【作者】 魏忠彬；

【导师】 钱爱东；

【作者基本信息】 吉林农业大学 ， 预防兽医学， 2007， 硕士

【摘要】 乳酸菌为益生菌，主要应用于近年来出现的新型绿色饲料添加剂，它具有无毒副作用，无残留，无抗药性，不污染环境等特点。其主要作用是在数量或种类上补充肠道内减少或缺乏的正常微生物，恢复肠道内的微生态平衡，以达到保健的目的。细菌活的非可培养状态(viable but nonculturable state，VBNC)指细菌处于不良环境条件下的一种特殊存活形式，用常规方法培养不能生长繁殖，但仍然是活的，具有代谢活性的一种特殊的细菌存活形式，并且这种状态的细菌在适当的条件下可以恢复为可培养状态。本试验旨在研究乳酸杆菌的VBNC状态，将其引入到益生素的研究之中，为研制新型实用的防病治病的益生菌活菌制剂开辟一条新径。试验对沈阳辉山乳液有限责任公司提供的乳酸杆菌，实验室分离的鸡源乳酸杆菌和猪源乳酸杆菌进行了活的非可培养状态的初步研究，并且结合多种检测方法，摸索出了乳酸杆菌进入非可培养状态的稳定条件，也探索到了非可培养状态乳酸杆菌的复苏条件。将乳酸杆菌悬浮于液体MRS，PH2-3中，置于4℃的低温下厌氧静止培养，三株乳酸杆菌均可进入活的非可培养状态。虽然同是乳酸杆菌，但三株菌进入VBNC状态的时间不同，从最短的一株R2的35天到最长的一株R3的46天。将活的非可培养状态的乳酸杆菌的菌悬液中加入7％的Tween80，在37℃的全温振荡培养箱中震荡培养，200rpm，可使三株非可培养状态的乳酸杆菌都恢复到可培养状态。在复苏过程中加入的Tween80还是一种常见的饲料添加剂，它可以大大增加微生物细菌总数和瘤胃细菌产量，并还可使奶牛的产奶量大大增加，减少仔猪下痢，改善发育效果，提高肉鸡增重率，从而使饲料添加剂的性能得以更大的发挥。益生特性对比试验发现，正常状态的乳酸杆菌R2不但对酸有较强的耐受性，而且还对0.2％的胆盐和1％的胃蛋白酶有耐受性。相比之下，VBNC乳酸杆菌R2对强酸、1％胃蛋白酶都具有较强耐受性，但对胆盐则不耐受。更多还原

【Abstract】 The lactobacillus lactis is probiotics, mainly be applied in the new green feed additive which appearedin the recent years. It has the characteristic of the non-toxic side effect, non-residual, non-drug resistance,non-pollute-environment and so on. Its main function is supplementing microorganism in the quantityeither the type which the intestinal tract reduced ,to be restore microbiology balance of the intestinaltract,achieve the health care the goal. "The viable but nonculturable state"is a kind of special survival statewhen the bacterium is under the poor surroundings condition.The bacterium could not grow by normalcultured, but it is still viable and possesses metabolic activity.On the adequate condition,the bacteriumcould recover to culturable state. "VBNC" is short for it. The purpose of the experiment is researchingviable but non-culturable state lactobacillus lactis, take it into the research area of probiotics, find a kind ofmethod in producing new feed additive which prevents and treats illness.lactobacillus lactis R2 was offered by ShenYang Huishan Milk Co.Ltd, lactobacillus lactis R1,R3were separated from chicken and pig.Through researching lactobacillus lactis R1,R2,R3 and uniting themany kinds of detected way, in the end, the stability condition that the lactobacillus lactis was inducedinto VBNC was finded out, also the condition that viable but non-culturable state was resuscitation inlactobacillus has been explored. lactobacillus lactis was added into liquid MRS ,PH2-3,suspension, viable but nonculturable state wasinduced in lactobacillus lactis by 4℃low temperature. The time that lactobacillus lactis entered intoviable but non-culturable state was different, the shortest was R2,35 days, but the longest was R3,46 days.7% tween80 was added in viable but nonculturable state lactobacillus lactis suspension, which wasshocked at 37℃,200rpm, viable but nonculturable state lactobacillus lactis could become culturablestate.Tween80 is also a sort of familiar feed additive,which can add the amount of probiotics.Thecapability of feed additive is exerted more effectively. And normal R2 could resist strong acid, 0.2% bilesalt, and 1% pepsin. Contrastly, VBNC R2 could only resist strong acid and 1% pepsin except for bile salt.更多还原