance from a conspecific individual collected at Ranomafana (COI: JQ909382). Family Gekkonidae Subfamily Gekkoninae Hemidactylus mercatorius Gray, 1842 Fig. 5A This species 1s widely distributed in Madagascar, com- monly found in human-dominated areas (Glaw and Venc- es 2007). Hemidactylus mercatorius was found at Ian- taranomby, Ambalavao, Ambatomainty, Anja—Ambilo, Iarintsena, Sakaviro and Tsaranoro (Suppl. material 4: Table S4), in the western part of the Region (between ca. 930 and 1,580 m a.s.l.), mostly in human settlements and sometimes also within semi-arid deciduous forest. The individuals were normally found in nocturnal activity on rocks, despite one individual being sampled during the day in Ambalavao. The species was common in Tsaranoro. The analysed samples show limited genetic differentiation (Suppl. material 6: Table S6). To the best of our knowl- edge, the population from Isalo is genetically the closest to the samples analysed here (ca. 2.70%, MH063351). Lygodactylus pictus (Peters, 1883) Fig. 5B This diurnal gecko is known from south-eastern Mada- gascar and a few other localities in the central highlands. It is commonly found in degraded and secondary forest patches and in human-dominated environments. The Andringitra Massif and the surrounding areas are at the southern limit of the distributional range of the species (Puente et al. 2005; Glaw and Vences 2007). We sampled Lygodactylus pictus in degraded and secondary forest at Asaramanitra, Belambo, Fivahona—Ambavanala and Tsaranoro, both in the eastern and western parts of the surveyed region and in the town of Ambalavao at an ele- vational range between ca. 970 and 1,610 ma.s.l. (Suppl. material 4: Table S4). This gecko was mostly found in human-associated environments (e.g. roofs, houses) and only rarely on tree trunks. The individuals were mostly found active during the day, despite two being spotted at night, probably sleeping. The species was common in Be- lambo, while it seemed rarer in the other sites. The anal- ysed samples are genetically homogeneous (COI, Suppl. material 6: Table S6) and almost identical to an individual from Ambositra (JQ909452, COI). There is some genetic difference (in 16S) between the individuals collected in Tsaranoro and Fivahona—Ambavanala. The sample from Tsaranoro (ACZC10950) is ca. 1% distant (16S) from an zse.pensoft.net Belluardo, F. et al.: Herpetological survey in Haute Matsiatra Figure 5. Geckos species identified in this study. Sampling localities for each photographed individual are indicated. ANP ES — Andringitra National Park Eastern Slopes; ANP WS — Andringitra National Park Western Slopes (Fig. 1; Suppl. material 1: Table S1). A. Hemidactylus mercatorius from Tsaranoro; B. Lygodactylus pictus from Belambo (ANP ES); C. Lygodactylus sp. aff. pictus Ca01 “Isalo” from Ambatomainty; D. Paragehyra felicitae from Anja; E. Paragehyra sp. aff. felicitae “Tsaranoro” from Tsaranoro; F. Paroedura rennerae from Anja; G. Paroedura sp. aff. bastardi Lineage D from Anja; H. Phelsuma barbouri from Belambo (ANP ES); I. Phelsuma gouldi from Tsaranoro; J. Phelsuma lineata elanthana from Fivahona—Velotsoa. Photographs by Javier Lobon-Ro- vira (A—G, I—J) and Gongalo M. Rosa (H). individual from Ambositra (AY653269), while the sam- ple from Fivahona—Ambavanala (ACZC11175) is identi- cal (16S) with two individuals of L. pictus from Ambos- itra (GU593455) and Sendrisoa (AY653270), a locality only ca. 6 km away from Fivahona—Ambavanala. Lygodactylus sp. aff. pictus Ca01 “Isalo” UCS Fig. 5G We sampled a juvenile within the semi-arid deciduous forest of Ambatomainty (ca. 970 ma.s.1.) (Suppl. material 4: Table S4). The collected individual was found at night on arock along a trail. This sample shows limited genetic distance from an individual collected at Analalava forest in Isalo (16S: 2.7%, AY653238; COI: 4.50%, JQ909445). The new record significantly extends the known distribu- tional range of this taxon by ca. 180 km towards the east from the Isalo Massif. Paragehyra felicitae Crottini, Harris, Miralles, Glaw, Jenkins, Randrianantoandro, Bauer & Vences, 2015 Fig. 5D This gecko was known only from Anja Community Re- serve and from a site a few km away from Anja. This species can be observed on granitic boulders associat- ed with both semi-arid deciduous forest and grasslands. Even though the other geckos of the genus Paragehyra zse.pensoft.net are nocturnal, these animals can also be spotted during the day (Crottini et al. 2015). We sampled this species in Anja and Sakaviro, the latter record representing a distributional range extension (ca. 8 km). The collected samples were active at night on granitic boulders and rocks within semi-arid deciduous forest, sometimes in clearings within the forest, at ca. 950-990 m a.s.l. (Suppl. material 4: Table S4). The species was present in high densities in both sites. Collected samples show some genetic differentiation at the COI fragment (Suppl. material 6: Table S6) and they are almost identical (16S) to individuals from Anja (e.g. KP025816). Paragehyra sp. aff. felicitae “Tsaranoro” CCS Fig. 5E This taxon was collected at Iantaranomby, Ambatomain- ty and Tsaranoro, all localities in the western part of the surveyed region (between ca. 910 and 1,610 m as.l.) (Suppl. material 4: Table S4). Animals were found active at night on granitic boulders both in open habitats and semi-arid deciduous forest. The species was particularly abundant in Tsaranoro and, to a less extent, in Ambato- mainty, whereas only one individual was sampled in Iantaranomby. The