Microbiota Research Articles

Validation of a Microbiome Dependent Colitis Model via Fecal Microbial Transplant in Germ-Free IL-10 Deficient Mice

Abstract Inflammatory Bowel Diseases (IBD) are immune-mediated intestinal tract diseases with undefined etiologies. Proposed pathogenic mechanisms include abnormal inflammatory response to the intestinal microbiome. Mice deficient in IL-10, a critical cytokine for mucosal immune homeostasis, spontaneously develop enterocolitis; phenotypes are microbiome-sensitive. To establish a microbiome-dependent colitis model with improved construct validity and translational clinical relevance, we assessed colitis development in Germ-Free IL-10 knockout mice (GF IL-10 KO; Taconic Biosciences) following inoculation with fecal microbial transplant (FMT) from wild-type C57BL/6NTac mice at the Murine Pathogen Free™ (MPF™) health standard and assessed clinical responses to anti-IL-12/23p40 (anti-p40). Compared to control GF IL-10 KO mice, FMT-inoculated mice demonstrated reduced weight gain, elevated levels of fecal lipocalin 2 and calprotectin, and increased colon weight:length ratio at 8 weeks following FMT. FMT-inoculated mice also showed elevated histopathology scores, increased colonic CD3+ T cells and CD4+ TH cells, and epithelial hyperplasia by Ki-67 immunolabeling. Several colon cytokines were elevated in FMT-inoculated mice. Anti-p40 treatment significantly reduced all of these disease indicators with the exception of colon weight:length. These data provide a validated colitis model with relevant mechanisms for assessing the role of the microbiome and response to therapeutics in IBD.

Examining the Role of the Gut-Brain Axis in Alcohol Use Disorder in Zebrafish

14.5 million people struggle with Alcohol Use Disorder (AUD), characterized by uncontrolled drinking and withdrawal. Both chronic and acute alcohol use are known to disrupt the intestinal microbiome and are associated with mood disorders. The gut-brain axis is a bidirectional communication pathway between the brain and the intestines. Probiotic modulation has already been shown to alter brain chemistry in zebrafish. Recent research indicates the gut-brain axis is involved in the behavior modifications associated with AUD, however the mechanism by which it does so remains unknown. The objective of this study is to examine the impact that alcohol has on the gut-brain axis. We hypothesize that a reduced intestinal diversity will lead to heightened behavioral responses and alter biomarkers associated with chronic alcohol exposure. In order to examine this hypothesis, zebrafish (Danio rerio) were exposed to amoxicillin and erythromycin for fifteen days to clear their microbiome. Following treatment, fish were immersed in 0.50% ethanol for one hour and in escalating dosage up to 0.25% for two weeks to mimic acute and chronic alcohol exposure. Fish were recorded for 10 minutes following treatment to analyze behavioral patterns. Bacterial populations were significantly knocked down following treatment in both groups (n= 6, p < 0.05). Behavioral results from the acute ethanol treatment indicate fish exposed to ethanol displayed reduced vertical and horizontal transitions in both antibiotic and control groups, with more pronounced shifts in antibiotic-ethanol- treated group (n= 6). This reduction was not observed in the chronically treated fish, suggesting an adaptation effect. In order to further assess cognitive function, levels of NMDA Glut1 receptor mRNA were assessed, we observed no significant changes in receptor gene expression in the brains of zebrafish in the treatment groups relative to control groups. Collectively these results indicate that acute alcohol induced behavior correlates to changes in the microbiome, though this correlation is not observed in chronic alcohol exposure. Future studies are needed to better understand what role the gut brain axis plays in alcohol mediated-behavioral observations. Glen Raven Endowed Grant, Elon University. This is the full abstract presented at the American Physiology Summit 2024 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

Colostrum Supplementation to Rescue Antibiotic-Induced Dysbiosis and Reduce Long-Term Obesity Risk

