Molecular Techniques For Resistance Detection

Currently Employed Detection Method

The current method used in the detection of QRDR mutations causing fluoroquinolone resistance is automated DNA sequencing. Most studies amplify only the QRDRs of ParC and GyrA, whereas other studies also examine the QRDRs of ParE and GyrB. There are various published primer sets and amplification conditions used in

these studies. " 1 After purification and quantitation of the amplification products, the sequencing reactions are conducted.1-10,12,131 DNA sequencing is performed using an ABI PRISM BigDye Terminator kit and run on an ABI PRISM sequencer.12,7,10,12,131

Novel Detection Methods

As previously mentioned, the utility of susceptibility studies for fluoroquinolone resistance mutation detection has been evaluated. Although this method was able to correctly identify isolates without resistance mutations and those with mutations in both ParC and GyrA, it was unable to identify isolates with only ParC mutations.[71 Susceptibility testing as a method of fluoroquinolone resistance detection is not sufficient for clinical use as S. pneumoniae isolates with ParC mutations are highly clinically significant.

A novel detection method for QRDR mutations has recently been developed: PCR-oligonucleotide ligation assays. Similar to sequencing studies, this method uses PCR to amplify the QRDRs of ParC and GyrA.[21 Oligonucleotide ligation assays are conducted and the specific mutations are detected colometrically.[21 This method correctly identified the majority (98%) of wildtype isolates and isolates with QRDR mutations in one or both target genes.[21 The substitutions correctly identified by this technique include the most common QRDR mutations (ParC: Ser79Phe/Tyr, Asp83Tyr/Val/ Asn; GyrA: Ser81Phe/Ala/Tyr, Glu85Lys).[21 Rare mutations were also reported to be identifiable by this method.[21 The PCR-oligonucleotide ligation assay is much more rapid than DNA sequencing because it requires 12 hr for completion as opposed to 33 hr.[21 This novel, rapid, and accurate method is a good advancement in the detection of fluoroquinolone resistance mutations. However, the robustness of PCR-oligonucleotide ligation assays must be confirmed with studies of many more isolates to become as accepted as the DNA sequencing gold standard.

Getting Started With Dumbbells

Getting Started With Dumbbells

The use of dumbbells gives you a much more comprehensive strengthening effect because the workout engages your stabilizer muscles, in addition to the muscle you may be pin-pointing. Without all of the belts and artificial stabilizers of a machine, you also engage your core muscles, which are your body's natural stabilizers.

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