Spatial ecology of Haemophilus and Aggregatibacter in the human oral cavity.

Haemophilus and Aggregatibacter are two of the most common bacterial genera in the human oral cavity, encompassing both commensals and pathogens of substantial ecological and medical significance. In this study, we conducted a metapangenomic analysis of oral Haemophilus and Aggregatibacter species to uncover genomic diversity, phylogenetic relationships, and habitat specialization within the human oral cavity. Using three metrics-pangenomic gene content, phylogenomics, and average nucleotide identity (ANI)-we first identified distinct species and sub-species groups among these genera. Mapping of metagenomic reads then revealed clear patterns of habitat specialization, such as Aggregatibacter species predominantly in dental plaque, a distinctive Haemophilus parainfluenzae sub-species group on the tongue dorsum, and H. sp. HMT-036 predominantly in keratinized gingiva and buccal mucosa. In addition, we found that supragingival plaque samples contained predominantly only one out of the three taxa, H. parainfluenzae, Aggregatibacter aphrophilus, and A. sp. HMT-458, suggesting independent niches or a competitive relationship. Functional analyses revealed the presence of key metabolic genes, such as oxaloacetate decarboxylase, correlated with habitat specialization, suggesting metabolic versatility as a driving force. Additionally, heme synthesis distinguishes H. sp. HMT-036 from closely related Haemophilus haemolyticus, suggesting that the availability of micronutrients, particularly iron, was important in the evolutionary ecology of these species. Overall, our study exemplifies the power of metapangenomics to identify factors that may affect ecological interactions within microbial communities, including genomic diversity, habitat specialization, and metabolic versatility. IMPORTANCE: Understanding the microbial ecology of the mouth is essential for comprehending human physiology. This study employs metapangenomics to reveal that various Haemophilus and Aggregatibacter species exhibit distinct ecological preferences within the oral cavity of healthy individuals, thereby supporting the site-specialist hypothesis. Additionally, it was observed that the gene pool of different Haemophilus species correlates with their ecological niches. These findings shed light on the significance of key metabolic functions in shaping microbial distribution patterns and interspecies interactions in the oral ecosystem.

Comparative analysis of 3 pollen sterilization methods for feeding bumble bees.

Pollen is an essential component of bee diets, and rearing bumble bees (Bombus spp.) for commercial use necessitates feeding pollen in mass quantities. This pollen is collected from honey bee (Apis mellifera L.) colonies because neither an artificial diet nor an economical, large-scale pollen collection process from flowers is available. The provenance of honey bee-collected pollen is often unknown, and in some cases has crossed international borders. Both deformed wing virus (DWV) and the fungal pathogen Ascosphaera apis (Claussen) Olive & Spiltoir (cause of chalkbrood disease); occur in honey bee-collected pollen, and infections have been observed in bumble bees. We used these pathogens as general surrogates for viruses and spore-forming fungal diseases to test the efficacy of 3 sterilization methods, and assessed whether treatment altered pollen quality for the bumble bee. Using honey bee-collected pollen spiked with known doses of DWV and A. apis, we compared gamma irradiation (GI), ozone fumigation (OZ), and ethylene oxide fumigation (EO) against an untreated positive control and a negative control. Following sterilization treatments, we tested A. apis spore viability, detected viral presence with PCR, and tested palatability to the bumble bee Bombus impatiens Cresson. We also measured bacterial growth from pollens treated with EO and GI. GI and EO outperformed OZ treatment in pathogen suppression. EO had the highest sterilizing properties under commercial conditions and retained palatability and supported bee development better than other treatments. These results suggest that EO sterilization reduces pathogen risks while retaining pollen quality as a food source for rearing bumble bees.

Differential bumble bee gene expression associated with pathogen infection and pollen diet.

BACKGROUND: Diet and parasitism can have powerful effects on host gene expression. However, how specific dietary components affect host gene expression that could feed back to affect parasitism is relatively unexplored in many wild species. Recently, it was discovered that consumption of sunflower (Helianthus annuus) pollen reduced severity of gut protozoan pathogen Crithidia bombi infection in Bombus impatiens bumble bees. Despite the dramatic and consistent medicinal effect of sunflower pollen, very little is known about the mechanism(s) underlying this effect. However, sunflower pollen extract increases rather than suppresses C. bombi growth in vitro, suggesting that sunflower pollen reduces C. bombi infection indirectly via changes in the host. Here, we analyzed whole transcriptomes of B. impatiens workers to characterize the physiological response to sunflower pollen consumption and C. bombi infection to isolate the mechanisms underlying the medicinal effect. B. impatiens workers were inoculated with either C. bombi cells (infected) or a sham control (un-infected) and fed either sunflower or wildflower pollen ad libitum. Whole abdominal gene expression profiles were then sequenced with Illumina NextSeq 500 technology. RESULTS: Among infected bees, sunflower pollen upregulated immune transcripts, including the anti-microbial peptide hymenoptaecin, Toll receptors and serine proteases. In both infected and un-infected bees, sunflower pollen upregulated putative detoxification transcripts and transcripts associated with the repair and maintenance of gut epithelial cells. Among wildflower-fed bees, infected bees downregulated immune transcripts associated with phagocytosis and the phenoloxidase cascade. CONCLUSIONS: Taken together, these results indicate dissimilar immune responses between sunflower- and wildflower-fed bumble bees infected with C. bombi, a response to physical damage to gut epithelial cells caused by sunflower pollen, and a strong detoxification response to sunflower pollen consumption. Identifying host responses that drive the medicinal effect of sunflower pollen in infected bumble bees may broaden our understanding of plant-pollinator interactions and provide opportunities for effective management of bee pathogens.

