Genetic characterization of fluoroquinolone resistant Escherichia coli from urban streams and municipal and hospital effluents.

Escherichia coli with reduced susceptibility to ciprofloxacin, isolated from urban streams, wastewater treatment plants and hospital effluent between 2004 and 2012, were compared based on multilocus sequence typing (MLST), quinolone and beta-lactam resistance determinants and plasmid replicon type. Isolates from the different types of water and isolation dates clustered together, suggesting the persistence and capacity to propagate across distinct aquatic environments. The most prevalent MLST groups were ST10 complex and ST131. Almost all isolates (98%) carried mutations in the chromosomal genes gyrA and/or parC, and 10% possessed the genes qepA, aac(6('))-Ib-cr and/or qnrS1. Over 80% of the isolates were resistant to three or more classes of antibiotics (MDR ≥ 3). The most prevalent beta-lactamase encoding gene was blaTEM, followed by blaCTX-M-15, co-existing with plasmid mediated quinolone resistance. The plasmid replicon types of the group IncF were the most prevalent and distributed by different MLST groups. The genes aac(6('))-Ib-cr and/or qnrS1 could be transferred by conjugation in combination with the genes blaTEM,blaSHV-12 or blaOXA-1 and the plasmid replicon types I1-Iγ, K, HI2 and/or B/O. The potential of multidrug resistant E. coli with reduced susceptibility to ciprofloxacin, harboring mobile genetic elements and with ability to conjugate and transfer resistance genes, to spread and persist across different aquatic environments was demonstrated.

[1]  J. Campos,et al.  Extended-spectrum beta-lactamase-producing Escherichia coli in Spain belong to a large variety of multilocus sequence typing types, including ST10 complex/A, ST23 complex/A and ST131/B2. , 2009, International journal of antimicrobial agents.

[2]  D. Gally,et al.  Multidrug-resistant Escherichia coli from canine urinary tract infections tend to have commensal phylotypes, lower prevalence of virulence determinants and ampC-replicons , 2014, Veterinary microbiology.

[3]  C. Manaia,et al.  Bacterial diversity and antibiotic resistance in water habitats: searching the links with the human microbiome. , 2014, FEMS microbiology reviews.

[4]  Célia M Manaia,et al.  Antimicrobial resistance patterns in Enterobacteriaceae isolated from an urban wastewater treatment plant. , 2007, FEMS microbiology ecology.

[5]  P. H. Roy,et al.  Precise insertion of antibiotic resistance determinants into Tn21-like transposons: nucleotide sequence of the OXA-1 beta-lactamase gene. , 1987, Proceedings of the National Academy of Sciences of the United States of America.

[6]  S. Bertilsson,et al.  Quinolone-resistant Escherichia coli isolated from birds of prey in Portugal are genetically distinct from those isolated from water environments and gulls in Portugal, Spain and Sweden. , 2014, Environmental microbiology.

[7]  D. Paterson,et al.  Escherichia coli O25b-ST131: a pandemic, multiresistant, community-associated strain. , 2011, The Journal of antimicrobial chemotherapy.

[8]  P. Courvalin,et al.  Transferable Resistance to Aminoglycosides by Methylation of G1405 in 16S rRNA and to Hydrophilic Fluoroquinolones by QepA-Mediated Efflux in Escherichia coli , 2007, Antimicrobial Agents and Chemotherapy.

[9]  A. Boudabous,et al.  Prevalence and characterisation of extended-spectrum beta-lactamase (ESBL)-producing Escherichia coli isolates in healthy volunteers in Tunisia , 2011, European Journal of Clinical Microbiology & Infectious Diseases.

[10]  G L French,et al.  The continuing crisis in antibiotic resistance. , 2010, International journal of antimicrobial agents.

[11]  J. Campos,et al.  AmpC beta-lactamases in Escherichia coli: emergence of CMY-2-producing virulent phylogroup D isolates belonging mainly to STs 57, 115, 354, 393, and 420, and phylogroup B2 isolates belonging to the international clone O25b-ST131. , 2010, Diagnostic microbiology and infectious disease.

[12]  Ronald N. Jones,et al.  Evolution and dissemination of extended-spectrum beta-lactamase-producing Klebsiella pneumoniae: epidemiology and molecular report from the SENTRY Antimicrobial Surveillance Program (1997-2003). , 2005, Diagnostic microbiology and infectious disease.

[13]  M. Wittenbrink,et al.  ESBL-producing uropathogenic Escherichia coli isolated from dogs and cats in Switzerland. , 2013, Veterinary microbiology.

[14]  A. Moura,et al.  Co-resistance to different classes of antibiotics among ESBL-producers from aquatic systems. , 2014, Water research.

