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. [1] (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.

Specific components found in circulating immune complexes (CIC) in paracoccidioidomycosis

Circulating immune complexes (CIC) from 15 paracoccidioidomycosis (PCM) patient sera and from 20 healthy control sera were analysed. After CIC precipitation, solubilization and acid treatment, only a little reactivity to P. brasiliensis antigens was found in the free antibodies from PCM-CIC. This result has suggested that there were antibodies with a high affinity bound to fungus components. Dissociated CIC were fractionated in a column of Sephacryl S300 and the fractions that probably contained antigens were pooled and applied to an affinity column, prepared with mouse anti-gp43 monoclonal antibody. Using ECL-Western blotting assay two polypeptide with apparent mass of 43 and 62 kDa were found.

Paracoccidioides brasiliensis antigen batches from the same isolate show immunological and biochemical differences

We investigated the occurrence of antigenic and biochemical variability among Paracoccidioides brasiliensis antigen batches prepared according to the same protocol. Initially (experiment 1), we analyzed two antigen lots of two human isolates (Bt1 and Bt2), cultured in two media (PYG: bactopeptone, yeast extract, glucose; MMM: McVeigh and Morton medium) in SDS-PAGE and in two immunological tests (immunodiffusion-ID and footpad swelling test-FPT). Afterwards (experiment 2), we compared the antigenic profile of three antigen hatches from three human isolates (Bt1, Bt2 and Bt3) by two-dimensional immunoelectrophoresis (2 D-IEP) against a reference system for P. brasiliensis antigens. In experiment 1, there were important intra- and inter-strain antigenic differences between batches of the fungal isolates cultured on both media. The block titration of the antigen batches for the immunological tests revealed correlation between protein concentration and biological activity in ID and no correlation in FPT. In experiment 2, the reference system for P brasiliensis showed 26 antigen peaks. There were important differences between batches prepared from the same isolate and between batches from different isolates. Our data suggested the occurrence of instability in the synthesis of antigenic components by a same P. brasiliensis isolate, under controlled incubation conditions.

Paracoccidioides brasiliensis-gp43 used as paracoccidioidin

A purified glycoprotein of 43 000 daltons from Paracoccidioides brasiliensis (gp43) was tested as paracoccidioidin in delayed-type hypersensitivity (DTH) tests in both experimental animals (guinea pig and mice) and patients with paracoccidioidomycosis (PCM). The gp43 paracoccidioidin was compared with the traditional Fava Netto antigen (AgFN). In guinea pigs, the intradermal injection of 2 μg of gp43 showed a similar response to those obtained with AgFN, showing in histological sections a population of lymphoid cells that participate in DTH. In mice, gp43 at a dose of 3.75μg showed positive DTH response. The use of gp43 as paracoccidioidin in humans showed that this molecule can be used to evaluate the DTH response in patients with PCM. Of 25 PCM patients studied, 48% were positive to gp43 while only 28% were positive to AgFN; 12 PCM patients were completely anergic to both antigens. Considering only those 13 PCM patients who were responsive to gp43 and/or to AgFN, 92.3% reacted against gp43 and 53.8% reacted against AgFN (P < 0.05). Gp43 skin test responses (13.67 ± 9.56 mm) were significantly larger than those obtained with AgFN (8.43 ±3.69 mm). Immunohistochemical study of the human skin showed a perivascular inflammatory response constituted predominantly by T lymphocytes, macrophages and polymorphonuclear leukocytes. © 1996 ISHAM.

The deep mycoses in HIV infection

The deep mycoses are uncommon infections, usually acquired from the inhalation or ingestion of fungal spores, sometimes from the soil in areas of endemicity, such as in the Americas and south-east Asia, or from decaying vegetable matter. They are also seen in immunocompromised persons and, increasingly, in HIV-infected persons. Respiratory involvement is frequent, with granuloma formation, and mucocutaneous involvement may be seen. Oral lesions of the deep mycoses are typically chronic but non-specific, though nodular or ulcerative appearances are common. Person-to-person transmission is rare. In HIV disease, the most common orofacial involvement of deep mycoses has been in histoplasmosis, cryptococcosis, aspergillosis and zygomycosis. Diagnosis is usually confirmed by lesional biopsy although culture may also be valuable. Treatment is with amphotericin or an azole.

