Comparison of DNA Microarray, Loop-Mediated Isothermal Amplification (LAMP) and Real-Time PCR with DNA Sequencing for Identification of Fusarium spp. Obtained from Patients with Hematologic Malignancies.
The performance of three molecular biology techniques, i.e., DNA microarray, loop-mediated isothermal amplification (LAMP), and real-time PCR were compared with DNA sequencing for properly identification of 20 isolates of Fusarium spp. obtained from blood stream as etiologic agent of invasive infections in patients with hematologic malignancies. DNA microarray, LAMP and real-time PCR identified 16 (80%) out of 20 samples as Fusarium solani species complex (FSSC) and four (20%) as Fusarium spp. The agreement among the techniques was 100%. LAMP exhibited 100% specificity, while DNA microarray, LAMP and real-time PCR showed 100% sensitivity. The three techniques had 100% agreement with DNA sequencing. Sixteen isolates were identified as FSSC by sequencing, being five Fusarium keratoplasticum, nine Fusarium petroliphilum and two Fusarium solani. On the other hand, sequencing identified four isolates as Fusarium non-solani species complex (FNSSC), being three isolates as Fusarium napiforme and one isolate as Fusarium oxysporum. Finally, LAMP proved to be faster and more accessible than DNA microarray and real-time PCR, since it does not require a thermocycler. Therefore, LAMP signalizes as emerging and promising methodology to be used in routine identification of Fusarium spp. among cases of invasive fungal infections.
Department of Internal Medicine, School of Medicine, University of Campinas, Campinas, São Paulo, Brazil.|LIM 46 - Laboratory of Parasitology, HC/FMUSP, São Paulo, Brazil.
Marcela
M
de Souza
Medical Mycology Research Center, Chiba University, Chiba, Japan.|Tetsuhiro Matsuzawa, University of Nagasaki, Nagasaki, Japan.
Tetsuhiro
T
Matsuzawa
Medical Mycology Research Center, Chiba University, Chiba, Japan.
Kanae
K
Sakai
Medical Mycology Research Center, Chiba University, Chiba, Japan.
Yasunori
Y
Muraosa
Department of Clinical Pathology, School of Medicine, University of Campinas, Campinas, São Paulo, Brazil.
Luzia
L
Lyra
Department of Internal Medicine, School of Medicine, University of Campinas, Campinas, São Paulo, Brazil.
Ariane Fidelis
AF
Busso-Lopes
Division of Infectious Diseases, School of Medicine, University of São Paulo, São Paulo, Brazil.
Anna Sara Shafferman
ASS
Levin
Department of Clinical Pathology, School of Medicine, University of Campinas, Campinas, São Paulo, Brazil.
Angélica Zaninelli
AZ
Schreiber
Medical Mycology Research Center, Chiba University, Chiba, Japan.
Yuzuru
Y
Mikami
Medical Mycology Research Center, Chiba University, Chiba, Japan.
Tohoru
T
Gonoi
Medical Mycology Research Center, Chiba University, Chiba, Japan.
Katsuhiko
K
Kamei
Department of Internal Medicine, School of Medicine, University of Campinas, Campinas, São Paulo, Brazil.
Maria Luiza
ML
Moretti
Department of Internal Medicine, School of Medicine, University of Campinas, Campinas, São Paulo, Brazil. trabasso@fcm.unicamp.br.
Plínio
P
Trabasso
D016469:Fungemia / Q000175:diagnosis / Q000382:microbiology; D060585:Fusariosis / Q000175:diagnosis* / Q000382:microbiology; D005670:Fusarium / Q000145:classification / Q000235:genetics / Q000302:isolation & purification*; D019337:Hematologic Neoplasms / Q000150:complications*; D006801:Humans; D025202:Molecular Diagnostic Techniques / Q000379:methods*; D021141:Nucleic Acid Amplification Techniques / Q000379:methods*; D009693:Nucleic Acid Hybridization / Q000379:methods*; D012680:Sensitivity and Specificity; D017422:Sequence Analysis, DNA / Q000379:methods*
Filamentous fungi; Fungal infection; Fusarium; Molecular methods