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Cell Wall

Protects and maintains the shape of the organism.

There are 2 basic cell walls, Gram positive and Gram negative.
Membrane Filter
When there is a small number of organisms in a large amount of liquid, the liquid is filtered through a membrane and then the membrane is placed on agar in a petri dish.
Lag Phase
Organisms are adjusting to the environment ( little or no division). They are synthesizing DNA, ribosomes and enzymes to breakdown nutrients, and to be used for growth.

Size varies from 0.2 - 60 µm.
Extra Cellular Matrix
It is attached to the cell wall and is made of polysaccharide or polypeptide, or a combination of both, to form a viscous layer. If it is thick, structured and adheres strongly to the cell wall, it is called a Capsule. If it is disorganized and loosely attached to the cell wall it is called a Slime layer. Both capsule and slime layer do not take up simple stains but can be observed by negative staining with India Ink or Negrosin.
Functions of Capsules - (a). Adhere to surfaces to form colonies. (b). Antiphagocytic. (c). Antigenic. (d). Protect the organism from dehydration.

Cytoplasmic Membrane
Made of a phospholipid bilayer with integral and peripheral proteins embedded. It maintains the selective permeability of the cell, has respiratory enzymes and during cell division the chromosome is linked to the cell membrane at a site called Mesosome.

Media Preparation

The powder is weighed and dissolved in a specified amount of water. pH is checked and adjusted. Then the medium is sterilized at 121º C at 15 lbs/sq. in. pressure for 15 minutes. Heat labile compounds that are to be added to the medium should be sterilized by filtration, and then added to the medium at 50 º C.


Binomial with genus and species.
1. Genus is suggestive of morphology,configuration or structure or other characteristics.

2. Species is suggestive of metabolic features, biochemical characteristics or disease e.g. Staphylococcus epidermidis.
Sex or conjugation Pili
for the transfer of extrachromosomal DNA between donor and recipient.
4 basic shapes - spherical, rods, spiral and square.

spherical, rods, spiral and square
Increase in the population of cells is called a Culture. Cell division is by an asexual process called Binary Fission, and the time it takes to divide (double) is called Generation time. Growth of culture goes through 4 phases with time, when plotted on a graph.
Under environmental stress (lack of water, nutrients etc.) some vegetative cells produce endospores e.g. Clostridium and Bacillus. Spores can be dormant for many years. They can survive extreme heat, desiccation, radiation and toxic chemicals. However, when conditions become favorable they revert to a vegetative state. Spore germination is activated by heat in the presence of moistures but the endospore must degrade the layers around the spore.
Gram Positive cells
Has a thick peptidoglycan (murein) layer and 2 classes of teichoic acids. Lipoteichoic acid which is on the surface, embedded in the peptidoglycan layer and is linked to the cytoplasmic membrane. Wall teichoic acid is on the surface and is linked to only the peptidoglycan layer. Teichoic acid is responsible for the antigenic determinant of the organism.
Made of the protein pilin and project from the cell surface. There are 2 types:

1.Sex or conjugation Pili

2. Attachment Pili or Fimbriae.

Stationary phase
Dying and dividing organisms are at an equilibrium. Death is due to reduced nutrients, pH changes, toxic waste and reduced oxygen. Cells are smaller and have fewer ribosomes. In some cases cells do not die but they are not multiplying.
Made of the protein flagellin and consists of a filament and basal region. The basal region has a hook and a basal body which has a rod and rings. Gram positive organisms have 2 rings, one in the cell wall and one in the cell membrane. Gram negative organisms have 4 rings, 2 in the cell wall and 2 in the cell membrane.

Spread Plate
Liquid specimen is spread on the surface of solid agar with a sterile bent glass rod.
Axial Filaments
Found in Spirochetes and are similar to flagella, but are located between the cell wall and an outer sheath, and are attached to one end of the organism.

