The surface of the cuticle of adult Nippostrongylus brasiliensis has been studied by means of the freeze-fracture technique and by transmission electron microscopy. Some of the surface coat appears to have been shed from the surface of the cuticle of adults fixed in situ in the intestine of its host and from the surface of individuals removed from the intestine and freeze-fractured. Freeze-fracturing the cuticle of individuals removed from the host has shown that this surface coat varies in thickness from 30 to 90 nm. The epicuticle is about 20 nm thick and cleaves readily to expose E- and P-faces. The P-face of the epicuticle possesses a small number of particles, similar to intra-membranous particles, whilst the E-face possesses a few, widely scattered depressions. Despite the presence of these particles the epicuticle is not considered to be a true membrane. Freeze-fracturing the remainder of the cuticle has confirmed its structure as described by conventional transmission electron microscopy. Clusters of particles on the P-face of the outer epidermal (hypodermal) membrane and corresponding depressions on the E-face of the membrane are thought to be associated with points of attachment of the cuticle to the epidermis (hypodermis). No differences in appearance of the cuticle and its surface layers were observed in individuals taken from 7-, 10-, 13- and 15-day infections.

Toxocara canis infective larvae are known to produce abundant glycosylated molecules which may be found associated with the surface or secreted into their environment. Using a range of fluorescein-conjugated and gold-conjugated lectins, the localization of particular carbohydrates was defined on the surface of live parasites, and internally at the ultrastructural level. Surface exposure of N-acetyl galactosamine and N-acetyl glucosamine was deduced by binding of FITC-conjugated Helix pomatia (HPA) and wheat-germ agglutinins (WGA). These sugars appear to be associated with a densely staining surface coat as conventional immuno-electron microscopy procedures dissipate this coat and reveal no surface binding site for these lectins. However, by using cryo-immuno-electron microscopical (C-IEM) techniques, the surface coat is retained and can be shown to bind WGA. The fluorescent lectins also revealed strong WGA binding to the secretory and amphidial pores, while the buccal opening and the cuticular alae bound HPA. Corresponding results were obtained at the ultrastructural level. Thus, HPA bound to the electron-dense area of the cuticle, areas of local cuticular thickening such as the alae and buccal labia, as well as to the oesophageal lumen. WGA also bound to the thickened cuticle of the alae and the buccal opening, but showed no reaction to either the electron-dense layer of the cuticle or the oesophageal lumen. Unlike HPA, WGA did bind specifically to the secretory column contents and the electron-dense regions of the lips associated with the chemosensory amphids. The compartmentalization of the sugars N-acetyl galactosamine and N-acetyl glucosamine, their sources and routes of surface expression and the possible association with the TES glycoprotein antigens are discussed.

Clavibacter sp. (syn. Corynebacterium rathayi) adhered to both Anguina funesta (syn. Anguina agrostis) and Anguina tritici, but differences in the nature of adhesion were noted. Similar patterns of binding of the bacteria and of anti-wheat germ agglutinin antibody initially led us to believe that the mechanism of bacterial adhesion was related to the presence of wheat-germ agglutinin (WGA) on the outer cuticle of both species of nematodes and its complementary carbohydrate on the bacterial capsule. However, treatment of either species of nematode with sodium metaperiodate inhibited bacterial adhesion but not the binding of anti-WGA antibody. Bacterial adhesion, therefore, is not mediated by WGA on the nematodes' surface. Moreover, differences in patterns of bacterial adhesion to Anguina species, both before and after treatments with NaCl and detergents, suggest basic interspecific differences in the nature of adhesion. Electron microscopy confirmed the contribution of the nematodes' cuticular surface coat (SC) to the process of adhesion, but it is still not clear how the SC interacts with the bacterial capsule or which of its components are involved. While complete removal of the SC with periodate prevented bacterial adhesion, juveniles that naturally resisted bacterial adhesion did not lack a SC. One explanation could be that the SC of individuals, to which bacteria do not adhere naturally, lacks crucial components that cannot be defined by conventional EM.

