Nghiên cứu đa dạng thành phần loài và quan hệ di truyền của họ Tắc kè (Gekkonidae) ở một số khu vực núi đá vôi tại nước Cộng hòa Dân chủ Nhân dân Lào tt

MINISTRY OF EDUCATION
AND TRAINING

MINISTRY OF AGRICULTURE
AND RURAL DEVELOPMENT

VIETNAM NATIONAL UNIVERSITY OF FORESTRY

SALY SITTHIVONG

SPECIES DIVERSITY AND PHYLOGENETIC RELATIONSHIPS
OF THE FAMILY GEKKONIDAE IN LIMESTONE AREAS
FROM LAO PEOPLE’S DEMOCRATIC REPUBLIC

MAJOR: FOREST MANAGEMENT
CODE: 9 62 02 11

SUMMARY OF THE DOCTORAL DISSERTATION

HANOI, 2022


The thesis was completed at the Vietnam National University of Forestry

Supervisor:
Assoc. Prof. Dr. Luu Quang Vinh – Vietnam National University of Forestry
Prof. Dr. Nguyen The Nha – Vietnam National University of Forestry

Reviewer 1: ......................................................................................
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Reviewer 2: ......................................................................................

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Reviewer 3: ......................................................................................
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The dissertation will be defended at the Doctoral Evaluation
Dissertation Council:
- Date and time:
- Place: Vietnam National University of Forestry

The dissertation can be found at:
- National Library of Vietnam
- Library of Vietnam National University of Forestry


1
INTRODUCTION
1. Justification for the research
Knowledge on the composition of reptiles in the world during a recent decade has
increased significantly from 9,300 species in the begining of 2011 to 11,570 species in 2021
(Uetz et al. 2021). However, according to Boehm et al. (2013), an estimated 20% of reptiles
globally are threatened with extinction. Besides, the conservation of reptiles also faces many
difficulties due to the limited understanding of their diversity and bio-ecological
characteristics.
Lao People's Democratic Republic (hereafter Laos) is one of the countries with a high
diversity of flora and fauna, due to its geographical location, climate and mountainous terrain
accounts for most of the area compared to other countries. However, studies on reptiles
(Reptilia) are limited, especially the Gecko family (Gekkonidae). In recent years, species
discovery studies in the family Gekkonidae are being known with many species discovered
and described in Laos, and these findings are mainly concentrated in the limestone karst
ecosystem. However, the number of known species is still underestimated in comparision

with these species from neibouring countries such as Vietnam and Thailand, creating the
question that how many species of geckos are in Laos?
The limestone ecosystem is considered an ideal natural laboratory for the study of
taxonomy, ecology, evolution and zoology. The limestone mountain contains many different
habitats and caves, so the fauna is often highly endemic. In Laos, limestone mountains are
mostly distributed in the central region and some northern provinces (Viossanges, 2017).
As a result, this thesis focuses on karst areas of the central and northern provinces of
Laos where have been poorly studied.
For mentioned reasons, I carried out the thesis “Species diversity and phylogenetic
relationships of the family Gekkonidae in limestone areas from Lao People’s Democratic
Republic”. The study’s results have been provided a raw evidence on the diversity of species
composition, taxonomy and genetic relationships of some genera of the Gecko family,
information on the distribution characteristics of species in the Gecko family. In addtion, the
similarity relationship in species composition among the study areas and propose solutions for
Gecko conservation in Laos have been evaluated.
2. Goal and objectives
2.1. Goal
Provide scientific data on the species diversity and distribution characteristics, in order to
propose solutions for the Gecko family (Gekkonidae) conservation in the limestone areas of
Laos.
2.2. Objectives
- To determine the species diversity level of the family Gekkonidae in the study area.
- To identify species composition and genetic relationships of species Cyrtodactylus,
Dixonius and Gekko .
- To evaluate the similarity in the composition of gecko species among the study sites.
- To assess the current status of Gecko species distribution according to habitat, altitude
and location.
- To identify threat factors and conservation status of gecko species in Laos.
- To propose solutions to conserve the family Gekkonidae in the limestone ecosystem of
Laos.

3. Subject and scope
- Research subject: Species belonging to Gekkonidae.


2
- Scope: Limestone areas in some provinces of Northern and Central Laos namely:
Vientiane, Luangprabang, Khammuane, Udomxay , Huaphane and Xiengkhoang provinces.
4. Contributions
- 28 gecko species have been recorded in the study areas.
- Specific morphological characteristics for 28 collected species have been described
with specific data on Gekko species distribution in the study areas as well as genetic
relationships of the 3 genera in the Gecko family.
- Species composition similarity and distribution characteristics according to habitat,
altitude and location have been evaluated.
- Current status, threats and proposing solutions for gecko conservation in the
limestone mountains of Laos have been assessed.
5. Scientific and practical significance
5.1. Scientific significance
- Described 6 new species to science and recorded 2 new provincial records of geckos
from Udomxay Province .
- Updated information on species composition, distribution characteristics of geckos in
6 forest areas on limestone mountains in Northern and Central Laos: Vientiane,
Luangprabang, Khammuone, Udomxay, Huaphane and Xiengkhoang provinces.
- Provided information on morphological characteristics and evaluated genetic
relationships of the 3 genera Cyrtodactylus, Dixonius and Gekko .
5.2. Practical significance
The dissertation has provided information as a scientific basis for the planning and
management of biodiversity conservation in Northern and Central Laos depending on:
1) Confirming prioritized areas for conservation.
2) Identifying prioritized species for conservation.

3) Establishing crucial activities for gecko species conservation.
4. Thesis outline
The dissertation consists of 142 pages, including: Introduction 4 pages; Chapter 1:
Overview 14 pages; Chapter 2: Contents, methods and study sites 18 pages; Chapter 3:
Results and discussion 67 pages; Conclusions, existences and recommendations 3 pages;
References 10 pages. Moreover, there are 15 tables and 65 figures in this dissertation.
Chapter 1. Overview
The dissertation has overviewed and summarized about 3 main relevant issues in the world, in
Vietnam and Laos: (1). Overview of reptiles research in Laos; (2). Taxonomy and distribution
system of the Gecko family (Gekkonidae); (3). Other related studies.
1. Overview of reptiles research in Laos
The overview of the research problems has helped to have a correct and comprehensive
understanding of the situation of reptile research in Laos. Accordingly, the total number of
recorded and described reptile species in Laos is 181 species in February 2018 (Uetz et al.
2018), there is an increasing number of total reptile species from 212 species in 2020 to 225
species in May 2021.
2. Taxonomy and distribution system of the Gecko family (Gekkonidae)
A research overview provided how to identify the taxonomic features and distribution
of Gekkonidae. The outstanding taxonomic features can be summarized as: Gekkonidae
(Animalia), Vertebrata (Chordata), Reptilia and Squamata. They live mainly in warm climates
around the world. Geckos (Gekkonidae) have unique vocalizations among lizards, they use
sound to communicate with each other. Most geckos do not have eyelids but have transparent


