Atovaquone exposure and Pneumocystis jirovecii cytochrome b mutations: French data and review of the literature.

Pneumocystis jirovecii is a transmissible fungus responsible for severe pneumonia (Pneumocystis pneumonia [PCP]) in immunocompromised patients. Missense mutations due to atovaquone selective pressure have been identified on cytochrome b (CYB) gene of P. jirovecii. It was recently shown that atovaquone prophylaxis can lead to the selection of specific P. jirovecii CYB mutants potentially resistant to atovaquone among organ transplant recipients. In this context, our objectives were to provide data on P. jirovecii CYB mutants and the putative selective pressure exerted by atovaquone on P. jirovecii organisms in France. A total of 123 patients (124 P. jirovecii specimens) from four metropolitan hospitals and two overseas hospitals were retrospectively enrolled. Fourteen patients had prior exposure to atovaquone, whereas 109 patients did not at the time of P. jirovecii detection. A 638 base-pair fragment of the CYB gene of P. jirovecii was amplified and sequenced. A total of 10 single nucleotide polymorphisms (SNPs) were identified. Both missense mutations C431T (Ala144Val) and C823T (Leu275Phe), located at the Qo active site of the enzyme, were significantly associated with prior atovaquone exposure, these mutations being conversely incidental in the absence of prior atovaquone exposure (P < 0.001). Considering that the aforementioned hospitals may be representative of the national territory, these findings suggest that the overall presence of P. jirovecii CYB mutants remains low in France.

The mutations C431T (Ala144Val) and C823T (Leu275Phe) at the cytochrome b (CYB) active site of Pneumocystis jirovecii are associated with patient prior exposure to atovaquone. Conversely, these mutations are incidental in the absence of exposure. Overall, the presence of P. jirovecii CYB mutants remains low in France.

Apparent Absence of Selective Pressure on Pneumocystis jirovecii Organisms in Patients with Prior Methotrexate Exposure.

Pneumocystis jirovecii infections occur in patients treated with methotrexate (MTX) because of immunosuppressive effects of this highly potent dihydrofolate reductase (DHFR) inhibitor. Conversely, MTX may act as an anti-P. jirovecii drug and consequently may exert a selective pressure on this fungus. In this context, we compared the sequences of the dhfr gene of P. jirovecii isolates obtained from two groups of patients with P. jirovecii infections. The first group, with systemic diseases or malignancies, had prior exposure to MTX (21 patients), whereas the second group (22 patients), the control group, did not. Three single nucleotide polymorphisms (SNPs) were observed at positions 278, 312, and 381. The first one was located in the intronic region and the two others were synonymous. Based on these SNPs, three P. jirovecii dhfr alleles, named A, B, and C, were specified. Allele A was the most frequent, as it was observed in 18 patients (85.7%) and in 16 patients (72.7%) of the first and second groups, respectively. No significant difference in P. jirovecii dhfr gene diversity in the two patient groups was observed. In conclusion, these original results suggest that MTX does not exert an overt selective pressure on P. jirovecii organisms.

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

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.