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Four stains for the detection of Pneumocystis carinii in respiratory specimens were compared for sensitivity, specificity, preparation time, and ease of interpretation. One hundred specimens were collected. Of these, 50 were induced sputum specimens and 50 were bronchoalveolar lavage fluid. All specimens were stained with Diff-Quik (DQ) (a modified Giemsa stain), a quick silver stain, and direct and indirect immunofluorescence stains. A positive specimen was defined as any smear positive by two or more of the methods. Fifty-eight percent of specimens were positive. Seventy-four percent of the sputum specimens and 42% of the bronchoalveolar lavages were positive. The sensitivities for detection of P. carinii in sputum were 92% with silver stain, 97% with direct immunofluorescence assay (DFA), 97% with indirect immunofluorescence assay (IFA), and 92% with DQ. The sensitivities for detection in bronchoalveolar lavage were 86% with silver stain, 90% with DFA, 86% with IFA, and 81% with DQ. Preparation times varied from 90 min for the silver stain and IFA to 3 min for DQ. Costs of the tests varied from $1.50 per slide for DQ to $10.00 per slide for the silver stain and DFA. Reading times varied from 10 to 30 min for the silver stain and DQ to less than 5 min for the immunofluorescence assays. We conclude that all of these tests are viable options for the clinical laboratory, and the choice will be influenced by factors such as clinical volume, ability to batch specimens, and expertise of technological support. A reasonable option may be to use the quick and inexpensive DQ as a screening test and to confirm negative smears with a more sensitive assay.

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Two patients with AIDS presented with pulmonary toxoplasmosis. Diagnosis was achieved by the visualization, in Giemsa-stained smears of bronchoalveolar lavage fluid, of tachyzoites of Toxoplasma gondii that were extracellular and in the process of invading or within pneumocytes. The intracellular localization of tachyzoites facilitated diagnosis by obviating potential confusion of extracellular tachyzoites with cellular debris or platelets.

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Disseminated toxoplasmosis is a life-threatening infection in transplant recipients, which results either from reactivation of latent infection or from organ-transmitted primary infection. Preventive measures and diagnostic screening methods differ between countries and are related to the seroprevalence of Toxoplasma spp. in the general population. Here we report a case of disseminated toxoplasmosis in a heart transplant recipient with previous immunity that occurred after cotrimoxazole prophylaxis for the prevention of Pneumocystis jirovecii pneumonia was stopped. Quantitative PCR proved useful for the diagnosis and monitoring of Toxoplasma infection. Decreasing parasitic burdens in sequential samples of cerebrospinal fluid, blood, and bronchoalveolar lavage fluid correlated with a favorable outcome and allowed modulation of the immunosuppressive drug regimen. The duration of anti-Toxoplasma treatment and the need for maintenance prophylaxis are discussed, as well as prophylaxis for solid-organ transplant recipients. Although a rare event in heart transplant recipients, Toxoplasma reactivation must be investigated promptly, since early treatment improves the prognosis.

The most common presentation of symptomatic postnatally acquired toxoplasmosis in immunocompetent patients is painless cervical adenopathy. Acute visceral manifestations are associated in rare cases. We report 16 cases of severe primary toxoplasmosis diagnosed in French Guiana during a 6.5-year period. All of the subjects were immunocompetent adults hospitalized with clinical presentations consisting of a marked, nonspecific infectious syndrome accompanied by an altered general status with at least one visceral localization, mainly pulmonary involvement (14 cases). Acute toxoplasmosis was diagnosed according to the results of serological tests suggestive of recent primary infection and the absence of an alternative etiology. Recovery was rapid following specific antitoxoplasmosis treatment. Thirteen of the 16 patients had consumed game in the 2 weeks before the onset of the symptoms, and in eight cases the game was considered to have been undercooked. Toxoplasma strains, which were virulent in mice, were isolated from three patients. Microsatellite analysis showed that all of these isolates exhibited an atypical multilocus genotype, with one allele found only for isolates of this region.

