Microsatellite DNA Identification and Genotyping of Candida albicans from Lebanese Clinical Isolates

Abstract

The present study involves collecting 125 isolates labeled as C. albicans from five different Lebanese hospitals and utilizing the microsatellite genotyping test to determine the following: first, the accuracy of hospital identification by comparing microsatellite results to hospital results. Second, the frequency and genotypes of infectious strains present relative to tissue and hospital location- a possible indicator of nosocomial infection, and third, a possible relationship between lack of microsatellite heterozygosity to azole resistance. Our results showed that the error in hospital identification varied from 2 to 33%, averaging at 7%, with the highest identification error in stool. Misidentified isolates were mainly Candida tropicalis followed by C. glabrata and C. parapsilosis. Strains with similar genotypes were also found to occur within certain hospitals suggesting the possibility of nosocomial infection. Finally, a relationship between lack of heterozygosity and azole resistance was observed since nine out of 10 homozygous isolates sharing a common allele with a heterozygote strain were sensitive to all drugs tested, whereas the homozygous genotype was resistant to at least one drug.

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References

  1. 1.

    Cowen LE, Anderson JB, Kohn LM. Evolution of drug resistance in Candida albicans. Ann Rev Microbiol. 2002;56:139–65.

    Article  CAS  Google Scholar 

  2. 2.

    Odds FC, Gow NA, Brown AJ. Fungal virulence studies come of age. Genome Biol. 2001;2:1009.1–.4.

    Article  Google Scholar 

  3. 3.

    Bougnoux ME, Morand S, D’enfert C. Usefulness of multilocus sequence typing for characterization of clinical isolates of Candida albicans. J Clin Microbiol. 2002;40:1290–97.

    PubMed  Article  CAS  Google Scholar 

  4. 4.

    Regan DR, Pfaller MA, Hollis RJ, Wenzel RP. Evidence of nosocomial spread of Candida albicans causing blood stream infection in a neonatal intensive care unit. Diagn Microbiol Infect Dis. 1995;21:191–4.

    Article  Google Scholar 

  5. 5.

    Robert F, Lebreton F, Bougnoux ME, Paugam A, Wassermann D, Schlotterer M, et al. Use of random amplified polymorphic DNA as a typing method for Candida albicans in epidemiological surveillance of a burn unit. J Clin Microbiol. 1995;33:2366–71.

    PubMed  CAS  Google Scholar 

  6. 6.

    Hajjeh RA, Sofair AN, Harrison LH, Lyon GM, Arthington-Skaggs BA, Mirza SA, et al. Incidence of bloodstream infections due to Candida species and in vitro susceptibilities of isolates collected from 1998 to 2000 in a population-based active surveillance program. J Clin Microbiol. 2004;42:1519–27.

    PubMed  Article  Google Scholar 

  7. 7.

    Sandven P, Bevanger L, Digranes A, Haukland HH, Mannsaker T, Gaustad P, The Norwegian Yeast Study Group (2006) Candidemia in Norway (1991 to 2003): results from a nationwide study. J Clin Microbiol. 2006;44:1977–81.

    PubMed  Article  Google Scholar 

  8. 8.

    Edmond MB, Wallace SE, McClish DK, Pfaller MA, Jones RN, Wenzel RP. Nosocomial bloodstream infections in United States hospitals: a three-year analysis. Clin Infect Dis. 1999;29:239–44.

    PubMed  Article  CAS  Google Scholar 

  9. 9.

    Gudlaugsson O, Gillespie S, Lee K, Vandeberg J, Hu J, Messer S, et al. Attributable mortality of nosocomial candidemia, revisited. Clin Infect Dis. 2003;37:1172–77.

    PubMed  Article  Google Scholar 

  10. 10.

    Vazquez JA, Sanchez C, Dmuchowski C, Dembry L, Sobel JD, Zervos MJ. Nosocomial acquisition of Candida albicans: an epidemiologic study. J Infect Dis. 1993;168:195–201.

    PubMed  CAS  Google Scholar 

  11. 11.