Northwestern overthrusting and related lateral escape during the Brasiliano orogeny north of the Patos lineament, Borborema province, northeast Brazil

The Borborema province (BP) of northeastern Brazil, located between the São Luís and São Francisco cratons, represents a branching system of Precambrian orogens of the South American platform. It is composed of segments of Archean and Proterozoic crust that were deformed by the convergence of the West African and São Francisco-Congo cratons during assembly of the Brasiliano collage (650 to 500 Ma), a period of intense orogenic activity considered to be the strongest and most pervasive tectonic event that affected the Precambrian of the South American platform. The tectonic and kinematic history of the Brasiliano/Panafrican orogeny is fundamental for reconstructing South American and African Precambrian geology. The correlation between Neoproterozoic tectonic processes occurring in both continents should use structural elements, of regional or local character, with identical kinematic and metamorphic conditions manifested in both basement and supracrustal units. North of the Patos shear zone, subhorizontal Brasiliano thrusts (0.65 to 0.58 Ga) affected the basement and the supracrustal Seridó belt with such related regional D1/D2 structures as foliation, lineation, isoclinal folds, and related metamorphism. Overprinting the previous structures, regional folding with a vertical S3 foliation and an associated strike-slip shear zone were developed (0.58 to 0.52 Ga). The metamorphism is similar for all deformation phases, ranging from upper-greenschist to amphibolite facies with mineral assemblages including biolite and garnet throughout the Seridó fold belt. We propose, on the basis of deformational and kinematic reconstructions, that the structural evolution of the Seridó fold belt was characterized by transition from a syn-collisional to a strike-slip regime. The transition between regimes occurred, progressively or instantaneously, by the switching of the maximum and intermediary strain axes of the strain ellipsoid. The entire tectonic history can be related to a frontal or oblique collision and lateral escape tectonics, with local, syn-collisional transpression and transtension. The Patos shear zone represents a final vertical shearing, juxtaposing different terranes of the northern and southern Borborema province.

IgG, IgM and IgA antibody response for the diagnosis and follow-up of paracoccidioidomycosis: Comparison of counterimmunoelectrophoresis and complement fixation

IgG, IgM and IgA antibodies to GP43 (glycoprotein fraction of Paracoccidioides brasiliensis) were measured by ELISA in 63 samples from 23 patients with paracoccidioidomycosis before and twice after chemotherapy was started. Antibodies against P. brasiliensis were detected by indirect immunofluorescence (IF) (IgG, IgM and IgA isotypes), counterimmunoelectrophoresis (CIE) and complement fixation. Two control groups composed of 19 healthy individuals and 12 patients with other diseases (six with histoplasmosis, three with tuberculosis and three with other mycoses). The highest efficiency percentages were found with IgG and IgA- ELISA (100%), IgG-IF (96.2%), CIE (94.4%) and the lowest with CF (75.9%). Highest positive and negative predictive values (100%) were observed for IgG and IgA ELISA. IgG and IgM-ELISA antibodies are more often found in patients with acute than chronic disease (P = 0.01). Four to six months after treatment follow-up showed decreased levels of IgG and IgM-ELISA for acute cases and decreased titres of CIE for chronic cases in relation to pretreatment levels. This study suggests that IgG-ELISA anti-GP43 represents a good marker to monitor clinical response to therapy.

Penfigo foliaceo no Hospital das Clinicas da Faculdade de Medicina de Botucatu, SP

BACKGROUND - Pemphigus foliaceus is an autoimmune bullous disease that is endemic in Brazil and in other South American countries such as Paraguay, Bolivia, Peru and Colombia. OBJECTIVES - To compare the observed clinical and epidemiological data with those referred in the literature. METHOD - Retrospective study in patients with pemphigus foliaceus admitted to and treated at University Hospital of the School of Medicine of Botucatu, SP, Brazil, from 1976 to 1993, analyzing the following variables: age, sex, race, origin and onset of disease. RESULTS - From 1976 to 1993, 63 patients were treated, with an average of 3.7 cases/year. Patients within the age range of 20 to 60 years old were the most affected. Most of the patients were white, females being slightly more affected than males. The majority of male patients were rural workers. Some towns presented higher incidence of cases. Most of the patients from rural areas became sick mainly in summer and in autumn. Occurrence of similar cases was observed in blood-related family members and neighbors. CONCLUSIONS - Clinical and epidemiological data were similar to those referred in the literature.

Crystallization and X-ray diffraction data analysis of oxyhemoglobin-I from the catfish Liposarcus anisitsi (pisces)

Hemoglobin remains, despite the enormous amount of research involving this molecule, as a prototype for allosteric models and new conformations. Functional studies carried out on Hemoglobin-I from the South-American Catfish Liposarcus anisitsi [1] suggest the existence of conformational states beyond those already described for human hemoglobin, which could be confirmed crystallographically. The present work represents the initial steps towards that goal.

Detection of circulating Paracoccidioides brasiliensis antigen in urine of paracoccidioidomycosis patients before and during treatment

For the diagnosis and follow-up of paracoccidioidomycosis patients undergoing therapy, we evaluated two methods (immunoblotting and competition enzyme immunoassay) for the detection of circulating antigen in urine samples. A complex pattern of reactivity was observed in the immunoblot test. Bands of 70 and 43 kDa were detected more often in urine samples from patients before treatment. The immunoblot method detected gp43 and gp70 separately or concurrently in 11 (91.7%) of 12 patients, whereas the competition enzyme immunoassay detected antigenuria in 9 (75%) of 12 patients. Both tests appeared to be highly specific (100%), considering that neither fraction detectable by immunoblotting was present in urine samples from the control group. gp43 remained present in the urine samples collected during the treatment period, with a significant decrease in reactivity in samples collected during clinical recovery and increased reactivity in samples collected during relapses. Reactivity of some bands was also detected in urine specimens from patients with 'apparent cure.' The detection of Paracoccidioides brasiliensis antigens in urine appears to be a promising method for diagnosing infection, for evaluating the efficacy of treatment, and for detecting relapse.

