Unraveling the epidemiology of urinary tract infections in neonates: Perspective from a Brazilian NICU.

BACKGROUND: Urinary tract infections (UTIs) can lead to neonatal complications like sepsis, worsened by empirical treatment, contributing to antimicrobial resistance (AMR). This study examined the incidence, etiology, risk factors, and antimicrobial susceptibility of uropathogens in a Neonatal Intensive Care Unit (NICU) in Brazil. METHODS: Medical records of neonates hospitalized in the NICU from January 2015 to June 2022 were retrospectively analyzed through the National Healthcare Safety Network system. RESULTS: Among 1,474 neonates, 3.9% developed UTI, with an alarming 24-fold increase in incidence from 2015 to 2021. Genitourinary complications (odds ratio = 4.8) were a major risk factor. Of the 71 uropathogens, 74.6% were Gram-negative bacteria (GNB), 21.2% Gram-positive bacteria (GPB), and 4.2% Candida albicans. AMR was notable, with 13.3% of GPB and 20.7% of GNB exhibiting multidrug-resistant (MDR), while 6.6% of GPB and 1.9% of GNB showed extensive drug-resistant (XDR). UTI was associated with prolonged hospitalization (16-59 days). In 57 neonates with UTI, 40.3% had bloodstream infections, elevating the risk of death (odds ratio = 1.8). CONCLUSIONS: The study underscores the urgency of implementing infection prevention and control measures in the NICU to curb rising UTI incidences, combat AMR, and mitigate severe complications in critically ill neonates.

Epidemiological indicators and predictors of lethality associated with fungal infections in a NICU: an historical series.

OBJECTIVES: Fungal infections (FI) pose a public health concern and significantly increase mortality rates, especially within Neonatal Intensive Care Units (NICU). Thus, this study aimed to investigate epidemiological indicators, risk factors, and lethality predictors associated with FI in a NICU. METHODS: This study included 1,510 neonates admitted to the NICU of a reference hospital in Brazil between 2015 and 2022. Demographic data, such as sex, birth weight, gestational age, and use of invasive devices were analyzed. RESULTS: Thirty neonates developed invasive FI, totaling 33 episodes and an incidence of 1.2 per 1,000 patient days. Candida albicans was the most frequent species (52.9 %), the bloodstream was the most affected site (78.9 %), and 72.7 % of infections occurred between 2015 and 2018. The lethality rate associated with FI was 33.3 %, and 90 % of deaths occurred within 30 days of diagnosis of infection. Weight < 750 g, prolonged hospital stay, use of parenteral nutrition, and broad-spectrum antimicrobials were independent risk factors for infection occurrence, especially glycopeptides and 4th generation cephalosporins, having a considerable role in the increase in fungal infections. Weight < 750 g was considered a significant predictor of lethality, and C. albicans had the highest lethality rate (40 %). CONCLUSION: These findings highlight the elevated lethality rate associated with these infections, reinforcing the importance of developing strategies to control FI within NICU.

Impact of intestinal colonization by Gram-negative bacteria on the incidence of bloodstream infections and lethality in critically ill neonates.

BACKGROUND: Early detection of antimicrobial-resistant microorganisms is crucial to prevent subsequent invasive infections and contain their spread in the Neonatal Intensive Care Unit (NICU). This study aims to investigate the association between intestinal colonization (IC) by Gram-negative bacteria and the risk of bloodstream infection (BSI) in critically ill neonates. METHODS: Data from the electronic medical records of 678 newborns admitted to a NICU Brazilian between 2018 and 2022 were retrospectively analyzed. Participants were monitored by the National Health Security Network. RESULTS: Among neonates, 6.9 % had IC (56.9 % attributed to Acinetobacter baumannii); of these, 19.1 % developed BSI (66.7 % by Staphylococcus spp.). Within the A. baumannii colonization, 34.5 % occurred during an outbreak in September 2021. Colonized individuals had a longer mean length of stay (49.3 ± 26.4 days) and higher mortality rate (12.8 %) compared to non-colonized individuals (22.2 ± 16.9 days; 6.7 %, respectively). Previous use of antimicrobials and invasive devices significantly increased the risk of colonization. Colonization by drug-resistant microorganisms, along with the occurrence of BSI, was associated with increased mortality and reduced survival time. CONCLUSIONS: IC contributed to the incidence of BSI, leading to more extended hospital stays and higher mortality rates. Its early detection proved to be essential to identify an outbreak and control the spread of resistant microorganisms within the NICU.

Antimicrobial, Antivirulence, and Antiparasitic Potential of Capsicum chinense Jacq. Extracts and Their Isolated Compound Capsaicin.

