Novel biovar (biovar 6) of Pseudomonas syringae pv. actinidiae causing ­bacterial canker of kiwifruit ( Actinidia deliciosa) in Japan.

Abstract

"Typical symptoms of bacterial canker disease (e.g., cankers with red exudates, white ooze exuding from pruning cuts, browning and discoloration beneath bark) were observed on trunks, branches and twigs of Actinidia deliciosa ‘Hayward’ in Nagano Prefecture, Japan, in April 2015. Thirteen strains of the causal bacterium were isolated from the diseased twigs and demonstrated by inoculation and reisolation to be pathogenic on A. deliciosa. The strains were gram-negative, aerobic rods with one to two polar flagella, and formed opaque, pale yellowish circular colonies. On the basis of biochemical and physiological characterization, PCR assays targeting ITS, hopZ3 and hopO1-2, and a multilocus sequence analysis (MLSA) using concatenated sequences of seven housekeeping genes (acnB, cts, gapA, gyrB, pfk, pgi and rpoD), we identified the pathogen as Pseudomonas syringae pv. actinidiae (Psa). However, the pathogen proved to be different from the four existing biovars (biovars 1, 2, 3 and 5) of Psa based on phenotypic characterization using API 20NE, and additional PCR assays, which showed that the pathogen did not possess hopH1, hopH3 or hopZ5 genes. Also, the results of the MLSA revealed that they clustered separately from the other Psa biovars. In addition, bioassays confirmed that they produced both phaseolotoxin and coronatine, although no plant pathogenic bacteria have ever been identified as producing both toxins. The combined results of genotypic and phenotypic analyses therefore support the classification of these strains as a novel biovar of Psa, for which the name “biovar 6” is proposed. In addition to biovar 6, three other biovars (biovars 1, 3 and 5) of Psa were confirmed as bacterial canker pathogens of kiwifruit in Japan. Although the distribution of biovar 6 is extremely limited at present, it may play an important role in helping us to understand the origin and evolution of Psa. Our results also show that toxin-related genes, which could be transmissible among bacteria, are not adequate markers to discriminate Psa biovars, although phaseolotoxin and coronatine biosynthetic genes have been utilized as PCR targets to identify biovars 1 and 2, respectively."