The research objectives of the thesis determine the diversity and distribution characteristics of amphibian species (EN) in some limestone areas of Northern Vietnam. Species composition and genetic relations of two species of Limnonectes and Odorrana frogs in Vietnam. Conservation values and threats to frog species in the study area and recommendations for conservation. To know the detailed research content, invite you to consult the thesis.
MINISTRY OF EDUCATION AND VIETNAM ACADEMY TRAINING OF SCIENCE AND TECHNOLOGY GRADUATE UNIVERSITY SCIENCE AND TECHNOLOGY PHAM THE CUONG STUDY ON THE DIVERSITY AND DISTRIBUTION PATTERNS OF AMPHIBIANS (AMPHIBIA) IN KARST FORESTS OF THE NORTH VIETNAM AND CONSERVATION MEASURES Research field: Zoology Code: 62.42.01.03 SUMMARY OF BIOLOGY DOCTORAL THESIS HA NOI – 2018 This dissertation is submitted to the Committee of the Graduate University of Science and Technology Supervisors: Dr Nguyen Quang Truong Dr Le Duc Minh Referee 1: Ass Prof Dr Le Nguyen Ngat Referee 2: Ass Prof Dr Hoang Ngoc Thao Referee 3: Dr Hoang Van Ngoc The research has been carried out at the Graduate University of Science and Technology Date of Oral Presentation:……,… /…… /2018 Copy of this dissertation is available: National Library of Viet Nam Graduate University of Science and Technology INTRODUCTION The necessity of the research Vietnam is recognized as one of the countries with a high level of biodiversity in the world (Conservation International 2014) In terms of amphibian diversity (Class Amphibia), the species number recorded from Vietnam has been rapidly increased in recent decades: from 82 species in 1996 to 162 species in 2005 and up to 176 species in 2009 and the current species number is ca 230 (Nguyen & Ho 1996, Nguyen et al 2005, 2009, Frost 2017) Karst ecosystem is considered as a natural laboratory for taxonomic, ecological, evolutionary and biogeographical research (Clements et al 2006) Karst forests provide a variety of distinct microhabitats like “isolated islands” and are noted for their high levels of endemism (Clements et al 2006) In Vietnam, limestone forests are extensive in the northeastern, northwestern regions and northern Truong Son Range (Sterling et al 2006) However, limestone forest is a sensitive ecosystem to the human impacts and it is very difficult to restore Limestone in Vietnam has been exploited for infrastructure and economic development, for example, quarrying for road construction, and cement industry Moreover, the fauna in karst forests is threatened by decline because of forest destruction, overexploitation, tourism development and climate change, in particular, the poikilotherm like amphibians (Sterling et al 2006) The research entitled "Study on the diversity and distribution patterns of amphibians (Amphibia) in karst ecosystem of the North Vietnam and conservation measures” aims to assess the diversity of amphibians in limestone forests of northern Vietnam with special focus on two group complexes of anuran frogs, Limnonectes and Odorrana, in Vietnam In addition, recommendations for conservation measures are also provided Objectives - To assess the diversity of amphibians in some selected limestone forests of northern Vietnam; - To review taxonomic and phylogenetic relationships of two genus complexes, Limnonectes and Odorrana, in Vietnam; 10 - To assess conservation potential and threats to the amphibian fauna and to provide recommendations for conservation Contents Task 1: Inventory of the species diversity of amphibians in some karst forests in northern Vietnam: Hoa Binh Province in the northwestern region, Cao Bang and Ha Giang provinces in the northeastern region and Cat Ba Island, an offshore island in the Gulf of Tonkin This research also focuses on the discovery of new species and new country records Task 2: Evaluating distribution patterns of amphibian species across altitudinal gradient (from to 1300 m), habitat types and life modes (on the tree, on the ground, and associated with water) Task 3: Evaluating biogeographic relationships of amphibian fauna amongstudy sites, between mainland and offshore islands, and between northeastern and northwestern regions to verify the evolutionary barrier of the Red River Task 4: Evaluating taxonomy and phylogenetic relationships of two genus complexes, Odorrana and Limnonectes, based on the morphological comparison and molecular analyses Task 5: Assessment of conservation potential based on the following criteria: species diversity, number of endemic and threatened species, possibility of population restoration Evaluation of threats to the amphibian populations and provide recommendations for conservation measure Scientific and conservation significances Scientific significance: The research provides updated data about the diversity of amphibians in the karst forests of northern Vietnam The research also provides novel data on taxonomy and phylogenetic relationships of Limnonectes and Odorrana from Vietnam Conservation significance: The research provides baseline data for biodiversity conservation planning and management in northern Vietnam New findings A total 65 