STERILIZATION IN DENTISTRY

Dr.Madhu Billa2nd year PG

SIBAR Institute Of Dental Sciences

Sterilization:- Is defined as the process by which an article, surface or medium is freed of all living microorganisms either in the vegetative or in the spore form.

Disinfection:- It is the destruction or removal of all pathogenic organisms to a level which seems to be no longer harmful to health

TERMS & DEFINITIONS

Contamination:-The presence of microorganisms on a body surface or on inanimate articles or substances.

Sanitation:-The process by which the number of microorganisms on inanimate objects is reduced to a safe level. It does not imply freedom from microorganisms, and generally refers to a cleaning process.

Asepsis:- A condition in which living pathogenic microorganisms are absent.

Antiseptic:-A substance that prevents or arrests the growth of action of

microorganisms either by inhibiting their activity or by destroying them.Term

used especially for preparations applied topically to living tissues.

.

Infection Control:-The selection and use of procedures and

products to prevent the spread of infectious diseases.

Bactericidal:- These are agents which are able to kill bacteria.

Bacteriostatic:- These agents prevent the multiplication of

bacteria

HISTORY

The concept of asepsis & sterilization and its role in the prevention of

infection was put forward nearly Two centuries ago.

The general principles for asepsis were laid down by Hungarian gynecologist,

Ignaz semmelweis in europe in early 1850’s and Oliver Holmes in USA.

These principles were accepted after Joseph Lister’s studies on prevention of

wound infection carried out between 1865-91.

Lister, working on antisepsis, initially used phenol (dilute carbolic acid) for contaminated wounds, later applied it in all surgical wounds, also in operating room by nebulization of the solution. Further developments occurred with the introduction of steam sterilization.

Anthony Van Leuwenhock first described microorganisms (1667).

Von Bremann introduced autoclave Infection control program recommended by the CDC Federal OSHA published standards on occupational

exposure to blood-borne pathogens in Dece. 1991 Required in every dental office by July 6, 1992

CLASSIFICATION OF STERILIZATION

Physical agents Chemical agents

Sunlight Alcohols

Drying Dyes

Dry heat Halogens

Moist heat Phenols

Filteration Surface active agents

Radiation Metallic salts

Ultrasonic and sonic vibration gases

SUNLIGHT

It has bactericidal activity and plays an important role in spontaneous sterilization that occurs under natural conditions.

Action is due to its content of ultraviolet rays, most of which, however are screened out by glass and the presence of ozone layer.

Direct sunlight as in tropical countryside where it is not filtered by impurities in the atmosphere, has an active germicidal effect due to the combined effect of ultraviolet and heat rays.

DRYING

Moisture is essential for bacteria

Drying therefore has a deleterious effect on most bacteria

Viruses and spores are unaffected by drying

So drying is a very unreliable method.

HEAT It is the most popular, reliable and widely used method of sterilization

Two methods - Dry heat

Flaming

Incineration

Hot Air Oven

- Moist heat

Temp below 100o C

Temp at 100o C

Steam at normal pressure

Steam under pressure

FLAMING

A simple & effective method

Loops or wires ,glass slides, cover slips the tips of the instruments are held in a Bunsen flame till they become red-hot. These materials may be dipped in a disinfectant before flaming

INCENERATION This is an excellent method for safely destroying

materials such as contaminated cloth, animal carcasses and pathologic materials

Plastics such as PVC and polythene can be dealt with similarly

Polystyrene materials emit clouds of dense black smoke and hence should be autoclaved in appropriate containers.

HOT AIR OVEN

• This is the most widely used method of sterilization by dry heat.

• This type of energy does not penetrate materials easily and thus, long periods of exposure to high temperatures are necessary.

• A holding period of 160oC( 320oF) for 1 hr is used to sterilize glassware, swabs, liquid paraffin, dusting powder, fats and grease. It is suitable for dry powders and water free oily substances.

• Since hot air is a bad conductor of heat its penetrating power is low.

• The oven is usually heated by electricity, with heating elements in the wall of the chamber.

• It must be fitted with a fan to ensure even distribution of air and elimination of air pockets.

• The material should be arranged so as to allow free circulation of air in between the objects.

Glassware should be perfectly dry before being placed in

the oven.

The British pharmacopoeia recommends a holding period

of 1hr at 1500C for oils, glycerol, and dusting powder.

