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Identification of medically important Candida and non-Candida yeast species by an oligonucleotide array
Shiang Ning Leaw1, Hsien Chang Chang, Richard Barton
1Institute of Biomedical Engineering, National Cheng Kung University, Tainan, Taiwan, Republic of China.
Abstract:
The incidence of yeast infections has increased in the recent decades, with Candida albicans still being the most common cause of infections. However, infections caused by less common yeasts have been widely reported in recent years. Based on the internal transcribed spacer 1 (ITS 1) and ITS 2 sequences of the rRNA genes, an oligonucleotide array was developed to identify 77 species of clinically relevant yeasts belonging to 16 genera. The ITS regions were amplified by PCR with a pair of fungus-specific primers, followed by hybridization of the digoxigenin-labeled PCR product to a panel of oligonucleotide probes immobilized on a nylon membrane for species identification. A collection of 452 yeast strains (419 target and 33 nontarget strains) was tested, and a sensitivity of 100% and a specificity of 97% were obtained by the array. The detection limit of the array was 10 pg of yeast genomic DNA per assay. In conclusion, yeast identification by the present method is highly reliable and can be used as an alternative to the conventional identification methods. The whole procedure can be finished within 24 h, starting from isolated colonies.
Insights
A new oligonucleotide array accurately identifies 77 yeast species using internal transcribed spacer (ITS) DNA sequences. This rapid molecular method offers a reliable alternative for clinical yeast identification within 24 hours.
Area of Science:
- Medical Mycology
- Molecular Diagnostics
- Microbiology
Background:
- Rising incidence of yeast infections, including those caused by less common species.
- Need for accurate and rapid identification of clinically relevant yeasts.
- Limitations of conventional yeast identification methods.
Purpose of the Study:
- To develop and evaluate an oligonucleotide array for identifying 77 clinically relevant yeast species.
- To utilize internal transcribed spacer (ITS) regions of rRNA genes for species-specific detection.
- To provide a rapid and reliable alternative to traditional yeast identification techniques.
Main Methods:
- PCR amplification of ITS 1 and ITS 2 regions using fungus-specific primers.
- Hybridization of digoxigenin-labeled PCR products to immobilized oligonucleotide probes on a nylon membrane.
- Testing with 452 yeast strains (419 target, 33 non-target).
Main Results:
- The oligonucleotide array achieved 100% sensitivity and 97% specificity.
- The detection limit was as low as 10 pg of yeast genomic DNA.
- The entire procedure, from colony to identification, can be completed within 24 hours.
Conclusions:
- The developed oligonucleotide array is a highly reliable method for yeast identification.
- This molecular approach serves as a valuable alternative to conventional diagnostic methods.
- The rapid turnaround time makes it suitable for clinical settings.
