INNATE (NON-SPECIFIC) IMMUNITY

The elements of the innate (non-specific) immune system include anatomical barriers, secretory molecules and cellular components. 

Among the mechanical anatomical barriers are the skin and internal epithelial layers, the movement of the intestines and the oscillation of broncho-pulmonary cilia. 

Associated with these protective surfaces are chemical and biological agents.

A. Anatomical barriers to infections

1. Mechanical factors

The epithelial surfaces form a physical barrier that is very impermeable to most infectious agents. Thus, the skin acts as our first line of defense against invading organisms. The desquamation of skin epithelium also helps remove bacteria and other infectious agents that have adhered to the epithelial surfaces. 

2. Chemical factors

Fatty acids in sweat inhibit the growth of bacteria. Lysozyme and phospholipase found in tears, saliva and nasal secretions can breakdown the cell wall of bacteria and destabilize bacterial membranes. The low pH of sweat and gastric secretions prevents growth of bacteria. Defensins (low molecular weight proteins) found in the lung and gastrointestinal tract have antimicrobial activity. Surfactants in the lung act as opsonins (substances that promote phagocytosis of particles by phagocytic cells). 

3. Biological factors

The normal flora of the skin and in the gastrointestinal tract can prevent the colonization of pathogenic bacteria by secreting toxic substances or by competing with pathogenic bacteria for nutrients or attachment to cell surfaces.

B. Humoral barriers to infection

Humoral factors play an important role in inflammation, which is characterized by edema and the recruitment of phagocytic cells. These humoral factors are found in serum or they are formed at the site of infection.

1. Complement system – The complement system is the major humoral non-specific defense mechanism (see complement chapter). Once activated complement can lead to increased vascular permeability, recruitment of phagocytic cells, and lysis and opsonization of bacteria. 

2. Coagulation system – Depending on the severity of the tissue injury, the coagulation system may or may not be activated. Some products of the coagulation system can contribute to the non-specific defenses because of their ability to increase vascular permeability and act as chemotactic agents for phagocytic cells. In addition, some of the products of the coagulation system are directly antimicrobial. For example, beta-lysin, a protein produced by platelets during coagulation can lyse many Gram positive bacteria by acting as a cationic detergent.

3. Lactoferrin and transferrin – By binding iron, an essential nutrient for bacteria, these proteins limit bacterial growth.

4. Interferons – Interferons are proteins that can limit virus replication in cells.

5. Lysozyme – Lysozyme breaks down the cell wall of bacteria. 

6. Interleukin -1 – Il-1 induces fever and the production of acute phase proteins, some of which are antimicrobial because they can opsonize bacteria.

C. Cellular barriers to infection

Part of the inflammatory response is the recruitment of polymorphonuclear eosinophiles and macrophages to sites of infection. These cells are the main line of defense in the non-specific immune system.

1. Neutrophils – Polymorphonuclear cells  are recruited to the site of infection where they phagocytose invading organisms and kill them intracellularly. In addition, PMNs contribute to collateral tissue damage that occurs during inflammation.

2. Macrophages – Tissue macrophages  and newly recruited monocytes , which differentiate into macrophages, also function in phagocytosis and intracellular killing of microorganisms. In addition, macrophages are capable of extracellular killing of infected or altered self target cells. Furthermore, macrophages contribute to tissue repair and act as antigen-presenting cells, which are required for the induction of specific immune responses.

3. Natural killer (NK) and lymphokine activated killer (LAK) cells – NK and LAK cells can nonspecifically kill virus infected and tumor cells. These cells are not part of the inflammatory response but they are important in nonspecific immunity to viral infections and tumor surveillance. 

4. Eosinophils – Eosinophils  have proteins in granules that are effective in killing certain parasites.

Precipitation Reaction

This reaction takes place only when antigen is in soluble form. Such an antigen when
comes in contact with specific antibody in a suitable medium results into formation of an insoluble complex which precipitates. This precipitate usually settles down at the bottom of the tube. If it fails to sediment and remains suspended as floccules the reaction is known as flocculation. Precipitation also requires optimal concentration of NaCl, suitable temperature and appropriate pH.