The oven must be allowed to cool slowly for about 2hrs

before the door is opened, since the glassware may crack

due to sudden or uneven cooling.

ADVANTAGES • Effective and safe sterilization of metal instruments and

mirrors• No corrosion of Carbon steel instruments and burs DISADVANTAGES• Long cycles• Poor penetration• Uneven heating • Damage to heat sensitive items

GLASS BEAD AND HOT SALT STERILISER

The glass bead steriliser uses a metal cup with glass beads of 1 mm diameter in it

The hot salt container uses ordinary table salt The temperature range for both varies from 425oF TO 475oF Both are used to sterilise endodontic instruments The hot salt steriliser is better than the glass bead steriliser

because the glass beads stick to the instruments and clog the root canal.

Also the material used in the hot salt steriliser is ordinary table salt which is readily available

Electric glass bead or hot salt steriliser

Moist heat

Moist heat kills microorganisms by denaturing proteins. Denaturation involves changes in the chemical or physical

properties of proteins. It includes structural alterations due to destruction of the chemical bonds holding proteins in a three dimensional form. As proteins revert to a two dimensional form they coagulate & become nonfunctional.

Moist heat penetrates material much more rapidly than dry heat because water molecules conduct heat better than air.

Temperatures below 1000CPASTEURIZATION

•Its purpose is to reduce the bacterial population of a liquid

such as milk and destroy organisms that may cause spoilage

•Spores are not affected by this method

Holder method – It involves heating up to 62.90C for 30 min.

although thermophilic bacteria thrive at this temperature; they are of little consequence because they cannot grow at body temperature.

Flash pasteurization -This method uses a temperature of 71.60 C for 1sec

followed by cooling to 130C Ultra pasteurization -This method uses a temperature of 820 C for 3 sec.

Temperatures at 1000C (boiling)

Vegetative bacteria are killed almost immediately at 90-1000C, but sporing bacteria require prolonged periods of boiling.

Boiling water is not considered as a sterilizing agent because destruction of bacterial spores and inactivation of viruses cannot always be assured. It is considered as a method for disinfection

A minimum exposure period of 30min. is recommended to kill vegetative bacteria.

Sodium bicarbonate 2% conc. is added to increase the efficiency of process.

Advantages

Rapidity, economic , no elaborate equipment, good penetration

and harmless to wide range of dental materials.

Diasadvantages

dulls the cutting edges

can also cause corrosion

Steam at atmospheric pressure Also known as compressed or saturated steam This is an inexpensive method using a Koch or

Arnold steamer. Principle used is steam under pressure is hotter and higher

the pressure the higher the temperature Liquids were sterilized by this method at 1000C for 30min

on each of 3 successive days. Also called fractional sterilization, because a

fraction was accomplished on each day. Also called Tyndallization after its developer John

Tyndall, and intermittent sterilization because it was a stop and start operation.

During the first day’s exposure, steam kills virtually all organisms except bacterial spores and it stimulates spores to germinate vegetative cells.

During overnight incubation the cells multiply and are killed on second day.

Again the material is cooled and a few remaining spores germinate only to be killed on the 3rd day.

This method also fails because certain spores ( ex; some anaerobes ) fail to germinate. A suitable medium for germination such as broth is required.

STEAM UNDER PRESSURE

Moist heat in the form of pressurized steam is regarded as the most dependable method for destruction of all forms of bacterial life including spores.

This method is incorporated into a device called the autoclave.

Over a hundred years ago, French & German microbiologists developed the autoclave.

The basic principle is that when the pressure of a gas increases the temperature increases

As the water molecules in steam become more energized, their penetration also increases

Same principle is used in home pressure cooker.

It is important to note that sterilizing agent is moist heat but not the pressure.

Pressure in autoclave –15lbs/sq. inch

Temperature - 121.50C

Time - 3 to 30 min

Autoclave

This method can be used for a broad variety of items such as instruments, clothing, glassware and intravenous solutions etc.

