Abstract
Pseudomonas savastanoi pv. savastanoi NCPPB 3335 is a model for the study of the molecular basis of disease production and tumor formation in woody hosts, and its draft genome sequence has been recently obtained. Here we closed the sequence of the plasmid complement of this strain, composed of three circular molecules of 78,357 nt (pPsv48A), 45,220 nt (pPsv48B), and 42,103 nt (pPsv48C), all belonging to the pPT23A-like family of plasmids widely distributed in the P. syringae complex. A total of 152 coding sequences were predicted in the plasmid complement, of which 38 are hypothetical proteins and seven correspond to putative virulence genes. Plasmid pPsv48A contains an incomplete Type IVB secretion system, the type III secretion system (T3SS) effector gene hopAF1, gene ptz, involved in cytokinin biosynthesis, and three copies of a gene highly conserved in plant-associated proteobacteria, which is preceded by a hrp box motif. A complete Type IVA secretion system, a well conserved origin of transfer (oriT), and a homolog of the T3SS effector gene hopAO1 are present in pPsv48B, while pPsv48C contains a gene with significant homology to isopentenyl-diphosphate delta-isomerase, type 1. Several potential mobile elements were found on the three plasmids, including three types of MITE, a derivative of IS801, and a new transposon effector, ISPsy30. Although the replication regions of these three plasmids are phylogenetically closely related, their structure is diverse, suggesting that the plasmid architecture results from an active exchange of sequences. Artificial inoculations of olive plants with mutants cured of plasmids pPsv48A and pPsv48B showed that pPsv48A is necessary for full virulence and for the development of mature xylem vessels within the knots; we were unable to obtain mutants cured of pPsv48C, which contains five putative toxin-antitoxin genes.
Highlights
The gamma proteobacterium Pseudomonas savastanoi pv. savastanoi causes olive (Olea europaea L.) knot disease, one of the most economically relevant diseases of the olive crop [1]
Native plasmids from strains of the P. syringae group generally share a large amount of repeated sequences [17,21,24], and our initial analyses showed that this was the case with the plasmids from strain NCPPB 3335
We previously identified two native plasmids, pPsv48A (73 kb) and pPsv48B (42 kb), in strain NCPPB 3335 [21]; after mutagenesis, we were able to visualize a new plasmid comigrating with pPsv48B, designated pPsv48C, which had a lower copy number and that was only evident in mutants with a transposon insertion in either plasmid B or plasmid C (Figure 1 and not shown)
Summary
The gamma proteobacterium Pseudomonas savastanoi pv. savastanoi causes olive (Olea europaea L.) knot disease, one of the most economically relevant diseases of the olive crop [1]. Savastanoi is part of the P. syringae complex, which includes at least 10 Pseudomonas species and 60 pathovars of P. syringae, most of which are pathogenic to plants, and whose taxonomy is confusing and still unresolved [2,3,4]. DNA-DNA hybridization studies indicate that the P. syringae complex could be split in nine different genomospecies [2]. Savastanoi has been assigned to the species P. amygdali (genomospecies 2) together with 16 other P. syringae pathovars, including P. syringae pv. Savastanoi infects woody hosts, but it is significant in that it is one of a few closely-related pathovars that cause aerial tumors in their plant hosts. The disease is considered to reduce both olive yield and productivity [10,11], and few commercial cultivars are significantly tolerant to olive knot disease [12]
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