title: Application of different molecular techniques for characterization of catalase-positive cocci isolated from sucuk.
abstract: This study was carried out for the characterization and discrimination of the indigenous Gram positive, catalase-positive cocci (GCC) population in sucuk, a traditional Turkish dry-fermented sausage. Sucuk samples, produced by the traditional method without starter culture were collected from 8 local producers in Kayseri/Turkey and a total of 116 GCC isolates were identified by using different molecular techniques. Two different molecular fingerprinting methods; namely, randomly amplified polymorphic DNA-PCR (RAPD-PCR) and repetitive extragenic palindrome-PCR (rep-PCR), were used for the clustering of isolates and identification at species level was carried out by full length sequencing of 16S rDNA. Combining the results obtained from molecular fingerprinting and 16S rDNA sequencing showed that the dominant GCC species isolated from the sucuk samples was Staphylococcus saprophyticus followed by Staphylococcus succinus and Staphylococcus equorum belonging to the Staphylococcus genus. Real-time PCR DNA melting curve analysis and high-resolution melting (HRM) analysis targeting the V1 + V3 regions of 16S rDNA were also applied for the discrimination of isolates belonging to different species. It was observed statistically different Tm values and species-specific HRM profiles for all except 2 species (S. saprophyticus and Staphylococcus xylosus) that have high 16S rDNA sequence similarity. The combination of rep-PCR and/or PCR-RAPD with 16S rRNA gene sequencing was an efficient approach for the characterization and identification of the GCC population in spontaneously fermented sucuk. On the other hand, intercalating dye assays were found to be a simple and very promising technique for the differentiation of the GCC population at species level.
authors.affiliation: Faculty of Engineering, Food Engineering Dept., Erciyes Univ, 38039, Kayseri, Turkey; Faculty of Engineering, Food Engineering Dept., Bingol Univ, Bingol, Turkey.
authors.forename: Zülal
authors.initials: Z
authors.lastname: Kesmen
authors.forename: Burcu
authors.initials: B
authors.lastname: Yarimcam
authors.forename: Hakiye
authors.initials: H
authors.lastname: Aslan
authors.forename: Esra
authors.initials: E
authors.lastname: Ozbekar
authors.forename: Hasan
authors.initials: H
authors.lastname: Yetim
mesh_terms: D000818:Animals; D001426:Bacterial Proteins / Q000378:metabolism*; D002374:Catalase / Q000378:metabolism*; D002417:Cattle; D055598:Chemical Phenomena; D016172:DNA Fingerprinting; D004269:DNA, Bacterial / Q000737:chemistry / Q000378:metabolism; D004275:DNA, Ribosomal / Q000737:chemistry / Q000378:metabolism; D004032:Diet / Q000208:ethnology; D005285:Fermentation; D063427:Food Quality; D057141:Food, Preserved / Q000032:analysis / Q000382:microbiology*; D006094:Gram-Positive Bacteria / Q000145:classification / Q000201:enzymology / Q000302:isolation & purification*; D006863:Hydrogen-Ion Concentration; D008461:Meat Products / Q000032:analysis / Q000382:microbiology*; D058889:Molecular Typing; D009691:Nucleic Acid Denaturation; D017422:Sequence Analysis, DNA; D034561:Sheep, Domestic; D013045:Species Specificity; D057790:Staphylococcus saprophyticus / Q000145:classification / Q000201:enzymology / Q000302:isolation & purification; D014421:Turkey
chemical_list: D001426:Bacterial Proteins; D004269:DNA, Bacterial; D004275:DNA, Ribosomal; D002374:Catalase
keywords: RAPD-PCR; catalase-positive cocci; intercalating dye assays; rep-PCR; sucuk