cjrABC-senB hinders survival of extraintestinal pathogenic E. coli in the bloodstream through triggering complement-mediated killing.

BACKGROUND: Extraintestinal pathogenic E. coli (ExPEC) is a common gram-negative organism causing various infections, including urinary tract infections (UTIs), bacteremia, and neonatal meningitis. The cjrABC-senB gene cluster of E. coli contributes to ExPEC virulence in the mouse model of UTIs. Consistently, the distribution of cjrABC-senB is epidemiologically associated with human UTIs caused by E. coli. cjrABC-senB, which has previously been proposed to encode an iron uptake system, may facilitate ExPEC survival in the iron availability-restricted urinary tract. Given that the bloodstream is also an iron limited environment to invading bacteria, the pathogenic role of cjrABC-senB in ExPEC bacteremia, however, remains to be investigated. METHODS: The ability of ExPEC RS218 strains with and without cjrABC-senB to survive in the mouse bloodstream and human serum was evaluated. Subsequently, the role of this gene cluster in the ExPEC interaction with the complement system was evaluated. Finally, the distribution of cjrABC-senB in human clinical E. coli isolates was determined by PCR. The frequency of cjrABC-senB in bacteremia isolates that were not associated with UTIs (non-UTI bacteremia isolates) was compared with that in UTI-associated isolates and fecal isolates. RESULTS: Expression of cjrABC-senB attenuated the survival of RS218 in the mouse bloodstream and human serum. The cjrABC-senB-harboring strains triggered enhanced classical- and alternative-complement pathway activation and became more vulnerable to complement-mediated killing in serum. cjrA was identified as the major gene responsible for the attenuated serum survival. Expressing cjrABC-senB and cjrA increased bacterial susceptibility to detergent and induced periplasmic protein leakage, suggesting that the expression of these genes compromises the integrity of the outer membrane of ExPEC. In addition, the frequency of cjrABC-senB in non-UTI bacteremia isolates was significantly lower than that in UTI-associated isolates, while the frequencies in non-UTI bacteremia isolates and fecal isolates showed no significant difference. Consistently, this epidemiological investigation suggests that cjrABC-senB does not contribute to E. coli bacteremia in humans. CONCLUSION: The contribution of cjrABC-senB to the pathogenesis of ExPEC is niche dependent and contradictory because the genes facilitate ExPEC UTIs but hinder bacteremia. The contradictory niche-dependent characteristic may benefit the development of novel strategies against E. coli-caused infections.

Genetic structure, antimicrobial resistance and frequency of human associated Escherichia coli sequence types among faecal isolates from healthy dogs and cats living in Canberra, Australia.

Extraintestinal pathogenic Escherichia coli (ExPEC) cause clinical infections in humans. Understanding the evolution and dissemination of ExPEC strains via potential reservoirs is important due to associated morbidity, health care costs and mortality. To further understanding this survey has examined isolates recovered from the faeces of 221 healthy dogs and 427 healthy cats. The distribution of phylogroups varied with host species, and depended on whether the animal was living in a shelter or a home. The human associated STs 69, 73, 95, 131 and 127 were prevalent, with 30.5% of cat isolates and 10.3% of dog isolates representing these ExPEC sequence types. Resistance to the antibiotics ampicillin and tetracycline was common, but resistance to other antimicrobials was negligible.

Escherichia coli Pathobionts Associated with Inflammatory Bowel Disease.

Gut bacteria play a key role in initiating and maintaining the inflammatory process in the gut tissues of inflammatory bowel disease (IBD) patients, by supplying antigens or other stimulatory factors that trigger immune cell activation. Changes in the composition of the intestinal microbiota in IBD patients compared to that in healthy controls and a reduced diversity of intestinal microbial species are linked to the pathogenesis of IBD. Adherent invasive Escherichia coli (AIEC) has been linked to Crohn's disease (CD) patients, while diffusely adherent E. coli (DAEC) has been associated with ulcerative colitis (UC). Bacteriological analysis of intestinal biopsy specimens and fecal samples from IBD patients shows an increased number of E. coli strains belonging to the B2 phylogenetic group, which are typically known as extraintestinal pathogenic E. coli (ExPEC). Results from studies of both cell cultures and animal models reveal pathogenic features of these E. coli pathobionts, which may link them to IBD pathogenesis. This suggests that IBD-associated E. coli strains play a facilitative role during IBD flares. In this review, we explain IBD-associated E. coli and its role in IBD pathogenesis.

