Phaeohyphomycosis caused by Chaetomium globosum in an allogeneic bone marrow transplant recipient.

Bone marrow transplant recipients are highly susceptible to opportunistic fungal infections. This is the report, of the first case of a Chaetomium systemic infection described in Brazil. A 34 year-old patient with chronic myeloid leukemia underwent an allogeneic sibling matched bone marrow transplant. Seven months later, he developed systemic infection with enlargement of the axillary and cervical lymph nodes. Culture of the aspirates from both lymph nodes yielded Chaetomium globosum. The infection was successfully treated with amphotericin B. The increasing population of immunosupressed patients requires a careful microbiologic investigation for uncommon fungal infections.

Susceptibility profile of 200 bloodstream isolates of Candida spp. collected from Brazilian tertiary care hospitals.

We evaluated the antifungal susceptibility profile of 200 recent bloodstream isolates of Candida spp. sequentially obtained from patients admitted to five tertiary care hospitals in Brazil. Isolates were identified by classical methods and the antifungal susceptibility profile was determined by the NCCLS microbroth assay method. Candida albicans was the most frequent species (41.5%); followed by C. tropicalis (24%) and C. parapsilosis (20.5%). The frequency of C. glabrata and C. krusei was low (nine and two isolates, respectively). Only three strains were resistant to fluconazole (two C. krusei and one C. glabrata) and only one was resistant to itraconazole (the same C. glabrata strain that was resistant to fluconazole). Two strains were considered susceptible dose-dependent (SDD) to fluconazole and 13 isolates (6.5%) were SDD to itraconazole. Overall, the MIC50 value of non-C. albicans isolates for fluconazole was two dilutions higher than that of C. albicans isolates, and for itraconazole was one dilution higher. Resistance to amphotericin B (MIC > or = 2 microg ml(-1)) was observed in 2.5% of isolates (two strains of C. albicans, two of C. parapsilosis and one of C. krusei). This study showed that episodes of candidemia in Brazilian public hospitals are represented mainly by fluconazole-susceptible non-C. albicans species. This finding is probably related to the low use of fluconazole in these hospitals.

Pichia anomala outbreak in a nursery: exogenous source?

BACKGROUND: Pichia anomala is a rare cause of fungemia. From February to April, 1998, eight cases of fungemia occurred in the intensive care and high risk units of the Nursery. There were four infants with P. anomala infection, one of whom also had Candida parapsilosis infection, two cases with C. parapsilosis infection and two with Candida albicans infection. OBJECTIVE: To determine factors associated with fungemia in the intensive care and high risk units of the Nursery, especially P. anomala. METHODS: A cohort study with 59 newborns. RESULTS: Factors associated with fungemia were: central venous catheter (CVC) (P = 0.0006); total parenteral nutrition (TPN) (P = 0.0005); lipid emulsion (P = 0.002); previous antimicrobial use (P = 0.002); and other invasive procedures (P = 0.002). Factors associated with P. anomala fungemia were: CVC (P = 0.004); TPN (P = 0.018); previous antibiotic use (P = 0.037); and other invasive procedures (P = 0.037). Evaluation of the units demonstrated that there were several technical problems involving administration of TPN that was manipulated in the Nursery without precautions. Changes in TPN formulation and education as to adequate technique were implemented. During follow-up (1998 to 1999) only two fungemias occurred that were caused by C. albicans. Cultures of hands of personnel were negative for P. anomala. Electrophoretic karyotyping of P. anomala showed three profiles. CONCLUSIONS: Factors associated with fungemia were catheter use, invasive procedures and total parenteral nutrition, suggesting that the acquisition of P. anomala was exogenous.

Atypical Cryptococcus neoformans isolate from an HIV-infected patient in Brazil.

