“DETERMINING THE ANTIFUNGAL SUSCEPTIBILITY PATTERN OF CANDIDA ALBICANS AND THE MOLECULAR PROFILING OF ERG11 GENE IN FLUCONAZOLE RESISTANT CANDIDA ALBICANS ISOLATES AT A TERTIARY CARE CENTRE, UTTAR PRADESH”.
Main Article Content
Keywords
Candida albicans, Antifungal, Molecular Characterization, ERG11, DNA, PCR
Abstract
Introduction: Candida is the most opportunistic fungal pathogen, which results in various manifestations ranging from mucocutaneous lesions to life threatening invasive diseases. The treatment of choice is the use of azoles, such as fluconazole (FLC). The mutations in ERG11 and amino acid substitutions in the target enzyme ERG11 leads to changes in the tertiary structure of enzyme and subsequently alter the abilities of azole antifungals.
Aim and Objective: To detect antifungal drug-resistant Antibiotic Susceptibility Pattern of Candida albicans and the Molecular Characterization of ERG11 gene of Fluconazole Resistant Candida albicans isolates.
Material and Methods: This was a cross sectional study carried out in the Department of Microbiology for a period of 1 year i.e, June 2022 to June 2023. A total of 72 consecutive Candida species were isolated from 764 clinical specimens. Growths on Sabouraud dextrose agar were evaluated for colony appearance, microscopic examination, Gram staining, germ tube test and urea hydrolysis test. Further, they were processed for Candida speciation on CHROMagar. Antifungal susceptibility testing was performed as recommended by Clinical and Laboratory Standards Institute (CLSI) guidelines 2022. The DNA isolation was done using the Qiagen DNA extraction kit followed by the conventional PCR for the molecular detection of ERG11 resistant gene.
Results: Out of the total 72 Candida isolates, Candida albicans 33 (45.83%) was the most common species. Among the non-albicans Candida species, Candida tropicalis 20 (27.7%) was the predominant isolate followed by Candida glabrata 12(16.6%) and least by C. krusie 7 (9.72%). The ratio of Males 21 (63.6%) was more as compared to that of the Females 12( 36.3%) with the maximum age of 31-40 years and least in the age group above 61 years of age. The number of isolates was maximum in the urine sample. The ERG11 gene was detected in all the fluconazole resistant 5 (15.15%) strain of candida albicans.
Conclusion: In the current study nearly 5(15.15) of the Candida albicans indicates a reduced sensitivity to the effects of azole drugs. Therefore, understanding how all Candida spp. display resistance to fluconazole is crucial if we are to maintain the efficacy of this essential antifungal treatment.
References
2. Kaur J, Sharma P, Sharma S. Emergence of non albicans Candida species in critical care patients of tertiary care hospital. India J Microbical Research. 2016; 3(3):398–400.
3. Kashid RA, Belawadi S, Devi G, Indumati. Incidence of Non-Candida albicans in patients with urinary tract infection with special reference to antifungal sensitivity. J Evolution Med Dent Sci. 2014; 1(4):572-77.
4. PfallerMA,DiekemaDJ,Epidemiologyofinvasivecandidiasis:apersistentpublichealthproblem.Clin Microbiol Rev. 2007; 20(1):133–163.
5. Centers for Disease Control and Prevention. Candidiasis. USA: 2015 June. Available from: https://www.cdc.gov/ fungal/diseases/candidiasis/
6. Lopes, J.P, Lionakis, M.S. Pathogenesis and virulence of Candida albicans. Virulence 202;, 13, 89–121.
7. de Oliveira Santos, G.C., Vasconcelos, C.C., Lopes, A.J.O., de Sousa Cartágenes, M.D.S., Filho, A.K.D.B., do Nascimento, F.R.F., Ramos, R.M.; Pires, E.R.R.B., de Andrade, M.S., Rocha, F.M.G. et al. Candida infections and therapeutic strategies: Mechanisms of action for traditional and alternative agents. Front. Microbiol. 2018; 3, 1351.
8. Flowers, S.A., Colón, B., Whaley, S.G., Schuler, M.A., Rogers, P.D. Contribution of clinically derived mutations in ERG11 to azole resistance in Candida albicans. Antimicrob. Agents Chemother. 2015; 59: 450–460.
9. Fumiyoshi, A.; Toshiki, H. Mechanistic role of ergosterol in membrane rigidity and cycloheximide resistance in Saccharomyces cerevisiae. Biochim. Biophys. Acta (BBA)-Biomembr. 2009; 1788, 743–752.
