Abstract: In vibrios and enterobacteria lateral gene transfer is often facilitated by Integrating Conjugative Elements (ICEs) of the SXT/R391 family. SXT/R391 ICEs integrate by site-specific recombination into prfC and transfer by conjugation, a process that is initiated at a specific locus called the origin of transfer (oriT(SXT) ). We identified genomic islands (GIs) harboring a sequence that shares >63% identity with oriT(SXT) in three species of Vibrio. Unlike SXT/R391 ICEs, these GIs are integrated into a gene coding for a putative stress-induced protein and do not appear to carry any gene coding for a conjugative machinery or for mobilization proteins. Our results show that SXT/R391 ICEs trigger the excision and mediate the conjugative transfer in trans of the three Vibrio GIs at high frequency. GIs' excision is independent of the ICE-encoded recombinase and is controlled by the ICE-encoded transcriptional activator SetCD, which is expressed during the host SOS response. Both mobI and traI, two ICE-borne genes involved in oriT-recognition, are essential for GIs' transfer. We also found that SXT/R391 ICEs mobilize in trans over 1Mb of chromosomal DNA located 5' of the GIs' integration site. Together these results support a novel mechanism of mobilization of GIs by ICEs of the SXT/R391 family.
Abstract: Integrating and conjugative elements (ICEs) are one of the three principal types of self-transmissible mobile genetic elements in bacteria. ICEs, like plasmids, transfer via conjugation; but unlike plasmids and similar to many phages, these elements integrate into and replicate along with the host chromosome. Members of the SXT/R391 family of ICEs have been isolated from several species of gram-negative bacteria, including Vibrio cholerae, the cause of cholera, where they have been important vectors for disseminating genes conferring resistance to antibiotics. Here we developed a plasmid-based system to capture and isolate SXT/R391 ICEs for sequencing. Comparative analyses of the genomes of 13 SXT/R391 ICEs derived from diverse hosts and locations revealed that they contain 52 perfectly syntenic and nearly identical core genes that serve as a scaffold capable of mobilizing an array of variable DNA. Furthermore, selection pressure to maintain ICE mobility appears to have restricted insertions of variable DNA into intergenic sites that do not interrupt core functions. The variable genes confer diverse element-specific phenotypes, such as resistance to antibiotics. Functional analysis of a set of deletion mutants revealed that less than half of the conserved core genes are required for ICE mobility; the functions of most of the dispensable core genes are unknown. Several lines of evidence suggest that there has been extensive recombination between SXT/R391 ICEs, resulting in re-assortment of their respective variable gene content. Furthermore, our analyses suggest that there may be a network of phylogenetic relationships among sequences found in all types of mobile genetic elements.
Abstract: Circulation of mobile genetic elements linked to drug resistance spread was studied in Vibrio strains isolated from surface urban water (river and sea) and shellfish samples in 2002-2003 in Maputo, Mozambique. Class 1 integrons and integrating conjugative elements (ICE) were investigated by PCR and mating experiments in strains of major health interest: 10 Vibrio cholerae, six Vibrio parahaemolyticus, two Vibrio alginolyticus and one Vibrio fluvialis. Resistance to at least two antibiotics (predominantly beta-lactams) was detected in all the strains, with additional resistances to sulfamethoxazole, spectinomycin, streptomycin and/or trimethoprim. Class 1 integrons contributed partially to the expression of drug resistance and were found in five isolates: four V. cholerae (blaP1 cassette, one strain also contained the dfrA15 cassette) and one V. alginolyticus (aadA2 cassette). ICEs, apparently devoid of resistance genes, were found in eight V. cholerae, three V. parahaemolyticus and one V. fluvialis isolates. A wide variability was observed by molecular characterization of ICEs. Five ICEs were included in the SXT/R391 family and seven ICEs were not classified. Our results indicate that the SXT/R391 family and related ICEs comprise a large class of polymorphic genetic elements widely circulating in environmental Vibrio strains in Africa, beside those evidently linked to drug resistance in clinical isolates.