Limitations: Plastic ware melts in high heat Sharp instruments become dull ( corrosion ) Many chemicals breakdown during the sterilization process, and

oily substances cannot be treated since they do not mix with water. A new form of autoclave called the Prevacuum autoclave has been

developed, which draws air out of the chamber at the beginning of the cycle. The major advantage of this is minimal exposure

Unsaturated Chemical Vapor Sterilization

This system depends on heat, water and chemical combination for it’s efficacy

The temperature pressure required is greater than that for autoclave

It is known as a chemiclave Instead of distilled water a solution of alcohol,

formaldehyde, ketone, acetone and water is used to produce the sterilizing vapor

Temperature 2700 F(1310C )at 20 pounds pressure for 20 to 40

minutes

Advantages

short cycle, no rusting or corrosion, dry instruments at the

end of the cycle, automatic preset cycle timing

Disadvantages

heat sensitivity of instruments, loading dry instruments,

chemical odors and constant monitoring

OIL BATHS

Can be used to sterilise metal instruments

It can be heated for a period of 1750C for 15 minutes

Disadvantages include poor sporicidal activity, fire hazard,

difficult to remove and constant monitoring

FILTRATION

Filters have been used since time immemorial

It is the method of choice for heat labile liquids and

solutions

Early pioneer was Charles Chamberland

Filters are used to purify IV solutions, culture media,

toxoids, beverages etc

Types of Filters

CANDLE FILTERS They are manufactured in different grades of porosity & have been widely used to purify water for industrial and drinking purposes. They are of 2 types

Unglazed ceramic filter

Ex; Chamberland & Doulton filters

Diatomaceous earth filter

Ex; Berkefeld & Mandler filters

ASBESTOS FILTERS They are in the form of disposable, single use discs. They have high absorbing capacity and tend to alkalinize filtered liquids They are carcinogenicEx; Seitz filter

SINTERED GLASS FILTERS They contain heat fusing finely powdered glass particles of graded sizes. They have low absorbing capacity and can be easily cleaned.

MEMBRANE FILTERS

They are made up of cellulose esters (acetate & polycarbonate) or other

polymers. They are routinely used in water purification

This is particularly valuable because bacteria multiply and form colonies on

a plate of culture medium.

Pore size 10 to 0.22μ

NUCLEOPORE FILTER It is a very thin polycarbonate film with etched hole pore space of 0.1 to 0.08μ

NON IONISING RADIATION

This include infrared rays and ultraviolet rays

Infra red rays is used for mass sterilization of syringes

ULTRAVIOLET LIGHT

When microorganisms are subjected to UV light, cellular DNA absorbs energy & adjacent thymine molecules link together.

Linked thymine molecules are unable to position adenine on m RNA molecules during the process protein synthesis thereby replication of chromosome will be impaired.

The damaged organism can no longer produce critical proteins or reproduce.

UV light is used to limit airborne or surface contamination in a hospital room, pharmacy food service operation.

UV light does not penetrate liquids or solids and it may cause damage to the human skin.

IONIZING RADIATION

X-rays and gamma rays have wave length shorter than UV light

As these rays can pass through microbial molecules, they force electrons out of their shells, thereby causing ionisation.

The ions quickly combine with & destroy proteins and nucleic acid such as DNA, causing death. Gram +ve are sensitive than gram –ve bacteria.

Can be used for syringe sterilization

MICROWAVE

Microwaves have a wavelength longer than UV light.

In a microwave oven waves are absorbed by water molecules.

The molecules are set into a high speed motion, and the heat

of friction is transmitted to food, which become hot rapidly.

LASER

LASER – Light Amplification by Stimulated Emission of

Radiation

Resent experiments indicate that laser beams can be used to

sterilize instruments & the air in operating rooms, as well as for

a wound surface.

Various types include CO2 , Argon ,NdYAG etc

Ultrasonic vibrations They are high frequency sound waves beyond the range of

human ear. When propagated in fluids ultrasonic vibrations cause formation

of microscopic bubbles or cavities and the water appears to boil. Some observers call this cold boiling. The cavities rapidly collapse & send out shock waves. The

formation and implosion of the cavities is known as cavitation. Microorganisms in the fluid are quickly disintegrated by the external pressures.

The current trend is to use ultrasonic as a cleaning agent to follow the process by sterilization in an autoclave.

FREEZING

Freezing can both kill and preserve depending on various factors

It’s primary use has been in the preservation of bacterial cultures.

Repeated freezing and thawing are much more destructive to bacteria than prolonged storage at freezing temperature.

If bacteria are rapidly subjected to temperature below –350C, ice crystals that form within the cell produce a lethal effect during defreezing.

