|
|
Background: Acinetobacter baumannii (Ab) is an important opportunistic pathogen in a hospital setting. The virulence varies among different clinical isolates, which can be measured by cytotoxicity assay or Galleria mellonella killing assay. However, these measurements are not suitable for routine clinical laboratory application. Thus, a putative marker for early prediction and intervention of these virulent isolates is desirable.
Methods: PgaB is part of the pgaABCD cluster, which is considered to be essential for the biofilm formation and pathogenesis of Ab. Sixty-five non-repetitive Ab clinical isolates were randomly collected from respiratory tract specimens in our hospital, which were pgaB positive as determined by PCR. The mRNA expression level of pgaB was measured by quantitative real-time PCR, with Taqman probe targeting the conserved region of pgaB. 16S rRNA was used as an internal control. The virulence was determined by the ability to kill A549 human lung epithelial cells with different MOIs and different time points. The virulence was also determined by Galleria mellonella killing assay and analyzed by Kaplan-Meier survival analysis. The statistical analysis was performed with SPSS 13.0 and Graphpad Prism 5.0.
Results: Sixty-five isolates were divided into pgaB high and low expression groups by the values of adjusted pgaB expression, the median values of which were 25.81 ± 6.34 (n = 41) and 4.16 ± 2.82 (n = 24), respectively (p < 0.05). The Ab virulence determined by cytotoxicity assay was significantly higher in the pgaB high expression group than in the low expression group (MOI = 100, incubated for 18 hours, p < 0.05). The killing rate of Galleria mellonella was significantly higher in the pgaB high expression group than in the low expression group (inoculum of 106 CFU/larva at 37°C, p < 0.05), with the median survival time of 40 hours and 75 hours, respectively.
Conclusions: The adjusted expression level of pgaB was in agreement with the virulence of Ab in cytotoxicity assays and Galleria mellonella killing assay, thus it may be used potentially as a putative marker for early prediction of the virulence of Ab clinical isolates.
DOI: 10.7754/Clin.Lab.2018.180916
|
