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Keywords:

  • Bacteria;
  • Gas chromatography-mass spectrometry;
  • Pan-bacterial PCR;
  • Rheumatoid arthritis

Abstract

Objective

To study the presence of bacterial components in the synovial tissue (ST) of patients with advanced rheumatoid arthritis (RA).

Methods

ST was collected during joint surgery from 41 RA patients. Tissue from 39 patients with osteoarthritis (OA), 4 patients with undifferentiated inflammatory arthritis (UA), and 3 cases of accidental deaths served as controls. The pan-bacterial polymerase chain reaction (PCR) with primers for the 23S ribosomal RNA (rRNA) and 16S rRNA genes was used to detect bacterial DNA. In addition, synovial fluid (SF) samples from patients with chlamydial reactive arthritis (ReA) were also examined by the same method. The positive controls, bacterial DNA or ST spiked with different living bacteria, were analyzed alongside clinical samples. Most of the ST samples were also analyzed by gas chromatography-mass spectrometry (GC-MS) for determining the presence of bacteria-derived muramic acid. Strict precautions were followed in the clinics and the laboratory to prevent contamination.

Results

In GC-MS analysis, muramic acid was observed in the ST from 4 of 35 RA patients and from 2 of 14 OA patients, but not in ST from 2 patients with UA and 3 cadavers. Bacterial DNA was not detected by either one of the PCR primers used in ST from 42 patients with RA and 39 patients with OA. However, 5 of 15 SF samples from ReA patients were PCR positive. The sensitivity of GC-MS to detect muramic acid was 2 pg/injected amount (227 pg muramic acid/mg ST), and that of the pan-bacterial PCR was 2–20 bacteria colony forming units/reaction.

Conclusion

These results indicate that a bacterial component, muramic acid, is detectable by GC-MS in ST from a few patients with advanced RA or OA. However, no bacterial DNA was detectable by PCR.