Monoclonal antibody 2A6 binds to the surface of the Dictyocaulus viviparus L3 sheath and identifies a molecule which is highly antigenic in both infected and vaccinated cattle. Immunogold electron microscopy reveals that the antibody binds to a highly stable surface coat overlying the epicuticle of the L3 sheath. The binding to the cuticular surface is stage specific, being restricted to the surface of the L3 sheath, although the antigen can be detected on Western blots of parasite homogenates from the L1 stage. Immunofluorescence on live parasites demonstrates that the antibodies also bind to the L3 sheath surface of many other strongylid nematodes. The antigen detected by 2A6 differs in molecular weight between the species; in D. viviparus the antigen is poorly resolved between 29 and 40 kDa by SDS-PAGE whereas in nematodes from the family Trichostrongyloidea a single band or doublet of approximately 20 kDa is detected, suggesting that a related molecule is present in these species. Although infection and vaccination with D. viviparus elicits a strong immune response to the L3 sheath antigen this does not appear to be the case following infection with other strongylid nematodes.

Glycans at the surface of adult Schistosoma mansoni were investigated with gold-labelled lectins. The fragile complex of the glycans with the outer membranes could be preserved for electron microscopy by avoiding extensive pre-fixation with aldehydes and by introducing osmium-ferrocyanide as a membrane fixative. Male and female worms were entirely covered with glycans that intensely bound lectins from Erythrina cristagalli and Datura stramonium, suggesting that galactose(β1–4)N-acetylglucosamine residues occur in high numbers in the surface glycans. Similar staining was obtained with lectins from Triticum vulgaris, Glycine max and Ricinus communis agglutinin I, which react with N-acetylglucosamine or terminal galactose residues and bind non-selectively with high affinity to N-acetyllactosamine. Fucose, N-acetylgalactose and sialic acid were not detected with lectins and sialidase treatment. The tegument contained an abundance of glycans with the same lectin reactivities as the surface-expressed molecules, indicating that the worms synthesize and replenish their surface glycans and do not merely adsorb host substances. Glycomimesis is discussed as a mechanism of immune evasion in view of N-acetyllactosamine being a common and weakly immunogenic component in glycans of vertebrate hosts. S. mansoni might disguise themselves with the glycans against attack by immune effectors.

Secretions of plant-parasitic nematodes which are released into plant tissue may play critical roles in plant-nematode interactions. The identification and characterization of these molecules are of fundamental importance and may help to facilitate the development of novel strategies to interfere with nematode infection of plants and thereby decrease nematode-induced damage to crops. An antibody-based approach was used to isolate molecules present on the nematode surface and in nematode secretions. Monoclonal antibodies (MAbs) were produced to secretions and to whole Heterodera avenue 2nd-stage juveniles; several of these MAbs recognized molecules present in nematode secretions produced in vitro. Three of these molecules have been partly characterized in H. avenae, Globodera rostochiensis, G. pallida and Meloidogyne incognita. A MAb reacting with the surfaces of these nematodes recognized antigens of different molecular weight in each of the species tested. This difference in antigenicity might be related to specific functions in these nematodes. Preliminary results show that this antibody also localized the antigen in root cells surrounding the feeding site induced by M. incognita in Arabidopsis thaliana.

