摘要
为探究亚抑菌浓度(Sub-MIC)强力霉素或庆大霉素对猪链球菌(SS)生物被膜形成能力和毒力因子表达的影响,本研究采用96孔板法测定强力霉素或庆大霉素对SS的最小抑菌浓度(MIC)。利用结晶紫染色和活细胞计数法分别测定亚抑菌浓度(1/2 MIC、1/4 MIC、1/8 MIC和1/16 MIC)的强力霉素或庆大霉素作用下SS生物被膜形成能力、SS生长能力及其被膜内SS活菌数量;采用革兰染色观察亚抑菌浓度强力霉素(1/4 MIC和1/8 MIC)或(1/2 MIC和1/4 MIC)庆大霉素作用下SS分布情况;采用qRT-PCR检测亚抑菌浓度(1/4 MIC和1/8 MIC)作用下的SS毒力因子转录水平。结果显示:亚抑菌浓度强力霉素(1/2 MIC^1/16 MIC)或庆大霉素(1/2 MIC和1/4 MIC)均能够增强SS生物被膜形成能力(p<0.001)和增加被膜内SS活菌数量(p<0.001),且SS具有明显集聚成团现象;不同亚抑菌浓度强力霉素和庆大霉素均减缓了SS的早期生长速率,但随着培养时间延长,其生长特性恢复;1/4 MIC和1/8 MIC的强力霉素或庆大霉素均能够显著降低SS毒力因子cps、ef、sly、fbps、gdh和gapdh mRNA的转录水平(p<0.01)。结果表明:亚抑菌浓度的强力霉素和庆大霉素均可以增强SS生物被膜形成能力并降低其毒力因子的转录水平从而形成持续性感染,本研究为抗生素在治疗或预防相关疾病中的正确应用提供参考依据。
To evaluate the effect of sub-MIC(Minimum inhibitory concentration)of doxycycline and gentamicin on biofilm formation and virulence factor expression of Streptococcus suis(S.suis),a 96-well plate method was used to determine the MIC of doxycycline and gentamicin against S.suis.The capability of S.suis in biofilm production,viable bacterial number in biofilm and growth curve at sub-MIC(1/2 MIC,1/4 MIC,1/8 MIC and 1/16 MIC)of doxycycline and gentamicin were performed using crystal violet assays and viable cell counting.Gram staining was used to observe the distribution of S.suis at sub-MIC of doxycycline(1/4 MIC and 1/8 MIC)or gentamicin(1/2 MIC and 1/4 MIC).The transcription levels of S.suis virulence factors were confirmed by q RT-PCR at sub-MICs(1/4 MIC and 1/8 MIC)of doxycycline and gentamicin.It was found that the biofilm formation increased at1/2-1/16 MIC of doxycycline or 1/2 and 1/4 MIC of gentamicin(p<0.001),the viable bacterial number in biofilm also increased(p<0.001)under these sub-MICs,and the agglomeration of S.suis was obvious.The early growth rates of S.suis were inhibited at sub-MIC of gentamicin and doxycycline,but its growth state recoverd with the cultivation time.Furthermore,real-time PCR results showed that the m RNA level of S.suis virulence factors:cps,ef,sly,fbps,gdh,and gapdh decreased significantly at 1/4 and 1/8 MIC of doxycycline or gentamicin(p<0.01).These results demonstrated that sub-MICs of doxycycline and gentamicin increase biofilm formation of S.suis and decrease transcription levels of S.suis virulence factors to the benefit of its persistent infection,thus providing correct treatments modalities of antibiotics in treatment or prevention of the related diseases.
作者
王嘉怡
刘宝宝
张港琛
李金朋
毛晨龙
樊擎莹
汪洋
WANG Jia-yi;LIU Bao-bao;ZHANG gang-chen;LI Jin-peng;MAO Chen-long;FAN Qing-ying;WANG Yang(College ofAnimal Science and Technology,Henan University of Science and Technology,Key Laboratory ofMolecular Pathogen and Immunology of Animal of Luoyang,Luoyang 471003,China)
出处
《中国预防兽医学报》
CAS
CSCD
北大核心
2020年第5期494-499,共6页
Chinese Journal of Preventive Veterinary Medicine
基金
国家自然科学基金项目(31902309、31772761)
河南省自然科学基金(182300410047、162300410067)
关键词
猪链球菌
亚抑菌浓度
生物被膜
毒力因子
Streptococcus suis
sub-minimum inhibitory concentration
biofilm
virulence factors
