EE JONES¹, SP BUDGE¹, MP MCQUILKEN², RH WILLIAMS³ and JM WHIPPS¹

¹HRI, Wellesbourne, Warwick, CV35 9EF, UK; ²SAC, Auchincruive, KA6 5HW, Scotland, UK; ³IACR-Rothamsted, Harpenden, Hertfordshire, AL5 2JQ, UK

Background and objectives
Coniothyrium minitans is a sclerotial mycoparasite of Sclerotinia sclerotiorum and has been well-documented as a biocontrol agent of this pathogen in field and glasshouse trials on a range of crops. For commercial use, isolates need to be easy and cost-effective to produce and must work reproducibly when applied over defmed environmental conditions for each target crop. This paper describes work on isolate type and physiology, inoculum production and ecology of C. minitans, aimed at achieving this commercial goal.

Results and conclusions
All four C. minitans isolates tested exhibited similar temperature, pH and culture medium requirements for hyphal extension, conidia production and germination. Light had no effect on conidial germination or hyphal extension, but significantly increased pycnidial production, suggesting that this maybe a technique to improve the pycnidial formation by this fungus. However, of the isolates tested, two showed sparse pycnidial production on all media, and under all environmental conditions, illustrating the need to choose isolates that sporulate well for future development.

Ten solid-substrates were tested for their ability to support the growth of C. minitans and these inocula were assessed for ability to infect and inhibit carpogenic germination of sclerotia of S. sclerotiorum using simple glasshouse and field pot bioassays. Coniothyrium minitans grew on all solid substrates tested, with little difference between inocula in ability to infect sclerotia or reduce apothecial production. Soil incorporations of five inocula (barley-rye-sunflower, maizemeal-perlite, peatbran, rice and wheat) decreased sclerotinia disease in lettuce in a large-scale glasshouse trial, with only small differences between the inocula tested. This suggests any of these substrate could be suitable to produce commercial inocula of C. minitans.

Both potato dextrose broth and molasses-yeast liquid medium supported mycelial growth and conidial production of C. minitans in static culture. In shaken cultures conidial production in potato dextrose broth reached 10⁶ conidia per ml. Air-dried biomass-kaolin dust preparations derived from static liquid cultures reduced sclerotial viability and apothecial production in glasshouse and field pot bioassays, and decreased sclerotinia disease in glasshouse lettuce trials, to a similar extent to the standard maizemeal-perlite inoculum of C. minitans. This suggests that liquid formulations maybe an approach for future inoculum production of this fungus.

Conidia of C. minitans were applied to S. sclerotiorum-infected sunflower seeds and sclerotia by polymer film coating to mimic treatment of S. sclerotiorum infested seed lots. Seed germination was increased, and recovery of S. sclerotiorum from seed was decreased in an agar plate test, although in seedling tests C. minitans failed to increase survival of seedlings. However, following burial in soil, sclerotia coated with C. minitans conidia were killed indicating a use for C. minitans to clean up sclerotia infested seed batches. Coniothyrium minitans did not spread by myceliogenic growth in soil, but was transmitted from infected to uninfected sclerotia by mites (Acarus siro) and collembolans (Folsomia candida). Both A. siro and F. candida transmitted C. minitans at least 55 mm in soil at water potentials ranging from saturation to -3.6 MPa. Combining soil mesofauna with antagonists may be a way to enhance biocontrol in general. Water splash from overhead irrigators was also found to disperse C. minitans to at least 2 m. This information has identified more effective strategies for biocontrol of S. sclerotiorum disease using C. minitans and should aid commercialisation.