Schistosoma mansoni, S. haematobium, and S. japonicum: identification of genus- and species-specific antigenic egg glycoproteins

Immunoreactive egg glycoproteins of Schistosoma mansoni, S. haematobium, and S. japonicum which are genus- and species-specific, or react with sera of patients infected with other parasites, have been identified. Egg proteins were labeled with Iodine-125, and the concanavalin A-binding glycoproteins were immunoprecipitated with sera of patients infected with one of four species of Schistosoma or Trichinella spiralis, Taenia solium, Echinococcus granulosus, Entamoeba histolytica, or Wuchereria bancrofti. These immunoprecipitates were analyzed by two-dimensional sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Despite the strikingly different patterns of glycoproteins of the African species, the antibody immune responses of patients infected with S. mansoni and S. haematobium were found to be so similar that differentiation could not be established. In contrast, sera of patients infected with S. japonicum, S. mekongi, or parasites not of the genus Schistosoma, immunoprecipitated fewer of the major S. mansoni or S. haematobium glycoproteins. Likewise, antibody immune responses of patients infected with the Oriental schistosomes (S. japonicum and S. mekongi) could not be differentiated. Only a few quantitative differences were noted between our S. mansoni egg glycoprotein extract and a standardized soluble egg antigen extract. This study provides an explanation for the extensive cross-reactivity observed in diagnostic assays which utilize various fractions of schistosomal egg extracts as the antigen.     

Schistosoma mansoni: circulating antigens and immune complexes in infected mice.

Circulating schistosome antigens (CSA) and circulating immune complexes (CIC) were investigated during the course of Schistosoma mansoni infection in mice. The radioimmunoprecipitation-polyethylene glycol (PEG) assay (RIPEGA) with [¹²⁵I]anti-S. mansoni antibodies or [¹²⁵I] anti-antigen “4” antibodies detected, respectively, total CSA and antigen “4” in serum and in 3% polyethylene glycol-precipitated CIC from infected mice. Complement fixation test and [¹²⁵I] C_1q-binding test revealed, respectively, an anticomplementary activity and the presence of C_1q-binding CIC. All these substances appeared in infected mice at approximately the same period, i.e., between the 40th- and the 55th- day postinfection. No correlation was observed between the detection of anticomplementary active substances and C_1q-binding CIC. In contrast, a close relationship was noticed between CSA and complement-activating material during the course of the infection. This suggests that substances with anticomplementary activity in serum from infected mice could be one or various CSA. A close correlation was also observed between C_1q-binding CIC and free or “complexed” antigen “4.” This observation supports well the possibility that antigen “4” is one of the major complexed circulating antigen present in schistosomiasis. The immunoglobulins G₁, G_2a, M, and A were also characterized in 3% PEG-precipitated CIC from infected mice during the period in which we detected C_1q-binding CIC. The roles played by specific S. mansoni CIC in either schistosomal nephropathy or protective mechanisms to a challenge infection in mice are discussed.     

Lower faecal egg excretion in chemically-induced diabetic mice infected with Schistosoma mansoni due to impaired egg maturation

The effect of streptozotocin-induced diabetes mellitus was studied in mice infected with Schistosoma mansoni. Faecal egg excretion was lower in diabetic mice but worm load and total amount of eggs in the intestine tissue were equal to the control group. Evaluation of an oogram showed a great number of immature dead eggs and a low number of mature eggs in diabetic mice. It was therefore concluded that faecal egg excretion was lower in diabetic mice due to impaired egg maturation.     

Nicarbazin in schistosome infections: I. Antibody formation in mice and hamsters infected with Schistosoma mansoni.

The immunoprecipitin response of mice infected with Schistosoma mansoni and treated with nicarbazin, an egg suppressive agent, is significantly lower than in untreated-infected mice. The precipitin response to cercarial extract is virtually abolished in treated mice indicating common antigenic determinants with eggs of the same species. Haemagglutinins to adult worms are also significantly diminished in treated mice. Finally, circumoval precipitins are absent in treated mice when the drug is given continuously to infected mice in order to prevent egg laying by the female adult parasite. The results suggest that a significant portion of the precipitating antibody produced in schistosome infections reactive with cercariae and adult worms, as well as eggs, is probably a secondary antibody response due to common antigenic determinants found in eggs.     

