Microsatellite based typing of Aspergillus fumigatus: Strengths, pitfalls and solutions

• Published in: Journal of microbiological methods Vol. 69; no. 2; pp. 268 - 272
• Main Authors: de Valk, Hanneke A., Meis, Jacques F.G.M., Klaassen, Corné H.W.
• Format: Journal Article
• Language: English
• Published: Shannon Elsevier B.V 01.05.2007; Elsevier Science
• Subjects: Allelic ladder; Aspergillus fumigatus; Aspergillus fumigatus - classification; Aspergillus fumigatus - genetics; Biological and medical sciences; DNA, Fungal - chemistry; DNA, Fungal - genetics; Electrophoresis, Capillary; Fundamental and applied biological sciences. Psychology; Growth, nutrition, metabolism, transports, enzymes. Molecular biology; Microbiology; Microsatellite Repeats; Mycology; Polymerase Chain Reaction - methods; Reproducibility of Results; Short tandem repeat; Sizing; Standardization; Typing; Typing; Aspergillus fumigatus; Sizing; Allelic ladder; Short tandem repeat; Standardization; Fungi; Allele; Typing; Standardization; Short tandem repeat; Allelic ladder; Sizing; Microsatellite DNA; Size; Fungi Imperfecti; Tandemly repeated sequence; Thallophyta
• ISSN: 0167-7012; 1872-8359
• DOI: 10.1016/j.mimet.2007.01.009

Microsatellites, or short tandem repeats (STR's), are popular tools to discriminate between microbial isolates. Here, we report on the robustness of a microsatellite panel for discrimination of Aspergillus fumigatus isolates. Two major PCR artefacts (stutter peaks and minus-A peaks) can complicate correct interpretation of STR data. We investigated the effect of alterations to the various components of the PCR amplification mixtures on these PCR artefacts and on the reproducibility of this assay. Some extreme conditions led to a loss of signal, but, under all conditions where a signal was obtained, identical typing results were produced. Furthermore, pitfalls with the exchange of results between labs are discussed. These pitfalls are primarily associated with sizing of the obtained PCR fragments.