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Previous research on stabilization methods for microbiome investigations has largely focused on human fecal samples. There are a few studies using feces from other species, but no published studies investigating preservation of samples collected from cattle. Given that microbial taxa are differentially impacted during storage it is warranted to study impacts of preservation methods on microbial communities found in samples outside of human fecal samples. Here we tested methods of preserving bovine fecal respiratory specimens for up to 2 weeks at four temperatures (room temperature, 4°C, -20°C, and -80°C) by comparing microbial diversity and community composition to samples extracted immediately after collection. Importantly, fecal specimens preserved and analyzed were technical replicates, providing a look at the effects of preservation method in the absence of biological variation. We found that preservation with the OMNIgene ® •GUT kit resulted in community structure most like that of fresh samples extracted immediately, even when stored at room temperature (~20°C). Samples that were flash-frozen without added preservation solution were the next most representative of original communities, while samples preserved with ethanol were the least representative. These results contradict previous reports that ethanol is effective in preserving fecal communities and suggest for studies investigating cattle either flash-freezing of samples without preservative or preservation with OMNIgene ® •GUT will yield more representative microbial communities.

The gastrointestinal tract (GIT) of cattle plays a vital role in nutrient absorption, immune function, and microbial homeostasis. While the importance of the GIT microbiome and epithelial barrier integrity has been increasingly recognized, the typical composition of microbial communities and the expression of tight junction proteins (TJPs) in feedlot cattle remains poorly characterized. We investigated microbial community structure and TJP expression at three GIT sites: the rumen (RU), small intestine (SI), and large intestine (LI) in 21 finish-fed feedlot steers sourced from 21 commercial feedyards in the Texas Panhandle. Samples of luminal contents and GIT tissue were collected from each region, as well as feces and liver abscess material. Microbial communities were characterized using 16S rRNA gene sequencing. TJP gene expression was quantified by RT-qPCR using synthetic standards, and protein expression was evaluated by immunohistochemistry (IHC) with both computer-generated and pathologist-generated scoring. Microbial community structures varied primarily by GIT region rather than by individual animals raised at different locations. Nine bacterial families were identified as core microbiome members, with Lachnospiraceae being the most abundant across the GIT. TJP gene expression varied considerably by site, with RU having significantly lower Claudin 1, Claudin 2, and E-Cadherin expression than the SI and LI. IHC results paralleled qPCR findings, with region-specific patterns of protein localization and intensity. Computerized and pathologist-generated H-scores showed moderate agreement but differed notably between epithelial and lamina propria regions. This study provides a comprehensive baseline of microbial and host factors associated with gut health in a uniquely diverse population of feedlot cattle. The identification of regional microbial communities and distinct TJP expression patterns offers foundational insights into gastrointestinal physiology and barrier function. This work establishes baseline data to support future investigations into the relationships among microbial ecology, epithelial barrier function, and cattle health and productivity.

Adherent-invasive Escherichia coli (AIEC) colonization has been strongly implicated in the pathogenesis of Crohn's disease. Environmental triggers such as vitamin D deficiency have emerged as key factors in the pathogenesis of inflammatory bowel diseases. The aim of this study was to investigate the effects of 1,25(OH)2D3 on AIEC infection-induced changes in vivo and in vitro.MethodsBarrier function was assessed in polarized epithelial Caco-2-bbe cells grown in medium with or without vitamin D and challenged with AIEC strain LF82. Weaned C57BL/6 mice were fed either a vitamin D–sufficient or –deficient diet for 5 weeks and then infected with AIEC, in the absence and presence of low-dose dextran sodium sulphate. Disease severity was assessed by histological analysis and in vivo intestinal permeability assay. Presence of invasive bacteria was assessed by transmission electron microscopy.ResultsCaco-2-bbe cells incubated with 1,25(OH)2D3 were protected against AIEC-induced disruption of transepithelial electrical resistance and tight-junction protein redistribution. Vitamin D–deficient C57BL/6 mice given a course of 2% dextran sodium sulphate exhibited pronounced epithelial barrier dysfunction, were more susceptible to AIEC colonization, and showed exacerbated colonic injury. Transmission electron microscopy of colonic tissue from infected mice demonstrated invasion of AIEC and fecal microbiome analysis revealed shifts in microbial communities.ConclusionsThese data show that vitamin D is able to mitigate the deleterious effects of AIEC on the intestinal mucosa, by maintaining intestinal epithelial barrier homeostasis and preserving tight-junction architecture. This study highlights the association between vitamin D status, dysbiosis, and Crohn's disease.

