The immunological response of CBA mice to Trypanosoma musculi: Elimination of the parasite from the blood

Agglutinating antibodies against Trypanosoma musculi could not be demonstrated in sera from parasitaemic, immune or immunized mice.Immune, non-adherent cells from peritoneal exudates of mice which had recovered from infection, accelerated the elimination of parasites when adoptively transferred to infected mice in which the parasitaemia was stable (plateau phase) and the parasites had reached the adult (non-dividing) stage. This effect was not influenced by the simultaneous administration of immune adherent cells and/or immune serum and, although the mechanism by which the blood infection is e radicated has not been established, the action of the sensitized cells does not appear to be due to formation of antibody which has a direct, trypanocidal effect.     

Trypanosoma musculi: Passive hemagglutination technique to measure antibody in mice

A passive hemagglutination assay was developed to measure Trypanosoma musculi-specific antibody in mice. Indicator-erythrocyte donor mice received 550 rad ⁶⁰Co 24 hr before intraperitoneal injection of 3 × 10⁴T. musculi. T. musculi antigen-coated erythrocytes were obtained from these mice on Day 9 postinfection. T. musculi antigen-coated erythrocytes obtained in this manner were used as indicator erythrocytes in a passive hemagglutination procedure. Serum from hyperimmunized mice (three consecutive infections at 21-day intervals) gave titers as high as 1:1024. Titers of 1:256 and 1:512 were obtained from singly infected mice on Days 18 and 28 postinfection, respectively. In marked contrast, nude mice infected with T. musculi did not produce a detectable agglutinating antibody response. Erythrocytes obtained from either irradiated (550 rad ⁶⁰Co) uninfected mice, nonirradiated infected mice, or normal mice did not agglutinate when combined with any of the sera tested. These data suggest the usefulness of this passive hemagglutination assay for the measurement of antibody to T. musculi in the serum of infected mice.     

Plasmodium berghei: The spleen in sporozoite-induced immunity to mouse malaria

A/J mice were splenectomized (Splx) or sham-splenectomized (SSplx) prior to administration of a single injection of irradiated sporozoites. Following challenge 7 days after immunization, it was found that none of the splenectomized mice were protected whereas 50% of the sham-splenectomized and intact animals were resistant to challenge. In another series of experiments similar groups, along with mice splenectomized just prior to challenge, were injected with 1.5 × 10⁵ irradiated sporozoites over a 5 week period. This resulted in protection of (1) 60–100% of the animals splenectomized before immunization, and (2) 90–100% protection of the animals splenectomized prior to challenge, as well as the intact and sham-splenectomized mice. Serum levels of antisporozoite antibodies (CSP and SNA) increased during immunization of the intact animals. Only 15–20% of the animals splenectomized prior to immunization presented positive CSP reactions and little if any sporozoite neutralizing activity (SNA) was detected. Serum from intact animals immunized and found resistant to sporozoite challenge was used for passive transfer studies. Immune serum recipients were challenged with small numbers of sporozoites. Only one out of 18 recipients was protected against sporozoite challenge.     

Leishmania mexicana and Leishmania tropica major: Adoptive transfer of immunity in mice

The transfer of lymphocytes from NIH mice infected with Leishmania tropica major to nonimmune syngeneic mice, which had received 600 rad irradiation, markedly altered the course of subsequent Leishmania mexicana and L.t. major infections in these animals, when compared with infections in irradiated mice reconstituted with lymphocytes from uninfected animals. Some resistance to L. mexicana could be transferred with unfractionated lymphocytes, mixed nylon wool adherent, and nonadherent lymphocytes and to a lesser extent by nylon wool nonadherent cells alone but not by the adherent population. When the recipients of “immune” lymphocytes were infected with L.t. major the resulting lesions were either small and did not ulcerate, or grew rapidly, ulcerated, and healed. The course of L.t. major in the recipients depended on whether the donor of “immune” lymphocytes had a healed or a healing lesion. Immunity to L.t. major was largely associated with the nylon wool nonadherent population.     

