Intra-strain Variation of Lipopolysaccharides from Citrobacter freundii E9750 by in vitro Passages


Rosa E. Reyes*, Hugo Ramírez Saad , Julia Pérez , Enrique Meléndez , Armando Navarro Chiharu Murata and Alejandro Cravioto

The genus Citrobacter is closely related to Salmonella and Escherichia coli, and there are some serological cross-reactions existing among these species. These cross-reactivities are caused by Ospecific polysaccharide antigens in the LPS, which contain similar epitopes. In some cross-reactive strains, O-specific polysaccharides exhibit structural homology. In order to determinate the frequency of intra-strain variation of lipopolysaccharides (LPS) from Citrobacter freundii E9750 by in vitro passages at two different growth temperatures, isolates obtained from Citrobacter freundii E9750 in vitro passages at 37 and 42°C were analyzed for carbohydrate concentration, reactions to serological tests, and heterogeneity of lipopolysaccharides using SDS-PAGE. The genetic relatedness of the isolates was investigated using the PCR-RFLP profile of rfb and Pulsed Field Gel Electrophoresis (PFGE) profiles. Variation in expression of LPS was observed as a change in the concentration of carbohydrates. In addition, low reactivity with C. freundii E9750 O-antiserum, serological cross reactivity with S. senftenberg O-antiserum, and heterogeneity in electrophoretic mobility, were found. The PFGE and RFLP patterns of all isolates were indistinguishable from each other, including those that presented a serological reaction with the O -antigen sera from S. senftenberg. C. freundii E9750 NCTC isolated strains showed a high frequency in LPS variability. Analysis of the results suggested that heterogeneity was due to changes in the structure of the LPS carbohydrates. Since lipopolysaccharides are important factors in virulence and determine the basis for serotyping, the high frequency of intra-strain O-antigen heterogeneity in C. freundii E9750 is important

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