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Pathogenic Yersinia enterocolitica is involved in yersiniosis through expression of chromosome-borne or plasmid-borne virulence factors. Yersinia enterocolitica is a cold-tolerant pathogen frequently isolated from refrigerated or frozen foods. However, little attention has been focused on the prevalence of pathogenic Y. enterocolitica in refrigerated or frozen dairy samples in China. In this study, we developed a new duplex PCR targeting the plasmid-borne virF gene and chromosome-borne ail gene for detection of pathogenic Y. enterocolitica isolates. We established a detection limit for the duplex PCR of 6.5 × 10(2)cfu/mL in artificially contaminated dairy samples. In addition, the duplex PCR could detect directly 4.5 to 5.7 cfu of Y. enterocolitica in 5 mL of brain heart infusion broth after 6 h of enrichment at 28 °C. A newly developed dot hybridization assay further confirmed specificity of the duplex PCR for detection of virulent Y. enterocolitica. Furthermore, 13 Y. enterocolitica and 5 pathogenic strains, from 88 commercial frozen or refrigerated dairy products, were detected successfully by the China National Standard method (GB/T4789.8-2008) and the duplex PCR, respectively. Finally, biotypes and serotypes of pathogenic Y. enterocolitica strains were further characterized. The duplex PCR developed here is reliable for large-scale screening, routine monitoring, and risk assessment of pathogenic Y. enterocolitica in refrigerated or frozen dairy products.

Vibrio parahaemolyticus is an important seafood‐borne pathogen associated with gastrointestinal disorders in humans. In this study, the efficacy of a loop‐mediated isothermal amplification (LAMP) combined with immunocapture assay was firstly developed for detection of V. parahaemolyticus in pure culture, artificially and naturally contaminated seafood samples. The detection limit was 7.3 × 101 cfu/mL for pure culture and artificially contaminated samples by IC‐LAMP, compared with 7.3 × 102 cfu/mL by LAMP. Furthermore, among 156 natural seafood samples, 13, 13 and 11 of samples were positive for V. arahaemolyticus by IC‐LAMP, LAMP and culture‐based method, respectively. V. parahaemolyticus was isolated from 11 of the 156 seafood samples by culture‐based method, all of which were positive by IC‐LAMP assay. A comparison between LAMP, IC‐LAMP and culture‐based method indicated that IC‐LAMP was sensitive, specific and reliable for monitoring V. parahaemolyticus contamination in seafood samples. Practical Application Development of rapid, sensitive and specific methods for detection of V. parahaemolyticus is of utmost importance for monitoring and controlling contaminated seafood. In this study, loop‐mediated isothermal amplification combined with the newly developed immunocapture showed higher detection limit than that by LAMP and more specific compared with that by culture‐based method. The IC‐LAMP was fit for monitoring V. parahaemolyticus contamination in seafood samples.

Biofilm of Cronobacter sakazakii on food contact surfaces is the important source of persistent contamination in powdered infant formula. This study aimed to assess the effects of Ca 2+ and Mg 2+ on biofilm formation. Results indicated that the biofilm-forming ability was strain specific and divalent cation concentrations dependent. Most of C. sakazakii formed the highest amount of biomass at 1.50% MgCl2 (17 strains, 74%) and 0.25% CaCl2 (10 strains, 43.4%). Finally, confocal laser scanning microscope (CLSM) further indicated that effects of divalent cation were attributable to changing properties of extracellular matrix. The findings presented here provide new insights for control of biofilm formation in dairy products. PRACTICAL APPLICATIONSC. sakazakii is an important foodborne pathogen involved in infections of infant. Control of C. sakazakii contamination in powdered infant formula is an intractable problem in production of powdered infant formula. In this study, effects of Ca 2+ and Mg 2+ on the biofilm formation were performed. Furthermore, the changes of extracellular matrix in biofilm were revealed by CLSM. The results contribute to useful information for development of essential measures for precaution and control of biofilm of C. sakazakii.