A new serologic test for antibodies to Toxoplasma is described, which is based upon inhibition of specific staining with fluorescent antibody. In performing the test, a mixture of the test serum and known fluorescein-labelled antiserum is added to a dried smear of toxoplasms for 1 hour at 37°C. The smear is then rinsed and examined with a fluorescence microscope. Reduction in the brightness of fluorescence, as compared to that of a negative control slide, indicates the presence of antibody in the test serum. A comparison of the results of this test with those of the methylene blue dye test showed a strong parallelism between the two sets of results. On the other hand, the complement-fixation test for toxoplasmosis did not yield nearly as many positives as the inhibition test. The specificity of the new test was studied by comparing it with dye test results and clinical histories in human patients, and by testing a group of animals immunized with a variety of non-Toxoplasma antigens. No evidence of cross-reactions was obtained in the latter series. Some advantages and disadvantages of the inhibition test are discussed.

PCR was used to evaluate the occurrence of Toxoplasma gondii parasitemia by detection of the B1 gene in blood samples in two groups of immunosuppressed patients (148 subjects) suspected of having cerebral or extracerebral infection, respectively. Group I consisted of 52 patients with AIDS with suspected cerebral toxoplasmosis. The diagnosis was clinically proven in 15 cases. Parasitemia was detected by PCR in only two of these patients (13.3%), both showing evidence of disseminated infection. Group II consisted of 96 immunocompromised patients, either with AIDS or receiving iatrogenic immunosuppressive therapy. Of these patients, 65 (34 with AIDS and 31 others) showed abnormalities only in chest radiography and were first screened for the presence of Toxoplasma DNA in bronchoalveolar lavage fluid. Blood was then analyzed when the parasite was detected in the bronchoalveolar lavage fluid. The remaining 31 subjects (22 with AIDS and 9 others) were suspected of having extracerebral, pulmonary, or disseminated toxoplasmosis, and blood was studied directly in these cases. Among the nine patients with clinically diagnosed extracerebral infection in group II, the parasite was detected by PCR in the blood of five patients (55.5%), all having pulmonary toxoplasmosis. If all patients with clinical manifestations of extracerebral toxoplasmosis (from both groups) who had not received antitoxoplasma therapy when the samples were collected are considered, PCR detected parasitemia in seven of the nine cases (77.8%). The present study indicates that examination of blood by PCR may be valuable in cases of extracerebral toxoplasmosis because of the disseminated nature of the disease. Since most cases of cerebral toxoplasmosis result from the local reactivation of latent brain cysts, detection of parasitemia by PCR is useful only in cases associated with severe cerebral infection or dissemination of this disease.

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Pulmonary Kaposi's sarcoma may contribute to respiratory dysfunction in patients with acquired immune deficiency syndrome (AIDS) and features of pneumonitis. Opportunistic infections are readily recognised in endoscopic material, but pulmonary Kaposi's sarcoma is easily missed, so that patients are deprived of specific treatment. The clinical and pathological findings from nine cases of pulmonary Kaposi's sarcoma have been reviewed; these were found among 84 patients with AIDS and pneumonitis undergoing fibreoptic bronchoscopy and bronchoalveolar lavage. Diagnosis was established before death in eight patients (in five by bronchial biopsy and in three by open lung biopsy). Examination of lavage fluid showed alveolar haemorrhage in six patients. It is concluded that: (1) fibreoptic bronchoscopy may be useful in the diagnosis of endobronchial lesions of Kaposi's sarcoma; (2) alveolar haemorrhage in patients with AIDS is suggestive of pulmonary Kaposi's sarcoma. Factors that may cause difficulties in diagnosis include the focal nature of some lesions and the pleural or parenchymatous location of others. In addition, in the lung as in the skin, the early stages of Kaposi's sarcoma resemble granulation tissue. Such lesions are far more difficult to recognise than is the late nodular stage.