Site Specialization of Human Oral Veillonella Species.

Veillonella species are abundant members of the human oral microbiome with multiple interspecies commensal relationships. Examining the distribution patterns of Veillonella species across the oral cavity is fundamental to understanding their oral ecology. In this study, we used a combination of pangenomic analysis and oral metagenomic information to clarify Veillonella taxonomy and to test the site specialist hypothesis for the Veillonella genus, which contends that most oral bacterial species are adapted to live at specific oral sites. Using isolate genome sequences combined with shotgun metagenomic sequence data, we showed that Veillonella species have clear, differential site specificity: Veillonella parvula showed strong preference for supra- and subgingival plaque, while closely related V. dispar, as well as more distantly related V. atypica, preferred the tongue dorsum, tonsils, throat, and hard palate. In addition, the provisionally named Veillonella sp. Human Microbial Taxon 780 showed strong site specificity for keratinized gingiva. Using comparative genomic analysis, we identified genes associated with thiamine biosynthesis and the reductive pentose phosphate cycle that may enable Veillonella species to occupy their respective habitats. IMPORTANCE Understanding the microbial ecology of the mouth is fundamental for understanding human physiology. In this study, metapangenomics demonstrated that different Veillonella species have clear ecological preferences in the oral cavity of healthy humans, validating the site specialist hypothesis. Furthermore, the gene pool of different Veillonella species was found to be reflective of their ecology, illuminating the potential role of vitamins and carbohydrates in determining Veillonella distribution patterns and interspecies interactions.

Sunflower pollen reduces a gut pathogen in the model bee species, Bombus impatiens, but has weaker effects in three wild congeners.

Commercial bumblebees have become popular models to understand stressors and solutions for pollinator health, but few studies test whether results translate to other pollinators. Consuming sunflower pollen dramatically reduces infection by the gut parasite Crithidia bombi in commercially reared Bombus impatiens. We assessed the effect of sunflower pollen on infection in wild B. impatiens, Bombus griseocollis, Bombus bimaculatus and Bombus vagans. We also asked how pollen diet (50% sunflower pollen versus wildflower pollen) and infection (yes/no) affected performance in wild B. impatiens microcolonies. Compared to controls, sunflower pollen dramatically reduced Crithidia infection in commercial and wild B. impatiens, had similar but less dramatic effects in B. bimaculatus and B. vagans, and no effect in B. griseocollis. Bombus impatiens, B. bimaculatus and B. vagans are in the same subgenus, suggesting that responses to sunflower pollen may be phylogenetically conserved. In microcolonies, 50% sunflower pollen reduced infection compared to wildflower pollen, but also reduced reproduction. Sunflower pollen could control Crithidia infections in B. impatiens and potentially close relatives, but may hinder reproduction if other resources are scarce. We caution that research using managed bee species, such as B. impatiens, be interpreted carefully as findings may not relate to all bee species.

Sunflower pollen induces rapid excretion in bumble bees: Implications for host-pathogen interactions.

Host diet can have a profound effect on host-pathogen interactions, including indirect effects on pathogens mediated through host physiology. In bumble bees (Bombus impatiens), the consumption of sunflower (Helianthus annuus) pollen dramatically reduces infection by the gut protozoan pathogen Crithidia bombi. One hypothesis for the medicinal effect of sunflower pollen is that consumption changes host gut physiological function, causing rapid excretion that flushes C. bombi from the system. We tested the effect of pollen diet and C. bombi infection on gut transit properties using a 2x2 factorial experiment in which bees were infected with C. bombi or not and fed sunflower or wildflower pollen diet. We measured several non-mutually exclusive physiological processes that underlie the insect excretory system, including gut transit time, bi-hourly excretion rate, the total number of excretion events and the total volume of excrement. Sunflower pollen significantly reduced gut transit time in uninfected bees, and increased the total number of excretion events and volume of excrement by 66 % and 68 %, respectively, in both infected and uninfected bees. Here we show that a sunflower pollen diet can affect host physiology gut function, causing more rapid and greater excretion. These results provide important insight into a mechanism that could underlie the medicinal effect of sunflower pollen for bumble bees.

Medicinal value of sunflower pollen against bee pathogens.

Global declines in pollinators, including bees, can have major consequences for ecosystem services. Bees are dominant pollinators, making it imperative to mitigate declines. Pathogens are strongly implicated in the decline of native and honey bees. Diet affects bee immune responses, suggesting the potential for floral resources to provide natural resistance to pathogens. We discovered that sunflower (Helianthus annuus) pollen dramatically and consistently reduced a protozoan pathogen (Crithidia bombi) infection in bumble bees (Bombus impatiens) and also reduced a microsporidian pathogen (Nosema ceranae) of the European honey bee (Apis mellifera), indicating the potential for broad anti-parasitic effects. In a field survey, bumble bees from farms with more sunflower area had lower Crithidia infection rates. Given consistent effects of sunflower in reducing pathogens, planting sunflower in agroecosystems and native habitat may provide a simple solution to reduce disease and improve the health of economically and ecologically important pollinators.