[15]  A. Carattoli,et al.  Identification of plasmids by PCR-based replicon typing. , 2005, Journal of microbiological methods.

[16]  F. Tenover,et al.  gyrA Mutations Associated with Fluoroquinolone Resistance in Eight Species ofEnterobacteriaceae , 1998, Antimicrobial Agents and Chemotherapy.

[17]  M. Upton,et al.  Prevalence and distribution of plasmid-mediated quinolone resistance genes in clinical isolates of Escherichia coli lacking extended-spectrum beta-lactamases. , 2008, The Journal of antimicrobial chemotherapy.

[18]  N. Woodford,et al.  Emergence of a new antibiotic resistance mechanism in India, Pakistan, and the UK: a molecular, biological, and epidemiological study , 2010, The Lancet. Infectious diseases.

[19]  C. Manaia,et al.  Urban wastewater treatment plants as hotspots for antibiotic resistant bacteria and genes spread into the environment: a review. , 2013, The Science of the total environment.

[20]  Torsten Seemann,et al.  Genomic Insights to Control the Emergence of Vancomycin-Resistant Enterococci , 2013, mBio.

[21]  D. Andersson,et al.  Evolution of antibiotic resistance at non-lethal drug concentrations. , 2012, Drug resistance updates : reviews and commentaries in antimicrobial and anticancer chemotherapy.

[22]  A. Alves,et al.  Analysing diversity among β-lactamase encoding genes in aquatic environments , 2006 .

[23]  A. Robicsek,et al.  Plasmid-Mediated Quinolone Resistance: a Multifaceted Threat , 2009, Clinical Microbiology Reviews.

[24]  I. Luzzi,et al.  Escherichia coli of human and avian origin: detection of clonal groups associated with fluoroquinolone and multidrug resistance in Italy. , 2012, The Journal of antimicrobial chemotherapy.

[25]  G. Jacoby Mechanisms of resistance to quinolones. , 2005, Clinical infectious diseases : an official publication of the Infectious Diseases Society of America.

[26]  Munirul Alam,et al.  Antimicrobial Drug–Resistant Escherichia coli in Wild Birds and Free-range Poultry, Bangladesh , 2012, Emerging infectious diseases.

[27]  Brian Austin,et al.  Selective Pressure of Antibiotic Pollution on Bacteria of Importance to Public Health , 2012, Environmental health perspectives.

[28]  I. Scaletsky,et al.  Variants of astA gene among extra‐intestinal Escherichia coli of human and avian origin , 2016, FEMS microbiology letters.

[29]  A. Robicsek,et al.  Fluoroquinolone-modifying enzyme: a new adaptation of a common aminoglycoside acetyltransferase , 2006, Nature Medicine.

[30]  Koichiro Tamura,et al.  MEGA6: Molecular Evolutionary Genetics Analysis version 6.0. , 2013, Molecular biology and evolution.

[31]  A. Pühler,et al.  Genomics of IncP-1 antibiotic resistance plasmids isolated from wastewater treatment plants provides evidence for a widely accessible drug resistance gene pool. , 2007, FEMS microbiology reviews.

[32]  C. Manaia,et al.  Factors influencing antibiotic resistance burden in municipal wastewater treatment plants , 2010, Applied Microbiology and Biotechnology.

[33]  L. Nilsson,et al.  Molecular identification of CTX-M and blaOXY/K1 β-lactamase genes in Enterobacteriaceae by sequencing of universal M13-sequence tagged PCR-amplicons , 2009, BMC infectious diseases.

[34]  Mark R. Liles,et al.  Identification of Diverse Antimicrobial Resistance Determinants Carried on Bacterial, Plasmid, or Viral Metagenomes from an Activated Sludge Microbial Assemblage , 2010, Applied and Environmental Microbiology.

[35]  L. McDonald,et al.  Emergence of Reduced Susceptibility and Resistance to Fluoroquinolones in Escherichia coliin Taiwan and Contributions of Distinct Selective Pressures , 2001, Antimicrobial Agents and Chemotherapy.

[36]  P. Nordmann,et al.  Emergence of Plasmid-Mediated Quinolone Resistance in Escherichia coli in Europe , 2005, Antimicrobial Agents and Chemotherapy.

[37]  R. Cantón,et al.  Emergence and spread of antibiotic resistance following exposure to antibiotics. , 2011, FEMS microbiology reviews.

[38]  M. Ferraro Performance standards for antimicrobial susceptibility testing , 2001 .

[39]  J. Acar,et al.  Consequences of bacterial resistance to antibiotics in medical practice. , 1997, Clinical infectious diseases : an official publication of the Infectious Diseases Society of America.

[40]  T. Jukes CHAPTER 24 – Evolution of Protein Molecules , 1969 .

