Enzyme production by Candida albicans and Candida dubliniensis in periodontal HIV-positive patients receiving and not receiving antiretroviral therapy / Producción enzimática de Candida albicans y Candida dubliniensis de pacientes con periodontitis VIH+ con y sin tratamiento antirretroviral

ABSTRACT Candida dubliniensis (Cd) and Candida albicans (Ca) are the most frequently isolated yeasts in HIV+ patients. Some of the enzymes produced by these yeasts are considered virulence factors since they contribute to pathogenicity of Candida spp. The aim of the present study was to compare production of enzymes such as phospholipase (Ph), proteinase (P), and hemolysin (H) by Cd and Ca strains isolated from periodontal HIV-positive patients receiving and not receiving highly active antiretroviral therapy (HAART). Subgingival biofilm samples were obtained using paper points, and a sample of oral mucosa was taken using a swab. Phenotypic and molecular methods were used to isolate 39 strains of Candida, including 25 strains of Cd and 14 strains of Ca, obtained from 33 periodontal pocket samples and 6 oral mucosa samples collected from 15 HIV+ patients (8 receiving and 7 not receiving HAART). Malt egg-yolk agar, albumin agar and blood agar were used to evaluate pH, P and H production respectively. The strains were inoculated in duplicate and incubated at 37 ºC. Colony and halo diameters were measured. A greater proportion of Ca was observed in patients not receiving HAART, and a higher proportion of Cd was observed in those under HAART, Chi2 p< 0.001. Phospholipase production was observed in 92.9% percent of isolated Ca strains but in none of the isolated Cd strains. Proteinase production was high in Ca and Cd strains isolated from patients not receiving HAART. Hemolysin production was observed in all the studied strains, though it was significantly higher (p=0.04) in Ca and Cd strains isolated from patients not receiving HAART. To sum up, the proportion of Candida dubliniensis strains was highest in the subgingival biofilm of patients receiving HAART, and Cd strains were found to express fewer virulence factors than Ca strains.

RESUMEN Las levaduras más aisladas en pacientes VIH+ son Candida dubliniensis (Cd) y Candida albicans (Ca). Algunas de sus enzimas constituyen factores de virulencia ya que favorecen la diseminación tisular. El objetivo fue comparar la producción de enzimas como fosfolipasa (F), proteinasa (P) y hemolisina (H) en cepas de Cd y Ca aisladas de pacientes VIH+ tratados y no tratados con antirretrovirales (TARGA). Se realizó la toma del biofilm de placa subgingival con conos de papel y la muestra de la mucosa bucal con hisopo. Se aislaron y tipificaron por métodos fenotípicos y moleculares 39 cepas: 25 de Cd y 14 Ca, obtenidas 33 de bolsas periodontales y 6 de mucosa bucal de 15 pacientes VIH+ (8 con y 7 sin tratamiento). Se utilizó agar malta con yema de huevo, agar albúmina y agar sangre para demostrar la producción de F, P y H, respectivamente. Se inocularon por duplicado e incubaron a 37°C. Se midieron los diámetros de las colonias y los de hidrólisis alrededor de las mismas. Se observó mayor proporción de Ca en los pacientes sin tratamiento y mayor proporción de Cd en los con tratamiento; Chi2 p< 0.001. El 92,9% de las Ca estudiadas, fueron productoras de fosfolipasa. En tanto que ninguna Cd produjo la enzima. En cuanto a la producción de proteinasa se observa una alta producción tanto en las cepas de Ca, como en las Cd aisladas en los pacientes no tratados. Todas las cepas estudiadas produjeron hemolisina, observándose una diferencia estadísticamente significativa (p=0,04) en ambas especies a favor de la alta producción de la enzima en las cepas obtenidas de pacientes no tratados. Podemos concluir que en el biofilm subgingival, en los pacientes bajo TARGA, se aíslan mayor proporción de Candida dubliniensis las cuales expresan menos factores de virulencia.

In vitro evaluation of hydrolytic enzyme activity and biofilm formation of Candida parapsilosis species complex from a nosocomial environment

Abstract INTRODUCTION: Candida parapsilosis complex species, frequently found in hospital environments, have gained importance as etiological agents of candidemia. METHODS: Candida parapsilosis complex isolates from a nosocomial environment were identified and their hydrolitic enzyme activity and ability to form biofilm were characterized. RESULTS: Twenty-two C. parapsilosis sensu stricto isolates produced proteinase and three produced phospholipase. Most Candida metapsilosis isolates produced proteinase and one also produced phospholipase. All 29 isolates formed biofilms. CONCLUSIONS: The nosocomial environment may act as a reservoir for C. parapsilosis complex isolates with phenotypic features that could possibly lead to nosocomial infections and health complications in hospital patients.

