Evaluation of antimicrobial susceptibility testing of Nocardia spp. isolates by broth microdilution with resazurin and spectrophotometry.

BACKGROUND: Nocardia species are ubiquitous in natural environments and can cause nocardiosis. In the present study, the use of Resazurin salt and Spectrophotometry were proposed as alternative methods to reduce subjectivity in the interpretation of susceptibility results to antimicrobials by the broth microdilution method for Nocardia spp. RESULTS: The susceptibility of Nocardia spp. isolates to Amikacin, Ciprofloxacin, Minocycline and Trimethoprim-Sulfamethoxazole was evaluated by Minimum Inhibitory Concentration (MIC) determinations by the broth microdilution method. To verify cellular growth, the colour-changing dye Resazurin was applied, the Optical Densities were measured on a spectrophotometer, and both were compared to Clinical and Laboratory Standards Institute (CLSI) Gold Standard method (visual MIC determination). Percentages of essential and categorical agreements and interpretative categorical errors were calculated within each method (intra-reading) and between them (inter-reading). The Gold Standard visual reading demonstrated 100% of essential and categorical intra-reading agreements for Amikacin, and there was no error when compared with the alternative methods. For Ciprofloxacin, the comparison between the Gold Standard and the Spectrophotometric reading showed 91.5% of essential agreement. In the categorical intra-reading analysis for Minocycline, there were 88.1 and 91.7% in the Gold Standard and in the Spectrophotometric readings, respectively, and 86.4% of concordance between them. High rates of categorical agreement were also observed on the Trimethoprim-Sulfamethoxazole analyses, with 93.7% for the Gold Standard, 84.9% for the Resazurin readings, and 80.5% between them. CONCLUSIONS: The alternative methods with Resazurin and Spectrophotometric readings showed high agreement rates with the Gold Standard.

Hemodialysis and Kidney Transplantation as Predisposing Conditions to Onychomycosis.

BACKGROUND/AIM: Onychomycosis (OM) is one of the commonest superficial fungal infections. Patients undergoing hemodialysis (HD) treatment and kidney transplant recipients (KTR) are considered at risk of contracting fungal infections, but the few published data do not reach the conclusion of whether they are predisposed to OM. This study aimed to determine the prevalence and etiology of OM in these patients and to determine the antifungal susceptibility profile of the isolated fungal species. METHODS: We recruited 149 HD patients, 187 KTR, and a control group comprising 174 patients attending an internal medicine service with other diseases than renal diseases. All patients underwent an examination of all toenails to check for the presence of OM. Antifungal susceptibility tests were performed following the Clinical and Laboratory Standards Institute (CLSI) recommendations. RESULTS: The prevalence rates of OM in HD patients (23.4%) and KTR (23.0%) were significantly higher than those in age- and sex-matched control groups (13.2%). In HD patients, OM was associated with diabetes but not with the duration of dialysis. In KTR, OM was more prevalent in those without diabetes and likely also in those using mycophenolate mofetil or azathioprine but was not associated with the duration of transplantation. Trichophyton rubrum was the most prevalent species (45.9%) followed by T. mentagrophytes (24.5%) and Candida parapsilosis (18.0%). Fluconazole, itraconazole, voriconazole, and terbinafine were all efficient against the isolates of dermatophyte, with terbinafine showing the lowest and fluconazole the highest minimal inhibitory concentrations. All isolates of C. parapsilosis were sensitive to the antifungals according to the CLSI criteria. CONCLUSION: We found a high prevalence of OM in HD and KTR patients and suggest that these conditions should be considered a risk factor of OM. All 4 antifungals evaluated in the study showed good in vitro activity against the etiologic agents.

Rhizopus arrhizus and Fusarium solani Concomitant Infection in an Immunocompromised Host.

Neutropenic patients are at risk of the development of hyalohyphomycosis and mucormycosis. Correct identification is essential for the initiation of the specific treatment, but concomitant mold infections are rarely reported. We report one unprecedented case of concomitant mucormycosis and fusariosis in a neutropenic patient with acute myeloid leukemia. The patient developed rhino-orbital infection by Rhizopus arrhizus and disseminated infection by Fusarium solani. The first culture from a sinus biopsy grew Rhizopus, which was consistent with the histopathology report of mucormycosis. A second sinus biopsy collected later during the patient's clinical deterioration was reported as hyalohyphomycosis, and the culture yielded F. solani. Due to the discordant reports, the second biopsy was reviewed and two hyphae types suggestive of both hyalohyphomycetes and mucormycetes were found. The dual mold infection was confirmed by PCR assays from paraffinized tissue sections. Increased awareness of the existence of dual mold infections in at-risk patients is necessary. PCR methods in tissue sections may increase the diagnosis of dual mold infections. In case of sequential biopsies showing discrepant results, mixed infections have to be suspected.

