Azole Resistance and erg11 Gene Expression in Non-albicans Candida Strains Isolated from Raw Milk and Human Samples: Cross-sectional Study from 14 Farms and 2 Hospitals, Iran, 2021-2022

Namvar, Zahra; Akhavan Sepahy, Abbas; Rafiei Tabatabaei, Robab; Sharifynia, Somayeh; Rezaie, Sassan

doi:10.30699/ijmm.17.4.447

year 17, Issue 4 (July - August 2023) Iran J Med Microbiol 2023, 17(4): 447-456 | Back to browse issues page

‎ 10.30699/ijmm.17.4.447

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Namvar Z, Akhavan Sepahy A, Rafiei Tabatabaei R, Sharifynia S, Rezaie S. Azole Resistance and erg11 Gene Expression in Non-albicans Candida Strains Isolated from Raw Milk and Human Samples: Cross-sectional Study from 14 Farms and 2 Hospitals, Iran, 2021-2022. Iran J Med Microbiol 2023; 17 (4) :447-456
URL: http://ijmm.ir/article-1-2075-en.html

Azole Resistance and erg11 Gene Expression in Non-albicans Candida Strains Isolated from Raw Milk and Human Samples: Cross-sectional Study from 14 Farms and 2 Hospitals, Iran, 2021-2022

Zahra Namvar¹

, Abbas Akhavan Sepahy¹

, Robab Rafiei Tabatabaei¹

, Somayeh Sharifynia²

, Sassan Rezaie

1- Department of Microbiology, School of Biological Sciences, North Tehran Branch, Islamic Azad University, Tehran, Iran
2- Clinical Tuberculosis and Epidemiology Research Center, National Research Institute of Tuberculosis and long Diseases (NRITLD), Shahid Beheshti University of Medical Sciences, Tehran, Iran
3- Department of Medical Mycology and Parasitology, Division of Molecular Biology, School of Public Health, Tehran University of Medical Sciences, Tehran, Iran , srezaie@tums.ac.ir

Abstract: (754 Views)

Background and Aim: Nowadays, non-albicans Candida are common in human pathogens, and some of these cases were found in milk. Therefore, as well as the lack of accurate estimates of its global prevalence and severity, the present study aims to assess the demographic features of non-albicans Candida (NAC) spp. and determine the species distribution of NAC. It was also evaluating the in vitro Azole susceptibility of NAC species and identified the erg11 gene and erg11 expression in fluconazole-resistant isolates of NAC spp., in Iran.
Materials and Methods: In the present study, non-albicans Candida, including Candida glabrata, Candia krusei, Candida parapsilosis, and Candida tropicalis, were isolated and identified from 14 farms (raw milk) and human patients using culture methods, Real-Time PCR and sequencing. The resistance and susceptibility of the samples to azole were examined and erg11 expression was evaluated by RT-qPCR. The results were analyzed by REST Software to compare the levels of erg11 gene expression involved in drug resistance of NAC.
Results: 74 and 52 NAC strains were isolated in 262 collected milk samples and human samples. Based on ITS sequencing, 0.76% were identified as C. glabrata, 2.29% C. tropicalis, 4.19% C. parapsilosis, and 19.8% C. krusei. The expression of erg11 gene in the NAC was increased in samples isolated from humans compared to samples isolated from livestock (P>0.05), while no significant difference was found in the case of Candida glabrata isolated from both sources (P<0.05). All NAC isolates were sensitive to flucytosine.
Conclusion: NAC isolates from cows' milk have antifungal resistance genes while they had not taken any antifungal drugs. The resistance gene is transferred from antifungal agents in crop protection medications. Clinical isolates also had increased resistance to antifungal activity. Also, using Azole antibiotics can increase resistance gene level activity. This phenomenon should be considered for treatment program protocols.

