Macrophage Migration Inhibition Test Research Articles

Protective activity of specific transfer factor against Salmonella typhimurium infection

Specific transfer factor (TF) extracted from spleens of sensitized and non-sensitized guinea pigs to study the efficacy of transfer of cellular immunity specific for salmonella . Two groups each of five guinea pigs were used for in vivo TF preparation. The First group was inoculated with 1 ml of aromatic dependent Salmonella typhimurium SL 1479 vaccine at a dose of 10' cfu/ml intramuscularly twice at two weeks intervals. The second group was injected with trypticase soy broth similarly . These two groups used as a donor for TF, and TFn respectively. Twenty one recipient guinea pigs were divided into three groups, the first group was TF at a dose of 1 ml equivalent to 5X10 cell intramuscularly three times/2 days intervals, Similarly the second group was given TF, where as the third group was given PBS- Cell mediated immunity in recipient animal was evaluated by delayed type hypersensitivity - skin test, Macrophage migration inhibition test (MIF) and then challenged with virulent Salmonella typhimurium. The TFt recipient group induced skin test and showed migration indices less than 0.8 and overcome the challenge organism. Contrary to TFn & PBS recipient groups which did not show any response for skin test and given migration indices more than 0.8 and did not show resistance for virulent Salmonella typhimurium .

Immunologic and testicular response in guinea pigs after unilateral traumatic orchitis.

An acute and intense trauma was inflicted in one of the testes of adult guinea pigs. The immunological response and histological changes developed both in the injured and contralateral glands were studied at different intervals of time. It was observed: 1) Necrotic seminiferous tubules and a granulomatous cell reaction were seen in the injured gonad at 15 days. This was followed by a reduction of the mononuclear cell infiltration at 48 days. Finally a fibrotic tissue replaced the damaged areas at around 100 days. 2) Foci showing the classical picture of immunologic orchitis appeared in the contralateral gland at 15 and 48 days. Less frequent lesions were detected in the final period of the experiment. 3) These lesions were accompanied by cellular immunity detectable by the macrophage migration inhibition test at 15 and 48 days. With the exception of cytophilic activity also detected in same periods, no antibodies of hemagglutinating and sperm-agglutinating types could be found.

The Study of Zinc’s Effect on Immunological Response in Guinea Pigs

The purpose of this study is to determine the effect of zinc on immunity, by use some immunological tests about humeral and cellular immunity. 18 guinea pigs put in three Groups (6 animals for each Group). the first group take a zinc (0.08 milligram),while the second take (0.16).the last treatment take a distill water as a control. (The doses given daily by stomach tube for tow months). ELISA test used to determine the antibodies level after vaccination with S typhi vaccine, Therese significant difference in response of different treatments compare to control, where this difference not found between treatments. The delayed type hypersensitivity test show that the highest effect is for treatment with Zn1 more than Zn2. Also with the phagocytic activity, highest phagocytic rate in treatment with Zn1. this effect decrease with heightened the dose. The macrophage migration inhibition test shows a significant difference in effect of zinc in compare with control.

Boophilus annulatus (Acari: Ixodidae): cattle-acquired humoral and cell-mediated immunity against tick gut and salivary gland antigens

Assays of antibodies and cell-mediated immunity (CMI) against specific antigens in Boophilus (B.) annulatus Say gut and salivary glands were performed using Bos indicus cattle infested for prolonged periods as a model for naturally acquired immunity to ticks. It was shown that cattle sera contained several antibodies that were reactive with gut proteins (45,66,116,150, and 274 KDa Mol. Wt.) and with salivary gland proteins (66,97,150 and 205 KDa Mol. Wt.) by an enzyme-linked immunosorbent assay (ELISA) and dot immunoblot assays. CMI positive responses to both gut proteins (2000 and 150 KDa Mol. Wt.) and salivary gland protein (150 KDa Mol. Wt) were observed by a macrophage migration inhibition (MMI) test. The potential usefulness of B. annulatus antigenic proteins for vaccination of local cattle breeds is the subject of an ongoing study.

