Association of Verticillium chlamydosporium and Paecilomyces lilacinus with Root-knot Nematode Infested Soil

Population densities of Meloidogyne incognita and the nematophagous fungi, Paecilomyces lilacinus and Verticillium chlamydosporium, were determined in 20 northern California tomato fields over two growing seasons. Paecilomyces lilacinus was isolated from three fields, V. chlamydosporium was isolated from one field, and both fungi were isolated from 12 fields. Verticillium chlamydosporium numbers were positively correlated with numbers of M. incognita and P. lilacinus. Paecilomyces lilacinus numbers were positively correlated with V. chlamydosporium numbers, but they did not correlate with M. incognita numbers. The correlation coefficients were low (R < 0.5) but significant (P < 0.05). All P. lilacinus and V. chlamydosporium field isolates parasitized M. incognita eggs in vitro. In a greenhouse study, numbers of V. chlamydosporium and P. lilacinus increased more in soils with M. incognita-infected tomato plants than in soil with uninfected tomato plants. After 10 weeks, the Pf/ Pi of second-stage juveniles in soils infested with P. lilacinus, V. chlamydosporium, and M. incognita was 47.1 to 295.6. The results suggest V. chlamydosporium and P. lilacinus are not effectively suppressing populations of M. incognita in California tomato fields.     

Benomyl Tolerance of Ten Fungi Antagonistic to Plant-parasitic Nematodes

Ten strains of fungi were tested for tolerance to the fungicide benomyl. Verticillium chlamydosporium strain 2 did not grow in the presence of benomyl; Drechraeria coniospora strains 1 and 2 and Chaetomium sp. tolerated only 0.1 μg benomyl/ml medium; Acremonium bacillisporum, an unidentified fungus, and Phoma chrysanthemicola uniformly grew at 1 μg/ml, but some hyphae grew at higher benomyl concentrations; Fusarium sp. tolerated 475 μg/ml, but some hyphae grew on medium amended with 1,000 μg/ml; Verticillium lecanii and V. chlamydosporium strain 1 routinely tolerated 1,000 μg/ml. Fungi generally grew more slowly at higher than at lower benomyl concentrations. Strains with elevated tolerance to benomyl were selected from Acremonium bacillisporum, Drechmeria coniospora, Fusarium sp., and an unidentified fungus. These strains retained the increased tolerance after repeated transfers on unamended medium.     

Effects of a Mutant Strain and a Wild Type Strain of Verticillium lecanii on Heterodera glycines Populations in the Greenhouse

A wild type strain ofVerticillium lecanii and a mutant strain with increased tolerance to the fungicide benomyl were evaluated in greenhouse experiments for effects on Heterodera glycines populations. Nematodes were applied at 300 eggs and juveniles per 4,550-cm³ pot (two soybean plants in 4,990 g loamy sand per pot) and at both 300 and 10,000 eggs and juveniles per 1,720-cm³ pot (one soybean plant in 2,060 g sand per pot). With 300 nematodes added per pot, both V. lecanii strains significantly reduced nematode populations in loamy sand (fungus applied at 0.02% dry weight per dry weight loamy sand) and sand (0.006% and 0.06% fungus application rates). The mutant strain applied at 0.002% to sand also significantly reduced cyst numbers. When 10,000 nematodes were added per pot, only the mutant strain at 0.06% significantly decreased population. Various media were tested for isolation of the fungus strains from prills, loamy sand, and sand, but the fungi were recovered from few of the greenhouse pots.     

Application of a Sex Pheromone,Pheromone Analogs,and Verticillium lecanii for Management of Heterodera glycines

A mutant strain of the fungus Verticillium lecanii and selected bioregulators of Heterodera glycines were evaluated for their potential to reduce population densities of the nematode on soybean under greenhouse conditions. The bioregulators tested were the H. glycines sex pheromone vanillic acid and the pheromone analogs syringic acid, isovanillic acid, ferulic acid, 4-hydroxy-3-methoxybenzonitrile, and methyl vanillate. A V. lecanii-vanillic acid combination and a V. lecanii-syringic acid combination were also applied as treatments. Syringic acid, 4-hydroxy-3-methoxybenzonitrile, V. lecanii, V. lecanii-vanillic acid, and V. lecanii-syringic acid significantly reduced nematode population densities in the greenhouse tests. Results with vanillic acid, isovanillic acid, and ferulic acid treatments were variable. Methyl vanillate did not significantly reduce cyst nematode population densities in the greenhouse tests.     