Does the Capsule Interfere with Performance of Matrix-Assisted Laser Desorption Ionization-Time of Flight Mass Spectrometry for Identification of Cryptococcus neoformans and Cryptococcus gattii?

We described the impact of the capsule size for Cryptococcus neoformans and Cryptococcus gattii identification at the species level by Bruker matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS). After experimental capsule size modulation, we observed that reducing the capsule size resulted in improved identification by Bruker MALDI-TOF MS across all of the reference strains analyzed.

Microbial colonization affects the efficiency of photovoltaic panels in a tropical environment.

Sub-aerial biofilm (SAB) development on solar panels was studied in São Paulo. After 6, 12 and 18 months' exposure, photovoltaic panels were covered by increasing proportions of organic matter (42%, 53% and 58%, respectively). Fungi were an important component of these biofilms; very few phototrophs were found. Major microorganisms detected were melanised meristematic ascomycetes and pigmented bacterial genera Arthrobacter and Tetracoccus. While diverse algae, cyanobacteria and bacteria were identified in biofilms at 6 and 12 months, diversity at a later stage was reduced to that typical for SAB: the only fungal group detected in 18 month biofilm was the meristematic Dothideomycetes and the only phototrophs Ulothrix and Chlorella. Photovoltaic modules showed significant power reductions after 6, 12 (both 7%) and 18 (11%) months. The lack of difference in power reduction between 6 and 12 months reflects the dual nature of soiling, which can result from the deposition of particulates as well as from SAB fouling. Although 12-month old SAB demonstrated an almost 10-fold increase in fungal colonization and a higher organic content, the larger non-microbial particles (above 10 µm), which were important for efficiency reduction of lightly-biofilmed panels, were removed by high rainfall just before the 12-month sampling.

An outbreak of invasive fusariosis in a children's cancer hospital.

Fusarium is considered an emerging pathogen, and there are few reports of fusariosis in children. The objective of this study was to describe an outbreak of invasive fusariosis in a children's cancer hospital. A neutropenic 17-year-old male patient hospitalized for 10 days for a relapse of acute myeloid leukaemia, under chemotherapy, presented fever without any other symptoms; a thoracic computerized tomography showed bilateral pulmonary nodules. During voriconazole treatment, 1-cm reddened and painful subcutaneous nodules appeared on arms and legs and the culture of a skin biopsy revealed F. solani. Another case occurred 11 days later and started an outbreak investigation. Water samples for cultures were collected from taps, showers and water reservoirs. Air from all patient rooms was sampled. Faucets and the drains of sinks and showers were swabbed and cultured. Environmental and clinical isolates were typed. There were 10 confirmed cases of infection caused by Fusarium spp. F. oxysporum and F. solani were isolated from water, swabs and air in patient rooms. Many control measures were instituted, but the outbreak was only controlled 1 year after the first case, when water filters filtering 0.2 µm were installed at the exit of all faucets and showers in all patient rooms (points-of-use). Typing demonstrated that clinical isolates of F. oxysporum were similar to those of the environment. In conclusion, to our knowledge this is the first reported outbreak of invasive fusariosis in children with oncohaematologic disease. It was controlled using 0.2-µm filters in all tap faucets and showers.

In vitro activity of potential old and new drugs against multidrug-resistant gram-negatives.

BACKGROUND: The aim of this study was to evaluate the in vitro susceptibility of MDR gram-negatives bacteria to old drugs such as polymyxin B, minocycline and fosfomycin and new drugs such as tigecycline. METHODS: One hundred and fifty-three isolates from 4 Brazilian hospitals were evaluated. Forty-seven Acinetobacter baumannii resistant to carbapenens harboring adeB, blaOxA23, blaOxA51, blaOxA143 and blaIMP genes, 48 Stenotrophomonas maltophilia including isolates resistant to levofloxacin and/or trimethoprim-sulfamethoxazole harboring sul-1, sul-2 and qnrMR and 8 Serratia marcescens and 50 Klebsiella pneumoniae resistant to carbapenens harboring blaKPC-2 were tested to determine their minimum inhibitory concentrations (MICs) by microdilution to the following drugs: minocycline, ampicillin-sulbactam, tigecycline, and polymyxin B and by agar dilution to fosfomycin according with breakpoint criteria of CLSI and EUCAST (fosfomycin). In addition, EUCAST fosfomycin breakpoint for Pseudomonas spp. was applied for Acinetobacter spp and S. maltophilia, the FDA criteria for tigecycline was used for Acinetobacter spp and S. maltophilia and the Pseudomonas spp polymyxin B CLSI criterion was used for S. maltophilia. RESULTS: Tigecycline showed the best in vitro activity against the MDR gram-negative evaluated, followed by polymyxin B and fosfomycin. Polymyxin B resistance among K. pneumoniae was detected in 6 isolates, using the breakpoint of MIC > 8 ug/mL. Two of these isolates were resistant to tigecycline. Minocycline was tested only against S. maltophilia and A. baumannii and showed excellent activity against both. CONCLUSIONS: Fosfomycin seems to not be an option to treat infections due to the A. baumannii and S. maltophilia isolates according with EUCAST breakpoint, on the other hand, showed excellent activity against S. marcescens and K. pneumoniae.