species of amphibans were recorded in karst forests of the North Vietnam, comprising 44 species recored from Ngoc Son-Ngo 11 Luong NR, 32 species recorded from Hang Kia-Pa Co NR, 33 species recorded from Bac Me NR, 21 species recorded from Ha Lang District, and 23 species recorded from Cat Ba NP New species: Three new species were described as new for science, Odorrana mutschmanni from Cao Bang Province, Theloderma annae and Rhacophorus hoabinhensis from Hoa Binh Province New records: Three species, Leptolalax minimus, Odorrana hainanensis, and O lipuensis, were recorded for the first time from Vietnam and some other new provincial records from northern Vietnam: One species from Cao Bang Province; two species from Ha Giang Province; three species from Hai Phong city; and five species from Hoa Binh Province Biogeographic relationships: Based on the list of amphibian species recorded from northern Vietnam, the amphibian fauna of the northeastern region was distinctly separated from that of the northwestern region (Similarity Index = 0.6087) Moreover, the amphibian fauna of the karst forests in mainland also differs from that of the offshore islands (Similarity Index = 0.4706) Distribution patterns: Most of amphibian species were recorded at elevations below 800 m (58 species), whereas 34 species were found at elevations above 800 m In terms of habitat types, most of amphibian species were recorded in the undisturbed natural forests (58 species), while the species number recorded in the disturbed forest was 33 species and agricultural and residential areas was 16 species The highest number of record species were found on the ground (42 species) In this study, we also provide taxonomic review of two poorly known genera of amphibians, Limnonectes (8 species) and Odorrana (25 species), based on morphological and molecular data with descriptions of two new species for science and three new country records for Vietnam Phylogenetic relationships of these genera were also discussed with remarks on some species complexes In addition, we evaluated the threats to the amphibian fauna and provided some recommendations for conservation in northern Vietnam 12 Chapter LITERATURE REVIEW 1.1 Overview of amphibian research in region Currently, a total 7.697 species of amphibians was recorded in the world (Frost 2017) In the neigboring countries, 432 species have been recorded from China; 152 species from Lao and 79 species from Cambodia (Frost 2017) In terms of conservation concern, Stuart et al (2004) stated that 168 species of amphibian were extinction and at least 2,500 species are under threatened in the last 20 years (1984-2004) 1.2 Overview of amphibian research in Vietnam 1.2.1 Faunistic inventory and new discoveies Bourret (1942) reported 171 species and subspecies of amphibians in Indochina (Vietnam, Laos, Cambodia) Nguyen and Ho (1996) published a checklist of reptiles and amphibians of Vietnam with 82 recorded species of amphibians Nguyen et al (2005) listed 162 species of amphibians In the recent book, Nguyen et al (2009) recorded a total of 176 species of amphibians from Vietnam Since 2010 more than 40 new species of amphibians have been described from Vietnam (Forst 2017) A number of studies on amphibians were conducted in the Northeast, Northwest, Central, Central Highlands, Southern and some offshore islands of Vietnam 1.2.2 Research on biology and ecology Research on biological and ecological characters of amphibians focused on few species that were kept in captivity, the widespread species or endemic to Vietnam: Duttaphrynus melanostictus, Hoplobatrachus rugulosus, Limnonectes bannaensis, Hylarana maosonensis, Quasipaa verrucospinosa, Polypedates mutus, Paramesotriton deloustali 1.2.3 Research on tadpoles and bioacoustics Descriptions of the tadpoles of the following species were mentioned in previous studies (Hendrix et al 2007, Gawor et a 2009, Wildenhues et al 2010, Rauhaus et al 2012, Pham et al 2015): Rhacophorus annamensis, Hynarana nigrovittata, Rhacophorus 13 maximus, Theloderma corticale, Amolops cremnobatus Le Thi Quy (2015) described the tadpoles of 21 amphibian species from Bach Ma National Park, Thua Thien Hue Province In addition, the bioacoustic analysis is a useful approach to study biology and taxonomy of amphibians Le Trung Dung (2016) described the biaocoustics of 11 amphibians species 1.2.4 Research on phylogenetic relationships Frost et al (2006) provided a comprehensive phylogenetic tree of amphibian species of the world Li et al (2008, 2009), Biju et al (2010), Orlov et al (2012), Kuraishi et al (2012), Li et al (2012), Yu et al (2010, 2013), Nguyen et al (2015), Poyarkov et al (2015) studied on taxonomy and phylogenetic relationships of various groups of Rhacophoridae 1.2.5 Research on pathology and conservation issues Rowley et al (2013), Nguyen et al (2013), Martel et al (2014), Laking et al (2017) and Nguyen et al (2017) have identified a number of amphibian populations that infected with Batrachochytrium dendrobatidis and B salamandrivorans Rowley et al (2010, 2016) assessed threats to the populations of amphibians in Southeast