Prime Journal of Microbiology Research
2, Issue 5, pp.
© Prime Journals
New Rapid Method for
Differentiation of MRSA and SSA by PCR Restriction Analysis of
920 bp of Dnaj gene
Zeinab H. Helal
Fatma Alzahraa M. Gomaa and Sahar Radwan
Microbiology and Immunology Department, Faculty
of Pharmacy, Al-Azhar University, Nasr City, Cairo, Egypt.
Accepted 18th October, 2012
Methicillin-resistant S. aureus (MRSA) and sensitive
S. aureus (SSA) are responsible for a high proportion of
nosocomial infections, which makes difficulty in treatment. MRSA
infections are responsible for increased mortality rates, longer
lengths of hospital stay, and higher rates of treatment failure
compared to SSA infections. Detection of MRSA by conventional
method is time consuming, influenced by culture medium,
concentration of NaCl, temperature, time of incubation and
antibiotics. Various PCR methods had been applied for the rapid
detection and identification of Staphylococcus species.
The dnaJ gene sequence is potentially useful for the
identification of genetically related Staphylococcus
species and subspecies. While, with other bacterium PCR-restriction
analysis is preferred, as a simple and cost-effective method
that does not involve radioisotopes. For that reason in this
study, we established and evaluated a rapid new protocol of
identifying clinically relevant MRSA species by PCR- restriction
analysis of the dnaJ gene. SSA and MRSA strains were
isolated during a one year period from patients with bacterimia.
Identification of S. aureus was performed by
standard laboratory methods. Resistance to methicillin was
detected by disc diffusion susceptibility test. DNA extraction
was performed for both clinical blood samples as well as from
isolated SSA and MRSA. Primers were designed to amplify specific
dnaJ gene target and confirming the presence of S.
aureus. MRSA was speciated by PCR-restriction analysis of
dnaJ gene using XapI restriction enzyme. Our results
showed two distinguished patterns of PCR-restriction analysis
for SSA and MRSA. Only SSA is known to have a XapI restriction
sites. Using our protocol, we were able to demonstrate the
existence of staphylococcus and to identify their methicillin
resistance. Therefore, we suggest that it would be very useful
to apply PCR amplification restriction analysis to dnaJ
gene directly to clinical specimens early in the diagnostic
process. This would save several days that are required for
conventional culture. Thus, this established protocol is
suggested as a simple and useful method for the rapid detection
and simultaneous identification of MRSA in primary clinical
specimens or for the identification of culture isolates. This
rapid detection would allow clinicians initially to avoid
potentially inappropriate treatment options.
Djna gene, Methicillin-resistant Staphylococcus aureus (MRSA),
Sensitive Staphylococcus aureus (SSA).
See Full Article [pdf]