Isolation Of Microsatellite Marker And Asexual Queen Succession System In Reticulitermes Aculabialis

In this study,the genomic sequenece library and microsatellite library of Reticulitermes aculabialis was set up on the basis of Illumina sequencing.The microsatellite primers were designed and the genetic diversity of R.aculabialis populations was analyzede by using 6 pairs of microsatellite markers.Meanwhile,we also used microsatellite markers to study the breeding system of nymphoids in Xi’an and Nanjing.By doing this we indicate that the asexual queen succession exists in R.aculabialis.The main findings are as follows:(1)A total of 147 microsatellite primers were designed to amplify their polymorphism in R.aculabialis and the related species of R.labralis.The amplified results showed that 22 pairs and 21 pairs of primers were able to obtain stable and effective amplified bands,among which 16 pairs and 18 pairs showed good polymorphism,respectively.(2)A total of 55 and 115 alleles were detected in 16 microsatellite loci of R.aculabialis and 18 microsatellite loci of R.labralis,respectively.The mean number of alleles was 3.438 and 6.389 respectively,and the mean observed heterozygosity was 0.481 and 0.368 respectively.The mean expected heterozygosity was 0.542 and 0.702 respectively,and the average polymorphic information content(PIC)was 0.478 and 0.663 respectively.The alleles ranged from 2 to 6 were found in 16 pairs of primers,and the most allele loci was 6 in Ra116 and Ra144,while alleles ranged from 3 to 9 at 18 pairs of primers,and the most allele loci was 9 in Ra 50 and Ra 91,indicating that these microsatellite loci have high polymorphism in R.aculabialis and R.labralis,respectively.These microsatellite markers will provide an effective molecular tool for the future study of genetic diversity of termites.(3)The genetic diversity of R.aculabialis in 2 natural populations was also analyzed using the 3 pairs of primers we screened and the published three pairs of primers.The results showed that the number of alleles(Na)ranged from 1.710 to 3,the average was 2.075,the number of effective alleles ranged from 1.431 to 2.441,the average was 1.747,and the observed heterozygosity(Ho)ranged from 0.363 to 0.774,the average observed heterozygosity was 0.483,the expected heterozygosity(He)ranged from 0.252 to 0.558,the average expected heterozygosity was 0.366,the diversity index(1)ranged from 0.379 to 0.939,the average was 0.566,indicating that the genetic diversity of population in Xi’an was at a moderate but relatively low level,and the degree of genetic variation was not high.The number of alleles(Na)ranged from 1.552 to 2.793,the average was 2.011,the number of effective alleles(Ne)ranged from 1.326 to 2.337,the average was 1.705,and the observed heterozygosity(Ho)ranged from 0.190 to 0.638,the average observed heterozygosity was 0.445,the expected heterozygosity(He)ranged from 0.190 to 0.502,the average expected heterozygosity was 0.366,the polymorphism index(I)ranged from 0.335 to 0.850,the average was 0.534,indicating that there was moderate and low genetic diversity within the Nanjing’s population,and the degree of genetic variation was not high.In summary,the genetic diversity within the population of Xi’an and Nanjing were basically the same.There was significant genetic differentiation(Fst=0.409)between the two populations,but the gene flow between the two populations was low(Nm=0.375)in the populations of Xi’an and Nanjing,which were analyzed by GenalEx software.(4)The frequency of parthenogenesis were analyzed in the 4 natural colonies of R.aculabialis in Xi’an and Nanjing,respectively.We found 85 and 30 female nymphoid neotenics in the 2 natural colonies in Xi’an and Nanjing,respectively.Out of a total of 59 secondary queens genotyped,and fifty-five individual were homozygous at all six microsatellite loci,indicating that the frequency of parthenogenesis was 93.2%.Of the four individuals classified as parthenogens,which were heterozygous at only ranged from one to three locus,respectively.Thus,it is possible that these secondary queens were actually parthenogenetically produced and were heterozygous owning to crossing over at these loci.Only one of 40(2.5%)workers and only two of 30(6.7%)alates were parthenogens,while all of 20 nymphs and 20 soldiers were heterozygous at six loci and were sexually produced,It can be considered that large colonies of R.aculabialis are indeed headed by numerous neotenic females,nearly all of which are parthenogenetic daughters of the queen.These female neotenics mate with a single king to produce the other colony members through sexual reproduction,and the AQS system is only available when the parthenogens is preferred to become a brachypterous female neotenics.Thus,this is the sixth termite species demonstrated to have the remarkerable AQS system.