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This article was published in 1969-70
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Bovine Brucellosis
COMPARISON OF THE RAPID WHOLE BLOOD AGGLUTINATION TEST AND MILK RING TEST WITH THE SERUM AGGLUTINATION TEST
P. CLAXTON, B.V.Sc., Veterinary Research Officer, Glenfield
Mr. Gill previously reported on a small preliminary trial with whole blood antigens conducted at Glenfield on 1/3/'68 on a few cattle reacting serologically to Brucellosis as a result of prior Strain 19 vaccination. In that trial the antigen composed of 4 per cent cells was thought to be superior to the antigens containing 8 and 11 per cent cells (concentration) so that Mr. Gill asked C.S.L. whether small quantities of both 11 per cent (C.S.L. standard line) and 4 per cent could be prepared for distribution to laboratories in the various States.
Following more extensive trials on naturally infected herds reported in this submission both the 11 per cent and 8 per cent cell antigens proved more efficient than the 4 per cent cell suspension so that Mr. Gill has now asked C.S.L. to make supplies of all three antigens available to the State Laboratories for further comparative work.
Three milking herds in the Bathurst district of N.S.W. were selected by Mr. B. P. Healy, the D.V.O. at Orange, because of current Brucella abortions, because there was no prior history of Strain 19 vaccination and because calving and abortion histories were available. The herds were tested on 10/4/'68 using the same three antigens as used at Glenfield on 1/3/'68. Mr. C. A. W. Jackson with considerable experience in pullorum testing and pullorum antigen examination and Mr. P. D. Claxton the officer responsible for serological tests at Glenfield independently read the whole blood agglutination tests (W.B.A.T.) while Mr. Healy and Mr. Irvine, the Veterinary Inspector, collected blood samples for subsequent serum examination at Glenfield (S.A.T.), milk quarter samples for Milk Ring Tests (M.R.T.) and vaginal mucus for V.M.A. tests. Unfortunately insufficient mucus could be collected from many cows so that V.M.A. tests have not been included in the results.
Mr. Healy carried out the Milk Ring Tests on quarter samples, on Cow samples and on cow samples diluted 1 in 20 with non-reacting milk. Only the "cow" and "diluted cow" results are included in Tables 1-3.
Although M.R.T. results look very promising when compared with serum tube results, they have not yet been summarised or analysed.
The W.B.A.T. was conducted with a similar technique to that used for pullorum disease in poultry, 0.04 ml antigen was delivered accurately by dropper on to an illuminated acrylic plate and 0.04 ml blood collected with a loop and adequately mixed with the antigen. Reactions, which were read after two minutes, were recorded as - Trace, 1, 2, 3 and 4 as with pullorum tests. Intermediate reactions were recorded as 1+, 2- etc., in Tables 1-3 but the + - signs were ignored in subsequent tables. Reactions appearing after two minutes were recorded as late reactors but they have been excluded from the results as they did not appear to improve the efficiency of the test.
The S.A.T. were performed according to the modified W.H.O. technique as recommended for veterinary laboratories in Australia, but with the maximum or final titres being recorded.
The total number of animals tested in the three herds was 121. The results from six animals (four in Herd "B" and two in Herd "C") were rejected from the analysis due to clotting in the W.B.A.T. Of the remaining 115 animals, 38 reacted (either positive or doubtful) in the S.A.T. For the purpose of correlation the S.A.T. was considered to be 100 per cent accurate in identifying infected animals.
The individual animal test results in the three herds are given in Tables 1-3 with separate observations of the two W.B.A.T. operators. The summarised results of each of the three herds, again with both operators, are given in Tables 4-6. There was little variation in the reading of the tests by the two operators so that the mean reading only has been recorded in Tables 7-9. These last three tables summarise the results of testing of the total 115 animals, the final Table No. 9 giving the per centage correlation or agreement between the W.B.A.T. and the S.A.T.
INTERPRETATION OF THE BRUCELLA ABORTUS SERUM AGGLUTINATION TEST
Female Elgible Animals
The Tables of interpretation are based on three international unit standards:
1. Animals vaccinated under the Ministry's Calf Vaccination Scheme (Official Vaccinates) and with a Brucella antibody content of less than 50 1.U. will be classified as "Pass" (Vacc. at 3-6 months of age).
2. Animals which are not Official Vaccinates (or are of unknown vaccinal status) and with a Brucella antibody content of less than 30 1.U. will be classified as "Pass".
3. Animals with a Brucella antibody content of 100 I.U. or more will be classified as "Fail".
TABLE 1. Serum Agglutination Test
| Pass | Inconclusive | Fail | |
|---|---|---|---|
| Official Vaccinates | Negative to +++ 1/20 | ++++ 1/20 to +++ 1/40 | ++++ 1/40 and over |
| Non-vaccinated or of unknown vaccinal status | Negative to ++++ 1/10 | + 1/20 to +++ 1/40 | ++++ 1/40 and over |
| Bulls | Negative to ++++ 1/10 | + 1/20 to ++++ 1/20 | + 1/40 and over |
TABLE III. Table of International Units (1.U.) of Antibody S.A.T.
| Final dilution of serum | End point reading | I.U. | |
|---|---|---|---|
| 1/10 | — | 0 | <30 |
| + | 16.75 | ||
| ++ | 20.00 | ||
| +++ | 23.25 | ||
| ++++ | 26.5 | ||
| 1/20 | + | 33.5 | <50 |
| ++ | 40.00 | ||
| +++ | 46.5 | ||
| ----- | |||
| ++++ | 53.0 | ||
| 1/40 | + | 67.0* | <100 |
| ++ | 80.00 | ||
| +++ | 93.0 | ||
| ----- † | |||
| ++++ | 106.0 ‡ | ||
| 1/80 | + | 134 | <200 |
| ++ | 160.00 | ||
| +++ | 186 | ||
| ----- | |||
| ++++ | 212 |
* (4/4/1)
† U.K. Fail point and now Australian Standard
‡ (4/4/4)
Male Eligible Animals Other Than Those at A.I. Centre
Animals with a Brucella antibody content of less than 30 I.U. will be classified at "Pass".
Animals with a Brucella antibody content of 67 I.U. or more will be classified as "Fail".