Bacterial, fungal, and parasitic infections increase morbimortality rates and hospital costs. This study aimed to assess the antimicrobial and antiparasitic activities of the crude extract from the seeds and peel of the pepper Capsicum chinense Jacq. and of the isolated compound capsaicin and to evaluate their ability to inhibit biofilm formation, eradicate biofilm, and reduce hemolysin production by Candida species. The crude ethanolic and hexane extracts were obtained by maceration at room temperature, and their chemical compositions were analyzed by liquid chromatography coupled to mass spectrometry (LC-MS). The antimicrobial activity of the samples was evaluated by determining the minimum inhibitory concentration. Inhibition of biofilm formation and biofilm eradication by the samples were evaluated based on biomass and cell viability. Reduction of Candida spp. hemolytic activity by the samples was determined on sheep blood agar plates. The antiparasitic action of the samples was evaluated by determining their ability to inhibit Toxoplasma gondii intracellular proliferation. LC-MS-ESI analyses helped to identify organic and phenolic acids, flavonoids, capsaicinoids, and fatty acids in the ethanolic extracts, as well as capsaicinoids and fatty acids in the hexane extracts. Antifungal action was more evident against C. glabrata and C. tropicalis. The samples inhibited biofilm formation and eradicated the biofilm formed by C. tropicalis more effectively. Sub-inhibitory concentrations of the samples significantly reduced the C. glabrata and C. tropicalis hemolytic activity. The samples only altered host cell viability when tested at higher concentrations; however, at non-toxic concentrations, they reduced T. gondii growth. In association with gold standard drugs used to treat toxoplasmosis, capsaicin improved their antiparasitic activity. These results are unprecedented and encouraging, indicating the Capsicum chinense Jacq. peel and seed extracts and capsaicin display antifungal and antiparasitic activities.

Inappropriate empirical antimicrobial treatment in bloodstream infections patients in the era of multidrug resistance / Terapia Empírica Inapropriada no Tratamento de Infecções de Corrente Sanguínea na Era da Multirresistência Antimicrobiana / Tratamiento antimicrobiano empírico inadecuado en pacientes con infecciones del torrente sanguíneo

Background and objectives: Bloodstream infection (BSI) by multidrug-resistant Pseudomonas aeruginosa is a severe infection. This study aimed to evaluate and identify the predictors of mortality in patients who had bloodstream infection by carbapenem-resistant P. aeruginosa. Methods: This is a retrospective cohort study, approved by Committee of Ethics in Research with Human Participants, which included 87 consecutive patients hospitalized in a referral hospital in Brazil. Clinical and demographic information about each patient were obtained from hospital records. The Student's T-test was used to compare continuous variables and x2 or Fisher's exact tests to compare categorical variables. To determine independent risk factors for 30-day mortality, a multiple logistic regression model was used. A survival curve was constructed using the Kaplan­Meier method. Results: Among the patients, 87.3% use antibiotics previously, 60.9% received inadequate empirical treatment, and the 30-day mortality rate was 57.5%. Inappropriate antibiotic empirical therapy was independently associated with a 30-days death and mortality rate. Conclusion: These findings can show some insights about the relationship between higher mortality and inappropriate empirical therapy for patients with BSI by P. aeruginosa. There is a need for better diagnostic tests and infection control programs should focus on de-escalation the antibiotic inappropriate therapy, mainly in BSI caused by carbapenem-resistant P. aeruginosa.(AU)

Justificativa e objetivos: Infecção da corrente sanguínea (ICS) por Pseudomonas aeruginosa multirresistente é grave. Este estudo teve como objetivo avaliar e identificar os preditores de mortalidade em pacientes admitidos em uma Unidade de Terapia Intensiva que apresentaram infecção da corrente sanguínea por P. aeruginosa resistente aos carbapenêmicos. Métodos: Trata-se de um estudo de coorte retrospectivo, aprovado pelo Comitê de Ética em Pesquisa com Seres Humanos, que incluiu 87 pacientes consecutivos internados em um hospital de referência no Brasil. As informações clínicas e demográficas de cada paciente foram obtidas através de análise dos prontuários dos pacientes. O teste T de Student foi usado para comparar variáveis contínuas e o teste x2 ou exato de Fisher para comparar variáveis categóricas. Para determinar fatores de risco independentes para mortalidade em 30 dias, foi utilizado um modelo de regressão logística múltipla. Uma curva de sobrevida foi construída pelo método de Kaplan-Meier. Resultados: Do total de pacientes, 87,3% faziam uso prévio de antibióticos, 60,9% receberam tratamento empírico inadequado e a mortalidade em 30 dias foi de 57,5%. A terapia empírica inadequada foi fator de risco independente para mortalidade. Conclusão: Esses achados revelam alguns insights sobre a relação entre maior mortalidade e terapia empírica inadequada para pacientes com ICS por P. aeruginosa. Além disso, destacam a necessidade de melhores testes diagnósticos e os programas de controle de infecção devem se concentrar na redução da terapia inadequada com antibióticos, principalmente na ICS causada por P. aeruginosa resistente a carbapenêmicos.(AU)

