Effect of phenotypic switching on the biological properties and susceptibility to chlorhexidine in Candida krusei ATCC 14243.
Department of Oral Biology, University of Malaya, Kuala Lumpur, Malaysia.
Phenotypic switching is characterized as a virulence factor of Candida spp. This study was carried out to evaluate the phenotypic switching ability of C. krusei ATCC 14243 and to determine its effect on the biological properties, adherence capacity and susceptibility towards chlorhexidine digluconate (CHX). To induce switched generations C. krusei was cultured under nitrogen-depleted growth conditions by adding phloxine B. These phenotypically switched colonies were designated as the 1st generation. Subsequent sub-culturing was performed to produce the 2nd, 3rd and 4th switched generations. The recovery of the 3rd generation was the highest at 85.7% while that of the 4th generation was lower at 70.8%, and the recovery of the 1st and 2nd generations gradually reduced to 46.6% and 36.4%, respectively. All generations of C. krusei were susceptible towards CHX. The unswitched C. krusei was the most susceptible but the least adherent to coated hard surfaces. The 2nd generation was the least susceptible, but with the highest adherent ability. The minimum inhibition concentration and minimal fungicidal concentration of C. krusei of all generations were determined at 0.4 mg mL(-1). These observations suggest that the switching activity of C. krusei induces changes to its biological properties and susceptibility towards CHX.
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J Dent. 2012 Jul ;40 (7):609-15 22521700
The antifungal properties of chlorhexidine digluconate and cetylpyrinidinium chloride on oral Candida.
Department of Oral Biology, Faculty of Dentistry, University of Malaya, 50603 Kuala Lumpur, Malaysia.
INTRODUCTION C. tropicalis and C. krusei have emerged as virulent species causing oral infections. Both have developed resistance to commonly prescribed azole antifungal agents. OBJECTIVE The study aimed to determine the effect of mouth rinses containing chlorhexidine digluconate (CHX), cetylpyridinium chloride (CPC) and their combination (CHX-CPC) on the growth of these strains. METHODS The minimal inhibition concentrations (MIC) of the mouth rinses were determined. The growth curves of the strains produced under the mouth rinse-treated and untreated conditions, as well as alterations to the morphology of the growth colonies and cells following the treatments were compared and analysed. RESULTS The MICs of CPC compared to CHX mouth rinses were found to be lower for both Candida sp. In the mixed formulation, CPC doubled the inhibitory effect of CHX towards both Candida sp., while CHX quadrupled the activity of CPC towards C. tropicalis. The growth colonies also appeared coarse, wrinkled and dried. CONCLUSION The profound effects shown may suggest the fungicidal activities of the mouth rinses incorporated with CHX, CPC or their combination on both C. tropicalis and C. krusei. Gargling using mouth rinses with such fungicidal activity would enhance a rapid reduction in the candidal population of patients with fungal infection.
E1210, a new broad-spectrum antifungal, suppresses Candida albicans hyphal growth through inhibition of glycosylphosphatidylinositol biosynthesis.
Next Generation Systems Core Function Unit, Eisai Product Creation Systems, Eisai Co., Ltd., Tsukuba, Ibaraki, Japan. email@example.com
Continued research toward the development of new antifungals that act via inhibition of glycosylphosphatidylinositol (GPI) biosynthesis led to the design of E1210. In this study, we assessed the selectivity of the inhibitory activity of E1210 against Candida albicans GWT1 (Orf19.6884) protein, Aspergillus fumigatus GWT1 (AFUA_1G14870) protein, and human PIG-W protein, which can catalyze the inositol acylation of GPI early in the GPI biosynthesis pathway, and then we assessed the effects of E1210 on key C. albicans virulence factors. E1210 inhibited the inositol acylation activity of C. albicans Gwt1p and A. fumigatus Gwt1p with 50% inhibitory concentrations (IC(50)s) of 0.3 to 0.6 μM but had no inhibitory activity against human Pig-Wp even at concentrations as high as 100 μM. To confirm the inhibition of fungal GPI biosynthesis, expression of ALS1 protein, a GPI-anchored protein, on the surfaces of C. albicans cells treated with E1210 was studied and shown to be significantly lower than that on untreated cells. However, the ALS1 protein levels in the crude extract and the RHO1 protein levels on the cell surface were found to be almost the same. Furthermore, E1210 inhibited germ tube formation, adherence to polystyrene surfaces, and biofilm formation of C. albicans at concentrations above its MIC. These results suggested that E1210 selectively inhibited inositol acylation of fungus-specific GPI which would be catalyzed by Gwt1p, leading to the inhibition of GPI-anchored protein maturation, and also that E1210 suppressed the expression of some important virulence factors of C. albicans, through its GPI biosynthesis inhibition.