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AIMS: For the diagnosis of pulmonary alveolar proteinosis from bronchoalveolar lavage specimens it is normally necessary to make an ultrastructural examination. However, this is thought to be impractical for bronchoalveolar lavage specimens that have been routinely fixed in ethanol. In the present study, bronchoalveolar lavage cytology smears on slide glasses were examined directly ultrastructurally to make a diagnosis of pulmonary alveolar proteinosis. METHODS: Bronchoalveolar lavage smears from three pulmonary alveolar proteinosis patients were stained with Papanicolaou and periodic acid-Schiff (PAS) for identification of amorphous globular structures. Subsequently, they were refixed with glutaraldehyde and osmium tetroxide, and embedded in epoxy resin. Ultrathin sections were cut and examined ultrastructurally. RESULTS: Papanicolaou stained specimens from pulmonary alveolar proteinosis patients contained scattered amorphous or granular globules, 20-50 microns in diameter, which were PAS positive. Ultrastructural examination of the globules revealed multilamellated structures, characteristic of pulmonary alveolar proteinosis, in all cases. CONCLUSIONS: In general, it is thought that the morphological diagnosis of pulmonary alveolar proteinosis from bronchoalveolar lavage specimens requires both cytological and ultrastructural examination. However, the amorphous globules evident on cytology smears proved to contain multilamellated structures so that they can themselves be used as diagnostic evidence.

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Optimum growth conditions for human cytomegalovirus (HCMV) include the use of subconfluent, actively growing cultures of human embryonic fibroblasts. Many clinical virology laboratories, however, use tissue culture cells from commercial sources. These cells are usually confluent, static cultures that tend to be less sensitive to viral infection. To determine whether dimethyl sulfoxide (DMSO) or dexamethasone (DEX), which are known enhancers of HCMV, facilitates the detection of the virus in confluent cells, we tested both HCMV AD169 and a number of clinical specimens suspected to contain HCMV on MRC-5 cells in both shell vials and conventional tube cultures. We found that, in the shell vial test, treatment of the cultures with either DMSO or DEX before and after inoculation increased the number of cells staining positive by three- to sixfold compared with untreated controls. Best results were obtained by pretreating the cultures with DEX alone and by treating the cultures with a combination of DEX and 1% DMSO postinfection. In the conventional MRC-5 culture tubes, treatment with the reagents resulted in the more rapid appearance of cytopathic effect and a more extensive infection of the cell sheet. The experimental findings indicate that the enhancing effect of DEX is attributable mainly to the increased production of a cellular mRNA during the period preceding viral infection.

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Since the first report of human infection with Pneumocystis carinii in 1942, cases of pneumonia due to this opportunistic pathogen have become increasingly common. Animal studies and clinical observations show that a significant depletion or dysfunction of T helper lymphocytes predisposes to clinical disease. Individuals with damaged T helper cells secondary to malignancies (eg, Hodgkin’s lymphoma), drugs (eg, cyclosporine, steroids), or certain infections (eg, human immunodeficiency virus) are at particular risk. Serological studies suggest that disease is most often secondary to the reactivation of an asymptomatic infection, usually acquired during childhood. Increasing shortness of breath, a nonproductive cough and hypoxia often preceded by several weeks of lethargy, fever and weight loss are the classical features of P carinii pneumonia in acquired immune deficiency syndrome. Bronchoalveolar lavage is usually the optimal diagnostic test. Immunofluorescent staining on liquified sputum induced by nebulized saline appears to be a promising and noninvasive test. Early empiric therapy with trimethoprim-sulphamethoxazole (trimethoprim 5 mg-sulphamethoxazole 25 mg/kg/day every 6 h) or intravenous pentamidine (4 mg/kg/day) for 21 days is usually effective, but infection is not eradicated, and clinical disease is likely to recur. Prophylaxis using aerosolized pentamidine reduces the risk of pulmonary disease but can predispose to extrapulmonary infection. Improved in vitro and in vivo models of human pneumocystis infection would significantly increase understanding of the molecular biology of the organism, the pathogenesis of disease, and the optimal therapeutic regimens.

