Ruminants harbor a ruminal microbiome that converts their host-indigestible diet into nutrients. This microbiome contains three distinct communities: liquid, solid, and epithelial (or epimural). Currently, there is limited research examining the diet-dependent responses of all three microbiomes within the same animal. Here, we used next-generation 16S rRNA sequencing to characterize the ruminal solid (RS), liquid (RL), and epimural (RE) microbiotas of 13 lactating and cannulated Holstein dairy cows fed either a high- or low-starch diet in a crossover experimental design. Independent of diet, we found that all sample types were dominated by the phyla Firmicutes and Bacteroidetes. Proteobacteria and Epsilonbacteraeota were also highly abundant but only in the RE. Although the total VFA molar abundance did not differ between diets, propionate and valerate were found to increase with the high-starch diet, while acetate was increased with the low-starch diet. Overall, we found that the RE microbiota was more diverse than the RS and RL communities, with diet impacting community diversity in the RS. We found that sample type, diet treatment, and their interaction significantly impacted community structure and composition. Notably, Prevotella was most abundant in the RL and RS, particularly on the low-starch diet. We also found that Lachnospiraceae were significantly enriched in the RS and RL with a high-starch diet, whereas Succiniclasticum was highly abundant in the RE with a low-starch diet. These data suggest that diet influences all three ruminal microbiomes and provides a useful framework for understanding the role of these microbiomes in mediating host production.

Oral swabs of dairy cows have been suggested as a proxy for direct ruminal sampling, and this approach can identify the presence of up to 70% of the rumen microbial community. Here, we further extend the utility of this approach by correlating the bacterial community of swabs collected from 226 dairy cows on a research farm in Wisconsin, USA, with average milk yield and days in milk, two phenotypes previously associated with differences in the ruminal microbiome. We then obtained milk production efficiency data for a subset of these animals (gross feed efficiency [GFE] and residual feed intake [RFI]) and correlated these metrics against their associated microbial data. We found that when using the oral swabs, we could identify correlations between bacterial genera and days in milk (P < 0.05). We further show that the ruminal microbiota was associated with average milk yield and days in milk for animals in their first lactation. Differential abundance testing identified amplicon sequence variants (ASVs) associated with these metrics (P < 0.05). Our comparison of bacterial communities between high and low efficiency groups, as determined by GFE and RFI, identified a significant difference in Shannon's diversity in second lactation cows (P < 0.05). We also found that RFI was significantly correlated with the bacterial community in second lactation animals (P < 0.05). Differential abundance analysis identified multiple oral- and rumen-associated ASVs correlated with GFE and RFI (P < 0.05). This study further establishes the utility of oral swabs as a ruminal proxy.IMPORTANCEImproving milk production efficiency is a key goal in the dairy industry and is traditionally pursued through genetic selection, diet optimization, and herd management practices. The ruminal microbiome, essential for digesting feed, has been linked to milk production efficiency, suggesting that microbiome modulation could improve efficiency. However, the integration of rumen microbiology into current management practices is hampered by the difficulty of large-scale rumen sampling, as proxies like fecal samples do not accurately reflect the ruminal microbiota. Traditional methods, like cannulation and stomach tubing, are labor-intensive and impractical for extensive sampling. Our research demonstrates the potential use of oral swabs as a scalable, effective method for characterizing the microbiome and its associations with milk production metrics, recapitulating established associations obtained through traditional ruminal sampling methods.