Viability of Heterodera glycines Exposed to Fungal Filtrates

Filtrates from nematode-parasitic fungi have been reported to be toxic to plant-parasitic nematodes. Our objective was to determine the effects of fungal filtrates on second-stage juveniles and eggs of Heterodera glycines. Eleven fungal species that were isolated from cysts extracted from a soybean field in Florida were tested on J2, and five species were tested on eggs in vitro. Each fungal species was grown in Czapek-Dox broth and malt extract broth. No toxic activity was observed for fungi grown in Czapek-Dox broth. Filtrates from Paecilomyces lilacinus, Stagonospora heteroderae, Neocosmospora vasinfecta, and Fusarium solani grown in malt extract broth were toxic to J2, whereas filtrates from Exophiala pisciphila, Fusarium oxysporum, Gliocladium catenulatum, Pyrenochaeta terrestris, Verticillium chlamydosporium, and sterile fungi 1 and 2 were not toxic to J2. Filtrates of P. lilacinus, S. heteroderae, and N. vasinfecta grown in malt extract broth reduced egg viability, whereas F. oxysporum and P. terrestris filtrates had no effect on egg viability.     

Hirsutella rhossiliensisand Verticillium chlamydosporium as Biocontrol Agents of the Root-knot Nematode Meloidogyne hapla on Lettuce

Hirsutella rhossiliensis and Verticillium chlamydosporium infected second-stage juveniles (J2) and eggs of Meloidogyne hapla, respectively, in petri dishes and in organic soil in pots planted to lettuce in the greenhouse. In vitro, H. rhossiliensis produced 78 to 124 spores/infected J2 of M. hapla. The number of J2 in roots of lettuce seedlings decreased exponentially with increasing numbers of vegetative colonies of H. rhossiliensis in the soil. At an infestation of 8 M. hapla eggs/cm³ soil, 1.9 colonies of H. rhossiliensis/cm³ soil were needed for a 50% decrease in J2 penetration of lettuce roots. Egg-mass colonization with V. chlamydosporium varied from 16% to 43% when soil was infested with 8 M. hapla eggs and treated with 5,000 or 10,000 chlamydospores of V. chlamydosporium/cm³ soil. This treatment resulted in fewer J2 entering roots of bioassay lettuce seedlings planted in the infested soils after harvesting the first lettuce plants 7 weeks after infestation with M. hapla. Hirsutella rhossiliensis (0 to 4.3 colonies/cm3 soil), V. chlamydosporium (500 to 10,000 chlamydospores/cm3 soil), or their combination, added to organic soils with 8 M. hapla eggs/cm³ soil, generally did not affect lettuce weight, root galling, or egg production of M. hapla. However, when lettuce was replanted in a mix of infested and uninfested soil (1:3 and 1:7, v:v), egg production was lower in soils with V. chlamydosporium than in soils without the fungus. Both fungi have potential to reduce the M. hapla population, but at densities below 8 eggs/cm³ soil.     

Pathogenicity of Fungi to Eggs of Heterodera glycines

Twenty-one isolates of 18 fungal species were tested on water agar for their pathogenicity to eggs of Heterodera glycines. An egg-parasitic index (EPI) for each of these fungi was recorded on a scale from 0 to 10, and hatch of nematode eggs was determined after exposure to the fungi on water agar for 3 weeks at 24 C. The EPI for Verticillium chlamydosporium was 7.6, and the fungus reduced hatch 74%. Pyrenochaeta terrestris and two sterile fungi also showed a high EPI and reduced hatch 42-73%. Arthrobotrys dactyloides, Fusarium oxysporum, Paecilomyces lilacinus, Stagonospora heteroderae, Neocosmospora vasinfecta, Fusarium solani, and Exophiala pisciphila were moderately pathogenic to eggs (EPI was 2.0-4.5, and hatch was reduced 21-56%). Beauveria bassiana, Hirsutella rhossiliensis, Hirsutella thompsonii, Dictyochaeta heteroderae, Dictyochaeta coffeae, Gliocladium catenulatum, and Cladosporium sp. showed little parasitism of nematode eggs but reduced hatch. A negative correlation was observed between hatch and fungal parasitism of eggs. Fusarium oxysporum, H. rhossiliensis, P. lilacinus, S. heteroderae, V. chlamydosporium, and sterile fungus 1 also were tested in soil in a greenhouse test. After 3 months, the nematode densities were lower in soil treated with H. rhossiliensis and V. chlamydosporium than in untreated soil. The nematode population densities were correlated negatively with the EPI, but not with the percentage of cysts colonized by the fungi. Plant weights and heights generally increased in the soil treated with the fungi.     

