Abstract
Listeria innocua is an important hygiene indicator bacterium in food industries because it behaves similar to Listeria monocytogenes, which is pathogenic to humans. PFGE is often used to characterize bacterial strains and to track contamination source. However, because PFGE is an expensive, complicated, time-consuming protocol, and poses difficulty in data sharing, development of a new typing method is necessary. MLVA is a technique that identifies bacterial strains on the basis of the number of tandem repeats present in the genome varies depending on the strains. MLVA has gained attention due to its high reproducibility and ease of data sharing. In this study, we developed a MLVA protocol to assess L. innocua and evaluated it by tracking the contamination source of L. innocua in an actual food manufacturing factory by typing the bacterial strains isolated from the factory. Three VNTR regions of the L. innocua genome were chosen for use in the MLVA. The number of repeat units in each VNTR region was calculated based on the results of PCR product analysis using capillary electrophoresis (CE). The calculated number of repetitions was compared with the results of the gene sequence analysis to demonstrate the accuracy of the CE repeat number analysis. The developed technique was evaluated using 60 L. innocua strains isolated from a food factory. These 60 strains were classified into 11 patterns using MLVA. Many of the strains were classified into ST-6, revealing that this MLVA strain type can contaminate each manufacturing process in the factory. The MLVA protocol developed in this study for L. innocua allowed rapid and easy analysis through the use of CE. This technique was found to be very useful in hygiene control in factories because it allowed us to track contamination sources and provided information regarding whether the bacteria were present in the factories.
Highlights
Listeria is a genus of gram-positive, facultatively anaerobic, nonspore forming bacteria [1]
We developed an multilocus variable number of tandem repeat analysis (MLVA) method effective for distribution assessment and tracking the contamination sources of
L. monocytogenes, which is pathogenic to humans, is strictly controlled for in food factories, and a number of methods for tracking contamination sources of this pathogen have been published [21], [22]
Summary
Listeria is a genus of gram-positive, facultatively anaerobic, nonspore forming bacteria [1]. Listeria spp. have been isolated from a variety of environmental sources, such as soil [2], river water [3], farm environments [4], and animal feed [5]. Listeria spp. have been isolated from some foods [6], [7], [8] and food processing environments [9]. Because Listeria monocytogenes, a member of Listeria spp., can cause serious listeriosis infections in humans and ruminants, this pathogen is of significant health concern. L. ivanovii can cause listeriosis infections in ruminants but rarely in humans [10]. Human listeriosis results from ingestion of L. monocytogenes via contaminated foods. The United States has a zerotolerance policy for contamination in processed foods, with certain
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