analysed samples show limited ge- netic differentiation amongst sampled localities (Suppl. material 6: Table S6). The genetic distance between this taxon and P. felicitae sampled in Anja is ca. 15% (COI: Zoosyst. Evol. 97 (2) 2021, 315-343 ACZC10432) and 7.6% (16S: KP025811), while it has 17% genetic difference with Paragehyra petiti Angel, 1929 (COI: JQ909497). This taxon is morphologically similar to P. felicitae. However, the analysis of the col- lected specimens highlighted the presence of a distinct number of longitudinal rows of enlarged turbercles on the dorsolateral surfaces of the body relative to the sister species P. felicitae (which is a morphological diagnostic character of this species), confirming its distinction also at the morphological level and determining the status of confirmed candidate species. The description of this can- didate species is currently in progress. Paroedura rennerae Miralles, Bruy, Crottini, Rako- toarison, Ratsoavina, Scherz, Schmidt, Kéhler, Glaw & Vences, 2021 Fig. SF The Paroedura bastardi (Mocquard, 1900) species com- plex has been recently revised (Miralles et al. 2021). This work identified at least three evolutionary lineages with- in this species group. Following this finding, the authors propose a new definition for P. bastardi sensu stricto. They resurrected the binomen Paroedura guibeae Dixon & Kroll, 1974 and formally described a third lineage as P. rennerae. The latter species is currently known from Miandrivazo, Kirindy, Marofandilia, Anja and Isalo. We sampled this gecko at Ambatomainty, Anja, Sakaviro and Tsaranoro, all sites in the western part of the surveyed region (at ca. 930-990 m a.s.1.) (Suppl. material 4: Table S4), extending the known distribution of this species by ca. 25 km towards the south from Anja. This species was abundant in all visited sites. The animals were always found active at night on boulders, mostly within semi-ar- id deciduous forest, but sometimes also at the edge of the forest and in human settlements. The collected sam- ples are molecularly uniform (Suppl. material 6: Table S6) and almost identical to an individual collected from Anja (COI: MG734947). They are slightly differentiated from conspecific populations from other localities (COI: 2.60%, MG734948, Kirindy; 16S: 1.4%, GU128989, Miandrivazo; 1.8%, GU129005, Marofandilia). Paroedura sp. aff. bastardi Lineage D UCS Fig. 5G Individuals belonging to this lineage of the P. bastardi species complex were previously known only from Anja (Miralles et al. 2021). We collected this taxon at Anja and Tsaranoro, in the western part of the Region (at ca. 930- 970 maz.s.l.), extending the distributional range by ca. 25 km towards the south (Suppl. material 4: Table S4). In both sites, this lineage was found in syntopy with Paroed- ura rennerae, which is very similar in morphology, and in high densities The individuals were found active at night on boulders within semi-arid deciduous forest and show a limited degree of genetic differentiation (Suppl. mate- 33:1 rial 6: Table S6). The collected samples are almost 100% identical to ZCMV 12790 (COI: MW311368), which is the only individual molecularly characterised for this tax- on before our study (Miralles et al. 2021). Phelsuma barbouri Loveridge, 1942 Pig. SE: This diurnal gecko is known from a few montane areas on the central highlands and in south-eastern Madagas- car. The area of Andringitra represents the southern limit of its distributional range. Unlike most Phelsuma geck- os, which are arboreal, this species is normally found in montane rocky habitats on the ground and boulders (Glaw and Vences 2007; Goodman et al. 2018). We sam- pled a single individual of Phelsuma barbouri active during the day on a small wooden bridge at Belambo in the eastern slopes of the Massif, at ca. 1,570 m as.l. (Suppl. material 4: Table S4). This sample is 96% similar to a conspecific individual from Tsiafajavona (Ankaratra) (COI: JQ909518). Phelsuma gouldi Crottini, Gehring, Glaw, Harris, Lima & Vences, 2011 Fig. 5] This species was reliably known only from Anja, where a single individual (the holotype) was spotted on a trunk within the forest patch of the Reserve (Crottini et al. 2011b). The species is confirmed to be quite rare. Two in- dividuals were sampled at night on lianas in the semi-ar- id deciduous forest of Tsaranoro, in the western part of the Region, at an elevation range of ca. 910-950 m a.s.l. (Suppl. material 4: Table S4). This finding extends the known distributional range of the species by ca. 25 km to the south. The two analysed samples are identical to each other (Suppl. material 6: Table S6) and identical to the holotype (16S: JF810252). Phelsuma lineata elanthana Kriiger, 1996 Fig. 5J Phelsuma lineata elanthana is distributed in the central highlands and the northern part of central-eastern Mad- agascar (Boumans et al. 2007; Gehring et al. 2013). It inhabits a great variety of habitats, from the rainforest to bushes in more arid areas and it is also commonly found in human settlements (Glaw and Vences 2007). This gecko was rare. We sampled P. /ineata only in Fivahona— Velotsoa, in the eastern part of the Region (at ca. 1,290 m_a.s.l.), where the animals were found active during the day on Pandanus trees within the rainforest (Suppl. ma- terial 4: Table S4). This record represents a range exten- sion for this taxon by ca. 350 km towards the south. The analysed samples show a 3% genetic distance (16S) from samples from Andasibe (EF210615) and Ambohitantely (EF210617). zse.pensoft.net 832 