Gene Cassette PCR: Sequence-Independent Recovery of Entire Genes from Environmental DNA

Stokes, H. W.; Holmes, Andrew; Nield, Blair; Holley, Marita; Nevalainen, Helena; Mabbutt, Bridget C.; Gillings, Michael R.

doi:10.1128/aem.67.11.5240-5246.2001

Cited by 169 publications

(213 citation statements)

References 29 publications

Supporting

Mentioning

207

Contrasting

Unclassified

Order By: Relevance

“…However, these techniques are unable to identify specific resistance determinants and their origins, and this knowledge gap limits understanding of dispersal of antibiotic resistance in wild marine populations (Stokes et al 2001).…”

Section: Introductionmentioning

confidence: 99%

MOLECULAR DETECTION OF ANTIBIOTIC-RESISTANCE DETERMINANTS INESCHERICHIA COLIISOLATED FROM THE ENDANGERED AUSTRALIAN SEA LION (NEOPHOCA CINEREA)

Delport

Harcourt

Beaumont

et al. 2015

Journal of Wildlife Diseases

View full text Add to dashboard Cite

ABSTRACT:Greater interaction between humans and wildlife populations poses significant risks of anthropogenic impact to natural ecosystems, especially in the marine environment. Understanding the spread of microorganisms at the marine interface is therefore important if we are to mitigate adverse effects on marine wildlife. We investigated the establishment of Escherichia coli in the endangered Australian sea lion (Neophoca cinerea) by comparing fecal isolation from wild and captive sea lion populations. Fecal samples were collected from wild colonies March 2009-September 2010 and from captive individuals March 2011-May 2013. Using molecular screening, we assigned a phylotype to E. coli isolates and determined the presence of integrons, mobile genetic elements that capture gene cassettes conferring resistance to antimicrobial agents common in fecal coliforms. Group B2 was the most abundant phylotype in all E. coli isolates (n537), with groups A, B1, and D also identified. Integrons were not observed in E. coli (n521) isolated from wild sea lions, but were identified in E. coli from captive animals (n516), from which class I integrases were detected in eight isolates. Sequencing of gene cassette arrays identified genes conferring resistance to streptomycin-spectinomycin (aadA1) and trimethoprim (dfrA17, dfrB4). Class II integrases were not detected in the E. coli isolates. The frequent detection in captive sea lions of E. coli with resistance genes commonly identified in human clinical cases suggests that conditions experienced in captivity may contribute to establishment. Identification of antibiotic resistance in the microbiota of Australian sea lions provides crucial information for disease management. Our data will inform conservation management strategies and provide a mechanism to monitor microorganism dissemination to sensitive pinniped populations.

show abstract

Section: Introductionmentioning

confidence: 99%

MOLECULAR DETECTION OF ANTIBIOTIC-RESISTANCE DETERMINANTS INESCHERICHIA COLIISOLATED FROM THE ENDANGERED AUSTRALIAN SEA LION (NEOPHOCA CINEREA)

Delport

Harcourt

Beaumont

et al. 2015

Journal of Wildlife Diseases

View full text Add to dashboard Cite

show abstract

“…Of the 46 sequenced gene cassettes obtained from total community DNA extracted from each sample type, 43 had predicted ORFs with start and stop codons in typical gene cassette orientation (Stokes et al, 2001), while the three with no obvious ORF were all less than 209 bp in length. Gene cassette sequences have been deposited in GenBank under the following accession numbers: EU435312-EU435354.…”

Section: Gene Cassette Functionmentioning

confidence: 99%

“…Class 1 integrons were first identified due to their role in the dissemination of antibiotic resistance gene cassettes in clinical bacteria (Stokes and Hall, 1989), but have since been found in a phylogenetically broad range of clinical and environmental bacteria including Gram-negative and Gram-positive bacteria (Nandi et al, 2004;Stokes et al, 2006;Boucher et al, 2007). Environmental surveys of integron-associated gene cassettes indicate that the majority of cassettes include novel sequences unrelated to known antibiotic resistance genes (Stokes et al, 2001;Holmes et al, 2003), with one conservative estimate of cassette richness being at least 2343 different gene cassettes in a 50-m 2 sediment plot (Michael et al, 2004). Integrons therefore represent a potential genetic resource by which bacteria in a variety of environments can adapt through the acquisition of new genetic material.…”