The prevalence of metabolic disease in the United States has increased from 30.4% to 41.9% in the past decades, with rising rates among children. Emerging evidence implicates intestinal microbiome function as contributing to metabolic disease risk, with dysbiosis being associated with overweight and obese status. While sometimes lifesaving, antibioticssuch as azithromycin cause microbiota ecological instability and are significantly associated with a greater risk ofmetabolic disease and obesity in children. Since antibiotics are often necessary, the identification of nutritional interventions to restore microbial membership and function has a high potential value for millions of patients. Colostrum is the first milk produced by mammals immediately after birth and shapes intestinal colonization patterns, specifically enriching Bifidobacterium, Lactobacillus, and limiting the colonization of Proteobacteria. Bovine colostrum containsimmunomodulators, lactoferrin, oligosaccharides, bioactive compounds, and macronutrients. The compounds are similar across mammalian species. Considering the impact of colostrum in the sterile early-life gut, we hypothesized that bovine colostrum provided as a supplemental nutraceutical therapy would augment gut microbial recolonization following antibiotic exposures and mitigate long-term metabolic consequences. Specific pathogen-free (SPF) C57BL/6 mice (male, three weeks old, Jackson Laboratory) were housed under standard laboratory conditions with a 45% high-fat diet. Tylosintartrate antibiotic (ABX) was administered to some mice in drinking water 3 days/week over three weeks, with four days washout/week. To examine colostrum, some animals were provided colostrum gavage during or immediately following ABX exposures. After intervention, body composition and metabolic phenotypes were monitored for 16 weeks.Compared with ABX alone, colostrum following ABX maintained similar body composition and glucose tolerance as controls, while colostrum during ABX appeared similar to ABX alone. Examination of the gut microbiome showed unique enrichment of Akkermansia and Sutterella with decreased Enterobacteriaceae in mice treated with ABX followed by colostrum. Analysis of the gut mucosa demonstrated ABX decreased goblet cell numbers per villi (7.2/villus) compared with control (9.8/villus), and colostrum following ABX elevated goblet cells (8.6/villus). These data suggest colostrum may stimulate epithelial responses that foster host-microbial symbiosis at the mucosal surface and play a role in intestinal recolonization to benefit long-term homeostasis. Dairy Innovation Hub, UW-Madison. This is the full abstract presented at the American Physiology Summit 2024 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

Intestinal Microbiome and Antimicrobial Resistivity: A Study in Recent Times

The world has recently witnessed a pandemic and is now challenged by continuous environmental changes. Human health is in constant jeopardy with the rise in bacterial, fungal, and viral infections. Antibiotics are the most widely used medication for the treatment of any infection. The urban population does not always maintain a healthy lifestyle due to the fast-paced nature of life. West Bengal the eastern state of India is no exception to this directive. Rampant use of antibiotics and their frequent consumption has widely increased over the years. This is posing a serious threat to the healthy microbiome of the human intestine. It is resulting in a decline of the healthy microbiota and the development of antibiotic resistance. The study aims to understand the effect of antibiotic consumption in the post-COVID scenario on the gut microbiome of the West Bengal population. The population includes both rural and urban as it takes into account the different lifestyles.

Factors that Increase the Risk of Inflammatory Bowel Disease

Inflammatory bowel disease (IBD) is a chronic disorder of the gastrointestinal tract. It comprises Crohn’s disease (CD) and ulcerative colitis (UC) The disorder begins in young adulthood. It is characterised by remission and relapse. IBD significantly affects a person’s life more so, their productivity and ability to perform activities of daily living. Although IBD is prevalent in Western countries, the recent epidemiology data shows a rising global prevalence and incidence. Addressing IBD is important. Although it is not significantly associated with high morbidity rates, it is a significant burden to healthcare. Identifying the risk factors of IBD can help in improving preventative measures. Since IBD does not have a known cure, controlling for these factors can minimize the impact of the disease. Some of the risk factors that increase the risk of IBD are genetics, environmental factors such as smoking, diet, lifestyle changes, medications, and intestinal microbiome. Identifying the factors that contribute to the risk of developing IBD can help in creating effective measures that can prevent the pathogenesis of the disease.

Effects of Alhagi camelorum Fisch polysaccharide from different regions on growth performance and gastrointestinal microbiota of sheep lambs.

