Paracoccidioides lutzii Formamidase Contributes to Fungal Survival in Macrophages
Abstract
:1. Introduction
2. Materials and Methods
2.1. Ethics Statement
2.2. Fungal Strains and Growth Conditions
2.3. Construction of fmd P. lutzii Silenced Strains with Antisense RNA Technology
2.4. Validation of P. lutzii AsFmd Transformants and Expression Analysis by RT-qPCR
2.5. Growth Curve and Viability of P. lutzii Silenced Cells
2.6. Total Protein Extraction and Fmd Immunoblotting Assay
2.7. Measurement of Formamidase and Urease Activity
2.8. Urease Heterologous Expression
2.9. Ure Polyclonal Antibodies Production and Immunoblotting
2.10. Ex-Vivo Infection of AsFmd in J774 Macrophages
2.11. Statistical Analysis
3. Results
3.1. Production of Fmd P. lutzii Silenced Strains
3.2. Reduced Plfmd Gene Expression Does Not Impair Fungal Growth Rate and Cell Viability
3.3. Decreased PlFmd Expression Levels Influence PlUre Expression and Activity
3.4. PlFmd Is Important for P. lutzii Survival during Macrophage Infection
4. Discussion
5. Conclusions
Author Contributions
Funding
Data Availability Statement
Conflicts of Interest
References
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Silva, L.O.S.; Moreira, T.R.; Gonçales, R.A.; Tomazett, M.V.; Parente-Rocha, J.A.; Mattos, K.; Paccez, J.D.; Ruiz, O.H.; Pereira, M.; Soares, C.M.d.A.; et al. Paracoccidioides lutzii Formamidase Contributes to Fungal Survival in Macrophages. Microorganisms 2022, 10, 2011. https://doi.org/10.3390/microorganisms10102011
Silva LOS, Moreira TR, Gonçales RA, Tomazett MV, Parente-Rocha JA, Mattos K, Paccez JD, Ruiz OH, Pereira M, Soares CMdA, et al. Paracoccidioides lutzii Formamidase Contributes to Fungal Survival in Macrophages. Microorganisms. 2022; 10(10):2011. https://doi.org/10.3390/microorganisms10102011
Chicago/Turabian StyleSilva, Lana O’Hara Souza, Thalison Rodrigues Moreira, Relber Aguiar Gonçales, Mariana Vieira Tomazett, Juliana Alves Parente-Rocha, Karine Mattos, Juliano Domiraci Paccez, Orville Hernandez Ruiz, Maristela Pereira, Célia Maria de Almeida Soares, and et al. 2022. "Paracoccidioides lutzii Formamidase Contributes to Fungal Survival in Macrophages" Microorganisms 10, no. 10: 2011. https://doi.org/10.3390/microorganisms10102011