亚抑菌浓度茅苍术挥发油对金黄色葡萄球菌毒力因子表达的抑制作用初步研究

Preliminary study on the inhibitory effects of volatile oils from the dried rhizome of Atractylodes lancea or Atractylodes chinensis on virulence factors of Staphylococcus aureus

目的探索亚抑菌浓度茅苍术挥发油对金黄色葡萄球菌(Staphylococcus aureus)毒力因子如α溶血素等表达的抑制作用及其机制。方法采用微量肉汤稀释法测定茅苍术挥发油对4株金黄色葡萄球菌的最小抑菌浓度(minimum inhibitory concentration,MIC);吸光度测量等方法体外检测亚抑菌浓度茅苍术挥发油作用下金黄色葡萄球菌毒力因子产量的变化;采用Real-Time PCR法检测亚抑菌浓度挥发油对AgrA和Hla mRNA表达的影响。结果茅苍术挥发油可抑制金黄色葡萄球菌生长并呈现浓度依赖性,标准菌株ATCC25923的MIC为0.0625%(v/v),临床菌株的MIC为0.03125%(v/v)。亚抑菌浓度茅苍术挥发油作用后4株菌株凝固酶的分泌受到抑制,标准菌株ATCC25923的凝固酶效价只有对照组的1/4,差异有统计学意义(P<0.05),临床菌株的凝固酶效价都只有对照组的1/8,差异有统计学意义(P<0.05)。亚抑菌浓度茅苍术挥发油减弱了四株菌株的粘附能力,ATCC25923、SA1.5、SA2.3和SA4.12的粘附能力分别只有对照组的(53.71±6.56)%(P<0.01)、(53.10±9.39)%(P<0.01)、(74.70±9.00)%(P<0.01)和(45.62±21.22)%(P<0.01)。标准菌株ATCC25923和SA2.3的溶血活力受到亚抑菌浓度挥发油的显著抑制,溶血活力分别只有对照组的(18.48±23.88)%(P<0.01)和(1.63±4.18)%(P<0.01)。α-溶血素毒力基因Hla及其调控基因AgrA的表达受到了亚抑菌浓度挥发油的抑制,其表达水平分别只有对照组的(6.17±2.23)%(P<0.01)和(24.79±9.19)%(P<0.01)。结论茅苍术挥发油能抑制金黄色葡萄球菌的生长,并且对多种毒力因子的表达有抑制作用。

Objective The purpose of this study was to investigate the inhibition of virulence factors of Staphylococcus aureus by volatile oils from the dried rhizome of Atractylodes lancea or Atractylodes chinensis and the mechanism by which volatile oils inhibit the a-toxin of S. aareus. Methods Broth dilution was used to measure the minimum inhibitory concentration （MIC） of volatile oils from A. lancea or A. chinensis on S. aureus. Absorbance was measured to detect changes in the production of the virulence factors when S. aureus was exposed to the volatile oils from A. lancea or A. chinensis in vitro. Real-time PCR was used to detect the effect of volatile oils on the level of AgrA and Hla mRNA. Result Results indicated that volatile oils from A. lancea or A. chinensis inhibited the growth of S. aureus in a dose-dependent manner. Volatile oils from A. lancea or A. chinensis had a MIC of 0. 062 5% （v/v） with respect to a standard strain of S. aureus （ATCC25923）, and those volatile oils had a MIC of 0. 03125 % （v/v） with respect to three clinical isolates of S. aureus . The sub-MIC of volatile oils from A. lancea or A. chinensis reduced the production of coagulase by the four S. aureas strains. The coagulase titer of ATCC25923 was only 1/4 the titer of the control （P〈0.05） while the coagulase titers of clinical isolates were only 1/8 the titer of the control （P〈0.05）. The sub-MIC of volatile oils from A. lancea or A. chinensis also inhibited the adhesive ability of ATCC25923 to 53.71 ±6.56% corn pared to the control group （P〈0.01）, it inhibited the adhesive ability of SA1.5 to 53.10±9.39% （P〈0.01）, it inhibited the adhesive ability of SA2.3 to 74.70±9.00% （P〈0.01）, and it inhibited the adhesive ability of SA4.12, to 45.62_=21.22% （p 〈0.01）. The sub-MIC of volatile oils from A. lancea or A. chinensis markedly inhibited the hemolytic activity of the standard strain （ATCC25923） to 18.48±23.88% （P〈0.01） and the hemolytic activity of SA 2.3 to 1.63±4.18）G （P〈0.01） compared to the control group. The sub-MIC of volatile oils from A. lancea or A. chinensis inhibited the expression of the virulence gene Hla, which encodes a-toxin, to 6.17±2.23% （P〈0.01） compared to the control group and it inhibited the expression of the gene that regulates Hla, AgrA, to 24.79±9.19% （P〈0.01）. Conclusion Volatile oils from A. lancea or A. chinensis inhibit the growth of S. aureus and can effectively inhibit the expression of several virulence factors.