Brasiliensis Antibodies Research Articles

Inducible nitric oxide synthase blockade with aminoguanidine, protects mice infected with Nocardia brasiliensis from actinomycetoma development

Mycetoma is a chronic infectious disease that can be caused by fungi or bacteria, Madurella mycetomatis and Nocardia brasiliensis are frequent etiologic agents of this disease. Mycetoma produced by bacteria is known as actinomycetoma. In mycetoma produced by fungi (eumycetoma) and actinomycetoma, diagnosis of the disease is based on clinical findings: severe inflammation, with deformities of affected tissues, abscesses, fistulae, sinuses and discharge of purulent material that contains micro colonies of the etiologic agent. Microscopic examination of infected tissue is similar regardless of the offending microbe; hallmark of infected tissue is severe inflammation with abundant neutrophils around micro colonies and granuloma formation with macrophages, lymphocytes, dendritic and foamy cells. Even though medical treatment is available for mycetoma patients, amputation, or surgical intervention is frequently needed. The pathogenesis of actinomycetoma is little known, most information was obtained from experimental animal models infected with bacteria. In other experimental mice infections with different microbes, it was demonstrated that nitric oxide is responsible for the intracellular killing of Mycobacterium tuberculosis by activated macrophages. Nitric oxide is a free radical with potent stimulatory and suppressive effects in innate and adaptive immunity. The unstable nitric oxide molecule is produced by action of nitric oxide synthases on L-arginine. There are three nitric oxide synthases expressed in different cells and tissues, two are constitutively expressed one in neurons, and the other in endothelial cells and one that is inducible in macrophages. Aminoguanidine is a competitive inhibitor of inducible nitric oxide synthase. Its administration in experimental animals may favor or harm them. We used aminoguanidine in mice infected with Nocardia brasiliensis, and demonstrated that all treated animals were protected from actinomycetoma development. Anti N. brasiliensis antibodies and T cell proliferation were not affected, but inflammation was reduced.

First report of Paracoccidioides brasiliensis infection in fish.

The thermodimorphic fungus Paracoccidioides brasiliensis is the etiological agent of paracoccidioidomycosis (PCM), a deep mycosis endemic in Latin American countries that affects mainly male rural workers. Infection by P. brasiliensis has also been reported in several species of terrestrial animals; however, the capacity of the fungus to infect aquatic organisms is poorly known. The aim of this study was to detect P. brasiliensis in a fish species, Nile tilapia (Oreochromis niloticus), the most farmed and widely distributed fish in endemic areas for human PCM in Brazil. As a first step, the humoral immune response against the fungus was evaluated in an experimental group of three fish immunized with inactivated P. brasiliensis yeast cells. For the seroepidemiological study, serum samples of Nile tilapia raised in cages (n=109) and in ponds (n=105), collected from a fish slaughterhouse, were analyzed for P. brasiliensis antibodies by ELISA using gp43 as antigen. All the inoculated fish produced antibodies against the fungus. The seropositivity observed in fish raised in cages and ponds was 17.4 and 5.7%, respectively. Due to the higher seropositivity observed in caged fish, 100 tissue samples (encephalon, liver, and kidney), from another group of tilapia raised in cages, were analyzed by polymerase chain reaction (PCR; Pb-ITSR and Pb-ITSE). Three tissue samples (liver n=1, kidney n=1, and enchepahlon n=1) from three different fish resulted positive to PCR. This is the first report to show serological and molecular evidence of P. brasiliensis infection in a fish species.

Evaluation of (1→3)-β-D-glucan assay for diagnosing paracoccidioidomycosis.

Paracoccidioidomycosis (PCM) is highly prevalent in Latin America, but no commercial system is available for diagnosing this endemic mycosis. To check the performance of (1→3)-β-D-glucan assay (BDG) for diagnosing PCM in 29 patients with proven fungal disease and compared with double immunodiffusion assay for detecting anti-Paracoccidioides antibodies. We selected 52 serum samples sequentially obtained from 29 patients with active PCM (12 chronic and 17 acute form). Samples were collected at baseline, and for16patients, additional serum levels were obtained after 3 and 6months of antifungal treatment. Detection of BDG in serum was performed by using the Fungitell® assay. For the double immunodiffusion assay, Paracoccidioides exoantigen was used in latex agglutination tests to detect serum anti-Paracoccidioides antibodies. Despite exhibiting good sensitivity in the diagnosis of patients with PCM, we failed to demonstrate any correlation between the postdiagnosis kinetic profile of BDG serum levels and clinical response to antifungal therapy. This finding may be related to the maintenance of quiescent foci of fungal infection in several organs and tissues, a phenomenon that has been previously reported by other authors and helps to understand why so many relapses are documented in patients treated for short periods of time. Finally, we did not find any correlation between BDG quantification and specific anti-P brasiliensis antibodies serum titres in patients with PCM. In conclusion, BDG is detected in serum samples of most patients with PCM but is probably not useful for predicting clinical response to antifungal therapy.

