Intact Microbiota Research Articles

Bee microbiomes in a changing climate: Investigating the effects of temperature on solitary bee life history and health.

Climate change is rapidly warming thermal environments, an important abiotic stimulus governing interactions between microbial symbionts and their hosts. Increasing evidence suggests that solitary bees rely on pollen provision microbes for successful development. However, the effects of heat stress on provision microbiota and the resulting consequences for larval health and development remain to be examined. We performed an in vitro study to investigate the effects of the thermal environment on provision microbiome composition and measured fitness outcomes for Osmia lignaria larvae. While pollen sterilisation removed bacteria from microbe-rich provisions, larval survivorship did not significantly differ between bees reared on microbe-rich (unmanipulated) diets and provisions treated with ethylene oxide (EO) gas. In contrast to previous research in solitary bees, larvae reared on EO-treated provisions weighed more and had higher total fat content, with temperature moderating the degree of difference. As anticipated, we observed a negative relationship between the duration of larval development and temperature. Our results indicated that an intact provision microbiota may not always improve bee fitness and that bee-microbe interactions during larval development may contribute to the size-shrinking effect observed for cavity-nesting bees under warming conditions.

Salmonella re-engineers the intestinal environment to break colonization resistance in the presence of a compositionally intact microbiota

The gut microbiota prevents harmful microbes from entering the body, a function known as colonization resistance. The enteric pathogen Salmonella enterica serovar (S.) Typhimurium uses its virulence factors to break colonization resistance through unknown mechanisms. Using metabolite profiling and genetic analysis, we show that the initial rise in luminal pathogen abundance was powered by a combination of aerobic respiration and mixed acid fermentation of simple sugars, such as glucose, which resulted in their depletion from the metabolome. The initial rise in the abundance of the pathogen in the feces coincided with a reduction in the cecal concentrations of acetate and butyrate and an increase in epithelial oxygenation. Notably, these changes in the host environment preceded changes in the microbiota composition. We conclude that changes in the host environment can weaken colonization resistance even in the absence of overt compositional changes in the gut microbiota.

FBS-based cryoprotective compositions for effective cryopreservation of gut microbiota and key intestinal microorganisms

ObjectiveThe need for innovative techniques to preserve microbiota for extended periods, while maintaining the species composition and quantitative balance of the bacterial community, is becoming increasingly important. To address this need, we propose an efficient approach to cryopreserve human gut microbiota using a two-component cryoprotective composition comprising fetal bovine serum (FBS) and 5% dimethyl sulfoxide (DMSO). Fetal serum is a commonly utilized component in the freezing media for eukaryotic cells, however, its effects on prokaryotic cells have not been extensively researched.ResultsIn our study, we demonstrated the high efficiency of using a two-component cryoprotective medium, FBS + 5% DMSO, for cryopreservation of human gut microbiota using three different methods. According to the obtained results, the intact donor microbiota was preserved at a level of 85 ± 4% of the initial composition based on fluorescent analysis using the LIVE/DEAD test. No differences in survival were observed when comparing with pure DMSO and FBS media. The photometric measurement method for growth of aerobic bacteria (A. johnsoni), facultative anaerobes (E. coli, E. faecalis), microaerophilic (L. plantarum), and obligate anaerobic bacterial cultures (E. barkeri, B. breve) also demonstrated high viability rates of 94–98% in the two-component protective medium, reaching intact control levels. However, for anaerobic microflora representatives, serum proved to be a more suitable cryoprotectant. Also, we demonstrated that using cryoprotective media with 50–75% FBS content is enough to preserve a significant level of bacterial cell viability, from an economic standpoint.

Myeloid Cells and Sphingosine-1-Phosphate Are Required for TCRαβ Intraepithelial Lymphocyte Recruitment to the Colon Epithelium.

Intraepithelial lymphocytes (IELs) are T cells important for the maintenance of barrier integrity in the intestine. Colon IELs are significantly reduced in both MyD88-deficient mice and those lacking an intact microbiota, suggesting that MyD88-mediated detection of bacterial products is important for the recruitment and/or retention of these cells. Here, using conditionally deficient MyD88 mice, we show that myeloid cells are the key mediators of TCRαβ+ IEL recruitment to the colon. Upon exposure to luminal bacteria, myeloid cells produce sphingosine-1-phosphate (S1P) in a MyD88-dependent fashion. TCRαβ+ IEL recruitment may be blocked using the S1P receptor antagonist FTY720, confirming the importance of S1P in the recruitment of TCRαβ+ IELs to the colon epithelium. Finally, using the TNFΔARE/+ model of Crohn's-like bowel inflammation, we show that disruption of colon IEL recruitment through myeloid-specific MyD88 deficiency results in reduced pathology. Our results illustrate one mechanism for recruitment of a subset of IELs to the colon.

