Candida's arranged marriage.

Biologists who study the fungus Candida albicans have always assumed that this organism reproduces asexually because they have not found evidence of mating, meiosis, or a haploid stage of the life cycle. However, as Gow et al. explain in a Perspective, sequencing of the C. albicans genome has revealed the existence of a possible mating type locus. This finding has now been extended to demonstrate actual mating in the fungus (Hull et al., Magee and Magee).

Candida albicans GRX2, encoding a putative glutaredoxin, is required for virulence in a murine model.

Resistance of Candida albicans to reactive oxygen species is thought to enhance its virulence in mammalian hosts. Genes such as SOD1, which encodes the anti-oxidant, superoxide dismutase, are known virulence factors. We disrupted the gene GRX2, which encodes a putative glutathione reductase (glutaredoxin) in C. albicans, and we compared the mutant with an sod1Deltamutant. In vitro, the grx2Deltastrain, but not the sod1Delta strain, was defective in hypha formation. The grx2Deltastrain, but not sod1Delta, was significantly more susceptible to killing by neutrophils. When exposed to two compounds that generate reactive oxygen species, both mutants were susceptible to 1 mM menadione, but grx2Deltanull alone was resistant to diamide. Both mutants were attenuated in a murine intravenous challenge model, and a GRX2 reintegrant regained partial virulence. Emphasis on the putative function of products of genes such as SOD1 and GRX2 in resistance to oxidative stress may oversimplify their functions in the virulence process, since the grx2Deltastrain also gave defective hypha formation. Both mutants were sensitive to menadione and were slow to form germ tubes, though growth rates matched controls once the lag phase was passed.

Molecular mechanisms of drug resistance in fungi.

Failures of treatment in fungal infections have drawn attention recently to the problem of antifungal resistance and its underlying mechanisms. The number of fungal isolates that are resistant to the orally active azole antifungals, especially fluconazole, is growing. Amphotericin-B-resistant isolates have been recovered during treatment of patients with candidiasis, and resistance to flucytosine is so common that this antifungal is no longer recommended as a single-drug therapy.

Detection of Candida precipitins. a comparison of double diffusion and counter immunoelectrophoresis.

The optimum conditions for detection of Candida precipitins by double diffusion and counter immunoelectrophoresis have been established. Counter immunoelectrophoresis was shown to be more sensitive than double diffusion, and its application to routine tests for Candida precipitins to cytoplasmic antigen is recommended.

Intraconazole--a new oral antifungal agent with a very broad spectrum of activity in superficial and systemic mycoses.

Itraconazole is a triazole-derivative antifungal agent with an extremely broad spectrum of action in vitro, in experimental animals in vivo and in clinical trials. When taken orally, itraconazole achieves high and sustained levels of active drug in many tissues, including skin, nail and most deep organs. Its clinical value has been proved in all forms of mycosis affecting the skin and mucous membranes and in onychomycosis. In systemic infections, clinical trials have led to recommendations that itraconazole is the current drug of choice for blastomycosis, histoplasmosis, paracoccidioidomycosis, lymphocutaneous sporotrichosis, chronic disseminated aspergillosis and many cases of phaeohyphomycosis. It is also finding application in deep-seated Candida infections and in cryptococcosis. Its safety profile is extremely good, with very few non-trivial adverse experiences noted in a large series of patients treated and monitored. Itraconazole therefore represents a useful therapeutic advance for the management of most forms of fungal infection.

Distribution of pathogenic yeasts and humoral antibodies to candida among hospital inpatients.

