Population Genetic Structure and Genetic Diversity in ...NongKhaiProvinceThailand(MKNK) Mekong 18 01...

Research ArticlePopulation Genetic Structure and Genetic Diversity inTwisted-Jaw Fish Belodontichthys truncatus Kottelat amp Ng1999 (Siluriformes Siluridae) from Mekong Basin

Surapon Yodsiri12 Komgrit Wongpakam12 Adisak Ardharn12

Chadaporn Senakun12 and Sutthira Khumkratok12

1Walai Rukhavej Botanical Research Institute MahasarakhamUniversity Kantharawichai District Maha Sarakham 44150Thailand2Biodiversity and Conservation Research Unit Walai Rukhavej Botanical Research Institute Mahasarakham UniversityKantharawichai District Maha Sarakham 44150 Thailand

Correspondence should be addressed to Komgrit Wongpakam komgrit wyahoocom

Received 15 February 2017 Accepted 11 July 2017 Published 16 August 2017

Academic Editor Marco Cucco

Copyright copy 2017 Surapon Yodsiri et alThis is an open access article distributed under theCreative CommonsAttribution Licensewhich permits unrestricted use distribution and reproduction in any medium provided the original work is properly cited

TheMekong River and its tributaries possess the second highest diversity in fish species in the world However the fish biodiversityin this river is threatened by several human activities such as hydropower plant construction Understanding the genetic diversityand genetic structure of the species is important for natural resource management Belodontichthys truncatus Kottelat amp Ng isendemic to theMekongRiver basin and is an important food source for people in this area In this study the genetic diversity geneticstructure and demographic history of the twisted-jaw fish B truncatus were investigated using mitochondrial cytochrome b genesequences A total of 124 fish specimens were collected from 10 locations in the Mekong and its tributaries Relatively high geneticdiversity was found in populations of B truncatus compared to other catfish species in the Mekong River The genetic structureanalysis revealed that a population from the Chi River in Thailand was genetically significantly different from other populationswhich is possibly due to the effect of genetic drift Demographic history analysis indicated that B truncatus has undergone recentdemographic expansion dating back to the end of the Pleistocene glaciation

1 Introduction

The Mekong is the second most biodiverse river for fishspecies It has been estimated that more than 877 fishspecies can be recorded in the Mekong and its tributaries[1] However many species are under threat due to human-mediated environmental change such as hydropower damconstruction [1 2]

The twisted-jaw catfish (Belodontichthys truncatus Kotte-lat amp Ng) is endemic to the Mekong basin [3] Two speciesof the genus Belodontichthys are found in the Mekong andits tributaries including B dinema Bleeker 1851 and Btruncatus The former species occur in central and southernThailand Malaysia Sumatra and Borneo while the latterspecies is found in northeast Thailand Lao PDR Cambodiaand Vietnam [3] Belodontichthys truncatus is a very impor-tant species for fisheries in Lao PDR andCambodia where the

fish is caught and exported toThailand [4] However there isno information on the genetic diversity and genetic structureof this important fish despite being important for naturalresource management [5]

In this study we used the mitochondrial cytochrome b(cyt b) sequences to examine the genetic diversity geneticstructure and demographic history of B truncatus in theMekong and two of its tributaries the Chi and Mun Riversin northeastern Thailand Previous studies indicated thatcyt b sequences can be successfully used to infer geneticstructure and demographic history of freshwater fishes [6ndash8] The information presented in this study will be useful forthe management of B truncatus In addition because thisspecies is widely distributed in theMekong and its tributariesunderstanding its genetic structure and demographic historywould shed some light on the effect of historical change

HindawiInternational Journal of ZoologyVolume 2017 Article ID 5976421 7 pageshttpsdoiorg10115520175976421

2 International Journal of Zoology

0 50 100 150 20025(km)

N

106∘00E105

∘00E104

∘00E103

∘00E102

∘00E101

∘00E

106∘00E105

∘00E104

∘00E103

∘00E102

∘00E101

∘00E

10∘00N

11∘00N

12∘00N

13∘00N

14∘00N

15∘00N

16∘00N

17∘00N

18∘00N

19∘00N

10∘00N

11∘00N

12∘00N

13∘00N

14∘00N

15∘00N

16∘00N

17∘00N

18∘00N

19∘00N

TributaryCountry boundaryLocation

Dam

Mekong River

Figure 1 Sampling locations for twisted-jaw fish Belodontichthys truncatus are used in this study Details of sampling sites are provided inTable 1