Justificación y objetivos: La infección del torrente sanguíneo por Pseudomonas aeruginosa multirresistente es grave. Este estudio tuvo como objetivo evaluar e identificar predictores de mortalidad en pacientes ingresados en una Unidad de Cuidados Intensivos que presentaban infección del torrente sanguíneo por P. aeruginosa resistente a carbapenémicos. Métodos: Se trata de un estudio de cohorte retrospectivo, aprobado por el Comité de Ética en Investigación con Participantes Humanos, que incluyó 87 pacientes consecutivos ingresados en un hospital de referencia en Brasil. La información clínica y demográfica de cada paciente se obtuvo mediante el análisis de las historias clínicas de los pacientes. Se utilizó la prueba t de Student para comparar variables continuas y x2 o prueba exacta de Fisher para comparar variables categóricas. Para determinar los factores de riesgo independientes para la mortalidad a los 30 días, se utilizó un modelo de regresión logística múltiple. Se construyó una curva de supervivencia utilizando el método de Kaplan-Meier. Resultados: Del total de pacientes, el 87,3% utilizaba antibióticos previamente, el 60,9% recibió tratamiento empírico inadecuado y la tasa de mortalidad a los 30 días fue del 57,5%. La terapia empírica inadecuada fue un factor de riesgo independiente de mortalidad. Conclusión: Estos hallazgos revelan algunos conocimientos sobre la relación entre el aumento de la mortalidad y la terapia empírica inadecuada para los pacientes con infección del torrente sanguíneo por P. aeruginosa. Además, destacan la necesidad de mejores pruebas de diagnóstico y los programas de control de infecciones deben centrarse en reducir la terapia con antibióticos inapropiados, particularmente en infección del torrente sanguíneo causados por P. aeruginosa resistente a carbapenémicos.(AU)

Antibacterial Activity of Isobavachalcone (IBC) Is Associated with Membrane Disruption.

Isobavachalcone (IBC) is a natural prenylated chalcone with a broad spectrum of pharmacological properties. In this work, we newly synthesized and investigated the antibacterial activity of IBC against Gram-positive, Gram-negative and mycobacterial species. IBC was active against Gram-positive bacteria, mainly against Methicillin-Susceptible Staphylococcus aureus (MSSA) and Methicillin-Resistant Staphylococcus aureus (MRSA), with minimum inhibitory concentration (MIC) values of 1.56 and 3.12 µg/mL, respectively. On the other hand, IBC was not able to act against Gram-negative species (MIC > 400 µg/mL). IBC displayed activity against mycobacterial species (MIC = 64 µg/mL), including Mycobacterium tuberculosis, Mycobacterium avium and Mycobacterium kansasii. IBC was able to inhibit more than 50% of MSSA and MRSA biofilm formation at 0.78 µg/mL. Its antibiofilm activity was similar to vancomycin, which was active at 0.74 µg/mL. In order to study the mechanism of the action by fluorescence microscopy, the propidium iodide (PI) and SYTO9 fluorophores indicated that IBC disrupted the membrane of Bacillus subtilis. Toxicity assays using human keratinocytes (HaCaT cell line) showed that IBC did not have the capacity to reduce the cell viability. These results suggested that IBC is a promising antibacterial agent with an elucidated mode of action and potential applications as an antibacterial drug and a medical device coating.

Inanimate Surfaces and Air Contamination with Multidrug Resistant Species of Staphylococcus in the Neonatal Intensive Care Unit Environment.

BACKGROUND: Contamination of the hospital environment with multi-resistant (MDR) Staphylococcus increases the risk of infection. The aim of this study is to identify the MDR species of Staphylococcus on inanimate surfaces, in air, and in clinical samples, and analyze the risk factors that correlate with the occurrence of infections in a Neonatal Intensive Care Unit. METHODS: Samples of inanimate surfaces and air were taken using a premoistened swab (0.9% sodium chloride) and spontaneous air sedimentation, respectively. The clinical isolates were recovered from infected neonates. The isolates (environmental and clinical) were identified by matrix-assisted laser desorption ionization-time of flight and the resistance profile was calculated using the disk diffusion agar technique. RESULTS: In total, 181 isolates were obtained, 93 from (surfaces), 18 from the air, and 70 clinical samples. S. epidermidis was the most frequent species (66.8%), and the failure rate in air cleaning was 100%. More than 60% of the isolates were MDR, and the majority of clinical isolates (60.4%) had a resistance profile identical to that of the environmental isolates. CONCLUSION: Staphylococcus spp. were found in most of the analyzed samples, with a high frequency of MDR isolates, demonstrating the importance of the hospital environment as a reservoir, and the need for infection control measures, and rational use of antimicrobials.

Risk factors associated with mortality among patients who had candidemia in a university hospital.

INTRODUCTION: Bloodstream infection due to Candida spp. is a primary cause of morbidity and mortality in tertiary hospitals. METHODS: In this retrospective study, we included patients with a positive blood culture for Candida spp. after 48 h of hospitalization. RESULTS: A total of 335 patients who had candidemia were included in this study. Risk factors associated with mortality were hospitalization in internal medicine units and surgical clinics, age >60 years, mechanical ventilation, orotracheal intubation, hemodialysis, corticosteroids use, and C. parapsilosis infection. CONCLUSIONS: This study highlights the importance of health care related to invasive procedures and actions to improve patient immunity.

