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Microbiology  Microbiology: The study of microorganisms  Micro means “microscopically small,” and bio means “living organisms”  Dental assistant needs a foundation in microbiology to understand the nature of pathogens and how to prevent the transmission of disease in the dental office  The two major oral diseases are bacterial infections:  Dental caries (decay) and periodontitis 2  Bacteria  Rickettsiae  Algae  Protozoa  Fungi  Prions  Viruses 3  A large group of one-celled microorganisms that vary in shape, size, and arrangement  Pathogenic bacteria usually grow best at 98.6˚ F (37˚ C) in a moist, dark environment  Skin, respiratory tract, and gastrointestinal tract are inhabited by a great variety of harmless bacteria, called the normal flora  Infection occurs when bacteria occurring naturally in one part of the body invade another part of the body and become harmful 4  Bacteria is not visible with the naked eye.  it must be viewed through a microscope  For optimal viewing bacteria should be stained with special dyes for better visualization  Both gram-negative and gram-positive  Gram-positive  appear blue or purple  Gram-negative  Appear pink or red  Both gram-negative and gram-positive bacteria are present in the oral cavity and are involved in the development of caries and periodontal disease.  Oral bacteria can cause infections in soft tissues and the bloodstream 5  Spherical (coccus, plural cocci)  Reproduces by dividing in two  Cocci that form chains as they divide are called streptococci  Cocci that form irregular groups or clusters are called staphylococci  Rod-shaped (bacillus, plural bacilli)  Spiral (spirillum, plural spirilla) 6 From Stepp CA, Woods M: Laboratory procedures for medical office personnel, Philadelphia, 1998, Saunders. 7  The common diameter of a coccus is about one micrometer or micron (µm). A micrometer is one-millionth of a meter or one-thousandth of a millimeter.  The common size of a bacilli are about one mm wide and five to ten mm long.  Spirilla are .2 to 1.0 µm wide and up to 30 µm long. From Stepp CA, Woods M: Laboratory procedures for medical office personnel, Philadelphia, 1998, Saunders. 8  Hans Christian Gram (1853–1938) developed a four-step staining process for separating bacteria into two groups  Gram staining requires the sequential use of a crystal violet dye, iodine solution, alcohol solution, and a safranin dye  The bacteria that are stained by the dye are classified as Gram positive (they appear dark purple under the microscope)  The bacteria that are not consistently stained are classified as Gram variable (e.g., Mycobacterium tuberculosis) 9 Gram-positive bacteria are stained during the Gram stain process, making them appear darker in color or purple under microscopic examination. G: Textbook of diagnostic microbiology, ed 4, Philadelphia, 2011, Saunders. 10 Gram-negative bacteria are almost colorless or a light pink in color under microscopic examination. From VanMeter KC, VanMeter WG, Hubert RJ: Microbiology for the healthcare professional, St Louis, 2010, Mosby. 11  Aerobes: A variety of bacteria that require oxygen to grow  Anaerobes: Bacteria that grow in the absence of oxygen and are destroyed by oxygen  Facultative anaerobes: Organisms that can grow in the presence or the absence of oxygen 12  Some types of bacteria form a capsule or protective layer that covers the cell wall (e.g, Streptococcus mutans)  Generally virulent (capable of causing serious disease)  Capsule increases the bacteria’s ability to resist the defense mechanisms of the body  Capsule may also prevent antibiotic agents from having an effect on the bacteria 13  Some bacteria change into a highly resistant form called spores  Bacteria remain alive in the spore form but are inactive  Spores represent the most resistant form of life known  Can survive extremes of heat and dryness and even the presence of disinfectants and radiation  Harmless spores are used to test the effectiveness of the techniques for sterilizing dental instruments 14  Tuberculosis  Mycobacterium tuberculosis  Legionnaires’ disease  Legionella pneumophila  Tetanus  Tetanospasmins  Syphilis  Treponema pallidum  MRSA  Methicillin-resistant staphylococcus aureus 15  Antibiotics  Chemicals that interfere with some metabolic activity of the bacteria without affecting our body cells  Bacteriostatic  Agents that prevent bacteria growth without killing them  Bactericidal  Agents that kill bacteria  Disinfectants  Strong chemicals used on inanimate objects to kill or reduce bacteria levels  Antiseptics  Chemicals used on tissues to kill bacteria 16  Fungi are plants, such as mushrooms, yeasts, and molds that lack chlorophyll  Candida is a common yeast found in the oral cavity of about half of the population  It also is found in the gastrointestinal tract, female genital tract, and sometimes the skin  Cross-infection may occur between mother and baby or among infant siblings 17  Caused by the yeast Candida albicans  All forms of candidiasis are considered opportunistic infections; they are found in patients who are very young, very old, or very ill  Characterized by white membranes on the surface of the oral mucosa, tongue, and elsewhere in the oral cavity  Lesions may look like thin cottage cheese and can be wiped off to reveal a raw, red, and sometimes bleeding base 18  Candida infections of the mouth and throat must be treated with prescription antifungal medication.  