Development of safe food using lactic acid bacteria

利用乳酸菌开发安全食品

• 批准号: 14560221
• 负责人: TOBA Takahiro
• 金额: $ 1.98万
• 依托单位: Hirosaki University
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (C)
• 财政年份: 2002
• 资助国家: 日本
• 起止时间: 2002 至 2003
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-14560221/
• 关键词: Lactobacillus crispatus; hydrogen peroxide; bifidobacteria; PCR; Lactobacillus gasseri; S-layer protein

Adhesion assay was performed using Lactobacillus and Bifidobacterium strains isolated from chicken and human faeces to select the strains which strongly adhered to chicken crop epithelium, human intestinal Caco-2 cells and HT-29 cells, and immobilized extracellular matrix proteins. For rapid identification of Lactobacillus crispatus we newly developed PCR primers targeting S-layer protein gene(Horie et al.2002). Adhesive lactobaciller and bifidobacterial strains inhibited the adhesion of pathogens such as Salmonella to Caco-2 cells Adhesion of L.crispatus and L.gasseri strains tested were induced on plate cultures under anaerobic conditions. A 46.5kDa S-layer protein(named as LbsA) was identified as an adhesin of highly adhesive L.crispatus MH 3 1 5 strain. The DNA region coding for LbsA was cloned and sequenced. Alignment comparisons of the predicted amino acid sequence with S-layer proteins of other Lactobacillus strains suggested the presence of laminin-binding site in LbsA (Mukai et al.2003). An 58.7kDa surface protein was identified an putative adhesin of L.gasseri MH 411 by Western bolting assay.We also selected hydrogen peroxide(H_2O_2)-producing Lactococcus lactis subsp.lactis and L.plantarum strains from foods. They produced >300ppm H_2O_2 under the condition tested. One of these strains, L.lactis subsp.lactis AI62 inhibited pathogens such as Listeria ivanovii, Yersinia enterocolitica and Aeromonas hydrophila efficiently(Ito et al.2003).

利用从鸡和人粪便中分离的乳杆菌和双歧杆菌菌株进行粘附试验，以选择对鸡作物上皮、人肠Caco-2细胞和HT-29细胞以及固定化细胞外基质蛋白具有强烈粘附的菌株。为了快速鉴定卷曲乳杆菌，我们新开发了靶向S层蛋白基因的PCR引物（Horie等，2002）。粘附性乳杆菌和双歧杆菌菌株抑制病原体如沙门氏菌对Caco-2细胞的粘附。在厌氧条件下在平板培养物上诱导所测试的卷曲乳杆菌和加氏乳杆菌菌株的粘附。结果表明，卷曲乳杆菌MH 315菌株的S层蛋白（LbsA）分子量为46.5kDa，是一种粘附素。对编码LbsA的DNA区域进行克隆和测序。预测的氨基酸序列与其他乳杆菌属菌株的S层蛋白的比对比较表明LbsA中存在层粘连蛋白结合位点（Mukai等人，2003）。通过Western筛选，确定了加氏乳球菌MH 411的表面蛋白为58.7kDa的粘附素，并从食品中筛选了产过氧化氢的乳酸乳球菌乳酸亚种和植物乳球菌。在试验条件下，它们产生的H_2O_2> 300 ppm。其中一种菌株，乳酸乳球菌乳酸亚种AI 62，可有效抑制伊氏李斯特菌、小肠结肠炎耶尔森氏菌和嗜水气单胞菌等病原体（Ito等人，2003年）。

项目成果

Horie M, Kajikawa HS, Toba T: "Identification of Lactobacillus crispatus by polymerase chain reaction targeting S-layer protein gene."Letters in Applied Microbiology. 35. 57-61 (2002)

Horie M、Kajikawa HS、Toba T：“通过针对 S 层蛋白基因的聚合酶链反应鉴定卷曲乳杆菌。”应用微生物学快报。

Ito A, Sato Y, Kudo S, Sato S, Nakajima H, Toba T: "The screening of hydrogen peroxide-producing lactic acid bacteria and their application to inactivating psychrotrophic food-borne pathogens"Current Microbiology. 46. (2003)

Ito A、Sato Y、Kudo S、Sato S、Nakajima H、Toba T：“产生过氧化氢的乳酸菌的筛选及其在灭活耐冷食源性病原体中的应用”当前微生物学。

Ito A, Sato Y, Kudo S, Sato S, Nakajima H, Toba T: "The screening of hydrogen peroxide-producing lactic acid bacteria and their application to inactivating psychrotrophic food-borne pathogens."Current Microbiology. 47. 231-236 (2003)

Ito A、Sato Y、Kudo S、Sato S、Nakajima H、Toba T：“产生过氧化氢的乳酸菌的筛选及其在灭活耐冷食源性病原体中的应用。”当前微生物学。

向井孝夫, 戸羽隆宏, 大堀均: "乳酸菌における細胞接着の分子機構"ミルクサイエンス. 52. 167-174 (2003)

Takao Mukai、Takahiro Toba、Hitoshi Ohori：“乳酸菌细胞粘附的分子机制”《牛奶科学》52. 167-174 (2003)。

Horie M, Kajikawa HS, Toba T: "Identification of Lactobacillus crispatus by polymerase chain reaction targeting S-layer protein gene."Letters in Applied Microbiology. 38. 57-61 (2002)

Horie M、Kajikawa HS、Toba T：“通过针对 S 层蛋白基因的聚合酶链反应鉴定卷曲乳杆菌。”应用微生物学快报。