单核细胞增生李斯特菌SigmaB、PrfA因子对生物被膜形成影响的研究

Effect of SigmaB,PrfA on Biofilm Formation in Listeria Monocytogenes

目的探讨单核细胞增生李斯特菌(listeria monocytogenes,LM)sigmaB、prfA基因与生物被膜(biofilm,BF)形成的关系。方法利用微孔板技术,体外建立LM野生菌株EGD、ΔsigmaB、ΔprfA及无害李斯特菌(listeria innocua)生物被膜模型,经1%结晶紫溶液染色,间接反映生物被膜的形成情况,并用倒置显微镜直接观察4种菌株生物被膜的形成情况。结果①利用结晶紫染色与倒置显微镜观察相结合的方法,实现了对上述4种菌株生物被膜形成差异的比较;②结晶紫染色结果显示:在595nm波长下,EGD光吸收值最高,ΔsigmaB与ΔprfA菌株次之,无害李斯特菌最低。统计学分析结果显示,EGD与ΔsigmaB、ΔprfA菌株差异不显著(P>0.05),EGD与无害李斯特菌之间存在显著差异(P<0.01);③倒置显微镜观察结果显示:EGD能形成致密的链网状膜结构,交联度较高;ΔsigmaB、ΔprfA形成的链网状膜结构相对比较疏松,交联度稍低;无害李斯特菌几乎没有成型的链网状膜结构形成。结论实验结果表明LM生物被膜的形成与调控因子sigmaB(σB)、prfA的作用有关,并且LM与无害李斯特菌之间生物被膜的形成差异暗示着生物被膜的形成可能与LM的致病性有关。

Objective To investigate the relationship between SigmaB, PrfA and biofilm formation. Methods Biofilm model of Listeria monocytogenes in vitro was established by microtiter plate, and its biofiim development was indirectly reflected by staining with crystal violet. The stained biofilms were also observed under inverted microscope. Results （1）We achieved the differences among the above - mentioned four kinds in comparison of BF by using the combination of the crystal violet staining and the inverted microscope method. （2）Crystal violet staining results showed that in the 595nm wavelength, EGD had the maximum value of optical absorption, and followed by AsigmaB, AprfA and L. innocua. Statistical analysis showed that EGD, AsigmaB and AprfA strains had no significant difference （P 〉 0.05） , but there were significant differences between EGD and L. innocua （P 〈 0.01 ）. （3）The results obtained by inverted microscope showed that EGD could form a network of knitted chains with high degree of cross - linking. AsigmaB, AprfA also formed a network of knitted chains, but the structure had lower degree of cross - linking. L. innocua barely formed a network of knitted chains structure. Conclusion PrfA and SigmaB were associated with biofilm formation. The formation of biofilms may be related to the pathogenicity.