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2.
Mem Inst Oswaldo Cruz ; 115: e200504, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32578684

RESUMO

BACKGROUND: Biodiversity screens and phylogenetic studies are dependent on reliable DNA sequences in public databases. Biological collections possess vouchered specimens with a traceable history. Therefore, DNA sequencing of samples available at institutional collections can greatly contribute to taxonomy, and studies on evolution and biodiversity. METHODS: We sequenced part of the glycosomal glyceraldehyde phosphate dehydrogenase (gGAPDH) and the SSU rRNA (V7/V8) genes from 102 trypanosomatid cultures, which are available on request at www.colprot.fiocruz.br. OBJECTIVE: The main objective of this work was to use phylogenetic inferences, using the obtained DNA sequences and those from representatives of all Trypanosomatidae genera, to generate phylogenetic trees that can simplify new isolates screenings. FINDINGS: A DNA sequence is provided for the first time for several isolates, the phylogenetic analysis allowed the classification or reclassification of several specimens, identification of candidates for new genera and species, as well as the taxonomic validation of several deposits. MAIN CONCLUSIONS: This survey aimed at presenting a list of validated species and their associated DNA sequences combined with a short historical overview of each isolate, which can support taxonomic and biodiversity research and promote culture collections.


Assuntos
Biodiversidade , Código de Barras de DNA Taxonômico , Trypanosomatina/classificação , Trypanosomatina/genética , Filogenia
3.
Braz J Microbiol ; 51(2): 427-436, 2020 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-31736016

RESUMO

Scedosporium spp. and Lomentospora prolificans are filamentous fungi that emerged as human pathogens; however, their mechanisms of virulence/pathogenesis are still largely unknown. In the present work, we have evaluated the interaction of S. apiospermum, S. minutisporum, S. aurantiacum, and L. prolificans with lung epithelial cells (A549 line). The results showed that conidia were able to interact with A549 cells, displaying association indexes of 73.20, 117.98, 188.01, and 241.63 regarding S. apiospermum, L. prolificans, S. minutisporum, and S. aurantiacum, respectively. Light microscopy images evidenced morphological changes in epithelial cells, including rounding and detachment, especially during the interaction with L. prolificans. Plasma membrane injuries were detected in A549 cells after 1 h of co-culturing with S. aurantiacum and S. minutisporum and after 4 h with S. apiospermum and L. prolificans, as judged by the passive incorporation of propidium iodide. After 24 h of fungi-epithelial cells interaction, only mycelia were observed covering the A549 monolayer. Interestingly, the mycelial trap induced severe damage in the A549 cells, culminating in epithelial cell death. Our results demonstrate some relevant events that occur during the contact between lung epithelial cells and Scedosporium/Lomentospora species, including conidial adhesion and hyphal growth with consequent irreversible injury on A549 cells, adding light to the infection process caused by these opportunistic and multidrug-resistant fungi.

4.
Curr Top Med Chem ; 19(28): 2527-2553, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31654512

RESUMO

Fungal infections are a veritable public health problem worldwide. The increasing number of patient populations at risk (e.g. transplanted individuals, cancer patients, and HIV-infected people), as well as the use of antifungal agents for prophylaxis in medicine, have favored the emergence of previously rare or newly identified fungal species. Indeed, novel antifungal resistance patterns have been observed, including environmental sources and the emergence of simultaneous resistance to different antifungal classes, especially in Candida spp., which are known for the multidrug-resistance (MDR) profile. In order to circumvent this alarming scenario, the international researchers' community is engaged in discovering new, potent, and promising compounds to be used in a near future to treat resistant fungal infections in hospital settings on a global scale. In this context, many compounds with antifungal action from both natural and synthetic sources are currently under clinical development, including those that target either ergosterol or ß(1,3)-D-glucan, presenting clear evidence of pharmacologic/pharmacokinetic advantages over currently available drugs against these two well-known fungal target structures. Among these are the tetrazoles VT-1129, VT-1161, and VT-1598, the echinocandin CD101, and the glucan synthase inhibitor SCY-078. In this review, we compiled the most recent antifungal compounds that are currently in clinical trials of development and described the potential outcomes against emerging and rare Candida species, with a focus on C. auris, C. dubliniensis, C. glabrata, C. guilliermondii, C. haemulonii, and C. rugosa. In addition to possibly overcoming the limitations of currently available antifungals, new investigational chemical agents that can enhance the classic antifungal activity, thereby reversing previously resistant phenotypes, were also highlighted. While novel and increasingly MDR non-albicans Candida species continue to emerge worldwide, novel strategies for rapid identification and treatment are needed to combat these life-threatening opportunistic fungal infections.