Med Mycol. 2011 Nov 25;: 22114891
Genetic and phenotypic evaluation of Candida albicans strains isolated from subgingival biofilm of diabetic patients with chronic periodontitis.
* Department of Oral Diagnosis, Microbiology and Immunology, Piracicaba Dental School, University of Campinas , Piracicaba , Brazil.
Candida spp. are commensal microorganisms that are part of the microflora of different sites within the oral cavity. In healthy subjects, who have an unaltered immunological status, these yeasts do not cause disease. However, in immunosuppressed individuals whose condition may have been caused by diabetes mellitus, Candida spp. can express different virulence factors and may consequently become pathogenic. Studies have detected the presence of Candida spp. in periodontal sites of patients with chronic periodontitis, especially those that are immunologically compromised. However, the role of these microorganisms in the pathogenesis of periodontal disease is still unknown. The objectives of this study were:(1) to isolate and identify Candida albicans strains from subgingival sites of diabetic patients with chronic periodontitis;(2) to evaluate the following virulence factors; colony morphology, proteinase, phospholipase and hemolysin activities and cell surface hydrophobicity (CSH) under different atmospheric conditions; and (3) to determine the genetic patterns of these C. albicans isolates. Microbial samples were collected from subgingival sites and seeded on CHROMagar for subsequent identification of C. albicans by polymerase chain reaction (PCR). For the phenotypic tests, all strains of C. albicans were grown under reduced oxygen (RO) and anaerobiosis (ANA) conditions. Genotypes were defined by the identification through PCR of the transposable introns in the 25S rDNA. The results obtained relative to virulence factors were analyzed according to the atmospheric condition or genetic group, using Chi-square and Wilcoxon non-parametric tests. In this study, 128 strains were identified as C. albicans and of these, 51.6% were genotype B, 48.4% were genotype A and Genotype C was not found. Most of the strains were alpha-hemolytic in both atmospheric conditions, without a statistical difference. However, when comparing the genotypes, 46.1% of the genotype A strains were beta-hemolytic. In relation to colony morphology, 100% of the strains under ANA showed rough colonies, which were especially prevalent in genotype A isolates. In contrast, most of the colonies were smooth under RO. C. albicans strains did not produce proteinase and phospholipase activity in the total absence of oxygen. In RO, most strains had high proteinase activity and were positive by phospholipase tests (P < 0.05). Hydrophobicity was higher in anaerobiosis and was noted mainly for genotype A isolates. In conclusion, environmental oxygen concentration influenced the virulence factors of C. albicans strains isolated from subgingival sites of diabetic and periodontal patients. In addition, genotype A seems to be more virulent based on the phenotypic tests evaluated in this study.
Antimicrobial activity of Calendula officinalis, Camellia sinensis and chlorhexidine against the adherence of microorganisms to sutures after extraction of unerupted third molars.
Raquel Lourdes Faria, Lincoln Marcelo Lourenço Cardoso, Gokithi Akisue, Cristiane Aparecida Pereira, Juliana Campos Junqueira, Antonio Olavo Cardoso Jorge, Paulo Villela Santos Júnior
São José dos Campos Dental School, Univ. Estadual Paulista, São José dos Campos, SP.
