Site Published:  16th April 2010
Last updated:   18th April 2010
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Testing
'Not Detected' does NOT mean 'Negative'
I have spoken with vets that have had what they considered 'highly suspect' cases of HeV.  Terminal cases, that have been tested by Biosecurity Queensland (in the last 12 months), and have been told they were negative.  On several cases, they have stated NEGATIVE, based purely on Nasal or Oral Swabs subjected to PCR (Tagman) testing here in Queensland.

Some of you reading this may have had similar experiences (we would like to hear from you!).   Of particular concern is the fact that vets are putting absolute faith in this testing and this WILL lead to more deaths if adequate precautions are not maintained when treating sick animals...
Page 94 of the AusVet/Nigel Perkins Review 2008
I have repeatedly pointed out to the authorities they are potentially exposing people to this deadly virus by giving false sense of security that they were not (or are not) dealing with Hendra virus infection on a property, by stating negative on limited testing.
The authorities have ALWAYS known that the PCR testing is of limited diagnostic value.  It has provided fast results and early detection in the early stages of clinical disease, but NOT ALWAYS!!

A Good EARLY indicator

In the recent (2008) Initial-experimental Characterisation Trials conducted at AAHL using the Redlands strain of HeV gives tables of samples tested and detecting the virus (using PCR Taqman) as early as two days post infection on nasal swabs and a full day prior to established fever on blood.  Once clinical symptoms were established the detection in nasal swab and blood was consistent.  However, all these samples were collected, by highly experienced scientists in a controlled environment and stored under optimum conditions until testing was conducted... 

extract from page 2
Extract from an email dated 2nd October 2009 from Ms Kareena Arthy - Managing Director Biosecurity Qld
They also knew exactly which sequence they were working with, having infected the animals with a known strain of the virus.

Samples being collected 'in the field' so to speak, and transported, would hardly be handled in the same optimal conditions.  There is no doubt about the fact this testing is the best currently available for early detection, but not detected does not mean negative.


NOT bullet proof!!

Every version of the veterinary guidelines has acknowledged the fact that multiple samples will improve the likelihood of a conclusive diagnosis.


Page 11 of the 2007 Guidelines stated:


(h) Diagnostic difficulties

The previous 2005 version had virtually the same “diagnostic difficulty”, wording which has now been removed from the current Veterinary Guidelines.  The warnings (to cover the authorities backsides) are there, but not quite as obvious as they have been previously!!

Page 24 of the current Veterinary Guidelines issued in April 2009 after considerable pressure being applied to Biosecurity Qld, state:  

Limited necropsy
A limited necropsy will increase the likelihood of reaching a conclusive diagnosis.


Next page (25) last para states:

Note: because of the serious zoonotic potential of HeV infection, if human exposure is thought possible then an accurate rule in/rule out diagnosis is required on public health grounds.  If Queensland Health deem this is required, then it is unlikely such a diagnosis can be confidently made without the laboratory testing of appropriate specimens - blood, lung, spleen, liver, kidney and lymph nodes of the head.  Other tissues such as brain and spinal cord are useful but are of secondary importance.  Note previous warnings : a necropsy of this level will require specialist training.


To demonstrate for you the dangers in blindly placing faith in a ‘Negative’ result given to you by Biosecurity Qld on limited samples, I have uploaded some files.

