Staphylococcus aureus, Staphylococcus epidermidis and Staphylococcus haemolyticus: methicillin-resistant isolates are detected directly in blood cultures by multiplex PCR.
In this study, we standardized and evaluated a multiplex-PCR methodology using specific primers to identify Staphylococcus aureus, Staphylococcus epidermidis and Staphylococcus haemolyticus and their methicillin-resistance directly from blood cultures. Staphylococci clinical isolates (149) and control strains (16) previously identified by conventional methods were used to establish the multiplex PCR protocol. Subsequently, this methodology was evaluated using a fast and cheap DNA extraction protocol from 25 staphylococci positive blood cultures. A wash step of the pellet with 0.1% bovine serum albumin (BSA) solution was performed to reduce PCR inhibitors. Amplicons of 154bp (mecA gene), 271bp (S. haemolyticus mvaA gene) and 108 and 124bp (S. aureus and S. epidermidis species-specific fragments, respectively) were observed. Reliable results were obtained for 100% of the evaluated strains, suggesting that this new multiplex-PCR combined with an appropriate DNA-extraction method could be useful in the laboratory for fast and accurate identification of three staphylococci species and simultaneously their methicillin resistance directly in blood cultures.
Pereira
Eliezer M
EM
Laboratório de Infecções Hospitalares, Departamento de Microbiologia Médica, Instituto de Microbiologia, Cidade Universitária Rio de Janeiro, Brazil.
Schuenck
Ricardo P
RP
Malvar
Karoline L
KL
Iorio
Natalia L P
NL
Matos
Pricilla D M
PD
Olendzki
André N
AN
Oelemann
Walter M R
WM
dos Santos
Kátia R N
KR
D001431:Bacteriological Techniques / Q000379:methods* / Q000592:standards; D001769:Blood / Q000382:microbiology*; D017931:DNA Primers / Q000235:genetics; D006801:Humans; D016106:Methicillin Resistance; D016133:Polymerase Chain Reaction / Q000379:methods* / Q000592:standards; D012680:Sensitivity and Specificity; D013203:Staphylococcal Infections / Q000382:microbiology; D013211:Staphylococcus aureus / Q000187:drug effects / Q000235:genetics / Q000302:isolation & purification*; D013212:Staphylococcus epidermidis / Q000187:drug effects / Q000235:genetics / Q000302:isolation & purification*; D041162:Staphylococcus haemolyticus / Q000187:drug effects / Q000235:genetics / Q000302:isolation & purification*
D017931:DNA Primers