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The Skin Test

 Added by  Sally
 3 Apr 2010, 7:55 PM
There is an interesting statement in 'Testing for Bovine Tuberculosis in California' by John H Kirk. The report is describing the skin test and warns 'be aware that in non-infected herds it is considered normal to have 1-5% false positives or reactions at the site not due to Bovine Tuberculosis, on a whole herd test. This response may be due to previous exposure to Avian Tuberculosis or the Johne’s Disease bacteria, another Mycobacterium disease.'
'Estimating the Hidden Burden of Bovine Tuberculosis in Great Britain'
Abstract from the Cambridge paper referred to in the above post.
The number of cattle herds placed under movement restrictions in Great Britain (GB) due to the suspected presence of bovine tuberculosis (bTB) has progressively increased over the past 25 years despite an intensive and costly test-and-slaughter control program. Around 38% of herds that clear movement restrictions experience a recurrent incident (breakdown) within 24 months, suggesting that infection may be persisting within herds. Reactivity to tuberculin, the basis of diagnostic testing, is dependent on the time from infection. Thus, testing efficiency varies between outbreaks, depending on weight of transmission and cannot be directly estimated. In this paper, we use Approximate Bayesian Computation (ABC) to parameterize two within-herd transmission models within a rigorous inferential framework. Previous within-herd models of bTB have relied on ad-hoc methods of parameterization and used a single model structure (SORI) where animals are assumed to become detectable by testing before they become infectious. We study such a conventional within-herd model of bTB and an alternative model, motivated by recent animal challenge studies, where there is no period of epidemiological latency before animals become infectious (SOR). Under both models we estimate that cattle-to-cattle transmission rates are non-linearly density dependent. The basic reproductive ratio for our conventional within-herd model, estimated for scenarios with no statutory controls, increases from 1.5 (0.26–4.9; 95% CI) in a herd of 30 cattle up to 4.9 (0.99–14.0) in a herd of 400. Under this model we estimate that 50% (33–67) of recurrent breakdowns in Britain can be attributed to infection missed by tuberculin testing. However this figure falls to 24% (11–42) of recurrent breakdowns under our alternative model. Under both models the estimated extrinsic force of infection increases with the burden of missed infection. Hence, improved herd-level testing is unlikely to reduce recurrence unless this extrinsic infectious pressure is simultaneously addressed.
Email dated 18/10/12 from Prof PT.
As I have said before - I think about 25,000 cattle test positive (by the skin test) each year in the UK. Assuming the test is close to 100% specific (actually about 99.7% specific) and 80% sensitive (DEFRA would agree on this). That means that there are around 6000 cattle which have bTB which test negative EVERY year. And they will not be the same 6000 each year as not every herd is tested each year, so the true number of infected bTB cattle at any one time not being detected will be somewhat higher. The main danger with these false negatives is in two area. First in herds where the rest of the herd is also negative so they can be traded and spread bTB around the country. Secondly in those herds under restrictions because at least one animal had a positive test. There will also be false negatives, not removed from the herd which continue to infect other animals... this will mean that the disease may remain on the farm, even if the herd then has a clean test - not really surprising but it is good to see academics confirming this. Whole herd slaughter would in fact deal with this as you correctly point out, but then that would mean killing a lot of cattle (25% of herds in the SW) and of course is totally unacceptable.
So I agree much of the problem is probably caused by the 6000 and not having a herd slaughter policy....you need a much more sensitive test to have a policy designed at the individual animal....
But I think we all know the present policy is not working and has not worked for a long time.
We now have more Defra-funded) research (www.cam.ac.uk/research/news/scientists-build-a-clearer-picture-of-the-spread-of-bovine-tuberculosis/) confirming just how unreliable the skin test is - the skin test is the backbone of the existing test/cull system currently used to 'eradicate' bTB.
Scientists at Cambridge have used mathematical tools to develop models for estimating the efficiency of cattle-based controls for bovine tuberculosis (bTB).
The models built by the team represent an advance over previous models as they are informed directly by extensive data on reported incidence and spread of the disease, rather than expert opinion. Importantly, they provide a first estimate of the quantity of infection missed by cattle testing and the contribution of this hidden burden of infection to the persistence of bTB within herds.