(eg Pleistocene climatic and environmental change) on fishbiodiversity

2 Materials and Methods

21 Specimen Collection and Identification A total of 124 fishspecimens were collected from seven locations in Thailandand Cambodia (Table 1 and Figure 1) Among these locationsonewas from theChi River and one from theMunRiver bothof these are in northeastern Thailand Three sites were fromthe Mekong River along the Thailand-Lao PDR border onefrom theMekong River in Cambodia and one fromTonle Sap

Lake in Cambodia Specimens were identified following thedescription of Belodontichthys truncatus by Kottelat amp Ng [3]

22 DNA Extraction Polymerase Chain Reaction andSequencing Genomic DNA was extracted from the tissueusing the Genomic DNA Extraction mini kit (RBC Bio-Science Xindian City Taiwan) A fragment of the cytochromeb (cyt b) gene was amplified using the primers Glu31 (51015840GTG-ACTTGAAAAACCACCGTT31015840) and CatThr29 (51015840ACC-TTCGATCTCCTGATTACAAGAC31015840) [9] The amplifi-cation reaction with a total volume of 50 120583l contained

International Journal of Zoology 3

Table1Detailsof

samplinglocatio

nsforB

elodontich

thys

truncatus

used

inthisstu

dyandhaplotypea

ndnu

cleotided

iversity

Locatio

n(cod

e)River

Geographic

region

119873Collection

date

Haplotype

diversity

Nucleotided

iversity

Yasothon

ProvinceTh

ailand

(CHYT

)Ch

i15∘471015840248210158401015840N

104∘081015840340410158401015840E

21011

2014

00

MueangDistric

tMahaS

arakham

Province

(CHMK)

Chi

16∘131015840182910158401015840N

103∘20101584031310158401015840E

214082014

00

AtSamatDistric

tRo

iEtP

rovince(CH

RO1)

Chi

15∘401015840381710158401015840N

104∘061015840488710158401015840E

46112014

08333plusmn02224

0001958plusmn0001641

Phanom

PhraiD

istric

tRo

iEtP

rovince(CH

RO2)

Chi

15∘411015840415010158401015840N

103∘071015840386410158401015840E

141011

2014

05055plusmn01581

0003413plusmn0002077

Ubo

nRa

tchathaniP

rovinceTh

ailand

(MUUB)

Mun

15∘141015840297110158401015840N

104∘571015840206410158401015840E

101111

2014

08667plusmn00850

000205plusmn000141

Non

gKhaiP

rovinceTh

ailand

(MKN

K)Mekon

g18∘011015840282910158401015840N

101∘ 5

21015840543410158401015840E

138112014

09487plusmn00506

000

405plusmn000242

Kho

ngCh

iam

Distric

tUbo

nRa

tchathani

ProvinceTh

ailand

(MKU

B)Mekon

g15∘231015840268210158401015840N

105∘291015840391410158401015840E

131710

2014

09359plusmn00507

000

401plusmn

000240

KhemaratDistric

tUbo

nRa

tchathaniP

rovince

Thailand

(MKK

R)Mekon

g16∘021015840321610158401015840N

105∘131015840323910158401015840E

281410

2014

08810plusmn00367

000271plusmn

000165

Phno

mPenh

ProvinceC

ambo

dia(

MKC

DPP

)Mekon

g11∘331015840281810158401015840N

104∘561015840141310158401015840E

121811

2014

08788plusmn00751

000242plusmn000158

TonleS

apLakeP

ursatP

rovinceCa

mbo

dia

(MKC

DPO

)To

nleS

apLake

12∘021015840192410158401015840N

104∘431015840435410158401015840E

261511

2014

08062plusmn00594

000365plusmn000212

Total

124

09765plusmn00051

0003179plusmn0001826


2 120583lMgCl2(50mM) 5 120583l of 10x PCR buffer 16 120583l of mixed

dNTPs (10 120583M) 2 120583l of each primer (10 120583M) 04 120583l of TaqDNA polymerase (5 u120583l) and 2 120583l DNA temple The tem-perature profile was as follows 94∘C for 3min followed by35 cycles of 94∘C for 30 seconds 48∘C for 1min and 72∘Cfor 130min with a final extension at 72∘C for 7min [10] ThePCR products were checked by 1 agarose gel electrophoresisand purified using a High Yield GelPCR DNA fragmentextraction kit (RBC BioScience Taiwan) Sequencing wasperformed at the Macrogen DNA sequencing service (SeoulKorea) using the same primers as in the PCR