    Vazquez JA, Dembry LM, Sanchez V, Vazquez MA, Sobel JD, Dmuchowski C, Zervos MJ. Nosocomial Candida glabrata colonization: an epidemiologic study. J Clin Microbiol. 1998;36:421–6.

    PubMed  CAS  Google Scholar 

  12. 12.

    Fidel PL, Vazquez JA, Sobel JD. Candida glabrata: review of epidemiology, pathogenesis, and clinical disease with comparison to C. albicans. Clin Microbiol Rev. 1999;12:80–96.

    PubMed  Google Scholar 

  13. 13.

    Isenberg HD, Tucci V, Cintron F, Singer C, Weinstein GS, Tyras D. Single source outbreak of Candida tropicalis complicating coronary bypass surgery. J Clin Microbiol. 1989;27:2426–28.

    PubMed  CAS  Google Scholar 

  14. 14.

    Banerjee SN, Emori TG, Culver DH, Gaynes RP, Jarvis WR, Horan T, et al. Secular trends in nosocomial primary bloodstream infections in the United States, 1980–1989: National Nosocomial Infections Surveillance System. Am J Med 1991;91:86S–9S.

    PubMed  Article  CAS  Google Scholar 

  15. 15.

    Abi-Said D, Anaissie E, Uzun O, Raad I, Pinzcowski H, Vartivarian S. The epidemiology of hematogenous candidiasis caused by different Candida species. Clin Infect Dis. 1997;24:122–1128, Erratum in: Clin Infect Dis 25:352.

    Article  Google Scholar 

  16. 16.

    Pappas PG, Rex JH, Sobel JD, Filler SG, Dismukes WE, Walsh TJ et al. Guidelines for treatment of candidiasis. Clin Infect Dis. 2004;15:161–89.

    Article  Google Scholar 

  17. 17.

    Denning DW, Kibbler CC, Barnes RA. British society for medical microbiology proposed standards of care for patients with invasive fungal infections. Lancet Infect Dis. 2003;3:230–40.

    PubMed  Article  Google Scholar 

  18. 18.

    Ellepola ANB, Morrison CJ. Laboratory diagnosis of invasive candidiasis. J Microbiol. 2005;43:65–84.

    PubMed  Google Scholar 

  19. 19.

    Rabkin JM, Oroloff SL, Corless CL, Benner KG, Flora KD, Rosen HR, et al. Association of fungal infection and increased mortality in liver transplant recipients. Am J Surg. 2000;179:426–30.

    PubMed  Article  CAS  Google Scholar 

  20. 20.

    Sampaio P, Gusmão L, Correia A, Alves C, Pina-Vaz C, Amorim A, et al. Highly polymorphic microsatellite for identification of Candida albicans strains. J Clin Microbiol. 2001;41:552–57.

    Article  CAS  Google Scholar 

  21. 21.

    Hedden DM, Buck JD. A re-emphasis germ tubes diagnostic for Candida albicans have no constrictions. Mycopathologia 1980;70:95–101.

    PubMed  Article  CAS  Google Scholar 

  22. 22.

    Pincus DH, Coleman DC, Pruitt WR. Rapid identification of Candida dubliniensis with commercial yeast identification systems. J Clin Microbiol. 1999;37:3533–39.

    PubMed  CAS  Google Scholar 

  23. 23.

    Gow NAR. Cell biology and cell cycle of Candida. In: Calderone RE, editors. Candida and candidiasis. Washington: ASM; 2002. p. 145–158.

    Google Scholar 

  24. 24.

    Gilfillan GD, Sullivan DJ, Haynes K, Parkinson T, Coleman DC, Gow NA. Candida dubliniensis: phylogeny and putative virulence factors. Microbiology 1998;144:829–38.

    PubMed  CAS  Google Scholar 

  25. 25.

    Rinaldi MG. Biology and pathogenicity of Candida species. In: Bode GP, editors. Candidiasis: pathogenesis, diagnosis and treatment. (2nd ed.), New York: Raven Press; 1993. p. 1–20.

  26. 26.

    Salkin IF, Land GA, Hurd NJ, Goldson PR, McGinnis MR. Evaluation of YeastIdent and Uni-Yeast-Tek yeast identification systems. J Clin Microbiol. 1987;25:624–27.

