Phylogenomic analyses and comparative genomics of Pseudomonas syringae associated with almond (Prunus dulcis) in California

Authors: MAGUVU, TAWANDA - University Of California, Davis; FRIAS, ROSA - University Of California, Davis; HERNANDEZ-ROSAS, ALEJANDRO - University Of California, Davis; HOLTZ, BRENT - University Of California - Cooperative Extension Service; NIEDERHOLZER, FRANZ - University Of California - Cooperative Extension Service; DUNCAN, ROGER - University Of California - Cooperative Extension Service; YAGHMOUR, MOHAMMAND - University Of California - Cooperative Extension Service; CULUMBER, CATHERINE - University Of California - Cooperative Extension Service; GORDON, PHOEBE - University Of California - Cooperative Extension Service; VIEIRA, FLAVIA - University Of California, Riverside; ROLSHAUSEN, PHILIPPE - University Of California, Riverside; ADASKAVEG, JAMES - University Of California, Riverside; Burbank, Lindsey; LINDOW, STEVEN - University Of California Berkeley; TROUILLAS, FLORENT - University Of California, Davis

Submitted to: PLOS ONE
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 1/10/2024
Publication Date: 4/11/2024
Citation: Maguvu, T.E., Frias, R.J., Hernandez-Rosas, A.I., Holtz, B.A., Niederholzer, F.J., Duncan, R.A., Yaghmour, M.A., Culumber, C.M., Gordon, P.E., Vieira, F.C., Rolshausen, P.E., Adaskaveg, J.E., Burbank, L.P., Lindow, S.E., Trouillas, F.P. 2024. Phylogenomic analyses and comparative genomics of Pseudomonas syringae associated with almond (Prunus dulcis) in California. PLOS ONE. 19(4). Article e0297867. https://doi.org/10.1371/journal.pone.0297867.
DOI: https://doi.org/10.1371/journal.pone.0297867

Interpretive Summary: Bacterial canker and bacterial blast of almond are two phases of a disease that can affect most parts of the almond tree, including flowers, leaves, trunk, and scaffold branches. In severe cases, bacterial blast can lead to more than 50% reduction in yield. Moreover, cankers can girdle scaffold branches leading to branch dieback; girdling of the trunk might lead to the death of the entire tree. The disease has been mainly attributed to Pseudomonas syringae pv. syringae but this pathogen species has not been well described in almond. In this project, 98 fluorescent pseudomonads isolated from different almond tissues and other Prunus sp. were sequenced and characterized. Pseudomonas syringae pv. syringae, P. cerasi, and P. viridiflava were identified as almond canker pathogens. Degree of pathogenicity was compared in laboratory assays, as well as presence of fitness-related genes, ice nucleation activity, and sensitivity to copper and kasugamycin antibiotics. Isolates that showed resistance to copper also contained a copy of the ctpV resistance gene. All isolates were susceptible to kasugamycin at the label-recommended rate of 100 ppm.

Technical Abstract: Bacterial canker and bacterial blast of almond are two phases of a disease that can affect most parts of the almond tree, including flowers, leaves, trunk, and scaffold branches. In severe cases, bacterial blast can lead to more than 50% reduction in yield. Moreover, cankers can girdle scaffold branches leading to branch dieback; girdling of the trunk might lead to the death of the entire tree. The disease has been mainly attributed to Pseudomonas syringae pv. syringae however, there are limited studies that comprehensively characterized Pseudomonas species affecting almonds. To address this, we sequenced and comprehensively analysed the genomic architecture of 98 fluorescent pseudomonads isolated from different symptomatic and asymptomatic almond tissues and a few other Prunus spp. Phylogenomic analysis, genome mining, field pathogenicity tests, and in vitro ice nucleation and antibiotic sensitivity tests were integrated to improve knowledge of the biology and management of bacterial blast and bacterial canker of almond. We identified P. syringae pv. syringae, P. cerasi, and P. viridiflava as almond canker pathogens. P. syringae pv. syringae caused both canker and foliar (blast) symptoms. In contrast, P. cerasi and P. viridiflava only caused canker, although P. viridiflava appeared to be a weak pathogen of almond. Isolates belonging to P. syringae pv. syringae were the most frequently isolated among the pathogenic species/pathovars, contributing 75% of all pathogenic isolates. P. cerasi and P. viridiflava isolates contributed to 8.3 and 16.7% of the pathogenic isolates, respectively. Laboratory leaf infiltration bioassay produced results distinct from experiments in the field with both P. cerasi and P. syringae pv. syringae, causing significant necrosis and browning of detached leaves, whereas P. viridiflava conferred moderate effects. Genome mining revealed the absence of key epiphytic fitness-related genes in P. cerasi and P. viridiflava genomic sequences, which could explain the contrasting field and laboratory bioassay results. P. syringae pv. syringae and P. cerasi isolates harboured the ice nucleation protein, which correlated with the ice nucleation phenotype. Results of sensitivity tests to copper and kasugamycin showed a strong linkage to putative resistance genes. Isolates harbouring the ctpV gene showed resistance to copper up to 600 ppm. In contrast, isolates without the ctpV gene could not grow on nutrient agar amended with 200 ppm copper, suggesting ctpV can be used to phenotype copper resistance. All isolates were susceptible to kasugamycin at the label-recommended rate of 100 ppm.