Genesis of a novel Shigella flexneri serotype by sequential infection of serotype-converting bacteriophages SfX and SfI

Abstract

Background: Shigella flexneri is the major pathogen causing bacillary dysentery. Fifteen serotypes have been recognized up to now. The genesis of new S. flexneri serotypes is commonly mediated by serotype-converting bacteriophages. Untypeable or novel serotypes from natural infections had been reported worldwide but have not been generated in laboratory.

Results: A new S. flexneri serotype-serotype 1 d was generated when a S. flexneri serotype Y strain (native LPS) was sequentially infected with 2 serotype-converting bacteriophages, SfX first and then SfI. The new serotype 1 d strain agglutinated with both serotype X-specific anti-7;8 grouping serum and serotype 1a-specific anti- I typing serum, and differed from subserotypes 1a, 1b and 1c. Twenty four S. flexneri clinical isolates of serotype X were all converted to serotype 1 d by infection with phage SfI. PCR and sequencing revealed that SfI and SfX were integrated in tandem into the proA-yaiC region of the host chromosome.

Conclusions: These findings suggest a new S. flexneri serotype could be created in nature. Such a conversion may be constrained by susceptibility of a strain to infection by a given serotype-converting bacteriophage. This finding has significant implications in the emergence of new S. flexneri serotypes in nature.

Figure 1

Construction of a novel serotype, 1 d, of S. flexneri with serotype-converting bacteriophages SfX and SfI. (A) Illustration of construction road map of S. flexneri 036_1d strain from a serotype Y strain 036, by sequential infection of phages SfX and SfI. (B) Serological identification of S. flexneri 036_1d as serotype 1 d with agglutination test using monovalent diagnostic sera. The constructed strain S. flexneri 036_1d agglutinated with both of typing sera I and grouping sera7;8. (C) Serological identification of S. flexneri 036_1d by Western-blot assay. The LPS extracted from the tested strains was separated by SDS-PAGE and hybridized with monovalent grouping sera 7;8 (a) and 3;4 (b), and typing sera I (c), respectively. LPS of serotype X strain 014 and serotype 1a strain 019 were used as positive controls for group specific antigen 7;8 and type specific antigen I. After strain name in brackets is the serotype of the strain.

Figure 2

Genetic organization of prophage genomes of SfX and/or SfI in S. flexneri 036_X and 036_1d. The prophage genomes of SfX and/or SfI are highlighted in yellow and pink respectively. The conserved genes of the host strain were shown in different colors: proA, gray; yaiC, yellow; IS600 ORF1 and ORF2, brown; IS629 ORF1 and ORF2, orange; the putative integrase gene (int), white. The integration sites attB, attL and attR are indicated in thick line. After strain name in brackets is the serotype of the strain.