Asia, including Vietnam Key impacts include habitat loss, overexploitation for food, medicinal and pet trade 1.2.6 Research on amphibians in northern Vietnam Although numerous field excursions have been conducted in northern Vietnam, most of surveys were conducted in evergreen forests such as Tay Yen Tu, Tay Con Linh, Phia Oac-Phia Den, Hoang Lien, Muong Nhe, Sop Cop, and Copia nature reserves (Hecht et al 2013, Bain & Nguyen 2004, Nguyen Thien Tao 2009, Orlov et al 2001, Le Trung Dung 2016, Pham Van Anh 2016) Research in limestone karst ecosystems are still limited with descriptions of new species, namely Tylototrion ziegleri from Cao Bang and Ha Giang provinces, Liuixalus calcarius and Philautus catbaensis from Cat Ba Island (Nishikawa et al 2013, Milto et al 2013) 1.2.7 Research on Limnonectes and Odorrana in Vietnam Genus Limnonectes: Limnonectes is one of the most diverse groups of amphibians to date with 68 currently recognized species and 14 16 of which have been described in the last ten years (Frost 2017) According to Nguyen et al (2009), five species of the genus Limnonectes are currently recorded from Vietnam However, some taxonomic changes have been published in recent years Due to their morphological similarity, species that were previously thought to be widespread are now being recognized as complexes of species with much narrower distributions Genus Odorrana: Odorrana is one of the most diverse groups of amphibians to date with 58 currently recognized species, more than 20 of which have been described in the last ten years (Frost 2017) Due to morphological similarity in the genus, species that were formally thought to be widespread are now being recognized as complexes of species with much narrower distributions Further study on taxonomic and genetic relationships should be continued 1.3 Natural conditions of the study area 1.3.1 Natural conditions of North Vietnam The North Vietnam bordered by China’s Guangxi Province to the northeast and Yunnan Province to the northwest, and by Laos along its western border (Sterling et al 2006) Limestone kart forests: Most of the limestone mountain area in Vietnam is distributed in northern Viet Nam, partly in Quang Binh province In the Gulf of Tonkin, there are thousands of limestone islands in Ha Long Bay next to each other with Cat Ba Island as the center (Sterling et al 2006) Vegetation: The dominant forest type in northern Vietnam is evergreen, including both broad-leaved and coniferous plants, with pockets of semi-evergreen forest Coastal mangrove formations and forests growing on limestone are also important elements of the north’s natural landscape diversity (Sterling et al 2006) Fauna: Northern Vietnam’s animal communities are a mixture of tropical and subtropical species, many of which are unique to the northern third of the country Within the region, the Red River may be an effective barrier to movement (or coincide with one) for some groups, such as amphibians and reptiles, resulting in different species and communities in the northeast and northwest (Sterling et al 2006) 15 1.3.2 Natural conditions of survey sites Field surveys were conducted in the limestone forestin northern Vietnam Northwest: Ngoc Son-Ngo Luong Nature Reserve (Hoa Binh Province): This NR is located in Tan Lac and Lac Son districts with a total area of 19.254 The area is characterized by limestone mountains, with steep terrain and rugged terrain Between the limestone mountains there are several flat valleys running in the north-west direction The altitude ranges from 100-1.065 m (Birdlife International 2004) Hang Kia-Pa Co Nature Reserve (Hoa Binh Province): Hang Kia-Pa Co Nature Reserve is located in Mai Chau District with a total area of 5.257 There are many high and pointed limestone blocks, the highest peak of which is 1.536m in the Northwest of the area, the height decreasing to the East Most of the nature reserve stands at over 500 m (Birdlife International 2004) Northeast: Bac Me Nature Reserve (Ha Giang Province): This NR is located in Bac Me District with a total area of 9.042 ha, Bac Me Nature Reserve is located in the depression of the Gam River to the south-east of Ha Giang Province, with steep terrain and strong splinters with many peaks above 1000 m along the Song Gam gulf (Biodiversity Report Bac Me Nature Reserve 2009) Ha Lang Forest (Cao Bang Province): Ha Lang District has a limestone karst topography that occupies most of the area, at an altitude of 100-750 m above sea level According to the biodiversity conservation planning of Cao Bang Province, a nature reserve will be established in this district with an area of 7.343 (Decree No 697/QĐ-UBND) Cat Ba National Park: Cat Ba is the largest island of the Cat Ba Archipelago with a total area of 26.241 Cat Ba National Park is located in the limestone archipelago of hundreds of large and small islands at the height of 100-150 m, the highest point is Cao Vong peak 331m and the area of this protected area is 16.196 (www.vuonquocgiacatba.com.vn) 16 CHAPTER MATERIALS AND METHODS 2.1 Field survey and study sites Fifteen field surveys with 119 days were conducted from 2014 to 