Spontaneous regression of bovine cutaneous leukosis

Biopsies of skin from a 2-year-old heifer with spontaneously regressing dermal leukosis were examined. The heifer was not infected with bovine leukemia virus and was negative for tumor-associated antigens of enzootic bovine lymphosarcoma. By hematological standards for bovine leukemia, the heifer was positive at about 60 days post-occurrence of the disease (POD). At 53 days POD, lymphoblastic neoplastic cells in the dermis reacted with anti-T lymphocyte monoclonal antibody by the avidin biotin peroxidase complex method. The cells had intracytoplasmic clustered dense bodies under electron microscopy. From 53 to 83 days POD, figures of the transepithelial elimination (TE) against neoplastic cells and perivascular infiltration of small lymphocytes were in the dermis. Small lymphocytes reacted with anti-T lymphocyte monoclonal antibody. At necropsy there were no neoplastic lesions; there were flat lymph node-like tissues in the subcutis. Many germinal centers were seen in the lymphatic organs. Blood lymphocytes at 46 days POD were stimulated by phytohemagglutinin-P and concanavalin A. Sera, taken until 75 days POD and at necropsy, showed an inhibitory effect on mitogen-induced blastogenesis of normal bovine lymphocytes. These results suggested the existence of a spontaneous regressive mechanism against neoplastic lesions by TE and tumor immunity.     

‘Plantibodies’: a flexible approach to design resistance against pathogens

Engineering resistance against various diseases and pests is hampered by the lack of suitable genes. To overcome this problem we started a research program aimed at obtaining resistance by transfecting plants with genes encoding monoclonal antibodies against pathogen specific proteins. The idea is that monoclonal antibodies will inhibit the biological activity of molecules that are essential for the pathogenesis. Potato cyst nematodes are chosen as a model and it is thought that monoclonal antibodies are able to block the function of the saliva proteins of this parasite. These proteins are, among others, responsible for the induction of multinucleate transfer cells upon which the nematode feeds. It is well documented that the ability of antibodies to bind molecules is sufficient to inactivate the function of an antigen and in view of the potential of animals to synthesize antibodies to almost any molecular structure, this strategy should be feasible for a wide range of diseases and pests.Antibodies have several desirable features with regard to protein engineering. The antibody (IgG) is a Y-shaped molecule, in which the domains forming the tips of the arms bind to antigen and those forming the stem are responsible for triggering effector functions (Fc fragments) that eliminate the antigen from the animal. Domains carrying the antigen-binding loops (Fv and Fab fragments) can be used separately from the Fc fragments without loss of affinity. The antigen-binding domains can also be endowed with new properties by fusing them to toxins or enzymes. Antibody engineering is also facilitated by the Polymerase Chain Reaction (PCR). A systematic comparison of the nucleotide sequence of more than 100 antibodies revealed that not only the 3′-ends, but also the 5′-ends of the antibody genes are relatively conserved. We were able to design a small set of primers with restriction sites for forced cloning, which allowed the amplification of genes encoding antibodies specific for the saliva proteins ofGlobodera rostochiensis. Complete heavy and light chain genes as well as single chain Fv fragments (scFv), in which the variable parts of the light (V_L) and heavy chain (V_H) are linked by a peptide, will be transferred to potato plants. A major challenge will be to establish a correct expression of the antibody genes with regard to three dimensional folding, assembly and intracellular location.     

Effects of Latent Infection of Stock Plants and Abundance of Thrips on the Occurrence of <Emphasis Type="Italic">Tomato spotted wilt virus </Emphasis>in Chrysanthemum Fields

DAS-ELISA proved to be reliable enough to detect a latent infection by Tomato spotted wilt virus (TSWV) in asymptomatic stock plants of chrysanthemum. A high density of Frankliniella occidentalis, the predominant vector, in the presence of latently infected stock plants resulted in a high incidence of disease in the chrysanthemum production field. The incidence of disease was low when the vector thrips were not abundant in spite of the presence of latently infected stock plants. These results suggest that an infestation of the vector thrips causes severe secondary spread of TSWV originating from latently infected stock plants in chrysanthemum production fields. Received 27 July 2001/ Accepted in revised form 27 November 2001     

Ultrastructural Localization of Hydrogen Peroxide in Host Leaves Treated with AK-toxin I Produced by Alternaria alternata Japanese Pear Pathotype

The rapid generation of reactive oxygen species (ROS), called the oxidative burst, is one of the earliest host responses to pathogen infection or elicitor treatments. Therefore, we looked for the induction of ROS generation in Japanese pear leaves by the host-specific toxin, AK-toxin I using a cytochemical method for detecting H₂O₂. A small amount of non-specific generation of H₂O₂ was found in the cell walls in toxin- and water-treated susceptible and resistant leaves. Thus, the generation of H₂O₂ at cell walls appears to be caused by wounding stress during sampling. Specific generation of ROS, however, was found only in the membrane fragments and extended desmotubules characteristic of modified sites of the plasma membrane in the toxin-treated susceptible leaves. In addition, generation of H₂O₂ at plasma membranes was observed with higher frequency in toxin-treated susceptible leaves. This result indicates that the H₂O₂ generation was associated with damaged sites in the plasmalemma after toxin treatment and perhaps with the formation of membrane fragments from altered portions of the invaginated plasma membrane. Received 21 September 2001/ Accepted in revised form 25 October 2001