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Probe-based qPCR Assay for Rapid Detection of Predominant Candida glabrata Sequence Type in Korea

  • Bae, Jinyoung (Department of Clinical Laboratory Science, College of Health Sciences, Catholic University of Pusan) ;
  • Lee, Kyung Eun (Department of Clinical Laboratory Science, College of Health Sciences, Catholic University of Pusan) ;
  • Jin, Hyunwoo (Department of Clinical Laboratory Science, College of Health Sciences, Catholic University of Pusan)
  • Received : 2019.10.28
  • Accepted : 2019.11.28
  • Published : 2019.12.31

Abstract

Recent years have seen an increase in the incidence of candidiasis caused by non-albicans Candida (NAC) species. In fact, C. glabrata is now second only to C. albicans as the most common cause of invasive candidiasis. Therefore, the rapid genotyping specifically for C. glabrata is required for early diagnosis and treatment of candidiasis. A number of genotyping assays have been developed to differentiate C. glabrata sequence types (STs), but they have several limitations. In the previous study, multi-locus sequence typing (MLST) has performed with a total of 101 C. glabrata clinical isolates to analyze the prevalent C. glabrata STs in Korea. A total of 11 different C. glabrata STs were identified and, among them, ST-138 was the most commonly classified. Thus, a novel probe-based quantitative PCR (qPCR) assay was developed and evaluated for rapid and accurate identification of the predominant C. glabrata ST-138 in Korea. Two primer pairs and hybridization probe sets were designed for the amplification of internal transcribed spacer 1 (ITS1) region and TRP1 gene. Analytical sensitivity of the probe-based qPCR assay was 100 ng to 10 pg and 100 ng to 100 pg (per 1 μL), which target ITS1 region and TRP1 gene, respectively. This assay did not react with any other Candida species and bacteria except C. glabrata. Of the 101 clinical isolates, 99 cases (98%) were concordant with MLST results. This novel probe-based qPCR assay proved to be rapid, sensitive, highly specific, reproducible, and cost-effective than other genotyping assay for C. glabrata ST-138 identification.

