Search

Mucin glycoproteins are a significant source of carbon for the gut bacteria. Various gut microbial species possess diverse hydrolytic enzymes and catabolic pathways for breaking down mucin glycans, resulting in competition for the limited nutrients within the gut environment. Adherence to mucin glycans represents a crucial strategy used by gut microbes to access nutrient reservoirs. Understanding these properties is pivotal for comprehending the survival mechanisms of bacteria in the gastrointestinal tract. However, characterization of individual strains within the vast array of coexisting bacteria in the microbiome is challenging. To investigate this, we developed mucin-immobilized particles by immobilizing porcine gastric mucin (PGM) onto glass beads chemically modified with boronic acid. These PGM-immobilized particles were then anaerobically cultured with human fecal microbiota, and the bacteria adhering to PGM were isolated. Interestingly, the microbiome composition remained largely unchanged irrespective of PGM immobilization. Nonetheless, bacteria isolated from PGM-immobilized glass particles exhibited notably higher N-acetylgalactosaminidase activity compared to the control beads. Furthermore, Bacteroides strains isolated from PGM-immobilized glass particles displayed enhanced adhesive and metabolic properties to PGM. These findings underscore the utility of PGM particles in enriching and isolating specific microbes. Moreover, they highlight substantial differences in microbial properties at the strain level. We anticipate that PGM-immobilized particles will advance culture-based microbiome research, emphasizing the significance of strain-level characterization. [ABSTRACT FROM AUTHOR]

Introduction: Drought is one of the biggest problems for crop production and also affects the survival and persistence of soil rhizobia, which limits the establishment of efficient symbiosis and endangers the productivity of legumes, the main source of plant protein worldwide. Aim: Since the biodiversity can be altered by several factors including abiotic stresses or cultural practices, the objective of this research was to evaluate the effect of water availability, plant genotype and agricultural management on the presence, nodulation capacity and genotypic diversity of rhizobia. Method: A field experiment was conducted with twelve common bean genotypes under irrigation and rain-fed conditions, both in conventional and organic management. Estimation of the number of viable rhizobia present in soils was performed before the crop establishment, whereas the crop yield, nodule number and the strain diversity of bacteria present in nodules were determined at postharvest. Results: Rainfed conditions reduced the number of nodules and of isolated bacteria and their genetic diversity, although to a lesser extent than the agrochemical inputs related to conventional management. In addition, the effect of water scarcity on the conventional management soil was greater than observed under organic conditions. Conclusions: The preservation of diversity will be a key factor to maintain crop production in the future, as problems caused by drought will be exacerbated by climate change and organic management can help to maintain the biodiversity of soil microbiota, a fundamental aspect for soil health and quality. [ABSTRACT FROM AUTHOR]

Donor and recipient human cytomegalovirus (HCMV) seropositive (D+R+) lung transplant recipients (LTRs) often harbor multiple strains of HCMV, likely due to transmitted donor (D) strains and reactivated recipient (R) strains. To date, the extent and timely occurrence of each likely source in shaping the post‐transplantation (post‐Tx) strain population is unknown. Here, we deciphered the D and R origin of the post‐Tx HCMV strain composition in blood, bronchoalveolar lavage (BAL), and CD45+ BAL cell subsets. We investigated either D and/or R formalin‐fixed paraffin‐embedded blocks or fresh D lung tissue from four D+R+ LTRs obtained before transplantation. HCMV strains were characterized by short amplicon deep sequencing. In two LTRs, we show that the transplanted lung is reseeded by R strains within the first 6 months after transplantation, likely by infiltrating CD14+ CD163+/− alveolar macrophages. In three LTRs, we demonstrate both rapid D‐strain dissemination and persistence in the transplanted lung for >1 year post‐Tx. Broad inter‐host diversity contrasts with intra‐host genotype sequence stability upon transmission, during follow‐up and across compartments. In D+R+ LTRs, HCMV strains of both, D and R origin can emerge first and dominate long‐term in subsequent episodes of infection, indicating replication of both sources despite pre‐existing immunity. [ABSTRACT FROM AUTHOR]

Feline calicivirus (FCV) is a significant viral pathogen causing upper respiratory tract and oral diseases in cats. The emergence of the virulent systemic FCV variant (VS-FCV) has raised global concern in the past decade. This study aims to explore the epidemiology, genetic characterization, and diversity of FCV strains circulating among Thai cats. Various sample types, including nasal, oral, and oropharyngeal swabs and fresh tissues, were collected from 184 cats across different regions of Thailand from 2016 to 2021. Using reverse transcription realtime polymerase chain reaction (RT-qPCR), FCV infection was investigated, with additional screening for feline herpesvirus-1 (FHV-1) by qPCR. The detection rates for FCV, FHV-1, and co-infection were 46.7, 65.8, and 31.5%, respectively. Significantly, the odds ratio (OR) revealed a strong association between the detection of a single FCV and the presence of gingivostomatitis lesions (OR: 7.15, 95% CI: 1.89-26.99, p = 0.004). In addition, FCV detection is notably less likely in vaccinated cats (OR: 0.22, 95% CI: 0.07-0.75, p = 0.015). Amino acid sequence analysis based on the VP1 major capsid protein gene of the 14 FCV-Thai (FCV-TH) strains revealed genetic diversity compared to the other 43 global strains (0 to 86.6%). Intriguingly, a vaccine-like FCV variant was detected in one cat. In summary, this study provides insights into the epidemiology and molecular characteristics of FCV diversity within the Thai cat population for the first time. The identification of unique physicochemical characteristics in the capsid hypervariable region of some FCV-TH strains challenges previous hypotheses. Therefore, further exploration of vaccine-like FCV variants is crucial for a comprehensive understanding and to improve viral prevention and control strategies. [ABSTRACT FROM AUTHOR]

ABSTRACT While research on the sourdough microbiome has primarily focused on lactic acid bacteria (LAB) and yeast, recent studies have found that acetic acid bacteria (AAB) are also common members. However, the ecology, genomic diversity, and functional contributions of AAB in sourdough remain unknown. To address this gap, we sequenced 29 AAB genomes, including three that represent putatively novel species, from a collection of over 500 sourdough starters surveyed globally from community scientists. We found variations in metabolic traits related to carbohydrate utilization, nitrogen metabolism, and alcohol production, as well as in genes related to mobile elements and defense mechanisms. Sourdough AAB genomes did not cluster when compared to AAB isolated from other environments, although a subset of gene functions was enriched in sourdough isolates. The lack of a sourdough-specific genomic cluster may reflect the nomadic lifestyle of AAB. To assess the consequences of AAB on the emergent function of sourdough starter microbiomes, we constructed synthetic starter microbiomes, varying only the AAB strain included. All AAB strains increased the acidification of synthetic sourdough starters relative to yeast and LAB by 18.5% on average. Different strains of AAB had distinct effects on the profile of synthetic starter volatiles. Taken together, our results begin to define the ways in which AAB shape emergent properties of sourdough and suggest that differences in gene content resulting from intraspecies diversification can have community-wide consequences on emergent function.IMPORTANCEThis study is a comprehensive genomic and ecological survey of acetic acid bacteria (AAB) isolated from sourdough starters. By combining comparative genomics with manipulative experiments using synthetic microbiomes, we demonstrate that even strains with >97% average nucleotide identity can shift important microbiome functions, underscoring the importance of species and strain diversity in microbial systems. We also demonstrate the utility of sourdough starters as a model system to understand the consequences of genomic diversity at the strain and species level on multispecies communities. These results are also relevant to industrial and home-bakers as we uncover the importance of AAB in shaping properties of sourdough starters that have direct impacts on sensory notes and the quality of sourdough bread.

