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Multiple sclerosis (MS) is a complex disease of the CNS thought to require an environmental trigger. Gut dysbiosis is common in MS, but specific causative species are unknown. To address this knowledge gap, we used sensitive and quantitative PCR detection to show that people with MS were more likely to harbor and show a greater abundance of epsilon toxin- producing (ETX-producing) strains of C. perfringens within their gut microbiomes compared with individuals who are healthy controls (HCs). Isolates derived from patients with MS produced functional ETX and had a genetic architecture typical of highly conjugative plasmids. In the active immunization model of experimental autoimmune encephalomyelitis (EAE), where pertussis toxin (PTX) is used to overcome CNS immune privilege, ETX can substitute for PTX. In contrast to PTX- induced EAE, where inflammatory demyelination is largely restricted to the spinal cord, ETX-induced EAE caused demyelination in the corpus callosum, thalamus, cerebellum, brainstem, and spinal cord, more akin to the neuroanatomical lesion distribution seen in MS. CNS endothelial cell transcriptional profiles revealed ETX-induced genes that are known to play a role in overcoming CNS immune privilege. Together, these findings suggest that ETX-producing C. perfringens strains are biologically plausible pathogens in MS that trigger inflammatory demyelination in the context of circulating myelin autoreactive lymphocytes., Introduction Multiple sclerosis (MS) lesions are centered on postcapillary venules (1, 2), and impairment of blood-brain barrier (BBB) function is considered the earliest event in lesion evolution, paving the way [...]

Fusobacterium nucleatum (Fn) and enterotoxigenic Bacteroides fragilis (ETBF) are two pathobionts consistently enriched in the gut microbiomes of patients with colorectal cancer (CRC) compared to healthy counterparts and frequently observed for their direct association within tumors. Although several molecular mechanisms have been identified that directly link these organisms to features of CRC in specific cell types, their specific effects on the epithelium and local immune compartment are not well-understood. To fill this gap, we leveraged single-cell RNA sequencing (scRNA-seq) on wildtype mice and mouse model of CRC. We find that Fn and ETBF exacerbate cancer-like transcriptional phenotypes in transit-amplifying and mature enterocytes in a mouse model of CRC. We also observed increased T cells in the pathobiont-exposed mice, but these pathobiont-specific differences observed in wildtype mice were abrogated in the mouse model of CRC. Although there are similarities in the responses provoked by each organism, we find pathobiont-specific effects in Myc-signaling and fatty acid metabolism. These findings support a role for Fn and ETBF in potentiating tumorigenesis via the induction of a cancer stem cell-like transit-amplifying and enterocyte population and the disruption of CTL cytotoxic function. [ABSTRACT FROM AUTHOR]

Human microbiome engineering is increasingly proposed as a way to modulate health outcomes. However, one of the current limitations to engineering microbial communities in situ is delivery of a genetic payload for introducing or modifying genes. Indeed, there is a need to identify novel broad-host delivery vectors for microbiome engineering. Therefore, in this study, we characterized conjugative plasmids from a publicly available dataset of antibiotic-resistant isolate genomes in order to identify potential broad-host vectors for further applications. From the 199 closed genomes available in the CDC & FDA AR Isolate Bank, we identified 439 plasmids, of which 126 were predicted to be mobilizable and 206 conjugative. Various characteristics of the conjugative plasmids, such as size, replication origin, conjugation machinery, host defense mechanisms, and plasmid stability proteins, were analyzed to determine these plasmids' potential host-range. Following this analysis, we clustered plasmid sequences and chose 22 unique, broad-host range plasmids that would be suitable for use as delivery vectors. This novel set of plasmids will provide a valuable resource for engineering microbial communities. [ABSTRACT FROM AUTHOR]

The protein interaction network involved in suppressing homologous recombination within the rDNA locus is analyzed using affinity purification combined with highly sensitive mixture mass spectrometry. A model is suggested in which regulator of nucleolar silencing and telophase exit [RENT], Tof2 and Lrs4/Csm1 physically clamp rDNA to the cohesin ring, thereby restricting the movement of rDNA sister chromatids relative to each other to inhibit unequal exchange.