Applied to recent data, the models suggest that around one in five of British herds that have been cleared of restrictions, following testing for bTB, may harbour the infection. It also points to a higher incidence, and faster spread, of the disease in large herds. The research focuses on the cattle-to-cattle transmission of bTB within herds.
“Many aspects of bTB, and its transmission, remain a mystery – and it’s long been known that the protocol and testing methods used to diagnose the disease are far from perfect. One of our models suggests that up to 21 per cent of herds may be harbouring infection after being cleared from movement restrictions,” said James Wood, Alborada Professor of Equine and Farm Animal Science at Cambridge University.
“However, our models also estimate a high rate of re-introduction of infection into herds, particularly in high incidence areas. This rate of re-infection is high enough that even if improvements in testing eliminated the hidden burden of infection, rates of recurrence would not be reduced.”
An article by farmers Jolyon and Alex Higgs in Farmers Weekly today will probably strike a chord with many cattle owners having to suffer the zero tolerance policy in Wales. They refer to the unhelpful and threatening letters received from the AHVLA.
The article is re-printed below.
On Gower we rear bucket calves from dairy herds, and during the past year have carried out 37 follow-up TB tests on calves, whose original herd has gone down with TB. This meant at least 10 separate testing days.
We are happy to carry out these tests, but feel that the Animal Health and Veterinary Laboratories send out notices that are unnecessarily threatening.
Last October, a full clear test was carried out on time. However, we still received a notice prohibiting the movement of bovine animals because the computer had not been updated.
At the end of the month, another restriction was imposed for failing to test a calf that was ineligible to be tested as it had been tested in the previous 60 days.
In March, a further notice was served on us for failing to test a dead calf.
In the past six weeks, a new computer system has been set up linking AHVLA with our vets. We are now receiving 'Zero Tolerance Advisory Letters', one month before a TB test is due, if a test date has not been booked with the vet.
As the AHVLA is not infallible, reminders sent out to farmers – who have not yet done anything wrong – could perhaps be worded less severely.
Finally, we recently asked to postpone a test, due on October, on four calves – two of which have been on our holding over a year – until our full herd test, which is due on 22 October. This was not allowed.
Excellent letter from a farmer published in the Independent that sums up the situation very well (www.independent.co.uk/voices/letters-badger-culling-is-emotive-but-debate-over-scientific-evidence-is-ignoring-established- facts-8182022.html)
www.independent.co.uk/voices/letters-badger-culling-is-emotive-but-debate-over-scientific-evidence-is-ignoring- established- facts-8182022.html
'We've allowed the facts about bovine TB to be buried by the furore surrounding the proposed badger cull and negative reporting on the part of farming bodies has painted a picture of an entire cattle industry brought to its knees by the effects of the disease. But this is misleading. Defra reports on its website that "11.5 per cent of herds were restricted in 2011". They could equally report the positive side, which is that 88.5 per cent of herds were not restricted in 2011. Only a small proportion of the national herd is affected by bovine TB.
The impact on an infected herd is not caused by the disease itself. The heartache is caused by the "test and cull" policy deployed by our government in response to an EU directive which demands eradication of bTB and simultaneously bans the use of cattle vaccine, thereby making it impossible to achieve the goal they set.
But rather than tackle the EU to allow cattle vaccination, ministers have sanctioned a mass badger-shoot to placate a minority of vociferous farmers who seem hellbent on decimating the badger population. The fact that a badger cull spells disaster in PR terms for the entire farming industry must have eluded them all'.
G E Purser (A badger-friendly farmer), Clapton-on-the-Hill, Gloucestershire
From the Alpaca website at http://www.alpacatb.com where alpaca owners are having to fund their own research, we have the initial results of the pilot study re PCR testing alpacas. The skin test is even less reliable for camelids.