Risk factors associated with mortality among patients who had candidemia in a university hospital

Abstract INTRODUCTION: Bloodstream infection due to Candida spp. is a primary cause of morbidity and mortality in tertiary hospitals. METHODS: In this retrospective study, we included patients with a positive blood culture for Candida spp. after 48 h of hospitalization. RESULTS A total of 335 patients who had candidemia were included in this study. Risk factors associated with mortality were hospitalization in internal medicine units and surgical clinics, age >60 years, mechanical ventilation, orotracheal intubation, hemodialysis, corticosteroids use, and C. parapsilosis infection. CONCLUSIONS: This study highlights the importance of health care related to invasive procedures and actions to improve patient immunity.

Isolation of Cryptococcus species from the external environments of hospital and academic areas.

INTRODUCTION: Fungi of the genus Cryptococcus are cosmopolitan and may be agents of opportunistic mycoses in immunocompromised and sometimes immunocompetent individuals. Cryptococcus species are frequently isolated from trees and bird excreta in the environment and infection occurs by inhalation of propagules dispersed in the air. The aim was to investigate Cryptococcus species in bird excreta and tree hollows located in a university hospital area and in an academic area of a university campus. METHODOLOGY: A total of 40 samples of bird excreta and 41 samples of tree hollows were collected. The identification of the isolates was done by classical methodology and matrix-assisted laser desorption/ionization time-of-flight mass spectrometry. RESULTS: Twenty (62.5%) isolates of Cryptococcus were found in bird excreta and 12 (37.5%) in tree hollows. C. laurentii (currently Papiliotrema laurentii) was the most frequent species in both samples, being found in 5 samples of excreta and in 8 tree hollows. The diversity of species found in excreta (C. laurentii, C. albidus [currently Naganishia albida], C. liquefaciens [currently N. liquefaciens], C. friedmanii [currently N. friedmannii] and others) was higher than in tree hollows (C. laurentii, C. flavescens [currently Papiliotrema flavescens], and other yeasts). CONCLUSION: Many Cryptococcus species were isolated from excreta and tree hollows, and this fact is important for understanding the environmental epidemiology of those emerging pathogens for public health, as a way to implement surveillance actions and control of cryptococcosis.

RELATED FACTORS FOR COLONIZATION BY Candida SPECIES IN THE ORAL CAVITY OF HIV-INFECTED INDIVIDUALS.

UNLABELLED: The colonization of the oral cavity is a prerequisite to the development of oropharyngeal candidiasis. AIMS: The aims of this study were: to evaluate colonization and quantify Candida spp. in the oral cavity; to determine the predisposing factors for colonization; and to correlate the levels of CD4+ cells and viral load with the yeast count of colony forming units per milliliter (CFU/mL) in HIV-positive individuals treated at a University Hospital. Saliva samples were collected from 147 HIV patients and were plated on Sabouraud Dextrose Agar (SDA) and chromogenic agar, and incubated at 30 ºC for 72 h. Colonies with similar morphology in both media were counted and the result expressed in CFU/mL. RESULTS: Of the 147 HIV patients, 89 had positive cultures for Candida spp., with a total of 111 isolates, of which C. albicans was the most frequent species (67.6%), and the mean of colonies counted was 8.8 × 10³ CFU/mL. The main predisposing factors for oral colonization by Candida spp. were the use of antibiotics and oral prostheses. The use of reverse transcriptase inhibitors appears to have a greater protective effect for colonization. A low CD4+ T lymphocyte count is associated with a higher density of yeast in the saliva of HIV patients.

RELATED FACTORS FOR COLONIZATION BY Candida SPECIES IN THE ORAL CAVITY OF HIV-INFECTED INDIVIDUALS / Fatores relacionados a colonização da cavidade bucal de indivíduos portadores do HIV por espécies de Candida

The colonization of the oral cavity is a prerequisite to the development of oropharyngeal candidiasis. Aims: The aims of this study were: to evaluate colonization and quantify Candida spp. in the oral cavity; to determine the predisposing factors for colonization; and to correlate the levels of CD4+ cells and viral load with the yeast count of colony forming units per milliliter (CFU/mL) in HIV-positive individuals treated at a University Hospital. Saliva samples were collected from 147 HIV patients and were plated on Sabouraud Dextrose Agar (SDA) and chromogenic agar, and incubated at 30 ºC for 72 h. Colonies with similar morphology in both media were counted and the result expressed in CFU/mL. Results: Of the 147 HIV patients, 89 had positive cultures for Candida spp., with a total of 111 isolates, of which C. albicans was the most frequent species (67.6%), and the mean of colonies counted was 8.8 × 10³ CFU/mL. The main predisposing factors for oral colonization by Candida spp. were the use of antibiotics and oral prostheses. The use of reverse transcriptase inhibitors appears to have a greater protective effect for colonization. A low CD4+ T lymphocyte count is associated with a higher density of yeast in the saliva of HIV patients.