3
membranes, which are cleaned by licking. The majority of gekko emit not only foul odors but
also droppings on their predators in self-defense. Besides, many species have suckers under
their toes that allow them to cling to tree trunks, ceilings and walls easily. In terms of species
diversity, Laos has 45 gecko species belonging to 7 genera in 2018, a described species in
2010 as Cnemaspis laoensis; the genus Cyrtodactylus is the genus with the largest number of

species in Laos. In addition, there are 58 genera of Gekkonidae with 1,430 species in the
world updated in May 2021.
3. Other related studies
Previous studies only selected a few representative species but did not include all
species of Gekkonidae as for species diversity and genetic relationships in the limestone areas
of Laos. In this study is to investigate and collect specimens in limestone mountains in
Northern and Central Laos, in order to gather available data serving to Gecko family
(Gekkonidae) diversity and genetic relationships.
Chapter 2: CONTENTS, METHODS AND STUDY SITES
2.1. Time and place of the study
The field surveys have been conducted from June 2018 to April 2020 in a total of 112
days for 7 field survey times, 13 sites, 38 survey transects . Survey efforts are shown in Table
2.1.
Table 2.1: Time and place for field trips
Stage
Location
1 Vangvieng District
2 Phuang District
Namor District
Xay District
3
Ngoi District
Luangprabang District
Phu Kut District
4
Nonghet District
5 Khunham District
6 Bualapha District
Hiem District
7 Xon District

Viengxay District

Coordinates
18°54’44” N; 102°27’05” E
18°39’21” N; 102°06’55” E
20°54’43” N; 101°45’37” E
20°41’05” N; 101°59’21” E
20°42’33” N;102°40’32” E
19°52’32” N; 102°08’36” E
19°34’25” N’;103°04’55” E
19°29’52” N; 103°59’05” E
18°12’25” N;104°31’36” E
17°29’03” N; 105°35’24” E
20°04’59” N; 103°22’10” E
20°27’25” N;103°20’57” E
20°25’08” N;104°13’49” E

Altitude (m)
230-684
232-675
620-827
642-782
348-702
298-439
1.080-1.205
1.105-1.397
168-524
170-214
687-1.270
715-926

698-985

Time
25/7-9/8/2018
23/10-6/11/2018
21/4-28/4/2019
29/4-6/5/2019
7/5-13/5/2019
14/5-20/5/2019
10/7-16/7/2019
17/7-23/7/2019
26/10-2/11/2019
25/2-3/3/2020
10/3-16/3/2020
17/3-23/3/2020
26/3-2/4/2020

2.2. Research contents
- Investigation of species diversity of Gekkonidae in limestone mountain areas of
Northern and Central Laos: karst mountains and caves in Vientiane, Luangprabang,
Khammuone, Udomxay, Huaphane and Xiengkhoang.
- Determination of species composition and evaluation of genetic relationships between
species and populations in the Bent-toed Gecko (Cyrtodactylus), Leaf-toed Gecko (Dixonius)
and Gecko (Gekko) recorded in Laos.
- Comparison of the similarity in species composition of the gecko family among the
study sites and between the North and the Central region.
- Evaluation of the species distribution characteristics in the Gecko family by altitude
ranges, habitat types, by micro-habitat (location: on trees, on the ground, cliffs).
- Assessment of threat factors to populations of Gekkonidae species at the study sites
and solutions for conservation.

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2.3. Materials and Methods
2.3.1. Field survey
Point and transect surveys have been applied in this study. To be more specific, the
transect surveys were made based on the topographic map, vegetation and habitat of the
Gecko species. These transects were designed through different habitat types and elevations
of the study area, paying special attention to the limestone mountains with caves and cliffs,
the valleys between the limestone mountain ranges in the forest. Each transect survey has be
recorded by GPS using trackmaker. Survey time was at night from 17h00 to 23h00.
2.3.2 Research materials
1) Chemicals
The chemicals used to separate the total DNA including: Dneasy Blood Kit and Tissue
(Qiagen, Germany); GenJet Genomic DNA Purification (ThermoFisher Scientific,
Lithuania); ethanol (Merck, Germany).
PCR reactions were performed using a mixture of HotStar Taq mastermix (Qiagen,
Germany) and DreamTaq Mastermix (ThermoFisher Scientific, Lithuania). PCR products
were visualized by electrophoresis using the following reagents: agarose, ethidium bromide,
tris base, EDTA, marker 1 kb, marker 100 bp (1st Base, Malaysia) and dye 6x (ThermoFisher
Scientific, Lithunia). Successful PCR products were purified using the GeneJET PCR
Purification Kit (ThermoFisher Scientific, Lithuania).
2) Primers for PCR reactions
Primers used in the genetic relationship study of the Bent-toed Gecko (Cyrtodactylus),
the Leaf-toed Gecko (Dixonius) and the Gecko (Gekko) were referenced to the previous study
of Ivanova et al. 2006, Macey et al. 1997 and Greenbaum et al. 2007. The catalyst process
was shown in tables 2.3-2.5.
Table 2.3. Primers used in studying genetic relationships of Bent-toed Gecko (Cyrtodactylus)
Primer name
Primer sequences

Source
VF1-d
5’- TTCTCAACCAACCACAARGAYATYGG -3’ Ivanova et al. 2006
VR1-d
5’- TAGACTTCTGGGTGGCCRAARAAYCA -3’ Ivanova et al. 2006
Table 2.4. Primers used in studying genetic relationships of Leaf-toed Gecko (Dixonius)
Primer name
Primer sequences
Source
MetF1
5’- AAGCTTTCGGGCCCATACC -3’
Macey et al. 1997
COIR1
5’- AGRGTGCCAATGTCTTTGTGRTT -3’
Macey et al. 1997
Table 2.5. Primers used in studying genetic relationships of Gekko genus (Gekko)
Primer name
Primer sequences
Source
GF1
GR1

5'- CAAGCACHATYATYACYATAT -3'
5'-CCTATGTGTGCGATTGATGA-3’

Greenbaum et al. 2007
Greenbaum et al. 2007

3) Bioinformatics softwares
The bioinformatics software used in the study included: Sequencher v5.4.6 (Gene Codes

Corp., AnnArbor, MI, USA), ClustalX v2.1 (Thompson et al., 1997), jModeltest v2.1.4
(Darriba et al., 2012), Modeltest v3.7 (Posada and Crandal, 1998), MrBayes v3.2 (Ronquist
et al., 2012), Tracer v1.5 (Rambaut and Drummond, 2009), Figtree v1.3 (Rambaut, 2009),
PAUP v4.0b10 (Swofford, 2001).
4) Lab equipment
Research equipment for genetic analyses belong to the Laboratory of Genetics
Department, Ha Noi University of Natural Sciences at Vietnam National University; Wildlife
Department in Faculty of Forest Resources and Environmental Management at Vietnam
National University of Forestry.
5) Field equipment