Cryptococcus neoformans is an important fungal pathogen in immunocompromised hosts. Capsulation, urease and melanin synthesis activity of the fungus are well known virulence factors. Although artificial melanin-deficient mutants of Cr. neoformans have been investigated, the clinical mutant is rare. We found a Cr. neoformans isolate in the cerebrospinal fluid of an AIDS patient which produced a light tan colony on a caffeic acid cornmeal agar (CACA) plate. The mycological feature of the isolate was as follows; normal capsulation, defective inositol assimilation ability, serotype A; urease-positive; mating type alfa; haploid; extremely slow growth in RPMI 1640 medium, Sabouraud dextrose broth, brain heart infusion broth and yeast nitrogen base; lower production of melanin with L-DOPA substrate; and low virulence to ddY mice. We also investigated the partial DNA sequence of CNLAC1 gene between the 3085th to 3623rd base. There were many substitutions, 3 insertions and 3 deletions in the isolate compared with GenBank accession number L22866. The result indicated some functional disorder in the gene. Although the CACA plate is an excellent selective medium for Cr. neoformans, other identification methods should also be used.

Trichosporon species infection in bone marrow transplanted patients.

Trichosporon species are emerging as opportunistic agents that cause systemic diseases in immunocompromised patients. Patients undergoing bone marrow transplant are submitted to intense and prolonged periods of neutropenia and consequently to several risk factors to fungal infections as the use of broad spectrum antibiotics and invasive devices. Two cases of fungal infections caused by Trichosporon asahii var. asahii and T. inkin in patients with bone marrow transplant are described T. asahii var. asahii was responsible for fungemia and the identification of this microorganism was later performed. T. inkin caused vascular accesses infection and was recovered from an implanted Hickman-Broviac catheter. Both patients were under oral fluconazole prophylaxis. The patient with systemic infection died despite the therapy with amphotericin B and the patient with catheter-related infection recovered from the fungal infection after catheter removal. Difficulties in the identification of this microorganism lead to delays in treatment and post-mortem diagnosis.

First isolation of Candida dubliniensis in Rio Grande do Sul, Brazil.

Candida dubliniensis is a newly recognized species closely phylogenetically related to Candida albicans and is commonly associated with oral candidiasis in human immunodeficiency virus-positive patients. In this paper we report the isolation of three strains of C. dubliniensis, from AIDS patients, in the state of Rio Grande do Sul (Brazil). The phenotypic identification was based on germ tube emission, abundant production of chlamydospores, assimilation of sucrose but not of xylose and the inability to grow at 42 degrees C. Randomly amplified polymorphic DNA (RAPD) analysis and genomic DNA sequencing confirmed the distinct genetic nature C. dubliniensis. Topics related to the epidemiology, isolation, phenotypical and genotypical identification of C. dubliniensis are also discussed.

Antifungal susceptibilities, varieties, and electrophoretic karyotypes of clinical isolates of Cryptococcus neoformans from Brazil, Chile, and Venezuela.

One hundred clinical isolates of Cryptococcus neoformans from human immunodeficiency virus (HIV)-infected and non-HIV-infected patients from Brazil, Chile, and Venezuela were separated according to varieties and tested for antifungal susceptibility. A high susceptibility to antifungal agents was observed among all the isolates. The electrophoretic karyotyping of 51 strains revealed good discrimination among Cryptococcus neoformans var. neoformans strains.

Extrusion of fluorescein diacetate by multidrug-resistant Candida albicans.

Over-expression of multidrug efflux transporters causes Candida albicans cells to be resistant to azole antifungal agents. There are several kinds of indicator for multidrug resistance (MDR) phenotype of higher eukaryotic cells. Calcein AM is a prefluorochrome that is known as a substrate for multidrug efflux transporters of mammalian cells. We investigated whether calcein AM was also extruded by the ATP-dependent multidrug transporter (cdr1p) of C. albicans. There was no significant difference in the accumulation of calcein AM between MDR cells and drug-susceptible cells of C. albicans even with sodium azide, suggesting that calcein AM may not be associated with the CDR1-gene-related multidrug efflux system of C. albicans. However, a structurally related prefluorochrome derivative, fluorescein diacetate (FDA), was shown to be extruded by the CDR1 mRNA-overexpressing yeast cells. In comparison with drug-susceptible cells, the resistant cells emitted very weak fluorescence when stained with FDA. Furthermore sodium azide increased the fluorescence of the resistant cells more than 20 times, whereas the fluorescence in the drug-susceptible cells with FDA and sodium azide was three to four times stronger. These results suggested that FDA might be extruded by the CDR1-related multidrug efflux transporter of C. albicans.

