Bacterial endosymbionts are common across insects, but we often lack a deeper knowledge of their prevalence across most organisms. Next-generation sequencing approaches can characterize bacterial diversity associated with a host and at the same time facilitate the fast and simultaneous screening of infectious bacteria. In this study, we used 16S rRNA tag encoded amplicon pyrosequencing to survey bacterial communities of 310 samples representing 221 individuals, 176 colonies and 95 species of ants. We found three distinct endosymbiont groups—Wolbachia (Alphaproteobacteria: Rickettsiales), Spiroplasma (Firmicutes: Entomoplasmatales), and relatives of Asaia (Alphaproteobacteria: Rhodospirillales)—at different infection frequencies (at the ant species level: 22.1%, 28.4%, and 14.7%, resp.) and relative abundances within bacterial communities (1.0%–99.9%). Spiroplasma was particularly enriched in the ant genus Polyrhachis, while Asaia relatives were most prevalent in arboreal ants of the genus Pseudomyrmex. While Wolbachia and Spiroplasma have been surveyed in ants before, Asaia, an acetic acid bacterium capable of fixing atmospheric nitrogen, has received much less attention. Due to sporadic prevalence across all ant taxa investigated, we hypothesize facultative associations for all three bacterial genera. Infection patterns are discussed in relation to potential adaptation of specific bacteria in certain ant groups. 1. Introduction Recent studies have shown that insects are associated with a broad range of unrelated microbial taxa [1, 2]. These interactions shape the ecology and evolution of hosts and bacterial symbionts and often heavily impact host biology [3, 4]. Congruent evolutionary histories between some symbiotic partners show the likely obligate nature of this relationship [5], while other associations occur sporadically and can vary both spatially and temporally [6]. Bacterial endosymbionts sometimes inhabit specialized host cells or structures [7, 8] and might even share metabolic pathways with their hosts [9], while others occur loosely in unspecific tissues or hemolymph [10]. Microbes associated with insects are extremely diverse and span-wide taxonomic groups, even within individual hosts. One of the best-characterized endosymbiont groups is comprised of insect-associated bacteria that increase the nutritive value of their hosts’ diets. These bacteria are often highly specialized and coevolved associates, playing particularly important roles in insects with nutritionally limited or deficient diets. Some well-known examples of such
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