Diploid formation of Candida tropicalis via protoplast fusion.

Haploid auxotrophic mutants were produced from Candida tropicalis, and protoplast usion was induced by polyethylene glycol. The resulting nutritional complementation was due to heterokaryon formation and, at a much lower frequenty, to spontaneous diploidization. During cultivation, heterokaryotic clones regularly gave rise to heterozygous diploids from which, in turn, haploids could be isolated. The technique of protoplast fusion gives an opportunity for genetic analysis of this and similarly asexual fungal species.     

Metabolic formation of dodecanedioic acid from n-dodecane by a mutant of Candida tropicalis

Summary These experiments studied the metabolic formation of a,-dodecanedioic acid using the mutant S 76 developed from the wild strain Candida tropicalis 1230 (capable of producing large amounts of a,-dodecanedioic acid).Our results show for the first time that 12-hydroxydodecanoic acid was excreted into the medium as a free acid.n-Dodecanol and n-dodecanoic acid were also detected in the n-dodecane medium. The mutant S 76 was able to produce a,-dodecanedioic acid using either n-dodecanol or dodecanoic acid as the sole carbon source. Quantitative cahnges in the concentrations of 12-hydroxy-dodecanoic acid and other intermediates were recorded during the formation of a,-dodecanedioic acid. S 76 was rapidly able to convert large amounts of 12-hydroxy-dodecanoic acid to a,-dodecanedioic acid.The formation of a,-dodecanedioic acid from n-dodecane via the sequence n-dodecanoln-dodecanoic acid 12-hydroxy-dodecanoic acid was confirmed.     

Microbial transformation of primaquine by Candida tropicalis

The microbial metabolism of primaquine, a 6-methoxy-8-aminoquinoline antimalarial agent, was investigated. The yeast Candida tropicalis was found to convert primaquine to the previously reported N-acetylated derivative. On continued incubation of C. tropicalis in the presence of the N-acetylated derivative, a minor dimeric metabolite was formed. The proposed structure of the metabolite was based primarily on the analysis of its spectroscopic properties (1H and 13C nuclear magnetic resonance spectra and field-desorption mass spectrum). The structure of the metabolite was proven by direct comparison with an authentic sample of the minor dimeric metabolite prepared by treatment of the N-acetylated derivative with formaldehyde in the presence of formic acid in methanol.     

Transglycosyl-Amylase of Candida tropicalis

Transglucosyl-amylase was purified 96-fold and partially characterized. The K_m value with dextrin as substrate was 9.1 mg/ml. Glycerol, an acceptor of d-glucose, appeared to inhibit dextrin hydrolysis noncompetitively. The energy of activation of the enzyme was 7,920 cal/mole. Indirect determinations showed that synthesis of d-glucosyl glycerol was significantly affected by the nature of the amylaceous substrate. Glucosyl-glycerol synthesis did not increase as incubation temperature was raised from 50 to 60 C. Direct determinations by gas-liquid chromatography indicated that the synthesis of glucosyl glycerol, as a function of the concentration of either enzyme, substrate, or glycerol, traced a curvilinear path approaching 15 mg/ml as the maximum. When enzyme, substrate, and glycerol at high concentrations were varied in all possible combinations, however, conditions for producing as much as 47.5 mg/ml of glucosyl glycerol were established.     

Trnasglucosyl-amylase of Candida tropicalis

Transglucosyl-amylase was purified 96-fold and partially characterized. The K(m) value with dextrin as substrate was 9.1 mg/ml. Glycerol, an acceptor of d-glucose, appeared to inhibit dextrin hydrolysis noncompetitively. The energy of activation of the enzyme was 7,920 cal/mole. Indirect determinations showed that synthesis of d-glucosyl glycerol was significantly affected by the nature of the amylaceous substrate. Glucosyl-glycerol synthesis did not increase as incubation temperature was raised from 50 to 60 C. Direct determinations by gas-liquid chromatography indicated that the synthesis of glucosyl glycerol, as a function of the concentration of either enzyme, substrate, or glycerol, traced a curvilinear path approaching 15 mg/ml as the maximum. When enzyme, substrate, and glycerol at high concentrations were varied in all possible combinations, however, conditions for producing as much as 47.5 mg/ml of glucosyl glycerol were established.     