Correlation among immune response, morphogenesis of the granulomatous reaction and spleen lymphoid structure in murine experimental paracoccidioidomycosis

We studied the correlation among cellular immune response, the pattern of lung granulomatous lesions and alterations in spleen lymphoid structure in Swiss mice inoculated intravenously with Paracoccidioides brasiliensis strain 18. The animals were evaluated at 24, 48 and 96 h after infection and further studied weekly for 18 weeks by: (i) the macrophage migration inhibition test with phytohemagglutinin (PHA) and P. brasiliensis antigen (PbAg); and (ii) histopathology of the lung and spleen lesions. One group of animals was γ-irradiated (8 Gy), infected under the same conditions and evaluated for the pattern of lung granulomatous lesions and spleen lymphoid structure at 24, 48 and 96 h after infection. During the first week of infection, the non-irradiated animals presented a positive response to PHA and PbAg, compact granulomas in the lungs and a typical hyperplasia of the spleen white pulp. However, from weeks 2 to 5, a depression of the cell-mediated immunity (CMI) response to PHA and PbAg was observed ...

Correlation among immune response, morphogenesis of the granulomatous reaction and spleen lymphoid structure in murine experimental paracoccidioidomycosis.

We studied the correlation among cellular immune response, the pattern of lung granulomatous lesions and alterations in spleen lymphoid structure in Swiss mice inoculated intravenously with Paracoccidioides brasiliensis strain 18. The animals were evaluated at 24, 48 and 96 h after infection and further studied weekly for 18 weeks by: (i) the macrophage migration inhibition test with phytohemagglutinin (PHA) and P. brasiliensis antigen (PbAg); and (ii) histopathology of the lung and spleen lesions. One group of animals was gamma-irradiated (8 Gy), infected under the same conditions and evaluated for the pattern of lung granulomatous lesions and spleen lymphoid structure at 24, 48 and 96 h after infection. During the first week of infection, the non-irradiated animals presented a positive response to PHA and PbAg, compact granulomas in the lungs and a typical hyperplasia of the spleen white pulp. However, from weeks 2 to 5, a depression of the cell-mediated immunity (CMI) response to PHA and PbAg was observed in association with granulomas presenting only large mononuclear cells and lacking both giant cells and a peripheral halo of small mononuclear cells. This pattern of granuloma formation was similar to that seen in gamma-irradiated animals, whose cells involved in CMI were absent. After week 7, the non-irradiated animals showed granulomas characterized by the presence of giant cells and a peripheral halo of small mononuclear cells. This type of granuloma was formed concomitantly with recovery of the CMI and of the lymphoid structure of the spleen. The results showed a correlation among granulomas composed of large mononuclear cells, hypoplasia of the splenic tissue and impaired CMI. This correlation indicated that although granuloma morphogenesis per se does not depend on the activation of CMI, this response is important at later stages during modulation of the cellular composition of the granulomas.

Immunotoxicity of beryllium

The lymphocyte transformation test and the macrophage migration inhibition test are quantitative methods invaluable for examination of beryllium (hereafter referred to as Be) effects on cell-mediated immunity. We recognized that the Be sensitizing ability was related to active as well as passive cell-mediated immunity in mice subcutaneously injected with Be once a week over a 6-week period. Be also affects B cells, and it increases the amount of immunoglobulins in sera. In the study of immunological health surveys of Be workers in a copper-beryllium casting factory, the serum complement titer tended to be lower in Be workers than in the controls. In mice, injected with Be once a week over a 12-week period, serum complement titers decreased. Correlation coefficients of the experimental parameters showed a significant negative correlation between the complement titers and the prothrombin time or the coagulation time for factor VII, using mice injected with 5 micrograms of Be. It was suggested that increases in the complement titers after Be administration may be induced by temporarily-activated plasma serin protease, which is a component of blood coagulation factor VII. The delta-aminolevulinic acid dehydratase and porphobilinogen deaminase activities were significantly elevated in the pregnant untreated group, compared with the nonpregnant mice (the control group). However, it was noted that these values in the pregnant mice injected with 50 micrograms of Be were almost the same as the values of the controls. It suggests that Be suppressed the expected pregnancy-induced increase in hematopoietic function. There are at least two risk factors induced in the effects of beryllium on organisms-exposure to the metal and inheritance of the genetic marker. It is necessary to reduce exposure, to give preventive education and to carry out periodic health examinations for the prevention of disease induced by Be.

Evaluation of Paracoccidioides brasiliensis exoantigen in the detection of delayed dermal hypersensitivity in experimental and human paracoccidioidomycosis.

