A polymicrobial infection must be suspected if a mixed sequence is observed on the DNA pherogram, and a cloning step with commercialized http://www.selleckchem.com/products/Sunitinib-Malate-(Sutent).html
kits should be performed . Sequencing of all of the clones is time-consuming, but it may help to identify new pathogens. Indeed, a sequence similarity of <97% of the 16S rRNA sequence is the criterion used to define a potentially new bacterial species [43�C45]. The main primers that have been published for the diagnosis of implant-associated bone and joint infections are summarized in Table?1 [5,9]. Although most studies agree about the good specificity of broad-range PCR in the diagnosis of implant-associated bone and joint infections, discrepancies have been reported about its sensitivity, with several studies showing poor sensitivity (<50%) [5,9,40,46�C48]. Several biases could have influenced these data, such as the lack of pretreatment prior to DNA extraction, the lack of control of the extracted DNA quality, or the small number of studied patients . Among the three studies that have involved more than 50 patients, the sensitivity of 16S rRNA PCR for osteoarticular infections (both prosthetic and not) <a href="https://en.wikipedia.org/wiki/Fossariinae
">Fossariinae was determined to be 92.5% vs. 89% for culture  in the first study. The second study reported a PCR sensitivity of 73.3% (53.8% for prosthetic joint infections and 88.2% for infections without prostheses), whereas for cultures, the sensitivity was 96.7% . In the third study on prosthetic joint infections, culture and PCR had similar sensitivities (72.6% and 70.4%) and specificities (98.3% and 97.8%) . For pathogen-specific PCR, the assays should be performed with quantitative real-time PCR in a closed system, in which not only the amplification but also the identification of amplified products with DNA probes with specific annealing within the target-amplified region are coupled in a single vessel, reducing the risk of contamination (Fig.?2) . If fluorescence-labelled oligonucleotide hybridization probes are not used, the identification must be confirmed by sequencing when an amplified product is detected. Overall, this technique has other advantages over conventional PCR, including speed, simplicity, reproducibility, and quantitative capacity. Pathogen-specific PCR http://www.selleckchem.com/screening/pi3k-signaling-inhibitor-library.html
has previously been shown to be more sensitive than 16S rDNA PCR in osteoarticular infections for S.?aureus and M.?tuberculosis [5,50]. Moreover, all of the currently available sequenced bacterial genomes (2848) allowed for the best DNA targets to be chosen for specific assays, with greater sensitivity than that of broad-range PCR . Thus, pathogen-specific PCR is useful not only when a more sensitive diagnosis is needed, but also to confirm the results obtained with broad-range PCR . PCR assays offer several advantages in the diagnosis of implant-associated bone and joint infection, but its use should be restricted, as proposed in Fig.?3.