Comparison of standard phenotypic assays with a PCR method to discriminate Candida albicans and C. dubliniensis

In 1995, Candida dubliniensis was described as a new species in the genus Candida. Its close relationship to C. albicans has proved problematic in the identification of C. dubliniensis in clinical specimens. The objective of this study was to determine if reproducible differentiation between both species can be obtained by phenotypic assays. Therefore, 100 strains from 86 patients with the ability to produce chlamydospores were examined with different methods including API ID 32 C, colour development on CHROMagar, chlamydospore formation on Staib agar, growth at different temperatures and germ tube formation at 39 degrees C. Additionally, polymerase chain reaction (PCR) was used as gold standard. Six of the investigated strains were C. dubliniensis. The results suggest that there is still no single phenotypic method satisfactory to distinguish between C. albicans and C. dubliniensis.     

Chlamydosporulation of Candida albicans and Candida dubliniensis on mustard agar

Chlamydospores are distinctive morphological forms characteristic of Candida albicans, a phenomenon shared only with the closely related species Candida dubliniensis. The production of chlamydospores has remained an important diagnostic criterion for the differentiation of C. albicans and C. dubliniensis from other yeast. We herein describe a new medium, mustard agar, for chlamydosporulation in Candida. All the strains of C. dubliniensis and C. albicans tested produced chlamydospores on mustard agar, whereas none of the other five species produced chlamydospores.     

Liquid growth conditions for abundant chlamydospore formation in Candida dubliniensis

Staib liquid medium (syn. Guizotia abyssinica creatinine medium) was evaluated for the induction of chlamydospores in the yeast species Candida dubliniensis. During growth in this liquid medium C. dubliniensis produced abundant chlamydospores at the tips of pseudohyphal elements, whereas C. albicans produced only round to oval blastospores. Chlamydospore formation could be induced with the same efficiency by use of a liquid growth medium in which Helianthus annuus (sunflower) seeds, another plant of the family Compositae, were substituted for Guizotia abyssinica plant seeds. Liquid growth culture conditions for rapid and abundant formation of chlamydospores in C. dubliniensis should facilitate their enrichment and their biochemical and genetic analysis.     

Effects of oestrogen on vulvovaginal candidosis

As a frequently occurring infectious disease mainly caused by Candida albicans, vulvovaginal candidosis (VVC) affects more than 100 million women worldwide every year. Multiple factors that influence C. albicans colonisation have been linked to the incidence of VVC, including high levels of circulating oestrogen due to pregnancy, the use of oral contraceptives, and hormone replacement therapy. This review provides an overview of the current understanding of the mechanism(s) by which oestrogen contributes to VVC, which might provide meaningful guidance to the prevention and treatment of this disease.     

Prevalence of Candida dubliniensis among germ tube-positive Candida isolates in a maternity hospital in Kuwait

In this study, 1644 germ tube-positive Candida isolates from a maternity hospital was prospectively examined for the prevalence of Candida dubliniensis. Candida species were isolated from different clinical specimens, but majority (>90%) of them came from high vaginal swabs and urine specimens. The phenotypic and molecular identification methods for C. dubliniensis included production of rough colonies and chlamydospores on simplified sunflower seed agar, determination of assimilation profile by Vitek 2 yeast identification system, specific amplification of rDNA of internally transcribed spacer (ITS)-2 region by semi-nested PCR and direct DNA sequencing of the ITS-1 and ITS-2 regions. Three germ tube-positive Candida isolates were identified as C. dubliniensis with an overall prevalence of 0.2%. Of these, two came from urine specimens and one from a vaginal swab. None of the C. dubliniensis isolates showed resistance against fluconazole, voriconazole and amphotericin B. The study reinforces the usefulness of sunflower seed agar in presumptive identification of C. dubliniensis and confirms the prevailing view that this species forms only a minor constituent of Candida species occurring in vagina or other anatomic sites of non-HIV/AIDS-infected individuals.     

Guideline vulvovaginal candidosis: guideline of the German Dermatological Society, the German Speaking Mycological Society and the Working Group for Infections and Infectimmunology of the German Society for Gynecology and Obstetrics

The Guideline vulvovaginal candidosis, as passed by three German medical societies, is presented.     

