Search

In the present study, 67 patients suspected to be cases of visceral leishmaniasis (VL) were each checked for leishmanial infection by the microscopical evaluation of various biological specimens, in-vitro culture, serology and an assay based on nested PCR. Most (35) of the subjects were immunocompetent (IC) but 32 were immunodeficient (ID) as the result of HIV infection (18 cases), treatment to prevent transplanted organs being rejected (six) or haematological malignancies (eight). Forty-one (61.2%) of the subjects (19 IC subjects, 12 HIV-positive patients, four transplant patients and six patients with malignancies) were considered true cases of VL. For the IC subjects, only the production and microscopical examination of leucocytoconcentrates and cultures of Buffy coats gave sensitivities of80%, the results of the other methods showing higher sensitivities and almost perfect agreement with the 'gold-standard' diagnoses. For the ID subjects, however, only the serological tests and the PCR gave reasonable sensitivities (of80%). For the initial diagnosis of leishmaniasis in ID patients, IFAT and western blots may be useful, as, among the present ID patients, they gave sensitivities (of 80.9% and 88.2%, respectively) that were almost as high as that for the PCR, and specificities of 100%. In the diagnosis of VL in either IC or ID patients, the assay based on a nested PCR appeared to be particularly reliable, with sensitivities of 88.9% and 95.2%, respectively, and a specificity of 100% in both groups of patients. The testing of bone-marrow aspirates by PCR revealed very few VL cases who were not found positive when samples of their peripheral blood were checked in the same assay. For both IC and ID subjects therefore, the use of the PCR-based method to test samples of peripheral blood (which can be collected much more easily than bone-marrow aspirates and with much less pain for the subject) is recommended.

An epidemiological survey to characterize Entamoeba histolytica/E dispar isolates from 123 human subjects was carried out in the Wonji area of Central Ethiopia, where an increased incidence of amoebic infection has been reported. In a randomized, coproparasitological study, 93 (75.6%) of the subjects were found positive for at least one species of intestinal parasite: 14 (15.1%) harboured only one species and 79 (84.9%) were infected with at least two species. In-vitro culture in Robinson's medium revealed amoebic parasites in 52 (82.5%) of the 63 cases tested. Of the 29 amoebic isolates successfully stabilized, cloned and characterized by Sargeaunt's electrophoretic technique, 27 (93.1%) were of E. dispar zymodemes (19 of zymodeme I, two each of zymodemes III, V and XI, and one each of zymodemes X and XV) and two (6.9%) were of E. histolytica (zymodeme XIII).

Objective: To estimate the prevalence of bacterial vaginosis, Candida albicans, and Trichomonas vaginalis infections in a population of postmenopausal women with symptoms of vaginitis seen at a vaginitis clinic either as self-referred or clinician referred patients. Methods: A cross-sectional study of 148 postmenopausal women (cases) and 1564 controls of reproductive age attending a vaginitis clinic. C. albicans and T. vaginalis infections were diagnosed by culture techniques. Bacterial vaginosis was diagnosed on the basis of clinical findings. Results: Fifty-six (37.8%) postmenopausal women and 834 (53.3%) controls were diagnosed with T. vaginalis or C. albicans infection, or bacterial vaginosis, or mixed infection (odds ratio (OR) 0.53, 95% confidence interval (CI) 0.37–0.75). C. albicans and T. vaginalis infection were diagnosed in 34.1% (5341564) and 1.92% (301564) of women of childbearing age and in 3.5% (20148) and 10.8% of postmenopausal women, respectively. (P < 0.05 for both comparisons). The prevalence of bacterial vaginosis was similar between the two groups (14148 in postmenopausal patients and 2101564 in controls of reproductive age; P = 0.22). Conclusions: Among postmenopausal women attending a vaginitis clinic, a defined diagnosis of bacterial vaginosis, C. albicans or T. vaginalis infection can be made in about one third of such patients. Concerning the two thirds of symptomatic women lacking such a microbiologic diagnosis, alternative causes (e.g., estrogen deficiency, nonanaerobic bacterial infections, local irritants or allergenes, and dermatologic conditions) need to be considered.