Trypanosoma congolense: Genetic control of resistance to infection in mice

Primary isolates of Trypanosoma congolense show a range of virulence in NMRI mice. Stabiliates derived from an isolate (Dinderesso/ 80/CRTA/3) which showed moderate virulence in most NMRI mice (moderate parasitemia and survival) were used in inbred mice. C57B1/6 were resistant with low parasitemia and survival. Parasitemias were higher in males than females. BALB/c were the most sensitive of the strains tested and died with fulminating parasitemia. Inheritance of resistance, defined as low parasitemia, was studied using these two strains. Male F1 showed high parasitemia; the backcrosses of F1 to the resistant parent had a ratio of one susceptible to one resistant product; the product of F1 to susceptible parent were all susceptible; and the F2 crosses showed a ratio of three susceptible to one resistant product. The results obtained with female F1, backcrosses, and F2 mice showed similar segregation to that found using males, but the range of parasitemia was always 1–2 log₁₀ lower, except for the F1 backcrossed to BALB/c, where female and male parasitemia were undistinguishable. The segregation ratios were identical whether resistant females were crossed with sensitive males or vice-versa. The results obtained are compatible with resistance being a recessive trait controlled by a single autosomal gene (or gene cluster). In addition, sex-associated factors appear to confer higher resistance in females.     

Trypanosoma cruzi: Responses by cells from infected mice to alloantigens

In unidirectional mixed lymphocyte cultures containing (as responders, stimulators, or regulators) spleen cells from mice infected with Trypanosoma cruzi, alloantigen responses were less than in cultures containing normal spleen cells only. Depletion of plastic adherent cells from infected spleen cells (stimulators or regulators) reversed their inhibitory effect on normal spleen cells (responders); removal of adherent responder cells and/or B lymphocytes did not alter the low alloantigen responses of normal spleen cells (stimulated by infected spleen cells) or infected spleen cells (stimulated by normal spleen cells). Infected spleen cells were effective in regulating mixed lymphocyte cultures only when added at the initiation of the culture. Serum from infected mice suppressed mixed lymphocyte cultures containing responder spleen cells syngeneic to the serum donor if added up to 24 hr after initiation of cultures, whereas the “suppressor serum” had to be present at the initiation of cultures when responder cells were allogeneic to the serum donor. Cultures of infected spleen cells (whole or macrophage enriched) produced a factor which was suppressive when added to mixed lymphocyte cultures containing syngeneic responder cells at initiation. It is proposed that the serum suppressor substance regulates cell-mediated immune responses directly by suppressing the response-potential of cells and indirectly by triggering the release of a factor from adherent splenic cells which induces a hyporesponsive state in T lymphocytes.     

Trypanosoma cruzi: Early immune responses in infected mice

The influence of an active Trypanosoma (Schizotrypanum) cruzi infection in mice and the subsequent immune response to an administered unrelated, erythrocyte antigen was studied. When mice were infected with blood forms of the parasite, a depression of the humoral immune response to injected burro erythrocytes (BE), was observed. The suppression became evident at a time when the magnitude of parasitemia and tissue forms was increasing in the sensitized and infected mice. The immunosuppressive effect induced by the infection to BE was demonstrated by a diminished direct (19S) and indirect (7S) antibody-forming cell response. Additionally, a significant increase in phagocytic activity was observed in T. cruzi-infected mice using colloidal carbon uptake experiments. Probable mechanisms of suppression are discussed and related to accepted lymphocyte activities.     

Trypanosoma cruzi: Immunoperoxidase antibody test for serologic diagnosis

An indirect antiglobulin immunoperoxidase test was developed for the serological diagnosis of American Trypanosomiasis. Purified rabbit antihuman IgG was labeled with the enzyme and the conjugate so obtained was characterized according to its immune and enzymatic activities, with the help of such parameters as the authors have recently described. For a maximal sensitivity in the tests, high antibody levels and a high-labeling ratio were chosen, as well as dilutions of conjugate ensuring maximal reactivity. Positive tests were found in all 90 serum samples from patients with Chagas' disease and titers did not differ significantly from those observed in immunofluorescence tests done in parallel. The specificity of both tests was also similar, as indicated by the results found for serum samples from 60 patients with other diseases, parasitic or not, showing high levels of antibodies against other infective agents or autoantibodies, and in 15 normals.     

