RESUMO
The objective of this study was to describe phenotypically and biochemically the symbiotic bacteria associated with three populations of Heterorhabditis amazonensis Andalo et al., 2006 (isolates: UEL-n 01, UEL-n 07, and UEL-n 08) and evaluate their pathogenicity on Galleria mellonella L. (Lepidoptera: Pyralidae) larvae. Bacteria were isolated by maceration of infective juveniles (IJs) and grown in culture medium (NBTA and MacConkey). The characterization of the bacteria was evaluated by employing motility test and biochemical tests like Gram staining, lipase activity, protease, and lecithinase. The production of antibiotics and bioluminescence was also evaluated. The pathogenicity was evaluated on the last instar larvae of G. mellonella at a concentration of 104 cells/mL. The bacteria from the three entomopathogenic nematodes isolates were positive for all biochemical tests except for lecithinase, and have presented bioluminescence when subjected to ultraviolet light, indicating that they belong to the genus Photorhabdus sp. Both were pathogenic to G. mellonella larvae causing 93.3 to 100.0% mortality.(AU)
O objetivo desse trabalho foi descrever fenotípica e bioquimicamente as bactérias simbiontes associadas a três isolados do nematoide Heterorhabditis amazonensis Andalo et al., 2006 (isolates: UEL-n 01, UEL-n 07, and UEL-n 08) e avaliar sua patogenicidade sobre lagartas de Galleria mellonella L. (Lepidoptera: Pyralidae). As bactérias foram isoladas por meio de macerado de juvenis infectantes (JIs) e cultivadas em meio de cultura (NBTA e MacConkey). A caracterização das bactérias foi realizada por meio de testes de motilidade e bioquímicos de coloração de Gram, atividade de lipase, protease e lecitinase. Avaliou-se também a produção de antibióticos e de bioluminescência. A patogenicidade foi avaliada em lagartas de G. mellonella na concentração de 104 células/mL. Observou-se que as bactérias dos três isolados de nematoides entomopatogênicos foram positivas para todos os testes bioquímicos, com exceção da lecitinase, e apresentaram bioluminescência quando submetidas a luz ultravioleta, indicando que estas cepas pertencem ao gênero Photorhabdus sp. Ambas foram patogênicas às lagartas de G. mellonellacausando mortalidade entre 93,3 e 100,0%.(AU)
Assuntos
Rabditídios/isolamento & purificação , Lepidópteros , Fenótipo , Virulência , BioquímicaRESUMO
Although hemolytic activity is known to be a putative virulence factor contributing to candidal pathogenesis, its production by Candida tropicalis, a species closely related to Candida albicans, is poor understood. The present study was undertaken to evaluate the hemolytic activity and the expression level of a putative haem oxygenase encoding gene by blood isolates of C. tropicalis following growth in iron deprivation, and in the presence of hemoglobin and erythrocytes. The lowest values of hemolytic activity were observed in cell-free culture supernatants of isolates growing in iron-restricted medium (RPMI medium and RPMI medium supplemented with iron chelator bathophenanthrolindisulphonic acid). Hemolysis was increased in the presence of either hemoglobin or erythrocytes. Reverse transcriptase PCR analysis showed that the putative haem oxygenase encoding gene (CtHMX1), potentially related with iron uptake, was up-regulated (p < 0.001) following growth in iron deprivation and in the presence of hemoglobin; CtHMX1 was repressed in the presence of human erythrocytes (p < 0.001). Our data suggest that hemoglobin had positive effect in the production of hemolytic factor and gene expression related to iron uptake in C. tropicalis.
Assuntos
Sangue/microbiologia , Candida tropicalis/enzimologia , Candida tropicalis/genética , Eritrócitos/metabolismo , Heme Oxigenase (Desciclizante)/genética , Heme Oxigenase (Desciclizante)/metabolismo , Hemoglobinas/metabolismo , Ferro/metabolismo , Candida tropicalis/crescimento & desenvolvimento , Candida tropicalis/ultraestrutura , Candidíase/sangue , Candidíase/microbiologia , Meios de Cultura , DNA Fúngico/isolamento & purificação , Proteínas Fúngicas/biossíntese , Proteínas Fúngicas/genética , Fungos/crescimento & desenvolvimento , Proteínas Hemolisinas , Hemólise , Humanos , RNA Fúngico/isolamento & purificação , Regulação para Cima , Fatores de Virulência/metabolismoRESUMO
BACKGROUND: Candida tropicalis is an increasingly important human pathogen associated with high mortality rates; however, little is known regarding the virulence properties of C. tropicalis, particularly the production of haemolytic factor. Although Candida spp may acquire iron from human blood red cells (RBCs) by producing a haemolytic factor that promotes cell lyses, at present there are no data regarding the effect of RBCs on the production of haemolytic molecules. The present study was undertaken to evaluate the role of human red blood cells on the production haemolytic factor by C. tropicalis; in addition, the transcription levels of a putative haemolysin-like protein gene (HLPt) were also analysed. RESULTS: C. tropicalis isolates produced a haemolytic factor following growth in either the absence or presence of RBCs; however, distinct levels of haemolysis were observed, with 60% of the isolates exhibiting a significant increase in the production of haemolytic factor when grown in the presence of human RBCs. All isolates in which the putative HLPt gene was up-regulated in presence of human RBCs, ranging from 1.044 to 6.965-fold, also exhibited higher haemolytic activity following growth in the presence of RBCs compared to that observed in the absence of RBCs. CONCLUSIONS: We propose that human RBCs may induce changes in the phenotypic expression of haemolytic factor and in transcriptional levels of the putative C. tropicalis HLPt gene in an isolate-dependent fashion.
