Cyclophosphamide effect on coccidioidomycosis in the rat

Coccidioidomycosis is a systemic mycosis, endemic in arid areas of the American continent. The rat was employed as an experimental host, since it had been shown to reproduce human lesions and present a chronic course of disease with granulomas mainly restricted to lungs. Given the influence of immunosuppressive therapy on the clinical course of human coccidioidomycosis, we studied the effect of cyclophosphamide (CY) in the experimental rat model. Accordingly, animals were inoculated with 400 Coccidioides immitis arthroconidia of the Acosta strain, by intracardiacal route. As single CY doses failed to alter the course of disease, three schedules were used: A) 4 daily doses of 20 mg/kg each, prior to C. immitis inoculation; B) 4 similar daily doses after infection; and C); 6 doses of 20 mg/kg each, given from day +1 to +4, then on days +8 and +9, post infection (pi), taking day 0 as the time of fungal inoculation. The first two schedules inhibited antibody formation up to day 28 pi, without modifying cellular response to coccidioidin as measured by foodpad swelling. Initially, there was greater fungal spread than in controls receiving C. immitis alone, which proved self-limiting in the latter. In contrast, schedule C led to 559r mortality, with both humoral and cellular response abrogation, accompanied by extensive C. immitis dissemination. Histology disclosed significant alterations, such as the persistence of primary infection sporangia, corresponding to the acute stage of coccidioidomycosis in the absence of granuloma development. Therefore, the observed depression in cellular immunity seems responsible for the lack of inflammatory reaction capable of restricting sporangia proliferation in tissues which, in turn, enhances pathogen spread and mortality rate.

Pulmonary cavities colonized by actinomycetes: report of six cases

Six cases of a cavitary pulmonary ball formed by Actinomycetes are reported. They were observed in the state of Bahia, Brazil. All patients complained of cough and hemoptysis and the pathological study showed bronchiectasis and small cavities in the lungs. The lesions contained micro-colonies of Actinomyces, identified by morphology, staining properties and culture in two cases (thioglycolate media). In the six patients the disease was limited to the lungs. In one patient grains were found, within micro-abscesses in the surrounding parenchyma. Probably the invasion occurred due to ulceration of bronchial mucosa that was covered by granulation tissue. The author suggests that as in nocardiosis actinomycosys may have an invasive form, a saprophytic one may and colonize pulmonary cavities.

Kinectics of the pulmonary phase of Schistosoma mansoni in mice treated with dexamethasone

In the experimental schistosomiasis mansoni glucocorticoids cause a reduction in the worm burden when administered in the week of infection or, the longest, at the next week. In order to determinate the probable(s) site(s) of reduction of the worm burden, mice were infected with cercariae of LE strain of S. mansoni and dexamethasone was administered daily (50 mg/kg, subcutaneously) starting 1 hour before infection until the eighth day. Mice were sacrificed daily starting on the third day after infection until the ninth day, and schistosomula from lungs were collected. Six weeks after infection, the remaining mice were sacrificed and perfused for adult worm recovery. Analysis of the results showed that the non-treated mice presented larger numbers of lung larvae than the treated ones, and this difference was also found later in the worm burden in the portal system. This difference may reflect the early death of larvae in treated animals, before or after reaching the lungs.

Experimental life cycle of Lagochilascaris minor Leiper, 1909

The life cycle of Lagochilascaris minor was studied using material collected from human lesion and applying the experimental model: rodents (mice, hamsters), and carnivorae (cats, dogs). In mice given infective eggs, orally, hatch of the third stage larvae was noted in the gut wall, with migration to liver, lungs, skeletal musculature and subcutaneous tissue becoming, soon after, encysted. In cats infected with skinned carcasses of mice (60 to 235 days of infection) it was observed: hatch of third stage larvae from the nodules (cysts) in the stomach, migration through the oesophagus, pharynx, trachea, related tissues (rhino-oropharynx), and cervical lymphonodes developing to the mature stage in any of these sites on days 9-20 post inoculation (P.I.). There was no parasite development up to the mature stage in cats inoculated orally with infective eggs, which indicates that the life cycle of this parasite includes an obligatory intermediate host. In one of the cats (fed carcass of infected mice) necropsied on day 43 P.I., it was observed the occurence of the self-infective cycle of L. minor in the lung tissues and in the cervical region which was characterized by the finding of eggs in different stages of development, third stage larvae and mature worms. It's believed that some component of the carnivorae gastrointestinal tracts may preclude the development of third stage larvae from L. minor eggs what explains the interruption of the life cycle in animals fed infective eggs. It's also pointed out the role of the intermediate host in the first stages of the life cycle of this helminth.

