•Adult schistosome worms do not produce many viable eggs after being placed in culture.•Female worms do not produce eggs in the absence of males.•Lack of the host-derived cytokine transforming growth factor-beta (TGF-β) may be a reason for poor female worm fecundity.•TGF-β can complex with the host blood-borne protease inhibitor alpha 2-macroglobulin (A2M).•TGF-β complexed with host A2M may be the source of the cytokine needed by schistosomes to enable egg production.

The surfaces of plant and animal parasitic nematodes share certain lipids, which seem to be important in the infection process. The surfaces of 2 parasitic nematodes, Meloidogyne incognita and Haemonchus contortus , were activated by different pH buffers to allow the insertion of different fluorescent probes. The lipid analogue PKH26 and the surface charge indicator, cationized ferritin, were used as probes with these nematodes but labelled only the retaining 2nd-stage moulted cuticle of H. contortus 3rd-stage larvae (L 3 ). Shedding of the second moult of H. contortus L 3 was also visualized with PKH26 and cationized ferritin. The fluorescent anionic lipid probe 5- N -(octadecanoyl)-aminofluorescein (AF18) was inserted into the epicuticle layer of M. incognita 2nd-stage juveniles (J 2 ) and H. contortus L 3 , and also of the second moult of H. contortus L 3 . Incubation with tomato root diffusate caused modifications of the M. incognita surface allowing the insertion of AF18. Fluorescence with AF18 was significantly decreased after treating M. incognita J 2 with amiloride, a potent blocker of hydrogen and sodium (H + /Na + ) antiporter. No surface fluidity was observed in M. incognita J 2 and H. contortus L 3 pre-treated with alkaline buffer when the lipid analogue AF18 was used in fluorescence recovery after photobleaching experiments. The significance of these findings to host infection processes is discussed.

Trichinella spiralis larvae incubated with a rabbit antiserum raised against the larval surface coat bound murine macrophages to the parasite surface. Cell binding was not observed without the antisurface coat serum, or with incubation of larvae in normal rabbit serum, or with antibodies to keyhole limpet haemocyanin which identify a cryptic T. spiralis larval antigen. Cell adherence to the larval surface was lost by treatment of the cells with the lysosomotropic drug primaquine, implicating a receptor‐mediated mechanism. Cells adhering to the parasite surface internalized parasite surface coat material, which was subsequently concentrated into endosomes. Culture supernatants from these cells contained enhanced levels of IL‐12. Thus, the initial Th1 response to T. spiralis infection may be explained by these data.

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 degrees 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, C14:2) 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 (C48:4), PE (C40:4 and C38:4), PS (C40:4), lysoPC (C20:2 and C18:2) and lysoPA (C14:2). These observations are discussed with respect to the role of the surface coat and its shedding in the T. spiralis host-parasite relationship.

As is the case in many parasite infections, research into schistosomiasis has not yet yielded a vaccine and, although chemotherapy with praziquantel is very effective, the mechanism of action of this drug is unknown. John Kusel and colleagues here suggest that an understanding of basic biological phenomena, such as the role of Ca(2+) in skin penetration and the function of the adult excretory system, might lead to important breakthroughs. Other crucial questions are also addressed, with the hope of stimulating debate. They invite suggestions and correspondence from others working in related fields.