Abstract Background The rumen microbiome enables ruminants to digest otherwise indigestible feedstuffs, thereby facilitating the production of high-quality protein, albeit with suboptimal efficiency and producing methane. Despite extensive research delineating associations between the rumen microbiome and ruminant production traits, the functional roles of the pervasive and diverse rumen virome remain to be determined. Results Leveraging a recent comprehensive rumen virome database, this study analyzes virus-microbe linkages, at both species and strain levels, across 551 rumen metagenomes, elucidating patterns of microbial and viral diversity, co-occurrence, and virus-microbe interactions. Additionally, this study assesses the potential role of rumen viruses in microbial diversification by analyzing prophages found in rumen metagenome-assembled genomes. Employing CRISPR–Cas spacer-based matching and virus-microbe co-occurrence network analysis, this study suggests that the viruses in the rumen may regulate microbes at strain and community levels through both antagonistic and mutualistic interactions. Moreover, this study establishes that the rumen virome demonstrates responsiveness to dietary shifts and associations with key animal production traits, including feed efficiency, lactation performance, weight gain, and methane emissions. Conclusions These findings provide a substantive framework for further investigations to unravel the functional roles of the virome in the rumen in shaping the microbiome and influencing overall animal production performance. Video Abstract

Background: The rumen microbiome enables ruminants to digest otherwise indigestible feedstuffs, thereby facilitating the production of high-quality protein, albeit with suboptimal efficiency and producing methane. Despite extensive research delineating associations between the rumen microbiome and ruminant production traits, the functional roles of the pervasive and diverse rumen virome remain to be determined. Results: Leveraging a recent comprehensive rumen virome database, this study analyzes virus-microbe linkages, at both species and strain levels, across 551 rumen metagenomes, elucidating patterns of microbial and viral diversity, co-occurrence, and virus-microbe interactions. Additionally, this study assesses the potential role of rumen viruses in microbial diversification by analyzing prophages found in rumen metagenome-assembled genomes. Employing CRISPR–Cas spacer-based matching and virus-microbe co-occurrence network analysis, this study suggests that the viruses in the rumen may regulate microbes at strain and community levels through both antagonistic and mutualistic interactions. Moreover, this study establishes that the rumen virome demonstrates responsiveness to dietary shifts and associations with key animal production traits, including feed efficiency, lactation performance, weight gain, and methane emissions. Conclusions: These findings provide a substantive framework for further investigations to unravel the functional roles of the virome in the rumen in shaping the microbiome and influencing overall animal production performance. 3QAbSJqYZPn2njEBJ23VpA Video Abstract [ABSTRACT FROM AUTHOR]

Once rodents have been successfully eradicated from Lord Howe Island, Australia, the critically endangered Lord Howe Island stick insect (Dryococelus australis (Montrouzier)) may be reintroduced, a century after it was thought to have become extinct. In captive populations of D. australis, elevated mortalities have been associated with bacterial pathogens. To better define the infectious risk posed by entomopathogens to the reintroduction program, we investigated the bacteria isolated from captive D. australis kept at Melbourne Zoo and on Lord Howe Island and from environmental samples and free-living invertebrates collected on various parts of the island. At Melbourne Zoo, Serratia and Pseudomonas spp. were the bacteria most frequently isolated between 2013 and 2019. Serratia spp. were also the organisms most frequently isolated from insects sampled in April 2019 from the captive population on Lord Howe Island. In addition, Serratia spp. were isolated from a range of environmental samples collected on Lord Howe Island during March–April 2019. These environmental isolates had a broader range of biochemical and molecular characteristics than those obtained from the captive insect populations. A large proportion of these isolates were urease positive and had biochemical profiles previously not described for Serratia spp. This study highlights the need for better surveillance for potential pathogens in understudied regions and sites. We conclude that infections caused by Serratia spp. might pose a problem to the captive breeding program for D. australis but that the risk of introducing novel pathogens to Lord Howe Island through infected insects is low. Our study explores some of the potential risks involved in captive breeding and provides a valuable example of using pathogen surveillance to better inform an invertebrate conservation program. [ABSTRACT FROM AUTHOR]

ABSTRACT Fungi were some of the earliest organismal systems used to explore mutational processes and its phenotypic consequences on members of a species. Yeasts that cause significant human disease were quickly incorporated into these investigations to define the genetic and phenotypic drivers of virulence. Among Candida species, Candida albicans has emerged as a model for studying genomic processes of evolution because of its clinical relevance, relatively small genome, and ability to tolerate complex chromosomal changes. Here, we describe major recent findings that used evolution of strains from defined genetic backgrounds to delineate mutational and adaptative processes and include how nascent exploration into naturally occurring variation is contributing to these conceptual frameworks. Ultimately, efforts to discern adaptive mechanisms used by C. albicans will continue to divulge new biology and can better inform treatment regimens for the increasing prevalence of fungal disease.

IntroductionDrought is one of the biggest problems for crop production and also affects the survival and persistence of soil rhizobia, which limits the establishment of efficient symbiosis and endangers the productivity of legumes, the main source of plant protein worldwide.AimSince the biodiversity can be altered by several factors including abiotic stresses or cultural practices, the objective of this research was to evaluate the effect of water availability, plant genotype and agricultural management on the presence, nodulation capacity and genotypic diversity of rhizobia.MethodA field experiment was conducted with twelve common bean genotypes under irrigation and rain-fed conditions, both in conventional and organic management. Estimation of the number of viable rhizobia present in soils was performed before the crop establishment, whereas the crop yield, nodule number and the strain diversity of bacteria present in nodules were determined at postharvest.ResultsRainfed conditions reduced the number of nodules and of isolated bacteria and their genetic diversity, although to a lesser extent than the agrochemical inputs related to conventional management. In addition, the effect of water scarcity on the conventional management soil was greater than observed under organic conditions.ConclusionsThe preservation of diversity will be a key factor to maintain crop production in the future, as problems caused by drought will be exacerbated by climate change and organic management can help to maintain the biodiversity of soil microbiota, a fundamental aspect for soil health and quality.

Abstract Background Strain-level diversity is widespread among bacterial species and can expand the functional potential of natural microbial communities. However, to what extent communities undergo consistent shifts in strain composition in response to environmental/host changes is less well understood. Results Here, we used shotgun metagenomics to compare the gut microbiota of two behavioral states of the Western honeybee (Apis mellifera), namely nurse and forager bees. While their gut microbiota is composed of the same bacterial species, we detect consistent changes in strain-level composition between nurses and foragers. Single nucleotide variant profiles of predominant bacterial species cluster by behavioral state. Moreover, we identify strain-specific gene content related to nutrient utilization, vitamin biosynthesis, and cell–cell interactions specifically associated with the two behavioral states. Conclusions Our findings show that strain-level diversity in host-associated communities can undergo consistent changes in response to host behavioral changes modulating the functional potential of the community.

In the initial stage of respiratory infection, Mycobacterium tuberculosis traverses from alveolar macrophages to phenotypically diverse monocyte-derived phagocytes and neutrophils in the lung parenchyma. Here, we compare the in vivo kinetics of early bacterial growth and cell-to-cell spread of two strains of M. tuberculosis: a lineage 2 strain, 4334, and the widely studied lineage 4 strain H37Rv. Using flow cytometry, live cell sorting of phenotypic subsets, and quantitation of bacteria in cells of the distinct subsets, we found that 4334 induces less leukocyte influx into the lungs but demonstrates earlier population expansion and cell-to-cell spread. The earlier spread of 4334 to recruited cells, including monocyte-derived dendritic cells, is accompanied by earlier and greater magnitude of CD4+ T cell activation. The results provide evidence that strain-specific differences in interactions with lung leukocytes can shape adaptive immune responses in vivo. IMPORTANCE Tuberculosis is a leading infectious disease killer worldwide and is caused by Mycobacterium tuberculosis. After exposure to M. tuberculosis, outcomes range from apparent elimination to active disease. Early innate immune responses may contribute to differences in outcomes, yet it is not known how bacterial strains alter the early dynamics of innate immune and T cell responses. We infected mice with distinct strains of M. tuberculosis and discovered striking differences in innate cellular recruitment, cell-to-cell spread of bacteria in the lungs, and kinetics of initiation of antigen-specific CD4 T cell responses. We also found that M. tuberculosis can spread beyond alveolar macrophages even before a large influx of inflammatory cells. These results provide evidence that distinct strains of M. tuberculosis can exhibit differential kinetics in cell-to-cell spread which is not directly linked to early recruitment of phagocytes but is subsequently linked to adaptive immune responses.