Polysaccharides derived from Alhagi camelorum Fisch possess diverse activities, making them a potential prebiotic candidates for enhancing lamb health. This study investigated the immunomodulatory effects of Alhagi camelorum Fisch polysaccharides from Aksu (AK) and Shanshan (SS) regions on sheep lambs. The results showed that sheep lambs in the SS group exhibited significantly increased (p < 0.05) average daily gain, levels of growth hormone (GH), insulin (INS), IgA and IgM, and cytokines IL-4, IL-10, IL-17, TNF-α and IFN-γ compared to those in the control check (CK) group. Moreover, the SS treatment significantly increased the diversity and abundance of beneficial bacteria, while concurrently diminishing the prevalence of harmful bacteria. Additionally, it modulated various metabolic pathways, promoted lamb growth, improved immunity, reduced the risk of gastrointestinal disease and improved the composition of gastrointestinal microbiota. In summary, our findings highlight the potential of SS treatment in enhancing gastrointestinal health of sheep lambs by improving intestinal function, immunity, and gut microbiome. Consequently, these results suggest that Alhagi camelorum Fisch polysaccharides derived from Shanshan regions holds promising potential as a valuable intervention for optimizing growth performance in sheep lambs.

Xylans as a promising prebiotic agent: a brief review

Xylans are the most abundant hemicelluloses in nature, constituting components of the secondary cell wall of plant cells. They are polysaccharides with versatile properties, which can be used in various industrial sectors. Several biological activities have been identified in different xylans, such as antioxidant, antitumor, antimicrobial, emulsifying, anticoagulant, and immunomodulatory activities. Among these applications of xylans, its use as a prebiotic stands out, composing functional foods. Xylans form non-digestible dietary fibers that serve as a source of energy for beneficial microorganisms, modifying the intestinal microbiome and bringing a general improvement to the health status of those who consume them. The literature reports the use of xylooligosaccharides as promoters of the growth of probiotic microorganisms, such as species of Lactobacillus and Bifidobacterium, in addition to treating dysbiosis and reducing intestinal inflammation. Due to their structural diversity, abundance in nature, and a wide range of biological properties, xylans are valuable components that can be used to create various products aimed at enhancing human health.

Nanopore and Illumina sequencing reveal different viral populations from human gut samples.

The advent of viral metagenomics, or viromics, has improved our knowledge and understanding of global viral diversity. High-throughput sequencing technologies enable explorations of the ecological roles, contributions to host metabolism, and the influence of viruses in various environments, including the human intestinal microbiome. However, bacterial metagenomic studies frequently have the advantage. The adoption of advanced technologies like long-read sequencing has the potential to be transformative in refining viromics and metagenomics. Here, we examined the effectiveness of long-read and hybrid sequencing by comparing Illumina short-read and Oxford Nanopore Technology (ONT) long-read sequencing technologies and different assembly strategies on recovering viral genomes from human faecal samples. Our findings showed that if a single sequencing technology is to be chosen for virome analysis, Illumina is preferable due to its superior ability to recover fully resolved viral genomes and minimise erroneous genomes. While ONT assemblies were effective in recovering viral diversity, the challenges related to input requirements and the necessity for amplification made it less ideal as a standalone solution. However, using a combined, hybrid approach enabled a more authentic representation of viral diversity to be obtained within samples.

Intestinal Microbiome Associated with Efficacy of Atezolizumab and Bevacizumab Therapy for Hepatocellular Carcinoma.

The combination of atezolizumab and bevacizumab has become the first-line treatment for patients with unresectable hepatocellular carcinoma (HCC). However, no studies have reported on specific intestinal microbiota associated with the efficacy of atezolizumab and bevacizumab. In this study, we analyzed fecal samples collected before treatment to investigate the relationship between the intestinal microbiome and the efficacy of atezolizumab and bevacizumab. A total of 37 patients with advanced HCC who were treated with atezolizumab and bevacizumab were enrolled. Fecal samples were collected from the patients, and they were divided into responder (n = 28) and non-responder (n = 9) groups. We compared the intestinal microbiota of the two groups and analyzed the intestinal bacteria associated with prognosis using QIIME2. The alpha and beta diversities were not significantly different between both groups, and the proportion of microbiota was similar. The relative abundance of Bacteroides stercoris and Parabacteroides merdae was higher in the responder group than in the non-responder group. When the prognosis was analyzed by the presence or absence of those bacteria, patients without both had a significantly poorer prognosis. Differences in intestinal microbiome are involved in the therapeutic effect of atezolizumab and bevacizumab.

