Impact of TR34/L98H, TR46/Y121F/T289A and TR53 Alterations in Azole-Resistant Aspergillus fumigatus on Sterol Composition and Modifications after In Vitro Exposure to Itraconazole and Voriconazole
Abstract
:1. Introduction
2. Materials and Methods
2.1. Isolates of A. fumigatus
2.2. Culture Conditions
2.3. Total Sterol Extraction
2.4. Sterol Derivatization
2.5. Sterol Content Analysis
2.6. Statistical Analysis
3. Results and Discussion
3.1. Identification of Sterols
3.2. Qualitative Composition and Relative Amount of Sterols
3.2.1. Under Basal Conditions
3.2.2. Under Itraconazole or Voriconazole Exposure
3.3. Statistical Comparison of Ergosterol Content
3.4. Statistical Comparison of 14α-Methylsterols Content
4. Conclusions
Author Contributions
Funding
Institutional Review Board Statement
Informed Consent Statement
Data Availability Statement
Acknowledgments
Conflicts of Interest
References
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Name | Origin | CYP51A Alteration | MIC Itraconazole mg/L | MIC Voriconazole mg/L |
---|---|---|---|---|
ATCC 204305 | NA | 0.25 (S) | 0.5 (S) | |
AF1861 | C | 0.125 (S) | 1 (S) | |
AF1799 | C | 0.5 (S) | 1 (S) | |
AF899 | C | TR34/L98H | >32 (R) | 8 (R) |
AF1897 | C | TR34/L98H | >32 (R) | 16 (R) |
AF23 | E | TR34/L98H | >32 (R) | 16 (R) |
AF1168 | C | TR46/Y121F/T289A | 1 (S) | >32 (R) |
AF2226 | C | TR46/Y121F/T289A | 8 (R) | >32 (R) |
AF1468 | C | TR46/Y121F/T289A | 8 (R) | >32 (R) |
AF84 | E | TR53 | >32 (R) | 3 (R) |
AF112 | E | TR53 | >32 (R) | 16 (R) |
AF124 | E | TR53 | 6 (R) | 2 (R) |
Identified Sterols | IUPAC Name | MW(TMS) | RRT | Reference |
---|---|---|---|---|
9-dehydroergosterol | 24-methylcholesta-5,7,9(11),22-tetraen-3ß-ol or ergosta -5,7,9(11),22-tetraen-3ß-ol | 466 | 1.05 | [24,25] |
Unknown 1 | 480 | 1.06 | ||
Ergosterol Z Ergosterol E | 24-methylcholesta-5,7,22-trien-3ß-ol or ergosta-5,7,22Z-trien-3ß-ol or ergosta-5,7,22E-trien-3ß-ol | 468 | 1.06 1.10 | [26] |
Ergosta-7,22-dienol | 24-methylcholesta-7,22-dien-3ß-ol or ergosta-7,22-dien-3ß-ol | 470 | 1.13 | [24] |
14α-methylepisterol | 14α,24- dimethylcholesta-7,24(28)-dien-3ß-ol or 14-methylergosta-7,24(28)-dien-3ß-ol | 484 | 1.14 | |
Unknown 2 | 470 | 1.18 | ||
7-dehydrostigmasterol | 24 ethyl-cholesta-5,7,22-trienol | 482 | 1.19–1.20 | [27] |
14α-methylfecosterol | 14α,24- dimethylcholesta-8,24(28)-dien-3ß-ol or 14α-methylergosta -8,24(28)-dien-3ß-ol | 484 | 1.20 | [28] |
Obtusifolione | 4α,14α,24-trimethylcholesta-8,24(28)-dien-3ß-one or 4α,14α-dimethylergosta-8,24(28)-dien-3ß-one | 424 | 1.22 | [29] |
Obtusifoliol | 4α,14α,24-trimethylcholesta-8,24(28)-dien-3ß-ol or 4α,14α-dimethylergosta-8,24(28)-dien-3ß-ol | 498 | 1.23 | [24] |
4-methylfecosterol | 4α,24- dimethylcholesta-8,24(28)-dien-3ß-ol or 4α- methylergosta-8,24(28)-dien-3ß-ol | 484 | 1.25 | [24] |
