【摘要】 【目的】本研究旨在明确共生菌Wolbachia属中特定菌株对宿主适合度的影响及分子机制。【方法】以20 mg/mL盐酸四环素蜂蜜溶液连续饲喂处理短管赤眼蜂Trichogramma pretiosum孤雌产雌品系3代成蜂，计算寄生率、羽化率和雌性比，并利用绝对定量qPCR(absolute quantitation qPCR, AQ-PCR)对短管赤眼蜂孤雌产雌品系成蜂体内Wolbachia属的wsp的lg(绝对拷贝数)进行检测，分析Wolbachia的滴度；利用Illumina平台以Wolbachia基因组作为参考基因组进行短管赤眼蜂孤雌产雌品系成蜂体内Wolbachia属的转录组测序，盐酸四环素处理前后短管赤眼蜂孤雌产雌品系成蜂体内Wolbachia转录组中筛选差异表达基因并进行GO/KEGG/COG数据库分析；利用RT-qPCR检测短管赤眼蜂孤雌产雌品系成蜂中Wolbachia属的5个基因(w1260,w0535,w3585,w4570和w2910)的表达量。【结果】盐酸四环素处理组短管赤眼蜂孤雌产雌品系成蜂寄生率、羽化率、雌性比和体内wsp的lg(绝对拷贝数)均显著低于对照组且随世代增加显著下降。从盐酸四环素处理前后的短管赤眼蜂孤雌产雌品系成蜂体内Wolbachia转录组中共鉴定出28个差异表达基因(6个下调)且显著富集于代谢过程、氧化磷酸化、能量产生与转换等多个与生长发育相关的通路，6个下调基因经NCBI Blast比对到3个Wolbachia菌株wCIFem,wfur和wSca。RT-qPCR验证表明，盐酸四环素处理后孤雌产雌成蜂体内5个差异表达基因(w1260,w0535,w3585,w4570和w2910)表达趋势与转录组测序结果一致。【结论】Wolbachia滴度降低通过抑制关键代谢通路(如氧化磷酸化)及清除功能菌株(wCIFem,wfur和wSca),削弱宿主能量供给与发育效率，导致生殖适合度下降。本研究首次以Wolbachia基因组为参考基因组解析共生菌-宿主互作的分子机制，研究结果表明Wolbachia通过菌株特异性基因网络维持共生稳态，揭示了Wolbachia中不同菌株通过多通路调控宿主生殖适合度的分子机制，为建立基于共生菌基因组的靶向调控技术体系提供理论支持。更多还原

【Abstract】 【Aim】 The aim of this study is to clarify the effect of specific strains in the symbiotic bacterium Wolbachia on the host fitness and its molecular mechanism. 【Methods】 The 3rd generation of parthenogenetic adult Trichogramma pretiosum was continuously fed with 20 mg/mL tetracycline hydrochloride honey solution. The parasitism rate, emergence rate and proportion of females were calculated. The lg(absolute copy number) of wsp of Wolbachia in parthenogenetic adult T. pretiosum was detected by absolute quantitation qPCR(AQ-PCR) to analyze the Wolbachia titer. The Illumina platform was used to sequence the transcriptome of Wolbachia in the parthenogenetic adult T. pretiosum with the Wolbachia genome as the reference genome, and the differentially expressed genes in the transcriptome of Wolbachia in parthenogenetic adult T. pretiosum before and after tetracycline hydrochloride treatment were screened and analyzed by databases GO/KEGG/COG. RT-qPCR was used to detect the expression levels of five genes(w1260, w0535, w3585, w4570 and w2910) of Wolbachia in the parthenogenetic adult T. pretiosum. 【Results】 The parasitism rate, emergence rate, proportion of females and the lg(absolute copy number) of wsp in the parthenogenetic adult T. pretiosum in the tetracycline hydrochloride treatment group were significantly lower than those in the control group and decreased significantly with the increase of generation. A total of 28 differentially expressed genes(6 down-regulated) were identified from the transcriptome of Wolbachia in parthenogenetic adult T. pretiosum before and after tetracycline hydrochloride treatment and significantly enriched in multiple pathways related to growth and development, such as metabolic process, oxidative phosphorylation, energy production and conversion. Six down-regulated genes were aligned to three Wolbachia strains, wCIFem, wfur and wSca by NCBI Blast. RT-qPCR verification showed that the expression trends of five differentially expressed genes(w1260, w0535, w3585, w4570 and w2910) in the parthenogenetic adults after tetracycline hydrochloride treatment were consistent with the transcriptome sequencing results.【Conclusion】 The decreased titer of Wolbachia weakens the energy supply and development efficiency of the host by inhibiting the key metabolic pathways(such as oxidative phosphorylation) and scavenging functional strains(wCIFem, wfur and wSca), resulting in a decrease in reproductive fitness. In this study, the Wolbachia genome was used as a reference genome to analyze the molecular mechanism of symbiotic bacteria-host interaction for the first time. The results showed that Wolbachia maintained symbiotic homeostasis through strain-specific gene network, revealing the molecular mechanism of different strains of Wolbachia regulating host reproductive fitness through multiple pathways, and providing theoretical support for the establishment of targeted regulation technology system based on symbiotic bacterial genome.更多还原