VARIATION BETWEEN STRAINS OF THE NEMATOPHAGOUS FUNGUS, VERTICILLIUM CHLAMYDOSPORIUM GODDARD. II. FACTORS AFFECTING PARASITISM OF CYST NEMATODE EGGS

BY

F. IRVING and B. R. KERRY Rothamsted Experimental Station, Harpenden, Herts, AL5 2JQ, England

The infection of Heterodera avenae and H. schachtii eggs by six strains of Verticillium chlamydosporiumon water agar was studied. Dead eggs and those that were immature were most readily colonised by all strains of the fungus. Strains differed in their pathogenicity but all were parasitic and capable of colonising viable eggs including those containing second-stage juveniles. Infection rates were no greater when hyphal growth around the eggs was stimulated by increas- ing the nutrient status of the agar medium, or when the eggs were surface sterilised. Eggs were more susceptible to parasitism within cysts and females of H. schachtiibut not in those of H. avenae.In general, H. avenaeeggs were more susceptible to parasitism than those of H. schachtii but strains differed in their virulence. The most likely strains to be effective pathogens of H. avenaeeggs were I > II > VI whereas those for H. schachtiiwere V > I > VI. The method used was reliable in determining the relative pathogenicity of the six V. chlamydosporiumstrains in a number of experiments and could be used to screen large numbers of similar fungi for their potential as biological control agents. Keywords: biological control, egg parasitism, Heterodera avenae, Heterodera schachtii, pathogenicity, screening technique.

Few studies have been made to determine whether the fungi isolated from some root knot and cyst nematode females and eggs are parasitic or saprophytic colonisers or to determine the factors affecting colonisation. Immature eggs of Meloidogyne incognita Kofoid & White were markedly more susceptible to Dactylaria oviparasitica Stirling & Mankau than those that had completed their development and contained second-stage juveniles (Stirling & Mankau, 1979). Similarly, Acremonium strictum Gams and Fusarium oxysporum Schlecht readily infected immature eggs of Heterodera schachtii Schmidt but few mature eggs (Nigh et al., 1980). Cayrol et al. (1982) reported that Heterodera avenae Woll. eggs were not susceptible to parasitism by herticillium chlamydosporium Goddard after they had developed beyond the three-cell stage. However, infection of mature eggs has been observed in both root knot and cyst nematodes (Morgan-Jones et al., 1983; Kerry, 1975a). D. oviparasitica infected more eggs of M. incognita in vitro if the eggs were retained within the mucilage of the egg masses than if the eggs were dispersed on the fungal culture (Stirling & Mankau, 1979). The authors concluded that the fungus derived lit- tle nutrition from the mucilage but spread more rapidly and killed more eggs when they were aggregated. However, the amount of inoculum surrounding 475 eggs greatly affected the rate of parasitism of some strains of h chlamydosporium (Kerry et al., 1986) and if the fungus can proliferate in the mucilage then kills are likely to be increased. The range of temperatures favouring hyphal growth of D. oviparasitica also supported the greatest rates of parasitism and at 15° and 27°C eggs of M. incognita were destroyed in 30 and 8.8 days respectively (Stirling, 1979). Condi- tions that favour hyphal growth may not necessarily lead to greater parasitism. In an earlier study (Kerry et al., 1986) of variation in h chlamydosporium, strain II grew most rapidly at 25°C whereas most other strains had temperature optima of about 18°C. However, strain II was still the most effective parasite of H. avenae eggs on agar at 12 ° C, but grew more slowly than the others at this temperature. Sometimes, examination of the influence of temperature on the parasitism of eggs is made difficult by the effects on hatch; egg parasitic fungi do not parasitise active second-stage juveniles and conditions which support a rapid hatch reduce the chances of infection. Strains of h chlamydosporium differed in their ability to infect H. avenae eggs in soil (Kerry et al., 1984) and on agar (Kerry et al., 1986), but the methods used were unsuitable for screening the variation in pathogenicity of large numbers of fungi. A quantitative method for measuring such differences in parasitic fungi, including h chlamydosporium was developed using A. lum- bricoides (Chalupova & Lysek, 1979). However, their method required very large densities of eggs (c. 2 x 105/ml) which were infected by the fungi in water. In preliminary studies using few cyst nematode eggs, infection by h chlamydosporium could not be reliably obtained in aqueous suspensions, and tests were made using suspensions of H. schachtii and H. avenae eggs (c. 200/Petri dish) which were added to cultures of the fungus on distilled water agar. Such a technique had detected differences in pathogenicity between strains of D. oviparasitica (Stirling & Mankau, 1978) and A. strictum and F. oxysporum (Nigh et al., 1980) and would be useful for screening large numbers of egg parasitic fungi for the selection of virulent strains that may have poten- tial as biological control agents. We have continued our studies on the variation between six strains of h chlamydosporium and here report on experiments that examined some factors affecting their parasitism of cyst nematode eggs.

MATERIALS AND METHODS

Six strains of h chlamydosporium were isolated from eggs of H. avenae in December, 1979 and cultured on corn meal agar (CMA) at 18°C using the methods described by Kerry et al. (1986). Populations of H. avenae were main- tained on oats cv. Peniarth and H. schachtii on cabbages cv. Greyhound in pots in the glasshouse. Cysts were extracted from 100 ml aliquots of the nematode- infested soil in a fluidising column (Trudgill et al., 1972) as described by Kerry