Role of the Gut Microbiota-Brain Axis in Brain Damage in Preterm Infants.

The greatest repository of microbes in the human body, the intestinal microbiome, is involved in neurological development, aging, and brain illnesses such as white matter injury (WMI) in preterm newborns. Intestinal microorganisms constitute a microbial gut-brain axis that serves as a crucial conduit for communication between the gut and the nervous system. This axis controls inflammatory cytokines, which in turn influence the differentiation of premyelinating oligodendrocytes (pre-OLs) and influence the incidence of WMI in premature newborns through the metabolites generated by gut microbes. Here, we describe the effects of white matter injury (WMI) on intestinal dysbiosis and gut dysfunction and explain the most recent research findings on the gut-brain axis in both humans and animals. We also emphasize the delicate relationship that exists between the microbiota and the brain following acute brain injury. The role that the intestinal microflora plays in influencing host metabolism, the immune system, brain health, and the course of disease is becoming increasingly clear, but there are still gaps in the field of WMI treatment. Thus, this review demonstrates the function of the gut microflora-brain axis in WMI and elucidates the possible mechanisms underlying the communication between gut bacteria and the developing brain via the gut-brain axis, potentially opening up new avenues for microbial-based intervention and treatment for preterm WMI.

Synergistic Effect of Dietary Supplementation with Sodium Butyrate, β-Glucan and Vitamins on Growth Performance, Cortisol Level, Intestinal Microbiome and Expression of Immune-Related Genes in Juvenile African Catfish (Clarias gariepinus).

The effect of dietary supplementation with sodium butyrate, β-glucan and vitamins (A, D3, E, K, C) on breeding indicators and immune parameters of juvenile African catfish was examined. The fish were fed with unenriched (group C) and enriched feed with a variable proportion of sodium butyrate/β-glucan, and constant content of vitamins (W1-W3). After the experiment, blood and the middle gut were collected. The microbiome of the gut was determined using Next Generation Sequencing (NGS). Liver tissue was collected for determination of expression of immune-related genes (HSP70, IL-1β, TNFα). W2 and W3 were characterized by the most favorable values of breeding indicators (p < 0.05). The highest blood cortisol concentration was in group C (71.25 ± 10.45 ng/mL), and significantly the lowest in W1 (46.03 ± 7.01 ng/ mL) (p < 0.05). The dominance of Cetobacterium was observed in all study groups, with the largest share in W3 (65.25%) and W1 (61.44%). Gene expression showed an increased number of HSP70 genes in W1. IL-1β and TNFα genes peaked at W3. The W3 variant turns out to be the most beneficial supplementation, due to the improvement of breeding and immunological parameters. The data obtained can be used to create a preparation for commercial use in the breeding of this species.

Early Life Exposures and the Development of the Infant Gut Microbiome: A Review

The influence of the intestinal microbiota on metabolic, nutritional, and immunological processes is widely reported. With increasing literature associating altered microbial compositions with adverse health outcomes, it is important to understand how early life exposures may impact the development of gut microbial colonization and subsequent risk of altered metabolic and immune regulation. The purpose of this review is to describe factors in the maternal prenatal and perinatal period that may impact the intestinal microbiome over the first 2 years of life. A comprehensive search of MEDLINE, EMBASE, and the Cochrane Library using Medical Subject Headings (MeSH) and keywords for studies reporting on determinants of gut microbiota in infants (0 to 24 months) born at full term was conducted. Articles using culture techniques were included but not those that exclusively used molecular techniques that lacked sensitivity. Each citation title and abstract was independently assessed for inclusion for full text review. Findings related to the maternal prenatal period, mode of birthing, infant diet and antibiotics were included. Bifidobacteria and Bacteroides abundance were consistently greater in the first weeks of life in children born vaginally, and increased Bacteroides presence persisted throughout the first year. Bifidobacteria abundance was greater in breastfed children. Introduction of solid food was associated with greater presence of bacteria of the Firmicutes phylum. Although these studies advance our knowledge of how exposures in prenatal, intrapartum, and early life may impact colonization, larger studies with longitudinal follow-up are needed to improve our understanding of how perturbations may contribute to early origins of disease. This article has been peer reviewed.