Fungal Parasitism of Heterodera glycines Eggs as Influenced by Egg Age and Pre-colonization of Cysts by Other Fungi

The objective of this study was to determine the effect of egg age and pre-colonization of cysts by a saprophytic or parasitic fungus on parasitism of Heterodera glycines eggs by other parasitic fungi. In agar and in soil tests, fungi generally parasitized more eggs in early developmental stages than eggs containing a juvenile. The effect of pre-colonization of cysts by a fungus on parasitism of eggs by other fungi depended on the fungi involved. In most cases, pre-colonization of cysts by an unidentified, saprophytic fungal isolate (A-1-24) did not affect parasitism of eggs in the cysts subsequently treated with other fungi. However, pre-colonization of cysts by A-1-24 reduced fungal parasitism of eggs in cysts subsequently treated with Cylindrocarpon destructans isolate 3. In agar tests, pre-colonization of cysts by Chaetomium cochliodes, a saprophytic or weakly parasitic fungus, reduced parasitism of eggs in cysts subsequently treated with Verticillium chlamydosporium Florida isolate, Fusarium oxysporum, Fusarium solani, ARF18, and another sterile fungus. However, in soil tests, pre-colonization of cysts by C. cochliodes had no effect on parasitism of eggs by subsequent fungal parasites. In another test, parasitism of eggs by V. chlamydosporium in cysts was not affected by pre-colonizing fungi C. destructans, F. oxysporum, and F. solani but was reduced by Mortierella sp., Pyrenochaeta terrestris, and C. cochliodes. Parasitism of eggs in cysts by ARF18 was reduced by pre-colonizing fungi C. destructans, F. oxysporum, F. solani, P. terrestris, and C. cochliodes but not Mortierella sp.     

Colonization of Soybean Cyst Nematode Females,Cysts, and Gelatinous Matrices by the Fungus Verticillium lecanii

Heterodera glycines was grown in monoxenic culture on soybean roots and then inoculated with the antagonistic fungus Verticillium lecanii. Use of root explant cultures allowed evaluation of the fungus-nematode interaction with the nematode attached to roots or removed from the host, and avoided contamination with other fungi. From 16 hours to 14 days following inoculation, female and cyst samples were examined with the light microscope, or prepared for either conventional or low-temperature scanning electron microscopy. Within 16 hours, hyphae had begun colonizing the gelatinous matrices (GM). The fungus proliferated in the GM of some specimens within a week, but was rarely seen in unhatched eggs. Fungus penetration holes in female and cyst walls were observed 3 days after inoculation; penetration through nematode orifices was not seen at that time. More cysts than females were colonized at the earliest sampling dates. Specimens associated with external hyphae exhibited variable internal colonization, ranging from no fungal penetration to extensive mycelial growth.     

Field Efficacy of Verticillium lecanii,Sex Pheromone,and Pheromone Analogs as Potential Management Agents for Soybean Cyst Nematode

A soybean cyst nematode sex pheromone (vanillic acid), chemical analogs of the pheromone, and the fungus Verticillium lecanii were applied in alginate prills (340 kg/ha) to microplots and small-scale field plots as potential management agents for Heterodera glycines on soybean. In 1991 microplot tests, treatment with V. lecanii, vanillic acid, syringic acid plus V. lecanii, or vanillic acid plus V. lecanii lowered midseason cyst numbers compared with the untreated susceptible cultivar control, autoclaved V. lecanii treatment, or aldicarb treatment, At-harvest cyst numbers were lowest with V. lecanii and with vanillic acid treatments. Aldicarb treatment reduced midseason cyst numbers in 1992. There were no differences among seed yields either year. In the field trials, numbers of cysts were reduced one or both years with aldicarb, ferulic acid, syringic acid, vanillic acid, or 4-hydroxy-3-methoxybenzonitfile treatments, or with a resistant cultivar, compared to an untreated susceptible cultivar. Highest yields were recorded after treatment with 4-hydroxy-3-methoxybenzonitrile (1991), methyl vanillate (1992), and aldicarb (1992). These studies indicate that some chemical analogs of vanillic acid have potential for use in soybean cyst nematode management schemes.     

