Practical Food Microbiology PDF

Culture medium Liquid Solid medium medium Liquid medium: Diffused growth No characteristics for identification Difficult to isolate Earliest liquid medium: urine...

Culture medium Liquid Solid medium medium Liquid medium: Diffused growth No characteristics for identification Difficult to isolate Earliest liquid medium: urine or meat broth used by Louis Pasteur Solid medium: Distinct colony morphology Characteristics → easy to identify 17  Colony – macroscopically visible collection of millions of bacteria originating from a single bacterial cell Earliest solid medium: Cooked cut potato by Robert Koch Gelatin - not satisfactory - liquefy at 24oC Agar Universally used for preparing solid medium Obtained from seaweed: Gelidium No nutritive value Not affected by the growth of the bacteria. Melts at 98°C & sets at 42°C 2% agar is employed in solid medium Types of culture media: Based on their consistency a) Solid medium b) Liquid medium c) Semi solid medium 18 Based on the constituents/ ingredients a) Simple medium b) Complex medium c) Synthetic or defined medium d) Special media Special media – Enriched media – Enrichment media – Selective media – Indicator media – Differential media – Sugar media – Transport media – Media for biochemical reactions III. Based on Oxygen requirement - Aerobic media - Anaerobic media Solid media (solid-reversible to liquid) – contains 2% agar Colony morphology, pigmentation, hemolysis can be appreciated. 19  Eg: Nutrient agar, Blood agar Liquid media – no agar. For inoculum preparation, Blood culture, continuous culture. Eg: Nutrient broth Semi solid medium – 0.5% agar. Eg: Motility medium Simple media / basal media: Most common in routine diagnostic laboratories Eg: Nutrient Broth, Nutrient Agar NB consists of peptone, meat extract, NaCl, water NB + 0.5% Glucose = Glucose Broth NB + 2% agar = Nutrient agar Agar conc. Reduced (0.2 - 0.5%) = Semi-solid medium Complex media Media other than basal media. They have added complex ingredients such as yeast extract or casein hydrolysate, which consist of a mixture of many chemical species in unknown proportions Provide special nutrients Synthetic or defined media Media prepared from pure chemical substances exact composition is known 20  Used for special studies, eg. metabolic requirements Eg: peptone water- (1% peptone + 0.5% NaCl in water) Enriched media Liquid media used to isolate pathogens from a mixed culture, contain the nutrients required to support the growth of a wide variety of organisms, including some of the more fastidious ones. They are commonly used to harvest as many different types of microbes as are present in the specimen. Blood agar is an enriched medium in which nutritionally rich whole blood supplements the basic nutrients. Chocolate agar is enriched with heat-treated blood (40–45 °C), which turns brown and gives the medium the color for which it is named. Eg: Selenite F Broth – for the isolation of Salmonella, Shigella Tetrathionate Broth – inhibit coliforms Alkaline Peptone Water – for Vibrio cholerae Selective media The inhibitory substance is added to a solid media Increase in number of colonies of desired bacterium Eg: Desoxycholate citrate medium for dysentery bacilli Mac Conkey’s medium for gram negative bacteria TCBS – for V. cholerae LJ medium – M. tuberculosis 21 Indicator media contain an indicator which changes its colour when a bacterium grows in them Eg: Wilson-Blair medium – S. typhi forms black colonies McLeod’s medium (Potassium tellurite)– Diphtheria bacilli Urease medium (yellow oranged): Urease producing bacteria Urease Urea → CO2 + NH3 NH3 → Medium turns pink Blood agar: shows three types of Hemolysis α Hemolysis β Hemolysis γ Hemolysis Differential media Substances incorporated in it enabling it to distinguish between bacteria. Eg: Mac Conkey’s medium – Peptone – Lactose – Agar – Neutral red 22  – Taurocholate Distinguish between lactose fermenters & non lactose fermenters. MacConkey agar: Lactose fermenters – Pink colonies Non lactose fermenters – colourless colonies Sugar media Media containing any fermentable substance Eg: glucose, arabinose, lactose, starch etc. Media consists: 1% of the sugar in peptone water + Indicator Contain a small tube (Durham’s tube) for the detection of gas by the bacteria Transport media Media used for transporting the samples. Delicate organisms may not survive the time taken for transporting the specimen without a transport media. Eg: – Stuart’s medium – non nutrient soft agar gel containing a reducing agent & charcoal used for Gonnococci – Buffered glycerol saline – enteric bacilli 23 Anaerobic media These media are used to grow anaerobic organisms. Eg: Robertson’s cooked meat medium, Thioglycolate medium. CULTURE METHODS Culture methods employed depend on the purpose for which they are intended. Purposes: – To isolate bacteria in pure cultures. – To demonstrate their properties. – To obtain sufficient growth for the preparation of antigens and for other tests. – For bacteriophage & bacteriocin susceptibility. – To determine sensitivity to antibiotics. – To estimate viable counts. – Maintain stock cultures. Culture methods include: Streak culture Lawn culture Stroke culture Stab culture 24