23 Data Analysis A fragment of 1024 bp of the cytb gene was obtained from 124 specimens Sequenceswere deposited in GenBank under the accession numbersKY607016ndashKY607139 Genealogical relationships betweenhaplotypes were estimated using a median joining (MJ)network (Bandelt et al 1999) calculated using Networkv 4610 (httpwwwfluxus-engineeringcom) Haplotypediversity and nucleotide diversity were calculated usingArlequin ver 35 [11] The population pairwise 119865ST calculatedin Arlequin was used to infer the genetic structure Thesignificance test statistic was obtained from 1023 permu-tations To avoid bias due to a small sample size popu-lations with less than five specimens were omitted fromthe genetic structure analysis A Mantel test [12] was usedto determine the relationship between the genetic distance(119865ST from Arlequin) and the geographical distance (km) foran isolation-by-distance (IBD) model The Mantel test wasimplemented in IBDver 152 [13] using 1000 randomizationsThe mismatch distribution was used to test the signature ofthe population expansion Populations that have undergonea recent past demographic expansion show a unimodalmismatch distribution [14] The sum-of-squares deviationand Harpendingrsquos raggedness index [15] were used to test thedeviation from the sudden expansion model A mismatchdistribution was estimated using Arlequin Fursquos 119865S [16] andTajimarsquosD [17] statistical tests were used to test the populationequilibrium A large negative value from these tests wasexpected for the demographic expansion

3 Results

31 Genetic Diversity A total of 124 sequences from sevensampling locations were obtained in this study and 40 hap-lotypes were identified Haplotype diversity in each locationranged between 0 in two populations (CHYT and CHMK) ofthe Chi River in northeastern Thailand and 09487 in NongKhai Province (MKNK) for theMekong River with an overallaverage of 09765 (Table 1) The nucleotide diversity in eachpopulation ranged from 0 in two populations (CHYT andCHMK) of the Chi River to 00041 in Nong Khai Province(MKNK) for the Mekong River with an overall average of00032 (Table 1)

32 Mitochondrial DNA Genealogy Themedian joining net-work (Figure 2) revealed no major phylogeographic breaksamong the 124 sequences included in the analysisThere is no

Mekong RiverChi RiverMun River

Figure 2 Median joining (MJ) network of 124 mitochondrialcytochrome b (cyt b) sequences of Belodontichthys truncatus Eachhaplotype is represented by a circle and sizes of circles are relative tonumber of individuals sharing a specific haplotype Haplotypes arelabelled according to the river

0

500

1000

1500

2000

2500

0 5 10 15 20 25 30

ObservedSimulation

Tau = 0838

Fursquos FS = minus262649lowastlowast

lowastlowastTajimarsquos D = minus25364lowastlowast

P lt 0001

Figure 3Mismatch distribution of 124mitochondrial cytochrome b(cyt b) sequences ofBelodontichthys truncatus representing observedand expected pairwise differences under sudden population expan-sionmodelMismatch distribution forB truncatus is consistent withpredictions of sudden population expansion model (SSD = 00047119875 = 04750 Harpendingrsquos raggedness index = 00349 119875 = 05270)

evidence of geographic association between the haplotypesThe core haplotype that has the highest frequency is sharedby 35 specimens from all threemajor rivers (Mekong Chi andMun) of the Mekong Basin Overall the haplotype is a star-like shape which is the characteristic of a recent expansion inthe population

33 Genetic Structure The population pairwise 119865ST revealedan overall low level of genetic structuring between the popu-lations of Belodontichthys truncatus No significant 119865ST valueswere observed except for comparisons between a population(CHRO2) from the Chi River and the other populationswhere most were significantly different (Table 2) A Manteltest revealed no significant relationships between the geneticand geographic distances (119903 = minus00528 119875 = 03590)

34 Demographic History A mismatch distribution analysisrevealed a unimodal mismatch graph (Figure 3) which isa characteristic of a recently expanding population that is


