Despite advances in the treatment of fungal infections, vulvovaginal candidiasis (VVC) remains a common problem among women of childbearing age worldwide (Sustr et al. 2020). This disease affects women to varying degrees, regardless of ethnic origin, and social or living conditions. This ailment can be induced by various factors ranging from host factors such as age, estrogen levels, sexual activity, exposure to stress, allergic diseases, or medications (Farr et al. 2021). However, most cases do not have an identifiable trigger. It is estimated that 70–75% of women will be diagnosed with vulvovaginal candidiasis at least once in their lifetime (Hedayati et al. 2015), and 5–8% of adult women have recurrent vulvovaginal candidiasis (RVVC), which is defined as four or more episodes per year (Sobel 1992). VVC affects approximately 138 million women annually (in the range of 103 to 172 million), with a global annual prevalence of 3,871 per 100,000 women; 372 million women are affected by RVVC during their lifetime (Denning et al. 2018). More than 40 species of the
The taxonomy of some
The available clinical and epidemiological data show that
The including criteria were as follows:
Two (2) vaginal swabs were taken from the posterior vaginal vault, using a sterile speculum. The first vaginal swab was used for Gram staining with PREVI® Color Gram stainer (bioMérieux, France).
The second swab was used to perform cultures of aerobic bacteria (TSA with 5% sheep blood; GRASO, Poland),
Vulvovaginal candidiasis (VVC) was diagnosed by positive
Initial species identification of strains was carried out based on the appearance of the colonial morphology on chromogenic medium (chromID®, bioMérieux, France). As a control of chromogenic agar, the reference strains were included:
Afterward, further identification was performed by an automatic biochemical method using the YST Card, VITEK®2 compact (bioMérieux, France) according to the manufacturer’s instruction. The results were approved when the acceptable score was generated using the database system, covering 27
The species identification of
The PCR conditions were as follows: initial denaturation at 94°C for 2 min, 30 cycles of denaturation at 94°C for 15 s, annealing at 63°C for 30 s, and extension at 72°C for 1 min, followed by a final extension at 72°C for 10 min, in a T-Personal Thermal Cycler (Biometra, Germany). The reaction mixture of the final volume 25 μl contained: 5 μl of GoTaq® Flexi Buffer, 0.5 μl of PCR Nucleotide Mix (10 mM each) (Promega, USA), 1 μl of each primer (Genomed, Poland), 2 μl of genomic DNA, 2 μl of MgCl2 (25 mM), 0.125 μl of GoTaq® DNA Polymerase (5 u/μl) (Promega, USA) and Nuclease-Free Water (Promega, USA). The PCR products of the size of 941 bp, 569 bp, and 700 bp for
The PCR amplicons were separated by 2% agarose gel electrophoresis and visualized with ethidium bromide staining (Sigma-Aldrich Chemie, Germany).
Reference strains included
According to the manufacturer’s recommendation of Fungitest™ (BioRad, France), we use the following criteria of drug susceptibility: susceptible (S), intermediate (I) (referring to “susceptible, increased exposure” according to the EUCAST recommendations (EUCAST 2020), and resistant (R).
Prevalence of single VVC and co-occurrence with bacteria.
Type of co-occurrence | No. (%) |
---|---|
Single VVC | 197 (60.43) |
VVC + bacteria: | 129 (39.57) |
|
60 (18.40) |
|
41 (12.58) |
|
11 (3.37) |
|
9 (2.76) |
|
3 (0.92) |
|
2 (0.61) |
Methicillin susceptible |
1 (0.31) |
|
1 (0.31) |
|
1 (0.31) |
VVC – vulvovaginal candidiasis
Molecular discrimination among
Species | No. (%) n = 326 |
---|---|
307 (94.17) | |
17 (5.21) | |
2 (0.61) |
The reference method for this study found as “a gold standard”, was molecular identification based on the
The VITEK®2 system (bioMérieux, France) showed agreement with the molecular assay in 97.85% of clinical strains. Most
Percentage of compatible results obtained by the methods used to discriminate species among
The compliance with PCR assay |
||
---|---|---|
VITEK 2 | 319 (97.85) | 7 (2.15) |
MALDI-TOF MS | 323 (99.08) | 3 (0.92) |
Discrepancies between species identification among
Isolate No. | VITEK 2 | MALDI-TOF MS | PCR assay based on the |
||||
---|---|---|---|---|---|---|---|
Species identification | Confidence level (probability) | Species identification | Confidence level | Species identification | Product size (bp) | ||
CA67 | clinical strain | low (< 85%) | 99.90% | 569 | |||
CA95 | clinical strain | excellent (96%) | 99.90% | 941 + 569 | |||
CA98 | clinical strain | not identified | – | 99.90% | 569 | ||
CA125 | clinical strain | acceptable (86%) | 99.90% | 569 | |||
CA13 | clinical strain | low (< 85%) | 99.90% | 569 | |||
CA137 | clinical strain | good (91%) | 99.90% | 569 | |||
CA313 | clinical strain | very good |
99.90% | 941 + 569 | |||
reference strain | excellent (98%) | 99.90% | 941 | ||||
reference strain | very good |
99.90% | 569 | ||||
reference strain | low (< 85%) | 99.90% | 700 |
The protein analysis by MALDI-TOF MS (VITEK®2 MS Biotyper; bioMérieux, France) did not require lengthy biochemical reactions. It was more accurate method than the biochemical analysis with the VITEK®2 system. Based on MALDI-TOF MS, 99.08% of clinical isolates were correctly identified as
In total, 100% of
Vulvovaginal candidiasis (VVC) remains a threatening problem among women (Sustr et al. 2020). All 326
One of the most commonly utilized, rapid, and cost-effective tests are chromogenic media, which are frequently used to detect and identify yeasts of the
The other method commonly used in routine laboratory practice for yeast identification is the VITEK®2 system. This automated system is based on kinetic analysis detecting metabolic changes and continuous colorimetric-based monitoring of reactions which provides reliable identification of 27 medically relevant species of
Due to the limited ability of phenotypic approaches, such as chromogenic media and the VITEK®2 system, to identify rare species and to confirm the identification of
According to the performed molecular assay based on the
The meta-analysis provided by Ghareghbolagh et al. (2020), showed that the distribution of
Epidemiological data indicated that the incidence of
Multiple global studies have shown that
According to the Centers for Disease Control and Prevention recommendations (CDC), treatment of uncomplicated VVC (which constitutes the majority of VVC) is based on empirical therapy. Culture and antifungals susceptibility testing should be considered only for patients who remain symptomatic after failure of the empirical treatment (Workowski et al. 2021). However, according to the demands reported by the physicians, susceptibility tests are performed. Currently, there are few commercial tests available that can be used to assess the susceptibility testing of drugs applicable in the treatment of VVC. Fungitest™ (BioRad, France) was the only method used to test antifungal susceptibility in this study and is still routinely used in many microbiological laboratories in Poland. However, the concentrations of drugs available in this kit do not meet the current European (EUCAST 2020) and American criteria (CLSI 2022). According to the manufacturer’s instructions, the interpretation of drug susceptibility is based only on scientific reports. Moreover, this semi-quantitative method has many advantages: it does not require complex handling and is cost-effective. Furthermore, the agreement with the European Committee on Antibiotic Susceptibility Testing (EUCAST) data for Fungitest™ was high, with correlation indices of 0.95 (p < 0.01) (Cuenca-Estrella et al. 2005).
The global prevalence of
A PCR assay targeting the