Wolbachia pipientis is the most widespread endosymbiotic bacterium, known for its ability to alter host reproduction. In Naupactini weevils, their reproductive mode appears to be linked to Wolbachia density, whereby species with low levels or an absence of Wolbachia reproduce sexually, while high-density infections would promote parthenogenesis. Although the factors driving high Wolbachia loads remain unclear, evidence suggests that the diversity of the host’s microbiota may influence Wolbachia abundance. This study explored the bacterial microbiota of Naupactini species with contrasting reproductive modes and Wolbachia densities using targeted metagenomics. The species analysed included Naupactus dissimulator and Naupactus xanthographus (sexually reproducing), as well as Naupactus versatilis and Pantomorus postfasciatus (parthenogenetic), all of which were infected with the dominant strain within the tribe, i.e., wNau1. Notably, P. postfasciatus comprises both parthenogenetic (PpP) and sexually reproducing populations (PpS), providing unique insights into factors influencing Wolbachia colonization.

A total of 1,630,618 high-quality reads yielded 923 operational taxonomic units (OTUs) across 106 bacterial genera. The four most abundant genera, Wolbachia (26.47%), Spiroplasma (20.35%), Enterobacter (13.88%), and Rickettsia (11.49%), accounted for 72% of all reads. Parthenogenetic populations showed a marked predominance of Wolbachia and Rickettsia, whereas Spiroplasma and Enterobacter were the most prevalent symbionts in sexual populations, which also exhibited a richer microbiota in terms of the genera identified. Principal Coordinate Analysis based on Bray–Curtis distances clearly distinguished populations by reproductive mode. PpS’ bacterial microbiota differed significantly from both PpP and other sexually reproducing species, exhibiting a profile that combined features of both groups. The low relative abundance of Wolbachia, coupled with the increased levels of Enterobacter and Spiroplasma, distinguished it from parthenogenetic populations, while elevated Rickettsia levels set it apart from other sexually reproducing species. These observations suggest that Enterobacter and Spiroplasma may inhibit Wolbachia proliferation, whereas Rickettsia might promote its growth.