Table 2 Pairwise 119865ST (below diagonal) and associated 119875 value (above diagonal) between populations of Belodontichthys truncatus collectedduring AugustndashNovember 2014 inThailand and Cambodia Population codes are according to Table 1

CHRO2 MKKR MKNK MKUB MKCDPO MKCDPP MUUBCHRO2 lt00001 lt00001 00090 lt00001 00090 00631MKKR 01107 00450 01982 04594 05405 00450MKNK 00970 00605 01441 01892 00631 03063MKUB 00517 00148 00128 05676 06036 02883MKCDPO 00963 minus00029 00162 minus00065 05946 01531MKCDPP 00577 minus00085 00435 minus00106 minus00079 00631MUUB 00397 00756 00108 00098 00267 00618Bold characters indicate statistically significant differences at 119875 values adjusted by Bonferronirsquos correction

consistent with the star-like shape of the mtDNA genealogyHarpendingrsquos raggedness index (00349 119875 = 05270) and thesum-of-squares deviation (SSD = 00047 119875 = 04750) werenot significantly different from the simulated data under thesudden population expansion model Population expansionwas also supported by highly significant negative values forboth Tajimarsquos D (minus25364 119875 lt 0001) and Fursquos 119865S (minus262649119875 lt 0001) The expansion time estimated based on thefish cyt b evolutionary rate of 1 Myr [18] and assuming ageneration time of two years forB truncatus was 20458 yearsago

4 Discussion

The levels of genetic diversity observed in B truncatus (ℎ =09765 120587 = 0003179) based on the cyt b mtDNA sequencesare similar to other fish species of the family Siluridae Forexample butter catfish Ompok bimaculatus in India (ℎ =08270 120587 = 000391) [19] and Trichomycterus areolatus fromChile [8] However the genetic diversity found inB truncatusis higher than other catfish species from the Mekong River[20]

Mismatch distribution analysis indicated a recent demo-graphic expansion in B truncatus that dated back to 20458years ago which is at the end of the last glaciation Previousstudies found a significant influence from the Pleistoceneglaciation on genetic diversity genetic structure and demo-graphic history in many Southeast Asian floras and faunas[eg [21ndash23]] During the Pleistocene glaciations climaticand environmental conditions in Southeast Asia were coolerand drier [24] Evidence indicated that the water level andwater flow in large rivers in Southeast Asia including theMekong reduced during the Pleistocene glaciation [25]After the climatic conditions recovered at the end of thePleistocene the water level and water flow increased whichcould trigger the population expansion in B truncatus

The population genetic structure analysis based on the119865ST values revealed that a population from the Chi Riverwas genetically significantly different from almost all otherpopulations from the Mekong but not from the Mun RiverMitochondrial genealogy indicated no evidence of divergentlineages for B truncatus from different rivers thus indicatingthat the genetic differentiation between the Chi and MekongRiver populations is likely to be a recent event The sharing

of the mtDNA haplotypes between fish from these riverssuggests that there is some gene flow between these popula-tions There are two possible explanations for the significantgenetic differentiations between the populations from theChiRiver and the Mekong River The Chi River is part of theMekong and it originates from a mountainous area in theupper part of northeastern Thailand The Chi River joins theMunRiver inUbon Ratchathani Province and then flows intothe Mekong Near the location (approximately 55 km) of theMun andMekong confluence there is a dam (PakMun dam)that was constructed in 1994 This dam could be a barrierto gene flow between B truncatus populations in the Chiand Mun Rivers and the Mekong The effects of dams ongene flow have been investigated in other fish species [26ndash28] However given that this dam is only 23 years old it isunlikely to have had an impact on the genetic structure Themore likely explanation for the genetic differentiation of theChi River populations from the Mekong River is the effect ofgenetic drift It has been suggested that the random samplingof the alleles from the source population of the colonizercould lead to the genetic differentiation of the population bythe effect of genetic drift [29] Although populations from theChi River possess considerable genetic diversity the relativelylowhaplotype diversity in this population (05055) comparedto the others that contributed to the significant pairwise 119865STvalues supports genetic drift playing a role in the geneticdifferentiation