2017 Survey sites: Cat Ba National Park (Hai Phong City), Ha Lang Disstrict (Cao Bang Province), Bac Me Nature Reserve (Ha Giang Province), Hang Kia-Pa Co and Ngoc Son-Ngo Luong nature reserves (Hoa Binh Province) Sampling of Limnonectes and Odorrana: Twenty field surveys with 200 days were conducted in more than twenty provinces in Vietnam from 2014 to 2017 Molecular analysis was carried out at the Institute of Ecology and Biological Resources and Vietnam National Museum of Nature, and Hanoi University of Science 2.2 Methods 2.2.1 Sampling Excursions were carried out during the day and at night in different habitat types Survey transects were set up along the streams, pools, small ponds or along the forest paths, under the leaf litter or on the tree branches, we thus also survey the forest near the cave entrances and cliffs 2.2.2 Laboratory work 2.2.2.1 Morphological examination Taxonomic identification: Morphological comparisons were made with the voucher specimens which are deposited in the collections of the Institute of Ecology and Biological Resources and Vietnam National Museum of Nature For taxonomic identification, we used the following documents: Bourret (1942), Bain et al (2003), Ohler et al (2011), Orlov et al (2003, 2006, 2012), Suwannapoom et al (2016), Taylor (1962), and othe related documents that were cited in each species account Species names followed Nguyen et al (2009), Frost (2017) and recently published papers 2.2.2.2 Molecular analysis A fragment of mitochondrial gene (cytochrome b, 12S, 16S) was sequenced to investigate phylogenetic relationships of all species of the genus Limnonectes and Odorrana in Vietnam 17 42 Ếch suối meng-la Họ Ếch 43 Nhái sọc 44 Nhái wa-za 45 Nhái quang 46 Nhái nhỏ đá vôi 47 Nhái tay-lo 48 Nhái cát bà 49 50 51 52 53 54 55 Ếch đầu to Ếch mi-an-ma Nhái tí hon Ếch xanh đốm Ếch phê Ếch ki-ơ Ếch hòa bình 56 57 58 59 Ếch lớn Ếch oóc-lốp Ếch màng bơi đỏ Ếch sần an-na 60 61 62 63 Ếch sần đốm trắng Ếch sần bắc Ếch sần go-don Ếch sần đỏ Bộ Có Họ cá cóc 64 Cá cóc zig-lơ Bộ Không chân Họ Ếch giun 65 Ếch giun ban-na Sylvirana menglaensis (Fei, Ye & Xie, 2008)(*) Rhacophoridae Hoffman, 1932 Feihyla vittata (Boulenger, 1887) Gracixalus waza Nguyen, Le, Pham, Nguyen, Bonkowski & Ziegler, “2012” 2013 Gracixalus quangi Rowley, Dau, Nguyen, Cao & Nguyen, 2011(*) Liuixalus calcarius Milto, Poyarkov, Orlov & Nguyen, 2013 Kurixalus bisacculus (Taylor, 1962) Philautus catbaensis Milto, Poyarkov, Orlov & Nguyen, 2013 Polypedates megacephalus Hallowell, 1861 Polypedates mutus (Smith, 1940) Raorchestes parvulus (Boulenger, 1893) Rhacophorus dennysi Blanford, 1881 Rhacophorus feae (Boulenger, 1893) Rhacophorus kio Ohler & Delorme, 2006 Rhacophorus hoabinhensis Nguyen, Pham, Nguyen, Eto & Ziegler, 2017(***) Rhacophorus maximus Günther, 1858(*) Rhacophorus orlovi Ziegler & Kohler, 2001 Rhacophorus rhodopus Liu & Hu, 1962(*) Thelodema annae Nguyen, Pham, Nguyen, Ngo & Ziegler, 2016 (***) Theloderma albopunctatum (Liu & Hu, 1962) Theloderma corticale (Boulenger, 1903)(*) Theloderma gordoni Taylor, 1962(*) Theloderma lateriticum Bain, Nguyen & Doan, 2009(*) Caudata Fischer von Waldheim, 1813 Salamandridae Goldfuss, 1820 Tylototriton ziegleri Nishikawa, Matsui & Nguyen, 2013 Gymnophiona Müller, 1831 Ichthyophiidae Taylor, 1968 Ichthyophis bannanicus Yang, 1984 Total + + + + + + + + + + + + + + + + + + + + + + + + + + + + + + + + + + + + + + + + + + + + + + + + + + + + 2 + + + + 4 3 Notes: (*)-New record for provinces, (**)-New record for Vietnam, (***)-New species Study sites: 1Ngoc Son-Ngo Luong NR, 2-Hang Kia-Pa Co NR, 3-Bac Me NR, 4-Ha Lang District, 5-Cat Ba NP 3.1.2 New discoveries - New species: Three new species were described as new for science: Odorrana mutschmanni from Cao Bang Province, Theloderma annae, Rhacophorus hoabinhensis from Hoa Binh Province - New records for Vietnam: Three species were recorded for the first time from Vietnam: Leptolalax minimus from Hoa Binh Province, Odorrana hainanensis and O lipuensis from Cao Bang Province - New provincial records: One species (Rhacophorus maximus) was recorded for the first time from Cao Bang Province, two species (Leptolalax minimus, Rhacophurus rhodopus) were recorded for first 21 time from Ha Giang Province, three species (Microhyla fissipes, Hylarana macrodactyla, Theloderma corticale) were recorded for first time from Hai Phong City, five species (Limnonectes linborgi, Sylvirana menglaensis, Gracixalus quangi, Theloderma lateriticum, T gordoni) were recorded for first time Hoa Binh Province 3.1.3 Taxonomic accounts In this section, we described the morphological character of 40 species based on novel data obtained from examined specimens Below in an example of the species description Sylvirana menglaensis (Fei, Ye & Xie, 2008) Specimens examined (n=5): Three adult males IEBR 3924, 3925, 3927 (HB 2014.190, 198, HB 2015.70) and two adult females IEBR 3926, 3928 (HB 2014.199, HB 2015.28) collected from Hoa Binh Province Morpholigical characters: The specimens agreed well with the description of Fei et al (2008), Le et al (2014) Males slightly smaller than females (SVL 40-50 