Microbial Production of d-Arabitol by n-Alkane-grown Candida tropicalis

In the course of study on citric acid fermentation by Candida tropicalis KY6224, in which n-alkane mixture (C–12 to C–15) was used as the sole source of carbon, we found that a arabitollike substance was accumulated when the medium-pH was controlled at low level (3.0 to 4.0). This substance was isolated in crystalline forms and identified as d-arabitol.

d-Arabitol production was also observed with ethanol, acetic acid and glucose as the sole source of carbon. Important factors for efficient production of d-arabitol were keeping the medium-pH at low-level (3.0 to 4.0) and the concentration of NaCl or KCl at high level (1 to 5%). This strain produced 75 mg/ml of d-arabitol in 120 hr incubation under optimal culture conditions; this corresponds to 50 % of n-alkane consumed.     

Microbial transformation of primaquine by Candida tropicalis.

The microbial metabolism of primaquine, a 6-methoxy-8-aminoquinoline antimalarial agent, was investigated. The yeast Candida tropicalis was found to convert primaquine to the previously reported N-acetylated derivative. On continued incubation of C. tropicalis in the presence of the N-acetylated derivative, a minor dimeric metabolite was formed. The proposed structure of the metabolite was based primarily on the analysis of its spectroscopic properties (1H and 13C nuclear magnetic resonance spectra and field-desorption mass spectrum). The structure of the metabolite was proven by direct comparison with an authentic sample of the minor dimeric metabolite prepared by treatment of the N-acetylated derivative with formaldehyde in the presence of formic acid in methanol.     

Targeted integrative transformation of Candida tropicalis by electroporation

Summary A method for integrative transformation of the diploid yeast Candida tropicalis by electroporation has been developed. By linearizing the transforming plasmid DNA containing the URA3 gene prior to electroporation of recipient cells, its integration was targeted to a specific locus in the genome, resulting in single or multiple tandem integrations. The optimal electroporation conditions for this yeast were established and include an electric pulse of 2.25 kV/cm for a duration of 50 ms. Using these conditions, Ura⁺ transformants were readily obtained at a high frequency (45 transformants/g DNA) as the result of targeted integration of the URA3 gene containing plasmid DNA at the chromosomal ura3 locus. The number of transformants resulting from this procedure is comparable to that achieved with a recently reported spheroplast transformation procedure for C. tropicalis; in addition, it offers the advantages of being simple, rapid and reproducible.     

Phagocyte-mediated killing of Candida tropicalis

Human peripheral monocytes (MO), neutrophils (PMN), and lymphocytes (PBL) were tested for their ability to kill Candida tropicalis. With incubation times between 30 min and 2 h, unstimulated MO and PMN, but not PBL, were efficient killers of C. tropicalis. Both leukocyte subsets were able to kill at minimum 2.5 1 effector to target ratios. Pre-incubation of MO for 24 h with interferon-gamma or tumor necrosis factor (TNF) increased their ability to kill yeast targets. TNF alone had no effect on C. tropicalis targets at concentrations up to 1000 U/ml. PBL activated for 4 d with interleukin-2 did not kill yeast targets. PMN exhibited more cytocidal efficiency per cell than MO in these assays. Direct contact of effectors and targets was required; no significant killing by PMN or MO supernatants was measured. PMN-mediated killing, but not MO killing, was inhibited by a mixture of catalase and Superoxide dismutase suggesting that oxygen-dependent killing mechanisms were partially responsible for candidacidal activity.     

Hydrocarbon emulsification by Candida tropicalis and Debaryomyces polymorphus

Potentiality of C. tropicalis and D. polymorphus, to produce surface active compounds (bioemulsifiers/biosurfactants) during shake cultivation on hexadecane and oily waste was studied. Better emulsification activity, specific towards aromatic hydrocarbons, was observed with C. tropicalis culture broth. Emulsification activity of culture broth was quite stable and was unaffected by change in pH and by increasing the concentration of NaCl up to 5%. The activity was marginally affected by heating in boiling water bath for 15 min, but inhibited to the tune of 90% by 0.3% CaCl2. The isolated bioemulsifying factor contained 40, 22 and 17.5% lipid, protein and carbohydrate, respectively.     

1例热带念珠菌脓肿误诊分析

报告1例热带念珠菌引起的腹腔脓肿瘘管真菌感染。患者女,78岁,因十二指肠球部穿孔并发腹膜炎,行腹腔镜术,术后伤口反复未完全愈合,有少量分泌物溢出。发现腹壁瘘管及腹腔脓肿入院,两次细菌培养均无菌生长,再次取分泌物直接镜检阳性,真菌培养鉴定为热带念珠菌。手术清理组织,病理示感染性肉芽肿改变,查见大量假菌丝及孢子。经口服伊曲康唑和两性霉素B溶液纱条引流治疗3个月后基本痊愈。