Variant Forms of Bacteria

Prortoplast ; surface is completely devoid of cell wall component,

Spheroplast : Some residual cell wall component is present 

Autoplast: protoplasts which are produced by the action of organisms’ own autolytic enzymes.

L Form: replicate as pleomorphic filtrable elements with defective or no cell wall These are designated as L forms after the Lister Institute where these were discovered by Klineberger-Nobel.

Bacterial Spores: Gram positive bacilli and actinomycetes form highly resistant and dehydrated forms which are called as endospores. The surrounding mother.cell which give rise to them is known as Sporangium. These endospores are capable of survival under adverse conditions
Structure :smooth walled and ovoid or spherical. 

In bacilli, spores usually fit into the normal cell diameter except in Clostridium where these may cause a terminal bulge. (drum stick ) or central. , these look like areas of high refractilitv under light microscope.

Germination : This is the process of converting a spore into the vegetative cell. It occurs in less than 2 hours and has three stages:Activation, Germination, Outgrowth
 

NITRIC OXIDE-DEPENDENT KILLING

Binding of bacteria to macrophages, particularly binding via Toll-like receptors, results in the production of TNF-alpha, which acts in an autocrine manner to induce the expression of the inducible nitric oxide synthetase gene (i-nos ) resulting in the production of nitric oxide (NO) . If the cell is also exposed to interferon gamma (IFN-gamma) additional nitric oxide will be produced (figure 12). Nitric oxide released by the cell is toxic and can kill microorganism in the vicinity of the macrophage.

NORMAL MICROBIAL FLORA 

A. Properties. Normal microbial flora describes the population of microorganisms that usually reside in the body. The microbiological flora can be defined as either 
1) Resident flora - A relatively fixed population that will repopulate if disturbed, 

2) Transient flora - that are derived from the local environment. These microbes usually reside in the body without invasion and can
even prevent infection by more pathogenic organisms, a phenomenon known as bacterial interference. 
The flora have commensal functions such as vitamin K synthesis. However, they may cause invasive disease in immunocompromised hosts or if displaced from their normal area. 

B. Location. Microbial flora differ in composition depending on their anatomical locations and microenvironments. The distribution of normal microbial flora.

DISINFECTION AND STERILIZATION

•    Sterilization is the best destruction or com removal_of all forms of micro organisms.
•    Disinfection is the destruction of many microorganisms but usually the b spores.
•    Antisepsis is the destruction or inhibition of microorganisms in living tissues thereby limiting or preventing the harmful effect of infection.
•    Astatic Agent  would only inhibit the growth of microorganisms (bacteriostatic, fungistatic, sporostatic).
•    Acidal agent would kill the microorganism (bactericidal. virucidal, fungicidal)
•    Sterilants are the chemicals which under controlled conditions can kill sporinQ bacteria.
 

CHEMICAL AGENTS

Chlorine and iodine are most useful disinfectant Iodine as a skin disinfectant and chlorine as a water disinfectant have given consistently magnificent results. Their activity is almost exclusively bactericidal and they are effective against sporulating organisms also. 
Mixtures of various surface acting agents with iodine are known as iodophores and these are used for the sterilization of dairy products.

Apart from chlorine, hypochlorite, inorganic chioramines are all good disinfectants but they act by liberating chlorine. 

Hydrogen peroxide in a 3% solution is a harmless but very weak disinfectant whose primary use is in the cleansing of the wound.
 
Potassium permanganate is another oxidising agent which is used in the treatment of urethntzs. 

Formaldehyde — is one of the least selective agent acting on proteins. It is a gas that is usually employed as its 37% solution, formalin. 

When used in sufficiently high concentration it destroys the bacteria and their spores.

Classification of chemical sterilizing agents

Chemical disinfectant

Interfere with membrane functions

•    Surface acting agents : Quaternary ammonium, Compounds, Soaps and fatty acids

•    Phenols : Phenol, cresol, Hexylresorcinol

•    Organic solvent : Chloroform, Alcohol

Denatures proteins

•    Acids and alkalies : Organic acids, Hydrochloric acid , Sulphuric acid

Destroy functional groups of proteins

•    Heavy metals :  Copper, silver , Mercury

•    Oxidizing agents: Iodine, chlorine, Hydrogen peroxide

•    Dyes : Acridine orange, Acriflavine

•    Alkylating agents : Formaldehyde, Ethylene oxide

Applications and in-use dilution of chemical disinfectants

Alcohols : Skin antiseptic Surface disinfectant, Dilution used 70%

Mercurials : Skin antiseptic Surface disinfectant Dilution Used 0.1 %

Silver nitrate : Antiseptic (eyes and burns)  Dilution Used 1 %

Phenolic compound : Antiseptic skin washes  Dilution Used .5 -5 %

Iodine : Disinfects inanimate object, Skin antiseptic Dilution used  2%

Chlorine compounds  : Water treatment Disinfect inanimate objects , Dillution used 5 %

Quaternary ammonium Compounds : Skin antiseptic , Disinfects inanimate object, Dilution Used < 1 %

Glutaraldehyde: Heat sensitve instruments, Dilution used 1-2 %

Cold sterilization can be achieved by dipping the precleaned instrument in 2% solution of gluteraldehyde for 15-20 minutes. This time is sufficient to kill the vegetative form as well as spores ofthe organisms that are commonly encountered in the dentistry.

Ethylene oxide is an a agent extensively used in gaseous sterilization. It is active against all kinds of bacteria and their spores. but its greatest utility is in sterilizing those objects which are damaged by heat (e.g. heart lung machine). It is also used to sterlise fragile, heat sensitive equipment, powders as well as components of space crafts.

Evaluation of Disinfectants

Two methods which are widely employed are:

 Phenol coefficient test, Kelsey -Sykes test
 
These tests determine the capacity of disinfectant as well as their ability to retain their activity.
 

BACTERIAL GROWTH

The conversion of a parental cell into two daughters constitutes the bacterial life cycle and the time taken to complete cell cycle is known as generation_time. This is around 15 minutes in vegetative bacteria except mycobacteria.

Bacterial Growth Curve

In the presence of fresh growth medium a bacterium shows following four phases;

The Lag phase -> The Log phase -> The Stationary phase  -> The Decline phase

The Lag Phase : short duration , bacteria adapt themselves to new environment 

The Log Phase (Exponential Phase) : Regular growth of bacteria occurs The morphology of bacteria is best developed in this phase and organisms manifest typical biochemical characters. 

- Most of the cidal Abx work best in this phase
•    i.e. Ampicillin
- Best phase for staining bacterial cultures

Chemostat and turbidostat are examples of technique by which this phase can be prolonged.

Stationary Phase : balanced growth and cell division cannot be sustained. The total cell Count remains static till lysis supervenes, but the viable cell count quickly declines.

Decline Phase: death phase. Dyeing bacteria exceed the dividing bacterias.