mm), head longer than wide, tympanum distinct, about 2/3 half of eye diameter, males with vocal sacs Webbing formula I0–1II1/3– 1III1/2–1IV1–0V Skins: Dorsum slightly shagreened, dorsolateral fold present, ventral surface smooth Coloration in life: Dorsum brown, flanks grey-yellow with some large black spots, belly white Ecological notes: The specimens were found between 19:00 and 22:00, on the bank of rocky stram The surrounding habitat was mixed secondary karst forest, composed of large, medium and small hardwoods, shrubs and vines Distribution: Ngoc Son-Ngo Luong and Hang Kia-Pa Co nature reserves, Hoa Binh Province 3.2 Comparison of species composition of amphibians 3.2.1 Between study sites The highest Similarity Index is between Ngoc Son-Ngo Luong NR and Hang Kia-Pa Co NRs (Similarity Index = 0.7733), followed by that between Bac Me NR and Ha Lang District (Similarity Index = 0.6038) The species compostion of the amphibian fauna of Cat Ba NP was distinctly separated from those of other sites (Figure 3.2) 22 Figure 3.2 Cluster analysis of the species composition of the amphibians between study sites (bootstrap value: 1000 replications) Notes: NS-NL = Ngoc Son-Ngo Luong NR, HK-PC = Hang Kia-Pa Co NR, BM = Bac Me NR, HL = Ha Lang District, CB = Cat Ba NP 3.2.2 Between mainland and Cat Ba island The amphibian fauna of the sites in mainland was distinctly separated from that of Cat Ba island (Similarity Index = 0.4706) 3.2.3 Between northeastern and northwestern regions The species composition of amphibians between the northeastern and northwestern regions is disctinct from each other (Similarity Index = 0.6087) This is also supported by the distinctness in morphological characteristics and genetic distance of some species groups found on each side of the Red River 3.2.4 Between forest areas in northeastern and northwestern Figure 3.3 Cluster analysis of the species composition of the amphibians between forest areas in northwestern (bootstrap value: 1000 replications) 23 Figure 3.4 Cluster analysis of the species composition of the amphibians between forest areas in northeastern (bootstrap value: 1000 replications) Figures 3.3 and 3.4 showed the separation of species composition between limestone forest and evergreen forest in the northwestern and northeastern regions 3.3 Distribution patterns of amphibians 3.3.1 Altitudinal gradient Figure 3.5 Species composition according altitudinal gradient Most of amphibian species were recorded at elevations below 800 m (58 species), whereas 34 species were found at elevations above 800 m (Figure 3.5) 3.3.2 Habitat types In terms of habitat types, most of amphibian species were recorded in the undisturbed natural forests (58 species), while the species number 24 recorded in the disturbed forest was 33 species and agricultural and residential areas was 16 species 3.3.3 Life modes The highest number of record species were found on the ground (42 species), 25 species found on the trees, and 22 found in associated with water 3.4 Species diversity and phylogenetic relationships of Limnonectes and Odorrana from Vietnam 3.4.1 Species diversity and phylogenetic relationships of the genus Limnonectes from Vietnam 3.4.1.1 Species diversity Eight species of Limnonectes were recorded from Vietnam (Table 3.5) One species was described as new for science, Limnonectes quangninhensis, from Quang Ninh Province New provincial records: Limnonectes dabanus was recorded for the first time from Binh Thuan, Ninh Thuan, Khanh Hoa provinces; L gyldenstolpei was recorded for the first time from Dong Nai Province; L limgorgi was recorded for the first time from Quang Ninh, Hoa Binh, Kon Tum; L nguyenorum was recorded for the first time from Vinh Phuc, Son La; L poilani was recorded for the first time from Binh Thuan, Ninh Thuan, Khanh Hoa Two species, Limnonectes nguyenorum and L quangninhensis are currently known only from Vietnam Table 3.5 List of the genus Limnonectes recorded from Vietnam No Scientific name Limnonectes bannaensis Ye, Fei & Jiang, 2007 L dabanus (Smith, 1922) (*) L gyldenstolpei (Andersson, 1916) (*) L khammonensis (Smith, 1929) L limborgi (Sclater, 1892) (*) L nguyenorum Mcleod, Kurlbaum & Hoang, 2015(*) L poilani (Bourret, 1942) (*) L quangninhensis Pham, Le, Nguyen, Ziegler, Wu & Nguyen, 2017(**) Vietnamese name Ếch nhẽo ban-na Ếch gáy dô Ếch gyl-den-s-tol-pe Ếch nhẽo khăm muộn Ếch lim-boc Ếch nhẽo nguyễn Endemic + Ếch nhẽo poi-lan Ếch nhẽo quảng ninh + Ghi chú: (*)-New provincial records, (**)-New species 3.4.1.2 Taxonomic accounts We described species of Odorrana recorded from Vietnam, including additional data about distribution and ecological characteristics 25 A key to species of the genus Limnonectes from Vietnam was also provided 3.4.1.3 Phylogenetic relationships Species of genus Limnonectes from Vietnam are placed in six independent clades, with strong support values: the Clade Icontaining L dabanus, L gyldenstolpei, L hascheanus, L lauhachindai, L limborgi, L macrognathus, and L kohchangae; the clade IIconsisting of L sp1., L sp2., and L fragilis; the clade III containing only L poilani; the clade IV consisting of L