| The question of how Wolbachia manipulate the thelytokous phenotype by a molecular mechanism is still unresolved,mainly for the reason that it is difficult to answer whether Wolbachia or the hosts are responsible for process of diploidization or feminization when the thelytoky is the result of interaction between Wolbachia and hosts.Moreover,what makes this discrimination even harder is that hosts might rely on or inherit the sex manipulation?i.e.,diploidization and feminization?of Wolbachia for the perennial co-evolution.The study of Wolbachia-induced thelytoky may serve to the biocontrol.Thelytokous parasitoid wasps are generally considered to be better biocontrol agents,they,however,could also have decreased fitness caused by Wolbachia and thus affecting the biocontrol efficiency.If the thelytokous mechanism applied by Wolbachia is elucidated,it might be possible to create excellent parasites with both the merits of being thelytoky and the avoidance of negative effects from Wolbachia.The Trichogramma dendrolimi,used in this study,is one of the sublimely successful biocontrol agents.Therefore,studying its thelytokous mechanism manipulated by Wolbachia might help improve its biocontrol potential and then elevates the efficiency and efficacy of biocontrol.Under the background above,current study explored the mechanism of Wolbachia-manipulated thelytoky and the sex determination mechanism employed by T.dendrolimi.Corresponding results were obtained.1.Since the variation of ploidy can reflect how Wolbachia manipulate hosts' sex mechanism,current study firstly determined the approach to the ploidy detection of T.dendrolimi,to provide technique that supported the subsequent study.q PCR was a more proper and efficient tool for detecting the ploidy of T.dendrolimi.When Fox O was as reference gene,the relative contents of COI in haploid and diploid wasps can be distinguished by different and stable ranges of 2-?Cq.Moreover,the ploidy of T.dendrolimi and Wolbachia-infection did not affect the content of mitochondrion.2.An efficient tool without non-target effects to explore the interaction between Wolbachia and their hosts was then decided.Tetracycline or rifampicin didn't affect factors like brood size,emergence rate,sex ratio,longevity and overall development duration when the concentrations was 0.5 mg/ml or 0.1 mg/ml respectively,but they caused substantial deaths under the concentration of 1 mg/ml.Within 0.25-0.5mg/ml,rifampicin showed significantly negative effects on brood size,emergence rate and sex ratio.Either in comparison to rifampicin or to heat,tetracycline was the most efficient method to induce thelytokous wasps to produce abnormal individuals?male +/ intersex?.Furthermore,there was a significantly negative correlation between either antibiotic concentration or heat gradient and the expression level of thelytokous phenotype.Wolbachia brought off manipulation of sex mechanism before meiosis.3.Current study confirmed that thelytoky of T.dendrolimi was manipulated by Wolbachia and also found that: bisexual and thelytokous strain from a same population had congenetic background;Wolbachia in thelytokous strain belonged to Sib group in B supergroup and caused the diversity of ITS-2 in current population of T.dendrolimi,i.e.,Wolbachia drived the evolution of ITS-2.The thelytoky was very stable and therefore provided possibility of controlling first and second generation of Asian corn borer.4.Current study demonstrated that Wolbachia manipulated thelytoky of T.dendrolimi by a strict two-step mechanism,whereby each step correlated with a threshold of titer: under lower titer the diploidization of unfertilized egg was firstly induced and followed by feminization under a higher titer.In addition,the thelytokous wasps neither inherited nor depended on the sex manipulation of Wolbachia.5.This study manifested that Wolbachia employed a gamete duplication mechanism to induce the diploidization of unfertilized eggs,by which all of the loci of hosts will become homozygous.The thelytokous wasps could use the sperm,and then the consequent zygotes conformed a normal development in sexual reproduction,instead of going through the diploidization manipulated by Wolbachia.6.Current study reported the third case of PSR chromosome in Hymenopetrans and proved that this chromosome was the prime factor that constrained the spread of Wolbachia?or thelytokous wasps?in T.dendrolimi population of this study.The copulation from male wasps was as the secondary factor by attenuating the fertility of thelytokous wasps and furthermore,the males from same population had significantly mightier ability of inhibition than that of those from our laboratory,which suggested a longer-term co-existence of the former and Wolbachia.7.Provided the sex determination mechanism of T.dendrolimi is complementary sex determination?CSD?,it will undermine the conclusion of two-step mechanism by Wolbachia.Therefore,current study further investigated whether CSD was present in T.dendrolimi.Through 10 generations of continuous inbreed,the CSD up to 10 sex loci was rejected safely.Therefore,the sex determination mechanism of T.dendrolimi was not CSD.Briefly,the study determined q PCR as the appropriate approach to the ploidy detection of T.dendrolimi,decided tetracycline as the efficient tool without non-target effects to induce the production of abnormal individuals in thelytokous strain,demonstrated a two-step mechanism of thelytoky in T.dendrolimi employed by Wolbachia starting with diploidization followed by feminization,proved PSR chromosome and male copulation to be factors restraining the spread of Wolbachia in T.dendrolimi population,and excluded the possibility of CSD in T.dendrolimi. |
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