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Background  Kissing bugs (Triatominae) are blood-feeding insects best known as the vectors of  Trypanosoma cruzi , the causative agent of Chagas’ disease. Considering the high epidemiological relevance of these vectors, their biology and bacterial symbiosis remains surprisingly understudied. While previous investigations revealed generally low individual complexity but high among-individual variability of the triatomine microbiomes, any consistent microbiome determinants have not yet been identified across multiple Triatominae species.    Methods  To obtain a more comprehensive view of triatomine microbiomes, we investigated the host-microbiome relationship of five  Triatoma  species sampled from white-throated woodrat ( Neotoma albigula ) nests in multiple locations across the USA. We applied optimised 16S rRNA gene metabarcoding with a novel 18S rRNA gene blocking primer to a set of 170  T .  cruzi -negative individuals across all six instars.    Results  Triatomine gut microbiome composition is strongly influenced by three principal factors: ontogeny, species identity, and the environment. The microbiomes are characterised by significant loss in bacterial diversity throughout ontogenetic development. First instars possess the highest bacterial diversity while adult microbiomes are routinely dominated by a single taxon. Primarily, the bacterial genus  Dietzia  dominates late-stage nymphs and adults of  T .  rubida ,  T .  protracta , and  T .  lecticularia  but is not present in the phylogenetically more distant  T .  gerstaeckeri  and  T .  sanguisuga . Species-specific microbiome composition, particularly pronounced in early instars, is further modulated by locality-specific effects. In addition, pathogenic bacteria of the genus  Bartonella , acquired from the vertebrate hosts, are an abundant component of  Triatoma  microbiomes.    Conclusion  Our study is the first to demonstrate deterministic patterns in microbiome composition among all life stages and multiple  Triatoma  species. We hypothesise that triatomine microbiome assemblages are produced by species- and life stage-dependent uptake of environmental bacteria and multiple indirect transmission strategies that promote bacterial transfer between individuals. Altogether, our study highlights the complexity of Triatominae symbiosis with bacteria and warrant further investigation to understand microbiome function in these important vectors.Video abstract.

Ontogeny, species identity, and environment dominate microbiome dynamics in wild populations of kissing bugs (Triatominae)