CHEMICAL METHODS OF STERILIZATION

Infection control with chemicals

IDEAL REQUIREMENTS of a chemical disinfectant

1. Broad spectrum

2. Fast acting

3. Non toxic

4. Surface compatibility

Should not corrode instruments and other metallic surfaces and cause disintegration of cloth, rubber or other materials

5. Easy to use6. Odourless7. Economical 8. Be stable9. High penetrating power10. Should not cause local irritation and sensitivity11. High solubility Should be soluble in water and have a substantial shelf

life

Infection control in dentistry requires the use of disinfectants in

several forms

1.Surface disinfectants

2.Immersion sterilants

3.Immersion disinfectants

4.Hand antimicrobials

• Surface disinfection is the treatment of envioronmental surfaces such as cabinets, chairs, lights, x-rays and similar surfaces where the items are too large or sensitive to be immersed in disinfecting chemicals. It is accomplished by spraying or wiping the solution on the surface

• Immersion disinfection also called cold sterilisation is the immersion of instruments, plastics, in a liquid disinfectant. Time is usually 5 to 30 minutes

• Immersion sterilization is the use of an agent that

has the capability of killing all living microorganisms and

infective agents in 6 to 10 hours. The items should be

completely immersed in the solution

• Hand antimicrobial treatment it is the specific

art of washing or otherwise treating hands with a chemical

soap or lotion with resulting reduction in the number of

hand microbes

A classification of chemical disinfectants was proposed by Spaulding in 1972

It was originally developed for classifying hospital instruments but now has been modified for dental instruments

Patient care items are classified -- • critical• semicritical • non critical

Three levels of disinfection are -- High Intermediate Low

PATIENT CARE ITEMS

CRITICAL:-Penetrate or touch broken skin or mucous membranes.Needles, Scalpels, surgical instruments, mirrors, dental explorers.SEMICRITICAL:-Touch intact mucous membranes.Amalgam condensers, handpieces ultrasonic cleaners.NON CRITICAL:-That do not touch mucous membranes. Counter tops, light handles, chair surfaces.

LEVELS OF DISINFECTION

1. High level disinfection – these disinfectants must inactivate resistant bacterial spores and all other microbial forms

E.g. ethylene oxide gas, immersion gluteraldehyde solutions2. Intermediate level disinfection– these disinfectants do not

inactivate spores during routine use but they destroy other forms of microbes

E.g. formaldehyde, chlorine compounds, iodophors, alcohols, phenolic compounds

3. Low level disinfectants - provide the narrowest antimicrobial range

E.g. quaternary ammonium compounds, simple phenols and detergents. Such compounds are suitable for cleaning environmental surfaces

ALCOHOL Effective skin antiseptics and valuable disinfectants for medical

instruments. Ethyl and isopropyl alcohol are most frequently used Isoprpyl alcohol is prefered to ethyl alcohol as it is a better fat

solvent ,more bactericidal and less volatile It is active against vegetative bacterial cells, including the tubercle

bacillus, It denatures proteins & lipids, and leads to cell membrane disintegration It is also a strong dehydrating agent. It is used to sterilize the skin prior to cutaneous injections .

It is active against gram +ve, -ve, & acid fast organisms at a concentration of 50-70%

‘Isopropyl alcohol’ or ‘rubbing alcohol’ has high bactericidal activity in concentration as high as 99%.

DISADVANTAGES

Relatively inefficient in the presence of blood and saliva

Lacks sporicidal activity

Causes corrosion of metals

ALDEHYDES

GLUTERALDEHYDE

2% solution is effective. It destroys vegetative cells within 10 min to 30min and spores in 10 hrs

It is recommended for use on surgical instruments where residual blood may be present.

It does not damage delicate objects, and therefore it can be used to sterilize optical equipment such as the optic fiber endoscopes.

DISADVANTAGEIt can damage many metal items.E.g. nickel coated impression trays & carbon steel burs will often discolour and corrode, respectively when immersed in gluteraldehyde solution for prolonged periodsIt can cause hypersensitivity on repeated exposure so instruments immersed in gluteraldehyde should be thoroughly rinsed in sterile water prior to use

FORMALDEHYDE It is gas at high temperatures & solid at room temperature.In aqueous solutions, it is bactericidal, sporicidal and virucidal.When 37 gms. of the solid are suspended in 100ml of water a solution called formalin results. Formalin is utilized for inactivating virus in vaccines & producing toxoids from toxins. 10% Formalin containing 0.5% sodium tetraborate is used to sterilize clean metal instruments.