Family Gerrhosauridae Subfamily Zonosaurinae Zonosaurus aeneus (Grandidier, 1872) Fig. 6E This species is distributed at a wide latitudinal range be- tween the central highlands and the eastern and south-east- ern escarpment. It is typically found on the edges of rain- forest or in open areas within forest (Glaw and Vences 2007). Zonosaurus aeneus was previously reported from Andringitra (Goodman et al. 2018) and was found at Fi- vahona—Ambavanala and Namoly, in the eastern part of the studied region (between ca. 1,480 and 1,650 m a.s.l.). Zonosaurus aeneus was sampled during the day. The ani- mals were active on the ground in grassy clearings inside rainforest and in open areas immediately next to rainfor- est patches. One individual was found next to a ricefield (Suppl. material 4: Table S4). The analysed individuals are identical (Suppl. material 6: Table S6) and they show a 3.3% genetic distance from a sequenced individual from Ranomafana (COI: JQ909624) anda 1.5% genetic distance from individuals from Torotorofotsy (16S: KC515131). Zonosaurus laticaudatus (Grandidier, 1869) Fig. 6C This large plated lizard lives over a wide latitudinal range throughout western Madagascar (Glaw and Vences 2007). This species is reported from dry forest, rocky open ar- eas, degraded and human-disturbed habitats (Glaw and Vences 2007; Recknagel et al. 2013). This taxon was not previously reported from the area of Andringitra and was collected at Ambatomainty, Anja and Tsaranoro, all lo- calities in the western part of the Region (between ca. 870 and 960 m a.s.l.) (Suppl. material 4: Table S4). This record extends the known distributional range of the species by ca. 180 km to the East. The individuals were found both in open areas on rocky substrate and on large boulders next to semi-arid deciduous forest patches. They were active during the day. Analysed samples show 2.2% genetic distance from the population of Isalo (e.g. COI: MH063372) and 1% genetic distance from the population of Hazofotsy, close to Tolagnaro (16S: AY 167372). Zonosaurus ornatus (Gray, 1831) Fig. 6D This species is found in central-eastern Madagascar where it inhabits open habitats and forest edges at a wide eleva- tional range. In Ankaratra and Andringitra, this species is found in montane savannah and heathlands above the tree line (Glaw and Vences 2007; Goodman et al. 2018). We sampled Zonosaurus ornatus at Antanifotsy 2, Fiva- hona—Ambavanala and Namoly, in the eastern part of the Region (between ca. 1,450 and 1,650 maz.s.1.) in open en- vironments next to rainforest, often in human-disturbed areas (Suppl. material 4: Table S4). One of the individu- zse.pensoft.net Belluardo, F. et al.: Herpetological survey in Haute Matsiatra als was spotted next to a ricefield. The animals were all active during the day. The analysed samples are identical (Suppl. material 6: Table S6) and show a 2.3% genetic distance from individuals from Ambatolahy (e.g. COI: JQ909633). However, the population from Ambatolahy is almost identical at the 16S marker (e.g. KC515145). Family Opluridae Oplurus grandidieri Mocquard, 1900 Fig. 6A This species is distributed in the south-central part of the central highlands where it is often found in rocky environ- ments (Glaw and Vences 2007). We found this species at Andramena, Iantaranomby, Siranandambo, Ambatomain- ty, Anja, Sakaviro and Tsaranoro in the western portion of the studied area (Suppl. material 4: Table S4). The animals were normally found in high densities on boul- ders and outcrops at an elevation range between ca. 930 and 1,740 m a.s.l. They were all active during the day. The analysed samples show a limited degree of genetic differentiation (Suppl. material 6: Table S6) and low ge- netic distance to the population from Isalo (COI: 3.5%, MH063380; 16S: 3.2%, MH063315). In Iantaranomby, two adult males were observed displaying an aggressive interaction on a sunny boulder (Lobon-Rovira et al. 2019). Oplurus quadrimaculatus Duméril & Bibron, 1851 Fig. 6B Oplurus quadrimaculatus is distributed in the south and the south-eastern part of the central highlands, in- cluding the area of Andringitra (Glaw and Vences 2007; Munchenberg et al. 2008). It was also recently identified at Isalo (Cocca et al. 2018). Oplurus quadrimaculatus is a saxicolous species, normally observed on large boulders within arid environments and, in some cases, also next to forest patches. We found this species at Andramena, Asaramanitra, Imaitso, Ambatomainty, Anja, Fivaho- na—Velotsoa, Sakaviro and Tsaranoro in both the eastern and western parts of the surveyed area (Suppl. material 4: Table S4). The animals were present in high densities and active during the day on large boulders and outcrops both in open and forested areas at an elevation between ca. 870 and 1,740 m a.s.l. Within forest, they were nor- mally spotted in rocky clearings or next to large streams delimited by outcrops. Oplurus quadrimaculatus was sometimes found in syntopy with O. grandidieri and, in some cases, the two species shared the same rocks. Anal- ysed samples show limited genetic differentiation among them (Suppl. material 6: Table S6) and in relation to other individuals collected in the Andringitra Region (e.g. 16S: EU099752). They show a 4.4% genetic distance from the population from Andohahela (COI: JQ909486) and no differentiation from the population from Antoetra (e.g. 16S: EU099737) and Ambositra (16S: EU099742). Zoosyst. Evol. 97 (2) 2021, 315-343 Figure 6. Scincid, gerrhosaurid and oplurid species identified in this study. Sampling