Development of Secondary Resistance to Fluconazole in Cryptococcus neoformans Isolated from a Patient with AIDS

Cryptococcus neoformans is the fifth most common opportunistic agent of infection in patients with AIDS in the USA, exceeded only by Candida species, Pneumocystis carinii, cytomegalovirus and Mycobacterium avium1, 2, 6, 10, 11. In Brazil is the sixth, exceeded by Candida species, P. carinii, Mycobacterium species, Toxoplasma gondii, and herpes simplex virus (AIDS, Boletim Epidemiológico, set/nov 96, Ministério da Saúde, Brasil). During 30 years, the treatment of C. neoformans meningitis was based on the use of amphotericin B with or without flucytosine13. Nowadays, with the immunodepression caused by human immunodeficiency virus (HIV) infection and the availability of new antifungal drugs as the triazoles, the concept related to cure and relapses of cryptococcosis has been altered7, 20. Patients are treated with amphotericin B with or without flucytosine as initial therapy, but maintenance therapy is always necessary in AIDS patients with C. neoformans infections

Toxocara spp. eggs in public squares of Sorocaba, São Paulo State, Brazil

The visceral larva migrans (VLM) is a syndrome observed in human infection with helminth larval eggs such as the Toxocara spp. that usually infects dogs and cats. Among the risk factors involved in the occurrence of VLM, particularly important is the size of these animal populations. Sorocaba is a city with a dog population twice as large as that recommended by the World Health Organization. This fact has led to a survey of the presence of Toxocara spp. eggs in public square soils of this city. Thirty squares were selected, fifteen located in the outskirts of the city and fifteen downtown. Soil samples were collected from five distinct sites in the same area. The material was homogenized and drained and 100 g was mixed with a saturated solution of magnesium sulfate and 5% potassium iodine. The floating material was analyzed under the light microscope. Toxocara spp. eggs were found in 16 squares, nine of which were located in the outskirts of the city and seven downtown. It was concluded that Sorocaba squares present a high rate of contamination with Toxocara spp. eggs. The squares in the outskirts of the city presented a higher occurrence of these eggs in comparison with those downtown, although the difference was not statistically significant (p > 0.05).

Evaluation of Opuntia spp. bioactive products as promising natural chemotherapeutical agents- an in vitro approach

Dissertation to obtain a Master Degree in Biotechnology

In vitro susceptibility to antifungal agents of clinical and environmental Cryptococcus neoformans isolated in Southern of Brazil

The purpose of the present study was to compare the susceptibility to four antifungal agents of 69 Cryptococcus neoformans strains isolated from AIDS patients with that of 13 C. neoformans strains isolated from the environment. Based on the NCCLS M27-A methodology the Minimal Inhibitory Concentrations (MICs) obtained for amphotericin B, itraconazole and ketoconazole were very similar for clinical and environmental isolates. Clinical isolates were less susceptible to fluconazole than environmental isolates. The significance of these findings and aspects concerning the importance, role and difficulties of C. neoformans susceptibility testing are also discussed.

Snakebites by Bothrops spp in children in Campinas, São Paulo, Brazil

From January, 1984 to March, 1999, 73 children under 15 y old (ages 1-14 y, median 9 y) were admitted after being bitten by snakes of the genus Bothrops. Twenty-six percent of the children were classified as mild envenoming, 50.7% as moderate envenoming and 20.6% as severe envenoming. Two patients (2.7%) showed no signs of envenoming. Most of the patients presented local manifestations, mainly edema (94.5%), pain (94.5%) ecchymosis (73.9%) and blisters (11%). Local and/or systemic bleeding was observed in 28.8% of the patients. Before antivenom (AV) administration, blood coagulation disorders were observed in 60.7% (incoagulable blood in 39.3%) of the 56 children that received AV only in our hospital. AV early reactions, most of which were considered mild, were observed in 44.6% of these cases (in 15/30 patients not pretreated and in 10/26 patients pretreated with hydrocortisone and histamine H1 and H2 antagonists). The main clinical complications observed were local infection (15.1%), compartment syndrome (4.1%), gangrene (1.4%) and acute renal failure (1.4%). No deaths were recorded. There were no significant differences with regard to severity of envenoming versus the frequency of blood coagulation disorders among the three categories of envenoming (p = 0.75) or in the frequency of patients with AV early reactions between the groups that were and were not pretreated (p = 0.55). The frequency of local infection was significantly greater in severe cases (p < 0.001). Patients admitted more than 6 h after the bite had a higher risk of developing severe envenoming (p = 0.04).

