Extraction, identification and relative quantification of outer membrane vesicles of Klebsiella pneumoniae / 中华检验医学杂志

ABSTRACT

Objective:Based on the modified ultracentrifugation method, the outer membrane vesicles (OMV) secreted by Klebsiella pneumoniae were rapidly separated, identified and quantified. Methods:Standard strains of classic Klebsiella Pneumoniae (cKP) purchased from the Clinical Laboratory Center of the National Health Commission, and hypervirulent Klebsiella pneumoniae (hvKP) which was donated by Taiwan University were cultured in M9 basal media for 9 hours, and the OMV were extracted by modified ultracentrifugation. The shape and size of OMV were identified by transmission electron microscopy (TEM), relative quantification by Stewart phospholipids analysis method. Two groups were compared using independent samples t test. Results:It was observed under the TEM that most of the OMV secreted by cKP and hvKP showed spherical vesicle structure and a small part were irregular. The diameter of OMV ranged from 20 to 250 nm, multiple vesicles could be seen in clusters. Relative quantification found that the number of OMV secreted by hvKP were more than cKP ( P<0.05). Conclusions:This study successfully achieved the extraction, identification and quantification of OMV from Klebsiella pneumoniae through the modified ultracentrifugation method, which provided a foundation for further study about the function and mechanism of OMV, and also provided new ideas for the treatment of bacteria. Based on the ultracentrifugation method, the OMV secreted by Klebsiella pneumoniae were rapidly separated and extracted, then identified and quantified.