摘要
研究拟确定亚抑制浓度牛至精油对金黄色葡萄球菌生物被膜形成的影响,为牛至精油在细菌生物被膜群落中的作用及在牦牛乳房炎治疗方面的替抗应用提供理论依据。采用微量结晶紫染色法筛选出具有强成膜能力的金黄色葡萄球菌株,并通过PCR测定生物被膜形成关键基因携带情况;利用微量二倍稀释法测定牛至精油最小抑菌浓度;分别以最小抑菌浓度的1/2、1/4、1/8、1/16、1/32作用于筛选出的菌株8 h、24 h、48 h、72 h后,通过微量结晶紫染色法测定其生物被膜形成总量。结果表明:①42株牦牛源金黄色葡萄球菌均能够形成生物被膜,其中5株(No.35、No.30、No.26、No.2和No.42)成膜能力强(+++);②5株均携带sar和sigB,3株携带cidA,1株同时携带icaA和cidA,1株携带icaA和icaD;③牛至精油对5株金黄色葡萄球菌的MIC为0.5%;④No.35菌株成膜能力最强,其生物被膜形成始终受到亚抑制浓度牛至精油的抑制,该作用随浓度下降和时间延长而减弱。No.30、No.26、No.2菌株成膜能力次之,1/2和1/4 MIC牛至精油对其生物被膜在形成阶段(8 h和24 h)有抑制作用,1/16和1/32 MIC始终表现为促进作用。No.42菌株成膜能力最低,牛至精油对生物被膜形成始终有促进作用,且促进作用随浓度下降和时间延长而增强。因此,金黄色葡萄球菌以eDNA释放和/或PIA合成途径生成生物被膜时,亚抑制浓度牛至精油对其有抑制作用;但形成成熟生物被膜(48~72 h)后,极低浓度的牛至精油主要表现为促进作用。
The effect of sub-minimal inhibitory concentration(sub-MIC)oregano essential oil on biofilm formation of S.aureus was studied to provide the theoretical basis for the role of the oil in bacterial biofilm and the application as a substitute for antibiotics in the treatment of yak mastitis.Trace crystal violet staining was used to evaluate the ability of S.aureus to form biofilms and to select those isolates with the strongest ability,and the key genes for biofilm formation of those isolates were detected by PCR;MIC of oregano essential oil on those selected isolates was determined by doubling dilution;after treatments with 1/2,1/4,1/8,1/16,and 1/32 MIC for 8,24,48 and 72 h,respectively,the total amount of biofilm of selected isolates was determined also by trace crystal violet staining.The results showed as follows:①42 isolates from yak could form the biofilm,among which 5 isolates had the strongest ability(+++);②all 5 isolates carried sar and sigB;3 isolate carried cidA;1 isolate carried icaA and cidA;1 isolate carried icaA and icaD;③the MIC of oregano essential oil against 5 isolates was 0.5%;④although it had the strongest biofilm-forming ability,the total amount of biofilm of isolate No.35 was lower than that of the control group in all testing groups with extremely significant difference(P<0.01),which mean its biofilm-forming ability was inhibited by oregano essential oil at sub-MIC,and the effect decreased as the declining concentration and increasing time.Isolate No.30,No.26,and No.2 had the second-highest biofilm-forming ability,which was inhibited by the oil at the formation stage(8 h and 24 h)at 1/2 and 1/4 MIC,but promoted at 1/16 and 1/32 MIC in all testing groups with different differences compared with the control group.The forming ability of isolate No.42 was lower than the other 4,which was always promoted with the decreasing concentration of oregano essential oil and the increasing time;there were extremely significant differences between all test groups and the control group(P<0.01),respectively,except for the 1/2 MIC group at 8 h.In conclusion,when S.aureus formed biofilms by eDNA release and/or PIA synthesis pathway,oregano essential oil at sub-MIC could inhibit it.However,after the formation of mature biofilm(24-48 h),oregano essential oil at 1/32 MIC mainly showed promoting effect.
作者
王斐
陈平
何振富
谢建鹏
李红英
丁泽
WANG Fei;CHEN Ping;HE Zhenfu;XIE Jianpeng;LI Hongying;DING Ze(Animal Husbandry-Pasture and Green Agriculture Institute,Gansu Academy of Agricultural Sciences,Lanzhou 730070,China;Agricultural and Rural Comprehensive Service Center of Zhongxing Town,Minqin 733300,China;Animal Husbandry and Veterinary Station of Jiudun Town,Wuwei 733000,China)
出处
《中国草食动物科学》
CAS
2022年第5期50-55,79,共7页
China Herbivore Science
基金
甘肃省科技计划项目(20JR10RA463)
甘肃省农业科学院农业科技创新专项项目(2021GAAS45)
甘肃省现代农业科技支撑体系区域创新中心重点科技项目(2020GAAS05)。