Keywords

References

  1. Abbes S, Amouri I, Sellami H, Sellami A, Makni F, Ayadi A. A review of molecular techniques to type Candida glabrata isolates. Mycoses. 2010. 53: 463-467. https://doi.org/10.1111/j.1439-0507.2009.01753.x
  2. Abbes S, Sellami H, Sellami A, Hadrich I, Amouri I, Mahfoudh N, Neji S, Makni F, Makni H, Ayadi A. Candida glabrata strain relatedness by new microsatellite markers. Eur J Clin Microbiol Infect Dis. 2012. 31: 83-91. https://doi.org/10.1007/s10096-011-1280-4
  3. Becker K, Badehorn D, Deiwick S, Peters G, Fegeler W. Molecular genotyping of Candida species with special respect to Candida (Torulopsis) glabrata strains by arbitrarily primed PCR. J Med Microbiol. 2000. 49: 575-581. https://doi.org/10.1099/0022-1317-49-6-575
  4. Berila N, Subik J. Molecular analysis of Candida glabrata clinical isolates. Mycopathologia. 2010. 170: 99-105. https://doi.org/10.1007/s11046-010-9298-1
  5. Bineshian F, Yadegari MH, Sharifi Z, Akbari Eidgahi M, Nasr R. Identification of Candida Species Using MP65 Gene and Evaluation of the Candida albicans MP65 Gene Expression in BALB/C Mice. Jundishapur J Microbiol. 2015. 8: e18984.
  6. Byun SA, Won EJ, Kim MN, Lee WG, Lee K, Lee HS, Uh Y, Healey KR, Perlin DS, Choi MJ, Kim SH, Shin JH. Multilocus Sequence Typing (MLST) Genotypes of Candida glabrata Bloodstream Isolates in Korea: Association With Antifungal Resistance, Mutations in Mismatch Repair Gene (Msh2), and Clinical Outcomes. Front Microbiol. 2018. 9: 1523. https://doi.org/10.3389/fmicb.2018.01523
  7. Camacho-Cardoso JL, Martinez-Rivera MA, Manzano-Gayosso P, Mendez-Tovar LJ, Lopez-Martinez R, Hernandez-Hernandez F. Molecular detection of Candida species from hospitalized patient's specimens. Gac Med Mex. 2017. 153: 581-589.
  8. Da Silva-Rocha WP, Lemos VL, Svidizisnki TI, Milan EP, Chaves GM. Candida species distribution, genotyping and virulence factors of Candida albicans isolated from the oral cavity of kidney transplant recipients of two geographic regions of Brazil. MC Oral Health. 2014. 14: 20. https://doi.org/10.1186/1472-6831-14-20
  9. Dodgson AR, Pujol C, Denning DW, Soll DR, Fox AJ. Multilocus sequence typing of Candida glabrata reveals geographically enriched clades. J Clin Microbiol. 2003. 41: 5709-5717. https://doi.org/10.1128/JCM.41.12.5709-5717.2003
  10. Enache-Angoulvant A, Bourget M, Brisse S, Stockman-Pannier C, Diancourt L, Francois N, Rimek D, Fairhead C, Poulain D, Hennequin C. Multilocus microsatellite markers for molecular typing of Candida glabrata: application to analysis of genetic relationships between bloodstream and digestive system isolates. J Clin Microbiol. 2010. 48: 4028-4034. https://doi.org/10.1128/JCM.02140-09
  11. Essendoubi M, Toubas D, Lepouse C, Leon A, Bourgeade F, Pinon JM, Manfait M, Sockalingum GD. Epidemiological investigation and typing of Candida glabrata clinical isolates by FTIR spectroscopy. J Microbiol Methods. 2007. 71: 325-331. https://doi.org/10.1016/j.mimet.2007.09.018
  12. Foongladda, S., Mongkol, N., Petlum, P, Chayakulkeeree M. Multiprobe Real-Time PCR Identification of Four Common Candida Species in Blood Culture Broth. Mycopathologia. 2014. 177: 251-261. https://doi.org/10.1007/s11046-014-9743-7
  13. Gohar AA, Badali H, Shokohi T, Nabili M, Amirrajab N, Moazeni M. Expression Patterns of ABC Transporter Genes in Fluconazole-Resistant Candida glabrata. Mycopathologia. 2017. 182: 273-284. https://doi.org/10.1007/s11046-016-0074-8
  14. Ho HL, Haynes K. Candida glabrata: new tools and technologiesexpanding the toolkit. FEMS Yeast Res. 2015. pii: fov066.
  15. Kang MJ. Sequence Type Analysis of Candida glabrata Clinical Isolates using Multi-locus Sequence Typing in Korea. Catholic University of Pusan. 2017. (Dissertation)
  16. Katiyar S, Shiffrin E, Shelton C, Healey K, Vermitsky J-P, Edlind T. Evaluation of Polymorphic Locus Sequence Typing for Candida glabrata Epidemiology. J Clin Microbiol. 2016. 54: 1042-1050. https://doi.org/10.1128/JCM.03106-15