Novel small molecules affecting cell membrane as potential therapeutics for avian pathogenic Escherichia coli.

Avian pathogenic Escherichia coli (APEC), a most common bacterial pathogen of poultry, causes multiple extra-intestinal diseases in poultry which results in significant economic losses to the poultry industry worldwide. In addition, APEC are a subgroup of extra-intestinal pathogenic E. coli (ExPEC), and APEC contaminated poultry products are a potential source of foodborne ExPEC infections to humans and transfer of antimicrobial resistant genes. The emergence of multi-drug resistant APEC strains and the limited efficacy of vaccines necessitate novel APEC control approaches. Here, we screened a small molecule (SM) library and identified 11 SMs bactericidal to APEC. The identified SMs were effective against multiple APEC serotypes, biofilm embedded APEC, antimicrobials resistant APECs, and other pathogenic E. coli strains. Microscopy revealed that these SMs affect the APEC cell membrane. Exposure of SMs to APEC revealed no resistance. Most SMs showed low toxicity towards chicken and human cells and reduced the intracellular APEC load. Treatment with most SMs extended the wax moth larval survival and reduced the intra-larval APEC load. Our studies could facilitate the development of antimicrobial therapeutics for the effective management of APEC infections in poultry as well as other E. coli related foodborne zoonosis, including APEC related ExPEC infections in humans.

O-specific polysaccharide confers lysozyme resistance to extraintestinal pathogenic Escherichia coli.

Extraintestinal pathogenic Escherichia coli (ExPEC) is the leading cause of bloodstream and other extraintestinal infections in human and animals. The greatest challenge encountered by ExPEC during an infection is posed by the host defense mechanisms, including lysozyme. ExPEC have developed diverse strategies to overcome this challenge. The aim of this study was to characterize the molecular mechanism of ExPEC resistance to lysozyme. For this, 15,000 transposon mutants of a lysozyme-resistant ExPEC strain NMEC38 were screened; 20 genes were identified as involved in ExPEC resistance to lysozyme-of which five were located in the gene cluster between galF and gnd, and were further confirmed to be involved in O-specific polysaccharide biosynthesis. The O-specific polysaccharide was able to inhibit the hydrolytic activity of lysozyme; it was also required by the complete lipopolysaccharide (LPS)-mediated protection of ExPEC against the bactericidal activity of lysozyme. The O-specific polysaccharide was further shown to be able to directly interact with lysozyme. Furthermore, LPS from ExPEC strains of different O serotypes was also able to inhibit the hydrolytic activity of lysozyme. Because of their cell surface localization and wide distribution in Gram-negative bacteria, O-specific polysaccharides appear to play a long-overlooked role in protecting bacteria against exogenous lysozyme.

Metals Enhance the Killing of Bacteria by Bacteriophage in Human Blood.

Multidrug-resistant bacterial pathogens are a major medical concern. E. coli, particularly the pathotype extraintestinal pathogenic E. coli (ExPEC), is a leading cause of bloodstream infections. As natural parasites of bacteria, bacteriophages are considered a possible solution to treat patients infected with antibiotic resistant strains of bacteria. However, the development of phage as an anti-infective therapeutic is hampered by limited knowledge of the physiologic factors that influence their properties in complex mammalian environments such as blood. To address this barrier, we tested the ability of phage to kill ExPEC in human blood. Phages are effective at killing ExPEC in conventional media but are substantially restricted in this ability in blood. This phage killing effect is dependent on the levels of free metals and is inhibited by the anticoagulant EDTA. The EDTA-dependent inhibition of ExPEC killing is overcome by exogenous iron, magnesium, and calcium. Metal-enhanced killing of ExPEC by phage was observed for several strains of ExPEC, suggesting a common mechanism. The addition of metals to a murine host infected with ExPEC stimulated a phage-dependent reduction in ExPEC levels. This work defines a role for circulating metals as a major factor that is essential for the phage-based killing of bacteria in blood.

