Population Density, Thermal Requirement And Geographic Pattern Of Genetic Structure In The Reevese's Butterfly Lizard (Leiolepis Reevesii) From Hainan, Southern China

Butterfly lizards of the genus Leiolepis(Agamidae)are widely distributed in coastal regions of Southeast Asia and southern China.Of the seven species of Leiolepis lizards recognized worldwide,the Reevese's butterfly lizard L.reevesii has a most northerly distribution that ranges from southern China to Vietnam.Population densities and nest traits of L.reevesii throughout Hainan were studied during April and May of 2007.A Kruskal-Wallis test revealed between-region differences in population density,with valuses for the popuiation variable being apparently greater in Sanya,Lingshui.Dongfang and Ledong than in Wenchang,Ding'an,Tunchang,Qionghai and Wanning.The test also revealed population densities were highly habitat-dependent, with the values being greater in orchards,shrubs of prickly pear and grassplots than in forests of small-leaved gam and beefwood.Populations at different altitudes differed in density,but no clear-cut correlation between population density and altitude was found.Nest length and nest depth were both positively correlated with body size(SVL, snout-vent length)in juveniles but not in adults;the number of turnings of nest chanel was positively correlated with SVL in adults but not in juveniles.One-way ANOVAs revealed that adults from different localities differed in nest length and depth.One-way ANCOVAs (with nest temperature as the covariate)revealed between-region differences in body temperatures of adult lizards.One-way ANOVAs revealed between-age differences in body and nest temperatures,but did not show between-age differences in air temperature. One-way ANCOVAs(with nest temperature as the covariate)revealed between-age diffcrences in body and air temperatures.A partial correlation analysis on adults revealed that body temperature was positively correlated with nest temperature,and that body and nest temperatures were both not always positively correlated with air temperature in the regions under study.When data were pooled for all regions,body temperature was positively correlated with not only nest temperature but also air temperature in adults as well as in juveniles but,neither in adults nor in juveniles,was nest temperature correlated with air temperature.in late April of 2006 and 2007,we collected a total of 844 lizards from Hainan to study sexual dimorphism and reproductive output.The minimal reproductive female in our sample was 84.0 mm SVL.Individuals larger than this size were arbitrarily judged as adults. The lizard is sexually dimorphic,with males being larger in both body size and head size. The rate(slope)at which head size(both head length and head width)increased with increasing SVL was greater in male adults than in female adults,and was greater in male juveniles than in females of the same age.A two-way ANOVA,with sex and age(adult and juvenile)as the factors,on the residuals of the regressions of head length and head width on SVL,showed that heads were larger in males than in females,and that juveniles had larger heads relative to SVL than did adults.Females maintained in the laboratory laid eggs between late May and mid-July.Relative clutch mass was negatively correlated with maternal SVL,which indicates that smaller reproductive females have relatively greater reproductive output in the lizard.Given that a female's reproduction retards her growth,the relatively greater reproductive in smaller females at least partly explains why females have the smaller adult size than do males.We used the Reevese's Butterfly Lizard as a model animal to test the hypothesis that incubation temperatures should be relatively high but narrow in lizards depositing eggs in warm and thermally stable nests.Eggs were incubated under three constant(27,30 and 33℃)and one fluctuating temperature regimes.Hatchability was highest at 30℃and lowest at 27℃.The mean rather than the variance of temperatures affected incubation length,and the mean incubation length was 101.1 days at 27℃,69.6 days at 30℃and 55.3 days at 33℃.Incubation temperature did not affect the sexual phenotype of hatchlings.Hatchlings from different treatments showed little variation in morphological phenotypes,but differed significantly in locomotor phenotypes.Hatchlings incubated at 27℃performed much more poorly in the racetrack than did hatchlings from the other three treatments,and the variance of incubation temperatures negatively affected sprint speed of hatchlings.The incubation temperatures of 27℃and 33℃supported egg incubation,but these two temperatures were not ideal.The range of temperatures is suitable for incubation of L. reevesii eggs.Compared with results reported for other squamate reptiles either from cool or warm habitats but depositing eggs in shallow nests,temperatures suitable for egg incubation are relatively high but narrow in L.reevesii.Thus,our data support the above hypothesis.Adult males(37.7℃)selected body temperatures higher than did gravid females (36.8℃),but lower than did nongravid females(39.0℃)in a thermal gradient during the breeding season.The mean values for the lower(critical thermal minimum,CTMin)and upper(critical thermal maximum,CTMax)limits of thermal tolerance were 11.1℃and 45.7℃,respectively.We used LB-2T temperature-sensitive radiotransmitters(Holohil,Canada)and HOBO datalogges(Onset,USA)to study behavioral thermoregulation of adult L.reevesii that were active in an enclosure located at Ledong.Lizards in the enclosure could maintain relatively high and stable body temperatures when they had the opportunity to thermoregulate.Consistent with the results reported for individuals in the laboratory thermal gradient,gravid females in the enclosure selected lower body temperatures than did nongravid females and males.Body temperatures were higher during the time intervals of 11:00-15:00h and 15:00-19:00h than during the time interval of 07:00-11:00h,and were higher in sunny than in cloudy days.The interaction between time interval and weather condition was a significant source of variation in body temperature. To assess the genetic diversity within L.reevesii,its population structure and its evolutionary history,we sequenced 1004 bp of cytochrome b for 254 individuals collected from 14 localities throughout Hainan including Dongdao Islet and two localities on mainland China.One hundred and twenty-two variable sites were observed and 73 haplotypes were defined.The statistical parsimony network suggested two distinct clades(A and B).The divergence time of the two clades was estimated to be 1.00-1.06 million years ago.Clade A suggests restricted gene flow with isolation by distance,whereas clade B suggests restricted gene flow but with some long distance dispersal.Leiolepis reevesii experienced population expansion in the recent past(0.60 million years ago).Results from nested clade analysis (NCA)and AMOVA reveal that L.reevesii is characterized by restricted gene flow and strong population genetic structure.Wuzhishan Mountains and Yinggeling Mountains in Hainan served as an important barrier to gene flow in the recent past,whereas Qiongzhou Strait between Hainan and mainland China did not.Although being very abundant in many coastal localities of southern China some thirty years ago.L.reevesii is currently regarded as a vulnerable species according to a recent volume of China Red Data Book of Endangered Animals.Overexploitation by local people for food and medicine and loss and fragmentation of habitats resulting from an increasingly rapid growth of sight-seeing and agricultural facilities and highways are the main causes of the population decline over the past three decades,and there are realistic threats of local extinction in some places.Furthermore,the high nest fidelity demonstrated by the lizard makes it much more susceptible to adverse effects of loss and fragmentation of habitats on genetic diversity.Therefore,measures are urgently wanted to protect the lizard.Data generated from this study,focusing primarily on the current situation,life-histories,genetic variability and population genetic structure can be usuful in improving management and conservation of the lizard.