Use of Dermasilk briefs in recurrent vulvovaginal candidosis: safety and effectiveness

Despite the generally excellent results achieved with fluconazole 150 mg weekly in recurrent vulvovaginal candidosis (RVVC), some patients with a long history of disease do not achieve complete resolution of symptoms following antimycotic treatment. It is thought that use of tight synthetic fabric underwear could be a significant factor in causing recurrence. We decided to compare underwear made of Dermasilk^®, a pure fibroin fabric impregnated with a permanent antimicrobial protection, with a cotton placebo to see whether it could be a useful adjunctive tool in the management of RVVC. We recruited 96 women who had a long‐term history of RVVC and had not responded to oral antimycotics with complete satisfaction. The patients were randomly divided into two groups and instructed to use either white cotton placebo briefs or Dermasilk^® briefs. Both groups were treated with fluconazole 150 mg once weekly for 6 months. After 6 months, the Dermasilk group showed a statistically significant greater decrease of itching, burning, erythema and a smaller number of recurrences than the cotton group. Our work suggests that Dermasilk^® briefs could be a useful adjunctive tool in addition to antimycotic treatment to help relieve the discomfort of recurrent vulvovaginitis.     

Experimental systemic virulence of oral Candida dubliniensis isolates in comparison with Candida albicans,Candida tropicalis and Candida krusei

There are no previous studies on the comparative virulence of Candida dubliniensis with other non‐albicans species. The aim of this study was to compare the virulence and infection kinetics of C. dubliniensis and other species. Candida albicans, C. dubliniensis, Candida tropicalis and Candida krusei (reference strains) were inoculated intravenously in mice. For infection kinetics evaluation, a group of five animals were sacrificed after 6 h, 3, 7, 14 and 21 days. Microbiological evaluations (liver, spleen, kidneys, lungs and brain) and histopathological examination of the kidney were performed. The results of virulence evaluation were analysed using Kaplan–Meier survival analysis (5%). Candida dubliniensis‐inoculated mice survived for longer periods compared with those with C. albicans (P = 0.005). No differences were detected in relation to C. tropicalis (P = 0.326) and C. krusei (P = 0.317). Most of the organs were persistently colonised by C. albicans and C. dubliniensis even by day 21. Tendency of C. krusei clearance was observed in all organs. Fungal masses and renal lesions were observed after inoculation of C. albicans, C. dubliniensis and C. tropicalis. Within the limits of the study, data on survival rate and dissemination capacity suggest that C. dubliniensis is less virulent than C. albicans.     

Candida carriage and Candida dubliniensis in oropharyngeal samples of type-1 diabetes mellitus patients

The carriage of Candida dubliniensis in the oral cavities of type-1 diabetic patients were investigated. Of 230 patients 81 (35%) had Candida spp. in their oral cavity; C. albicans was the most frequently isolated species (58%). No C. dubliniensis was found in the study population.     

Stress as a cause of chronic recurrent vulvovaginal candidosis and the effectiveness of the conventional antimycotic therapy

Chronic Candida vulvovaginitis cannot, at present, be treated effectively with local or oral antimycotic medication. Göttlicher & Meyer [Vulvovaginalmykose. Klinische Ergebnisse einer epidemiologischen Langzeitstudie, Thieme, Stuttgart (1998); Mycoses 41 (1998) 49] formulated a hypothesis to explain this fact. They say that stress is the main cause of vulvovaginal mycosis, and that frequently cited predisposing somatic risk factors are not causing the illness. Two research projects were carried out to test this hypothesis. The first project was conceived to provide direct evidence for two empirical statements derived from the Göttlicher–Meyer hypothesis: (i) stress as a predictor of vulvovaginitis is least valid as the commonly assumed somatic factors and (ii) combinations of factors that reliably discriminate index women from symptom‐free women point to aspects of psychosocial development, particularly stress, and not to somatic factors. Between March and November 1999, 309 successive patients were randomly chosen from those successively treated in one in‐patient and two out‐patient settings. Of those chosen, 117 had had at least one episode of vaginal candidosis within the 2 years prior to enlistment in the study. The remaining 192 patients had experienced no such illness. For the 117 index patients both of the above empirical statements were confirmed by statistical analyses. The second project was designed to test predictions concerning the ineffectiveness of traditional antimycotic treatment derived from the Göttlicher–Meyer hypothesis. Three different empirical statements were derived from the hypothesis – each based on the assumption of a Poisson distribution of relapses that remains uninfluenced by treatment attempts. Each of the statements was tested and confirmed in independent samples – the first in the sample used in project one, the second in a sample of 206 women insured by public Healths Management Organizations (HMOs) and the third in a sample of 179 women insured by private HMOs. Each patient had evidenced Candida albicans vulvovaginitis at least once in the time interval between 1996 and 2000. Statistical analyses confirmed each of the empirical statements, thereby substantiating the Göttlicher–Meyer hypothesis. The authors conclude that (i) psychosocial factors, particularly stress, are the primary causes of Candida albicans vulvovaginitis. Accompanying somatic factors are of little statistical significance in explaining occurrence and relapse. (ii) Traditional antimycotic treatment influences only the symptoms of the illness, not its causes, and has no effect on the probability of a relapse. Increased treatment effectiveness can be achieved only at the cost of a complete re‐orientation concerning the causes of vaginal mycosis. Interventions designed to strengthen the patients’ immune response are among the most promising.     

