Lakhasly

Step 1: Handling the test material obtained from a patient.Aspects of bacterial growth in liquid media: o Diffuse turbidity o Near-bottom and sidewall growth in a clear medium o Surface film formation o Cotton ball-like growth Aspects of bacterial growth in semi-liquid media: o Diffuse growth around the stab-inoculation site (motile bacteria) o Growth along the way of stab inoculation (non-motile bacteria) o Formation of gas bubblesAspects of bacterial growth on solid media: o Growth as individual colonies o Growth as a biofilm Microbial culture properties: 1.The saccharolytic activity (ability to break down sugars) and proteolytic activity (ability to break down proteins) are studied.To study the saccharolytic enzymes a pure bacterial culture is inoculated in Hiss medium.Preliminary microscopic examination of the smear is performed using the Gram staining to identify microbial genus present in the test material.Examine the pattern of the bacterial culture growth on the agar slant and confirm the colony purity by microscopy (Gram staining).As the colonies are typical of each bacterial species, they are an important diagnostic aspect that enables to identify the bacterial genus and species.Examination of the culture properties of isolated bacteria and description of the colony structure.Step 3: There are three main methods for the determination of microbial species (identification): 1.Inoculation of the test material on the culture medium using the streak plate method.Colony size is determined by its diameter (large colonies -- over 4 mm in diameter, medium colonies -- 2-4 mm, small colonies -- 1-2 mm, pin-point colonies -- less than 1 mm).Colony shape -- round, amoeboid, root-shaped, etc.Degree of transparency -- opaque, clear.2.3.4.

Step 1:
Handling the test material obtained from a patient. Preliminary microscopic
examination of the smear is performed using the Gram staining to identify
microbial genus present in the test material.
Inoculation of the test material on the culture medium using the streak plate
method.
Examination of the culture properties of isolated bacteria and description of
the
colony structure. As the colonies are typical of each bacterial species, they are
an
important diagnostic aspect that enables to identify the bacterial genus and
species.
Aspects of bacterial growth in liquid media:
• Diffuse turbidity
• Near-bottom and sidewall growth in a clear medium
• Surface film formation
• Cotton ball-like growth
Aspects of bacterial growth in semi-liquid media:
• Diffuse growth around the stab-inoculation site (motile bacteria)
• Growth along the way of stab inoculation (non-motile bacteria)
• Formation of gas bubblesAspects of bacterial growth on solid media:
• Growth as individual colonies
• Growth as a biofilm
Microbial culture properties:

1. Colony size is determined by its diameter (large colonies — over 4 mm in
   diameter, medium colonies — 2–4 mm, small colonies — 1–2 mm, pin-point
   colonies — less than 1 mm).


2. Colony shape — round, amoeboid, root-shaped, etc.


3. Degree of transparency — opaque, clear.


4. Colony color is determined by a pigment produced by the bacterial culture
   (buff,
   yellow, orange, blue, red, black, etc.).


5. Surface pattern — smooth, rough, glistening, dull, dry, moist, radially or
   concentrically wrinkled.


6. Colony edge/margin pattern — even, uneven.


7. Colony consistency — paste-like, mucoid, leather-like, etc.


8. Odor — many bacterial species release specific aromatic substances during
   the
   growth in the culture media. For example, Pseudomonas aeruginosa cultures
   smell like strawberries, and Proteus cultures have a putrid odor.
   Prepare a smear preparation from each colony type for Gram staining and
   examination of morphological and tinctorial properties.
   Step 2:


9. Subculture each type of the colonies onto agar slant to obtain pure bacterial
   cultures. To achieve this dip a sterile inoculation loop with the collected colony
   material in the condensed liquid at the bottom of the slant agar tube and, by
   slightly touching the agar surface, make zigzag movements in the upward
   direction. Briefly hold the edge of the tube over the flame of a burner, close the
   tube with a stopper, and sterilize the loop. Great caution should be exercised to
   collect the intended colonies without touching the adjacent ones.


10. Examine the pattern of the bacterial culture growth on the agar slant and
    confirm
    the colony purity by microscopy (Gram staining).
    Step 3:
    There are three main methods for the determination of microbial species
    (identification):


11. Biochemical identification. In this method microbial species are identified
    based
    on their exoenzymes. The saccharolytic activity (ability to break down sugars)
    and proteolytic activity (ability to break down proteins) are studied.To study the saccharolytic enzymes a pure bacterial culture is inoculated in Hiss
    medium. These media are intended for differential diagnostics and contain a
    certain carbohydrate and an indicator. The formation of acid and gaseous
    products
    due to bacterial fermentation of carbohydrates results in the change of colour of
    the medium and appearance of air bubbles. A Hiss media set containing
    different
    carbohydrates is also known as the “coloured row”. The Hiss “coloured row”
    containing mono- and di-saccharides is called a short row; and the one
    containing
    mono-, di-, polysaccharides, and higher alcohols is called a long row.
    Inoculation
    is carried out by stabbing microorganisms deep into the culture medium.
    To determine proteolytic enzymes a pure culture is stab-inoculated into a gelatin
    column. The process of gelatin liquefaction is microbial species-specific. The
    formation of indole, ammonia and hydrogen sulphide indicates more profound
    protein breakdown. To detect these substances a pure culture is inoculated into