The surface coat of the infective larvae of the parasitic nematode Trichinella spiralis was characterized with respect to its biophysical properties, morphology and composition. Labelling of larvae with the fluorescent surface probe PKH26 was lost after activation (by incubation in mammalian medium containing trypsin and bile), or following pronase treatment. Electron microscopical examination revealed that pronase treatment resulted in the loss of an amorphous surface layer only, further demonstrating the specificity of PKH26 for the larval surface coat. Surface coat shedding was inhibited by sodium azide and carbonyl cyanide, or by incubation of larvae at 4°C, suggesting the shedding process required metabolic energy. Pre-labelled, unactivated larvae demonstrated continuous slow surface coat shedding and could be re-labelled with PKH26, indicating that the shed coat is replaced in these parasites. However, pre-labelled larvae which were activated failed to re-label with the probe, suggesting that activation provides an irreversible trigger for surface changes. PKH26, therefore, is a useful marker for larval activation. Examination of the shed coat material by scanning electron microscopy revealed 2 types of morphologies; one comprising thin multilaminate sheets and the other of amorphous material with ridges producing a fingerprint-like motif. Western- and lectin-blotting of the shed coat material demonstrated 2 prominent entities; a 90 kDa glycoprotein, which bound Datura stramonium agglutinin and was resistant to N- and O-glycanase treatment and a 47–60 kDa set of protein(s). Analysis of the surface lipids by electrospray mass spectometry revealed the presence of lysophosphatidic acid (lysoPA, C14[ratio ]2) and an unidentifiable component of 339·4 Da. These two lipids constituted 36·9% and 36% by mass of surface coat lipids respectively. The presence of lysoPA was confirmed by thin layer chromatography, which also detected phosphatidic acid (PA). The polar lipids detected in solvent rinses of intact parasites by electrospray mass spectrometry were PI (C48[ratio ]4), PE (C40[ratio ]4 and C38[ratio ]4), PS (C40[ratio ]4), lysoPC (C20[ratio ]2 and C18[ratio ]2) and lysoPA (C14[ratio ]2). These observations are discussed with respect to the role of the surface coat and its shedding in the T. spiralis host–parasite relationship.

Nematode surface coat (SC) proteins and excreted–secreted products (E–S) are likely to play important roles in the host–parasite interaction and considerable similarities can be found in SC proteins and E–S products from certain plant and animal parasitic nematodes. Monoclonal antibodies raised to E–S products of plant-parasitic nematodes were shown to cross-react with E–S products and the surface coats of the animal parasites Trichinella spiralis and Haemonchus contortus. Most of the antibodies recognized carbohydrate epitopes but the activity of 2 MAbs (IACR–CCNj.2a.15 and IACR-Misec.8D.3) which recognized proteic epitopes in these nematodes were further characterized. Antibody 2a.15 recognized the SC and oral exudate of Meloidogyne incognita, T. spiralis and H. contortus. This antigen was also immunolocalized in the lining of the oesophagus and gut and in the exudate present during ecdysis of H. contortus L3. Antibody 8D.3 reacted with the SC of these nematodes on cryosections but on live nematodes the immunofluorescence was very patchy and was shed from the nematode SC.

Characterization of surface coat (SC) proteins including carbohydrate-binding proteins and glycoproteins of the plant-parasitic nematode Meloidogyne javanica 2nd-stage juvenile (J2) is reported. Extraction of surface proteins with sodiumdodecyl sulfate (SDS) and separation by denaturing polyacrylamide gel electrophoresis (SDS–PAGE) results with bandsat 6, 9, 14, 22, 26, 31, 46, 49, 58, 66, 80, 205 and 250 kDa. On Western blots, the neoglycoprotein, fucosylated-,mannosylated- and glucosylated-bovine serum albumin, reacted with the 14, 22, 26, 58 and 66 kDa bands. The lectins,Concanavalin A and wheat-germ agglutinin (WGA) labelled surface protein bands of 6, 9, 14, 31, 58 and 66 kDa; WGAalso labelled the 22 and 26 kDa bands. Biotin reagents were used to specifically trace surface proteins on live J2.SDS–PAGE of biotinylated J2 extracts revealed only 2 specific biotin-protein bands at 46 and 49 kDa. The labile andtransitory nature of Meloidogyne javanica SC was demonstrated by the dynamics of human red blood cells (HRBC)adherence to J2 of different ages. HRBC adherence was also used to demonstrate the SC recovery of detergent-treatedJ2, which was further exhibited in the SDS–PAGE profiles.