Development of a sensitive DNA microarray suitable for rapid detection of Campylobacter spp.

Keramas, Georgios; Bang, Dang Duong; Lund, Marianne; Madsen, Mogens; Rasmussen, Svend Erik; Bunkenborg, H.; Telleman, Pieter; Christensen, Claus Bo Vöge

doi:10.1016/s0890-8508(03)00052-5

Cited by 83 publications

(53 citation statements)

References 40 publications

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“…Recently, a DNA microarray suitable for use for the mass screening of Campylobacter directly from chicken fecal samples was developed (14). In Denmark samples are collected from broiler chickens as individual fecal swabs and 10 swabs are pooled for testing, as recommended by the Danish National Surveillance for Campylobacter in Broiler Production program.…”

Section: Discussionmentioning

confidence: 99%

“…1). The multiplex PCR was shown to be highly specific, since no PCR amplicons were detected when the method was applied to DNA from a set of bacterial reference strains, including different Campylobacter species, Campylobacter-related bacteria, and other enterobacteria (14). By use of the multiplex PCR in combination with capillary electrophoresis for PCR amplicon analysis, 23 more samples were found to be positive for members of the genus Campylobacter, and 2 of these samples were positive for both C. jejuni and C. coli (Table 3).…”

Section: Discussionmentioning

confidence: 99%

“…However, it has been shown that PCR-based methods can detect not only viable Campylobacter cells but also noncultivable and dead Campylobacter cells (2,20), an advantage in, e.g., food testing, in which noncultivable forms present a potential risk to humans. Recently, we showed that the use of the DNA microarray increased the sensitivity by a factor of 100 in comparison to the sensitivity of capillary chip electrophoresis with the same PCR samples (14).…”

Section: Discussionmentioning

confidence: 99%

“…The sequences of the PCR primers, the oligonucleotide capture probes, and an oligonucleotide used as a positive hybridization control were described previously (14) and are summarized in Fig. 1 and Table 1.…”

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confidence: 99%

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Use of Culture, PCR Analysis, and DNA Microarrays for Detection of Campylobacter jejuni and Campylobacter coli from Chicken Feces

Keramas

Bang²,

Lund³

et al. 2004

J Clin Microbiol

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A DNA microarray for detection of Campylobacter spp. was recently developed and applied to detect Campylobacter spp. directly from chicken feces. Sixty-five pooled chicken cloacal swab samples from 650 individual broiler chickens were included in the study. The results of Campylobacter sp. detection obtained with DNA microarrays were compared to those obtained by conventional culture and gel electrophoresis. By conventional culture, 60% of the samples were positive for either Campylobacter jejuni or Campylobacter coli. By PCR and capillary electrophoresis, 95% of the samples were positive for Campylobacter spp., whereas with DNA microarrays all samples were positive for Campylobacter spp. By application of DNA microarray analysis, the isolates in 4 samples (6%) could not be identified to the species level, whereas by PCR-capillary electrophoresis, the isolates in 12 samples (19%) remained unidentified. Interestingly, PCR-capillary electrophoresis analysis revealed that two (3%) of the samples were positive for both C. jejuni and C. coli, while DNA microarray analysis revealed that nine (14%) of the samples were positive for both species. Of 65 samples, 2 samples were identified to contain C. coli by conventional culture but were positive for C. jejuni by both PCR-capillary electrophoresis and DNA microarray analysis. The discrepancy between the methods is discussed.Campylobacter is the most common cause of intestinal disorders in humans in many industrial countries. An incidence of 86 campylobacteriosis cases per 100,000 inhabitants in 2001 (3) makes Campylobacter infection the most common food-borne pathogen in Denmark.Poultry and poultry products are considered important sources of human campylobacteriosis and play important roles in disease transmission (5,6,8,11). In Denmark, a systematic sampling procedure for continued evaluation of the prevalence rates and epidemiology of Campylobacter in poultry has been in place since 1998 (3). At the retail level, 30 to 40% of poultry at slaughter have been reported to be contaminated with Campylobacter (31). Isolation and identification of Campylobacter by the conventional culture method are laborious due to the slow growth rate, the lack of phenotypic differences in the bacteria, and often, the failure to identify Campylobacter to the species level by the available culture methods. There is a need for the development of a sensitive, rapid method for Campylobacter detection and identification to the species level.Several PCR assays have successfully been applied to the detection of Campylobacter spp. in water (15, 26), some dairy products (9,12,29,32), and chicken litter (13). The PCR method allows detection not only of viable bacteria but also of noncultivable forms of Campylobacter (12, 32). A multiplex PCR assay suitable for mass screening to detect Campylobacter directly from chicken feces has been developed (1). Agarose gel electrophoresis is often used for the PCR assays. Gel electrophoresis has a number of drawbacks, however, such as a poor ability to different...

