title: A simple polymerase chain reaction/restriction fragment length polymorphism assay capable of identifying medically relevant filamentous fungi.
abstract: Because of the accumulating evidence that suggests that numerous unhealthy conditions in the indoor environment are the result of abnormal growth of the filamentous fungi (mold) in and on building surfaces, it is necessary to accurately reflect the organisms responsible for these maladies and to identify them in precise and timely manner. To this end, we have developed a method that is cost effective, easy to perform, and accurate. We performed a simple polymerase chain reaction/restriction fragment length polymorphism (PCR/RFLP) analysis on multiple members of species known to negatively influence the indoor environment. The genera analyzed were Stachybotrys, Penicillium, Aspergillus, and Cladosporium. Each organism underwent PCR with universal primers that amplified ribosomal sequences generating products from 550 to 600 bp followed by enzymatic digestion with EcoRI, HaeIII, MspI, and HinfI. Our results show that using this combination of restriction enzymes enables the identification of these fungal organisms at the species level.
authors.affiliation: National Risk Management Research Laboratory, U.S. Environmental Protection Agency, 109 T.W. Alexander Drive, Research Triangle Park, NC 27711, USA. dean.timothy@epa.gov
authors.forename: Timothy R
authors.initials: TR
authors.lastname: Dean
authors.forename: Michael
authors.initials: M
authors.lastname: Kohan
authors.forename: Doris
authors.initials: D
authors.lastname: Betancourt
authors.forename: Marc Y
authors.initials: MY
authors.lastname: Menetrez
mesh_terms: D001230:Aspergillus / Q000235:genetics; D002963:Cladosporium / Q000235:genetics; D004262:DNA Restriction Enzymes; D005658:Fungi / Q000145:classification / Q000235:genetics* / Q000302:isolation & purification; D010407:Penicillium / Q000235:genetics; D016133:Polymerase Chain Reaction / Q000379:methods*; D012150:Polymorphism, Restriction Fragment Length; D017422:Sequence Analysis, DNA; D018877:Sick Building Syndrome; D013172:Spores, Fungal / Q000235:genetics* / Q000302:isolation & purification; D013191:Stachybotrys / Q000235:genetics
chemical_list: D004262:DNA Restriction Enzymes