Assuntos
Antifúngicos/farmacologia , Candida/efeitos dos fármacos , Farmacorresistência Fúngica/efeitos dos fármacos , Micoses/tratamento farmacológico , Micoses/microbiologia , Animais , Antifúngicos/química , Candida/classificação , Humanos , Testes de Sensibilidade Microbiana , Especificidade da Espécie
5.
Front Microbiol ; 10: 1791, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31456762

RESUMO

Enteroaggregative Escherichia coli (EAEC) is an emerging pathotype responsible for acute and persistent diarrhea. It can be classified as typical and atypical strains, respectively, based on the presence or absence of the AggR regulon, suggesting a higher virulence for typical EAEC. This study aims to evaluate in the Galleria mellonella model if there are differences in the virulence profiles among clinical strains of typical and atypical EAEC, prototype strains EAEC C1096, 042 and its aggR mutant. The clinical EAEC strains (n = 20) were analyzed for the presence of 22 putative virulence factors of EAEC or extraintestinal E. coli by PCR, as well as phenotypic characteristics of virulence (enzymes, siderophore, and biofilm). The survival of the larvae was analyzed after inoculation of 104-107 CFU/larva; the monitoring of bacterial growth in vivo and hemocyte quantification was determined after inoculation of the prototype strains (105 CFU/larva) at different periods after infection. The strains of typical and atypical EAEC presented the same virulence profile for the larva, regardless of the amount or type of genes and phenotypic aspects of virulence analyzed. In addition, the EAEC 042 aggR mutant strain showed a significant reduction in the mortality of the inoculated larvae compared to the wild-type strain. In conclusion, the results obtained herein demonstrate that the virulence of EAEC seems to be related to the AggR regulon, but not exclusively, and atypical EAEC strains may be as virulent as typical ones in vivo in the G. mellonella model.

6.
Biometals ; 32(4): 671-682, 2019 08.
Artigo em Inglês | MEDLINE | ID: mdl-31230149

RESUMO

Hydrazide ligand, (Z)-N'-(6-oxo-1,10-phenanthrolin-5(6H)-ylidene)isonicotinohydrazide, 1 forms from a 1:1 Schiff base condensation reaction between isoniazid (INH) and 1,10-phenanthroline-5,6-dione (phendione). Ag+ and Mn2+ complexes with 1:2 metal:ligand stoichiometry are prepared: [Ag(1)2]NO3, [Ag(1)2]BF4 and [Mn(1)2](NO3)2. Polymeric {[Ag(1)(NO3)]}n has 1:1 stoichiometry and forms upon infusion of CH2Cl2 into a DMSO solution of [Ag(1)2]NO3. {[Ag(1)(NO3)]}n was structurally characterized using X-ray crystallography. Metal-free 1 and its 1:2 complexes exhibit very good, broad-spectrum antimicrobial activity and are not excessively toxic to mammalian cells (A549 lineage).


Assuntos
Anti-Infecciosos/química , Complexos de Coordenação/química , Isoniazida/química , Manganês/química , Fenantrolinas/química , Prata/química , Células A549 , Anti-Infecciosos/farmacologia , Sobrevivência Celular/efeitos dos fármacos , Humanos , Testes de Sensibilidade Microbiana
8.
PLoS One ; 13(9): e0203159, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-30231045

