Effects of supplemental bacteriophage on the gut microbiota and nutrient digestibility of ileal-cannulated pigs.

This study measured the potential changes of the microbiota in the gastrointestinal tract and energy and nutrient digestibility by supplemental bacteriophages in pigs. Twelve castrated male pigs (initial mean body weight = 29.5 ± 2.3 kg) were surgically cannulated using T-cannula. The animals were housed individually in pens equipped with a feeder and a nipple waterer. The pigs were allotted to 1 of 3 experimental diets in a quadruplicated 3 × 2 Latin square design with 3 experimental diets, 2 periods, and 12 pigs resulting in 8 replicates per diet. The 3 diets were a control mainly based on corn and soybean meal with no antibiotics or bacteriophages, a diet containing 0.1% antibiotics, and a diet containing 0.2% bacteriophages. On day 5 of the experimental period, feces were collected and on days 6 and 7, ileal digesta were collected. Genomic DNA for bacteria were extracted from the ileal digesta and feces and the V4 region of the 16S rRNA gene was amplified. The ileal and fecal digestibility of energy, dry matter, organic matter, crude protein, and fiber was unaffected by dietary antibiotics or bacteriophages. At the phylum level, the supplemental antibiotic or bacteriophage tended to result in a higher proportion of Firmicutes (p = 0.059) and a lower proportion of Bacteroidetes (p = 0.099) in the ileal digesta samples compared with the control group with no difference between the antibiotic and bacteriophage groups. At the genus level, the supplemental antibiotic or bacteriophage tended to result in a higher proportion of Lactobacillus (p = 0.062) and a lower proportion of Bacteroides (p = 0.074) and Streptococcus (p = 0.088) in the ileal digesta compared with the control group with no difference between the antibiotic and bacteriophage groups. In the feces, supplemental antibiotics or bacteriophages reduced the proportion of Bifidobacterium compared with the control group (p = 0.029) with no difference between the antibiotic and bacteriophage groups. Overall, supplemental antibiotics and bacteriophages showed positive effect on the microbiota of in the ileal digesta without largely affecting energy or nutrient digestibility, with no differences between the antibiotic and bacteriophage groups in growing pigs.

Complement-ary protection for all ages.

Complement proteins form a proteolytic cascade to clear invading microbes. In a recent issue of Cell, Wu et al. and Xu et al. demonstrate two distinct complement systems operating in the gut, independent of circulating complement, and protecting against intestinal pathogens.

FMRP deficiency leads to multifactorial dysregulation of splicing and mislocalization of MBNL1 to the cytoplasm.

Fragile X syndrome (FXS) is a neurodevelopmental disorder that is often modeled in Fmr1 knockout mice where the RNA-binding protein FMRP is absent. Here, we show that in Fmr1-deficient mice, RNA mis-splicing occurs in several brain regions and peripheral tissues. To assess molecular mechanisms of splicing mis-regulation, we employed N2A cells depleted of Fmr1. In the absence of FMRP, RNA-specific exon skipping events are linked to the splicing factors hnRNPF, PTBP1, and MBNL1. FMRP regulates the translation of Mbnl1 mRNA as well as Mbnl1 RNA auto-splicing. Elevated Mbnl1 auto-splicing in FMRP-deficient cells results in the loss of a nuclear localization signal (NLS)-containing exon. This in turn alters the nucleus-to-cytoplasm ratio of MBNL1. This redistribution of MBNL1 isoforms in Fmr1-deficient cells could result in downstream splicing changes in other RNAs. Indeed, further investigation revealed that splicing disruptions resulting from Fmr1 depletion could be rescued by overexpression of nuclear MBNL1. Altered Mbnl1 auto-splicing also occurs in human FXS postmortem brain. These data suggest that FMRP-controlled translation and RNA processing may cascade into a general dys-regulation of splicing in Fmr1-deficient cells.

Epithelial organoid supports resident memory CD8 T cell differentiation.

Resident Memory T cells (TRM) play a vital role in regional immune defense in barrier organs. Although laboratory rodents have been extensively used to study fundamental TRM biology, poor isolation efficiency, sampling bias and low cell survival rates have limited our ability to conduct TRM-focused high-throughput assays. Here, we engineered a murine vaginal epithelial organoid (VEO)-CD8 T cell co-culture system that supports CD8 TRM differentiation in vitro. The three-dimensional VEOs established from murine adult stem cells resembled stratified squamous vaginal epithelium and induced gradual differentiation of activated CD8 T cells into epithelial TRM. These in vitro generated TRM were phenotypically and transcriptionally similar to in vivo TRM, and key tissue residency features were reinforced with a second cognate-antigen exposure during co-culture. TRM differentiation was not affected even when VEOs and CD8 T cells were separated by a semipermeable barrier, indicating soluble factors' involvement. Pharmacological and genetic approaches showed that TGF-ß signaling played a crucial role in their differentiation. We found that the VEOs in our model remained susceptible to viral infections and the CD8 T cells were amenable to genetic manipulation; both of which will allow detailed interrogation of antiviral CD8 T cell biology in a reductionist setting. In summary, we established a robust model which captures bonafide TRM differentiation that is scalable, open to iterative sampling, and can be subjected to high throughput assays that will rapidly add to our understanding of TRM.

Microbial underdogs: exploring the significance of low-abundance commensals in host-microbe interactions.