OBJECTIVE The objective of this study was to compare the antimicrobial effect of mouthwashes containing Calendula officinalis L., Camellia sinensis (L.) Kuntze and 0.12% chlorhexidine digluconate on the adherence of microorganisms to suture materials after extraction of unerupted third molars. MATERIAL AND METHODS Eighteen patients with unerupted maxillary third molars indicated for extraction were selected (n=6 per mouthwash). First, the patients were subjected to extraction of the left tooth and instructed not to use any type of antiseptic solution at the site of surgery (control group). After 15 days, the right tooth was extracted and the patients were instructed to use the Calendula officinalis, Camellia sinensis or chlorhexidine mouthwash during 1 week (experimental group). For each surgery, the sutures were removed on postoperative day 7 and placed in sterile phosphate-buffered saline. Next, serial dilutions were prepared and seeded onto different culture media for the growth of the following microorganisms: blood agar for total microorganism growth; Mitis Salivarius bacitracin sucrose agar for mutans group streptococci; mannitol agar for Staphylococcus spp.; MacConkey agar for enterobacteria and Pseudomonas spp., and Sabouraud dextrose agar containing chloramphenicol for Candida spp. The plates were incubated during 24-48 h at 37ºC for microorganism count (CFU/mL). RESULTS The three mouthwashes tested reduced the number of microorganisms adhered to the sutures compared to the control group. However, significant differences between the control and experimental groups were only observed for the mouthwash containing 0.12% chlorhexidine digluconate. CONCLUSIONS Calendula officinalis L. and Camellia sinensis (L.) Kuntze presented antimicrobial activity against the adherence of microorganisms to sutures but were not as efficient as chlorhexidine digluconate.
Pyridine-derived thiosemicarbazones and their tin(IV) complexes with antifungal activity against Candida spp.
Gabrieli L Parrilha, Jeferson G da Silva, Ludmila F Gouveia, Alan K Gasparoto, Roberta P Dias, Willian R Rocha, Daniel A Santos, Nivaldo L Speziali, Heloisa Beraldo
Departamento de Química, Instituto de Ciências Exatas, Universidade Federal de Minas Gerais, 31270-901 Belo Horizonte, MG, Brazil.
[(n-Bu)Sn(2Ac4oClPh)Cl2](1),[(n-Bu)Sn(2Ac4oFPh)Cl2](2),[(n-Bu)Sn(2Ac4oNO2Ph)Cl2](3),[(n-Bu)Sn(2Bz4oClPh)Cl2](4),[(n-Bu)Sn(2Bz4oFPh)Cl2](5) and [(n-Bu)Sn(2Bz4oNO2Ph)Cl2](6) were obtained by reacting [(n-Bu)SnCl3] with 2-acetylpyridine-N4-orthochlorophenyl thiosemicarbazone (H2Ac4oClPh), 2-acetylpyridine-N4-orthofluorphenyl thiosemicarbazone (H2Ac4oFPh), 2-acetylpyridine-N4-orthonitrophenyl thiosemicarbazone (H2Ac4oNO2Ph), and with the corresponding 2-benzoylpyridine-derived thiosemicarbazones (H2Bz4oClPh, H2ABz4oFPh and H2Bz4oNO2Ph). The antifungal activity of the studied compounds was evaluated against several Candida species. Upon coordination of H2Bz4oNO2Ph to tin in complex (6) the antifungal activity increased three times against Candida albicans and Candida krusei and six times against Candida glabrata and Candida parapsilosis. The minimum inhibitory concentration (MIC) values of H2Ac4oNO2Ph and its complex (3) against C. albicans, C. parapsilosis and C. glabrata are similar to that of fluconazole. All studied compounds were more active than fluconazole against C. krusei.
In vitro activity of isavuconazole against 140 reference fungal strains and 165 clinically isolated yeasts from Japan.
