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 Kumeda, Y.
Right arrow Articles by Asao, T.
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by Kumeda, Y.
Right arrow Articles by Asao, T.
Agricola
Right arrow Articles by Kumeda, Y.
Right arrow Articles by Asao, T.

 Previous Article  |  Next Article 

Applied and Environmental Microbiology, September 2001, p. 4084-4090, Vol. 67, No. 9
0099-2240/01/$04.00+0   DOI: 10.1128/AEM.67.9.4084-4090.2001
Copyright © 2001, American Society for Microbiology. All rights reserved.

Heteroduplex Panel Analysis, a Novel Method for Genetic Identification of Aspergillus Section Flavi Strains

Yuko Kumeda and Tsutomu Asao*

Osaka Prefectural Institute of Public Health, Nakamichi 1-3-69, Higashinari-ku, Osaka 537-0025, Japan

Received 10 April 2001/Accepted 3 July 2001

For genetic identification of Aspergillus Section Flavi isolates and detection of intraspecific variation, we developed a novel method for heteroduplex panel analysis (HPA) utilizing fragments of the internal transcribed spacer (ITS) regions (ITS1-5.8S-ITS2) of the rRNA gene that was PCR amplified with universal primers. The method involves formation of heteroduplexes with a set of reference fragments amplified from Aspergillus flavus, A. parasiticus, A. tamarii, and A. nomius and subsequent minislab vinyl polymer gel electrophoresis. The test panel is compared with species-specific standard panels (F-1, P-1, T-1, and N-1) generated by pairwise reannealing among four reference fragments. Of 90 test panels, 89 succeeded in identifying the species and 74 were identical to one of the four standard panels. Of the 16 new panels, 11 A. flavus/A. oryzae panels were identical and typed as F-2 and 4 of 5 A. nomius panels were typed as N-2 or N-3. The other strain, A. nomius IMI 358749, was unable to identify the species because no single bands were formed with any of the four reference strains. DNA sequencing revealed that our HPA method has the highest sensitivity available and is able to detect as little as one nucleotide of diversity within the species. When Penicillium or non-Section Flavi Aspergillus was subjected to HPA, the resulting bands of heteroduplexes showed apparently lower mobility and poor heteroduplex formation. This indicates that HPA is a useful identification method without morphological observation and is suitable for rapid and inexpensive screening of large numbers of isolates. The HPA typing coincided with the taxonomy of Section Flavi and is therefore applicable as an alternative to the conventional methods (Samson, R. A., E. S. Hoekstra, J. C. Frisvad, and O. Filtenborg, p. 64-97, in Introduction to Food- and Airborne Fungi, 6th ed., 2000).

* Corresponding author. Mailing address: Osaka Prefectural Institute of Public Health, Nakamichi 1-3-69, Higashinari-ku, Osaka, 537-0025, Japan. Phone: 81-6-6972-1321. Fax: 81-6-6972-1329. E-mail: asao{at}iph.pref.osaka.jp.

Applied and Environmental Microbiology, September 2001, p. 4084-4090, Vol. 67, No. 9
0099-2240/01/$04.00+0   DOI: 10.1128/AEM.67.9.4084-4090.2001
Copyright © 2001, American Society for Microbiology. All rights reserved.


This article has been cited by other articles:

  • Manikandan, P., Varga, J., Kocsube, S., Samson, R. A., Anita, R., Revathi, R., Doczi, I., Nemeth, T. M., Narendran, V., Vagvolgyi, C., Manoharan, C., Kredics, L. (2009). Mycotic Keratitis Due to Aspergillus nomius. J. Clin. Microbiol. 47: 3382-3385 [Abstract] [Full Text]  
  • Gibbons, J. G., Rokas, A. (2009). Comparative and Functional Characterization of Intragenic Tandem Repeats in 10 Aspergillus Genomes. Mol Biol Evol 26: 591-602 [Abstract] [Full Text]  
  • Kredics, L., Varga, J., Kocsube, S., Doczi, I., Samson, R. A., Rajaraman, R., Narendran, V., Bhaskar, M., Vagvolgyi, C., Manikandan, P. (2007). Case of Keratitis Caused by Aspergillus tamarii. J. Clin. Microbiol. 45: 3464-3467 [Abstract] [Full Text]  
  • Rokas, A., Payne, G., Fedorova, N.D., Baker, S.E., Machida, M., Yu, J., Georgianna, D. R., Dean, R. A., Bhatnagar, D., Cleveland, T.E., Wortman, J.R., Maiti, R., Joardar, V., Amedeo, P., Denning, D.W., Nierman, W.C. (2007). What can comparative genomics tell us about species concepts in the genus Aspergillus?. SIM 59: 11-17 [Abstract] [Full Text]  
  • Hinrikson, H. P., Hurst, S. F., Lott, T. J., Warnock, D. W., Morrison, C. J. (2005). Assessment of Ribosomal Large-Subunit D1-D2, Internal Transcribed Spacer 1, and Internal Transcribed Spacer 2 Regions as Targets for Molecular Identification of Medically Important Aspergillus Species. J. Clin. Microbiol. 43: 2092-2103 [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»