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 10 5 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 pre-screening 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. HIGHLIGHTS Potential novel in vivo model for the study of Clostridium perfringens infection. Novel time-lapse approach to data collection. First report of the pathogenicity of C. perfringens toward G. mellonella . First report of the efficacy of antibiotic therapy in response to C. perfringens infection in G. mellonella .

23Important research progress into the mechanisms of Clostridium perfringens associated 24 diseases (CPAD) has been slowed by the lack of a reliable infection model. Wax moth 25 larvae (Galleria mellonella) have emerged as a viable alternative to traditional mammalian 26 organisms since they are economic, survive at 37°C and require no specialist equipment. 27This study aims to establish whether G. mellonella larvae can be developed as a viable 28 model for the study of CPAD and their suitability for studying novel treatment strategies. In 29 addition, the study demonstrates a novel time-lapse approach to data collection. Mortality 30 and morbidity rates of larvae challenged with 10 5 CFU of C. perfringens isolates from 31 various sources were observed over 72h and dose response data obtained using inoculum 32 sizes of 10 -10 5