Toshikazu Yamazaki, Yukiko Inagaki, Toshihiko Fujii, Jun Ohwada, Masao Tsukazaki, Isao Umeda, Kazuko Kobayashi, Nobuo Shimma, Malcolm G P Page, Mikio Arisawa
Chugai Pharmaceutical Co. Ltd., Kamakura Research Laboratories, 200 Kajiwara Kamakura, Kanagawa 247-8530, Japan. firstname.lastname@example.org
The in vitro susceptibilities of 140 laboratory reference strains of fungi, including type strains, and 165 clinical yeast isolates from Japan towards isavuconazole compared with fluconazole (FLC), itraconazole (ITC), voriconazole and amphotericin B were measured. Broth microdilution methods based on Clinical and Laboratory Standards Institute (CLSI) methods were used for yeasts, and RPMI-MOPS medium semi-solidified with 0.2% low-melting-point agarose based on CLSI guidelines was used for moulds. The range of isavuconazole minimum inhibitory concentrations (MICs) was 0.0004-0.21 mg/L for Candida albicans, 0.0036-0.4 mg/L for Candida glabrata, 0.023-0.058 mg/L for Candida krusei, 0.0026-0.032 mg/L for Cryptococcus neoformans, 0.1-0.39mg/L for Aspergillus fumigatus and 0.2-0.39 mg/L for Aspergillus terreus. Isavuconazole was as active as ITC against the dimorphic true pathogenic fungi, with a range of MICs from <0.0004 mg/L to 0.0063 mg/L for Blastomyces dermatitidis and Histoplasma capsulatum. It was also active against uncommon dematiaceous fungi such as Exophiala spp. and Phialophora spp. as well as against dermatophytic species. Isavuconazole showed very good in vitro antifungal activity with a broad spectrum, including against FLC-resistant Candida spp., Aspergillus spp. and uncommon opportunistic fungal species. This is the first report of the in vitro susceptibility of Japanese clinical yeast isolates to isavuconazole. No cross-resistance was found to isavuconazole amongst FLC-resistant strains.
Role of Candida albicans Aft2p transcription factor in ferric reductase activity, morphogenesis and virulence.
Key Laboratory of Molecular Microbiology and Technology, Ministry of Education, Department of Microbiology, Nankai University, Tianjin, PR China.
The ability of Candida albicans to act as an opportunistic fungal pathogen is linked to its ability to switch among different morphological forms. This conversion is an important feature of C. albicans and is correlated with its pathogenesis. Many conserved positive and negative transcription factors regulate morphogenetic transition of C. albicans. Here, we show the results of functional analysis of CaAFT2, which is an orthologue of the Saccharomyces cerevisiae AFT2 gene. We have cloned CaAFT2 which has an ability to complement the S. cerevisiae aft1Δ mutant strain growth defect. Interestingly, although disruption of the AFT2 gene did not affect cell growth in solid and liquid iron-limited conditions, the cell surface ferric reductase activity was significantly decreased. Importantly, deletion of AFT2 in C. albicans led to growth of a smooth colony with no peripheral hyphae. Moreover, virulence of an aft2Δ/aft2Δ mutant was markedly attenuated in a mouse model. Our results suggest that CaAft2p represents a novel activator and that it functions in ferric reductase activity, morphogenesis and virulence in C. albicans.
Mycoses. 2009 Dec 17;: 20028461
Laboratorio de Micología Médica, Departamento de Inmunología, Microbiología y Parasitología, Facultad de Medicina y Odontología, Universidad del País Vasco-Euskal Herriko Unibertsitatea, Bilbao, Spain.