10. Dupont, S., Lemetais, G., Ferreira, T, Cayot, P. Gervais, P. Beney, L. Ergosterol biosynthesis: A fungal pathway for life on land? Evolution .2012; 66, 2961–2968.
11. Xiang, M.J. Liu, J.Y. Ni, P.H. Wang, S. Shi, C. Wei, B. Ni, Y.X. Ge, H.L. Erg11 mutations associated with azole resistance in clinical isolates of Candida albicans. FEMS Yeast Res. 2013; 13, 386–393.
12. Marichal, P. Koymans, L. Willemsens, S. Bellens, D. Verhasselt, P. Luyten, W. Borgers, M. Ramaekers, F.C.S. Odds, F.C. Vanden Bossche, H. Contribution of mutations in the cytochrome P450 14 alpha-demethylase (Erg11p, Cyp51p) to azole resistance in Candida albicans. Microbiology 1999; 145, 2701–2713.
13. Wang H, Kong F, Sorrell TC, Wang B, McNicholas P, Pantarat N et al. Rapid detection of ERG11 gene mutations in clinical Candida albicans isolates with reduced susceptibility to fluconazole by rolling circle amplification and DNA sequencing. BMC Microbiol. 2009; 14:167.
14. Clinical and Laboratory Standards Institute (CLSI).Reference method for broth dilution antifungal susceptibility testing of yeasts: approved standard, 3rd ed. CLSI document M27-A3. Clinical and Laboratory Standards Institute, Wayne, PA. 2022.
15. Aneta K. Urbanek 1 , Zofia Łapi ´ nska , Daria Derkacz 1 and Anna Krasowska . The Role of ERG11 Point Mutations in the Resistance of Candida albicans to Fluconazole in the Presence of Lactate. Pathogens. 2022; 11, 1289.
16. Patil, S. Rao, R.S.Majumdar, B. Anil, S. Clinical appearance of oral Candida infection and therapeutic strategies. Front. Microbiol.2015; 6, 1391.
17. Nabili M, Abdollahi Gohar A, Badali H, Mohammadi R, Moazeni M. Amino acid substitutions in Erg11p of azoleresistant Candida glabrata: possible effective substitutions and homology modelling. J Glob Antimicrob Resist. 2016; 5:42-6.
18. Goldman GH, da Silva Ferreira ME, dos Reis Marques E, Savoldi M, Perlin D, Park S, et al. Evaluation of fluconazole resistance mechanisms in Candida albicans clinical isolates from HIV-infected patients in Brazil. Diagn Microbiol Infect Dis. 2004; 50(1):25-32.
19. Chongtham, Urvashi & Athokpam, Debina & Singh, Rajkumar. (2022). Isolation, Identification and Antifungal Susceptibility Testing of Candida Species: A Cross-sectional Study from Manipur, India. JOURNAL OF CLINICAL AND DIAGNOSTIC RESEARCH. 16. 10.7860/JCDR/2022/55695.16248.
20. Shweta R. Sharma, Amit Mishra, Sudhir Singh, Prachi Singh. Distribution of Candida infection in clinical samples and their antifungal susceptibility pattern in hospital of western U.P.Int J Acad Med Pharm 2023; 5 (6); 440-443.
21. Hazrat Bilal, Muhammad Shafiq, Bing Hou, Rehmat Islam, Muhammad Nadeem Khan, Rahat Ullah Khan & Yuebin Zeng. Distribution and antifungal susceptibility pattern of Candida species from mainland China: A systematic analysis, Virulence. 2022; 13:1, 1573-1589.
22. Paul S, Kannan I, MohanramK. Extensive ERG11 mutations associated with fluconazole-resistant Candida albicans isolated from HIV-infected patients. Curr Med Mycol. 2019; 5(3): 1-6. DOI: 10.18502/cmm.5.3.1739.
23. Kaur, Pavneet & Rawat, Tania & Sharma, Sarabjeet & Kaur, Parmeet. (2021). Speciation of candida species isolated in clinical samples in a tertiary health care centre in Northern India. IP International Journal of Medical Microbiology and Tropical Diseases. 7. 262-268. 10.18231/j.ijmmtd.2021.054.
24. S U and Sumana MN (2023) Retrospective analysis on distribution and antifungal susceptibility profile of Candida in clinical samples: a study from Southern India. Front. Public Health 11:1160841. doi: 10.3389/fpubh.2023.1160841.