5
Equipment for field investigation: GPS, digital caliperto the nearest 0.1 mm, monitoring
sheet, thermohygrometer, digital camera, headlight, measuring tape, etiket, lighter, marker,
alcohol, tongs, DNA sample tube, gloves.
2.3.3. Research specimens
Sampling method: Samples are collected by hand and specialized tools such as rubbercoated clamps are used to avoid harming the animals. After measuring, photographing,
collecting DNA samples, presentative samples will be kept as research specimens.
Specimen making: Specimens were anaesthetized and euthanized in a closed vessel
with a piece of cotton wool containing ethyl acetate (Simmons, 2002) and fixed in
approximately 85% ethanol, then later transferred to 70% ethanol for permanent storage.
Sampling for DNA analyses: Muscle, liver or tail tissue samples were collected and stored
separately in 70% alcohol (Merk, Germany) at Wildlife Department, Faculty of Forest
Resources and Environmental management at Vietnam National Umiversity of Forestry.
2.3.4. Analysis of morphological characteristics and Gecko identification
After collection, specimens were brought to the laboratory of the Vietnam National
University of Forestry for analysis. The morphological measurements were taken with an
Etopoo digital caliper to the nearest 0.1 mm. Scale counts were taken using a stereo
microscope (Olympus SZ61). Measurements and scale counts followed Nguyen (2013), Luu

(2015), and Ziegler (2016)
Table 2.6. List of abbreviations and explanations of measurement and scale count
characters
Abbreviation
Explanation
Measurements (mm)
1
SVL
2
TaL
3
TrunkL/AG
4
HL
5
HW
6
HH
7
SE
8
EyeEar
9
ForeL
10
FemurL
11
CrusL
12
LD4A

13
LD4P
14
OD
15
EarL
16
RW
17
RH
18
MW
19
ML
Scale counts (1, 2, 3, ...)
20
CS
21
N
22
I/IN
23
SL
24
IL
25
IO

Snout-vent length
Tail length

Trunk length
Head length
Maximum head width
Maximum head height
Snout to orbit distance
Orbit to ear distance
Forearm length
Femur length
Crus length
Length of finger IV
Length of toe IV
Greatest diameter of orbit
Ear length
Maximum rostral width
Maximum rostral height
Maximum mental width
Maximum mental length
Ciliary spines
Nasals (nasorostrals,supranasals, postnasals)
Intersupranasals
Supralabials
Infralabials
Interorbitals


6
Abbreviation
26
27
28

29
30
31
32
33
34
35
36
37
38
39

PO
PM
GP
DTR
GSDT
SBL
SR
V
LF1
LF4
LT1
LT4
PP
PAT

Explanation
Preorbitals
Postmentals

Gulars bordering the postmentals
Dorsal tubercle rows at midbody
Granular scales surrounding dorsal tubercles
Number of scales along the midbody
Number of scale rows around midbody
Ventral scales in longitudinal rows at midbody
Subdigital lamellae under whole first finger
Subdigital lamellae under whole fourth finger
Subdigital lamellae under whole first toe
Subdigital lamellae under whole fourth toe
Precloacal pores
Postcloacal tubercles

After analyzing morphological data, detailed identifications of gecko specimens were
followed Smith (1935), Taylor (1963), Dao (1979), Nguyen et al. (2010, 2011), Hartmann et
al. (2013), Ziegler et al. (2013, 2016), Nguyen et al. (2014), Luu et al. (2014, 2015, 2016),
Vassilieva et al. (2016), Schneider et al. (2020) and other documents.
Furthermore, when the process of classifying samples finished, specimens were
subsequently deposited in the collections of the Vietnam National University of Forestry
(VNUF), the National University of Laos (NUOL), the Institute of Ecology and Biological
Resources (IEBR), and the Zoologisches Forschungsmuseum Alexander Koenig, Bonn,
Germany (ZFMK).,
Systematics and nomenclature followed Nguyen et al. (2009) and Uetz et al. (2011) and
some recently published documents are main references for the classification common names
of taxonomic levels.

Figure 2.3. Measuring and counting indicators for gecko specimens


7

2.3.5. DNA extraction and sequencing
Tissue samples were stored at 4°C prior to extraction. The tissue used for extraction is
taken from deep within the specimen mass to limit the risk of contamination. The Dneasy
Blood and Tissue Kit (Qiagen, Germany) is used for low-volume or long-collected samples
and stored under uncertain temperature, solution conditions and GeneJet Genomic DNA
Purification (ThermoFisher Scientific, Lithuania) for newly collected tissue samples, large
amount of samples stored in guaranteed conditions (Merck alcohol 70%, Germany). The
extraction was performed according to the manufacturer's instructions, with process-based
adjustments. The extraction process is carried out according to the following specific steps:
sample pretreatment (cut the sample into small pieces, allow to dry and put in 1.5 ml
eppendorf tube); membrane disruption and protein removal (using buffer ATL, AL-Qiagen,
Germany; Digestion Solution, Lysis solution-ThermoFisher Scientific, Lithuania và protein
K-Qiagen, Germany); DNA precipitation (using Merck alcohol 50-100%, Germany);
separation of DNA from other cell components (filter column containing a silica membrane);
DNA purification (using buffer AW1, AW2-Qiagen, Germany; Wash 1, Wash 2ThermoFisher, Lithuania); DNA dissolution (buffer AE-Qiagen, Germany; Elution BufferThermoFisher Scientific, Lithuania). After extraction of total DNA, the obtained total DNA
concentration was checked by electrophoresis on 1% agarose gel, 1X TBE buffer (Tris base,
Boric acid, EDTA pH 8) at 70V for 30 min. Total DNA was compared with a 1 kb marker
and then visualized by ultraviolet light on an Alphamager MINI (Protein Simple, USA).
PCR reaction was performed to diffuse the COI gene fragment of the mitochondrial
genome with primer pairs. In this study, with purpose for the next evolution research, the
priority is to use the mastermix finished-product in order to save time, optimize the conditions
for the PCR reaction and save the meager sample resources to be able to conduct amplification
of gene fragments. Two mastermixes finished-products were used in the study namely
HotStarTaq Mastermix for samples with low DNA concentrations and DreamTaq Mastermix
for amplification of samples with high DNA concentrations.
The total volume of each PCR reaction was 21 µl, consisting of 1-2 µl of DNA template
depending on the quality of DNA in the final extraction solution, 2 µl of each primer (10
µM/µl), 5 µl water, 10 µl of Taq mastermix. PCR conditions were: 95oC for 15min for Qiagen
mastermix and 5’ for ThermoFisher mastermix; followed by 35 cycles, at 95oC in 30s, 48oC60oC in 45s, 72oC in 1min; final elongation step at 72oC in 6min. Negative controls, each
extraction and each PCR were carried out at the same time.

