Human fluids alter DNA-acquisition in Acinetobacter baumannii

Abstract

Transformation is one of the mechanisms of acquisition of foreign genetic material leading to the emergence of multidrug resistant (MDR) bacteria. Recently, human serum albumin (HSA) was shown to specifically increase transformation frequency in the nosocomial pathogen Acinetobacter baumannii. To further assess the relevance of HSA as a possible modulator of A. baumannii transformation in host-pathogen interactions, in this work we examined the effect of different human fluids. We observed a significant increase in transformation frequencies in the presence of pleural fluid, whole blood cells and liquid ascites, and to a lesser extent with urine. The observed effects correlate with both HSA and bacterial content found in the assayed patient fluids. Taken together, these results are in agreement with our previous findings that highlight HSA as a possible host signal with the ability to trigger natural transformation in A. baumannii.

Keywords: Acinetobacter baumannii; Antibiotic resistance; DNA-acquisition; Human serum albumin; Pleural fluid; Transformation.

Figure 1.. Effects of pleural fluid on A. baumannii natural transformation and differential expression of competence genes PilQ and comEA.

A) Transformation assays were performed with LB and LB plus PF 4%. Cultures were transformed with either plasmid (black) or genomic (grey) DNA and plated onto LB agar plates supplemented with 10ug/ml of kanamycin. Experimental controls are presented as solely LB broth. Data is presented as means and error bars represent standard deviation. Asterisks represent statistical significance with a p- value <0.05 (Mann Whitney t test (n=3 to 13). B) Real-time Quantitative PCR was performed to identify the variance in expression levels of A. baumannii strains A118 and A42 competence genes PilQ and comEA in exposure to PF 4%. Fold changes were calculated using the double △Ct analysis. Data are presented as the mean and the errors bars represent standard deviation. At least three independent samples were used, and four technical replicates were performed from each sample. Asterisks represent statistical significance (P-value <0.05) as determined by the Mann Whitney t-test (n= 3 to 4).

Figure 2.. Natural transformation frequencies with host human fluids.

Transformation assays of A. baumannii strain A) A118 and B) A42 were performed with LB broth, LB plus HWB 0.2%, LB plus AF 4%, LB plus UR 4%, and LB plus NF 0.2%. Experimental controls were transformed solely with LB broth. Cultures were transformed with plasmid DNA (black) or genomic DNA (grey) and plated on LB agar supplemented with 10ug/mL of kanamycin. CFUs were plated on LB agar. The data is presented as the mean and error bars represent standard deviation. At least three independent replicates were conducted and asterisks above represent statistical significance with a P-value <0.05 (Mann Whitney t test (n=3 to 13).

Figure 3.

Schematic representation of albumin effect on natural transformation.