RESUMO A colonização da cavidade oral pode ser considerada um pré-requisito para o desenvolvimento de candidíase orofaríngea. Os objetivos deste estudo foram: avaliar e quantificar espécies de Candidaisoladas da cavidade oral, para determinar os fatores predisponentes para a colonização, e correlacionar os níveis de células CD4+ e carga viral em indivíduos HIV-positivos atendidos em um hospital universitário. Foram coletadas amostras de saliva de 147 pacientes portadores do HIV, as quais foram semeadas em Ágar Sabouraud Dextrose (ASD) e ágar cromogênico e incubadas a 30 °C por 72 horas. As colônias com morfologia semelhante em ambos os meios foram contadas e o resultado expresso em unidade formadora de colônias por mililitro (UFC/mL). Dos 147 pacientes HIV positivos, 89 apresentaram culturas positivas para Candidaspp., totalizando 111 isolados, e C. albicansfoi a espécie mais frequente (67,6%). A contagem média de colônias foi de 8.8 × 10³ UFC/mL. Os principais fatores predisponentes para colonização oral por Candidaspp. foram a utilização de antibióticos e de próteses orais. O uso de antirretroviral da classe de inibidores da transcriptase reversa pareceu ter maior efeito protetor para a colonização. Baixa contagem de linfócitos T CD4+ está relacionada com maior densidade de leveduras na saliva de indivíduos HIV positivos.

FREQUENCY OF Candida SPECIES IN A TERTIARY CARE HOSPITAL IN TRIANGULO MINEIRO, MINAS GERAIS STATE, BRAZIL.

Infections by Candida species are a high-impact problem in public health due to their wide incidence in hospitalized patients. The goal of this study was to evaluate frequency, susceptibility to antifungals, and genetic polymorphism of Candida species isolated from clinical specimens of hospitalized patients. The Candida isolates included in this study were obtained from blood cultures, abdominal fluids, and central venous catheters (CVC) of hospitalized patients at the Clinical Hospital of the Federal University of Uberlândia during the period of July 2010 - June 2011. Susceptibility tests were conducted by the broth microdilution method. The RAPD-PCR tests used employed initiator oligonucleotides OPA09, OPB11, and OPE06. Of the 63 Candida isolates, 18 (28.5%) were C. albicans, 20 (31.7%) were C. parapsilosis complex species, 14 (22.2%) C. tropicalis, four (6.4%) C. glabrata, four (6.4%) C. krusei, two (3.3%) C. kefyr, and one (1.6%) C. lusitaniae. In vitro resistance to amphotericin B was observed in 12.7% of isolates. In vitro resistance to azoles was not detected, except for C. krusei. The two primers, OPA09 and OPB11, were able to distinguish different species. Isolates of C. albicans and C. parapsilosis complex species presented six and five clusters, respectively, with the OPA09 marker by RAPD-PCR, showing the genetic variability of the isolates of those species. It was concluded that members of the C. parapsilosis complex were the most frequent species found, and most isolates were susceptible to the antifungals amphotericin B, flucozanole, and itraconazole. High genetic polymorphisms were observed for isolates of C. albicans and C. parapsilosis complex species, mainly with the OPA09 marker.

FREQUENCY OF Candida SPECIES IN A TERTIARY CARE HOSPITAL IN TRIANGULO MINEIRO, MINAS GERAIS STATE, BRAZIL / Frequência de espécies de Candida em hospital terciário do Triângulo Mineiro, Minas Gerais, Brasil

Infections by Candida species are a high-impact problem in public health due to their wide incidence in hospitalized patients. The goal of this study was to evaluate frequency, susceptibility to antifungals, and genetic polymorphism of Candida species isolated from clinical specimens of hospitalized patients. The Candida isolates included in this study were obtained from blood cultures, abdominal fluids, and central venous catheters (CVC) of hospitalized patients at the Clinical Hospital of the Federal University of Uberlândia during the period of July 2010 - June 2011. Susceptibility tests were conducted by the broth microdilution method. The RAPD-PCR tests used employed initiator oligonucleotides OPA09, OPB11, and OPE06. Of the 63 Candida isolates, 18 (28.5%) were C. albicans, 20 (31.7%) were C. parapsilosis complex species, 14 (22.2%) C. tropicalis, four (6.4%) C. glabrata, four (6.4%) C. krusei, two (3.3%) C. kefyr, and one (1.6%) C. lusitaniae. In vitro resistance to amphotericin B was observed in 12.7% of isolates. In vitro resistance to azoles was not detected, except for C. krusei. The two primers, OPA09 and OPB11, were able to distinguish different species. Isolates of C. albicans and C. parapsilosis complex species presented six and five clusters, respectively, with the OPA09 marker by RAPD-PCR, showing the genetic variability of the isolates of those species. It was concluded that members of the C. parapsilosis complex were the most frequent species found, and most isolates were susceptible to the antifungals amphotericin B, flucozanole, and itraconazole. High genetic polymorphisms were observed for isolates of C. albicans and C. parapsilosis complex species, mainly with the OPA09 marker.