Species of Root-knot Nematodes and Fungal Egg Parasites Recovered from Vegetables in Almería and Barcelona,Spain

Intensive vegetable production areas were surveyed in the provinces of Almería (35 sites) and Barcelona (22 sites), Spain, to determine the incidence and identity of Meloidogyne spp. and of fungal parasites of nematode eggs. Two species of Meloidogyne were found in Almería—M. javanica (63% of the samples) and M. incognita (31%). Three species were found in Barcelona, including M. incognita (50%), M. javanica (36%), and M. arenaria (14%). Solanaceous crops supported larger (P < 0.05) nematode numbers than cucurbit crops in Almería but not in Barcelona. Fungal parasites were found in 37% and 45% of the sites in Almería and Barcelona, respectively, but percent parasitism was never greater than 5%. Nine fungal species were isolated from single eggs of the nematode. The fungi included Verticillium chlamydosporium, V. catenulatum, Fusarium oxysporum, F. solani, Fusarium spp., Acremonium strictum, Gliocladium roseum, Cylindrocarpon spp., Engiodontium album, and Dactylella oviparasitica. Two sterile fungi and five unidentified fungi also were isolated from Meloidogyne spp. eggs.     

Multivariate Analysis of Selected Edaphic Factors and Their Relationship to Heterodera glycines Population Density

The influence of selected soil physical and chemical factors on population density of Heterodera glycines was investigated in 1988 and 1989 in two different locations of a soybean (Glycine max) field. Soil variables of a Norfolk loamy sand were measured after planting soybeans susceptible to H. glycines. Cyst and egg populations were determined after harvest. Nematode population density was found to be clustered. Up to 91% of the eggs were parasitized by a sterile fungus. Principal component analysis with orthogonal VARIMAX rotation grouped 12 variables into five uncorrelated factors in 1988 and three in 1989. In 1988, the factor "pH and Mg" was positively correlated (P < 0.001) with cyst and egg population density. Also, the factor "fine texture and Cu" was negatively correlated (P < 0.05) with egg population density. In 1989, the factor "pH, Mg and Cu" was positively correlated (P < 0.05) with levels of cysts and percentage of parasitized eggs, but not with total egg number. Across 2 years, factors containing soil pH and Mg were positively associated with cyst nematode population density. Copper appeared to be negatively associated with populations of H. glycines.     

Fungal Parasites of the Potato Cyst Nematode Globodera rostochiensis: Isolation and Reinfection

Fungal parasitism of eggs of the potato cyst nematode Globodera rostochiensis was < 1, 3, and 17% at three sites in Sweden. The fungi isolated most frequently from infected eggs were a Septocylindrium-like fungus ( 19 %), Exophiala spp. (17 %), and Cylindrocarpon spp. (13 %). Verticillium suchtasporium was isolated from infected eggs at a low frequency (4%). In laboratory experiments V. suchlasporium infected 93% of the eggs within cysts after 10 days on dilute corn meal agar. This species showed chitinase and protease activity. Infection of eggs by the Septocylindrium-like fungus was moderate, whereas Cylindrocarpon destructans and Cladosporium cladosporoides did not infect eggs. No chitinase activity was found in these fungi, but protease activity was recorded in all. Growth of the fungi in cysts did not influence the number of physiologically disordered eggs.     

Seasonal Biochemical Changes in Eggs of Heterodera glycines in Missouri

Changes in the carbohydrate (glucose, trehalose, and glycogen) and total protein contents of eggs retained within Heterodera glycines cysts were monitored monthly in a field microplot experiment conducted from March 1993 to March 1995. Treatments included two near-isogenic lines of soybean cv. Clark differing for date of maturity, and one corn hybrid. The soybean lines were planted in microplots infested with H. glycines at a high average initial population density (Pi) (23,810 eggs/100 cm³ soil), and the corn was planted in microplots infested at high (24,640) and low (5,485) Pi. Soil temperatures at 15 cm depth and rainfall were monitored. Carbohydrate contents varied in the same pattern, with the highest levels measured before planting (May) and after harvest (October) in both years. Neither Pi nor soybean isoline had an effect on any measured response, but the carbohydrate contents of eggs from corn and soybean microplots differed during the overwinter (October-May) periods (P < 0.0001). Trehalose accumulation was negatively correlated with soil temperature (r = -0.78 and r = -0.84, P = 0.0001, July through November 1993 and 1994, respectively), which reflects its role as a cryoprotectant. In contrast to the pattern for carbohydrates, total protein was lowest before planting and after harvest, and highest (>20 μg/1,000 eggs) June through October. Protein content was unaffected by plant cultivar or species. Protein and carbohydrate levels in H. glycines eggs showed seasonal changes that appeared to be primarily temperature-dependent.     