The exoantigen of Paracoccidioides brasiliensis standardized by Camargo et al. (AgR) was used to evaluate the in vivo and in vitro cell immune response of experimental animals and of patients with paracoccidioidomycosis (PBM). Fava Netto antigen (AgF) was tested in parallel as a control antigen. The study was conducted with mice and guinea pigs infected with P. brasiliensis or immunized with its fungal antigens, on patients with PBM and on their respective control groups. The cell immune response was analysed by skin tests, and by the macrophage and leucocyte migration inhibition tests (MMIT and LMIT) in the animals and in the patients, respectively. The skin test with AgR as paracoccidioidin was positive in infected or immunized mice and guinea pigs and negative in control animals. The skin tests with AgR (24 h) showed 96.7% positivity in patients with PBM and were negative in control individuals. Histopathological study of the in vivo tests in the different experimental models was consistent with a delayed hypersensitivity response (DHR). Immunohistochemical study of the skin tests of PBM patients demonstrated a predominance of T lymphocytes, confirming the nature of a DHR to the fungal antigens. The in vitro cell immune response showed variable results for the various experimental models, i.e. significant rates of MMIT in immunized mice, a tendency to positivity in infected guinea pigs, and the absence of migration inhibition in PBM patients. Taken together, the data indicate that the AgR is efficient as paracoccidioidin in the evaluation of DHR in PBM, with an optimum time of reading the test of 24 h.

Cellular immune responsiveness in rabbits with Setaria Digitata filarial antigen and TDM adjuvant

The purified surface antigens of the bovine filarial parasite Setaria digitata were used as an antigen to immunize rabbits. The aqueous suspensions of trehalose 6-6′ dimycolate (TDM) has been successfully used as an effective immunomodulator in experimental studies on filariasis. The effectiveness of such an antigen — TDM combination was demonstrated by enhanced humoral and cellular immunity. Administration of antigen alone shows only humoral immunity. The detectable cellular immune responses further confirm the effect of filarial antigen — TDM combination. The cell-mediated immunity was expressed in vivo by delayed skin reaction and in vitro by leukocyte and macrophage migration inhibition tests.

Dialysable leukocyte extracts modify the course of experimental paracoccidioidomycosis in the Syrian hamster.

The effect of dialysable leukocyte extracts (DLE) obtained from hamsters immunized with Paracoccidioides brasiliensis (immune DLE) and from non-immunized hamsters (non-immune DLE) was studied in hamsters inoculated with P. brasiliensis by the intratesticular route. Treatment with immune or non-immune DLE was started during the third week of infection and was repeated at 7, 11, 15 and 19 weeks. A group of untreated infected animals was used as control. Animals were submitted to the delayed hypersensitivity skin test to P. brasiliensis antigen (PbAg) in vivo and assayed in vitro by the macrophage migration inhibition test in the presence of Phytohemagglutinin (PHA) and PbAg and by immunodiffusion for specific antibody. The animals were sacrificed at 4, 8, 12, 16 and 20 weeks. The morphology and extension of the lesions were studied at the inoculation site, and in lymph nodes, lungs, liver, spleen and kidneys. In contrast to the controls, animals treated with both DLEs maintained a positive cell-mediated immune response throughout the experiment and developed less extensive infection with a significantly lower number of fungi in the lesions. The results suggest that immune and non-immune DLE preparations modified the evolution of experimental paracoccidioidomycosis with equal efficiency. This similarity may be explained by the immunoregulatory activities of both extracts.

Probable role of Streptococcus pyogenes in Kawasaki disease.