Understanding the mechanisms of early clearance of Mycobacterium tuberculosis (Mtb) may illuminate new therapeutic strategies for tuberculosis (TB). We previously found genetic, epigenetic, and transcriptomic signatures associated with resistance (resister, RSTR) to tuberculin skin test (TST)/interferon gamma release assay (IGRA) conversion among highly exposed TB contacts. We hypothesized that integration of these datasets with multi-omic latent factor methods would detect pathways differentiating RSTR patients from those with asymptomatic TB infection (TBI, also known as latent TB infection or LTBI) that were not detected in individual dataset analyses. We pre-filtered and scaled features with the largest change between TBI and RSTR groups for 126 patients with data in at least two of five data modalities: single nucleotide polymorphisms (SNP), monocyte RNAseq (baseline and Mtb-stimulated conditions), and monocyte epigenetics (methylation and ATAC-seq). Using multiomic latent factor analysis (MOFA), we generated ten latent factors on the subset of 33 patients with all five datasets available, four of which differed by RSTR status (FDR < 0.1). Factor 4 showed the greatest difference between RSTR and TBI groups (FDR < 0.001). Three additional latent factor integration methods also distinguished the RSTR and TBI groups and identified overlapping features with MOFA. Using pathway analysis and a cluster-based enrichment method, we identified functions associated with latent factors and found that MOFA Factors 2–4 include functions related to cell-cell adhesion, cell shape, and multicellular structure development. In summary, latent variable integration methods uncovered signatures associated with resistance to TST/IGRA conversion that were not detected by individual dataset analyses and included pathways associated with cellular interactions and multicellular structures.

Abstract Asthma has striking disparities across ancestral groups, but the molecular underpinning of these differences is poorly understood and minimally studied. A goal of the Consortium on Asthma among African-ancestry Populations in the Americas (CAAPA) is to understand multi-omic signatures of asthma focusing on populations of African ancestry. RNASeq and DNA methylation data are generated from nasal epithelium including cases (current asthma, N = 253) and controls (never-asthma, N = 283) from 7 different geographic sites to identify differentially expressed genes (DEGs) and gene networks. We identify 389 DEGs; the top DEG, FN1, was downregulated in cases (q = 3.26 × 10−9) and encodes fibronectin which plays a role in wound healing. The top three gene expression modules implicate networks related to immune response (CEACAM5; p = 9.62 × 10−16 and CPA3; p = 2.39 × 10−14) and wound healing (FN1; p = 7.63 × 10−9). Multi-omic analysis identifies FKBP5, a co-chaperone of glucocorticoid receptor signaling known to be involved in drug response in asthma, where the association between nasal epithelium gene expression is likely regulated by methylation and is associated with increased use of inhaled corticosteroids. This work reveals molecular dysregulation on three axes – increased Th2 inflammation, decreased capacity for wound healing, and impaired drug response – that may play a critical role in asthma within the African Diaspora.

Using oral swabs to collect the remnants of stomach content regurgitation during rumination in dairy cows can replicate up to 70% of the ruminal bacterial community, offering potential for broad-scale population-based studies on the rumen microbiome. The swabs collected from dairy cows often vary widely with respect to sample quality, likely due to several factors such as time of sample collection and cow rumination behavior, which may limit the ability of a given swab to accurately represent the ruminal microbiome. One such factor is the color of the swab, which can vary significantly across different cows. Here, we hypothesize that darker-colored swabs contain more rumen contents, thereby better representing the ruminal bacterial community than lighter-colored swabs. To address this, we collected oral swabs from 402 dairy cows and rumen samples from 13 cannulated cows on a research farm in Wisconsin, United States and subjected them to 16S rRNA sequencing. In addition, given that little is known about the ability of oral swabs to recapitulate the ruminal fungal community, we also conducted ITS sequencing of these samples. To correlate swab color to the microbiota we developed and utilized a novel imaging approach to colorimetrically quantify each swab from a range of light to dark. We found that swabs with increasing darkness scores were significantly associated with increased bacterial alpha diversity (p < 0.05). Lighter swabs exhibited greater variation in their community structure, with many identified amplicon sequence variants (ASVs) categorized as belonging to known bovine oral and environmental taxa. Our analysis of the fungal microbiome found that swabs with increasing darkness scores were associated with decreased alpha diversity (p < 0.05) and were also significantly associated with the ruminal solids fungal community, but not with the ruminal liquid community. Our study refines the utility of oral swabs as a useful proxy for capturing the ruminal microbiome and demonstrates that swab color is an important factor to consider when using this approach for documenting both the bacterial and fungal communities.

We introduce kimma (Kinship In Mixed Model Analysis), an open-source R package for flexible linear mixed effects modeling of RNA-seq including covariates, weights, random effects, covariance matrices, and fit metrics. In simulated datasets, kimma detects differentially expressed genes (DEGs) with similar specificity, sensitivity, and computational time as limma unpaired and dream paired models. Unlike other software, kimma supports covariance matrices as well as fit metrics like AIC. Utilizing genetic kinship covariance, kimma revealed that kinship impacts model fit and DEG detection in a related cohort. Thus, kimma equals or outcompetes current DEG pipelines in sensitivity, computational time, and model complexity.

