Abstract
BackgroundThe fliC and fljB genes in Salmonella code for the phase 1 (H1) and phase 2 (H2) flagellin respectively, the rfb cluster encodes the majority of enzymes for polysaccharide (O) antigen biosynthesis, together they determine the antigenic profile by which Salmonella are identified. Sequencing and characterisation of fliC was performed in the development of a molecular serotyping technique.ResultsFliC sequencing of 106 strains revealed two groups; the g-complex included those exhibiting "g" or "m,t" antigenic factors, and the non-g strains which formed a second more diverse group. Variation in fliC was characterised and sero-specific motifs identified. Furthermore, it was possible to identify differences in certain H antigens that are not detected by traditional serotyping. A rapid short sequencing assay was developed to target serotype-specific sequence motifs in fliC. The assay was evaluated for identification of H1 antigens with a panel of 55 strains.ConclusionFliC sequences were obtained for more than 100 strains comprising 29 different H1 alleles. Unique pyrosequencing profiles corresponding to the H1 component of the serotype were generated reproducibly for the 23 alleles represented in the evaluation panel. Short read sequence assays can now be used to identify fliC alleles in approximately 97% of the 50 medically most important Salmonella in England and Wales. Capability for high throughput testing and automation give these assays considerable advantages over traditional methods.
Highlights
The fliC and fljB genes in Salmonella code for the phase 1 (H1) and phase 2 (H2) flagellin respectively, the rfb cluster encodes the majority of enzymes for polysaccharide (O) antigen biosynthesis, together they determine the antigenic profile by which Salmonella are identified
O antigen synthesis and assembly is encoded by the rfb gene cluster which typically contains 12 open reading frames, and ranges in size between serotypes, from approximately 8 kbp to 23 kbp
Alignment of 106 fliC sequences generated in this study and 32 phase 1 flagellin sequences previously published, representing 35 phase 1 flagellar serotypes revealed a clear division of sequences into two groups
Summary
The fliC and fljB genes in Salmonella code for the phase 1 (H1) and phase 2 (H2) flagellin respectively, the rfb cluster encodes the majority of enzymes for polysaccharide (O) antigen biosynthesis, together they determine the antigenic profile by which Salmonella are identified. Sequencing and characterisation of fliC was performed in the development of a molecular serotyping technique. Salmonella express flagellar (H), polysaccharide (O) and capsular (Vi) antigens which determine strain pathogenicity and variation of these antigens has formed the basis for Salmonella serotyping. O antigen synthesis and assembly is encoded by the rfb gene cluster which typically contains 12 open reading frames, and ranges in size between serotypes, from approximately 8 kbp to 23 kbp. 20,000 repeating flagellin proteins polymerise to form the flagellar filament. The ends of the protein are (page number not for citation purposes)
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