Nguyet Kong

Complete genome sequences of a clinical isolate and an environmental isolate of Vibrio parahaemolyticus

ABSTRACT Vibrio parahaemolyticus is the leading cause of seafood-borne infections in the United States. We report complete genome sequences for two V. parahaemolyticus strains isolated in 2007, CDC_K4557 and FDA_R31 of clinical and oyster origin, respectively. These two sequences might assist in the investigation of differential virulence of this organism.

Gastrointestinal Microbes Interact with Canine Adipose-Derived Mesenchymal Stem Cells In Vitro and Enhance Immunomodulatory Functions

Mesenchymal stem cells (MSCs) are somatic, multipotent stromal cells with potent immunomodulatory and regenerative properties. Although MSCs have pattern recognition receptors and are modulated by Toll-like receptor ligands, MSC-microbial interactions are poorly defined. The objectives of this study were to determine the effect of bacterial association on MSC function. We hypothesized that gastrointestinal bacteria associate with MSCs and alter their immunomodulatory properties. The effect of MSC-microbial interactions on MSC morphology, viability, proliferation, migration, and immunomodulatory functions was investigated. MSCs associated with a remarkable array of enteric pathogens and commensal bacteria. MSC interactions with two model organisms, the pathogen Salmonella typhimurium and the probiotic Lactobacillus acidophilus, were further investigated. While ST readily invaded MSCs, LB adhered to the MSC plasma membrane. Neither microbe induced MSC death, degeneration, or diminished proliferation. Microbial association did not upregulate MHC-II, CD80/86, or CD1 expression. MSC-microbial interaction significantly increased transcription of key immunomodulatory genes, including COX2, IL6, and IL8, coupled with significantly increased prostaglandin E2 (PGE2), interleukin (IL)6, and IL8 secretion. MSC-ST coincubation resulted in increased MSC expression of CD54, and significant augmentation of MSC inhibition of mitogen-induced T-cell proliferation. T-cell proliferation was partially restored when PGE2 secretion was blocked from ST-primed MSCs. MSC-microbe interactions have a profound effect on MSC function and may be pivotal in a variety of clinical settings where MSCs are being explored as potential therapeutics in the context of microbial communities, such as Crohns disease, chronic nonhealing wounds, and sepsis.

Salmonella Degrades the Host Glycocalyx Leading to Altered Infection and Glycan Remodeling

Complex glycans cover the gut epithelial surface to protect the cell from the environment. Invasive pathogens must breach the glycan layer before initiating infection. While glycan degradation is crucial for infection, this process is inadequately understood. Salmonella contains 47 glycosyl hydrolases (GHs) that may degrade the glycan. We hypothesized that keystone genes from the entire GH complement of Salmonella are required to degrade glycans to change infection. This study determined that GHs recognize the terminal monosaccharides (N-acetylneuraminic acid (Neu5Ac), galactose, mannose, and fucose) and significantly (p < 0.05) alter infection. During infection, Salmonella used its two GHs sialidase nanH and amylase malS for internalization by targeting different glycan structures. The host glycans were altered during Salmonella association via the induction of N-glycan biosynthesis pathways leading to modification of host glycans by increasing fucosylation and mannose content, while decreasing sialylation. Gene expression analysis indicated that the host cell responded by regulating more than 50 genes resulting in remodeled glycans in response to Salmonella treatment. This study established the glycan structures on colonic epithelial cells, determined that Salmonella required two keystone GHs for internalization, and left remodeled host glycans as a result of infection. These data indicate that microbial GHs are undiscovered virulence factors.

Genomic Comparison of Campylobacter spp. and Their Potential for Zoonotic Transmission between Birds, Primates, and Livestock