Nasal MRSA colonization of AIDS Patients cared for in a Brazilian university hospital.

Weekly culture surveillance was conducted over a 2-year period to determine the incidence of methicillin-resistant Staphylococcus aureus nasal colonization among acquired immunodeficiency syndrome patients cared for in a day-care unit and in an infectious diseases unit. Analysis of genomic DNA profiles showed a predominant pattern in both units.

A new PCR primer for the identification of Paracoccidioides brasiliensis based on rRNA sequences coding the internal transcribed spacers (ITS) and 5 x 8S regions.

Internal transcribed spacer (ITS) genes including the 5.8S ribosomal (r)RNA of Paracoccidioides brasiliensis were amplified and the DNA sequences were determined. Based on a comparison of the sequence information, a new polymerase chain reaction (PCR) primer pair was designed for specific amplification of DNA for P. brasiliensis. This primer pair amplified a 418-bp DNA sequence and was 100% successful in identifying 29 strains of P. brasiliensis (including the reference strains) isolated from the regions of Brazil, Costa Rica, Japan, Argentina or from different sources. The results of specificity tests of these primers to compare the fungus with those of Aspergillus fumigatus, Blastomyces dermatitidis, Candida albicans, Cryptococcus neoformans, Histoplasma capsulatum and Penicillium marneffei are also reported.

Geographic grouping of Cryptococcus neoformans var. gattii by random amplified polymorphic DNA fingerprint patterns and ITS sequence divergence.

Eleven reference strains of Cryptococcus neoformans var. gattii were analyzed by random amplified polymorphic DNA (RAPD) patterns using three oligo primers. Three major RAPD pattern profiles (profiles I, II and III) were identified with A-1 oligo primer. Each profile was found to relate to a geographic region. Since the strains which belong to profiles I and II were mainly the isolates from America, and profile III from Asia with A-1 primer, these three profiles were assigned to the geographic grouping of America-1, America-2 and Asia, respectively. Analysis of rDNA sequences coding an internal transcribed spacer (ITS) region of C. neformans var. gattii revealed that the fungus with each RAPD profile has characteristic base sequences at the four positions (10 and 15 positions in ITS1 and 8 and 56 positions at ITS2) of the ITS regions. On the basis of the combinations of the four bases specific for the ITS regions, four ITS types, AAGG (America-1), AAAC (America-2), GGGC (Asia-1) and AGGC (Asia-2) were identified: the geographic group of Asia was further classified into two subgroups of Asia-1 and Asia-2 based on the ITS typing. Clinical isolates from Thailand (6 strains) and Brazil (7 strains) were found to belong to the geographic group of America-1 nd America-2, respectively. Five reference strains of C. neoformans var. gattii from the CBS culture collection were classified into two America-2, one Asia-1 and two Asia-2 groups. This ITS region analysis allowed us to distinguish all isolates of C. neoformans var. gattii into four geographic groups based on the ITS base sequence, and further molecular epidemiological and ecological research on this fungus is recommended.

Enterobacter cloacae sepsis outbreak in a newborn unit caused by contaminated total parenteral nutrition solution.

OBJECTIVE: The study aimed to investigate an outbreak caused by Enterobacter cloacae in a neonate intensive care unit. DESIGN: A descriptive study of an outbreak of sepsis in high-risk neonates was used. SETTING: The study was set in a tertiary care university teaching hospital. PATIENTS: The patients were 11 neonates infected with Enterobacter cloacae whose symptoms and signs of sepsis developed during a 16-hour period. All but one neonate received parenteral nutrition. Isolates from blood cultures, in-use parenteral nutrition solutions, and control aliquots of parenteral nutrition solution were typed by pulsed-field gel electrophoresis. RESULTS: Enterobacter cloacae was found in the refrigerated aliquots of parenteral nutrition solution, in blood cultures from infected newborns, and from in-use parenteral nutrition solutions. All these strains of Enterobacter cloacae had the same antibiotic susceptibility pattern and the same genomic DNA profile. The strain isolated from the one patient who did not receive parenteral nutrition presented a different susceptibility profile and genotype. CONCLUSION: The source of the nosocomial sepsis was the parenteral nutrition solution in 10 neonates. This contamination apparently occurred during preparation of the parenteral solution.