The frequencies of the carriage of yeast pathogens and of serum precipitins to a variety of candida antigens among 254 patients generally tended to increase with the length of the patient's stay in hospital. This trend was observed even though none of the patients investigated showed signs or symptoms of superficial or systemic candidosis. The extent of the general trend varied considerably between subgroups of patients within the general categories of 'surgical' and 'nonsurgical' inpatients. Increases in both frequencies and quantities of yeasts in the mouth were most evident postoperatively among patients who underwent open-heart surgery and among nonsurgical patients who received antibiotics or steroids in hospital. The frequency of precipitins to Candida albicans cytoplasmic antigens in the absence of candidosis rose overall from 11% of 217 sera obtained within 24 hours of admission to 35% of 85 sera obtained five to 11 days after admission or operation. These 'false positive' antibodies were thought to arise after transient yeast overgrowth in the gut at the time of an acute illness or immediately after surgery. The study adds further data to documented examples of 'false positive' candida antibodies and indicates the need for care in the diagnostic interpretation of candida precipitin test results among groups of patients at risk of yeast overgrowth during their hospital stay.

Prevalence of pathogenic yeasts and humoral antibodies to candida in diabetic patients.

The prevalence of oral yeasts and humoral precipitating antibodies to candida was estimated in 204 unselected diabetic patients (172 outpatients and 32 inpatients). Yeasts, mainly Candida albicans, were isolated from the mouths of 41% of the outpatients and precipitins were found in 17.5% although none of the patients had clinically overt candidiasis. The extent of oral yeast colonisation and incidence of antibodies was not related to their antidiabetic treatment or to the duration of their diabetes. It was, however, related to the blood glucose and urine sugar levels at the time they were sampled, the highest incidence being among the diabetic inpatients with high blood glucose levels at the time of sampling and the lowest among outpatients with normal blood glucose levels at the time of sampling. There was no such correlation when diabetic control over the previous 12-month period was considered.

Carriage of Candida species and C albicans biotypes in patients undergoing chemotherapy or bone marrow transplantation for haematological disease.

Six hundred and seventy four yeast isolates obtained from routine microbiological screening of 153 patients with haematological disease were identified and Candida albicans isolates biotyped over nine months to determine longitudinal and cross sectional patterns of yeast colonisation. A yeast microflora persisted in many patients despite the routine prophylactic use of oral antifungal agents. Analysis of the yeast species isolated on a cross sectional basis showed that C albicans accounted for 65% of yeasts isolated from the oral cavity but only 45% of the faecal yeast flora. Longitudinal changes in yeast flora occurred significantly more often in faecal samples than in oral samples and significantly less often in sites colonised with C albicans than in sites colonised with other species. No associations were found between the yeasts isolated and the nature of antifungal prophylaxis used, or the extent of a patient's stay in hospital.

"Room temperature" use of CHROMagar Candida.

The color of colonies of 9 Candida species was examined on the chromogenic medium CHROMagar Candida incubated for 24-72 h at 25 degrees C, 30 degrees C or 37 degrees C. Colors and colony forms characteristic of C. albicans, C. dubliniensis, C. krusei and C. tropicalis were formed most rapidly and with the deepest hues at 37 degrees C. After 48 h incubation at 25 degrees C, 9 of 48 C. albicans isolates gave pink colonies instead of the green colonies characteristic for the species, and the blue-purple colony color characteristic of C. tropicalis isolates was not formed until 48 h at 25 degrees C. Incubation of the chromogenic medium at temperatures below 30 degrees C cannot be recommended for reliable presumptive identification of Candida spp., and pink colonies of C. glabrata would not be reliably distinguished from pink colonies formed by other species under any of the incubation conditions used.

Clonality structure in Candida dubliniensis.

Multilocus enzyme electrophoresis was performed on 76 European strains of Candida dubliniensis. Ten of the 20 enzyme-encoding loci were polymorphic, giving rise to 10 electrophoretic types within the sample studied. Investigation of the population genetics of a subset of 36 strains from HIV-infected patients in London showed the existence of strong heterozygote deficits and excesses associated with significant linkage disequilibria between pairs of loci. These findings, together with the predominance of multilocus genotypes, strongly suggest that C. dubliniensis is mainly (if not totally) clonal. Analysis of genotypes of a larger number of strains should confirm this conclusion and improve our understanding of the epidemiology of this pathogen.