Production of extracellular enzymes in the entomopathogenic fungus Verticillium lecanii

This study investigates the mechanisms as well as strategies for purification and characterization of potential enzymes involved in pathogenesis of entomopathogenic fungi. The test strain of Verticillium lecanii that was screened, during the present investigation, proved to be an efficient producer of protein and polysaccharide degrading enzymes (amylase, protease, and lipase), hence indicating versatility in biochemical mechanisms. Halo zones produced colony growth of V. lecanii on agar confirmed activity of protease, amylase and lipase enzyme by the V. lecanii isolate. Enzymatic Index (EI) observed were: Protease – 2.195, Amylase- 2.196, Lipase- 2.147. Spectrophotometric analysis of enzymatic activity of V.lecanii at five different pH – 3, 5, 7, 9, 11 revealed that highest proteolytic activity of the V. lecanii isolate was reported at pH 7 and 9 whereas proteolytic activity was minimum at acidic pH 3. Maximum amylolytic activity of V. lecanii on the 7^th day of inoculation was at pH 3 i.e. in an acidic environment in contrast to neutral pH 7. Maximum lipolytic activity of V. lecanii was found at pH 7. Since enzyme production in entomopathogenic fungi is specific and forms an important criterion for successful development as well as improvement of mycoinsecticides, hence a significant conclusion from the present analysis is the degree of variation in secretion of enzymes in test strain of Verticillium lecanii.     

Fungal Parasites of the Cereal Cyst Nematode Heterodera avenae in Southern Sweden

Fungal parasites of the cereal cyst nematode Heterodera avenae were isolated from four sites in southern Sweden. In all, 15 different fungi were isolated from different stages of the nematode life cycle. Among the egg parasites, Verticillium chlamydosporium was common in young cysts on roots, whereas an unidentified species of Verticillium (Verticillium sp. 1) was the dominating species in cysts from soil, especially if the soll had been stored for 8-12 months. V. chlamydosporium was frequently isolated from eggs in cysts from soil, when analyzed shortly after sampling. Verticillium sp. 1 is distinct from V. chlamydosporium because it does not produce dichtyo-chlamydospores in the aerial mycelium and because it grows at 6 C where V. chlamydosporium fails to grow. Paecilomyces lilacinus, Microdochium bolleyi, Cylindrocarpon sp., and several nonsporulating fungi were also isolated from eggs in cysts from soil. Between 10 and 20% of the eggs in cysts collected in the field were infected with fungi. In a pot test between < 1 and 29%, with a mean of 13%, of females on roots became infected, always by Nematophthora gynophila. Resting spores of N. gynophila extracted directly from field soil, collected at the four sites, varied from 3 to 49 spores/gram of air dried soil.     

Dormancy of Heterodera glycines in Missouri

A 2-year study was conducted in field microplots to determine the relative importance of soybean phenology and soil temperature on induction of dormancy in Heterodera glycines in Missouri. Four near-isogenic soybean lines differing for maturity date were planted in microplots infested with a race 5 isolate of H. glycines. Soil temperature was monitored at a depth of 15 cm. Eggs of H. glycines, extracted from cysts collected monthly from each microplot, were used in hatching tests and bioassays to determine dormancy. Egg hatching and second-stage juvenile (J2) infectivity rates decreased sharply from their highest levels in midsummer (July-August) to a low level by October of each year and remained low (< 10% hatching and < 0.2 J2/cm root) until May or June of the following year. The patterns of numbers of females and eggs in the bioassays were similar. The decreases were not related to soil temperature and did not differ consistently among soybean isolines. The monophasic changes in all nematode responses with peak midsummer rates suggest that H. glycines produces one primary generation per year in central Missouri. Changes in hatching rates and the timing of minimum and maximum rates suggested that H. glycines eggs exhibit more than one type of dormancy.     

