Isolation, identification and molecular characterization of (Cronobacter spp.) Cronobacter sakazaki virulence factors from varied foods and environmental samples

Cronobacter sakazakii is an opportunistic pathogen that causes life-threatening infections, especially in preterm and low birth weight infants. Although it has been detected in various nutrients and other environmental sources, only contaminated powder infant formulas have been epidemiologically associated with infections caused by C. sakazakii. Exposure to environmental and foodborne C. sakazakii species pose a serious human health risk. A sufficient number of studies on the detection of C. sakazakii species by rapid and sensitive molecular methods are not available in our country. Molecular identification methods are one of the most important innovations of developing technology in the field of microbiology. The addition of new molecular identification techniques day by day is of great benefit to scientific studies carried out for different purposes. The aim of this study was to genotypically identify C. sakazakii species in various nutrients and environmental samples in a sensitive and reliable way by using fingerprints of OmpA, OmpX, zpx and Cpa genes which enable C. sakazakii to easily reach organs and tissues in all mammals. Within the scope of the study, a total of 120 nutrients and environmental samples, including 20 unpackaged milk powder, 20 packaged starch, 20 packaged rice flour, 20 packaged semolina, 20 packaged baby formulas and 20 environmental samples, collected in the provincial borders of Istanbul in 2015-2016, were analysed using the rapid test kit API 20E (BioMérieux, France) and real-time polymerase chain reaction (RT-PCR) in order to compare the cultural, biochemical and molecular methods. As a result, RT-PCR gene characterization was performed for 7 unpackaged milk powder, 2 packaged rice flour, 1 packaged semolina and 3 environmental samples identified with API 20E. OmpA, OmpX, zpx and Cpa virulence genes were detected in all suspected isolates. The parameters between different sample groups in which C. sakazakii were detected were examined and Fisher Freeman Halton test was used for the comparison of the data between the parameters based on the findings. Incidence rate ratio of C. sakazakii in unpackaged milk powder (35%) was significantly higher than that of starch (0%), semolina (5%), rice flour (%10), environmental (15%), and baby formula (0%) (p<0.05). Based on the findings, the presence of OmpA, OmpX, zpx and Cpa genes, which display virulent characteristics, and are among the genotypic factors of C. sakazakii species of nutrient and environmental origin were shown. Keywords: Cronobacter sakazakii, virulence factors, Real-Time PCR