Examination of milk :
Inflammation of udder called mastitis which lead to change in
characteristics of milk .
Mastitis
clinical mastitis
Sub clinical mastitis
(without clinical signs)
Three types of Clinical Mastitis :
1. Acute clinical mastitis
2. Sub acute clinical mastitis
3. Chronic clinical mastitis
Etiology of mastitis :
1. Causes of contagious mastitis , particularly Streptococcus
agalactia , Staphylococcus aureus
2. Causes of environmental mastitis : E.coli, Streptococcus
dysgalactiae , S. uberis
3. Normal teat flora : Staphylococcus hyicus , Staph. Epidermidis ,
Coagulase negative staphylococci and Corynebacterium bovis
Samples collection : Samples should be collected carefully as follows :
1- Clean the udder well by brushing off any dirt.
2- Wash the udder thoroughly with warm water then soaked in
disinfectant solution .
3- Allow the udder to dry and treat the teat orifice with tincture of
iodine solution or alcohol 70% and allow it to dry.
4- While the tincture of iodine is drying, label the tubes as to cow
and quarter from which the sample will be taken .
5- The cap of the sterile tube is carefully removed and held
between the fingers in such a manner that the inside of the cap
is facing down ward. The tube should be held at a slight angle
to prevent contamination of the sample by falling particles .
6- Immediately following collection, refrigerate the samples for
transportation to the laboratory .
Methods of examination of milk : including into
A- Physical methods of diagnosis
B- Chemical methods of diagnosis
C- Culture methods of diagnosis
A- Physical methods of diagnosis :(colour and consistence of udder,odor
or smell of milk ).
B- Chemical methods of diagnosis : determination specially sub-clinical
mastitis including into( Card field test, pH test , Chloride test, White
side test, Calfornia mastitis test , Catalase test , Somatic cell count ,
Microscopic examination of incubated milk )
1- pH determination : Normal pH of milk (6.4-6.8) paper strips
contian Bromthymol blue or Bromcresol purple .
2- Chloride test : The chloride test is dependent upon the
determination of an abnormal quantity of chloride in the milk.
Normal milk contains 0.08-0.14 % chloride .The test is conducted
by adding 5ml of silver nitrate solution to 1ml of milk, followed
by the addition of two drops of a potassium chromate solution and
mixing by inversion of the tube .The appearance of yellow color
indicator that more than 0.14% chlorides are present in the sample
and brownish red color indicates that the sample contain less than
that amount. Cows in either early or late lactation may give false
positive reaction to the chloride test because of normal physiologic
processes of the udder .
3- Catalase test : Most living cells (RBCs catalase help to release
oxygen from peroxide hydrogen therefore action determination
that amount catalase in WBCs milk) catalase isn’t found in milk
normally except in the early and finally period lactation .2ml of
milk incubated for 1-2 hours and then add drops from H2O2 and
the positive test appearance bubble airs and negative test
disappearance bubble airs .
4- White side test : Put 5 drops of milk on dark smooth surface on
which the test can be conducted and then add 2 drops of sodium
hydroxide 4% solution for 20-25 second. Milk from
normal
animals will have no change after the addition of the NaOH, but
mastitis milk appear as the following :
 Nagative (-) The mixture is milky and opaque and entirely
free of precipition .In such animals ,the leukocyte count is
generally under 500,000 cells /cm3 .
 1 + The background is less opaque but still somewhat
milky, with large particles of coagulated material being
present and thickly scattered throughout the area .A slight
degree of clumping is observed .The leukocyte count is
usually between 1 and 2 million .
 2 ++ The background is more watery and large clump of
coagulated material are present. If the stirring has been
rapid ,fine threads or strings may be present .In such
specimens the total leukocyte count is usually over 2
million.
 3 +++ The background is very watery and whey like with
large masses of coagulated material forming into strings and
shreds. the total leukocyte count is usually several million .
5- California mastitis test : take 2ml of milk with 2ml CMT solution
(2:2) mixing rotation movement and then for 10 second read
results:
 Negative
(-)
the
mixture
remains
liquid
without
precipitation
 1+ distinct precipitate but no tendency toward gel formation
 2++ the mixture thickens immediately, with mucous
formation moving slowly on the paddle .
 3+++distinct gel forms that tends to adhere to the bottom of
the paddle and during swirling distinct central peak forms .
6- Direct microscopic total leukocytic count : taken drop of milk put
in slide spreading and dryness staining by (Newman's-lampert
stain) for 2 minute and dryness by filter paper and then washing by
distill water and dryness after that count by microscopic and
calculated in 30 fields = N/30 X400.000
N= number of cells
400.000= Zoom factor
7- Microscopic examination of incubated milk : taken precipitate
milk which incubated 12-16 hours and putting in slide spread drop
and left air for dry and then staining by gram stain
diagnosis )
 Take loopful for primary culture .
 After 24 hours at incubator then make gram stain .
(rapid
C- Bacterial Culture methods of diagnosis :
Primary culture
Culture
Secondary culture
Specimens (Milk)
Primary culture (Nutrient agar ,Blood agar , Macconkey agar.)
After 24h.incubated
Gram stain
(Genus of M.O.)
Secondary culture
Biochemical test
(oxidase, catalase, indol test.. etc.)
(Differential diagnosis for Species of M.O.)