The pilot project demonstrated that a two-stage PCR test on camelid clinical samples has the potential to detect Mycobacterium bovis with reasonable sensitivity. Samples were taken post-mortem from 21 alpacas with gross lesions of tuberculosis, most of the alpacas selected had severe lesions. Fifteen faeces samples were positive and ten nasal swabs were positive. Samples that were negative in the PCR tests were from alpacas with less severe pathology
We also saw research into PCR testing for badgers at (http://www.plosone.org/article/info%3Adoi%2F10.1371%2Fjournal.pone.0027369) is the first example of a multi-laboratory validation of a real time PCR assay for the detection of mycobacteria in environmental samples ...
www.idexx.com/view/xhtml/en_us/livestock-poultry/ruminant/bovine-tuberculosis.jsf Details of the test the subject of the last posting is as follows:
Control of bTB is complicated by the lack of sensitive tests. Although tuberculosis tests based on cell-mediated responses (skin and gamma interferon) can detect animals in the early stages of infection, they still fail to detect up to 20% of truly infected animals. Using antibody testing for bTB control programs can significantly improve the detection of bTB-infected herds. In bTB-negative areas, antibody testing can be used on other sample streams as an easy, cost-effective surveillance tool.
The IDEXX M. bovis Ab Test is intended for the detection of Mycobacterium bovis (M. bovis) antibody in cattle serum and plasma samples. Adding IDEXX M.bovis Ab test to bTB control programs will increase detection by identifying infections other tests miss. This test is a robust and reproducible tool that offers a fast, 2-hour protocol for objective, quantitative results. IDEXX easy-to-use M.bovis ELISA format requires no specialized training or handling requirements and can be repeated as often as necessary, allowing more rapid bTB control.
'IDEXX announced that the World Organization for Animal Health (OIE) has added the IDEXX M. bovis Ab Test to its registry of tests. OIE develops and maintains international standards for diagnostic tests and vaccines, and this marks the first time the organization has certified an antibody test for the detection of bovine tuberculosis (TB). Based on a stringent independent review, the IDEXX M. bovis Ab Test was validated and certified by the OIE as fit for the purposes defined in the kit insert.'
'Released in 2011, the IDEXX M. bovis test offers a "fast, three-hour protocol that delivers objective, quantitative results compatible with typical laboratory data management systems," IDEXX said. Its "easy-to-use" ELISA format requires no specialized training or handling requirements and can be automated for high-volume use. The test also can utilize serum or plasma samples already collected for other disease surveillance purposes.'
“The IDEXX M. bovis test is already used in several countries, including Chile and Ireland,” said John Lawrence, Senior Research Scientist, IDEXX Livestock and Poultry Diagnostics. “We’re also participating in trials to gain specific country-level approvals, as well as undergoing research studies in France, the United Kingdom and New Zealand.”
More info from idexx.com/mbovis.
'Improved Skin Test for Differential Diagnosis of Bovine Tuberculosis by the Addition of Rv3020c-Derived Peptides' by Gareth J. Jonesa, Adam Whelana, Derek Cliffordb, Mick Coada and H. Martin Vordermeiera.
A peptide cocktail derived from the mycobacterial antigens ESAT-6, CFP-10, and Rv3615c allowed differentiation between Mycobacterium bovis-infected and M. bovis bacillus Calmette-Guérin (BCG)-vaccinated cattle when used as a skin test reagent for a “DIVA” test (i.e., a test capable of differentiating infected and uninfected vaccinated animals). Addition of the antigen Rv3020c improves the diagnostic sensitivity without compromising specificity in the face of BCG or Johne's disease vaccination.
I have heard that a dairy farmer in South West Wales has on 4 seperate occasions lost a number of cows that were inconclusive reactors.None of these animals showed any sign of TB on post mortem.He has now refused to have any more of his cows slaughtered and is mounting a legal challenge against his latest order to cull cows.
According to the Holyhead and Anglesey Mail (www.theonlinemail.co.uk/bangor-and-anglesey-news/local-bangor-and-anglesey-news/2012/04/18/anglesey-vets-pracice-banned-from- testing-for-bovine-tb-66580-30786655/) an Anglesey veterinary practice has been banned from testing for Bovine TB after it was found out it wasn’t conducting the process properly. The practice is the Gaerwen based vets Bennett-Williams which sent out letters to farmers and farming unions explaining the disruption to TB testing.
Cattle farmers who use the practice for TB testing have been told by Department of Environment, Food and Rural Affairs (DEFRA) to make alternative testing arrangements after an audit of TB testing procedures identified the problems.