[41]  A. Robicsek,et al.  Prevalence in the United States of aac(6′)-Ib-cr Encoding a Ciprofloxacin-Modifying Enzyme , 2006, Antimicrobial Agents and Chemotherapy.

[42]  P. Nordmann,et al.  Multiplex PCR for detection of plasmid-mediated quinolone resistance qnr genes in ESBL-producing enterobacterial isolates. , 2007, The Journal of antimicrobial chemotherapy.

[43]  D. Hooper,et al.  New Plasmid-Mediated Quinolone Resistance Gene, qnrC, Found in a Clinical Isolate of Proteus mirabilis , 2009, Antimicrobial Agents and Chemotherapy.

[44]  Marnie L Campbell The pointy end of the stick: managing biological invasions , 2013 .

[45]  K. Smalla,et al.  Broad diversity of conjugative plasmids in integron-carrying bacteria from wastewater environments. , 2012, FEMS microbiology letters.

[46]  F. Baquero,et al.  Spread of blaCTX-M-14 Is Driven Mainly by IncK Plasmids Disseminated among Escherichia coli Phylogroups A, B1, and D in Spain , 2009, Antimicrobial Agents and Chemotherapy.

[47]  T. Jukes Transferable resistance. , 1968, British medical journal.

[48]  A. Robicsek,et al.  The worldwide emergence of plasmid-mediated quinolone resistance. , 2006, The Lancet. Infectious diseases.

[49]  A. Ivens,et al.  Clinically Relevant Mutant DNA Gyrase Alters Supercoiling, Changes the Transcriptome, and Confers Multidrug Resistance , 2013, mBio.

[50]  A. Oliver,et al.  Corrigendum to Extended-spectrum β-lactamase-producing Escherichia coli in Spain belong to a large variety of multilocus sequence typing types, including ST10 complex/A, ST23 complex/A and ST131/B2 [Int. J. Antimicrob. Agents 34 (2009) 173-176] , 2010 .

[51]  Jian Sun,et al.  Plasmid-mediated quinolone resistance determinants oqxAB and aac(6')-Ib-cr and extended-spectrum β-lactamase gene blaCTX-M-24 co-located on the same plasmid in one Escherichia coli strain from China. , 2011, The Journal of antimicrobial chemotherapy.

[52]  S. Schwarz,et al.  Analysis of extended-spectrum-β-lactamase-producing Escherichia coli isolates collected in the GERM-Vet monitoring programme. , 2013, The Journal of antimicrobial chemotherapy.

[53]  Alessandra Carattoli,et al.  Resistance Plasmid Families in Enterobacteriaceae , 2009, Antimicrobial Agents and Chemotherapy.

[54]  F. Baquero,et al.  Antibiotics and antibiotic resistance in water environments. , 2008, Current opinion in biotechnology.

[55]  J. Martínez,et al.  Environmental pollution by antibiotics and by antibiotic resistance determinants. , 2009, Environmental pollution.

[56]  M. Falagas,et al.  Multidrug-resistant, extensively drug-resistant and pandrug-resistant bacteria: an international expert proposal for interim standard definitions for acquired resistance. , 2012, Clinical microbiology and infection : the official publication of the European Society of Clinical Microbiology and Infectious Diseases.

[57]  E. Kuijper,et al.  Emergence of reduced susceptibility to metronidazole in Clostridium difficile. , 2008, The Journal of antimicrobial chemotherapy.

[58]  Luis Martõ Plasmid-mediated quinolone resistance: an update , 2011 .

[59]  C. Manaia,et al.  Insights into the relationship between antimicrobial residues and bacterial populations in a hospital-urban wastewater treatment plant system. , 2014, Water research.

[60]  G. Jacoby,et al.  Quinolone resistance from a transferable plasmid , 1998, The Lancet.

[61]  Y. Carmeli,et al.  Treatment with Fluoroquinolones or with β-Lactam-β-Lactamase Inhibitor Combinations Is a Risk Factor for Isolation of Extended-Spectrum-β-Lactamase-Producing Klebsiella Species in Hospitalized Patients , 2010, Antimicrobial Agents and Chemotherapy.

[62]  H. Christensen,et al.  Genetic diversity and virulence profiles of Escherichia coli causing salpingitis and peritonitis in broiler breeders. , 2013, Veterinary microbiology.

[63]  E. Wellington,et al.  Waste water effluent contributes to the dissemination of CTX-M-15 in the natural environment , 2014, The Journal of antimicrobial chemotherapy.

[64]  F. Baquero,et al.  Dissemination of Clonally Related Escherichia coli Strains Expressing Extended-Spectrum β-Lactamase CTX-M-15 , 2008, Emerging infectious diseases.

