Comprehensive survey of copper resistance and analysis of responsible genes in Pseudomonas syringae pv. actinidiae biovar 1 and biovar 3 isolates from Japan

The control of kiwifruit canker, caused by Pseudomonas syringae pv. actinidiae (Psa), largely relies on copper pesticides. After epidemics of kiwifruit canker in Ehime Prefecture, Japan, we comprehensively surveyed isolates of Psa biovars 1 (Psa 1) and 3 (Psa 3) for copper resistance from 2002 to 2019. In Psa 1, copper-resistant isolates drastically increased in 2006 and onward during the survey from 2002 to 2008 and were frequently found in 2017. By contrast, copper-resistant Psa 3 isolates emerged soon after its first identification in 2014 and spread to different orchards with an isolation ratio of about 10%. Identification of copper resistance-related genes by whole-genome resequencing revealed that cop genes from Psa 1 isolates in 2017 were almost identical to those from Psa 1 isolates from the late 1980s. On the contrary, in Psa 3 isolates, we found two different sets of cop genes. One of them was closely related to those from other Pseudomonas species, and the other to those from P. syringae pv. tomato. Notably, copper-sensitive Psa1 and 3 isolates had copAB and copRS homologs but no homologs of copCD. Despite the diversity in cop gene sequences, PCR detection of copCD sequences from different lineages matched perfectly with their copper resistance.