Schistosoma mansoni: evaluation of the activity of oxamniquine on schistosomules, at 24 hours after infection

Mice transcutaneously infected with about 400 cercariae were submitted to treatment with oxamniquine (400 mg/kg), 24 hours after infection. The recovery of schistosomules, at 4, 24, 48 and 72 hours and 35 days after treatment, showed the activity of the drug on the parasites, thus practically preventing their migration from the skin to the lungs. Worm recovery performed in the lungs (96 hours after treatment) showed recovery means of 0.6 worms/mouse in the treated group and 53.8 in the control group (untreated). The perfusion of the portal system carried out at 35 days after treatment clearly showed the elimination of all the parasites in the treated group, whereas a recovery mean of 144.7 worms/mouse was detected in the control group (untreated). These findings confirm the efficacy of oxamniquine at the skin phase of infection, and also show similarity with the immunization method that uses irradiated cercariae. The practical application of these findings in the medical clinic is discussed too

Flucytosine + fluconazole association in the treatment of a murine experimental model of cryptococcosis

The efficacy of flucytosine (5-FC) and fluconazole (FLU) association in the treatment of a murine experimental model of cryptococcosis, was evaluated. Seven groups of 10 Balb C mice each, were intraperitoneally inoculated with 10(7) cells of Cryptococcus neoformans. Six groups were allocated to receive 5-FC (300 mg/kg) and FLU (16 mg/ kg), either combined and individually, by daily gavage beginning 5 days after the infection, for 2 and 4 weeks. One group received distilled water and was used as control. The evaluation of treatments was based on: survival time; macroscopic examination of brain, lungs, liver and spleen at autopsy; presence of capsulated yeasts in microscopic examination of wet preparations of these organs and cultures of brain homogenate. 5-FC and FLU, individually or combined, significantly prolonged the survival time of the treated animals with respect to the control group (p<0.01). Animals treated for 4 weeks survived significantly longer than those treated for 2 weeks (p<0.01). No significant differences between the animals treated with 5-FC and FLU combined or separately were observed in the survival time and morphological parameters. The association of 5-FC and FLU does not seem to be more effective than 5-FC or FLU alone, in the treatment of this experimental model of cryptococcosis.

Central nervous system virion detection in acute measles: histopathological, ultrastructural and pathogenetic aspects

Histopathological and ultrastructural studies of 23 patients who died with clinical diagnosis of measles were carried out. In 12 cases viral nucleocapsids were searched by electron microscopy and detected in 100% of the cases in the lungs and in 50% of the cases in the central nervous system. They were mostly intranuclear. Histopathological changes associated to neurological alterations and the detection of virion are discussed in relation to acute and delayed clinical manifestations.

Human pulmonary dirofilariasis: a review

The authors presented a detailed summary of the geographical distribution, clinical and pathological aspects of human pulmonary dirofilariasis. Although benign, this zoonosis, of which Dirofilaria immitis is the major etiological agent, represents a medical problem since it produces symptoms which may be confused with neoplasia and thus may subject patients to unnecessary thoracic surgery. Of 229 cases cited in the literature, only 17 were reported in Brazil, despite the existence of highly favorable conditions for the transmission of this infection in man. Thus it may well be that this parasitic infection remains underdiagnosed. Finally, the importance of a differential diagnosis between dirofilariasis and pulmonary neoplasia is emphasized in cases where there is a solitary subpleural nodule ("coin lesion") present. In addition, the development and improvement of modern immunological diagnostic techniques are essential to distinguish this benign disease from other pathological conditions and thus avoid unneccessary surgery. These techniques may reveal the true prevalence of this parasitic infection in our environment.