ABSTRACT Campylobacter is the leading cause of human gastroenteritis worldwide. Wild birds, including American crows, are abundant in urban, suburban, and agricultural settings and are likely zoonotic vectors of Campylobacter. Their proximity to humans and livestock increases the potential spreading of Campylobacter via crows between the environment, livestock, and humans. However, no studies have definitively demonstrated that crows are a vector for pathogenic Campylobacter. We used genomics to evaluate the zoonotic and pathogenic potential of Campylobacter from crows to other animals with 184 isolates obtained from crows, chickens, cows, sheep, goats, humans, and nonhuman primates. Whole-genome analysis uncovered two distinct clades of Campylobacter jejuni genotypes; the first contained genotypes found only in crows, while a second genotype contained “generalist” genomes that were isolated from multiple host species, including isolates implicated in human disease, primate gastroenteritis, and livestock abortion. Two major β-lactamase genes were observed frequently in these genomes (oxa-184, 55%, and oxa-61, 29%), where oxa-184 was associated only with crows and oxa-61 was associated with generalists. Mutations in gyrA, indicative of fluoroquinolone resistance, were observed in 14% of the isolates. Tetracycline resistance (tetO) was present in 22% of the isolates, yet it occurred in 91% of the abortion isolates. Virulence genes were distributed throughout the genomes; however, cdtC alleles recapitulated the crow-only and generalist clades. A specific cdtC allele was associated with abortion in livestock and was concomitant with tetO. These findings indicate that crows harboring a generalist C. jejuni genotype may act as a vector for the zoonotic transmission of Campylobacter. IMPORTANCE This study examined the link between public health and the genomic variation of Campylobacter in relation to disease in humans, primates, and livestock. Use of large-scale whole-genome sequencing enabled population-level assessment to find new genes that are linked to livestock disease. With 184 Campylobacter genomes, we assessed virulence traits, antibiotic resistance susceptibility, and the potential for zoonotic transfer to observe that there is a “generalist” genotype that may move between host species.

Fallacy of the Unique Genome: Sequence Diversity within Single Helicobacter pylori Strains

ABSTRACT Many bacterial genomes are highly variable but nonetheless are typically published as a single assembled genome. Experiments tracking bacterial genome evolution have not looked at the variation present at a given point in time. Here, we analyzed the mouse-passaged Helicobacter pylori strain SS1 and its parent PMSS1 to assess intra- and intergenomic variability. Using high sequence coverage depth and experimental validation, we detected extensive genome plasticity within these H. pylori isolates, including movement of the transposable element IS607, large and small inversions, multiple single nucleotide polymorphisms, and variation in cagA copy number. The cagA gene was found as 1 to 4 tandem copies located off the cag island in both SS1 and PMSS1; this copy number variation correlated with protein expression. To gain insight into the changes that occurred during mouse adaptation, we also compared SS1 and PMSS1 and observed 46 differences that were distinct from the within-genome variation. The most substantial was an insertion in cagY, which encodes a protein required for a type IV secretion system function. We detected modifications in genes coding for two proteins known to affect mouse colonization, the HpaA neuraminyllactose-binding protein and the FutB α-1,3 lipopolysaccharide (LPS) fucosyltransferase, as well as genes predicted to modulate diverse properties. In sum, our work suggests that data from consensus genome assemblies from single colonies may be misleading by failing to represent the variability present. Furthermore, we show that high-depth genomic sequencing data of a population can be analyzed to gain insight into the normal variation within bacterial strains. IMPORTANCE Although it is well known that many bacterial genomes are highly variable, it is nonetheless traditional to refer to, analyze, and publish “the genome” of a bacterial strain. Variability is usually reduced (“only sequence from a single colony”), ignored (“just publish the consensus”), or placed in the “too-hard” basket (“analysis of raw read data is more robust”). Now that whole-genome sequences are regularly used to assess virulence and track outbreaks, a better understanding of the baseline genomic variation present within single strains is needed. Here, we describe the variability seen in typical working stocks and colonies of pathogen Helicobacter pylori model strains SS1 and PMSS1 as revealed by use of high-coverage mate pair next-generation sequencing (NGS) and confirmed by traditional laboratory techniques. This work demonstrates that reliance on a consensus assembly as “the genome” of a bacterial strain may be misleading. IMPORTANCE Although it is well known that many bacterial genomes are highly variable, it is nonetheless traditional to refer to, analyze, and publish “the genome” of a bacterial strain. Variability is usually reduced (“only sequence from a single colony”), ignored (“just publish the consensus”), or placed in the “too-hard” basket (“analysis of raw read data is more robust”). Now that whole-genome sequences are regularly used to assess virulence and track outbreaks, a better understanding of the baseline genomic variation present within single strains is needed. Here, we describe the variability seen in typical working stocks and colonies of pathogen Helicobacter pylori model strains SS1 and PMSS1 as revealed by use of high-coverage mate pair next-generation sequencing (NGS) and confirmed by traditional laboratory techniques. This work demonstrates that reliance on a consensus assembly as “the genome” of a bacterial strain may be misleading.