Osteoarthritis is increasingly viewed as a heterogeneous disease with multiple phenotypic subgroups. Obesity enhances joint degeneration in mouse models of posttraumatic osteoarthritis (PTOA). Most models of PTOA involve damage to surrounding tissues caused by surgery/fracture; it is unclear if obesity enhances cartilage degeneration in the absence of surgery/fracture. We used a nonsurgical animal model of load‐induced PTOA to determine the effect of obesity on cartilage degeneration 2 weeks after loading. Cartilage degeneration was caused by a single bout of cyclic tibial loading at either a high or moderate load magnitude in adult male mice with severe obesity (C57Bl6/J + high‐fat diet), mild obesity (toll‐like receptor 5 deficient mouse [TLR5KO]), or normal adiposity (C57Bl6/J mice + normal diet and TLR5KO mice in which obesity was prevented by manipulation of the gut microbiome). Two weeks after loading, cartilage degeneration occurred in limbs loaded at a high magnitude, as determined by OARSI scores (P <.001). However, the severity of cartilage damage did not differ among groups. Osteophyte width and synovitis of loaded limbs did not differ among groups. Furthermore, obesity did not enhance cartilage damage in limbs evaluated 6 weeks after loading. Constituents of the gut microbiota differed among groups. Our findings suggest that, in the absence of surgery/fracture, obesity may not influence cartilage loss after a single mechanical insult, suggesting that either damage to surrounding tissues or repeated mechanical insult is necessary for obesity to influence cartilage degeneration. These findings further illustrate heterogeneity in PTOA phenotypes and complex interactions between mechanical/metabolic factors in cartilage loss. [ABSTRACT FROM AUTHOR]

Bactericidal antibiotics are powerful agents due to their ability to convert essential bacterial functions into lethal processes. However, many important bacterial pathogens are remarkably tolerant against bactericidal antibiotics due to inducible damage repair responses. The cell wall damage response two‐component system VxrAB of the gastrointestinal pathogen Vibrio cholerae promotes high‐level β‐lactam tolerance and controls a gene network encoding highly diverse functions, including negative control over multiple iron uptake systems. How this system contributes to tolerance is poorly understood. Here, we show that β‐lactam antibiotics cause an increase in intracellular free iron levels and collateral oxidative damage, which is exacerbated in the ∆vxrAB mutant. Mutating major iron uptake systems dramatically increases ∆vxrAB tolerance to β‐lactams. We propose that VxrAB reduces antibiotic‐induced toxic iron and concomitant metabolic perturbations by downregulating iron uptake transporters and show that iron sequestration enhances tolerance against β‐lactam therapy in a mouse model of cholera infection. Our results suggest that a microorganism's ability to counteract diverse antibiotic‐induced stresses promotes high‐level antibiotic tolerance and highlights the complex secondary responses elicited by antibiotics. SYNOPSIS: The cholera pathogen Vibrio cholerae is highly tolerant against β‐lactam antibiotics due to its sensing system VxrAB. Upon sensing antibiotic exposure, VxrAB upregulates cell wall synthesis functions and downregulates iron acquisition systems, which are both required for antibiotics tolerance. Upon beta lactam exposure, V. cholerae experiences iron influx, hydrogen peroxide production and cell wall damage.The two‐component system VxrAB is induced by cell wall acting antibiotics.VxrAB induces cell wall synthesis, iron acquisition systems and other cell envelope functions.Removing VxrAB control over cell wall synthesis only partially reduces tolerance in the wild‐type.Reducing iron influx partially restores β‐lactam tolerance of ΔvxrAB.Therapeutic reduction of in vivo iron levels by iron chelators increases V. cholerae tolerance to a β‐lactam antibiotic in a mouse model. [ABSTRACT FROM AUTHOR]

Shotgun metagenomic sequencing has revolutionized our ability to detect and characterize the diversity and function of complex microbial communities. In this review, we highlight the benefits of using metagenomics as well as the breadth of conclusions that can be made using currently available analytical tools, such as greater resolution of species and strains across phyla and functional content, while highlighting challenges of metagenomic data analysis. Major challenges remain in annotating function, given the dearth of functional databases for environmental bacteria compared to model organisms, and the technical difficulties of metagenome assembly and phasing in heterogeneous environmental samples. In the future, improvements and innovation in technology and methodology will lead to lowered costs. Data integration using multiple technological platforms will lead to a better understanding of how to harness metagenomes. Subsequently, we will be able not only to characterize complex microbiomes but also to manipulate communities to achieve prosperous outcomes for health, agriculture, and environmental sustainability. [ABSTRACT FROM AUTHOR]