Isolation of Microsporum gypseum in soil samples from different geographical regions of Brazil, evaluation of the extracellular proteolytic enzymes activities (keratinase and elastase) and molecular sequencing of selected strains

A survey of Microsporum gypseum was conducted in soil samples in different geographical regions of Brazil. The isolation of dermatophyte from soil samples was performed by hair baiting technique and the species were identified by morphology studies. We analyzed 692 soil samples and the recuperating rate was 19.2%. The activities of keratinase and elastase were quantitatively performed in 138 samples. The sequencing of the ITS region of rDNA was performed in representatives samples. M. gypseum isolates showed significant quantitative differences in the expression of both keratinase and elastase, but no significant correlation was observed between these enzymes. The sequencing of the representative samples revealed the presence of two teleomorphic species of M. gypseum (Arthroderma gypseum and A. incurvatum). The enzymatic activities may play an important role in the pathogenicity and a probable adaptation of this fungus to the animal parasitism. Using the phenotypical and molecular analysis, the Microsporum identification and their teleomorphic states will provide a useful and reliable identification system.

Isolation of Microsporum gypseum in soil samples from different geographical regions of brazil, evaluation of the extracellular proteolytic enzymes activities (keratinase and elastase) and molecular sequencing of selected strains.

A survey of Microsporum gypseum was conducted in soil samples in different geographical regions of Brazil. The isolation of dermatophyte from soil samples was performed by hair baiting technique and the species were identified by morphology studies. We analyzed 692 soil samples and the recuperating rate was 19.2%. The activities of keratinase and elastase were quantitatively performed in 138 samples. The sequencing of the ITS region of rDNA was performed in representatives samples. M. gypseum isolates showed significant quantitative differences in the expression of both keratinase and elastase, but no significant correlation was observed between these enzymes. The sequencing of the representative samples revealed the presence of two teleomorphic species of M. gypseum (Arthroderma gypseum and A. incurvatum). The enzymatic activities may play an important role in the pathogenicity and a probable adaptation of this fungus to the animal parasitism. Using the phenotypical and molecular analysis, the Microsporum identification and their teleomorphic states will provide a useful and reliable identification system.

Malassezia spp. in acoustic meatus of bats (Molossus molossus) of the Amazon Region, Brazil.

The yeasts of the Malassezia genus are opportunistic microorganisms and can cause human and animal infections. They are commonly isolated from the skin and auricular canal of mammalians, mainly dogs and cats. The present study was aimed to isolate Malassezia spp. from the acoustic meatus of bats (Molossus molossus) in the Montenegro region, "Rondônia", Brazil. From a total of 30 bats studied Malassezia spp. were isolated in 24 (80%) animals, the breakdown by species being as follows (one Malassezia sp. per bat, N = 24): 15 (62.5%) M. pachydermatis, 5 (20.8%) M. furfur, 3 (12.5%) M. globosa and 1 (4.2%) M. sympodialis. This study establishes a new host and anatomic place for Malassezia spp., as it presents the first report ever of the isolation of this genus of yeasts in the acoustic meatus of bats.

Candiduria in a public hospital of São Paulo (1999-2004): characteristics of the yeast isolates.

The study involved 100 yeast isolates, obtained from urine samples provided by a Public Pediatric Hospital of São Paulo, Brazil, from 1999 to 2004. The most frequent species was Candida albicans, followed by C. tropicalis, C. glabrata and C. parapsilosis. In regard to virulence, 97% of the isolates showed index 3 for proteinase and 63% index 2 for phospholipase. The most frequent killer biotypes were 511 and 888.

Candiduria in a public hospital of São Paulo (1999-2004): characteristics of the yeast isolates

The study involved 100 yeast isolates, obtained from urine samples provided by a Public Pediatric Hospital of São Paulo, Brazil, from 1999 to 2004. The most frequent species was Candida albicans, followed by C. tropicalis, C. glabrata and C. parapsilosis. In regard to virulence, 97 percent of the isolates showed index 3 for proteinase and 63 percent index 2 for phospholipase. The most frequent killer biotypes were 511 and 888.

