title: The use of FTA cards for transport and detection of gyrA mutation of Campylobacter jejuni from poultry.
abstract: The purpose of the present study was to evaluate a technique involving the use of commercially available FTA classic card (Whatman) for transporting and detection of DNA to use in PCR analysis and genetic sequencing of Campylobacter jejuni of poultry origin. Fifty isolates of Campylobacter jejuni were obtained from broiler carcasses in Rio Grande do Sul, Brazil. Antimicrobial susceptibility testing to ciprofloxacin revealed that all 50 isolates were resistant to ciprofloxacin. Each isolate was transferred to Brucella broth tubes and incubated overnight at 41.5°C. Cell cultures were diluted to match a McFarland Turbidity Standard 0.5, and 110 μL of the cell suspension were applied to one circle on Whatman FTA classic cards. The samples were then covered and allowed to dry at room temperature. Cards were identified and stored at room temperature until further use (3 mo after collection). FTA cards were shipped for analysis to the Department of Poultry Science, University of Arkansas. Amplification of the Campylobacter gyrA gene was successful and demonstrated strong bands for a large amplicon for all 50 samples preserved on FTA cards. Mutations present in each gene were confirmed by DNA sequencing. Then, 7 samples were chosen for the sequencing. The detection of a mutation regarding ciprofloxacin-resistant isolates revealed that 7 samples had a mutation in the gyrA gene. In conclusion, the characteristics of the profiles suggest that the DNA has maintained its integrity after 3 mo of storage at room temperature and is a suitable template for PCR and sequencing from Campylobacter samples. The application of this technology has potential in numerous methodologies, especially when working in remote areas and in developing countries where access to laboratory facilities and equipment is limited.
authors.affiliation: Center for Diagnostics and Research in Avian Pathology, College of Veterinary Medicine, Federal University of Rio Grande do Sul, Av. Bento Gonçalves, 8824, Agronomia, Porto Alegre, Rio Grande do Sul, Brazil yuli_melisasierra@yahoo.com.
authors.forename: Y M
authors.initials: YM
authors.lastname: Sierra-Arguello
authors.affiliation: Department of Poultry Science, University of Arkansas, Fayetteville.
authors.forename: O
authors.initials: O
authors.lastname: Faulkner
authors.affiliation: Department of Poultry Science, University of Arkansas, Fayetteville.
authors.forename: G
authors.initials: G
authors.lastname: Tellez
authors.affiliation: Department of Poultry Science, University of Arkansas, Fayetteville.
authors.forename: B M
authors.initials: BM
authors.lastname: Hargis
authors.affiliation: Center for Diagnostics and Research in Avian Pathology, College of Veterinary Medicine, Federal University of Rio Grande do Sul, Av. Bento Gonçalves, 8824, Agronomia, Porto Alegre, Rio Grande do Sul, Brazil.
authors.forename: V
authors.initials: V
authors.lastname: Pinheiro do Nascimento
mesh_terms: D000818:Animals; D001426:Bacterial Proteins / Q000235:genetics* / Q000378:metabolism; D016123:Campylobacter jejuni / Q000235:genetics*; D002645:Chickens / Q000382:microbiology*; D027081:DNA Gyrase / Q000235:genetics* / Q000378:metabolism; D009154:Mutation; D016133:Polymerase Chain Reaction / Q000662:veterinary*; D017422:Sequence Analysis, DNA / Q000662:veterinary*
chemical_list: D001426:Bacterial Proteins; D027081:DNA Gyrase
keywords: Campylobacter jejuni; DNA; FTA cards; gyrA; sequencing