ABSTRACT
The common hop (Humulus lupulus L., Cannabaceae) is a perennial plant cultivated in the temperate climate zone and used in the brewing and pharmaceutical industry. In June 2021, symptoms of wilting and subsequent drying of shoots were observed on hop plants (cv. Lubelski) in Lubelskie Province, in Poland (50°55'30.5"N, 22°10'35.4"E). Wilted shoots showed no symptoms of chlorosis. Usually both healthy and wilted shoots were present on the same plant. Approx. 20% of the plants in the 2 hectare hop garden showed these symptoms. An inspection of the underground parts of the infected shoots revealed a brown necrosis of the tissues partly covered by white mycelium with black sclerotia. These symptoms were characteristic for Sclerotinia sclerotiorum (Lib.) de Bary (Bolton et al. 2006), but this pathogen has never been detected in hop gardens in Poland before. In order to confirm the preliminary diagnosis, shoots from five different plants with symptoms of necrosis were collected, disinfected with 1% sodium hypochlorite for 1 min, rinsed with sterile water and dried. Twelve fragments from partially necrotized tissue were placed on potato dextrose agar (PDA) amended with chlortetracycline hydrochloride. After three days of incubation at room temperature and daylight, 75% of the explants had white or light grey mycelia present. Pure cultures were obtained by transplanting mycelium to fresh PDA. Within 4 days, the mycelium has reached the diameter of Petri dishes (90 mm). On PDA at room temperature and daylight, black spherical or cylindrical sclerotia formed after 6-14 days. Sclerotial size was 1.0 to 10.0 x 1.0 to 5.0 mm (average 2.8 x 2.0 mm; n = 57 from 3 plates). Five isolates were subjected to DNA extraction. Then ITS region was amplified and sequenced using primers ITS1/ITS4 (White et al. 1990). The sequences obtained from the five fungal isolates were identical with each other and BLAST analysis revealed also 100% identity with the GenBank records of S. sclerotiorum (e.g. KT224645, Baturo-Ciesniewska et al. 2017). One representative sequence of isolate Ss5HL was deposited to GenBank (OQ998981). Pathogenicity of the isolate Ss5HL was confirmed in inoculation tests on approx. 30 cm high, young shoots of hop plants (cv. Lubelski) grown from rootstock for 4 weeks in the greenhouse. Five plants were inoculated by placing mycelial discs from a 7-day-old culture on shoots and covering them with wet sterilized cotton pads and aluminum foil. Two control plants were treated in the same way but instead of mycelial discs pure agar discs were placed on the shoots. Then plants were kept in a climate chamber (80% relative humidity, 15-h light at 22°C, 9-h dark at 18°C). All fungal inoculated shoots wilted within 3-4 days, then necrosis developed and spread from the place of inoculation up the shoot. No tissue necrosis was observed below the place of inoculation. The control plants remained asymptomatic. The inoculation test was carried out twice with the same result. Fungal cultures reisolated from the inoculated shoots were morphologically identical with the culture used as inoculum. Isolate Ss5HL was deposited in the collection of the Westerdijk Fungal Biodiversity Institute (CBS 150077).To our knowledge, this is the first case of Sclerotinia wilt of hops in Poland and also in Europe, with the exception of one, nearly 100 years old report from the United Kingdom (Salmon and Ware 1936). S. sclerotiorum rarely occurs on hop but it can cause severe yield reduction if pathogen accumulates in the soil.
ABSTRACT
Black root rot (BRR) caused by Thielaviopsis basicola as well as Tomato spotted wilt virus (TSWV) are the most serious problems in tobacco growing regions. We crossed the breeding line WGL 3 carrying BRR resistance derived from N.glauca with the line PW-834 the resistance of which to TSWV was transferred from cultivar Polalta. Anthers obtained from F(1) hybrid plants were cultured to induce haploids combining resistance to Th. basicola and TSWV. Flow cytometry analysis revealed 242 haploids and 2 spontaneous doubled haploids among regenerants. All haploids were cloned and then evaluated for BRR as well as TSWV resistance. The presence of pathogens was detected by microscopic evaluation of roots or using DAS-ELISA test. Microscopic assessment showed that, 132 haploids had no symptoms of Th. basicola which, together with the absence of symptoms in the F(1) hybrids, indicated a dominant monogenic mode of inheritance. At the same time only 30 haploids demonstrated resistance to TSWV. SCAR markers associated with TSWV resistance gene detection was applied. The results indicate that small proportion of TSWV-resistant haploids is probably due to the influence of deleterious genes flanking the resistance factor that reduced vitality of gametophytes. Altogether, 24 haploids showed multiple resistance to Th. basicola and TSWV.