Monday 17 November 2014

Ebola testing: 48-72 hours for a negative to turn positive

Currently, some fraction of the people who present very early after they may have been infected by Ebola virus for testing, return a negative result. This is probably a rare event because the majority of cases arrive for care with Ebola virus disease (EVD) already well underway.

The latest Centers for Disease Control and Prevention (CDC) guidance in these instances is to wait (48 to) 72 hours and see if the patient remains ill, or becomes more unwell. If they do either of these, a second test is performed.[1] If the suspect case recovers from illness, no repeated testing is indicated. 

The test we rely on to confirm a clinically suspected EVD case is called a reverse transcriptase polymerase chain reaction (RT-PCR). RT-PCR is a technique designed to  seek out a tiny but very specific region of the Ebola virus's RNA genome, copy it into DNA then amplify those DNA copies a billion-fold by making more copies. Somewhere during that exponential amplification process, the technology of the day (currently fluorescence detection but formerly agarose gel detection, radiation and chemiluminescence) allows us to identify that the specific DNA we seek is appearing above an arbitrary threshold...we have a positive test result for Ebola virus. 

RT-PCR is a very sensitive technique. It was not that long ago - the 80s, not that long for some of us anyway - that clinicians and scientists were complaining that PCR methods were too sensitive. This was in large part because PCR was too successful at finding infectious agents where, and when, they had not been previously found. Change to dogma was in the wind. Fast forward to today and now we're lamenting that PCR isn't always sensitive enough. Very early on after acquiring what we later know to be a true infection, even exquisitely sensitive PCR methods can fail to detect those earliest of viruses while they are struggling to gain a foothold in our cells and replicate themselves to levels that outstrip our immune system's capacity to contain. Whether this is because the virus is hidden away in organs during its early replication or whether too few circulating viruses yet exist to surpass the necessary threshold of the RT-PCR assay's sensitivity at these early stages is unclear.

EVD patients who are not yet showing signs and symptoms of disease may present early for testing and care because they they are healthcare workers with a suspected or known exposure, or they may be the contact of a known EVD case or infected animal being tested early on to exclude infection. But as we have seen and read anecdotally, that first test can sometimes be negative; not due to inhibition of the RT-PCR (which can also happen, just not so much with today's purification methods) nor because they are truly uninfected, but simply because we're testing too early. These are examples of false negative results.

For the past few weeks I have been trying to find he evidence that underpins why the world chooses to use a 48-72 hour window in its guidelines. I've been asking a lot of people-and I thank those who replied. Tonight the very diligent and extremely tolerant folks at the World Health Organization got back to me with a quote from Dr Pierre Formenty, team leader Emerging and Dangerous Pathogens. A hard man to get hold of sometimes-as you might imagine. He said (lightly edited)...
There is at least one documented case during an outbreak in Africa; a contact with fever = a suspect case; he was negative at day 1 with RT-PCR (CDC Lab) and was found positive at day 3 (when retested).
So the 48-72 hours come from this incident. We want to be on the safe side and limit the number of false negative that are inevitable with any test.
And so there you have it. If anyone has anything further to add to this story, I'd be most happy put it here.


Sunday 2 November 2014

Influenza A(H7N9) virus: detection numbers and graphs...

This is a static page that houses my graphs of influenza A(H7N9) virus ("H7N9") numbers produced by the various Ministries of Health for the provinces and municipalities of China, Hong Kong's Centre for Health Protection (CHP) the World Health Organization and FluTrackers.

Please be aware that these numbers are in flux during the winter epidemic. They are a guide only.

There is also an accompanying map page which for now is located here.

  • The graphs above, as with all on VDU, are made for general interest only. They are also freely available for anyone's use, just cite the page and me please. The data can be downloaded by clicking on the "Download" link at the bottom-right of each dashboard. It may be that I have misinterpreted the language in the reports (sometimes a little tricky to wade through) or miscalculated some totals based on the way data have been presented.
  • In any outbreak, epidemic or pandemic caused by a know or emerging pathogen, the numbers presented publicly, and used in these graphs, are expected to represent only a fraction of all the cases that have and are occurring. This is just the nature of the imperfect biological'ness of these events.
  • I am only able to plot what is publicly available-you could do this too. No secret associations or back-room deals provide me with these data.