Inducible proteinase of Candida albicans in diagnostic serology and in the pathogenesis of systemic candidosis.

The extracellular acid proteinase of Candida albicans was purified from culture filtrates by a single-column chromatographic step. The purity of the enzyme and its unique antigenic properties were confirmed by polyacrylamide-gel electrophoresis and by reaction with homologous and heterologous anti-sera. The purified enzyme (PP), which was a carboxyl proteinase, contained mannan as an integral part of the molecule. C. albicans proteinase was detected in experimental candida kidney lesions by indirect immunoflourescence. Precipitating antibodies to PP and to cytoplasmic extract (CE) were detected in sera from rabbits with chronic, experimental, systemic candidosis; however precipitins to PP were not found in sera from infected rabbits in which tissue invasion was prevented by antifungal treatment. In retrospective tests with sera from healthy subjects and from patients with and without proven systemic candidosis a qualitative distinction between true and false-positive precipitins to PP was not found; however, whereas 72% of sera from proven cases of deep-seated candida infection had anti-PP titres greater than 4 and greater than or equal to anti-CE titres, these same quantitative criteria were met by only 15% of sera from patients for whom information of a diagnosis of candidosis was not available. The purified proteinase was therefore a more specific antigen than the widely used cytoplasmic extract for detection of antibodies in cases of candidosis.

Disseminated Candida infection syndrome in heroin addicts--dominance of a single Candida albicans biotype.

Among 21 intravenous heroin abusers with cutaneous and ocular manifestations of disseminated Candida infection, a single C. albicans strain type (serotype A, biotype 153/7) was isolated from skin lesions in 14 cases. This suggests that central contamination of the heroin with C. albicans is less likely to be the source of infection than an endogenous source, and that one particular strain type is either better adapted than others to grow in the lemon juice used as a heroin solvent, or more likely than others to cause the specific pathology seen in these patients.

Candida species and C. albicans biotypes in women attending clinics in genitourinary medicine.

Yeasts were isolated from two or more anatomical sites in 198 women attending genitourinary clinics on at least two occasions. The yeast biotypes isolated concurrently from the vagina and urethra were the same in 138 (99%) of 140 instances, and 94% of 124 concurrent genital and anal isolates were of matching types, whereas only 75% of concurrent genital and oral isolates were of the same type. Mixtures of Candida spp. or C. albicans biotypes were encountered only five times among 545 yeast-positive samples. In instances where Candida spp. were isolated at successive times from the same site in a patient, the same yeast type was encountered on 97 (87%) of 112 occasions when the interval between samples was less than 15 weeks, and on 19 (66%) of 29 occasions when the interval was 15 weeks or more. These data indicate a tendency to carriage of phenotypically consistent types of Candida among most women attending genitourinary clinics.

Reaction of Candida albicans cells of different morphology index with monoclonal antibodies specific for the hyphal form.

Two monoclonal antibodies (MAbs), 3D9 with reported specificity for Candida albicans hyphae, and 3B7 with reported specificity for morphological forms of C. albicans found in vivo, were tested by indirect immunofluorescence with C. albicans cells that were grown in 12 different environments (four different culture media incubated at various temperatures) and whose cellular morphology was estimated in terms of morphology index (Mi). Both MAbs reacted strongly with cells with Mi greater than 3.0, i.e., with pseudohyphal and hyphal forms, but in Eagle's medium at 26 degrees C and in a modified Sabouraud's broth medium at 30 degrees C, some reactivity was also found with cells of lower Mi (i.e., yeast forms). Therefore, it was concluded that the hyphal phenotype and the epitopes reactive with the MAbs were co-expressed but that the epitopes could also be expressed independently of the hyphal phenotype. The results confirm the propensity of C. albicans for variation of its surface antigenic composition.

Antifungal pulse therapy for onychomycosis. A pharmacokinetic and pharmacodynamic investigation of monthly cycles of 1-week pulse therapy with itraconazole.

