Diarrhea, including soft and loose stools, was observed in three guinea-pigs among two groups of this study, but no animal developed diarrhea that was persistent or severe. The morphological changes observed in the colonic mucosa of the virulent Shigella-infected

guinea-pigs are characteristic of an acute inflammatory response check details with progressively increasing severity during the first 48 h of observation, whereas such results were not seen in the avirulent challenge group. However, after 72–96-h postinfection, the severity tended to decline, finally disappearing after 120 h (data not shown). Studies with candidate live invasive and noninvasive Shigella vaccines showed that underattenuation was responsible for excessive reactogenicity and overattenuation led to the poor immunogenicity in humans. In this respect, the killed vaccines are getting importance and gaining confidence (Chakrabarti et al., 1999; Sur et al., 2009). The efficacy study showed complete protection against wild-type S. dysenteriae 1 (NT4907) and S. flexneri 2a (B294) after four doses of oral immunization with heat-killed see more shigellae. Significantly higher

levels of lipopolysaccharide-specific IgG and IgA antibodies were detected in both serum and mucosal secretions of immunized guinea-pigs. During oral immunization, an exponential increase of serum IgG was also observed. The protective immunity to shigellae may be conferred by serum IgG antibodies to the O-specific polysaccharide of their lipopolysaccharide (Robbins et al., 1992). Although the bacterial colonization was detected in the distal colon of immunized animals, their levels were far lower when compared also with the control group. Histopathological

features of the distal colon also revealed protection against homologous virulent live Shigella. Over the years, several approaches have been explored using mice, guinea-pigs, rabbits, macaques and piglets as a suitable animal model for shigellosis. The mice model of pulmonary pneumonia with the intranasal inoculation of Shigella (Voino-Yasenetsky & Voino-Yasenetskaya, 1962) was used to determine the virulence attenuation, immunization efficacy and protection against infection (Mallett et al., 1995). However, this model lacked clinical relevance with respect to the infection site of the pathogen. Fernandez et al. (2003) demonstrated a murine infection model with newborn mice in which inflammatory destruction of the mucosa and substantial infiltration of polymorphonuclear neutrophils into the gut were observed. Because of the narrow window of time (3–4 days after birth), this model was not applicable for the evaluation of protective immunity.