Fungi are an essential part of the normal collection of intestinal microorganisms, even though their collective abundance comprises only 0.1–1% of all fecal microbes. The composition and role of the fungal population is often studied in relation to early-life microbial colonization and development of the (mucosal) immune system. The genus Candida is frequently described as one of the most abundant genera, and altered fungal compositions (including elevated abundance of Candida spp.) have been linked with intestinal diseases such as inflammatory bowel disease and irritable bowel syndrome. These studies are performed using both culture-dependent and genomic (metabarcoding) techniques. In this review, we aimed to summarize existing data on intestinal Candida spp. colonization in relation to intestinal disease and provide a brief overview of the biological and technical challenges in this field, including the recently described role of sub-species strain variation of intestinal Candida albicans. Together, the evidence for a contributing role of Candida spp. in pediatric and adult intestinal disease is quickly expanding, even though technical and biological challenges may limit full understanding of host-microbe interactions. [ABSTRACT FROM AUTHOR]

One of the largest concerns with world health today is still antibiotic resistance, which is making it imperative to find efficient alternatives as soon as possible. It has been demonstrated that microbes are reliable sources for the creation of therapeutic antibiotics. This research intends to investigate the endophytic microorganisms from several medicinal plants in Fenghuang Mountain (Jiangsu Province, China) and to discover new antibiotics from their secondary metabolites. A total of 269 endophytic strains were isolated from nine distinct medicinal plants. Taxonomic analysis revealed that there were 20 distinct species among these endophytes, with Streptomyces being the most common genus. Three of the target strains were chosen for scale-up fermentation after preliminary screening of antibacterial activities and the metabolomics investigation using LC-MS. These strains were Penicillium sp. NX-S-6, Streptomyces sp. YHLB-L-2 and Streptomyces sp. ZLBB-S-6. Twenty-three secondary metabolites (1–23), including a new sorbicillin analogue (1), were produced as a result of antibacterial activity-guided isolation. Through spectroscopic analysis using MS and NMR, the structures of yield compounds were clarified. According to antibacterial data, S. aureus or B. subtilis were inhibited to varying degrees by sorrentanone (3), emodic acid (8), GKK1032 B (10), linoleic acid (14), toyocamycin (17) and quinomycin A (21). The most effective antimicrobial agent against S. aureus, B. subtilis, E. coli and A. baumannii was quinomycin A (21). In addition, quinomycin A showed strong antifungal activity against Aspergillus fumigatus, Cryptococcus neoformans, and two clinical isolated strains Aspergillus fumigatus #176 and #339, with MIC as 16, 4, 16 and 16 µg/mL, respectively. This is the first time that bioprospecting of actinobacteria and their secondary metabolites from medicinal plants in Fenghuang Mountain was reported. The finding demonstrates the potential of endophytic microbes in medical plants to produce a variety of natural products. Endophytic microbes will be an important source for new antibiotics. [ABSTRACT FROM AUTHOR]

Wolbachia are among the most prevalent and widespread endosymbiotic bacteria on Earth. Wolbachia's success in infecting an enormous number of arthropod species is attributed to two features: the range of phenotypes they induce in their hosts, and their ability to switch between host species. Whilst much progress has been made in elucidating their induced phenotypes, our understanding of Wolbachia host‐shifting is still very limited: we lack answers to even fundamental questions concerning Wolbachia's routes of transfer and the importance of factors influencing host shifts. Here, we investigate the diversity and host‐shift patterns of Wolbachia in scale insects, a group of arthropods with intimate associations with other insects that make them well suited to studying host shifts. Using Illumina multitarget amplicon sequencing of Wolbachia‐infected scale insects and their direct associates we determined the identity of all Wolbachia strains. We then fitted a generalized additive mixed model to our data to estimate the influence of host phylogeny and the geographical distribution on Wolbachia strain sharing among scale insect species. The model predicts no significant contribution of host geography but strong effects of host phylogeny, with high rates of Wolbachia sharing among closely related species and a sudden drop‐off in sharing with increasing phylogenetic distance. We also detected the same Wolbachia strain in scale insects and several intimately associated species (ants, wasps and flies). This indicates putative host shifts and potential routes of transfers via these associates and highlights the importance of ecological connectivity in Wolbachia host‐shifting. [ABSTRACT FROM AUTHOR]

Microcystis spp. produce diverse secondary metabolites within freshwater cyanobacterial harmful algal blooms (cyanoHABs) around the world. In addition to the biosynthetic gene clusters (BGCs) encoding known compounds, Microcystis genomes harbor numerous BGCs of unknown function, indicating a poorly understood chemical repertoire. While recent studies show that Microcystis produces several metabolites in the lab and field, little work has focused on analyzing the abundance and expression of its broader suite of BGCs during cyanoHAB events. Here, we use metagenomic and metatranscriptomic approaches to track the relative abundance of Microcystis BGCs and their transcripts throughout the 2014 western Lake Erie cyanoHAB. The results indicate the presence of several transcriptionally active BGCs that are predicted to synthesize both known and novel secondary metabolites. The abundance and expression of these BGCs shifted throughout the bloom, with transcript abundance levels correlating with temperature, nitrate, and phosphorus concentrations and the abundance of co-occurring predatory and competitive eukaryotic microorganisms, suggesting the importance of both abiotic and biotic controls in regulating expression. This work highlights the need for understanding the chemical ecology and potential risks to human and environmental health posed by secondary metabolites that are produced but often unmonitored. It also indicates the prospects for identifying pharmaceutical-like molecules from cyanoHAB-derived BGCs. IMPORTANCE Microcystis spp. dominate cyanobacterial harmful algal blooms (cyanoHABs) worldwide and pose significant threats to water quality through the production of secondary metabolites, many of which are toxic. While the toxicity and biochemistry of microcystins and several other compounds have been studied, the broader suite of secondary metabolites produced by Microcystis remains poorly understood, leaving gaps in our understanding of their impacts on human and ecosystem health. We used community DNA and RNA sequences to track the diversity of genes encoding synthesis of secondary metabolites in natural Microcystis populations and assess patterns of transcription in western Lake Erie cyanoHABs. Our results reveal the presence of both known gene clusters that encode toxic secondary metabolites as well as novel ones that may encode cryptic compounds. This research highlights the need for targeted studies of the secondary metabolite diversity in western Lake Erie, a vital freshwater source to the United States and Canada. [ABSTRACT FROM AUTHOR]

Abstract Background Symbiotic gut microbes have a rich genomic and metabolic pool and are closely related to hosts’ health. Traditional sequencing profiling masks the genomic and phenotypic diversity among strains from the same species. Innovative droplet-based microfluidic cultivation may help to elucidate the inter-strain interactions. A limited number of bacterial phylotypes colonize the honeybee gut, while individual strains possess unique genomic potential and critical capabilities, which provides a particularly good model for strain-level analyses. Results Here, we construct a droplet-based microfluidic platform and generated ~ 6 × 108 droplets encapsulated with individual bacterial cells from the honeybee gut and cultivate in different media. Shotgun metagenomic analysis reveals significant changes in community structure after droplet-based cultivation, with certain species showing higher strain-level diversity than in gut samples. We obtain metagenome-assembled genomes, and comparative analysis reveal a potential novel cluster from Bifidobacterium in the honeybee. Interestingly, Lactobacillus panisapium strains obtained via droplet cultivation from Apis mellifera contain a unique set of genes encoding l-arabinofuranosidase, which is likely important for the survival of bacteria in competitive environments. Conclusions By encapsulating single bacteria cells inside microfluidic droplets, we exclude potential interspecific competition for the enrichment of rare strains by shotgun sequencing at high resolution. The comparative genomic analysis reveals underlying mechanisms for host-specific adaptations, providing intriguing insights into microbe-microbe interactions. The current approach may facilitate the hunting for elusive bacteria and paves the way for large-scale studies of more complex animal microbial communities. Video Abstract

Ecological and evolutionary dynamics are intrinsically entwined. On short timescales, ecological interactions determine the fate and impact of new mutants, while on longer timescales evolution shapes the entire community. Here, we study the evolution of large numbers of closely related strains with generalized Lotka Volterra interactions but no niche structure. Host-pathogen-like interactions drive the community into a spatiotemporally chaotic state characterized by continual, spatially-local, blooms and busts. Upon the slow serial introduction of new strains, the community diversifies indefinitely, accommodating an arbitrarily large number of strains in spite of the absence of stabilizing niche interactions. The diversifying phase persists — albeit with gradually slowing diversification — in the presence of general, nonspecific, fitness differences between strains, which break the assumption of tradeoffs inherent in much previous work. Building on a dynamical-mean field-theory analysis of the ecological dynamics, an approximate effective model captures the evolution of the diversity and distributions of key properties. This work establishes a potential scenario for understanding how the interplay between evolution and ecology — in particular coevolution of a bacterial and a generalist phage species — could give rise to the extensive fine-scale diversity that is ubiquitous in the microbial world.