Mepolizumab (anti-IL5) reduces asthma exacerbations in urban children. We previously utilized nasal transcriptomics to identify inflammatory pathways (gene co-expression modules) associated with exacerbations despite this therapy. To understand mepolizumab's precise impact on these pathways, we assess gene co-expression and loss of correlation, "decoherence," using differential co-expression network analyses. 290 urban children (6-17 years) with exacerbation-prone asthma and blood eosinophils ≥150/microliter were randomized (1:1) to q4 week placebo or mepolizumab injections added to guideline-based care for 52 weeks. Nasal lavage samples were collected before and during treatment for RNA-sequencing. Differential co-expression of gene networks was evaluated to assess interactions and regulatory aspects of type-2 and eosinophilic airway inflammation. Mepolizumab, but not placebo, significantly reduced the overall expression of an established type-2 inflammation gene co-expression module (fold change=0.77, p=0.002) enriched for eosinophil, mast cell, and epithelial IL-13 response genes (242 genes). Mepolizumab uncoupled co-expression of genes in this pathway. During mepolizumab, but not placebo treatment, there was significant loss of correlation among eosinophil-specific genes including RNASE2 (EDN), RNASE3 (ECP), CLC, SIGLEC8, and IL5RA contrasting a reciprocal increase in correlation among mast cell-specific genes (TPSAB1, CPA3, FCER1A), T2 cytokines (IL4, IL5, and IL13), and POSTN. These results suggest mepolizumab disrupts the regulatory interactions of gene co-expression among airway eosinophils, mast cells and epithelium by interrupting transcription regulation in eosinophils with enhancement in mast cell and epithelial inflammation. This paradoxical effect may contribute to an incomplete reduction of asthma exacerbations and demonstrates how differential co-expression network analyses can identify targets for more precise therapies.

Identification of airway inflammatory pathways in asthma has proven essential to understanding mechanisms of disease and has led to effective personalized treatment with biologic therapies. However, relatively little is known about patterns of airway inflammation at the time of respiratory illnesses and how such patterns relate to responsiveness to biologic therapies. The MUPPITS-1 (n=106) and MUPPITS-2 (n=290) studies investigated asthma exacerbations in urban children with exacerbation-prone asthma and ≥150/microliter blood eosinophils. Children in both studies received guidelines-based asthma care; in MUPPITS-2, participants were additionally randomized (1:1) to placebo or mepolizumab. Nasal lavage samples were collected during respiratory illnesses for RNA-sequencing and analyzed by modular analysis to assess genome-wide expression patterns associated with exacerbation illnesses. Among 284 illnesses, exacerbations that occurred in the absence of mepolizumab therapy showed significantly higher upregulation of eosinophil associated inflammatory pathways (fold change values [FC]=1.27-1.43, p-values<0.05), including a Type-2 inflammation module composed of eosinophil, mast cell, and IL-13 response genes. In contrast, exacerbations that occurred while on mepolizumab therapy showed significantly higher upregulation of several epithelial inflammatory pathways (FC=1.36-1.64, p-values<0.05) including TGF-β/Smad3 signaling, extracellular matrix production, and epidermal growth factor receptor signaling. These results indicate that novel inflammatory pathways, likely originating from the airway epithelium and distinct from Type-2 or eosinophilic inflammation, drive residual exacerbations that occur in children treated with mepolizumab therapy added to guideline-based care. These findings identify likely mechanisms of persistent disease expression in these children despite significant depletion of eosinophils and can identify novel treatment targets for future studies.