Production and characterization of an extracellular lipase from Candida guilliermondii

Extracellular lipases from the endophytic yeast Candida guilliermondii isolated from castor leaves (Ricinus communis L.) were produced using low-cost raw materials such as agro-industrial residues and applying them in the esterification of oleic acid for evaluating their potential use in biodiesel production. After partial purification using ammonium sulfate, the enzyme was characterized and presented higher activity (26.8 ± 1.5 U mL-1) in the presence of 5 mmol L-1 NaCl at 30 ºC and pH 6.5. The production through submerged fermentation was formerly performed in 150 mL erlenmeyer flasks and, once the enzyme production was verified, assays in a 14 L bioreactor were conducted, obtaining 18 ± 1.4 U mL-1. The produced enzyme was applied in the oleic acid esterification under different solvents: hexane, cyclohexane or cyclohexanone) and different acid:alcohol molar ratios. Higher ester conversion rate (81%) was obtained using hexane and the molar ratio of 1:9 was the best conditions using methanol. The results suggest the potential for development of endophytic yeast in the production of biocatalyst through submerged fermentation using agroindustrial residues as culture medium.

Fluconazole and amphotericin-B resistance are associated with increased catalase and superoxide dismutase activity in Candida albicans and Candida dubliniensis

Introduction Candida dubliniensis, a new species of Candida that has been recovered from several sites in healthy people, has been associated with recurrent episodes of oral candidiasis in AIDS and HIV-positive patients. This species is closely related to C. albicans. The enzymatic activity of C. dubliniensis in response to oxidative stress is of interest for the development of drugs to combat C. dubliniensis. Methods Fluconazole- and amphotericin B-resistant strains were generated as described by Fekete-Forgács et al. (2000). Superoxide dismutase (SOD) and catalase assays were performed as described by McCord and Fridovich (1969) and Aebi (1984), respectively. Results We demonstrated that superoxide dismutase (SOD) and catalase activities were significantly higher (p<0.05) in the fluconazole- and amphotericin B-resistant strains of C. dubliniensis and C. albicans than in the sensitive strains. The catalase and SOD activities were also significantly (p<0.01) higher in the sensitive and resistant C. albicans strains than in the respective C. dubliniensis strains. Conclusions These data suggest that C. albicans is better protected from oxidative stress than C. dubliniensis and that fluconazole, like amphotericin B, can induce oxidative stress in Candida; oxidative stress induces an adaptive response that results in a coordinated increase in catalase and SOD activities. .

Oral colonization by yeasts in HIV-positive patients in Brazil / Colonização oral por leveduras em pacientes HIV-positivos no Brasil

INTRODUCTION: In HIV-infected patients, colonization of the oral cavity by potential pathogenic yeast may lead to development of systemic fungemia. We evaluated the prevalence of yeast in the oral cavity of Brazilian HIV-positive patients and verified whether or not the species characterized were enzymatically active. Furthermore, the species identified were tested for their susceptibility to antifungal treatment. METHODS: Patient saliva and oropharyngeal candidiasis samples were collected from 60 seropositive HIV patients and identified by the API20C system. Enzymatic activity was evaluated by the production of proteinase and phospholipase. Susceptibility to antifungal treatments were determined using the broth microdilution method. RESULTS: the most commonly isolated species were C. albicans (51.56 percent) followed by non-albicans Candida species (43.73 percent), Trichosporon mucoides (3.12 percent) and Kodamaea ohmeri (1.56 percent). Oral colonization by association of different species was observed in 42 percent of the patients. Enzymatic activity was verified in most of species isolated, except for C. glabrata, C. lusitaniae and C. guilliermondii. Resistance to Fluconazole and Amphotericin B was observed in isolates of C. albicans, C. glabrata, C. parapsilosis, C. krusei, and K. ohmeri. CONCLUSION: HIV-positive patients are orally colonized by single or multiple species of yeast that are occasionally resistant to Fluconazole or Amphotericin B.