localities for each photographed individual are provided. ANP ES — Andringitra National Park Eastern Slopes; ANP WS — Andringitra National Park Western Slopes (Fig. 1; Suppl. material 1: Table $1). A. Oplurus grandidieri from Anja; B. Oplurus quadrimaculatus from Anja; C. Zonosaurus laticau- datus from Anja; D. Zonosaurus ornatus from Antanifotsy 2; E. Zonosaurus aeneus from Namoly; F. Trachylepis elegans from Iantaranomby (ANP WS); G. Trachylepis gravenhorstii from Anja; H. Trachylepis sp. aff. vato from Asaramanitra (ANP ES); I. Trachylepis boettgeri from Antanifotsy 3. Photographs by Javier Lobén-Rovira. Family Scincidae Subfamily Mabuyinae Trachylepis boettgeri (Boulenger, 1887) Fig. 61 Trachylepis boettgeri is found in central-eastern Mada- gascar where it inhabits the open habitats of the central highlands, including the Region of Andringitra (Goodman et al. 2018). We sampled one individual at Antanifotsy 3, in an area dominated by grasslands and rice fields in the eastern part of the Region at ca. 1,440 ma.s.l. (Suppl. ma- terial 4: Table S4). The animal was active on the ground during the day. The analysed sample is genetically almost identical to a population from Ankaratra (COI: JQ909591; 16S: DQ238879) and Ambatolampy (16S: AY070355). Trachylepis elegans (Peters, 1854) Lineage A Fig. 6F This skink is distributed throughout Madagascar, except for the eastern escarpment (Glaw and Vences 2007). It mostly inhabits open habitats in both dry and humid environments and can be found in cities and other hu- man-dominated environments (Glaw and Vences 2007; Vences et al. 2014). We found this taxon in high densities at Iantaranomby, Ambalavao, Ambatomainty, Anja—Am- bilo and Fivahona—Velotsoa (in the western part of the surveyed region) and in Antanifotsy 4 (in the eastern) be- tween ca. 830 and 1,670 m a.s.l. (Suppl. material 4: Ta- ble S4). All individuals were found in grassy open areas, sometimes next to small boulders, either close to forest patches or in human-dominated areas. They were found active during the day. The analysed samples show a lim- ited degree of genetic differentiation (Suppl. material 6: Table S6) and they show a 2% genetic distance from the population of Isalo (e.g. COI: KF250670) attributed to lineage A (sensu Vences et al. 2014), which is the most widespread lineage in Madagascar. Trachylepis gravenhorstii (Duméril & Bibron, 1839) Lineage 4A Fig. 6G Trachylepis gravenhorstii is found almost everywhere in Madagascar up to 1,400 m a.s.l. (Glaw and Vences 2007; Vences et al. 2014; Goodman et al. 2018). We sampled this taxon at Belambo, Iantaranomby, Riandahy and Anja, in both the eastern and western part of the Region at an elevation between ca. 960 and 1,640 m a.s.l. (Suppl. ma- terial 4: Table S4). Trachylepis gravenhorstii was often found in syntopy with 7: elegans, although less common than the latter species. Where 7: elegans was not pres- ent, 7? gravenhorstii was found in less human-disturbed habitats. The individuals were sampled during the day, both in grassy open areas with presence of small boulders and within forest active on the ground. The analysed sam- ples show some degree of intra-populational variability zse.pensoft.net 334 (Suppl. material 6: Table S6) and are identical to a sample collected at Ranomafana (COI: KF250703) attributed to lineage 4A (sensu Vences et al. 2014) distributed in cen- tral-eastern Madagascar. Previously available data of this species from the area (Ambalavao) belong to the lineage 4B (COI: KF250708). As previously reported for Isalo (Cocca et al. 2018), it seems that these two lineages occur sympatrically in the surveyed area. Trachylepis sp. aff. vato UCS Fig. 6H Trachylepis vato (Nussbaum & Raxworthy, 1994) is dis- tributed in central and southern Madagascar where it in- habits arid environments with boulders and rocks (Glaw and Vences 2007). It was described from a locality close to Andohahela (Type locality: Mananara River between Bev- ia and Hazofotsy, 24°51.00'S, 46°31.00'E), in south-east- ern Madagascar (Nussbaum and Raxworthy 1994). We collected this lineage at Asaramanitra, Iantaranomby, Anja, Fivahona—Velotsoa, Iarintsena and Sakaviro, both in the eastern and western parts of the surveyed region (be- tween ca. 990 and 1,660 ma.s.1.), mostly on boulders, both in open environments and within forest (Suppl. material 4: Table S4). The animals were all active during the day. This skink was not common in the sampling sites we visited. These records represent a slight range extension within the area of Andringitra. Lima et al. (2013) assigned individu- als from Ambalavao (16S: KC345435; ND1: KC345095), Andringitra (16S: KC345394; ND1: KC345053) and Col des Tapias (16S: KC345432; ND1: KC345092) to a can- didate species 7! cf. vato. Our samples show some sign of genetic differentiation (Suppl. material 6: Table S6) and the genetic distance between our samples and the samples of 7: cf. vato included in Lima et al. (2013) is between 1% and 2.3% (ND1: KC345092, KC345053, KC345095). Our samples are also almost identical to individuals from Ibity (16S: AY159097), close to Col des Tapias. Family Psammophiidae Mimophis mahfalensis (Grandidier, 1867) Fig. 7G This snake 1s widely distributed across almost all of the southern half of Madagascar, where it can be found in rainforest, dry forest, arid spiny thornbush savannah and human-dominated areas (Glaw and Vences 2007; Ruane et al. 2017). We sampled this species at lakanga, Anja, Fivahona—Velotsoa and Sakaviro, both in the western and eastern parts of the surveyed