A Multiplex Polymerase Chain Reaction for the Differentiation of Campylobacter jejuni and Campylobacter coli from a Swine Processing Facility and Characterization of Isolates by Pulsed-Field Gel Electrophoresis and Antibiotic Resistance Profiles

Cloak, Orla M.; Fratamico, Pina M.

doi:10.4315/0362-028x-65.2.266

Cited by 58 publications

(31 citation statements)

References 38 publications

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“…tant species of Campylobacter besides C. jejuni that causes food-borne gastroenteritis (6,8,44). Another technique used routinely for typing both C. jejuni and C. coli is PFGE (3,4,13,22,41,42). This method is being used by PulseNet within the United States for the nationwide surveillance of this pathogen along with other food-borne pathogens like Salmonella and Shigella (43).…”

Section: Discussionmentioning

confidence: 99%

Campylobacter coliin Swine Production: Antimicrobial Resistance Mechanisms and Molecular Epidemiology

Thakur

Gebreyes

2005

J Clin Microbiol

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The aim of this study was to determine antimicrobial resistance, to evaluate and compare the use of two genotyping methods for molecular epidemiology purposes, and to determine the genotypic diversity of Campylobacter coli of porcine origin. A total of 100 C. coli isolates from swine were tested for susceptibility to six antimicrobials using the agar dilution method and genotyped using two high-resolution fingerprinting approaches: multilocus sequence typing (MLST) and pulsed-field gel electrophoresis (PFGE). Evaluation of the methods was based on their resistance patterns, discriminatory indexes (DI), high test throughputs, costs, and turnaround times. Resistance to erythromycin and tetracycline was the most common. Both genotypic methods were found to have high discriminatory power, although MLST had a higher DI (0.936) than PFGE (DI ‫؍‬ 0.889). It also had a higher throughput than PFGE. Isolates were clustered into 27 groups by MLST compared to 11 by PFGE. MLST was able to further discriminate the isolates grouped under the same cluster by PFGE. Out of the 65 MLST sequence types (STs) identified among the total isolates, 50 were reported for the first time. Most STs were found to be specific to the farm (n ‫؍‬ 38) and to slaughter (n ‫؍‬ 22). Resistance against tetracycline and erythromycin was encoded by the tet(O) gene and a A2075G point mutation in the 23S rRNA gene, respectively. A high ciprofloxacin MIC (>64 g/liter) was conferred by a point mutation in the gyrA gene. The weak clonal structure of the C. coli population among swine was further highlighted by the index of association value of 0.293. The findings of this study indicate that multidrug-resistant diverse C. coli strains exhibiting resistance to ciprofloxacin and erythromycin are concerning, since these are the drugs of choice for treating invasive campylobacteriosis cases in humans.

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Section: Discussionmentioning

confidence: 99%

Campylobacter coliin Swine Production: Antimicrobial Resistance Mechanisms and Molecular Epidemiology

Thakur

Gebreyes

2005

J Clin Microbiol

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“…The expected lpxA amplification product sizes are 391 bp for C. coli and 331 bp for C. jejuni; the expected hipO amplification product size is 176 bp. NADC strains were speciated using a multiplex PCR method as described by Cloak and Fratamico (2002). Additionally, all NADC strains were typed by PFGE of SmaI-digested genomic DNA.…”

Section: Methodsmentioning

confidence: 99%

Identification of host-associated alleles by multilocus sequence typing of Campylobacter coli strains from food animals