Terapia fotodinâmica na artrite experimental induzida por Paracoccidioides brasiliensis

AIMS: To evaluate the effectiveness of photodynamic therapy with Brilliant Blue G in the treatment of an experimental model of arthritis by Paracoccidioides brasiliensis (P. brasiliensis). METHODS: After the induction of experimental arthritis with isolated from P. brasiliensis of lineage Pb18 in the knees of Wistar rats, the animals were divided into groups and submitted to photodynamic therapy with intra-articular Brilliant Blue G photosensitizer and laser therapy only, without Brilliant Blue G. All groups received their respective treatments from the seventh to the 11th day. For edema analysis, the knee lateral-lateral diameter of each animal was measured daily and after the treatment period the animals were sacrificed for experimental knee dissection and blood collection for analysis by ELISA, in order to quantify levels of anti-P. brasiliensis antibodies. RESULTS: The results showed that the application of photodynamic therapy was able to prevent the formation of edema when compared to the control (p>0.005), as well as the production of anti-Gp-43 antibodies from P. brasiliensis (p=0.001). In the anatomopathological examination it was possible to observe a higher degree of synovitis and a greater presence of granulomas with the fungus inside the group that did not receive treatment when compared to the groups that received the photodynamic therapy. CONCLUSIONS: Photodynamic therapy was effective in attenuating the experimental arthritis induced by P. brasiliensis in the proposed joint model.

Estrogen Receptor-α Correlates with Higher Fungal Cell Number in Oral Paracoccidioidomycosis in Women.

Paracoccidioidomycosis is a neglected tropical fungal infection with great predilection for adult men, indicating the participation of female hormone estrogen in preventing paracoccidioidomycosis development in women. Estrogen has an immunologic effect leading to polarization toward the Th2 immune response, which favors the disease evolution. To evaluate estrogen and progesterone receptors in oral paracoccidioidomycosis lesions and to verify any association with tissue fungi counting in women and men. Thirty-two cases of chronic oral paracoccidioidomycosis were included. Immunohistochemical analyses for anti-estrogen receptor-α, anti-progesterone receptor and anti-Paracoccidioides brasiliensis antibodies were performed. The differences between women and men and the relations among the immunomarkers for each gender were also evaluated. A significant positive correlation was observed between estrogen receptor-α and the amount of fungi in women. In addition, estrogen receptor-α was mildly expressed in the inflammatory cells of female patients, while progesterone receptor was expressed in both genders, with similar expression between women and men. Moreover, fungi counting revealed no differences between genders. Estrogen receptor-α was expressed only in women and showed a positive correlation with the amount of fungi in oral paracoccidioidomycosis, while progesterone receptor was observed in both genders and exhibited no correlation with estrogen receptor-α or fungi counting.

Quantitative Analysis of AntiHevea brasiliensis Antibody Cross-Reactivity against Taraxacum kok-saghyz Latex Proteins Demonstrates Significantly Reduced Antibody Recognition

The annual plant Taraxacum kok-saghyz, commonly called Russian dandelion, has recently gained commercial attention as an alternative source to the rubber tree, Hevea brasiliensis, for natural rubber latex. While producing rubber of equivalent quality, the potential application of TKS latex for hypoallergenic products remains to be determined. In order to quantify the allergenicity of TKS latex proteins, we compared the extent of anti-Hevea latex polyclonal and monoclonal antibody cross-reactivity toward Hevea latex proteins to that of TKS latex proteins by quantitative ELISA methods and semi-quantitative image densitometry. ELISA measurement of polyclonal antibody recognition toward TKS latex proteins was on the order of one-tenth relative to equal amounts of Hevea latex protein, while recognition by the monoclonal antibody was below 2%. Immunoblots confirmed significantly reduced polyclonal antibody recognition toward TKS latex proteins, in the same range as the cross-reactivity exhibited toward the rubber-producing plant, Lactuca sativa and Glycine max, a non-rubber-producing plant. Despite the presence of cross-reactivity, these quantitative results support TKS as an alternative source of latex that may require lessintensive processing for the reduction of antigenic proteins in the manufacture of hypoallergenic natural rubber latex products.