Controlling Candida: immune regulation of commensal fungi in the gut.

The intestinal microbiome harbors fungi that pose a significant risk to human health as opportunistic pathogens and drivers of inflammation. Inflammatory and autoimmune diseases are associated with dysbiotic fungal communities and the expansion of potentially pathogenic fungi. The gut is also the main reservoir for disseminated fungal infections. Immune interactions are critical for preventing commensal fungi from becoming pathogenic. Significant strides have been made in defining innate and adaptive immune pathways that regulate intestinal fungi, and these discoveries have coincided with advancements in our understanding of the fungal molecular pathways and effectors involved in both commensal colonization and pathogenesis within the gut. In this review, we will discuss immune interactions important for regulating commensal fungi, with a focus on how specific cell types and effectors interact with fungi to limit their colonization or pathogenic potential. This will include how innate and adaptive immune pathways target fungi and orchestrate antifungal immune responses, in addition to how secreted immune effectors, such as mucus and antimicrobial peptides, regulate fungal colonization and inhibit pathogenic potential. These immune interactions will be framed around our current understanding of the fungal effectors and pathways regulating colonization and pathogenesis within this niche. Finally, we highlight important unexplored mechanisms by which the immune system regulates commensal fungi in the gut.

Single-cell transcriptomics unveiled that early life BDE-99 exposure reprogrammed the gut-liver axis to promote a proinflammatory metabolic signature in male mice at late adulthood.

Polybrominated diphenyl ethers (PBDEs) are legacy flame retardants that bioaccumulate in the environment. The gut microbiome is an important regulator of liver functions including xenobiotic biotransformation and immune regulation. We recently showed that neonatal exposure to polybrominated diphenyl ether-99 (BDE-99), a human breast milk-enriched PBDE congener, up-regulated proinflammation-related and down-regulated drug metabolism-related genes predominantly in males in young adulthood. However, the persistence of this dysregulation into late adulthood, differential impact among hepatic cell types, and the involvement of the gut microbiome from neonatal BDE-99 exposure remain unknown. To address these knowledge gaps, male C57BL/6 mouse pups were orally exposed to corn oil (10 ml/kg) or BDE-99 (57 mg/kg) once daily from postnatal days 2-4. At 15 months of age, neonatal BDE-99 exposure down-regulated xenobiotic and lipid-metabolizing enzymes and up-regulated genes involved in microbial influx in hepatocytes. Neonatal BDE-99 exposure also increased the hepatic proportion of neutrophils and led to a predicted increase of macrophage migration inhibitory factor signaling. This was associated with decreased intestinal tight junction protein (Tjp) transcripts, altered gut environment, and dysregulation of inflammation-related metabolites. ScRNA-seq using germ-free (GF) mice demonstrated the necessity of a normal gut microbiome in maintaining hepatic immune tolerance. Microbiota transplant to GF mice using large intestinal microbiome from adults neonatally exposed to BDE-99 down-regulated Tjp transcripts and up-regulated several cytokines in large intestine. In conclusion, neonatal BDE-99 exposure reprogrammed cell type-specific gene expression and cell-cell communication in liver towards proinflammation, and this may be partly due to the dysregulated gut environment.

Enterotype-Dependent Probiotic-Mediated Changes in the Male Rat Intestinal Microbiome In Vivo and In Vitro.

