SEMINAR ON MICROBIAL TECHNIQUE

8/7/2019 SEMINAR ON MICROBIAL TECHNIQUE

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B Y ,

KASTURI PANDA

M. PHARM

SEMINAR ON MICROBIAL TECHNIQUESUSED IN TESTING OF WATER & AIR.


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INTRODUCTION.

Water is elixir for life. It is an essential part of protoplasm & creates a state for metabolicactivities to occur smoothly. No life can exists without water. Major areas (about ) ofearth surface is covered by water , mainly by oceans, & to some extent by lakes , river ,streams, ponds etc. water receives micro-organism from air, soil, air, sewage, organicwastes, dead plants & animals etc.

Types of water-

1-atmospheric water.

2-surface water.

3-stored water.

4-ground water.


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WATER MICRO-ORGANISM

A large no of micro-organism are found in water like fungi, bacteria, protozoa. some ofthe aquatic fungi in different water bodies are-Rhizophydium sp., Achlya

americana., A. debaryana, A. laevis.Some of the water borne pathogenic microbes are-

Bacteria- Mycobacterium tuberculosis, Pseudomonas aeroginosa, Salmonellaenteriditis.

Protozoa- Giardia lamblia, Entamoeba histolytica, Cryptosporium.

Microbiological analysis of water purity.

The Faecal coliform-these are intestinal bacteria which are present in water due tofaecal contamination. Coliform are mambers of family Enterobacteriaceae whichincludes E. coli, Enterobacter aerogens, Salmonella & Klebsiella pneumoniae. Ifsuch organism are found in water it is not suitable for drinking.


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Sanitary test for coliform.

The original test for the presence of coliform in water is done by standardmultiple tube fermentation techniques. These are

1-The presumptive test.

2-The complete test.

Presumptive Test- A series of fermentation tubes each containinglactose broth or lauryl tryptose broth of known conc. are incubatedwith known amount of water. The tubes are incubated for 24 to 48hr at 35 c. Here 15 tubes are used,5 tubes with 10 ml, 5 with 1 ml, 5with 0.1ml. At the end of incubation , the indicates the presence orabsence of coliform.

Complete test This test is to ascertain about the presence of coliformin water.Each colonies from positive confirmed test is transferred tolactose fermentation tube and to nutrient agar slants. The tubes are

incubated at 35c for 48 hr. Production of gas in the fermentedtubes shows positive test.


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The most probable no of (MPN) of coliform

MPN is used to calculate the no. of coliform micro-organism in 100ml ofwater. Techniques used are-

1-Membrane filtration techniques- it is also known as millipore filter tech.A sterile membrane filter (0.45 m) disk is placed in the sterilizedholding apparatus. A volume of water is passed through filter disk.

Bacteria present in the water sample are retained in filter disk. Thesides of the funnel & membrane are rinsed with sterile water.Thereafter the membrane filter disk is aseptically removed & placed onadsorbent disk saturated with cultural media & contained in petridish.The medium passes through the pores of membrane & nourish thebacteria present in it.After incubation at 35c for 24hr each bacterium

multiplies to form visible colony, which can be easily counted.


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MPN of Coliform.

2-The defined substrate test- it is used for both coliform & E. coli in asingle 100ml water sample.

A specialized medium containing O-nitrophenyl-beta-D-galactopyranoside(ONPG) & 4- methylumbelliferyl-beta-D-glucuronide (MUG) is added with water & incubated for 24 hr at 35c.If coliform are present the medium will turns yellow within 24 hr dueto breakdown of ONGP.When E. coli are present, the MUG is changed

to yield a fluorescent product. The fluorescence is observed underlong wavelength UV light for the presence of E. coli.

3-IMViC test the IMViC represents indole, methyl red, Voges-Proskauer & citrate test. This test is used to distinguish between E.coli & Aerobacter aerogens .


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Characteristic IMViC reaction on E. coli & A. aerogens

Test E. coli A. aerogens

Indole Indole is producedfromtryptophan(+)

Indole is not produced(-)

Methyl

red

E. Coli produces acid that drops

pH below 4.5.hence methylred

turned red(+).

Methyl red remains yellow

due to production of less acid

(-).

Voges-

Proskau

er

E. Coli does not produce acetyl-methylcarbinol in glucose-peptone

medium (-).

Acetylmethylcarbinol isproduced by this bacterium(+)

Citrate Citrate does not support thegrowth of E. coli.(-)

Citrate support the growth of

bacterium(+).


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Purification of water (water treatment).

There are several methods for purification of water the use of whichdepends upon amount & quantity of water. There are three chief

methods which are used for the purification of water.

1-Sedimantation.

2-Filtration.

3-Disinfection.


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Testing compressed airlines for microbialcontamination

A micro-organism that is capable of forming a viable colony forming unit(CFU) & which exist within the compressed air lines system is calledthe microbiological particle (MP) as per ISO. The microbial load on aproduct is referred to as BIOBURDEN. The bioburden came fromdifferent sources such as human contact, air conditioning system, themanufacturing process itself, raw materials etc.

Routinely compressed air lines should be tested on a routine basis inorder to determine the microbiological cleanliness of the air suppply.


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Test equipment.


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Methodology.

The slit sampler (also known as slit-to agar sampler) is a device whichutilizes a rotating stage which holds the petri-plate. The air impacts thesurface of agar with whatever organism are present, the organismbecame impinged onto the agar, the plate is incubated, & the organismare allow to grow. Care should be be taken to insure that theequipment used has been properly disinfected in order to minimize theintroduction of bacteria not associated with the compressed air supply.A negative control should be used as a qualitative measure, and the endpoint to be tested should be purged & aseptically cleaned to minimizefalse result.


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THANK YOU.


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SEMINAR ON MICROBIAL TECHNIQUE