乳酸菌所產生抗菌物質抑制多重抗藥性金黃色葡萄球菌能力之分析

本研究主要目的為從動物消化道及人類糞便中分離能夠結抗多重抗藥性金黃色葡萄球菌 (Multi-resistant Staphylococcus aureus, MRSA) 之乳酸菌株。試驗首先從臨床病患及牛乳房炎檢體所分離之 107 株 S. aureus 以抗生素感受性試驗篩選出 50 株 MRSA。針對 MRSA 我們以 Agarose/agar assay 方法由豬及犬隻的腸道中篩選出 5 株能夠產生非乳酸之抗菌物質並可有效抑制 MRSA 生長之乳酸菌 (lactic acid bacteria, LAB)。利用生化試驗、16S rDNA-RFLP 及 16S-23S intergenic spacer region DNA sequence 得知 Lactococcus lactis 5-12H 及 Lactococcus lactis 6-3H 等 2 株能產生 nisin A；另 3 株 Lactobacillus animalis 30a-2、Weissella confusa C34 及 Lactobacillus reuteri 4-12E則能產生過氧化氫，其中 L. animalis 30a-2 及 W. confusa C34 為本研究首次發現能有效對抗 MRSA 之乳酸菌株，並經由 catalase test 及 TMB–MRS agar plate assay 進一步證實它們所產生之抗菌物質確實為過氧化氫，當濃度為 1010 cfu/ml 時，此二株乳酸菌所產生的 H2O2 濃度分別為 30 mM 與 15.3 mM。此外，L. animalis 30a-2 及 W. confusa C34 均能耐酸及耐膽鹽，其中 L. animalis 30a-2 可產生膽鹽水解?並對於 HeLa cells 及 Int-407 cells具有良好的吸附能力，頗具優良益生菌之開發潛力。

關鍵字

乳酸菌；多重抗藥性金黃色葡萄球菌； Lactobacillus animalis ； Weissella confusa ；過氧化氫

並列摘要

The object of this study was to isolate lactic acid bacteria (LAB), which could produce anti-microbe agents against multi-resistant Staphylococcus aureus (MRSA) from animals intestines and children faeces. We isolated 107 S. aureus from hospitals and from bovine suffering from mastitis. Among these, 50 oxacillin-resistant S. aureus (ORSA) are identified as MRSA by the antibiotics susceptibility test. By means of agarose/agar assay, we successfully obtained 5 LABs, which could produce non-lactic acid anti-microbes agents to inhibit MRSA proliferation. Utilization of biochemistry test, 16S rDNA-RFLP and analysis of 16S-23S intergenic spacer region DNA sequence, we identified two LABs, Lactococcus lactis 5-12H and Lactococcus lactis 6-3H, which could produce nisin A and three LABs, Lactobacillus animalis 30a-2, Weissella confusa C34, and Lactobacillus reuteri 4-12E, which could produce hydrogen peroxide. L. animalis 30a-2 and W. confusa C34 were the first found LAB with activity against MRSA via the actior of hydrogen peroxide that have been provided by catalase test and TMB–MRS agar plate assay. 1010 cfu/ml of L. animalis 30a-2 and W. confusa C34 could produce approximate 30 mM and 15.3 mM hydrogen peroxide by Agarose/agar assay. In addition, L. animalis 30a-2 and W. confusa C34 are also acid tolerant and bile-salt resistant. Further more, L. animalis 30a-2 had bile-salt hydrolase (BSH) activity and could adhere excellently on HeLa cells and Int-407 cells. Taken together the nearly isolated LABs harbor the anti-MRSA and probiotic properties.

並列關鍵字

lactic acid bacteria ； multi-resistant Staphylococcus aureus ； hydrogen peroxide ； Lactobacillus animalis ； Weissella confusa

參考文獻

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Anonymous (2001) European Antimicrobial Resistance Surveillance System (EARSS) Annual Report. National Institute of Public Health and the Environment, Bilthoven.

Antonio, M.A., Hawes, S.E., and Hillier, S.L. (1999) The identification of vaginal Lactobacillus species and the demographic and microbiologic characteristics of women colonized by these species. J Infect Dis 180: 1950-1956.

Arques, J.L., Fernandez, J., Gaya, P., Nunez, M., Rodriguez, E., and Medina, M. (2004) Antimicrobial activity of reuterin in combination with nisin against food-borne pathogens. Int J Food Microbiol 95: 225-229.

Aslim, B., and Kilic, E. (2006) Some probiotic properties of vaginal lactobacilli isolated from healthy women. Jpn J Infect Dis 59: 249-253.

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乳酸菌所產生抗菌物質抑制多重抗藥性金黃色葡萄球菌能力之分析

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