A Probiotic Targets Bile Acids Metabolism to Alleviate Ulcerative Colitis by Reducing Conjugated Bile Acids.

Gut microbiota (GM) dysbiosis and dysregulated bile acids (BAs) metabolism have been linked to ulcerative colitis (UC) pathogenesis. The possibility of utilizing live probiotics with a defined BAs-metabolizing capability to modify the composition BAs for UC treatment remains unexplored. In this study, Strain GR-4 is sourced from traditional Chinese fermented food, "Jiangshui," and demonstrated the ability to deconjugate two common conjugated BAs by over 69% and 98.47%, respectively. It administers strain GR-4 to dextran sulfate sodium (DSS)-induced UC mice, and observes an overall alleviation of UC symptoms, as evidence by improved colon morphology, reduces inflammation and oxidative stress, and restores intestinal barrier function. Importantly, these effects are reliant on an intact commensal microbiota, as depletion of GM mitigated GR-4s efficacy. Metabolomics analysis unveils a decline in conjugated BAs and an increase in secondary BAs following GR-4 administration. GM analysis indicates that GR-4 selectively enriches bacterial taxa linked to BAs metabolism, enhancing GM's capacity to modify BAs. This research demonstrates the potential for natural fermented foods and probiotics to effectively manipulate BAs composition, including conjugated and secondary BAs, to alleviate UC symptoms, underscoring the benefits of these approaches for gut health.

Intestinal persistence of Bifidobacterium infantis is determined by interaction of host genetics and antibiotic exposure.

Probiotics have gained significant attention as a potential strategy to improve health by modulating host-microbe interactions, particularly in situations where the normal microbiota has been disrupted. However, evidence regarding their efficacy has been inconsistent, with considerable inter-individual variability in response. We aimed to explore whether a common genetic variant that affects the production of mucosal α(1,2)-fucosylated glycans, present in around 20% of the population, could explain the observed interpersonal differences in the persistence of commonly used probiotics. Using a mouse model with varying α(1,2)-fucosylated glycans secretion (Fut2WT or Fut2KO), we examined the abundance and persistence of Bifidobacterium strains (infantis, breve, and bifidum). We observed significant differences in baseline gut microbiota characteristics between Fut2WT and Fut2KO littermates, with Fut2WT mice exhibiting enrichment of species able to utilise α(1,2)-fucosylated glycans. Following antibiotic exposure, only Fut2WT animals showed persistent engraftment of Bifidobacterium infantis, a strain able to internalise α(1,2)-fucosylated glycans, whereas B. breve and Bifidobacterium bifidum, which cannot internalise α(1,2)-fucosylated glycans, did not exhibit this difference. In mice with an intact commensal microbiota, the relationship between secretor status and B. infantis persistence was reversed, with Fut2KO animals showing greater persistence compared to Fut2WT. Our findings suggest that the interplay between a common genetic variation and antibiotic exposure plays a crucial role in determining the dynamics of B. infantis in the recipient gut, which could potentially contribute to the observed variation in response to this commonly used probiotic species.

Vaccination with an HIV T-Cell Immunogen (HTI) Using DNA Primes Followed by a ChAdOx1-MVA Boost Is Immunogenic in Gut Microbiota-Depleted Mice despite Low IL-22 Serum Levels.

Despite the important role of gut microbiota in the maturation of the immune system, little is known about its impact on the development of T-cell responses to vaccination. Here, we immunized C57BL/6 mice with a prime-boost regimen using DNA plasmid, the Chimpanzee Adenovirus, and the modified Vaccinia Ankara virus expressing a candidate HIV T-cell immunogen and compared the T-cell responses between individuals with an intact or antibiotic-depleted microbiota. Overall, the depletion of the gut microbiota did not result in significant differences in the magnitude or breadth of the immunogen-specific IFNγ T-cell response after vaccination. However, we observed marked changes in the serum levels of four cytokines after vaccinating microbiota-depleted animals, particularly a significant reduction in IL-22 levels. Interestingly, the level of IL-22 in serum correlated with the abundance of Roseburia in the large intestine of mice in the mock and vaccinated groups with intact microbiota. This short-chain fatty acid (SCFA)-producing bacterium was significantly reduced in the vaccinated, microbiota-depleted group. Therefore, our results indicate that, although microbiota depletion reduces serum levels of IL-22, the powerful vaccine regime used could have overcome the impact of microbiota depletion on IFNγ-producing T-cell responses.