Leishmania mexicana and Leishmania tropica: Cross immunity in C57BL/6 mice

Leishmania mexicana and Leishmania tropica infection were comparatively studied in C57BL/6 mice. Infection with 10⁴ amastigotes of L. mexicana was followed by the appearance of a single lesion which ulcerated in 8 weeks and healed in 24 weeks. Mice infected with 10⁴ amastigotes of L. tropica developed less severe lesions which healed in 18 weeks. In both cases healing was accompanied by a delayed hypersensitivity response and an in vitro lymphocyte reactivity to leishmanial antigens. Mice recovered from a primary infection with L. mexicana or L. tropica were resistant to both homologous and heterologous challenge. In vitro and in vivo immunological tests indicated that L. mexicana and L. tropica share antigenic determinants which are involved in cell-mediated immune responses to these parasites.     

Babesia bigemina,Babesia argentina, and Anaplasma marginale: Coinfectious immunity in bovines

Forty-eight intact and eight splenectomized cattle were used to evaluate different systems of coinfectious immunization against Babesia bigemina, Babesia argentina, and Anaplasma marginale. Coinfectious immunity was induced by two methods: (1) blood of cattle acutely infected with B. bigemina, B. argentina and A. marginale was used as the source of inoculum and the post vaccination reactions were chemotherapeutically controlled with Imidocarb, Ganaseg, Gloxazone, and Liquamycin, and (2) by artificially inducing babesiosis with the blood of carrier cattle with chronic infections of B. bigemina and B. argentina without chemotherapy. The degree of resistance was determined by bloodborne and tick-borne challenges. Ticks were collected from cattle and identified as Boophilus microplus and Dermacentor nitens. Vaccinated cattle demonstrated a high degree of resistance to babesiosis and anaplasmosis; however, cattle without coinfectious immunity were treated chemotherapeutically to prevent death losses.     

Trypanosoma cruzi: Lactate dehydrogenase isoenzymes and infections in mice

Total plasma LDH isoenzyme (EC 1.1.1.27) levels increased significantly over the normal level in mice infected with strains of Trypanosoma cruzi from three different geographic locations, but some strain differences were observed. The most rapid increase was exhibited by the blood-induced Tulahuen strain, but this strain, unlike the House 510 or House 11, did not elicit an increase during the early period of infection. Overall increases in LDH-1 and LDH-2, heart isoenzymes, were most marked in vector-derived House 510 infections, but, as in the Tulahuen strain, a considerable increase was also observed in blood-induced infections. The House 510 strain also elicited significant increases in LDH-4; these were particularly high during the early period of the blood-induced infection. By contrast, the vector-derived Tulahuen strain elicited a higher increase in LDH-4 during the early period than the House 510 or House 11 strains. Comparable similarites and differences were also observed in regard to LDH-3, 5, and “X.” The most marked isoenzyme increases were those of “LDH-X” exhibited by the blood-induced House 510 and vector-derived Tulahuen strains.Parallel histopathologic studies of liver, heart, and skeletal muscle disclosed significant pathology in all the infections. Animals with blood-induced Tulahuen strain infections characteristically showed extensive necrosis with marked multiplication of parasites throughout the liver, but little or no evident damage to the heart and skeleal muscle. Animals infected with House 510 and House 11 strains exhibited minimal pathology in the liver but severe damage to the heart and skeletal muscle. Increases in LDH-4 and LDH-5, isoenzymes which represent both liver and skeletal muscle, in blood-induced Tulahuen infections were attributed largely to liver damage, but in the House 510 and House 11 infections were related more to skeletal muscle.     

Trypanosoma brucei: The peptidases of bloodstream trypanosomes

Peptidases (EC.3.4.11 and EC.3.4.13.9) from bloodstream Trypanosoma brucei were examined by starch gel electrophoresis and substrate specificities and relative activities of six peptidases (S,B,E,A,F,D,) determined. The substrate specificities corresponded closely to those of the peptidases of human blood cells and tissues although human peptidase C appeared not to have a T. brucei equivalent.     

Plasmodium berghei: Heat-treated sporozoite vaccination of mice

Repeated intravenous (IV) immunizing injections with 25,000 Plasmodium berghei heat-treated sporozoites gave an average protection of 13% in five experiments (0–53%). Four injections of 10⁵ sporozoites gave 50% protection, seven injections of 10⁵ gave 37% protection, and six injections of 10⁵ gave 11% protection. Viable spleen cells (1.2 × 10⁸) from twice challenged immune syngenic mice did not protect naïve mice against iv challenge. Six ip injections of the supernatant of Parr Bomb disintegrated sporozoites gave no protection against ip challenge, but 7 ip injections gave 40% protection. Centrifuged pellets from French Pressure Cell-disintegrated sporozoites gave almost no protection either iv, sc, or ip. The supernatant was disc-electrophoresed and compared to normal mosquito heads and salivary glands in order to select sporozoite bands for immunizing injections. Results were discussed with respect to uniformity of antigen batches.     