To regain the right to deal with TB, vets at the practice will need to be re-trained.
An angry farmer told the Mail: “There are deadlines and legal requirements about testing that we as farmers need to meet.
“If this is not done right then it could put the whole TB control plan in jeopardy. We could believe that our cattle are healthy and then find that they have to be destroyed, which would put you out of business.”
A spokesman for the Animal, Health and Veterinary Laboratories Agency (AHVLA) part of government’s DEFRA department said: “Following an audit of TB testing procedures, which identified a failure to follow standard operating procedures, AHVLA has suspended a private veterinary surgeon working in North Wales from operating as a panel 1a (bovine TB testing) official veterinarian, pending re-training.
“Owners of cattle served by the suspended practice have been notified and advised to make alternative testing arrangements.”
The Mail apparently contacted the Bennett-Williams practise about the issue and was told they were too busy to comment.

An interesting piece on the Bovine TB Blog - entitled That EU elephant again and posted on February 12, 2012 (http://bovinetb.blogspot.com/). It reports on the costs of bTB testing - some £20 million a year and the existing contract with veterinary practices, the involvement of the EU and the usual confusion and recently announced measures to tighten up on controls. Well worth a read. What a mess.
Some very interesting admissions from Defra in this document - see extracts below:
'No diagnostic test, including the tuberculin skin test, is 100% accurate, but the current skin test is effective (and is the primary diagnostic test required under EU legislation). On the one hand, the comparative skin test used in the UK and Ireland can be expected to vary around approximately 80% detection rate of all the infected cattle in a herd at any one test (at standard interpretation, range 52-100%). On the other hand, reactions to the tuberculin test can sometimes be caused by exposure to other mycobacteria which do not cause bovine TB.'
'Studies evaluating the sensitivity of the test suggest that its sensitivity lies between 52% and 100%.'
'Do animals with fluke show a stronger reaction to the skin test and result in false positive reactions? ... A review of the veterinary literature on this topic provides somewhat conflicting evidence. On one hand, the cattle TB pathogenesis study conducted in GB (Defra project SE3013) reported that skin test reactors and contacts with antibodies to liver fluke (Fasciola hepatica) were less likely to show with evidence of M. bovis infection at post-mortem examination. The effect was most significant in dairy reactors.... In Ireland, Flynn et al. (2007) established an experimental model of co-infection of F. hepatica and M. bovis BCG to examine the impact of liver fluke infestation on correct diagnosis of TB in cattle. They found that the sensitivity of skin and gamma interferon tests was compromised in co-infected animals and that F. hepatica infection altered macrophage function. Their results raise the question of whether F. hepatica infection can
affect the predictive capacity of tests for the diagnosis of bovine TB and possibly also influence
susceptibility to bovine TB and other bacterial diseases. In summary, this is a hypothesis that merits
further investigation.'
'Is TB testing compromised by the presence of Johne‟s disease?... Yes. Exposure to Johne‟s disease can cause cross reactivity when using the skin and gamma interferon tests for bovine TB.'
'If the skin test for bovine TB can be compromised by other mycobacteria (e.g. avium, microti),is the gamma interferon test compromised in the same way? ... Yes. In cattle, false positive reactions to the gamma interferon test can sometimes be caused by exposure to mycobacteria other than M. bovis.'
Interesting presentation papers from the 2009 M Bovis conference in Wellington, New Zealand: 'How does the ‘field’ SICTT performance of Tuberculin PPDs compare? - from different manufacturers? - with different potencies? What if only the bovine site reading is considered – the SIT?'
Some of the conclusions:
A potency change in tuberculin would have repercussions for the TB eradication programme
Importance of having the injection sites in the same plane relative to the shoulder blade was confirmed
Tuberculin X failed to detect 20% of reactors with visible lesions and 9% of the visible lesion animals passed the SIT – the use of tuberculin of this potency could have serious repercussions and damage the reputation of the exporting country
Sensitivity and specificity is highly dependent on potency and the relative potency of both avian and bovine PPDs in the case of the SICTT
The World Organisation for Animal Health (OIE) and/or the EU in setting standards might consider if checks on tuberculin potency should be performed independently of the checks performed by the manufacturer so as to ensure quality of surveillance data and security of disease freedom certification based on the use of a variety of commercially available tuberculin PPDs.
