Journal of the Korean Society of Food Culture (한국식생활문화학회지)

Korean Society of Food Culture (한국식생활문화학회)
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Strain-specific Detection of Kimchi Starter Leuconostoc mesenteroides WiKim33 using Multiplex PCR

  • Lee, Moeun (Advanced Process Technology and Fermentation Research Group, World Institute of Kimchi) ;
  • Song, Jung Hee (Advanced Process Technology and Fermentation Research Group, World Institute of Kimchi) ;
  • Park, Ji Min (Advanced Process Technology and Fermentation Research Group, World Institute of Kimchi) ;
  • Chang, Ji Yoon (Advanced Process Technology and Fermentation Research Group, World Institute of Kimchi)
  • Received : 2019.03.21
  • Accepted : 2019.04.22
  • Published : 2019.04.30
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Abstract

Leuconostoc spp. are generally utilized as kimchi starters, because these strains are expected to have beneficial effects on kimchi fermentation, including improvement of sensory characteristics. Here, we developed a detection method for verifying the presence of the kimchi starter Leuconostoc mesenteroides WiKim33, which is used for control of kimchi fermentation. A primer set for multiplex polymerase chain reaction was designed based on the nucleotide sequence of the plasmids in strain WiKim33, and their specificity was validated against 45 different strains of Leuconostoc spp. and 30 other strains. Furthermore, the starter strain consistently tested positive, regardless of the presence of other bacterial species in starter kimchi during the fermentation period. Our findings showed that application of a strain-specific primer set for strain WiKim33 presented a rapid, sensitive, and specific method for detection of this kimchi starter strain during natural kimchi fermentation.

Keywords

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Agarose gel electrophoresis of multiplex PCR products obtained from pure cultures of Leuconostoc mesenteroides WiKim33 in MRS broth. Lane M, size marker (100-bpladder); lane 1, primer 1; lane 2, primer 2; lane 3, primer 3; lane 4, primer 4; lane 5, integrated primers.

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Specificity test of the designed primer set. Lane M, size marker (100-bp ladder); lane 1, WiKim33; lanes 2-45, Leuconstoc spp.; lanes 46-76, other strains .

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Determination of the detection limit of the WiKim33-specific primer set. (A) Dilution with distilled water at the indicated concentrations. Lane M, size marker (100-bpladder); lanes 1-10, 1.8×109 CFU/mL (277 ng of DNA) to 1.8×100 CFU/mL (0.000277 pg of DNA). (B) Diluted with kimchi filtrate (10 ng of DNA for each sample). Lane M, size marker (100-bp ladder); lanes 1-10, kimchi filtrate with 1.8×109 CFU/mL to 1.8×100 CFU/mL of WiKim33.

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Fermentation characteristics of starter kimchi containing Leuconostoc mesenteroides WiKim33. (A) pH profiles and titratable acidity. (B) Total viable bacteria and LAB counts of kimchi according to storage temperature.

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Application of kimchi to the WiKim33-specific detection system. (A) Analysis of the dominance ratio by measuring total viable bacteria and WiKim33 counts in kimchi. (B) Application of the multiplex PCR system to starter kimchi.

Summarya of bacterial strains used in the PCR specificity test.

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Multiplex PCR primer sets used in this study

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Bacterial strains used in the PCR specificity test

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Bacterial strains used in the PCR specificity test (continued)

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