The type and duration of treatment depends on the severity of the infection and patient-specific factors such as age and immune status. Untreated infections can lead to a more serious form of invasive candidiasis.  Oral candidiasis usually responds to topical treatments such as clotrimazole troches and nystatin suspension (nystatin “swish and swallow”). Systemic antifungal medication such as fluconazole or itraconazole may be necessary for oropharyngeal infections that do not respond to these treatments. 19 From Regezi JA, Sciubba JJ, Pogrel MA: Atlas of oral and maxillofacial pathology, St Louis, 2000, Saunders. 20  Oral thrush occurs most frequently among babies less than one month old, the elderly, and groups of people with weakened immune systems. Other factors associated with oral and esophageal candidiasis include:  HIV/AIDS  Cancer treatments  Organ transplantation  Diabetes  Corticosteroid use  Dentures  Broad-spectrum antibiotic use 21  Denture-related stomatitis (also termed denture sore mouth, denture stomatitis, chronic atrophic candidiasis, Candida-associated denture induced stomatitis, and denture-associated erythematous stomatitis) is a common condition where mild inflammation and redness of the oral mucous membrane occurs beneath a denture. 22  Good oral hygiene practices may help to prevent oral thrush in people with weakened immune systems.  Some studies have shown that chlorhexidine (CHX) mouthwash can help to prevent oral candidiasis in people undergoing cancer treatment.  People who use inhaled corticosteroids (used to help treat asthma) may be able to reduce the risk of developing thrush by washing out the mouth with water or mouthwash after using an inhaler. 23  Much smaller than bacteria  Despite their tiny size, many viruses cause fatal diseases  New and increasingly destructive viruses are being discovered and have caused the creation of a special area within microbiology called virology  Viruses can live and multiply only inside an appropriate host cell  The host cells may be human, animal, plant, or bacterial  A virus invades a host cell, replicates (produces copies of itself), and then destroys the host cell so that the viruses are released into the body 24  Viruses can exhibit specificity (preference) for particular cell types in which to replicate.  For example, HIV infects cells known as CD4+ cells, whereas the hepatitis virus infects only liver cells.  Some other viruses are able to cause disease in more than one organ.  Unfortunately, some viruses can cross the placenta and infect the fetus. 25  Some viruses establish a latent (dormant) state in host cells  A latent virus can be reactivated in the future and produce more infective viral particles, followed by signs and symptoms of the disease  Stress, another viral infection, and exposure to ultraviolet light can reactivate the virus  HIV has a latency period of months to years  Hepatitis C is known to have a latency period of 15 to 25 years 26  Viruses cause many clinically significant diseases in human beings  General antibiotics are ineffective in preventing or curtailing viral infections, and even the few drugs that are effective against some specific viruses have limitations  Viruses are also capable of mutation  It is very difficult to develop vaccines against viruses because of the ability of viruses to change their genetic code 27  Viral diseases are transmitted by the following means:  Direct contact  Insects  Blood transfusions  Contaminated food or water  Inhalation of droplets expelled by coughing or sneezing 28  Viruses are easily destroyed in the external environment  Chemicals such as chlorine (bleach), iodine, phenol, and formaldehyde easily and effectively destroy viruses on surfaces and objects  These agents, however, are too toxic to be used internally 29 Viral hepatitis  There are at least six types of viral hepatitis, each of which is caused by a different virus:  Hepatitis  Hepatitis  Hepatitis  Hepatitis  Hepatitis  Hepatitis A virus (HAV) B virus (HBV) C virus (HCV) D virus (HDV) E virus (HEV) G virus (HGV) 30  Can affect anyone  Spread from person to person when something is put in the mouth that has been contaminated with the stool of a person with hepatitis A (fecal-oral transmission)  Good personal hygiene and proper sanitation can also help prevent hepatitis A  Always wash your hands after changing a diaper or using the bathroom  The least serious form of viral hepatitis  A vaccine is available that provides