Estudou-se 100 amostras de leveduras, isoladas de urina, provenientes de Hospital Público Infantil de São Paulo Brasil, no período de 1999-2004. A espécie mais freqüente foi Candida albicans, seguida de C. tropicalis, C. glabrata e C. parapsilosis. Em relação à virulência, 97 por cento dos isolados apresentaram índice 3 para proteinase e, 63 por cento índice 2 para fosfolipase. Os biótipos "killer" mais freqüentes foram o 511 e 888.

Caracterizaçäo de amostras de Microsporum gypseum quanto ao seu perfil eletroforético e produçäo de enzimas extracelulares / The characterization of samples of Microsporum gypseum about its electrophoetic patterns and extracellular enzyme production

Objetivou-se caracterizar as amostras de M. gyseum isoladas de diferentes fontes quanto ao seu perfil eletroforético e à produçäo de enzimas extracelulares. A partir de fonte ambiental, animal e humana foram isoladas outo amostras de M. gypseum. Estas amostras foram avaliadas por curva de crescimento, caracterizaçäo de perfis eletroforéticos quanto ao número e intensidade de bandas e a produçäo de enzimas extracelulares empregando-se método em placas para avaliaçäo da presença de fosfolipase e proteinase e por eletroforese em gel-substrato (caseína e gelatina), além da determinaçäo quantitativa da atividade azocolítica e caseinolítica. Diferenças morfológicas näo foram observadas entre as amostras. As curvas de crescimento de duas amostras, uma de solo e outra de paciente foram bastante semelhantes, principalmente no período do 12§ ao 20§ dias de incubaçäo. O 19§ dia foi escolhido como o ideal para a obtençäo dos filtrados de uma cultura de todas as amostras. Os filtrados apresentaram concentraçäo protéica média de 2,15 mg/mL. Quanto ao perfil eletroforético, a análise revelou existir diferentes padröes protéicos entre as amostras de M. gypseum. A produçäo de enzimas extracelulares em placas foi baixa e näo permitiu observar diferenças entre as amostras nem tampouco relacioná-la com a patogenicidade. Os filtrados de cultura das diferentes amostras apresentaram variaçöes no número de enzimas expressas. As proteinases observadas em gel de gelatina variaram de 180 a 27 kDa e também variaram dependendo da amostra. As diferentes amostras de M. gypseum expressaram componentes protéicos com e sem atividade enzimática. As enzimas proteolíticas foram expressas sem uso de substratos indutores, significando que sua produçäo é independente da fonte protéica e portanto podem ser proteinases de amplo espectro de atividade

Aspectos gerais das micoses com caráter de zoonoses em pacientes com Aids / General aspects of zoonotic mycoses in Aids patients

As infecçöes oportunistas em pacientes com Aids representam a principal complicaçäo desta síndrome. As micoses, por sua vez, têm sempre estado presentes como uma manifestaçäo relevante. A criptococose, a histoplasmose e as dermatofitoses zoofílicas säo discutidas, em seus aspectos gerais, destacando-se o caráter zoonótico destas doenças

Rhodotorula Rubra em cabelos humanos simulando quadro de piedra branca / Similar white piedra by Rodotorula rubra

Resumo: Rhodotorula rubra é levedura oportunista, esando associada a uma variedade de processos patológicos no homem, é frequêntemente encontrada como contaminante da pele, pulmäo, urina, fezes e sistema nervoso central e com grande significado quando presente no sangue. As determinantes humanas causadas por Rhodotorula têm sido relatadasmas näo documentadas. O presente trabalho relata pela primeira vez em nosso meio, o isolamento desse microrganismo, como agente etiológico de infecçäo de cabelos humanos que ao exame clínico, simula pidra branca. Cabelos apresentando nódulos claros foram colhidos de paciente com 18 anos de idade. Em seguida classificados com KOH a 20 (por cento) entre lâmina e lamínula e observados no microscópio óptico comum e na microscopia eletrônicade varredura. Os cabelos foram semeados em ágar Sabnouraud-glicose contendo 200 mg/l de clonranfenicol, incubados à 25 graus C por 5 dias. Após o isolamento da levedura, a mesma foi identificada utilizando-se técnicas micológicas usuais. O exame microscópico dos pêlos infectados revelou em algumas regiöes o aumento do volume do cabelo, característico da tricorrexe nodosa. Em outras regiöes os pêlos já se apresentavam fragmentados e, com relativa facilidade podia-se observar leveduras arredondadas com brotamento em base estreita. A microscopia eletrônica de varredura pode-se observar mais detalhadamente a descriçäo acima. A levedura isolada foi identificada como Rhodotorula rubra, sem atividade lipolítica. É importante ressaltar que a referida levedura foi isolada repetidas vezes em cultura pura dos cabelos infectados. Este achado clínicoe laboratorial deve seer considerado, assim como deve merecer a devida atençäo dos pesquisadoresos isolamentos de R. rubra os quais permitiräo um correto diagnóstico micológico e terapêutico adequada (au)