As infecções causadas por espécies de Candida são problema de grande impacto para a saúde pública, devido à alta incidência em pacientes hospitalizados e como causa de mortalidade. O presente estudo teve como objetivo avaliar a frequência de Candida spp. isoladas de pacientes hospitalizados, assim como a sensibilidade aos antifúngicos e o polimorfismo genético por RAPD-PCR. Os microrganismos incluíram isolados de hemocultura, líquido abdominal e ponta de cateter venoso central de pacientes internados no Hospital de Clínicas da Universidade Federal de Uberlândia, região do Triângulo Mineiro, Minas Gerais, Brasil, no período de julho de 2010-junho de 2011. Os testes de sensibilidade aos antifúngicos foram realizados por microdiluição em caldo e na análise por RAPD-PCR foram utilizados os oligonucleotídeos OPA09, OPB11, e OPE06. Dos 63 isolados, 18 (28,5%) foram C. albicans, 20 (31,7%) C. parapsilosis, 14 (22,2%) C. tropicalis, quatro (6,4%) C. glabrata, quatro (6,4%) C. krusei, dois (3,3%) C. kefyr, e um (1,6%) C. lusitaniae. Resistência in-vitro à anfotericina B foi observada em 12,7% dos isolados. Não foi observada resistência in-vitro aos azólicos, exceto para os isolados de C. krusei. Os oligonucleotídeos OPA09 e OPB11 possibilitaram distinguir diferentes espécies. Isolados de C. albicans apresentaram seis clusters e o complexo C. parapsilosis, cinco clusters, com o iniciador OPA09, por RAPD-PCR, mostrando a variabilidade genética daquelas espécies. Conclui-se que o complexo C. parapsilosis foi a espécie mais frequente, e a maioria dos isolados foi sensível in vitro aos antifúngicos testados. Alto polimorfismo genético foi observado para os isolados de C. albicans e complexo C. parapsilosis, principalmente com o oligonucleotídeo OPA09.

Enzymatic and hemolytic activity in different Candida species / La actividad enzimática y hemolítica en diferentes especies de Candida

Background: Candida species, in conditions of microbiota imbalance or decreased immune defenses, may be one of the main human fungal pathogens. Virulence factors constitute the mechanisms used by the fungus to avoid host defenses. Aims: This study aimed to investigate the in vitro production of virulence factors, such as hemolytic activity, and deoxyribonuclease (DNase), proteinase, and phospholipase activities in Candida spp. Methods: Fifty clinical isolates were analyzed for virulence factors: Candida albicans (15), Candida tropicalis (15), Candida parapsilosis (10), Candida glabrata (5), and Candida krusei (5). Hemolytic activity was determined in Sabouraud dextrose agar plates containing 3% glucose and 7% sheep red cells. Culture media containing, respectively, agar-base DNA, egg yolk, and bovine albumin were used to determine DNase, phospholipase and proteinase activities, respectively. Results: Forty-eight (96%) of 50 isolates showed hemolytic activity, with 10 (20%) positive for DNase, 19 (38%) for proteinase, and 16 (32%) for phospholipase. Statistically significant differences were observed between species for phospholipase (p < 0.0001) and proteinase (p < 0.05) production. Conclusions: It is concluded that all species had hemolytic activity. DNase activity was detected in all species except in C. glabrata; proteinase activity was detected in C. albicans, C. tropicalis, and C. parapsilosis; and phospholipase activity was observed in C. albicans and C. tropicalis (AU)

Antecedentes: Las levaduras del género Candida, en condiciones de desequilibrio de la microbiota o de disminución de las defensas inmunológicas, pueden ser uno de los principales patógenos fúngicos del hombre. Los factores de virulencia constituyen los mecanismos utilizados por el hongo para evadir las defensas del huésped. Objetivos: Este estudio tiene como objetivo investigar la producción in vitro de algunos factores de virulencia, como la actividad hemolítica, y las actividades desoxirribonucleasa (DNasa), proteinasa y fosfolipasa en Candidaspp. Métodos: Se analizaron 50 aislamientos clínicos: Candida albicans (15), Candida tropicalis (15), Candida parapsilosis (10), Candida glabrata (5), y Candida krusei (5). La actividad hemolítica fue determinada en placas de agar glucosado de Sabouraud, con glucosa al 3% y un 7% de hematíes de oveja. Los medios de cultivo de agar-ADN, yema de huevo y albúmina bovina fueron utilizados para determinar las actividades DNasa, fosfolipasa y proteinasa, respectivamente. Resultados: De los 50 aislamientos, 48 (96%) presentaron actividad hemolítica, 10 (20%) fueron positivos para DNasa, 19 (38%) para proteinasa y 16 (32%) para fosfolipasa. Se observaron diferencias estadísticamente significativas entre las especies para las actividades fosfolipasa (p < 0,0001) y proteasa (p < 0,05). Conclusiones: Se concluye que todas las especies estudiadas poseen actividad hemolítica. La actividad DNasa fue detectada en todas las especies, excepto en Candida glabrata; la actividad proteinasa fue detectada en C. albicans, C. tropicalis y C. parapsilosis, y la actividad fosfolipasa se observó en C. albicans y C. tropicalis (AU)

Enzymatic and hemolytic activity in different Candida species.

