BackgroundUnsafe water supplies continue to raise public health concerns, especially in urban areas in low resource countries. To understand the extent of public health risk attributed to supply water in Dhaka city, Bangladesh, Escherichia coli isolated from tap water samples collected from different locations of the city were characterized for their antibiotic resistance, pathogenic properties and genetic diversity.Methodology/Principal FindingsA total of 233 E. coli isolates obtained from 175 tap water samples were analysed for susceptibility to 16 different antibiotics and for the presence of genes associated with virulence and antibiotic resistance. Nearly 36% (n = 84) of the isolates were multi-drug(≥3 classes of antibiotics) resistant (MDR) and 26% (n = 22) of these were positive for extended spectrum β-lactamase (ESBL). Of the 22 ESBL-producers, 20 were positive for bla CTX-M-15, 7 for bla OXA-1-group (all had bla OXA-47) and 2 for bla CMY-2. Quinolone resistance genes, qnrS and qnrB were detected in 6 and 2 isolates, respectively. Around 7% (n = 16) of the isolates carried virulence gene(s) characteristic of pathogenic E. coli; 11 of these contained lt and/or st and thus belonged to enterotoxigenic E. coli and 5 contained bfp and eae and thus belonged to enteropathogenic E. coli. All MDR isolates carried multiple plasmids (2 to 8) of varying sizes ranging from 1.2 to >120 MDa. Ampicillin and ceftriaxone resistance were co-transferred in conjugative plasmids of 70 to 100 MDa in size, while ampicillin, trimethoprim-sulfamethoxazole and tetracycline resistance were co-transferred in conjugative plasmids of 50 to 90 MDa. Pulsed-field gel electrophoresis analysis revealed diverse genetic fingerprints of pathogenic isolates.SignificanceMulti-drug resistant E. coli are wide spread in public water supply in Dhaka city, Bangladesh. Transmission of resistant bacteria and plasmids through supply water pose serious threats to public health in urban areas.

The main objective of this study was to investigate the prevalence of bla (NDM-1) in Gram-negative bacteria in Bangladesh. In October 2010 at the International Centre for Diarrhoeal Disease Research, Bangladesh (ICDDR,B) laboratories, 1,816 consecutive clinical samples were tested for imipenem-resistant Gram-negative organisms. Imipenem-resistant isolates were tested for the bla (NDM-1) gene. Among 403 isolates, 14 (3.5 %) were positive for bla (NDM-1), and the predominant species were Klebsiella pneumoniae, Acinetobacter baumannii, and Escherichia coli. All bla (NDM-1)-positive isolates were resistant to multiple antibiotics. Among β-lactamase genes, bla (CTX-M-1-group) was detected in ten isolates (eight bla (CTX-M-15)), bla (OXA-1-group) in six, bla (TEM) in nine, bla (SHV) in seven, and bla (VIM) and bla (CMY) in two isolates each. The 16S rRNA methylase gene, armA, was detected in five K. pneumoniae isolates and in one E. coli isolate. rmtB and rmtC were detected in a Citrobacter freundii and two K. pneumoniae isolates, respectively. qnr genes were detected in two K. pneumoniae isolates (one qnrB and one qnrS) and in an E. coli isolate (qnrA). Transferable plasmids (60-100 MDa) carrying bla (NDM-1) were detected in 7 of the 11 plasmid-containing isolates. Pulsed-field gel electrophoresis (PFGE) analysis grouped K. pneumoniae isolates into three clusters, while E. coli isolates differed significantly from each other. This study reports that approximately 3.5 % of Gram-negative clinical isolates in Bangladesh are NDM-1-producing.

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Campylobacter jejuni, a foodborne pathogen, is one of the most common bacterial causes of gastroenteritis in the world. Undercooked poultry, raw (unpasteurized) dairy products, untreated water, and contaminated produce are the most common sources associated with infection. C. jejuni establishes a niche in the gut by adhering to and invading epithelial cells, which results in diarrhea with blood and mucus in the stool. The process of colonization is mediated, in part, by surface-exposed molecules (adhesins) that bind directly to host cell ligands or the extracellular matrix (ECM) surrounding cells. In this review, we introduce the known and putative adhesins of the foodborne pathogen C. jejuni. We then focus our discussion on two C. jejuni Microbial Surface Components Recognizing Adhesive Matrix Molecule(s) (MSCRAMMs), termed CadF and FlpA, which have been demonstrated to contribute to C. jejuni colonization and pathogenesis. In vitro studies have determined that these two surface-exposed proteins bind to the ECM glycoprotein fibronectin (FN). In vivo studies have shown that cadF and flpA mutants exhibit impaired colonization of chickens compared to the wildtype strain. Additional studies have revealed that CadF and FlpA stimulate epithelial cell signaling pathways necessary for cell invasion. Interestingly, CadF and FlpA have distinct FN-binding domains, suggesting that the functions of these proteins are nonredundant. In summary, the binding of FN by C. jejuni CadF and FlpA adhesins has been demonstrated to contribute to adherence, invasion, and cell signaling.

