CRISPR/Cas9-mediated gene deletion of the ompA gene in symbiotic Cedecea neteri impairs biofilm formation and reduces gut colonization of Aedes aegypti mosquitoes
Hegde, Shivanand and Nilyanimit, Pornjarim and Kozlova, Elena and Anderson, Enyia R. and Narra, Hema P. and Sahni, Sanjeev K. and Heinz, Eva and Hughes, Grant L. (2019) CRISPR/Cas9-mediated gene deletion of the ompA gene in symbiotic Cedecea neteri impairs biofilm formation and reduces gut colonization of Aedes aegypti mosquitoes. PLOS Neglected Tropical Diseases, 13 (12). e0007883. ISSN 1935-2727 (https://doi.org/10.1371/JOURNAL.PNTD.0007883)
Preview |
Text.
Filename: Hedge-etal-PLOS-NTD-2019-Crispr-Cas9-mediated-gene-deletion-of-the-ompA-gene.pdf
Final Published Version License: Download (2MB)| Preview |
Abstract
Background Symbiotic bacteria are pervasive in mosquitoes and their presence can influence many host phenotypes that affect vectoral capacity. While it is evident that environmental and host genetic factors contribute in shaping the microbiome of mosquitoes, we have a poor under-standing regarding how bacterial genetics affects colonization of the mosquito gut. The CRISPR/Cas9 gene editing system is a powerful tool to alter bacterial genomes facilitating investigations into host-microbe interactions but has yet to be applied to insect symbionts. Methodology/Principal findings To investigate the role of bacterial genetic factors in mosquito biology and in colonization of mosquitoes we used CRISPR/Cas9 gene editing system to mutate the outer membrane protein A (ompA) gene of a Cedecea neteri symbiont isolated from Aedes mosquitoes. The ompA mutant had an impaired ability to form biofilms and poorly infected Ae. aegypti when reared in a mono-association under gnotobiotic conditions. In adult mosquitoes, the mutant had a significantly reduced infection prevalence compared to the wild type or complement strains, while no differences in prevalence were seen in larvae, suggesting genetic factors are particularly important for adult gut colonization. We also used the CRISPR/Cas9 system to integrate genes (antibiotic resistance and fluorescent markers) into the symbionts genome and demonstrated that these genes were functional in vitro and in vivo. Conclusions/Significance Our results shed insights into the role of ompA gene in host-microbe interactions in Ae. aegypti and confirm that CRISPR/Cas9 gene editing can be employed for genetic manipulation of non-model gut microbes. The ability to use this technology for site-specific integration of genes into the symbiont will facilitate the development of paratransgenic control strategies to interfere with arboviral pathogens such Chikungunya, dengue, Zika and Yellow fever viruses transmitted by Aedes mosquitoes.
ORCID iDs
Hegde, Shivanand, Nilyanimit, Pornjarim, Kozlova, Elena, Anderson, Enyia R., Narra, Hema P., Sahni, Sanjeev K., Heinz, Eva ORCID: https://orcid.org/0000-0003-4413-3756 and Hughes, Grant L.;-
-
Item type: Article ID code: 90683 Dates: DateEvent2 December 2019Published26 October 2019AcceptedSubjects: Medicine > Pharmacy and materia medica Department: Faculty of Science > Strathclyde Institute of Pharmacy and Biomedical Sciences Depositing user: Pure Administrator Date deposited: 24 Sep 2024 11:53 Last modified: 25 Nov 2024 04:04 Related URLs: URI: https://strathprints.strath.ac.uk/id/eprint/90683