The role of the spleen in protective immunity against Angiostrongylus cantonensis in rats: Splenectomy and passive spleen cell transfers

Splenectonuzed rats infected with Angiostrongylus cantonensis had more and longer worms and lower anti-A. cantonensis antibody titres compared to intact and sham operated rats. Splenectomy, however, had no effect on the capacity of rats to mount a cell-mediated immune response, as assessed by the in vitro uptake of tritiated thymidine by peripheral blood lymphocytes following stimulation by PHA and adult worm antigen. Adoptive protection and antibody production against A. cantonensis could be transferred with immune spleen cells. This protection was strongly dose and time dependent. Protection was adoptively conferred with 3 × 10⁹ spleen cells transferred 7 days before infection.     

Mucosal and systemic immunity in mice after intranasal immunization with recombinant Lactococcus lactis expressing ORF6 of PRRSV

The purpose of the study was to construct mucosal vaccine of a recombinant Lactococcus lactis expressing PRRSV ORF6 gene and evaluate mucosal and systemic immune response against PRRSV in mice after intranasal immunization. The result show that the vaccine can stimulate mice to produce specific IgG in serum and remarkable special s-IgA in lung lavage fluid, at the same time, the contents of cytokines IL-2 and IFN-γ of the experimental group were significant higher than those of the control group (P < 0.01), however, the contents of cytokines IL-4 was not different to the all groups. In summary, the constructed mucosal vaccine can significantly induce mucosal immune, humoral immunity and cellular immunity involved Th1 type cytokines, which will lay a theoretical foundation on immune mechanism and new efficient vaccines for PRRSV.     

Trypanosoma equiperdum: Movement from the dermis

Twelve dogs were injected intradermally with 352,770 to 14,391,660 Trypanosoma equiperdum and afferent and efferent lymph, lymph nodes, and blood examined by mouse inoculation at minute, hourly, and daily intervals following inoculation. The log dosage of trypanosomes given each dog was closely related to their body weight (P < 0.01). Afferent lymph contained trypanosomes as soon as 5 and 27 min after inoculation. Lymph nodes on the side of injection became positive within 5 min of injection, while those on the contralateral side remained negative for at least 120 min after injection. Blood contained trypanosomes as soon as 5 min after injection, although the average time for all dogs, before trypanosomes were demonstrated in the blood, was 40 min postinjection. Efferent lymph did not contain organisms until 25–76 hr after inoculation. We consider this sequence to indicate that T. equiperdum can leave the dermis in afferent lymphatics, reach the local lymph node, invade the blood stream from this site, and only after a day or longer do they leave the node via the efferent lymphatics.     

Trypanosoma musculi and Trichinella spirails: Concomitant infections and selection for resistance genotypes in mice

Trypanosoma musculi infections were given to mice of different strains before, at the same time, and after an infection with 400 Trichinella spiralis. Examined parameters of the host response to T. spiralis were worm rejection, antifecundity responses, development of immunological memory, and muscle larvae burden. After dual infection, each mouse strain showed characteristic effects on resistance to T. spiralis. This was due to a dynamic interaction between the genes controlling rejection of T. spiralis and those influencing T. musculi growth. C3H mice develop high trypanosome parasitemias. This impairs worm expulsion and the development of memory to T. spiralis when Trypanosoma infections take place on the same day or 7 days before. The C57B1/6 mouse develops low parasitemias and T. musculi infections on the same day, or 7 days before T. spiralis, delaying worm rejection only slightly despite the overall weak capacity of B6 mice to expel worms. NFR-strain mice are strong responders to T. spiralis and also develop low parasitemias. Trypanosome infections on the same day, or after T. spiralis, produce a delay in worm rejection; the former is comparable to C3H mice. However, NFR mice alone showed enhanced rejection of worm when T. musculi infections preceded T. spiralis by 7 days. An unusual feature of C3H mice was that T. musculi infections 7 days before T. spiralis increased antifecundity responses at the same time that worm expulsion was inhibited. Trypanosome infections can therefore modulate distinct antihelminth immune responses in different directions simultaneously. The different outcomes of dual infections compared with single infections provides another selective mechanism by which genetic polymorphisms can be established and maintained in the vertebrate host.     

