BRUCELLA

AGGLUTINATION TEST
(BAT).

CLT 13 2023
BY

MR SHADRACH BMLS, Masters fellow MUST

Brucellosis a zoonotic disease
 INTRODUCTION
Brucellosis is a bacterial disease caused by
various Brucella species, which mainly infect cattle,
swine, goats, sheep and dogs.
Humans generally acquire the disease through direct
contact with infected animals, by eating or drinking
contaminated animal products or by inhaling airborne
agents.
 Most cases are caused by ingesting unpasteurized milk
or cheese from infected goats or sheep.
It’s a zoonotic disease (infect both animals and humans)
 4 major Species of Brucella
 Brucella abortus (cattle)
 Brucella melitensis (goats, sheep,
and camels)
Brucella suis (pigs)
 Brucella canis (dogs
 Laboratory tests for Brucellosis
  Brucella agglutination test
Brucella microagglutination test
Isolation of bacteria (brucella) culture
ELISA (enzyme linked immunosorbent
assay) brucella test
Polymerase chain reaction DNA
quantitative Brucella test
PRINCIPLE OF AGGLUTINATION TESTS
In agglutination tests, an antigen reacts with its corresponding
antibody, resulting in visible clumping of bacterial cells.
With latex agglutination tests, latex particles are coated with
antibodies that agglutinate specific antigens and form a more
easily visible precipitate. 
Agglutination tests are frequently used for initial confirmation
of specific pathogens. Since antibodies to the target organism
may cross-react with other organisms and autoagglutination
may occur,
these must be considered as screening tests and further
confirmation will usually be necessary
 BRUCELLA AGGLUTINATION TEST (BAT)
Introduction:
The Brucella agglutination test detects total antibodies to
the two commonly encountered
Brucella species, i.e. Brucella abortus and Brucella
melitensis.
This test is routinely used in
diagnostic laboratory and is generally considered to be
poor in terms of both specificity and
sensitivity.
 BRUCELLA AGGLUTINATION TEST (BAT)
 Purpose:
To aid in diagnosis of brucellosis and monitoring of
treatment.
Principle:
Standardized suspension of killed and stained Brucella
abortus or Brucella melitensis antigen agglutinate with
antibody present in the serum
Specimen required:
Serum
 
 Materials and reagents recquired for BAT
1.Agglutinating antigens for both B.abortus and B.melitensis.
2.Agglutination slides or cards
3.Khan tubes.
4.Test tube rack.
5. Water bath (370 C).
6. Pipettes (0.1 – 2ml).
7. 50µl Pipette.
8.Brucella Standard serum.
9.Centrifuge
10.Mechanical rotator/VDRL shake
 
 1. Slide agglutination test
This is a useful rapid screening test to detect a reactive serum
that requires titration.
Procedure:
1. Bring sample and reagent to room temperature.
2. Place a drop of the undiluted serum into a ring of the card.
3. Resuspend the Brucella antigen and add a drop onto the serum.
4. Mix and spread over the ring.
5. Rotate for not more than 3 minutes at 80 – 100 rpm.
6. Examine for agglutination.
 
Slide agglutination test for Brucella
 Interpreation of results

1. Visible agglutination …positive test

2. No agglutination …….Negative
Some times titer could be done on a slide though
not recommended
 2. Tube agglutination test
Performed when slide agglutination test is positive and in all
strongly suspected cases of
brucellosis even when slide agglutination is negative
1. Make serial dilutions of the serum: 1:20, 1:40, 1:80, 1:160,
1:320 and 1:640.
2. Add antigen and mix.
3. Incubate tubes in 370 C water bath.
4. Read tubes over a mirror after 24 and 48 hours of
incubation.
Note: Manufacturers instructions should be followed when
performing titration.
2. Tube agglutination test
 Reporting of results
Reading of Results:
1. Examine for agglutination in the base of the tube.
2. Record results from ++ for strong agglutination, +
for weak agglutination and - for no agglutination.
3. The result is recorded as a titre and is the
reciprocal of the highest dilution showing
agglutination.
 Slide agglutination test
1. Titres of 20 – 40 are equivocal( not indicative
off an active infection)
2. A titre of 1:160 is often taken as indicative of
active brucellosis when clinical signs and
symptoms are suggestive