Studies On Population Ecology And Population Genetic Differentiation Of Phrynocephalus Guinanensis(Agamidae)

The toad-headed lizard(Phrynocephalus guinanensis)exclusively distributes in an area of sand dunes in Guinan County of Qinghai Province.Currently,relative little is known about the ecological information for P.guinanensis.Reccent molecular phylogenetic study indicated that two closely related lizards,P.guinanensis and P.putjatia were phylogenetically clustered as a species,however,possessed significant difference on sexual color and distribution.The obtained result of the food resources,sexual size dimorphism(SSD),population demography,potential geographic distribution of P.guinanensis strengthened our understanding on population ecology of the species.In addition,analyses on the population genetic divergence between P.putjatia and P.guinanensis was applied to confirm the systematic status of P.guinanensis.(1)Study the spatial distribution of active caves and potential food resources in P.guinanensisThree line transects with 150 traps were used to study the spatial distribution of active caves and the potentially available food resources for P.guinanensis in both sides of the grassland and sand dunes contact area.The relationship between cave geographical distribution and insect resource availability was analyzed.We successfully captured 1236 individual insects belonging to 2 classes,10 orders,26 families,and 42 species.Among them,978 Insecta individuals were observed,comprising 7 orders,21 families,and 37 species,while the remaining 258 Arachnoidea individuals comprised 3 orders,5 families,and 5 species.We also discovered that among the 570 individuals captured in the grassland,2 classes,9 orders,22 families,and 37 species were present,whereas 666 individuals were captured in sand dunes and included 2 classes,9 orders,21 families,and 33 species.The Shannon-Wiener diversity index,Simpson diversity index,and an evenness index for grassland insects were all larger than the corresponding indexes for the insects in sand dunes,which indicated that the insect resources were more abundant and homogeneous in grasslands than in sand dunes.Our results also showed that more active caves existed in sand dunes than in grasslands,and their quantity in sand dunes increased with a decrease in the distance to the grassland-sand dune boundary;however,no correlation was found between the quantity of active caves in grasslands and their distance to the boundary,which supports the idea that a homogeneous spatial distribution of caves exists in grasslands.Moreover,we found no correlation between the distance to the boundary,and the number of insect species and individuals in grassland and the number of insect individuals in sand dunes;however,a significantly negative correlation was obtained between the distance to the boundary and the number of insect species in sand dunes.Therefore,our results concluded that a close relationship exists between the spatial distribution of active caves of P.guinanensis and the distribution of insect species in sand dunes.(2)Growth,survivorship,sex ratio and sexual size dimorphism of P.guinanensisUsing mark-recapture data from 2014 to 2016,we investigated the development of sex-specific survivorship,sex ratios,growth rates and SSD in P.guinanensis.The results indicated that the adult sex ratio(male: female)in this population was about 1:2.8.Male hatchlings suffered lower survival rate(8%)than females(38%),while male juveniles experienced higher survival rate(75%)than females(37%);the life expectancy of male hatchlings was lower than females,but male juveniles had a higher life expectancy than females.This suggested that the female-biased sex ratio might be related to the obviously higher mortality in male hatchlings.Male-biased SSD in tail length and head width were found only in adults rather than in hatchling or juvenile lizards.Size increases in body dimensions were undistinguishable between both sexes during the period from hatchling to juvenile,but increases of head length were significantly greater in males during development from juveniles to adults.Mean growth rates of all measurements during the period from hatchling to juvenile were relatively larger than that from juvenile to adult.However,only increases in tail length,head width,abdomen length,abdomen width in females and snout-vent length in males were significantly different between two stages.Our results concluded that female-biased sex ratio of this population is correlated with higher morality in hatchlings males while male biased SSD in head size occurred in adult males.(3)Sequence variation in the MC1 R gene between two closely related lizards,P.putjatia and P.guinanensisWe cloned and sequenced the total MC1 R gene CDS of two closely related lizards,P.putjatia and P.guinanensis,which were regarded as the same species in recent molecular systematic study,and possessed significant color and geographical variation between them.Our results showed that the overall length of MC1 R gene was 942 bp,41 nucleotide variable sites and 8 amino acid variable sites were identified in 76 individuals.Fisher exact test indicated that only the 28 th amino acid variable site(Arg,Gln)was statistically significantly associated(P=0.004)with the color variation between P.putjatia and P.guinanensis.The MC1 R gene network revealed there wasn't significant divergence between P.putjatia and P.guinanensis.Our results confirmed that a specific coding locus in MC1 R gene was statistically significant associated with color variation between two species,but the variation could not well explain the color variation in P.putjatia and P.guinanensis due to the absence of absolute relationship between the residual polymorphism and color phenotypes for many individuals.The coloration of two species might be result from multiple genetic factors or other mechanism.(4)Prediction of potential geographic distribution for P.guinanensis and P.putjatia based on MAXENTNine distributing sites of P.guinanensis and nineteen distributing sites of P.putjatia were collected from field investigation and literatures,and the MAXENT ecological niche model was utilized to predict the potential distribution of P.guinanensis and P.putjatia by importing corresponding data on 19 biological environmental variables and elevational variable downloaded from WORLDCLIM database for above each area.The suitable habitat(0.5<P<1.0)for P.guinanensis was mainly concentrated in the junction between Qinghai,Gansu and Sichuan.The suitable habitat(0.5<P<1.0)for P.putjatia mainly included northeastern part of Qinghai and some portion of southeast Gansu.Since Last Inter-glacial,suitable habitats of P.guinanensis and P.putjatia were in related with the dispersion of Qinghai-Tibet Plateau desert,while the suitable habitats in Last Glacial Maximum and Mid-Holocene were similar to current habitats.The AUC values of two models for both training and test data predicted by MAXENT were larger than 0.9,indicating high accuracy and reliability of our predicted result.Jackknife test in MAXENT ecological niche model showed that the contribution of different environmental variables to two models were different,the altitude(contribution,36.2%)and monthly mean diurnal temperature(contribution,20.9%)were primary factors that affect the distribution of P.guinanensis,while the altitude(contribution,50.3%)and precipitation of direst quarter(contribution,24.9%)were main factors for P.putjatia.(5)Analysis of population differentiation between P.guinanensis and P.putjatia based on ddRAD-seqWe used ddRAD-seq(double digest restriction-site associated DNA sequencing)analyses to examine the population divergence between two putative species with morphological divergence,P.guinanensis and P.putjatia.A total of 20361 alleles were obtained from 18 samples of 7 P.guinanensis populations and 25 samples of 10 P.putjatia populations in this study.The result of cross-validation on variance of sequences showed that the proportion of successful outcome prediction achieved the highest value when retained the first 2 principal components.The result of Bayesian Information Criterion analysis showed that the value of BIC reached the lowest value when retained 3 clusters.Discriminant analysis of principal components showed that 17 populations were separated into 3 groups.Furthermore,the group of P.putjatia was splitted into two population clusters,distributing in Qinghai Lake Basin(QLB),two populations of P.putjatia from Guide and eastern Gonghe basin were clustering with all P.guinanensis populations as a single cluster.Moreever,two individuals respectively from the eastern Gonghe basin and the southern distribution area of P.guinanensis were identified with P.putjatia mtDNA haplotypes and P.vlangalii nuclear genetypes.Our result supported an early phylogeographic and systematic study with the conclusion that P.guinanensis and P.putjatia were not genetic independent species.