RESUMO

Flavonoids are one of the most important and diversified phenolic groups among products of natural origin. An important property of this metabolite class is the antioxidant action. This study evaluated the antioxidant and cytotoxic activities and oxidative stress of transesterification products of the flavonoid rutin, catalyzed by Novozym® 435. The presence of monoacetate and diacetate was confirmed by quantitative evaluation of the retention times (rutin, 15.68 min; rutin monoacetate, 18.14 min; and rutin diacetate, 18.57 min) and by the data from LC-MS and NMR 1H and 13C. The experiment showed excellent conversion values of 96% in total acetates (rutin monoacetate and diacetate). These results confirmed that rutin derivatives have antioxidant potential, as evaluated by the ORAC method (rutin standard: 0.53 ± 0.08 µM Trolox/g and rutin derivatives: 2.33 ± 1.08 µM Trolox/g) and also show low cytotoxicity in human and animal cells. Rutin derivatives reduced the production of reactive oxygen species in RAW macrophages as well. Many qualities attributed to rutin derivatives make them promising potential candidates for use as nutraceuticals, including their high amounts of antioxidants, biological potential and low toxicity, which contribute to the reduction of oxidative stress.


Assuntos
Lipase/metabolismo , Rutina/análogos & derivados , Animais , Antioxidantes/química , Antioxidantes/metabolismo , Antioxidantes/farmacologia , Sobrevivência Celular/efeitos dos fármacos , Chlorocebus aethiops , Esterificação , Células Hep G2 , Humanos , Macrófagos/citologia , Macrófagos/efeitos dos fármacos , Macrófagos/metabolismo , Camundongos , Ressonância Magnética Nuclear Biomolecular , Estresse Oxidativo , Células RAW 264.7 , Espécies Reativas de Oxigênio/metabolismo , Rutina/metabolismo , Rutina/farmacologia , Células Vero
9.
Parasitol Res ; 117(7): 2085-2094, 2018 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-29728827

RESUMO

Despite the available drug options, leishmaniasis treatment remains unsatisfactory. The repurposing of calpain inhibitors originally developed for human diseases became an interesting alternative, since Leishmania cells express calpain-related proteins. The susceptibility of six Leishmania species (L. amazonensis, L. braziliensis, L. major, L. mexicana, L. chagasi, and L. donovani) to the calpain inhibitor MDL28170 was determined. Promastigote and intracellular amastigote viability in the presence of MDL28170 was evaluated. MDL28170 was able to reduce promastigote proliferation in a dose-dependent manner for all the parasites. A significant reduction on the general parasite metabolism was detected, as judged by resazurin assay, as well as induced important morphological alterations, including rounding promastigotes and loss of the flagellum. MDL28170 was also able to reduce the number of intracellular amastigotes in RAW macrophages. The susceptibility of both parasite stages (promastigotes and amastigotes) to MDL28170 was similar for all Leishmania species tested. MDL28170 showed a much higher toxicity to Leishmania amastigotes when compared with mammalian macrophages, displaying selectivity index values varying from 13.1 to 39.8. These results suggest that the development of calpain inhibitors may represent an interesting alternative in the treatment of leishmaniasis.


Assuntos
Antiprotozoários/farmacologia , Inibidores de Cisteína Proteinase/farmacologia , Dipeptídeos/farmacologia , Glicoproteínas/farmacologia , Leishmania/crescimento & desenvolvimento , Leishmania/metabolismo , Animais , Linhagem Celular , Humanos , Leishmania/efeitos dos fármacos , Leishmaniose/tratamento farmacológico , Macrófagos/parasitologia , Camundongos , Camundongos Endogâmicos BALB C , Testes de Sensibilidade Parasitária , Células RAW 264.7
10.
Mediators Inflamm ; 2017: 8952878, 2017.
Artigo em Inglês | MEDLINE | ID: mdl-28814823