area, at an elevation between ca. 900 and 1,270 ma.s.l. (Suppl. material 4: Table S4). The animals were found during the day active on the ground in a wide variety of habitats: rainforest, semi-ar- id deciduous forest and grassland. The analysed samples are molecularly similar to each other (Suppl. material 6: Table S6) and 0.8% distant (COI) to the population from zse.pensoft.net Belluardo, F. et al.: Herpetological survey in Haute Matsiatra Ibity (JQ909481) and Isalo (MH063403). The individ- ual collected at Fivahona—Velotsoa was found with a Furcifer willsii chameleon (ACZC11200) in its stomach (Lobon-Rovira et al. 2020). Family Pseudoxyrhophiidae Compsophis infralineatus (Ginther, 1882) Fig. 7A This semi-arboreal snake is distributed in eastern and south-eastern Madagascar where it is normally found along ponds and small streams (Glaw and Vences 2007). Comp- sophis infralineatus was already reported from Andringitra (Goodman et al. 2018). Two individuals were found at night in Fivahona—Velotsoa and Namoly, in the eastern part of the Region, within the rainforest at an elevational range of ca. 1,270—1,640 m a.s.l. (Suppl. material 4: Table S4). In Fiva- hona—Velotsoa, this snake was found active on the ground along the banks of a small stream. The analysed individuals are genetically identical amongst each other (Suppl. mate- rial 6: Table S6) and they are ca. 1% distant (COI) from the population from Manjakatompo (e.g. JQ909355). Leioheterodon modestus (Giinther, 1863) Fig. 7B This snake has a patchy distribution in central, western and southern Madagascar, where it is generally found in dry areas, both inside and outside the forest and in anthropo- genic environments (Glaw and Vences 2007). This species has not yet been reported from the study area and we found it at Antanifotsy 1, Fivahona—Ambavanala and Fivahona— Velotsoa in the eastern part of the Region, contributing to the extension of its known distributional range by ca. 200 km towards the east. Two individuals were spotted during the day active on the ground close to rice fields and vil- lages near forest patches at an elevational range between ca. 1,280 and 1,460 ma.s.l. (Suppl. material 4: Table S4). The analysed samples are genetically identical to each oth- er (Suppl. material 6: Table S6) and 0.60 % distant to an individual collected at Zazafotsy (COI: MH063415). Liophidium torquatum (Boulenger, 1888) Fig. 7F This terrestrial snake is distributed in eastern and north- ern Madagascar where it mostly inhabits rainforest, de- spite being also found in dry deciduous forest (Glaw and Vences 2007). This species is reported from Andringitra (Goodman et al. 2018) and we sampled a single individu- al inside the semi-arid deciduous forest fragment of Anja, in the western part of the surveyed region (at ca. 990 m a.s.1.) (Suppl. material 4: Table S4). The animal was ac- tive during the day. This sample is almost 100% identi- cal to the population of L. torquatum from Ranomafana (Cytb: DQ979984). Zoosyst. Evol. 97 (2) 2021, 315-343 $35 cds - a. : po ae i 5 Ps “ty . - om Se be x Pie surat ‘ k= Figure 7. Psammophiid, pseudoxyrhophiid and sanziniid snakes identified in this study. Sampling localities for each photographed hes individual are provided. ANP ES — Andringitra National Park Eastern Slopes; ANP WS — Andringitra National Park Western Slopes (Fig. 1; Suppl. material 1: Table S1). A. Compsophis infralineatus from Namoly; B. Leioheterodon modestus from Antanifotsy 1; C. Madagascarophis meridionalis from Anja; D. Thamnosophis lateralis from Anja; E. Pseudoxyrhopus sp. Ca2 from Ambatomainty; F. Liophidium torquatum from Anja G. Mimophis mahfalensis from Sakaviro; H. Sanzinia cf. volontany from Anja; I. Acrantophis dumerili from Sakaviro. Photographs by Javier Lobon-Rovira (A-E, G, I), Goncgalo M. Rosa (F) and Franco Andreone (H). Madagascarophis meridionalis Domergue, 1987 Fig. 7C This species inhabits arid environments in southern and south-western Madagascar (Glaw and Vences 2007). Madagascarophis meridionalis is known to inhabit the Andringitra Region (Nagy et al. 2007) and was sampled at Ambatomainty, Anja, Iantaranomby and Tsaranoro, in the western part of the Region at an elevation range between ca. 930 and 1,580 m a.s.l. (Suppl. material 4: Table S4). The animals were mostly found active on the ground in open areas next to semi-arid deciduous forest, sometimes along streams and in human-disturbed envi- ronments. They were mostly spotted at night, although one individual was found during the day. The analysed samples are genetically uniform (Suppl. material 6: Ta- ble S6). They are identical to a published sequence from the area (16S: AY586213) and almost identical to the population from Antoetra (16S: AY586212). They are 1% distant (COI) from the population from Andranovo- rivato (KU925345). Pseudoxyrhopus sp. Ca2 UCS Fisv7E This taxon was sampled in Ambatomainty, in the western part of the Region (at ca. 960 m a.s.l.) (Suppl. material 4: Table S4). The only collected individual was moving on the ground at night in an open area with a few scattered trees next to a small fragment of semi-arid deciduous forest. This specimen is genetically very similar (2.7% distance at COI) to a candidate species collected at Zom- bitse—Vohibasia National Park (RAN 43545-UMMZ 203648, Burbrink et al. 2019). Our finding extends the known distribution of this taxon by ca. 230 km to the north-east. Thamnosophis lateralis (Duméril, Bibron & Duméril, 1854) Fig. 7D This is one of Madagascar’s most common snakes (Glaw and Vences 2007), being