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Section: Discussionmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

mentioning

confidence: 99%

See 2 more Smart Citations

Use of Culture, PCR Analysis, and DNA Microarrays for Detection of Campylobacter jejuni and Campylobacter coli from Chicken Feces

Keramas

Bang²,

Lund³

et al. 2004

J Clin Microbiol

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“…Oligonucleotide microarrays based on 16S or 23S rDNA sequences have been used for detection of Campylobacter from fecal cloacal swabs (13), for detection of intestinal bacterial species from human fecal samples (33), and for detection of bacteremia-causing bacteria from positive blood culture broths (1). Other approaches have been developed in addition to these oligonucleotide-based methods targeting bacterial rRNA sequences.…”

Section: Discussionmentioning

confidence: 99%

Use of an Oligonucleotide Array for Laboratory Diagnosis of Bacteria Responsible for Acute Upper Respiratory Infections

Roth¹,

Jalava

Ruuskanen

et al. 2004

J Clin Microbiol

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We developed a diagnostic array of oligonucleotide probes targeting species-specific variable regions of the genes encoding topoisomerases GyrB and ParE of respiratory bacterial pathogens. Suitable broad-range primer sequences were designed based on alignment of gyrB/parE sequences from nine different bacterial species. These species included Corynebacterium diphtheriae, Fusobacterium necrophorum, Haemophilus influenzae, Legionella pneumophila, Moraxella catarrhalis, Mycoplasma pneumoniae, Staphylococcus aureus, Streptococcus pneumoniae, and Streptococcus pyogenes. Specific probe sequences were selected by comparative analysis against the European Bioinformatics Database, as well as gyrB/parE sequences generated for this study. To verify specificity, at least six initial oligonucleotide probe sequences per bacterial species were tested by hybridization on a solid glass support using culture collection strains as templates. Finally, three oligonucleotide probes per bacterial species were utilized to examine 65 middle ear fluid and 29 throat swab samples. The sensitivities of the developed assay compared to classic culture from middle ear fluid samples for H. influenzae, M. catarrhalis, and S. pneumoniae were 96 (93 for culture), 73 (93 for culture), and 100% (78% for culture), respectively. No cross-reactivity with bacterial species belonging to the normal oral flora was observed when the 29 throat swab samples were studied. The sensitivity of the assay to detect S. pyogenes from these samples was 93% (80% for culture). These results provide a proof of concept for the diagnostic use of microarray technology based on broad-range topoisomerase gene amplification, followed by hybridization and specific detection of bacterial species.Modern high-density arrays of DNA probes (microarrays) have proved to be powerful tools to study complex genetic profiles of cellular life. However, only a few experimental assays based on this valuable tool have been introduced for microbiological diagnostics. We present here novel data demonstrating the application of microarray technology combined with broad-range PCR amplification of topoisomerase genes to the diagnosis of bacterial respiratory infections.Broad-range bacterial PCR is based on the use of primers that recognize conserved sequences of bacterial genes encoding essential molecules. The resulting DNA amplicons contain variable regions that provide a basis for the analysis of phylogenetic relationships and identification of different bacterial species. Furthermore, some genetically hypervariable regions provide targets for bacterial-species-specific oligonucleotide sequences. These oligonucleotides that target hypervariable regions may be used in a diagnostic array of probes for detection of bacterial pathogens.The use of broad-range ribosomal DNA (rDNA) PCR technologies for identification of bacterial and fungal culture isolates in the clinical laboratory is well established (3, 28), and there are commercial kits available for that purpose. However, the kits are not widely...

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Molecular Technologies for Detecting and Characterizing Pathogens

Duffy¹,

Catarame²

2007

Advances in Food Diagnostics

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Development of a sensitive DNA microarray suitable for rapid detection of Campylobacter spp.

Cited by 83 publications

References 40 publications

Use of Culture, PCR Analysis, and DNA Microarrays for Detection of Campylobacter jejuni and Campylobacter coli from Chicken Feces

Use of Culture, PCR Analysis, and DNA Microarrays for Detection of Campylobacter jejuni and Campylobacter coli from Chicken Feces

Use of an Oligonucleotide Array for Laboratory Diagnosis of Bacteria Responsible for Acute Upper Respiratory Infections

Molecular Technologies for Detecting and Characterizing Pathogens

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