PCR products were visualized by electrophoresis through a 1% agarose gel, 2pg/ml
ethidium-bromide, in 1X TBE buffer (Tris base, Boric acid, EDTA pH 8) at 90V for 30min.
After that, I was visualized by ultraviolet light on an Alphamager MINI (Protein Simple,
USA). Successful PCR products were purified using the GeneJET PCR Purification kit
(ThermoFisher Scientific, Lithuania). The procedure was performed according to the
manufacturer's instructions including the following steps: attaching DNA to the membrane
(using Binding Buffer and silica membrane filter column containing); clean DNA (using 350
µl Wash Buffer); Dissolve the DNA (using 30 µl of Elution Buffer). After purification, PCR
products were stored at -4 oC and then sent for two-dimensional sequencing at First Base
company (Malaysia).
2.3.6. Building phylogenetic tree
The obtained sequences were aligned in Sequencher v5.4.6 software (Gene Codes Corp,
Ann Arbor, MI, USA). The obtained sequences were checked using the Basic Local
Alignment Search Tool (BLAST) available on the website of the National Center for
Biotechnology Information. The sequences were then solved together with those from the


8
Gen Bank (Genbank). After the sequences were aligned using Clustal X v2 software
(Thompson et al. 1997), data were analyzed using maximum parsing (MP) as implemented in
PAUP* 4.0b10 (Swofford 2001), maximum likelihood (ML) as implemented in IQ-TREE
v1.6.7.1 (Nguyen et al. 2015), and Bayesian Inference (BI) as implemented in MrBayes v3
.2.7 (Ronquist et al. 2012).
The settings for MP and BI analyses followed Le et al. (2006), except that the number
of generations in Bayes analysis was increased to 1×107. Both BI and ML are run using a
best-of-breed model, TVM+I+G, as chosen by jModelTest v2.1.4 (Posada 2008). The limition
for memory function is 54 in Bayes analysis because the -lnL score reached a stable level
after 54,000 generations in two times runs. Nodal support was evaluated using 1,000 bootstrap
replication (BP) as calculated in PAUP, 10,000 superfast bootstrap clones (UFB) in IQ-TREE
v1.6.7.1 and posterior probabilities (PP) in IQ-TREE v1.6.7.1. MrBayes v3.2. BP ≥ 70 and

PP, UFBP ≥ 95% were considered strong support for a clade (Hillis & Bull 1993; Ronquist
et al. 2012; Nguyen et al. 2015). The uncorrected pairwise distance (p) were calculated in
PAUP* 4.0b10.
2.3.7. Statistical analysis
The study used MS-Excel software in Microsoft Office 2010 to analyze data and
compare the similarity in species composition between provinces with similar habitats in the
North and Central regions based on data collected during the field survey and combined with
the reference of published works. In addition, in the study, the researcher also compared the
correlation of species composition between regions, and used PAST Statistic software
(Hammer et al. 2001) for statistical analysis.The data was coded in symmetric form. (1:
Present; 0: Not present). The Sorensen-Dice index was calculated as follows:
djk = 2M/(2M+N)
M: the number of species recorded in two regions; N: the total number of species
recorded in a region.
2.3.8. Gecko distribution characters
1). Gecko species distribution along habitat types
Based on the classification of vegetational cover types and the level of human impact
on the vegetation according to the document “Guideline for biodiversity investigation and
monitoring” (Pham Nhat et al. 2003), and based on the current status of limestone forest in
the studied area, the distribution of gecko species is divided into 3 main habitat types
including:
- Limestone habitat in farming area (SC1): An area where people use land to grow
agricultural and industrial crops or seasonal crops, including interlaced limestone ranges here
(Fig. 2.4).
- Limestone habitat in secondary forest (SC2): A forest where it has been affected or not
developed with few big trees, poor forest type, in which there is a distribution of limestone
mountains (Fig. 2.4).
- Limestone habitat in primary forest (SC3): A forest that is low-impact rich with many
big trees (Fig. 2.4).

9
Figure 2.4. Types of habitat distribution A: (SC1); B: (SC2); C: (SC3)

2). Gecko species distribution along altitudinal gradients
According to Bain & Hurley (2011), there are 2 kinds of altitude belts below 800 m and
above 800 m in Indochina based on natural conditions. In principle, this research still followed
the provided method to collect data for comparison.
Nevertheless, the actual characteristics of the study area showed that habitat distribution
types in many high-altitude belts, the geographical topography from both plains and hills, and
the level of human impacts are quite similar. To be more specific, altitude is categorized by
each level of 200m including: less than 200m, from 200 to under 400m, from 400 to under
600m, from 600 to under 800m, from 800 to under 1,000m and over 1,000m.
3). Gecko species distribution along habitat types
To assess the species habitat of Gekko (Gekkonidae), this research based on
encountered points to divided into 3 locations:
- Cliff: slits, holes, small caves (Fig. 2.5).
- On tree: all positions of tree species such as trunks, branches or vines... (Fig. 2.5).
- Ground: slits, holes, ground surface, ravines of mountain foot and cliffs close to the
ground (Fig. 2.5).

Figure 2.5.Encounter points, A: Cliff; B: Ground; C: On tree.
2.3.9. Conservation status assessment
Threats to the gecko fauna have been identified based on field observation. Habitat loss
is directly threatened through quarrying of limestone and converting forest to agriculture.
There are some scientific and legal documents for gecko fauna conservation of Laos,
including the International Union for Conservation of Nature's Red List (IUCN, 2021); Law
on Wildlife Protection No. 07/QH Laos dated 24/12/2007 Lao Government Decree No.
08/CP/2021, Appendix CITES (2021). Priority areas for conservation are identified through
scoring and overlapping assessment classes. Five criteria are used for identifying priority

areas for conservation (Nguyen Quang Truong et al. 2011): (1) species richness, (2) the
number of rare/or threatened species, (3) habitat quality, (4) levels of human disturbance, (5)
prioritized levels of protected areas. In each criterion, a higher number shows a better score
(ranking score from 1 to 13). High rank areas are identified as priorities for conservation.
.


10
Chapter 3. RESULTS AND DISCUSSION
3.1. Diversity of species composition of the Gecko family (Gekkonidae)
Based on morphological and/or molecular analyses of 138 gecko specimens,28 gecko species
of 6 genera have been identified in the study areas consisting of 9 species of the Bent-toed
gecko (Cyrtodactylus); the Leaf-toed Gecko (Dixonius), House gecko (Hemidactylus), and
Half leaf-fingered geckos (Hemiphyllodactylus) each of these has 4 species; and 3 Web-toed
geckos (Gehyra) (Table 3.1).
Table 3.1. List of Geckos (Gekkonidae) species recorded in study sites
TT
I
1
2
3
4
5
6
7
8
9
II
1
2

3
4
III
1
2
3
4
IV
1
2
3
V
1
2
3
4
VI
1
2
3
4

Name

Scientific name

Cyrtodactylus
Cyrtodactylus houaphanensis***
Cyrtodactylus interdigitalis
Cyrtodactylus muangfuangensis***

Cyrtodactylus ngoiensis***
Cyrtodactylus pageli
Cyrtodactylus teyniei
Cyrtodactylus wayakonei*
Cyrtodactylus sp.1
Cyrtodactylus sp.2
Leaf-toed Gecko Dixonius
Dixonius lao***
Dixonius siamensis
Dixonius somchanhae***
Dixonius sp.
Gecko
Gekko
Gekko aaronbaueri
Gekko kabkaebin
Gekko khunkhamensis***
Gekko gecko
Web-toed geckos Gehyra
Gehyra mutilata
Gehyra sp.1
Gehyra sp.2
House geckos
Hemidactylus
Hemidactylus bowringii
Hemidactylus frenatus
Hemidactylus ganotii
Hemidactylus platyurus
Half leaf-fingered
Hemiphyllodactylus
geckos

Hemiphyllodactylus kiziriani*
Hemiphyllodactylus fc.serpispecus
Hemiphyllodactylus sp1.
Hemiphyllodactylus sp2.