mostly found outside dense forest and often in degraded areas (Vences 2011). We zse.pensoft.net 336 sampled this species at Antanifotsy 3, Anja, Tsaranoro, Fivahona—Ambavanala and Fivahona—Velotsoa in both the western and eastern parts of the surveyed region at an elevation between ca. 930 and 1,480 m a.s.l. (Suppl. material 4: Table S4). We found this species active on the ground during the day, both within forest and in an- thropogenic environments (next to ricefields). Analysed samples are genetically uniform (Suppl. material 6: Ta- ble S6) and show 1% distance from the population from Isalo (COI: MH063410). Family Sanziniidae Acrantophis dumerili Jan, 1860 Fig. 7] Acrantophis dumerili is distributed in central and south- ern Madagascar where it inhabits dry forest, savannah, as well as open and cultivated areas (Glaw and Vences 2007). Acrantophis dumerili is reported from Ambalavao (Glaw and Vences 2007) and was sampled at Sakaviro and Vidia, in the western part of the surveyed region (at ca. 850-990 m a.s.1.) (Suppl. material 4: Table S4). While the specimen sampled in Sakaviro was found after dusk active on the ground at the edge of semi-arid deciduous forest, the other was found dead on the Route Nationale 7. The samples are identical to each other (Suppl. mate- rial 6: Table S6) and to an individual from an imprecise sampling locality (COI: JQ909244, “300 km from Tana’) and the population from Ambositra (16S: AY336072). They are also almost identical to the population from Isa- lo (16S: EU419793). Sanzinia cf. volontany Vences & Glaw, 2004 Fig. 7H Following a recent taxonomic revision, the genus San- zinia comprises two species, Sanzinia volontany and S. madagascariensis (Dumeéril & Bibron, 1844) (Reynolds et al. 2014). The two species are genetically divergent and show some degree of morphological differentiation in colouration and pholidosis (Vences and Glaw 2003; Orozco-terWengel et al. 2008; Reynolds et al. 2014). They are divergently distributed, with S. madagascar- iensis occurring in eastern Madagascar and S. volontany throughout the west. An individual of Sanzinia sp. was photographed at night in Anja while moving on the flo- rest floor, in the western part of the Region, but no tis- sues were collected from that specimen (Suppl. material 4: Table S4). The analysis of the photographic material would lead to assign the individual to S. volontany, al- though the record requires further confirmation. Sanzin- ia madagascariensis is known from Ivohibe (Glaw and Vences 2007), whereas S. volontany from Isalo (Oroz- co-terWengel et al. 2008). Given the geographic proxim- ity of these records and the presence of both rain and dry zse.pensoft.net Belluardo, F. et al.: Herpetological survey in Haute Matsiatra forests in the Region, there is the possibility that both species inhabit this area. Discussion We provided the first list and barcoding reference da- tabase for 28 amphibians and 38 reptiles of the area surrounding the Andringitra Massif and extended the known distributional range of nine amphibians and twelve reptiles. Species composition is probably in- fluenced by the environmental diversity of the Region (Goodman 1996; Goodman et al. 2018). Besides the several microendemics (Boophis laurenti, Mantidacty- lus bourgati, Brookesia brunoi, Paragehyra felicitae, Paragehyra sp. aff. felicitae “Tsaranoro”, Paroedura sp. aff. bastardi Lineage D and Phelsuma gouldi), many taxa are distributed only in the eastern part dominated by rainforest (e.g. Boophis ankaratra, B. boppa, B. majori, B. popi, B. rhodoscelis, Aglyptodactylus madagascar- iensis, Gephyromantis blanci, Mantidactylus delormei, Spinomantis elegans, Calumma crypticum, Phelsuma lineata, Zonosaurus ornatus and Compsophis infralin- eatus), while others are only present in the western dry habitats (e.g. Laliostoma labrosum, Mantella betsileo, Scaphiophryne_ calcarata, Heterixalus luteostriatus, Furcifer major and Madagascarophis meridionalis) (Glaw and Vences 2007). Surveyed sites are highly fragmented and embedded within a matrix of anthropogenically-modified land- scape. Despite the likely loss in species richness and the alteration of species composition, forest fragments retain high levels of diversity in Madagascar (Crottin1 et al. 2011la; Durkin et al. 2011; Jenkins et al. 2014; Rie- mann et al. 2015). Such diversity can be irreplaceable when it includes microendemics. These species are reli- ably known only from a few geographically close local- ities, therefore fragments alteration and destruction may lead to their extinction. As already reported by Jenkins et al. (2014), Anja Community Reserve stands out at the national level for its multiple microendemic taxa and, in our sampling, Anja resulted as the most species-rich (24 species) amongst the surveyed fragments (Fig. 1A; Sup- pl. material 9: Table S7). Meaningful comparisons of the number of recorded species amongst sampling sites are hampered by the non-standardised sampling effort and the limited time spent in each locality, which was probably not enough to sample the actual total diversity. However, we can still notice an expected positive re- lationship between fragment area and species richness. We sampled 21 and 19 species in Tsaranoro and Fiva- hona—Velotsoa (Fig. 1G), respectively, the two largest fragments along with Anja. In the smallest patches, we found a lower number of species. In both Sakaviro and Ambatomainty (Fig. 1B and C), we found 14 species and, in Fivahona—Ambavanala, we recorded 12 species. Beyond species numbers, it is