A total of 77 mentally retarded male inpatients residing in a psychiatric institution in northern Italy were screened for the presence of stool parasites, Entamoeba histolytica particularly. Parasitological stool examination showed Entamoeba spp. (E. histolytica and/or E. dispar) in 26 cases (33.7%). In vitro culture on Robinson's medium was positive in 16 cases (61.1%); in 11 cases we could stabilize and clone the isolates and proceed to electrophoretic assays. In all cases, patterns of pathogenic zymodemes were found (zymodeme II, 3 isolates; zymodeme XII, 4 isolates; zymodeme XIV, 4 isolates). All isolates were therefore identified as E. histolytica.

A sample of 300 sexually-active adults was selected at random from patients, from the rural area of Malenga Makali, Tanzania, who were attending a dispensary because they had diarrhoea of at least 2 weeks' duration. The potential associations between the patient's health (in terms of the World Health Organization's clinical definition of AIDS), HIV-1 seroprevalence and malaria and other parasitic infections were then investigated. Although, HIV-1 seroprevalence was 20.6% overall, the level of seroprevalence was directly correlated with the distance between the patients' home villages and the nearest main road. Strict application of the clinical definition of AIDS gave 98.7% specificity, 46% sensitivity and a predictive value of 90.6% when validated by HIV-1 seropositivity. Although malaria infection was more common in HIV-1 seropositives than in the seronegatives, the intensity of the Plasmodium falciparum infections, intestinal amoebiasis and giardiasis did not appear to be correlated with HIV-1 infection. In contrast, intestinal infections with Cryptosporidium parvum and Isospora belli were virtually restricted to HIV-1 seropositive individuals who had had diarrhoea for a relatively long time.

A cohort of 51 homosexuals who were either HIV-positive or had AIDS was followed prospectively with parasitologic stool examination and in vitro culture in order to determine the incidence of E. histolytica infection. Amoebic isolates were further characterized by electrophoretic isoenzyme study. Five subjects (9.8%) were found to be infected with E. histolytica. None of the amoebic isolates were found to be pathogenic by isoenzyme analysis.

Three autochthonous cases of Entamoeba histolytica infection in institutionalized mentally-retarded patients are reported. Isoenzyme analysis by starch-gel electrophoresis shows the pathogenicity of the three isolates: two belong to zymodeme II, and one to zymodeme XIX. The study shows that invasive E. histolytica strains occur in Italy and can be isolated from institutionalized oligophrenic patients.

The review summarizes the results in the main parasitological topics of our Lab: amoebic infections due to Entamoeba histolytica/Entamoeba dispar complex and to Acanthamoeba spp. respectively, and human infections caused by microsporidia. Different rapid and advanced techniques have been included in the standardized diagnostic protocols for each topic, and a critical comparison among them was made, in order to define the gold standard diagnostic method: a) E. histolytica/E. dispar: in vitro culture, zymodeme typization, biomolecular identification (PCR), immunoenzymatic assay (ELISA) for direct detection in stools of specific surface antigenic lectins; b) Acanthamoeba spp.: in vitro culture, light and ultrastructural characterization, species identification by immunofluorescence method with monoclonal antibodies, in vitro pharmacological studies; c) Microsporidia: ultrastructural (TEM), biomolecular (PCR), biochemical and immunological (SDS-PAGE, Immunoblotting) studies for species identification, use of advanced ultrastructural techniques ("freeze-etching", "deep-etching") in order to deepen the spore wall structure, to study the cytoskeletal function of actin and to define the mode of infection, in vitro pharmacological assays on some inhibitors of chitin-synthases.

Of 550 mentally retarded patients in an Italian institution, 125 (23%) were found to be infected with intestinal parasites. The infections were most frequent in young men, those with severe mental retardation, the chronically institutionalized and those living in older wards. Ninety-four (75.2%) of the parasitised subjects were infected only with protozoa, 25 (20%) only with helminths, and six (4.8%) with protozoa and helminths. Entamoeba histolytica and E. dispar infections were detected, but at low prevalences; in-vitro culture in Robinson's medium and isoenzyme electrophoresis of the cloned amoebic isolates indicated one infection with E. histolytica (zymodeme XII) and two infections with E. dispar (zymodemes I and III). All three Entamoeba-positive subjects were asymptomatic cyst-passers. Antibodies to E. histolytica were detected in seven (1%) of the sera from the 550 patients examined; only one of these was a carrier of an E. dispar strain at the time of investigation. The low prevalences of all the parasitic infections and of the amoebic infections in particular (compared with those observed previously in institutions for the mentally retarded) reflect relatively good facilities and sanitary conditions, an adequate number of well trained staff and good control of the more susceptible subjects.

