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Attempts to control bacterial pathogens have led to an increase in antibiotic-resistant cells and the genetic elements that confer resistance phenotypes. These cells and genes are disseminated simultaneously with the original selective agents via human waste streams. This might lead to a second, unintended consequence of antimicrobial therapy; an increase in the evolvability of all bacterial cells. The genetic variation upon which natural selection acts is a consequence of mutation, recombination and lateral gene transfer (LGT). These processes are under selection, balancing genomic integrity against the advantages accrued by genetic innovation. Saturation of the environment with selective agents might cause directional selection for higher rates of mutation, recombination and LGT, producing unpredictable consequences for humans and the biosphere. © 2012 .

A Tn402-like class 1 integron was recovered from a prawn-associated bacterium. One of its cassettes included methionine sulfoxide reductase genes, the first example of such genes being captured by an integron. The integron was flanked by direct repeats that resemble miniature inverted-repeat transposable element sequences. Excision of the integron by homologous recombination through these sequences was demonstrated. Copyright © 2009, American Society for Microbiology. All Rights Reserved.

OBJECTIVES: To determine clonality and identify plasmid-mediated resistance genes in 11 multidrug-resistant Escherichia coli (MDREC) isolates associated with opportunistic infections in hospitalized dogs in Australia. METHODS: Phenotypic (MIC determinations, modified double-disc diffusion and isoelectric focusing) and genotypic methods (PFGE, plasmid analysis, PCR, sequencing, Southern hybridization, bacterial conjugation and transformation) were used to characterize, investigate the genetic relatedness of, and identify selected plasmid-mediated antimicrobial resistance genes, in the canine MDREC. RESULTS: Canine MDRECs were divided into two clonal groups (CG 1 and 2) with distinct restriction endonuclease digestion and plasmid profiles. All isolates possessed bla(CMY-7) on an approximately 93 kb plasmid. In CG 1 isolates, bla(TEM), catA1 and class 1 integron-associated dfrA17-aadA5 genes were located on an approximately 170 kb plasmid. In CG 2 isolates, a second approximately 93 kb plasmid contained bla(TEM) and unidentified class 1 integron genes, although a single CG 2 strain carried dfrA5. Antimicrobial susceptibility profiling of E. coli K12 transformed with CG 2 large plasmids confirmed that the bla(CMY-7)-carrying plasmid did not carry any other antimicrobial resistance genes, whereas the bla(TEM)/class 1 integron-carrying plasmid carried genes conferring resistance to tetracycline and streptomycin also. CONCLUSIONS: This is the first report on the detection of plasmid-mediated bla(CMY-7) in animal isolates in Australia. MDREC isolated from extraintestinal infections in dogs may be an important reservoir of plasmid-mediated resistance genes.

Integrons are best known for assembling antibiotic resistance genes in clinical bacteria. They capture genes by using integrase-mediated site-specific recombination of mobile gene cassettes. Integrons also occur in the chromosomes of many bacteria, notably beta- and gamma-Proteobacteria. In a survey of Xanthomonas, integrons were found in all 32 strains representing 12 pathovars of two species. Their chromosomal location was downstream from the acid dehydratase gene, ilvD, suggesting that an integron was present at this site in the ancestral xanthomonad. There was considerable sequence and structural diversity among the extant integrons. The majority of integrase genes were predicted to be inactivated by frameshifts, stop codons, or large deletions, suggesting that the associated gene cassettes can no longer be mobilized. In support, groups of strains with the same deletions or stop codons/frameshifts in their integrase gene usually contained identical arrays of gene cassettes. In general, strains within individual pathovars had identical cassettes, and these exhibited no similarity to cassettes detected in other pathovars. The variety and characteristics of contemporary gene cassettes suggests that the ancestral integron had access to a diverse pool of these mobile elements, and that their genes originated outside the Xanthomonas genome. Subsequent inactivation of the integrase gene in particular lineages has largely fixed the gene cassette arrays in particular pathovars during their differentiation and specialization into ecological niches. The acquisition of diverse gene cassettes by different lineages within Xanthomonas has contributed to the species-genome diversity of the genus. The role of gene cassettes in survival on plant surfaces is currently unknown.

Integron-encoded integrases recognize two distinct types of recombination site: attI sites, found in integrons, and members of the 59-base element (59-be) family, found in the integron-associated gene cassettes. The class 1 integron integrase, IntI1, catalyses recombination between attI1 and a 59-be, two 59-be, or two attI1 sites, but events involving two attI1 sites are less efficient than the reactions in which a 59-be participates. The full attI1 site is required for high-efficiency recombination with a 59-be site. It is 65 bp in length and includes a simple site, consisting of a pair of inversely oriented IntI1-binding domains, together with two further directly oriented IntI1-binding sites designated strong and weak. However, a smaller region that contains only the simple site is sufficient to support a lower level of recombination with a complete attI1 partner and the features that determine the orientation of attI1 reside within this region. An unusual reaction between the attI1 site and a 59-be appears to be responsible for the loss of the central region of a 59-be to create a potential fusion of two adjacent gene cassettes.

Site-specific insertion of gene cassettes into the insert region of integrons has been demonstrated. Insertion was only observed if the integron DNA integrase was expressed in the recipient cell and if the cassette DNA was ligated prior to transformation. The essential ligation products were resistant to treatment with exonuclease III, indicating that they were closed circular molecules. Insertion of cassettes into integron fragments containing either no insert (one recombination site), or one gene cassette (two recombination sites), was demonstrated. In the latter case, insertion occurred predominantly at the core site located 5' to the resident cassette, which corresponds to the only site available when no insert is present in the recipient. When DNA molecules including two gene cassettes were used, insertion of only one of the gene cassettes was generally observed, suggesting that resolution of the circular molecule to generate two independent circular cassettes occurred more rapidly than insertion into the recipient integron.