ISOLATION,

IDENTIFICATION&
PRESERVATIONOF
INDUSRTIAL
MICRO-ORGANISMS.
PresentedBy,
PRANAVH.NAKHATE
M.TechBiochemicalEngg.
AmityUniversity,Jaipur

POINTSTOBEDISCUSS
Why microbes are
important industrially ?
Isolation of microbes
Identificationof
microbes
Microbial
preservation
INTRODUCTION
Themicrobewillhavethelastword.
LouisPasteur
Industrialmicrobiologyassociateswiththe
commercialexploitationofmicrobesforthebenefit
ofmankind.
Thesemicrobialproductsmayhavedirector
indirectimpactontheeconomics,environmentand
socialparametersofthesociety.
Microorganismsareusedtocreateawidevariety
ofproductsandtoassistinmaintainingand
improvingtheenvironment

Microbeshavewidescopeindifferentareas.Some
ofthemare,
starterculturesincommercialfoodandbeverage
production.
Productsofalcoholicfermentation.
ProductionofEnzymesandotherindustrial
products.
ProductionofAlternativefuels.
ProductionofPharmaceuticals,likeantimicrobials,
recombinanthormones,andothercellregulators.
Pesticidesandagriculturalproducts
Biosensorsandbioreporters.
INDUSTRIALSTRATEGYFORUSAGEOF
MICROBES
Isolationofmicrobesfromnaturalsource.

Identificationofdesiredmicrobes.

Characterisationofmicrobes

Screeningofdesiredmicrobes.

Inoculumpreparation

SrainImprovement

Fermentation

ISOLATIONOFINDUSTRIALIMPORTANT
MICROBES
Isolationishavingmainly2strategies:
1)fromtheenvironmentbycollectingsamplesof
freelivingmicroorganismfromnaturalhabitats.
2)Bysamplingofmicrobeswithdesired
characteristicsfromspecificsitessuchas
foundamongthenaturalmicroflora.
Hence,itsimportanttoisolatethatspecific
microbefromtheenvironment.
Thesiteforisolationofmicrobesisdecidedby
thetypeofmicrobeoneisinterestedin.

Sr.No. PRODUCT SITE

1 Thermophiles Hotspring,thermophilicvents,
compost
2 Alkalophiles Highsalinitysoilsandwells
3 Acidophiles Pinegrowingsites,fruits

4 Osmophiles Seasandoceans

5 Yeast Wineyards
6 Phosphatesolubilizers Agriculturalsoil,rhizosphere

7 Proteaseproducers Dairyindustry,meat
processingunits.
8 Enzymeslikecellulase,
pectinase,xylanaseetc.
Agriculturalsoil,paperand
pulpindustry,waste
disposalareaoffruitjuice
plants,grainmarket.
9 Biogas(methane)producers Gober-gasplants,effluent
collectionsites.
Cont.(Ref.2)
Aftersamplingoftheorganismthenextstepisof
enrichment.
Thisincreasesthequantityofthedesiredorganism
priortoisolationandscreening.
Althoughnumerousmediahavebeendevisedfor
thegrowthofmicrobes,thereisnosinglemedia
thatsupportsallkindsofmicrobes.
E.g.Asoilsampleplatedonnutrientagar(pH7.0),
incubatedat37Cfor24-48hrs,willgrowonly
aerobicmesophiles,butanaerobes,thermophiles,
psychrophiles,alkalophilesandacidophileswillnot
grow
IDENTIFICATIONOFINDUSTRIALIMPORTANT
MICROBES(Ref3)

Aftertheisolationofbacteria,itisimportant
toidentifythespecifiedbacteria.
Identificationismostlydonewiththehelpof
followingcharacteristics
1)Morphology,
2)Selectiveanddiagnosticmedia,
3)Culturalcharacteristics,
4)Additionalrecognizedbiochemicaltests,
5)Profileofmicrobialstains,and
6)Rapididentificationmethods.

1)Morphology:-
Itisthesciencedealingwiththe
structuresandformsoforganisms.
Itexclusivelytrustsconfidentlyupona
numberoffactors,suchasshape,length
andbreadth,endsoforganism,body
arrangement,motility,formsspores,
capsule

2)Selectiveanddiagnosticmedia:-
Itspecificallyfavourthegrowthof
particularmicroorganisms.
Eg.MacConkeysagarmedium
forE.coli.
Differentialmedia,Enrichment
media,andCharacteristicmedia
aretypesofthismedia.
3)Additionalrecognizedbiochemicaltests:-
Thesetestsareextensivelyusedfor
identificationpurpose.Thesetestsare,
Carbohydrate(Sugar)Fermentation,
LitmusMilk,IndoleProduction,MethylRed
Test[MR-Test],Voges-ProskauerTest
[VP-Test],CitrateUtilization,etc.

4)Profileofmicrobialstains:-
microbesareextremelytoosmallinsize,
hencetovisualisethem,stainingistakes
place,inwhicheithercellinterioror
exteriorpartstainsincolour.
DifferentStainingtechniqueincludes,
Gramstaining,Differentialstaining,
Capsulestaining;Endosporestaining;
Flagellarstaining.

5)Culturalcharacteristics:-
Baseduponabroadspectrumofintensiveandconclusive
researchcarriedoutduringthepastfewdecade
Thesecharacteristicsare,Shape,Size,ElevationMargins,
Surface,Edges,Colour,Structure,Consistency,etc.

6)Rapididentificationmethods:-
TheseincludesadvancedtechniqueslikeSEM,
TEM.
CULTUREPRESERVATION
(Ref4)
Theconventionalmicrobiologyistotally
dependentontheavailabilityofpureandstable
cultures.
serialtransferisoftenthefirstpreservation
techniqueusedbymicrobiologists,butcultureis
pronetocontaminationbythis.
Inordertopreventthelossofproductionofa
desiredmetaboliteandtopreserveculture,a
varietyofculturepreservationtechniqueshave
beendeveloped.

1)CultureTransfer:-Toavoidcontamination,genetic
change
2)Refrigerationfrom0
o
to5
o
C:-
shorttermstorage
3)LowTemperatureFreezing:-
ultralowtemp.freezer(-80
o
C),
liquidnitrogen(196
o
C)

4)Lyophilization:freezewithdryice
andacetonesublimeoffwater(dries
cellswithoutdisruption)useof
glycerol,orsucrosetoprotectcells
5)MineralOil
6)DrySpores

USESOFPROPERISOLATION,IDENTIFICATION
&PRESERVATION
ToImproveindustrialproduction.
Tostudypropermorphology&physiology
ofmicrobes.
Tounderstandmicrobialkinetics.
Microbialbiosensor.
BiopesticidesoverChemicalpesticides.

REFERENCE
Ref1-biotechnology.editor:u.satyanarayanan.publishers:
booksandallied(p)ltd.calcutta.india.2005.
Ref2PharmaceuticalMicrobiology,sixtheditioneditedby
w.b.hugo&a.d.russell,professorofpharmaceutical
microbiologyuniversityofwalescardiff,cardiff
Ref3-Aamand,J.,T.AhlandE.Spieck.2006.Monoclonal
antibodiesrecognizingnitriteoxidoreductaseofNitrobacter
hamburgensis,N.winogradskyi,andN.vulgaris.Appl.
Environ.Microbiol.
Ref4-BeckerW.M.,KleinsmithL,andHardinJ.TheWorldof
theCell,4thed.,BenjaminCummings,RedwoodCity,Calif.,
2008.