Abstract Wolbachia is a maternally inherited intracellular bacterium that infects a wide range of arthropods including mosquitoes. The endosymbiont is widely used in biocontrol strategies due to its capacity to modulate arthropod reproduction and limit pathogen transmission. Wolbachia infections in Culex spp. are generally assumed to be monoclonal but the potential presence of genetically distinct Wolbachia subpopulations within and between individual organs has not been investigated using whole genome sequencing. Here we reconstructed Wolbachia genomes from ovary and midgut metagenomes of single naturally infected Culex pipiens mosquitoes from Southern France to investigate patterns of intra- and inter-individual differences across mosquito organs. Our analyses revealed a remarkable degree of intra-individual conservancy among Wolbachia genomes from distinct organs of the same mosquito both at the level of gene presence–absence signal and single-nucleotide polymorphisms (SNPs). Yet, we identified several synonymous and non-synonymous substitutions between individuals, demonstrating the presence of some level of genomic heterogeneity among Wolbachia that infect the same C. pipiens field population. Overall, the absence of genetic heterogeneity within Wolbachia populations in a single individual confirms the presence of a dominant Wolbachia that is maintained under strong purifying forces of evolution.