Association of the Plant-beneficial Fungus Verticillium lecanii with Soybean Roots and Rhizosphere

Colonization of soybean roots by the biocontrol fungus Verticillium lecanii was studied in vitro and in situ. For in vitro experiments, V. lecanii was applied to soybean root tip explant cultures. Four weeks after inoculation, the fungus grew externally on at least half of the roots (all treatments combined), colonizing 31% to 71% of root length (treatment means). However, when a potato dextrose agar plug was available as a nutrient source for the fungus, root tips inoculated soon after transfer were not colonized by V. lecanii unless Heterodera glycines was present. Scanning electron microscopy of colonized roots from in vitro cultures revealed a close fungus-root association, including fungal penetration of root cells in some specimens. In the greenhouse, soybeans in sandy soil and in loamy sand soil were treated with V. lecanii applied in alginate prills. The fungus was detected at greater depths from the sandy soil than from the loamy sand soil treatment, but fungus population numbers were small and variable in both soils. Root box studies coupled with image processing analysis of the spatial distribution of V. lecanii in sandy soil supported these findings. Verticillium lecanii was detected randomly in the rhizosphere and soil of root boxes, and was rarely extensively distributed. These in vitro and in situ experiments indicate that V. lecanii can grow in association with soybean roots but is a poor colonizer of soybean rhizosphere in the soil environment.     

Plantago lanceolata and Plantago rugelii Extracts are Toxic to Meloidogyne incognita but not to Certain Microbes

Extracts from the plants Plantago lanceolata and P. rugelii were evaluated for toxicity to the root-knot nematode Meloidogyne incognita, the beneficial microbes Enterobacter cloacae, Pseudomonas fluorescens and Trichoderma virens, and the plant-pathogenic fungi Fusarium oxysporum f. sp. gladioli, Phytophthora capsici, Pythium ultimum, and Rhizoctonia solani. Wild plants were collected, roots were excised from shoots, and the plant parts were dried and ground to a powder. One set of extracts (10% w/v) was prepared in water and another in methanol. Treatments included extract concentrations of 25%, 50%, 75% and 100%, and water controls. Meloidogyne incognita egg hatch was recorded after 7-day exposure to the treatments, and second-stage juvenile (J2) activity after 48 hours. All extracts were toxic to eggs and J2, with P. lanceolata shoot extract tending to have the most activity against M. incognita. Numbers of active J2 remained the same or decreased in a 24-hour water rinse following the 48-hour extract treatment, indicating that the extracts were lethal. When data from water- and methanol-extracted roots and shoots of both plant species were combined for analysis, J2 tended to be more sensitive than eggs to the toxic compounds at lower concentrations, while the higher concentrations (75% and 100%) were equally toxic to both life stages. The effective concentrations causing 50% reduction (EC₅₀) in egg hatch and in J2 viability were 44.4% and 43.7%, respectively. No extract was toxic to any of the bacteria or fungi in our assays.     

Meloidogyne incognita Survival in Soil Infested with Paecilomyces lilacinus and Verticillium chlamydosporium

Meloidogyne incognita-infected tomato seedlings were transplanted into sterilized soil or unsterilized soil collected from 20 California tomato fields to measure suppression caused by Paecilomyces lilacinus, Verticillium chlamydosporium, and other naturally occurring antagonists. Unsterilized soils Q, A, and H contained 35, 39, and 55% fewer M. incognita second-stage juveniles (J2) than did sterilized soil 1 month after infected tomato seedlings were transplanted to these soils and placed in a greenhouse. Three months after infected seedlings were transplanted to unsterilized or sterilized soil, unsterilized soils K, L, and Q had 97, 62, and 86% fewer J2 than the corresponding sterilized soils. Unsterilized soils of M. incognita-infected seedlings that were maintained 1 month in a greenhouse followed by 1 or 2 months of post-harvest incubation contained J2 numbers equal to, or greater than, numbers in the corresponding sterilized soil. The most suppressive of the unsterilized soils, K and Q, were not infested with V. chlamydosporium. Paecilomyces lilacinus and V. chlamydosporium increased in colony forming units in unsterilized soil of all bioassays, but they were not associated with lower numbers of J2.