The fungal biocontrol agent, Trichoderma harzianum, was evaluated for its potential to control the root-knot nematode Meloidogyne javanica. In greenhouse experiments, root galling was reduced and top fresh weight increased in nematode-infected tomatoes following soil pretreatment with Trichoderma peat-bran preparations. The use of a proteinase Prb1-transformed line (P-2) that contains multiple copies of this gene improved biocontrol activity in the greenhouse experiments compared with the nontransformed wild-type strain (WT). All the Trichoderma strains showed the ability to colonize M. javanica-separated eggs and second-stage juveniles (J2) in sterile in vitro assays, whereas P-2 also penetrated the egg masses. This protease-transformed line presented the same nematicidal and overall proteolytic activity as the WT in in vitro tests in which concentrated soil extracts from Trichoderma-treated soils immobilized the infective J2. However, the J2 immobilization and proteolytic activities of both P-2 and the WT were higher than those obtained with strain T-203. Characterization of the activity of all Trichoderma strains soil extracts on J2 showed that it was heat resistant and restricted to the low-molecular-weight fraction (less than 3 kDa). It is suggested that improved proteolytic activity of the antagonist may be important for the biological control of the nematodes.

New nematode control measures are required due to the likely removal of effective nematicides from the market because of increasing concerns of possible effects on human health and the environment. Biological control, organic and inorganic soil amendments, naturally occurring nematicides, induced resistance, interruption of host recognition and transgenic plants will be a part of integrated management of plant-parasitic nematodes in the near future. The paper reports work carried out in Israel on these topics.

Trichoderma (T. asperellum-203, 44 and GH11; T. atroviride-IMI 206040 and T. harzianum-248) parasitism on Meloidogyne javanica life stages was examined in vitro. Conidium attachment and parasitism differed beween the fungi. Egg masses, their derived eggs and second-stage juveniles (J2) were parasitized by Trichoderma asperellum-203, 44, and T. atroviride following conidium attachment. Trichoderma asperellum-GH11 attached to the nematodes but exhibited reduced penetration, whereas growth of T. harzianum-248 attached to egg masses was inhibited. Only a few conidia of the different fungi were attached to eggs and J2s without gelatinous matrix; the eggs were penetrated and parasitized by few hyphae, while J2s were rarely parasitized by the fungi. The gelatinous matrix specifically induced J2 immobilization by T. asperellum-203, 44 and T. atroviride metabolites that immobilized the J2s. A constitutive-GFP-expressing T. asperellum-203 construct was used to visualize fungal penetration of the nematodes. Scanning electron microscopy revealed the formation of coiling and appressorium-like structures upon attachment and parasitism by T. asperellum-203 and T. atroviride. Gelatinous matrix agglutinated T. asperellum-203 and T. atroviride conidia, a process that was Ca 2+ -dependent. Conidium agglutination was inhibited by carbohydrates, including fucose, as was conidium attachment to the nematodes. All but T. harzianum could grow on the gelatinous matrix, which enhanced conidium germination. A biomimetic system based on gelatinousmatrix-coated nylon fibers demonstrated the role of the matrix in parasitism: T. asperellum-203 and T. atroviride conidia attached specifically to the gelatinous-matrix-coated fibers and parasitic growth patterns, such as coiling, branching and appressorialike structures, were induced in both fungi, similarly to those observed during nematode parasitism. All Trichoderma isolates exhibited nematode biocontrol activity in pot experiments with tomato plants. Parasitic interactions were demonstrated in planta: females and egg masses dissected from tomato roots grown in T. asperellum-203-treated soil were examined and found to be parasitized by the fungus. This study demonstrates biocontrol activities of

The surface coat of the 2nd-stage juveniles (J2) of plant-parasitic nematodes is considered to be involved in interactions with microorganisms in the soil and rhizosphere, as well as with the host plant. Characterization of surface antigens might be important in the development of new nematode control strategies. In this study, polyclonal and monoclonal antibodies raised against Meloidogyne javanica, M. incognita and other plant-parasitic nematodes were tested for their binding to the surface coat and secreted-excreted products of M. javanica. Some of the monoclonal and polyclonal antibodies raised against M. incognita showed cross-reactivity with the surface coat of M. javanica. Western blot analysis of M. javanica surface coat extracts revealed labelling of several polypeptides with a 48 kDa main band for the polyclonal antibody IACR-PC Mi 373, and a 55 kDa main band for PC Mj E2. Further characterization of the antigens recognized by the polyclonal antibody PC Mj E2, in planta, showed that they were present in the parasitic stages J2 and J3 and that the surface coat was shed during root penetration. The hypodermis of the infective juveniles was labelled by PC Mj E2 and the monoclonal antibody IACR-Misec 3F.4, suggesting that these surface antigens are produced in the hypodermis. Nematode behaviour was affected by all the antibodies that bound to the surface coat of the pre-parasitic J2, and we demonstrated that the movement pattern of the M. javanica J2 was affected by these antibodies. Continuous binding of the antibodies to the M. javanica surface inhibited the infection of Arabidopsis thaliana roots on agar plates.

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