Summary The aim of the present study was to characterise phospholipase and proteinase activities of oral Candida isolates from 100 denture wearers and to study the relationship of these activities with denture stomatitis. Of 100 patients studied, 44 suffered from denture stomatitis. Specimens were collected by swabbing the denture and underlying mucosa. Isolates were previously identified by conventional mycological and genotypic methods. The phospholipase and proteinase activities were evaluated by agar plate methods. A total of 152 isolates were recovered from denture and underlying mucosa, including 101 Candida albicans, 18 Candida tropicalis, 14 Candida glabrata, 11 Candida guilliermondii, four Candida parapsilosis, two Saccharomyces cerevisiae and one isolate each of Candida dubliniensis and Candida krusei. Most C. albicans (97%) showed phospholipase activity; furthermore, the unique C. dubliniensis isolate showed a moderate phospholipase activity. The isolation of C. albicans (chi-square test, P = 0.0016) and phospholipase production by Candida spp.(chi-square test, P = 0.0213) was found to be significantly associated with denture stomatitis. Proteinase production was observed in <30% of isolates, and it was not related to the presence of denture stomatitis (P = 0.7675). Candida albicans isolates may produce both virulence factors, although the proteinase production was only observed in <30% of the isolates. Phospholipase production was exclusive of C. albicans and C. dubliniensis.
Characterising the post-antifungal effects of micafungin against Candida albicans, Candida glabrata, Candida parapsilosis and Candida krusei isolates.
Katherine T Nguyen, Philip Ta, Bich Thu Hoang, Shaoji Cheng, Binghua Hao, M Hong Nguyen, Cornelius J Clancy
Department of Medicine, University of Florida College of Medicine, Gainesville, FL, USA.
In this study, we measured time-kills and post-antifungal effects (PAFEs) for micafungin against Candida albicans (n=4), Candida glabrata (n=3), Candida parapsilosis (n=3) and Candida krusei (n=2) isolates and further characterised the PAFEs. Minimum inhibitory concentrations (MICs) were 0.5-1.0 mg/L against C. parapsilosis and 0.008-0.125 mg/L against the other species. Micafungin caused kills >1 log at 1 x MIC, 4 x MIC (range 1.19-3.10 log) and 16 x MIC (2.27-3.68 log), achieving fungicidal levels (> or = 3 log) against nine isolates. One-hour drug exposure during PAFE experiments resulted in kills of 0.73-2.88 log and 1.72-3.55 log at 4 x and 16 x MIC, respectively, achieving fungicidal levels against five isolates. Isolates of each species collected 8 h after a 1-h exposure to micafungin (4 x and 16 x MIC) were hypersusceptible to sodium dodecyl sulphate (SDS) and Calcofluor White. Cells tested during the PAFE period demonstrated cell wall disturbances as evident on electron micrographs as well as significant reductions in adherence to epithelial cells. Phagocytosis by J774 macrophages was significantly enhanced for three PAFE isolates tested. Micafungin is fungicidal and exerts PAFEs that kill diverse Candida spp., disturb cell walls of viable organisms, reduce adherence and enhance susceptibility to phagocytosis.
[In vitro activities of posaconazole, fluconazole, itraconazole, ketoconazole and voriconazole against Candida glabrata.]
This study has been conducted to asses the in vitro activity of the novel triazole antifungal agent posaconazole against 123 clinically important isolates of yeasts. Susceptibility was tested using the Sensititre YeastOne microdilution commercial method. Minimun inhibitory concentrations (MICs) were determined at the recommended endpoints and time intervals. The activity of posaconazole against Candida glabrata was compared with those of fluconazole, itraconazole, ketoconazole and voriconazole. The most susceptible species to posaconazole were C. albicans, C. parapsilosis, C. tropicalis and C. dubliniensis. Candida glabrata was the least susceptible. The percentage of strains with MIC for posaconazole >/= 1 mg/L was 9%, all of them were C. glabrata. The species with MIC for itraconazole >/= 0.5 mg/L were 36%(41 C. glabrata, 1 C. krusei, 1 C. guilliermondii, 1 C. ciferrii). Candida glabrata strains resistant to fluconazole, ketoconazole and voriconazole were 8%, 4% and 4%, respectively. Posaconazole exhibited good activity to the majority of Candida species. However, it was similar to itraconazole and less active than ketoconazole and voriconazole against C. glabrata. Key words: Posaconazole. Yeasts. Candida glabrata. Rev Esp Quimioter 2009;22(3):139-143.