[65]  C. Dierikx,et al.  Increased detection of extended spectrum beta-lactamase producing Salmonella enterica and Escherichia coli isolates from poultry. , 2010, Veterinary microbiology.

[66]  P. Heisig,et al.  Genetic evidence for a role of parC mutations in development of high-level fluoroquinolone resistance in Escherichia coli , 1996, Antimicrobial agents and chemotherapy.

[67]  M. Achtman,et al.  Genome Sequences and Phylogenetic Analysis of K88- and F18-Positive Porcine Enterotoxigenic Escherichia coli , 2011, Journal of bacteriology.

[68]  F. Aarestrup,et al.  Molecular Characterization and Antimicrobial Susceptibility Testing of Escherichia coli Isolates from Patients with Urinary Tract Infections in 20 Chinese Hospitals , 2011, Journal of Clinical Microbiology.

[69]  A. Mellmann,et al.  Analysis of Collection of Hemolytic Uremic Syndrome–associated Enterohemorrhagic Escherichia coli , 2008, Emerging infectious diseases.

[70]  M. Kuskowski,et al.  Escherichia coli sequence type ST131 as the major cause of serious multidrug-resistant E. coli infections in the United States. , 2010, Clinical infectious diseases : an official publication of the Infectious Diseases Society of America.

[71]  F. Baquero,et al.  Emergence and spread of antibiotic resistance: setting a parameter space , 2014, Upsala journal of medical sciences.

[72]  A. Brisabois,et al.  Emergence of Extended-Spectrum-β-Lactamase (CTX-M-9)-Producing Multiresistant Strains of Salmonella enterica Serotype Virchow in Poultry and Humans in France , 2004, Journal of Clinical Microbiology.

[73]  C. Manaia,et al.  Differential patterns of antimicrobial resistance in population subsets of Escherichia coli isolated from waste- and surface waters. , 2011, The Science of the total environment.

[74]  G. Kahlmeter,et al.  High genetic diversity of nitrofurantoin- or mecillinam-resistant Escherichia coli indicates low propensity for clonal spread. , 2013, The Journal of antimicrobial chemotherapy.

[75]  Alfred Pühler,et al.  Detection of 140 clinically relevant antibiotic-resistance genes in the plasmid metagenome of wastewater treatment plant bacteria showing reduced susceptibility to selected antibiotics. , 2009, Microbiology.

[76]  Alessandra Carattoli,et al.  Replicon sequence typing of IncF plasmids carrying virulence and resistance determinants. , 2010, The Journal of antimicrobial chemotherapy.

[77]  F. Aarestrup,et al.  qnrD, a Novel Gene Conferring Transferable Quinolone Resistance in Salmonella enterica Serovar Kentucky and Bovismorbificans Strains of Human Origin , 2008, Antimicrobial Agents and Chemotherapy.

[78]  D. Livermore,et al.  High prevalence of acquired quinolone-resistance genes among Enterobacteriaceae from Saudi Arabia with CTX-M-15 β-lactamase. , 2012, Diagnostic microbiology and infectious disease.

[79]  C. Manaia,et al.  Diversity and antibiotic resistance of Aeromonas spp. in drinking and waste water treatment plants. , 2011, Water research.

[80]  J. Campos,et al.  Extended-Spectrum- (cid:1) -Lactamase-Producing Escherichia coli as a Cause of Pediatric Infections: Report of a Neonatal Intensive Care Unit Outbreak Due to a CTX-M-14-Producing Strain , 2011 .

[81]  N. Woodford,et al.  Multiresistant Gram-negative bacteria: the role of high-risk clones in the dissemination of antibiotic resistance. , 2011, FEMS microbiology reviews.

[82]  M. Nei,et al.  Prospects for inferring very large phylogenies by using the neighbor-joining method. , 2004, Proceedings of the National Academy of Sciences of the United States of America.

[83]  R. Koczura,et al.  Phylogenetic groups, virulence genes and quinolone resistance of integron-bearing Escherichia coli strains isolated from a wastewater treatment plant , 2011, Antonie van Leeuwenhoek.

[84]  C. Manaia,et al.  Antibiotic resistance, antimicrobial residues and bacterial community composition in urban wastewater. , 2013, Water research.

[85]  Daniel Falush,et al.  Sex and virulence in Escherichia coli: an evolutionary perspective , 2006, Molecular microbiology.

[86]  J. Blanco,et al.  Poultry as reservoir for extraintestinal pathogenic Escherichia coli O45:K1:H7-B2-ST95 in humans. , 2013, Veterinary microbiology.

[87]  J. Blanco,et al.  Intercontinental emergence of Escherichia coli clone O25:H4-ST131 producing CTX-M-15. , 2007, The Journal of antimicrobial chemotherapy.