Detection of salmonella in poultry faeces by molecular means in comparison to traditional bacteriological methods | Primerjava molekularnih in klasicnih bakterioloskih metod za ugotavljanje salmonel v kokosjem blatu
2010
Kusar, D., Ljubljana Univ. (Slovenia). Veterinary Fac., Microbiology and Parasitology Inst. | Pate, M. | Mićunović, J. | Bole-Hribovsek, V. | Ocepek, M.
Comparison of traditional (cultivation-dependent) and molecular (nucleic acid-based) bacteriological methods was performed to detect Salmonella in reference capsules containing quantified amounts of Salmonella enterica subsp. enterica serovars Panama, Typhimurium or Enteritidis and in poultry faeces that was naturally contaminated with Salmonella or Salmonella-negative but spiked with reference materials. Traditional techniques were performed according to ISO 6579 using different enrichment (MSRV, MKTTn and RVS, respectively) and isolation plating media (XLD, BGA and Rambach agar, respectively). Molecular detection was preceded by the pre-enrichment step. Detection efficiency of two DNA isolation kits, namely High Pure foodproof I Kit (Roche Diagnostics, Germany) and QIAamp DNA Stool Mini Kit (Qiagen, Germany), in combination with classical and real-time PCR assay was compared. Results showed that traditional and molecular detection of Salmonella was unambiguous for reference control capsules, but was hindered for faecal samples. RVS medium was less appropriate than MSRV and MKTTn. Combination of MKTTn with Rambach agar plates generated the highest number of positive results with traditional approach. However, recommendation of using the semisolid MSRV medium was confirmed as it enabled detection of Salmonella in high proportion of samples, which was the least variable depending on the selection of isolation plating media. In contrast to culture-based methods, the molecular approach, especially a combination of High Pure foodproof I Kit and real-time PCR assay, enabled successful detection in all Salmonella-positive samples and should therefore be considered an important supplement to traditional protocol for Salmonella detection in foodstuffs.
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