Comparison of pathogenicity of various Candida albicans and C. stellatoidea strains

In order to clarify the pathogenicity and the pathogenic factors of various Candida species strains, three strains, NIH A-207 and J-1012 (serotype A), and NIH B-792 (serotype B) of Candida albicans and two strains, ATCC 20408 (karyotype II) and ATCC 36232 (karyotype I) of C. stellatoidea, a synonym for C. albicans, were tested for their lethality to mice, adherence to Hela cells, hydrophobicity, and cell growth under acidic conditions, pH 2.0-5.9. The pathogenicity for mice of all the strains was observed in the order NIH B-792, ATCC 36232, J-1012, NIH A-207, and ATCC 20408. The pathogenicity for mice by all the strains used was well correlated with adherence to the Hela cells, the hydrophobicity, and the cell growth under the acidic condition, pH 2.0. These results emphasize that these specific properties of the C. albicans and C. stellatoidea strains play an important role in the pathogenesis of candidosis.     

Adherence of Candida albicans is inhibited by yeast mannan

The adherence of Candida albicans strain NIH A207 to plastic plates was inhibited by the addition of mannan, and plates coated with mannan also inhibited the adherence of C. albicans strains TIMM1768, TIMM2640, and JCM2076. Mannan coated the plastic plates under neutral and acidic conditions, but not under alkaline conditions. These results indicate that C. albicans cannot attach to mannan. Thus mannan-coated medical equipment might be useful to prevent C. albicans adherence.     

Antigenicity of cell wall mannans of Candida albicans and Candida stellatoidea cultured at high temperatures in BACTEC medium

The study of the antigenicity of pathogenic Candida albicans and Candida stellatoidea cells grown in BACTEC fungal medium (BFM) is useful for clinical analysis so as accurately to diagnose candidiasis. When C. albicans NIH A-207 was grown in BFM and fetal bovine serum-added BFM at the high temperatures of 36 and 40 degrees C, the cell density increased, with a mixture of yeast cells, pseudohyphae, and hyphae and with full hyphal development in the cultures compared with cultivation (mostly cells in yeast form) at 27 degrees C in both media. The mannans produced when cells were grown at these high temperatures were less reactive by enzyme-linked immunosorbent assay with factor sera 4, 5, and 6 in the commercially available kit 'Candida Check' than were the mannans obtained following growth at 27 degrees C. Based on 1H-nuclear magnetic resonance analysis, the mannans from cells grown at high temperatures had lost a phosphate group and a beta-1,2-linked mannopyranose unit, and had increased the number of non-reducing terminal alpha-1,3-linked mannopyranose units. We obtained similar results for mannans produced by C. albicans J-1012, C. albicans NIH B-792, C. albicans JCM 9061, C. stellatoidea ATCC 20408, and C. stellatoidea ATCC 36232 strains cultivated in BFM at 36 degrees C. These results suggest that both C. albicans and C. stellatoidea cells cultured at high temperatures, irrespective of the medium and shape of the cells, alter their antigenicity and chemical structure of cell wall mannans.     

Influence of Fungicidal Activity against Candida tropicalis on the Efficacy of Micafungin and Liposomal Amphotericin B in a Neutropenic Murine Lethal Infection Model

Objectives: To investigate the correlation between in vitro killing activity and in vivo efficacy of micafungin (MCFG) and liposomal amphotericin B (L-AMB) against Candida tropicalis in a neutropenic murine lethal infection model. Methods: Candida albicans (one strain) and C. tropicalis (three strains) were tested in time-kill studies. Cyclophosphamide-treated mice were inoculated intravenously with each strain. One day after inoculation, antifungals were administered intravenously once daily for 1 or 3 days. Results: MCFG exhibited fungicidal activity against C. albicans ATCC 90029 and C. tropicalis SP-20142, and fungistatic activity against C. tropicalis ATCC 42678 and SP-20047. The ED(50)s (dosage that results in 50% survival) of MCFG for C. tropicalis ATCC 42678 and SP-20047 (4.1-50 mg/kg) were higher than those for other strains (1.6-12 mg/kg). A 1-day course of MCFG was not effective against C. tropicalis ATCC 42678 and SP-20047 at the clinical dose (5 mg/kg), which achieved an AUC level almost equal to that of 100 mg in humans, whereas a 3-day course of 5 mg/kg MCFG was efficacious against all strains. In contrast, L-AMB showed fungicidal activity against all strains tested and the ED(50)s of L-AMB were 0.08-0.65 mg/kg. In both treatment regimens, the minimum effective doses of L-AMB (≤0.5 mg/kg) were less than the clinical dosage (≤5 mg/kg). Conclusions: The in vivo efficacy of MCFG and L-AMB showed a correlation with the in vitro killing activity. At the clinical dose, L-AMB exerted anti-C. tropicalis activity within a shorter treatment period than MCFG.     

