Maurice J. Schoenbechler

In vitro histamine release from blood cellular elements of rabbits infected with Schistosoma mansoni.

Summary In vitro antigen-induced histamine release from blood platelets of rabbits infected with Schistosoma mansoni is described. Histamine release was detectable both in the presence and absence of passive cutaneous anaphylactic antibodies. Ion exchange chromatography, used to separate some of the antibodies in the rabbit antisera, revealed that fractions containing most of the IgG immunoglobulin had all the detectable flocculating activity. Electrophoretically faster immunoglobulins contained passive cutaneous anaphylactic activity but no detectable flocculating titer. A pure platelet suspension from sensitized rabbits did not release histamine when challenged with antigen. Histamine release did occur in the presence of whole washed blood when similarly challenged. A significant increase in histamine release was noted when normal blood platelets in a suspension of washed sensitized cells were added to antigen.

Demonstration of a homologous anaphylactic antibody in rabbits infected with Schistosoma mansoni.

Groups of rabbits were exposed to 1000, 5000, or 25,000 Schistosoma mansoni cercariae. Flocculating antibodies were detected by the use of somatic antigens from eggs and cercariae, and by metabolic antigens from adult worms. Flocculating antibodies appeared in all the animals by the third week and persisted throughout the experiment. Highest titers were obtained with the adult metabolic antigen. Nine of eleven rabbits developed an antibody which produced passive cutaneous anaphylaxis in normal rabbits challenged with cercarial antigen. This antibody appeared 7–12 weeks following infection. It remained elevated for 3–6 weeks, often in high titers, and then either could no longer be demonstrated, or was detected at very low titers. Reinfection with 5000 cercariae at 36 weeks elicited an abrupt return of anaphylactic antibodies in half of the animals. The anaphylactic antibody was inactivated by heating at 56 °C and by mercaptoethanol. No obvious correlation was observed between antibodies detected by slide flocculation tests and those detected by passive cutaneous anaphylaxis.

Trypanosoma rhodesiense: variant specificity of immunity induced by irradiated parasites.

Abstract Rats immunized with irradiated Trypanosoma rhodesiense resisted infection with the homologous strain. When similarly immunized rats were challenged with parasites obtained from rhesus monkeys infected with the same strain, resistance depended on when parasites were obtained from the donor monkeys. Immunized rats challenged with trypanosomes obtained from a monkey during the first peak of parasitemia were solidly immune; immunized rats challenged with trypanosomes obtained from monkeys after their initial peak of parasitemia all succumbed to the challenging infection. These observations indicate that parasites of a variant antigenic specificity arose during the course of the monkey infections. Neutralization tests performed on the various isolates from rats and monkeys using antiserum obtained from immunized rats confirmed that the immunity produced by irradiated trypanosomes was variant specific.

Cross absorption studies performed with Schistosoma mansoni and Trichinella spiralis antigens in sera from patients with trichinosis.

Abstract The apparent one-way cross reaction between Schistosoma mansoni antigens and sera from patients with trichinosis was studied by absorption technics. When the Trichinella antiserum was absorbed with Trichinella slide flocculation test antigen, the homologous antibody was removed but the antibody reactive against cercarial slide flocculation test antigen remained. The converse was also true in that absorption of the Trichinella antiserum with schistosome antigen removed antibody serologically reactive with the schistosome antigen but failed to remove antibody reactive with Trichinella antigen.

CHOLESTEROL-LECITHIN SLIDE (TSSF) AND CHARCOAL CARD (TSCC) FLOCCULATION TESTS USING AN ACID SOLUBLE FRACTION OF TRICHINELLA SPIRALIS LARVAE.