Specific tyrosine phosphorylation induced in Schistosoma mansoni miracidia by haemolymph from schistosome susceptible, but not resistant, Biomphalaria glabrata

Molecular interplay during snail-schistosome interactions is poorly understood and there is much to discover concerning the effect of snail host molecules on molecular processes in schistosomes. Using the Biomphalaria glabrata - Schistosoma mansoni host-parasite system, the effects of exposure to haemolymph, derived from schistosome-resistant and susceptible snail strains, on protein tyrosine phosphorylation in miracidia have been investigated. Western blotting revealed several tyrosine phosphorylated proteins in this larval stage. Exposure of miracidia to haemolymph from susceptible snails for 60 min resulted in a striking, 5-fold, increase in the tyrosine phosphorylation of a 56 kDa (p56) S. mansoni protein. In contrast, haemolymph from resistant snails had little effect on protein tyrosine phosphorylation levels in miracidia. Confocal microscopy revealed that tyrosine phosphorylation was predominantly associated with proteins present in the tegument. Finally, treatment of miracidia with the tyrosine kinase inhibitor genistein significantly impaired their development into primary sporocysts. The results open avenues for research that focus on the potential importance of phospho-p56 to the outcome of schistosome infection in snails, and the significance of protein tyrosine kinase-mediated signalling events to the transformation of S. mansoni larvae.     

Schistosoma mansoni: control of extramedullar eosinophil myelopoiesis in chronically infected mice by inflammatory macrophages

Extramedullar proliferation of eosinophil granulocytes can be induced in mice chronically infected with Schistosoma mansoni, by intraperitoneal implants of glass coverslips. Immature eosinophils are located in discrete foci on glass implants; they are not correlated with the eosinophil population of the peritoneal cavity, where only mature eosinophils can be observed. The same induction of eosinophil proliferation can be obtained in normal mice, by the transfer of macrophages elicited by glass implants in mice with chronic schistosomiasis. This induction could not be done with cells mobilized in normal mice, either after transplant into normal mice or into schistosome infected ones. Stimulated macrophages of mice with chronic schistosomiasis have a capacity to induce peripherical proliferation of eosinophil granulocytes. This capacity is independent of the quality of the intraperitoneal environment. It can be expressed after transferring macrophages elicited in schistosome infected mice into normal mice.     

Schistosoma mansoni heat shock protein 70 elicits an early humoral immune response in S. mansoni infected baboons

A study was undertaken to search for DNA recombinant Schistosoma mansoni proteins responsible for eliciting an antibody response from the host at a very early phase after infection. A S. mansoni adult worm cDNA expression library was screened using pooled sera from baboons with four weeks of infection. Based on their specific reactivity with the S. mansoni infected sera and no reactivity when tested against the pre-infection sera from the same baboons, four clones were selected for further studies. Sequence analysis revealed that they were homologous to the S. mansoni heat shock protein 70 (hsp70). The insert sizes of the four selected clones varied from 1150 to 2006 bp. The preliminary characterization for antibody reactivity against a panel of baboon sera showed that the longest clone was the most reactive, eight out of eight acute and three out of four chronic sera reacting positively to this clone. The shortest clone was the least reactive. Our results suggest that the S. mansoni hsp70 elicits an early and strong antibody response in baboons and that antibodies to this protein can be detected in chronically infected animals. Therefore S. mansoni hsp70 may be a valid target for immunodiagnosis. However further studies are needed to identify the portion of the hsp70 that best fits the requirements for a valuable diagnostic antigen.     

Schistosoma mansoni: reduced efficacy of chemotherapy in infected T-cell-deprived mice

The effect of host immunosuppression on the efficacy of schistosomicidal chemotherapy has been tested in T-cell-deprived CBA mice infected with Schistosoma mansoni. The drugs hycanthone, oxamniquine, and praziquantel were found to kill fewer adult S. mansoni worms in deprived mice than in comparably infected strain-, age-, and sex-matched, immunologically intact controls. Inconsistent results were obtained with niridazole, and amoscanate was as effective in deprived mice as in controls. The possibility that hycanthone, oxamniquine, praziquantel, and previously studied antimony act synergistically with immune effector mechanisms in killing adult schistosomes is discussed.     

Alterations of the intestinal innervation in mice infected with Schistosoma mansoni.