High rate of non-albicans candidemia in Brazilian tertiary care hospitals.

In order to evaluate the epidemiology of candidemia in Brazil, we performed a prospective multicenter study conducted in six general hospitals from São Paulo and Rio de Janeiro, We enrolled a total of 145 candidemic patients (85 males) with a median age of 32 years. Non-albicans species accounted for 63% of all episodes and the species most frequently causing candidemia were C. albicans (37%), C. parapsilosis (25%), C. tropicalis (24%), C. rugosa (5%), and C. glabrata (4%). Systemic azoles were used before the onset of candidemia in only six patients. There were no differences in the coexisting exposures or underlying diseases associated with the species most frequently causing candidemia. The overall crude mortality rate was 50%. Nosocomial candidemias in our tertiary hospitals are caused predominantly by non-albicans species, which are rarely fluconazole resistant. This predominance of non-albicans species could not be related to the previous use of azoles.

Infection control in a Brazilian regional multihospital system.

OBJECTIVE: This study describes the organization of infection control committees in a Brazilian multihospital system, identifying their major problems. Our goal is to discuss the main deficiencies in infection control and to target some interventions that can improve the efficiency of these actions in Brazilian hospitals. DESIGN AND SETTING: We used a descriptive epidemiologic design. We interviewed the chairs and the nurses of the infection control committees and visited the main areas to observe infection control in a multihospital system with 3146 beds. For analysis of the results, we performed a standardization process, establishing a score for each hospital by using infection control organization as a surrogate marker for quality outcome. The mean hospital scores for infection control, existence of policies, and infrastructure at each hospital were compared by using different stratification and multivariate analysis. RESULTS: Statistically significant differences were found among surveyed hospitals by using stratification by size, funding status, and presence of teaching activities. CONCLUSIONS: Diverse patterns of infection control organization were found among surveyed hospitals. Small hospitals represented the major problem in providing effective infection control. Chiefly for these hospitals, the epidemiologic indicators and the surveillance and control system proposed by the Brazilian Ministry of Health and based on the Centers for Disease Control and Prevention model showed poor suitability.

New PCR primer pairs specific for Cryptococcus neoformans serotype A or B prepared on the basis of random amplified polymorphic DNA fingerprint pattern analyses.

Thirty-three strains of Cryptococcus neoformans were isolated from clinical specimens, including specimens from AIDS patients in Brazil, and were classified into two serotypes; we detected 31 and 2 strains of serotypes A and B, respectively. Random amplified polymorphic DNA (RAPD) fingerprint pattern analyses of these strains of serotypes A and B showed that the patterns were similar for strains of each serotype when three 10-mer primers were used as the RAPD primers. Comparative studies of the fingerprint patterns of the study isolates with those of the reference strains also showed that the RAPD patterns for strains of each serotype were related and that most of the fingerprint bands existed commonly for all strains of each serotype tested. The common RAPD bands (an approximately 700-bp band for serotype A and an approximately 450-bp band for serotype B) were extracted and the DNA sequences were determined. Using this information, we prepared two and one PCR primer pairs which were expected to be specific for C. neoformans serotypes A and B, respectively. Use of each PCR primer combination thus prepared for serotype A or B was 100% successful in identifying the respective C. neoformans serotypes, including the 33 clinical isolates tested in the present study. Among these combinations, one for serotype A was found to amplify DNA from C. neoformans serotype B as well as serotype A. Serotype B-specific PCR primer pairs amplified DNA from not only serotype B strains but also from serotype C strains. The usefulness of other serotype-specific PCR primers for clinical C. neoformans isolates is discussed.