Similarity of acid proteinases secreted by Candida albicans NIH A-207 and NIH B-792 strains cultured in BSA-supplemented medium

Candida albicans NIH A-207 (serotype A) and NIH B-792 (serotype B) strains secreted one acid proteinases (AP) each in a yeast carbon-based medium supplemented with bovine serum albumin (BSA) as the sole nitrogen source. Isolation of AP from the culture filtrates was achieved by dialysis, followed by DEAE-Sepharose and Biogel P-100 column chromatographies. It was found that both enzymes from the two strains had very similar properties when examined. The molecular weights and isoelectric points were found to be 43 kDa and pH 4.0, respectively. The amino acid components and first 12 N-terminal amino acid sequences were virtually identical in both enzymes. The optimum pH of the enzymes was 3.5-4.0. The enzymes were heat-labile, and decreases in their activities were found above 37 degrees C. The AP activities were completely inhibited by the addition of pepstatin. No other inhibitor among those tested had any effect. The enzymes degraded all proteins examined, especially host defense factors such as immunoglobulin G and the granulocyte colony stimulating factor. The enzymes also caused similar degrees of enhancement of vascular permeability when they were injected into the dorsal skin of guinea pigs.     

Adhesion of Candida albicans to HeLa cells: studies using polystyrene beads

The adherence to HeLa cells by the yeast-type cells of the Candida albicans NIH A-207 strain cultivated for 2 d at 27 degrees C in the yeast extract-added Sabouraud liquid medium (YSLM) and the 500 mM galactose-added yeast nitrogen base medium (YNB-Gal) was compared. The yeast cells cultured in the YNB-Gal clearly increased the adherence to the HeLa cells compared to the YSLM. The adherence drastically decreased by pronase treatment of the yeast cells. Next, to define the sugar receptors of the yeast cells, lactosylceramide (LC)-, the H type 1 antigen (HA)-, Lewis(a) antigen (Le(a))-, mannan- and BSA-coated polystyrene beads were used for the adherence to the yeast cells. Only the LC- and HA-beads were bound to the yeast cells. The adherence of the two beads to the yeast cells cultured in the YNB-Gal was higher than that to the yeast cells cultured in the YSLM. The yeast cell wall mannan-coated polystyrene beads did not adhere at all to the Hela cells. Based on these results, it is evident that the protein parts involving the LC and HA receptors on the surface of the yeast cells correlate with the adherence to the HeLa cells.     

Production and biological activities of a new antifungal antibiotic, TAN-950 A.

A novel antifungal antibiotic, TAN-950 complex, was isolated from the culture filtrate of Streptomyces platensis A-136 (IFO 14603, FERM BP-1786). The water-soluble amphoteric substances in this complex were purified by chromatography using ion-exchange resins, QAE-Sephadex and adsorptive resins and were designated TAN-950 A and TAN-950 A-E mixture. The molecular formula of TAN-950 A was determined to be C6H7N2O4Na for the sodium salt. This new amino acid antibiotic showed antifungal activity against Candida albicans in vitro and in vivo, and had low toxicity in mice.     

血液真菌感染的菌群分布及耐药性分析

目的分析本院近3年来血液真菌的分离率、菌种分布和耐药特性。方法收集自2004年8月～2007年7月3年间住院患者的血液标本,经BacT／ALERT3D血培养仪培养,沙保罗培养基分离培养真菌,柯玛嘉显色培养基和VITEK-60YBC卡进行菌种鉴定,Rosco纸片扩散法测定药敏。结果8707份血液标本中共分离出93株真菌,分离率为1．07％,其中白念珠菌41株,占44．09％,热带念珠菌16株,占17．20％,光滑念珠菌13株,占13．98％,对氟康唑的耐药率分别为：2．4％、6．3％和23．1％,对两性霉素B和制霉菌素均敏感。结论我院血液真菌感染的病原菌以白念珠菌为主,热带念珠菌和光滑念珠菌次之,白念珠菌和热带念珠菌对唑类药物敏感性较高,光滑念珠菌对唑类药物的耐药率较高,两性霉素B和制霉菌素的抗菌活性最强。     