Keywords: erg11, non-albicans Candida, Real time PCR, genes Expression, Azole

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Type of Study: Original Research Article | Subject: Molecular Microbiology
Received: 2023/05/7 | Accepted: 2023/08/3 | ePublished: 2023/09/27

References

1. Rayens E, Norris KA. Prevalence and Healthcare Burden of Fungal Infections in the United States, 2018. Open Forum Infect Dis. 2022;9(1):ofab593. [DOI:10.1093/ofid/ofab593] [PMID] [PMCID]

2. Khodadadi H, Zomorodian K, Nouraei H, Zareshahrabadi Z, Barzegar S, Zare MR, et al. Prevalence of superficial-cutaneous fungal infections in Shiraz, Iran: A five-year retrospective study (2015-2019). J Clin Lab Anal. 2021;35(7):e23850. [DOI:10.1002/jcla.23850] [PMID] [PMCID]

3. Colombo AL, de Almeida Júnior JN, Slavin MA, Chen SCA, Sorrell TC. Candida and invasive mould diseases in non-neutropenic critically ill patients and patients with haematological cancer. Lancet Infect Dis. 2017;17(11):e344-e56. [DOI:10.1016/S1473-3099(17)30304-3] [PMID]

4. Mendonça A, Santos H, Franco-Duarte R, Sampaio P. Fungal infections diagnosis - Past, present and future. Res Microbiol. 2022;173(3):103915. [DOI:10.1016/j.resmic.2021.103915] [PMID] [PMCID]

5. Agnelli C, Valerio M, Bouza E, Guinea J, Sukiennik T, Guimarães T, et al. Prognostic factors of Candida spp. bloodstream infection in adults: A nine-year retrospective cohort study across tertiary hospitals in Brazil and Spain. Lancet Reg Health Am. 2022;6:100117. [DOI:10.1016/j.lana.2021.100117] [PMID] [PMCID]

6. Pfaller MA, Huband MD, Flamm RK, Bien PA, Castanheira M. Antimicrobial activity of manogepix, a first-in-class antifungal, and comparator agents tested against contemporary invasive fungal isolates from an international surveillance programme (2018-2019). J Glob Antimicrob Resist. 2021;26:117-27. [DOI:10.1016/j.jgar.2021.04.012] [PMID]

7. Zastempowska E, Grajewski J, Twarużek M. Food-Borne Pathogens and Contaminants in Raw Milk - A Review. Ann Anim Sci. 2016;16(3):623-39. [DOI:10.1515/aoas-2015-0089]

8. Du J, Wang X, Luo H, Wang Y, Liu X, Zhou X. Epidemiological investigation of non-albicans Candida species recovered from mycotic mastitis of cows in Yinchuan, Ningxia of China. BMC Vet Res. 2018;14(1):251. [DOI:10.1186/s12917-018-1564-3] [PMID] [PMCID]

9. Ghannoum MA. Potential role of phospholipases in virulence and fungal pathogenesis. Clin Microbiol Rev. 2000;13(1):122-43. [DOI:10.1128/CMR.13.1.122] [PMID] [PMCID]

10. Arastehfar A, Gabaldón T, Garcia-Rubio R, Jenks JD, Hoenigl M, Salzer HJF, et al. Drug-Resistant Fungi: An Emerging Challenge Threatening Our Limited Antifungal Armamentarium. Antibiotics. 2020;9(12):877. [DOI:10.3390/antibiotics9120877] [PMID] [PMCID]

11. Fromtling RA. Overview of medically important antifungal azole derivatives. Clin Microbiol Rev. 1988;1(2):187-217. [DOI:10.1128/CMR.1.2.187] [PMID] [PMCID]

12. Berkow EL, Lockhart SR. Fluconazole resistance in Candida species: a current perspective. Infect Drug Resist. 2017;10:237-45. [DOI:10.2147/IDR.S118892] [PMID] [PMCID]

13. Mukherjee PK, Chandra J, Kuhn DM, Ghannoum MA. Mechanism of fluconazole resistance in Candida albicans biofilms: phase-specific role of efflux pumps and membrane sterols. Infect Immun. 2003;71(8):4333-40. [DOI:10.1128/IAI.71.8.4333-4340.2003] [PMID] [PMCID]

14. Krishnan S, Manavathu EK, Chandrasekar PH. Aspergillus flavus: an emerging non-fumigatus Aspergillus species of significance. Mycoses. 2009;52(3):206-22. [DOI:10.1111/j.1439-0507.2008.01642.x] [PMID]