Background

Department of Pathology was founded in 1960. With the establishment of the Institute for Pulmonary Diseases. Laboratories for histology, cytology, immunohistochemistry and autopsy unit are integral part of this department.

Patients and methods

In histopathologic laboratory over 10,000 endoscopical and surgical biopsies, with ex tempore analyzes annually, are technically prepared and processed by using standard as well as special stainings. Over 6000 samples per year obtained by exfoliative cytology: sputum, pleural, pericardial and abdominal effusions, aspiration cytology: transthoracic fine needle aspiration (FNA), and samples obtained during bronchoscopy: lavage, brushes and transbronchial fine needle aspiration biopsy (obtained during endobronchial ultrasound guided (EBUS) FNA) and bronchoalveolar lavages are processed in the laboratory for cytology. May Grunwald Giemsa and Papanicolaou stainings are used for all cytological specimens and in many cases cell blocks are prepared too for ancillary technics. Laboratory for immunohistochemistry disposes of 43 tumor markers for the diagnosis and differentiation of primary and secondary lung tumors, malignant mesothelioma, lymphoma and thymoma and annually performs over 300 analyzes. Over 200 autopsies per year are performed in the autopsy unit. Predominant field of work is thoracic pathology, but we are also dealing with cardiovascular, endocrine, gastrointestinal, hepatobiliary and gynecological pathology.

Results

Today The Department of Pathology employs 1 biologist, 6 laboratory technicians and 3 autopsy assistants as well as 2 pathologists, 3 cytopathologists and 1 resident. As the Institute for Pulmonary Diseases is university hospital all doctors, 4 PhD and 2 postgraduate students are engaged in the educational work. Teachers participate in undergraduate and postgraduate teaching at Medical Faculty in Novi Sad, Banja Luka and Foca (Serbian Republic). The Department of Pathology from the very beginning enforced specialization in Pathology and sub-specialization in Medical Cytology. In the cooperation with the Center for Continuing Education, several educational seminars in the field of pathology and cytology have been organized.

Conclusions

The future of this department is the automatization and standardization of working processes, control improvement, continuing education of all employees and greater engagement in the field of research. Introducing of genetic and molecular techniques for better diagnosis and individualized therapy is our task in the next few years.

Cerebral toxoplasmosis is an acquired immunodeficiency syndrome (AIDS)-related infection and is one of the causes of CNS mass lesions in AIDS. Toxoplasmosis is the most common cerebral mass lesion encountered in HIV-infected patients, and its incidence has increased markedly since the beginning of the AIDS epidemic. Cerebral toxoplasmosis is associated with high mortality and morbidity in patients with acquired immunocopromised state. We are reporting a case of cerebral toxoplasmosis presented with status epileptics and treated with cotrimoxazole. Refractory status epilepsy was controlled with intravenous levetiracetam, which has a unique drug profile.

Bronchoalveolar lavage and transbronchial biopsy have been used as adjuncts to the management of patients with pneumonia associated with the acquired immunodeficiency syndrome (AIDS) at the Middlesex Hospital and the experience gained and difficulties encountered in the first five cases are reported. Widely varying organisms were isolated from lavage aspirates, some of which may have been nasopharyngeal contaminants, and organisms cultured from the transbronchial biopsy specimens may offer a better guide to antimicrobial treatment. Pneumocystis carinii was found in two of the patients. In view of the potentially serious toxicity of high dose co-trimoxazole, continuation of this treatment may be inadvisable if Pneumocystis carinii is not identified by all available methods unless there are strong clinical grounds to suspect its presence.