Introduction The composition of the nasal microbiota in surgical patients in the context of general anesthesia and nasal povidone-iodine decolonization is unknown. The purpose of this quality improvement study was to determine: (i) if general anesthesia is associated with changes in the nasal microbiota of surgery patients and (ii) if preoperative intranasal povidone-iodine decolonization is associated with changes in the nasal microbiota of surgery patients. Materials and methods One hundred and fifty-one ambulatory patients presenting for surgery were enrolled in a quality improvement study by convenience sampling. Pre- and post-surgery nasal samples were collected from patients in the no intranasal decolonization group (control group, n = 54). Pre-decolonization nasal samples were collected from the preoperative intranasal povidone-iodine decolonization group (povidone-iodine group, n = 97). Intranasal povidone-iodine was administered immediately prior to surgery and continued for 20 minutes before patients proceeded for surgery. Post-nasal samples were then collected. General anesthesia was administered to both groups. DNA from the samples was extracted for 16S rRNA sequencing on an Illumina MiSeq. Results In the control group, there was no evidence of change in bacterial diversity between pre- and post-surgery samples. In the povidone-iodine group, nasal bacterial diversity was greater in post-surgery, relative to pre-surgery (Shannon’s Diversity Index (P = 0.038), Chao’s richness estimate (P = 0.02) and Inverse Simpson index (P = 0.027). Among all the genera, only the relative abundance of the genus Staphylococcus trended towards a decrease in patients after application (FDR adjusted P = 0.06). Abundant genera common to both povidone-iodine and control groups included Staphylococcus, Bradyrhizobium, Corynebacterium, Dolosigranulum, Lactobacillus, and Moraxella. Conclusions We found general anesthesia was not associated with changes in the nasal microbiota. Povidone-iodine treatment was associated with nasal microbial diversity and decreased abundance of Staphylococcus. Future studies should examine the nasal microbiota structure and function longitudinally in surgical patients receiving intranasal povidone-iodine.

A major goal for the dairy industry is to improve overall milk production efficiency (MPE). With the advent of next-generation sequencing and advanced methods for characterizing microbial communities, efforts are underway to improve MPE by manipulating the rumen microbiome. Our previous work demonstrated that a near-total exchange of whole rumen contents between pairs of lactating Holstein dairy cows of disparate MPE resulted in a reversal of MPE status for ∼10 days: historically high-efficiency cows decreased in MPE, and historically low-efficiency cows increased in MPE. Importantly, this switch in MPE status was concomitant with a reversal in the ruminal bacterial microbiota, with the newly exchanged bacterial communities reverting to their pre-exchange state. However, this work did not include an in-depth analysis of the microbial community response or an interrogation of specific taxa correlating to production metrics. Here, we sought to better understand the response of rumen communities to this exchange protocol, including consideration of the rumen fungi. Rumen samples were collected from 8 days prior to, and 56 days following the exchange and were subjected to 16S rRNA and ITS amplicon sequencing to assess bacterial and fungal community composition, respectively. Our results show that the ruminal fungal community did not differ significantly between hosts of disparate efficiency prior to the exchange, and no change in community structure was observed over the time course. Correlation of microbial taxa to production metrics identified one fungal operational taxonomic unit (OTU) in the genus Neocallimastix that correlated positively to MPE, and several bacterial OTUs classified to the genus Prevotella . Within the Prevotella , Prevotella_1 was found to be more abundant in high-efficiency cows whereas Prevotella_7 was more abundant in low-efficiency cows. Overall, our results suggest that the rumen bacterial community is a primary microbial driver of host efficiency, that the ruminal fungi may not have as significant a role in MPE as previously thought, and that more work is needed to better understand the functional roles of specific ruminal microbial community members in modulating MPE.

This study aimed to investigate the changes in abomasum transcriptome and the associated microbial community structure in young calves with artificially dosed, adult rumen contents. Eight young bull calves were randomly dosed with freshly extracted rumen contents from an adult cow (high efficiency (HE), n = 4), or sterilized rumen content (Con, n = 4). The dosing was administered within 3 days of birth, then at 2, 4, and 6 weeks following the initial dosing. Abomasum tissues were collected immediately after sacrifice at 8 weeks of age. Five genera (Tannerella, Desulfovibrio, Deinococcus, Leptotrichia, and Eubacterium; p < 0.05) showed significant difference in abundance between the treatments. A total of 975 differentially expressed genes were identified (p < 0.05, fold-change > 1.5, mean read-counts > 5). Pathway analysis indicated that up-regulated genes were involved in immune system process and defense response to virus, while the down-regulated genes involved in ion transport, ATP biosynthetic process, and mitochondrial electron transport. Positive correlation (r > 0.7, p < 0.05) was observed between TRPM4 gene and Desulfovibrio, which was significantly higher in the HE group. TRPM4 had a reported role in the immune system process. In conclusion, the dosing of adult rumen contents to calves can alter not only the composition of active microorganisms in the abomasum but also the molecular mechanisms in the abomasum tissue, including reduced protease secretion and decreased hydrochloric acid secretion.