Beneficial properties of lactic acid bacteria have been known long ago, but particular interest in probiotics has arisen in the last two decades due to the understanding of the important role of intestinal microflora in human life. Thus, the ability of probiotics to support healthy homeostasis of gut microbiomes has received particular attention. Here, we evaluated the effect of a probiotic consisting of Bifidobacterium longum and Lacticaseibacillus paracasei on the gut microbiome of male rats, assessed their persistence in the fecal biota, and compared probiotic-mediated changes in vitro and in vivo. As expected, microbiomes of two enterotypes were identified in the feces of 21 animals, and it turned out that even a single dose of the probiotic altered the microbial composition. Upon repeated administration, the E1 biota temporarily acquired properties of the E2 type. Being highly sensitive to the intervention of probiotic bacteria at the phylum and genus levels, the fecal microbiomes retained the identity of their enterotypes when transferred to a medium optimized for gut bacteria. For the E2 biota, even similarities between probiotic-mediated reactions in vitro and in vivo were detected. Therefore, fecal-derived microbial communities are proposed as model consortia to optimize the response of resident bacteria to various agents.

Characteristics of intestinal microbiome in very low birth weight preterm neonatal intensive care unit patients

characteristics of intestinal microbiome in very low birth weight preterm neonatal intensive care unit patients

Single-cell calcium monitoring of Caco-2 cell co-cultured with intestinal microbiome through carbon fiber based potentiometric microelectrode

The Caco-2 cells were used as intestinal epithelial cell model to illustrate the hyperuricemia (HUA) mechanism under the co-culture of the imbalanced intestinal microbiome in this work. The uric acid (UA) concentration in the HUA process was monitored, and could be up to 425 μmol/L at 8 h co-cultured with the imbalanced intestinal microbiome. Single-cell potentiometry based on ion-selective microelectrode was used to study extracellular calcium change, which is hypothesized to play an important role in the UA excretion. The potential signal of the calcium in the extremely limited microenvironment around single Caco-2 cell was recorded through the single-cell analysis platform. The potential signal of sharp decrease and slow increase followed within a few seconds indicates the sudden uptake and gradually excretion process of calcium through the cell membrane. Moreover, the value of the potential decrease increases with the increase of the time co-cultured with the imbalanced intestinal microbiome ranging from 0 to 8 h. The Ca2+ concentration around the cell membrane could decrease from 1.3 mM to 0.4 mM according to the potential decrease of 27.0 mV at the co-culture time of 8 h. The apoptosis ratio of the Caco-2 cells also exhibits time dependent with the co-culture of the imbalanced intestinal microbiome, and was 39.1 ± 3.6 % at the co-culture time of 8 h, which is much higher than the Caco-2 cells without any treatment (3.9 ± 2.9 %). These results firstly provide the links between the UA excretion with the apoptosis of the intestinal epithelial cell under the interaction of the imbalanced intestinal microbiome. Moreover, the apoptosis could be triggered by the calcium signaling.

Effect of different levels of single cell protein and probiotic microorganisms on performance, immunological responses, and intestinal histology in laying hens

AbstractAn experiment was conducted to evaluate the effects of single‐cell protein (SCP), as a valuable by‐product and substitute for soybean meal, and multistrain probiotics on the performance, blood biochemistry, intestinal histomorphology, ileal microbiota, and immune responses in laying hens. Leghorn laying hens were randomly assigned to nine experimental treatments with eight replicates of five birds each. Dietary treatments consisted of three levels of replacement of soybean meal with SCP (0.0%, 30%, and 60%) and three levels of probiotics (0.0, 50, 100 mg/kg) offered through 70 days of main recording. The result indicated that replacing 30% soybean meal with SCP in the diet significantly increased the eggshell weight, thickness, and strength; however, hens receiving 60% SCP replacement showed lower productive performance. During the second 35‐d trial, hens receiving at least 50 mg/kg probiotics produced higher eggshell weight, thickness, and strength than the control hens. Remarkable lower levels of serum cholesterol content were observed in the SCP and probiotic‐supplemented diets. The significant reduction in the ileal pH, Escherichia coli and Clostridium spp. enumerations and a remarkable increase in Lactobacillus count was observed after feeding incremental levels of probiotics. The highest villus height to crypt depth ratio, antibody titer against sheep red blood cells (in secondary response) and Newcastle disease vaccine, as well as egg production and egg mass, were found in the hens that received 30% SCP replacement in combination with 50 or 100 mg/kg probiotic (SCP × Probiotic). The present study concluded that feeding SCP, as a cheap protein alternative source, along with probiotics while reducing production costs can support productive performance via modulating the intestinal microbiomes and health indices and fortifying the immune system in laying hens.