BACKGROUND: Candida species, in conditions of microbiota imbalance or decreased immune defenses, may be one of the main human fungal pathogens. Virulence factors constitute the mechanisms used by the fungus to avoid host defenses. AIMS: This study aimed to investigate the in vitro production of virulence factors, such as hemolytic activity, and deoxyribonuclease (DNase), proteinase, and phospholipase activities in Candida spp. METHODS: Fifty clinical isolates were analyzed for virulence factors: Candida albicans (15), Candida tropicalis (15), Candida parapsilosis (10), Candida glabrata (5), and Candida krusei (5). Hemolytic activity was determined in Sabouraud dextrose agar plates containing 3% glucose and 7% sheep red cells. Culture media containing, respectively, agar-base DNA, egg yolk, and bovine albumin were used to determine DNase, phospholipase and proteinase activities, respectively. RESULTS: Forty-eight (96%) of 50 isolates showed hemolytic activity, with 10 (20%) positive for DNase, 19 (38%) for proteinase, and 16 (32%) for phospholipase. Statistically significant differences were observed between species for phospholipase (p<0.0001) and proteinase (p<0.05) production. CONCLUSIONS: It is concluded that all species had hemolytic activity. DNase activity was detected in all species except in C. glabrata; proteinase activity was detected in C. albicans, C. tropicalis, and C. parapsilosis; and phospholipase activity was observed in C. albicans and C. tropicalis.

Sensibilidade de isolados de candida spp. a antifúngicos por disco-difusão em ágar e microdiluição em caldo / Antifungal susceptibility of isolates of candida spp. by disk diffusion and broth microdilution

Os testes de sensibilidade aos antifúngicos realizados pelo método de disco-difusão em ágar são práticos e bem conhecidos pelos profissionais do laboratório de microbiologia, entretanto apresentam particularidades que os diferem dos testes realizados para bactérias. O objetivo deste trabalho foi comparar as técnicas de disco-difusão em ágar e microdiluição em caldo na determinação da sensibilidade in vitro de isolados de Candida spp. a antifúngicos. Foram analisados 63 isolados clínicos de leveduras, que incluíram as espécies Candida parapsilosis complex (n = 20), Candida albicans (n = 18), Candida tropicalis (n = 14), Candida glabrata (n = 4), Candida krusei (n = 4), Candida kefyr (n = 2) e Candida lusitaniae (n = 1). As técnicas de disco-difusão em ágar e de microdiluição em caldo foram utilizadas para testar a sensibilidade em relação aos antifúngicos fluconazol, itraconazol e anfotericina B. A sensibilidade ao voriconazol foi determinada somente pela técnica de disco-difusão. Os halos ao redor dos discos de fluconazol variaram de 14 mm a 50 mm, e a CIM de 0,125 µg/mL a 32 µg/mL; para itraconazol, os halos variaram de 9 mm a 27 mm e a CIM de 0,03 µg/mL a 0,25 µg/mL; para anfotericina B, 9 mm a 21mm e 0,5 µg/mL a 2 µg/mL, respectivamente; para voriconazol, o diâmetro dos halos variaram de 19 mm a 50 mm. Para as três espécies, C. albicans, C. parapsilosis e C. tropicalis, a técnica de disco-difusão apresentou boa concordância com a microdiluição, especialmente em relação ao fluconazol, representando, assim, um recurso importante para os laboratórios reportarem os resultados dos testes de sensibilidade dos isolados dessas espécies ao fluconazol.

Antimicrobial susceptibility tests performed by disk diffusion method are practical and well known by professionals that work in the microbiology laboratory. The disk diffusion methodology used to verify the susceptibility of fungi to antifungal agents, however, has characteristics that differ from the tests for bacteria. The objective of this study was to evaluate the disk diffusion method to determine the in vitro susceptibility to antifungal agents of Candida species. We analyzed 63 clinical isolates of yeasts, which included Candida parapsilosis complex species (n = 20), Candida albicans (n = 18), Candida tropicalis (n = 14), Candida glabrata (n = 4), Candida krusei (n = 4), Candida kefyr (n = 2) and Candida lusitaniae (n = 1). The susceptibility tests to antifungal drugs was performed by disk diffusion methods and broth microdilution for antifungal fluconazole, itraconazole and amphotericin B. Voriconazole was used to test the susceptibility only by the disk diffusion method. The inhibition halos of growth around disks of fluconazole ranged from 14 mm to 50 mm and the MIC from 0.125 µg/mL to 32 µg/mL, for itraconazole, halos ranged from 9 mm to 27 mm and the MIC from 0.03 µg/mL to 0.25 µg/mL, for amphotericin B, 9 mm to 21 mm and 0.5 µg/mL to 2 µg/mL, respectively. The diameter of voriconazole disks varied from 19 mm to 50 mm. For the three species, C. albicans, C. parapsilosis and C. tropicalis, the disk diffusion method showed good agreement with the microdilution, especially to fluconazole, thus representing an important resource for medical laboratories reporting results of susceptibility testing of isolates of these species to fluconazole.

