PMC9537450_01
(A) Growth on Sabouraud Dextrose Agar after incubation at 30 C for 48 h in aerobic conditions. (B,C) Growth on CHROMagar  Candida after incubation at 35 C for 48 h in aerobic conditions. (D,E) Growth on CHROMagar Candida Plus after incubation at 35 C for 48 h in aerobic conditions.
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fig1
No bacterial growth was detected, and fungal colonies with the same morphology and characteristics were observed from all clinical samples (Figure 1A). SDA was subcultured onto CHROMagar Candida (BD Difco) and CHROMagar Candida Plus (CHROMagar ) and colonies were interpreted according to the manufacturer's instructions. Small pink colonies, suggesting C. guilliermondii, were observed in both media (Figures 1B-E) from all clinical samples.
Neighbor-Joining tree based on Pearson correlation constructed with MALDI-TOF MS spectra of the clinical isolate and of reference strains of Candida albicans (ATCC 18804), Candida glabrata (ATCC 2001), Candida guilliermondii (ATCC 7350), Candida inconspicua (ATCC 16783), Candida kefyr (ATCC 4135), Candida krusei (ATCC 6258), Candida lipolytica (ATCC 18942), Candida metapsilosis (ATCC 96143), Candida orthopsilosis (ATCC 96141), Candida parapsilosis (ATCC 22019), and Candida rugosa (ATCC 10571).
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fig2
The clinical isolate was clustered together with C. guilliermondii reference strain (Figure 2) confirming the automated identification by MALDI-TOF MS.
Phylogenetic relationship between the clinical isolate and reference strains of Candida species inferred from ITS sequences by Neighbor-Joining method. The optimal tree is shown. The percentages of replicate trees in which the associated taxa clustered together in the bootstrap test (1,000 replicates) are shown next to the branches. Branches corresponding to partitions reproduced in less than 50% bootstrap replicates are collapsed. The evolutionary distances were computed using the Maximum Composite Likelihood method and are in the units of the number of base substitutions per site. This analysis involved 11 nucleotide sequences. There were a total of 78 positions in the final dataset. Evolutionary analyses were conducted in MEGA X.
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fig3
Aligner (Genes Code Corporation, Ann Arbor, United States), and phylogenetic analysis was performed using Blast software for comparison with the sequences deposited in the NCBI/GenBank database. An identity of 100% of the clinical isolate when compared to the reference sequence of C. guilliermondii (ATCC6260) KU729068.1 deposited at NCBI/GenBank was observed (Figure 3).