Over the past 25 years, the clinical course of Kawasaki disease has been defined, the prevalence and nature of the cardiovascular effects widely understood, and pathological changes in the most severe cases well described. However, the aetiology and pathogenesis of this puzzling disease have remained unclear, thus specific therapy is not yet available. Because of some close clinical similarities between this disease and streptococcal scarlet fever, particular attention has been paid to the possible role of Streptococcus pyogenes as an aetiological agent in this illness. Until now, however, group A beta-haemolytic streptococci have never been consistently isolated from any patients; in addition, the titre of anti-streptolysin 0 is not raised, and lack of response to antibiotics is a feature of this disease. Our long series of investigations over more than 10 years, which will be covered in the present review, were performed in an attempt at elucidating causative agent(s) of Kawasaki disease. This has led to our firm belief in the probable role of S. pyogenes in the pathogenesis of this disease, despite the lack of fulfillment of Koch's postulates, on the basis of the following findings. Patients with Kawasaki disease recovering from the acute, febrile phase of the illness exhibited an exaggerated cell-mediated reactivity, as measured by the macrophage migration inhibition test, to group A beta-haemolytic streptococci, their pyrogenic exotoxin and streptolysin 0 as well as to several mammalian muscle cell extracts which are allegedly related antigenically to the cell wall and/or cytoplasmic membrane of S. pyogenes. Protoplast-like "spherical bodies" varying in diameter from 0.5 to 1.5 microns, and devoid of cell walls, were detected in the buffy coats of peripheral blood from patients with this disease, and stained distinctly by immuno-electron microscopy using, as a primary antibody, a rabbit antiserum to S. pyogenes- derived protoplasts, and followed by absorption with protoplasts from Staphylococcus aureus and Escherichia coli. Newborn mice infected with S. pyogenes having no capacity to confer cell-mediated immunity even in adult murine hosts, and reinfected 4-6 weeks later with another strain of the same species of bacteria which is able to elicit cellular immunity, showed a lack of humoral response to streptococcal antigens, leaving intact cell-mediated immunity. Such a biased immunological characteristic is an exact counterpart of that of Kawasaki disease patients.(ABSTRACT TRUNCATED AT 400 WORDS)

Occupational diseases caused by exposure to sensitizing metals

Diseases caused by occupational exposure to sensitizing metals including platinum (Pt), rhodium (Rh), nickel (Ni), chromium (Cr), cobalt (Co), gold (Au), mercury (Hg), zirconium (Zr) and beryllium (Be) are reviewed. Allergic reactions induced by the metals are described according to the classification by Coombs and Gell. Metals with unproven sensitizing potential are not discussed if reports on these are either very rare or devoid of convincing evidence for allergic involvement. The sensitizing metals are haptens which are not themselves able to act as antigens. There is evidence that combination of the metals with circulating or tissue protein gives rise to new antigens. An alternative hypothesis is that these metals interfere with the antigen recognition step of the immune response. Immunomodulatory effects or immunotoxicity of the metals may be also involved in metal-induced hypersensitivity. Occupational exposure to Pt, Rh, Ni, Cr, and Co causes allergic asthma via type I allergic reaction in which serum from affected individuals shows specific IgE antibodies against mental-human serum albumin conjugates. Some rheumatoid arthritis patients treated with gold salt therapy develop glomerulonephritis, thrombocytopenia, or agranulocytosis, which arise from type II and/or type III allergic reactions. Occupational exposure to mercury causes glomerulonephritis in which involvement of type III reaction is suggested. Type IV hypersensitivity reaction of the skin also takes place following exposure to the metals: allergic contact dermatitis is evoked by exposure to Ni, Cr, Co, Rh, and Hg; cutaneous granuloma is formed by contact with Zr and Be. Be is also a sensitizer of the lungs, resulting in granulomatous disease. Diagnosis of metal-induced allergic diseases is made on the basis of allergological tests with metal antigens including skin tests, radioallergosorbent test for specific antibody, lymphocyte transformation test, macrophage migration inhibition test, and provocation test. Atopy is a predisposing factor and smoking is a risk factor for developing metal-induced asthma. Evidence for genetic factors in the development of metal contact dermatitis is conflicting, although animal models implicate genetic factors in skin sensitization with some metals and respiratory sensitization with Be. Skin irritation, forearm injury, complication with atopic dermatitis and concomitant sensitization to other agents are determinants for prognosis of the dermatitis. Epidemiological reports of occupational diseases from allergic reactions to metals in industries are reviewed with respect to prevalence and allergic manifestations. There is a report on a clinical trial of hyposensitization with Pt in a platinum asthma patient. Predictive methods for evaluating sensitization potential of metals have been developed and new methods, which quantify potential more objectively, are sought.

Is injectable collagen truly safe?

Most patients have no response to injectable collagen or silicone, but some cases may have positive or 'undersea' (= clinically negative but immunologically positive) response to collagen. From the results of the Macrophage migration inhibition test, the relative immunogenicity was augmented most when we used implants with the following combination. The first immunization was collagen and the second one was collagen with silicone. The augmented antigenicity might be enough to cause an allergic reaction to the patients who had no response to each implant alone.

