title: Discrimination of methicillin-resistant Staphylococcus aureus from methicillin-susceptible Staphylococcus aureus or coagulase-negative staphylococci by detection of penicillin-binding protein 2 and penicillin-binding protein 2' using a bioluminescent enzyme immunoassay.
abstract: For the discrimination of methicillin-resistant S. aureus (MRSA) from methicillin-susceptible S. aureus (MSSA) or coagulase-negative staphylococci (CNS), we developed a bioluminescent enzyme immunoassay (BLEIA) for detecting penicillin-binding protein 2 (PBP2) and penicillin-binding protein 2' (PBP2') using biotinylated firefly luciferase. The BLEIA was able to detect recombinant PBP2 at 50 pg/ml and recombinant PBP2' at 500 pg/ml. PBP2 and PBP2' present in the membranes of S. aureus were extracted by acid and detergent treatment. The method was able to detect PBP2 or PBP2' extracted from 10(6) colony forming units of S. aureus because of efficient extraction and the high sensitivity of luciferase. In a study of clinical isolates previously characterized as either MRSA or MSSA by antibiotic susceptibility testing, all 34 specimens identified as MRSA were both PBP2 and PBP2' positive. The 34 MSSA specimens were PBP2 positive and PBP2' negative. Moreover, the BLEIA could detect PBP2' extracted from four species of methicillin-resistant CNS, but not PBP2 extracted from four species of methicillin-resistant and methicillin-susceptible CNS. This result suggested that PBP2 could be a unique marker for discrimination of S. aureus from CNS. A BLEIA that is able to detect PBP2 and PBP2' may be useful in clinical diagnostics.
authors.affiliation: Research and Development Division, Kikkoman Corporation, 399 Noda, Noda City, Chiba 278-0037, Japan. ksiga@mail.kikkoman.co.jp
authors.forename: Kazuki
authors.initials: K
authors.lastname: Shiga
authors.forename: Keiko
authors.initials: K
authors.lastname: Gomi
authors.forename: Motoi
authors.initials: M
authors.lastname: Nishimura
authors.forename: Masaharu
authors.initials: M
authors.lastname: Watanabe
authors.forename: Fumio
authors.initials: F
authors.lastname: Nomura
authors.forename: Naoki
authors.initials: N
authors.lastname: Kajiyama
mesh_terms: D003030:Coagulase / Q000172:deficiency / Q000378:metabolism; D006801:Humans; D007124:Immunoenzyme Techniques / Q000379:methods*; D008156:Luciferases / Q000737:chemistry*; D008163:Luminescent Measurements / Q000379:methods; D055624:Methicillin-Resistant Staphylococcus aureus / Q000737:chemistry / Q000187:drug effects / Q000201:enzymology / Q000302:isolation & purification*; D046915:Penicillin-Binding Proteins / Q000032:analysis*; D013203:Staphylococcal Infections / Q000382:microbiology; D013211:Staphylococcus aureus / Q000187:drug effects / Q000201:enzymology / Q000302:isolation & purification
chemical_list: D003030:Coagulase; D046915:Penicillin-Binding Proteins; D008156:Luciferases