isanensis, L jarujini, L megastomias, L longchuanensis, L nguyenorum, L sp4., L taylori; the clade V containing only L bannaensis; and the clade VI conssisting of L fujianensis and L quangninhensis (Figure 3.6) Figure 3.6 Phylogram based on Bayesian analysis of the genus Limnonectes in Vietnam Number above branches are Bayesian posterior probabilities 26 3.4.2 Species diversity and phylogenetic relationships of the genus Odorrana from Vietnam 3.4.2.1 Species diversity A total of 25 species of genus Odorrana was recorded from Vietnam (Table 3.12) One new species was described as new for science: Odorrana mutschmanni from Cao Bang Province; three species were recorded for the first time from Vietnam: Odorrana hainanensis, O lipuensis, and O versabilis In addition, we provide the first descriptions of females of two species: Oodorran gigatympana and O trankieni New provincial records: Odorrana bacboensis was recorded for the first time from Ha Tinh; O gigatympana was recorded for the first time from Thua Thien-Hue, Quang nam, Gia Lai, and Phu Yen provinces; O jingdongensis was recorded for the first time from Lai Chau and Dien Bien provinces; O khalam was recorded for the first time from Quang Nam Province; O morafkai was recorded for the first time from Thanh Hoa, Kon Tum, and Lam Dong provinces; O tiannanensis was recorded for the first time from Cao Bang and Thanh Hoa provinces; O trankieni was recorded for the first time from Hoa Binh and Bac Giang provinces; and O yentuensis was recorded for the first time from Quang Ninh Province Among 25 recorded species of Odorrana from Vietnam, 11 species are globally or nationally threatened at different levels (Table 3.12) Table 3.12 List of the genus Odorrna recorded from Vietnam No Scientific name Vietnamese name Odorrana absita (Stuart & Chan-ard, 2005) O andersonii (Boulenger, 1882) O bacboensis (Bain, Lathrop, Murphy, Orlov & Ho, 2003)(*) O banaorum (Bain, Lathrop, Murphy, Orlov & Ho, 2003) O chapaensis (Bourret, 1937) O chloronota (Günther, 1876) O geminata Bain, Stuart, Nguyen, Che & Rao, 2009 O gigatympana (Orlov, Ananjeva & Ho, 2006) (*) O grahami (Boulenger, 1917) O graminea (Boulenger, 1900) Odorrana hainanensis Fei, Ye & Li, 2001(**) Ếch mõm ap-si-ta Chàng an-dec-sơn 10 11 27 IUCN RL 2017 RBVN 2007 Ende -mic VU Ếch bắc Ếch ba na Ếch bám đá sa pa Ếch xanh NT Ếch bám đá hoa VU Ếch màng nhĩ khổng lồ Ếch g-ra-ham Ếch g-ra-mi-ne Ếch hải nam + NT VU 12 13 14 15 16 17 18 19 20 21 22 23 24 25 Ếch ging-đông Ếch giun-li-an Ếch ta-bu O jingdongensis Fei, Ye & Li, 2001(*) O junlianensis Huang, Fei &Ye, 2001 O khalam (Stuart, Orlov & Chan-ard, 2005) (*) O lipuensis Mo, Chen, Wu, Zhang & Zhou, 2015(**) O margaretae (Liu, 1950) O morafkai (Bain, Lathrop, Murphy, Orlov & Ho, 2003) (*) O mutschmanni Pham, Nguyen, Le, Bonkowski & Ziegler, 2016(***) O nasica (Boulenger, 1903) O orba (Stuart & Bain, 2005) O schmackeri (Boettger, 1892) O tiannanensis (Yang & Li, 1980) (*) O trankieni (Orlov, Le & Ho, 2003) (*) O versabilis (Liu & Hu, 1962) (**) O yentuensis Tran, Orlov & Nguyen, 2008(*) VU VU Ếch li-pu Ếch mac-ga-ret Ếch mo-rap-kai Ếch đá mut-x-man Ếch mõm dài Ếch mồ côi Ếch s-mac-ko Ếch ti-an-nan Ếch trần kiên Ếch đá Ếch yên tử + + EN Notes (*)-New provincial records, (**)-New records for Vietnam, (***)-New species RBVN (2007) = Vietnam Red Data Book (2007) Part I Animals: VU = Vulnerabale; IUCN (2017) = IUCN Red List of Threatened Species: EN = Endangered, VU = Vulnerabale, NT = Near Threatened 3.4.2.2 Taxonomic accounts We described 18 species of Odorrana recorded from Vietnam, including additional data about distribution and ecological characteristics A key to 25 species of the genus Odorrana from Vietnam was also provided 3.4.2.3 Phylogenetic relationships Species of the genus Odorrana from Vietnam are placed in four indipendent clades: the clade I consisting of O banaorum, O chloronota, O graminea, O leporipes, O morafkai, O sp1, O sp2, O tiannanensis, O nasica, O yentuensis, O trankieni, O nasuta, O bacboensis O fengkaiensis, O hainanensis, O gigatympana, and O sp3; the clade II containing O chapaensis and O geminata; the clade III consisting of O andersonii, O grahami, O jingdongensis, O junlianensis, O margaretae, O mutschmanni, and O wuchuanensis; and the clade IV containing only O lipuensis (Figure 3.7) 28 Hình 3.7 Phylogram based on the Bayesian analysis of the genus Odorrana in Vietnam Number above branches are Bayesian posterior probabilities 29 3.5 Conservation issues 3.5.1 Species of conservation concern The amphibian fauna of karst forests of the North Vietnam contain a high level of conservation concern with six species listed in the Red Data Book of Vietnam (2007), six species listed in the IUCN Red list (2017), and nine species are currently known only from Vietnam (Table 3.16) Table 3.16 List of threatened and endemic species No 10 11 12 13 14 15 16 17 18 19 Species name Ingerophrynus galeatus Quasipaa delacouri Quasipaa boulengeri Quasipaa verucospinosa Odorrana geminate Odorrana hainanensis Odorrana mustchmanni Gracixalus quangi Gracixalus waza Liuixalus catbaensis Philautus catbaensis Rhacophorus feae