  17. Lin CY, Chen YC, Lo HJ, Chen KW, Li SY. Assessment of Candida glabrata strain relatedness by pulsed-field gel electrophoresis and multilocus sequence typing. J Clin Microbiol. 2007. 45: 2452-2459. https://doi.org/10.1128/JCM.00699-07
  18. Lott TJ, Frade JP, Lockhart SR. Multilocus sequence type analysis reveals both clonality and recombination in populations of Candida glabrata bloodstream isolates from U.S. surveillance studies. Eukaryot Cell. 2010. 9: 619-625. https://doi.org/10.1128/EC.00002-10
  19. Navarro E, Serrano-Heras G, Castano MJ, Solera J. Real-time PCR detection chemistry. Clin Chim Acta. 2015. 439: 231-250. https://doi.org/10.1016/j.cca.2014.10.017
  20. Ortiz B, Perez-A E, Galo C, Fontecha G. Molecular identification of Candida species from urinary infections in Honduras. Rev Iberoam Micol. 2018. 35: 73-77. https://doi.org/10.1016/j.riam.2017.07.003
  21. Paluchowska P, Tokarczyk M, Bogusz B, Skiba I, Budak A. Molecular epidemiology of Candida albicans and Candida glabrata strains isolated from intensive care unit patients in Poland. Mem Inst Oswaldo Cruz. 2014. 109: 436-441. https://doi.org/10.1590/0074-0276140099
  22. Perez-Losada M, Cabezas P, Castro-Nallar E, Crandall KA. Pathogen typing in the genomics era: MLST and the future of molecular epidemiology. Infect Genet Evol. 2013. 16: 38-53. https://doi.org/10.1016/j.meegid.2013.01.009
  23. Pujol C, Joly S, Lockhart SR, Noel S, Tibayrenc M, Soll DR. Parity among the randomly amplified polymorphic DNA method, multilocus enzyme electrophoresis, and Southern blot hybridization with the moderately repetitive DNA probe Ca3 for fingerprinting Candida albicans. J Clin Microbiol. 1997. 35: 2348-2358. https://doi.org/10.1128/jcm.35.9.2348-2358.1997
  24. Rezazadeh E, Moazeni M, Sabokbar A. Use of cost effective and rapid molecular tools for identification of Candida species, opportunistic pathogens. Curr Med Mycol. 2016. 2: 1-4. https://doi.org/10.18869/acadpub.cmm.2.3.1
  25. Rozen SG, Untergasser A, Cutcutache I, Koressaar T, Ye J, Faircloth BC, Remm M, Rozen SG. Primer3 - new capabilities and interfaces. Nucleic Acids Research. 2012. 40: e115. https://doi.org/10.1093/nar/gks596
  26. Sadeghi G, Ebrahimi-Rad M, Mousavi SF, Shams-Ghahfarokhi M, Razzaghi-Abyaneh M. Emergence of non-Candida albicans species: Epidemiology, phylogeny and fluconazole susceptibility profile. J Mycol Med. 2018. 28: 51-58. https://doi.org/10.1016/j.mycmed.2017.12.008
  27. Shahrokhi S, Noorbakhsh F, Rezaie S. Quantification of CDR1 Gene Expression in Fluconazole Resistant Candida glabrata Strains Using Real-time PCR. Iran J Public Health. 2017. 46: 1118-1122.
  28. Shin JH, Chae MJ, Song JW, Jung SI, Cho D, Kee SJ, Kim SH, Shin MG, Suh SP, Ryang DW. Changes in karyotype and azole susceptibility of sequential bloodstream isolates from patients with Candida glabrata candidemia. J Clin Microbiol. 2007. 45: 2385-2391. https://doi.org/10.1128/JCM.00381-07
  29. 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: 288-305. https://doi.org/10.1111/j.1574-6976.2011.00278.x
  30. Takahashi H, Saito R, Miya S, Tanaka Y, Miyamura N, Kuda T, Kimura B. Development of quantitative real-time PCR for detection and enumeration of Enterobacteriaceae. Int J Food Microbiol. 2017. 246: 92-97. https://doi.org/10.1016/j.ijfoodmicro.2016.12.015
  31. Whaley SG, Caudle KE, Simonicova L, Zhang Q, Moye-Rowley WS, Rogers PD. Jjj1 Is a Negative Regulator of Pdr1-Mediated Fluconazole Resistance in Candida glabrata. mSphere. 2018. pii: e00466-17.
  32. Zhao Y, Nagasaki Y, Kordalewska M, Press EG, Shields RK, Nguyen MH, Clancy CJ, Perlin DS. Rapid Detection of FKSAssociated Echinocandin Resistance in Candida glabrata. Antimicrob Agents Chemother. 2016. 60: 6573-6577. https://doi.org/10.1128/AAC.01574-16