BACKGROUND AND DESIGN: In the treatment of onychomycosis, oral therapies have generally been given as a continuous-dosing regimen. For example, the suggested dose of itraconazole for the treatment of onychomycosis has thus far been 200 mg/d for 3 months. Based on the advances in our understanding of the pharmacokinetics of itraconazole, we investigated the efficacy and nail kinetics of intermittent pulse-dosing therapy with oral itraconazole in patients who were suffering from onychomycosis. Fifty patients with confirmed onychomycosis of the toenails, predominantly Trichophyton rubrum, were recruited and randomly assigned to three (n = 25) or four (n = 25) pulses of 1-week itraconazole therapy (200 mg twice daily for each month). Clinical and mycological evaluation of the infected toenails, and determination of the drug levels in the distal nail ends of the fingernails and toenails, were performed at the end of each month up to month 6 and then every 2 months up to 1 year. RESULTS: In the three-pulse treatment group, the mean concentration of itraconazole in the distal ends of the toenails ranged from 67 (month 1) to 471 (month 6) ng/g, and in the distal ends of the fingernails, it ranged from 103 (month 1) to 424 (month 6) ng/g. At month 11, the drug was still present in the distal ends of the toenails at an average concentration of 186 ng/g. The highest individual concentrations of 1064 and 1166 ng/g were reached at month 6 for toenails and fingernails, respectively. At end-point follow-up, toenails in 84% of the patients were clinically cured with a negative potassium hydroxide preparation and culture in 72% and 80% of the patients, respectively. In the four-pulse treatment group, the mean concentration of itraconazole in the distal ends of the toenails ranged from 32 (month 1) to 623 (month 8) ng/g, and in the distal ends of the fingernails, it ranged from 42 (month 1) to 380 (month 6) ng/g. The highest individual concentrations of 1549 and 946 ng/g were reached at month 7 for toenails and at month 9 for fingernails, respectively. At month 12, the drug was still present in the distal ends of the toenails at an average concentration of 196 ng/g. At end-point follow-up, toenails in 76% of the patients were clinically cured with a negative potassium hydroxide preparation and culture in 72% and 80% of the patients, respectively. There were no significant intergroup differences between the three- and four-pulse treatment groups for the primary efficacy parameters. The drug was well tolerated with no significant side effects in either patient group. CONCLUSIONS: Following pulse therapy with itraconazole (400 mg/d given for 1 week each month for 3 to 4 months), the drug has been detected in the distal ends of nails after the first pulse, and it has reached therapeutic concentrations with further therapy. After stopping the last pulse, the drug remains in the nail plate at levels above 300 ng/g for several months. Clinical cure rates between 76% and 84% and negative mycological examination findings between 72% and 80%, respectively, were observed in toenail onychomycosis. The data suggest that pulse therapy with itraconazole is an effective and safe treatment option for onychomycosis.

Epidemiological shifts in opportunistic and nosocomial Candida infections: mycological aspects.

Increases in the prevalence of yeast species other than C. albicans as agents of disseminated mycoses in immunosuppressed patients and of fluconazole-resistant C. albicans isolates from patients infected with HIV indicate changes in the epidemiology of Candida infections. The precise reasons for alterations in prevalence of various agents are difficult to pinpoint but changes in the types of host populations at risk of Candida infection and selection of resistant yeast populations by widescale usage of certain antifungal agents seem to be factors involved in the process. Greater attention to speciation of clinical yeasts and standardized susceptibility test methods are needed for future epidemiological surveillance.

Resistance of clinically important yeasts to antifungal agents.

Resistance of yeasts to antifungal agents was a relatively minor clinical problem for many years. Recently Candida albicans isolates resistant to fluconazole have been reported with increasing frequency in the setting of oral infections in HIV-positive patients. Improved standardization of fluconazole susceptibility testing has resulted in demonstrable correlations between yeast resistance in vitro and in vivo for this agent in the AIDS setting. Known resistance mechanisms for azole antifungals include reduced access of the drug to the intracellular sterol demethylase target, probably because of the action of multidrug resistance efflux pumps, and overproduction of that target. Management and prevention of future resistance development requires greater vigilance for surveillance than has been the practice in the past.