15. Sanglard D, Odds FC. Resistance of Candida species to antifungal agents: molecular mechanisms and clinical consequences. Lancet Infec Dis. 2002;2(2):73-85. [DOI:10.1016/S1473-3099(02)00181-0] [PMID]

16. Villasmil ML, Barbosa AD, Cunningham JL, Siniossoglou S, Nickels JT, Jr. An Erg11 lanosterol 14-α-demethylase-Arv1 complex is required for Candida albicans virulence. PLoS One. 2020;15(7):e0235746. [DOI:10.1371/journal.pone.0235746] [PMID] [PMCID]

17. Jensen RH. Resistance in human pathogenic yeasts and filamentous fungi: prevalence, underlying molecular mechanisms and link to the use of antifungals in humans and the environment. Dan Med J. 2016;63(10):1-34.

18. Namvar Z, Sepahy AA, Tabatabaei RR, Rezaie S. Antifungal susceptibility of non-albicans Candida spp. isolated from raw milk and human blood in Alborz and Tehran provinces. Iran J Microbiol. 2019;11(6):520-6. [DOI:10.18502/ijm.v11i6.2224]

19. Pfaffl MW, Horgan GW, Dempfle L. Relative expression software tool (REST) for group-wise comparison and statistical analysis of relative expression results in real-time PCR. Nucleic Acids Res. 2002;30(9):e36. [DOI:10.1093/nar/30.9.e36] [PMID] [PMCID]

20. Han G, Liu N, Li C, Tu J, Li Z, Sheng C. Discovery of Novel Fungal Lanosterol 14α-Demethylase (CYP51)/Histone Deacetylase Dual Inhibitors to Treat Azole-Resistant Candidiasis. J Med Chem. 2020;63(10):5341-59. [DOI:10.1021/acs.jmedchem.0c00102] [PMID]

21. Aslam S, Qazi JI. Isolation of acidophilic lactic acid bacteria antagonistic to microbial contaminants. Pak J Zool. 2010;42(5):567-73.

22. Lee SC, Billmyre RB, Li A, Carson S, Sykes SM, Huh EY, et al. Analysis of a food-borne fungal pathogen outbreak: virulence and genome of a Mucor circinelloides isolate from yogurt. mBio. 2014;5(4):e01390-14. [DOI:10.1128/mBio.01390-14] [PMID] [PMCID]

23. Goncalves SS, Souza ACR, Chowdhary A, Meis JF, Colombo AL. Epidemiology and molecular mechanisms of antifungal resistance in Candida and Aspergillus. Mycoses. 2016;59(4):198-219. [DOI:10.1111/myc.12469] [PMID]

24. De Aguiar Cordeiro R, Pereira de Alencar L, Nogueira Brilhante RS, de Souza Collares Maia Castelo-Branco D, Cordeiro Teixeira CE, de Brito Macedo R, et al. Antifungal susceptibility of emerging opportunistic yeasts and yeast-like fungi from Rhea americana. Can J Microbiol. 2013;59(8):577-80. [DOI:10.1139/cjm-2013-0176] [PMID]

25. Voidarou C, Antoniadou Μ, Rozos G, Tzora A, Skoufos I, Varzakas T, et al. Fermentative Foods: Microbiology, Biochemistry, Potential Human Health Benefits and Public Health Issues. Foods. 2020;10(1):69. [DOI:10.3390/foods10010069] [PMID] [PMCID]

26. Silva S, Negri M, Henriques M, Oliveira R, Williams DW, Azeredo J. Candida glabrata, Candida parapsilosis and Candida tropicalis: biology, epidemiology, pathogenicity and antifungal resistance. FEMS Microbiol Rev. 2012;36(2):288-305. [DOI:10.1111/j.1574-6976.2011.00278.x] [PMID]

27. Demir KK, Butler-Laporte G, Del Corpo O, Ekmekjian T, Sheppard DC, Lee TC, et al. Comparative effectiveness of amphotericin B, azoles and echinocandins in the treatment of candidemia and invasive candidiasis: A systematic review and network meta-analysis. Mycoses. 2021;64(9):1098-110. [DOI:10.1111/myc.13290] [PMID]