Just because two things are related does not mean they are the same. In analyzing microbiome data, we are often limited to species-level analyses, and even with the ability to resolve strains, we lack comprehensive databases and understanding of the importance of strain-level variation outside of a limited number of model organisms. The bacterial genome is highly plastic with gene gain and loss occurring at rates comparable or higher than de novo mutations. As such, the conserved portion of the genome is often a fraction of the pangenome which gives rise to significant phenotypic variation, particularly in traits which are important in host microbe interactions. In this review, we discuss the mechanisms that give rise to strain variation and methods that can be used to study it. We identify that while strain diversity can act as a major barrier in interpreting and generalizing microbiome data, it can also be a powerful tool for mechanistic research. We then highlight recent examples demonstrating the importance of strain variation in colonization, virulence, and xenobiotic metabolism. Moving past taxonomy and the species concept will be crucial for future mechanistic research to understand microbiome structure and function. [ABSTRACT FROM AUTHOR]

Abstract Background Short read sequencing has been used extensively to decipher the genome diversity of human cytomegalovirus (HCMV) strains, but falls short to reveal individual genomes in mixed HCMV strain populations. Novel third-generation sequencing platforms offer an extended read length and promise to resolve how distant polymorphic sites along individual genomes are linked. In the present study, we established a long amplicon PacBio sequencing workflow to identify the absolute and relative quantities of unique HCMV haplotypes spanning over multiple hypervariable sites in mixtures. Initial validation of this approach was performed with defined HCMV DNA templates derived from cell-culture enriched viruses and was further tested for its suitability on patient samples carrying mixed HCMV infections. Results Total substitution and indel error rate of mapped reads ranged from 0.17 to 0.43% depending on the stringency of quality trimming. Artificial HCMV DNA mixtures were correctly determined down to 1% abundance of the minor DNA source when the total HCMV DNA input was 4 × 104 copies/ml. PCR products of up to 7.7 kb and a GC content

Just because two things are related does not mean they are the same. In analyzing microbiome data, we are often limited to species-level analyses, and even with the ability to resolve strains, we lack comprehensive databases and understanding of the importance of strain-level variation outside of a limited number of model organisms. The bacterial genome is highly plastic with gene gain and loss occurring at rates comparable or higher than de novo mutations. As such, the conserved portion of the genome is often a fraction of the pangenome which gives rise to significant phenotypic variation, particularly in traits which are important in host microbe interactions. In this review, we discuss the mechanisms that give rise to strain variation and methods that can be used to study it. We identify that while strain diversity can act as a major barrier in interpreting and generalizing microbiome data, it can also be a powerful tool for mechanistic research. We then highlight recent examples demonstrating the importance of strain variation in colonization, virulence, and xenobiotic metabolism. Moving past taxonomy and the species concept will be crucial for future mechanistic research to understand microbiome structure and function.

While research on the sourdough microbiome has primarily focused on lactic acid bacteria (LAB) and yeast, recent studies have found that acetic acid bacteria (AAB) are also common members. However, the ecology, genomic diversity, and functional contributions of AAB in sourdough remain unknown. To address this gap, we sequenced 29 AAB genomes, including three that represent putatively novel species, from a collection of over 500 sourdough starters surveyed globally from community scientists. We found variations in metabolic traits related to carbohydrate utilization, nitrogen metabolism, and alcohol production, as well as in genes related to mobile elements and defense mechanisms. Sourdough AAB genomes did not cluster when compared to AAB isolated from other environments, although a subset of gene functions was enriched in sourdough isolates. The lack of a sourdough-specific genomic cluster may reflect the nomadic lifestyle of AAB. To assess the consequences of AAB on the emergent function of sourdough starter microbiomes, we constructed synthetic starter microbiomes, varying only the AAB strain included. All AAB strains increased the acidification of synthetic sourdough starters relative to yeast and LAB by 18.5% on average. Different strains of AAB had distinct effects on the profile of synthetic starter volatiles. Taken together, our results begin to define the ways in which AAB shape emergent properties of sourdough and suggest that differences in gene content resulting from intraspecies diversification can have community-wide consequences on emergent function., Importance: This study is a comprehensive genomic and ecological survey of acetic acid bacteria (AAB) isolated from sourdough starters. By combining comparative genomics with manipulative experiments using synthetic microbiomes, we demonstrate that even strains with >97% average nucleotide identity can shift important microbiome functions, underscoring the importance of species and strain diversity in microbial systems. We also demonstrate the utility of sourdough starters as a model system to understand the consequences of genomic diversity at the strain and species level on multispecies communities. These results are also relevant to industrial and home-bakers as we uncover the importance of AAB in shaping properties of sourdough starters that have direct impacts on sensory notes and the quality of sourdough bread., Competing Interests: The authors declare no conflict of interest.

Mucin glycoproteins are a significant source of carbon for the gut bacteria. Various gut microbial species possess diverse hydrolytic enzymes and catabolic pathways for breaking down mucin glycans, resulting in competition for the limited nutrients within the gut environment. Adherence to mucin glycans represents a crucial strategy used by gut microbes to access nutrient reservoirs. Understanding these properties is pivotal for comprehending the survival mechanisms of bacteria in the gastrointestinal tract. However, characterization of individual strains within the vast array of coexisting bacteria in the microbiome is challenging. To investigate this, we developed mucin-immobilized particles by immobilizing porcine gastric mucin (PGM) onto glass beads chemically modified with boronic acid. These PGM-immobilized particles were then anaerobically cultured with human fecal microbiota, and the bacteria adhering to PGM were isolated. Interestingly, the microbiome composition remained largely unchanged irrespective of PGM immobilization. Nonetheless, bacteria isolated from PGM-immobilized glass particles exhibited notably higher N -acetylgalactosaminidase activity compared to the control beads. Furthermore, Bacteroides strains isolated from PGM-immobilized glass particles displayed enhanced adhesive and metabolic properties to PGM. These findings underscore the utility of PGM particles in enriching and isolating specific microbes. Moreover, they highlight substantial differences in microbial properties at the strain level. We anticipate that PGM-immobilized particles will advance culture-based microbiome research, emphasizing the significance of strain-level characterization., Importance: Metabolism of mucin glycans by gut bacteria represents a crucial strategy for accessing nutrient reservoirs. The efficacy of mucin glycan utilization among gut bacteria hinges on the metabolic capabilities of individual strains, necessitating meticulous strain-level characterization. In this investigation, we used glass beads chemically immobilized with mucins to selectively enrich bacteria from fecal fermentation cultures, based on their superior adhesion to and metabolism of mucin glycoproteins. These findings lend support to the hypothesis that the physical interactions between bacteria and mucin glycoprotein components directly correlate with their capacity to utilize mucins as nutrient sources. Furthermore, our study implies that physical proximity may significantly influence bacterial nutrient acquisition within the ecosystem, facilitating gut bacteria's access to carbohydrate components., Competing Interests: The authors declare no conflict of interest.

Abstract Background Microbial acquisition and development of the gut microbiota impact the establishment of a healthy host-microbes symbiosis. Compared with other animals, the eusocial bumblebees and honeybees possess a simple, recurring, and similar set of gut microbiota. However, all bee gut phylotypes have high strain-level diversity. Gut communities of different bee species are composed of host-specific groups of strains. The variable genomic regions among strains of the same species often confer critical functional differences, such as carbon source utilization, essential for the natural selection of specific strains. The annual bumblebee colony founded by solitary queens enables tracking the transmission routes of gut bacteria during development stages. Results Here, we first showed the changes in the microbiome of individual bumblebees across their holometabolous life cycle. Some core gut bacteria persist throughout different stages of development. Gut microbiota of newly emerged workers always resembles those of their queens, suggesting a vertical transmission of strains from queens to the newborn workers. We then follow the dynamic changes in the gut community by comparing strain-level metagenomic profiles of queen-worker pairs longitudinally collected across different stages of the nest development. Species composition of both queen and worker shifts with the colony’s growth, and the queen-to-worker vertical inheritance of specific strains was identified. Finally, comparative metagenome analysis showed clear host-specificity for microbes across different bee hosts. Species from honeybees often possess a higher level of strain variation, and they also exhibited more complex gene repertoires linked to polysaccharide digestion. Our results demonstrate bacterial transmission events in bumblebee, highlighting the role of social interactions in driving the microbiota composition. Conclusions By the community-wide metagenomic analysis based on the custom genomic database of bee gut bacteria, we reveal strain transmission events at high resolution and the dynamic changes in community structure along with the colony development. The social annual life cycle of bumblebees is key for the acquisition and development of the gut microbiota. Further studies using the bumblebee model will advance our understanding of the microbiome transmission and the underlying mechanisms, such as strain competition and niche selection. Video Abstract