VT

Study sites
LPB UDX XK

HP KM

Bent-toed Gecko

+
+
+
+
+
+
+
+
+
+
+
+
+

+

+

+

+
+

+

+

+

+

+
+
+
+
+

+
+
+
+
+

+
+
+

+

+

+

+

+

+

+
+
+
+

+
+
+
+

+
+
+
+

Notes: (*): new species recorded for the province; (**): new species for science; (sp.):
undetermined species; (cf.): closely related species. Research location: VC= Vientiane, LPB=

LuangPrabang, UDX=Udomxay, XK= Xiengkhoang, HP = Huaphane, KM = Khammuone.

Diversity of species: The genus Cyrtodactylus has the most diverse number of species
with 9 species, the genera as Leaf-toed Gecko (Dixonius), Gecko (Gekko), House geckos


11
(Hemidactylus), Half leaf-fingered geckos (Hemipyllodactylus) have 4 species of each and
Gehyra has 3 species.
3.1.1. New species discoveries.
- New species to science: Within the framework of this research, 06 new species have
been described as following:
1). Cyrtodactylus muangfuangensis Sitthivong, Luu, Ha, Nguyen, Le & Ziegler, 2019.
The phylogenetic tree also shows the new species as a sister species to C. pageli, this
species is also found in Vientiane province with detection sites about 50 km apart,
morphologically, the new species may be clearly differentiated from C. pageli in the number
of ventral and anterior foraminal scales in both sexes. In addition, the genetic difference
between C. muangfuangensis and C. pageli was 18% based on a mitochondrial COI gene
fragment (Fig. 3.1).

Figure 3.1. New species standard specimen Cyrtodactylus muangfuangensis
(VNUF R.2018.32) male. Photo: S. Sitthivong.
2). Cyrtodactylus houaphanensis Schneider, Luu, Sitthivong, Teynié, Le, Nguyen &
Ziegler, 2020.
Cyrtodactylus houaphanensis differs from all species in the genus, in the C. wayakonei
group having at least 3.3% genetic variation in the COI gene. This new species is
morphologically similar to C. chauquangensis and is a sister taxon to C. puhuensis by genetic
analysis, and differs from the latter species by the absence of femoral fores (Fig. 3.2).

Figure 3.2. New species standard specimen Cyrtodactylus houaphanensis

(IEBR A.2013.109) Male specimen. Photo: A. Teynie.
3). Cyrtodactylus ngoiensis Schneider, Luu, Sitthivong, Teynié, Le, Nguyen & Ziegler, 2020.
Cyrtodactylus ngoiensis differs from other closely related congeners by at least 11.6%
genetic variation in COI genes. The new species is believed to be a member of the C.
wayakonei species group, but is morphologically close to C. dumnuii from Thailand (Fig.
3.3).


12
Figure 3.3. New species Cyrtodactylus ngoiensis. A: Male specimen (IEBR 4548);
B: Female model (IEBR A.2013.110). Photos: A. Teynié and T.Q. Nguyen.
4). Dixonius lao Nguyen, Sitthivong, Ngo, Luu, Nguyen, Le & Ziegler, 2020.
In phylogenetic analyses, the new species was shown to be a sister taxon with two
undescribed taxa from Thailand but differing by at least 8.6% in pairwise genetic distances.
with, the second is based on the complete sequence of the mitochondrial ND2 gene with
partial or complete sequencing of six adjacent tRNAs (Fig. 3.4).

Figure 3.4. New species standard specimen Dixonius tuberculosis A: Male specimen (VNUF
R.2016.2); B: Female model (IEBR A.2019.6). Photos: L. Q. Vinh

5). Dixonius somchanhae Nguyen, Luu, Sitthivong, Ngo, Nguyen, Le & Ziegler, 2021.
Genetically, the new species and the closely related species D. siamensis
areapproximately 9.4% divergent from each other based on the complete mitochondrial
ND2 gene with six partial or complete adjacent tRNAs. It is the 12th known species of the
genus Dixonius and the second Dixonius species to be described from Laos. (Fig. 3.5).

Figure 3.5. New species Dixonius somchanhae A: Standard male specimen (VNUF R.2020.3);
B: Male specimen (VNUF R.2020.2). Photos: L. Q. Vinh.
6). Gekko khunkhamensis Sitthivong, Lo, Nguyen, Le, Ngo, Khotpathoom, Ziegler & Luu, 2021.

This area is adjacent to Quang Binh province, Vietnam, where G. scientiadventura was
discovered. New species differs from its congeners by at least 13% in pairwise distances based
on a segment of the mitochondrial ND2 gene. (Fig. 3.6).

Figure 3.6. New species Gekko khunkhamensis (A) Male specimen; (B) Female
specimen. Photos: S. Sitthivong
3.1.2. New provincial records
1) Cyrtodactylus wayakonei Nguyen, Kingsada, Rosler, Auer & Ziegler, 2010.

This species was originally described from Luang Nam Tha Province in Northern Laos
in 2010, and in 2011 it was reported in Yunnan Province, China. In this study, Cyrtodactylus
wayakonei was encountered in Na Mo district, Udomxay Province, northern Laos. The new
sample has sequences close to the original description.


13

Figure 3.7. (A) The holotype (IEBR A.2010.01); (B) New recorded specimen (VNUF
R.2021.50). Photos: T.Q. Nguyen and S. Sitthivong.
2) Hemiphyllodactylus kizirianii Nguyen, Botov, Le, Nophaseud, Zug, Bonkowski & Ziegler,
2014.

Originally described in Luangprabang province in Northern Laos in 2014, in this
study, the species Hemiphyllodactylus kiziriani was encountered in Namo district, Udomxay
province, northern Laos. The new sample has sequences close to the original description.

Figure 3.8. (A) The holotype (IEBR A.2014.3); (B) New recorded specimen (VNUF R.2021.52).
Photos: T.Q. Nguyen and S. Sitthivong.

3.1.3. Species morphological characteristics have not been identified

In this study, 07 species have not been identified yet and are in the stage of molecular
analysis, the species morphological characteristics are as followed:
1). Cyrtodactylus sp.1: Due to the lack of genetic comparison, the species has not been
temporarily identified .

Figure 3.9. Unidentified specimen Cyrtodactylus sp.1 (A) Dorsolateral view of a female
specimen; (B) A female specimen is molting. Photos: S. Sitthivong.

2). Cyrtodactylus sp.2: Due to the lack of genetic comparison, the species has not been
temporarily identified .

Figure 3.10. Unidentified specimen Cyrtodactylus sp.1 (A) Dorsolateral view of a male
specimen; (B) Dorsal view of afemale specimen. Photos: S. Sitthivong

3). Gehyra sp.1: Because there has not been a genetic comparison, the species has not been
accurately identified.


14

Figure 3.11. Unidentified specimen Gehyra sp.1 (A) Dorsal view;
(B) Ventral view. Photos: S. Sitthivong.

4). Gehyra sp.2: Because there has not been a genetic comparison, the species has not been
accurately identified.