Two simple sensitive flocculation tests were developed employing acid soluble T. spiralis larvae antigen absorbed onto cholesterol-lecithin crystals. Washing of the emulsion greatly increased the sensitivity of the antigen in the slide flocculation test. The addition of charcoal to the antigen emulsion permitted its use in a simple card test using serum or minute amounts of plasma collected from finger puncture. Absorption of reactive sera with a single dose of washed, packed, and essentially dry antigen-cholesterol-lecithin complex resulted in a complete removal of serologically detectable antibody indicating that the tests were based upon a true antigen-antibody system. Sensitivity of both tests was excellent. In the slide flocculation test, 64 sera from 68 patients were reactive, 2 were weakly reactive and 2 were nonreactive. All 21 sera from trichinosis patients were reactive in the charcoal card test. The tests also appear to be relatively specific. Only 3 sera from a total of 96 normal persons and patients with other diseases were reactive in the slide flocculation test; 13 sera showed weak reactions. With the charcoal card test 3 reactions were obtained with sera from a total of 64 normal individuals and patients with other diseases. The tests are especially well suited to small laboratory and field conditions because they are simple to perform and utilize antigenic emulsions which may be kept for prolonged periods without apparent loss of reactivity. Slide flocculation tests are among the simplest procedures for the serodiagnosis of infections. These techniques have not yet found wide application in the field of parasitology primarily because of the difficulty in many instances of combining the parasitic antigens with the carrier (cholesterol, latex, or bentonite). Suessenguth and Kline (1944) found that an aqueous extract of Trichinella larvae coated onto cholesterol crystals could be used as antigen in a simple and rapid flocculation slide test for trichinosis. They reported that this test was more sensitive than the complement fixation test (Suessenguth et al., 1957). However, other investigators have presented evidence indicating that this test lacks the specificity of other serological tests for trichinosis (Bozicevich et al., 1951; Sadun and Norman, 1955a; Greene and Brazeale, 1951). Recently Anderson (1960) developed a slide Received for publication 26 April 1963. flocculation test for schistosomiasis using antigen prepared by coating a buffered saline extract of cercariae onto cholesterol-lecithin particles and subsequently washing this complex to remove the excess antigen. This method has been shown to be one of the most sensitive and specific serodiagnostic procedures for schistosomiasis (Anderson, 1960; Anderson and Naimark, 1960; Jachowski and Anderson, 1961). Following the development of the rapid plasma reagin card test (Portnoy et al., 1962) for syphilis, the schistosome slide flocculation te t antigen was adapted to the card procedure and evaluated under laboratory and field conditions (Sadun et al., 1963). The card test in schistosomiasis gave results comparable to those obtained with the standard slide flocculation procedure and permitted the test to be completed within a few minutes on plasma obtained by finger puncture. The purpose of the current study was to determine whether similar procedures would provide a slide flocculation test for trichinosis

Trypanosoma rhodesiense: Protection in mice by inoculations of homologous parasite products

Abstract Mice were immunized against Trypanosoma rhodesiense (Wellcome strain) with whole lyophilized trypanosomes, with antigens produced by disrupting lyophilized trypanosomes under pressure, and with excretions and secretions of the living parasites. The survival rate in groups of 40 mice inoculated with disrupted trypansomes and challenged with the homologous strain was 48% with a soluble fraction, and 70% with a particulate fraction of the parasites. There was 95% survival after challenge in a group immunized with lyophilized trypanosomes; none of the controls survived. Results were essentially the same whether or not an aluminum hydroxide adjuvant was used. In subsequent experiments, complete protection was obtained with either crude excretion-secretion (ES) antigens or the particulate fraction of the ES antigen, while 40% of the mice survived challenge after inoculations of ES supernatant fluid. Mice immunized with crude ES antigen failed to survive challenge with a heterologous strain, although their mean survival time was prolonged several days beyond that of the controls.

Immune Damage to Liposomes Containing Lipids from Schistosoma mansoni Worms

Summary A protein-free lipid extract (“F2”) consisting mainly of glycolipids, a small amount of phospholipids and an unidentified brown pigment, was obtained from Schistosoma mansoni adult worms. The F2 was tested for the presence of haptenic molecules by incorporation into liposomal model membranes containing trapped glucose. Sera from five infected monkeys were assayed for serological activity against the F2 in liposomes at different times following infection. Complement-dependent damage leading to liposomal glucose release was observed with sera from four out of five monkeys. Glucose release did not occur when the F2 was omitted from the liposomes nor when heat-inactivated complement was used. All of the activity was removed by adsorption of monkey serum with adult schistosomes. The IgM-containing fraction of serum accounted for all of the antibody activity. It was concluded that a complement-dependent immune response against lipids may be observed during the course of schistosomiasis in monkeys. The antibody activity can be detected by utilizing liposomal model membranes which contain schistosomal lipids.

A plasma card test for rapid serodiagnosis of schistosomiasis (SPC).

Summary and conclusions Use of a schistosomiasis card test as an indicator of antibody is described. This test is performed by using a device for obtaining plasma from 3 drops of finger blood in a rapid and simple manner, a stable antigen suspension containing charcoal, and a plastic coated card surface. Preliminary findings contained in this report suggest that the SPC test possesses adequate sensitivity and specificity. Because of the economy, rapidity and simplicity of this test, and the fact that it provides a permanent record for future reference, the procedure may be particularly useful as a screening procedure for field epidemiologic investigations.

Trichinella spiralis and Schistosoma mansoni: specificity of in vitro, leukocyte-mediated histamine release from rabbit platelets.

Abstract The specificity of the allergic release of histamine from rabbits infected with Trichinella spiralis or Schistosoma mansoni was examined using five different helminth antigens. Significant release could be demonstrated with homologous antigen in all 20 infected rabbits. Although several cross reactions in the infected animals were observed with non-Trichinella and Schistosoma antigens, the frequency of the cross reactions was considerably less than that observed in complement fixation or soluble antigen fluorescent antibody tests.

In vivo Passive Sensitization of Normal Rabbit Leukocytes with Sera Demonstrating Homocytotropic Antibody Activity

Normal rabbit leukocytes were passively sensitized, as determined by in vitro antigen-induced histamine release, by the i.v. administration of homologous sera demonstrating homocyto