There is a surge in studies of the bee microbiome that is driven primarily by a growing concern for global bee decline (Potts et al., 2010, Zattara and Aizen, 2021, Ghisbain et al., 2023). This decline could be partly explained by health issues associated to dysbiosis (Engel et al., 2016, Voulgari-Kokota et al., 2019, Rothman et al., 2019). Social bees such as honey bees, bumble bees and stingless bees, host core and non-core microbiota that were reported to contribute to food digestion, detoxification, protection against parasites and pathogens, as well as to the provision of essential nutrients that support host fitness and homeostasis (Zhang and Zheng, 2022). While bacteria are present in the crop, midgut and hindgut of their bee hosts, around 99 % of bacterial colonization occurs in the hindgut where undigested remnants of pollen, sugars and nitrogenous waste products serve as nutrient sources (Zheng et al., 2017, Martinson et al., 2012). Compared to honey bee symbionts, bumble bee symbionts such as Gilliamella spp., Bifidobacterium spp. and Lactobacillus sensu lato have a more restricted polysaccharide-degrading enzyme repertoire, and therefore several roles of bumble bee symbiont bacteria remain speculative (Hammer et al., 2021, Ellegaard et al., 2019, Zheng et al., 2019).

Fructobacillus bacteria are generally considered non-core microbiota in social bees, but have been reported in bumble bee (Hammer et al., 2021), honey bee (Filannino et al., 2016) and carpenter bee (Handy et al., 2023) samples including the larval gut (Vojvodic et al., 2013, Rokop et al., 2015), brood cells and bee bread of honey bees (Rokop et al., 2015), honey of stingless bees (Andrade-Velasquez et al., 2023) and they dominated the microbial community in Brassica rapa floral nectar (Russell and McFrederick, 2022). Additionally, Fructobacillus fructosus has been reported as a core symbiont in another social insect, the Asian hornet (Hettiarachchi et al., 2023). Shifts in the gut microbiomes of bumbleANTbee workers have been reported to be characterized by the replacement of core symbionts by Fructobacillus bacteria (Krams et al., 2022, Zhang et al., 2021, Villabona et al., 2023) and such shifts have been considered a form of dysbiosis (Hammer et al., 2021). While some bumble bee workers with Fructobacillus-dominated microbiomes have been collected from habitats with higher levels of anthropogenic disturbances which may have provoked gut dysbiosis (Villabona et al., 2023), others were collected in forest meadows, which represent a habitat with the least anthropogenic impact (Krams et al., 2022).

Fructobacilli are fructophilic lactic acid bacteria (FLAB), which highlights their preference for fructose over glucose as a growth substrate. They are obligate FLAB, which implies that they prefer aerobic growth conditions and grow poorly on glucose unless an external electron acceptor such as pyruvate, oxygen or fructose is present (Endo et al., 2018). Although they are heterofermentative lactic acid bacteria, FLAB do not produce ethanol from glucose, because of a complete or partial deletion of the adhE gene that encodes a bifunctional alcohol/acetaldehyde dehydrogenase (Endo et al., 2014, Endo et al., 2018). To date, twelve Fructobacillus species have been formally named, of which F. fructosus is the type species (Endo and Okada, 2008). The latter was first described in 1956 and has been reported in a large variety of sources including insect samples, flowers, fruits and fermented foods (Mesas et al., 2011, Koch and Schmid-Hempel, 2011, Praet et al., 2018, Yaacob et al., 2018, He et al., 2011, Thaochan et al., 2010, Rodriguez et al., 2019, Veron et al., 2017, Antunes et al., 2002, Endo and Okada, 2008). Most other Fructobacillus species were described more recently and these too have been isolated from insect samples, flowers, fruits and fermented foods (Endo and Okada, 2008, Leisner et al., 2005, Rodriguez et al., 2019, Antunes et al., 2002, Chambel et al., 2006, Endo et al., 2009, Janashia and Alaux, 2016, Endo et al., 2011, Praet et al., 2016, Snauwaert et al., 2013, Lin et al., 2022, Gallus et al., 2022, Chen et al., 2022, Oliphant et al., 2023).

There is limited information on the diversity and functional role of Fructobacillus isolates in their insect hosts. Metagenetic studies based on the analysis of partial 16S rRNA gene fragments generally fail to provide species level discrimination, and several groups of closely related Fructobacillus species have virtually identical 16S rRNA gene sequences (Endo et al., 2011, Gallus et al., 2022, Oliphant et al., 2023). A comparative genomic and phylogenetic analysis of publicly available Fructobacillus genome sequences showed that these genomes represented two clades that differed considerably in amino acid biosynthetic potential (Mohamed et al., 2023). F. fructosus isolates from stingless bees and honey bees exhibited antagonistic activity against several pathogenic bacteria (Yaacob et al., 2018, Zeid et al., 2022). Heptyl 2-methylbutyrate, di-isobutyl phthalate, d-turanose, heptakis (trimethylsilyl), di-isooctyl phthalate, and hyodeoxycholic acid compounds were identified in fractions with biological activity against Paenibacillus larvae (Zeid et al., 2022). F. fructosus isolates from honey bee gut samples metabolized phenolic acids commonly found in pollen (Filannino et al., 2016) and Fructobacillus sp. isolates from honey bee hives promoted growth of honey bee core bacteria (Rokop et al., 2015). In the present study, we generated draft genome sequences of 21 Fructobacillus isolates previously isolated from five bumble bee species (Praet et al., 2018) and three flowers (unpublished data) and performed a comprehensive functional genomic analysis.