Circulating immune complexes in experimental syphilis and their relation to immunological response againstTreponema pallidum

It was found that circulating immune complexes (CIC) were formed in rabbits at different times after infection with Treponema pallidum. The CIC which appeared at the beginning of the disease were short-lived (2-6 weeks) but those appearing later than 20 weeks after infection remained for 10-25 weeks. CIC contained both IgM and IgG classes of immunoglobulin. The antibodies present in CIC were found to be specific and nonspecific for T. pallidum. The presence of CIC led to a marked decline of treponemal antibodies in rabbit sera. The cell-mediated immune response measured by the macrophage migration inhibition (MMI) test at the beginning of the disease (up to 12 weeks) was not decreased. However, when syphilis lasted for more than 14 weeks and when CIC were formed mainly from IgG, a distinct decrease in the ability of lymphocytes to cause MMI was observed. These findings strongly suggest that IgG-complexes suppress the immunological responsiveness of lymphocytes against T. pallidum which in turn facilitates the multiplication of treponemes in the host.

Comparison of cross-reactivities of imipenem and other beta-lactam antibiotics by delayed-type hypersensitivity reaction in guinea pigs.

The cross-reactivity of imipenem (IPM) in a delayed-type hypersensitivity (DTH) reaction was investigated by intradermal skin reaction, macrophage migration inhibition tests (MIT) and lymphocyte stimulation tests (LST) in guinea pigs. The animals were immunized with IPM, ampicillin (ABPC) or cephalexin (CEX) using Freund's complete adjuvant. Three sensitized-drugs exhibited the same immunogenicity in the skin reaction. The cross-reactivities in skin reaction among IPM, sulbactam, aztreonam, ABPC and 6-aminopenicillanic acid as penam, and CEX, ceftizoxime, 7-aminocephalosporanic acid as cephem were examined. IPM did not show cross-reactions with any of the tested drugs in three sensitized groups, indicating that the drugs possessed no cross-reactivity with tested beta-lactams in DTH. In MIT, 4 or 5 of 6 animals reacted to be sensitized drugs in each group. The cross-reactivity of IPM- and ABPC-sensitized groups in MIT was similar to that of a skin reaction, however, the CEX-sensitized animals showed broad cross-reactions. Using a kind of test drug, one or two animals in the ABPC-sensitized group showed migration enhancement, but IPM- or CEX-sensitized animals did not exhibit migration enhancement. In LST, most of the animals tested exhibited lymphocyte prolifration by sensitized drugs but a cross-reaction was scarcely observed. The above results suggest that among beta-lactams, IPM has a very low cross-reactivity. LST is considered to be more a useful assay than MIT for in vitro identification of causative drugs in animal models.

Immunostimulatory Effect of Rothia dentocariosa in Mice

In mice killed Rothia dentocariosa cells in doses of about 1.5 mg dry weight activated anti-infection immunity to Listeria antigens and anti-tumour immunity to the ascitic form of mouse sarcoma S-180. Their probable target site is the macrophage. The Rothia-activated macrophages in human gingiva may take part in the pathogenesis of periodontal disease. Three models were employed to verify the immunostimulating properties of preventively administered Rothia dentocariosa bacterin-1) a spleen macrophage migration test, using mice immunized with Listeria innocua, with the soluble listeria Ei antigen as the antigenic signal, 2) determination of the increase in the Listeria monocytogenes LD50 for mice and 3) the prolongation of survival of mice carrying the S-180 tumour. In all three cases, the administration of Rothia bacterin stimulated the immune response to the later administration of other antigens. Furthermore, in the macrophage migration inhibition test, the chemotaxis of non-immune mouse macrophages was found to be stimulated. This gives evidence of the fact that Rothia bacterin has an activating effect on these macrophages.

A possible case of drug-induced familial pemphigus.

Two sisters developed pemphigus vulgaris and pemphigus erythematosus within 3 years. The diagnosis was confirmed by clinical, histologic and immunofluorescent antibody studies. One of the sisters experienced a common cold before the pemphigus developed and displayed a positive macrophage migration inhibition (MIF) test to a combination drug compounded of paracetamol, caffeine, chlorpheniramine maleate and phenylephrine HCl, which she had received 2 weeks prior to the appearance of the cutaneous lesions. It is suggested that her pemphigus was triggered by the drug. Although the patient had a strong genetic and familial predisposition to pemphigus, her clinical symptoms did not become evident until they were activated through an exogenous factor, namely, the causative drug. This case offers an example of a possible interaction between endogenous, genetic factors, and exogenous, triggering factors in the development of full-blown disease.