Role for FimH in Extraintestinal Pathogenic Escherichia coli Invasion and Translocation through the Intestinal Epithelium.

The translocation of bacteria across the intestinal epithelium of immunocompromised patients can lead to bacteremia and life-threatening sepsis. Extraintestinal pathogenic Escherichia coli (ExPEC), so named because this pathotype infects tissues distal to the intestinal tract, is a frequent cause of such infections, is often multidrug resistant, and chronically colonizes a sizable portion of the healthy population. Although several virulence factors and their roles in pathogenesis are well described for ExPEC strains that cause urinary tract infections and meningitis, they have not been linked to translocation through intestinal barriers, a fundamentally distant yet important clinical phenomenon. Using untransformed ex situ human intestinal enteroids and transformed Caco-2 cells, we report that ExPEC strain CP9 binds to and invades the intestinal epithelium. ExPEC harboring a deletion of the gene encoding the mannose-binding type 1 pilus tip protein FimH demonstrated reduced binding and invasion compared to strains lacking known E. coli virulence factors. Furthermore, in a murine model of chemotherapy-induced translocation, ExPEC lacking fimH colonized at levels comparable to that of the wild type but demonstrated a statistically significant reduction in translocation to the kidneys, spleen, and lungs. Collectively, this study indicates that FimH is important for ExPEC translocation, suggesting that the type 1 pilus is a therapeutic target for the prevention of this process. Our study also highlights the use of human intestinal enteroids in the study of enteric diseases.

Characterization and distinction of two flagellar systems in extraintestinal pathogenic Escherichia coli PCN033.

Extraintestinal pathogenic Escherichia coli (ExPEC) can invade and colonize multiple extraintestinal tissues and can cause a wide range of infections; however the mechanisms of its pathogenicity are not well understood. Flagella contribute to the infection of E. coli strains by mediating adhesion and invasion. Our previous bioinformatic analysis revealed two flagella gene clusters in the genome of an ExPEC isolate, PCN033. One encodes the conventional flagellum system (Flag-1) and the other encodes the Flag-2 system, whose function is uncharacterized. Here we aimed to characterize these two flagellum systems and determine their contributions to the flagellum formation and certain pathogenicity-associated phenotypes. Our observations support the involvement of Flag-1 system, but not Flag-2 system, in the synthesis and maturation of the flagellum structure, and in mediating bacterial swimming and swarming. Moreover, flgD, which encodes a flagellar-hook scaffolding protein in the Flag-1 system, is required for flagellum assembly by influencing the production of FliC (flagellin). Deletion of flgD attenuated ExPEC strain PCN033 invasion and colonization in vivo, probably by affecting bacterial adhesion and invasion, and by reducing resistance to phagocytosis by circulating monocytes. In contrast, these phenotypes were not observed in the strain with deletion of lfgD, encoding the FlgD-like protein in the Flag-2 system. Taken together, these findings indicate that Flag-1 flagellum system is the determinative component of bacterial flagella that contributes to the infection.

Oxygen-Free Condition Inhibited Biofilm Formation in Extraintestinal Pathogenic Escherichia coli Strain PPECC42 Through Preventing Curli Production.

Extraintestinal pathogenic Escherichia coli (ExPEC) is an important zoonotic and foodborne pathogen. Biofilms are specially structured communities for bacteria to survive in different hostile environments and can protect the bacteria from eradication by the host and external factors. In this study, we found that oxygen is definitely required for biofilm formation in ExPEC strain PPECC42. Aerobically growing ExPEC showed a bdar (brown, dry, and rough) morphotype, whereas anaerobically growing ExPEC showed a saw (smooth and white) morphotype. Under anaerobic condition, curli fimbriae did not accumulate and the expression levels of curli biosynthesis-related genes including csgB, csgD, and rpoS decreased significantly; in contrast, the expression level of h-ns, of which the encoding protein is a repressor for csgD transcription, increased significantly. Taken together, the results suggested that oxygen-free condition limited ExPEC strain PPECC42 biofilm formation mainly through preventing curli accumulation by affecting the transcriptional levels of curli biosynthesis-related genes.