One of the largest concerns with world health today is still antibiotic resistance, which is making it imperative to find efficient alternatives as soon as possible. It has been demonstrated that microbes are reliable sources for the creation of therapeutic antibiotics. This research intends to investigate the endophytic microorganisms from several medicinal plants in Fenghuang Mountain (Jiangsu Province, China) and to discover new antibiotics from their secondary metabolites. A total of 269 endophytic strains were isolated from nine distinct medicinal plants. Taxonomic analysis revealed that there were 20 distinct species among these endophytes, with Streptomyces being the most common genus. Three of the target strains were chosen for scale-up fermentation after preliminary screening of antibacterial activities and the metabolomics investigation using LC-MS. These strains were Penicillium sp. NX-S-6, Streptomyces sp. YHLB-L-2 and Streptomyces sp. ZLBB-S-6. Twenty-three secondary metabolites (1–23), including a new sorbicillin analogue (1), were produced as a result of antibacterial activity-guided isolation. Through spectroscopic analysis using MS and NMR, the structures of yield compounds were clarified. According to antibacterial data, S. aureus or B. subtilis were inhibited to varying degrees by sorrentanone (3), emodic acid (8), GKK1032 B (10), linoleic acid (14), toyocamycin (17) and quinomycin A (21). The most effective antimicrobial agent against S. aureus, B. subtilis, E. coli and A. baumannii was quinomycin A (21). In addition, quinomycin A showed strong antifungal activity against Aspergillus fumigatus, Cryptococcus neoformans, and two clinical isolated strains Aspergillus fumigatus #176 and #339, with MIC as 16, 4, 16 and 16 µg/mL, respectively. This is the first time that bioprospecting of actinobacteria and their secondary metabolites from medicinal plants in Fenghuang Mountain was reported. The finding demonstrates the potential of endophytic microbes in medical plants to produce a variety of natural products. Endophytic microbes will be an important source for new antibiotics.

Background: Symbiotic gut microbes have a rich genomic and metabolic pool and are closely related to hosts' health. Traditional sequencing profiling masks the genomic and phenotypic diversity among strains from the same species. Innovative droplet-based microfluidic cultivation may help to elucidate the inter-strain interactions. A limited number of bacterial phylotypes colonize the honeybee gut, while individual strains possess unique genomic potential and critical capabilities, which provides a particularly good model for strain-level analyses. Results: Here, we construct a droplet-based microfluidic platform and generated ~ 6 × 108 droplets encapsulated with individual bacterial cells from the honeybee gut and cultivate in different media. Shotgun metagenomic analysis reveals significant changes in community structure after droplet-based cultivation, with certain species showing higher strain-level diversity than in gut samples. We obtain metagenome-assembled genomes, and comparative analysis reveal a potential novel cluster from Bifidobacterium in the honeybee. Interestingly, Lactobacillus panisapium strains obtained via droplet cultivation from Apis mellifera contain a unique set of genes encoding l-arabinofuranosidase, which is likely important for the survival of bacteria in competitive environments. Conclusions: By encapsulating single bacteria cells inside microfluidic droplets, we exclude potential interspecific competition for the enrichment of rare strains by shotgun sequencing at high resolution. The comparative genomic analysis reveals underlying mechanisms for host-specific adaptations, providing intriguing insights into microbe-microbe interactions. The current approach may facilitate the hunting for elusive bacteria and paves the way for large-scale studies of more complex animal microbial communities. 6MWJv9N2gCCgq2b2j_Wic- Video Abstract [ABSTRACT FROM AUTHOR]

Approximately half of the world's population is infected with Helicobacter pylori, which causes gastric disease. Recent systematic reviews and meta-analyses have reported that H. pylori may also have extragastric manifestations such as hematologic diseases, including chronic immune thrombocytopenia (cITP). However, the molecular mechanisms by which H. pylori induces cITP remain unclear, and may involve the host immune response, bacterial strain diversity, and delivery of bacterial molecules to the host blood vessels. This review discusses the important pathophysiological mechanisms by which H. pylori potentially contributes to the development of cITP in infected patients. [ABSTRACT FROM AUTHOR]

Undefined strain starters are used for the production of many traditional and artisanal cheeses. Composition of undefined starters depends on several factors, and the diversity in strains and species significantly affects cheese quality and features. Culture-dependent approaches have long been used for the microbial profiling and functionalities of undefined cultures but underestimate their diversity due to culturability biases. Recently, culture-independent methods, based on high-throughput sequencing (HTS), have been preferred, with a significant boost in resolution power and sensitivity level. Amplicon targeted (AT) metagenomics, based on 16S rRNA sequencing, returned a larger microbiota diversity at genus and, sometimes, at species levels for artisanal starters of several PDO cheeses, but was inappropriate for populations with high strain diversity, and other gene targets were tested in AT approaches. Shotgun metagenomics (total DNA) and metatranscriptomics (total RNA), although are more powerful in depicting diversity and functionality of undefined cultures, have been rarely applied because of some limitations (e.g., high costs and laboriousness, need for bioinformatics skills). The advantages of HTS technologies are undoubted, but some hurdles need to be still overcame (e.g., resolution power, discrepancy between active and inactive cells, robust analytic pipelines, cost and time reduction for integrated approaches) so that HTS become routinary and convenient for defining complexity, microbial interactions (including host-phage relationships) and evolution in cheeses of undefined starters. [ABSTRACT FROM AUTHOR]

To test the prevailing dogma that Streptococcus pyogenes emm -types that cause pharyngitis are the same as those associated with the carriage, using a global dataset. Drawing on our systematic review of the global distribution of S. pyogenes emm -types and emm -clusters from 1990 to 2023, we compared the distribution and diversity of strains associated with pharyngitis and pharyngeal carriage, in the context of local United Nations Development Programme Human Development Index (HDI) values. We included 20 222 isolates from 71 studies done in 34 countries, with the vast majority of carriage strain data from studies in 'Low HDI' settings (550/1293; 43%). There was higher emm -type diversity for carriage than pharyngitis strains (Simpson Reciprocal Index of diversity 28.9 vs. 11.4). Compared with pharyngitis strains, carriage emm -types were disproportionately from emm -clusters E and D, usually described as 'generalist' or 'skin' strains. A limited number of studies have compared S. pyogenes strains from cases of pharyngitis compared with carriage. Our understanding of strains associated with carriage is the poorest for high-income settings. In low and medium HDI countries, we found greater strain associated with pharyngeal carriage than pharyngitis. Improving our understanding of S. pyogenes carriage epidemiology in the pre-vaccine era will help to decipher the direct and potential indirect effects of vaccines. [ABSTRACT FROM AUTHOR]

Background: Short read sequencing has been used extensively to decipher the genome diversity of human cytomegalovirus (HCMV) strains, but falls short to reveal individual genomes in mixed HCMV strain populations. Novel third-generation sequencing platforms offer an extended read length and promise to resolve how distant polymorphic sites along individual genomes are linked. In the present study, we established a long amplicon PacBio sequencing workflow to identify the absolute and relative quantities of unique HCMV haplotypes spanning over multiple hypervariable sites in mixtures. Initial validation of this approach was performed with defined HCMV DNA templates derived from cell-culture enriched viruses and was further tested for its suitability on patient samples carrying mixed HCMV infections. Results: Total substitution and indel error rate of mapped reads ranged from 0.17 to 0.43% depending on the stringency of quality trimming. Artificial HCMV DNA mixtures were correctly determined down to 1% abundance of the minor DNA source when the total HCMV DNA input was 4 × 104 copies/ml. PCR products of up to 7.7 kb and a GC content < 55% were efficiently generated when DNA was directly isolated from patient samples. In a single sample, up to three distinct haplotypes were identified showing varying relative frequencies. Alignments of distinct haplotype sequences within patient samples showed uneven distribution of sequence diversity, interspersed by long identical stretches. Moreover, diversity estimation at single polymorphic regions as assessed by short amplicon sequencing may markedly underestimate the overall diversity of mixed haplotype populations. Conclusions: Quantitative haplotype determination by long amplicon sequencing provides a novel approach for HCMV strain characterisation in mixed infected samples which can be scaled up to cover the majority of the genome by multi-amplicon panels. This will substantially improve our understanding of intra-host HCMV strain diversity and its dynamic behaviour. [ABSTRACT FROM AUTHOR]

Abstract Background Populations of closely related microbial strains can be simultaneously present in bacterial communities such as the human gut microbiome. We recently developed a de novo genome assembly approach that uses read cloud sequencing to provide more complete microbial genome drafts, enabling precise differentiation and tracking of strain-level dynamics across metagenomic samples. In this case study, we present a proof-of-concept using read cloud sequencing to describe bacterial strain diversity in the gut microbiome of one hematopoietic cell transplantation patient over a 2-month time course and highlight temporal strain variation of gut microbes during therapy. The treatment was accompanied by diet changes and administration of multiple immunosuppressants and antimicrobials. Methods We conducted short-read and read cloud metagenomic sequencing of DNA extracted from four longitudinal stool samples collected during the course of treatment of one hematopoietic cell transplantation (HCT) patient. After applying read cloud metagenomic assembly to discover strain-level sequence variants in these complex microbiome samples, we performed metatranscriptomic analysis to investigate differential expression of antibiotic resistance genes. Finally, we validated predictions from the genomic and metatranscriptomic findings through in vitro antibiotic susceptibility testing and whole genome sequencing of isolates derived from the patient stool samples. Results During the 56-day longitudinal time course that was studied, the patient’s microbiome was profoundly disrupted and eventually dominated by Bacteroides caccae. Comparative analysis of B. caccae genomes obtained using read cloud sequencing together with metagenomic RNA sequencing allowed us to identify differences in substrain populations over time. Based on this, we predicted that particular mobile element integrations likely resulted in increased antibiotic resistance, which we further supported using in vitro antibiotic susceptibility testing. Conclusions We find read cloud assembly to be useful in identifying key structural genomic strain variants within a metagenomic sample. These strains have fluctuating relative abundance over relatively short time periods in human microbiomes. We also find specific structural genomic variations that are associated with increased antibiotic resistance over the course of clinical treatment.

