Identifying Fungi Quickly and Accurately: The Strengths of Molecular Biology Technology
Fungi are organisms classified under the kingdom Fungi and play a crucial role in ecosystems as decomposers or recyclers of organic matter that has died, such as leaves, wood, and animals.Fungi can be microscopic, like yeasts and molds, or macroscopic, like mushrooms commonly found in nature. They reproduce through spores, which can be dispersed through the air, water, or other organisms.
Although fungi play an important role in ecosystems, many types of fungi can be harmful to humans, causing various health issues, damage to crops, and degradation of building materials. For example, fungi like Aspergillus flavus and Aspergillus parasiticus can grow and contaminate food, producing mycotoxins that are highly dangerous to human health, as well as reducing the quality and safety of food products. Some fungi can also become invasive species threatening local biodiversity by attacking native species. For instance, the fungus Batrachochytrium dendrobatidis, which causes chytridiomycosis, has significantly reduced biodiversity and even wiped out amphibian populations worldwide.
Therefore, identifying fungi is a crucial step in various fields such as food safety, agriculture, medicine, and biotechnology. In medicine, undetected or misidentified fungal infections can be fatal, especially for patients with weakened immune systems. In biotechnology, the use of fungi as sources of enzymes, antibiotics, and other bioactive compounds requires a deep understanding of the fungal species used.
Molecular Biology Testing Methods for Fungi
Traditionally, fungal identification has been done by observing morphological characteristics such as spore shape, colony color, and hyphal structure. Although this method has been used for many years, there are significant limitations, particularly concerning accuracy. Many fungal species have very similar morphologies, leading to identification errors. Moreover, some fungi require specific conditions to exhibit their morphological characteristics, which can take a long time to observe in the laboratory.
These limitations have driven the development of molecular biology-based identification methods that enhance the accuracy and speed of fungal identification. The process generally involves several stages and can vary depending on the specific methods used. However, the common steps include isolating DNA from the fungal sample, amplifying the DNA using PCR, and sequencing the DNA, which can be done using Sanger sequencing or other techniques such as Next-Generation Sequencing (NGS). The obtained DNA sequences are then analyzed using bioinformatics to provide specific information for identifying fungi down to the species level.
One well-known molecular biology technique for identifying fungi is DNA barcoding, which works by identifying a specific DNA segment that varies significantly among different fungal species but remains consistent within the same species. For fungi, the most commonly used DNA segment is the Internal Transcribed Spacer (ITS) region of ribosomal DNA because it has high variability among fungal species, allowing accurate identification to the species level. To isolate ITS from a fungal sample, DNA extraction must be performed first. After obtaining the DNA isolate, the next step is to amplify the ITS segment using Polymerase Chain Reaction (PCR). Universal primers, such as ITS1 and ITS4, are used to specifically target, bind, and amplify the ITS segment from the fungal DNA. Sequencing is then performed to obtain the specific nucleotide sequence. This sequencing result is compared with reference data in genetic databases, such as GenBank or specialized fungal databases, to determine the match and accurately identify the species.
Validation of the results obtained through additional tests or confirmation methods, such as PCR with more specific primers after the fungal species is known, is also crucial to ensure that the identification is accurate and consistent.
With advancements in molecular biology techniques, particularly DNA barcoding, the process of fungal identification has become more efficient and precise. This technique enables high-accuracy detection and classification of fungal species, addressing the limitations of traditional methods and ensuring that the information obtained is reliable.
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REFERENCE
Campbell, C. K., & Johnson, E. M. (2013). Identification of pathogenic fungi. John Wiley & Sons.
Fisher, M. C., Garner, T. W., & Walker, S. F. (2009). Global emergence of Batrachochytrium dendrobatidis and amphibian chytridiomycosis in space, time, and host. Annual review of microbiology, 63(1), 291-310.
Humber, R. A. (1997). Fungi: identification. In Manual of techniques in insect pathology (pp. 153-185). Academic press.
Raja, H. A., Miller, A. N., Pearce, C. J., & Oberlies, N. H. (2017). Fungal identification using molecular tools: a primer for the natural products research community. Journal of natural products, 80(3), 756-770.
Xu, J. (2016). Fungal DNA barcoding. Genome, 59(11), 913-932.