Isoenzyme analysis by starch-gel electrophoresis has proved to be a useful method for the biochemical differentiation of pathogenic Entamoeba histolytica and non-pathogenic E. dispar isolates. Of the 24 known zymodemes, 3 are laboratory-made and have not previously been identified in humans. Parasitology screening was carried out in a psychiatric institution. Two amebic stocks were isolated and characterized that had never previously been found in humans and that have protein patterns identical to that of the laboratory-made zymodeme XX.

A limited outbreak of symptomatic intestinal and extraintestinal amoebiasis within a family complex is described. The infection was almost certainly transmitted by a Philippino housemaid, who was an asymptomatic carrier of Entamoeba histolytica infection acquired in her native country. Starch-gel electrophoresis showed isoenzyme patterns characteristic of pathogenic zymodeme XIX in all the amoebic isolates.

Cryptosporidiosis has emerged as one of the life-threatening opportunistic enteric infections in HIV-infected persons. To date, Cryptosporidium parvum is known to infect man via person-to-person or zoonotic transmission. We studied the sequential stages of the life cycle of C. parvum by Normarski interference-contrast microscopy in fresh gut specimens of newborn mice, infected with a strain derived from an AIDS patient with cryptosporidial diarrheal enteritis. Many 4- to 5-day-old suckling BALB/C mice were orally inoculated with 1 x 10(6) oocysts, obtained by acid flocculation of the patient's stools. The animals were sacrificed from 4 to 96 h post-infection and the ileum was examined microscopically. All stages of the asexual life cycle of C. parvum, from excysted sporozoites in the intestinal lumen through the development of type II mature meronts, 12- to 72-h post-infection, were documented by extemporaneous microscopic evaluation of fresh gut samples. The sexual cycle, characterized by the appearance of micro- and macrogametocytes, followed by a zygote developing into a sporulated oocyst, was documented as early 48-h post-infection. Our Nomarski interference-contrast observations on the life cycle of C. parvum yielded data comparable with those originally published by Current and Reese, and confirm the results of previous electron microscopic studies performed by several other authors.

Two new extemporaneous negative-staining methods are proposed to detect Cryptosporidium oocysts in stools, using light-green and merbromine in 1% and 2% aqueous solution, respectively. A Ziehl-Neelsen stain as modified by Henriksen and Polhenz was used as a reference technique. A comparison between these two new stains and the reference, a modified Ziehl-Neelsen mod. method gave almost identical sensitivity and specificity. We propose their use in routine diagnosis for enteric cryptosporidiosis.

A direct method for detection of Pneumocystis carinii was evaluated in 14 patients with impaired immune function (3 seropositive for HIV, 8 with AIDS and 3 with heart transplants) and signs and symptoms suggestive of Pneumocystis carinii pneumonia. Direct examination by phase-contrast and interference-contrast microscopy of fresh clinical specimens obtained by sputum induction, bronchoalveolar lavage or transbronchial lung biopsy was found to be a simple and rapid method for detection of Pneumocystis carinii, the sensitivity of the method being comparable to that of the classical toluidine blue O and Diff-Quik staining methods. These findings suggest that this direct microscopy technique could be considered for routine clinical application in patients with suspected Pneumocystis carinii pneumonia.

One hundred and eighteen cases of S. stercoralis infection were detected by stool examination among 4203 patients seen over a period of three years at the Division of Infectious Diseases (IRCCS, San Matteo, University of Pavia) of a University Hospital in Northern Italy. Because chronic strongyloidiasis is often asymptomatic, a 3 per cent prevalence in this selected population indicates that the parasitosis may be much more frequent in the general population of the area. Parasite-specific humoral responses were studied in 48 patients. Strongyloides -specific IgG and IgA antibodies were found by ELISA in the serum of 40 (83.3 per cent) and 43 (89.5 per cent) patients, respectively, and specific IgE antibodies were detected by RAST in 39 (81.3 per cent). The specific IgG antigen recognition patterns of these patients were compared by immunoblotting with those of North and South American and Thai patients, and differences among the groups suggested the existence of separate geographic strains of S. stercoralis . This was further supported by the virtual absence in our patients of the larva currens, a common and highly characteristic sign of strongyloidiasis in subjects infected in southeast Asia. More awareness of the epidemiologic and clinical features of strongyloidiasis, with the use of serologic tests to screen candidates for immunosuppression, may result in the early detection of chronic uncomplicated strongyloidiasis and in the prevention of fatal opportunistic hyperinfections.