Two prospective studies were undertaken to evaluate a commercial indirect fluorescent-antibody (IFA) stain for the detection of Pneumocystis carinii in respiratory specimens from individuals at risk for or with the acquired immunodeficiency syndrome. The first study compared IFA with Diff-Quik (DQ; a rapid Giemsa-like stain) for detecting P. carinii in 95 induced sputa obtained from 77 asymptomatic patients who had survived one previous episode of P. carinii pneumonia and who were being treated prophylactically with aerosolized pentamidine. Only one induced sputum specimen was found to contain P. carinii; organisms were detected by both stains. The second study compared the performance of the IFA stain versus DQ, modified toluidine blue O, and Gomori methenamine silver stains for detecting P. carinii in symptomatic individuals at risk for or with acquired immunodeficiency syndrome. Of 182 specimens examined, P. carinii was detected in 105 by one or more stains; the DQ stain detected 73 (70%), the modified toluidine blue O stain detected 75 (71%), the Gomori methenamine silver stain detected 76 (72%), and the IFA stain detected 95 (90%). The IFA stain was more sensitive (P less than 0.01) than the other traditional stains for detecting P. carinii; however, a subsequent clinical evaluation revealed that a subset of IFA-positive-only specimens were from patients whose clinical symptoms resolved without specific anti-P. carinii therapy.

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1. Eosinate of thionin gives very satisfactory staining for blood smears. It is easily prepared and dissolves readily in methylated spirits. 2. In the methods heretofore employed for making mixtures of eosin and methylene blue derivatives, the eosinates of methylene violet and methylene azure are present in very small quantities or are altogether absent. 3. Thionol and thionin are probably formed in methylene blue which has been long boiled with dilute alkalies and silver oxide. 4. Good stains of eosin and methylene blue derivatives can be obtained by a variety of manipulations. 5. Methylated spirits is more economical as a solvent for this stain and better adapted for the simple technique above described than is methyl alcohol. There is some evidence that the staining act is of a chemical nature.

The technique of immunoblotting for detecting soluble Pneumocystis carinii antigen(s) in bronchoalveolar lavage fluid specimens from patients with AIDS and Pneumocystis pneumonia was evaluated. A soluble 67 kilodalton polypeptide that was immunoreactive with an anti-P carinii monoclonal antibody (2G2) was found in the supernatants of 26 lavage samples from patients with pneumocystosis. Intact organisms in lavage sediments were detected by methenamine silver or immunofluorescence staining procedures. The diagnostic use of this technique was shown in four cases in which lavage sediments proved negative for intact Pneumocystis carinii organisms on first examination; 2G2 reactive soluble antigen, however, was identified in the immunoblots of the supernatants from the same samples. It is concluded that immunoblotting of bronchoalveolar lavage specimens using 2G2 monoclonal antibody as a detection probe may be a useful adjunct to the morphological demonstration of organisms by special staining procedures.

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By using a conventional tissue culture method as a standard, four shell vial centrifugation culture (SVC) formats were compared for herpes simplex virus (HSV) detection in 300 clinical samples. Both MRC-5 and primary rabbit kidney (PRK) cells were used in the conventional and SVC systems. In addition, both a direct monoclonal fluorescent antibody to HSV (MAb-FA; Syva Corporation, Palo Alto, Calif.) and an indirect HSV polyclonal antibody-horseradish peroxidase stain (poly-HRP; Difco Laboratories, Detroit, Mich.) were used to stain shell vials of both cell types. Conventional tubes were incubated for up to 7 days with daily examination for cytopathic effect, which was confirmed as HSV by staining with an MAb-FA. Shell vials were inoculated, centrifuged, incubated for 16 to 24 h, and stained directly with MAb-FA or indirectly with a poly-HRP stain. Of the 300 specimens examined, 82 (27%) were HSV positive by conventional tissue culture. PRK cells detected 81 (99%) positive specimens, compared with 74 (90%) specimens detected with MRC-5 cells. Of the 82 positive specimens by conventional culture, the SVC formats detected 68 by MRC-5 and MAb-FA, 74 by MRC-5 and poly-HRP, 64 by PRK and MAb-FA, and 77 by PRK and poly-HRP. Therefore, PRK stained by an indirect method with poly-HRP was the most sensitive of the SVC formats tested, detecting 94% of the positive specimens.