ABSTRACT Plastic is a ubiquitous pollutant in the marine environment. Here, we investigated how temporal changes in environmental factors affect the microbial communities formed on plastic (polyethylene terephthalate; PET) versus a ceramic substrate. In situ mesocosms (N = 90 replicates) were deployed at the sediment–water interface of a coastal lagoon and sampled every 4 weeks for 424 days. Sequencing data (16S rRNA) was parsed based on variation in temperature with the exposure starting in fall 2016 and remaining in situ through the next four seasons (winter, spring, summer and fall 2017). PET biofilms were distinct during the summer when salinity and temperature were highest. In particular, a significant shift in the relative abundance of Ignavibacteriales and Cytophagales was observed during the summer, but PET and ceramic communities were again indistinguishable the following fall. Water temperature, salinity and pH were significant drivers of PET biofilm diversity as well as the relative abundance of plastic-discriminant taxa. This study illustrates the temporal and successional dynamics of PET biofilms and clearly demonstrates that increased water temperature, salinity, pH and exposure length play a role in the formation of a plastic-specific microbial community, but this specificity can be lost with a change in environmental conditions. Temperature and salinity drive the formation of a reversible plastic-specific community.

Closed recirculating aquatic systems (aquariums) offer a multitude of benefits including the ability to observe and research aquatic animals ex situ, and under controlled environmental conditions [...]

Nutritional programming (NP) is a concept in which early nutritional events alter the physiology of an animal and its response to different dietary regimes later in life. The objective of this study was to determine if NP via broodstock with dietary plant protein (PP) has any effect on the gut microbiome of the progeny fish and whether this modified gut microbiome leads to better utilization of PP diet. The experiment consisted of four different treatments as follows: (1) progeny that received FM diet obtained from fishmeal (FM)-fed broodstock (FMBS-FM, +control); (2) progeny that received PP diet obtained from FM-fed parents (FMBS-PP); (3) progeny that received PP diet obtained from “nutritionally programmed” parents (PPBS-PP; −control); and (4) progeny that received FM diet obtained from “nutritionally programmed” parents (PPBS-FM). Zebrafish was used as a model species. This study found that parental programming seems to have some positive effect on dietary PP utilization in progeny. However, the influence of NP with PP through broodstock on gut microbiota of the offspring fish was not detected.

Background Liver abscesses (LAs) are one of the most common and important problems faced by the beef industry. The most efficacious method for the prevention of LAs in North America is through dietary inclusion of low doses of antimicrobial drugs such as tylosin, but the mechanisms by which this treatment prevents LAs are not fully understood. LAs are believed to result from mucosal barrier dysfunction in the gastrointestinal tract (GIT) allowing bacterial translocation to the liver via the portal vein, yet differences in the GIT microbiome of cattle with and without LAs have not been explored. Here, we characterized microbial communities from LAs, rumen, ileum, and colon from the same cattle for the first time. Results Results demonstrate that tylosin supplementation was associated with differences in microbial community structure in the rumen and small intestine, largely because of differences in the predominance of Clostridia. Importantly, we show for the first time that microbial communities from multiple LAs in one animal’s liver are highly similar, suggesting that abscesses found at different locations in the liver may originate from a localized source in the GIT (rather than disparate locations). A large portion of abscesses were dominated by microbial taxa that were most abundant in the hindgut. Further, we identified taxa throughout the GIT that were differentially abundant between animals with and without liver abscesses. Bifidobacterium spp.—a bacteria commonly associated with a healthy GIT in several species—were more abundant in the rumen and ileum of animals without LAs compared to those with LAs. Conclusions Together these results provide the first direct comparison of GIT and LA microbial communities within the same animal, add considerable evidence to the hypothesis that some LA microbial communities arise from the hindgut, and suggest that barrier dysfunction throughout the GIT may be the underlying cause of LA formation in cattle.

