, 2006). A LuxR-like domain is present in the pPAA3-0024 protein. Proteins that contain Lux-R

domains are involved in response regulation and can act as transcriptional activators or repressors. The plasmid also contains two proteins, which are mobB-like and mobC-like, which have been implicated in conjugative plasmid mobilization (Zhang & Meyer, 1997). The gene designated pPAA3_001 shows homology to mpr, a zinc metalloproteinase with an SprT domain that is predicted to play a role in transcription elongation. The pPAA3 plasmid is displayed in Fig. 4, with annotated genes coloured according to putative function. Plasmid pPAA3 encodes a 10-gene cluster highly homologous to the Type IV secretion system genes encoded on the cryptic Yersinia plasmid pCRY (see Table 2). We speculate that these genes could play a role in intracellular invasion by the Australian P. asymbiotica PCI-32765 mouse isolates. Nevertheless, we cannot rule out the possibility that these pPAA2 genes encode a Type IV DNA conjugation system for horizontal plasmid transfer. Pathogenic T4SS are used by many pathogens to infect eukaryotic cells and have been implicated in the transport of essential

virulence factors that establish bacterial infection in the eukaryotic host (de Paz et al., 2005). Most T4SS are formed by 11 proteins, named virB1 to virB11. The overall architecture of the transporter is conserved in the family and there is evidence to suggest that a ‘core’

through complex made up of proteins virB7, virB8, virB9 and virB10 is responsible for the central transmembrane channel. These proteins Idelalisib mw are located mostly in the periplasm. The T4SS of the pPAA3 and pCRY plasmids lack the virB7 gene. The pCRY plasmid also lacks a virB3 gene, whereas in pPAA3, it is fused with the virB4 gene. While virB7 was required in Agrobacterium tumefaciens for the formation of a functional secretion apparatus, den Hartigh et al. (2008) showed that the deletion of virB7 from the Brucella abortus chromosome did not reduce the ability of the Gram-negative bacterium to persist in the spleens of mice, suggesting that virB7 was not an essential component of the secretion system. The T4SS in pPAA3 is also lacking a virD4 component, which is a substrate recognition receptor known as the Type IV secretion coupling protein. It was thought that the virD4 protein was important for the correct functioning of the secretion apparatus; however, a T4SS in Bartonella species, which lacks this virD4 component, still allows the bacterium to invade erythrocytes (Dehio, 2008). The acquisition of the pPAA3 plasmid in the Kingscliff isolate may account for the increased virulence of the Australian isolate both against tissue culture cells and infected patients. Fig. S1. Three different workflows were followed, that combined different types of sequence data with different assembly algorithms, to look for the optimal de novo assembly. Fig. S2.