Influential factors of saliva microbiota composition

Background: The oral microbiota is emerging as an influential factor of host physiology and disease state. Factors influencing oral microbiota composition have not been well characterised. In particular, there is a lack of population-based studies. We undertook a large hypothesis-free study of the saliva microbiota, considering potential influential factors of host health (frailty; diet; periodontal disease), demographics (age; sex; BMI) and sample processing (storage time), in a sample (n=679) of the TwinsUK cohort of adult twins. Results: Alpha and beta diversity of the saliva microbiota was associated most strongly with frailty (alpha diversity: Q = 0.003, Observed; Q=0.002, Shannon; Q=0.003, Simpson; Beta diversity: Q = 0.002, Bray Curtis dissimilarity ) and age (alpha diversity: Q=0.006, Shannon; Q=0.003, Simpson; beta diversity: Q=0.002, Bray Curtis dissimilarity; Q= 0.032, Weighted UniFrac) in multivariate models including age, frailty, sex, BMI, frailty and diet, and adjustment for multiple testing. Those with a more advanced age were more likely to be dissimilar in the saliva microbiota composition than younger participants (P = 5.125e-06, ANOVA). In subsample analyses, including consideration of periodontal disease (total n=138, periodontal disease n=66), the association with frailty remained for alpha diversity (Q=0.002, Observed ASVs; Q= 0.04 Shannon Index), but not beta diversity, whilst age was not demonstrated to associate with alpha or beta diversity in this subsample, potentially due to insufficient statistical power. Length of time that samples were stored prior to sequencing was associated with beta diversity (Q = 0.002, Bray Curtis dissimilarity). Six bacterial taxa were associated with age after adjustment for frailty and diet. Conclusions: Frailty and age emerged as the most influential factors of saliva microbiota composition. Whilst frailty and age are correlates, the associations were independent of each other, suggesting that both biological and chronological ageing are key drivers of saliva microbiota composition.

Diversity of Site-Specific Microbes of Occlusal and Proximal Lesions in Severe- Early Childhood Caries (S-ECC)

Background: Severe-early childhood caries (S-ECC) is a global problem of significant concern, commonly manifest as a dentinal lesion on the occlusal and proximal surfaces of the affected deciduous dentition. Although there are major ecological differences between these two niches, it is unclear whether these are reflected in the composition of their dysbiotic cariogenic microbiome. Therefore, we compared the compositional differences in the microbiota of occlusal and proximal caries lesions in S-ECC. Methods: Deep-dentine caries samples (19-occlusal and 19-proximal) from asymptomatic primary molars of children with S-ECC (n=19) belonging to caries-code 5/6, according to ICDAS classification, were evaluated. Employing two primer pools, we amplified and compared the bacterial 16S rRNA gene sequences of the seven hypervariable regions (V2 to V4 and V6 to V9) using a next-generation sequencing based assay.Results: Bray-Curtis dissimilarity data indicated that occlusal lesions had a more homogeneous microbial community structure than the proximal lesions with significant compositional differences at species level (p=0.01; R-value of 0.513). Together, the occlusal and proximal niches harbored 263 species, of which 202 (76.8%) species were common to both locales, while 49 (18.6%) and 12 (4.6%) disparate species were exclusively isolated from the proximal and occlusal niches, respectively. The most commonly found genera at both locales included Streptococcus, Prevotella, and Lactobacillus, with 33, 27, and 22 species each, respectively. In addition, Streptococcus mutans predominated in the proximal cavities (p≤0.05), as opposed to Atopobium parvulum (p=0.01) in the occlusal niches, while Vellonella alcalescens was present in similar proportions in both habitats (p≥0.05). Conclusions: Distinct differences between the caries microbiota of occlusal and proximal caries in S-ECC exist. The former niche appears to provide a habitat for a more homogeneous growth of communal microbiota than the latter. This may be due to the conditions prevalent in relatively quiescent inter-proximal regions, as opposed to the occlusal regions exposed to the ebb and flow of salivary and masticatory forces, and/or the anatomical and structural differences in the two locales. The clinical implications of these findings in terms of the rate and severity of caries progression remain to be determined.