Paracoccidioides brasiliensis interacts with dermal dendritic cells and keratinocytes in human skin and oral mucosa lesions.

Paracoccidioidomycosis (PCM) is a systemic disease caused by the fungus Paracoccidioides brasiliensis and Paracoccidioides lutzii. In PCM the skin and oral mucosa are often affected. Dendritic cells and keratinocytes of the integument play a role in innate and adaptive immune response against pathogens, due to their function as antigen presenting cells. Aiming to verify the interaction of P. brasiliensis with these cell populations, we studied 52 skin and 47 oral mucosa samples taken from patients with proven diagnosis of PCM. The biopsies were subjected to immunohistochemical and/or immunofluorescence staining with anti-factor XIIIa (marker of dermal dendrocytes), anti-CD207 (marker of mature Langerhans cells), anti-pan cytokeratins (AE1-AE3) and anti-P. brasiliensis antibodies. Analyses with confocal laser microscopy were also performed for better visualization of the interaction between keratinocytes and the fungi. In sum, 42% of oral mucosa samples displayed yeast forms in Factor XIIIa dermal dendrocytes cytoplasm. Langerhans cells in skin and oral mucosa samples did not show yeast cells in their cytoplasm. In sum, 54% of skin and 60% of mucosal samples displayed yeast cells in the cytoplasm of keratinocytes. The parasitism of keratinocytes may represent a possible mechanism of evasion of the fungus to local immune mechanisms. Factor XIIIa dendrocytes and keratinocytes may be acting as antigen-presenting cells to fulfill the probably impaired function of Langerhans cells in skin and oral mucosa of human PCM.

Seroepidemiology of Paracoccidioides brasiliensis infection in horses from Rio Grande do Sul, Brazil.

Paracoccidioides brasiliensis is the etiological agent of the major systemic mycosis in Brazil, called paracoccidioidomycosis. Although the Rio Grande do Sul is considered an endemic area of the disease, there are few studies on the ecology of P. brasiliensis in the state. Therefore, this study aimed to evaluate the infection of P. brasiliensis in horses from the mesoregion of Southwest Riograndense, using these animals as sentinels. Serological techniques, such as double immunodiffusion in agar gel (AGID) and indirect ELISA, were performed to detect the anti-gp43 P. brasiliensis antibody in horses from five different farms in the region of Bagé, RS, Brazil. Serology was performed in 200 Pure Blood English horses up to two years of age that were born and raised exclusively at the farms. Of these horses, 12% had anti-gp43 antibodies according to the ELISA results, with rates ranging from 0 to 30% according to the farm of origin (p < 0.001). Based on the immunodiffusion results, all equine serum samples were negative. These results indicate the presence of the fungus P. brasiliensis in the middle region of the southwestern state of Rio Grande do Sul, Brazil.

IgG anti nocardia brasiliensis antibodies block IgM mediated protective effect in BALB/c mice. (MPF2P.814)

Abstract Intracellular bacteria such as, Nocardia brasiliensis, produce a chronic human disease named actinomycetoma. Diagnosis of mycetoma is made on clinical findings as swelling and deformation of infected tissue and confirmed by culturing the offending microbe or by an ELISA test. Pathogenesis of mycetoma is little known and role of antibodies is controversial but it has been published that that cell mediated immunity is responsible of host protection in other intracellular bacterial infections. We used Nocardia brasiliensis to infect BALB/c mice and studied the role of both monoclonal and polyclonal antigen-specific IgM and IgG antibodies. We investigated antibodies role in host protection using passive immunization. Mice recipients of anti N. brasiliensis antibodies were then infected to assess actinomycetoma development. We found that IgM but not IgG anti N. brasiliensis antibodies from sera of infected animals and IgM, but not IgG anti N. brasiliensis monoclonal antibodies were able to confer protection and prevented actinomycetoma infection in experimental animals. IgG antibodies blocked the IgM protective effect in vivo. Implications of these finding in vaccine development are presented.