Monoclonal antibodies (Syva Co., Palo Alto, Calif.) were used for the detection and serotyping of herpes simplex virus (HSV) isolates by immunofluorescence 16 h after inoculation of MRC-5 monolayers in 3.7-ml shell vials and after low-speed centrifugation. A total of 119 specimens were inoculated into conventional tube cell cultures and shell vials. Of 98 specimens inoculated on the same day of receipt in the laboratory (fresh specimens), all 23 (23.5%) HSV-positive specimens were identified by serotype in 16 h in shell vials by immunofluorescence, whereas only 8 of 23 HSV-positive specimens (34.8%) produced cytopathic effects in conventional tube cell cultures in this time period. Similarly, of 21 original specimen extracts previously determined to be culture positive for HSV (stored specimens), all were detected and serotyped by the immunofluorescence test with monoclonal antibodies 16 h postinoculation compared with the recognition of only 8 of these isolates (38.1%) by cytopathic effect that soon. This technique of centrifugal inoculation of HSV in shell vials containing MRC-5 cells permitted detection of this virus in all positive specimens with serotype determination within 16 h postinoculation.

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A modified toluidine blue O staining technique for Pneumocystis carinii is described. An easily prepared sulfation reagent made with sulfuric and acetic acids was used. The stain can be employed for bronchoalveolar lavages and lung tissue touch preparations. Most background material was removed by the sulfation reagent, slides were generally easy to read, and time from receipt of a specimen to reporting of results was approximately 1 h. P. carinii cysts were more easily visualized with this stain than in slides stained with modified methylene blue, Gram, Gram-Weigert, and standard toluidine blue O procedures. The importance of certain procedural aspects of subsegmental bronchoalveolar lavage, by which most of the specimens were obtained, is also emphasized.

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The production of gamma interferon in acute acquired and congenital toxoplasmosis was studied. Gamma interferon was produced at significant titers (P less than 0.001) in the course of both congenital toxoplasmosis and acquired toxoplasmosis at an early stage of infection, when Toxoplasma gondii was multiplying. Its presence in fetal blood was correlated with the positive inoculation of fetal blood or amniotic fluid into mice (95%). The data suggest that the fetus is able to synthesize gamma interferon as early as week 21 of pregnancy. This test, easily and rapidly performed, could be included among those useful for diagnosing fetal toxoplasmic disease.

From January 1981 to January 1983 acquired immune deficiency syndrome (AIDS) was diagnosed in 90 patients admitted to Kings County Hospital-Downstate Medical Center. CNS involvement occurred in 18 patients of whom 12 had toxoplasmosis confirmed by biopsy or necropsy. Pathological specimens from these 12 patients were notable for a marked diminution or absence of cellular inflammation. Each patient had elevated serological studies for toxoplasma. AIDS presented with symptoms referable to CNS toxoplasma in eight patients. In the remaining four patients, toxoplasma was found late in the course of the illness. CT showed either ring enhancing lesions or solid nodules. The course was uniformly fatal, though patients treated continuously with pyrimethamine and sulfadiazine survived longer.

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BACKGROUND:

Smear-negative tuberculosis occurs more frequently in human immunodeficiency virus (HIV)-infected patients than in non-HIV-infected patients. Besides, there are substantial numbers of patients who cannot produce sputum, making the diagnosis of pulmonary tuberculosis (PTB) difficult.

AIMS:

To evaluate the relative yield of pre- and post-bronchoscopy sputum and bronchoalveolar lavage (BAL) in ‘sputum smear’-negative, HIV-positive patients.

SETTINGS:

A tertiary care referral hospital in Addis Ababa.

MATERIALS AND METHODS:

Acid-fast stain (AFS) using the concentration technique was done on 85 pre-bronchoscopy sputum and 120 BAL samples. Direct AFS was done on all BAL and 117 post-bronchoscopy sputum samples. Culture for Mycobacterium tuberculosis (MTB) was done for all sputa and BAL.