INTRODUÇÃO: Em pacientes infectados pelo HIV, a colonização da cavidade bucal por leveduras patogênicas pode levar ao desenvolvimento de fungemias. No presente estudo, avaliamos a prevalência de leveduras na cavidade bucal de pacientes HIV-positivos e verificamos se as espécies isoladas foram enzimaticamente ativas. Além disso, as espécies identificadas foram testadas quanto à suscetibilidade a antifúngicos. MÉTODOS: Amostras de saliva e de candidose orofaríngea foram coletadas de 60 pacientes soropositivos para HIV e identificados pelo sistema API20C. A atividade enzimática foi avaliada pela produção de proteinase e fosfolipase. A suscetibilidade a antifúngicos foi determinada utilizando o método de microdiluição em caldo. RESULTADOS: As espécies mais comumente isoladas foram C. albicans (51,56 por cento), seguido por espécies de Candida não-albicans (43,73 por cento), Trichosporon mucoides (3,12 por cento) e Kodamaea ohmeri (1,56 por cento). A colonização bucal por associação de diferentes espécies foi observada em 42 por cento dos pacientes. A atividade enzimática foi verificada na maioria das espécies isoladas, com exceção de C. glabrata, C. lusitaniae e C. guilliermondii. Resistência ao fluconazol e anfotericina B foi observada em isolados de C. albicans, C. glabrata, C. parapsilosis, C. krusei, e K. ohmeri. CONCLUSÃO: Os pacientes HIV-positivos são colonizados por espécies únicas ou múltiplas de levedura que ocasionalmente são resistentes ao fluconazol ou anfotericina B.

In vitro differential activity of phospholipases and acid proteinases of clinical isolates of Candida / Atividade diferencial in vitro de fosfolipases e proteinases ácidas de isolados clínicos de Candida

INTRODUCTION: Candida yeasts are commensals; however, if the balance of normal flora is disrupted or the immune defenses are compromised, Candida species can cause disease manifestations. Several attributes contribute to the virulence and pathogenicity of Candida, including the production of extracellular hydrolytic enzymes, particularly phospholipase and proteinase. This study aimed to investigate the in vitro activity of phospholipases and acid proteinases in clinical isolates of Candida spp. METHODS: Eighty-two isolates from hospitalized patients collected from various sites of origin were analyzed. Phospholipase production was performed in egg yolk medium and the production of proteinase was verified in a medium containing bovine serum albumin. The study was performed in triplicate. RESULTS: Fifty-six (68.3 percent) of isolates tested were phospholipase positive and 16 (44.4 percent) were positive for proteinase activity. C. tropicalis was the species with the highest number of positive isolates for phospholipase (91.7 percent). Statistically significant differences were observed in relation to production of phospholipases among species (p<0,0001) and among the strains from different sites of origin (p=0.014). Regarding the production of acid protease, the isolates of C. parapsilosis tested presented a larger number of producers (69.2 percent). Among the species analyzed, the percentage of protease producing isolates did not differ statistically (χ2=1.9 p=0.5901 (χ2=1.9 p=0.5901). CONCLUSIONS: The majority of C. non-albicans and all C. albicans isolates were great producers of hydrolytic enzymes and, consequently, might be able to cause infection under favorable conditions.

INTRODUÇÃO: Candida são leveduras comensais, porém, se o equilíbrio da flora normal for interrompido ou as defesas imunitárias estiverem comprometidas, espécies de Candida podem causar manifestações de doença. Vários atributos contribuem na virulência e patogenicidade de Candida, inclusive a produção de enzimas extracelulares hidrolíticas, especialmente fosfolipases e proteinases. O objetivo deste estudo foi verificar a atividade in vitro de fosfolipases e proteinases ácidas em isolados clínicos de Candida spp. MÉTODOS: Oitenta e dois isolados provenientes de pacientes hospitalizados coletados a partir de sítios de origem diversos foram analisados. A produção de fosfolipase foi verificada em meio egg yolk e a de proteinase em meio contendo soro albumina bovina. O estudo foi feito em triplicata. RESULTADOS: Cinquenta e seis (68,3 por cento) dos isolados testados apresentaram atividade de fosfolipase positiva e 16 (44,4 por cento) foram positivos para atividade de proteinase. C. tropicalis foi a espécie que apresentou o maior número de isolados positivos para fosfolipases (91,7 por cento). Diferenças estatisticamente significantes em relação à produção de fosfolipases entre as espécies e entre as cepas provenientes de diferentes sítios de origem foram detectadas. Quanto à produção de proteinases ácidas, os isolados de C. parapsilosis testados foram os maiores produtores (69,2 por cento). Entre as espécies analisadas, a porcentagem de produção de proteinase entre os isolados não diferiu estatisticamente (χ2=1.9 p=0.5901 (χ2=1.9 p=0.5901). CONCLUSÕES: A maioria dos isolados de C. não-albicans, assim como os de C. albicans, foram grandes produtores de enzimas hidrolíticas e, consequentemente, podem ser capazes de causar infecção em condições adequadas.