Background Bovine respiratory disease (BRD) is caused by interactions among host, environment, and pathogens. One standard method for antemortem pathogen identification in cattle with BRD is deep-guarded nasopharyngeal swabbing, which is challenging, costly, and waste generating. The objective was to compare the ability to recover Mannheimia haemolytica and compare microbial community structure using 29.5 inch (74.9 cm) deep-guarded nasopharyngeal swabs, 16 inch (40.6 cm) unguarded proctology swabs, or 6 inch (15.2 cm) unguarded nasal swabs when characterized using culture, real time-qPCR, and 16S rRNA gene sequencing. Samples for aerobic culture, qPCR, and 16S rRNA gene sequencing were collected from the upper respiratory tract of cattle 2 weeks after feedlot arrival. Results There was high concordance of culture and qPCR results for all swab types (results for 77% and 81% of sampled animals completely across all 3 swab types for culture and qPCR respectively). Microbial communities were highly similar among samples collected with different swab types, and differences identified relative to treatment for BRD were also similar. Positive qPCR results for M. haemolytica were highly concordant (81% agreed completely), but samples collected by deep-guarded swabbing had lower amounts of Mh DNA identified (Kruskal–Wallis analysis of variance on ranks, P  < 0.05; Dunn-test for pairwise comparison with Benjamini–Hochberg correction, P  < 0.05) and lower frequency of positive compared to nasal and proctology swabs (McNemar’s Chi-square test, P  < 0.05). Conclusions Though differences existed among different types of swabs collected from individual cattle, nasal swabs and proctology swabs offer comparable results to deep-guarded nasopharyngeal swabs when identifying and characterizing M. haemolytica by culture, 16S rRNA gene sequencing, and qPCR.

Aside from two samples collected nearly 50 years ago, little is known about the microbial composition of wind tidal flats in the hypersaline Laguna Madre, Texas. These mats account for ~42% of the lagoon’s area. These microbial communities were sampled at four locations that historically had mats in the Laguna Madre, including Laguna Madre Field Station (LMFS), Nighthawk Bay (NH), and two locations in Kenedy Ranch (KRN and KRS). Amplicon sequencing of 16S genes determined the presence of 51 prokaryotic phyla dominated by Bacteroidota, Chloroflexi, Cyanobacteria, Desulfobacteria, Firmicutes, Halobacteria, and Proteobacteria. The microbial community structure of NH and KR is significantly different to LMFS, in which Bacteroidota and Proteobacteria were most abundant. Twenty-three cyanobacterial taxa were identified via genomic analysis, whereas 45 cyanobacterial taxa were identified using morphological analysis, containing large filamentous forms on the surface, and smaller, motile filamentous and coccoid forms in subsurface mat layers. Sample sites were dominated by species in Oscillatoriaceae (i.e., Lyngbya) and Coleofasciculaceae (i.e., Coleofasciculus). Most cyanobacterial sequences (~35%) could not be assigned to any established taxa at the family/genus level, given the limited knowledge of hypersaline cyanobacteria. A total of 73 cyanobacterial bioactive metabolites were identified using ultra performance liquid chromatography-Orbitrap MS analysis from these commu nities. Laguna Madre seems unique compared to other sabkhas in terms of its microbiology.

Background. Vitamin D, an important modulator of the immune system, has been shown to protect mucosal barrier homeostasis. This study investigates the effects of vitamin D deficiency on infection-induced changes in intestinal epithelial barrier function in vitro and on Citrobacter rodentium–induced colitis in mice. Methods. Polarized epithelial Caco2-bbe cells were grown in medium with or without vitamin D and challenged with enterohemorrhagic Escherichia coli O157:H7. Barrier function and tight junction protein expression were assessed. Weaned C57BL/6 mice were fed either a vitamin D-sufficient or vitamin D-deficient diet and then infected with C. rodentium. Disease severity was assessed by histological analysis, intestinal permeability assay, measurement of inflammatory cytokine levels, and microbiome analysis. Results. 1,25(OH)₂D₃ altered E. coli O157:H7-induced reductions in transepithelial electrical resistance (P<.01), decreased permeability (P<.05), and preserved barrier integrity. Vitamin D-deficient mice challenged with C. rodentium demonstrated increased colonic hyperplasia and epithelial barrier dysfunction (P<.0001 and P<.05, respectively). Vitamin D deficiency resulted in an altered composition of the fecal microbiome both in the absence and presence of C. rodentium infection. Conclusions. This study demonstrates that vitamin D is an important mediator of intestinal epithelial defenses against infectious agents. Vitamin D deficiency predisposes to more-severe intestinal injury in an infectious model of colitis.