Assuntos
Candida tropicalis/fisiologia , Candidíase/microbiologia , Eritrócitos/metabolismo , Regulação da Expressão Gênica , Proteínas Hemolisinas/genética , Hemólise , Proteínas Hemolisinas/metabolismo , Humanos , RNA Mensageiro/genética , RNA Mensageiro/metabolismoRESUMO
INTRODUCTION: In this study, we aimed at identifying Candida isolates obtained from blood, urine, tracheal secretion, and nail/skin lesions from cases attended at the Hospital Universitário de Londrina over a 3-year period and at evaluating fluconazole susceptibilities of the isolates. METHODS: Candida isolates were identified by polymerase chain reaction (PCR) using species-specific forward primers. The in vitro fluconazole susceptibility test was performed according to EUCAST-AFST reference procedure. RESULTS: Isolates were obtained from urine (53.4%), blood cultures (19.2%), tracheal secretion (17.8%), and nail/skin lesions (9.6%). When urine samples were considered, prevalence was similar in women (45.5%) and in men (54.5%) and was high in the age group >61 years than that in younger ones. For blood samples, prevalence was high in neonates (35%) and advanced ages (22.5%). For nail and skin samples, prevalence was higher in women (71.4%) than in men (28.6%). Candida albicans was the most frequently isolated in the hospital, but Candida species other than C. albicans accounted for 64% of isolates, including predominantly Candida tropicalis (33.2%) and Candida parapsilosis (19.2%). The trend for non-albicans Candida as the predominant species was noted from all clinical specimens, except from urine samples. All Candida isolates were considered susceptible in vitro to fluconazole with the exception of isolates belonging to the intrinsically less-susceptible species C. glabrata. CONCLUSIONS: Non-albicans Candida species were more frequently isolated in the hospital. Fluconazole resistance was a rare finding in our study.
Assuntos
Antifúngicos/farmacologia , Candida/efeitos dos fármacos , Fluconazol/farmacologia , Adulto , Candida/classificação , Candida/isolamento & purificação , Candidíase/microbiologia , Infecção Hospitalar/microbiologia , Farmacorresistência Fúngica , Feminino , Humanos , Masculino , Testes de Sensibilidade Microbiana , Pessoa de Meia-Idade , Reação em Cadeia da Polimerase , Especificidade da EspécieRESUMO
INTRODUCTION: In this study, we aimed at identifying Candida isolates obtained from blood, urine, tracheal secretion, and nail/skin lesions from cases attended at the Hospital Universitário de Londrina over a 3-year period and at evaluating fluconazole susceptibilities of the isolates. METHODS: Candida isolates were identified by polymerase chain reaction (PCR) using species-specific forward primers. The in vitro fluconazole susceptibility test was performed according to EUCAST-AFST reference procedure. RESULTS: Isolates were obtained from urine (53.4 percent), blood cultures (19.2 percent), tracheal secretion (17.8 percent), and nail/skin lesions (9.6 percent). When urine samples were considered, prevalence was similar in women (45.5 percent) and in men (54.5 percent) and was high in the age group >61 years than that in younger ones. For blood samples, prevalence was high in neonates (35 percent) and advanced ages (22.5 percent). For nail and skin samples, prevalence was higher in women (71.4 percent) than in men (28.6 percent). Candida albicans was the most frequently isolated in the hospital, but Candida species other than C. albicans accounted for 64 percent of isolates, including predominantly Candida tropicalis (33.2 percent) and Candida parapsilosis (19.2 percent). The trend for non-albicans Candida as the predominant species was noted from all clinical specimens, except from urine samples. All Candida isolates were considered susceptible in vitro to fluconazole with the exception of isolates belonging to the intrinsically less-susceptible species C. glabrata. CONCLUSIONS: Non-albicans Candida species were more frequently isolated in the hospital. Fluconazole resistance was a rare finding in our study.
