Role of attP in Integrase-Mediated Integration of the Shigella Resistance Locus Pathogenicity Island of Shigella flexneri

Abstract

The Shigella resistance locus (SRL) pathogenicity island (PAI) in Shigella spp. mediates resistance to streptomycin, ampicillin, chloramphenicol, and tetracycline. It can be excised from the chromosome via site-specific recombination mediated by the P4-related int gene. Here, we show that SRL PAI attP is capable of RecA-independent, site-specific, int-mediated integration into two bacterial tRNA attB sites.Shigella spp. are the causative agents of bacillary dysentery, a disease responsible for the deaths of over 1.1 million people annually (10). The infection, which is spread via the fecal-oral route, is commonly treated with a combination of rehydration and antimicrobial therapy. However, over the past few decades, treatment has become increasingly difficult as resistance to most of the widely used therapeutic antibiotics has emerged (9, 16).Multiple-antibiotic resistance to Shigella spp. was first reported as early as the 1950s (21), and since that time, multiresistance gene clusters have been identified on R plasmids, transposons, and integrons. Recently, a 16-kb cluster of genes known as the Shigella resistance locus (SRL) was identified on the chromosome of Shigella flexneri 2a strain YSH6000. In this strain, the SRL, which confers resistance to the antibiotics streptomycin, ampicillin, chloramphenicol, and tetracycline, is present on a 66-kb pathogenicity island (PAI) designated the SRL PAI (11). Recent findings show that the SRL PAI is present in numerous Shigella strains, and it is hypothesized that the SRL PAI may be involved in the spread of multiple-antibiotic resistance in Shigella spp. (17).PAIs, which are believed to be acquired by horizontal gene transfer, frequently carry phage-related integrase genes, are often integrated adjacent to tRNA genes, and are flanked by short direct repeats (DRs) which resemble phage attachment (att) sites (1). It is thought that the genetic similarities of PAIs and phages may extend to related mechanisms of integration and excision (7). However, genetic instability of PAIs has been observed in only a small number of cases, with only two PAIs having been shown to be mobilizable, and very little is known about mechanisms of PAI transfer in general (5). In contrast, the mechanism by which the SRL PAI is excised from the serX tRNA gene in S. flexneri 2a YSH6000 is known to be integrase mediated and site specific. These data, in conjunction with the presence of the SRL PAI in other Shigella strains (17), suggest that the SRL PAI, and therefore the antibiotic resistance genes it carries, may still be mobile. However, nothing is known of the excised form of the SRL PAI or its ability to integrate into a new host.Circular form of the SRL PAI. The SRL PAI of strain YSH6000 is flanked by 14-bp sequences that correspond to the terminal 3Ј 14 bp of the serX tRNA gene at the right boundary and a directly repeated sequence at the left boundary. Integrase-mediated excision results in the loss of the PAI and one copy of the DR, while an intact serX gene and a single copy of the DR r...

Excision of the Shigella Resistance Locus Pathogenicity Island in Shigella flexneri Is Stimulated by a Member of a New Subgroup of Recombination Directionality Factors

“…However, there was no statistically significant change (P value ϭ 0.385; two-tailed t test) in int transcription when rox was overexpressed following Little is known about the mechanisms of PAI mobility. However, Turner et al (27) showed that the SRL PAI int gene is required for excision of the SRL PAI, and it has been demonstrated that HPI int and SRL PAI int modules are capable of site-specific integration (19,26). Together, these results support the significance of int genes in PAI mobility.…”

“…Like many PAIs, the SRL PAI encodes an integrase that is homologous to members of the P4 family of integrases (13). Integrases mediate PAI integration into the chromosome (19,26) and are also essential for PAI excision (11,20,27). However, little is known about how the excision of PAIs is regulated.…”

“…The SRL PAI also encodes a functional ferric citrate transport system, a number of insertion sequences, and a large number of phage-like genes. Turner et al have shown that the SRL PAI can undergo precise insertion and deletion in an integrase-dependent manner (26,27).…”

Detection of CTX-M-14 extended-spectrum β-lactamase in Shigella sonnei isolates from China

“…1 Shigella flexneri was the most predominant Shigella species in China, but Shigella sonnei has been more and more common, recently. Antimicrobial resistance of Shigella species is increasing because of antibacterial agents used widely in clinical medicine.…”

Regulation, Integrase-Dependent Excision, and Horizontal Transfer of Genomic Islands in Legionella pneumophila

“…Furthermore, it is likely that an integration host factor (IHF) is involved in excision/integration of LpcGI-2, since we found four putative IHF sites on the genomic island directly associated with the attP/attR sites. The presence of putative IHF-binding sites and the role of the IHFs for the function of integrative and conjugative elements (ICEs) are known (50)(51)(52)(53)(54).…”