Effect of two shampoo formulations on the prokaryotic and eukaryotic microbiota composition of the human scalp.

The human scalp is characterized by a moderately diverse microbial community, comprising prokaryotic (bacteria) and eukaryotic (fungi) members. Although the details are far from being fully understood, the human scalp microbiota is implicated in several scalp disorders, in particular dandruff formation. Hence, the protection of an intact and diverse scalp microbiota can be regarded as a quality criterion for hair and scalp care formulations. In this study, we investigated the influence of two commercially available, non-antimicrobial shampoo formulations on the structure of the scalp microbiota. Scalp microbiota samples, obtained by swab sampling from two cohorts of probands (n = 25, each), were analysed before and after daily use of two different shampoo formulations for 2 weeks, respectively. A polyphasic approach was used, comprising quantitative cultivation of bacteria and fungi on selective media as well as sequencing of PCR-amplified 16S rRNA and 18S rRNA genes, respectively. All analyses revealed a microbiota composition typical for the human scalp. While in particular fungal germ numbers increased significantly during the treatments, overall bacterial and fungal community composition was not affected, based on alpha- and beta-diversity measures. However, we observed an increase in structural bacterial diversity with the age of the probands. Over an application period of 2 weeks, the investigated shampoo induced quantitative but no qualitative changes in the scalp microbial community structure of the investigated probands, suggesting no adverse but rather preserving or even stimulating effects of the underlying formulations on the scalp microbiota. Further investigation will have to clarify if this is also true for longer application periods and if the formulations might affect community functionality, for example microbial gene expression, rather than community composition.

MHCII+CD80+ thymic eosinophils increase in abundance during neonatal development in mice and their accumulation is microbiota dependent.

Eosinophils are present in the thymus of mammals, yet their function at this site during homeostatic development is unknown. We used flow cytometry to determine the abundance and phenotype of eosinophils (here defined as SSchigh SiglecF+ CD11b+ CD45+ cells) in the thymus of mice during the neonatal period, the later postnatal period, and into adulthood. We show that both the total number of thymic eosinophils and their frequency among leukocytes increase over the first 2 wk of life and that their accumulation in the thymus is dependent on the presence of an intact bacterial microbiota. We report that thymic eosinophils express the interleukin-5 receptor (CD125), CD80, and IDO, and that subsets of thymic eosinophils express CD11c and major histocompatibility complex II (MHCII). We found that the frequency of MHCII-expressing thymic eosinophils increases over the first 2 wk of life, and that during this early-life period the highest frequency of MHCII-expressing thymic eosinophils is located in the inner medullary region. These data suggest a temporal and microbiota-dependent regulation of eosinophil abundance and functional capabilities in the thymus.

Experimental evidence root‐associated microbes mediate seagrass response to environmental stress

Abstract Below‐ground microbiota play an important role in mediating environmental conditions with important consequences for plant performance. Micro‐organisms involved in plant–soil interactions may be associated with roots or bulk soil; however, the relative influence of these below‐ground microbial assemblages on plant performance is poorly known, particularly for marine plants. We separately manipulated the root and sediment microbial assemblages of the seagrass Zostera muelleri in a fully factorial experiment to determine how these assemblages determined plant response (e.g. growth) to nutrient enrichment, a major stressor in marine systems. Under ambient nutrient conditions, seagrass growth was maintained regardless of root microbial assemblage disruption. Under high nutrient stress, however, seagrasses with disrupted root microbiota had reduced growth, whereas growth was maintained in seagrasses with an intact root microbiota. Disruption of bulk‐sediment microbiota did not affect seagrass growth. Nutrient elevation was correlated with enhanced abundances of several putatively beneficial microbial taxa (e.g. sulphide‐oxidising Beggiatoaceae and denitrifying Geofilum rubicundum) associated with roots. Synthesis. Our results suggest that under ambient nutrient conditions micro‐organisms play a reduced role in influencing plant performance, but under more stressful conditions positive plant–root micro‐organism interactions strengthened. These results are among the first to experimentally determine that interactions between marine plants and the root‐associated microbiota are key drivers of seagrass performance under human‐induced environmental changes. This suggests that as in terrestrial systems, marine plant resilience depends on the stress‐mitigating functions of their root‐associated microbiota and disturbance to those plant–microbiota interactions can be deleterious for plant performance. Improving our understanding of these plant–micro‐organism interactions may be critical for understanding the functioning and resilience of threatened marine plants and developing more effective restoration strategies for them.