Risk factors for death in patients with candidemia.

OBJECTIVE: To analyze possible risk factors for death among patients with nosocomial candidemia. To identify risk factors for death in patients with candidemia, we analyzed demographic, clinical, and microbiological data. SETTING: Six tertiary hospitals in Brazil. PATIENTS: A cohort of 145 patients with candidemia. DESIGN: 26 possible risk factors for death, including age, underlying disease, signs of deep-seated infection, neutropenia, number of positive blood cultures, removal of a central venous catheter, etiologic agent of the candidemia, susceptibility pattern of the isolate to amphotericin B, and antifungal treatment were evaluated by univariate stepwise logistic regression analysis. RESULTS: Non-albicans species accounted for 63.4% of the candidemias. Risk factors for death in univariate analysis were older age, catheter retention, poor performance status, candidemia due to species other than Candida parapsilosis, hypotension, candidemia due to species other than Candida parapsilosis, and no antifungal treatment. In multivariate analysis, older age and nonremoval of a central venous catheter were the only factors associated with an increased risk for death. CONCLUSIONS: These data suggest that patients with candidemia and a central venous catheter should have the catheter removed.

Candida parapsilosis fungemia associated with implantable and semi-implantable central venous catheters and the hands of healthcare workers.

A cluster of six cases of fungemia among hematology, bone marrow transplant, and oncology patients was investigated in a case-control study (18 controls). The use of implantable and semi-implantable central venous catheters was significantly associated with cases (p = 0.016). The hands of three healthcare workers (HCWs) were positive for Candida parapsilosis. Electrophoretic karyotyping showed two profiles among patients and HCWs, and five among six unrelated strains. The profiles of two HCWs matched the ones of the patients they had handled. The patients' strains were moderate or strong slime producers, whereas none of the HCWs' were strong producers. In conclusion, our results indicated the occurrence of an outbreak C. parapsilosis fungemia related to long-term central venous catheters in which the hands of HCWs were implicated. The amount of slime production might be associated with the pathogenicity of the strains.

Colorimetric MTT assessment of antifungal activity of D0870 against fluconazole-resistant Candida albicans.

The in vitro antifungal activity of D0870 against eight isolates of fluconazole-resistant Candida albicans was compared with that of itraconazole, ketoconazole and miconazole. The colorimetric MTT [3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2H tetrazolium bromide] assay was used to assess the antifungal activities. The 50% minimum inhibitory concentration (MIC50) of D0870 was below 0.031 microgram ml-1 for seven isolates and 0.25 microgram ml-1 for one isolate. The activity of D0870 was superior to that of the other azoles. Ketoconazole was the most effective azole next to D0870. Therefore, the new bis-triazole, D0870, is expected to be promising for the therapy of fluconazole-resistant candidosis. The present data also confirmed that the MTT assay may be useful for evaluation of resistance and detection of resistant C. albicans.

Relative frequency of nosocomial microorganisms at UNICAMP University Hospital from 1987 to 1994.

The frequency of microorganisms identified in nosocomial infections at Unicamp University Hospital from 1987 to 1994 was analysed. The most common microorganism was S. aureus (20.9%), which was found in surgical wound, bloodstream and arterial-venous infections. In urinary tract infections (UTI), gram-negative rods (56.5%) and yeasts (9%) predominated. A. baumannii isolates were observed to have increased in the last three years. There was a gradual increase in the frequency of coagulase-negative staphylococci and A. baumannii in bloodstream infections but there wasn't any change in Candida sp.

Gastrointestinal translocation as a possible source of candidemia in an AIDS patient.

Apart from cryptococcosis and histoplasmosis, which are mycoses contained by T cell-mediated mechanisms of host defense, fungemia is rarely found in AIDS patients. The frequency of fungemia due to Candida spp. has been reported to be as low as 1%. We report a non-neutropenic AIDS patient who presented a candidemia which probably arose from her gastrointestinal tract.