et al. 2006

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Campylobacter coli is a food-borne pathogen associated increasingly with human gastroenteritis. C. coli has a high prevalence in swine, but is isolated also from cattle and poultry. Multilocus sequence typing (MLST) systems have been developed to differentiate C. coli strains. Although substantial allelic diversity was identified across all seven C. coli MLST loci, no correlations were made in two previous studies between allele or sequence type (ST) and the source of the organism. However, this may be due to either the relatively small number or the low diversity of C. coli strains used to validate both MLST studies. This study describes the typing of 488 C. coli strains from 4 different food animal sources (cattle, chickens, swine and turkeys), collected at different times over a 6 year period from different USA geographical locations. A total of 149 STs were identified. The 185 swine strains were the most diverse, possessing 82 STs. The cattle strains were the most clonal; 52/63 (83 %) strains possessed a single ST (ST-1068). A subpopulation of C. coli strains, collected primarily from turkeys, was identified, containing both C. coli-and Campylobacter jejuni-associated MLST alleles, specifically the C. jejuni allele aspA103. The majority of STs and alleles were host associated, i.e. found primarily in strains from a single food-animal source. Only 12/149 (8 %) STs were found in multiple sources. Additionally, the majority (34/46, 74 %) of major (n>5) alleles were more prevalent in certain hosts (swine, poultry). The presence of host-associated C. coli MLST alleles could lead potentially to more efficient source tracking in this species, especially in the trace-back of both sporadic and outbreak human clinical C. coli strains to animal sources.

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“…Oligodeoxynucleotide primers and PCR assays that distinguish between one or more combinations of the thermotolerant Campylobacter species have been described (1,4,5,9,11,13,16,21,48). The advantages of PCRbased assays are that they can be performed quickly and relatively cheaply (29).…”

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confidence: 99%

Differentiation of Campylobacter coli , Campylobacter jejuni , Campylobacter lari , and Campylobacter upsaliensis by a Multiplex PCR Developed from the Nucleotide Sequence of the Lipid A Gene lpxA

et al. 2004

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We describe a multiplex PCR assay to identify and discriminate between isolates of Campylobacter coli, Campylobacter jejuni, Campylobacter lari, and Campylobacter upsaliensis. The C. jejuni isolate F38011 lpxA gene, encoding a UDP-N-acetylglucosamine acyltransferase, was identified by sequence analysis of an expression plasmid that restored wild-type lipopolysaccharide levels in Escherichia coli strain SM105 [lpxA(Ts)]. With oligonucleotide primers developed to the C. jejuni lpxA gene, nearly full-length lpxA amplicons were amplified from an additional 11 isolates of C. jejuni, 20 isolates of C. coli, 16 isolates of C. lari, and five isolates of C. upsaliensis. The nucleotide sequence of each amplicon was determined, and sequence alignment revealed a high level of species discrimination. Oligonucleotide primers were constructed to exploit species differences, and a multiplex PCR assay was developed to positively identify isolates of C. coli, C. jejuni, C. lari, and C. upsaliensis. We characterized an additional set of 41 thermotolerant isolates by partial nucleotide sequence analysis to further demonstrate the uniqueness of each species-specific region. The multiplex PCR assay was validated with 105 genetically defined isolates of C. coli, C. jejuni, C. lari, and C. upsaliensis, 34 strains representing 12 additional Campylobacter species, and 24 strains representing 19 non-Campylobacter species. Application of the multiplex PCR method to whole-cell lysates obtained from 108 clinical and environmental thermotolerant Campylobacter isolates resulted in 100% correlation with biochemical typing methods.

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A Multiplex Polymerase Chain Reaction for the Differentiation of Campylobacter jejuni and Campylobacter coli from a Swine Processing Facility and Characterization of Isolates by Pulsed-Field Gel Electrophoresis and Antibiotic Resistance Profiles

Cited by 58 publications

References 38 publications

Campylobacter coliin Swine Production: Antimicrobial Resistance Mechanisms and Molecular Epidemiology

Campylobacter coliin Swine Production: Antimicrobial Resistance Mechanisms and Molecular Epidemiology

Identification of host-associated alleles by multilocus sequence typing of Campylobacter coli strains from food animals

Differentiation of Campylobacter coli , Campylobacter jejuni , Campylobacter lari , and Campylobacter upsaliensis by a Multiplex PCR Developed from the Nucleotide Sequence of the Lipid A Gene lpxA

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