Analysis of 525 Samples To Determine the Usefulness of PCR Amplification and Sequencing of the 16S rRNA Gene for Diagnosis of Bone and Joint Infections

Abstract

The 16S rRNA gene PCR in the diagnosis of bone and joint infections has not been systematically tested. Five hundred twenty-five bone and joint samples collected from 525 patients were cultured and submitted to 16S rRNA gene PCR detection of bacteria in parallel. The amplicons with mixed sequences were also cloned. When discordant results were observed, culture and PCR were performed once again. Bacteria were detected in 139 of 525 samples. Culture and 16S rRNA gene PCR yielded identical documentation in 475 samples. Discrepancies were linked to 13 false-positive culture results, 5 false-positive PCR results, 9 false-negative PCR results, 16 false-negative culture results, and 7 mixed infections. Cloning and sequencing of 16S rRNA gene amplicons in 6 of 8 patients with mixed infections identified 2 to 8 bacteria per sample. Rarely described human pathogens such as Alcaligenes faecalis, Comamonas terrigena, and 21 anaerobes were characterized. We also detected, by 16S rRNA gene PCR, four previously identified bacteria never reported in human infection, Alkanindiges illinoisensis, dehydroabietic acid-degrading bacterium DhA-73, unidentified Hailaer soda lake bacterium, and uncultured bacterium clone HuCa4. Seven organisms representing new potential species were also detected. PCR followed by cloning and sequencing may help to identify new pathogens involved in mixed bone infection.

Diagnosis of prosthetic joint infections using UMD-Universal Kit and the automated multiplex-PCR Unyvero i60 ITI® cartridge system: a pilot study

“…16S rRNA gene real-time PCR results were in line with the conventional culture results in about 96 % of the studied patient cases. These findings seem to be consistent with previously published data on NAAT techniques, indicating a relatively wide range of sensitivity and specificity [8][9][10][11][12]. NAATs may reasonably expand the diagnostic panel and contribute to a higher recovery rate especially after antimicrobial use.…”

“…Only few studies have assessed the diagnostic utility of nucleic acid amplification test (NAAT) techniques in this context. The reported findings for different PJI-associated materials indicate a wide range of sensitivity (50-92 %) and specificity (65-94 %) [8][9][10][11][12][13]. Results from a recent study comparing 16S rRNA gene PCR and consecutive sequencing of sonication fluid with culture of synovial aspirates, tissue samples and sonication fluids reports that 16S rRNA gene PCR improved sensitivity [14].…”

Propionobacter acnes Infection as an Occult Cause of Postoperative Shoulder Pain: A Case Series

“…At the time of the initial débridement, all specimens sent for intraoperative Gram stain in our series were negative. There are recent reports on PCR in combination with sequencing of 16S rDNA [16] and multiplex PCR of sonification fluid in detecting periprosthetic P. acnes joint infections [2] in patients who had false-negative P. acnes cultures. A limitation of this testing technique is the time it takes to confirm a diagnosis; the results come in too late to help in the clinical diagnosis.…”

Detection of Periprosthetic Infections With Use of Ribosomal RNA-Based Polymerase Chain Reaction

“…In the last decade, many studies have investigated the use of traditional polymerase chain reaction, a technology that enzymatically amplifies deoxyribonucleic acid (DNA) by means of sequence-specific oligodeoxynucleotide primers, to detect bacterial DNA in order to identify joint infections [13][14][15] . The results have shown high sensitivity but have been limited by the number of potential false-positive results 16 .…”