Difficulties in the molecular diagnosis of Helicobacter rodent infections.
Molecular diagnostic methods using the polymerase chain reaction (PCR) are the gold standard in Helicobacter diagnostics. Most rely on the amplification of parts of the 16S rRNA gene sequence. Therefore, the validity and accuracy of results depends heavily on the PCR design and the time of its publication because new sequences are continually being submitted to databases. Here we report the presence of helicobacter in commercially bred mice supposedly free of this infection. Furthermore, three out of six different commercial laboratories performing helicobacter testing on the same spiked faecal samples failed to detect and identify H. hepaticus. We designed a simple generic PCR assay that amplifies a 261bp amplicon spanning two of the seven variable regions in the 16S rRNA of helicobacter. Using this assay together with an established generic assay designed by Bohr [Bohr, U.R., Primus, A., Zagoura, A., Glasbrenner, B., Wex, T., Malfertheiner, P., 2002. A group-specific PCR assay for the detection of Helicobacteraceae in human gut. Helicobacter 7, 378-383] and then cloning and sequencing their products, we detected the H. hepaticus used in the study that three commercial laboratories failed to detect. We think these assays together could detect all the currently known species of helicobacter and hopefully the new ones as well. In addition, we have been able to identify different species of helicobacter and their relative proportions infecting a single animal. This information has also shown that some helicobacters may have a much broader host range than originally reported.
Laboratory Animal Science, GlaxoSmithkline, The Frythe, Welwyn, Herts AL6 9AR, UK.
Scott
S
Poynter
Julian D
JD
Phipps
Angel
A
Naranjo-Pino
Jose M
JM
Sanchez-Morgado
D000818:Animals; D001426:Bacterial Proteins / Q000145:classification / Q000235:genetics; D001483:Base Sequence; D003001:Cloning, Molecular; D016998:Helicobacter / Q000145:classification* / Q000235:genetics / Q000302:isolation & purification; D016481:Helicobacter Infections / Q000175:diagnosis / Q000662:veterinary*; D051379:Mice; D016133:Polymerase Chain Reaction / Q000662:veterinary*; D012336:RNA, Ribosomal, 16S / Q000235:genetics; D012376:Rodent Diseases / Q000175:diagnosis* / Q000382:microbiology; D013045:Species Specificity
D001426:Bacterial Proteins; D012336:RNA, Ribosomal, 16S