Partial Substitution of Meat with Insect (Alphitobius diaperinus) in a Carnivore Diet Changes the Gut Microbiome and Metabolome of Healthy Rats

Insects are suggested as a sustainable protein source of high nutritional quality, but the effects of insect ingestion on processes in the gastrointestinal tract and gut microbiota (GM) remain to be established. We examined the effects of partial substitution of meat with insect protein (Alphitobius diaperinus) in a four-week dietary intervention in a healthy rat model (n = 30). GM composition was characterized using’ 16S rRNA gene amplicon profiling while the metabolomes of stomach, small intestine, and colon content, feces and blood were investigated by 1H-NMR spectroscopy. Metabolomics analyses revealed a larger escape of protein residues into the colon and a different microbial metabolization pattern of aromatic amino acids when partly substituting pork with insect. Both for rats fed a pork diet and rats fed a diet with partial replacement of pork with insect, the GM was dominated by Lactobacillus, Clostridium cluster XI and Akkermansia. However, Bray-Curtis dissimilarity metrics were different when insects were included in the diet. Introduction of insects in a common Western omnivore diet alters the gut microbiome diversity with consequences for endogenous metabolism. This finding highlights the importance of assessing gastrointestinal tract effects when evaluating new protein sources as meat replacements.

Macroalgal canopies reduce beta diversity in intertidal communities

Species diversity in a habitat is often termed alpha diversity. As it influences various community properties, many studies have investigated its drivers. For instance, intertidal macroalgal canopies limit understory thermal stress during low tides and thus often increase alpha diversity. More recently, beta diversity has also become of interest. Beta diversity measures the change in species composition across space and is another important attribute of communities because it influences their multifunctionality, productivity, and resilience. Using data from a field experiment done in Atlantic Canada, we tested the hypothesis that fucoid macroalgal canopies limit beta diversity in intertidal communities. This prediction stems from previous evidence that such canopies limit thermal variation across the substrate during low tides, an important consideration because spatial thermal changes influence spatial variability in species composition. To test our hypothesis, we compared two treatments (full canopy cover and canopy removal) created the year before on intertidal areas that were originally all fully covered by canopies. Separately for each treatment, we calculated beta diversity as the Bray-Curtis dissimilarity between nearby quadrats using species abundance data. Overall, fucoid macroalgal canopies significantly reduced beta diversity, showing that these foundation species can have opposing effects on alpha and beta diversity.

Tracking Microbial Communities Across Aedes Albopictus Life Stages and Larval Habitat Types

Aedes albopictus, the Tiger Mosquito, has been hailed as one of the most invasive arbovirus-transmitting mosquitoes globally. With the growing potential of microbial methods for mosquito control, it has become increasingly imperative to understand the factors that contribute to naturally-occurring microbiome communities. Here, we analyzed the impact of larval water type and life stage on the microbial community of Aedes albopictus. The field-collected water samples from tree holes and tires that were used to rear larval mosquitoes in the laboratory were significantly different from each other in terms of sOTU (bacterial species) richness, with tree holes having a far greater number of sOTUs. For beta diversity measures (Bray-Curtis dissimilarity) Aedes albopictus were not significantly different from adult Aedes aegypti, but mosquito sex, life stage, and overall treatment group were significantly different when analyzed by ANOSIM. Based on our findings, the environment surrounding larval mosquitoes (and subsequent adult mosquitoes from those habitats) and the life stage of mosquitoes (regardless of species) shapes mosquito microbiome assemblages. This work further supports the idea that mosquito adults maintain a microbiome specific to larval habitat, despite major reductions to their microbiome prior to eclosion, which could shape the success of implemented microbial engineering or control methods.

