492015 Seminar, Institute of Food Science & Biotechnology, NCHU
Title:
Activity of lysozyme on Lactobacillus hilgardii strains isolated from Port wine
Authors:
Rita Dias, Eduardo Vilas-Boas, Francisco M. Campos, Tim Hogg, José António Couto*
Source: Food Microbiology 49(2015)6-11
Date:2015/06/14
Speaker:蔣紹元
Advisor: Chen Chin-Shuh Professor
Abstract
This work evaluated the effect of lysozyme on lactobacilli isolated from Port wine.
Bacterial growth experiments were conducted in MRS/TJ medium and inactivation
studies were performed in phosphate buffer (KH2PO4), distilled water and wine
supplemented with different concentrations of lysozyme. The response of bacteria to
lysozyme was found to be highly strain dependent. Some strains of Lactobacillus
hilgardii together with Lactobacillus collinoides and Lactobacillus fructivorans were found
to be resistant to concentrations of lysozyme as high as 2000 mg/L. It was observed that
among the L. hilgardii taxon the resistant strains possess an S-layer coat. Apparently, the
strains of L. collinoidesand L. fructivoransstudied are also S-layer producers as suggested
by the total protein profile obtained by SDS-PAGE. Thus, the hypothetical protective
role of the S-layer against the action of lysozyme was investigated. From the various
treatments used to remove the protein from the surface of the cells, the one employing
LiCl (5 M) was the most effective. LiCl pre-treated cells exposed to lysozyme (2000 mg/L)
in KH2PO4 buffer maintained its resistance. However, when cells were suspended in
distilled water an increased sensitivity to lysozyme was observed. Moreover, it was found
that the addition of ethanol (20% v/v) to the suspension medium (distilled water)
triggered a strong inactivation effect especially on cells previously treated with LiCl
(reduction of>6 CFU log cycles). The results suggest that the S-layer exerts a protective
effect against lysozyme and that the cell suspension medium influences the bacteriolysis
efficiency. It was also noted that ethanol enhances the inactivation effect of lysozyme.
Introduction
The occurrence and activity of lactic acid bacteria (LAB) in wine is widely documented.
LAB can interact with a wine, changing its composition in a beneficial way through
malolactic fermentation (MLF) and associated metabolic transformations that occur
during this process. Besides the desirable MLF in wines, some LAB strains may
contribute to a decrease in wine quality due to the production of acetic and lactic acids
from the metabolism of sugars. The heterolactic degradation of sugars may lead to a
detectable increase in the oenological parameter“volatile acidity”. Serious wine spoilage
of this type may occur in table as well as in fortified wines at some time after the
completion of the alcoholic fermentation.
The microbiological stability of wines is mostly assured by the use of SO2. SO2 exhibits
good antimicrobial and antioxidant capacities but can cause allergic-type reactions in
sensitive individuals, especially asthmatics, and may have negative sensory properties.
Material and Methods
Bacteria and growth conditions
↓
Bacterial growth measurement
Influence of lysozyme on
isolated from growth of
lactobacilli strains
Inactivation of lactobacilli
Port wine by lysozyme
SDS-PAGE of whole-cell protein samples
S-layer extraction and analysis of cell viability
Statistical analysis
Results
Finally, the effect of lysozyme on L. hilgardii 35 was studied in Port wine at the
concentration of 500 mg/L, the maximum permitted by the EU regulation. Both LiCl
treated and non-treated cells exhibited low sensitivity to lysozyme, with only a slight
effect (<1 log cycle ) on cell survival being observed (data not shown).
Conclusions
This effect may be related to the recognized action of this agent on the cell membrane
and/or by the interference with the molecular bonds in the peptidoglycan inducing cell
lysis.
References
Couto, J.A., 1996. Studies on Diversity, Taxonomy and Physiology of Ethanol Tolerant
Lactobacilli Isolated from Douro Fortified Wine (PhD thesis). Escola Superior de
Biotecnologia da Universidade Católica Portuguesa, Porto, Portugal.
Ribéreau-Gayon, P., Glories, Y., Maujean, A., Dubordieu, D., 2006. Handbook of
Enologyethe Microbiology of Wine and Vinifications, second ed. John Wiley
and Sons, Chichester, UK.
Fugelsang, K.C., Edwards, C.G., 2007. Wine Microbiology, Practical Applications and
Procedures, second ed. Springer, New York.