Figure 3.12. Unidentified species Gehyra sp.2 (A) Dorsal side;
(B) Ventral side. Photo source: Saly Sitthivong.

5). Dixonius sp. Because there has not been a genetic comparison, the species has not been

accurately identified.

Figure 3.13. Unidentified species Dixonius sp. (A) Dorsal view;
(B) Ventral view. Photos: S. Sitthivong.

6). Hemiphyllodactylus sp.1: Due to the lack of genetic comparison, the species has not
been temporarily identified.

Figure 3.14. Unidentified species Dixonius sp. (A) Dorsal view;
(B) Ventral view. Photos: S. Sitthivong.

7). Hemiphyllodactylus sp.2: Because there has not been a genetic comparison, the species
has not been accurately identified.

Figure 3.15. Unidentified species Hemiphyllodactylus sp. (A) Dorsal view;
(B) Ventral view. Photos: S. Sitthivong.


15
3.1.4. Morphological characteristics of described gecko species in study sites
1). Cyrtodactylus interdigitalis Ulber, 1993. Head covered with small scales; prison muzzle;
the eye is covered by the third eyelid; eardrum round, shallow, smaller than eye diameter;
supralabials scales 9/9, infralabials scales 8/10; small, smooth dorsal scales; dorsal tubercle
rows at midbody 16, row of ventral scales 34, precloacal pores absent in female.
2). Cyrtodactylus pageli Schneider et al., 2011. Head covered with small scales; prison
muzzle; the eye is covered by the third eyelid; eardrum round, shallow, smaller than eye diameter;
supralabials scales 7-12, infralabials scales 7-9; dorsal tubercle rows at midbody 8-11, row of
ventral scales 41-48, number precloacal pores 4-6 for (♂) and 4-6 for (♀), femoral foramen
scale unknown; subdigital lamellae under whole fourth finger 19-23 and fourth toe 20-24.
3). Cyrtodactylus teyniei David et al., 2011. Some morphological features: supralabials

scales 9-13, infralabials scales 9-11; row of convex tubers on dorsal 17-22, row of scales
surrounding body 112-138, row of ventral scales 34-42, number of scales precloacal pores:
8-14 for (♂) and 7-14 for (♀); subdigital lamellae under whole fourth finger 17-20 and
fourth toe 18-22; dorsal and limbs brown.
4). Dixonius siamensis Boulenger, 1899. Some morphological features: rows of ventral
scales 22-26, supralabials scales 7-8, infralabials scales 6-9; row of dorsal convex tubercles
20-24, number precloacal pores scales: 7-8 for (♂) and absent in (♀); subdigital lamellae
under whole fourth finger 10-12. Body small and slender, dark spots on body small and not
round, rows of scales running along the body in alternating depressions.
5). Gehyra mutilata Wiegmann, 1834. Some other common features: supralabials scales 610, infralabials scales 6-9; plates around chin plate 8-11, row of scales around body 101-134,
row of ventral scales 36-43, precloacal pores scales: 36-42 for (♂) and absent in (♀);
subdigital lamellae under whole fourth finger 6-8 and fourth toe 6-9, scale under caudal
extended. Specimen with identifiable characteristics consistent with Taylor's (1963)
description: Scaled head; prison muzzle; the eye is covered by the third eyelid; round eardrum.
6). Gekko aaronbaueri Ngo et al., 2015. Morphological characteristics: Head covered with
small scales; prison muzzle; the eye is covered by the third eyelid; eardrum round, shallow,
smaller than eye diameter; dorsal form with five to six dirty white vertebral spots interspersed
with yellow, W-shaped spots between nape and spine and six to seven pairs of dirty white
spots interspersed with dark spots with yellowish margins on the flanks between the
extremities, the scales under the tail are extended.
7). Gekko kabkaebin Grismer et al., 2019. Morphological features: Body has dark brown
segments, separated by distinct wave-shaped black thread, light brown along the back. The
head is gray-brown, the nape has a W-shaped black streak. The two tail edges are serrated,
the eyes are black, the black line connects from the side of the neck running across the eye to
the nose, the toes are connected by membranes.
8). Gekko gecko Linnaeus, 1758. Pale greenish-gray back with yellow or bright red spots,
with many nodules. Males are more colorful than females. The abdomen is white or gray with
many small yellow dots. The five-toed toe has claws and thin skin that forms a grip. The eyes
have vertical pupils, can expand in the dark, and have very good focus. Living alone, only
finding each other during the mating season.

9). Hemidactylus bowringii Gray, 1845. On the dorsal and lateral surfaces, dark markings
form thin longitudinal stripes with the lateral line most clearly defined; This latter stripe
extends from the back of the eye to the shoulder, then is broken by circular cream dots, and a
solid stripe at the base of the tail. The dorsal stripe is always fragmented; begins on the front
neck and extends to the base of the tail.


16
10). Hemidactylus frenatus Duméril & Bibron, 1836. Usually gray or light brown to beige
with green iridescence and white underside. They also appear semi-transparent at times. Their
scales are usually uniform on the front, but increase in size along the back, and the large spiny
scales are arranged in bands around the tail. Has vertical pupils and is sensitive to darkness.
supralabials scales 9/9, infralabials scales 8/8, number of Subdigital lamellae under whole
fourth finger 13, under fourth toe 15.
11). Hemidactylus ganotii Duméril & Bibron, 1836. On the body is dark brown, tiny white
dots on the body form a vertical line with the body. The pupils of the eyes are brown and there
is a black thread along with the eye, and the belly is yellow. The flanks have alternating brown
and white dotted stripes extending from the base of the hind thigh to the base of the front
thigh, the scales under the tail are extended.
12). Hemidactylus platyurus (Schneider, 1797). Morphological characteristics of the
specimen are consistent with the previous description: Body length 50.1-59.4 mm, tail length
43.3-55.7 mm. This gecko is easy to identify by the skin folds on the body from the armpit to
the groin that is very clearly frilled, about 2mm wide at the widest place; the back of the thighs
and shins also have narrow frills; body flattened, muzzle slightly pointed, head longer than
wide, slightly differentiated from neck; round eye hole; triangular chin plate, with 10-11
supralabials scales and 8-9 infralabials scales .
13). Hemiphyllodactylus cf. serpispecus Eliades et al. 2019. On the dark brown body, the
black rings alternate in the middle, unevenly, the upper part of the middle of the tail base and
the waist has brown marks. The tail has brown patches interspersed with distinctly small black
bands. On the brown head, there are alternating black dots, with brown dotted stripes running

from the neck to the eyes and over the eyelids.
3.2. Genetic relationship of some genera in the Gecko family in Laos.
3.2.1. Genetic relationship of the Bent-Toed Gecko (Cyrtodactylus)
Most species of the genus Cyrtodactylus are distributed in limestone forests. They are
found on cliffs, rarely seen on the ground, there are a few species that are related to trees such
as Cyrtodactylus interdigitalis and Cyrtodactylus buchardi

Figure 3.30. Genetic tree of species in the genus Cyrtodactylus. (Analysis (>50%), (-)
unresolved, (*) represents a value of 100%).


