Biopreservation of fresh-cut salads using bacteriocinogenic lactic acid bacteria isolated from commercial produce
Deana Sharpe
Thesis, Master of Science( Food Science and Technology), Dalhousie University. 111 pages. 2009.
2009
บทคัดย่อ
The food industry is seeking novel biopreservatives such as active lactic acid bacteria (LAB) cultures and/or their bacteriocins to control psychrotrophic pathogens including Listeria monocytogenes. However, most bacteriocins have been extracted from and applied to meat and dairy products with little emphasis on fresh produce. The objective of this study was to isolate and characterize bacteriocinogenic LAB strains of plant origin for applications on fresh-cut fruit and vegetable products.
Fourteen different vegetable products were investigated for content of native LAB. From 218 potential LAB isolates, 92 were confirmed catalase negative and gram positive. The antimicrobial properties of cell-free supernatants from these isolates were screened using the agar diffusion bioassay with Lactobacillus sakei and Listeria innocua as indicator organisms. Eight strains isolated from mung bean sprouts, swiss chard and mini-seedless cucumbers inhibited both indicator organisms. The presence of bacteriocin-like substances (BLS) was indicated by inhibition zones following the neutralization of pH and elimination of H 2 O2. These zones were lost following the addition of proteolytic enzymes. The eight LAB isolates were identified using 16S rRNA gene sequencing as Lactococcocus lactis ssp. lactis (1) and Enterococcus faecium (7). Antimicrobial effectiveness was tested against common fresh-cut produce pathogens and spoilage organisms at 5 and 20 °C. The BLS showed significant antimicrobial effects against Li. innocua , with a mean inhibition zone diameter of 9.7 mm at 5 °C and 10.2 mm in diameter at 20 °C. However, only the organic acids and not the H2 O2 or BLS showed strong antimicrobial effects against Pseudomonas fluorescens, Erwinia carotovora, Bacillus cereus and weak effects against Leuconostoc mesenteroides . A further investigation of the eight BLS producing LAB against Penicillium expansum, Botrytis cinerea and Monilinia fructicola spores using the microtiter plate method showed that inhibition was due to mainlt to the production of organic acids and partially due to H2 O2 not the BLS.
The effects of both L. lactis and Ent. faecium on the natural microflora and on Listeria inoculated fresh-cut salads were investigated. Bacteriocinogenic LAB applied to fresh-cut salad inoculated with Li. innocua reduced Listeria counts significantly (ρ=0.005) when compared to the control plates with LAB, during a 10 day storage trial at 5 °C. The addition of LAB was able to significantly reduce the counts of natural Listeria sp. (ρ=0.033), yeasts (ρ=0.011), Pseudomonas sp. (ρ=0.010) and total conforms (ρ=0.011) in comparison to the controls. Overall, results suggest that L. lactis and Ent. faecium could be used as a successful biopreservative because they were effective at controlling both spoilage and pathogenic organisms when applied as protective cultures to fresh-cut salad during cold-storage, thereby improving the product shelf life and safety.