FUSARIUM DISCRIMINATION USING AUTOMATED REP PCR DNA FINGERPRINTING

Kontoyiannis DP 1 , Reece K 2 , Healy M

Author address: 

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Abstract: 

Background: Infections caused by Fusarium species are increasing in frequency among immunocompromised hosts. These infections are collectively referred to as fusariosis and are often associated with endophthalmitis, cutaneous infections, pulmonary infections, central venous catheter infections, hematological malignancies, and transplant patients. Confirmation of suspected clinical disease currently presents a challenge to the clinician, with difficulty in making diagnosis and frequently delaying treatment. The identification of these infections is very poor and death occurs in up to 80% of the cases despite antifungal therapy. The increase of infections and the severity of the outcome now requires a quick and accurate method of species discrimination. Traditional testing is based on colony morphology and serological testing and molecular methods, such as sequencing, are laborious. These methods of identifying Fusarium species do not differentiate between strains. This study uses automated repetitive sequence-based PCR (rep-PCR) to demonstrate discrimination of Fusarium species and strains. Materials and Methods: DNA was extracted from 15 Fusarium isolates; including several different species, such as F. solani, F. oxysporum, F. proliferatum, and F.pallidorum, previously characterized by 16S RNA gene sequencing. Fifty nanograms of DNA was amplified using the DiversiLab DNA Fingerprinting Kit. The amplicons were separated using lab-on-a-chip technology and the Caliper ® 1000 Analyzer. Results were analyzed using the system software. Results: The results showed multiple, visually distinct fingerprint patterns. The software classified the organisms into the appropriate Fusarium species groups, concordant with previous characterization. In addition, subtype differences within samples of the same species were also noted by variations in the patterns and as were displayed by the dendrogram indicating pattern similarities. Conclusions: The results indicate that the DiversiLab System can be a useful tool to differentiate Fusarium isolates to the species level as well as differentiate samples within a subspecies. Because of the differences seen within a species, this approach shows strong promise for studying the epidemiology of fusariosis.
2003

abstract No: 

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Full conference title: 

The 15 th Congress of the International Society for Human and Animal Mycology
    • ISHAM 15th (2003)