Subtyping and antifungal susceptibilities of Candida spp. in the intensive care unit of a Greek general hospital

This study identified the Candida spp., susceptibility to antifungal agents and the prevailing Candida albicans subtypes responsible for infections or colonization of 42 patients in the ICU over a 6-month period. Most isolates were C. albicans (66.1%) and Candida tropicalis (28.3%) all of which were susceptible in vitro to antifungal agents. Subtypes of the C. albicans isolates were identified by pulsed field gel electrophoresis Sfi I chromosomal digests. Two major C. albicans subtypes were identified, whereas subtype heterogeneity was found among strains of Candida glabrata and C. tropicalis. Sfi I PFGE restriction patterns were able to discriminate between sub-populations of C. albicans isolates, clustering them into distinct, epidemiologically congruous groups.     

In vitro antifungal activity of itraconazole: a comparative study with ketoconazole]

In vitro antifungal activities of itraconazole (ITZ), a new oral triazole antifungal agent, were studied against a wide range of medically important fungi including 16 genera, 37 species and 51 strains stocked in this center. The test was carried out using the agar dilution method on Sabouraud dextrose agar. ITZ showed equal or superior antifungal activities to ketoconazole against most strains of pathogenic yeasts, dimorphic fungi, non-pigmented hypomycetes, dermatophytes and dematiacious fungi. Although some strains of Candida albicans and Candida tropicalis were not completely inhibited by ITZ at concentrations up to 80 micrograms/ml, partial growth inhibitions were observed even at drug concentrations as low as 0.04 microgram/ml. The antifungal activity of ITZ against C. albicans was markedly influenced by medium composition, medium pH, inoculum size and incubation time.     

The antifungal activities of some 6-[N-(halophenyl)amino]-7-chloro-5,8-quinolinediones againstCandida species

A series of 6-[N-(halophenyl)amino]-7-chloro-5,8-quinolinedione derivatives 1-10 were tested for antifungal susceptibilities, in vitro, against pathogenic Candida species such as C. albicans, C. glabrata, C. krusei, C. parapsilosis and C. tropicalis. The MICs were determined by the standard macrodilution techniques, according to the NCCLS 1992 guidelines. The 6-[N-(halophenyl)amino]-7-chloro-5,8-quinolinedione derivatives showed generally potent antifungal activities against pathogenic Candida species. Among them, derivative 1, 2, 5 and 7 showed more potent antifungal activities than ketoconazole. All derivatives 1-10 had specially potent activities against C. tropicalis. Derivative 1 and 2 containing (N-3,4-dihalo-phenyl)amino moiety exhibited the potent antifungal activities. Derivative 2 with (3,4-dichlorophenyl)amino substituent was the most effective in preventing the growth of Candida species at MICs 4 micrograms/ml respectively.     

Susceptibility of several species of Candida and Torulopsis to fluconazole and ketoconazole

The authors compared the in vitro antifungal activity of fluconazole, a new triazole antifungal agent, and ketoconazole, an imidazole derivative. The MIC values were determined against 50 strains of Candida albicans, 10 strains of C. guilliermondii, 10 strains of C. krusei, 10 strains of C. parapsilosis, 10 strains of C. pseudotropicalis, 10 strains of C. tropicalis and 15 strains of Torulopsis glabrata. The fungistatic activity was evaluated by the agar dilution method using BHI and casitone media after incubation for 48 h at 28-30 degrees C. Both antifungal agents showed higher activity when tested on casitone medium; however, the G-MIC values for ketoconazole were lower than those for fluconazole.     

深部病原性真菌感染及耐药性分析

目的:探讨深部病原性真菌感染的特点及耐药趋势.方法:用回顾调查的方法对我院住院病人及门诊病人各类标本培养的68株病原性真菌对常用抗真菌药物药敏试验结果进行分析.结果:分离出的68株病原真菌以内科系统为主,共52株(占76.47%);检出病原性真菌最多的部位是呼吸道,占真菌标本的64.71%;真菌以白色假丝酵母菌、近平滑假丝酵母菌和类星型念球菌为主,分别占58.82%、8.82%、8.82%;药敏结果,氟胞嘧啶、两性霉素B 90%敏感,白色假丝酵母菌对氟康唑、伊曲康唑、伏立康唑的耐药率很高均在70%以上,5种常见抗真菌药物对其他病原真菌的敏感率均在90%以上.结论:加强临床病原性真菌分离株,对抗真菌药物的耐药性检测,然后根据药敏试验结果科学合理选用抗真菌药物十分重要.     