Background: Periprosthetic joint infection (PJI) is one of the most devastating complications of total joint arthroplasty. Given the mortality and morbidity associated with PJI and the challenges in treating it, there has been increased interest in risk factors that can be modified before surgery. In this study, we used a novel mouse model to consider the role of the gut microbiome as a risk factor for PJI.Questions/purposes: (1) Does the state of the gut microbiota before surgery influence the likelihood of developing an established infection in a mouse model of PJI? (2) How does the state of the gut microbiota before surgery influence the local and systemic response to the presence of an established infection in a mouse model of PJI?Methods: Male C57Bl/6 mice were divided into two groups: those with modified microbiome [INCREMENT]microbiome (n = 40) and untreated mice (n = 42). In [INCREMENT]microbiome mice, the gut flora were modified using oral neomycin and ampicillin from 4 weeks to 16 weeks of age. Mice received a titanium tibial implant to mimic a joint implant and a local inoculation of Staphylococcus aureus in the synovial space (10 colony forming units [CFUs]). The proportion of animals developing an established infection in each group was determined by CFU count. The local and systemic response to established infection was determined using CFU counts in surrounding joint tissues, analysis of gait, radiographs, body weight, serum markers of inflammation, and immune cell profiles and was compared with animals that received the inoculation but resisted infection.Results: A greater proportion of animals with disrupted gut microbiota had infection (29 of 40 [73%]) than did untreated animals (21 of 42 [50%]; odds ratio, 2.63, 95% CI, 1.04-6.61; p = 0.035). The immune response to established infection in mice with altered microbiota was muted; serum amyloid A, a marker of systemic infection in mice, was greater than in mice with disrupted gut microbiota with infection (689 µg/dL; range, 68-2437 µg/dL, p < 0.05); infection associated increases in monocytes and neutrophils in the spleen and local lymph node in untreated mice but not were not observed in mice with disrupted gut microbiota.Conclusions: The findings from this in vivo mouse model suggest that the gut microbiota may influence susceptibility to PJI.Clinical Relevance: These preclinical findings support the idea that the state of the gut microbiome before surgery may influence the development of PJI and justify further preclinical and clinical studies to develop appropriate microbiome-based interventions. [ABSTRACT FROM AUTHOR]

Transmission usually refers to the movement of pathogenic organisms. Yet, commensal microbes that inhabit the human body also move between individuals and environments. Surprisingly little is known about the transmission of these endogenous microbes, despite increasing realizations of their importance for human health. The health impacts arising from the transmission of commensal bacteria range widely, from the prevention of autoimmune disorders to the spread of antibiotic resistance genes. Despite this importance, there are outstanding basic questions: what is the fraction of the microbiome that is transmissible? What are the primary mechanisms of transmission? Which organisms are the most highly transmissible? Higher resolution genomic data is required to accurately link microbial sources (such as environmental reservoirs or other individuals) with sinks (such as a single person's microbiome). New computational advances enable strain-level resolution of organisms from shotgun metagenomic data, allowing the transmission of strains to be followed over time and after discrete exposure events. Here, we highlight the latest techniques that reveal strain-level resolution from raw metagenomic reads and new studies that are tracking strains across people and environments. We also propose how models of pathogenic transmission may be applied to study the movement of commensals between microbial communities. [ABSTRACT FROM AUTHOR]

Infectious diseases are economically detrimental to aquaculture, and with continued expansion and intensification of aquaculture, the importance of managing infectious diseases will likely increase in the future. Here, we use evolution of virulence theory, along with examples, to identify aquaculture practices that might lead to the evolution of increased pathogen virulence. We identify eight practices common in aquaculture that theory predicts may favor evolution toward higher pathogen virulence. Four are related to intensive aquaculture operations, and four others are related specifically to infectious disease control. Our intention is to make aquaculture managers aware of these risks, such that with increased vigilance, they might be able to detect and prevent the emergence and spread of increasingly troublesome pathogen strains in the future. [ABSTRACT FROM AUTHOR]

SNP-level analysis of 5,000+ metagenomic samples enabled Hildebrand and colleagues (2021) to identify human gut microbiota by their dispersal strategies in this issue of Cell Host & Microbe. This brings us a step closer in understanding how microbial communities are formed and maintained and also hints at ways in which microbiomes can be manipulated to improve human health. [ABSTRACT FROM AUTHOR]

