Review and application of pooled testing strategies

1
Review and application of pooled testing
strategies

• Jim Kennedy
• Colorado State University
• Veterinary Diagnostic Laboratory
• Rocky Ford Branch

2
Why pooled testing?

• Pooled testing offers advantages over individual
  testing
• Allows the diagnostician to take advantage of
  highly sensitive and specific tests while
  minimizing cost
• Diminishes cumulative testing error over
  individual tests

3
Where testing error happens

• Pre-analytical error sources, wrong sample,
  mishandled sample, improper sample collection,
  etc. Starts from collection and goes until
  analysis begins.
• Analytical error, analytic variation such as
  mechanical wear and tear or inherent error such
  as that seen with a set of spring type scales.
• Biological variation, an average means some are
  higher and some are lower.
• Post-analytical error, reporting errors misread
  values or misreported values transposition of
  figures, etc.

4
Why not pooled testing?

• Potential impact of dilution diminishing Se
• Logistical requirements for pooling samples
  (pooling of individual samples can be labor
  intensive)
• Loss of samples for follow-up testing on positive
  pools

5
Assumptions associated with pooled testing

• Pooled test Se must be approximately the same as
  individual test Se
• Samples must be easily obtainable
• Pools must represent a homogenous mixture of
  samples
• The outcome is binomially distributed, i.e. a
  discrete probability distribution of the number
  of successes in a sequence of independent yes/no
  events each yielding success with a probability p

6
Our human counterparts institute pooled testing
strategies

• During WW II the military implemented pooled
  testing strategies for inductees
• Pooled testing for HIV is common today.
• The commonality between pooled testing in human
  medicine and veterinary medicine lies in herd
  diagnostics, more can be accomplished at less
  expense.

7
Veterinarians and screening tests

• Limited applications of screening test strategies
• Salmonella contamination of eggs
• Johnes fecal pools
• BVD
• T. foetus

8
Application of pooled testing for BVD and T.
foetus

• Push for BVD control/eradication stimulated a
  search for a method to screen herds for
  persistently infected animals.
• The prevalence of PIs was/is quite low
• Individual tests are expensive
• The use of a pooled strategy would lower the cost
  of testing and encourage more to adopt BVD
  control

9
In the past four years what have we learned about
pooled BVD testing

• A total of 4039 pools have been tested those
  pools represent 265,388 animals with an average
  pool size of 66.
• 205 pools have been found positive (5.1)
• From the 205 positive pools 513 positive
  individuals have been identified for an overall
  prevalence of .19
• The cost of testing had pooling not been done
  would have been 1,061,552 while with pooling the
  cost was 319,508, 70 less.

10
Pooled T. foetus testing

• Several states have mandatory T. foetus testing
  programs.
• Programs were based on culture with PCR
  confirmation of positive culture.
• Published Se for a single culture range from .7
  to .9 while PCR Se is published to exceed .98.
• Diagnostic labs charge 5 for culture and 25 for
  individual PCR.
• By pooling the charge per PCR could be spread out
  over several animals, provided Se was not
  severely compromised.

11
Applying pooled testing for T. foetus

• Maximum pool size of 5 samples was established.
• 2370 pools have been tested with 103 pools found
  positive (from April 07 to July 08)
• The 2370 pools represent 8567 bulls, 143 bulls
  were identified as T. foetus infected.

12
No laboratory cost advantage to pool T. foetus
samples

• Using culture the lab charge for testing the
  8,567 bulls would have been 42,835, and assuming
  the 143 positives were found an additional charge
  for PCR confirmation at 25 each would be 3,575
  for a total of 45,960.
• Laboratory charges for pooling for the 2,370
  pools would have been 59,250 plus the additional
  charge of 10,300 for identifying the positive
  member/s of the pool, for a total of 69,550

13
Then why pool for T. foetus

• During the same period 4,693 requests for culture
  with a PCR confirmation of culture positives were
  received with 69 positives and 39 confirmed as T.
  foetus by PCR yielding an apparent prevalence of
  .8.
• This compares to 1.7 by pooled PCR for the 8567
  samples. Assuming that both the culture group
  and the PCR group are representative of the same
  population it would appear that 86 bulls would
  have been misclassified as not infected.
• A look from a different angle is that the cost
  per positive bull by culture was 667 versus 486
  for pooled PCR 181 less, minor to the cost of
  missing 86 infected bulls.