Rhacophorus kio Rhacophorus hoabinhensis Theloderma annae Theloderma corticale Theloderma lateriticum Tylototriton ziegleri Ichthyophis bannanicus Tổng số RBVN (2007) IUCN RL (2017) VU EN Ende -mic Location NS NL HK PC BM HL CB + + EN NT VU VU + + + + + + + + VU + + + + + EN EN + + + + + + EN VU VU + + + + + + + + + + + + + + + + + 3.5.2 Threats to the amphibian faunas in the study sites 3.5.2.1 Habitat loss and degradation Major causes of deforestation and degradation in the study areas are exploitation of forest products, forest fire, timber logging, and road construction in Hang Kia-Pa Co and Ngoc Son-Ngo Luong NRs (Hoa Binh Province), and Bac Me NR (Ha Giang Province) 3.5.2.2 Hunting of amphibians Amphibians were collected by local peoples for food and wildlife trade in Ngoc Son-Ngo Luong NR (Hoa Binh Province) and Bac Me NR (Ha Giang Province) 21 3.5.3 Conservation measures 3.5.3.1 Priority sites of conservation Priority sites of conservation for amphibians in study areas were evaluated on the basis of the following criteria: species richness, number of endemic and threatened species, forest area and habitat quality, and level of human disturbance The Ngoc Son-Ngo Luong NR and Cat Ba NP are two sites with a high level of conservation priority 3.5.3.2 Species of conservation concern Species of conservation consern comprises all threatened and endemic species, for instance the members of Quasipaa, Rhacophorus, Theloderma, and Tylototriton 3.5.3.3 Conservation activities The following conservation activities are recommended in order to protect the habitat and populations of amphibians in northern Vietnam: Establishment of the Ha Lang Nature Reserve in Cao Bang Province Protecting the existing fores area in all survey sites in northern Vietnam and linkage of isolated forest patches Sustainable use of amphibians for food consumption and control hunting activities in breeding season Protecting all endemic and threatened species Developing a breeding program for some amphibian species in order to provide breeding animals for food consumption or pet trade: Limnonectes, Quasipaa, Rhacophorus, and Theloderma Economic development and ecotourism: Developing a cultural tourism program, combining nature discovery and animal observation in Ngoc Son-Ngo Luong, Hang Kia-Pa Co, Ha Lang and on Cat Ba Island Raising awareness campaign about amphibian crisis and conservation 22 CONCLUSIONS AND RECOMMENDATIONS Conclusions Species diversity: A total 65 species of amphibans were recorded in karst forests of the North Vietnam, comprising 44 species recored from Ngoc Son-Ngo Luong Nature Reserve, 32 species recorded from Hang Kia-Pa Co Nature Reserve, 33 species recorded from Bac Me Nature Reserve, 21 species recorded from Ha Lang District, and 23 species recorded from Cat Ba National Park Three new species were described as new for science, Odorrana mutschmanni, Theloderma annae, and Rhacophorus hoabinhensis Three species, Leptolalax minimus, Odorrana hainanensis, O lipuensis, were recorded for the first time from Vietnam and some other new provincial records from northern Vietnam: one species from Cao Bang Province, two species from Ha Giang Province, three species from Hai Phong City, and five species from Hoa Binh Province Biogeographic relationships: The amphibian fauna of the northeastern region was distinctly separated from that of the northwestern region (Similarity Index = 0.6087) The amphibian fauna of the karst forests in mainland also differs from that of the offshore islands (Similarity Index = 0.4706) Distribution patterns: Most of amphibian species were recorded at elevations below 800 m (58 species), whereas 34 species were found at elevations above 800 m In terms of habitat types, most of amphibian species were recorded in the undisturbed natural forests (58 species), while the species number recorded in the disturbed forest was 33 species and agricultural and residential areas was 16 species The highest number of record species were found on the ground (42 species), 25 species were found on the trees, and 22 were found on associated with water Eight species of genus Limnonectes and 25 species genus Odorrana were recorded from Vietnam, based on morphological and molecular data with descriptions of two new species for science and three new country records for Vietnam In terms of phylogenetic 23 relationships, members of Limnonectes were splitted into six clades and members of Odorrana were placed in four clades The amphibian fauna of karst forests of the North Vietnam contain a high level of conservation concern with six species listed in the Red Data Book of Vietnam (2007), six species listed in the IUCN Red list (2017), and nine species are currently known only from Vietnam Major threats to the amphibian faunas in the study sites arehabitat loss and degradation and hunting of amphibians Recommendations Further studies: Additional field work should be conducted in the border areas of Lang Son, Cao Bang, and Ha Giang provinces Some unidentified populations of Limnonectes and Odorrana from Vietnam should be described as new species or subspecies Conservation issues