Our understanding of host-microbe interactions has broadened through numerous studies over the past decades. However, most investigations primarily focus on the dominant members within ecosystems while neglecting low-abundance microorganisms. Moreover, laboratory animals usually do not have microorganisms beyond bacteria. The phenotypes observed in laboratory animals, including the immune system, have displayed notable discrepancies when compared to real-world observations due to the diverse microbial community in natural environments. Interestingly, recent studies have unveiled the beneficial roles played by low-abundance microorganisms. Despite their rarity, these keystone taxa play a pivotal role in shaping the microbial composition and fulfilling specific functions in the host. Consequently, understanding low-abundance microorganisms has become imperative to unravel true commensalism. In this review, we provide a comprehensive overview of important findings on how low-abundance commensal microorganisms, including low-abundance bacteria, fungi, archaea, and protozoa, interact with the host and contribute to host phenotypes, with emphasis on the immune system. Indeed, low-abundance microorganisms play vital roles in the development of the host's immune system, influence disease status, and play a key role in shaping microbial communities in specific niches. Understanding the roles of low-abundance microbes is important and will lead to a better understanding of the true host-microbe relationships.

Mucin-binding adhesins: A key to unlocking the door of mutualism.

From corals to mammals, mucus is a conserved feature that prevents microbes from accessing the surfaces that produce it. However, interactions between mucus and microbes remain poorly understood. In this issue of Cell Host & Microbe, Smith et al. unveil that mucus binding by bacteria is crucial for host-microbe symbiosis.

Omega-3-Rich Fish-Oil-Influenced Mouse Gut Microbiome Shaped by Intermittent Consumption of Beef.

Beef consumption can provide various amino acids, lipids, vitamins, and minerals; however, excessive intake causes metabolic disorders and increases the probability of obesity, atherosclerosis, and colorectal cancer. The intake of omega-3 fatty acids can ameliorate metabolic disorders by lowering blood glucose and triglyceride levels. In the present study, we investigated the effect of omega-3-rich fish oil on body performance and the gut microbiome in a beef-rich diet. Four-week-old C57BL/6 mice were distributed into four groups (chow diet [Chow], chow with beef diet [Beef], chow with omega-3 diet [Cw3], and chow with beef and omega-3 diet [Bw3]). We observed that body weight was unaltered between groups, and serum triglyceride levels were reduced in the omega-3 supplemented groups. The beta diversity indices, unweighted UniFrac distance (P = 0.001), and Jaccard distance (P = 0.001) showed statistically significant differences, and the principal coordinates analysis plot showed a clear separation between groups. In addition, the taxonomic comparison revealed that beef consumption increased numerous potentially pathogenic bacteria, including Escherichia-Shigella, Mucispirillum, Helicobacter, and Desulfovibrio, which were decreased following omega-3 supplementation. Metabolic comparison based on 16S rRNA revealed that energy and glucose metabolism were higher in omega-3 supplemented groups. Our findings suggest that the omega-3 supplementation under intermittent beef consumption contributes to changes in the gut microbiome and microbial metabolic pathways.

Low abundance members of the gut microbiome exhibit high immunogenicity.

Studies identifying bacterial members that dictate host phenotype have focused mainly on the dominant members, and the role of low abundance microbes in determining host phenotypes and pathogenesis of diseases remains unexplored. In this study, we compared the gut bacterial community of mice with wide-ranging microbial exposure to determine if low abundance bacteria vary based on microbial exposure or remain consistent. We noted that similar to the high abundance bacterial community, a core community of low abundance bacteria made up a significant portion of the gut microbiome irrespective of microbial exposure. To determine the role of low abundance bacteria in regulating community composition and host gene expression, we devised a microbiome dilution strategy to "delete" out low abundance bacteria and engrafted the diluted microbiomes into germ-free mice. Our approach successfully excluded low abundance bacteria from small and large intestinal bacterial communities and induced global changes in microbial community composition in the large intestine. Gene expression analysis of intestinal tissue revealed that loss of low abundance bacteria resulted in a drastic reduction in expression of multiple genes involved MHCII antigen presentation pathway and T-cell cytokine production in the small intestine. The effect of low abundance bacteria on MHCII expression was found to be specific to the intestinal epithelium at an early timepoint post-colonization and correlated with bacteria belonging to the family Erysipelotrichaceae. We conclude that low abundance bacteria have a significantly higher immuno-stimulatory effect compared to dominant bacteria and are thus potent drivers of early immune education in the gut.

Gut commensals expand vitamin A metabolic capacity of the mammalian host.

Conversion of dietary vitamin A (VA) into retinoic acid (RA) is essential for many biological processes and thus far studied largely in mammalian cells. Using targeted metabolomics, we found that commensal bacteria in the mouse gut lumen produced a high concentration of the active retinoids, all-trans-retinoic acid (atRA) and 13-cis-retinoic acid (13cisRA), as well as the principal circulating retinoid, retinol. Ablation of anerobic bacteria significantly reduced retinol, atRA, and 13cisRA, whereas introducing these bacteria into germ-free mice significantly enhanced retinoids. Remarkably, cecal bacterial supplemented with VA produced active retinoids in vitro, establishing that gut bacteria encode metabolic machinery necessary for multistep conversion of dietary VA into its active forms. Finally, gut bacteria Lactobacillus intestinalis metabolized VA and specifically restored RA levels in the gut of vancomycin-treated mice. Our work establishes vitamin A metabolism as an emergent property of the gut microbiome and lays the groundwork for developing probiotic-based retinoid therapy.