A steamed broccoli sprout diet preparation that reduces colitis via the gut microbiota

Sulforaphane is a bioactive metabolite with anti-inflammatory activity and is derived from the glucosinolate glucoraphanin, which is highly abundant in broccoli sprouts. However, due to its inherent instability its use as a therapeutic against inflammatory diseases has been limited. There are few studies to investigate a whole food approach to increase sulforaphane levels with therapeutic effect and reduce inflammation. In the current study, using a mouse model of inflammatory bowel disease, we investigated the ability of steamed broccoli sprouts to ameliorate colitis and the role of the gut microbiota in mediating any effects. We observed that despite inactivation of the plant myrosinase enzyme responsible for the generation of sulforaphane via steaming, measurable levels of sulforaphane were detectable in the colon tissue and feces of mice after ingestion of steamed broccoli sprouts. In addition, this preparation of broccoli sprouts was also capable of reducing chemically-induced colitis. This protective effect was dependent on the presence of an intact microbiota, highlighting an important role for the gut microbiota in the metabolism of cruciferous vegetables to generate bioactive metabolites and promote their anti-inflammatory effects.

Androgens can modulate immunoregulation directly through gut cells and indirectly through alteration of microbiota composition/function in a lupus mouse model.

Abstract Hormones and gut microbiota affect lupus. In female NZBxNZW1(BWF1) mice, androgen treatment abrogates lupus; castration/treatment with androgen receptor blocker (flutamide) induces disease in BWF1 male mice. Gut microbiota alterations have been reported in lupus. Our goal is to understand the androgen/gut microbiota relationship, and its affect on lupus. We found male castration decreases peripheral, not thymic, derived Tregs; this correlates with decreased gut CD103 dendritic cells (CD103DC) function (induce Treg conversion) and RALDH2 [required for retinoic acid (RA) production) expression. CD103DC have very low androgen receptor (AR) expression, thus unlikely to be affected by androgens. We found that lamina propria stromal cells (LPSC), a major source of RA (via RALDH1/2 expression) required for normal gut CD103DC development, express very high AR levels. Castration/flutamide treatment in male mice decreased RALDH1/2 expression in LPSC, suggesting androgens may affect CD103DC function through LPSC. Castration/flutamide treatment also altered gut microbiota composition. Interestingly, intact male microbiota transfer to castrated male mice restored RALDH2 expression and CD103DC function, suggesting male microbiota factors may also alter CD103DC function. Metabolomic analysis of castrated male feces showed reduced phytanic acid (PA) levels. Administration of PA in vitro/in vivo restored castrated CD103DC function. These data strongly suggest that androgens alter (1) immunoregulatory function through gut cells and (2) gut microbiota composition/function that in turn modulate immunoregulatory function, and reveal how androgens may influence lupus and provide the basis for effective therapy development. Funded by a TIL grant from the Alliance for Lupus Research and the NIH (R01AR067188)

Antimicrobial Susceptibility of Microbiota in Bacterial Vaginosis Using Fluorescence In Situ Hybridization.

Background: Testing of antibiotic resistance of intact vaginal microbiota in pure culture is not feasible. METHODS: Metronidazole, antiseptic octenisept®, antimycotic ciclopirox, bacterial probiotic Lactobacillus crispatus, yeast probiotic Saccharomyces boulardii, Gardnerella-phage-endolysin named phagolysin and phagolysin in combination with probiotics were tested for bacteriolytic activity. Included were vaginal swabs from 38 random women with Amsel-confirmed bacterial vaginosis (BV). Test aliquots were incubated by 37° for 2 and 24 h. Gardnerella, low G+C, Atopobium, lactobacilli, Lactobacillus iners and crispatus, Prevotella-Bacteroides, and Gammaproteobacteria microbial groups were quantified using fluorescence in situ hybridization (FISH). Results: The probiotic strain Lactobacillus crispatus demonstrated the weakest bacteriolytical effects, followed by metronidazole. Both had no impact on Gardnerella species, instead lysing Prevotella-Bacteroides, Enterobacteriaceae (by L.crispatus) or LGC, Atopobium and Prevotella-Bacteroides (by metronidazole) groups of the microbiota. Cytolytic activity on Gardnerella was highly pronounced and increased from octenisept to ciclopirox, phagolysin, phagolysin with L.crispatus, being best in the combination of phagolysin with S.boulardii. Universally active ciclopirox and octenisept® suppressed nearly all microbial groups including those which are regarded as beneficial. Phagolysin had no effect on naturally occurring Lactobacillus crispatus. Conclusions: FISH susceptibility testing allows unique efficacy evaluation of individually adjusted topical therapy without microbial isolation facilitating optimal therapy choice.