RESUMO

Candida glabrata is a facultative intracellular opportunistic fungal pathogen in human infections. Several virulence-associated attributes are involved in its pathogenesis, host-pathogen interactions, modulation of host immune defenses, and regulation of antifungal drug resistance. This study evaluated the in vitro antifungal susceptibility profile to five antifungal agents, the production of seven hydrolytic enzymes related to virulence, and the relationship between these phenotypes in 91 clinical strains of C. glabrata. All C. glabrata strains were susceptible to flucytosine. However, some of these strains showed resistance to amphotericin B (9.9%), fluconazole (15.4%), itraconazole (5.5%), or micafungin (15.4%). Overall, C. glabrata strains were good producers of catalase, aspartic protease, esterase, phytase, and hemolysin. However, caseinase and phospholipase in vitro activities were not detected. Statistically significant correlations were identified between micafungin minimum inhibitory concentration (MIC) and esterase production, between fluconazole and micafungin MIC and hemolytic activity, and between amphotericin B MIC and phytase production. These results contribute to clarify some of the C. glabrata mechanisms of pathogenicity. Moreover, the association between some virulence attributes and the regulation of antifungal resistance encourage the development of new therapeutic strategies involving virulence mechanisms as potential targets for effective antifungal drug development for the treatment of C. glabrata infections.


Assuntos
Antifúngicos/farmacologia , Candida glabrata/enzimologia , Anfotericina B/farmacologia , Candida glabrata/efeitos dos fármacos , Farmacorresistência Fúngica , Equinocandinas/farmacologia , Fluconazol/farmacologia , Hidrólise , Itraconazol/farmacologia , Lipopeptídeos/farmacologia , Micafungina , Testes de Sensibilidade Microbiana , Virulência
11.
Dalton Trans ; 46(16): 5297-5307, 2017 Apr 19.
Artigo em Inglês | MEDLINE | ID: mdl-28382355

RESUMO

Trypanosoma cruzi and Leishmania amazonensis are the causative agents of Chagas' disease and leishmaniasis, respectively. These conditions affect millions of people worldwide, especially in developing countries. As such, there is an urgent need for novel, efficient and cost-effective treatments for these diseases, given the growing resistance and side-effects of current therapies. This work details the synthesis and evaluation of the anti-parasitic activity of novel amino- and iminopyridyl metal chelators, their glycosylated derivatives and some of their metal complexes. Our results revealed the potent and metal-dependent activity for the aminopyridyl compounds: Cu(ii) complexes were most effective against T. cruzi trypomastigotes, while Zn(ii) complexes presented excellent activity against L. amazonensis promastigotes. In addition, the compounds showed excellent selectivity indexes and very low relative toxicity as judged by in vitro and in vivo studies, respectively, using RAW macrophages and Galleria mellonella larvae model.


Assuntos
Antiparasitários/síntese química , Antiparasitários/farmacologia , Quelantes/síntese química , Quelantes/farmacologia , Leishmania/efeitos dos fármacos , Trypanosoma cruzi/efeitos dos fármacos , Aminopiridinas/química , Animais , Antiparasitários/química , Quelantes/química , Cobre/química , Glicosilação , Humanos , Interações Hidrofóbicas e Hidrofílicas , Larva/efeitos dos fármacos , Camundongos , Mariposas , Células RAW 264.7 , Sensibilidade e Especificidade , Zinco/química
12.
Front Microbiol ; 8: 76, 2017.
Artigo em Inglês | MEDLINE | ID: mdl-28194139

RESUMO

Phialophora verrucosa is a dematiaceous fungus able to cause chromoblastomycosis, phaeohyphomycosis and mycetoma. All these fungal diseases are extremely difficult to treat and often refractory to the current therapeutic approaches. Therefore, there is an urgent necessity to develop new antifungal agents to combat these mycoses. In this context, the aim of the present work was to investigate the effect of 1,10-phenanthroline-5,6-dione (phendione) and its metal-based derivatives [Ag(phendione)2]ClO4 = ([Ag(phendione)2]+) and [Cu(phendione)3](ClO4)2.4H2O = ([Cu(phendione)3]2+) on crucial physiological events of P. verrucosa conidial cells. Using the CLSI protocol, we have shown that phendione, [Ag(phendione)2]+ and [Cu(phendione)3]2+ were able to inhibit fungal proliferation, presenting MIC/IC50 values of 12.0/7.0, 4.0/2.4, and 5.0/1.8 µM, respectively. [Cu(phendione)3]2+ had fungicidal action and when combined with amphotericin B, both at sub-MIC (½ × MIC) concentrations, significantly reduced (~40%) the fungal growth. Cell morphology changes inflicted by phendione and its metal-based derivatives was corroborated by scanning electron microscopy, which revealed irreversible ultrastructural changes like surface invaginations, cell disruption and shrinkages. Furthermore, [Cu(phendione)3]2+ and [Ag(phendione)2]+ were able to inhibit metallopeptidase activity secreted by P. verrucosa conidia by approximately 85 and 40%, respectively. Ergosterol content was reduced (~50%) after the treatment of P. verrucosa conidial cells with both phendione and [Ag(phendione)2]+. To different degrees, all of the test compounds were able to disturb the P. verrucosa conidia-into-mycelia transformation. Phendione and its Ag+ and Cu2+ complexes may represent a promising new group of antimicrobial agents effective at inhibiting P. verrucosa growth and morphogenesis.