Itraconazole in the treatment of acute and recurrent vulvovaginal candidosis: comparison of a 1-day and a 3-day regimen

Itraconazole is a systemic triazole with a broad-spectrum antifungal effect which belongs to a standard choice for the treatment of vulvovaginal candidosis. Thirty-four patients with acute and 26 patients with recurrent form of culture documented attacks of vulvovaginal candidosis were evaluated. There was frequent discrepancy between severe symptoms and weaker signs in patients with recurrent form than in the group with acute sporadic form. Eighteen patients with the acute form and 13 patients with the recurrent form of vulvovaginal candidosis received 1-day therapy (twice 200 mg of itraconazole in 1 day). Sixteen patients with the acute form and 13 patients with the recurrent form received 3-day therapy (200 mg of itraconazole daily for three following days). The cure rates 1 month after treatment were 97.1 and 76.9% in patients with the acute sporadic and recurrent form of vulvovaginal candidosis, respectively. Positive cultures in the latter group documented relapses. These clinical failures were not related to the resistance of yeasts to itraconazole. With the exception of one case of alopecia areata, side effects were minimal. The application of the long-term regimens and suppressive therapy with azole drugs is suggested in patients with recurrent vulvovaginal candidosis.     

Epidemiology and antifungal susceptibility patterns of Candida isolates from Greek women with vulvovaginal candidiasis

Vulvovaginal candidiasis (VVC) is a common infection of the genital tract affecting millions of women worldwide. Data on epidemiological trends of VVC in Greece are scarce. This study was undertaken to evaluate the prevalence of VVC among symptomatic women in Crete, Greece, identify the Candida species involved and determine their susceptibility to antifungals. Over a 6‐year period (2012‐2017), 10 256 symptomatic women with vaginitis were evaluated. Isolation of yeasts was performed on Sabouraud dextrose agar with chloramphenicol, and the isolates were identified using the API 20 C AUX and/or the Vitek 2 YST card. Susceptibility of the isolates to amphotericin, fluconazole, voriconazole and flucytosine was determined by the Vitek 2 automated system. The results were interpreted according to Clinical and Laboratory Standards criteria. Vaginal swab cultures of 1217 (11.9%) women yielded Candida species. Recurrent VVC was documented in 62 (5.1%) of them. Candida albicans was the most frequently isolated species (75.6%), followed by Candida glabrata (13.6%). Overall, resistance rates to amphotericin B, fluconazole, voriconazole and flucytosine were 0.2%, 6.6%, 1.4% and 2.1%, respectively. Fluconazole resistance of C. albicans significantly increased in the second period of the study (2015‐2017) (P = 0.031). This study demonstrated that VVC is a common infection among women in our region, with C. albicans being the predominant species involved. Although resistance to antifungals was infrequent, resistance to fluconazole among C. albicans isolates was found to significantly increase with time. Continued surveillance of changes in species distribution and susceptibility to antifungals are necessary to guide treatment.     

Evaluation of Mueller-Hinton-agar as a simple medium for the germ tube production of Candida albicans and Candida dubliniensis

Candida albicans is the most frequently isolated yeast species from clinical specimens. A classical rapid presumptive differentiation from non- albicans species is based on its ability to produce germ tubes after incubation in human serum. The only non- albicans Candida species producing germ tubes is Candida dubliniensis. In this study, we evaluated Mueller-Hinton-agar (MH-agar) as a medium for germ tube formation of C. albicans and C. dubliniensis . A total of 859 yeast isolates from stool samples, including 632 strains of C. albicans , 10 C. dubliniensis and 217 other yeast strains from 20 different species, were grown on Sabouraud glucose (2%) agar at 37 °C for 24–72 h. Species were identified by standard methods. For the germ tube test (GTT), an inoculum from a single colony was streaked onto a MH-agar plate and covered by a sterile coverslip. After incubation at 37 °C for 2 h, the MH plates were examined using a light microscope at ×200. The GTT was positive in 578 of 632 C. albicans strains (sensitivity 91.5%), in six of 10 C. dubliniensis strains (sensitivity 60.0%), and in none of the other yeast strains. MH-agar is a suitable medium for the GTT and the presumptive identification of C. albicans . It is safer to use than human serum and is widely available in microbiology laboratories.     

