This Article
Right arrow Full Text
Right arrow Full Text (PDF)
Right arrow Alert me when this article is cited
Right arrow Alert me if a correction is posted
Services
Right arrow Similar articles in this journal
Right arrow Similar articles in PubMed
Right arrow Alert me to new issues of the journal
Right arrow Download to citation manager
Right arrowReprints and Permissions
Right arrow Copyright Information
Right arrow Books from ASM Press
Right arrow MicrobeWorld
Citing Articles
Right arrow Citing Articles via HighWire
Right arrow Citing Articles via Google Scholar
Google Scholar
Right arrow Articles by Bialek, R.
Right arrow Articles by González, G. M.
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by Bialek, R.
Right arrow Articles by González, G. M.

 Previous Article  |  Next Article 

Journal of Clinical Microbiology, February 2004, p. 778-783, Vol. 42, No. 2
0095-1137/04/$08.00+0     DOI: 10.1128/JCM.42.2.778-783.2004
Copyright © 2004, American Society for Microbiology. All Rights Reserved.

PCR Assays for Identification of Coccidioides posadasii Based on the Nucleotide Sequence of the Antigen 2/Proline-Rich Antigen

Ralf Bialek,1* Jan Kern,1 Tanja Herrmann,1 Rolando Tijerina,2 Luis Ceceñas,3 Udo Reischl,4 and Gloria M. González2

Institute for Tropical Medicine, University Hospital Tübingen,1 Institute of Medical Microbiology, University of Regensburg, Germany,4 Departamento de Microbiología,2 Departamento de Patología, Facultad de Medicina, Universidad Autónoma de Nuevo León, Monterrey, Nuevo León, México3

Received 25 July 2003/ Returned for modification 23 October 2003/ Accepted 8 November 2003

A conventional nested PCR and a real-time LightCycler PCR assay for detection of Coccidioides posadasii DNA were designed and tested in 120 clinical strains. These had been isolated from 114 patients within 10 years in Monterrey, Nuevo Leon, Mexico, known to be endemic for coccidioidomycosis. The gene encoding the specific antigen 2/proline-rich antigen (Ag2/PRA) was used as a target. All strains were correctly identified, whereas DNA from related members of the family Onygenaceae remained negative. Melting curve analysis by LightCycler and sequencing of the 526-bp product of the first PCR demonstrated either 100% identity to the GenBank sequence of the Silveira strain, now known to be C. posadasii (accession number AF013256), or a single silent mutation at position 1228. Length determination of two microsatellite-containing loci (GAC and 621) identified all 120 isolates as C. posadasii. Specific DNA was amplified by conventional nested PCR from three microscopically spherule-positive paraffin-embedded tissue samples, whereas 20 human tissue samples positive for other dimorphic fungi remained negative. Additionally, the safety of each step of a modified commercially available DNA extraction procedure was evaluated by using 10 strains. At least three steps of the protocol were demonstrated to sufficiently kill arthroconidia. This safe procedure is applicable to cultures and to clinical specimens.

* Corresponding author. Mailing address: Institut für Tropenmedizin, Universitätsklinikum Tübingen, Keplerstrasse 15, 72074 Tübingen, Germany. Phone: 49 7071 298 2367. Fax: 49 7071 29 5267. E-mail: ralf.bialek{at}med.uni-tuebingen.de.

Journal of Clinical Microbiology, February 2004, p. 778-783, Vol. 42, No. 2
0095-1137/04/$08.00+0     DOI: 10.1128/JCM.42.2.778-783.2004
Copyright © 2004, American Society for Microbiology. All Rights Reserved.



This article has been cited by other articles:

  • Brilhante, R. S. N., Cordeiro, R. A., Rocha, M. F. G., Fechine, M. A. B., Furtado, F. M., Nagao-Dias, A. T., Camargo, Z. P., Sidrim, J. J. C. (2008). Coccidioidal pericarditis: a rapid presumptive diagnosis by an in-house antigen confirmed by mycological and molecular methods. J Med Microbiol 57: 1288-1292 [Abstract] [Full Text]  
  • Martin-Davila, P., Fortun, J., Lopez-Velez, R., Norman, F., Montes de Oca, M., Zamarron, P., Gonzalez, M. I., Moreno, A., Pumarola, T., Garrido, G., Candela, A., Moreno, S. (2008). Transmission of Tropical and Geographically Restricted Infections during Solid-Organ Transplantation. Clin. Microbiol. Rev. 21: 60-96 [Abstract] [Full Text]  
  • Gonzalez, G. M., Gonzalez, G., Najvar, L. K., Graybill, J. R. (2007). Therapeutic efficacy of caspofungin alone and in combination with amphotericin B deoxycholate for coccidioidomycosis in a mouse model. J Antimicrob Chemother 60: 1341-1346 [Abstract] [Full Text]  
  • Saubolle, M. A., McKellar, P. P., Sussland, D. (2007). Epidemiologic, Clinical, and Diagnostic Aspects of Coccidioidomycosis. J. Clin. Microbiol. 45: 26-30 [Full Text]  
  • Binnicker, M. J., Buckwalter, S. P., Eisberner, J. J., Stewart, R. A., McCullough, A. E., Wohlfiel, S. L., Wengenack, N. L. (2007). Detection of Coccidioides Species in Clinical Specimens by Real-Time PCR. J. Clin. Microbiol. 45: 173-178 [Abstract] [Full Text]  
  • Umeyama, T., Sano, A., Kamei, K., Niimi, M., Nishimura, K., Uehara, Y. (2006). Novel Approach to Designing Primers for Identification and Distinction of the Human Pathogenic Fungi Coccidioides immitis and Coccidioides posadasii by PCR Amplification.. J. Clin. Microbiol. 44: 1859-1862 [Abstract] [Full Text]  
  • Espy, M. J., Uhl, J. R., Sloan, L. M., Buckwalter, S. P., Jones, M. F., Vetter, E. A., Yao, J. D. C., Wengenack, N. L., Rosenblatt, J. E., Cockerill, F. R. III, Smith, T. F. (2006). Real-Time PCR in Clinical Microbiology: Applications for Routine Laboratory Testing. Clin. Microbiol. Rev. 19: 165-256 [Abstract] [Full Text]  
  • Bialek, R, Konrad, F, Kern, J, Aepinus, C, Cecenas, L, Gonzalez, G M, Just-Nubling, G, Willinger, B, Presterl, E, Lass-Florl, C, Rickerts, V (2005). PCR based identification and discrimination of agents of mucormycosis and aspergillosis in paraffin wax embedded tissue. J. Clin. Pathol. 58: 1180-1184 [Abstract] [Full Text]  


Copyright © 2009 by the American Society for Microbiology.   For an alternate route to Journals.ASM.org, visit: http://intl-journals.asm.org | More Info»