13.
Curr Top Med Chem ; 2017 Jan 05.
Artigo em Inglês | MEDLINE | ID: mdl-28056742

RESUMO

The past decades have witnessed a dramatic increase in invasive fungal infections, especially caused by different species belonging to the Candida genus. Nowadays, even after many improvements in several medical procedures, Candida infections (candidiasis) still account for an unacceptable high rate of morbimortality in hospital settings. Corroborating this statement, fungal biofilms formed on both abiotic and living surfaces are responsible for an important medical and economic burden, since biofilm lifestyle confers numerous advantages to the pathogens, including high tolerance to environmental stresses such as antimicrobials and host immune responses. Aggravating this scenario, the currently used antifungal drugs have mostly been developed to target exponentially growing fungal cells and are poorly or not effective against biofilm structures. So, the challenges to inhibit biofilm formation (e.g., blocking the fungal adhesion and its fully development due to the changes of physicochemical properties of the inert substrates by covering or impregnating them with antimicrobial compounds, for example, silver nanoparticles) and/or to disarticulate mature biofilm architecture (e.g., by using compounds capable in destabilizing, weakening or destroying the extracellular matrix components, including inhibitors of quorum sensing signals, hydrolytic enzymes, surfactants, chelator agents and biocides) are stimulating researchers around the world to search novel strategies and new chemotherapeutic options to control fungal biofilm. In this context, the present review summarizes some promising approaches and/or strategies that could improve our ability to prevent or eradicate fungal biofilms in medical settings, focusing on the lessons learned with Candida model.

14.
Bioelectromagnetics ; 38(2): 95-108, 2017 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-27783424

RESUMO

Available treatments against human fungal pathogens present high levels of resistance, motivating the development of new antifungal therapies. In this context, the present work aimed to analyze direct electric current (DC) antifungal action, using an in vitro apparatus equipped with platinum electrodes. Candida albicans yeast cells were submitted to three distinct conditions of DC treatment (anodic flow-AF; electroionic flow-EIF; and cathodic flow-CF), as well as different charges, ranging from 0.03 to 2.40 C. Our results indicated C. albicans presented distinct sensibility depending on the DC intensity and polarity applied. Both the colony-forming unit assay and the cytometry flow with propidium iodide indicated a drastic reduction on cellular viability after AF treatment with 0.15 C, while CF- and EIF-treated cells stayed alive when DC doses were increased up to 2.40 C. Additionally, transmission electron microscopy revealed important ultrastructural alterations in AF-treated yeasts, including cell structure disorganization, ruptures in plasmatic membrane, and cytoplasmic rarefaction. This work emphasizes the importance of physical parameters (polarity and doses) in cellular damage, and brings new evidence for using electrotherapy to treat C. albicans pathology process. Bioelectromagnetics. 38:95-108, 2017. © 2016 Wiley Periodicals, Inc.