Eburicol | 4α,4ß,14α,24-tetramethylcholesta-8,24(28)-dien-3ß-ol or 4α,4ß,14α-trimethylergosta-8,24(28)-dien-3ß-ol | 512 | 1.31 | [28] |
ATCC 204305 | AF1861 | AF1799 | |||||||
---|---|---|---|---|---|---|---|---|---|
Basal | ITC | VRC | Basal | ITC | VRC | Basal | ITC | VRC | |
ergosterol Z et E | 80.0 (±0.4) | 81.1 (±5.3) | 70.4 (±2.9) | 79.5 (±6.1) | 78.8 (±4.1) | 68.5 (±5.6) | 85.9 (±2.7) | 68.3 (±56.9) | 65.3 (±2.0) |
Unknown 1 | 1.3 (±1.5) | ||||||||
14α-methylepisterol | 1.6 (±0.2) | 1.8 (±0.4) | 2.4 (±0.5) | 3.0 (±0.8) | |||||
7-dehydrostigmasterol | 20.0 (±0.4) | 13.9 (±7.0) | 9.9 (±1.1) | 18.9 (±5.5) | 17.0 (±4.0) | 2.1 (±1.0) | 14.2 (±2.6) | 1.0 (±1.7) | 0.7 (±1.2) |
obtusifoliol + obtusifolione | 2.4 (±2.5) | 8.8 (±1.0) | 0.8 (±1.4) | 1.5 (±0.1) | 13.3 (±3.3) | 12.6 (±3.1) | 14.1 (±0.8) | ||
eburicol | 2.5 (±1.3) | 8.1 (±1.2) | 0.8 (±1.3) | 2.8 (±0.5) | 14.4 (±1.3) | 15.8 (±6.4) | 16.9 (±2.1) |
AF23 | AF0899 | AF1897 | |||||||
---|---|---|---|---|---|---|---|---|---|
Basal | ITC | VRC | Basal | ITC | VRC | Basal | ITC | VRC | |
9-dehydroergosterol | 0.4 (±0.7) | ||||||||
ergosterol Z et E | 82.9 (±2.7) | 74.0 (±5.5) | 68.8 (±3.4) | 88.0 (±1.0) | 72.8 (±3.3) | 69.5 (±3.1) | 84.6 (±4.3) | 69.5 (±5.5) | 44.4 (±9.7) |
ergosta-7,22-dienol | 0.3 (±0.6) | 0.4 (±0.7) | |||||||
14α-methylepisterol | 1.1 (±0.2) | 1.9 (±0.2) | 1.6 (±1.6) | ||||||
unknown 2 | 0.5 (±0.9) | ||||||||
7-dehydrostigmasterol | 16.7 (±2.1) | 16.4 (±6.8) | 5.7 (±1.7) | 12.0 (±0.9) | 12.8 (±5.0) | 4.9 (±3.9) | 15.0 (±3.6) | 14.6 (±4.4) | 0.7 (±1.2) |
obtusifoliol + obtusifolione | 2.0 (±0.6) | 10.4 (±2.8) | 4.4 (±0.4) | 10.0 (±2.1) | 4.4 (±0.6) | 20.3 (±4.3) | |||
4-methyl-fecosterol | 0.3 (±0.6) | ||||||||
eburicol | 8.0 (±0.4) | 16.3 (±5.4) | 10.0 (±2.3) | 13.2 (±4.6) | 11.1 (±2.6) | 33.0 (±7.9) |
AF1168 | AF2226 | AF1468 | |||||||
---|---|---|---|---|---|---|---|---|---|
Basal | ITC | VRC | Basal | ITC | VRC | Basal | ITC | VRC | |
ergosterol Z et E | 82.8 (±2.5) | 79.1 (±3.5) | 62.5 (±3.3) | 83.7 (±4.1) | 68.3 (±3.7) | 67.0 (±3.7) | 89.5 (±0.9) | 76.9 (±2.0) | 70.2 (±3.2) |
ergosta-7,22-dienol | 0.7 (±0.6) | ||||||||
14α-methylepisterol | 0.4 (±0.8) | 1.7 (±1.0) | 0.4 (±0.8) | * | |||||
7-dehydrostigmasterol | 17.2 (±2.5) | 11.7 (±3.8) | 0.8 (±1.3) | 16.3 (±4.1) | 18.3v (±1.1) | 7.3 (±1.2) | 10.5 (±0.9) | 9.2 (±1.7) | 7.3 (±2.9) |
obtusifoliol + obtusifolione | 3.5 (±1.9) | 13.6 (±3.4) | 1.7 (±0.8) | 6.9 (±0.8) | 2.5 (±0.7) | 5.4 (±0.6) | |||
eburicol | 5.2 (±3.5) | 21.5 (±4.3) | 11.1 (±3.0) | 18.5 (±1.4) | 11.5 (±3.4) | 17.0 (±4.2) |
AF84 | AF112 | AF124 | |||||||
---|---|---|---|---|---|---|---|---|---|
Basal | ITC | VRC | Basal | ITC | VRC | Basal | ITC | VRC | |
ergosterol Z et E | 89.1 (±2.9) | 67.2 (±3.6) | 74.2 (±5.1) | 85.9 (±4.1) | 70.3 (±3.6) | 53.7 (±3.9) | 89.1 (±2.9) | 61.8 (±12.8) | 58.9 (±6.6) |
ergosta-7,22-dienol | 1.1 (±1.0) | ||||||||
