Research Article
Volume 16 Issue 2 - 2021
Study of the Nature of Contamination of Raw Milk with Bacteria of the Genus Campylobacter Using Traditional Microbiological Methods and Quantitative PCR Analysis
Efimochkina Natalia Ramazanovna*, Stetsenko Valentina Valerievna, Markova Yulia Mikhailovna, Bykova Irina Borisovna, Pichugina Tatiana Viktorovna, Polyanina Anna Sergeevna, Minaeva Lyudmila Pavlovna and Sheveleva Svetlana Anatolyevna
Federal Research Center of Nutrition, Biotechnology and Food Safety, Russia
*Corresponding Author: Efimochkina Natalia Ramazanovna, Federal Research Center of Nutrition, Biotechnology and Food Safety, Russia.
Received: January 12, 2021; Published: Janauary 30, 2021




Abstract

Introduction: Campylobacter gastroenterocolitis is the most common foodborne acute infectious zoonotic disease. One of the important factors in the transmission of infection is contaminated dairy products, therefore, the assessment of contamination of dairy raw materials with Campylobacter is necessary to develop effective measures to suppress the growth of the pathogen and ensure the safety of products.

Aim of the Study: The aim of the study was to study the microbial background of raw milk samples and the nature of their contamination with thermophilic bacteria of the genus Campylobacter.

Materials and Methods: 60 samples of raw milk from the central regions of the Russian Federation and 48 samples of raw milk experimentally infected with Campylobacter were studied. To assess microbial contamination of milk, the amount of extraneous microflora, including coliform bacteria (BCB), was determined. The identification and determination of the number of bacteria of the genus Campylobacter was carried out by cultural methods in comparison with quantitative analysis by the method of polymerase chain reaction (PCR analysis). For PCR, primers were used that detected the species-specific sequence of 16s rRNA of C. jejuni, the presence of the cytolethal toxin cdtB gene and the ciaB invasion gene.

Results and Discussion: A significant part of the raw milk samples examined (31.6%) were characterized by high levels of microbial contamination, exceeding 106 CFU/cm3. Gram-negative bacteria are the dominant type of bacterial microflora, their levels were comparable to the revealed values of the total number of microorganisms. BGKP were found in all the samples studied, their number in 90% of the samples reached 105 CFU/cm3 and in some samples - 107 CFU/cm3. Detection rate of Campylobacter spp. in raw milk was 8.3% and their number ranged from 0.1 to 100 CFU/cm3 (on average 2.0 x 10 CFU/cm3). All the isolated Campylobacter strains were identified as C. jejuni by a complex of phenotypic characters. Comparative analysis of contamination of raw milk with Campylobacter by RT-PCR was carried out. Most of the samples (over 60%) were positive for the genome-specific genomic sequence of 16SrRNA, while they were characterized by the highest values of the total bacterial contamination and the number of BHCPs. The use of a multi-primer approach (simultaneous testing for the presence of 16SrRNA and the cytolethal toxin cdtB gene of C. jejuni) reduced the number of positive cases of Campylobacter DNA detection to 16.6%, which suggests that the cdtB gene is more informative in relation to the detection of viable, including uncultured, cells. with toxigenicity. A tentative assessment of the results in a quantitative format indicated high detectable levels of Campylobacter bacteria - 104 - 106-5 genomic equivalents/cm3, which indicates the possible presence of viable Campylobacter cells in the test material with a significantly higher frequency than that established by culture.

Conclusion: Microbiological methods of inoculation at low levels of contamination with Campylobacter do not ensure their reliable detection due to massive contamination of raw materials with extraneous microflora. Campylobacter spp. were detected by the culture method in 8.3% of cases, while the use of multi-primer PCR analysis with the cdtB and ciaB genes makes it possible to double the detection of C. jejuni in raw milk samples.

 

Keywords: Bacteria of the Genus Campylobacter; Raw Milk; PCR Analysis; Microbial Contamination

References

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Citation: Efimochkina Natalia Ramazanovna., et al. “Study of the Nature of Contamination of Raw Milk with Bacteria of the Genus Campylobacter Using Traditional Microbiological Methods and Quantitative PCR Analysis”. EC Nutrition 16.2 (2021): 86-92.

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