Identification of Cryptosporidium spp. oocysts in fecal smears stained with Heidenhain's iron hematoxylin

There is no paucity of methods for diagnosing Cryptosporidium spp. infection. The merits of immunoassays notwithstanding, microscopic identification of Cryptosporidium spp. oocysts in fecal samples remains an important diagnostic procedure. It owes the persistence of its use to such characteristics as dispensing with expensive equipment and kits, requiring only basic laboratory facilities, and having a low probability of false positive results when permanent slides are prepared, which can be re-examined in case of doubt. Cryptosporidium spp. oocysts can be readily identified in fecal smears prepared according to a regressive iron hematoxylin staining technique. The number of steps and their duration, as well as costs, were reduced to a minimum without loss of image quality and permanence of the preparations.

Characterization of Pinus spp. needles by gas chromatography and mass spectrometry: Application to plant-insect interactions

Dissertação apresentada para obtenção do Grau de Doutor em Ciências do Ambiente pela Universidade Nova de Lisboa, Faculdade de Ciências e Tecn

Primary cutaneous cryptococcosis due to Cryptococcus neoformans var. gattii serotype B, in an immunocompetent patient

The authors report a male patient, a seller with no detected immunosuppression, with an extensive ulcerated skin lesion localized on the left forearm, caused by Cryptococcus neoformans var. gattii serotype B. Oral treatment with fluconazole was successful. A review of the literature showed the rarity of this localization in HIV-negative patients. In contrast, skin lesions frequently occurs in HIV-positive patients, with Cryptococcus neoformans var. neoformans serotype A predominating as the etiological agent. In this paper, the pathogenicity of C. neoformans to skin lesions in patients immunocompromised or not, is discussed, showing the efficacy of fluconazole for the treatment of these processes.

Assessment of PCR in the detection of Leishmania spp in experimentally infected individual phlebotomine sandflies (Diptera: Psychodidae: Phlebotominae)

DNA amplification by the polymerase chain reaction (PCR) was applied in the investigation of the presence of Leishmania (Kinetoplastida: Trypanosomatidae) parasites in single phlebotomine sandflies. Three phlebotomine/parasite pairs were used: Lutzomyia longipalpis/Leishmania chagasi, Lutzomyia migonei/Leishmania amazonensis and Lutzomyia migonei/Leishmania braziliensis, all of them incriminated in the transmission of visceral or cutaneous leishmaniasis. DNA extraction was performed with whole insects, with no need of previous digestive tract dissection or pooling specimens. The presence of either mouse blood in the digestive tract of the sandflies or the digestive tract itself did not interfere in the PCR. Infection by as few as 10 Leishmania sp. per individual were sufficient for DNA amplification with genus-specific primers. Using primers for L. braziliensis and L. mexicana complexes, respectively, it was possible to discriminate between L. braziliensis and L. amazonensis in experimentally infected vectors (L. migonei).

The spectrum of computerized tomography (CT) findings in central nervous system (CNS) infection due to Cryptococcus neoformans var. gattii in immunocompetent children

Cranial CT scans of eleven immunocompetent children with central nervous system (CNS) infection due to Cryptococcus neoformans var. gattii were retrospectively reviewed. These children had an average age of 8.8 years and positive culture for C. n. var. gattii in cerebrospinal fluid. The most common signs and symptoms were headache, fever, nuchal rigidity, nausea and vomiting. No normal cranial CT was detected in any patient. Hypodense nodules were observed in all patients . The remaining scan abnormalities were as follows: nine had diffuse atrophy, six had hydrocephalus, and five had hydrocephalus coexistent with diffuse atrophy.

Serotype, mating type and ploidy of Cryptococcus neoformans strains isolated from patients in Brazil

Serotype, mating type and ploidy of 84 strains of Cryptococcus neoformans isolated from 61 AIDS and 23 non-AIDS patients admitted in a tertiary teaching hospital in São Paulo, Brazil were examined. Among 61 strains isolated from AIDS patients, 60 strains were var. grubii (serotype A). Only one strain was var. gattii (serotype B). No var. neoformans (serotype D) was found. Among 23 strains isolated from non-AIDS patients, 15 were var. grubii (serotype A) and the remaining 8 were var. gattii, all of which were serotype B. Seventy-three of the 75 serotype A strains were the heterothallic alpha type (MATalpha) and the remaining 2 were untypable (asexual). Most of the MATalpha strains (69/73) were haploid and the remaining 4 strains were diploid. Similarly, both of the 2 asexual strains among the 75 serotype A strains were haploid. There were no alpha-mating type (MATalpha) strains among the 84 isolates. All of the 8 var. gattii strains were serotype B and haploid. Among a total of 84 strains tested, neither serotype AD nor serotype D were found. Neither triploid nor tetraploid were found. These results suggest that the serological, sexual and ploidy characteristics in C. neoformans strains isolated from AIDS patients in São Paulo were rather simple, whereas strains isolated from non-AIDS patients presented serotype A and B with predominance of serotype A.