P685 Gut microbiota in patients with Inflammatory Bowel Disease during remission

Background Dysbiosis in patients with active IBD has been described in many studies and populations. There is, however, minimal data on whether dysbiosis persists during remission and the extent to which it does. The aim of this study was to assess the gut microbiota in Maltese IBD patients who are in remission as compared to a healthy control population. Methods Stool samples of patients with IBD in remission and healthy controls were analyzed by 16S rRNA sequencing. High quality Amplicon sequence variants (ASV) were derived and classified via DADA2. Results Ninety-eight patients with IBD (UC: 67.3%; CD: 32.7%) and 97 controls were recruited. Patients with IBD had a decrease in alpha diversity compared to healthy controls (Figure 1) with significant differences in beta diversity (Bray-Curtis dissimilarity, Jaccard distance and Generalized UniFrac distance: all p=0.0001). At the phylum level, abundances differed significantly with respect to health condition (Figure 2). Twenty-five ASVs that were differentially abundant between the different cohorts were identified (Table 1); 13 being over abundant in healthy individuals, while 6 being least abundant among controls. In both CD and UC, 3 different ASVs were found to be more abundant. Conclusion Despite remission, the faecal gut microbiota in IBD is dysbiotic. Future studies could be directed at assessing whether species identified as being more abundant in controls can be used as probiotics to either reduce or be able to stop the standard medications.

P479 Biological therapies and bio-microbial dynamics in inflammatory bowel diseases

Background Therapeutic aims in IBDs are to induce remission through rapidly effective treatment. Conventional therapeutic methods including immunosuppression have been available for more than half a century with biological therapies the treatment of choice in patients who fail immunosuppressive drugs. There has been little attempt to longitudinally examine the differences in biopsychological factors and their associations with treatment modalities in IBD patients. Methods 50 IBD participants (24 UC, 26 CD) in clinical remission were followed for 12 months. Complete longitudinal datasets and monthly stool and blood samples for inflammatory biomarkers were collected for analysis of association. Microbiome analysis was performed using V4 16SrRNA for identification of microbial phylogenetic relationships. Patients were grouped on whether or not they were in remission and on biological agents. Results At the baseline there were similar ecological indices between the two treatment options in both disease classes; (Shannon, p = 0.380; Pielou’s, p = 0.246; Chao1, p = 0.934). At family level, the CD group showed significant differences (Wilcoxon, p ≤ 0.05) between the treatment received (biologics vs non-biologic) in the abundance of Barnesiellaceae, Bifidobacteriaceae, unclassified Clostridia, and Clostridiaceae (Figure 1A). Results showed similar ecological index values in samples from the two treatment options. Samples in UC groups on biological and non-biologic treatment modes were found to have significantly different microbial communities based on Bray-Curtis dissimilarity (PERMANOVA, p = 0.0031). Microbial family composition in UC group and on two treatment modalities showed a total of 12 families that were significantly different (Wilcoxon, p ≤ 0.05): Acidaminococcaceae, unclassified Bacteroidales, Barnesiellaceae, Christensenellaceae, Clostridiales vadinBB60 group, Coriobacteriaceae, Defluvitaleaceae, Eggerthelaceaem Fusobacteriaceae, Muribaculaceae, Prevotellaceae, and Streptococcaceae (Figure1B). Results identified significant differences in microbial diversity (Shannon index, p = 0.041), and evenness (Pielou’s evenness, p = 0.045) between the two treatment modes. No differences were identified in richness (Chao1) in UC participants. Conclusion Conclusion: Baseline biological indices were mostly similarly distributed between the two treatment options in remissive IBD patients, however biological therapy significantly influenced longitudinal trends of some microbiome dynamics, especially in UC. This suggests an underlying interrelationship between mode of treatment and biopsychological trajectories which might overshadow response to the treatment, requiring further assessment.

Evaluating a Laboratory Flume Microbiome as a Window Into Natural Riverbed Biogeochemistry

