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Variation at the common polysaccharide antigen locus drives lipopolysaccharide diversity within the Pseudomonas syringae species complex.

ABSTRACT: The common polysaccharide antigen (CPA) of the lipopolysaccharide (LPS) from Pseudomonas syringae is highly variable, but the genetic basis for this is poorly understood. We have characterized the CPA locus from P. syringae pv. actinidiae (Psa). This locus has genes for l- and d-rhamnose biosynthesis and an operon coding for ABC transporter subunits, a bifunctional glycosyltransferase and an o-methyltransferase. This operon is predicted to have a role in the transport, elongation and termination of the CPA oligosaccharide and is referred to as the TET operon. Two alleles of the TET operon were present in different biovars (BV) of Psa and lineages of the closely related pathovar P. syringae pv. actinidifoliorum. This allelic variation was reflected in the electrophoretic properties of purified LPS from the different isolates. Gene knockout of the TET operon allele from BV1 and replacement with that from BV3, demonstrated the link between the genetic locus and the biochemical properties of the LPS molecules in Psa. Sequence analysis of the TET operon from a range of P. syringae and P. viridiflava isolates displayed a phylogenetic history incongruent with core gene phylogeny but correlates with previously reported tailocin sensitivity, suggesting a functional relationship between LPS structure and tailocin susceptibility.

SUBMITTER: Jayaraman J

PROVIDER: S-EPMC7820976 | biostudies-literature | 2020 Dec

REPOSITORIES: biostudies-literature

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Variation at the common polysaccharide antigen locus drives lipopolysaccharide diversity within the Pseudomonas syringae species complex.

Jayaraman Jay J Jones William T WT Harvey Dawn D Hemara Lauren M LM McCann Honour C HC Yoon Minsoo M Warring Suzanne L SL Fineran Peter C PC Mesarich Carl H CH Templeton Matthew D MD

Environmental microbiology 20201007 12

The common polysaccharide antigen (CPA) of the lipopolysaccharide (LPS) from Pseudomonas syringae is highly variable, but the genetic basis for this is poorly understood. We have characterized the CPA locus from P. syringae pv. actinidiae (Psa). This locus has genes for l- and d-rhamnose biosynthesis and an operon coding for ABC transporter subunits, a bifunctional glycosyltransferase and an o-methyltransferase. This operon is predicted to have a role in the transport, elongation and termination ...[more]

PMID: 32985740

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Project description:UnlabelledCommon polysaccharide antigen (CPA) is a conserved cell surface polysaccharide produced by Pseudomonas aeruginosa. It contains a rhamnan homopolymer and is one of the two forms of O polysaccharide attached to P. aeruginosa lipopolysaccharide (LPS). Our laboratory has previously characterized an eight-gene cluster (pa5447-pa5454 in P. aeruginosa PAO1) required for biosynthesis of CPA. Here we demonstrate that an adjacent five-gene cluster pa5455-pa5459 is also involved. Using reverse transcriptase PCR (RT-PCR), we showed that the original eight-gene cluster and the new five-gene cluster are both organized as operons. We have analyzed the LPS phenotypes of in-frame deletion mutants made in each of the five genes, and the results verified that these five genes are indeed required for CPA biosynthesis, extending the CPA biosynthesis locus to contain 13 contiguous genes. By performing overexpression experiments of different sets of these biosynthesis genes, we were able to obtain information about their possible functions in CPA biosynthesis.ImportanceLipopolysaccharide (LPS) is an important cell surface structure of Gram-negative bacteria. The human opportunistic pathogen Pseudomonas aeruginosa simultaneously produces an O-antigen-specific (OSA) form and a common polysaccharide antigen (CPA) form of LPS. CPA, the focus of this study, is composed of α-1-2, α1-3-linked d-rhamnose sugars and has been shown to be important for attachment of the bacteria to human airway epithelial cells. Genome sequencing of this species revealed a new five-gene cluster that we predicted to be involved in CPA biosynthesis and modification. In this study, we have generated chromosomal knockouts by performing in-frame deletions and allelic replacements. Characterizing the function of each of the five genes is important for us to better understand CPA biosynthesis and the mechanisms of chain length termination and regulation of this unique D-rhamnan polysaccharide.

Project description:The Pseudomonas syringae complex comprises different genetic groups that include strains from both agricultural and environmental habitats. This complex group has been used for decades as a "hodgepodge," including many taxonomically related species. More than 60 pathovars of P. syringae have been described based on distinct host ranges and disease symptoms they cause. These pathovars cause disease relying on an array of virulence mechanisms. However, P. syringae pv. syringae (Pss) is the most polyphagous bacterium in the P. syringae complex, based on its wide host range, that primarily affects woody and herbaceous host plants. In early 1990s, bacterial apical necrosis (BAN) of mango trees, a critical disease elicited by Pss in Southern Spain was described for the first time. Pss exhibits important epiphytic traits and virulence factors, which may promote its survival and pathogenicity in mango trees and in other plant hosts. Over more than two decades, Pss strains isolated from mango trees have been comprehensively investigated to elucidate the mechanisms that governs their epiphytic and pathogenic lifestyles. In particular, the vast majority of Pss strains isolated from mango trees produce an antimetabolite toxin, called mangotoxin, whose leading role in virulence has been clearly demonstrated. Moreover, phenotypic, genetic and phylogenetic approaches support that Pss strains producers of BAN symptoms on mango trees all belong to a single phylotype within phylogroup 2, are adapted to the mango host, and produce mangotoxin. Remarkably, a genome sequencing project of the Pss model strain UMAF0158 revealed the presence of other factors that may play major roles in its different lifestyles, such as the presence of two different type III secretion systems, two type VI secretion systems and an operon for cellulose biosynthesis. The role of cellulose in increasing mango leaf colonization and biofilm formation, and impairing virulence of Pss, suggests that cellulose may play a pivotal role with regards to the balance of its different lifestyles. In addition, 62-kb plasmids belonging to the pPT23A-family of plasmids (PFPs) have been strongly associated with Pss strains that inhabit mango trees. Further, complete sequence and comparative genomic analyses revealed major roles of PFPs in detoxification of copper compounds and ultraviolet radiation resistance, both improving the epiphytic lifestyle of Pss on mango surfaces. Hence, in this review we summarize the research that has been conducted on Pss by our research group to elucidate the molecular mechanisms that underpin the epiphytic and pathogenic lifestyle on mango trees. Finally, future directions in this particular plant-pathogen story are discussed.

Dataset Information

Variation at the common polysaccharide antigen locus drives lipopolysaccharide diversity within the Pseudomonas syringae species complex.

Publications

Variation at the common polysaccharide antigen locus drives lipopolysaccharide diversity within the Pseudomonas syringae species complex.

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