Schistosomiasis mansoni is a parasitic disease in which granulomas form around schistosome eggs in the liver and intestines. The purpose of this study was to determine the alterations in the intrinsic innervation of the distal ileum and proximal colon resulting from schistosomiasis. Using murine schistosomiasis mansoni, we examined light microscopic preparations stained with osmium-zinc iodide or the dihydronicotinamide adenine dinucleotide: nitro BT oxidoreductase (NADH) method. We also examined specific populations of peptidergic nerves (vasoactive intestinal polypeptide and substance P) using an avidin-biotin complex (ABC) immunohistochemical technique. We found that granulomas focally destroyed the enteric nerves. Occasionally nerves were found within granulomas, particularly at the periphery of the lesions. Nerve cell bodies close to granulomas had altered staining, which included increased staining for vasoactive intestinal polypeptide. The distribution of nerve injury varied between the 2 enteric segments studied. In the distal ileum, the principal injury was to the myenteric plexus; whereas, the submucous and mucosal plexuses were predominantly damaged in the proximal colon. The physiologic significance of this injury to the enteric nerves requires elucidation.     

Localization and identification of Schistosoma mansoni/KLH-crossreactive components in infected mice.

KLH (Keyhole limpet hemocyanin) is highly immunogenic, and crossreactive epitopes occur widely in nature. In schistosomiasis, infected hosts generate antibodies reactive with KLH. This is of diagnostic importance but we lack detailed information on the immunogen-carrying molecules and their distribution in the worm. We used anti-KLH antibodies to localize cross-reacting epitopes in the various developmental stages of the parasite in experimental Schistosoma mansoni infection. The staining results show KLH crossreactivity in the life stages of the parasite. By immunoblotting we show that KLH-crossreactive antigenic epitopes in the parasite eggs are carbohydrates, also recognized by antibodies against soluble schistosome egg antigens. The localizations in the larval stages and in adult worms suggest that crossreacting antigenic epitopes are secretory products.     

Ki-67 is expressed in multiplying forms of Schistosoma mansoni, but not in snail host tissues

Ki-67 is a protein expressed in the nucleus of several species during cell-division, being absent during the GO resting phase of the cellular cycle. During attempts to disclose mitosis in the so-called " amebocyte-producing organ " in Biomphalaria glabrata infected with Schistosoma mansoni, the parasite multiplying forms appeared strongly marked for Ki-67, while the snail tissues were completely negative. These data are worth registering to complement general data on Ki-67, and to help future studies on the relationship of the parasite and of its intermediate host.     

Schistosoma mansoni: peritoneal plasmacytogenesis and polypoid transformation of mesenteric milky spots in infected mice.

We have studied inflammatory reactions in the mesenteric tissue of mice infected with Schistosoma mansoni. Perivascular tissue contained diffuse infiltrates of macrophages, eosinophilic granulocytes and lymphocytes. Angiogenesis in the perivascular adipose tissue was associated with superficial plasmacytogenic foci. Polypoid structures were occasionally formed adjacent to inflammatory foci in the adipose tissue, organized around loops of capillaries, with terminal formation of a glomerular capillary network embedded in connective tissue, covered by plasmacytes. We conclude that these structures are specialized milky spots dedicated to active plasmacytogenesis and antibody secretion into the peritoneal cavity of schistosome-infected mice.     

Schistosoma mansoni: extramedullar eosinophil myelopoiesis induced by intraperitoneal glass implants in chronically infected mice

Intraperitoneal glass implants in mice with chronic Schistosoma mansoni infections induce intense local myeloid reactions involving essentially myeloid granulocytes. The same phenomenon is not observed in normal mice nor in mice with acute schistosomal infection. An association of myelopoiesis with differentiated macrophages mobilized on glass implants and with dense ameboid cells located inside myeloid foci was detected. A macrophage-dependent induction of this eosinophil reaction is postulated.     

Schistosoma mansoni: peripheral and tissue eosinophilia in infected rats.

Peripheral eosinophilia is induced in Sprague-Dawley rats following infection with cercariae of Schistosoma mansoni. Beginning 3 weeks after infection, peripheral eosinophil levels rise above the baseline range; they reach peak values during the fifth week. Following a decline from peak values, peripheral eosinophil levels remain elevated and are observed to fluctuate for the next 5 months. The magnitudes of both the initial peak response at 5 weeks and the subsequent chronic level of peripheral eosinophils are dependent upon dose of cercariae. The initial peak response phase of peripheral eosinophilia coincides in time with the period of adult worm elimination (Weeks 4–6) in the schistosome-infected rat. Histological examination of the liver at 5 weeks after infection reveals eosinophil-rich inflammatory reactions associated with both live and dead worms residing in the portal blood vessels. Around live worms the inflammatory cells are localized in a perivascular arrangement; around dead worms these cells are in the vascular lumen in contact with destroyed worms. The chronic phase of peripheral eosinophilia is associated, in part, with inflammatory reactions surrounding eggs deposited in the liver by the few worm pairs which survive more than 6 weeks and remain within the liver. Histological examination during this period reveals granulomatous lesions within the liver surrounding eggs and dead worms. The granulomas are predominately monocytic (lymphocytes, macrophages) at 11 and 16 weeks. The initial peak response phase of peripheral eosinophilia appears to be a marker for tissue-localized reactions of eosinophils with worms. There are relationships between inflammatory reactions and survival of adult worms.     