Riverbeds are hotspots for microbially-mediated reactions that exhibit pronounced variability in space and time. It is challenging to resolve biogeochemical mechanisms in natural riverbeds, as uncontrolled settings complicate data collection and interpretation. To overcome these challenges, laboratory flumes are often used as proxies for natural riverbed systems. Flumes capture spatiotemporal variability and thus allow for controlled investigations of riverbed biogeochemistry. These investigations implicitly rely on the assumption that the flume microbiome is similar to the microbiome of natural riverbeds. However, this assumption has not been tested and it is unknown how the microbiome of a flume compares to natural aquatic settings, including riverbeds. To evaluate the fundamental assumption that a flume hosts a microbiome similar to natural riverbed systems, we used 16s rRNA gene sequencing and publicly available data to compare the sediment microbiome of a single large laboratory flume to a wide variety of natural ecosystems including lake and marine sediments, river, lake, hyporheic, soil, and marine water, and bank and wetland soils. Richness and Shannon diversity metrics, analyses of variance, Bray-Curtis dissimilarity, and analysis of the common microbiomes between flume and river sediment all indicated that the flume microbiome more closely resembled natural riverbed sediments than other ecosystems, supporting the use of flume experiments for investigating natural microbially-mediated biogeochemical processes in riverbeds.

Effects of parity, blood progesterone, and non-steroidal anti-inflammatory treatment on the dynamics of the uterine microbiota of healthy postpartum dairy cows

This study evaluated the effects of treatment with meloxicam (a non-steroidal anti-inflammatory drug), parity, and blood progesterone concentration on the dynamics of the uterine microbiota of 16 clinically healthy postpartum dairy cows. Seven primiparous and 9 multiparous postpartum Holstein cows either received meloxicam (0.5 mg/kg SC, n = 7 cows) once daily for 4 days (10 to 13 days in milk (DIM)) or were untreated (n = 9 cows). Endometrial cytology samples were collected by cytobrush at 10, 21, and 35 DIM, from which the microbiota analysis was conducted using 16S rRNA gene sequence analysis. A radioimmunoassay was used to measure progesterone concentration in blood serum samples at 35 DIM and cows were classified as ˃ 1 ng/mL (n = 10) or ≤ 1 ng/mL (n = 6). Alpha diversity for bacterial genera (Chao1, Shannon-Weiner, and Camargo’s evenness indices) were not affected by DIM, meloxicam treatment, parity, or progesterone category. For beta diversity (genera level), principal coordinate analysis (Bray-Curtis) showed differences in microbiota between parity groups. At the phylum level, the relative abundance of Actinobacteria was greater in primiparous than multiparous cows. At the genus level, there was lesser relative abundance of Bifidobacterium, Lactobacillus, Neisseriaceae, Paracoccus, Staphylococcus, and Streptococcus and greater relative abundance of Bacillus and Fusobacterium in primiparous than multiparous cows. Bray-Curtis dissimilarity did not differ by DIM at sampling, meloxicam treatment, or progesterone category at 35 DIM. In conclusion, uterine bacterial composition was not different at 10, 21, or 35 DIM, and meloxicam treatment or progesterone category did not affect the uterine microbiota in clinically healthy postpartum dairy cows. Primiparous cows presented a different composition of uterine bacteria than multiparous cows. The differences in microbiota associated with parity might be attributable to changes that occur consequent to the first calving, but this hypothesis should be investigated further.

Fish Assemblage Structure in the Northwestern Hawaiian Islands Is Associated with the Architectural Complexity of Coral-Reef Habitats

The architectural complexity of coral-reef habitat plays an important role in determining the assemblage structure of reef fish. We investigated associations between the reef habitats and fish assemblages in the Northwestern Hawaiian Islands (NWHI) using in situ fish counts and data on habitat metrics and benthic community composition that were obtained from three-dimensional (3D) photogrammetric reconstructions of the surveyed sites. The structure of fish assemblage as a whole on the basis of Bray–Curtis dissimilarity, species richness and the abundances of herbivores and piscivores were associated with habitat metrics, with higher levels of architectural complexity generally supporting greater numbers of fish species and individuals. Benthic cover did not explain additional variation in these variables after the effects of habitat metrics were taken into account. Corallivorous fish was the only group that showed positive associations with both habitat metrics and benthic cover (Acropora and Pocillopora corals). The total fish abundance and the abundances of planktivores and invertivores did not show associations with either habitat metrics or benthic cover. This study suggests that an appropriate combination of habitat metrics can be used to account sufficiently for the effects of habitat architecture on fish assemblages in reef monitoring efforts in the NWHI.