A transmissible γδ intraepithelial lymphocyte hyperproliferative phenotype is associated with the intestinal microbiota and confers protection against acute infection.

Intraepithelial lymphocytes expressing the γδ T cell receptor (γδ IELs) serve as a first line of defense against luminal microbes. Although the presence of an intact microbiota is dispensable for γδ IEL development, several microbial factors contribute to the maintenance of this sentinel population. However, whether specific commensals influence population of the γδ IEL compartment under homeostatic conditions has yet to be determined. We identified a novel γδ IEL hyperproliferative phenotype that arises early in life and is characterized by expansion of multiple Vγ subsets. Horizontal transfer of this hyperproliferative phenotype to mice harboring a phenotypically normal γδ IEL compartment was prevented following antibiotic treatment, thus demonstrating that the microbiota is both necessary and sufficient for the observed increase in γδ IELs. Further, we identified two guilds of small intestinal or fecal bacteria represented by 12 amplicon sequence variants (ASV) that are strongly associated with γδ IEL expansion. Using intravital microscopy, we find that hyperproliferative γδ IELs also exhibit increased migratory behavior leading to enhanced protection against bacterial infection. These findings reveal that transfer of a specific group of commensals can regulate γδ IEL homeostasis and immune surveillance, which may provide a novel means to reinforce the epithelial barrier.

Psychological stress disrupts intestinal epithelial cell function and mucosal integrity through microbe and host-directed processes

ABSTRACT Psychological stress alters the gut microbiota and predisposes individuals to increased risk for enteric infections and chronic bowel conditions. Intestinal epithelial cells (IECs) are responsible for maintaining homeostatic interactions between the gut microbiota and its host. In this study, we hypothesized that disruption to colonic IECs is a key factor underlying stress-induced disturbances to intestinal homeostasis. Conventionally raised (CONV-R) and germ-free (GF) mice were exposed to a social disruption stressor (Str) to ascertain how stress modifies colonic IECs, the mucosal layer, and the gut microbiota. RNA sequencing of IECs isolated from CONV-R mice revealed a robust pro-inflammatory (Saa1, Il18), pro-oxidative (Duox2, Nos2), and antimicrobial (Reg3b/g) transcriptional profile as a result of Str. This response occurred concomitant to mucus layer thinning and signs of microbial translocation. In contrast to their CONV-R counterparts, IECs from GF mice or mice treated with broad spectrum antibiotics exhibited no detectable transcriptional changes in response to Str. Nevertheless, IECs from Str-exposed GF mice exhibited an altered response to ex vivo bacterial challenge (increased dual Oxidase-2 [Duox2] and nitric oxide synthase-2 (Nos2)), indicating that STR primes host IEC pro-oxidative responses. In CONV-R mice stress-induced increases in colonic Duox2 and Nos2 (ROS generating enzymes) strongly paralleled changes to microbiome composition and function, evidencing Str-mediated ROS production as a primary factor mediating gut-microbiota dysbiosis. In conclusion, a mouse model of social stress disrupts colonic epithelial and mucosal integrity, a response dependent on an intact microbiota and host stress signals. Together these preclinical findings may provide new insight into mechanisms of stress-associated bowel pathologies in humans.