Aphids have evolved tight relationships with heritable endosymbionts, i.e., bacteria hosted within their tissues. Besides the primary endosymbiont Buchnera aphidicola, aphids host many facultative secondary endosymbionts with functions they may or may not benefit from. The different phenologies, lifestyles, and natural enemies of aphid species are predicted to favor the selection for distinct endosymbiont assemblages, as well as the emergence of intra-specific genetic diversity in the symbiotic bacteria. In this study, we (1) investigated the diversity of endosymbionts associated with four species from the genus Aphis in the field, and (2) we characterized the genetic diversity of Hamiltonella defensa, an endosymbiont that protects aphids against parasitoid wasps. We observed strong differences in the composition of endosymbiont communities among the four aphid species. H. defensa was clearly the dominant symbiont, although its abundance in each species varied from 25 to 96%. Using a multilocus sequence-typing approach, we found limited strain diversity in H. defensa. Each aphid species harbored two major strains, and none appeared shared between species. Symbiont phylogenies can thus help to understand the (seemingly limited) mobility of endosymbionts in aphid communities and the selection forces driving strain diversification.

ABSTRACT Candida albicans is a pathobiont that colonizes multiple niches in the body including the gastrointestinal (GI) tract but is also responsible for both mucosal and systemic infections. Despite its prevalence as a human commensal, the murine GI tract is generally refractory to colonization with the C. albicans reference isolate SC5314. Here, we identify two C. albicans isolates, 529L and CHN1, that stably colonize the murine GI tract in three different animal facilities under conditions where SC5314 is lost from this niche. Analysis of the bacterial microbiota did not show notable differences among mice colonized with the three C. albicans strains. We compared the genotypes and phenotypes of these three strains and identified thousands of single nucleotide polymorphisms (SNPs) and multiple phenotypic differences, including their ability to grow and filament in response to nutritional cues. Despite striking filamentation differences under laboratory conditions, however, analysis of cell morphology in the GI tract revealed that the three isolates exhibited similar filamentation properties in this in vivo niche. Notably, we found that SC5314 is more sensitive to the antimicrobial peptide CRAMP, and the use of CRAMP-deficient mice modestly increased the ability of SC5314 to colonize the GI tract relative to CHN1 and 529L. These studies provide new insights into how strain-specific differences impact C. albicans traits in the host and advance CHN1 and 529L as relevant strains to study C. albicans pathobiology in its natural host niche. IMPORTANCE Understanding how fungi colonize the GI tract is increasingly recognized as highly relevant to human health. The animal models used to study Candida albicans commensalism commonly rely on altering the host microbiome (via antibiotic treatment or defined diets) to establish successful GI colonization by the C. albicans reference isolate SC5314. Here, we characterize two C. albicans isolates that can colonize the murine GI tract without antibiotic treatment and can therefore be used as tools for studying fungal commensalism. Importantly, experiments were replicated in three different animal facilities and utilized three different mouse strains. Differential colonization between fungal isolates was not associated with alterations in the bacterial microbiome but rather with distinct responses to CRAMP, a host antimicrobial peptide. This work emphasizes the importance of C. albicans intraspecies variation as well as host antimicrobial defense mechanisms in defining the outcome of commensal interactions.

Fungi were some of the earliest organismal systems used to explore mutational processes and its phenotypic consequences on members of a species. Yeasts that cause significant human disease were quickly incorporated into these investigations to define the genetic and phenotypic drivers of virulence. Among Candida species, Candida albicans has emerged as a model for studying genomic processes of evolution because of its clinical relevance, relatively small genome, and ability to tolerate complex chromosomal changes. Here, we describe major recent findings that used evolution of strains from defined genetic backgrounds to delineate mutational and adaptative processes and include how nascent exploration into naturally occurring variation is contributing to these conceptual frameworks. Ultimately, efforts to discern adaptive mechanisms used by C. albicans will continue to divulge new biology and can better inform treatment regimens for the increasing prevalence of fungal disease., Competing Interests: The authors declare no conflict of interest.

At the end of 2022 into early 2023, the UK Health Security Agency reported unusually high levels of scarlet fever and invasive disease caused by Streptococcus pyogenes (StrepA or group A Streptococcus ). During this time, we collected and genome-sequenced 341 non-invasive throat and skin S. pyogenes isolates identified during routine clinical diagnostic testing in Sheffield, a large UK city. We compared the data with that obtained from a similar collection of 165 isolates from 2016 to 2017. Numbers of throat-associated isolates collected peaked in early December 2022, reflecting the national scarlet fever upsurge, while skin infections peaked later in December. The most common emm -types in 2022-2023 were emm 1 (28.7 %), emm 12 (24.9 %) and emm 22 (7.7 %) in throat and emm 1 (22 %), emm 12 (10 %), emm 76 (18 %) and emm 49 (7 %) in skin. While all emm 1 isolates were the M1 UK lineage, the comparison with 2016-2017 revealed diverse lineages in other emm -types, including emm 12, and emergent lineages within other types including a new acapsular emm 75 lineage, demonstrating that the upsurge was not completely driven by a single genotype. The analysis of the capsule locus predicted that only 51 % of throat isolates would produce capsule compared with 78% of skin isolates. Ninety per cent of throat isolates were also predicted to have high NADase and streptolysin O (SLO) expression, based on the promoter sequence, compared with only 56% of skin isolates. Our study has highlighted the value in analysis of non-invasive isolates to characterize tissue tropisms, as well as changing strain diversity and emerging genomic features which may have implications for spillover into invasive disease and future S. pyogenes upsurges.

Background: Streptococcus pyogenes-related skin infections are increasingly implicated in the development of rheumatic heart disease (RHD) in lower-resourced settings, where they are often associated with scabies. The true prevalence of S. pyogenes-related pyoderma may be underestimated by bacterial culture., Methods: A multiplex qPCR for S. pyogenes, Staphylococcus aureus and Sarcoptes scabiei was applied to 250 pyoderma swabs from a cross-sectional study of children <5 years in The Gambia. Direct PCR-based emm-typing was used to supplement previous whole genome sequencing (WGS) of cultured isolates., Results: Pyoderma lesions with S. pyogenes increased from 51% (127/250) using culture to 80% (199/250) with qPCR. Compared to qPCR, the sensitivity of culture was 95.4% for S. pyogenes (95% CI 77.2-99.9) in samples with S. pyogenes alone (22/250, 9%), but 59.9% (95% CI 52.3-67.2) for samples with S. aureus co-infection (177/250, 71%). Direct PCR-based emm-typing was successful in 50% (46/92) of cases, identifying 27 emm-types, including six not identified by WGS (total 52 emm-types)., Conclusions: Bacterial culture significantly underestimates the burden of S. pyogenes in pyoderma, particularly when co-infected with S. aureus. Molecular methods should be used to enhance the detection of S. pyogenes in surveillance studies and clinical trials of preventative measures in RHD-endemic settings., (© The Author(s) 2024. Published by Oxford University Press on behalf of Infectious Diseases Society of America.)