Ig G and IgM opsonizing antibodies affect Il-12 and TNF-α cytokine production in macrophages infected with a Nocardia brasiliensis recombinant strain. (99.6)

Abstract Nocardia brasiliensis is is an intracellular microbe that replicates in host macrophages. We developed an actinomycetoma model in BALB/c mice and showed that IgM but not IgG anti-N. brasiliensis antibodies conferred protection and prevent ed actinomycetoma lesion. The survival or death of the bacterium, once in macrophages, possibly depends on the antibodies used to opsonize N. brasiliensis. We constructed a recombinant strain of N. brasiliensis that produces a green fluorescent protein (GFP) to evaluate virulence and role of IgG and IgM antibodies in cytokine production by macrophages in BALB/c mice. The recombinant N. brasiliensis strain expressing the green fluorescent protein constructed by adding a plasmidic vector containing the GFP gene was used to infect BALB/c mice and macrophages to evaluate phagocytosis and cytokine production with opsonizing IgG and IgM antibodies. N. brasiliensis GFP induces actinomycetoma as well as N. brasiliensis WT. The production of IL-12 with opsonized bacteria increased with IgG and decreased with IgM antibodies. TNF- α production was greater at 24 and 48 hours in non-opsonized bacteria. Opsonized bacteria with IgG and IgM antibodies decreased TNFα production but only IgM isotype antibodies decreased IL-12 production by macrophages. The modified N. brasiliensis strain conserves its property to induce disease.

Serological diagnosis of paracoccidioidomycosis in HIV-coinfected patients

Paracoccidioidomycosis should be differentiated from other opportunistic diseases in human immunodeficiency virus (HIV)/acquired immunodeficiency syndrome (AIDS) patients who live in Latin America. Laboratory investigation can begin with serological tests, which are rapid and efficient. In the present study, double immunodiffusion (DID), counterimmunoelectrophoresis (CIEP) and an enzyme linked immunosorbent assay (ELISA) tests were assessed for the detection of anti-Paracoccidioides brasiliensis antibodies in 40 patients coinfected with HIV. The results were compared to those obtained for 75 non-HIV-infected patients with endemic paracoccidioidomycosis. Anti-P. brasiliensis antibodies were detected in 65% (DID), 79% (CIEP) and 95% (ELISA) of the patients with HIV/AIDS, significantly lower rates than those detected in cases of endemic paracoccidioidomycosis, which were 89%, 99% and 100%, respectively. The reactive sera of HIV-infected patients also showed lower anti-P. brasiliensis antibody titres than those of non-HIV-infected patients. Despite the lower intensity of the specific humoral response, serological tests are useful for the diagnosis of opportunistic paracoccidioidomycosis in the HIV/AIDS population. We suggest optimization of the laboratory diagnosis by combining the ELISA test with CIEP or DID.

Detection of Anti-Paracoccidioides brasiliensis antibodies in suspected tuberculosis patients

Paracoccidioidomycosis (PCM) is an important systemic mycosis in Latin America that occurs as active disease in 1-2% of Paracoccidioides brasiliensis infected people. Like PCM, tuberculosis (TB) affects mainly the lungs and the clinical and radiological aspects do not always allow differentiation between them. The aim of this study was to carry out serological investigation for detecting anti-P. brasiliensis antibodies, by three serological methods, in patients with symptoms suggestive of pulmonary TB. From August 2005 to September 2006, 76 patients with pulmonary symptoms suspected for TB were attended at the Regional Specialties Center Laboratory in the city of Paranavaí, Paraná, Brazil and submitted to microbiological TB research, ELISA, immunodiffusion and immunoblotting for PCM. Of all the individuals, 21 (27.63%) were reactive to P. brasiliensis by ELISA and 11 (14.47%) showed a laboratory diagnosis of pulmonary TB. Of all the individuals serologically reactive to P. brasiliensis, by ELISA, none had positive results by immunodiffusion and one reacted with antigen 43 kDa when Immunobloting was carried out. Our results lead us to reflect a necessity to obtain a more specific serologic test for diagnosis of PCM disease in patients with respiratory symptoms considering the high number of individuals reactive to P. brasiliensis especially in endemic areas.

Complexo primário da paracoccidioidomicose e hipereosinofilia

Primary infection with Paracoccidioides brasiliensis has rarely been observed. A 28-year-old male patient presented with a three-month history of fever, respiratory symptoms and malaise. Chest X-rays revealed bilateral apical infiltrates, right pleuritis and hilar lymphadenomegaly. The patient presented with leukocytosis, severe eosinophilia and increasing titers of anti-P. brasiliensis antibodies in serum. To our knowledge, this is the first report of the primary pulmonary lymph node complex of paracoccidioidomycosis accompanied by hypereosinophilia and affecting a previously healthy adult.