Psychological Stress Disrupts Intestinal Epithelial Cell Function and Mucosal Integrity Through Microbe and Host-Directed Processes

Background: Psychological stress alters the gut microbiota and predisposes individuals to increased risk for enteric infections and chronic bowel conditions. Intestinal epithelial cells (IECs) are responsible for maintaining homeostatic interactions between the gut microbiota and its host. In this study, we hypothesized that disruption to colonic IECs is a key factor underlying stress-induced disturbances to intestinal homeostasis. Methods: Conventionally-raised (CONV-R) and germ-free (GF) mice were exposed to a social disruption stressor (Str) to ascertain how stress modifies colonic IECs, the mucosal layer, and the gut microbiota. Findings: RNA sequencing of IECs isolated from conventionally-reared (CONV-R) mice revealed a robust pro-inflammatory (Saa1, Il18), pro-oxidative (Duox2, Nos2), and antimicrobial (Reg3g, Reg3b) transcriptional profile as a result of Str. This response occurred concomitant to thinning of the mucus layer and signs of systemic endotoxemia. In contrast to their CONV-R counterparts, IECs from GF mice exhibited no detectable transcriptional changes in response to Str. Still, IECs from Str-exposed GF mice exhibited an altered response to ex vivo bacterial challenge (increased Dual Oxidase-2 [DUOX2]), indicating a priming event by host stress signals. Stress-induced increase in colonic DUOX2 in CONV-R mice paralleled changes to catalase positive bacteria and luminal catalase activity, evidencing Str-mediated disruptions in gut microbiota function through excessive IEC ROS production. Interpretation: Social stress disrupts colonic epithelial cells and mucosal integrity, a response dependent on an intact microbiota and endogenous stress signal priming. Funding: This work was supported by a Ruth L. Kirschstein National Research Service T32DE014320 to J. Allen, NIH grant AT006552-01A1 to M. Bailey, and internal funding from the Abigail Wexner Research Institute at Nationwide Children’s Hospital to J. Allen and M. Bailey. Declaration of Interest: The authors report no competing interests. Ethical Approval: All animals used in this study were approved by the IACUC at Nationwide Children’s Hospital.

Gut Microbiota Composition Affects Procyanidin A2-Attenuated Atherosclerosis in ApoE-/- Mice by Modulating the Bioavailability of Its Microbial Metabolites.

Procyanidin A2 (PCA2) has been shown to improve lipid metabolism. However, it remains to know whether it can play a role in preventing atherosclerosis (AS) through gut microbiota. This study examined the effect of PCA2 on high fat diet (HFD)-induced AS in ApoE-/- mice with an intact and antibiotic-depleted microbiota. PCA2 administration for 12 weeks attenuated HFD-induced AS in ApoE-/- mice, evidenced by obviously alleviating the histological abnormalities of the aorta, lipid accumulation, oxidative stress, and inflammation, which were accompanied by downregulating the expression of vascular cell adhesion molecule-1 and intracellular adhesion molecule-1 and upregulating peroxisome proliferator-activated receptor gamma, cholesterol 7 alpha-hydroxylase, and ATP-binding cassette transporter A1. Moreover, PCA2 treatment reshaped the gut microbiota imbalance caused by HFD, especially reducing the ratio of Firmicutes/Bacteroidetes and increasing the abundance of Verrucomicrobia. However, antibiotic intervention almost offset the alleviation of AS by PCA2 and prevented the biotransformation of PCA2 by gut microbiota, thus resulting in a 2327.21-6.27-fold decrease in its microbial metabolites of plasma. There was a marked correlation among the microbiota composition, the bioavailability of PCA2-derived microbial metabolites, and AS indicators. The findings indicate that the gut microbiota robustly influences the bioavailability of microbial metabolites that may partially drive the AS resilience property of PCA2.

Improvement of Gut Barrier Function by Potato Anthocyanins Is Dependent on Gut Microbiota

