Occurence of sclerotinia stem rot of fenugreek caused by Sclerotinia trifoliorum and S. sclerotiorum in Tunisia

Fenugreek is an annual leguminous crop grown for hay and grains in Tunisia. It is also considered a valuable rotation crop with cereals. Sclerotinia rot was observed in production fields since 2010. The survey conducted in 2013 revealed that the incidence of diseased plants varied between 5 and 20%. The identification of isolates of Sclerotinia obtained from fenugreek plants with symptoms of stem rot was determined using morphological and molecular criteria. The size, shape and abundance of sclerotia in potato dextrose agar (PDA) cultures were used to classify isolates as S. sclerotiorum or S. trifoliorum. A comparison of colony diameter on PDA after 24, 48 and 72 h at 25 °C, showed that one isolate grew faster (36 mm/day) than the other 10 isolates (14.8 mm/day). There was a significant difference in sclerotial size between the fast and the slow growing isolates, but there was no significant difference in the number of sclerotia produced after 3 weeks on PDA. Two of the slow growing isolates exhibited ascospore dimorphism, whereas the fast growing isolate did not. PCR amplification with the primer pair ITS5/ITS4 produced a fragment of 560 base pairs from the fast growing isolate and 1000 base pairs from all of the slow growing isolates. The ITS sequences of the fast growing isolate had 100% homology with S. sclerotiorum, whereas those of the slow growing isolates had 100% homology with S. trifoliorum. Isolates of both species were pathogenic on fenugreek seedlings in the greenhouse assay and there was no significant difference in the percentage of dead plants two weeks after inoculation between the two species.