Assuntos
Candida albicans/citologia , Eletricidade , Candida albicans/metabolismo , Candida albicans/fisiologia , Candida albicans/ultraestrutura , Adesão Celular , Linhagem Celular , Eletrodos , Células Epiteliais/citologia , Humanos , Masculino , Viabilidade Microbiana , Pessoa de Meia-Idade , Platina/química , Propídio/metabolismo
15.
Parasitology ; 144(2): 117-123, 2017 02.
Artigo em Inglês | MEDLINE | ID: mdl-27869056

RESUMO

Leishmaniasis is a neglected disease, which needs improvements in drug development, mainly due to the toxicity, parasite resistance and low compliance of patients to treatment. Therefore, the development of new chemotherapeutic compounds is an urgent need. This opinion article will briefly highlight the feasible use of calpain inhibitors as leading compounds to search for new therapeutic options to treat leishmaniasis. The milestone of this approach is to take advantage on the myriad of inhibitors developed against calpains, some of which are in advanced clinical trials. The deregulated activity of these enzymes is associated with several pathologies, such as strokes, diabetes and Parkinson's disease, to name a few. In Leishmania, calpain upregulation has been associated to drug resistance and virulence. Whereas the difficulties in developing new drugs for neglected diseases are more economical than biotechnological, repurposing approach with compounds already approved for clinical use by the regulatory agencies can be an interesting shortcut to a successful chemotherapeutic treatment for leishmaniasis.


Assuntos
Antiprotozoários/farmacologia , Calpaína/antagonistas & inibidores , Calpaína/metabolismo , Leishmania/metabolismo , Leishmaniose/tratamento farmacológico , Animais , Antiprotozoários/química , Humanos , Leishmania/efeitos dos fármacos , Doenças Negligenciadas/tratamento farmacológico
16.
BMC Microbiol ; 15: 188, 2015 Sep 29.
Artigo em Inglês | MEDLINE | ID: mdl-26415499

RESUMO

BACKGROUND: Angomonas deanei is a trypanosomatid parasite of insects that has a bacterial endosymbiont, which supplies amino acids and other nutrients to its host. Bacterium loss induced by antibiotic treatment of the protozoan leads to an aposymbiotic strain with increased need for amino acids and results in increased production of extracellular peptidases. In this work, a more detailed examination of A. deanei was conducted to determine the effects of endosymbiont loss on the host calpain-like proteins (CALPs), followed by testing of different calpain inhibitors on parasite proliferation. RESULTS: Western blotting showed the presence of different protein bands reactive to antibodies against calpain from Drosophila melanogaster (anti-Dm-calpain), lobster calpain (anti-CDPIIb) and cytoskeleton-associated calpain from Trypanosoma brucei (anti-CAP5.5), suggesting a possible modulation of CALPs influenced by the endosymbiont. In the cell-free culture supernatant of A. deanei wild type and aposymbiotic strains, a protein of 80 kDa cross-reacted with the anti-Dm-calpain antibody; however, no cross-reactivity was found with anti-CAP5.5 and anti-CDPIIb antibodies. A search in A. deanei genome for homologues of D. melanogaster calpain, T. brucei CAP5.5 and lobster CDPIIb calpain revealed the presence of hits with at least one calpain conserved domain and also with theoretical molecular mass consistent with the recognition by each antibody. No significant hit was observed in the endosymbiont genome, indicating that calpain molecules might be absent from the symbiont. Flow cytometry analysis of cells treated with the anti-calpain antibodies showed that a larger amount of reactive epitopes was located intracellularly. The reversible calpain inhibitor MDL28170 displayed a much higher efficacy in diminishing the growth of both strains compared to the non-competitive calpain inhibitor PD150606, while the irreversible calpain inhibitor V only marginally diminished the proliferation. CONCLUSIONS: Altogether, these results indicate that distinct calpain-like molecules are expressed by A. deanei, with a possible modulation in the expression influenced by the endosymbiont. In addition, treatment with MDL28170 affects the growth rate of both strains, as previously determined in the human pathogenic species Leishmania amazonensis and Trypanosoma cruzi, with whom A. deanei shares immunological and biochemical relationships.