This work analyzes the high-pressure (HP) germination of spores of the food-borne pathogen Clostridium perfringens (with inner membrane [IM] germinant receptors [GRs]) and the opportunistic pathogen Clostridium difficile (with no IM GRs), which has growing implications as an emerging food safety threat. In contrast to those of spores of Bacillus species, mechanisms of HP germination of clostridial spores have not been well studied. HP treatments trigger Bacillus spore germination through spores' IM GRs at ϳ150 MPa or through SpoVA channels for release of spores' dipicolinic acid (DPA) at >400 MPa, and DPA-less spores have lower wet heat resistance than dormant spores. We found that C. difficile spores exhibited no germination events upon 150-MPa treatment and were not heat sensitized. In contrast, 150-MPa-treated unactivated C. perfringens spores released DPA and became heat sensitive, although most spores did not complete germination by fully rehydrating the spore core, but this treatment of heat-activated spores led to almost complete germination and greater heat sensitization. Spores of both clostridial organisms released DPA during 550-MPa treatment, but C. difficile spores did not complete germination and remained heat resistant. Heat-activated 550-MPa-HP-treated C. perfringens spores germinated almost completely and became heat sensitive. However, unactivated 550-MPa-treated C. perfringens spores did not germinate completely and were less heat sensitive than spores that completed germination. Since C. difficile and C. perfringens spores use different mechanisms for sensing germinants, our results may allow refinement of HP methods for their inactivation in foods and other applications and may guide the development of commercially sterile low-acid foods. IMPORTANCESpores of various clostridial organisms cause human disease, sometimes due to food contamination by spores. Because of these spores' resistance to normal decontamination regimens, there is continued interest in ways to kill spores without compromising food quality. High hydrostatic pressure (HP) under appropriate conditions can inactivate bacterial spores. With growing use of HP for food pasteurization, advancement of HP for commercial production of sterile low-acid foods requires understanding of mechanisms of spores' interactions with HP. While much is known about HP germination and inactivation of spores of Bacillus species, how HP germinates and inactivates clostridial spores is less well understood. In this work we have tried to remedy this information deficit by examining germination of spores of Clostridium difficile and Clostridium perfringens by several HP and temperature levels. The results may give insight that could facilitate more efficient methods for spore eradication in food sterilization or pasteurization, biodecontamination, and health care.

Campylobacter jejuni, a zoonotic pathogen that frequently colonizes poultry, possesses two Microbial Surface Components Recognizing Adhesive Matrix Molecule(s) (MSCRAMMs) termed CadF and FlpA that bind to the glycoprotein fibronectin (FN). Previous to this study, it was not known whether the CadF and FlpA proteins were functionally redundant or if both were required to potentiate host cell binding and signaling processes. We addressed these questions by generating a complete repertoire of cadF and flpA mutants and complemented isolates, and performing multiple phenotypic assays. Both CadF and FlpA were found to be necessary for the maximal binding of C. jejuni to FN and to host cells. In addition, both CadF and FlpA are required for the delivery of the C. jejuni Cia effector proteins into the cytosol of host target cells, which in turn activates the MAPK signaling pathway (Erk 1/2) that is required for the C. jejuni invasion of host cells. These data demonstrate the non-redundant and bi-functional nature of these two C. jejuni FN-binding proteins. Taken together, the C. jejuni CadF and FlpA adhesins facilitate the binding of C. jejuni to the host cells, permit delivery of effector proteins into the cytosol of a host target cell, and aid in the rewiring of host cell signaling pathways to alter host cell behavior.

Clostridium perfringens is an important pathogen to human and animals and causes a wide array of diseases, including histotoxic and gastrointestinal illnesses. C. perfringens spores are crucial in terms of the pathogenicity of this bacterium because they can survive in a dormant state in the environment and return to being live bacteria when they come in contact with nutrients in food or the human body. Although the strategies to inactivate C. perfringens vegetative cells are effective, the inactivation of C. perfringens spores is still a great challenge. A number of studies have been conducted in the past decade or so toward developing efficient inactivation strategies for C. perfringens spores and vegetative cells, which include physical approaches and the use of chemical preservatives and naturally derived antimicrobial agents. In this review, different inactivation strategies applied to control C. perfringens cells and spores are summarized, and the potential limitations and challenges of these strategies are discussed.

Major foodborne bacterial pathogens, such as Campylobacter jejuni , have devised complex strategies to establish and foster intestinal infections. For more than two decades, researchers have used immortalized cell lines derived from human intestinal tissue to dissect C. jejuni -host cell interactions. Known from these studies is that C. jejuni virulence is multifactorial, requiring a coordinated response to produce virulence factors that facilitate host cell interactions. This study was initiated to identify C. jejuni proteins that contribute to adaptation to the host cell environment and cellular invasion. We demonstrated that C. jejuni responds to INT 407 and Caco-2 cells in a similar fashion at the cellular and molecular levels. Active protein synthesis was found to be required for C. jejuni to maximally invade these host cells. Proteomic and transcriptomic approaches were then used to define the protein and gene expression profiles of C. jejuni co-cultured with cells. By focusing on those genes showing increased expression by C. jejuni when co-cultured with epithelial cells, we discovered that C. jejuni quickly adapts to co-culture with epithelial cells by synthesizing gene products that enable it to acquire specific amino acids for growth, scavenge for inorganic molecules including iron, resist reactive oxygen/nitrogen species, and promote host cell interactions. Based on these findings, we selected a subset of the genes involved in chemotaxis and the regulation of flagellar assembly and generated C. jejuni deletion mutants for phenotypic analysis. Binding and internalization assays revealed significant differences in the interaction of C. jejuni chemotaxis and flagellar regulatory mutants. The identification of genes involved in C. jejuni adaptation to culture with host cells provides new insights into the infection process.