False positive PCR detection of Tropheryma whipplei in the saliva of healthy people.
BACKGROUND
Tropheryma whipplei, the agent of Whipple's disease (WD), has been recently isolated and the genomes of two isolates have been fully sequenced. Previous diagnosis tools for the diagnosis of the disease used sequence analysis of the 16S rRNA gene. Using this target gene, the high percentage of detection of the bacterium in saliva of healthy people was in contrast to the negative results obtained with specific target genes. The aim of our study was to compare previously published primers targeting the 16S rRNA gene to real-time PCR with Taqman* probes targeting specific repeat genes only found in the genome of T. whipplei in a series of 57 saliva from healthy people.


RESULTS
Although the specific real-time PCR assays with both primers and probes were negative for all the samples, 13 out of 57 samples were positive with different primers previously reported targeting the 16S rRNA gene. Among the positive samples, 8 yielded a 231-bp sequence that was 99.1% identical to that of Actinomyces odontolyticus, 2 yielded a 226-bp that was 99.6% identical to that of A. turicensis, and 3 yielded a 160-bp sequence that was 98.5% identical to that of Capnocytophaga gingivalis. We found that the C. gingivalis and A. odontolyticus 16S rRNA sequences obtained in our study share more than 80% homology with the corresponding 16S rRNA sequences of the T. whipplei genomes especially at 5' and 3' end.


CONCLUSIONS
Asymptomatic carriers of T. whipplei in saliva may exist but their prevalence is much lower than those previously reported. Testing the specificity of designed primers is critical to avoid false positive detection of T. whipplei. In atypical case we recommend to test two different specific target genes before concluding.
Rolain
Jean-Marc
JM
Université de la Méditerranée, Unité des Rickettsies, CNRS UMR 6020, IFR 48, Faculté de Médecine, Marseille cedex 05, France. jm.rolain@medecine.univ-mrs.fr <jm.rolain@medecine.univ-mrs.fr>
Fenollar
Florence
F
Raoult
Didier
D
D000190:Actinomyces / Q000235:genetics; D000192:Actinomycetales / Q000235:genetics / Q000302:isolation & purification*; D000328:Adult; D002206:Capnocytophaga / Q000235:genetics; D017931:DNA Primers / Q000235:genetics; D004269:DNA, Bacterial / Q000235:genetics; D004275:DNA, Ribosomal / Q000235:genetics; D005189:False Positive Reactions; D005798:Genes, Bacterial / Q000235:genetics; D006801:Humans; D008875:Middle Aged; D016133:Polymerase Chain Reaction / Q000379:methods*; D012336:RNA, Ribosomal, 16S / Q000235:genetics; D012463:Saliva / Q000382:microbiology*; D017422:Sequence Analysis, DNA; D012689:Sequence Homology, Nucleic Acid; D008061:Whipple Disease / Q000175:diagnosis
D017931:DNA Primers; D004269:DNA, Bacterial; D004275:DNA, Ribosomal; D012336:RNA, Ribosomal, 16S