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Bacteriological Examination of water, milk and air
Bacteriological Examination of water,
milk and air
Water
Milk
Air
Examination of water, milk and air
Importance of water examination for pathogens
Water intended for human consumption should not contain any pathogenic organisms.
Water is used for many applications either at home for cooking ,washing or drinking or in industries such as food and pharmaceuticals.
It is also important for hospitals for example haemodialysis unit
Testing of water samples are done regularly to make sure of its safety
Supplies of drinking water contaminated with sewage may cause diseases such as: typhoid fever and cholera.
All sources of water should be tested regularly.
Microorganisms which indicate the fecal pollution in water are usually common intestinal commensal bacteria.
Most important indicators of fecal pollution of water
Escherichia coli: The essential indicator of fecal pollution of human
/animal origin. It is an important member of the coliform bacteria.
Coliforms are members of the enterobacteriaceae family and they
1. grow in the presence of bile salts.2. produce acid and gas from fermentation of lactose at 37°C.
It is the commonly-used bacterial indicator of sanitary quality of food and water.
Enterococcus faecalis:• less numerous than E.coli in
human feces, but more resistant to chlorination.
Clostridium perfringens:• Less numerous in human feces• Its spores can survive in the
environment • Resist treatment processes than
most of the indicators.
Media used in bacteriological examination of water
1. For coliforms: MacConkey’s broth Containing bromocresol purple as the pH indicator.
To confirm the presence of E.coli : EMB agar + IMVC
Enterococcus faecalis: Glucose azide broth.
Clostridium perfringes: Differential reinforced clostridial medium.
Membrane Filtration Method
Determination of Most Probable Number (MPN) by dilution method
Pour plate technique
Methods Used in Bacteriological
Examination of Water
• Using Millipore Filter Apparatus
Membrane Filtration Method
MacConkey’s agar
Determination of MPN of Coliforms by Dilution Method
Water Sample
50 mlDSMB
5 x 10 mlDSMB
5 x 5 mlSSMB
50 ml watersample 10 ml water
sample1 ml water
sample
Results:
Positive tubes: showing production of acid or gas.
Acid production: change color of tube from purple to yellow
Gas production: detected in the Durham’s tube.
Purple Yellow
Gas
Determine no. of coliforms per 100 ml water sample (MPN) using the standard probability table.
1 3 2MPN = 14
i.e: No. of coliform bacilli per 100 ml water sample is 14 cells.
Most probable number of coliforms by McCrady’s table
Using 10 fold serial dilution methodViable Bacterial Count
9 ml Saline1 2 3Water sample
1 ml water
1 ml 1 ml
1/101/10 x 1/10
1/1001/100 x 1/101/1000
1 ml 1 ml 1 mlMelted NA
1 2 3
Results:
Dilution
factor 1 2 3 XX . y
10 x1 X1.y1
102 x2 X2.y2 103
x3 X3.y3
No. of colonies per plateY
No. of cells per 1 ml = X1.y1 + X2.y2 + X3.y3
3
Water
Milk
Air
Examination of water, milk and air
Human infections may be caused bytheingestion of animal milk whichcontains microorganisms derived from:
a. Animal e.g. by contamination with its feces
b. The environment c. Milk handlers such as dairy workers
Introduction:
Importance of milk examination for pathogens
It is important to examine milk for pathogens to ensure that it is safe to be consumed by man.
Milk is further used for
obtaining many milk products like cheese ,cream , butter and ice cream
E.coli Streptoccus pyogenesMycobacterium bovisBacillus anthracis Salmonella sp. Brucella sp.
Pathogenic bacteria present in milk
Determination of viable bacterial count:
Using the pour plate method after preparation of 10fold serial dilution from the milk sample with ringersolution.
Permissible number of bacterial flora in pasteurized milk is 5 x 104 cfu/ml
Permissible number of bacterial flora in long life milk is 10 cfu/ml
Methylene Blue Reduction Test
To determine quality of the milk Increasing the number of bacterial flora will reducethe color of methylene blue more rapidly due toincreasing consumption of oxygen.i.e.: The speed of reduction of methylene blue color isdirectly proportional to the number of bacteria presentin milk sample.
Methylene Blue Reduction Test
Results:
The shorter the decolorization time, the higherthe number of bacterial flora present in milk,and the poor quality of milk
Decolorization time Result
30 min – 2 hrs Poor quality 2 – 6 hrs fair quality 6 – 8 hrs good quality Over 8 hrs excellent quality
Test for coliforms Done by inoculation of MacConkey’s broth
with 0.1 ml of milk sample.
Examine for the production of acid detected by changing the color of the medium from purple to yellow.
+ve result with gas production
-ve result
Water
Milk
Air
Examination of water, milk and air
Importance of keeping the micro- organisms count low in air
Surgical theaters
Food preparations
Drug materials
Cross infection and out breaks in hospitals
Number on bacteria in air depends on
Number of persons
Body movement
Disturbance of clothing
Methods of examination of aira. Settle plate: Petri dishes containing an agar medium are left open
for a measured period of time. Large bacteria-carrying dust particles settle on the
medium. The plates are incubated and a count of the colonies is
formed
Blood agar is suitable for over all count
For detection of a particular microorganism suitable media is used .
Disadvantage of this method :Despite its simplicity it measures only therate of deposition of large particles fromthe air
b. Slit sampler It draws in air from the environment at a fixed rate and
causes the suspended particles to fall on the surface of the agar plate.
c. Air centrifuge
Centrifuging particles from the air on to a culture medium.
The sampled air passed along a tube lined with nutrient agar which was rotated on its long axis.
After sampling the strip is removed from the instrument and incubated then colonies can be counted.
Notice:
No level of contamination however low can be regarded as certainly safe.
Infection can be initiated by deposition of a single infected particle at a favorable site.
The probability of S. aureus initiated infection is low in comparison with Mycobacterium tuberculosis
Demonstration:
Air examination Settle plate Water examination Determination of MPN Milk examination Methylene blue reduction test
