Fluoroquinolone resistant potential extraintestinal Escherichia coli in canine faeces: a possible link to human extraintestinal infections

Guo, SiYu (2013). Fluoroquinolone resistant potential extraintestinal Escherichia coli in canine faeces: a possible link to human extraintestinal infections PhD Thesis, School of Veterinary Science, The University of Queensland.

       
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Author Guo, SiYu
Thesis Title Fluoroquinolone resistant potential extraintestinal Escherichia coli in canine faeces: a possible link to human extraintestinal infections
Formatted title
Fluoroquinolone resistant potential extraintestinal Escherichia coli in canine faeces: a possible link to human extraintestinal infections
School, Centre or Institute School of Veterinary Science
Institution The University of Queensland
Publication date 2013
Thesis type PhD Thesis
Supervisor Rowland Cobbold
Huub Brouwers
Darren Trott
Toni Chapman
Total pages 199
Total colour pages 23
Total black and white pages 176
Language eng
Subjects 0707 Veterinary Sciences
070704 Veterinary Epidemiology
Formatted abstract
Extraintestinal pathogenic E. coli (ExPEC) cause urinary tract infections in humans and companion animals, as well as neonatal meningitis, bacteraemia, and sepsis. Fluoroquinolones are often the drug of choice. Given the frequency of human-dog contact and evidences of inter-host transmission of E. coli, the hypothesis of a canine faecal reservoir of fluoroquinolone resistant (FQ-R) ExPEC was evaluated. Objectives included: estimate the prevalence of canine FQ-R E. coli in general practice and referral clinics; compare canine faecal with extraintestinal FQ-R E. coli; assess the use of FQs on resistance development and canine intestinal FQ-R population dynamics using an in vivo model; characterise isolates by phylogenetic associations and virulence potential.

Canine faecal FQ-R E. coli isolates were obtained from a major NSW referral veterinary hospital, and underwent phylogenetic grouping, Random Amplified Polymorphism DNA analysis (RAPD) and ExPEC virulence genotyping. Twenty three (18.7%) of 123 dogs were colonised with FQ-R E. coli and within-hospital transmission was demonstrated. An important global pandemic human ExPEC clonal group (B2-O25b-ST131) was found in two dogs, which demonstrated RAPD and virulence gene (VG) similarity to canine extraintestinal ST131 from the Australian east coast. 61% of dogs carried group D faecal FQ-R ExPEC, with some clonal groups sharing similar VG profiles with ST131 isolates.

Group D faecal isolates from this study were compared to clinical group D canine FQ-R ExPEC from the Australian east coast to determine if canine faeces was a reservoir of important international ExPEC clonal groups other than B2-ST131. In addition to molecular analysis and Otyping, strains were screened for clonal group A and O15:K52:H1. Faecal and clinical isolates differed considerably by RAPD. Sub-clusters containing similar faecal and clinical isolates by RAPD and VG from multiple states were identified by multilocus sequence typing (MLST) as internationally distributed human-associated clonal groups of public health concern: ST393-O15:K52:H1, O15-ST130 and ST354. Results indicated possible human-canine cross-over of these and may demonstrate common selection advantages favouring both gastrointestinal colonisation and extraintestinal infection in both host species.

Next, experimental dogs were used to assess the effects of oral FQs on gastrointestinal microbiota and FQ-R E. coli population dynamics. Treatment groups received a FQ for 21 days. On day 14, all dogs were challenged orally with two canine multi-drug resistant (also FQ-R) E. coli representing phylogenetic groups A and D. Total coliform counts and counts on counter-selecting PRA (0.25 and 8 mg/L) or chloramphenicol (CHL) (16 mg/L) supplemented agar were monitored. Isolates underwent RAPD, virulence genotyping, Pulsed Field Gel Electrophoresis (PFGE), and O-typing. The total faecal microbial communities were analysed by Terminal-Restriction Fragment Length Polymorphism. Major findings included: Challenge strains were transient in dogs with no FQ treatment; FQ treatment appeared to induce emergence of a FQ-R group D E. coli population distinct from the challenge strains; higher doses of PRA were more effective at suppressing this FQR E. coli population, but may have initiated the expression of CHL resistance and facilitated acquisition of additional virulence genes in one MDREC challenge strain C13c; FQ-treated dogs carried the group D FQ-R challenge strain until day 34, at which time CHL therapy eliminated the strain but induced significant populations of the group A challenge strain, which was CHL-resistant; PRA appeared to alter several species and the total faecal microbiota more compared to ENR. The two dosages of PRA did not show a significant difference in their degree of alteration.

Another survey evaluated the prevalence of FQ-R E. coli among dogs hospitalised at primary accession clinics in QLD and NSW. A total of 114 faecal samples were obtained from 72 hospitalised dogs in nine hospitals. FQ-R E. coli isolates underwent real-time PCR screening for ST131 and CTX-M-15, followed by routine molecular analysis. No CTX-M-15 producing and/or ST131 FQ-R isolates were detected. Overall, primary accession clinics demonstrated a lower prevalence rate of FQ-R E. coli (2/72 dogs, 2.8%), with strains found in only one hospital, which exhibited a similar prevalence rate of 14.3% to the 18.7% obtained in the pilot study in the referral hospital. Group D reduced FQ-susceptible E. coli isolates were found in the same hospital, and displayed a FQ MIC phenotype suggesting the presence of pradofloxacin-specific efflux. Reduced FQ-susceptible and FQ-R isolates exhibited very similar RAPD profiles to some FQ-R faecal isolates from the pilot study. The close proximity of two sites may indicate a local focus for FQ-R E. coli, or the reduced FQ-susceptible and FQ-R group D isolates may have a recent common ancestor.

In summary, canine faecal reservoirs of human FQ-R ExPEC strains, including global pandemic human clonal groups causing serious extraintestinal infections have been demonstrated. However, dogs presenting to primary accession clinics may be less involved in disseminating human FQ-R ExPEC, including pandemic clones, compared to dogs presenting to referral hospitals due to their lower carriage rate. The higher dosage of PRA suppressed certain FQ-R E. coli strains compared to lower dosage of PRA and ENR, which may be an important finding to reduce the chances of crossinfection between dogs within hospitals.
Keyword Escherichia Coli infections
Fluoroquinolone
Drug Resistance
Dogs
Bacterial DNA
Microbial sensitivity tests
Bacterial typing techniques
Drug effects
Pradofloxacin
T-RFLP

 
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Created: Wed, 17 Jul 2013, 21:18:04 EST by Ms Si Yu Guo on behalf of Scholarly Communication and Digitisation Service