Trypanosoma cruzi: Correlation of resistance and susceptibility in infected inbred mice with the in vivo primary antibody response to sheep red blood cells

Nonspecific immune responses during the course of murine Trypanosoma cruzi infection were examined in mouse strains genetically resistant or susceptible to the Brazil strain of T. cruzi. Spleen cells from infected susceptible (C3H) or resistant [C57 B1/10 and FI (C3H × C57)]mice at various points during the course of infection exhibited a reduced response to concanavalin A and lipopolysaccharide in vitro. Since this reduced response occurred in both susceptible and resistant mice, it was not predictive of resistance or susceptibility in vivo. We next examined the kinetics of in vivo primary antibody response to sheep red blood cells (SRBC) in infected C3H and C57 mice. C3H mice exhibited inhibition of the direct plaque-forming cell assay (d-PFC) which persisted until death. In contrast C57 mice exhibited no inhibition of the response at Day 5 and subsequently a markedly augmented response was observed. Other strains of mice were similarly investigated: all the susceptible mice examined (A/J, BALB/c) showed inhibition or depression of the primary antibody response and resistant mice [B10Br, C57B1/10, SJL, F1 (C3H × C57)]demonstrated either no inhibition or considerable augmentation of this response. CS7 mice resistant to the Brazil strain were susceptible to the Tulahuén strain. The mice in this latter group exhibited a markedly significant inhibition of the in vivo primary antibody response to SRBC. Culture forms of the Brazil strain protected C3H mice from a virulent challenge. This immunization resulted in a markedly augmented antibody response. The data reported herein are consistent with the notion that inhibition of the primary antibody response to SRBC correlates with susceptibility whereas no inhibition or, indeed, augmentation of the response correlates with natural as well as acquired resistance.     

Trypanosoma cruzi: Antibody-induced mobility of surface antigens

Antibody induces mobility of surface antigens of live blood forms of Trypanosoma cruzi when these cells are incubated with human or animal antisera. Parasites of one strain (Y) showed aggregation of surface antigens to form an anterior or more frequently a posterior cap. The proportion of cap forming trypomastigotes increased with time and was dependent on temperature and amount of antiserum. Cap formation was inhibited by sodium azide or iodoacetamide. Crosslinking of surface antigens by antibodies causing agglutination of trypomastigotes decreased antigenic mobility and capping. Capping was not affected by treatment of the parasites with colchicine or cytochalasin B. Antigen mobility was not related to the presence of antibodies against parasite and host tissue cross-reacting antigens. Aggregation of surface membrane antigens was also observed in parasites which survive after immune lysis. Results of light and electron microscopic studies suggested that at least part of the aggregated antigens was eliminated from the trypomastigote's surface. Cap formation was strain and stage dependent. It was not observed when Y-strain epimastigotes were used and it occurred less frequently in CL than in Y-strain trypomastigotes.     

Trypanosoma cruzi: Activity of heterocyclic cationic molecules in vitro

Chagas disease remains a serious public health problem in several Latin American countries. New chemotherapy is urgently needed since current drugs are limited in efficacy and exhibit undesirable side effects. Aromatic diamidines and analogs are well known anti-parasitic agents and in this study, we have evaluated the in vitro trypanocidal effect of several different heterocyclic cationic compounds, including diamidines (DB1195, DB1196 and DB1345), a monoamidine (DB824), an arylimidamide (DB613A) and a guanylhydrazone (DB1080) against amastigotes and bloodstream trypomastigotes of Trypanosoma cruzi, the etiological agent of Chagas disease. Our present findings showed that all compounds exerted, at low-micromolar doses, a trypanocidal effect upon both intracellular parasites and bloodstream trypomastigotes of T. cruzi. The activity of DB1195, DB1345, DB824 and DB1080 against bloodstream forms was reduced when these compounds were assayed in the presence of mouse blood possibly due to their association with plasma constituents and/or due to metabolic instability of the compounds. However, trypanocidal effects of DB613A and DB1196 were not affected by plasma constituents, suggesting their potential application in the prophylaxis of banked blood. In addition, potency and selectivity of DB613A, towards intracellular parasites, corroborate previous results that demonstrated the highly promising activity of arylimidamides against this parasite, which justify further studies in experimental models of T. cruzi infection.