Candida infections in AIDS patients.

PIP: In 1987, data from the Centers for Disease Control AIDS data base indicated a 50% prevalence of oropharyngeal Candida infection, a 10% rate of esophageal infection, and .5% rate of bronchopulmonary infection among AIDS patients. Candida-positive blood cultures were found in 13 of 903 AIDS patients, and disseminated Candida infection was ascertained in 11 of 101 post mortem examinations of AIDS victims. 5 of 12 patients with oral Candida infection progressed to AIDS within a 42-week investigation as opposed to only 1 of 17 patients without Candida. In the former group, CD4 counts and CD4/CD8 ratios were also significantly lower. Most infections were caused by Candida albicans. Genital Candida occurs in 5-20% of women in reproductive age. In a study of 66 HIV-infected women Candida vaginitis preceded oral Candida infections which preceded Candida esophagitis. 33 women had vaginal infection, 25 had oral Candida, and 9 had esophageal infection with reduced CD4 counts. Infections of the oropharynx and the vagina are reduced CD4 counts. Infections of the oropharynx and the vagina are treated with amphotericin B, nystatin, miconazole, and clotrimazole. Systemically effective compounds include ketoconazole, itraconazole, and fluconazole, although interactions with rifampicin, phenobarbital, and phenytoin used in HIV treatment occur. Fluconazole is contraindicated in C. glabrata and C. krusei infections as it selects for azole-resistant Candida strains. Iv amphotericin B and fluconazole are used in serious infections when oral treatment is ineffective.^ieng

Candida albicans strain types from the genitalia of patients with and without Candida infection.

The strain phenotypes of 266 C. albicans isolates from patients attending a genitourinary clinic were determined on the basis of 9 biochemical tests. Analysis of the strain patterns of isolates from the genitalia showed that there were no statistically significant differences between types associated with clinically overt Candida infection and types isolated in the absence of symptoms of candidosis. This finding accords with the traditional view of C. albicans as an opportunistic pathogen, rather than a species containing some strains of high virulence. In cases where isolations were made from the same patient at different times, or from different anatomical sites in the same patient, it was found that usually, but not always, a patient carried the same phenotype at different sites and different times. Similarly, the same strain type was isolated from the genitalis of both partners in a majority of instances where strains were isolated from consorts; however, this was not the case for a substantial minority of couples, particularly in those where high promiscuity appeared to promote considerable mixture and interchange of the C. albicans genital microflora.

Epidemiology of vaginal Candida infection: significance of numbers of vaginal yeasts and their biotypes.

The prevalence, quantity and biotypes of vaginal yeasts were determined for 1082 randomly selected nonpregnant patients attending two genitourinary medicine clinics. The overall yeast prevalence was 22.1% with a geometric mean of 40 and a median of 75 yeast colonies per positive isolate (on 5-cm Petri dishes). There was no statistically significant variation in prevalence, quantity or biotype of yeasts with the patients' age, season of the year, stage of the menstrual cycle, recent antibiotic history, contraceptive use or main diagnosis (excluding candidosis). A clinical score for Candida infection, based on symptoms of pruritus and signs of Candida vulvovaginitis, showed significant variation with the prevalence and quantity of yeast isolates. Unequivocal clinical evidence of candidosis was strongly associated with high concentrations of vaginal yeasts. There was also some association between certain groups of Candida albicans biotypes and the clinical score. These observations reemphasize the need for consideration of both clinical and mycological factors in establishing a diagnosis of vulvovaginal Candida infection. It is suggested that isolation of fewer than 10 yeast colonies from a vaginal swab is usually unlikely to indicate an infection requiring treatment.