Identification of Acinetobacter baumannii type VI secretion system effectors and characterisation of a novel effector/immunity pair

The type VI secretion system (T6SS) is a bacterial nanomachine utilised by many Gram-negative bacteria, including Acinetobacter baumannii, to deliver toxic effectors for microbial warfare. These toxic effectors are often delivered via specific non-covalent interactions with cognate VgrG proteins, which form part of the T6SS tip. In A. baumannii, each vgrG gene is usually located in the same locus as two other genes, one encoding the cognate effector and one encoding an immunity protein that protects against self-intoxication. Bioinformatic searches of ninety seven A. baumannii genomes using a highly conserved domain found within the VgrG proteins, enabled the identification of more than 250 genes encoding putative effectors and, in most cases, the gene encoding the corresponding immunity protein. Phylogenetic analysis revealed that the predicted effectors clustered into 33 distinct groups, some of which contained predicted amidases, chitinases, lipases, nucleases and deaminases. Two effectors, Tse5Ab, containing no toxic domains and Tse6Ab, containing a Tox-GHH nuclease domain characteristic of nucleases, were chosen for functional analysis. The C-terminal region encoding the predicted toxic domain of each effector was cloned and expressed in E. coli. Expression of this region of Tse5Ab did not perturb E. coli growth. In contrast, expression of Tse6Ab was toxic but toxicity could be neutralised by the co-expression of the cognate immunity protein. However, Tse6Ab did not exhibit DNase activity and instead may function as an RNase. Further characterisation of the diverse A. baumannii T6SS effectors may lead to the identification of antibacterial molecules with novel activities.