沙门氏菌H鞭毛抗原纯培养和位相变异方法改进及效果的分析

Study on the improvement of a novel pure culture and a novel phase variation method of Salmonella H flagella antigen

目的探讨一种新的纯培养和一种新位相变异方法对沙门氏菌H鞭毛抗原表达的改进效果。方法(一)30株沙门氏菌同时用下述两种方法纯培养后,进行H鞭毛抗原血清学鉴定再比较:⑴依据GB4789.4-2016中方法,采用营养琼脂平板直接培养后,检测H鞭毛抗原;⑵采用改进的沙门氏菌H鞭毛抗原纯培养法(即PCA琼脂平板表面加布氏肉汤1.5ml挑菌涂抹直接培养),36℃24h后检测H鞭毛抗原。(二)22株沙门氏菌同时用下述两种方法做H鞭毛抗原位相变异培养后,进行H鞭毛抗原血清学鉴定再比较:⑴参考GB4789.4-2016中的“简易平板法”添加诱导血清,进行位相变异培养,检测H鞭毛抗原;⑵采用改进的沙门氏菌位相变异方法(即接种沙门氏菌在含有布氏肉汤1小滴和沙门氏菌诊断血清1小滴的液体中)诱导培养36℃24h,再检测H鞭毛抗原。结果(一)两种纯培养方法中,改进的沙门氏菌鞭毛抗原纯培养方法两相H鞭毛抗原都检出的比率高于常规的纯培养方法,经χ^(2)检验显示,P<0.01,差异有统计学意义;(二)两种位相变异方法中,改进的沙门氏菌H鞭毛抗原位相变异试验方法检出的阳性率高于传统的“简易平板”位相变异H鞭毛诱导方法,χ^(2)检验显示,P<0.01,差异有统计学意义。结论改进的沙门氏菌纯培养和位相变异方法鉴定沙门氏菌H鞭毛抗原检出率高、诱导所用时间短、诱导细菌繁殖快,且简单便捷、廉价高效,不受贵重仪器设备的限制。新方法可为基层实验室开展沙门氏菌的血清分型工作提供省时、简单、高效的新途径。

Objective To investigate the effect of a novel pure culture and a new phase variation method on the expression of H flagella antigen in Salmonella.Methods(1) Serological identification of H flagella antigen was performed for comparison after the same 30 Salmonella strains were cultured by the following two methods:(A).According to the method in Chinese National Standard for Food Safety(Food Microbiology,Salmonella Detection,GB4789.4-2016),the H flagella antigen was detected after the Salmonella strains were directly cultured with nutrient agar.(B).The modified pure culture method of Salmonella H flagella antigen.The Salmonella was streaked and inoculated onto the surface of PC A agar plate with 1.5 ml of Brucella broth,and then the H flagella antigen was detected after cultivation at 36℃.for 24 hours.(2) Serological identification of H flagella antigen was performed for comparison after the same 22 Salmonella strains were cultured by the following two different H flagella antigen phase variation methods:(A).According to the "simple plate method in GB4789.4-2016,the H flagella antigen was detected after the Salmonella strains were cultured on semi-solid agar with anti-H phase serum;(B).The modified Salmonella phase variation method.The Salmonella strains were inoculated in a liquid broth containing 1 droplet of brucellosis soup and 1 droplet of Salmonella anti-H phase diagnostic serum,then the H flagella antigen was detected after cultivation at 36℃ for 24 hours.Results(1) After 24 hours of culture,among the two Salmonella pure culture methods,the H flagella antigen detection rate of the improved method was higher than that of the conventional pure culture method(P<0.01).(2) The improved method of pure culture and phase variation was suitable for identifying the H flagella antigen of Salmonella with advantages of high detection rate,short induction time,fast inducing bacteria reproduction,simple,convenient,cheap and efficient,and not limited by expensive equipment and equipment.The new method can provide a new time-saving,simple and efficient way for salmonella serotyping in basic laboratory.Conclusion The improved method of pure culture and phase variation method were suitable to identify the H flagella antigen of Salmonella with advantages of high detection rate,short induction time and fast reproduction of inducing bacteria,as well as simple,convenient,cheap,efficient,and expensive equipment free.Thus,the new method can provide a new time-saving,simple and efficient way for Salmonella serotyping in basic laboratory.