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Regional dissemination of a trimethoprim-resistance gene cassette via a successful transposable element.

ABSTRACT: Antimicrobial resistance is a growing international problem. We observed a 50% increase in the prevalence of trimethoprim resistance among fecal Escherichia coli from healthy Nigerian students between 1998 and 2005, a trend to increase that continued in 2009.A PCR-based screen revealed that 131 (43.1%) of isolates obtained in Nigeria in 2005 and 2009 carried integron-borne dfrA cassettes. In the case of 67 (51.1%) of these isolates, the cassette was a class 1-integron-borne dfrA7 gene, which has been reported at high prevalence from E. coli isolates from other parts of Africa. Complete sequencing of a 27 Kb dfrA7-bearing plasmid from one isolate located the dfrA7 gene within a Tn21-type transposon. The transposon also contained an IS26-derived bla/sul/str element, encoding resistance to ?-lactams, sulphonamides and streptomycin, and mercury resistance genes. Although the plasmid backbone was only found in 12 (5.8%) of trimethoprim-resistant isolates, dfrA7 and other transposon-borne genes were detected in 14 (16.3%) and 32 (26.3%) of trimethoprim resistant isolates collected in Nigeria in 2005 and 2009, respectively. Additionally, 37 (19.3%) of trimethoprim-resistant E. coli isolates collected between 2006 and 2008 from Ghana were positive for the dfrA7 and a transposon marker, but only 4 (2.1%) harbored the plasmid backbone.Our data point to transposition as a principal mechanism for disseminating dfrA7 among E. coli from Nigeria and Ghana. On-going intensive use of the affordable broad-spectrum antibacterials is likely to promote selective success of a highly prevalent transposable element in West Africa.

SUBMITTER: Labar AS

PROVIDER: S-EPMC3364232 | biostudies-literature | 2012

REPOSITORIES: biostudies-literature

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Regional dissemination of a trimethoprim-resistance gene cassette via a successful transposable element.

Labar Amy S AS Millman Jennifer S JS Ruebush Ellen E Opintan Japheth A JA Bishar Rima A RA Aboderin A Oladipo AO Newman Mercy J MJ Lamikanra Adebayo A Okeke Iruka N IN

PloS one 20120530 5

<h4>Background</h4>Antimicrobial resistance is a growing international problem. We observed a 50% increase in the prevalence of trimethoprim resistance among fecal Escherichia coli from healthy Nigerian students between 1998 and 2005, a trend to increase that continued in 2009.<h4>Methods and findings</h4>A PCR-based screen revealed that 131 (43.1%) of isolates obtained in Nigeria in 2005 and 2009 carried integron-borne dfrA cassettes. In the case of 67 (51.1%) of these isolates, the cassette wa ...[more]

PMID: 22666464

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Project description:BackgroundPowdery mildew (PM) is an important disease of cucumber (Cucumis sativus L.). CsaMLO8 was previously identified as a candidate susceptibility gene for PM in cucumber, for two reasons: 1) This gene clusters phylogenetically in clade V, which has previously been shown to harbour all known MLO-like susceptibility genes for PM identified in dicot species; 2) This gene co-localizes with a QTL on chromosome 5 for hypocotyl-specific resistance to PM.MethodsCsaMLO8 alleles from susceptible and resistant cucumber were cloned and transformed to mlo-mutant tomato. Cucumber seedlings were inoculated with Podosphaera xanthii, tissues were studied for CsaMLO8 expression at several timepoints post inoculation using qRT-PCR. The occurrence of the observed loss-of-function allele of CsaMLO8 in resequenced cucumber accessions was studied in silico.ResultsWe cloned CsaMLO8 alleles from susceptible and resistant cucumber genotypes, the latter carrying the QTL for hypocotyl resistance. We found that insertion of a non-autonomous Class LTR retrotransposable element in the resistant genotype leads to aberrant splicing of CsaMLO8 mRNA. Heterologous expression of the wild-type allele of CsaMLO8 in a tomato mlo-mutant restored PM susceptibility. However, heterologous expression of the CsaMLO8 allele cloned from the resistant cucumber genotype failed to restore PM susceptibility. Furthermore we showed that inoculation of susceptible cucumber with the PM pathogen Podosphaera xanthii induced transcriptional upregulation of CsaMLO8 in hypocotyl tissue, but not in cotyledon or leaf tissue. This coincides with the observation that the QTL at the CsaMLO8-locus causes full resistance in hypocotyl tissue, but only partial resistance in cotyledons and true leafs. We studied the occurrence of the loss-of-function allele of CsaMLO8 in cucumber germplasm by an in silico approach using resequencing data of a collection of 115 cucumber accessions, and found that this allele was present in 31 out of 115 accessions.ConclusionsCsaMLO8 was characterised as a functional susceptibility gene to PM, particularly in the hypocotyl where it was transcriptionally upregulated upon inoculation with the PM pathogen P. xanthii. A loss-of-function mutation in CsaMLO8 due to the insertion of a transposable element was found to be the cause of hypocotyl resistance to PM. This particular allele of CsaMLO8 was found to occur in 27 % of the resequenced cucumber accessions.

Project description:Reviewing the genetics underlying the arms race between bacteria and bacteriophages can offer an interesting insight into the development of bacterial resistance and phage co-evolution. This study shows how the natural development of resistances to the K1F bacteriophage, a phage which targets the K1 capsule of pathogenic Escherichia coli, can come about through insertion sequences (IS). Of the K1F resistant mutants isolated, two were of particular interest. The first of these showed full resistance to K1F and was found to have disruptions to kpsE, the product of which is involved in polysialic acid translocation. The second, after showing an initial susceptibility to K1F which then developed to full resistance, had disruptions to neuC, a gene involved in one of the early steps of polysialic acid biosynthesis. Both of these mutations came with a fitness cost and produced considerable phenotypic differences in the completeness and location of the K1 capsule when compared with the wild type. Sequential treatment of these two K1F resistant mutants with T7 resulted in the production of a variety of isolates, many of which showed a renewed susceptibility to K1F, indicating that these insertion sequence mutations are reversible, as well as one isolate that developed resistance to both phages. IMPORTANCE Bacteriophages have many potential uses in industry and the clinical environment as an antibacterial control measure. One of their uses, phage therapy, is an appealing alternative to antibiotics due to their high specificity. However, as with the rise in antimicrobial resistance (AMR), it is critical to improve our understanding of how resistance develops against these viral agents. In the same way as bacteria will evolve and mutate antibiotic receptors so they can no longer be recognized, resistance to bacteriophages can come about via mutations to phage receptors, preventing phage binding and infection. We have shown that Escherichia coli will become resistant to the K1F bacteriophage via insertion element reshufflings causing null mutations to elements of the polysialic acid biosynthetic cluster. Exposure to the T7 bacteriophage then resulted in further changes in the position of these IS elements, further altering their resistance and sensitivity profiles.

Dataset Information

Regional dissemination of a trimethoprim-resistance gene cassette via a successful transposable element.

Publications

Regional dissemination of a trimethoprim-resistance gene cassette via a successful transposable element.

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