25. Lulu Zhang,XiaodongShe,Daniel Merenstein et.al.Fluconazole Resistance patterns in Candida species that colonize women with HIV infection.Current Therapeutic research. 2014; 76(C):84-89.
26. Lamsal,S.,Adhikari,S.,Raghubhansi,B.R.,Sapkota,S.,et.al Antifungal Susceptibility and Biofilm formation of Casndida albicans isolated from different clinical specimens.Tribhuvan University Journal of Microbiology. 2021; 8(1),53-62.https://doi.org/10.3126/tujm.v8.41195.
27. Henry, K.W., Nickels, J.T., Edlind, T.D.(2000).Upregulation of Erg Genes in Candida Species by Azoles and other sterol Biosynthesis Inhibitors. Antimicrob.Agents chemother.44,2693-2700.doi:10.1128/AAC.44.10.2693-2700.2000.
28. Ahmad A, Khan A, Manzoor N, Khan LA. Evolution of ergosterol biosynthesis inhibitors as fungicidal against Candida. Microb Pathog. 2010; 48(1):35–41
29. Revie NM; Iyer KR; Robbins N; Cowen LE Antifungal Drug Resistance: Evolution, Mechanisms and Impact. Curr. Opin. Microbiol .2018, 45, 70–76.
30. Robbins N; Caplan T; Cowen LE Molecular Evolution of Antifungal Drug Resistance. Annu. Rev. Microbiol .2017, 71, 753–775
31. Hoot SJ; Smith AR; Brown RP; White TC An A643V Amino Acid Substitution in Upc2p Contributes to Azole Resistance in Well-Characterized Clinical Isolates of Candida albicans. Antimicrob. Agents Chemother .2011, 55, 940–2. [
32. Heilmann CJ; Schneider S; Barker KS; Rogers PD; Morschhauser J An A643T Mutation in the Transcription Factor Upc2p Cause Constitutive ERG11 Upregulation and Increased Fluconazole Resistance in Candida albicans. Antimicrob. Agents Chemother. 2010, 54, 353–9.
33. Dunkel N; Liu TT; Barker KS; Homayouni R; Morschhauser J; Rogers PD A Gain-of-Function Mutation in the Transcription Factor Upc2p Causes Upregulation of Ergosterol Biosynthesis Genes and Increased Fluconazole Resistance in a Clinical Candida albicans Isolate. Eukaryotic Cell. 2008, 7, 1180–90.
34. MacPherson S; Akache B; Weber S; De Deken X; Raymond M; Turcotte B Candida albicans Zinc Cluster Protein Upc2p Confers Resistance to Antifungal Drugs and is an Activator of Ergosterol Biosynthetic Genes. Antimicrob. Agents Chemother .2005, 49, 1745–52.
35. Silver PM; Oliver BG; White TC Role of Candida albicans Transcription Factor Upc2p in Drug Resistance and Sterol Metabolism. Eukaryotic Cell .2004, 3, 1391–7.
36. Flowers SA; Barker KS; Berkow EL; Toner G; Chadwick SG; Gygax SE; Morschhauser J; Rogers PD Gain-of-Function Mutations in UPC2 are a Frequent Cause of ERG11 Upregulation in Azole-Resistant Clinical Isolates of Candida albicans. Eukaryotic Cell .2012, 11, 1289–99.
37. Howard, K.C.; Dennis, E.K.; Watt, D.S.; Garneau-Tsodikova, S. A comprehensive overview of the medicinal chemistry of antifungal drugs: Perspectives and promise. Chem. Soc. Rev. 2020; 49, 2426–2480.
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Nilam Nigam
Professor and Head, Department of Pharmacology, Rama Medical College Hospital and Research Centre, Mandhana, Kanpur, Uttar Pradesh, India
Saif Anees
Assistant Professor, Department of Community Medicine, Government Medical College, Kannauj, India.
Tahseen Mazhar
Research Associate, Department of Microbiology, Integral University, Lucknow, India.
Nashra Afaq
Research Associate, Department of Microbiology and Central Research Laboratory, Rama Medical College Hospital and Research Centre, Mandhana, Kanpur, Uttar Pradesh, India.
Suraiya Khanam Ansari
Associate Professor and Head, Department of Microbiology, G.S.V.M. Medical College, Kanpur, Uttar Pradesh, India.
Deepak Kumar
Assistant Professor and Head, Department of Microbiology, Uma Nath Singh Autonomous State Medical College, Jaunpur, India.
Raziuddin Khan
Assistant Professor, Department of Microbiology, Al-Falah University, Faridabad, Haryana, India