Increased expression of keratinase and other peptidases by Candida parapsilosis mutants

Keratinases are enzymes of great importance involved in pathogenic processes of some fungi. They also have a widespread ecological role since they are responsible for the degradation and recycling of keratin. On the one hand, studying them furthers our knowledge of pathogenicity mechanisms, which has important implications for human health, and on the other hand, understanding their ecological role in keratin recycling has biotechnological potential. Here, a wild-type keratinolytic Candida parapsilosis strain isolated from a poultry farm was treated with ethyl methanesulfonate in order to generate mutants with increased keratinase activity. Mutants were then cultured on media with keratin extracted from chicken feathers as the sole source of nitrogen and carbon. Approximately 500 mutants were screened and compared with the described keratinolytic wild type. Three strains, H36, I7 and J5, showed enhanced keratinase activity. The wild-type strain produced 80 U/mL of keratinolytic activity, strain H36 produced 110 U/mL, strain I7, 130 U/mL, and strain J5, 140 U/mL. A 70 percent increase in enzyme activity was recorded for strain J5. Enzymatic activity was evaluated by zymograms with proteic substrates. A peptidase migrating at 100 kDa was detected with keratin, bovine serum albumin and casein. In addition, a peptidase with a molecular mass of 50 kDa was observed with casein in the wild-type strain and in mutants H36 and J5. Gelatinase activity was detected at 60 kDa. A single band of 35 kDa was found in wild-type C. parapsilosis and in mutants with hemoglobin substrate.

Use of Candida rugosa lipase immobilized on sepabeads for the amyl caprylate synthesis: batch and fluidized bed reactor study

Lipase from Candida rugosa was covalently immobilized on Sepabeads EC-EP for application for amyl caprylate synthesis in an organic solvent system. Several solvents were tested in terms of biocatalyst stability and the best result was obtained with isooctane. The lipase-catalyzed esterification in the selected system was performed in batch and fluidized bed reactor systems. The influence of several important reaction parameters including temperature, initial water content, enzyme loading, acid/alcohol molar ratio, and time of addition of molecular sieves is carefully analyzed by means of an experimental design. Almost complete conversion (> 99 percent) of the substrate to ester could be performed in a batch reactor system, using lipase loading as low as 37 mg g-1 dry support and in a relatively short time (24 hrs) at 37°C, when high initial substrate molar ratio of 2.2 is used. Kinetics in a fluidized bed reactor system seems to still have a slightly better profile than in the batch system (90.2 percent yields after 14 hrs). The fluidized bed reactor operated for up 70 hrs almost with no loss in productivity, implying that the proposed process and the immobilized system could provide a promising approach for the amyl caprylate synthesis at the industrial scale.

Differences in exoenzyme production and adherence ability of Candida spp. isolates from catheter, blood and oral cavity / Diferenças na produção de exoenzimas e habilidade de aderência entre isolados de espécies de Candida provenientes do cateter, sangue e cavidade bucal

Phospholipase and proteinase production and the ability of adhesion to buccal epithelial cells (BEC) of 112 Candida isolates originated from oral cavity of HIV infected patients and from blood and catheter of intensive care unit patients were investigated. The proteinase production was detected by inoculation into bovine serum albumin (BSA) agar and the phospholipase activity was performed using egg yolk emulsion. A yeast suspension of each test strain was incubated with buccal epithelial cells and the number of adherence yeast to epithelial cells was counted. A percentage of 88.1 percent and 55.9 percent of Candida albicans and 69.8 percent and 37.7 percent of non-albicans Candida isolates produced proteinase and phospholipase, respectively. Non-albicans Candida isolated from catheter were more proteolytic than C. albicans isolates. Blood isolates were more proteolytic than catheter and oral cavity isolates while oral cavity isolates produced more phospholipase than those from blood and catheter. C. albicans isolates from oral cavity and from catheter were more adherent to BEC than non-albicans Candida isolates, but the adhesion was not different among the three sources analyzed. The results indicated differences in the production of phospholipase and proteinase and in the ability of adhesion to BEC among Candida spp. isolates from different sources. This study suggests that the pathogenicity of Candida can be correlated with the infected site.