Vibrio spp. and Salmonella spp., presence and susceptibility in crabs Ucides cordatus

The presence of Vibrio spp. and Salmonella spp. in crabs marketed at the Bezerra de Menezes Ave., Fortaleza, State of Ceará, Brazil, was assessed between February and May, 2003. The number of individuals sampled in each one of the fifteen weekly samplings ranged between four and eight. Seven strains of Salmonella, from four different samplings, were identified, being five of them identified as serotype S. Senftenberg and two as S. Poona. All strains of Salmonella were sensitive to the tested anti-microbial drugs, with the exception of tetracycline and nalidixic acid, for which an intermediary sensibility was found. The MPN's for Vibrio ranged between 110/g and 110,000/g. Of the forty five Vibrio strains isolated from the crab samples, only 10 were identified up to the species level: two V. alginolyticus and eight V. parahaemolyticus. Bacteria belonging to the Enterobacteriaceae and Pseudomonaceae families were also identified, namely Escherichia coli, Klebsiella pneumoniae, Enterobacter cloacae, Pantoea agglomerans and Pseudomonas aeruginosa. The proper cooking of the animals is recommended in order to avoid problems for the consumers of this crustacean.

Environmental straits of Cryptococcus neoformans variety grubii in the city of Santos, SP, Brazil

This study involved a total of 116 samples, 79 taken from pigeon droppings and 37 of atmospheric air taken close to accumulations of excrement. Cryptococcus neoformans var. grubii was isolated from 11 (13.9%) of these samples. Other species of Cryptococcus were also isolated from these samples, such as C. albidus (12.6%) and C. laurentii (8.9%). C. neoformans was not isolated from the air samples, though C. albidus (5.4%) was. All the strains of C. neoformans were found to belong to the A serotype (C. neoformans var. grubii). In regard to the studies with the antifungal agents 5-fluorocytosine, fluconazole, itraconazole, amphotericin B and voriconazole, by means of the microdilution method (EUCAST), we point out that one sample demonstrated resistance to fluconazole, this being especially significant because this is an environmental strain.

Multiple brain abscesses due to Penicillium spp infection

We present a case of central nervous system (CNS) infection by a member of the Penicillium genera in a HIV-negative man in Brazil. The patient was admitted complaining of loss of visual fields and speech disturbances. CT scan revealed multiple brain abscesses. Stereothacic biopsies revealed fungal infection and amphotericin B treatment begun with initial improvement. The patient died few days later as a consequence of massive gastrointestinal bleeding due to ruptured esophageal varices. The necropsy and final microbiologic analyses disclosed infection by Penicillium sp. There are thousands of fungal species of the Penicillium genera. Systemic penicilliosis is caused by the P. marneffei and was formerly a rare disease, but now is one of the most common opportunistic infection of AIDS patients in Southeast Asia. The clinical presentation usually involves the respiratory system and the skin, besides general symptoms like fever and weight loss. Penicillium spp infection caused by species other than P. marneffei normally cause only superficial or allergic disease but rare cases of invasive disease do occur. We report the fourth case of Penicillium spp CNS infection.

Characterization of Cryptococcus neoformans isolated from urban environmental sources in Goiânia, Goiás State, Brazil

Cryptococcus neoformans is an opportunistic fungal pathogen that causes meningoencephalitis as the most frequent clinical presentation in immunocompromised patients, mainly in people infected by HIV. This fungus is an environmental encapsulated yeast, commonly found in soil enriched with avian droppings and plant material. A total of 290 samples of pigeon and the other avian droppings, soil, ornamental trees and vegetable material associated with Eucalyptus trees were collected to study environmental sources of Cryptococcus species in Goiânia, Goiás State. The determination of varieties, serotypes and the susceptibility in vitro to fluconazole, itraconazole and amphotericin B of C. neoformans isolates were performed. C. neoformans var. grubii (serotype A) was found in 20.3% (36/177) of pigeon dropping samples and in 14.3% (5/35) of samples of Eucalyptus. None of the environmental isolates of C. neoformans showed in vitro resistance to three antifungal agents. The knowledge of major route for human cryptococcal infection (inhalation of infectious particles from saprophytic sources) and a total of 60 C. neoformans isolates obtained from AIDS patients with cryptococcal meningitis between October 2001 and April 2002 justify the study of the habitats of these yeasts as probable sources of cryptococcosis in this city.