Complete genomic sequence of SfV, a serotype-converting temperate bacteriophage of Shigella flexneri

Abstract

Bacteriophage SfV is a temperate serotype-converting phage of Shigella flexneri. SfV encodes the factors involved in type V O-antigen modification, and the serotype conversion and integration-excision modules of the phage have been isolated and characterized. We now report on the complete sequence of the SfV genome (37,074 bp). A total of 53 open reading frames were predicted from the nucleotide sequence, and analysis of the corresponding proteins was used to construct a functional map. The general organization of the genes in the SfV genome is similar to that of bacteriophage lambda, and numerous features of the sequence are described. The superinfection immunity system of SfV includes a lambda-like repression system and a P4-like transcription termination mechanism. Sequence analysis also suggests that SfV encodes multiple DNA methylases, and experiments confirmed that orf-41 encodes a Dam methylase. Studies conducted to determine if the phage-encoded methylase confers host DNA methylation showed that the two S. flexneri strains analyzed encode their own Dam methylase. Restriction mapping and sequence analysis revealed that the phage genome has cos sites at the termini. The tail assembly and structural genes of SfV show homology to those of phage Mu and Mu-like prophages in the genome of Escherichia coli O157:H7 and Haemophilus influenzae. Significant homology (30% of the genome in total) between sections of the early, regulatory, and structural regions of the SfV genome and the e14 and KpLE1 prophages in the E. coli K-12 genome were noted, suggesting that these three phages have common evolutionary origins.

FIG. 1.

Genetic map of bacteriophage SfV. (A) Relative locations of the different ORFs. Filled rectangle, cos site; filled circles, putative Rho-independent terminators as predicted by the GCG Terminator Program (refer to Table 2 for the sequence). Functional modules are indicated above the genetic map and are based on sequence and experimental analyses. (B) Regions of significant contiguous similarity (>77% identity at the nucleotide level) to other sequences in the data banks.

FIG. 1.

Genetic map of bacteriophage SfV. (A) Relative locations of the different ORFs. Filled rectangle, cos site; filled circles, putative Rho-independent terminators as predicted by the GCG Terminator Program (refer to Table 2 for the sequence). Functional modules are indicated above the genetic map and are based on sequence and experimental analyses. (B) Regions of significant contiguous similarity (>77% identity at the nucleotide level) to other sequences in the data banks.

FIG. 2.

Intergenic region between the cI and cro genes of SfV. The putative operator, consisting of three regions of inverted repeats (IR1, IR2, and IR3), is boxed, and the inverted repeats are italicized. Ribosomal binding sites, predicted by WebGeneMark.HMM (32), are in boldface; putative −10 and −35 promoter sequences are underlined.

FIG. 3.

Functional analysis of orf-41 (A) and effect of SfV on host DNA methylation (B). The presence and absence of orf-41 or SfV are indicated by plus and minus signs, respectively. MboI and Sau3AI digests are represented by M and S, respectively. In panel A, plasmid DNAs were extracted from strains B1041 (lanes 1 and 4), B1045 (lanes 2 and 5), and B1046 (lanes 3 and 6). In panel B, total DNA was extracted from strains EW595/52 (lanes 1 and 2), SFL1337 (lanes 3 and 4), SFL1 (lanes 5 and 6), and SFL1338 (lanes 7 and 8). Ma, EcoRI-digested SPP-1 molecular weight marker.