interesting to note that even the smallest fragments could host taxa that were Zoosyst. Evol. 97 (2) 2021, 315-343 not detected in any other locality, some of which repre- sent relevant range extensions. Ambatomainty is prob- ably the most interesting example in this sense. In this highly degraded forest of only two hectares (Fig. 1C), we recorded two candidate new species: Lygodactylus sp. aff. pictus Ca0l1 “Isalo” and Pseudoxyrhopus sp. Ca2, to date reliably known only from a few other sites some hundreds of kilometres away (i.e. Isalo and Zom- bitse—Vohibasia National Parks, respectively). Deforestation and habitat fragmentation are more pro- nounced in the western part of the surveyed region where, even within the National Park borders, the forest cover is reduced (Goodman 1996). The taxa that inhabit these fragments may not find large forests at a similar eleva- tion within the Park and may consequently lack available legally protected habitat. These small forest fragments can thus play a fundamental role as refugia to the local herpetofauna. Their conservation should, therefore, be prioritised for the long-term survival of their unique her- petological diversity and, more in general, for the conser- vation of the biodiversity of the entire Region. Finally, the improved knowledge on the species distribution of the candidate taxa, identified in this study, will now likely enable their formal description (e.g. in the case of Par- agehyra sp. aff. felicitae “Tsaranoro” and Paroedura sp. aff. bastardi Lineage D). Conclusions In a country plagued by centuries of forest loss and fragmentation (Hornac 1943; Jarosz 1993; Vieilledent et al. 2018), species inventories of remnant forest frag- ments are of paramount importance to achieve a better understanding of Malagasy biodiversity. We highlight- ed the herpetological significance of the small forest patches surrounding the Andringitra Massif, where we identified several taxa that were previously unknown from this area and, in several instances, we contribut- ed to the extension of their known distributional ranges by hundreds of kilometres (e.g. Belluardo et al. 2021). Many of these taxa are candidate new species and the newly-collected specimens will enable future taxonom- ic evaluations and descriptions. We also identified one candidate species previously unknown to science and provided a better characterisation of the distribution of several microendemic species that inhabit the study area. We generated a first barcoding reference data- base for this area that will facilitate future systematic research, both at the regional and country level. These results emphasise the relevance of the Region of An- dringitra in terms of microendemic diversity hosted in highly altered habitats. Apart from three private re- serves managed by local communities, and despite their herpetological value, the other investigated fragments are not officially protected. Granting some legal pro- tection to these sites 1s highly desirable to warrant the conservation of this unique biodiversity. Say Acknowledgements This work was funded by the National Geographic So- ciety (grant number EC—50656R-18 to FB). We thank Malagasy authorities, in particular the Ministere de I’ En- vironment, de l’Ecologie et des Forests (now Ministere l’ Environnement et du Développement Durable) and the Direction Regionale de |’Environment, de |’Ecologie et des Forests Haute Matsiatra, for providing research, trans- port and export permits (222/18/MEEF/SG/DGF/DSAP/ SCB.Re; 1596/18/MEEF/SG/DREEF.HM; 390N—EA 12/ MG18; 529C—EA08/MG19). We also thank MICET for their logistic support. We are grateful to the several lo- cal guides who provided their invaluable support during fieldwork: Meja, Boba Joseph, Francky Rasolondraibe, Anycet Rambonivoatra, Arsene Ravelomantazafy, Nidra Rasolomamopia, Josef Samuel, Ratsia, Mobadoda and Jakob Jules. We thank Walter Cocca for his help in data analysis and Jesper Ostlund for useful discussions. We also thank reviewers for their helpful suggestions. 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Rosa, Malalatiana Rasoazanany, Franco Andreone, Angelica Crottini Data type: sampling sites coordinates Copyright notice: This dataset is made available under the Open Database License (http://opendatacommons. org/licenses/odbl/1.0/). The Open Database License (ODbL) is a license agreement intended to allow us- ers to freely share, modify, and use this Dataset while maintaining this same freedom for others, provided that the original source and author(s) are credited. Link: https://do1.org/10.3897/zse.97.63936.suppl1 Supplementary material 2 Table S2. Amplified genes, primers and PCR conditions used in this study Authors: Francesco Belluardo, Darwin Diaz Quiros, Javier Lobon-Rovira, Gon¢galo M. Rosa, Malalatiana Rasoazanany, Franco Andreone, Angelica Crottini Data type: Primers and PCR conditions Copyright notice: This dataset is made available under the Open Database License (http://opendatacommons. org/licenses/odbl/1.0/). The Open Database License (ODbL) is a license agreement intended to allow us- ers to freely share, modify, and use this Dataset while maintaining this same freedom for others, provided that the original source and author(s) are credited. Link: https://do1.org/10.3897/zse.97.63936.suppl2 Supplementary material 3 Table S3. Amphibian samples identified in this study Authors: Francesco Belluardo, Darwin Diaz Quiros, Javier Lobon-Rovira, Gon¢galo M. Rosa, Malalatiana Rasoazanany, Franco Andreone, Angelica Crottini Data type: occurrences