    PubMed  CAS  Google Scholar 

  27. 27.

    Ogletree FA, Abdlol AT, Ahearn DG. Germ tube formation in atypical strains of Candida albicans. Antonie van Leeuwenhoek 1978;44:15–24.

    PubMed  Article  CAS  Google Scholar 

  28. 28.

    Ellepola AN, Hurst SF, Elie CM, Morrison CJ. Rapid and unequivocal differentiation of Candida dubliniensis from other Candida species using species specific DNA probes: comparison with phenotypic identification methods. Oral Microbiol Immunol. 2003;18:379–88.

    PubMed  Article  CAS  Google Scholar 

  29. 29.

    McGinnis MR, Molina TC, Pierson DL, Mishra SK. Evaluation of the Biolog Microstation System for yeast identification. J Med Vet Mycol. 1996;34:349–52.

    PubMed  Article  CAS  Google Scholar 

  30. 30.

    Elie CM, Lott TJ, Reiss E Morrison CJ. Rapid identification of Candida species using species-specific DNA probes. J Clin Microbiol. 1998;36:3260–65.

    PubMed  CAS  Google Scholar 

  31. 31.

    Coignard C, Hurst SF, Benjamin LE, Brandt ME, Warnock DW, Morrison CJ. Resolution of discrepant results of Candida species identification by using DNA probes. J Clin Microbiol. 2004;42:858–61.

    PubMed  Article  CAS  Google Scholar 

  32. 32.

    Lunel FV, Licciardello L, Stefani S, Verbrugh HA, Melchers WJ, Meis JF, et al. Lack of consistent short sequence repeat polymorphisms in genetically homologous colonizing and invasive Candida albicans strains. J Bacteriol. 1998;180:3771–78.

    PubMed  CAS  Google Scholar 

  33. 33.

    Lott TJ, Effat MM. Evidence for a more recently evolved clade within a Candida albicans North American population. Microbiology 2001;147:1687–92.

    PubMed  CAS  Google Scholar 

  34. 34.

    Yazbek S, Barada G, Basma R, Mahfouz J, Khalaf RA. Significant discrepancy between real-time PCR identification and hospital identification of C. albicans from Lebanese patients. Med Sci Monit. 2007;13:MT7–12.

    PubMed  Google Scholar 

  35. 35.

    Tsuchimori N, Sharkey LL, Fonzi WA, French SW, Edwards JE, Filler SG. Reduced virulence of HWP1-deficient mutants of Candida albicans and their interactions with host cells. Infect Immun. 2000;68:1997–2002.

    PubMed  Article  CAS  Google Scholar 

  36. 36.

    Baumgartner C, Freydiere A, Gille Y. Direct identification and recognition of yeast species from clinical material by using Albicans ID and CHROMagar Candida plates. J Clin Microbiol. 1996;34:454–56.

    PubMed  CAS  Google Scholar 

  37. 37.

    Marot-Leblond A, Grimaud L, David S, Sullivan DJ, Coleman DC, Ponton J, Robert R. Evaluation of a rapid immunochromatographic assay for identification of Candida albicans and Candida dubliniensis. J Clin Microbiol. 2004;42:4956–60.

    PubMed  Article  CAS  Google Scholar 

  38. 38.

    Chong PP, Lee YL, Tan BC, Ng KP. Genetic relatedness of Candida strains isolated from women with vaginal candidiasis in Malaysia. J Med Microbiol. 2003;52:657–66.

    PubMed  Article  CAS  Google Scholar 

  39. 39.

    Lockhart SR, Reed BD, Pierson CL, Soll DR. Most frequent scenario for recurrent Candida vaginitis is strain maintenance with “subshuffling”: demonstration by sequential DNA fingerprinting with probes Ca3, C1, and CARE2. J Clin Microbiol. 1996;34:767–77.

    PubMed  CAS  Google Scholar 

  40. 40.

    Bretagne S, Costa JM, Besmond C, Carsique R, Calderone R. Microsatellite polymorphism in the promoter sequence of the elongation factor 3 gene of Candida albicans as the basis for a typing system. J Clin Microbiol. 1997;35:1777–80.