14α-methylepisterol | ** | ** | 0.5 (±0.9) | 2.5 (±2.4) | 2.3 (±0.6) | ||||
Unknown 2 | 1.2 (±0.1) | ||||||||
7-dehydrostigmasterol | 10.9 (±2.9) | 12.6 (±5.9) | 8.6 (±4.9) | 14.1 (±4.0) | 15.7 (±3.1) | 1.1 (±1.5) | 10.9 (±2.9) | 2.5 (±4.3) | 1.5 (±0.1) |
14α-methylfecosterol | *** | ||||||||
obtusifoliol + obtusifolione | 4.6 (±0.4) | 5.6 (±0.5) | 3.3 (±0.3) | 13.0 (±1.2) | 14.5 (±6.8) | 15.2 (±2.8) | |||
eburicol | 14.5 (±1.5) | 11.6 (±10.2) | 10.7 (±2.1) | 32.0 (±2.7) | 17.7 (±8.9) | 22.5 (±3.4) |
Reference | Isolates Name | Condition of Growth | Relative Composition of Sterols | |||||||
---|---|---|---|---|---|---|---|---|---|---|
Time | T °C | Liquid Medium | 1 | 2 | 3 | 4 | 5 | 6 ** | ||
[12,16] | CM237 | 18 h | 37 °C | MM | 73.5–75.8 | 4.7–6.1 | 1.9–2.7 | 1.3–1.6 | 1.1 | 13.8–17.0 |
[13] | ATCC46645 | Overnight | 37 °C | RPMI 1640 | 95.0 | 5.0 | ||||
[14] | ASFU1112 | 24 h | 35 °C | Sabouraud | 89.9 | 0.9 | 0.3 | 8.9 | ||
ASFU1119 | 87.1 | 0.3 | 12.6 | |||||||
ASFU1463 * | 75.5 | 19.4 | 5.1 | |||||||
[19] | ATCC MYA-4609 | 48 h | 37 °C | Sabouraud | 90.8 | 0.6 | 8.6 | |||
This study | 48 h | 37 °C | YPD | 79.5–89.5 | 10.5–20.0 | <1.0 | <1.0 | <1.0 |
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Lavergne, R.-A.; Albassier, M.; Hardouin, J.-B.; Alvarez-Moreno, C.; Pagniez, F.; Morio, F.; Le Pape, P.; Ourliac-Garnier, I. Impact of TR34/L98H, TR46/Y121F/T289A and TR53 Alterations in Azole-Resistant Aspergillus fumigatus on Sterol Composition and Modifications after In Vitro Exposure to Itraconazole and Voriconazole. Microorganisms 2022, 10, 104. https://doi.org/10.3390/microorganisms10010104
Lavergne R-A, Albassier M, Hardouin J-B, Alvarez-Moreno C, Pagniez F, Morio F, Le Pape P, Ourliac-Garnier I. Impact of TR34/L98H, TR46/Y121F/T289A and TR53 Alterations in Azole-Resistant Aspergillus fumigatus on Sterol Composition and Modifications after In Vitro Exposure to Itraconazole and Voriconazole. Microorganisms. 2022; 10(1):104. https://doi.org/10.3390/microorganisms10010104
Chicago/Turabian StyleLavergne, Rose-Anne, Marjorie Albassier, Jean-Benoît Hardouin, Carlos Alvarez-Moreno, Fabrice Pagniez, Florent Morio, Patrice Le Pape, and Isabelle Ourliac-Garnier. 2022. "Impact of TR34/L98H, TR46/Y121F/T289A and TR53 Alterations in Azole-Resistant Aspergillus fumigatus on Sterol Composition and Modifications after In Vitro Exposure to Itraconazole and Voriconazole" Microorganisms 10, no. 1: 104. https://doi.org/10.3390/microorganisms10010104
APA StyleLavergne, R.-A., Albassier, M., Hardouin, J.-B., Alvarez-Moreno, C., Pagniez, F., Morio, F., Le Pape, P., & Ourliac-Garnier, I. (2022). Impact of TR34/L98H, TR46/Y121F/T289A and TR53 Alterations in Azole-Resistant Aspergillus fumigatus on Sterol Composition and Modifications after In Vitro Exposure to Itraconazole and Voriconazole. Microorganisms, 10(1), 104. https://doi.org/10.3390/microorganisms10010104