The lateral diffusion (D L ) properties of the fluorescent lipid probe 5- N (octadecanoyl) aminofluorescein (AF18) inserted into the surface of muscle-stage larvae of Trichinella spiralis were investigated by fluorescence recovery after photobleaching. AF18 was not free to diffuse laterally in dormant larvae, and this remained unchanged after larval activation in vitro with trypsin and bile. However, a significant increase in surface fluidity of the probe was demonstrated (%R = 74·5; D L = 11·5 × 10 −9 cm 2 /sec) when larvae invaded intestinal epithelial tissue following oral infection of mice. Membrane-permeant photoactivatable caged cyclic AMP was used to analyse the putative mechanism responsible for this increase in lateral diffusion in the parasite surface. Although incubation of larvae with 1–50 μ M caged cAMP had no effect on surface fluidity, incubation with 100 μ M caged cAMP induced a substantial increase in the lateral mobility of AF18 (%R = 64·3; D L = 8·3 × 10 −11 cm 2 /sec) immediately following photo-activation of the caged messenger. This induced fluidity, however, was transient and the larval surface reverted to immobility within 15 min. These observations constitute the first reported measurement of the fluid properties of the surface of intracellular parasites, the first demonstration of the parasite surface fluidity altering as a result of host cell invasion and the first indication of a mechanism underlying changes in surface fluidity in parasitic helminths.

The surface of parasitic nematodes has been well studied with respect to its structural and immunological properties, but little is known about its biophysical nature and the role this plays in the host-parasite relationship. In this article, Clare Roberts and Jay Modha highlight some biophysical features of nematode surfaces and discuss their recent findings regarding mechanisms controlling surface-associated biophysical phenomena observed in parasitic nematodes during infection or culture in medium simulating the mammalian host environment. The nematode surface is distinct from the plasma membrane, nevertheless some parallel features exist and are described.

Molecular MicrobiologyVolume 20, Issue 1 p. 233-238 Electrospray ionization mass spectrometric analysis of phospholipids of Escherichia coli G. Sweetman, G. Sweetman MRC Toxicology Unit, Hodgkin Building, University of Leicester, Leicester LE1 9HN, UK.Search for more papers by this authorM. Trinei, M. Trinei Laboratory of Theoretical Biology, Department of Microbiology and Immunology, School of Medicine, PO Box 138, University of Leicester, Leicester LE1 9HN, UK.Search for more papers by this authorJ. Modha, J. Modha Division of Biochemistry and Molecular Biology, The Davidson Building, University of Glasgow, Glasgow G12 8QQ, UK.Search for more papers by this authorJ. Kusel, J. Kusel Division of Biochemistry and Molecular Biology, The Davidson Building, University of Glasgow, Glasgow G12 8QQ, UK.Search for more papers by this authorP. Freestone, P. Freestone Laboratory of Theoretical Biology, Department of Microbiology and Immunology, School of Medicine, PO Box 138, University of Leicester, Leicester LE1 9HN, UK.Search for more papers by this authorI. Fishov, I. Fishov Department of Life Sciences, Ben Gurion University of the Negev, Beer-Sheva 84105, Israel.Search for more papers by this authorD. Joseleau-Petit, D. Joseleau-Petit Institut Jacques Monod, 2 place Jussieu, Paris, France.Search for more papers by this authorC. Redman, C. Redman Division of Biochemistry and Molecular Biology, The Davidson Building, University of Glasgow, Glasgow G12 8QQ, UK.Search for more papers by this authorP. Farmer, P. Farmer MRC Toxicology Unit, Hodgkin Building, University of Leicester, Leicester LE1 9HN, UK.Search for more papers by this authorV. Norris, Corresponding Author V. Norris Laboratory of Theoretical Biology, Department of Microbiology and Immunology, School of Medicine, PO Box 138, University of Leicester, Leicester LE1 9HN, UK. *E-mail vjn@leicester.ac.uk; Tel. (116) 2525094; Fax (116) 2525030.Search for more papers by this author G. Sweetman, G. Sweetman MRC Toxicology Unit, Hodgkin Building, University of Leicester, Leicester LE1 9HN, UK.Search for more papers by this authorM. Trinei, M. Trinei Laboratory of Theoretical Biology, Department of Microbiology and Immunology, School of Medicine, PO Box 138, University of Leicester, Leicester LE1 9HN, UK.Search for more papers by this authorJ. Modha, J. Modha Division of Biochemistry and Molecular Biology, The Davidson Building, University of Glasgow, Glasgow G12 8QQ, UK.Search for more papers by this authorJ. Kusel, J. Kusel Division of Biochemistry and Molecular Biology, The Davidson Building, University of Glasgow, Glasgow G12 8QQ, UK.Search for more papers by this authorP. Freestone, P. Freestone Laboratory of Theoretical Biology, Department of Microbiology and Immunology, School of Medicine, PO Box 138, University of Leicester, Leicester LE1 9HN, UK.Search for more papers by this authorI. Fishov, I. Fishov Department of Life Sciences, Ben Gurion University of the Negev, Beer-Sheva 84105, Israel.Search for more papers by this authorD. Joseleau-Petit, D. Joseleau-Petit Institut Jacques Monod, 2 place Jussieu, Paris, France.Search for more papers by this authorC. Redman, C. Redman Division of Biochemistry and Molecular Biology, The Davidson Building, University of Glasgow, Glasgow G12 8QQ, UK.Search for more papers by this authorP. Farmer, P. Farmer MRC Toxicology Unit, Hodgkin Building, University of Leicester, Leicester LE1 9HN, UK.Search for more papers by this authorV. Norris, Corresponding Author V. Norris Laboratory of Theoretical Biology, Department of Microbiology and Immunology, School of Medicine, PO Box 138, University of Leicester, Leicester LE1 9HN, UK. *E-mail vjn@leicester.ac.uk; Tel. (116) 2525094; Fax (116) 2525030.Search for more papers by this author First published: April 1996 https://doi.org/10.1111/j.1365-2958.1996.tb02504.xCitations: 28AboutPDF ToolsRequest permissionExport citationAdd to favoritesTrack citation ShareShare Give accessShare full text accessShare full-text accessPlease review our Terms and Conditions of Use and check box below to share full-text version of article.I have read and accept the Wiley Online Library Terms and Conditions of UseShareable LinkUse the link below to share a full-text version of this article with your friends and colleagues. Learn more.Copy URL Share a linkShare onFacebookTwitterLinked InRedditWechat No abstract is available for this article.Citing Literature Volume20, Issue1April 1996Pages 233-238 RelatedInformation

