Protocols for Studying Pathogenic Fungi

Ritu Pasrija, Deepika Kumari

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English

Springer Nature Switzerland AG
24 May 2025

This book is a compilation of protocols for conducting experiments with a variety of yeasts, including S. cerevisiae, Candida albicans, non-albicans Candida sp., Cryptococcus sp.; and molds, including Aspergillus sp., Fusarium sp., Mucor, Rhizopus. The chapters not only discuss National Committee for Clinical Laboratory Standards (NCCLS) accredited protocols but also emphasize the significance of Biosafety Level-2 (BSL-2) facilities. Subsequent chapters cover the essentials of handling, culturing, preserving, and reviving various fungal pathogens. Moreover, advanced techniques are explored, including counting colony-forming units (CFU), growth curve preparation, and microtiter plate assays. A separate chapter discusses the protocols for disc-diffusion, spot and kill-kinetics assays, addressing analyses like a checkerboard and Fractional Inhibitory Concentration (FIC) index assessments. Additionally, the book covers the formation of biofilm, capsule and melanin, fungal cell adhesion, and sterols extraction. Finally, it elaborates on the optimized impact achieved by assessing mitochondrial fitness through non-fermentable carbon.

This book is relevant for mycologists and scientists working in the field, providing insights into fungal pathogens and protocols for handling different fungal species.

By: Ritu Pasrija, Deepika Kumari
Imprint: Springer Nature Switzerland AG
Country of Publication: Switzerland
Dimensions: Height: 235mm, Width: 155mm,
ISBN: 9789819670468
ISBN 10: 9819670462
Pages: 262
Publication Date: 24 May 2025
Audience: Professional and scholarly , College/higher education , Undergraduate , Further / Higher Education
Format: Hardback
Publisher's Status: Active

1. Introduction to BSL-2 Facilities.- 2. Handling, Culturing, Preservation and Revival of Fungal Pathogens.- 3. Counting Colony Forming Units (CFU) to Estimate Viable Fungal Cells in a Sample.- 4. Growth Curve Preparation for the Calculation of Fungal Cell Count Over Time.- 5. Microtiter Assay for Drug Susceptibility Testing of Pathogenic Fungi in Broth.- 6. Disk Diffusion Assay for Testing the Drug Sensitivity of Pathogenic Fungi in Agar.- 7. Spot Assay for Comparing Drug Susceptibility of Pathogenic Fungi in Agar.- 8. Time-Kill Kinetics to Determine the Number of Living Cells Over a Period of Time.- 9. Checkerboard Analysis to Calculate the Fractional Inhibitory Concentration (FIC) Index of Pathogenic Fungi for Drug Interaction.- 10. Hyphal Induction in Yeasts and Molds.- 11. Isolation of Genomic DNA from Pathogenic Fungi.- 12. Exopolysaccharide Secretion by Pathogenic Fungi.- 13. Biofilm Formation Test and Determination of the Effect of Drugs on their Formation.- 14. Induction of Capsule Formation in Cryptococcus sp.- 15. Melanin Formation in Cryptococcus sp.- 16. Urease Assay for Cryptococcus sp.- 17. Phospholipase Assay for the Detection of Virulence of Pathogenic Fungi.- 18. Hydrophobicity Test for Virulence of Pathogenic Fungi.- 19. Adhesion of Fungal Cells to Buccal Epithelial Cells.- 20. Sterols Extraction, Analysis and Quantification in Pathogenic Fungi.- 21. Assessing Mitochondrial Fitness by Assessing Microbial Growth on Non-Fermentable Carbon Sources.- 22. In Vivo Assay using Caenorhabditis elegans.

Ritu Pasrija is an esteemed Associate Professor in the Department of Biochemistry at Maharshi Dayanand University (MDU), where she has made significant contributions to the field of biochemistry and microbiology. With over 20 years of experience in the study of drug resistance, her research primarily focuses on pathogenic fungi, including Candida sp., Cryptococcus, and Fusarium. Dr. Pasrija completed her PhD at Jawaharlal Nehru University, one of India’s leading institutions, where she deepened her understanding of molecular biology and microbial pathogenesis. Before her academic career, she worked at Ranbaxy Pharmaceuticals, where she gained valuable industry experience, particularly in pharmaceutical research and development. Her research expertise lies in the mechanisms of drug resistance, with a particular interest in antifungal resistance. Over the years, she has undertaken several research projects funded by prestigious agencies such as the University Grants Commission (UGC), the Department of Science and Technology (DST), and the Department of Biotechnology (DBT). These projects have furthered our understanding of the pathogenicity and resistance mechanisms of fungi that are responsible for various infections, particularly in immunocompromised individuals. In addition to her research work, she has published numerous research papers, review articles, and book chapters in reputable journals, contributing extensively to the scientific community. Her dedication to scientific excellence has been recognized by her peers, and she was honored with the Research Excellence Award at Maharshi Dayanand University. Her continued work in the field of drug resistance, particularly in the context of fungal pathogens, remains at the forefront of efforts to develop more effective treatments for infections caused by resistant organisms. Deepika Kumari is a dedicated PhD student in the Department of Biochemistry at Maharshi Dayanand University, Rohtak, India. In recognition of her exceptional academic potential and in support of her doctoral research, she has been awarded the prestigious Junior Research Fellowship (JRF) and Senior Research Fellowship (SRF) by the Council of Scientific & Industrial Research (CSIR), India. She has made significant contributions to the scientific community and has published five research papers in high-impact, peer-reviewed journals. She has also authored six book chapters and a comprehensive review article, demonstrating her expertise and tireless commitment to advancing the field of mycology. Her research focuses on the biology of fungi and their wider biochemical implications, demonstrating her deep passion for the subject and her potential to significantly contribute to the field. Her work is a testament to her dedication and growing stature as a scientist in biochemistry and mycology.