Crude protein content in diets associated with intestinal microbiome and metabolome alteration in Huanjiang mini-pigs during different growth stages.

Adequate crude protein (CP) content in diets plays a crucial role in the intestinal health of the animal. This study investigated the impacts of CP content in diets on the intestinal microbiome and metabolome profiles in growing Huanjiang mini-pigs. A total of 360 pigs with similar body weight (BW) were allocated for three independent feeding trials based on three different BW stages, including (i) 5-10 kg BW, diets consisting of 14, 16, 18, 20, and 22% CP content; (ii) 10-20 kg BW, diets consisting of 12, 14, 16, 18, and 20% CP content; and (iii) 20-30 kg BW, diets consisting of 10, 12, 14, 16, and 18% CP content. These experiments lasted 28, 28, and 26 days, respectively. The results showed that the Shannon and Simpson indices were decreased (p < 0.05) in the ileum of pigs in response to the 14-18% CP compared with the 20% CP content at 5-10 kg BW stage, while diets containing 12 and 14% CP had higher Chao1 (p < 0.05) and Shannon (p = 0.054) indices compared with 18% CP at 20-30 kg BW stage. Compared with the 20% CP, the diet containing 16% CP displayed an increasing trend (p = 0.089) of Firmicutes abundance but had decreased (p = 0.056) Actinobacteria abundance in the jejunum at 5-10 kg BW stage. In addition, a diet containing 16% CP had higher Lactobacillus abundance in the jejunum and ileum compared with the 18, 20, and 22% CP, while had lower Sphingomonas and Pelomonas abundances in the jejunum and Streptococcus abundance in the ileum compared with the diet containing 22% CP (p < 0.05). Diets containing lower CP content altered differential metabolites in the small intestine at the early stage, while higher CP content had less impact. These findings suggest that a diet containing lower CP content (16% CP) may be an appropriate dietary CP content for 5-10 kg Huanjiang mini-pigs, as 16% CP content in diet has shown beneficial impacts on the intestinal microbiome and metabolome profiles at the early growth stage of pigs.

Exploring the immune-inflammatory mechanism of Maxing Shigan Decoction in treating influenza virus A-induced pneumonia based on an integrated strategy of single-cell transcriptomics and systems biology.

Influenza is an acute respiratory infection caused by influenza virus. Maxing Shigan Decoction (MXSGD) is a commonly used traditional Chinese medicine prescription for the prevention and treatment of influenza. However, its mechanism remains unclear. The mice model of influenza A virus pneumonia was established by nasal inoculation. After 3days of intervention, the lung index was calculated, and the pathological changes of lung tissue were detected by HE staining. Firstly, transcriptomics technology was used to analyze the differential genes and important pathways in mouse lung tissue regulated by MXSGD. Then, real-time fluorescent quantitative PCR (RT-PCR) was used to verify the changes in mRNA expression in lung tissues. Finally, intestinal microbiome and intestinal metabolomics were performed to explore the effect of MXSGD on gut microbiota. The lung inflammatory cell infiltration in the MXSGD group was significantly reduced (p < 0.05). The results of bioinformatics analysis for transcriptomics results show that these genes are mainly involved in inflammatory factors and inflammation-related signal pathways mediated inflammation biological modules, etc. Intestinal microbiome showed that the intestinal flora Actinobacteriota level and Desulfobacterota level increased in MXSGD group, while Planctomycetota in MXSGD group decreased. Metabolites were mainly involved in primary bile acid biosynthesis, thiamine metabolism, etc. This suggests that MXSGD has a microbial-gut-lung axis regulation effect on mice with influenza A virus pneumonia. MXSGD may play an anti-inflammatory and immunoregulatory role by regulating intestinal microbiome and intestinal metabolic small molecules, and ultimately play a role in the treatment of influenza A virus pneumonia.