The first of these files are Queensland Health and AAHL pathology reports on the three confirmed Hendra horses involved in the Proserpine outbreak in 2008.  View these files by clicking on the thumbnails below or visit the Pathology Reports Page
Confirming the diagnosis in a live acutely-ill animal can be particularly difficult.  Virus isolation can be attempted on nasal swabs and antibody detection on serum samples, but the sensitivity of these techniques is likely to be lowConfirmation or exclusion of HeV disease will most likely only be obtained at necropsy.
Tests include Serum Neutralisation Test (SNT and ELISA on serum, virus isolation on body fluids and discharges, and polymerase chain reaction (PCR) on fresh and formalised tissues.
PCR tests are highly sensitive and can provide a good, early indication of infection.
PCR tests are of limited diagnostic value alone and should be considered in conjunction with other results when making a diagnosis.
Histological examination of lung (and kidney and spleen) tissue can be safely performed on formalised samples at a regional veterinary laboratory and will indicate whether lesions are consistent with HeV disease.
Histological findings of acute severe pulmonary oedema and interstitial pneumonia are indicative of HeV disease.  Fibrinoid degeneration of blood vessels throughout the body with syncytial cell formation in affected vessels is highly characteristic.  Histopathology may also confirm an alternative diagnosis, for example, inhalation pneumonia or purulent bronchopneumonia or Crofton weed poisoning.  Histology should therefore be undertaken as a matter of urgency.
Note that on each of these cases the PCR Tagman test was "not detected" on one of the blood samples.
As veterinarians, you are (presumably) only collecting samples to be submitted from the animal, once, either early in the clinical stages of illness or possibly near death (or after).  I hope this table demonstrates that trusting a PCR Negative on swab or blood alone, taken at one time, and trusting that it is negative means playing Russian Roulette IF adequate precautions are not maintained.

Most people have been brainwashed into thinking that horses infected with HeV will show severe clinical symptoms and die within 1-4 days.  Please bear in mind that of the known 'recorded' outbreaks 20% of the horses infected recovered.

Hendra must remain on the minds of all people working with animals, if suspected, it must remain on the list until proven otherwise, and one or two PCR tests on swab or blood alone is NOT conclusive!!
This page, the first confirmed horse in the Proserpine case, confirmed at AAHL, reveals at the bottom of the page

"Note the Hendra Taqman (this is the older assay) was not detecting the virus in horses from Redlands, but is in this case."
A disclaimer is also printed on each of the Queensland Health laboratory reports (for HeV testing) that states:

"Failure to detect organism-specific nucleic acids does not exclude the presence of disease due to this agent."

OK, now if you are thinking, this doesn't matter because those horses were confirmed anyway, or yes, well they have improved the testing and all the labs are now using the latest and greatest PCR Tagman assay for testing and you are placing absolute faith in being told NEGATIVE, read on....

It is well documented that Hendra virus has manifested as respiratory and neurological disease in both humans and horses.  The PCR Tagman testing detects viral genetic material whether it is viable or not. The testing requires that the sample taken actually contains viral genetic material, i.e. when the animal or human is shedding the virus at the time the samples are collected.

I have prepared the table below from pathology results on two human cases (which can be found in the Pathology Reports page), one fatal and one non-fatal.  The table represents the twenty days testing conducted on aspirate, blood and urine immediately after both cases were confirmed positive to HeV.  All tests conducted at the same lab, using the same test.
'Not Detected'  DOES NOT MEAN  'Negative'
Early detection is the key, and this information will be included in submission to Government to support the urgent need for a rapid stall-side test kit to be used in the field.    Whilst cases are going undetected and unrecorded, this virus will remain 'rare' and the political motivation to invest appropriate funding into finding a vaccine or cure will also be a low priority. 

Please support
the petition to show the Government the widespread concern about this virus.


Update to show further proof that
'Not Detected' does NOT mean 'Negative'
As at the 30th August 2011, there have been 17 separate confirmed outbreaks of Hendra, 9 in Queensland and 8 in NSW, involving 21 horse deaths and the first 'recorded' case of a domestic dog developing antibodies.

The bulletins released by the authorities in both states have been giving a variety of information, not consistent with each outbreak.  However, on at least 3 of the Queensland cases, I have spoken with vets that have had what they considered 'highly suspect' cases of HeV.  Terminal cases, that have been tested by Biosecurity
Queensland (in the last 12 months), and have been told they were negative.  On several cases, they have stated NEGATIVE, based purely on Nasal or Oral Swabs subjected to PCR (Tagman) testing here in Queensland.

Some of you reading this may have had similar experiences (we would like to hear from you!).   Of particular concern is the fact that vets are putting absolute faith in this testing and this WILL lead to more deaths if adequate precautions are not maintained when treating sick animals...