Cryostat sections of juvenile and adult female stages of the soybean cyst-nematode, Heterodera glycines, were incubated with 4 different naphthylamide-linked peptide substrates to localize and characterize proteinase activity within the animal. Detected activity was restricted to the intestine and 2 distinct classes of proteinase were identified on the basis of substrate specificity and sensitivity to plant proteinase inhibitors. A cathepsin L-like cysteine proteinase activity capable of hydrolysing the synthetic substrates Z-Ala-Arg-Arg-MNA and Z-Phe-Arg-MNA but not Z-Arg-Arg-MNA or L-Arg-NA was inhibited by an engineered variant of a cysteine proteinase inhibitor from rice (Oc-IδD86). The cleavage of Z-Phe-Arg-MNA was sensitive to inhibition by a combination of Oc-IδD86 and cowpea trypsin inhibitor (CpTI). Degenerate oligonucleotide primers were used to amplify fragments of cysteine proteinase genes from 2 cyst-nematodes, H. glycines and Globodera pallida. Comparison of the H. glycines fragment with known genes established highest homology to cathepsin L-like genes. In contrast, the amplified G. pallida fragment displayed greatest homology to cathepsin B-like genes from Caenorhabditis elegans.

The invasion and establishment of Heterodera glycines has been studied for 0–120 h after addition to host roots using a synchronized infection system. A high degree of synchrony in development was noted, with juvenile nematodes reaching their feeding sites adjacent to the root endodermis and discharging their subventral pharyngeal glands by 24 h post-invasion (p.i.). The moult to the 3rd-stage juvenile occurred approximately 89 h after entry to the root. Using an enzyme-linked immunosorbent assay with monoclonal antibodies (MAbs) whose tissue specificities had already been defined by immuno-cytochemistry, it was found that MAbs recognizing intestinal lipid droplets and granules showed a major increase in reactivity 18–24 h p.i. before declining by 39 h p.i. In contrast, the reactivity of two MAbs recognizing adult cuticle increased significantly only after 89 h p.i., following the second moult. MAbs to the two subventral pharyngeal gland cells showed a variety of patterns of changing reactivity during the experiment. One increased within 24 h of invasion before declining whereas a second fell without any initial rise. Two further MAbs specific for the subventral glands initially declined but showed a secondary increase in reactivity after 72 h and could be detected in adult females. This pattern was also seen in a MAb specific for the dorsal pharyngeal gland cell. These changes are discussed in the context of events taking place following invasion, with particular reference to the initiation and maintenance of a feeding syncytium by the developing nematode.

Low doses of the acetylcholinesterase-inhibiting carbamate nematicides disrupt chemoreception in plant-parasitic nematodes. Fluorescein isothiocyanate (FITC)/dextran conjugates up to 12 kDa are taken up from the external medium by certain chemosensory neurons in Caenorhabditis elegans. Similar chemoreceptive neurons of the non-feeding infective stage of Heterodera glycines (soybean cyst nematode) fill with FITC and the nuclei of their cell bodies selectively stain with bisbenzimide. The widely used nematicide aldicarb disrupts the chemoreceptive response of H. glycines with 50% inhibition at very low concentrations (ca 1 pM), some 10−6-fold lower than required to affect locomotion. Similarly, the anthelmintic levamisole had this effect at 1 nM. Peptides selected as mimetics of aldicarb and levamisole also disrupt chemoreception in H. glycines and Globodera pallida at 10−3-fold or lower concentration than required to inhibit locomotion. We propose an uptake pathway for aldicarb, levamisole, peptide mimetics and other soluble molecules by retrograde transport along dendrites of chemoreceptive neurons to the cell bodies and synapses where they act. This may prove to be a general mechanism for the low-dose effects of some nematicides and anthelmintics.