Protein–protein interactions between the microbiome and host organism play an important role in shaping host health. These host-modulating proteins have therapeutic potential in treating microbiome-linked disorders such as inflammatory bowel disease and obesity. Structural analysis of interacting proteins provides highly mechanistic insight into the domains driving these interactions and the resulting influence on host cell processes. Here, we briefly review recent publication of microbiome protein structures involved in host binding interactions, the effects of these interactions on host physiology, and the need for further study to increase the ability to detect proteins with therapeutic potential. • Structural analysis of proteins from probiotic and commensal bacteria reveals host-binding functions. • Proteins from these beneficial microbes modulate host physiology independently of the whole bacterium, indicating therapeutic potential. • Further structures of microbiome proteins will improve identification of host-interacting functions in large datasets. [ABSTRACT FROM AUTHOR]

Many microbial communities are characterized by high genetic diversity. 16S ribosomal RNA sequencing can determine community members, and metagenomics can determine the functional diversity, but resolving the functional role of individual cells in high throughput remains an unsolved challenge. Here, we describe epicPCR (Emulsion, Paired Isolation and Concatenation PCR), a new technique that links functional genes and phylogenetic markers in uncultured single cells, providing a throughput of hundreds of thousands of cells with costs comparable to one genomic library preparation. We demonstrate the utility of our technique in a natural environment by profiling a sulfate-reducing community in a freshwater lake, revealing both known sulfate reducers and discovering new putative sulfate reducers. Our method is adaptable to any conserved genetic trait and translates genetic associations from diverse microbial samples into a sequencing library that answers targeted ecological questions. Potential applications include identifying functional community members, tracing horizontal gene transfer networks and mapping ecological interactions between microbial cells.

Changes in neuronal activity modify the structure of dendritic spines and alter the function and protein composition of synapses. Regulated degradation of postsynaptic density (PSD) proteins by the ubiquitin-proteasome system is believed to play an important role in activity-dependent synaptic remodeling. Stimulating neuronal activity in vitro and in vivo induces the ubiquitination and degradation of GKAP/SAPAP and Shank, major scaffold proteins of the PSD. However, the specific ubiquitin ligases that regulate postsynaptic protein composition have not been identified. Here we identify the RING fingercontaining protein TRIM3 as a specific E3 ubiquitin ligase for the PSD scaffold GKAP/SAPAP1. Present in PSD fractions from rat brain, TRIM3 stimulates ubiquitination and proteasome-dependent degradation of GKAP, and induces the loss of GKAP and associated scaffold Shank1 from postsynaptic sites. Suppression of endogenous TRIM3 by RNA interference (RNAi) results in increased accumulation of GKAP and Shank1 at synapses, as well as enlargement of dendritic spine heads. RNAi of TRIM3 also prevented the loss of GKAP induced by synaptic activity. Thus, TRIM3 is a novel E3 ligase that mediates activity-dependent turnover of PSD scaffold proteins and is a negative regulator of dendritic spine morphology. [ABSTRACT FROM AUTHOR]

Klebsiella pneumoniae is a leading cause of severe infections in humans and dairy cows and is rapidly becoming untreatable due to the emergence of multidrug-resistant (MDR) strains. However, little is known about the relationship between bovine and human K. pneumoniae isolates at the genome population level. Here, we investigated the genomic structures, pan-genomic profiles, virulence determinants, and resistomes of 308 K. pneumoniae isolates from humans and dairy cows including 96 newly sequenced cow isolates. We identified 177 functional protein families that were significantly different across human and bovine isolates; genes expressing proteins related to metal ion metabolism (iron, zinc, and calcium) were significantly more prevalent among the bovine isolates. Siderophore systems were found prevalent in both the bovine and human isolates. In addition, we found that the Klebsiella ferric uptake operon kfuABC was significantly more prevalent in clinical mastitis cases compared with healthy cows. Furthermore, in two dairy farms we identified a unique IncN-type plasmid, pC5, co-harboring blaCTX-M-1 and mph(A) genes, which confers resistance to cephalosporins and macrolides, respectively. We provide here the complete annotated sequence of this plasmid. [ABSTRACT FROM AUTHOR]