The involvement of physico-chemical interactions in the adhesion of Candida albicans and Candida dubliniensis to epithelial cells

Candida albicans and Candida dubliniensis are two pathogenic yeasts particularly hazardous to immunocompromised patients. Adhesion of yeast cells to epithelium is considered one of the virulence factors and its study is of major importance. The main aim of this study was the comparison of the influence of physico-chemical properties on the adhesion of C. albicans and C. dubliniensis to epithelium. Two strains of each Candida species were used in the adhesion assays to HeLa cells. Adhered cells were enumerated by direct microscopic images observation. Yeast cell surface tension parameters and degree of hydrophobicity were determined by contact angle measurement. Pseudohyphae and hyphae formation was analysed by scanning electron microscopy. Yeast cells presented no statistical differences concerning their physico-chemical surface properties. However, the extent of adhesion to epithelium was different among the four strains. As general conclusion, yeast adhesion to epithelium seems to be strain-dependent and not directly correlated with pseudohyphae formation.     

Epidemiological survey of vulvovaginal candidosis in Sfax, Tunisia

Vulvovaginal candidosis (VVC) is a common infection of the female genital tract affecting 75% women at least once in their lifetime. The aim of this study was to determine the incidence and potential risk factors associated with VVC and recurrent vulvovaginal candidosis (RVVC). A prospective study of women with vaginitis symptoms was conducted over 2 years in the regional clinic of population and family education in Sfax. A discriminant analysis was used to evaluate the association between the incidence of Candida vaginitis and potential risk factors. Sporadic and recurrent VVC were documented respectively in 48% and 6.1%. The most frequent factors associated with positive Candida culture were employed women, uncontrolled diabetes, history of genital infection and intrauterine device contraception. Increased episode numbers of VVC and condom/spermicidal contraception were positively associated with recurrences. Candida albicans was the predominantly isolated species (76.3%) followed by Candida glabrata (19.3%). Infection with C. glabrata occurred in 34% and 17.5% of patients with RVVC and VVC respectively. The discriminant investigation had provided further insights into the basis for prevention and control of RVVC. Increased prevalence of C. glabrata in patients with RVVC and observed risk factors should be taken into consideration to achieve success in the management of this infection.     

Prevalence of Candida dubliniensis among germ tube-positive yeasts recovered from the respiratory specimens in HIV-negative patients

The present investigation was conducted to identify Candida dubliniensis, from respiratory specimens, recovered from HIV-negative patients. Over a 7-month period, 75 germ tube and chlamydospore-positive yeasts were screened for C. dubliniensis, using a variety of phenotypic characteristics. Their identification was based on sugar assimilation reactions using API 20 C Aux. A total of seven (9%) isolates recovered from sputum, bronchial lavage and nasopharyngeal aspirate were identified as C. dubliniensis. All the isolates were susceptible to amphotericin B. One isolate each showed resistance to fluconazole and ketoconazole, and two were resistant to itraconazole. A significantly high percentage (43%) of C. dubliniensis showed resistance to flucytosine.     

Prevalence of Candida dubliniensis among cancer patients in Kuwait: a 5‐year retrospective study

Despite close genetic and phenotypic relationship of Candida dubliniensis with Candida albicans, its role in human disease is mostly restricted to oral colonisation, particularly among HIV‐infected patients. The prevalence of C. dubliniensis in association with other disease conditions has been infrequently reported. In this study, we present data on the prevalence of C. dubliniensis among yeast species isolated from cancer patients over a 5‐year period. A total of 1445 yeast isolates recovered from respiratory specimens, blood, urine and oral swabs were analysed. Candida dubliniensis isolates were provisionally identified by phenotypic methods and their identity was further confirmed by species‐specific amplification and/or sequencing of internally transcribed spacer region of rDNA. Antifungal susceptibility for fluconazole was determined by Etest. The number of isolates identified as C. dubliniensis, C. albicans and other yeast species were 71 (4.9%), 862 (59.6%) and 512 (35%) respectively. All the C. dubliniensis isolates originated from respiratory (5.9%) or oral (3.2%) specimens with an overall prevalence of 4.9%, and were found to be susceptible to fluconazole. The isolation of C. dubliniensis from respiratory or oral specimens and not from blood or urine specimens suggests that this species has preference to colonise these sites of human body.     

The serum levels of calcium, magnesium, iron and zinc in patients with recurrent vulvovaginal candidosis during attack, remission and in healthy controls

The real cause of recurrent vulvovaginal candidosis (RVVC) is concealed and the etiopathogenesis of this disease remains to be determined. In a cohort study, concentrations of metals in 44 patients with RVVC and 30 healthy age-matched women were measured and compared. The concentrations of serum calcium (Ca), magnesium (Mg) and iron (Fe) were measured photometrically, the zinc (Zn) levels were determined using flame atomic absorption spectrometry. For statistical analysis were used the Student's t-tests (paired analysis for attack vs. remission; non-paired analysis for patient vs. control). Although all measured metals were within normal ranges the patients with RVVC had in contrast to the healthy controls significantly lower levels of serum Ca, Mg and Zn and insignificantly higher levels of Fe. These relative changes may contribute to the development of attacks in patients with RVVC.