The anthelmintic drug praziquantel has proved useful in the treatment of schistosomiasis. The precise mechanism by which praziquantel kills the parasites has yet to be elucidated. Here, John Kusel and colleagues review the current theories on praziquantel action and suggest future avenues for research, which becomes urgent in the light of some reports of drug resistance.

The view of the schistosome host-parasitic relationship has changed in the past two decades. Previously, it was thought the parasite simply defended itself in the face of a hostile host environment. However, it is now realized that the host-parasite interaction is much more of a dynamic interplay, where the parasite is able to exploit host homeostatic mechanisms for survival, maturity and transmission. Here, Jay Modha, Clare Roberts and John Kusel discuss the recent identification of serine protease inhibitors (serpins) on the schistosome surface and suggest how their properties might be exploited by the parasite.

The involvement of second messengers in the control of activation-induced changes to the surface of Trichinella spiralis infective larvae was investigated using membrane-permeant photo-activatable 'caged' compounds to alter intracellular levels of inositol trisphosphate (IP3), calcium ions (Ca2+) and cyclic AMP (cAMP). Activation of larvae by incubation in culture medium containing trypsin and bile was followed by the loss of the surface coat labelled with the fluorescent PKH26 lipid probe and this correlated with the reciprocal acquisition of surface lipophilicity detected using the fluorescent lipid probe octadecanoyl aminofluorescein (AF18). Optimal surface coat shedding and AF18 insertion was also achieved following photolysis of caged mediators liberating IP3, Ca2+ or cAMP within the parasite. Chelation of Ca2+, however, abolished the effects of larval activation. Nevertheless, addition of cAMP (but not IP3) to Ca(2+)-depleted larvae overcame this inhibition and restored AF18 insertion to levels achieved by activated parasites. Therefore, the existence of a linear second messenger pathway involving the sequential release of IP3, Ca2+ and then cAMP is likely.