Zone Phenomenon

Precipitation occurs most rapidly and abundantly when antigen and antibody are in optimal proportions or equivalent ratio. This is also known as zone of equivalence. When antibody is in great excess, lot of antibody remains uncombined. This is called zone of antibody excess or prozone. Similarly a zone of antigen excess occurs in which all antibody has combined with antigen and additional uncombined antigen is present.

Applications of Precipitation Reactions

Both qualitative determination as well as quantitative estimation of antigen and antibody can be performed with precipitation tests. Detection of antigens has been found to be more sensitive.

Agglutination

In agglutination reaction the antigen is a part of the surface of some particulate material such as erythrocyte, bacterium or an inorganic particle e.g. polystyrene latex which has been coated with antigen. Antibody added to a suspension of such particles combines with the surface antigen and links them together to form clearly visible aggregate which is called as agglutination.

Application of precipitation reactions

Precipitation reaction            Example

Ring test                             Typing of streptococci, Typing of pneumococci 
Slide test (flocculation)       VDRL test
Tube test (flocculation)       Kahn test
Immunodiffusion                 Eleks test
Immunoelectrophoresis      Detection Of HBsAg, Cryptococcal antigen in CSF
 

Immunofluorescence

This is precipitation or complement fixation tests. The technique can detect proteins at concentrations of around 1 µg protein per ml body fluid. Major disadvantage with this technique is frequent occurrence of nonspecific fluorescence in the tissues and other material.
The fluorescent dyes commonly used are fluorescein isothocyanate (FITC). These dyes exhibit fluorescence by absorbing UV light between 290 and 495 nm and emitting longer wavelength coloured light of 525 nm which gives shining appearance (fluorescence) to protein labelled with dye. Blue green (apple green) fluorescence is seen with FITC and orange red with rhodamine.

Enzyme Immunoassays

These are commonly called as enzyme linked immunosorbent assays or EL1SA. It is a simple and versatile technique which is as sensitive as radioimmunoassays. It is now the
technique for the detection of antigens, antibodies, hormones, toxins and viruses.

Identification of organisms by immunofluorescence

Type of agent         Examples

Bacterial            Neisseria gonorrhoeae, H. influenzae ,Strept pyogenes, Treponema pallidum
Viral                  Herpesvirus, Rabiesvirus, Epstein-Barr virus
Mycotic             Candida albicans

Enzymatic activity results in a colour change which can be assessed visibly or quantified in a simple spectrophotometer.

CROSS INFECTION AND STERLIZATION IN DENTISTRY

Cross infection is defined as the transmission of infectious agents amongst patients and staff with in hospital environment.

Routes of Infection 
Two routes are important : transdermal  and respiratory. 

 In transdermal route microorganisms enter the tissues of the recipient by means of injection through intact skin or mucosa (usually due to an accident involving a sharp instrument) or via defects in the skin e.g. recent cuts and abrasions.
 
Microorganisms causing cross infection in dentistry

Transmitted through skin 

Bacteria : Treponema pallidum, Staphylococcus aureus

Viruses :Hepatitis virus, HIV ,Herpes simplex virus, Mumps, Measles , Epstein-Barr virus

Fungi: Dermatomycoses, Candidiasis, 

Transmitted through aerosols

Bordetella pertussis, Myco.tuberculosis, Streptococcus pyogenes, Influenza virus
Rhinovirus,  Rubella 
 

Types of microscopy used in bacteriology

Light microscopy
Phase contrast microscopy
Fluorescence microscopy
Darkfield microscopy
Transmission electron microscopy
Scanning electron microscopy

Fluorescent microscopy in which ultraviolet rays are used to examine cells after treatment with fluorescent days.

Phase contrast microscope enhances the refractive index differences of the cell components. This microscopy can be used to reveal details of the internal structures as well as capsules, endospores and motility

Electron microscope The resolving power is more than 200 times that of light microscope.
 

Measurement of Bacterial of Growth

A convenient method is to determine turbidity by photoelectric colorimeter or spectrophotometer. 
The cell number can be counted as total cell number as well as viable count. Viable Count Viable number of bacteria can be counted by inoculating the suspension onto solid growth medium and counting the number of colonies. Since each colony is the end product of one viable bacterium, their count gives the number of viable bacteria in the suspension.
Total number of bacteria can be ascertained in specially designed chambers such as Coulter counter.