PRESENCE OF ANTI-TOXOCARA ANTIBODIES IN CHILDREN SELECTED AT HOSPITAL UNIVERSITÁRIO, CAMPO GRANDE, MS, BRAZIL

Visceral Larva Migrans syndrome (VLM) results from the presence or migration of helminth larvae in humans, who nonetheless only play the role of paratenic hosts in the helminths' life cycle. In humans, VLM can be caused by larvae of various nematode species, chiefly those of the ascarid Toxocara canis, which can then be found at a variety of body sites, such as the liver, lungs, heart, and brain. Clinical and pathological manifestations depend primarily on larvae number and location, infection duration, reinfection occurrence, and host's immunological condition. Signs and symptoms may range from asymptomatic infection to severe disease. In humans, infection is acquired through ingestion of T. canis eggs present in soil, containing larvae in the infective stage7, 8, 9. Indeed, eggs of Toxocara sp. have been found in sandboxes in several public places in the city of Campo Grande, Mato Grosso do Sul state2. This study was carried out to detect the presence of anti-Toxocara antibodies in children attending the Pediatrics division of Hospital Universitário of Universidade Federal de Mato Grosso do Sul at Campo Grande, Brazil. Over the years 1992-94, 454 serum samples, obtained from children of 5.25 ± 3.28 years of mean age and selected at that hospital on the basis of eosinophil count greater than or equal to 1000/mm3 of blood, were tested for the presence of antibodies by means of the ELISA technique employing Toxocara canis larvae excretory-secretory antigens5. A high prevalence rate for toxocariasis (35.55%) was found, which was observed to be associated with eosinophil levels lower than those usually reported in literature. Furthermore, a higher frequency of positive serology among boys was also observed (13 cases in contrast to only 3 among girls), a result also reported by other authors

HANTAVIRUS PULMONARY SYNDROME (HPS) IN GUARIBA, SP, BRAZIL: REPORT OF 2 CASES

Human infections caused by a hantavirus were reported in different regions of the State of São Paulo (SP), Brazil during the first six months of 1998. Two cases of fatal pulmonary syndrome occurred in May of 1998 in the City of Guariba, located in the Northeastern Region of SP. Both patients worked in a corn storage barn infested by rodents. These patients, after 2 or 3 days of non-specific febrile illness, developed a severe interstitial pneumonia spreading widely in both lungs, causing respiratory failure and death. At autopsy both patients showed lung interstitial edema with immunoblast-like mononuclear cell infiltrates, consistent with a viral etiology. Hantavirus infection was diagnosed by ELISA in both cases and by RT-PCR in one of the patients. Aspects of the clinical presentation, physiopathology and differential diagnosis of Hantavirus Pulmonary Syndrome are discussed.

Hantavirus pulmonary syndrome in Brazil: clinical aspects of three new cases

Hantavirus pulmonary syndrome (HPS) has been recognized recently in Brazil, where 28 cases have been reported as of September 1999. We report here the clinical and laboratory findings of three cases whose diagnoses were confirmed serologically. All the patients were adults who presented a febrile illness with respiratory symptoms that progressed to respiratory failure that required artificial ventilation in two of them. Laboratory findings were most of the time consistent with those reported in the United States in patients infected with the Sin Nombre virus, and included elevated hematocrit and thrombocytopenia; presence of atypical lymphocytes was observed in one patient. The chest radiological findings observed in all the patients were bilateral, diffuse, reticulonodular infiltrates. Two patients died. Histopathological examination of the lungs of these patients revealed interstitial and alveolar edema, alveolar hemorrhage, and mild interstitial pneumonia characterized by infiltrate of immunoblasts and mononuclear cells. In the epidemiologic investigation of one of the cases, serologic (ELISA) tests were positive in 3 (25%) out of 12 individuals who shared the same environmental exposure. HPS should be included in the differential diagnosis of interstitial pneumonia progressing to acute respiratory failure.

Fibrotic sequelae in pulmonary paracoccidioidomycosis: histopathological aspects in BALB/c mice infected with viable and non-viable Paracoccidioides brasiliensis propagules

Patients with paracoccidioidomycosis often present pulmonary fibrosis and exhibit important respiratory limitations. Based on an already established animal model, the contribution of viable and non-viable P. brasiliensis propagules to the development of fibrosis was investigated. BALB/c male mice, 4-6 weeks old were inoculated intranasally either with 4x10(6 )viable conidia (Group I), or 6.5x10(6) fragmented yeast cells (Group II). Control animals received PBS. Six mice per period were sacrificed at 24, 48, 72h (initial) and 1, 2, 4, 8, 12 and 16 weeks post-challenge (late). Paraffin embedded lungs were sectioned and stained with H&E, trichromic (Masson), reticulin and Grocott´s. During the initial period PMNs influx was important in both groups and acute inflammation involving 34% to 45% of the lungs was noticed. Later on, mononuclear cells predominated. In group I, the inflammation progressed and granulomas were formed and by the 12th week they fussed and became loose. Thick collagen I fibers were observed in 66.6% and 83.3% of the animals at 8 and 12 weeks, respectively. Collagen III, thick fibers became apparent in some animals at 4weeks and by 12 weeks, 83% of them exhibited alterations in the organization and thickness of these elements. In group II mice, this pattern was different with stepwise decrease in the number of inflammatory foci and lack of granulomas. Although initially most animals in this group had minor alterations in thin collagen I fibers, they disappeared by the 4th week. Results indicate that tissue response to fragmented yeast cells was transitory while viable conidia evoked a progressive inflammatory reaction leading to granuloma formation and to excess production and/or disarrangement of collagens I and III; the latter led to fibrosis.