SUMMARYContrapsin, a serine protease inhibitor (serpin) present in mouse serum, was compared with that found in adultSchistosoma mansoniworm homogenates, which although immunologically identical to contrapsin in mouse serum, had a higher molecular weight in Western blotting. Immunolocalization studies demonstrated parasite-associated contrapsin on the surface and interstitial cells of adult male worms. After extraction of these parasites with Triton X-114, contrapsin was found in the aqueous phase of the detergent, suggesting it is unlikely to be an integral membrane protein. Treatment of adult worms with deoxycholate resulted in a change in the electrophoretic behaviour of worm-derived contrapsin. Parallel studies with trypsin suggested this was due to interaction of the serpin with a protease. Using porcine pancreatic trypsin as a model for a putative schistosome protease reacting with contrapsin, we have shown that trypsin, following complex formation with contrapsin, loses immunogenicity. Thus, when contrapsin–trypsin complexes were used as immunogen, the resulting antisera contained antibodies to contrapsin and contrapsin–trypsin complexes only, and none to native trypsin. Thus, epitopes characterizing native trypsin were presumably either masked following complex formation with contrapsin, or their processing and presentation to antigen presenting cells was suppressed, so that an antibody response was not mounted against them. These observations encourage speculation thatS. mansonimay be elaborating an immune evasion strategy whereby immunologically sensitive proteases are first complexed with host serpins, which would render them immunogenically inert, and then cleared from the circulation by the host's reticulo-endothelial system. In this way the immune system would be unable to ‘see’ sensitive parasite proteases sufficiently to mount a response against the parasite.

Human alpha-1-antitrypsin (alpha 1-AT) was incubated with an extract of Schistosoma mansoni cercariae or porcine pancreatic elastase and analysed by immunoelectrophoresis and Western blotting. The inhibitor was shown to form complexes with components in S. mansoni cercariae in the same way as elastase. The role of alpha 1-AT in S. mansoni infection is discussed.

A multi-subunit antigen (native M(r) > 200 kDa, reduced M(r) 97-100 kDa) has been identified in homogenates of Trichinella spiralis larvae using affinity-purified rabbit anti-keyhole limpet haemocyanin (KLH) antibodies and its cross-reactivity with KLH was confirmed by competition blotting. The antigen was not present at the larval surface but was exposed after treatment of the larvae with the detergent cetyltrimethyl ammonium bromide (CTAB) which removed the surface coat. This correlated with a significant decrease in insertion of the surface-restricted fluorescent lipid probe AF18, indicating that the surface coat must be lipidic in nature. Unlike KLH, the larval antigen blotted onto nitrocellulose was itself periodate insensitive. Periodate treatment of whole larvae, however, resulted in shedding of the surface, to which anti-KLH antibodies then bound intensely. Anti-KLH antibodies also recognized three (49, 55, 108 kDa) of the four most dominant antigens in excretory-secretory (ES) products of cultured larvae, whose excretion-secretion was increased with CTAB. The nature, location and function of the antigen is discussed.

SUMMARYThe efficacy of praziquantel‐treatment of murine Schistosoma mansoni‐infections can be enhanced by concurrent administration of rabbit anti‐sera with specificity for parasite antigens. Monospecific rabbit serum raised against S. mansoni worm alkaline phosphatase, that was reactive with the enzyme on the drug‐treated female surface, was found to significantly and preferentially increase the mortality of female worms by PZQ. Immunoglobulins purified from the anti‐alkaline phosphatase antiserum inhibited 54% of schistosome alkaline phosphatase enzymatic activity on the surface of praziquantel‐treated worms. We propose that synergistic antibody‐mediated death of drug‐damaged worms is a consequence of the inhibition of drug‐exposed alkaline phosphatase on the female worm surface by passively transferred antibody.