In the gaseous form, formaldehyde is expelled into a closed chamber where it is used as a sterilant for surgical equipment, hospital gowns and medical instruments.

However penetration is poor and the surface must be exposed to the gas for up to 12hrs. for effective sterilization. ( Fumigation)

Formaldehyde leaves a residue and instruments must be rinsed before use.

Can cause hypersensitivity reactions

HALOGEN

These are a group of highly reactive elements whose

atoms have 7 electrons in the outer shell.

Two halogens chlorine & iodine are commonly used for

disinfection.

CHLORINE Available as -- gas

- organic compound - inorganic compounds Most commonly used as hypochlorites

The disinfectant action results from it’s ability to liberate free chlorine.

In aqueous solutions the liberated Cl ions reacts with water to form hypochlorous acid,

Used in water supplies and swimming baths

Chlorax & purex bleach used for household sanitation &

the disinfection of food utensils.

Chlorinated lime [NaOCl or Ca (OCl)2] is used as

bleaching agent in textile industry.

DAKIN’s solution

0.5% sodium hypochlorite

Extensively for wounds during the I & II World War

It is used to treat athlete’s foot

Chloramines Chloramines-T are organic compounds

Chlorine and amino groups.

Compounds release free chlorine more slowly than hypochlorite solutions

More stable.

Used for general wound antisepsis and root canal therapy

DISADVANTAGES

Corrosive to metals

Irritating to the skin

Destroys fabrics

Solution has to be prepared daily

Iodine Iodine is more reactive and more germicidal.

Iodine acts on the tyrosine portions of protein molecules.

Tincture of iodine2% iodine solution in ethyl alcohol used as an antiseptic for application over skin and mucous membrane

DISADVANTAGES Irritating Allergic Corrodes metal Stains skin and clothes

Iodophors

Compounds of iodine with non ionic surface active agents that release iodine over a long period of time

The detergent portion of the complex loosens the organisms from the surface & halogen kills them.

ADVANTAGESLess allergicLess irritating to tissuesDo not stain clothesProlonged activity

PHENOLS AND THEIR DERIVATIVES (Carbolic Acid)

Obtained by distillation of coal tar between temperatures of 1700C and 2700C.

Surgical antiseptic by Lister in 1865, phenol was widely used as a disinfectant.

Low solubility in water, they are formulated with emulsifying agents, such as soaps, which also increase their antimicrobial action.

They act as cytoplasmic poisons by penetrating and disrupting cell wall, thereby leading to denaturation of intracellular proteins

Phenol is active against gram +ve bacteria

Bactericidal at 1% and fungicidal at 1.3 %

Drawbacks

Its activity is reduced in the presence of organic matter

Expensive

Caustic to the skin

CRESOL

Phenol derivatives containing methyl groups have greater

germicidal activity and lower toxicity than the parent

compound.

Ortho, meta, para cresols are appreciably more active than

phenol and are employed as a mixture named “Tricresol”

They are not readily inactivated in the presence of organic

matter and so are good disinfectants

Bisphenols It is a combination of 2 phenol molecules Eg. .Hexachlorophene, Chlorhexidine Hexachlorophene It is highly effective against gram –ve organisms esp. staphylococci

& streptococci. In 1961 routine daily hexachlorophene bathing of newborn infants

became an accepted procedure in many nurseries in order to reduce colonization of umbilical stump & infection of scalp with streptococcus aureus.

In 1971, FDA placed strict controls on the use of hexachlorophene and curtailed its use in newborn nursery because numerous studies indicate that excessive amounts could be absorbed through the skin and cause neurological damage in newborn.

SURFACE ACTIVE AGENTS Substances that alter the energy relationships at interfaces

producing a reduction of surface tension are referred to as surface-active agents.

They are widely used in industry as wetting agents, detergents. Surface active agents are compounds that possesses both

water attracting (hydrophilic), and water repelling (hydrophobic) groups.

There are 4 types of surface active agents.– Cationic surface active agents– Anionic surface active agents– Nonionic surface active agents– Amphoteric surface active agents

Cationic Surface Active Agents (quaternary ammonium compounds )

They act on the phospholipid present in the cell membrane, then the non polar portion of the detergents penetrate into the hydrophobic interior of the membrane.