    PubMed  CAS  Google Scholar 

  41. 41.

    Metzgar D, Van Belkum A, Field D, Haubrich R, Wills C. Random amplification of polymorphic DNA and microsatellite genotyping of pre- and posttreatment isolates of Candida sp. from human immunodeficiency virus-infected patients on different fluconazole regimens. J Clin Microbiol. 1998;36:2308–13.

    PubMed  CAS  Google Scholar 

  42. 42.

    Benjamin DK Jr, Ross K, McKinney RE Jr, Benjamin DK, Auten R, Fisher RG. When to suspect fungal infection in neonates: a clinical comparison of Candida albicans and Candida parapsilosis fungemia with coagulase-negative Staphylococcal bacteremia. Pediatrics 2000;106:712–8.

    PubMed  Article  Google Scholar 

  43. 43.

    Trubenova D, Viragova S, Pilipcinec E, et al. Candidemia in cancer patients and in children in a neonatal intensive care unit. Folia Microbiol. 2001;46:161–4.

    CAS  Article  Google Scholar 

  44. 44.

    Magee BB, Magee PT. Induction of mating in Candida albicans by construction of MTLa and MTLalpha strains. Science 2000;289:310–13.

    PubMed  Article  CAS  Google Scholar 

  45. 45.

    Rustchenko EP, Howard DH, Sherman F. Chromosomal alterations of Candida albicans are associated with the gain and loss of assimilating functions. J Bacteriol 1994;176:3231–41.

    PubMed  CAS  Google Scholar 

  46. 46.

    Tavanti A, Gow NA, Maiden MC, Odds FC, Shaw DJ. Genetic evidence for recombination in Candida albicans based on haplotype analysis. Fungal Genet Biol. 2004;41:553–62.

    PubMed  Article  CAS  Google Scholar 

  47. 47.

    Tsang PW, Cao B, Siu PY, Wang J. Loss of heterozygosity, by mitotic gene conversion and crossing over, causes strain-specific adenine mutants in constitutive diploid Candida albicans. Microbiology 1999;145:1623–29.

    PubMed  CAS  Google Scholar 

  48. 48.

    Sampaio P, Gusmão L, Correia A, Alves C, Rodrigues G, Pina-Vaz C, et al. New microsatellite multiplex PCR for Candida albicans strain typing reveals microevolutionary changes. J Clin Microbiol. 2005;43:3869–76.

    PubMed  Article  CAS  Google Scholar 

  49. 49.

    Hilton C, Markie D, Corner B, Rikkerink E, Poulter R. Heat shock induces chromosome loss in the yeast Candida albicans. Mol Gen Genet. 1985;200:162–8.

    PubMed  Article  CAS  Google Scholar 

  50. 50.

    Janbon G, Sherman F, Rustchenko E. Appearance and properties of L-sorbose-utilizing mutants of Candida albicans obtained on a selective plate. Genetics 1999;153:653–64.

    PubMed  CAS  Google Scholar 

  51. 51.

    Perepnikhatka V, Fischer FJ, Niimi M, Baker RA, Cannon RD, Wang YK, et al. Specific chromosome alterations in fluconazole-resistant mutants of Candida albicans. J Bacteriol. 1999;181:4041–9.

    PubMed  CAS  Google Scholar 

  52. 52.

    Pasqualotto AC, Denning DW, Anderson MJ. A cautionary tale: lack of consistency in allele sized between two laboratories for a published multilocus multisatellite typing system. J Clin Microbiol. 2007;45:522–8.

    PubMed  Article  CAS  Google Scholar 

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Acknowledgements

I would like to thank Drs. Tokajian and Saab for their help and constructive criticism of the manuscript.

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Correspondence to Roy A. Khalaf.

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Barada, G., Basma, R. & Khalaf, R.A. Microsatellite DNA Identification and Genotyping of Candida albicans from Lebanese Clinical Isolates. Mycopathologia 165, 115–125 (2008). https://doi.org/10.1007/s11046-008-9089-0

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Keywords

  • Candida albicans
  • Microsatellite genotyping
  • Nosocomial infection
  • Drug resistance