We have been sent a link to www.fwi.co.uk/Articles/06/01/2010/119362/Strategy-to-eradicate-Johne39s-disease-needed-now.htm
The article (from Jan 2010) reports on Dairy UK's conference on Johne's disease. According to vet Keith Cutler if this disease is ignored 'it will rise unnoticed until it is almost impossible to do anything to control it'.
He also sai that it is rare you will see animals in severe stages of the disease becuase it's known that 70% of animals culled for 'other reasons' are culture positive for Mycobacterium paratuberculosis, the organism that causes Johne's disease.
Results from a Defra study released at the conference estimated that up to 42.5% of UK dairy herds were infected with this bacteria when sampled in 2006. This figure could have risen since then. Testing is problematic as they lack sensitivity with many false negatives.
In Denmark the disease is in 80-85% of cattle!
We have been sent this link http://www.agriview.com/news/dairy/patton-finds-teachable-moments-in-johne-s-program/article_b7a127c0-1b75-11e1-a128-001cc4c002e0.htm, which mentions Johne's disease.
Of particular interest is; 'The US Department of Agriculture has specific conditions we have to meet in order to vaccinate herds for Johne's, because it can lead to an increase in false positive tests for bovine tuberculosis, so our field staff used to do vaccinations, ...'
... and when you consider when you consider that (from same article) 'Johne's disease is a chronic, contagious bacterial infection that infects an estimated two-thirds of the nation's dairy herds', (relating to USA) just how many reactors/IRs are due to Johanes - which is caused by a Mycobacteria too?
Interesting paper from back in 2009 relating to Northern ireland. The Control of Bovine Tuberculosis in Northern IrelandREPORT BY THE COMPTROLLER AND AUDITOR GENERAL NIA 92/08-09, 18 March 2009 available at http://www.bovinetb.info/docs/Bovine_Final.pdf
Of particular interest:
Re testing carried out by private vets or in-house staff:
'However, within the same ‘at risk’ type of bTB test, PVPs detected markedly fewer reactors than in-house staff who were almost twice as likely to classify a herd as a breakdown herd. '
... and on fraud:
'It is a matter of concern that two herd owners successfully prosecuted for fraud received a total of £6,400 compensation for subsequent bTB outbreaks.'

According to This is Devon (www.thisisdevon.co.uk/President-aims-answer-bTB-cull-questions/story-13236670-detail/story.html) a farmer in the Westcountry is in the process of suing his own vet after one of his animals tested positive following the bTB skin test. The vet apparently injected the tuberculin into the animal's shoulder, for some reason, rather than on the neck, where it should have been done. The cow was in a crush and was not causing a problem.
The animal was subsequently found to be clear, and the farmer remonstrated to the AHVLA about the situation, who told him that careful siting of the test on the animal was critical: "because the sensitivity of the test varies in different areas."
As he said: "Vets are paid to do the tests – and should do them properly. Farmers should be aware of this. They could be 'going down' with TB when in reality they are clear."
The experiences of this Westcountry milk producer could be frighteningly common.
http://blog.lancashirebadgergroup.org.uk/wp-content/uploads/2011/07/Animal-Health-Official-Veterinarian-Newsletter-March-2011.pd... On page 10 of this 2008 document, is an example of a Friesian cow with chronic mastitis. It had passed the TB skin test 11 times since 2003. In February three calves suckling from her tested positive to TB - one of these was classed as a reactor when TB tested and had lesions at post-mortem. The cow thought once again passed the skin test! Consent was given for the cow to have a blood test. She was positive to both the gIFN test and a Rapid Antibody test, so was slaughtered. The carcass was condemned - it had so many lesions.
We have been sent details of this paper from 2008 which indicates some of the failings f the existing skin test. (http://veterinaryrecord.bmj.com/content/163/12/357.abstract):
Outbreak of bovine tuberculosis featuring anergy to the skin test, udder lesions and milkborne disease in young calves
M. G. Houlihan, MVB, MSc, MRCVS1, F. W. Dixon, BVM&S, MRCVS2 and N. A. Page, BVSc, MRCVS2
A severe outbreak of bovine tuberculosis in a 1300-head, multisite dairy herd in Great Britain had several unusual features, including anergy to the tuberculin skin test, milkborne disease in calves and a farm cat, and a risk of human infection. The outbreak was controlled by culling 221 cattle over 15 months, by using the γ-interferon (γ-IFN) test and by the examination of milk samples. The γ-IFN test detected infected animals that were not detected by the skin test.