17
The alignment contained no gaps. MP analysis of the dataset recovered seven most
parsimonious trees with 729 steps (CI = 0.5; RI = 0.78). In the ML analysis, the -Ln likelihood
score of the best tree found was 3920.996. The topology derived from the BA was similar to
those of Nguyen et al. (2017) and Brennan et al. (2017). Through the study, the genetic
relationships of the genus Cyrtodactylus in Laos is divided into 3 groups (Schneider et al.
2020) such as: The first group is C. wayakonei distributed in the North including 5 species,
the second group is C. wayakonei distributed in the North, including 5 species, the second
group is C. phongnhakebangensis distributed in the Central region includes 17 species and
the last group is C. irregularis distributed in the South including 3 species.
3.2.2. Genetic relationship of the genus Gekko
Species of the genus Gekko are mainly distributed in limestone forests, often found on cliffs,
only Tokay Gecko (Gekko gecko) species can live in many places including cliffs, on trees.
Farming areas are also available in people's houses from North to South.

Figure 3.32. Phylogenetic tree of species in the genus Gekko.
(-) unresolved node, (*) represents a value of 100%.

The alignment contained no gaps. MP analysis of the dataset recovered seven most

parsimonious trees with 1,296 steps (CI = 0.45; RI = 0.63). The topology derived from the
BA was similar to those of Nguyen et al. (2017) and Brennan et al. (2017) (Fig. 3.32).
According to Wood et al. (2021), the genus Gekko is divided into 7 groups of Gecko (Gekko)
in the world, including: Gekko (Gekko) gecko, Gekko (Japonigecko) japonicus, Gekko
(Ptychozoom) homalocephala, Gekko (Phacogecko) rhacophorus, and Gekko (Sundagecko)
vittatus, group Gekko (Pseudosundagecko) gulat and group Gekko (Archipelagecko)
mindorensis. In my research on the genus Gecko (Gekko) discovered 4 species, 1 new species
for science, in 3 groups including: group Gekko (Gekko) gecko with species Gekko gecko,
group Gekko (Japonigecko) japonicus with species Gekko aaronbaueri and Gekko
khunkhamensis, the group Gekko (Ptychozoom) homalocephala includes the species Gekko
kabkaebin.


18
3.2.3. Phylogenetic relationship of Leaf-toed Gecko (Dixonius)
Species of the Dixonius are mainly distributed in secondary forests, cultivated areas, near
populations with limestone mountains or large rocks on the ground, most species in this breed
all live on the ground or cliffs close to the ground.

Figure 3.33. Phylogenetic tree of species in the genus Dixonius.
((-) unresolved node, (*) represents a value of 100%).
The alignment contained no gaps. MP analysis of the dataset recovered seven most
parsimonious trees with 1,433 steps (CI = 0.70; RI = 0.65). The topology derived from the
BA was similar to those of Nguyen et al. (2017) and Brennan et al. (2017). Molecular data of
D. somchanhae from Laos, presented as sister taxon to D. siamensis from Laos, Thailand and
Vietnam with statistically significant values obtained from MP and BI analysis (Fig. 3.33).
3.3. Evaluation of similarity in the family Gekkonidae composition in study sites
3.3.1. Among study sites
Sorensen-Dice indexes among study sites are shown in Table 3.5.
Table 3.5. Similarity index (Sorensen-Dice) composition among study sites

Locations
VC
LPB
UDX
XK
HP
KM
1
VC
0,454
1
LPB
0,300
1
UDX
0,666
0,600
1
XK
0,222
0,750
0,461
0,555
0,500
0,428
1
HP
0,444
0,421
0,352

0,521
1
KM
0,266
Note: VC= Vientiane Province, LPB= Luong Pha Bang Province, EK= Xieng Khuang
Province, UDX= Udomxay province, HP= Huaphane Province and KM= Khammuone province.

Comparing the Sorensen-Dice index in Table 3.5, we see that Xiengkhoang province and
Udomxay province have the highest similarity in gecko species composition with (djk = 0.750),
followed by Luong Pha Bang and Udomxay province has a similar level of species composition
of gecko species (djk = 0.666) because Udomxay province, Luangprabang province and Xieng
Khuang province are all in the North with similar geographical borders, karst formations and
ecosystems with limestone mountains and covered soil with many trees and forests.


19
3.4. Distribution characteristics of Gecko species.
3.4.1. Habitat distribution
Based on the current status of karst forests in the study area, the distribution of
gecko species in 3 main habitat types has been assessed: (SC1) Limestone habitat in the
cultivation area; (SC2) Limestone karst habitat in secondary forest; (SC3) Limestone
mountain habitat in primeval forest. The results are shown above (Figure 3.37).
Hemiphyllodactylus

Hemidactylus
Gehyra
Gekko
Dixonius
Cyrtodactylus
0

5
10
15
20
Limestone habitat in the cultivation area

25

Limestone karst habitat in secondary forest
Limestone mountain habitat in primeval forest

Figure 3.37. Gecko species diversity and varieties based on habitat in study sites
3.4.2. Altitude belt distribution
Based on the collected, encountered and recorded specimens that have been
synthesized in in figure 3.38.
Species
7
6

6
5
4

4

4

4

4

4
33

3

3

3
2

2

2

2

2
1

11

1

1

1

1

1

1

1

1
0
0
0
Altitude (m) <200
Cyrtodactylus

0
200-400
Dixonius

Gekko

0

0

400-600

600-800

Gehyra

0

Hemidactylus

0

0

800-1000

00

0

0

>1000

Hemiphyllodactylus

Figure 3.38. Altitude belt distribution of Gecko species in stuy sites
Research results showed that there are 11 species recorded at the altitude below 200
m, most of which are distributed here are common species in some genera such as Gekko and
Hemidactylus, including species such as Dixonius siamensis; Gekko aaronbaueri; Gekko
gecko; Gekko kabkaebin; Gehyra mutilata; Hemidactylus bowringii; Hemidactylus frenatus;
Hemidactylus ganotii; Hemidactylus platyurus. In which, two new species were discovered
in this high altitude belt, such as Dixonius tubercle and Gekko khumkhamensis.


20

3.4.3. Residence distribution
The analysis results in 03 habitat types showed that the difference in the distribution
of species is clear, especially in the habitat type per kilometer, there are few species, this can
be explained because it is rare for species to live in trees. It is not the main habitat of many
species of geckos because only one species is recorded on the tree, the rest are common
species and live in many forms (Fig. 3.40).
Species

Individuals

83

21

34

26
ON THE CLIFF

8

5

ON THE TREE

ON THE GROUND

Figure 3.40. Records on the habitat of gecko species in the study sites

3.5. Related problems of Gekko conservation in study sites

3.5.1. Rare and endemic species
Precious and rare species in the study area account for a high proportion of up to 15 species
(accounting for 30% of the total number of gecko species in Laos), up to 33 endemic species
with a narrow distribution and are only distributed in Laos making up 66% of the total number
of gecko species in Laos), there are 06 newly discovered species for science (occupying 12%
of the total number of gecko species in Laos). Priority should be given to conserving their
habitat forms and conducting more research sessions to search for more precious, rare and
new species for science.
3.5.2 Threat factors to Gecko species
1). Habitat loss and degradation
Deforestation for swidden cultivation: Most of the forest area in the study area belongs
to limestone mountains characterized by mountain peaks interspersed with valleys, so people
often take advantage of arable land for reclamation. swidden cultivation, especially in the
Northern provinces because of the limestone mountains covered with soil and trees, people often
take advantage of these areas.
Quarrying: Quarrying in the provinces of Laos is emerging a lot, especially in the
provinces with many limestone mountains such as Khammuone province, Vientiane province,
Huaphane province and Luangprabang province.
Roadworks: The road construction project in Huaphane province from Muong Hiem
through Muong Xon to Pha Thi and Tu Muong Xon to the Laos-Vietnam border has destroyed a
large area of natural forest at the multi-section road. There are skewers in the Nam Et-Phu Loi
National Park.