The serial changes of antibody production, macrophage migration inhibition test (MIT) and histopathologic examination in an experimental hypersensitivity pneumonitis in rabbits

We developed a hypersensitivity pneumonitis model by sensitizing rabbits intratracheally (IT) twice after two subcutaneous sensitizations (SS), and investigated the serial changes in the production of antibody against micropolyspora faeni (Mf), macrophage migration inhibition test (MIT), and histopathologic features. On the third day after the last IT, the proportions of neutrophils in bronchoalveolar lavage fluid (BALF) increased. The antibody levels of Mf specific-antigen in BALF decreased transiently, but those in serum began to increase. The number of AFC in spleen increased rapidly. Histopathologic examination revealed alveolitis and prominent acute pneumonia. On the seventh day after the last IT, the proportion of lymphocytes in BALF, the antibody levels of Mf specific-antigen in BALF, the number of AFC in BALF, and the percent inhibition of MIT were highest. Histopathologic features were characterized by the formation of prominent epithelioid granulomas. On the fourteenth day after the last IT, these features returned to the state of untreated group. These results indicate that the lesions in this HP model are formed by both type III and type IV immune reactions (Coombs-Gall). It was concluded that the seventh day after the last IT was most appropriate for the study of pathophysiology of human HP.

In vitro tests with sensitized lymphocytes—Relevance for predictive allergenicity testing

The main obstacle to routine in vitro tests to identify contact sensitizers is the inappropriate application of the allergens in vitro, even when sufficient antigen-presenting cells are available. Some allergens may need the skin tissue for adequate conjugation with carrier molecules, whereas others may be poorly solubilized or toxic for the cultures. Methods, therefore, which make use of the in vivo system for allergen binding and presentation, but which assess in vitro the subsequent lymphocyte activation are most promising. For this reason, the local lymph node assay deserves more attention as a predictive assay in allergenicity testing and should be validated in more laboratories in the near future. The value of allergen-stimulated in vitro tests for predictive allergenicity screening will remain limited. This type of in vitro test is, however, invaluable for mechanistic studies on contact sensitivity in well defined allergen models. Also for clinical diagnosis of allergic contact sensitivity to selected allergens, the lymphocyte transformation test and macrophage migration inhibition test may be useful, particularly when skin tests fail to be conclusive.

犬の住血原虫Ｂａｂｅｓｉａ ｇｉｂｓｏｎｉの免疫に関する研究 Ｂｏｒｄｅｔｅｌｌａ ｂｒｏｎｃｈｉｓｅｐｔｉｃａの免疫増強効果

Four species of bacteria, Corynebacterium anaerobium 578, Actinobacillus pleuropneumoniae G-4, Mycobacterium bovis BCG, and Bordetella bronchiseptica A-2, were injected intravenously into mice (5 weeks old, ICR-SPF). The clearance of carbon from the blood stream and the weights of the spleen and liver were determined as indicators of RES stimulation. Mouse footpad reaction was assessed as an indicator of delayed-type hypersensitivity to each species of bacteria. The immuno-stimulative activity of each species of bacteria against bovine serum albumin was monitored by passive hemagglutination assay and the macrophage migration-inhibition test in guinea pigs. Based on the results of the experiments described above, B. bronchiseptica was selected as an immunostimulator (Ims) for immunization trials of the hemo-protozoan parasite, Babesia gibsoni, with inactivated merozoites of B. gibsoni (BgK). Twelve dogs, pointers about 6 months old, were divided into four groups of three dogs each. Group 1 dogs were initially injected with Ims, and later injected with BgK and Ims (BgK+Ims) after a 3-week interval. Group 2 and Group 3 dogs were injected twice, at a 3-week interval, with BgK+Ims and BgK, respectively, and Group 4 served as a control. As the results, the serum antibody titres of Group 1 and 2 were several times higher than that of Group 3, and the cell-mediated immunity to parasites was noticeably stimulated by immunization with BgK+Ims. The peak level of parasitemia following the challenge were over 10% for Group 4 and 4.5% for Group 3, while levels for Group 1 and 2 were 2.5% and less than 1%, respectively. No such major clinical signs of babesiosis as jaundice and anemia were observed in Group 1 or 2.