Pantoea ananatis Utilizes a Type VI Secretion System for Pathogenesis and Bacterial Competition

Abstract

Type VI secretion systems (T6SSs) are a class of macromolecular machines that are recognized as an important virulence mechanism in several Gram-negative bacteria. The genome of Pantoea ananatis LMG 2665 T , a pathogen of pineapple fruit and onion plants, carries two gene clusters whose predicted products have homology with T6SS-associated gene products from other bacteria. Nothing is known regarding the role of these T6SS-1 and T6SS-3 gene clusters in the biology of P. ananatis. Here, we present evidence that T6SS-1 plays an important role in the pathogenicity of P. ananatis LMG 2665T in onion plants, while a strain lacking T6SS-3 remains as pathogenic as the wild-type strain. We also investigated the role of the T6SS-1 system in bacterial competition, the results of which indicated that several bacteria compete less efficiently against wild-type LMG 2665 T than a strain lacking T6SS-1. Additionally, we demonstrated that these phenotypes of strain LMG 2665 T were reliant on the core T6SS products TssA and TssD (Hcp), thus indicating that the T6SS-1 gene cluster encodes a functioning T6SS. Collectively, our data provides the first evidence demonstrating that the T6SS-1 system is a virulence determinant of P. ananatis LMG 2665 T and plays a role in bacterial competition.

The impact of type VI secretion system, bacteriocins and antibiotics on competition amongst Soft-RotEnterobacteriaceae: Regulation of carbapenem biosynthesis by iron and the transcriptional regulator Fur

“…Inter and intra species bacterial competition assays were performed as previously described (25). The complete list of bacteria used in this assay is provided in Table 1.…”

“…The different Pcb 1692 mutant strains were generated by site-directed mutagenesis using the lambda red recombination technique (124). In summary, overlap extension polymerase chain reaction (PCR) was used to generate a gene knockout cassette by fusing the upstream and downstream regions flanking the targeted gene to the kanamycin resistance genes (25, 125). The fused PCR product was then electroporated into Pcb 1692 harbouring pKD20 and transformants were selected on nutrient agar supplemented with either 50 µg/ml kanamycin or 50 µg/ml chloramphenicol.…”

Pantoea ananatis: genomic insights into a versatile pathogen

“…In a study conducted by Shyntum et al . (), it was shown that the first locus is fully functional and plays a role in pathogenicity in plant hosts, specifically onion seedlings, and is also an important factor in intra‐ and interspecies bacterial competition.…”

Influence of the ferric uptake regulator (Fur) protein on pathogenicity in Pectobacterium carotovorum subsp. brasiliense

“…The kanamycin resistance gene was PCR amplified from plasmid pKD4. The three amplicons were fused by overlap extension PCR to produce a gene disruption cassette, as described previously [31]. The fused PCR product was then electroporated into Pcb1692 harboring pKD46 and transformants were selected on nutrient agar supplemented with 50 μg/ml kanamycin.…”