应用NaOH制备胸膜肺炎放线杆菌菌影方法的建立

Establishment of a novel preparation method for Actinobacillus pleuropneumoniae ghost by sodium hydroxide

将培养12、24、48和72 h的胸膜肺炎放线杆菌血清1型shope菌株的菌液浓度分别调至1×10~8和1×10~9CFU/m L,测定氢氧化钠(NaOH)对其最小抑菌浓度;将不同培养时间、2种浓度的菌液与对应的最小抑菌浓度NaOH混合,37℃孵育75 min;间隔取样,对细菌形态学、裂解情况进行观察,确定制备菌影的条件。结果显示,NaOH对各培养时长、2种浓度的胸膜肺炎放线杆菌血清1型菌液均有裂解效果,37℃作用75 min,均未能检测到活菌,裂解率可达到100%。通过扫描电镜观察发现,细菌菌体形态完好、外膜孔道明显。结果表明,用NaOH成功地制备了胸膜肺炎放线杆菌菌影,这种新型制备菌影的方法安全、高效,为日后该菌菌影疫苗的生产与应用奠定了基础。

To determine the minimum inhibitory concentratin（MIC）of sodium hydroxide（NaOH） against Actinobacillus pleuropneumoniae（APP）serotype 1 strain shope,bacteria were cultured for 12 h,24 h,48 h and 72 h,respectively,and each incubation period of bacteria concentration adjusted to 1×108CFU/m L and 1×109CFU/m L.Bacterial suspension mixed with NaOH incubated at 37℃.The optimal conditions for prepration were determined by studying the morphology,cell lysis rate and security.The results showed that NaOH had effects on all bacterial cultures of strain shope of different culture time and concentration.The cell lysis rate could reach 100% and no living bacteria survived when the APP serotype 1 strain shope lysis were induced for 75 min at 37 ℃.The bacteria ghosts were in good shape and the external mode was visualized obviously by scanning electron microscopy.In conclusion,nonliving APP bacterial ghosts were successfully generated by MIC of NaOH.This new preparation method for APP serotype 1 strain shope ghost is safe and effective.It provides the foundations for studying ghost vaccines of APP.