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Characterisation of class 1 and class 2 integrons among bacteria isolated from an urban waste water treatment plant in Italy

Abstract number: P1514

Pellegrini C., Celenza G., Forcella C., Sacchetti E., Segatore B., Bellio P., Setacci D., Di Lisio C., Rainaldi S., Amicosante G., Perilli M.

Objectives: The role of environmental bacteria as a reservoir for antibiotic resistance determinants is still poorly established. The spread of resistance genes is greatly enhanced when they form part of a mobile gene cassette, associated with integrons. The aim of this study was to investigate the presence and distribution of integron-carrying bacteria from a urban waste water treatment plant of L'Aquila city (Italy).

Methods: During two years (may 2005-december 2007), 628 Gram-negative bacteria were isolated at different stages of the waste water treatment process, and selected on selective medium supplemented with ceftazidime 6 mg/L and imipenem 2 mg/L. Resistant bacteria were screened for the presence of integrase genes by colony blot hybridisation. Genotyping of integrase-positive strains was carried out by RAPD analysis. Variable region was investigated by PCR using primers designed to conserved regions of the integron structure, and sequenced. Plasmid profile was performed on selected strains.

Results: Overall 40% (251/628) of strains harboured an integron. The predominant organism (37.5%) was represented by Escherichia coli. PCR analysis with specific primers for intI1and intI2 genes was performed on 32 isolates that showed different genotype profile. The intI1 gene was detected in 26 out of 32 isolates screened and intI2 gene in 6 out 32 isolates. Sequence analysis of variable regions showed five cassette arrays in class 1 integrons and two arrays in class 2 integrons, encoding for antibiotic resistance determinants, as shown in table 1. Most of the integrons were located in chromosomal DNA, whereas only two integrons were found to be into large plasmids. Several strains contained b-lactamase genes, such as blaTEM-1 gene, blaCTX-M-1-type gene (detected in Alcaligenes faecalis), blaIMP-22, blaVIM-1 and blaAmpC gene.

Conclusion: Our results support the hypothesis that waste water treatment might be an important antibiotic resistance reservoirs and highlight the risk of spreading of harmful gene cassettes through discharges in aquatic ecosystems.

OrganismsIntegrase type (location)*Variable region size (bp)Variable region contentResistance phenotypePlasmidb-lactamase
Escherichia coliI (C)1664dfr17, aadA5AMP, AMX, AMC, STX+blaTEM
Escherichia coliII (P)1227dfr1, sat1AMP, AMX, PIP, CAZ, STX+-
Serratia liquefaciensI (C)NDAMP, AMX, PIP, CAZ+-
Enterobacter agglomeransI (C)NDAMP, AMC, AMX, CEF, CAZ--
Acinetobacter baumanniiI (C)1664dfr17, aadA5AMP, AMX, PIP, CEF, CTX, CAZ, IMP, STX+-
Serratia odoriferaII (C)NDAMP, AMX, PIP, CEF+-
Escherichia coliI (C)880aadA10AMP, AMX, AMC, PIP, LVF, STX+blaTEM
Acinetobacter baumanniiI (C)NDAMP, AMX, AMC, PIP, CEF, CTX, CAZ, LVF--
Escherichia coliII (C)2158dfr1, sat1, aadA1AMP, AMX, AMC, PIP, CEF, CTX, STX, LVF+blaTEM
Escherichia coliI (C)NDAMP, AMX, AMC, PIP, CEF, CAZ, LVF-blaTEM
Escherichia coliI (C)NDAMP, AMX, PIP, CEF, CAZ, STX-blaTEM
Klebsiella pneumoniaeI (C)1664dfr17, aadA5AMP, AMX, AMC, CEF, STX, LVF+blaTEM
Klebsiella oxytocaII (C)2158dfr1, sat1, aadA1AMP, AMX, PIP, TZP, CEF, STX+blaTEM
Aeromonas hydrophilaI (C)NDAMP, AMX, CEF--
Escherichia coliI (C)NDAMP, AMX, PIP, CEF, CAZ-blaTEM
Escherichia coliI (C)1664dfr17, aadA5AMP, AMX, AMC, PIP, CEF, STX-blaTEM
Citrobacter freundiiI (C)880aadA10AMP, AMX, AMC, PIP, CEF, STX--
Klebsiella oxytocaI (C)1664dfr17, aadA5AMP, AMX, PIP, CEF, CTX, CAZ, LVF, STX+blaTEM
Acinetobacter baumanniiI (C)1664dfr17, aadA5AMP, AMX, PIP, CEF, CTX, CAZ, LVF, STX+blaTEM
Escherichia coliI (C)ND AMP, AMX, PIP, CEF, LVF+blaTEM
Klebsiella pneumoniaeI (C)1664dfr17, aadA5AMP, AMX, AMC, PIP, CEF, STX, LVF+blaTEM
Citrobacter freundiiII (P)2158dfr1, sat, aadA1AMP, AMX, AMC, PIP, TZP, CAZ, IMP, STX--
Enterobacter agglomeransI (C)NDAMP, AMX, CEF, CAZ, STX--
Enterobacter agglomeransII (C)2158dfr1, sat, aadA1AMP, AMX, AMC, PIP, CEF, CAZ, IMP, STX-blaTEM
Pseudomonas putidaI (C)930blaVIM-1AMP, AMX, AMC, PIP, CEF, CTX, CAZ, IMP-blaVM
Pseudomonas fluorescensI (C)1281blaIMP-22, ORFXXAMP, AMX, AMC, PIP, CEF, CTX, CAZ, IMP-blaIMP
Pseudomonas fluorescensI (C)1910blaIMP-22, ORFXX, aacA4AMP, AMX, AMC, PIP, CEF, CTX, CAZ, IMP, STX-blaIMP
Enterobacter intermediusI (C)1664dfr17, aadA5AMP, AMX, PIP, CEF, CAZ, STX-blaAmpC
Escherichia coliI (C)570mobAAMP, AMX, AMC, PIP, CEF, CTX, CAZ, STX+blaTEM
Escherichia coliI (C)NDAMP, AMX, PIP, CEF-blaTEM
Alcaligenes faecalisI (C)NDCEF, CTX, CAZ+blaCTX-M
Escherichia coliI (C)880aadA10AMP, AMX, PIP, CEF, CAZ, STX-blaAmpC
*C: chromosomal localisation; P: plasmidic localisation. ND: not detected.

Session Details

Date: 16/05/2009
Time: 00:00-00:00
Session name: 19th European Congress of Clinical Microbiology and Infectious Diseases
Subject:
Location: Helsinki, Finland, 16 - 19 May 2009
Presentation type:
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