Dataset Information

Genomic Characterization, Formulation and Efficacy in Planta of a Siphoviridae and Podoviridae Protection Cocktail against the Bacterial Plant Pathogens Pectobacterium spp.

ABSTRACT: In the face of global human population increases, there is a need for efficacious integrated pest management strategies to improve agricultural production and increase sustainable food production. To counteract significant food loses in crop production, novel, safe and efficacious measures should be tested against bacterial pathogens. Pectobacteriaceae species are one of the causative agents of the bacterial rot of onions ultimately leading to crop losses due to ineffective control measures against these pathogens. Therefore, the aim of this study was to isolate and characterize bacteriophages which could be formulated in a cocktail and implemented in planta under natural environmental conditions. Transmission electron microscopy (TEM) and genome analysis revealed Siphoviridae and Podoviridae family bacteriophages. To test the protective effect of a formulated phage cocktail against soft rot disease, three years of field trials were performed, using three different methods of treatment application. This is the first study to show the application of a phage cocktail containing Podoviridae and Siphoviridae bacteriophages capable of protecting onions against soft rot in field conditions.

SUBMITTER: Zaczek-Moczydlowska MA

PROVIDER: S-EPMC7077305 | biostudies-literature | 2020 Jan

REPOSITORIES: biostudies-literature

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Genomic Characterization, Formulation and Efficacy in Planta of a Siphoviridae and Podoviridae Protection Cocktail against the Bacterial Plant Pathogens Pectobacterium spp.

Zaczek-Moczydłowska Maja A MA Young Gillian K GK Trudgett James J Fleming Colin C CC Campbell Katrina K O'Hanlon Richard R

Viruses 20200128 2

In the face of global human population increases, there is a need for efficacious integrated pest management strategies to improve agricultural production and increase sustainable food production. To counteract significant food loses in crop production, novel, safe and efficacious measures should be tested against bacterial pathogens. Pectobacteriaceae species are one of the causative agents of the bacterial rot of onions ultimately leading to crop losses due to ineffective control measur ...[more]

PMID: 32012814

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Project description:Soft rot pathogenic bacteria from the genus Dickeya cause severe economic losses in orchid nurseries worldwide, and there is no effective control currently available. In the last decade, the genus Dickeya has undergone multiple changes as multiple new taxa have been described, and just recently a new putative Dickeya species was reported. This study reports the isolation of three bacteriophages active against putative novel Dickeya spp. isolates from commercially produced infected orchids that show variable host-range profiles. Bacteriophages were isolated through enrichment from Dickeya-infected orchid tissue. Convective interaction media monolith chromatography was used to isolate bacteriophages from wastewaters, demonstrating its suitability for the isolation of infective bacteriophages from natural sources. Based on bacteriophage morphology, all isolated bacteriophages were classified as being in the order Caudovirales, belonging to three different families, Podoviridae, Myoviridae, and Siphoviridae. The presence of three different groups of bacteriophages was confirmed by analyzing the bacteriophage specificity of bacterial hosts, restriction fragment length polymorphism and plaque morphology. Bacteriophage BF25/12, the first reported Podoviridae bacteriophage effective against Dickeya spp., was selected for further characterization. Its genome sequence determined by next-generation sequencing showed limited similarity to other characterized Podoviridae bacteriophages. Interactions among the bacteriophages and Dickeya spp. were examined using transmission electron microscopy, which revealed degradation of electron-dense granules in response to bacteriophage infection in some Dickeya strains. The temperature stability of the chosen Podoviridae bacteriophage monitored over 1 year showed a substantial decrease in the survival of bacteriophages stored at -20°C over longer periods. It showed susceptibility to low pH and UV radiation but was stable in neutral and alkaline pH. Furthermore, the stability of the tested bacteriophage was also connected to the incubation medium and bacteriophage concentration at certain pH values. Finally, the emergence of bacteriophage-resistant bacterial colonies is highly connected to the concentration of bacteriophages in the bacterial environment. This is the first report on bacteriophages against Dickeya from the Podoviridae family to expand on potential bacteriophages to include in bacteriophage cocktails as biocontrol agents. Some of these bacteriophage isolates also showed activity against Dickeya solani, an aggressive strain that causes the soft rot of potatoes, which indicates their broad potential as biocontrol agents.

Project description:Pectobacterium carotovorum subsp. carotovorum is a necrotrophic plant pathogen that secretes plant cell wall-degrading enzymes (PCWDEs) that cause soft rot disease in various crops. Bacteriophages have been under consideration as harmless antibacterial agents to replace antibiotics and copper-based pesticides. However, the emergence of bacteriophage resistance is one of the main concerns that should be resolved for practical phage applications. In this study, we developed a phage cocktail with three lytic phages that recognize colanic acid (phage POP12) or flagella (phages POP15 and POP17) as phage receptors to minimize phage resistance. The phage cocktail effectively suppressed the emergence of phage-resistant P. carotovorum subsp. carotovorum compared with single phages in in vitro challenge assays. The application of the phage cocktail to napa cabbage (Brassica rapa subsp. pekinensis) resulted in significant growth retardation of P. carotovorum subsp. carotovorum (P < 0.05) and prevented the symptoms of soft rot disease. Furthermore, phage cocktail treatments of young napa cabbage leaves in a greenhouse environment indicated effective prevention of soft rot disease compared to that in the nonphage negative control. We isolated 15 phage-resistant mutants after a phage cocktail treatment to assess the virulence-associated phenotypes compared to those of wild-type (WT) strain Pcc27. All mutants showed reduced production of four different PCWDEs, leading to lower levels of tissue softening. Ten of the 15 phage-resistant mutants additionally exhibited decreased swimming motility. Taken together, these results show that the phage cocktail developed here, which targets two different types of phage receptors, provides an effective strategy for controlling P. carotovorum subsp. carotovorum in agricultural products, with a potential ability to attenuate P. carotovorum subsp. carotovorum virulence. IMPORTANCE Pectobacterium carotovorum subsp. carotovorum is a phytopathogen that causes soft rot disease in various crops by producing plant cell wall-degrading enzymes (PCWDEs). Although antibiotics and copper-based pesticides have been extensively applied to inhibit P. carotovorum subsp. carotovorum, the emergence of antibiotic-resistant bacteria and demand for harmless antimicrobial products have emphasized the necessity of finding alternative therapeutic strategies. To address this problem, we developed a phage cocktail consisting of three P. carotovorum subsp. carotovorum-specific phages that recognize colanic acids and flagella of P. carotovorum subsp. carotovorum. The phage cocktail treatments significantly decreased P. carotovorum subsp. carotovorum populations, as well as soft rot symptoms in napa cabbage. Simultaneously, they resulted in virulence attenuation in phage-resistant P. carotovorum subsp. carotovorum, which was represented by decreased PCWDE production and decreased flagellum-mediated swimming motility. These results suggested that preparations of phage cocktails targeting multiple receptors would be an effective approach to biocontrol of P. carotovorum subsp. carotovorum in crops.

Dataset Information

Genomic Characterization, Formulation and Efficacy in Planta of a Siphoviridae and Podoviridae Protection Cocktail against the Bacterial Plant Pathogens Pectobacterium spp.

Publications

Genomic Characterization, Formulation and Efficacy in Planta of a <i>Siphoviridae</i> and <i>Podoviridae</i> Protection Cocktail against the Bacterial Plant Pathogens <i>Pectobacterium</i> spp.

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