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A Synergic Potential of Antimicrobial Peptides against Pseudomonas syringae pv. actinidiae.

ABSTRACT: Pseudomonas syringae pv. actinidiae (Psa) is the pathogenic agent responsible for the bacterial canker of kiwifruit (BCK) leading to major losses in kiwifruit productions. No effective treatments and measures have yet been found to control this disease. Despite antimicrobial peptides (AMPs) having been successfully used for the control of several pathogenic bacteria, few studies have focused on the use of AMPs against Psa. In this study, the potential of six AMPs (BP100, RW-BP100, CA-M, 3.1, D4E1, and Dhvar-5) to control Psa was investigated. The minimal inhibitory and bactericidal concentrations (MIC and MBC) were determined and membrane damaging capacity was evaluated by flow cytometry analysis. Among the tested AMPs, the higher inhibitory and bactericidal capacity was observed for BP100 and CA-M with MIC of 3.4 and 3.4-6.2 µM, respectively and MBC 3.4-10 µM for both. Flow cytometry assays suggested a faster membrane permeation for peptide 3.1, in comparison with the other AMPs studied. Peptide mixtures were also tested, disclosing the high efficiency of BP100:3.1 at low concentration to reduce Psa viability. These results highlight the potential interest of AMP mixtures against Psa, and 3.1 as an antimicrobial molecule that can improve other treatments in synergic action.

SUBMITTER: Mariz-Ponte N

PROVIDER: S-EPMC7962642 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Characterization of a Lytic Bacteriophage against Pseudomonas syringae pv. actinidiae and Its Endolysin.

Project description:Pseudomonas syringae pv. actinidiae (Psa) is a phytopathogen that causes canker in kiwifruit. Few conventional control methods are effective against this bacterium. Therefore, alternative approaches, such as phage therapy are warranted. In this study, a lytic bacteriophage (PN09) of Psa was isolated from surface water collected from a river in Hangzhou, China in 2019. Morphologically, PN09 was classified into the Myoviridae family, and could lyse all 29 Psa biovar 3 strains. The optimal temperature and pH ranges for PN09 activity were determined as 25 to 35 ∘C and 6.0 to 9.0, respectively. The complete genome of PN09 was found to be composed of a linear 99,229 bp double-stranded DNA genome with a GC content of 48.16%. The PN09 endolysin (LysPN09) was expressed in vitro and characterized. LysPN09 was predicted to belong to the Muraidase superfamily domain and showed lytic activity against the outer-membrane-permeabilized Psa strains. The lytic activity of LysPN09 was optimal over temperature and pH ranges of 25 to 40 ∘C and 6.0 to 8.0, respectively. When recombinant endolysin LysPN09 was combined with EDTA, Psa strains were effectively damaged. All these characteristics demonstrate that the phage PN09 and its endolysin, LysPN09, are potential candidates for biocontrol of Psa in the kiwifruit industry.

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Characterization of Bacteriophages against Pseudomonas Syringae pv. Actinidiae with Potential Use as Natural Antimicrobials in Kiwifruit Plants.

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Project description:Bacteriophages are viruses that specifically infect target bacteria. Recently, bacteriophages have been considered potential biological control agents for bacterial pathogens due to their host specificity. Pseudomonas syringae pv. actinidiae (Psa) is a reemerging pathogen that causes bacterial canker of kiwifruit (Actinidia sp.). The economic impact of this pest and the development of resistance to antibiotics and copper sprays in Psa and other pathovars have led to investigation of alternative management strategies. Phage therapy may be a useful alternative to conventional treatments for controlling Psa infections. Although the efficacy of bacteriophage φ6 was evaluated for the control of Psa, the characteristics of other DNA bacteriophages infecting Psa remain unclear. In this study, the PHB09 lytic bacteriophage specific to Psa was isolated from kiwifruit orchard soil. Extensive host range testing using Psa isolated from kiwifruit orchards and other Pseudomonas strains showed PHB09 has a narrow host range. It remained stable over a wide range of temperatures (4-50 °C) and pH values (pH 3-11) and maintained stability for 50 min under ultraviolet irradiation. Complete genome sequence analysis indicated PHB09 might belong to a new myovirus genus in Caudoviricetes. Its genome contains a total of 94,844 bp and 186 predicted genes associated with phage structure, packaging, host lysis, DNA manipulation, transcription, and additional functions. The isolation and identification of PHB09 enrich the research on Pseudomonas phages and provide a promising biocontrol agent against kiwifruit bacterial canker.

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Genome analysis of the kiwifruit canker pathogen Pseudomonas syringae pv. actinidiae biovar 5.

