Thursday, April 05, 2012

The 30th Anniversary of the Culture of Helicobacter pylori

On Easter Thursday 1982, which was the 8th of April, we performed endoscopy on a man in his 60's who I recall had a recurrent bleeding duodenal ulcer.  This was a major problem for him because he also had a heart valve problem and therefore was required to take anticoagulants (making him more likely to bleed from the ulcer).  Dr Warren and I were in the third week of a study of 100 consecutive endoscopy patients and we had already entered more than 30 patients in the study.  I took two gastric biopsies to the pathology lab and one to the micro lab (for Dr Pearman).
I have no idea what I did that Easter but I suspect that I was on-call as registrar in haematology with many very sick patients to attend to.  Luckily I lived only 15 minutes from Royal Perth Hospital.
In Perth, the Easter Break is four days (Good Friday to Easter Monday incl.) so the bacterial cultures which had been set up on the Thursday morning were not examined until Tuesday morning, at which time the typical "water spray" appearance of Helicobacter colonies were visible.
The next day, an excited John Pearman called me to come and see the bacteria they had grown from a patient cultured the previous Thursday.  He showed me the culture plates and we peered at the Gram stained smear of the bacteria through the lab microscope.  The bacteria were Gram-negative - pink (at least that part was correct) but were not obviously spiral - they were all shapes and sizes!  After 6 months of disappointment, I was not going to break out the champagne on such borderline evidence of success.  After all, why should we suddenly be able to grow the bacteria when all other attempts had failed?  What were we doing differently?  In the next week John's lab was able to culture Helicobacter (which we called Campylobacter in those days) from several more patients.  Reflecting on this, we realised that in the previous attempts, lab staff had been examining the plates after 48 hours then discarding them if nothing new was visible.  After all, biopsies covered in saliva and dragged up from the stomach through the channel of an endoscope would be severely contaminated with commensal organisms.  These irrelevant "commensal organisms" (fungus, oral streptococcus, bacillus and hundreds of other species) would be expected to completely cover the plates after 48 hours and obscure any new kind of bacteria.
But biopsy specimens taken from the human stomach for Helicobacter were a little different.  The wall of the stomach is exposed to a puddle of acid which kills most of the bacteria being swallowed from the mouth.  So the biopsy samples were sometimes almost sterile (except for Helicobacter pylori which lives under the protective mucus gel layer); and even non-selective blood agar plates could be incubated for 3-5 days with quite a few areas of clean agar visible upon which Helicobacter could slowly grow.
Normally, after two days, the lab technician would have discarded the cultures after seeing nothing worth keeping at the 48 hour inspection (Easter Saturday).  However, that Easter may have been particularly busy so that urgent cases took precedence over our clinical research; which was regarded more as a hobby than as science.  I recall that there was a MRSA (methicillin resistant staph-aureus) control program in the hospital that month so maybe the microbiology staff were overloaded.  In any case, on Easter Saturday, the technologist did not examine the research cultures and left them to sit for three more days until the Tuesday morning examination.  The first culture plate probably looked like this:

That first culture became the "type strain" of Helicobacter pylori, ATCC #11637.  This bacterial strain has been studied by thousands of research labs and costs £150 to obtain from the NCTC.  You can read about the type strain of Helicobacter pylori here: pdf file of NCTC citation copied on 2012-04-04