5 Conclusion

We found that the genetic diversity of B truncatus is consid-erable compared to other fishes in the Mekong This impliesa large effective population size for this species The resultsshowed significant genetic differentiation for the populationfrom a Mekong tributary the Chi River in Thailand that wasgenetically significantly different from the other populationsdue to the effect of genetic drift We also found that historicalenvironmental change during the Pleistocene has had aneffect on the demographic history of this species Giventhe recent rapid changes in the Mekong and its tributariesparticularly for the hydropower dam construction whichis predicted to have an effect on the fish productivity andbiodiversity [1] further studies should examine the effect ofthis on genetic structure and diversity on the fish species


Conflicts of Interest

The authors declare that they have no conflicts of interest

Acknowledgments

This study was financially supported by a grant fromMahasarakham University

References

[1] G Ziv E Baran S Nam I Rodrıguez-Iturbe and S A LevinldquoTrading-off fish biodiversity food security and hydropower inthe Mekong River Basinrdquo Proceedings of the National Academyof Sciences of the United States of America vol 109 no 15 pp5609ndash5614 2012

[2] D Dudgeon ldquoLarge-scale hydrological changes in tropical AsiaProspects for riverine biodiversityrdquo BioScience vol 50 no 9 pp793ndash806 2000

[3] M Kottelat and H H Ng ldquoBelodontichthys truncatus a newspecies of silurid catfish from Indochina (Teleostei SiluridaerdquoIchthyological Exploration of Freshwaters vol 10 pp 387ndash3911999

[4] A Termvidchakorn and K G Hortle ldquoA guide to larvae andjuveniles of some common fish species from the Mekong RiverBasinrdquo MRC Technical Paper 38 Mekong River CommissionPhnom Penh Cambodia 2013

[5] R DeSalle and G Amato ldquoThe expansion of conservationgeneticsrdquo Nature Reviews Genetics vol 5 no 9 pp 702ndash7122004

[6] L Yang and S He ldquoPhylogeography of the freshwater catfishHemibagrus guttatus (Siluriformes Bagridae) implications forSouth China biogeography and influence of sea-level changesrdquoMolecular Phylogenetics and Evolution vol 49 no 1 pp 393ndash398 2008

[7] M Habib W S Lakra V Mohindra et al ldquoEvaluation ofcytochrome b mtDNA sequences in genetic diversity studies ofChannamarulius (Channidae Perciformes)rdquoMolecular BiologyReports vol 38 no 2 pp 841ndash846 2011

[8] E G Gonzalez C Pedraza-Lara and I Doadrio ldquoGeneticdiversity and population history of the endangered killifishAphanius baeticusrdquo Journal of Heredity vol 105 no 5 pp 597ndash610 2014

[9] P J Unmack A P Bennin EMHabit P F Victoriano and J BJohnson ldquoImpact of ocean barriers topography and glaciationon the phylogeography of the catfish Trichomycterus areolatus(Teleostei Trichomycteridae) in Chilerdquo Biological Journal of theLinnean Society vol 97 no 4 pp 876ndash892 2009

[10] P J Unmack J P Barriga M A Battini E M Habit and JB Johnson ldquoPhylogeography of the catfish Hatcheria macraeireveals a negligible role of drainage divides in structuringpopulationsrdquo Molecular Ecology vol 21 no 4 pp 942ndash9592012

[11] L Excoffier and H E L Lischer ldquoArlequin suite ver 35 anew series of programs to perform population genetics analysesunder Linux and Windowsrdquo Molecular Ecology Resources vol10 no 3 pp 564ndash567 2010

[12] N Mantel ldquoThe detection of disease clustering and a general-ized regression approachrdquo Cancer Research vol 27 no 2 pp209ndash220 1967

[13] A J Bohonak ldquoIBD (isolation by distance) A program foranalyses of isolation by distancerdquo Journal of Heredity vol 93no 2 pp 153-154 2002

[14] A R Rogers and H Harpending ldquoPopulation growth makeswaves in the distribution of pairwise genetic differencesrdquoMolecular Biology and Evolution vol 9 no 3 pp 552ndash569 1992

[15] H C Harpending ldquoSignature of ancient population growth ina low-resolution mitochondrial DNA mismatch distributionrdquoHuman Biology vol 66 no 4 pp 591ndash600 1994