We tested in vitro amphotericin B (AMP-B), econazole (ECO), and 5-fluorocytosine (5-FC) on pathogenic Naegleria fowleri (KUL strain), Naegleria australiensis s.sp. italica (AB-T-F3, original strain) to assess their sensitivity to chemotherapeutic compounds. Previous reports have shown the polyene antibiotic AMP-B to be the most active agent. It was, however, much more active on N. fowleri than on N. australiensis and N. australiensis s.sp italica. 5-FC and ECO gave rise to non appreciable effect at non-toxic corresponding dosages in vivo. The results of these in vitro tests are discussed.

We performed an epidemiological survey of 17 thermal baths and the same number of mud-basins. This study aimed to ascertain the presence and incidence of small free-living amoebae, particularly species and/or strains of Naegleria and Acanthamoeba spp., occasional etiological agents of fatal meningoencephalitis and/or ocular infections in man. Over 51 samples of water and mud incubated at 37 degrees C and at 45 degrees C, 34 (66.7%) became positive at 37 degrees C and 33 (64.7%) at 45 degrees C. We isolated 7 (6%) strains of Naegleria spp., 6 (5.2%) of Acanthamoeba spp., 39 (33.6%) of Vahlkampfia spp., 28 (24.1%) of Hartmannella spp. and 36 (31.1%) strains of other species of free-living amoebae. 4 strains of Naegleria spp. and 6 of Acanthamoeba spp. proved pathogenic both in vivo, after experimental infection (meningoencephalitis) in the albino mouse, and in vitro, having previously contaminated monolayers of Vero cell line (cytopathic effect). Within the isolated pathogenic strains of Naegleria spp., a new strain of N. australiensis s.sp. italica was typified from an immunochemical point of view. This should be added to previous isolations reported by us.

A subspecies of Naegleria australiensis, N. australiensis italica, pathogenic for mice, was recently isolated and identified from an Italian thermal spa. We describe the histopathological changes of the central nervous system with experimental infection of albino mice. The histopathological patterns are intermediate to those seen with infection caused by N. fowleri and N. australiensis or Acanthamoeba spp. An acute inflammatory reaction was present within the choroid plexus, ependyma, midbrain, cerebellum, and basal ganglia. Occasional single amebic trophozoites were found within some microabscesses. Cysts were not identified. Involvement of the olfactory neuroepithelium and of the nasal mucosa was not detected.

The authors report a case-study of 20 subjects affected by visceral Larva migrans (VLM) and ocular Larva migrans (OLM). Of the 20 patients, 11 were male, and 9 female (mean age: 45 years); 7 were children, from 1 to 8 years old. Seventeen patients presented clinical and bio-humoral features which could be correlated with VLM syndrome; alongside cases with clearly suggestive symptomatology, in some patients the infection by Toxocara sp. was found to show few or minor symptoms, or even to be asymptomatic and occasionally it was suspected because of a marked peripheral eosinophilia. In 3 cases the infection was exclusively ocular. So far as immunology is concerned, serological tests confirmed in all 20 cases the clinico-epidemiological and/or bio-humoral suspicion; in particular the ELISA IgG test showed high titres in 16 cases, while the RAST showed significant levels of specific IgE in 17 cases. From the therapeutic point of view, the subjects treated were exclusively those presenting a manifest clinical picture: in 5 patients the first choice drug was thiabendazole, with clinical cure in only 2 cases; diethylcarbamazine resolved the clinico-parasitological picture in 1 case; 1 patient is still under treatment with albendazole.

Acanthamoeba keratitis is a severe ocular infection secondary to accidental macro- or microscopic trauma of the cornea. Starting in 1985, a dramatic increase of this infection was recorded along with the spread of contact lens use. This protozoal disease is difficult to treat because of the scarcity of efficacious topical and systemic drugs. We evaluated the in vitro effectiveness of povidone-iodine (PVP-I [Betadine]), an agent with broad antibacterial and antiviral activity, compared to that of chlorhexidine (CXD), a cationic antiseptic, on Acanthamoeba isolates from patients with amebic keratitis. The results showed that PVP-I solution from 0.5 to 2.5% has a better antiamebic activity both on trophic and cystic stages of Acanthamoeba spp. than does CXD.