Abstract Objectives Ulcerative colitis (UC) is characterized by chronic colonic inflammation, impaired barrier function and gut bacterial dysbiosis. Anthocyanin-containing potatoes have been shown to maintain the intestinal barrier function in colitic mice. However, the role of gut microbiota in the anti-colitic effects of anthocyanin-containing potatoes is not clear. This study evaluated the gut barrier protective efficacy of purple- and red-fleshed potatoes using a DSS-induced murine model of colitis with the intact and antibiotic-depleted microbiota. Methods Four-week-old mice (C57BL6) were randomized into four groups, receiving a standard control diet, or a 20% purple-/red-fleshed potato (PP/RP) supplemented diet. A broad-spectrum antibiotic cocktail was used to deplete the gut bacteria. After eight weeks, mice were treated with 2% DSS in their drinking water for five days to induce colitis. The polyphenol composition of potatoes was analyzed by LC-MS/MS. Intestinal permeability was measured using FITC-dextran. Colonic myeloperoxidase (MPO) activities were determined using the o-dianisidine dihydrochloride method. The fecal short-chain fatty acids (SCFA) content of mice was measured by GC-FID. RT-PCR was used to analyze the relative gene expression levels of cytokines and bacterial abundance. Results The predominant anthocyanin detected in RP was pelargonidin (93.13%). Whereas petunidin (75.57%) and malvidin (21.05%) were the two major anthocyanins present in PP. Administration of antibiotics resulted in a reduction in fecal SCFA. Antibiotic-treated mice were more prone to the DSS-induced increase in gut permeability. Anthocyanin-containing potato diets alleviated DSS-induced colonic damage, elevation in histology score and gut permeability only in non-antibiotic mice. RP alone suppressed the elevated colonic MPO activity and the gene expression levels of MUC2 in mice with intact gut microbiota. Conclusions In summary, anthocyanin-containing potatoes alleviated colitis-associated colonic epithelial damage and increased gut permeability in intact microbiota mice but not in microbiota-ablated mice. This suggests that the gut microbiome is essential for the gut barrier protective activity of anthocyanin-containing potatoes. Funding Sources USDA-NIFA awards 2016–67,017-29,285 and 2019–67,017-29,258.

The effect of intestinal microbiota dysbiosis on growth and detection of carbapenemase-producing Enterobacterales within an in vitro gut model

Carbapenemase-producing Enterobacterales (CPE) can colonize the gut and are of major clinical concern. Identification of CPE colonization is problematic; there is no gold-standard detection method, and the effects of antibiotic exposure and microbiota dysbiosis on detection are unknown. Based on a national survey we selected four CPE screening assays in common use. We used a clinically reflective invitro model of human gut microbiota to investigate the performance of each test to detect three different CPE strains under different, clinically relevant antibiotic exposures. Twelve gut models were seeded with a pooled faecal slurry and exposed to CPE either before, after, concomitant with, or in the absence of piperacillin-tazobactam (358 mg/L, 3× daily, seven days). Total Enterobacterales and CPE populations were enumerated daily. Regular screening for CPE was performed using Cepheid Xpert® Carba-R molecular test, and with Brilliance™ CRE, Colorex™ mSuperCARBA and CHROMID® CARBA SMART agars. Detection of CPE when the microbiota are intact is problematic. Antibiotic exposure disrupts microbiota populations and allows CPE proliferation, increasing detection. The performances of assays varied, particularly with respect to different CPE strains. The Cepheid assay performed better than the three agar methods for detecting a low level of CPE within an intact microbiota, although performance of all screening methods was comparable when CPE populations increased in a disrupted microbiota. CPE strains differed in their dynamics of colonization in an invitro gut model and in their subsequent response to antibiotic exposure. This affected detection by molecular and screening methods, which has implications for the sensitivity of CPE screening in healthcare settings.

A respiratory commensal bacterium acts as a risk factor for Mycoplasma gallisepticum infection in chickens

Commensal microbiota has been shown to play an important role in local infections. However, the correlation between host respiratory microbiota and Mycoplasma gallisepticum (MG) infection is not well characterized. Here, the results of 16S rRNA sequencing showed that MG infection correlated with alteration in respiratory microbiota of chickens characterized by decreased richness and diversity. To explore whether respiratory microbiota contributed to MG infection, an antibiotics cocktail was used to deplete respiratory microbiota. It has been found that depletion of respiratory Gram-positive and Gram-negative bacteria promoted MG infection, as reflected in the form of increased MG colonization, pro-inflammatory cytokines and proteins expression, and severe lung damage compared to the control group. Importantly, depletion of Gram-negative bacteria in respiratory tract mitigated MG infection, which indicated that certain Gram-negative bacteria may promote MG infection. By reconstitution of individual cultivable respiratory tract bacteria in antibiotic-treated chickens, a respiratory commensal microbe Serratia marcescens was identified to facilitate MG infection. We further found that Serratia marcescens may promote MG infection by downregulating Mucin 2 (MUC2) and tight junction related gene mRNA expression levels in trachea and lung tissues. Together, our data demonstrated that MG infection induced disturbed respiratory microbiota and the specific respiratory commensal bacterium Serratia marcescens could promote MG infection, and thus expand our understanding of the pathogenesis of MG infection.