Feline calicivirus (FCV) is a significant viral pathogen causing upper respiratory tract and oral diseases in cats. The emergence of the virulent systemic FCV variant (VS-FCV) has raised global concern in the past decade. This study aims to explore the epidemiology, genetic characterization, and diversity of FCV strains circulating among Thai cats. Various sample types, including nasal, oral, and oropharyngeal swabs and fresh tissues, were collected from 184 cats across different regions of Thailand from 2016 to 2021. Using reverse transcription real-time polymerase chain reaction (RT-qPCR), FCV infection was investigated, with additional screening for feline herpesvirus-1 (FHV-1) by qPCR. The detection rates for FCV, FHV-1, and co-infection were 46.7, 65.8, and 31.5%, respectively. Significantly, the odds ratio (OR) revealed a strong association between the detection of a single FCV and the presence of gingivostomatitis lesions (OR: 7.15, 95% CI: 1.89-26.99, p  = 0.004). In addition, FCV detection is notably less likely in vaccinated cats (OR: 0.22, 95% CI: 0.07-0.75, p  = 0.015). Amino acid sequence analysis based on the VP1 major capsid protein gene of the 14 FCV-Thai (FCV-TH) strains revealed genetic diversity compared to the other 43 global strains (0 to 86.6%). Intriguingly, a vaccine-like FCV variant was detected in one cat. In summary, this study provides insights into the epidemiology and molecular characteristics of FCV diversity within the Thai cat population for the first time. The identification of unique physicochemical characteristics in the capsid hypervariable region of some FCV-TH strains challenges previous hypotheses. Therefore, further exploration of vaccine-like FCV variants is crucial for a comprehensive understanding and to improve viral prevention and control strategies., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2024 Phongroop, Rattanasrisomporn, Piewbang, Tangtrongsup, Rungsipipat and Techangamsuwan.)

Background: Microbial acquisition and development of the gut microbiota impact the establishment of a healthy host-microbes symbiosis. Compared with other animals, the eusocial bumblebees and honeybees possess a simple, recurring, and similar set of gut microbiota. However, all bee gut phylotypes have high strain-level diversity. Gut communities of different bee species are composed of host-specific groups of strains. The variable genomic regions among strains of the same species often confer critical functional differences, such as carbon source utilization, essential for the natural selection of specific strains. The annual bumblebee colony founded by solitary queens enables tracking the transmission routes of gut bacteria during development stages. Results: Here, we first showed the changes in the microbiome of individual bumblebees across their holometabolous life cycle. Some core gut bacteria persist throughout different stages of development. Gut microbiota of newly emerged workers always resembles those of their queens, suggesting a vertical transmission of strains from queens to the newborn workers. We then follow the dynamic changes in the gut community by comparing strain-level metagenomic profiles of queen-worker pairs longitudinally collected across different stages of the nest development. Species composition of both queen and worker shifts with the colony's growth, and the queen-to-worker vertical inheritance of specific strains was identified. Finally, comparative metagenome analysis showed clear host-specificity for microbes across different bee hosts. Species from honeybees often possess a higher level of strain variation, and they also exhibited more complex gene repertoires linked to polysaccharide digestion. Our results demonstrate bacterial transmission events in bumblebee, highlighting the role of social interactions in driving the microbiota composition. Conclusions: By the community-wide metagenomic analysis based on the custom genomic database of bee gut bacteria, we reveal strain transmission events at high resolution and the dynamic changes in community structure along with the colony development. The social annual life cycle of bumblebees is key for the acquisition and development of the gut microbiota. Further studies using the bumblebee model will advance our understanding of the microbiome transmission and the underlying mechanisms, such as strain competition and niche selection. DndA2Lw_mPA4b2NMU2mTwy Video Abstract [ABSTRACT FROM AUTHOR]

ABSTRACT Liver cirrhosis (LC) has been associated with gut microbes. However, the strain diversity of species and its association with LC have received little attention. Here, we constructed a computational framework to study the strain heterogeneity in the gut microbiome of patients with LC. Only Faecalibacterium prausnitzii shows different single-nucleotide polymorphism (SNP) patterns between the LC and healthy control (HC) groups. Strain diversity analysis discovered that although most F. prausnitzii genomes are more deficient in the LC group than in the HC group at the strain level, a subgroup of 19 F. prausnitzii strains showed no sensitivity to LC, which is inconsistent with the species-level result. The functional differences between this subgroup and other strains may involve short-chain fatty acid production and chlorine-related pathways. These findings demonstrate functional differences among F. prausnitzii subgroups, which extend current knowledge about strain heterogeneity and relationships between F. prausnitzii and LC at the strain level. IMPORTANCE Most metagenomic studies focus on microbes at the species level, thus ignoring the different effects of different strains of the same species on the host. In this study, we explored the different microbes at the strain level in the intestines of patients with liver cirrhosis and of healthy people. Previous studies have shown that the species Faecalibacterium prausnitzii has a lower abundance in patients with liver cirrhosis than in healthy people. However, our results found multiple F. prausnitzii strains that do not decrease in abundance in patients with liver cirrhosis. It is more sensitive to select the appropriate strains as indicators to distinguish between the disease and the control samples than to use the entire species as an indicator. We clustered multiple F. prausnitzii strains and discuss the functional differences of different clusters. Our findings suggest that more attention should be paid to metagenomic studies at the strain level.

In high-transmission endemic regions, local populations of Plasmodium falciparum exhibit vast diversity of the var genes encoding its major surface antigen, with each parasite comprising multiple copies from this diverse gene pool. This strategy to evade the immune system through large combinatorial antigenic diversity is common to other hyperdiverse pathogens. It underlies a series of fundamental epidemiological characteristics, including large reservoirs of transmission from high prevalence of asymptomatics and long-lasting infections. Previous theory has shown that negative frequency-dependent selection (NFDS) mediated by the acquisition of specific immunity by hosts structures the diversity of var gene repertoires, or strains, in a pattern of limiting similarity that is both non-random and non-neutral. A combination of stochastic agent-based models and network analyses has enabled the development and testing of theory in these complex adaptive systems, where assembly of local parasite diversity occurs under frequency-dependent selection and large pools of variation. We show here the application of these approaches to theory comparing the response of the malaria transmission system to intervention when strain diversity is assembled under (competition-based) selection vs. a form of neutrality, where immunity depends only on the number but not the genetic identity of previous infections. The transmission system is considerably more persistent under NFDS, exhibiting a lower extinction probability despite comparable prevalence during intervention. We explain this pattern on the basis of the structure of strain diversity, in particular the more pronounced fraction of highly dissimilar parasites. For simulations that survive intervention, prevalence under specific immunity is lower than under neutrality, because the recovery of diversity is considerably slower than that of prevalence and decreased var gene diversity reduces parasite transmission. A Principal Component Analysis of network features describing parasite similarity reveals that despite lower overall diversity, NFDS is quickly restored after intervention constraining strain structure and maintaining patterns of limiting similarity important to parasite persistence. Given the described enhanced persistence under perturbation, intervention efforts will likely require longer times than the usual practice to eliminate P. falciparum populations. We discuss implications of our findings and potential analogies for ecological communities with non-neutral assembly processes involving frequency-dependence.

The recent outbreak of Elizabethkingia anophelis in the Midwestern states of USA caused a number of deaths. Notably, these deaths occurred due to E. anophelis causing neonatal meningitis, bacteraemia, sepsis, blood stream infections and respiratory infections. These infections may pose serious threats to public health. This systematic review is meant to develop a deeper insight into the current status of E. anophelis related evidence and to highlight areas that need further research. Reviewing the existing literature will help other researchers to identify and address the knowledge gaps. Various free access databases such as Google Scholar, Scopus, PubMed, and Science Direct were employed for literature survey. All articles published since 2011, when the outbreak was reported for the first time, were consulted for this systematic review. Research related to this subject is in its earlier stages and little information is currently available. Future studies must focus on the molecular basis, control, prevention, and therapeutics of E. anophelis to mitigate its increasing risk. This review is meant to provide baseline data for future research. Scientific community must carry out research on the infections caused by E. anophelis mosquito, else it may result in a disastrous outbreak. Copyright (c) 2021 NoshabaRani, BrairaWahid

The precise impact of species and strain diversity on fungal-bacterial interactions and the overall community functioning has remained unclear. First, our study revealed how Debaryomyces hansenii influences diverse bacteria to accumulate key metabolites in a simulated fermented food system. For flavor, D. hansenii promoted the accumulation of branched-chain esters in Staphylococcus xylosus by promoting growth and facilitating the precursor branched-chain acids transformations but hindered the accumulation of Staphylococcus equorum . Furthermore, fungal-bacterial interactions displayed diversity among S. equorum strains. For bioactive compounds, species and strain diversity of lactic acid bacteria (LAB) also influences the production of indole derivatives. Then, we investigated specific metabolic exchanges under reciprocal interaction. Amino acids, rather than vitamins, were identified as the primary drivers of the bacterial growth promotion. Moreover, precursor transformations by D. hansenii played a significant role in branched-chain esters production. Finally, a synthetic community capable of producing high concentrations of branched-chain esters and indole derivatives was successfully constructed. These results provide valuable insights into understanding and designing synthetic communities for fermented sausages.