A produção de proteinase e fosfolipase e habilidade de adesão à célula epitelial bucal de 112 isolados de Candida originadas da cavidade bucal de pacientes infectados pelo HIV e de sangue e cateter de pacientes hospitalizados foram investigados. A produção de proteinase foi detectada por inoculação em ágar soro albumina bovina e a atividade de fosfolipase foi realizada usando emulsão de gema de ovo. A suspensão de levedura de cada isolado foi incubada com célula epitelial e o número de leveduras aderidas a célula epitelial foi contada. Uma porcentagem de 88,1 e 55,9 por cento de C. albicans e 69,8 e 37,7 por cento de isolados de Candida não albicans produziram proteinase e fosfolipase, respectivamente. Candida não albicans obtidas do cateter foram mais proteolíticos que isolados de Candida albicans (p < 0,001). Isolados do sangue foram mais proteolíticos do que isolados do cateter e cavidade bucal, enquanto isolados da cavidade bucal produziram mais fosfolipase do que aqueles isolados do sangue e cateter. C. albicans isoladas da cavidade bucal e do cateter foram mais aderentes à célula epitelial bucal do que isolados de Candida não albicans, mas não houve diferença na adesão entre os três locais analisados. Os resultados indicaram diferenças na produção de fosfolipase e proteinase e na habilidade de adesão à célula epitelial bucal entre os isolados de Candida das diferentes fontes. Este estudo sugere que a patogenicidade de Candida spp pode estar correlacionada ao local infectado.

Artificial neural network modeling studies to predict the yield of enzymatic synthesis of betulinic acid ester

3 beta-O-phthalic ester of betulinic acid was synthesized from reaction of betulinic acid and phthalic anhydride using lipase as biocatalyst. This ester has clinical potential as an anticancer agent. In this study, artificial neural network (ANN) analysis of Candida antarctica lipase (Novozym 435) -catalyzed esterification of betulinic acid with phthalic anhydride was carried out. A multilayer feed-forward neural network trained with an error back-propagation algorithm was incorporated for developing a predictive model. The input parameters of the model are reaction time, reaction temperature, enzyme amount and substrate molar ratio while the percentage isolated yield of ester is the output. Four different training algorithms, belonging to two classes, namely gradient descent and Levenberg-Marquardt (LM), were used to train ANN. The paper makes a robust comparison of the performances of the above four algorithms employing standard statistical indices. The results showed that the quick propagation algorithm (QP) with 4-9-1 arrangement gave the best performances. The root mean squared error (RMSE), coefficient of determination (R²) and absolute average deviation (AAD) between the actual and predicted yields were determined as 0.0335, 0.9999 and 0.0647 for training set, 0.6279, 0.9961 and 1.4478 for testing set and 0.6626, 0.9488 and 1.0205 for validation set using quick propagation algorithm (QP).

Candida dubliniensis does not show phospholipase activity: true or false? / Candida dubliniensis não evidencia atividade fosfolipásica: verdadeiro ou falso?

INTRODUCTION: The phospholipase activity in Candida albicans and Candida dubliniensis isolated from oral candidiasis cases were studied. METHODS: The phospholipase activity was evaluated in egg yolk agar. RESULTS: All the C. albicans isolates (n = 48) showed phospholipase activity (mean Pz = 0.66). However, none of the C. dubliniensis isolates (n = 24) showed this activity. CONCLUSIONS: The authors discuss whether these findings are a true characteristic of C. dubliniensis or a consequence of the methodology employed, which includes the possibility that NaCl may have inhibited the enzymatic activity of C. dubliniensis.

INTRODUÇÃO: Avaliou-se a atividade fosfolipásica em Candida albicans e Candida dubliniensis isoladas de casos de candidíase oral. MÉTODOS: A atividade de fosfolipase foi avaliada em ágar gema de ovo. RESULTADOS: Todos os isolados de C. albicans (nº = 48) evidenciaram atividade fosfolipásica (média Pz = 0.66). Todavia, nenhum isolado de C. dubliniensis (nº= 24) demonstrou esta atividade. CONCLUSÕES: Os autores discutem se estes achados são uma característica verdadeira de C. dubliniensis ou uma conseqüência da metodologia empregada, a qual inclui a possibilidade de que o NaCl seja inibidor da atividade enzimática de C. dubliniensis.