Dairy cattle are globally important agricultural animals. Central to their biology is the rumen, which houses an essential microbial community, or microbiome, important for providing nutrition from otherwise host-inaccessible dietary components. The rumen environment is noted for its substantial spatial heterogeneity, as illustrated by the stratification into ruminal solid and liquid phases. A third microbiota found directly attached to the ruminal epithelium (the epimural microbiota) also exists but is less well understood because of challenges in sampling the ruminal epithelium. As a result, our understanding of the epimural microbiota is based on analyses of cannulated animals sampled at a single location-the ventral sac-and does not account for other ruminal locations, which may have importance for overall rumen function. To address this knowledge gap, we hypothesize that the epimural microbiota at different ruminal locations differs due to known morphological, physiological, and functional differences across the geographic spread of the rumen epithelium. Here, we characterized bacterial epimural communities at different sites within 8 lactating Holstein dairy cows using 16S rRNA gene sequencing. Four different sites were sampled via rumen tissue biopsy: cranial sac (CS), ventral sac (VS), caudodorsal blind sac (CDBS), and caudoventral blind sac (CVBS). We found that locations differed in both epimural bacterial community structure and composition, with the CDBS community displaying the greatest diversity. Across all sampling sites, epimural bacterial communities were dominated by members of the phyla Bacteroidetes, Firmicutes, and Proteobacteria. Bacteria within Prevotellaceae, Butyrivibrio, Campylobacter, Mogibacterium, and Desulfobulbus all showed high relative sequence abundance and differential distributions according to sample location. There appears to be a core epimural microbiota present across all locations in all cows, although relative abundance was highly variable. The difference in relative abundance in epimural microbial communities, perhaps influenced by host physiology and the diversity within rumen contents, likely has important consequences for nutrition acquisition and general health. To the best of our knowledge, this work represents the first characterization of the ruminal epimural microbiota across different epithelial locations for any bovine ruminant.

Development of a properly functioning gastrointestinal tract (GIT) at an early age is critical for the wellbeing and lifetime productivity of dairy cattle. The role of early microbial colonization on GIT development in neonatal cattle and the associated molecular changes remain largely unknown, particularly for the small intestine. In this study, we performed artificial dosing of exogenous rumen fluid during the early life of the calf, starting at birth through the weaning transition at 8 wk. Six calves were included in this study. At 8 wk of age, tissue from the ileum was collected and subjected to host transcriptome and microbial metatranscriptome analysis using RNA sequencing. A total of 333 genes showed significant differential expression (DE) (fold-change ≥2; adjusted P < 0.1, mean read-count ≥10) between the treated and control calves. Gene ontology analysis indicated that these DE genes are predominantly associated with processes related to the host immune response (P < 0.0001). Association analysis between the host gene expression and the microbial genus abundance identified 57 genes as having significant correlation with the ileum microbial genera (P < 0.0001). Of these, three genes showed significant association with six microbial genera: lysozyme 2 (LYZ2), fatty acid binding protein 5 (FABP5), and fucosyltransferase (FUT1). Specifically, the profound increase in expression of LYZ2 in treated calves suggests the initiation of antibacterial activity and innate response from the host. Despite the limitation of a relatively small sample size, this study sheds light on the potential impact of early introduction of microbes on the small intestine of calves.