Infective endocarditis (IE) first diagnosed at autopsy: analysis of 31 cases in Ribeirão Preto, Brazil

Thirty one infective endocarditis (IE) fatal cases whose diagnosis was first obtained at autopsy were studied. The clinical data of these patients (Group 1) showed significant differences compared to other 141 IE cases (Group 2). The average age of 53 years in Group 1 patients was 18 years higher than that of Group 2. The Group 1 patients had a low frequency of IE predisposing heart disease. Both patient groups presented fever (about 87%), but a significant low frequency of cardiac murmur (25.8%) was observed in Group 1 patients and echocardiography tests were performed in only 16.1%, suggesting that IE diagnosis was not suspected. Likewise, although most Group 1 patients appeared with severe acute illness, they did not present the classic IE clinical presentation. Blood cultures were performed in only 64.5% of the Group 1 patients. However, bacteria were isolated in 70% of these blood cultures and Staphylococcus aureus was isolated in 71.4%. The bacteria attacked mitral and aortic valves. Complications such as embolizations and cardiac failure occurred in almost half of the cases and they also presented with infections of the lungs, urinary tract, and central nervous system. Medical procedures were performed in practically all fatal cases whose diagnosis was first obtained at autopsy. Sepsis occurred in about half of the patients and it was followed by shock in more than 25%. This form of IE must be suspected in mature and in old febrile hospitalized patients having infection predisposing diseases, embolization, and suffering medical procedures.

A morphological protocol and guide-list on uterine cervix cytology associated to Papillomavirus infection

The present study was designed to further assess the validity of the cytological description of morphological lesions said to be related to Papillomavirus (HPV) infections in senior women. The casuistic comprised 196 cervical smears from a group of women with no clinical or morphological evidence of neoplasia, collected simultaneously with samples submitted to detection of HPV DNA by PCR in a previous study. Three experienced cytologists studied each slide in two different conditions, with an interval of 20 months between them. The first approach was performed under routine laboratory standards, whereas the second was guided by a list of 16 well-defined parameters indicative of HPV-related cytological lesions. When suspicious cases of HPV-related alterations were grouped with positive cases, they showed on average: sensitivity of 25.5%, specificity of 84.4% and positive predictive value (PPV) of 26.8%. When suspicious cases were grouped with negative cases, sensitivity decreased, whereas specificity and PPV increased, as expected. In the second reading, which followed a "guide-list", a decrease in sensitivity was observed, contrasting with a sharp increase of positive predictive value. Among the 16 cytomorphological criteria tested, "koilocytosis", "mild koilocytosis" and "condylomatous parabasal cells" yielded the best predictive value for HPV DNA detection by PCR. In conclusion, despite the low sensitivity, cytopathologic assessment of cervico-vaginal smears leads to a highly specific diagnosis of HPV infection in menopausal women, with PPV of 91.0% when directed by a guide-list of well-defined morphologic criteria.

Virulence profile of ten Paracoccidioides brasiliensis isolates: association with morphologic and genetic patterns

Ten isolates of Paracoccidioides brasiliensis were examined for differences in virulence in outbred mice intravenously inoculated with the fungus, associated with mycelial morphology, and genetic patterns measured by random amplified polymorphic DNA (RAPD). Virulence was evaluated by viable yeast cell recovery from lungs and demonstration of histopathologic lesions in different organs. The results showed that the isolates presented four virulence degrees: high virulence, intermediate, low and non-virulence. RAPD clustered the isolates studied in two main groups with 56% of genetic similarity. Strains with low virulence, Pb265 or the non-virulent, Pb192, showed glabrous/cerebriform morphology and high genetic similarity (98.7%) when compared to the other isolates studied. The same was observed with Bt79 and Bt83 that shared 96% genetic similarity, cottony colonies and high virulence. The RAPD technique could only discriminate P. brasiliensis isolates according to glabrous/cerebriform or cottony colonies, and also high from low virulence strains. Isolates with intermediate virulence such as Pb18, Pb18B6, Bt32 and Bt56 showed variability in their similarity coefficient suggesting that RAPD was able to detect genetic variability in this fungal specie. Virulence profile of P. brasiliensis demonstrated that both mycelial morphologic extreme phenotypes may be associated with fungal virulence and their in vitro subculture time. Thus, RAPD technique analysis employed in association with virulence, morphologic and immunologic aspects might prove adequate to detect differences between P. brasiliensis isolates.