Phylogeny and evolution of the aspartyl protease family from clinically relevant Candida species

Aspartyl proteases are a class of enzymes that include the yeast aspartyl proteases and secreted aspartyl protease (Sap) superfamilies. Several Sap superfamily members have been demonstrated or suggested as virulence factors in opportunistic pathogens of the genus Candida. Candida albicans, Candida tropicalis, Candida dubliniensis and Candida parapsilosis harbour 10, four, eight and three SAP genes, respectively. In this work, genome mining and phylogenetic analyses revealed the presence of new members of the Sap superfamily in C. tropicalis (8), Candida guilliermondii (8), C. parapsilosis(11) and Candida lusitaniae (3). A total of 12 Sap families, containing proteins with at least 50 percent similarity, were discovered in opportunistic, pathogenic Candida spp. In several Sap families, at least two subfamilies or orthologous groups were identified, each defined by > 90 percent sequence similitude, functional similarity and synteny among its members. No new members of previously described Sap families were found in a Candida spp. clinical strain collection; however, the universality of SAPT gene distribution among C. tropicalis strains was demonstrated. In addition, several features of opportunistic pathogenic Candida species, such as gene duplications and inversions, similitude, synteny, putative transcription factor binding sites and genome traits of SAP gene superfamily were described in a molecular evolutionary context.

Infecções oportunistas por leveduras e perfil enzimático dos agentes etiológicos / Opportunistic yeast infections and enzymatic profile of the etiological agents

Infecções por leveduras são freqüentes em imunocomprometidos, contudo espécies emergentes têm alterado o perfil epidemiológico. A habilidade de secretar proteases tem sido associada à patogenicidade do gênero Candida. Esta pesquisa teve como objetivos diagnosticar leveduroses em pacientes imunocomprometidos e avaliar a virulência dos agentes etiológicos baseado em teste de secreção de protease utilizando soro de albumina bovina como substrato. Do total de 104 pacientes estudados, 19 apresentaram episódios de leveduroses. O trato respiratório (63,2 por cento), seguido pelo trato urinário (10,5 por cento) foram os locais mais comuns de infecção. Candida albicans, Candida parapsilosis, Candida tropicalis e espécies emergentes como Candida krusei e Candida guilliermondii foram isoladas. Cinco isolados de Candida parapsilosis e um de Candida albicans e Candida guilliermondii exibiram alta atividade enzimática. Concluímos que a caracterização enzimática de isolados de Candida pode ser um útil marcador prognóstico, especialmente em imunocomprometidos, uma vez que leveduroses nestes pacientes são geralmente graves.

Yeast infections are common in immunocompromised patients, although emerging species have been changing the epidemiological profile. The ability to secrete proteinases has been associated with pathogenicity within the genus Candida. This study had the aims of diagnosing yeast infections in immunocompromised patients and evaluating the virulence of the etiological agents, based on a proteinase secretion assay using bovine serum albumin as a substrate. Out of a total of 104 patients studied, 19 presented episodes of yeast infection. The respiratory tract (63.2 percent), followed by the urinary tract (10.5 percent), were the most common sites of infection. Candida albicans, Candida parapsilosis, Candida tropicalis and emerging species such as Candida krusei and Candida guilliermondii were isolated. Five isolates of Candida parapsilosis and one of Candida albicans and Candida guilliermondii exhibited high enzymatic activity. We conclude that enzymatic characterization of Candida isolates may be a useful prognostic marker, especially among immunocompromised individuals, since yeast infections in such patients are generally serious.

Produção de fatores de virulência in vitro por espécies patogênicas do gênero Candida / Production of virulence factors in vitro by pathogenic species of the genus Candida

Avaliou-se, in vitro, a capacidade de crescimento em 39ºC e 42ºC, a produção de enzimas hidrolíticas e a atividade hemolítica de 21 cepas clínicas e de referência de sete espécies de Candida spp, Candida dubliniensis e Candida krusei demonstraram menor potencial de virulência e Candida albicans maior.