Log or Logarithmic phase
Division is at a constant rate (generation time) but varies with species, temperature and media. Cells are most susceptible to inhibitors.

Cytoplasmic Structures
(a). Nucleoid - An area of concentrated DNA with no nuclear membrane. The DNA is single circular, double stranded without proteins. (b). Ribosomes - 30S and 50S to form a 70S complex.
(c). Plasmids - Extrachromosomal loops of DNA, some code for drug resistance, toxins and other factors.

Microorganisms can be categorized based on their optimal temperature requirements for the shortest generation time.
(a) Psychrophiles (0 - 20 ºC).

(b) Mesophiles (20 - 40 ºC)

(c) Thermophiles (40 - 90 ºC).

Most bacteria are mesophiles especially pathogens that require 37 ºC.

Gram Negative Cells
Has a thin peptidoglycan (murein) layer with an outer membrane attached to the peptidoglycan layer by lipoproteins. The outer membrane is made of protein, phospholipid and lipopolysaccharide. In the lipopolysaccharide, the lipid portion is embedded in the phospholipid and the O antigen polysaccharide is on the surface. The lipid is called Lipid A and it is toxic, but the whole lipopolysaccharide is called an Endotoxin. The cell wall has channels called Porins for the transport of low molecular weight substances. Between the cytoplasmic membrane and the cell wall is a periplasmic space with hydrolytic enzymes, antibiotic inactivating enzymes and transport proteins.
Required for the construction of all organic molecules. Autotrophs use inorganic
carbon (CO2) as their carbon source, while heterotrophs use organic carbon.
Attachment Pili or Fimbriae
There are many and are used for attachment to surfaces. Pili are virulence factors
Enriched Medium
A base growth medium with special supplements added such as Blood, vitamins and amino acids e.g. Blood Agar.
Pour Plate
Organisms are serially diluted, then a small amount is added to an empty sterile petri dish, to which melted agar at 50 ºC is added. Then mix to distribute the organisms.

Selective Medium
It allows one species to grow and suppresses others by incorporating dyes, antibiotics, adjusting pH etc. e.g. Eosin methylene blue (EMB).
Obtained from an inorganic source e.g. Nitrogen gas (N2)
Nitrate (NO3), Nitrite(NO2), and Ammonia (NH3), or from an organic source e.g. Proteins, broken down to amino acids. Many organisms use nitrogen gas by nitrogen fixation to produce ammonia.

Types of Media
Liquid media is called broth. Solid media is called agar. When the exact chemical composition of media is known its called synthetic or defined media.
All purpose Medium
Supports growth of most organisms e.g. Trypticase soy broth and Nutrient Broth.
Used to dissolve materials to be transported across the cytoplasmic membrane.
Reduced Medium
A reducing agent (thioglycolate, cystine or ascorbate) is added to the medium to remove oxygen so anaerobes can grow.

Other nutrients
Required in small amounts are Iron, Sulfur, Phosphorus and Minerals.
Preservation of Isolated Organisms

(a) Freezing - Organisms are frozen rapidly (- 20 ºC to - 30 ºC) with liquid nitrogen.
(b) Freeze dry or Lyophilization - The specimen is frozen in dry ice and acetone, then water is removed by sublimation.
(c) Grow the organisms on a slant then cover the growth with sterile mineral oil and refrigerate.
Required for aerobic respiration and energy production.

Organisms are classified according to their gaseous requirements - Obligate aerobes, Microaerophilics, Facultative anaerobes, Obligate anaerobes and Aerotolerant anaerobes.

Differential Medium
More than one type of organism can grow but separation is based on color and other characteristic differences e.g. MacConkey and EMB agars.
Media can be selective and differential or enriched and differential at the same time.
Streak Plate
An inoculating loop is used to thin out organisms on the surface of the agar.
Death or Decline phase
The population is dying in a geometric fashion so there are more deaths than new cells. Deaths are due to the factors in stationary phase in addition to lytic enzymes that are released when bacteria lyse.

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