... and from another fed up farmer (email 3.9.11) who is appealing against a test that was not undertaken properly.
He says that from what he has read it is wholly wrong (and not in the law or the Order) that negative tests can be ignored (and imputed as positive). Nor is it provided that two subsequent clear tests should be imposed where there is no actual sign of infection. It is wrong in logic that all bumps are treated by default as tuberculin reactions. It is unsound that a reaction to M.avium can be treated by DEFRA as a reaction to M bovis for the purposes of extending movement restrictions. Moreover, it is unsafe in law that unless all bumps are recorded that subsequent bumps are per se caused by a reaction to an injection at a SCTT test. It is also not in the law that where a culture is negative the AHVLA can proceed as if it were positive. It requires reasonable belief that there is an infection! The vets are misinterpreting or ‘gold plating’ the legislation!
He is appealing against the movement restrictions imposed as the one reactor identified was wrongly identified as such. The test was not carried out properly and a formal complaint has been made.
So here (www.thisisdevon.co.uk/President-aims-answer-bTB-cull-questions/story-13236670-detail/story.html) we have an indication that yet another farmer is not happy with the procedure regarding testing.
The article says:
'Meanwhile arguments about the cattle-testing methods have gone quiet – though the experiences of one Westcountry milk producer could be frighteningly common.
He is in the process of suing his own vet after registering a positive to the TB test for the first time ever, the vet having made the test on the animal's shoulder, for some reason, rather than on the neck, where it should have been done. The cow evidently was in a crush and was not causing a problem.
The animal was subsequently found to be clear, and the farmer remonstrated to the AHVLA about the situation, who told him that careful siting of the test on the animal was critical: "because the sensitivity of the test varies in different areas."
As he said: "Vets are paid to do the tests – and should do them properly. Farmers should be aware of this. They could be 'going down' with TB when in reality they are clear."'
Email from Prof PT 16/8/11.
The skin test is a measure of the immunological response to infection. ie. a positive response tells you the animal has been exposed and mounted an immune response. That is not the same as being infected. However, the skin test as used in cattle is highly specific. The false positive rate is less than half of 1 percent of healthy animals. Indeed DEFRA do claim to find evidence of leisions or positive bacteria culture from most animals who skin test positive, so the test is probably quite reliable when it is positive....But the presence of lesions or even bacteria in the lesions does not mean the animal could not mount a successful immune response and clear the infection....
There is one theory that suggests that the present skin test is actually selecting for cattle who do not mount an immune response - all the cattle that response immunologically are culled, leaving the cattle that do not mount such a response infected and in the national herd...just a thought...
Email from MR 16/08/11
The stock Defra excuse for slaughtering every animal failing a BTB skin test is caution (culling 'just in case' – however unreliable the test the animal just might be infected.
However, when humans were routinely given BCG they were tested using essentially the same skin test. The reactors were declared immune and not vaccinated. Many people are telling us about their memory of this in childhood, but none seem to have any recollection of any immediate or ongoing investigation for symptoms of TB in the reactors.
Surely if cattle are killed as a precaution, humans in the same circumstances would be examined for signs of actual TB and then kept under observation in the future, yet this does not seem to happen?
Is there an explanation for this apparent inconsistency?
Media responses from Defra have regurgitated its usual responses regarding test reliability and has not even replied properly to some of the questions, including the one on costs. I find it hard to believe they can still claim the test was done properly when the court case confirmed it was not.
As usual Defra has failed to say that the test only indicates the animal has been exposed to the bacteria and (if we look at the human parallels) most would not be likely to succumb to the disease and are killed 'just in case'. It is a shame that more farmers do not challenge the testing as this would force change. Of course, the costs and stress preclude this for most farmers. The Jacksons were exemplary because Boxster is a champion and they are very fond of him but have obviously been put through hell and significant costs over the last 18 months - .

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