Ruminants harbor a rumen microbial community that convert their host-indigestible diet into nutrients. This rumen microbiome is known to contain three distinct communities: luminal solids, luminal liquids, and the epithelial, or epimural, communities. Currently, there is little research examining the diet-dependent responses of all three microbial communities within the same animals. To address this, we used next-generation 16S rRNA sequencing to characterize the bacterial communities associated with the ruminal solids, liquids, and epimural tissue of 13 lactating and cannulated Holstein dairy cows fed high-starch (diet A) and high-forage (diet B) diets in a crossover experimental design. This crossover design consisted of two groups, two diets, and two sampling periods, with animals receiving a single diet for 8 weeks. Ruminal solids and liquids were collected by separating whole rumen contents through cheesecloth, while rumen epithelial tissue was collected from the ventral-sac via biopsy. Samples were collected at the end of each diet-treatment period. Independent of diet, all sample-types were dominated by bacteria in the phyla Firmicutes and Bacteroidetes. Proteobacteria and Epsilonbacteraeota were also highly abundant, but only in epimural samples. Overall, the epimural microbiota was more diverse, relative to both luminal communities, and diet was found to impact community diversity in solids only. We also found that sample-type, diet-treatment, and their interactions, significantly impacted community structure (Bray-Curtis; PERMANOVA P < 0.05) and composition (Jaccard; PERMANOVA P < 0.05). Of the genera which differed in abundance according to diet and sample-type, Succiniclasticum was highly abundant in epimural samples only, and increased in abundance under a high-forage diet (P < 0.05). In contrast, Saccharofermentans was highly abundant in luminal samples only, but also increased in abundance under a high-forage diet (P < 0.05). These sample-type and diet-dependent differences in specific bacteria likely reflect the overall dietary and environmental factors shaping the heterogeneity of all three ruminal communities.

Crop rotation, the successive cultivation of different crops on the same field, has been practiced for centuries, and it is often associated with increased crop yields. Cover cropping is a less ubiquitous farming practice that also increases plant biodiversity over time. Cover crops are a soil conservation tool; they are grown between harvest and planting of the main crop to protect and enrich the soil. Increasing crop diversity with crop rotation and cover cropping may contribute to shifts in soil bacterial communities. Our first objective was to investigate the soil bacterial communities associated with growing corn (Zea mays L.) or soybean (Glycine max L.) continuously versus annually rotating these crops. Our second objective was to determine if the first season of cover cropping had an impact on soil bacteria in a corn-soybean system. Soil was collected from a long-term crop rotational study with continuous corn, continuous soybean, and annually rotated corn-soybean treatments. These rotation treatments had various cover crops established within each plot, which were sampled individually. Bacterial communities were estimated in each sample by extracting DNA and sequencing the V3-V4 region of the 16S rRNA gene. We found that soil pH, organic matter, and certain macronutrients were essential drivers in determining the composition of bulk soil bacterial communities. Continuously cropped corn and soybean had distinct bacterial communities, while annually rotated communities were similar in both crop phases. The incorporation of cover crops into the rotation system did not result in significant changes to the bulk soil bacterial community. This result was probably due to limited cover crop growth in the first year of establishment, and a limited amount of time for soil communities to respond to this change.

Analysis of the cow microbiome, as well as host genetic influences on the establishment and colonization of the rumen microbiota, is critical for development of strategies to manipulate ruminal function toward more efficient and environmentally friendly milk production. To this end, the development and validation of noninvasive methods to sample the rumen microbiota at a large scale are required. In this study, we further optimized the analysis of buccal swab samples as a proxy for direct bacterial samples of the rumen of dairy cows. To identify an optimal time for sampling, we collected buccal swab and rumen samples at six different time points relative to animal feeding. We then evaluated several biases in these samples using a machine learning classifier (random forest) to select taxa that discriminate between buccal swab and rumen samples. Differences in the inverse Simpson's diversity, Shannon's evenness, and Bray-Curtis dissimilarities between methods were significantly less apparent when sampling was performed prior to morning feeding (P < 0.05), suggesting that this time point was optimal for representative sampling. In addition, the random forest classifier was able to accurately identify nonrumen taxa, including 10 oral and putative feed-associated taxa. Two highly prevalent (>60%) taxa in buccal and rumen samples had significant variance in relative abundances between sampling methods but could be qualitatively assessed via regular buccal swab sampling. This work not only provides new insights into the oral community of ruminants but also further validates and refines buccal swabbing as a method to assess the rumen bacterial in large herds. IMPORTANCE The gastrointestinal tracts of ruminants harbor a diverse microbial community that coevolved symbiotically with the host, influencing its nutrition, health, and performance. While the influence of environmental factors on rumen microbes is well documented, the process by which host genetics influences the establishment and colonization of the rumen microbiota still needs to be elucidated. This knowledge gap is due largely to our inability to easily sample the rumen microbiota. There are three common methods for rumen sampling but all of them present at least one disadvantage, including animal welfare, sample quality, labor, and scalability. The development and validation of noninvasive methods, such as buccal swabbing, for large-scale rumen sampling is needed to support studies that require large sample sizes to generate reliable results. The validation of buccal swabbing will also support the development of molecular tools for the early diagnosis of metabolic disorders associated with microbial changes in large herds.

