Galleria mellonella Infection Model Identifies Both High and Low Lethality of Clostridium perfringens Toxigenic Strains and Their Response to Antimicrobials

Research progress into mechanisms of the anaerobe Clostridium perfringens and associated diseases has been frustrated by the lack of reliable infection models. Wax moth larvae (Galleria mellonella) have emerged as a viable alternative to other models of infection since they are economic, survive at 37�C and require no specialist equipment. This study aims to establish to what extent G. mellonella larvae can be used to study the virulence of C. perfringens strains and its suitability for studying novel treatment strategies by an improved time-lapse approach to data collection. Mortality and morbidity rates of larvae challenged with 105 CFU of C. perfringens isolates from various sources were observed over 72 h and dose response data obtained. Phenoloxidase enzyme activity was investigated as a marker for immune response and tissue burden assessed by histopathological techniques. Results demonstrate that C. perfringens is pathogenic toward G. mellonella although potency varies dramatically between C. perfringens isolates and the reference strain ATCC 13124 was shown to be avirulent. Infection with C. perfringens strains activated the melanisation pathway resulting in melanin deposition but no increase in enzyme activity was observed. Efficacy of antibiotic therapy (penicillin G, bacitracin, neomycin, and tetracycline) administered parenterally to some extent correlates with that of in vitro analysis. The findings suggest G. mellonella might be a useful in vivo model of infection and convenient as a prescreening assay for virulence of C. perfringens strains or as a simple, cheap and rapid in vivo assay in the first stage development of novel therapeutics against anaerobes

Crystal structure of the ADP-ribosylating component of BEC, the binary enterotoxin of Clostridium perfringens

AbstractBinary enterotoxin of Clostridium perfringens (BEC), consisting of the components BECa and BECb, was recently identified as a novel enterotoxin produced by C. perfringens that causes acute gastroenteritis in humans. Although the detailed mechanism of cell intoxication by BEC remains to be defined, BECa shows both NAD+-glycohydrolase and actin ADP-ribosyltransferase activities in the presence of NAD+. In this study, we determined the first crystal structure of BECa in its apo-state and in complex with NADH. The structure of BECa shows striking resemblance with other binary actin ADP-ribosylating toxins (ADPRTs), especially in terms of its overall protein fold and mechanisms of substrate recognition. We present a detailed picture of interactions between BECa and NADH, including bound water molecules located near the C1′-N glycosidic bond of NADH and the catalytically important ADP-ribosylating turn-turn (ARTT) loop. We observed that the conformational rearrangement of the ARTT loop, possibly triggered by a conformational change involving a conserved tyrosine residue coupled with substrate binding, plays a crucial role in catalysis by properly positioning a catalytic glutamate residue in the E-X-E motif of the ARTT loop in contact with the nucleophile. Our results for BECa provide insight into the common catalytic mechanism of the family of binary actin ADPRTs

Dissemination of Extended-Spectrum β-Lactamase- and AmpC β-Lactamase-Producing Escherichia coli within the Food Distribution System of Ho Chi Minh City, Vietnam

To investigate the dissemination of ESBL/pAmpC-producing E. coli within the food distribution system of Ho Chi Minh City (HCMC), Vietnam, the prevalence of ESBL/pAmpC-producing E. coli strains in chicken meat, pork, beef, and fish/shrimp samples obtained from slaughterhouses, a wholesale market, and supermarkets was examined. Among the total of 330 collected food samples, ESBL/pAmpC-producing E. coli was detected in 150 samples (45.5%). The highest prevalence of these isolates was in chicken meat (76/82, 92.7%), followed by pork (32/92, 34.8%), beef (18/74, 34.3%), and fish/shrimp (24/82, 29.3%). A total of 342 strains of ESBL/pAmpC-producing E. coli were isolated from 150 positive food samples. The most prevalent genes responsible for ESBL or pAmpC activity belonged to the CTX-M-9 (110/342, 31.2%), CTX-M-1 (102/342, 29.8%), and CIT (118/342, 34.5%) groups. To our knowledge, this is the first report of the high occurrence of pAmpC (37.1%) in animal-based food in Vietnam. Among the 342 total ESBL/pAmpC-producing E. coli isolates, 276 (80.7%) were resistant to at least 6 antibiotic agents. Notably, high percentages of resistance to ciprofloxacin and fosfomycin were found in isolates from chicken (80.5% and 50.8%, resp.). These findings demonstrate that animal-based food products in HCMC represent a major reservoir of ESBL/pAmpC-producing E. coli