[16] Y-X Fu ldquoStatistical tests of neutrality of mutations againstpopulation growth hitchhiking and background selectionrdquoGenetics vol 147 no 2 pp 915ndash925 1997

[17] F Tajima ldquoStatistical method for testing the neutral mutationhypothesis by DNApolymorphismrdquoGenetics vol 123 no 3 pp585ndash595 1989

[18] E Bermingham S McCafferty and A Martin ldquoFish biogeog-raphy andmolecular clocks perspectives from the PanamanianIsthmusrdquo in Molecular Systematics of Fishes pp 113ndash126 Aca-demic Press New York NY USA 1997

[19] R Kumar B K Pandey U K Sarkar et al ldquo Population geneticstructure and geographic differentiation in butter catfish rdquoMitochondrial DNA Part A vol 28 no 3 pp 442ndash450 2017

[20] U Na-Nakorn S Sukmanomon M Nakajima et al ldquoMtDNAdiversity of the critically endangered Mekong giant catfish(Pangasianodon gigas Chevey 1913) and closely related speciesImplications for conservationrdquo Animal Conservation vol 9 no4 pp 483ndash494 2006

[21] C H Cannon and P S Manos ldquoPhylogeography of the South-east Asian stone oaks (Lithocarpus)rdquo Journal of Biogeographyvol 30 no 2 pp 211ndash226 2003

[22] P Pramual C Kuvangkadilok V Baimai and C Walton ldquoPhy-logeography of the black fly Simulium tani (Diptera Simuliidae)from Thailand as inferred from mtDNA sequencesrdquo MolecularEcology vol 14 no 13 pp 3989ndash4001 2005

[23] K Morgan Y-M Linton P Somboon et al ldquoInter-specificgene flow dynamics during the Pleistocene-dated speciationof forest-dependent mosquitoes in Southeast Asiardquo MolecularEcology vol 19 no 11 pp 2269ndash2285 2010

[24] D Penny ldquoA 40000 year palynological record from north-east Thailand implications for biogeography and palaeo-environmental reconstructionrdquo Palaeogeography Palaeoclima-tology Palaeoecology vol 171 no 3-4 pp 97ndash128 2001

[25] V S Kale A Gupta and A K Singhvi ldquoLate PleistoceneHolocene palaeohydrology of monsoon Asiardquo in Palaeohydrol-ogy Understanding Global Change pp 213ndash232 JohnWiley andSons New York NY USA 2003

[26] H AnvariFar H Farahmand D M Silva R P Bastos AKhyabani and H AnvariFar ldquoFourteen years after the Shahid-Rajaei dam construction An evaluation of morphometric andgenetic differentiation between isolated up- and downstreampopulations of Capoeta capoeta gracilis (Pisces Cyprinidae) inthe Tajan River of Iranrdquo Genetics and Molecular Research vol12 no 3 pp 3465ndash3478 2013

[27] M M Hansen M T Limborg A-L Ferchaud and J-MPujolar ldquoThe effects of medieval dams on genetic divergenceand demographic history in brown trout populationsrdquo BMCEvolutionary Biology vol 14 no 1 article 122 2014

[28] L Zhao E L Chenoweth J Liu and Q Liu ldquoEffects of damstructures on genetic diversity of freshwater fish Sinibramamacrops in Min River Chinardquo Biochemical Systematics andEcology vol 68 pp 216ndash222 2016


[29] L Excoffier and N Ray ldquoSurfing during population expansionspromotes genetic revolutions and structurationrdquo Trends inEcology and Evolution vol 23 no 7 pp 347ndash351 2008

Submit your manuscripts athttpswwwhindawicom

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Anatomy Research International

PeptidesInternational Journal of


Hindawi Publishing Corporation httpwwwhindawicom

International Journal of

Volume 201


Molecular Biology International

GenomicsInternational Journal of


The Scientific World JournalHindawi Publishing Corporation httpwwwhindawicom Volume 2014


BioinformaticsAdvances in

Marine BiologyJournal of



Signal TransductionJournal of


BioMed Research International

Evolutionary BiologyInternational Journal of



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ArchaeaHindawi Publishing Corporationhttpwwwhindawicom Volume 2014


Genetics Research International


Advances in

Virolog y

Hindawi Publishing Corporationhttpwwwhindawicom

Nucleic AcidsJournal of

Volume 2014

Stem CellsInternational



Enzyme Research



Microbiology


0 50 100 150 20025(km)