Different culture media containing methyldopa for melanin production by Cryptococcus species.

INTRODUCTION: Melanin production by species of Cryptococcus is widely used to characterize C. neoformans complex in mycology laboratories. This study aims to test the efficacy of methyldopa from pharmaceutical tablet as a substrate for melanin production, to compare the production of melanin using different agar base added with methyldopa, and to compare the melanin produced in those media with that produced in Niger seed agar and sunflower seed agar by C. neoformans, C. laurentii, and C. albidus. Two isolates of each species, C. neoformans, C. laurentii, and C. albidus, and one of Candida albicans were used to experimentally detect conditions for melanin production. METHODS: The following media were tested: Mueller-Hinton agar (MHA), brain and heart infusion agar (BHIA), blood agar base (BAB), and minimal medium agar (MMA), all added with methyldopa, and the media Niger seed agar (NSA) and sunflower seed agar (SSA). RESULTS: All isolates grew in most of the culture media after 24h. Strains planted on media BAB and BHIA showed growth only after 48h. All isolates produced melanin in MMA, MHA, SSA, and NSA media. CONCLUSIONS: Methyldopa in the form pharmaceutical tablet can be used as a substrate for melanin production by Cryptococcus species; minimal medium plus methyldopa was more efficient than the BAB, MHA, and BHIA in the melanin production; and NSA and SSA, followed by MMA added with methyldopa, were more efficient than other media studied for melanin production by all strains studied.

Different culture media containing methyldopa for melanin production by Cryptococcus species / Avaliação de diferentes meios de cultura contendo metildopa para a produção de melanina por espécies de Cryptococcus

INTRODUCTION: Melanin production by species of Cryptococcus is widely used to characterize C. neoformans complex in mycology laboratories. This study aims to test the efficacy of methyldopa from pharmaceutical tablet as a substrate for melanin production, to compare the production of melanin using different agar base added with methyldopa, and to compare the melanin produced in those media with that produced in Niger seed agar and sunflower seed agar by C. neoformans, C. laurentii, and C. albidus. Two isolates of each species, C. neoformans, C. laurentii, and C. albidus, and one of Candida albicans were used to experimentally detect conditions for melanin production. METHODS: The following media were tested: Mueller-Hinton agar (MHA), brain and heart infusion agar (BHIA), blood agar base (BAB), and minimal medium agar (MMA), all added with methyldopa, and the media Niger seed agar (NSA) and sunflower seed agar (SSA). RESULTS: All isolates grew in most of the culture media after 24h. Strains planted on media BAB and BHIA showed growth only after 48h. All isolates produced melanin in MMA, MHA, SSA, and NSA media. CONCLUSIONS: Methyldopa in the form pharmaceutical tablet can be used as a substrate for melanin production by Cryptococcus species; minimal medium plus methyldopa was more efficient than the BAB, MHA, and BHIA in the melanin production; and NSA and SSA, followed by MMA added with methyldopa, were more efficient than other media studied for melanin production by all strains studied.

INTRODUÇÃO: A produção de melanina por espécies de Cryptococcus é uma característica amplamente utilizada em laboratórios de micologia para caracterização do complexoC. neoformans. O objetivo deste estudo foi verificar a eficácia da metildopa na forma farmacêutica de comprimido, como substrato para a produção de melanina por Cryptococcus, comparar diferentes bases de meios de cultura acrescidas de metildopa para produção de melanina e comparar o pigmento produzido nestes meios com o produzido em ágar Níger e ágar girassol por C. neoformans, C. laurentii e C. albidus. MÉTODOS: Foram testados dois isolados de cada uma das espécies, C. neoformans, C.laurentii e C.albidus, e um de C. albicans para avaliar a produção de melanina nos meios de cultura ágar Müeller-Hinton (MH), ágar brain heart infusion (BHI), ágar base sangue (BS), meio mínimo (MM), todos acrescidos de metildopa, e ainda ágar girassol e ágar Níger. RESULTADOS: Todos os isolados cresceram na maioria dos meios após 24h. O crescimento nos meios BS e BHI somente ocorreu após 48h. Todos os isolados produziram melanina nos meios MM, MH, girassol e Niger. CONCLUSÕES: A metildopa de origem farmacêutica pode ser utilizada como substrato para a produção de melanina por espécies de Cryptococcus; o MM acrescido de metildopa mostrou-se mais eficiente na produção de melanina do que os meios BS, MH e BHI; ágar girassol e ágar Níger seguidos de MM acrescido de metildopa foram os mais eficientes na produção de melanina pelos isolados estudados.