21
Wildfires spread: In the dry season, from December to April next year, there are many
areas of Laos where widespread forest fires occur during the survey period.
Logging and non-timber forest products: Talking about logging Lao PDR is very
famous after the liberation of the country, in 1960 Laos covered about 71% of the country's area
and by 2015 the coverage only about 40% remained, in the period 1980-2010 Laos' main income

was from forest exploitation.
2). Excessive hunting

3.5.3 Some proposed solutions for conservation work
1). Prioritized points for conservation
The prioritied conservation area is determined based on the priority conservation point
and the number of rare species present at that point. Accordingly, the priority area for the
conservation of gecko species has been identified, which is a high concentration of limestone
mountains such as the Khammuone Province research area Cyrtodactylus jaegeri;
Cyrtodactylus khammouanensis; Cyrtodactylus lomyenensis; Dixonius tuberculosis; Gekko
aaronbaueri; Gekko boehmei; Gekko bonkowskii; Gekko khunkhamensis; Gekko
sengchanthavongi; Gekko thakhekensis, Vientiane province includes: Cyrtodactylus
muangfuangensis and Dixonius somchanha, Huaphane province: Cyrtodactylus
houaphanensis, Luangprabang province has Cyrtodactylus ngoiensis and Udomxay Province
has Cyrtodactylus wayakonei species (Fig. 3.54).

Figure 3.54. Prioritied conservation areas in the study sites


22
2). Prioritized species for conservation
In the study area, there are a number of gecko species that are not on the list of rare
and precious species, but are also being exploited strongly for export to Vietnam and China
for food and medicinal wine, such as: Tokay Gekko. Conservation activities also need
attention to maintain and restore the population of this species
3). Priority conservation activities
- Protect the habitat of geckos;
- Propularize to raise awareness:
- Develop to ecotourism:
CONCLUSIONS - EXISTENCES - RECOMMENDATIONS

1. Conclusions
- The result has been recorded 28 species in 6 genera of the Gecko family. In which,
the genus Cyrtodactylus has 9 species; the genus Dixonius, Gekko, Hemidactylus,
Hemiphyllodactylus have all 4 species of each; and the genus Gehyra has 3 species.
Specifically, 14 species belonging to 5 genera have been recorded from Vientiane Province,
8 species belonging to 6 genera have been recorded from Luong Pha Bang Province, 5 species
belonging to 4 genera have been recorded from Udomxay Province, 3 species belonging to 3
genera have been recorded from Xiengkhoang Province, 11 species belonging to 5 genera
have been recorded from Huaphane Province and 12 species of 5 genera have been recorded
from Khammuone Province. Combined with previous studies bring, the total number of gecko
species from Laosto 45. Notably, the thesis results have described to the 6 new species of
geckos to science from Laos, including: Cyrtodactylus houaphanensis, Cyrtodactylus
muangfuangensis, Cyrtodactylus ngoiensis, Dixonius lao, Dixonius somchanhae and Gekko
khunkhamensis. Two species were recorded for the first time from Udomxay Province,
including Cyrtodactylus wayakonei and Hemiphyllodactylus kiziriani. There are 7 potential
species in the process of taxonomic analysis, including: Cyrtodactylus sp1, Cyrtodactylus sp2,
Gehyra sp1, Gehyra sp2, Hemiphyllodactylus sp1, Hemiphyllodactylus sp2 và Dixonius sp.
- The phylogenetic relationship of the genus Cyrtodactylus is divided into 3 groups
including C. phongnhakebangensis, C. wayakonei, and C. irregularis; the genus Gekko has
all belonging to the group Gekko japonicus; and the genus Dixonius has a member of the
Dixonius siamensis group.
- The species composition of geckos between Xieng Khouang and Udomxay provinces
has the highest degree of similarity (djk = 0.750), the lowest degree of similarity is from
Vientiane to Xieng Khouang provinces (djk = 0.222), and between the Central and the North
is (djk = 0.400).
- Distribution characteristics: Distributed by habitat, habitat 2 recorded the distribution
of the most species of geckos with 22 species (accounting for 78.6%). Distributed by altitude
belt, from 200 to less than 400 m altitude recorded the most species of geckos with 18 species
(64.3% of total species), especially 06 new species for science were discovered in the high
altitude belt. Distribution by location, karst cliffs recorded the most with 26 species (92.9%

of total species), 7 species were recorded in two locations, no species was recorded in all 3
locations.


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- Threats to gecko species and places that need to be prioritized for conservation:
Deforestation for farming; quarry mining; forest fires; construction of transport infrastructure;
harvesting timber and non-timber forest products; Hunted for food and for traditional
medicine. Regarding conservation status, 33 species are endemic to Laos, 9 species are
assessed as threatened in the Red List (IUCN, 2021) and 45 species are classified as Level II
in Decree No. 08/CP 2021 by the Government of Laos. Places that need to be prioritized for
conservation: The limestone mountains have a high concentration and distribution of rare and
precious species such as the research sites in Vientiane, Luangprabang, Khammuone,
Udomxay, Huaphane and Xiengkhoang provinces.
2. Existences
- As for taxonomy, 7 species have not been identified so far because DNA analysis is
in progress and furthermorphological data is needed to support species identification .
- Since the study area has an average altitude of less than 800 m, most of the field
studies are concentrated at an altitude of 150-650 m. Moreover, the altitude above 800 m has
dangerous terrain, difficult to access. Therefore, there is little research data at altitudes above
800 m for comparison and evaluation.
- Regarding the conservation of gecko species, Decree No. 08/CP/2021 of the Lao
Government on the list of wild animals in Laos has criteria to evaluate and rank the priority
of gecko species. All species are listed in List II, this is due to the lack of a complete research
database and the lack of experts on reptiles in general, geckos in particular.
3. Recommendations
1) The next research
- Conduct additional research on species composition, especially unidentified species,
conduct analysis and continue to investigate and collect more specimens.
- Conduct more research in the area with an altitude of over 800 m a.s.l., although the

habitat is difficult, difficult to access, and the cool climate rarely encounters geckos but rare
species.
2) Proposals for conservation
- Amending Decree No. 08/CP/2021 of the Government on the list of Lao wild animals,
the gecko species is based on legal and scientific basis to identify rare and precious species
prioritized for protection, international documents including IUCN Red List and CITES
Appendix.
- Focus on conserving locations with diverse species composition and habitats.
Implement conservation solutions for precious, rare and endemic species in priority
conservation areas.