Alcoholic fermentation is influenced by yeast strain, culture media, substrate concentration and fermentation conditions, which contribute to taste and aroma. Some non -Saccharomyces yeasts are recognized as volatile compound producers that enrich aromatic profile of alcoholic beverages. In this work, 21 strains of Pichia kluyveri isolated from different fermentative processes and regions were evaluated. A principal component analysis (PCA) showed statistical differences between strains mainly associated with the variety and concentration of the compounds produced. From the PCA, two strains (PK1 and PK8) with the best volatile compound production were selected to evaluate the impact of culture media (M12 medium and Agave tequilana juice), stirring speeds (100 and 250 rpm) and temperatures (20°C, 25°C and 30°C). Increased ester production was observed at 250 rpm. Greatest effect in alcohols and ester production was found with A. tequilana , identifying PK1 as higher alcohol producer, and PK8 as better ester producer. Regarding temperature, PK1 increased ester production with decreased fermentation temperature. PK8 presented maximum levels of ethyl acetate and ethyl dodecanoate at 20°C, and finally isoamyl acetate increased its production at 30°C. Therefore,  P. kluyveri strains are of great interest to produce different aromatic profiles that are affected by factors including medium, agitation and temperature. [ABSTRACT FROM AUTHOR]

ABSTRACT Host-associated microbiomes can be critical for the health and proper development of animals and plants. The answers to many fundamental questions regarding the modes of acquisition and microevolution of microbiome communities remain to be established. Deciphering strain-level dynamics is essential to fully understand how microbial communities evolve, but the forces shaping the strain-level dynamics of microbial communities remain largely unexplored, mostly because of methodological issues and cost. Here, we used targeted strain-level deep sequencing to uncover the strain dynamics within a host-associated microbial community using the honey bee gut microbiome as a model system. Our results revealed that amplicon sequencing of conserved protein-coding gene regions using species-specific primers is a cost-effective and accurate method for exploring strain-level diversity. In fact, using this method we were able to confirm strain-level results that have been obtained from whole-genome shotgun sequencing of the honey bee gut microbiome but with a much higher resolution. Importantly, our deep sequencing approach allowed us to explore the impact of low-frequency strains (i.e., cryptic strains) on microbiome dynamics. Results show that cryptic strain diversity is not responsible for the observed variations in microbiome composition across bees. Altogether, the findings revealed new fundamental insights regarding strain dynamics of host-associated microbiomes. IMPORTANCE The factors driving fine-scale composition and dynamics of gut microbial communities are poorly understood. In this study, we used metagenomic amplicon deep sequencing to decipher the strain dynamics of two key members of the honey bee gut microbiome. Using this high-throughput and cost-effective approach, we were able to confirm results from previous large-scale whole-genome shotgun (WGS) metagenomic sequencing studies while also gaining additional insights into the community dynamics of two core members of the honey bee gut microbiome. Moreover, we were able to show that cryptic strains are not responsible for the observed variations in microbiome composition across bees.

ABSTRACT Diversification can generate genomic and phenotypic strain-level diversity within microbial species. This microdiversity is widely recognized in populations, but the community-level consequences of microbial strain-level diversity are poorly characterized. Using the cheese rind model system, we tested whether strain diversity across microbiomes from distinct geographic regions impacts assembly dynamics and functional outputs. We first isolated the same three bacterial species (Staphylococcus equorum, Brevibacterium auranticum, and Brachybacterium alimentarium) from nine cheeses produced in different regions of the United States and Europe to construct nine synthetic microbial communities consisting of distinct strains of the same three bacterial species. Comparative genomics identified distinct phylogenetic clusters and significant variation in genome content across the nine synthetic communities. When we assembled each synthetic community with initially identical compositions, community structure diverged over time, resulting in communities with different dominant taxa. The taxonomically identical communities showed differing responses to abiotic (high salt) and biotic (the fungus Penicillium) perturbations, with some communities showing no response and others substantially shifting in composition. Functional differences were also observed across the nine communities, with significant variation in pigment production (light yellow to orange) and in composition of volatile organic compound profiles emitted from the rinds (nutty to sulfury). IMPORTANCE Our work demonstrated that the specific microbial strains used to construct a microbiome could impact the species composition, perturbation responses, and functional outputs of that system. These findings suggest that 16S rRNA gene taxonomic profiles alone may have limited potential to predict the dynamics of microbial communities because they usually do not capture strain-level diversity. Observations from our synthetic communities also suggest that strain-level diversity has the potential to drive variability in the aesthetics and quality of surface-ripened cheeses.

ABSTRACT Sporadic Creutzfeldt-Jakob disease (sCJD) cases are currently classified according to the methionine/valine polymorphism at codon 129 of the PRNP gene and the proteinase K-digested abnormal prion protein (PrPres) isoform identified by Western blotting (type 1 or type 2). Converging evidence led to the view that MM/MV1, VV/MV2, and VV1 and MM2 sCJD cases are caused by distinct prion strains. However, in a significant proportion of sCJD patients, both type 1 and type 2 PrPres were reported to accumulate in the brain, which raised questions about the diversity of sCJD prion strains and the coexistence of two prion strains in the same patient. In this study, a panel of sCJD brain isolates (n = 29) that displayed either a single or mixed type 1/type 2 PrPres were transmitted into human-PrP-expressing mice (tgHu). These bioassays demonstrated that two distinct prion strains (M1CJD and V2CJD) were associated with the development of sCJD in MM1/MV1 and VV2/MV2 patients. However, in about 35% of the investigated VV and MV cases, transmission results were consistent with the presence of both M1CJD and V2CJD strains, including in patients who displayed a “pure” type 1 or type 2 PrPres. The use of a highly sensitive prion in vitro amplification technique that specifically probes the V2CJD strain revealed the presence of the V2CJD prion in more than 80% of the investigated isolates, including isolates that propagated as a pure M1CJD strain in tgHu. These results demonstrate that at least two sCJD prion strains can be present in a single patient. IMPORTANCE sCJD occurrence is currently assumed to result from spontaneous and stochastic formation of a misfolded PrP nucleus in the brains of affected patients. This original nucleus then recruits and converts nascent PrPC into PrPSc, leading to the propagation of prions in the patient’s brain. Our study demonstrates the coexistence of two prion strains in the brains of a majority of the 23 sCJD patients investigated. The relative proportion of these sCJD strains varied both between patients and between brain areas in a single patient. These findings strongly support the view that the replication of an sCJD prion strain in the brain of a patient can result in the propagation of different prion strain subpopulations. Beyond its conceptual importance for our understanding of prion strain properties and evolution, the sCJD strain mixture phenomenon and its frequency among patients have important implications for the development of therapeutic strategies for prion diseases.

Biofilm production, hitherto an uncharacterized feature among circulating Pasteurella multocida strains, was studied along with the antibiotic susceptibility pattern. On the basis of biofilm formation ability, all the strains were categorized into four groups under six different culture conditions: strong biofilm-forming (22%), moderate (19%), weak (51%), and non-adherent (7%). Strains from serogroups A and B formed significant biofilms in at least one culture condition whereas strains from serogroup D were unable to form biofilms. All strains were found to be susceptible to tetracycline. In addition, the correlation between diverse factors (host, capsule type, clinical condition and the tadD gene) as well as antimicrobial susceptibility in biofilm production were analyzed by Joint distribution models, and showed that enrofloxacin and azithromycin resistant strains were positively correlated with strong biofilm production. [ABSTRACT FROM AUTHOR]

Background: Simian immunodeficiency virus (SIV) naturally infects African non‐human primates (NHPs) and poses a threat of transmission to humans through hunting and consumption of monkeys as bushmeat. This study investigated the as of yet unknown molecular diversity of SIV in free‐ranging Chlorocebus species (African green monkeys—AGMs) and Papio anubis (olive baboons) within Mombasa, Kisumu and Naivasha urban centres in Kenya. Methods: We collected blood samples from 124 AGMs and 65 olive baboons in situ, and detected SIV by high‐resolution melting analysis and sequencing of PCR products. Results: Simian immunodeficiency virus prevalence was 32% in AGMs and 3% in baboons. High‐resolution melting (HRM) analysis demonstrated distinct melt profiles illustrating virus diversity confirmed by phylogenetic analysis. Conclusions: There is persistent evolutionary diversification of SIVagm strains in its natural host, AGMs and cross‐species infection to olive baboons is occurring. Further study is required to establish pathogenesis of the diverse SIVagm variants and baboon immunological responses. [ABSTRACT FROM AUTHOR]