Dam deletion mutant of Salmonella Enteritidis: studies on virulence and protection

NOTO LLANA M, SARNACKI SH, GIACOMODONATO MN, CERQUETTI MC

Rosario, Santa Fe, Argentina

Congreso; XLII Reunion Anual de la Sociedad Argentina de Investigacion en Bioquimica y Biologia; 2006

Sociedad Argentina de Investigacion en Bioquimica y Biologia

S. Typhimurium null dammutants have been proposed as ideal vaccine strains. In contrast, S.Enteritidis mutants bearing a defective Dam protein are less protective and unable to agglutinate in the presence of antibodies against LPS O9 antigen. Here, we analyze the behaviour of SEÄdam, a damdeletion mutant of S. Enteritidis. This mutant was constructed by PCR-based mutagenesis. Attenuation and protection studies were performed in BALB/c mice. Epithelial and macrophage cell lines were used for in vitro studies. Nitric oxide (NO) production was assessed using Griess reagent. We found that SEÄdam mutant is moderately attenuated (LD 50 :107 cfu), induces significant  delayed-type hypersensitivity and improves bacterial clearance from spleen of immunized mice (range:44-200 vs. 4 x 105 - 2 x 108 cfu). However, its protective capacity is reduced (20 % survival rate). In vitro SEÄdam induces production of NO in macrophages but not in Hep2; concurrently, we found that it is unable to invade epithelial cells. In addition, SEÄdam LPS presents high proportion of reduced length O antigen polysaccharide. This defect does not affect the mutant serum resistance. Our results suggest that deletion of damgene in S.Enteritidis renders a modestly attenuated mutant with reduced protective capacity. Interestingly, we found that Dam protein is involved in the regulation of LPS synthesis.