Detection and genotyping of Arcobacter and Campylobacter isolates from retail chicken samples by use of DNA oligonucleotide arrays.
To explore the use of DNA microarrays for pathogen detection in food, we produced DNA oligonucleotide arrays to simultaneously determine the presence of Arcobacter and the presence of Campylobacter in retail chicken samples. Probes were selected that target housekeeping and virulence-associated genes in both Arcobacter butzleri and thermotolerant Campylobacter jejuni and Campylobacter coli. These microarrays showed a high level of probe specificity; the signal intensities detected for A. butzleri, C. coli, or C. jejuni probes were at least 10-fold higher than the background levels. Specific identification of A. butzleri, C. coli, and C. jejuni was achieved without the need for a PCR amplification step. By adapting an isolation method that employed membrane filtration and selective media, C. jejuni isolates were recovered from package liquid from whole chicken carcasses prior to enrichment. Increasing the time of enrichment resulted in the isolation of A. butzleri and increased the recovery of C. jejuni. C. jejuni isolates were further classified by using an additional subset of probes targeting the lipooligosaccharide (LOS) biosynthesis locus. Our results demonstrated that most of the C. jejuni isolates likely possess class B, C, or H LOS. Validation experiments demonstrated that the DNA microarray had a detection sensitivity threshold of approximately 10,000 C. jejuni cells. Interestingly, the use of C. jejuni sequence-specific primers to label genomic DNA improved the sensitivity of this DNA microarray for detection of C. jejuni in whole chicken carcass samples. C. jejuni was efficiently detected directly both in package liquid from whole chicken carcasses and in enrichment broths.
Quiñones
Beatriz
B
U.S. Department of Agriculture, Agricultural Research Service, Western Regional Research Center, Produce Safety and Microbiology Research Unit, Albany, CA 94710, USA.
Parker
Craig T
CT
Janda
John M
JM
Miller
William G
WG
Mandrell
Robert E
RE
D000818:Animals; D020640:Arcobacter / Q000145:classification / Q000235:genetics / Q000302:isolation & purification*; D015373:Bacterial Typing Techniques; D002167:Campylobacter / Q000145:classification / Q000235:genetics / Q000302:isolation & purification*; D002645:Chickens / Q000382:microbiology*; D004269:DNA, Bacterial / Q000235:genetics*; D005516:Food Microbiology; D005798:Genes, Bacterial; D005838:Genotype; D008070:Lipopolysaccharides / Q000096:biosynthesis; D008460:Meat / Q000382:microbiology*; D009693:Nucleic Acid Hybridization; D020411:Oligonucleotide Array Sequence Analysis; D012680:Sensitivity and Specificity
D004269:DNA, Bacterial; D008070:Lipopolysaccharides; C023023:lipid-linked oligosaccharides