RESULTS:

MTB was isolated from 26 (21.7%), 23 (19.7%) and 13 (15.3%) of BAL, post- and pre-bronchoscopy sputum cultures respectively. AFS on pre-bronchoscopy sputum using concentration technique and direct AFS on BAL together detected 11 (41%) of the 27 culture-positive cases. In patients who could produce sputum, the sensitivity of pre-bronchoscopy sputum culture (13/85, 15.3%) was comparable to BAL (12/85, 14%) and post-bronchoscopy sputum (12/85, 14%). In patients who could not produce sputum, however, both BAL (12/35, 40%) and post-bronchoscopy sputum (12/32, 31.4%) detected significantly more patients than those who could produce sputum (P=0.002, P=0.028 respectively).

CONCLUSION:

In HIV-infected patients, AFS by concentration method on pre-bronchoscopy sputum and direct AFS on BAL in patients who cannot produce sputum are the preferred methods of making a rapid diagnosis. BAL culture seems to add little value in patients who can produce sputum; therefore, bronchoscopy should be deferred under such circumstances.

After 30 years of the human immunodeficiency virus (HIV) epidemic, parasites have been one of the most common opportunistic infections (OIs) and one of the most frequent causes of morbidity and mortality associated with HIV-infected patients. Due to severe immunosuppression, enteric parasitic pathogens in general are emerging and are OIs capable of causing diarrhoeal disease associated with HIV. Of these, Cryptosporidium parvum and Isospora belli are the two most common intestinal protozoan parasites and pose a public health problem in acquired immunodeficiency syndrome (AIDS) patients. These are the only two enteric protozoan parasites that remain in the case definition of AIDS till today. Leismaniasis, strongyloidiasis and toxoplasmosis are the three main opportunistic causes of systemic involvements reported in HIV-infected patients. Of these, toxoplasmosis is the most important parasitic infection associated with the central nervous system. Due to its complexity in nature, toxoplasmosis is the only parasitic disease capable of not only causing focal but also disseminated forms and it has been included in AIDS-defining illnesses (ADI) ever since. With the introduction of highly active anti-retroviral therapy (HAART), cryptosporidiosis, leishmaniasis, schistosomiasis, strongyloidiasis, and toxoplasmosis are among parasitic diseases reported in association with immune reconstitution inflammatory syndrome (IRIS). This review addresses various aspects of parasitic infections in term of clinical, diagnostic and therapeutic challenges associated with HIV-infection.

The value of transbronchial biopsy and bronchoalveolar lavage was assessed in the diagnosis of pulmonary disease in patients infected with the human immunodeficiency virus (HIV). Seventy four transbronchial biopsy and 66 bronchoalveolar lavage specimens (60 paired specimens) from 80 examinations in 64 patients were reviewed. Pneumocystis carinii was the most common pathogen isolated (43 patients). Bronchoalveolar lavage was superior to transbronchial biopsy for the diagnosis of this pathogen, the sensitivities being 90% and 56%. Cytomegalovirus was identified three times by lavage and once by transbronchial biopsy. Neither method detected Kaposi's sarcoma in the one patient shown to have it by open lung biopsy. The complication rate in a concurrent study of bronchoscopy with transbronchial biopsy in 74 consecutive HIV positive patients was 22%. This study does not support the use of transbronchial biopsy in these patients.

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A combination of three monoclonal antibodies, two prepared against human and one against rat Pneumocystis carinii, was used in an indirect fluorescent-antibody stain (IFA) to diagnose P. carinii in both bronchoalveolar lavage and lung biopsy specimens. This combination of monoclonal antibodies was specific for P. carinii and yielded bright fluorescence of both P. carinii cysts and trophozoites. A total of 126 specimens from 93 patients were stained for P. carinii by a toluidine blue O stain and the monoclonal IFA. Forty-five (35.7%) of these were positive for P. carinii by toluidine blue O, and 43 (34.1%) were positive by IFA. There was 98.4% agreement between both methods, with no false-positive and two false-negative occurrences by IFA. An IFA procedure with monoclonal antibodies such as those used in these studies can provide a simple, fast, and sensitive method for diagnosing P. carinii pneumonia by microbiology laboratories.

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