title: A quorum sensing-defective mutant of Pectobacterium carotovorum ssp. brasiliense 1692 is attenuated in virulence and unable to occlude xylem tissue of susceptible potato plant stems.
abstract: Pectobacterium carotovorum ssp. brasiliense 1692 (Pcb1692) is an important emerging pathogen of potatoes causing blackleg in the field and soft rot during post-harvest storage. Blackleg diseases involve the bacterial colonization of vascular tissue and the formation of aggregates, also known as biofilms. To understand the role of quorum sensing in vascular colonization by Pcb1692, we generated a Pcb1692ΔexpI mutant strain. Inactivation of expI led to the reduced production of plant cell wall-degrading enzymes (PCWDEs), the inability to produce acyl homoserine lactone (AHL) and reduced virulence in potato tubers and stems. Complementation of the mutant strain with the wild-type expI gene in trans successfully restored AHL and PCWDE production as well as virulence. Transmission electron microscopy and in vitro motility assays demonstrated hyperpiliation and loss of flagella and swimming motility in the mutant strain compared with the wild-type Pcb1692. Furthermore, we noted that, in the early stages of infection, Pcb1692 wild-type cells had intact flagella which were shed at the later stages of infection. Confocal laser microscopy of PcbΔexpI-inoculated plants showed that the mutant strain tended to aggregate in intercellular spaces, but was unable to transit to xylem tissue. On the contrary, the wild-type strain was often observed forming aggregates within xylem tissue of potato stems. Gene expression analyses confirmed that flagella are part of the quorum sensing regulon, whereas fimbriae and pili appear to be negatively regulated by quorum sensing. The relative expression levels of other important putative virulence genes, such as those encoding different groups of PCWDEs, were down-regulated in the mutant compared with the wild-type strain.
authors.affiliation: Forestry, Agriculture and Biotechnology Institute, University of Pretoria, Lunnon Road, Pretoria, South Africa, 0028.|Department of Microbiology and Plant Pathology, University of Pretoria, Lunnon Road, Pretoria, South Africa, 0028.
authors.forename: Lucy Novungayo
authors.initials: LN
authors.lastname: Moleleki
authors.affiliation: Forestry, Agriculture and Biotechnology Institute, University of Pretoria, Lunnon Road, Pretoria, South Africa, 0028.
authors.forename: Rudolph Gustav
authors.initials: RG
authors.lastname: Pretorius
authors.affiliation: Forestry, Agriculture and Biotechnology Institute, University of Pretoria, Lunnon Road, Pretoria, South Africa, 0028.
authors.forename: Collins Kipngetich
authors.initials: CK
authors.lastname: Tanui
authors.affiliation: Forestry, Agriculture and Biotechnology Institute, University of Pretoria, Lunnon Road, Pretoria, South Africa, 0028.
authors.forename: Gabolwelwe
authors.initials: G
authors.lastname: Mosina
authors.affiliation: Department of Microbiology and Plant Pathology, University of Pretoria, Lunnon Road, Pretoria, South Africa, 0028.
authors.forename: Jacques
authors.initials: J
authors.lastname: Theron
mesh_terms: D001681:Biological Assay; D004198:Disease Susceptibility; D005407:Flagella / Q000378:metabolism / Q000648:ultrastructure; D020869:Gene Expression Profiling; D015964:Gene Expression Regulation, Bacterial; D005798:Genes, Bacterial; D009154:Mutation / Q000235:genetics*; D016973:Pectobacterium carotovorum / Q000235:genetics* / Q000472:pathogenicity* / Q000648:ultrastructure; D010935:Plant Diseases / Q000382:microbiology*; D018547:Plant Stems / Q000382:microbiology*; D035281:Plant Tubers / Q000382:microbiology; D053038:Quorum Sensing / Q000235:genetics*; D011198:Solanum tuberosum / Q000382:microbiology*; D014774:Virulence / Q000235:genetics; D052584:Xylem / Q000382:microbiology*
keywords: Pectobacterium brasiliense; biofilm; expI; soft rot Enterobacteriaceae