The growth capacity at 39ºC and 42ºC, production of hydrolytic enzymes and hemolytic activity of 21 clinical and reference strains of seven species of Candida spp were evaluated in vitro.Candida dubliniensis and Candida krusei demonstrated lower virulence potential and Candida albicans higher potential.

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.

Comparative esterification of phenylpropanoids versus hydrophenylpropanoids acids catalyzed by lipase in organic solvent media

The esterification of phenylpropanoid and hydrophenylpropanoid acids, catalyzed by candida antarctica lipase B (CAL-B), with several alcohols has demonstrated that the substitution pattern on the aromatic ring has a very significant influence on the reactivity of the carboxyl group due, mainly, to electronic effects, when compared to the unsaturated acids with the hydrogenated acids. It is also clear that in the saturated acids there still remain some unclear effects related to the aromatic substituents.

Enzymatic and hemolytic activities of Candida dubliniensis strains

Candida dubliniensis is an opportunistic yeast that has been recovered from several body sites in many populations; it is most often recovered from the oral cavities of human immunodeficiency virus-infected patients. Although extensive studies on epidemiology and phylogeny of C. dubliniensis have been performed, little is known about virulence factors such as exoenzymatic and hemolytic activities. In this study we compared proteinase, hyaluronidase, chondroitin sulphatase and hemolytic activities in 18 C. dubliniensis and 30 C. albicans strains isolated from AIDS patients. C. albicans isolates produced higher amounts of proteinase than C. dubliniensis (p < 0.05). All the tested C. dubliniensis strains expressed hyaluronidase and chondroitin sulphatase activities, but none of them were significantly different from those observed with C. albicans (p > 0.05). Hemolytic activity was affected by CaCl2; when this component was absent, we did not notice any significant difference between C. albicans and C. dubliniensis hemolytic activities. On the contrary, when we added 2.5 g percent CaCl2, the hemolytic activity was reduced on C. dubliniensis and stimulated on C. albicans tested strains (p < 0.05).

C. dubliniensis é uma levedura oportunista que, embora já tenha sido isolada de vários sítios anatômicos é, com maior frequência, encontrada na boca de pacientes infectados pelo HIV. Embora tenham sido realizados numerosos estudos sobre a epidemiologia e filogenia, seus fatores de virulência como atividade exoenzimática e atividade hemolítica, são, ainda, pouco conhecidos. Neste estudo comparou-se a atividade in vitro de proteinase, hialuronidase, condroitin sulfatase e atividade hemolítica de 18 cultivos de C. dubliniensis com 30 cultivos de C. albicans, todos isolados de pacientes com SIDA. Foi evidenciada maior atividade de proteinase em C. albicans em relação a C. dubliniensis (p < 0,05). Todos os isolados de C. dubliniensis evidenciaram atividade de hialuronidase e condroitin-sulfatase de forma similar ao observado com C. albicans (p > 0,05). Constatou-se que a atividade hemolítica foi influenciada pelo CaCl2; em sua ausência não foram observadas diferenças na atividade hemolítica das duas espécies; todavia, ao se agregar 2,5 por cento de CaCl2, a atividade hemolítica de C. dubliniensis foi reduzida enquanto a de C. albicans, estimulada (p < 0,05).

Freqüência e atividade enzimática de Candida spp. na cavidade oral de pacientes diabéticos do serviço de endocrinologia de um hospital de Fortaleza-CE / Frequency and enzymatic activity of Candida spp. oral cavity of diabetic patients of the service of endocrinology of a hospital of Fortaleza-CE

O diabetes mellitus (DM), uma doença endócrino-metabólica de alta e crescente prevalência, é citada como responsável pela ocorrência das candidíases orais. A candidíase constitui um espectro de infecções causadas por fungos do gênero Candida, sendo o seu agente mais comum a Candida albicans, embora outras espécies tenham sido identificadas (Candida tropicalis, Candida guillermondii, Candida glabrata, Candida krusei). O objetivo deste trabalho foi avaliar a freqüência e a atividade enzimática de Candida spp. na cavidade oral de pacientes diabéticos atendidos no Serviço de Endocrinologia do Hospital Universitário Walter Cantídio da Universidade Federal do Ceará (HUWC/UFC). Foram coletadas amostras de 48 pacientes diabéticos, de ambos os sexos, com situações variáveis de controle glicêmico. Os materiais clínicos foram colhidos com ajuda de swabs e semeados em placas de Petri contendo ágar-Sabouraud dextrose com cloranfenicol e incubado a 37ºC. Os crescimentos foram identificados pelas provas clássicas usadas em micologia. Depois, essas cepas de Candida foram submetidas a provas de detecção de enzimas fosfolipase e proteinase. Destas, 15 amostras (31,25 por cento) apresentaram cultura positiva para o gênero Candida. A espécie mais freqüente foi a C. albicans, com 80 por cento, seguida de C. tropicalis (13,3 por cento) e C. guillermondii (6,7 por cento). Quanto à pesquisa da atividade enzimática de Candida spp., foi observado que 86,6 por cento delas apresentaram atividade de proteinase e 80 por cento, de fosfolipase. Conclui-se com tais resultados que a C. albicans é a mais freqüente e que as espécies de Candida isoladas possuem fortes atividades enzimáticas.