Characteristics of Environmental Paracoccidioides brasiliensis Isolates

The ecological niche or exact habitat of the fungus Paracoccidioides brasiliensis is not known, and few isolates have been obtained from the environment. In this study, ten isolates were analyzed with respect to antigenic composition, serology, pathogenicity, and molecular aspects. Gp43 is considered to be the molecular basis for the serodiagnosis of paracoccidioidomycosis; however, in this study only six of the environmental isolates secreted this molecule (four in great amounts and two in small amounts). Other molecules were also produced. When exoantigens from these isolates were tested using immunodiffusion, only four preparations were positive by ID tests. However, when these exoantigens were tested by ELISA, all of them except one were able to detect anti-P. brasiliensis antibodies. In Western blot assays, these exoantigens showed different reactivities. Isolates that secreted gp43 presented positive reactions for this molecule, and isolates that did not secrete gp43 gave positive reactions for other minor molecules. RAPD analysis revealed that there is great genetic variation between these environmental isolates. These isolates were non-pathogenic: no mortality was observed among the inoculated mice during an 18-month follow-up period.

IgM but not IgG monoclonal anti-Nocardia brasiliensisantibodies confer protection against experimental actinomycetoma in BALB/c mice

Nocardia brasiliensis is a facultative intracellular microorganism that produces a human chronic infection known as actinomycetoma. Human and mouse anti-N. brasiliensis antibody response identify P24, P26 and P61 immunodominant antigens. In this work, we generated immunoglobulin M (IgM) and IgG monoclonal antibodies (mAbs) specific to immunodominant P61 antigen. The monoclonal IgM (NbM1) and IgG2a (NbG1) antibodies were assessed for their in vitro bactericidal activity, in vivo protective effect and ability to block catalase activity. These mAbs specifically recognized P61, but they did not inhibit its enzyme activity. The in vitro bactericidal effect of NbG1 was higher than the killing ability of the IgM mAb. In vivo experiments with a murine model of experimental infection with N. brasiliensis injected into rear footpads was used to test the effect of NbM1 and NbG1. The negative untreated group developed a chronic actinomycetoma within 4 weeks. IgM mAbs conferred protection to BALB/c mice infected with N. brasiliensis. IgG mAb lacked this protective effect. IgM mAb showed a dose-response correlation between antibody concentration and lesion size. These results demonstrate that humoral immune response mediated by antigen-specific IgM antibody protects against an intracellular bacterial infection.

Sensitivity of an immunoenzymatic test for detection of ant-L. brasiliensis antibodies compared to other tests used for the diagnosis of American cutaneous leishmaniasis

The diagnosis of American cutaneous leishmaniasis (ACL) is frequently based on clinical and epidemiological data associated with the results of laboratory tests. Some laboratory methods are currently being applied for the diagnosis of ACL, among them the indirect immunofluorescence reaction (IIFR), the Montenegro skin test (MST), histopathological examination, and the polymerase chain reaction (PCR). The performance of these methods varies in a considerable proportion of patients. After the standardization of an immunoenzymatic test (ELISA) for the detection of IgG in the serum of patients with ACL using a crude Leishmania braziliensis antigen, the results obtained were compared to those of other tests routinely used for the diagnosis. The tests revealed the following sensitivity, when analyzed separately: 85% for ELISA IgG, 81% for PCR, 64.4% for MST, 58.1% for IIFR, and 34% for the presence of parasites in the biopsy. ELISA was positive in 75% of patients with ACL presenting a negative MST, in 84.8% of ACL patients with negative skin or mucous biopsies for the presence of the parasite, and in 100% of cases with a negative PCR. Thus, ELISA presented a higher sensitivity than the other tests and was useful as a complementary method for the diagnosis of ACL.

Natural antibodies in paracoccidioidomycosis

Recent attention has been focused on the natural antibodies as a component of natural immunity and as integral part of the idiotypic network. However, their functional role in different infections has rarely been studied. This work was undertaken to investigate the presence of natural antibodies in paracoccidioidomycosis (PCM). In addition, we analyzed anti-P. brasiliensis antibodies and their distribution in IgG subclasses in order to acquire better knowledge about the humoral immune response in this mycosis. Our findings show that the natural antibody response is not very much increased in PCM when compared with other parasite infections and this response is restricted to a few specificities, suggesting that P. brasiliensis moderately triggers CD5+ B cells. The anti-actin antibody was the main antibody specificity found in PCM. Specific antibodies to P. brasiliensis were mainly found in the IgG1 subclass in chronic patients of PCM.