Priority sites of conservation: Ngoc Son-Ngo Luong NR and Cat Ba NP Species of conservation concern: Species of conservation consern comprises all threatened and endemic species, for instance the members of Quasipaa, Rhacophorus, Theloderma, and Tylototriton Conservation activities: Establishment of the Ha Lang Nature Reserve in Cao Bang Province Protecting the existing fores area in all survey sites in northern Vietnam and linkage of isolated forest patches Sustainable use of amphibians for food consumption and control hunting activities in breeding season Protecting all endemic and threatened species Developing a breeding program for some amphibian species in order to provide breeding animals for food consumption or pet trade: Limnonectes, Quasipaa, Rhacophorus, and Theloderma Economic development and ecotourism: Developing a cultural tourism program, combining nature discovery and animal observation in Ngoc Son-Ngo Luong, Hang Kia-Pa Co, Ha Lang and on Cat Ba Island Raising awareness campaign about amphibian crisis and conservation 24 PUBLISHED PAPERS RELATED TO THE DISSERTATION Pham The Cuong, Nguyen Quang Truong, Ngo Ngoc Hai (2015): Diversity of tree frogs (Amphibia: Anura: Rhacophoridae) from Hoa Binh Province Proceedings of the sixth National Scientific Conference on Ecology and Biological Resources Publishing House for Science and Technology: 498– 453 Ngo Thi Hanh, Le Duc Minh, Pham The Cuong, Nguyen Quang Truong (2016): Species diversity and phylogenetic relationships of the genus Odorrana (Amphibia: Anura: Ranidae) from northern Vietnam Proceeding of the 3rd National Scientific Conference on Amphibians and Reptiles in Vietnam Publishing House for Science and Technology: 67–76 Nguyen, T.Q., Pham, C.T., Nguyen, T.T., Ngo, H.N & Ziegler, T (2016): A new species of Theloderma (Amphibia: Anura: Ranidae) from Vietnam Zootaxa, 4168(1): 171–186 Pham, A.V., Le, D.T, Pham, C.T., Nguyen, S.L.H., Ziegler, T & Nguyen, T.Q (2016): Two additional records of megophryid species, Leptolalax minimus (Taylor, 1962) and Leptobrachium masatakasatoi Matsui, 2013, for the herpetofauna of Vietnam Revue suisse de Zoologie, 123(1): 35–43 Pham Van Anh, Tu Van Hoang, Nguyen Quang Truong, Pham The Cuong, Song Ba Nenh, Bui The Quyen, Hoang Le Quoc Thang (2016): Species diversity of the family Dicroglossidae (Amphibia: Anura) from Son La Province Proceeding of the 3rd National Scientific Conference on Amphibians and Reptiles in Vietnam Publishing House for Science and Technology: 133–139 Pham, C.T., Nguyen, T.Q., Bernades, M., Nguyen, T.T., Ziegler T (2016): First records of Bufo gargarizans Cantor, 1842 and Odorrana lipuensis Mo, Chen, Wu, Zhang & Zhou, 2015 (Anura: Bufonidae, Ranidae) from Vietnam Russian Journal of Herpetology, 23(2): 103–107 Pham, C.T., Nguyen, T.Q., Le, M.D., Bonkowski, M & Ziegler, T (2016): A new species of Odorrana (Amphibia: Anura: Ranidae) from Vietnam Zootaxa, 4084(3): 421–435 Pham The Cuong, Nguyen Quang Truong, Ngo Ngoc Hai (2016): Species composition of amphibians from Ngoc Son-Ngo 25 10 11 12 13 14 Luong Nature Rescerve, Hoa Binh Province Proceeding of the 3rd National Scientific Conference on Amphibians and Reptiles in Vietnam Publishing House for Science and Technology: 125–132 Pham, A.V., Pham, C.T, Hoang, N.V, Ziegler, T., & Nguyen, T.Q (2017): New records of amphibians and reptiles from Ha Giang Province, Vietnam Herpetology Notes, 10: 183–191 Pham, C.T., An, H.T., Herbst, S., Bonkowski, M., Ziegler, T., & Nguyen T.Q (2017): First report on the amphibian fauna of Ha Lang karst forest, Cao Bang Province, Vietnam Bonn zoological Bulletin, 66 (1): 37–53 Pham, C.T., Le, M.D., Nguyen, T.T., Ziegler, T., Wu, Z.J., & Nguyen, T.Q (2017): A new species of Limnonectes (Amphibia: Anura: Dicroglossidae) from Vietnam Zootaxa, 4269(4): 545–558 Nguyen, T.T., Pham, C.T., Nguyen, T.Q., Ninh H.T., & Zigler, T (2017): A new species of Rhacophorus (Amphibia: Anura: Rhacophoridae) from Vietnam Asian Herpetological Research, 8(4): 221-234 Pham, C.T., Nguyen, T.Q., Nguyen, T.T., Nguyen, T.V., van Schingen, M., & Ziegler, T (2018): First record of Liuixalus feii Yang, Rao & Wang, 2015 (Anura: Rhacophoridae) from Vietnam and taxonomic assignment of L jinxiuensis Li, Mo, Xie & Jiang, 2015 Russian Journal of Herpetology In press Nguyen, T.V., Pham, C.T., Do, D.T., Zielger, T & Nguyen, T.Q (2018): New records and first description of females of Large-eared frog, Odorrana gigatympana Orlov, Annajeva & Ho, 2006 (Anura: Ranidae) from Vietnam Russian Journal of Herpetology In press 26 ... like amphibians (Sterling et al 2006) The research entitled "Study on the diversity and distribution patterns of amphibians (Amphibia) in karst ecosystem of the North Vietnam and conservation measures ... 3.5 Conservation issues 3.5.1 Species of conservation concern The amphibian fauna of karst forests of the North Vietnam contain a high level of conservation concern with six species listed in the. .. Contents Task 1: Inventory of the species diversity of amphibians in some karst forests in northern Vietnam: Hoa Binh Province in the northwestern region, Cao Bang and Ha Giang provinces in the