Assuntos
Bactérias/crescimento & desenvolvimento , Calpaína/antagonistas & inibidores , Calpaína/biossíntese , Glicoproteínas/metabolismo , Simbiose , Trypanosomatina/crescimento & desenvolvimento , Trypanosomatina/microbiologia , Calpaína/genética , Trypanosomatina/efeitos dos fármacos , Trypanosomatina/genética
17.
Rev. iberoam. micol ; 32(2): 122-125, abr.-jun. 2015. tab
Artigo em Inglês | IBECS | ID: ibc-137315

RESUMO

Background: cases of superficial and invasive mycoses caused by emerging species of Candida have been increasingly reported over the last thirty years. The production of hydrolytic enzymes plays a central role in the fungal infective process. In Candida infections the secretion of both proteases and phospholipases are well-known virulence attributes. Aims: to determine the protease and phospholipase production from 58 human clinical isolates of Candida obtained from individuals with cutaneous candidiasis seen in the Human and Veterinary Diagnostic Mycology Sector from Universidade Federal Fluminense (UFF), Brazil, from November 2008 to August 2009. Methods: fungal identification was performed using biochemical tests. Proteolytic activity was detected on agar plates containing bovine serum albumin, and phospholipase production was determined on egg-yolk plates. Results: the Candida species isolated were Candida parapsilosis (27.59%), Candida famata (18.96%), Candida albicans (15.52%), Candida haemulonii (12.06%), Candida ciferri (8.62%), Candida guilliermondii (6.90%), Candida tropicalis (5.17%) and Candida lipolytica (5.17%). All isolates of C. albicans produced both protease and phospholipase. As regards the isolates of non-C. albicans Candida species, 53.06% and 4.08% were able to produce protease and phospholipase, respectively. For example, the majority of isolates of C. parapsilosis (15/16) produced protease, while 40% of C. ciferri isolates (2/5) were phospholipase producers. This study shows, for the first time, that C. ciferri and C. haemulonii strains were able to produce protease. Conclusions: collectively, our results showed that different species of Candida isolated from cutaneous lesions were able to produce proteases and/or phospholipases, which are multifunctional molecules directly involved in the infectious process of these fungi


Antecedentes: Los casos de micosis superficiales e invasoras relacionados con las especies emergentes de Candida se han reportado progresivamente durante las últimas tres décadas. La producción de enzimas hidrolíticas juega un papel central en varios contextos de la patogenicidad fúngica. Con respecto a la infección por Candida, la secreción de proteasas y fosfolipasas son atributos de virulencia bien conocidos. Objetivos: Determinar y comparar la producción de proteasa y fosfolipasa de 58 aislamientos clínicos humanos de diferentes especies de Candida obtenidos de pacientes con candidiasis cutánea, atendidos en el Sector de Diagnóstico Micológico Humano y Veterinario de la Universidad Federal Fluminense (UFF), durante el período de noviembre de 2008 a agosto de 2009. Métodos: La identificación de las especies de Candida se realizó mediante pruebas bioquímicas, la actividad proteolítica se detectó en placas de agar que contenían albúmina de suero bovino y la actividad fosfolipasa se determinó utilizando el método de la placa de yema de huevo semi-cuantitativa. Resultados: Las especies aisladas fueron Candida parapsilosis (27,59%), Candida famata (18,96%), Candida albicans (15.52%), Candida haemulonii(12,06%), Candida ciferri(8,62%), Candida guilliermondii(6,90%), Candida tropicalis (5,17%) y Candida lipolytica (5,17%). Todos los aislamientos de C. albicans produjeron tanto proteasa como fosfolipasa. El 53,06% de los aislamientos de Candida no-C. albicans fueron capaces de producir proteasa y el 4,08% fosfolipasa. La mayoría de los aislamientos de C. parapsilosis (15/16) produjo proteasa, mientras que el 40% de los aislamientos de C. ciferri (2/5) fueron productores de fosfolipasa. Se describe por primera vez en la literatura científica la producción de proteasas por cepas de C. haemulonii y C. ciferri. Conclusiones: Nuestros resultados muestran el potencial que tienen los aislamientos de Candida provenientes de lesiones cutáneas para producir proteasas y fosfolipasas (AU)


Assuntos
Humanos , Candidíase Cutânea/microbiologia , Candida/patogenicidade , Fosfolipases/análise , Peptídeo Hidrolases/análise , Candida/enzimologia , Fatores de Virulência
18.
Mycopathologia ; 179(3-4): 231-42, 2015 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-25502596