INTRODUÇÃO: Neste estudo objetivamos a identificação de isolados de Candida obtidos de sangue, urina, secreção traqueal e de lesões de unha/pele, de casos atendidos no Hospital Universitário de Londrina num período de três anos. Avaliamos também a suscetibilidade dos isolados ao fluconazol. MÉTODOS: Os isolados de Candida foram identificados pela reação em cadeia da polimerase (RCP) usando oligonucleotídeos iniciadores espécie-específicos. O teste de suscetibilidade in vitro ao fluconazol foi realizado segundo o procedimento de referência EUCAST-AFST. RESULTADOS: Isolados foram obtidos de urina (53,4 por cento), sangue (19,2 por cento), secreção traqueal (17,8 por cento) e lesões de unha/pele (9,6 por cento). Considerando as amostras de urina, a prevalência foi similar em mulheres (45,5 por cento) e em homens (54,5 por cento) e foi alta no grupo de idade > 61 anos do que em grupos mais jovens. Para amostras de sangue a prevalência foi alta em neonatos (35 por cento) e idades avançadas (22,5 por cento). Para amostras de unha e pele a prevalência foi maior em mulheres (71,4 por cento) do que em homens (28,6 por cento). Candida albicans foi a mais frequentemente isolada no hospital, mas outras espécies de Candida corresponderam a 64 por cento dos isolados, incluindo predominantemente Candida tropicalis (33,2 por cento) e Candida parapsilosis (19,2 por cento). A tendência de Candida não-albicans como espécie predominante foi observada para todas as amostras clínicas, exceto para amostras de urina. Todos isolados de Candida foram considerados suscetíveis, in vitro, ao fluconazol com exceção dos isolados pertencentes às espécies intrinsecamente menos suscetíveis C. glabrata. CONCLUSÕES: Espécies de Candida não-albicans foram mais frequentemente isoladas no hospital. Resistência ao fluconazol foi rara no nosso estudo.
Assuntos
Adulto , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Antifúngicos/farmacologia , Candida/efeitos dos fármacos , Fluconazol/farmacologia , Candida/classificação , Candida/isolamento & purificação , Candidíase/microbiologia , Infecção Hospitalar/microbiologia , Farmacorresistência Fúngica , Testes de Sensibilidade Microbiana , Reação em Cadeia da Polimerase , Especificidade da EspécieRESUMO
INTRODUCTION: Yeasts belonging to the genus Candida are responsible for the majority of fungal infections in humans. Candida tropicalis has been one of most commonly isolated non-albicans species. To analyze in vitro hemolysis promoted by clinical isolates of C. tropicalis obtained from blood and other clinical samples from hospitalized patients at the University Hospital of Londrina State University, Paraná, Brazil. METHODS: The hemolysis promoted by 28 clinical isolates of C. tropicalis was evaluated, and the isolates were grouped into classes according to the hemolysis levels. RESULTS: The majority of the blood isolates showed weak hemolysis (+), while the classes of strong hemolysis (+++) and very strong hemolysis (++++) predominated among isolates from other clinical samples such as urine, nail lesions and tracheal secretions. However, no statistical differences were detected (p> 0.05). CONCLUSIONS: Isolates of C. tropicalis obtained from different clinical samples showed a capacity to promote in vitro hemolysis.
Assuntos
Candida tropicalis/fisiologia , Hemólise , HumanosRESUMO
INTRODUÇÃO: Leveduras do gênero Candida são responsáveis pela maioria das infecções fúngicas em humanos. Candida tropicalis tem sido uma das mais comumente isoladas dentre as espécies não-albicans. O objetivo foi analisar a hemólise in vitro promovida por isolados clínicos de C. tropicalis provenientes de sangue e outras amostras clínicas de pacientes internados no Hospital Universitário da UEL, PR-Brasil. MÉTODOS: Foi avaliada a hemólise promovida por 28 isolados clínicos de C. tropicalis, sendo os isolados agrupados em classes de acordo com os níveis de hemólise. RESULTADOS: A maioria dos isolados de sangue apresentou hemólise fraca (+), enquanto as classes de hemólise forte (+++) e muito forte (++++) foram as predominantes nos isolados de outras amostras clínicas como urina, lesão de unha e secreção traqueal, embora não tenham sido detectadas diferenças estatísticas (p>0,05). CONCLUSÕES: Isolados de C. tropicalis, obtidos de diferentes amostras clínicas, apresentam capacidade de promover hemólise in vitro.
INTRODUCTION: Yeasts belonging to the genus Candida are responsible for the majority of fungal infections in humans. Candida tropicalis has been one of most commonly isolated non-albicans species. To analyze in vitro hemolysis promoted by clinical isolates of C. tropicalis obtained from blood and other clinical samples from hospitalized patients at the University Hospital of Londrina State University, Paraná, Brazil. METHODS: The hemolysis promoted by 28 clinical isolates of C. tropicalis was evaluated, and the isolates were grouped into classes according to the hemolysis levels. RESULTS: The majority of the blood isolates showed weak hemolysis (+), while the classes of strong hemolysis (+++) and very strong hemolysis (++++) predominated among isolates from other clinical samples such as urine, nail lesions and tracheal secretions. However, no statistical differences were detected (p> 0.05). CONCLUSIONS: Isolates of C. tropicalis obtained from different clinical samples showed a capacity to promote in vitro hemolysis.