Project description:Pseudomonas syringae pv. actinidiae (Psa) is a destructive pathogen of kiwifruit bacterial canker disease, causing severe economic losses to kiwifruit industry worldwide. Biovar 5 is the most recently reported biovar of Psa, and is found in only a local area of Japan at present. There is not much information of genetic characteristics of biovar 5. Thus, the genome of biovar 5 was sequenced and analyzed to clarify its detailed genetic characteristics. Here, the genomes of strain MAFF 212056 and MAFF 212061 of biovar 5 were estimated to be about 6.3 Mbp and 6.5 Mbp, respectively, and their phylogenetic positions were proved to be near that of biovar 2 in the phylogenetic tree. However, it was confirmed that biovar 5 had neither the coronatine biosynthetic genes conserved in biovar 2, its phylogenetic neighbor, nor the phaseolotoxin biosynthetic genes conserved in biovar 1, Japanese native pathogen. In addition, 45 genes of type III secreted effectors were identified in biovar 5 genomes, showing that their composition is different from that in the other biovars. Moreover, some biovar 5-specific regions were identified. Then, biovar 5-specific PCR primers for targeting these regions were designed, and proved to be applicable for detecting biovar 5 specifically.

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Comparative genomics reveal pathogenicity-related loci in Pseudomonas syringae pv. actinidiae biovar 3.

Project description:Bacterial canker of kiwifruit, is a severe global disease caused by Pseudomonas syringae pv. actinidiae (Psa). Here, we found that Psa biovar 3 (Psa3) was the only biovar consisting of three widely distributed clades in the largest Chinese kiwifruit cultivated area. Comparative genomics between the three clades revealed 13 polymorphic genes, each of which had multiple intra-clade variations. For instance, we confirmed that the polymorphic copA gene, which encodes a periplasmic protein CopA that is translocated by the Twin-arginine targeting (Tat) system, was involved in copper tolerance. We also found extensive variation in pathogenicity amongst strains within each genetically monomorphic clade. Accordingly, the pathogenic determinants of Psa3 were identified via a genomic comparison of phenotypically different strains within each clade. A case study of the high- and low-virulence strains in the clade 2 of Psa3 revealed that an hfq variant involved in in vitro growth and virulence, while a conserved locus 930 bp upstream of the hrpR gene in the Type III secretion system (T3SS) cluster was required for full pathogenicity on kiwifruit and elicitation of the hypersensitivity response on non-host Nicotiana benthamiana. The '-930' locus is involved in transcriptional regulation of hrpR/S and modulates T3SS function via the hierarchical 'HrpR/S-HrpL-T3SS/effector' regulatory cascade in Psa. Our results provide insights into the molecular basis underlying the genetic diversification and evolution of pathogenicity in Psa3 since kiwifruit canker emerged in China in the 1980s.

| S-EPMC6589868 | biostudies-literature

Identification of bacteriophages for biocontrol of the kiwifruit canker phytopathogen Pseudomonas syringae pv. actinidiae.

Project description:Pseudomonas syringae pv. actinidiae is a reemerging pathogen which causes bacterial canker of kiwifruit (Actinidia sp.). Since 2008, a global outbreak of P. syringae pv. actinidiae has occurred, and in 2010 this pathogen was detected in New Zealand. The economic impact and the development of resistance in P. syringae pv. actinidiae and other pathovars against antibiotics and copper sprays have led to a search for alternative management strategies. We isolated 275 phages, 258 of which were active against P. syringae pv. actinidiae. Extensive host range testing on P. syringae pv. actinidiae, other pseudomonads, and bacteria isolated from kiwifruit orchards showed that most phages have a narrow host range. Twenty-four were analyzed by electron microscopy, pulse-field gel electrophoresis, and restriction digestion. Their suitability for biocontrol was tested by assessing stability and the absence of lysogeny and transduction. A detailed host range was performed, phage-resistant bacteria were isolated, and resistance to other phages was examined. The phages belonged to the Caudovirales and were analyzed based on morphology and genome size, which showed them to be representatives of Myoviridae, Podoviridae, and Siphoviridae. Twenty-one Myoviridae members have similar morphologies and genome sizes yet differ in restriction patterns, host range, and resistance, indicating a closely related group. Nine of these Myoviridae members were sequenced, and each was unique. The most closely related sequenced phages were a group infecting Pseudomonas aeruginosa and characterized by phages JG004 and PAK_P1. In summary, this study reports the isolation and characterization of P. syringae pv. actinidiae phages and provides a framework for the intelligent formulation of phage biocontrol agents against kiwifruit bacterial canker.

| S-EPMC3993152 | biostudies-literature

Effector loss drives adaptation of Pseudomonas syringae pv. actinidiae biovar 3 to Actinidia arguta.

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| S-EPMC9182610 | biostudies-literature

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