我院深部真菌感染的菌群分布及药敏分析

目的了解深部真菌感染的菌群分布及其耐药性。方法用CHRoM agar显色培养基及ATB(法国梅里埃)半自动真菌鉴定及药敏分析仪进行菌种鉴定及药敏分析。结果 634例深部真菌中共检出白假丝酵母菌339株,分离率最高为53.5%;光滑假丝酵母菌220株,分离率为34.7%;热带假丝酵母菌42株,分离率为6.6%。分离率较高的白假丝酵母菌和光滑假丝酵母菌对两性霉素B保持了较高的敏感率,分别为97.9%和94.5%;对5-氟咆嘧啶的敏感率分别为96.8%和86.4%;对氟康唑和伊曲康唑的敏感率最低。结论深部真菌感染已成为医院感染的重要原因之一,不同真菌对常用的4种抗真菌药物的敏感性有所不同,故快速而准确的鉴定及药敏对临床上深部真菌感染的及时诊断和治疗有重要的意义。     

198例泌尿系统感染真菌耐药性分析

目的探讨医院近5年来泌尿系统感染真菌的耐药性,为临床合理使用抗真菌药物提供依据。方法对重庆市急救医疗中心临床标本中段尿真菌培养分离的菌株鉴定,用APIAUX20C真菌ID条板或Ryid Nicroscan真菌ID条板进行鉴定分型,纸片扩散法进行药物敏感试验。结果共分离到真菌198株,其中白色念珠菌129株（65.15%）,热带念珠菌34株（17.17%）;对两性霉素B、5-氟胞嘧啶、伊曲康唑、氟康唑的耐药率,白色念珠菌分别为8.53%,7.75%,41.86%,8.53%,热带念珠菌分别为5.88%,20.59%,61.76%,20.59%,克柔念珠菌分别为18.18%,18.18%,22.73%,18.18%,光滑珠念菌分别为0,0,42.86%,71.43%。结论泌尿系统感染真菌分离率高,以白色念珠菌和热带念珠菌为主;联合应用两性霉素B和5-氟胞嘧啶对于控制尿路感染具有重要意义。     

Effects of subinhibitory concentrations of amikacin and ciprofloxacin on the hydrophobicity and adherence to epithelial cells of uropathogenic Escherichia coli strains

The effect of subinhibitory concentrations of amikacin and ciprofloxacin on the hydrophobicity and adherence to uroepithelial cells of Escherichia coli strains was investigated. The hydrophobicity of the tested strains was evaluated by the bacterial adherence to hydrocarbon–xylene test and by the salt aggregation test of ammonium sulphate. The hydrophobic character of strains exposed to 1/2 to 1/8 minimal inhibitory concentration (MIC) of amikacin and 1/2 to 1/16 MIC of ciprofloxacin was altered to a hydrophilic state. Results of the SAT also correlated with these data. Moreover, comparisons were made between the number of bacteria attached to the epithelial cells before and after exposure to 1/2, 1/4 and 1/8 MIC of antibiotics. The greatest loss of adherence capability occurred at 1/2 MIC of ciprofloxacin. In conclusion, antibiotics are often present at sub-MICs and may still be effective in reducing bacterial virulence by interfering with bacterial cell functions.     

Synthesis and antifungal activity of 6,7-bis-[S-(aryl)thio]-5,8-quinolinediones

6,7-Bis-[S-(aryl)thio]-5,8-quinolinediones 4 and 5 were synthesized by the substitution of 6,7-dichloro-5,8-quinolinediones with appropriate arylthiols. Their antifungal activity were tested in vitro for their growth inhibitory activities against pathogenic fungi in comparison with flucytosine. The antifungal activities were significantly improved by S-(aryl)thio moieties of the compounds 4 and 5. The all tested compounds 4 and 5 showed generally good activities against C. albicans and A. niger ranging from 0.8 to 25 microg/ml. Among them, compounds 4d-4h and 5a-5c exhibited also good activities against C. krusei and C. tropicalis. The activities of compounds 4j and 4l were comparable to those of flucytosine against all tested fungi.     