Schistosoma mansoni: higher free proline levels in the livers of infected mice

The concentration of L-hydroxyproline in the liver of ICR female mice increased rapidly during the 8th to 11th weeks of Schistosoma mansoni infection. Free L-proline concentration began to increase about the 7th week and reached its maximum at the 8th to 9th weeks of the infection, when the granulomatous response to the schistosome eggs in the liver was most prominent, as indicated by the increase in liver wet weight and its deoxyribonucleic acid concentration. A significant increment in the total activity of ornithine-delta-transaminase (EC 2.6.1.13) and the decrease in the specific activity of proline oxidase (EC 1.4.3.2) became detectable in the liver homogenate of infected mice on the 8th week. However, changes in these enzymatic activities were not parallel to that of the hepatic free L-proline content. Intraperitoneal administration of S. mansoni egg granulomas or 15,000g x 30 min supernatant fluid of their extracts into uninfected, normal mice significantly increased the hepatic free L-proline content without any appreciable effect on the enzymatic activities of proline oxidase and ornithine-delta-transaminase. These findings suggest that S. mansoni egg granulomas contain a factor(s) which may be responsible for the elevation of free L-proline content in the fibrotic liver caused by experimental schistosomiasis mansoni.     

Schistosoma mansoni: ingestion of dextrans, serum albumin, and IgG by schistosomula.

Schistosomula of Schistosoma mansoni develop the ability to ingest and digest red blood cells after the fourth day post-transformation. Here, we have used fluorescently-labeled dextrans and two plasma proteins, albumin and IgG, to test whether day-old schistosomula can ingest and process macromolecules prior to the time that they eat red cells. Worms ingested dextrans of molecular weights 4,000, 70,000 and 2 x 10(6) in a time- and concentration-dependent manner. The dextran remained in the cecal lumen for up to 2 days after feeding. Parasites ingested both fluorescein-conjugated bovine serum albumin and rabbit IgG, but neither of these proteins remained confined to the cecum over time. Instead, fluorescence redistributed to the acetabular glands within a few hours. Thin-layer chromatography indicated that schistosomula degraded fluorescein-conjugated albumin to fluorescein-conjugated peptides approximately 10-15 amino acids long. The volume of the cecum was estimated to be 2431 microns 3 and the surface area 299 microns 2. These results demonstrate that larval schistosomes can ingest both proteins and complex carbohydrates shortly after transformation, before they can ingest red cells. Further, the gut apparently releases proteases that cleave plasma proteins, but not saccharidases that cleave dextran.     

Schistosoma mansoni: IL-4 is necessary for concomitant immunity in mice.

To ask whether type-2 immune responses serve an essential role in concomitant immunity, that is the prevention of superinfection with Schistosoma mansoni, we compared resistance to a challenge infection in infected wild-type (WT) mice and in infected IL-4-/- mice, which are unable to mount Th2 responses during schistosomiasis. Although WT mice are protected from superinfection, resistance is abrogated in the absence of interleukin (IL)-4. We conclude that IL-4 or IL-4-dependent responses, or both, are necessary for resistance to S. mansoni superinfection in mice.     

Schistosoma mansoni egg glycoproteins and C-type lectins of host immune cells: molecular partners that shape immune responses

Schistosome eggs and egg-derived molecules are potent immunomodulatory agents. There is increasing evidence that the interplay between egg glycoproteins and host C-type lectins plays an important role in shaping immune responses during schistosomiasis. As most experiments in this field so far have been performed using complex protein/glycoprotein mixtures or synthetic model glycoconjugates, it is still largely unclear which individual moieties of schistosome eggs are immunologically active. In this review we will discuss molecular aspects of Schistosoma mansoni egg glycoproteins, their interactions with C-type lectins, and the relevance to schistosome egg immunobiology.