Carboxy-Terminal Processing Protease Controls Production of Outer Membrane Vesicles and Biofilm in <i>Acinetobacter baumannii</i>

Carboxy-terminal processing protease (Ctp) is a serine protease that controls multiple cellular processes through posttranslational modification of proteins. Acinetobacter baumannii ATCC 17978 ctp mutant, namely MR14, is known to cause cell wall defects and autolysis. The objective of this study was to investigate the role of ctp mutation–driven autolysis in regulating biofilms in A. baumannii and to evaluate the vesiculation caused by cell wall defects. We found that in A. baumannii, Ctp is localized in the cytoplasmic membrane, and loss of Ctp function enhances the biofilm-forming ability of A. baumannii. Quantification of the matrix components revealed that extracellular DNA (eDNA) and proteins were the chief constituents of MR14 biofilm, and the transmission electron microscopy further indicated the presence of numerous dead cells compared with ATCC 17978. The large number of MR14 dead cells is potentially the result of compromised outer membrane integrity, as demonstrated by its high sensitivity to sodium dodecyl sulfate (SDS) and ethylenediaminetetraacetic acid (EDTA). MR14 also exhibited the hypervesiculation phenotype, producing outer-membrane vesicles (OMVs) of large mean size. The MR14 OMVs were more cytotoxic toward A549 cells than ATCC 17978 OMVs. Our overall results indicate that A. baumanniictp negatively controls pathogenic traits through autolysis and OMV biogenesis