Characterisation of Pectobacterium carotovorum subsp. brasiliense isolates causing blackleg and soft rot diseases of potato in South Africa

Abstract

Pectobacterium carotovorum subsp. brasiliense (Pcb) is a plant pathogenic bacterium that causes blackleg and tuber soft rot disease of potato worldwide. Pectobacterium spp. are characterized by the secretion of large quantities of plant cell wall degrading enzymes. As the name indicates, Pectobacteria are pectinolytic pathogens, producing enzymes such as pectate lyase, polygalacturonase, and many others that are used to catalyse the breakdown of pectin, the main plant cell wall component. Consequently, virulence of Pectobacteria is highly reliant upon the production and secretion of macerating enzymes. Hence these bacteria are also referred to as “brute-force” pathogens. Infection and disease symptoms on plants commonly result in the development of blackleg disease, a characteristic black-like decay extending on the stems of infected potato plants. Furthermore, the infection of tubers results in the development of soft rot disease. Pcb is of particular interest in that among Pectobacterium spp. such as Pectobacterium atrosepticum (Pa), P. carotovorum subsp. carotovorum (Pcc), and P. wasabiae, Pcb strains are reported to be the most aggressive and virulent pathogens causing blackleg and soft rot disease of potato in many growing regions across the world. The fact that strains of Pcb were recently reported and isolated in South Africa has necessitated that this work be undertaken in order to characterise this newly described important pathogen of potato in regard to its phenotypic, genetic diversity, virulence and host range traits. Therefore in this work Pcb strains were subjected to multilocus phylogenetic analyses (MLSA) in order to investigate and determine whether there is any interspecies and intraspecies genetic diversity among the South African Pcb isolates. It was thus established that there is a significant genetic diversity that exists both on an interspecies and intraspecies level among Pcb isolates. As a result we sought to investigate further if the level of genetic diversity observed can be reflected in terms of the pathogen’s virulence, biochemical, phenotypic as well as host range characteristics. The results of virulence assays on potato tubers and stems indicated that Pcb strains are significantly much more virulent on potato compared to closely related Pectobacterium spp. such as Pa and Pcc. Moreover, the level of intraspecies diversity observed through phylogeny was also evident and reflected on the phenotypic, virulence and host range characteristics of the pathogen. This study also focused on investigating virulence factors employed by Pectobacterium spp. during infection. Such factors include the ability to produce and secrete of various extracellular macerating enzymes, as well as screening for the presence of virulence associated effectors and phytotoxin genes. It was of interest to observe that Pcb strains have the ability to grow and produce substrate-degrading enzymes much more rapidly compared to Pa and Pcc. This phenomenon was also observed in virulence assays where Pcb strains were noted to cause more rapid and most severe maceration symptoms on potato tubers and stems. Thus in agreement with other studies, our results suggests that Pcb is a uniquely sophisticated but diverse plant pathogen which can be considered to be one of the most aggressive causal agents of blackleg and soft rot disease of potato in South Africa.