In mammals, microbial colonization of the digestive tract (GIT) occurs right after birth by several bacterial phyla. Numerous human and mouse studies have reported the importance of early gut microbial inhabitants on host health. However, few attempts have been undertaken to directly interrogate the role of early gut/rumen microbial colonization on GIT development or host health in neonatal ruminants through artificial manipulation of the rumen microbiome. Thus, the molecular changes associated with bacterial colonization are largely unknown in cattle. In this study, we dosed young calves with exogenous rumen fluid obtained from an adult donor cow, starting at birth, and repeated every other week until six weeks of age. Eight Holstein bull calves were included in this study and were separated into two groups of four: the first group was treated with rumen content freshly extracted from an adult cow, and the second group was treated with sterilized rumen content. Using whole-transcriptome RNA-sequencing, we investigated the transcriptional changes in the host liver, which is a major metabolic organ and vital to the calf's growth performance. Additionally, the comparison of rumen epimural microbial communities between the treatment groups was performed using the rRNA reads generated by sequencing. Liver transcriptome changes were enriched with genes involved in cell signaling and protein phosphorylation. Specifically, up-regulation of SGPL1 suggests a potential increase in the metabolism of sphingolipids, an essential molecular signal for bacterial survival in digestive tracts. Notably, eight genera, belonging to four phyla, had significant increases in abundance in treated calves. Our study provides insight into host liver transcriptome changes associated with early colonization of the microbial communities in neonatal calves. Such knowledge provides a foundation for future probiotics-based research in microbial organism mediated rumen development and nutrition in ruminants.

Bacterial contamination of corn-based ethanol biorefineries can reduce their efficiency and hence increase their carbon footprint. To enhance our understanding of these bacterial contaminants, we temporally sampled four biorefineries in the Midwestern USA that suffered from chronic contamination and characterized their microbiomes using both 16S rRNA sequencing and shotgun metagenomics. These microbiotas were determined to be relatively simple, with 13 operational taxonomic units (OTUs) accounting for 90% of the bacterial population. They were dominated by Firmicutes (89%), with Lactobacillus comprising 80% of the OTUs from this phylum. Shotgun metagenomics confirmed our 16S rRNA data and allowed us to characterize bacterial succession at the species level, with the results of this analysis being that Lb. helveticus was the dominant contaminant in this fermentation. Taken together, these results provide insights into the microbiome of ethanol biorefineries and identifies a species likely to be commonly responsible for chronic contamination of these facilities.

Early gut microbial colonization is important for postnatal metabolic and immune development. However, little is known about the effects of different feeding modes (suckling versus bottle-feeding) or microbial sources on this process in farm animals. We found that suckled and bottle-fed newborn lambs had their own distinct gut microbiota. Results from 16S rRNA gene sequencing and qPCR showed that, compared with suckling, bottle feeding significantly increased the abundances of Escherichia/Shigella, Butyricicoccus, and Clostridium XlVa, while significantly decreased the abundance of Clostridium XI. The higher levels of Escherichia/Shigella in bottle-fed lambs suggest that artificial feeding may increase the number of potential pathogens and delay the establishment of the anaerobic environment and anaerobic microbes. Feeding modes also affected the direct transmission of bacteria from the mother and the environment to newborns. The SourceTracker analysis estimated that the early gut microbes of suckled lambs were mainly derived from the mother's teats (43%) and ambient air (28%); whereas those of bottle-fed lambs were dominated by bacteria from the mother's vagina (46%), ambient air (31%), and the sheep pen floor (12%). These findings advance our understanding of gut microbiota in early life and may help design techniques to improve gut microbiota and health.