This changes the permeability of the cell It then enters the cell and causes denaturation of the protein More active at an alkaline pH. They are bactericidal for a wide range of organisms,

although gram +ve species are more susceptible.

Disadvantages

Inability to penetrate organic debris

Incompatible with anionic detergents, Ca, Mg, and iron of hard

water

E.g. cetyl trimethyl ammonium bromide (cetavlon or ctrimide)

& benzalkonium chloride

Anionic surface active agents

These are active at acid pH

These agents cause gross disruption of the lipoprotein

framework of the cell membrane and also have mechanical

cleansing action.

e.g. Soaps and fatty acids

Nonionic surface-active agents

These agents are relatively nontoxic and few promote

bacterial growth.

E.g. Tween 80 facilitates the diffused growth of

mycobacterium tuberculosis and provides a source of oleic

acid which is stimulatory to the organism.

Amphoteric surface active agents

These compounds are also called as TEGO’ compounds.

They are active against a wide range of gram +ve, -ve

organisms and some viruses.

Ethylene oxide Ethylene oxide is a gas at temperature above 10.80C It has excellent penetration capacity and is sporicidal as well as

virucidal. However, it is both toxic and highly explosive. Its explosiveness is reduced by mixture with Freon gas The gas is released into tightly sealed chamber where it

circulates for up to 4hrs. with carefully controlled humidity. The chamber then must be flushed with inert gas for 8-12hrs.

To ensure that all traces of ethylene oxide are removed otherwise, the chemical will cause “cold burns” on contact with skin.

It is used to sterilize paper, leather, wood, metal and rubber as

well as plastics.

In hospitals it is used to sterilize catheters, artificial heart

valves, heart-lung machine components and optical equipment.

Used in chemiclaves

Beta Propiolactone (BPL)

It is a condensation product of ketone and formaldehyde

Is less explosive than ethylene oxide, but its penetrating power

is more limited.

As a liquid it is used to sterilize vaccines, sera and surgical

ligatures.

However, it is carcinogenic & is therefore used only under

restricted conditions.

OXIDANTS

Included in this group are halogens, hydrogen peroxide,

and potassium permanganate.

The oxidizing agents inactivate enzymes by converting

functional –SH groups to the oxidized form.

Hydrogen Peroxide (H2O2)

Rinse wounds, scrapes and abrasions.

The area foams & effervescence as catalase in the tissue breaks down hydrogen peroxide to oxygen & water.

The furious bubbling removes microorganisms mechanically.

Anaerobic bacteria are sensitive to H2O2 because sudden release of oxygen gas inhibits their growth

CHLORHEXIDINE

This compound was approved in 1976 by FDA as surgical

scrub, hand wash, and superficial skin wound cleanser.

Chlorhexidine in a conc. of 0.2% is also most effective

antiplaque & antigingivitis agent

After each patient use, run any handpiece that is

connected to the dental air/water system, to discharge

water and/or air for at least 30 seconds after each patient

use

Leave the bur in place while you clean the outside of the

handpiece with detergent and warm water.

Sterilise in an autoclave.

If recommended by the manufacturer, lubricate the

handpiece with pressurised oil until clean oil appears from

the chuck.

STERILISATION OF HANDPIECES

Tests for sterilization

Chemical indicators:• External chemical indicator • Internal chemical indicator Biologic indicator Microorganisms usedSteam autocalve: Bacillus stearothermophillus in vials,strips Dry heat oven : Bacillus subtilis stripsChemical vapour: Bacillus stearothermophillus on strips

CONCLUSION “PREVENTION IS BETTER THAN CURE”- a proverb

well suited to sterilisation

A thorough understanding of the application of sterilisation

will help ensure safety from the invisible but deadly world of

microbial pathogens

Hence utilisation of proper sterilization, disinfection and

aseptic procedures helps us achieve the safety of our

professional demands

REFRENCES• Textbook of microbiology- R.Anantnarayan 7th Edition

Orient Longman

• Fundamentals of microbiology- Frobisher 9th Edition

Saunders

• Infection control and office safety- DCNA April 1991

• Clinical Practice of the Dental Hygienist- Esther M

Wilkins 7th Edition