28. Forastiero A, Mesa-Arango AC, Alastruey-Izquierdo A, Alcazar-Fuoli L, Bernal-Martinez L, Pelaez T, et al. Candida tropicalis antifungal cross-resistance is related to different azole target (Erg11p) modifications. Antimicrob Agents Chemother. 2013;57(10):4769-81. [DOI:10.1128/AAC.00477-13] [PMID] [PMCID]

29. Ghannoum MA, Rice LB. Antifungal agents: mode of action, mechanisms of resistance, and correlation of these mechanisms with bacterial resistance. Clin Microbiol Rev. 1999;12(4):501-17. [DOI:10.1128/CMR.12.4.501] [PMID] [PMCID]

30. Zhang J, Li L, Lv Q, Yan L, Wang Y, Jiang Y. The Fungal CYP51s: Their Functions, Structures, Related Drug Resistance, and Inhibitors. Front Microbiol. 2019;10:691. [DOI:10.3389/fmicb.2019.00691] [PMID] [PMCID]

31. Turner SA, Butler G. The Candida pathogenic species complex. Cold Spring Harb Perspect Med. 2014;4(9):a019778. [DOI:10.1101/cshperspect.a019778] [PMID] [PMCID]

32. Gómez-Gaviria M, Mora-Montes HM. Current Aspects in the Biology, Pathogeny, and Treatment of Candida krusei, a Neglected Fungal Pathogen. Infect Drug Resist. 2020;13:1673-89. [DOI:10.2147/IDR.S247944] [PMID] [PMCID]

33. West PT, Peters SL, Olm MR, Yu FB, Gause H, Lou YC, et al. Genetic and behavioral adaptation of Candida parapsilosis to the microbiome of hospitalized infants revealed by in situ genomics, transcriptomics, and proteomics. Microbiome. 2021;9(1):142. [DOI:10.1186/s40168-021-01085-y] [PMID] [PMCID]

34. Trofa D, Gácser A, Nosanchuk JD. Candida parapsilosis, an emerging fungal pathogen. Clin Microbiol Rev. 2008;21(4):606-25. [DOI:10.1128/CMR.00013-08] [PMID] [PMCID]

35. van Asbeck EC, Clemons KV, Stevens DA. Candida parapsilosis: a review of its epidemiology, pathogenesis, clinical aspects, typing and antimicrobial susceptibility. Crit Rev Microbiol. 2009;35(4):283-309. [DOI:10.3109/10408410903213393] [PMID]

36. Fernandes T, Silva S, Henriques M. Candida tropicalis biofilm's matrix--involvement on its resistance to amphotericin B. Diagn Microbiol Infect Dis. 2015;83(2):165-9. [DOI:10.1016/j.diagmicrobio.2015.06.015] [PMID]

37. Hazirolan G, Canton E, Sahin S, Arikan-Akdagli S. Head-to-head comparison of inhibitory and fungicidal activities of fluconazole, itraconazole, voriconazole, posaconazole, and isavuconazole against clinical isolates of Trichosporon asahii. Antimicrob Agents Chemother. 2013;57(10):4841-7. [DOI:10.1128/AAC.00850-13] [PMID] [PMCID]

38. Milici ME, Maida CM, Spreghini E, Ravazzolo B, Oliveri S, Scalise G, et al. Comparison between disk diffusion and microdilution methods for determining susceptibility of clinical fungal isolates to caspofungin. J Clin Microbiol. 2007;45(11):3529-33. [DOI:10.1128/JCM.00826-07] [PMID] [PMCID]

39. Jensen RH, Johansen HK, Soes LM, Lemming LE, Rosenvinge FS, Nielsen L, et al. Posttreatment Antifungal Resistance among Colonizing Candida Isolates in Candidemia Patients: Results from a Systematic Multicenter Study. Antimicrob Agents Chemother. 2015;60(3):1500-8. [DOI:10.1128/AAC.01763-15] [PMID] [PMCID]

40. Lupetti A, Danesi R, Campa M, Tacca MD, Kelly S. Molecular basis of resistance to azole antifungals. Trends Mol Med. 2002;8(2):76-81. [DOI:10.1016/S1471-4914(02)02280-3] [PMID]