N

106∘00E105

∘00E104

∘00E103

∘00E102

∘00E101

∘00E

106∘00E105

∘00E104

∘00E103

∘00E102

∘00E101

∘00E

10∘00N

11∘00N

12∘00N

13∘00N

14∘00N

15∘00N

16∘00N

17∘00N

18∘00N

19∘00N

10∘00N

11∘00N

12∘00N

13∘00N

14∘00N

15∘00N

16∘00N

17∘00N

18∘00N

19∘00N

TributaryCountry boundaryLocation

Dam

Mekong River

Figure 1 Sampling locations for twisted-jaw fish Belodontichthys truncatus are used in this study Details of sampling sites are provided inTable 1

(eg Pleistocene climatic and environmental change) on fishbiodiversity

2 Materials and Methods

21 Specimen Collection and Identification A total of 124 fishspecimens were collected from seven locations in Thailandand Cambodia (Table 1 and Figure 1) Among these locationsonewas from theChi River and one from theMunRiver bothof these are in northeastern Thailand Three sites were fromthe Mekong River along the Thailand-Lao PDR border onefrom theMekong River in Cambodia and one fromTonle Sap

Lake in Cambodia Specimens were identified following thedescription of Belodontichthys truncatus by Kottelat amp Ng [3]

22 DNA Extraction Polymerase Chain Reaction andSequencing Genomic DNA was extracted from the tissueusing the Genomic DNA Extraction mini kit (RBC Bio-Science Xindian City Taiwan) A fragment of the cytochromeb (cyt b) gene was amplified using the primers Glu31 (51015840GTG-ACTTGAAAAACCACCGTT31015840) and CatThr29 (51015840ACC-TTCGATCTCCTGATTACAAGAC31015840) [9] The amplifi-cation reaction with a total volume of 50 120583l contained


Table1Detailsof

samplinglocatio

nsforB

elodontich

thys

truncatus

used

inthisstu

dyandhaplotypea

ndnu

cleotided

iversity

Locatio

n(cod

e)River

Geographic

region

119873Collection

date

Haplotype

diversity

Nucleotided

iversity

Yasothon

ProvinceTh

ailand

(CHYT

)Ch

i15∘471015840248210158401015840N

104∘081015840340410158401015840E

21011

2014

00

MueangDistric

tMahaS

arakham

Province

(CHMK)

Chi

16∘131015840182910158401015840N

103∘20101584031310158401015840E

214082014

00

AtSamatDistric

tRo

iEtP

rovince(CH

RO1)

Chi

15∘401015840381710158401015840N

104∘061015840488710158401015840E

46112014

08333plusmn02224

0001958plusmn0001641

Phanom

PhraiD

istric

tRo

iEtP

rovince(CH

RO2)

Chi

15∘411015840415010158401015840N

103∘071015840386410158401015840E

141011

2014

05055plusmn01581

0003413plusmn0002077

Ubo

nRa

tchathaniP

rovinceTh

ailand

(MUUB)