A Syst-OMICS Approach to Ensuring Food Safety and Reducing the Economic Burden of Salmonellosis

The Salmonella Syst-OMICS consortium is sequencing 4,500 Salmonella genomes and building an analysis pipeline for the study of Salmonella genome evolution, antibiotic resistance and virulence genes. Metadata, including phenotypic as well as genomic data, for isolates of the collection are provided through the Salmonella Foodborne Syst-OMICS database (SalFoS), at https://salfos.ibis.ulaval.ca/. Here, we present our strategy and the analysis of the first 3,377 genomes. Our data will be used to draw potential links between strains found in fresh produce, humans, animals and the environment. The ultimate goals are to understand how Salmonella evolves over time, improve the accuracy of diagnostic methods, develop control methods in the field, and identify prognostic markers for evidence-based decisions in epidemiology and surveillance.

Large-Scale Release of Campylobacter Draft Genomes: Resources for Food Safety and Public Health from the 100K Pathogen Genome Project.

ABSTRACT Campylobacter is a food-associated bacterium and a leading cause of foodborne illness worldwide, being associated with poultry in the food supply. This is the initial public release of 202 Campylobacter genome sequences as part of the 100K Pathogen Genome Project. These isolates represent global genomic diversity in the Campylobacter genus.

Draft Genome Sequences of Campylobacter jejuni Strains That Cause Abortion in Livestock

ABSTRACT Campylobacter jejuni is an intestinal bacterium that can cause abortion in livestock. This publication announces the public release of 15 Campylobacter jejuni genome sequences from isolates linked to abortion in livestock. These isolates are part of the 100K Pathogen Genome Project and are from clinical cases at the University of California (UC) Davis.

Comparative genomics reveals the diversity of restriction-modification systems and DNA methylation sites in Listeria monocytogenes

ABSTRACT Listeria monocytogenes is a bacterial pathogen that is found in a wide variety of anthropogenic and natural environments. Genome sequencing technologies are rapidly becoming a powerful tool in facilitating our understanding of how genotype, classification phenotypes, and virulence phenotypes interact to predict the health risks of individual bacterial isolates. Currently, 57 closed L. monocytogenes genomes are publicly available, representing three of the four phylogenetic lineages, and they suggest that L. monocytogenes has high genomic synteny. This study contributes an additional 15 closed L. monocytogenes genomes that were used to determine the associations between the genome and methylome with host invasion magnitude. In contrast to previous findings, large chromosomal inversions and rearrangements were detected in five isolates at the chromosome terminus and within rRNA genes, including a previously undescribed inversion within rRNA-encoding regions. Each isolates epigenome contained highly diverse methyltransferase recognition sites, even within the same serotype and methylation pattern. Eleven strains contained a single chromosomally encoded methyltransferase, one strain contained two methylation systems (one system on a plasmid), and three strains exhibited no methylation, despite the occurrence of methyltransferase genes. In three isolates a new, unknown DNA modification was observed in addition to diverse methylation patterns, accompanied by a novel methylation system. Neither chromosome rearrangement nor strain-specific patterns of epigenome modification observed within virulence genes were correlated with serotype designation, clonal complex, or in vitro infectivity. These data suggest that genome diversity is larger than previously considered in L. monocytogenes and that as more genomes are sequenced, additional structure and methylation novelty will be observed in this organism. IMPORTANCE Listeria monocytogenes is the causative agent of listeriosis, a disease which manifests as gastroenteritis, meningoencephalitis, and abortion. Among Salmonella, Escherichia coli, Campylobacter, and Listeria—causing the most prevalent foodborne illnesses—infection by L. monocytogenes carries the highest mortality rate. The ability of L. monocytogenes to regulate its response to various harsh environments enables its persistence and transmission. Small-scale comparisons of L. monocytogenes focusing solely on genome contents reveal a highly syntenic genome yet fail to address the observed diversity in phenotypic regulation. This study provides a large-scale comparison of 302 L. monocytogenes isolates, revealing the importance of the epigenome and restriction-modification systems as major determinants of L. monocytogenes phylogenetic grouping and subsequent phenotypic expression. Further examination of virulence genes of select outbreak strains reveals an unprecedented diversity in methylation statuses despite high degrees of genome conservation.

100K Pathogen Genome Project: 306 Listeria draft genome sequences for food safety and public health

ABSTRACT Listeria monocytogenes is a food-associated bacterium that is responsible for food-related illnesses worldwide. This is the initial public release of 306 L. monocytogenes genome sequences as part of the 100K Pathogen Genome Project. These isolates represent global genomic diversity in L. monocytogenes.