Wolbachia and virus alter the host transcriptome at the interface of nucleotide metabolism pathways. Wolbachia and virus alter the host transcriptome at the interface of nucleotide metabolism pathways

Wolbachia is a maternally transmitted bacterium that manipulates arthropod and nematode biology in myriad ways. The Wolbachia strain colonizing Drosophila melanogaster creates sperm-egg incompatibilities and protects its host against RNA viruses, making it a promising tool for vector control. Despite successful trials using Wolbachia-transfected mosquitoes for Dengue control, knowledge of how Wolbachia and viruses jointly affect insect biology remains limited. Using the Drosophila melanogaster model, transcriptomics and gene expression network analyses revealed pathways with altered expression and splicing due to Wolbachia colonization and virus infection. Included are metabolic pathways previously unknown to be important for Wolbachia-host interactions. Additionally, Wolbachia-colonized flies exhibit a dampened transcriptomic response to virus infection, consistent with early blocking of virus replication. Finally, using Drosophila genetics, we show Wolbachia and expression of nucleotide metabolism genes have interactive effects on virus replication. Understanding the mechanisms of pathogen blocking will contribute to the effective development of Wolbachia-mediated vector control programs. Overall design: To determine the effect of Wolbachia and virus infection on fly gene expression, we established in vivo systemic viral infections in adult Drosophila, using a block design with a time series. Flies with or without Wolbachia (W+/W-), were injected with either virus or saline (SINV+/SINV-), and collected at 6, 24, and 48 hours post-injection (hpi). For each unique condition of W-SINV-time, we generated four biological replicates (A-D), with each replicate consisting of a pool of five virgin females. In total, 48 libraries were generated.

沃尔巴克氏体（Wolbachia）是一类母系传播的细菌，可通过多种方式调控节肢动物与线虫的生物学特性。侵染黑腹果蝇（Drosophila melanogaster）的沃尔巴克氏体菌株可引发精卵不相容性，并为宿主提供RNA病毒感染抗性，使其成为媒介生物防控的极具应用前景的工具。尽管利用转染沃尔巴克氏体的蚊子开展登革热防控的试验已取得成功，但学界对沃尔巴克氏体与病毒共同调控昆虫生物学的机制仍知之甚少。
本研究以黑腹果蝇为模型，通过转录组学与基因表达网络分析，鉴定出受沃尔巴克氏体定殖和病毒感染共同影响的表达与剪接异常通路，其中包含此前未被发现与沃尔巴克氏体-宿主互作相关的代谢通路。此外，携带沃尔巴克氏体的果蝇对病毒感染的转录组响应显著减弱，这与病毒复制早期被阻断的表型相符。最后，借助果蝇遗传学实验，我们证实沃尔巴克氏体与核苷酸代谢基因的表达在病毒复制过程中存在交互调控效应。阐明病原体阻断机制将有助于推动沃尔巴克氏体介导的媒介生物防控策略的高效开发。
总体实验设计：为探究沃尔巴克氏体与病毒感染对果蝇基因表达的影响，我们建立了成虫系统性病毒感染的体内模型，采用区组设计结合时间序列方案。将携带或不携带沃尔巴克氏体（W+/W-）的果蝇分别注射病毒或生理盐水（SINV+/SINV-），并于注射后6、24、48小时（hpi）收集样本。针对每一组包含沃尔巴克氏体定殖状态、病毒感染状态与采样时间点的独特实验条件，我们设置了4个生物学重复（A-D），每个重复由5只处女雌蝇混合制备。最终共构建48个测序文库。