Mun

15∘141015840297110158401015840N

104∘571015840206410158401015840E

101111

2014

08667plusmn00850

000205plusmn000141

Non

gKhaiP

rovinceTh

ailand

(MKN

K)Mekon

g18∘011015840282910158401015840N

101∘ 5

21015840543410158401015840E

138112014

09487plusmn00506

000

405plusmn000242

Kho

ngCh

iam

Distric

tUbo

nRa

tchathani

ProvinceTh

ailand

(MKU

B)Mekon

g15∘231015840268210158401015840N

105∘291015840391410158401015840E

131710

2014

09359plusmn00507

000

401plusmn

000240

KhemaratDistric

tUbo

nRa

tchathaniP

rovince

Thailand

(MKK

R)Mekon

g16∘021015840321610158401015840N

105∘131015840323910158401015840E

281410

2014

08810plusmn00367

000271plusmn

000165

Phno

mPenh

ProvinceC

ambo

dia(

MKC

DPP

)Mekon

g11∘331015840281810158401015840N

104∘561015840141310158401015840E

121811

2014

08788plusmn00751

000242plusmn000158

TonleS

apLakeP

ursatP

rovinceCa

mbo

dia

(MKC

DPO

)To

nleS

apLake

12∘021015840192410158401015840N

104∘431015840435410158401015840E

261511

2014

08062plusmn00594

000365plusmn000212

Total

124

09765plusmn00051

0003179plusmn0001826





3 Results





0

500

1000

1500

2000

2500

0 5 10 15 20 25 30

ObservedSimulation

Tau = 0838



P lt 0001









4 Discussion





5 Conclusion





Acknowledgments


References






































Volume 201






















Advances in

Virolog y



Volume 2014




Enzyme Research



Microbiology


Table1Detailsof

samplinglocatio

nsforB

elodontich

thys

truncatus

used

inthisstu

dyandhaplotypea

ndnu

cleotided

iversity

Locatio

n(cod

e)River

Geographic

region

119873Collection

date

Haplotype

diversity

Nucleotided

iversity

Yasothon

ProvinceTh

ailand

(CHYT

)Ch

i15∘471015840248210158401015840N

104∘081015840340410158401015840E

21011

2014

00

MueangDistric

tMahaS

arakham

Province

(CHMK)

Chi

16∘131015840182910158401015840N

103∘20101584031310158401015840E

214082014

00

AtSamatDistric

tRo

iEtP

rovince(CH

RO1)

Chi

15∘401015840381710158401015840N

104∘061015840488710158401015840E

46112014

08333plusmn02224

0001958plusmn0001641

Phanom

PhraiD

istric

tRo

iEtP

rovince(CH

RO2)

Chi

15∘411015840415010158401015840N

103∘071015840386410158401015840E

141011

2014

05055plusmn01581

0003413plusmn0002077

Ubo

nRa

tchathaniP

rovinceTh

ailand

(MUUB)

Mun

15∘141015840297110158401015840N

104∘571015840206410158401015840E

101111

2014

08667plusmn00850

000205plusmn000141

Non

gKhaiP

rovinceTh

ailand

(MKN

K)Mekon

g18∘011015840282910158401015840N

101∘ 5

21015840543410158401015840E

138112014

09487plusmn00506

000

405plusmn000242

Kho

ngCh

iam

Distric

tUbo

nRa

tchathani

ProvinceTh

ailand

(MKU

B)Mekon

g15∘231015840268210158401015840N

105∘291015840391410158401015840E

131710

2014

09359plusmn00507

000

401plusmn

000240

KhemaratDistric

tUbo

nRa

tchathaniP

rovince

Thailand

(MKK

R)Mekon

g16∘021015840321610158401015840N

105∘131015840323910158401015840E

281410

2014

08810plusmn00367

000271plusmn

000165

Phno

mPenh

ProvinceC

ambo

dia(

MKC

DPP

)Mekon

g11∘331015840281810158401015840N

104∘561015840141310158401015840E

121811

2014

08788plusmn00751

000242plusmn000158

TonleS

apLakeP

ursatP

rovinceCa

mbo

dia

(MKC

DPO

)To

nleS

apLake

12∘021015840192410158401015840N

104∘431015840435410158401015840E

261511

2014

08062plusmn00594

000365plusmn000212

Total

124

09765plusmn00051

0003179plusmn0001826





3 Results





0

500

1000

1500

2000

2500

0 5 10 15 20 25 30

ObservedSimulation

Tau = 0838



P lt 0001









4 Discussion





5 Conclusion





Acknowledgments


References






































Volume 201






















Advances in

Virolog y



Volume 2014




Enzyme Research



Microbiology





3 Results





0

500

1000

1500

2000

2500

0 5 10 15 20 25 30

ObservedSimulation

Tau = 0838



P lt 0001









4 Discussion





5 Conclusion





Acknowledgments


References






































Volume 201






















Advances in

Virolog y



Volume 2014




Enzyme Research



Microbiology





4 Discussion





5 Conclusion





Acknowledgments


References






































Volume 201






















Advances in

Virolog y



Volume 2014




Enzyme Research



Microbiology




Acknowledgments


References






































Volume 201






















Advances in

Virolog y



Volume 2014




Enzyme Research



Microbiology










Volume 201






















Advances in

Virolog y



Volume 2014




Enzyme Research



Microbiology

Population Genetic Structure and Genetic Diversity in ...NongKhaiProvinceThailand(MKNK) Mekong 18 01...

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