Section: Introductionmentioning

confidence: 99%

Influence of industrial contamination on mobile genetic elements: class 1 integron abundance and gene cassette structure in aquatic bacterial communities

et al. 2008

View full text Add to dashboard Cite

The acquisition of new genetic material via horizontal gene transfer allows bacteria to rapidly evolve. One key to estimating the contribution of horizontal gene transfer to bacterial evolution is to quantify the abundance of mobile genetic elements (MGEs) in bacterial communities under varying degrees of selective pressure. We quantified class 1 integrase (intI1) gene abundance in total community DNA extracted from contaminated and reference riverine and estuarine microhabitats, and in metal-or antibiotic-amended freshwater microcosms. The intI1 gene was more abundant in all contaminant-exposed communities indicating that relative gene transfer potential is higher in these communities. A second key to assessing the contributions of MGEs to bacterial evolution is to examine the structure and function of the MGE-associated gene pool. We determined that the gene cassette pool is a novel and diverse resource available for bacterial acquisition, but that contamination has no discernible effect on cassette richness. Gene cassette profiles were more similar within sites than among sites, yet bacterial community profiles were not, suggesting that selective pressures can shape the structure of the gene cassette pool. Of the 46 sequenced gene cassette products, 37 were novel sequences, while the 9 gene cassettes with similarity to database sequences were primarily to hypothetical proteins. That class 1 integrons are ubiquitous and abundant in environmental bacterial communities indicates that this group of MGEs can play a substantial role in the acquisition of a diverse array of gene cassettes beyond their demonstrated impact in mediating multidrug resistance in clinical bacteria.

show abstract

“…This reaction was performed using class 1-specific primers HS463A and HS464 . Colonies were also screened for gene cassettes with degenerate primers (HS287 and HS286) targeting attC regions (Stokes et al, 2001). PCR products by these reactions were cloned and sequenced as described above.…”

Section: Sample Collection and Cultivationmentioning

confidence: 99%

Coral-mucus-associated Vibrio integrons in the Great Barrier Reef: genomic hotspots for environmental adaptation

et al. 2011

View full text Add to dashboard Cite

Integron cassette arrays in a dozen cultivars of the most prevalent group of Vibrio isolates obtained from mucus expelled by a scleractinian coral (Pocillopora damicornis) colony living on the Great Barrier Reef were sequenced and compared. Although all cultivars showed 499% identity across recA, pyrH and rpoB genes, no two had more than 10% of their integron-associated gene cassettes in common, and some individuals shared cassettes exclusively with distantly-related members of the genus. Of cassettes shared within the population, a number appear to have been transferred between Vibrio isolates, as assessed by phylogenetic analysis. Prominent among the mucus Vibrio cassettes with potentially inferable functions are acetyltransferases, some with close similarity to known antibiotic-resistance determinants. A subset of these potential resistance cassettes were shared exclusively between the mucus Vibrio cultivars, Vibrio coral pathogens and human pathogens, thus illustrating a direct link between these microbial niches through exchange of integron-associated gene cassettes.

show abstract

Gene Cassette PCR: Sequence-Independent Recovery of Entire Genes from Environmental DNA

Cited by 169 publications

References 29 publications

MOLECULAR DETECTION OF ANTIBIOTIC-RESISTANCE DETERMINANTS INESCHERICHIA COLIISOLATED FROM THE ENDANGERED AUSTRALIAN SEA LION (NEOPHOCA CINEREA)

MOLECULAR DETECTION OF ANTIBIOTIC-RESISTANCE DETERMINANTS INESCHERICHIA COLIISOLATED FROM THE ENDANGERED AUSTRALIAN SEA LION (NEOPHOCA CINEREA)

Influence of industrial contamination on mobile genetic elements: class 1 integron abundance and gene cassette structure in aquatic bacterial communities

Coral-mucus-associated Vibrio integrons in the Great Barrier Reef: genomic hotspots for environmental adaptation

Contact Info

Product

Resources

About