Abstract: Integrating conjugative elements (ICEs) are self-transmissible, mobile elements that are widespread among bacteria. Following their excision from the chromosome, ICEs transfer by conjugation, a process initiated by a single-stranded DNA break at a specific locus called the origin of transfer (oriT). The SXT/R391 family of ICEs includes SXT(MO10), R391, and more than 25 related ICEs found in gammaproteobacteria. A previous study mapped the oriT locus of SXT(MO10) to a 550-bp intergenic region between traD and s043. We suspected that this was not the correct oriT locus, because the identical traD-s043 region in R391 and other SXT/R391 family ICEs was annotated as a gene of an unknown function. Here, we investigated the location and structure of the oriT locus in the ICEs of the SXT/R391 family and demonstrated that oriT(SXT) corresponds to a 299-bp sequence that contains multiple imperfect direct and inverted repeats and is located in the intergenic region between s003 and rumB'. The oriT(SXT) locus is well conserved among SXT/R391 ICEs, like R391, R997, and pMERPH, and cross-recognition of oriT(SXT) and oriT(R391) by R391 and SXT(MO10) was demonstrated. Furthermore, we identified a previously unannotated gene, mobI, located immediately downstream from oriT(SXT), which proved to be essential for SXT(MO10) transfer and SXT(MO10)-mediated chromosomal DNA mobilization. Deletion of mobI did not impair the SXT(MO10)-dependent transfer of the mobilizable plasmid CloDF13, suggesting that mobI has no role in the assembly of the SXT(MO10) mating pair apparatus. Instead, mobI appears to be involved in the recognition of oriT(SXT).
Abstract: We analyzed 28 epidemic Vibrio cholerae O1 strains isolated in the region of Thua Thien Hue (Vietnam) in 2003. Ubiquitous amoxicillin, prevalent aminoglycosides and sporadic erythromycin resistances were observed. All were devoid of plasmids, class 1 integrons and ICEs and showed the same BglI ribotype, irrespective of their site of isolation and resistance pattern. A strain isolated in 1990 in the same area was resistant to amoxicillin and aminoglycosides but characterized by a different ribotype. This strain contained ICEVchVie0, belonging to the SXT/R391 ICE family, devoid of any resistance cluster. The molecular analysis of three conserved and six variable regions outlined an original genetic profile. ICEs not coding for resistance to drugs seem to be more frequent than supposed, and this finding reinforces the idea that the SXT/R391 family of genetic elements is wide and composite. The clearance of ICEVchVie0 in the 2003 epidemic may be explained by the lack of any resistance determinant as a favorable selective marker.
Abstract: The resistance profile and its correlation with mobile genetic elements were investigated in 11 Vibrio cholerae O1 and 2 Vibrio parahaemolyticus clinical isolates, as well as in 1 V. cholerae O1 and 1 V. cholerae non-O1 environmental isolate, isolated between 1991 and 1996 in different provinces of Angola. All clinical isolates of V. cholerae O1 were resistant to ampicillin, chloramphenicol, trimethoprim, sulfamethoxazole, and tetracycline. They also contained a large conjugative plasmid (p3iANG) with a set of three class 1 integrons harboring dfrA15, blaP1, and qacH-aadA8 cassettes, which code for resistance to trimethoprim, beta-lactams, quaternary ammonium compounds, and aminoglycosides, clustered in a 19-kb region. Chloramphenicol (cat1), kanamycin (aph), sulfonamide (sul2), and tetracycline (tetG) resistance genes were also carried on the plasmid within the same 19-kb region. A chromosomal integron containing the dfrA15 cassette was also revealed in V. parahaemolyticus strains. SXT integrase genes were present in six V. cholerae isolates but apparently were not associated with known SXT-associated resistance genes. This study indicates that plasmids and integrons contributed mainly to the circulation of multiple-drug resistance determinants in Vibrio strains from Angola.
Abstract: This study reports the drug resistance and clonal relationship of 24 Staphylococcus aureus community acquired isolates from patients attending Maputo Central Hospital, Mozambique, during one year (2002-2003). All the isolates produced beta-lactamase, six strains were resistant to tetracycline alone, three were resistant to erythromycin alone and one was resistant to trimethoprim-sulfamethoxazole; 11 were susceptible to all other drugs tested. Only one strain showed a multiple resistance pattern, including methicillin resistance. To investigate the clonal relationships we applied the ERIC AP-PCR and the SmaI PFGE RFLP methods. Overlapping drug resistances with these two molecular profiles, no significant correlation was obtained. The emergence of methicillin resistance in a multiple resistant strain is of great concern for resistance spreading surveillance in Mozambique.
