Highly Conserved gsc1 Gene of Pneumocystis jirovecii in Patients with or without Prior Exposure to Echinocandins

Abstract

Echinocandins are noncompetitive inhibitors of the GSC1 subunit of the enzymatic complex involved in synthesis of 1,3-beta-d-glucan, a cell wall component of most fungi, including Pneumocystis spp. Echinocandins are widely used for treating systemic candidiasis and rarely used for treating Pneumocystis pneumonia. Consequently, data on P. jirovecii gsc1 gene diversity are still scarce compared to that for the homologous fks1 gene of Candida spp. In this study, we analyzed P. jirovecii gsc1 gene diversity and the putative selection pressure of echinocandins on P. jirovecii. gsc1 gene sequences of P. jirovecii specimens from two patient groups were compared. One group of 27 patients had prior exposure to echinocandins, whereas the second group of 24 patients did not, at the time of P. jirovecii infection diagnoses. Two portions of the P. jirovecii gsc1 gene, HS1 and HS2, homologous to hot spots described in Candida spp., were sequenced. Three single-nucleotide polymorphisms (SNPs) at positions 2204, 2243, and 2303 close to the HS1 region and another SNP at position 4540 more distant from the HS2 region were identified. These SNPs represent synonymous mutations. Three gsc1 HS1 alleles, A, B, and C, and two gsc1 HS2 alleles, a and b, and four haplotypes, Ca, Cb, Aa, and Ba, were defined, without significant difference in haplotype distribution in both patient groups (P = 0.57). Considering the identical diversity of P. jirovecii gsc1 gene and the detection of synonymous mutations in both patient groups, no selection pressure of echinocandins among P. jirovecii microorganisms can be pointed out so far.

Keywords: 1,3-beta-d-glucan; Pneumocystis jirovecii; echinocandins; genomic diversity; gsc1; hot spots.

FIG 1

Single-nucleotide polymorphisms in Pneumocystis jirovecii hot spot 1 (HS1) and HS2 DNA regions after sequencing. Analysis of 51 P. jirovecii specimens led to the identification of three SNPs close to the HS1 region at positions 2204, 2243, and 2303 and one SNP more distant from the HS2 region at position 4540. Only the surrounding regions of polymorphic positions are shown. Sequence logos were created using the WebLogo application (https://weblogo.berkeley.edu/logo.cgi). The height of each amino acid symbol indicates the relative frequency of each amino acid at that position. Dots indicate identical nucleotides to the reference sequence. Pjgsc1-Cisse, gsc1 gene sequence of P. jirovecii retrieved from Cissé and colleagues’ genome (20); Pjgsc1-Ma, gsc1 gene sequence of P. jirovecii retrieved from Ma and colleagues’ genome (21); Pjgsc1-B4, gsc1 gene sequence of P. jirovecii from patient B4; Pjgsc1-B20, gsc1 gene sequence of P. jirovecii from patient B20; Pjgsc1-N10, gsc1 gene sequence of P. jirovecii from patient N10.

FIG 2

Position of HS1 and HS2 of wild GSC1 protein types of C. albicans, C. tropicalis, C. krusei (Desnos-Olivier et al. [18]), and Saccharomyces cerevisiae (Luraschi et al. [22]) aligned with the GSC1 protein type of P. jirovecii (Cissé et al. [20], Ma et al. [21]). Boldfaced amino acids indicate known positions of mutations at HS1 and HS2 of GSC1 protein of Candida species. HS1 and HS2 GSC1 protein sequences of P. jirovecii in the present study were identical to those described by Cissé et al. (20) and Ma et al. (21).