The Infection And Transmission Mode And MLST Analysis Of Endosymbionts Wolbachia In Several Cynipini And Synergini Gall Wasps

Wolbachia are maternally inherited endosymbiotic bacteria. Recent study of Wolbachia incidence estimated that20-76%of insect species are infected with Wolbachia, and Wolbachia may be the most abundant and widespread endosymbiotic bacteria on earth. They could manipulate the reproduction of their hosts by causing cytoplasmic incompatibility, parthenogenesis induction, feminization of genetic males and male-killing. Cynipinae (Hymenoptera:Cynipoidea), which comprises about1400described species, are phytophagous species.29gall wasp species belonging to11genera had been recorded in China (including Taiwan).In this paper, I collected plant galls of gall wasps in southern China and used PCR to detect the infection of Wolbachia in gall wasps. The phylogeny of Wolbachia in gall wasps was analysed by wsp gene and Multi Locus Sequence Typing (MLST). The transmission mode of Wolbachia and its effect on the reproduction mode of gall wasps were discussed.The primary achievements of this dissertation are listed as follows:(1)14plant galls of gall wasps were collected from Changsha, Yueyang, Loudi, Changde, Shaoyang, Yiyang in Hunan, Jian in Jiangxi, Shaoguang in Guangdong, Anqing in Anhui, Ningde in Fujian and LinZhou in Henan of China. Their host plants belonged to Castanea, Quercus, Lithocarpus and Castanopsis of Fagaceae and Rosa of Rosaceae. The15gall wasps were reared from these plant galls, and10of15gall wasps were new species:Dryocosmus zhuili Liu, ms; Cynips crystaloides Liu, ms; Synergus intricatus Liu, ms; Trigonaspis acuminatus Liu, ms; Andricus elodeoides Liu, ms; Synergus henanensis Liu, ms; Synergus carinatus Liu, ms; Saphonecrus glaberus Liu, ms; Synergus brutus Liu, ms; Saphonecrus hupingshanensis Liu, ms. A. mukaigawae, Synergus japonicus and Diplolepis japonica were recorded for the first time in China. Andricus mairei and Dryocosmus kuriphilus had been recorded in China.(2) Our survey results showed Wolbachia infection rate Andricus mairei in Yueyang, Changsha, and Shaoyang populations:100%for the male wasps from all populations, while,100%,100%and80%for the Yueyang, Changsha and Shaoyang populations of the female wasps, respectively. The sequencing results showed that the wsp gene from all the three populations has the same sequence. Our study discovered strikingly male-biased sex ratios in3populations of A. mairei. Females only accounted for15.3%,12.1%and19.8%of the Yueyang, Changsha and Shaoyang populations, respectively. Thus, these results suggested that the Wolbachia did not induce parthenogenesis in A. mairei.(3) Plant galls of Andricus mukaigawae were collected from eight locations in China. A. mukaigawae (gall marker), Synergus japonicus (inquiline) and Torymus sp.(parasitic wasp) emerged from plant galls of A. mukaigawae. The infection rate of Wolbachia in A. mukaigawae, S. japonicus and Torymus sp. was60%,37.1%and100%, respectively. The high levels of multiple Wolbachia infection was found in A. mukaigawae and its associated inquiline S. japonicus, with5and8Wolbachia strains, respectively. The Torymus sp. was single infection with Wolbachia. The results showed obvious recombination of wsp gene and revealed3recombination groups for Wolbachia in A. mukaigawae communities. The results suggested the horizontal transmission of Wolbachia between A. mukaigawae and S. japonicus according to the fact that A. mukaigawae and S. japonicus shared5of9Wolbachia strains in A. mukaigawae communities. The results suggested that horizontal transmission and recombination played an important role in the high levels of multiple Wolbachia infection in A. mukaigawae and S. japonicus.(4) The results revealed Wolbachia infection in A. mukaigawae, A. mairei, Dryocosmus zhuili Liu, ms; Cynips crystaloides Liu, ms; Synergus japonicus, Synergus intricatus Liu, ms; Saphonecrus glaberus Liu, ms and Synergus brutus Liu, ms by PCR detection. The Wolbachia incidence of15gall wasps was53.3%. The infection ratio of A. mairei was the highest (100%), and that of S. brutus was lowest (26.7%).(5)11new Wolbachia strains were found in9gall wasps using conserved gatB-. coxA、hcpA、ftsL and fbpA genes according to MLST analysis. The11strains had39MLST alleles, and18of39MLST alleles were novel alleles. The gatB, coxA, hep A, ftsZ and fbpA genes of MLST genes had4,4,5,2and3novel alleles, respectively. The MLST phylogenetic tree of Wolbachia and CO I phylogenetic tree of gall wasps were constructed, and the MLST tree and CO I tree were incongruent. Some Wolbachia strains sharing same wsp gene had a high sequence differences of conserved MLST genes. Thus, the results suggested horizontal transmission of Wolbachia between gall wasps.