First case of superficial infection due to Naganishia albida (formerly Cryptococcus albidus) in Iran: A review of the literature

Background and Purpose: Naganishia albida (formerly Cryptococcus albidus) is a non-neoformans cryptococcal species rarely isolated as a human pathogen. Case report: Herein, we present the case of a 26-year-old Iranian man with a superficial cutaneous lesion in the axilla. The initial treatment for pityriasis versicolor by clotrimazole was unsuccessful. We performed skin sampling based on the standard protocol and conducted further investigations by the conventional laboratory tests and molecular analysis of the skin samples. All the mentioned analyses revealed N. albida as the causative agent of infection. The minimum inhibitory concentration (MIC) analysis was carried out for the isolated agent, and the patient was treated using 100 mg daily of oral itraconazole. Conclusion: N. albida can be the causative agent of some superficial infections. This is the first report on the successful detection and treatment of a superficial skin infection due to N. albida by oral itraconazole.

Introduction ryptococcus spp. are basidiomycetous yeasts considered as the responsible agents for a wide range of diseases, among which C. neoformans and C. gattii are mentioned as the most common pathogenic species [1][2][3]. However, the incidence of infection due to non-neoformans cryptococcal species such as Papiliotrema laurentii (formerly C. laurentii) and Naganishia albida has increased recently [4,5]. N. albida is an encapsulated yeast occasionally detected on human skin, air, and soil [6,7].
Some cases of infection caused by N. albida such as keratitis, pneumonia, encephalitis, and cutaneous and disseminated cryptococcosis have been reported [8][9][10]. The treatment regimen for systemic cryptococcosis is amphotericin B in combination with flucytosine followed by fluconazole as consolidation therapy [8,11,12]. To date, there is no defined treatment for superficial cryptococcal infection. In the current study, we presented the first case of superficial cutaneous infection caused by N. albida, as well as its successful treatment with 100 mg daily of oral itraconazole. To the best of our knowledge, this is the first report on superficial cutaneous infection due to N. albida. A 26-year-old Iranian man without any underlying diseases was referred to Razi Hospital, a referral center for skin diseases in Iran that is affiliated to Tehran University of Medical Sciences, with a hyperpigmented patch in the axilla. Based on clinical examination, the diagnosis was made as pityriasis versicolor. However, treatment with clotrimazole cream was not successful and he returned to the hospital after one month.

Case report
Direct microscopic examination of the scales after mixing with 10% potassium hydroxide revealed the presence of yeast cells. Furthermore, the scales cultured on Dixon Agar (Quelab, Canada) plates for five days at 32°C produced white and creamy colonies with smooth surfaces. By Lactophenol Cotton Blue Staining of the smears, yeast cells similar to Cryptococcus were observed under microscope ( Figure 1). Genomic DNA was extracted from culture using glass beads method [13], and polymerase chain reaction (PCR) was performed using universal primers [14] of ITS1 (5'-TCC GTA GGT GAA CCT GCG G-3') and ITS4 (5'-TCC TCC GCT TAT TGA TAT GC-3') (Sinaclon, Iran). The PCR product with the approximate size of 600 bp was applied for sequencing (Macrogen, South Korea). Alignment of the obtained sequence in BLAST revealed high homology (99%) with N. albida, which is indicated with GenBank ID: MG020697.1. In the following step, the minimum inhibitory concentration (MIC) values for itraconazole, voriconazole, and amphotericin B were determined using the microbroth dilution method according to the Clinical Laboratory Standards Institute (CLSI) standard protocols [15]. Briefly, the test was performed in 96-well round-bottom microtiter plates. Drug concentration ranges were 0.03 to 16 µg/ml for itraconazole, voriconazole, and amphotericin B. Yeast suspensions were prepared in RPMI-1640 medium and adjusted to provide a final inoculum concentration of about 0.5 × 10 3 to 2.5 × 10 3 cells/ml. The culture plates were then incubated at 35°C followed by reading after 48 h according to the M27-S3 supplement of the CLSI guideline [15]. The MIC results were then compared with a drug-free control culture plate.
The MIC values for itraconazole, voriconazole, and amphotericin B were 0.062 µg/ml, 0.062 µg/ml, and 0.062 µg/ml, respectively, revealing the sensitivity of the mentioned causative agent. The patient was finally treated successfully using 100 mg daily of oral itraconazole. The Ethics Committee of Tehran University of Medical Sciences approved this report with the code No. IR.TUMS.SPH.REC.1396.2400.

Discussion
N. albida is a non-neoformans species of the genus Cryptococcus with a similar morphology to C. neoformans. However, they can be differentiated by their reaction to biochemical tests such as phenol oxidase and color changes in Bird Seed Agar medium [9,[16][17][18][19]]. In addition, N. albida is an opportunistic and encapsulated yeast found on human skin [6,7]. Although N. albida rarely causes any diseases, there have been some case reports, including cases of meningitis, peritonitis, fungaemia, pulmonary and cutaneous infections, and keratitis (Table 1).
Moreover, cases of encephalitis, disseminated cryptococcosis, and pneumonia have been reported [4,8,[20][21][22][23][24][25][26][27][28][29]. However, N. albida has never been reported as the causative agent of a superficial skin infection ( Table 1). As can be observed in this table, the majority of reported cases were from the USA, whereas there was only one record from other countries such as Turkey. Besides, the most reported cases were isolated from blood (fungemia) and the most effective treatment protocol was with amphotericin B.
To the best of our knowledge, the case described here is the first report of a pityriasis versicolor-like superficial infection due to N. albida. In addition, treatment choices are limited for cryptococcal infections. The first choice of treatment for these infections is the combination of amphotericin B and flucytosine [11,12]. Nonetheless, the treatment for infections due to N. albida is not well-defined, and amphotericin B has been mentioned to have limited efficacy in the treatment of N. albida [9]. N. albida shows various responses to different antifungal treatments. Therefore, there is no common treatment protocol for infections caused by this fungus.  Recently, it has been indicated that N. albida is only sensitive to amphotericin B and itraconazole [4,30,31]. Other reports revealed side effects due to the conventional amphotericin B (1 mg/kg/day) therapy against N. albida, which led the therapy to change to itraconazole [32]. Besides, successful treatment with fluconazole has been reported in patients with cutaneous infections caused by N. albida using a mean treatment period of 56 days [16].

Curr
In the present case, after MIC approval, we chose 100 mg daily of itraconazole in a 10-day treatment period, which resulted in successful treatment. The three-month follow up of the patient revealed no relapse of the infection and no evidence of any clinical manifestations at the involved site. Furthermore, several factors such as the anatomical location of involvement, the power of immune system in host, and tissue damages can determine the proper medication and the required treatment duration for cryptococcal infections due to non-neoformans species [4,5,17,33].

Conclusion
In conclusion, N. albida was found to have the ability to cause superficial infections. The case presented here is the first report of successful detection and treatment of infection by this yeast. N. albida was