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Paper of the week: Polymerase chain reaction assay using the restriction fragment length polymorphism technique in the detection of prosthetic joint infections: A Multi-Centered Study

Paper of the week: Polymerase chain reaction assay using the restriction fragment length polymorphism technique in the detection of prosthetic joint infections: A Multi-Centered Study. Moshirabadi A, Razi M, Arasteh P, Sarzaeem MM, Ghaffari S, Aminiafshar S, Hosseinian Khosroshahy K, Sheikholeslami FM. J Arthroplasty. 2018 Oct 25. pii: S0883-5403(18)31057-X. doi: 10.1016/j.arth.2018.10.017. [Epub ahead of print].

Summary and Editorial by Sreeram Penna

The main aim of this prospective study was to assess the diagnostic accuracy of polymerase chain reaction (PCR) using RFLP (restriction fragment length polymorphism) method. Researchers also obtained bacterial cultures at the same time. The study assessed 76 samples using this technique. International consensus meeting criteria were used to identify prosthetic joint infection. 50% of the samples were deemed infected based on the above criteria. Results showed that using PCR RFLP Sensitivity and specificity was found to be 97.4% and 100% respectively. This was superior compared to the culture where sensitivity and specificity was 31.6% and 100%. Researchers isolated a broad range of bacteria including fastidious organisms like Chlamydophila pneumonia, Stenotrophomonas maltophilia, Brucella melitensis. One advantage of this technique is the amount of time required to get the pathogen identification is approximately 3 to 4 hours compared to multiple days for microbiological culture methods.

Restriction fragment length polymorphism (RFLP) is a difference in homologous DNA sequences which are identified by the different length of sequences after digestion of DNA samples using specific restriction endonucleases. RFLP probes are widely used in genome mapping and variation analysis such as genotyping, forensics, paternity tests, hereditary disease diagnostics, etc. This process requires a large amount of DNA and is labor intensive.[1] Combining PCR along with RFLP (also called cleaved amplified polymorphic sequences or CAPS) solves the problem of the requirement of a large sample.[2] Using PCR RFLP method with 16s bacterial DNA has been used in bacterial identification in clinical situations, food safety and also identify different strains of bacteria.[3–6] Rohit et al., used this technique to rapidly diagnose bacterial species in the setting of neonatal sepsis.[3] This study provides importance of such technique in PJI setting where it is very important to identify pathogens as it has huge implications in the management.

References

[1] Restriction Fragment Length Polymorphism (RFLP) n.d. https://www.ncbi.nlm.nih.gov/probe/docs/techrflp/ (accessed December 17, 2018).

[2] Cleaved Amplified Polymorphic Sequences (CAPS) n.d. https://www.ncbi.nlm.nih.gov/probe/docs/techcaps/ (accessed December 17, 2018).

[3] Rohit A, Maiti B, Shenoy S, Karunasagar I. Polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) for rapid diagnosis of neonatal sepsis. Indian J Med Res 2016;143:72–8. doi:10.4103/0971-5916.178613.

[4] Schütte UME, Abdo Z, Bent SJ, Shyu C, Williams CJ, Pierson JD, et al. Advances in the use of terminal restriction fragment length polymorphism (T-RFLP) analysis of 16S rRNA genes to characterize microbial communities. Appl Microbiol Biotechnol 2008;80:365–80. doi:10.1007/s00253-008-1565-4.

[5] Meyer R, Höfelein C, Lüthy J, Candrian U. Polymerase chain reaction-restriction fragment length polymorphism analysis: a simple method for species identification in food. J AOAC Int 1995;78:1542–51.

[6] González A, Moreno Y, González R, Hernández J, Ferrús MA. Development of a simple and rapid method based on polymerase chain reaction-based restriction fragment length polymorphism analysis to differentiate Helicobacter, Campylobacter, and Arcobacter species. Curr Microbiol 2006;53:416–21. doi:10.1007/s00284-006-0168-5.