Bacteriological Media:Cultivation of Microbes

Dr. Md. Abdullah YusufAssistant Professor of Microbiology

National Institute of Neurosciences & HospitalDhaka, Bangladesh

Email: ayusuf75@yahoo.com

Learning Objectives

• Definition of culture/cultivation and media• Types of culture• Types of microbes as per ability of culture• Basic equipment and logistic arrangement for

bacterial culture• Indication of culture• Classification, type and use of bacteriological

media

Definition

• Cultivation– Process of allowing bacteria to multiply in

artificial food providing optimum environment

• Culture Medium/media– Artificial food for bacteria for in vitro growth

Types of Culture

• Inanimate– Using nutritional ingredients but no living cells

or tissues– Used for culture of bacteria and fungus

• Animate– Using nutritional ingredients and metabolic

substrate for living cells or tissues to multiply (Tissue/Cell Culture)

– Used for in vitro multiplication of virus, rickettsia, chlamydia

Types of Bacteria as per ability to Culture

• Some bacteria can multiply easily– ordinary

• some with difficulties– fastidious

• some cannot– uncultivable

Conti….

• Fastidious– Neisseria gonorrhoea– Neisseria meningitides – Haemophilus influenzae

• Uncultivable– Mycobacterium leprae– Treponema pallidum

Culture technique

Materials Needed for Culture

• Specimen• Sterile Cotton Swab • Bacteriological Wire Loop • Suitable Culture Media• Incubator

• Aerobic• CO2/Anaerobic

Indication of Culture

• Isolation- diagnosis • Identification- diagnosis• Antimicrobial susceptibility- treatment• Genomic study- characterization• Progressive research- up to date information

Culture Media

ClassificationA. On the basis of consistency• Liquid• Solid• Semi-solid (gel)

Conti…

B. On the basis of nutritional ingredients– Simple or basal– Enriched– Selective– Indicator – Differential

Classification-contd

C. On the basis of use– Primary– Subculture– Transport– preservative

Important points

• Liquid media is also called broth• Solid media– Liquid media + solidifying agent in appropriate

concentration (5-10%)• Semisolid media– liquid media + solidifying agents at low

concentration (study bacterial motility)

Solidifying agents

• Agar agar (sea weeds)– melt at high temperature (55-600 C)– solidified at low temperature (40-450 C)

• Egg whole• Gelatin• Potatoes• Tomato powder• Serum

According to ingredients• Simple/basal– contain basic nutrient support

• Enriched– contain growth enhancing substances. e.g.- blood,

serum, haemoglobin• Selective– inhibitory substance that do not inhibit one

group/species, but inhibit all other• Semi-selective– if allow growth of one family-many members

• Highly selective– allow only one species of a particular genus

According to Ingredients (contd.)

• Indicator– contain chemical substance that changes it’s

colour with change of pH– indicate particular property of a genus of

bacteria• Differential– substance with change of colour (black colour

due metalic tellurium) or phenomenon (haemolysis) observed due to growth of a particular bacteria

According to Use

• Primary– first inoculation then transfer or not

• Subculture– subsequent inoculation from primary media

• Transport– Dissolve or mix clinical specimens or bacteria– allow little growth but no inhibition– transported from one location to another

• Preservative– dehydrated media containing metabolically inactive

bacteria – kept for prolonged period (lyophilized skimmed milk)

Subculture

How a medium can be made selective?

• Adding inhibitory substance– bile salt in MacConckey agar

• Adding antibiotic– Campybap for Campylobacter

• Adding alkali/ Changing pH– Sodium bi carbonate in alkaline peptone water

• Removing oxygen– by reducing chemicals or mechanically for

anaerobic bacteria• Changing temperature– thermophiles grow at high temperature

Advantages of solid media over liquid media

• Discrete growth as separate unit (colony)– hazy or opaque appearance in liquid

• Clear colony morphology• Easy to pick-up for subculture• Sensitivity test or further identification tests

Differential media

Haemolysis in blood agar

Blood agar

Thank You