Diabetes mellitus, a endocrine-metabolic disease, of high and increasing prevalence, is cited as responsible by the occurrence of oral candidiasis. Candidiasis constitutes a specter of infections caused by fungi of genera Candida; the most common agent is Candida albicans, but other species have also been identified (Candida tropicalis, Candida guillermondii, Candida glabrata, Candida krusei). The objective of this work was to evaluate frequency and enzymatic activity of Candida spp. in the oral cavity of diabetic patients taken care in the service of endocrinology of the University Hospital Walter Cantídio of the Federal University of the Ceará. Samples had been collected of 48 diabetic patients, men and women, with various situations of glicemic control. Clinical materials had been collected with aid of swab and harvested in plates of Petri contend Sabouraud agar dextrose with cloranfenicol and incubated to 37ºC. The grown were identified by the used classic tests in mycology. In the following, these Candida strains were submitted to tests to detect phospholipase and proteinase enzymes. Of these, 15 samples (31,35 percent) presented positive culture for the genera Candida. The species more frequent was C. albicans with 80 percent, followed by C. tropicalis (13.3 percent) and C. guilliermondii (6.7 percent). Asfor the research on the enzymatic activity of Candida sp. it was observed that 86.6 percent presented activity of proteinase and 80 percent of phospholipase. It was concluded with these results that C. albicans is more frequent and that Candida spp. isolated species have strong enzymatic activity.

Atividade enzimática, produção de slime e sensibilidade a antifúngicos de Candida sp / Enzymatic activity, slime production and antifungal agent sensitivity of Candida sp

A habilidade de Candida spp secretar enzimas extracelulares e slime tem sido associada como fatores de patogenicidade. Do total de 37 cepas de Candida sp, 100 por cento foram produtoras de proteinase, 83,8 por cento fosfolipase, 64,9 por cento slime e 100 por cento sensíveis ao fluconazol e itraconazol. Foram encontradas 17 tipagens (enzima/slime). Esta metodologia apresentou um bom índice discriminatório (D=0,93) podendo ser utilizado na caracterização fenotípica das leveduras.

Abilith of Candida spp to secrete extracellular enzymes and slime has been associated as pathogenicity factors. Out of a total of 37 strains of Candida sp, 100 percent were proteinase producers, 83.8 percent were phospholipase producers, 64.9 percent were slime producers and 100 percent were sensitive to fluconazole and itraconazole. Seventeen typings (enzymes/slime) were found. This methodology presented a good discrimination rate (D = 0.93) and could be used for phenotypic characterization of yeasts.

Yeast lipases: enzyme purification, biochemical properties and gene cloning

Lipases are placed only after proteases and carbohydrases in world enzyme market and share about 5% of enzyme market. They occur in plants, animals and microorganisms and are accordingly classified as plant, animal and microbial lipases. Wherever they exist, they function to catalyze hydrolysis of triglycerides to glycerol and fatty acid. Like carbohydrases and proteases, lipases of microbial origin enjoy greater industrial importance as they are more stable (compared to plant and animal lipases) and can be obtained in bulk at low cost. Majority of yeast lipases are extracelluar, monomericglycoproteins with molecular weight ranging between ~33 to ~65 kD. More than 50% reported lipases producing yeast, produce it in the forms of various isozymes. These lipase isozymes are in turn produced by various lipase encoding genes. Among many lipase producing yeasts Candida rugosa is most frequently used yeast as the source of lipase commercially. This review is aimed at compiling the information on properties of various yeast lipases and genes encoding them.