Paracoccidioidomycosis: Survey and clinical aspects from the Department of Dermatology of the School of Medicine of Botucatu (Sao Paulo - Brazil)

Background - Paracoccidioidomycosis is the most frequent among the systemic mycoses in Brazil. Objective: To study the cases of paracoccidioidomycosis diagnosed from 1976 to 1996 at the Department of Dermatology of the School of Medicine of Botucatu. Methods - Descriptive study obtained from specific protocols comprising anamnesis, clinical- dermatological-laboratorial examination, treatment and follow-up of the patients. Results - Paracoccidioidomycosis was diagnosed in 1.04% of the dermatologic outpatients assisted from 1976 to 1996. In 315 cases, 89.8% were male, 61.9% were over forty and 53.7% were rural workers. The complaints were related more to oropharyngolaryngeal (53.6%) or cutaneous (23.8%) lesions or adenopathy (10.2%). Seventy patients had already been treated in other health services. The chronic multifocal clinical from was the most common: 80.6% of the cases, followed by the acute-subacute (juvenile type) 15.5%. The disease was pulmonary in 80.0%, oropharyngolaryngeal in 69.2% and cutaneous in 45.7%. Th treatment: Amphotericin B in 146 patients, Ketoconazole - 88, Itraconazole - 56, Sulfonamide derivatives - 146 and Terbinafine in 3. The lethality index resulting from the disease or its treatment was 2.0%. Conclusions - The high number of cases, showing the regional relevance of the disease and the high percentage of tegumentary complaints and the high number of relapsing are remarkable.

Paracoccidioidomicose

The aspects related to the epidemiology, pathogenesis, clinical classification, clinical manifestations and treatment of Paracoccidioidomycosis, are presented. The disease is caused by Paracoccidioides brasiliensis and is the most frequent and the most important systemic mycosis in the South America, specifically in Brazil, Argentina, Colombia and Venezuela. Because of the high frequency of cutaneous-mucosal lesions presented by the patients, the potential severity of the disease and the socio-economic consequences to the patient, it is very important for a drematologist to be qualified to precisely diagnose the disease and render proper care.

Energetic cost of predation: Aerobic metabolism during prey ingestion by juvenile rattlesnakes, Crotalus durissus

We investigated the cost of prey ingestion in the South American rattlesnake, Crotalus durissus, to see if the capacity to generate energy aerobically could be a constraint on the size of the prey that can be ingested. To accomplish this goal, we measured time and aerobic metabolism (inferred from oxygen consumption) of juvenile C. durissus ingesting prey ranging from 10 to 50% of their own body mass. Time needed for prey ingestion increased with prey size, with prey representing 10 and 20% of snake size being ingested with the same effort. Whole animal rates of oxygen consumption increased linearly with prey size, but at a slower pace for snakes ingesting prey larger than 30% of their body mass. Aerobic factorial power input necessary for prey ingestion increased with prey size, and for snakes ingesting prey representing 50% of their body mass it equaled the aerobic factorial scope for exercise. For the maximum prey size tested, the aerobic derived energy necessary for prey ingestion represented 0.02% of the total energy content of the prey. Within the prey size range we studied, the cost of ingestion did not constitute any constraint on the size of the prey that can be ingested. These constraints are set by morphological (gape size), ecological (predation risk), and, probably, by physiological parameters, as suggested by the tendency of V̇O2 during ingestion to increase at a slower pace at relative larger prey sizes.

Re-evaluation of antibiotic and mercury resistance in Escherichia coli populations isolated in 1978 from Amazonian rubber tree tappers and Indians

A study was carried out to assess the stability of antimicrobial susceptibility of wild isolates upon long-term storage using fifty-three Escherichia coli strains isolated in 1978 from feces of healthy children from the Amazon region in Brazil, exposed to low levels of antimicrobial agents, and examined for resistance to mercury and four antibiotics. All of the strains were kept in Lignieres medium at room temperature and were transferred to fresh media four times during this period. Thirty-five out of the 53 strains analyzed in 1978 were viable. Upon recovery, antibiotic and mercury resistance was estimated. All of the 35 strains maintained their original phenotype in a stable fashion, except for one multiresistant strain which became susceptible to kanamycin. Fifty-four percent of the strains exhibited a resistance phenotype, among which 47% had conjugative plasmids.