{"doi":"10.1016/j.jip.2025.108346","title":"Targeted knockdown of in vitro candidates does not alter Wolbachia density in vivo","abstract":"• Wolbachia is used in biocontrol, but key host phenotypes it induces and the drivers of associated host fitness costs remain poorly understood. • We test candidate genes shown to alter Wolbachia density in an RNAi screen in cell culture in vivo in D. melanogaster using the Gal4-UAS system. • Out of 14 candidates tested, seven were lethal to the host, and two were successfully knocked down but did not alter Wolbachia density. • We find that variation in Wolbachia density across tissues may underlie the discordance between in vitro and in vivo results. • This area warrants further study, as modulating Wolbachia density in vivo would provide a powerful tool to dissect Wolbachia -host interactions. The bacterial endosymbiont Wolbachia has emerged as an effective biocontrol method to reduce arbovirus transmission. Transinfection of w Mel Wolbachia from Drosophila melanogaster to Aedes aegypti results in the transfer of important Wolbachia -induced phenotypes including the reproductive modification, cytoplasmic incompatibility, and inhibition of viruses including dengue and chikungunya. However, the mechanisms underlying these critical traits as well other Wolbachia -host interactions are still not fully understood. Recently an in vitro genome wide RNAi screen was performed on w Mel-infected Drosophila S2 cells and identified large cohorts of host genes that alter w Mel density when targeted. If these findings can be replicated in vivo , this would provide a powerful tool for modulating w Mel density both systemically and in a tissue-specific manner allowing for interrogation of w Mel-host interactions. Here, we used the GAL4/UAS system to express RNAi molecules targeting host gene candidates previously identified to dysregulate w Mel density in vitro . We found systemic knockdown of two candidate D. melanogaster genes does not lead to w Mel density dysregulation. To explore the lack of consistency between our study and previous work, we also examined native tissue-specific density of w Mel in D. melanogaster . We show density is varied between tissues and find that individual tissue densities are not reliable linear predictors of other tissue densities. Our results demonstrate the complexities of implementing in vitro findings in systemic applications.","journal":"Journal of Invertebrate Pathology","year":2025,"id":564379,"datarank":0.0,"base_score":0.0,"endowment":0.0,"self_citation_contribution":0.0,"citation_network_contribution":0.0,"self_endowment_contribution":0.0,"citer_contribution":0.0,"corpus_percentile":null,"corpus_rank":null,"citation_count":0,"citer_count":0,"citers_with_citation_signal":0,"citers_with_endowment":0,"datacite_reuse_total":0,"is_dataset":false,"is_dataset_confidence":0.9549,"is_data_producer":false,"deposit_databanks":null,"is_oa":true,"file_count":0,"downloads":0,"has_version_chain":false,"published_date":"2025-01-01","fair_score":null,"fair_percentile":null,"algorithm_id":"datarank_citation_only_1hop_v6","ranking_scope":"data_only","authors":[{"id":345178,"name":"Johanna M. Duyvestyn","orcid":"0000-0001-9163-2318","position":1,"is_corresponding":false},{"id":345179,"name":"Heather A. Flores","orcid":"0000-0001-5003-7547","position":2,"is_corresponding":false},{"id":1464699,"name":"Kimberley R. Dainty","orcid":"0000-0001-7960-1615","position":0,"is_corresponding":true}],"reference_count":41,"raw_metadata":null,"created_at":"2026-07-19T02:56:20.933088Z","pmid":"40252916","pmcid":null,"fwci":null,"citation_percentile":null,"influential_citations":0,"oa_status":null,"license":null,"views":0,"total_file_size_bytes":0,"version_count":0,"fair_f":null,"fair_a":null,"fair_i":null,"fair_r":null,"fair_zscore":null,"fair_rationale":null,"fair_model":null,"fair_agent_version":null,"fair_fulltext_source":null,"fair_has_llm":null,"fair_computed_at":null,"clinical_trials":[],"software_tools":[],"db_accessions":[],"linked_datasets":[],"topics":[]}