and species identification Copyright notice: This dataset is made available under the Open Database License (http://opendatacommons. org/licenses/odbl/1.0/). The Open Database License (ODbL) is a license agreement intended to allow us- ers to freely share, modify, and use this Dataset while maintaining this same freedom for others, provided that the original source and author(s) are credited. Link: https://do1.org/10.3897/zse.97.63936.suppl3 zse.pensoft.net Belluardo, F. et al.: Herpetological survey in Haute Matsiatra Supplementary material 4 Table S4. Reptile samples identified in this study Authors: Francesco Belluardo, Darwin Diaz Quiros, Javier Lobon-Rovira, Goncgalo M. Rosa, Malalatiana Rasoazanany, Franco Andreone, Angelica Crottini Data type: occurrences and species identification Copyright notice: This dataset is made available under the Open Database License (http://opendatacommons. org/licenses/odbl/1.0/). The Open Database License (ODbL) is a license agreement intended to allow us- ers to freely share, modify, and use this Dataset while maintaining this same freedom for others, provided that the original source and author(s) are credited. Link: https://doi.org/10.3897/zse.97.63936.suppl4 Supplementary material 5 Table SS. Within taxa uncorrected p—distances (16S) of amphibian taxa identified in this study Authors: Francesco Belluardo, Darwin Diaz Quiros, Javier Lobon-Rovira, Gon¢galo M. Rosa, Malalatiana Rasoazanany, Franco Andreone, Angelica Crottini Data type: genetic distances Copyright notice: This dataset is made available under the Open Database License (http://opendatacommons. org/licenses/odbl/1.0/). The Open Database License (ODbL) is a license agreement intended to allow us- ers to freely share, modify, and use this Dataset while maintaining this same freedom for others, provided that the original source and author(s) are credited. Link: https://doi.org/10.3897/zse.97.63936.suppl5 Supplementary material 6 Table S6. Within taxa uncorrected p—distances (COI) of reptile taxa identified in this study Authors: Francesco Belluardo, Darwin Diaz Quiros, Javier Lobon-Rovira, Goncgalo M. Rosa, Malalatiana Rasoazanany, Franco Andreone, Angelica Crottini Data type: genetic distances Copyright notice: This dataset is made available under the Open Database License (http://opendatacom- mons.org/licenses/odbl/1.0/). The Open Database License (ODbL) is a license agreement intended to allow users to freely share, modify, and use this Data- set while maintaining this same freedom for others, provided that the original source and author(s) are credited. Link: https://doi.org/10.3897/zse.97.63936.suppl6 Zoosyst. Evol. 97 (2) 2021, 315-343 Supplementary material 7 Figure Sl. Amphibians Neighbor joining tree of the 16S rRNA gene 3’ terminus Authors: Francesco Belluardo, Darwin Diaz Quiros, Javier Lobon-Rovira, Goncalo M. Rosa, Malalatiana Rasoazanany, Franco Andreone, Angelica Crottini Data type: phylogenetic tree Explanation note: The tree was computed with MEGA X 10.0.5 (Kumar et al. 2018) setting 1000 bootstrap repli- cates. The evolutionary distances were computed with the Kimura 2—parameter method, and ambiguous posi- tions were removed with the pairwise deletion option. Copyright notice: This dataset is made available under the Open Database License (http://opendatacommons. org/licenses/odbl/1.0/). The Open Database License (ODbL) is a license agreement intended to allow us- ers to freely share, modify, and use this Dataset while maintaining this same freedom for others, provided that the original source and author(s) are credited. Link: https://do1.org/10.3897/zse.97.63936.suppl7 Supplementary material 8 Reptiles Neighbor joining tree of the cytochrome oxidase I gene Authors: Francesco Belluardo, Darwin Diaz Quiros, Javier Lobon-Rovira, Goncalo M. Rosa, Malalatiana Rasoazanany, Franco Andreone, Angelica Crottini Data type: phylogenetic tree Explanation note: The tree was computed with MEGA X 10.0.5 (Kumar et al. 2018) setting 1000 bootstrap repli- cates. The evolutionary distances were computed with the Kimura 2—parameter method, and ambiguous positions were removed with the pairwise deletion option. Copyright notice: This dataset is made available under the Open Database License (http://opendatacommons. org/licenses/odbl/1.0/). The Open Database License (ODbL) is a license agreement intended to allow us- ers to freely share, modify, and use this Dataset while maintaining this same freedom for others, provided that the original source and author(s) are credited. Link: https://do1.org/10.3897/zse.97.63936.suppl8 343 Supplementary material 9 Table S7. Locality records of amphibian and reptile species identified in this study Authors: Francesco Belluardo, Darwin Diaz Quiros, Javier Lobon-Rovira, Goncalo M. Rosa, Malalatiana Rasoazanany, Franco Andreone, Angelica Crottini Data type: occurrences Explanation note: Species records from this study are marked with '+'. Records from previous publications are labelled with '*', and the source is reported in the column 'Reference'. Localities coordinates are avail- able in Suppl. material 1: Table S1. Copyright notice: This dataset is made available under the Open Database License (http://opendatacommons. org/licenses/odbl/1.0/). The Open Database License (ODbL) is a license agreement intended to allow us- ers to freely share, modify, and use this Dataset while maintaining this same freedom for others, provided that the original source and author(s) are credited. Link: https://do1.org/10.3897/zse.97.63936.suppl9 zse.pensoft.net