RESUMO

Phialophora verrucosa is one of the etiologic agents of chromoblastomycosis, a fungal infection that affects cutaneous and subcutaneous tissues. This disease is chronic, recurrent and difficult to treat. Several studies have shown that secreted peptidases by fungi are associated with important pathophysiological processes. Herein, we have identified and partially characterized the peptidase activity secreted by P. verrucosa conidial cells. Using human serum albumin as substrate, the best hydrolysis profile was detected at extreme acidic pH (3.0) and at 37 °C. The enzymatic activity was completely blocked by classical metallopeptidase inhibitors/chelating agents as 1,10-phenanthroline and EGTA. Zinc ions stimulated the metallo-type peptidase activity in a dose-dependent manner. Several proteinaceous substrates were cleaved, in different extension, by the P. verrucosa metallopeptidase activity, including immunoglobulin G, fibrinogen, collagen types I and IV, fibronectin, laminin and keratin; however, mucin and hemoglobin were not susceptible to proteolysis. As metallopeptidases participate in different cellular metabolic pathways in fungal cells, we also tested the influence of 1,10-phenanthroline and EGTA on P. verrucosa development. Contrarily to EGTA, 1,10-phenanthroline inhibited the fungal viability (MIC 0.8 µg/ml), showing fungistatic effect, and induced profound morphological alterations as visualized by transmission electron microscopy. In addition, 1,10-phenanthroline arrested the filamentation process in P. verrucosa. Our results corroborate the supposition that metallopeptidase inhibitors/chelating agents have potential to control crucial biological events in fungal agents of chromoblastomycosis.


Assuntos
Antifúngicos/farmacologia , Proteínas Fúngicas/metabolismo , Metaloproteases/metabolismo , Fenantrolinas/farmacologia , Phialophora/efeitos dos fármacos , Phialophora/enzimologia , Esporos Fúngicos/crescimento & desenvolvimento , Humanos , Micoses/microbiologia , Phialophora/crescimento & desenvolvimento , Sistemas de Translocação de Proteínas/metabolismo , Transporte Proteico , Esporos Fúngicos/efeitos dos fármacos , Esporos Fúngicos/enzimologia
19.
Rev Iberoam Micol ; 32(2): 122-5, 2015.
Artigo em Inglês | MEDLINE | ID: mdl-24853474

RESUMO

BACKGROUND: Cases of superficial and invasive mycoses caused by emerging species of Candida have been increasingly reported over the last thirty years. The production of hydrolytic enzymes plays a central role in the fungal infective process. In Candida infections the secretion of both proteases and phospholipases are well-known virulence attributes. AIMS: To determine the protease and phospholipase production from 58 human clinical isolates of Candida obtained from individuals with cutaneous candidiasis seen in the Human and Veterinary Diagnostic Mycology Sector from Universidade Federal Fluminense (UFF), Brazil, from November 2008 to August 2009. METHODS: Fungal identification was performed using biochemical tests. Proteolytic activity was detected on agar plates containing bovine serum albumin, and phospholipase production was determined on egg-yolk plates. RESULTS: The Candida species isolated were Candida parapsilosis (27.59%), Candida famata (18.96%), Candida albicans (15.52%), Candida haemulonii (12.06%), Candida ciferri (8.62%), Candida guilliermondii (6.90%), Candida tropicalis (5.17%) and Candida lipolytica (5.17%). All isolates of C. albicans produced both protease and phospholipase. As regards the isolates of non-C. albicans Candida species, 53.06% and 4.08% were able to produce protease and phospholipase, respectively. For example, the majority of isolates of C. parapsilosis (15/16) produced protease, while 40% of C. ciferri isolates (2/5) were phospholipase producers. This study shows, for the first time, that C. ciferri and C. haemulonii strains were able to produce protease. CONCLUSIONS: Collectively, our results showed that different species of Candida isolated from cutaneous lesions were able to produce proteases and/or phospholipases, which are multifunctional molecules directly involved in the infectious process of these fungi.


Assuntos
Candida/enzimologia , Candidíase Cutânea/microbiologia , Proteínas Fúngicas/análise , Peptídeo Hidrolases/análise , Fosfolipases/análise , Brasil , Candida/classificação , Candida/isolamento & purificação , Candida/patogenicidade , Humanos , Técnicas de Tipagem Micológica , Especificidade da Espécie , Virulência
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