Japanese Journal of Phytopathology

We improved an established one-step multiplex RT-PCR method to detect six orthotospoviruses that infect Solanaceae plants (tomato spotted wilt virus [TSWV], newly reported tomato zonate spot virus [TZSV], impatiens necrotic spot virus [INSV], chrysanthemum stem necrosis virus [CSNV], watermelon silver mottle virus [WSMoV], capsicum chlorosis virus [CaCV]). Specific primer sets for one-step multiplex RT-PCR were designed for TSWV, TZSV, INSV, CSNV, WSMoV, and CaCV, and this optimized RT-PCR amplified DNA fragments corresponding to the target virus species: TSWV (831 bp), TZSV (734 bp), INSV (592 bp), CSNV (502 bp), WSMoV (395 bp), CaCV (324 bp). All combinations of orthotospoviruses were detected in mixed samples. The method was applied to Solanaceae plants infected with orthotospoviruses under field conditions.

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Seven isogenic lines (ILs) of the rice cultivar Koshihikari with different true resistance genes to blast were previously developed to create the disease-resistant multiline Koshihikari Niigata BL in Niigata Prefecture. This multiline has been cultivated with seed mixtures of four of the seven ILs on 68,000~94,000 ha in the prefecture since 2005. The multiline, consisting of 70% resistant and 30% susceptible ILs to most of distributed blast races, has controlled blast for 18 years with systematic changes to its component resistant ILs to avoid resistance breakdown. To verify the validity of this method, we investigated the distribution of blast races in Niigata Prefecture before and after introduction of the multiline and monitored blast races in several areas where virulent races to the resistant ILs were isolated at high frequencies. Within 2–3 years of the multiline introduction, the dominant races changed rapidly from 001.0 and 003.0 to 007.0 and 037.1. The virulent races to the resistant ILs overwintered in areas where they were isolated at high frequency. Their frequencies increased in years when rice plants susceptible to these races were planted at high rates and decreased markedly in years when 30% of the multiline mixture were susceptible to the races, owing to replacement of component ILs in the multiline. Isolation probability of the virulent races was lower when isolation frequencies of the races had decreased in the previous years. After 2–3 consecutive years of cultivation rates of 30% susceptible ILs in the multiline, no virulent races were isolated, even after rates of the susceptible ILs in the multiline increased again. Our results show that virulent races did not increase when component ILs in the multiline are replaced. Thus, systematically changing component ILs in the rice multiline is an effective means to ensure the durability of true resistance genes to rice blast.

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