In vitro activity of a new semisynthetic echinocandin, micafungin, against clinical isolates of Candida species isolated in Tenri Hospital

We have examined the antifungal activities of the available antifungal agents including micafungin (MCFG), one of the echinocandin antifungal group, against 92 yeast-like fungi isolated at our hospital during a 3-month period from November 2002 to February 2003. Determination of the antifungal susceptibility was conducted in conformity with the Standards of the Japanese Society for Medical Mycology. The MIC 80% of the antifungal agents against 4 fungi species including C. albicans (55 strains), C. tropicalis (20 strains), C. glabrata (8 strains), C. krusei (5 strains) were as follows; MCFG: 0.03-0.125 microgram/ml, amphotericin-B: 0.125-0.25 microgram/ml, 5-fluorocytosine: 0.125-16 micrograms/ml, itraconazole: 0.25-2 micrograms/ml, fluconazole: 0.5-32 micrograms/ml. The isolation rate of the drug-resistant fungi was 20% for the fluconazole (FLCZ)-resistant C. tropicalis and 33% when including the susceptible dose dependent (S-DD) class. The rate was 5% for FLCZ-resistant strains of C. albicans and 11% when including the S-DD class. However, MCFG was shown to have an excellent antifungal activity against those azole-resistant strains of Candida species. An analysis of the randomly amplified polymorphic DNA pattern (RAPD) was carried out to assess the fingerprinting of the azole-resistant strains. The results demonstrated a common pattern in 3 of the 6 strains of C. tropicalis that showed MIC of > or = 16 micrograms/ml for fluconazole, while all of the 6 strains of C. albicans demonstrated their respective patterns.     

Antifungal resistance in Candida spp. isolated in Italy between 2002 and 2005 from children and adults

The activity of amphotericin B, fluconazole, flucytosine, itraconazole and voriconazole was tested in vitro against 618 clinical Candida spp. isolates, using the broth microdilution or the disk diffusion method (voriconazole). Amphotericin B and voriconazole were the most potent antifungal agents assayed (100% of susceptible strains). Resistance to fluconazole and itraconazole was detected in three (0.7%) and 11 (2.7%) isolates of Candida albicans and in four (3.7%) isolates of Candida glabrata. Flucytosine intermediate, resistant strains, or both, were observed in C. albicans (0.3% and 0.7%), C. glabrata (2.8% intermediate) and C. tropicalis (15.2% and 15.2%). C. krusei was the least susceptible species to azoles. No statistically significant differences in the rates of resistant isolates depending on site of infection and age of the patient were observed, with the exception of C. albicans and itraconazole (higher percentage of resistance in children). At present, acquired antifungal resistance represents an uncommon finding in most Candida spp. circulating in Northern Italy.     

In vitro antifungal activity of amorolfine, a new morpholine antimycotic agent

In vitro antifungal activities of amorolfine (MT-861), a newly developed morpholine antimycotic agent, against a wide range of pathogenic fungal strains were investigated using an agar-dilution method with an imidazole antifungal agent, clotrimazole (CTZ), as the reference drug. The results showed that MT-861 had a broad antifungal spectrum, and was active against all of the pathogenic fungi tested except nonpigmented filamentous fungi such as aspergilli and penicillia. Dermatophytes and Malassezia furfur was the most highly susceptible to MT-861. Antifungal activities of MT-861 against most strains of these fungi as well as those against most strains of dimorphic fungi were higher than those of CTZ. By contrast, MT-861 showed lower activities against pathogenic yeasts such as Candida albicans than CTZ. Several assay conditions, such as inoculum size of fungi, incubation time, media pH and compositions including serum supplementation, affected MT-861 activities against C. albicans and, to lesser extents, those against Trichophyton mentagrophytes. MIC values of MT-861 against C. albicans and other Candida spp., were the lowest on casitone agar. MT-861 exerted fungicidal actions toward susceptible fungi such as T. mentagrophytes and Sporothrix schenckii at relatively low concentrations, and these activities were increased when the time of incubation was extended beyond 24 hours after inoculation of testing organisms. Susceptibilities of any strains of C. albicans and C. glabrata to MT-861 were not reduced by as many as 15 successive transfers in MT-861 containing media.