Wza gene knockout decreases Acinetobacter baumannii virulence and affects Wzy-dependent capsular polysaccharide synthesis

Abstract

To investigate the virulence of capsular polysaccharide export protein (Wza) in carbapenemresistant Acinetobacter baumannii and its effect on capsule formation. wza gene knockout and complementation strains were constructed, and changes in bacterial virulence were observed using in vitro adhesion, antiserum complement killing, anti-oxidation experiments, and infections in Galleria mellonella and mice. The effect of wza knockout on the genes wzb and wzc and wzi were assessed by RT-PCR. We successfully constructed wza knockout and complementation strains. Compared with wildtype (WT) strains, wza knockout strains displayed lower adhesion to A549 cells (p = 0.044), lower antiserum complement killing ability (p = 0.001), and lower mortality of G. mellonella (p = 0.010) and mice (p = 0.033). Expression levels of wzb, wzc and wzi were decreased in wza knockout strains. The antioxidant capacity of Wza knockout bacteria was only slightly decreased. Complementation of the wza gene returned the adhesion ability, antiserum complement killing ability, and mortality of G. mellonella and mice to WT levels. Expression of wzb, wzc and wzi was also returned to WT levels following wza complementation. The results clearly demonstrate that Wza is toxic. Wza affects the expression of other proteins of the Wzy capsule polysaccharide synthesis pathway, which affects the assembly, export, and extracellular fixation of capsular polysaccharide, resulting in synergistic effects that decrease bacterial virulence.

Mechanisms Protecting Acinetobacter baumannii against Multiple Stresses Triggered by the Host Immune Response, Antibiotics and Outside-Host Environment

“…The CPS of different isolates are linear or branched and may possess non-carbohydrate substituents, including the most frequent O - and N -acyl groups [ 44 ]. The Wzy pathway encoded by the K locus is responsible for the export and extracellular assembly of CPS [ 45 ]. The K locus is regulated by the two-component signal transduction system BfmRS.…”

“…Several studies demonstrated that the production of capsules affects A. baumannii virulence and persistence in the host [ 50 , 51 ]. Capsule-enriched strains cause more severe disease or higher lethality [ 35 , 45 , 50 , 52 , 53 ]. These observations may be partly explained by the high hydrophilicity of CPS and negative charges of CPS monosaccharides that prevent phagocytosis by hindering interactions with the negatively charged surfaces of neutrophils and macrophages, preventing phagocytosis [ 35 ].…”

“…These observations may be partly explained by the high hydrophilicity of CPS and negative charges of CPS monosaccharides that prevent phagocytosis by hindering interactions with the negatively charged surfaces of neutrophils and macrophages, preventing phagocytosis [ 35 ]. However, it should be noted that the highly variable structure of the CPS may affect its capacity as a protective barrier against the immune response and other stresses, including complement-mediated killing, lysozyme degradation, and ROS [ 45 , 50 , 51 , 52 , 53 , 54 ].…”

The Toxin-Antitoxin Systems of the Opportunistic Pathogen Stenotrophomonas maltophilia of Environmental and Clinical Origin

“…The ability to form capsule is considered as an important virulence trait in bacteria, together with resistance to antibiotics, biofilm formation, adhesiveness, desiccation, and other features [ 35 , 36 , 37 ]. The role of capsular exopolysaccharides in S. maltophilia virulence is still unknown.…”

“…The ability to produce exopolysaccharide capsule is indicative of a strain’s ability to avoid killing by a host’s immune system [ 36 , 38 ]. We aimed to test if isolates differed in their ability to survive serum-mediated killing.…”