na INLIS000000000014376 20240213024808 0010-0521014376 240213 | | 0161-5890 en NONE MIVol70th2016p140 Kingston, Joseph J. Shreya, Das Majumder, Saugata Generation and characterization of recombinant bivalent fusionprotein r-Cpib for immunotherapy against Clostridium perfringens betaand iota toxemia / Kingston, Joseph J. Elsevier Publishing, 2016 tClostridium perfringens beta (CPB) and iota (CPI) toxaemias result in some of the most lethal forms ofhaemorrhagic and necrotic enteritis and sudden death syndrome affecting especially neonates. WhileCPB enterotoxemia is one of the most common forms of clostridial enterotoxemia, CPI enterotoxemiathough putatively considered to be rare is an emerging cause of concern. The similarities in clinical man-ifestation, gross and histopathology findings of both types of toxaemias coupled to the infrequency of CPItoxaemia might lead to symptomatic misidentification with Type C resulting in therapeutic failure due tohabitual administration of CPB anti-toxin which is ineffective against CPI. Therefore in the present study,to generate a composite anti-toxin capable of neutralizing both toxaemias, a novel bivalent chimera r-Cpib was constructed by splicing the non-toxic C terminal binding regions of CPB and CPI, via a flexibleglycine linker (G4S) by overlap-extension PCR. The fusion protein was characterized for its therapeuticabilities toward CPI and CPB toxin neutralizations. The r-Cpib was found to be non-toxic and could com-petitively inhibit binding of CPB to host cell receptors thereby reducing its cytotoxicity. Immunization ofmice with r-Cpib generated specific antibodies capable of neutralizing the above toxaemias both in vitroand in vivo. Caco-2 cells exposed to a mixture of anti-r-Cpib sera and native CPI or CPB, displayed signif-icantly superior protection against the respective toxins while passive challenge of mice with a similarmixture resulted in 83 and 91% protection against CPI and CPB respectively. Alternatively, mice exposedto a mixture of sham sera and native toxins died within 2–3 days. This work thus demonstrates r-Cpibas a novel bivalent fusion protein capable of efficient immunotherapy against C. perfringens CPI and CPBtoxaemia. CLOSTRIDIUM PERFRINGENS -- TOXINS -- IMMUNOTHERAPY