Prevalence of paracoccidioidomycosis infection determined by sorologic test in donors' blood in the Northwest of Paraná, Brazil

In this paper the classical ELISA test was used, which is a highly sensitive method, with the objective of estimating the anti-Paracoccidioides brasiliensis antibodies level in healthy people, residing in supposedly endemic regions for PCM, Northwest Paran State, Brazil. The sera was analyzed from 680 blood donors' who were approved by the classical serologic screening by blood banks. Four cities were studied (Campo Mour o, Maring , Cianorte and Umuarama) with a sample of 170 donors each. The antigen used was Ag7days, diluted serum 1/400, anti human IgG-conjugate marked with peroxidase and revealed with O-Phenylenediamine. It was possible to detect 181 (27%) sera with antibody level above the cut off threshold. The percentage varied between the cities. These results allied with the favorable climatic and geographical conditions and also previous reports of PCM indigenous to the region, indicate that the Northwest of Paran is an endemic region for PCM.

Serological follow-up of patients with paracoccidioidomycosis treated with itraconazole using Dot-blot, ELISA and western-blot.

Twenty-seven mycologically proven cases of paracoccidioidomycosis (PCM) were treated with itraconazole (100-200 mg/day in month 1 and 100 mg/day until month 6-8) and evaluated clinically and serologically, up to 3.5 years post-therapy, using Dot-blot and ELISA for measuring the titers of IgG, IgA and IgM anti-P.brasiliensis antibodies and Western-blot for determining IgG, IgA and IgM antibodies against the antigen components of the fungus. Before treatment, 81.5% (Dot-blot) and 84% (ELISA) of the patients presented elevated IgG anti-P.brasiliensis antibody titers which dropped slightly with treatment. On the other hand, the percentages of pre-treatment high-titered sera for IgA and IgM anti-P.brasiliensis were lower (51.9% and 51.8%: Dot-blot; 16.5 and 36%: ELISA, respectively) but the titers tended to become negative more frequently with treatment. Prior to treatment, the percentages of positivity for IgG, IgA and IgM anti-P.brasiliensis antibodies in Western-blot were 96%, 20.8% and 41.6%, respectively. Antigens with molecular weights varying from 16-78 kDa, from 21-76 kDa and from 27-78 kDa were reactive for IgG, IgA and IgM antibodies, respectively. The most frequently reactive antigenic components had molecular weights of 27, 33 and 43 kDa for IgG, and 70 for IgA and IgM antibodies. During the period of study, the patients responded well to treatment. The present data confirm the diversity and complexity of the humoral response in PCM, and the importance of utilizing different serological tests to detect IgG, IgA and IgM anti-P. brasiliensis antibodies.

Antigen-specific immunosuppression in paracoccidioidomycosis.

To characterize the immune dysfunction associated with paracoccidioidomycosis, we studied the in vitro lymphocyte reactivity to phytohemagglutinin (PHA), pokeweed mitogen (PWM), a Candida albicans antigen (CMA), and a Paracoccidioides brasiliensis antigen (PbAg) in 32 patients with the acute and the chronic form of the disease before or during the initial phase of treatment and after clinical cure. We also studied, as controls, 30 healthy individuals, 15 of them immune to P. brasiliensis. Results showed a strong hyporesponsiveness to the PbAg while responses to mitogens and CMA were comparable with those of controls. Patients with the acute form of the disease (usually more severe) had more marked PbAg hyporesponsiveness than those with the chronic form. After patients' clinical cure, PbAg proliferative responses were similar to controls and greater than those seen before pretreatment. Changes in other parameters were also seen in the treated patients; skin test anergy to paracoccidioidin, high levels of anti-P. brasiliensis antibodies, leukocytosis, and eosinophilia. These changes were usually more intense in patients with the acute form of the disease. The post-treatment CD4+, CD8+, and total lymphocyte counts were similar to those of controls. Correlation between these parameters and the lymphoproliferative responses to the various stimuli was only found with PbAg: PbAg responses correlated inversely with eosinophil and anti P. brasiliensis antibody levels. Overall, our results demonstrate an antigen-specific-cellular immunity defect, which is reversible with treatment and possibly related to a T helper cell-2 pattern of immune response during active disease.