long-term prevention in persons older than 2 years 31  May infect different blood cells or other tissues of the body  Pathogens may exist or be released into the blood or other body fluids  Diseases of concern in dentistry are:  HBV, HCV and HIV  Diseases can be spread from one person to another by contact with the fluids  Body fluids (OPIM’s) include:  Semen  Vaginal secretion  Intestinal secretions  Tears  Mother’s milk  Synovial fluid (joints)  Pericardial fluid  Amniotic fluid  Saliva 32  A very serious disease that may result in prolonged illness, liver cancer, cirrhosis of the liver, liver failure, and even death  Bloodborne disease that may also be transmitted by other body fluids, including saliva  Anyone who has ever had the disease, and some persons who have been exposed but have not been actually ill, may be carriers of HBV and may actually be spreading the infection to others  This presents a high risk for dental personnel because dental treatment brings them into contact with saliva and blood 33  It is a major health problem in the United States and is endemic in other parts of the world  Between 200,000 to 300,000 people are infected with the hepatitis B virus each year  Approximately 10,000 people require hospitalization  About 250 people will die of fulminant hepatitis  Approximately 15,000 will become chronic carriers  About 4,000 people die of hepatitis related cirrhosis of the liver and 800 from related liver cancer  CDC suggests that there are approximately 1 to 1.25 million HBV carriers in the United States that have potential to spread the disease 34 A milliliter of blood from an infected person can contain as many as 100 million virus particles A very small amount fluid can easily transmit the disease Three important parts: Surface antigen: HBsAg 2 inside antigens: HBcAg & HBeAg 35  About 90% of those infected recover completely  Approximately 5-10% become carriers; ½ of them eliminate the disease from their bodies within 2 ½ years  The other ½ become chronic carriers  The carrier state is called: HBsAg-positive on at least two occasions when tested at least 2 months apart or being HBsAg positive and IgM anti-HBc negative at a single test  HBsAg positive have a great chance to spread the disease  HBeAg positive have very high concentration of the virus in their blood and are considered very infectious  Persons with chronic hepatitis have greater chance of later developing liver cancer 36  Percutaneously  Permucosally  Spreading through blood transfusion or blood products is rare because of testing  High risk activities that increase chance of exposure:  Sharing contaminated needles during intravenous drug abuse  Homosexual, bisexual or heterosexual behaviors with multiple partners  Injuries with sharp objects contaminated with blood or other body fluids  Exposure of non-intact skin or mucous membranes to blood or other body fluids 37  If they develop they begin 2 ½ - 6 months after exposure  About 1/3 show regular symptoms  Regular symptoms:  Yellowing of skin & whites of eyes (jaundice)  Light colored stools, Dark urine, Joint pain  Fever, Rash, Itching  Additional 1/3 show mild symptoms:  Malaise  Loss of appetite  Nausea  Abdominal pain  Additional 1/3 show no symptoms  Because many bloodborne diseases show little or no symptoms the person can still spread the disease  Standard and Universal Precautions must always be followed 38 39 40  Highly effective vaccine is available to prevent hepatitis B  All dental personnel with a chance of occupational exposure should be vaccinated against hepatitis B  The OSHA Bloodborne Pathogen Standard requires that the employer offer the hepatitis B vaccination, at no cost to the employee, within 10 days of initial assignment to a position in which there is chance of occupational exposure to blood or other body fluids  The employee has the right to refuse the offer of vaccination; however, that employee must sign a release form indicating that the employer did offer the vaccine and that the employee understands the potential risks of contracting hepatitis B 41  Adult vaccine regimen consists of:  1 mL doses given at 0, 1, and 6 months  Injection given into deltoid muscle  Single antigen vaccines:  Recombivax HB (Merck & Co., Inc.)  Engerix-B (GlaxoSmithKline Biologicals) 42  Combination vaccines:  Twinrix: adults (HBsAg and inactive hepatitis A)  Comvax: infants and young children (hepatitis B and Haemophilus influenzae type b (HIB) )  Pediarix: infants and young children(hepatitis B, diphtheria. pertussis and tetanus  Postscreening for antibody level is recommended 1-2 months after 3 dose vaccination  Anti-HBsAg  Ifnd a person does not seroconvert they should have a 2 series of three injections and then be retested  If still no seroconversion then test for HBsAg the presence of which would indicate a hepatitis B carrier state or current infection  One indication of chronic HBV is inability to produce defensive antibody  Anti-HBsAg 43  Most efficiently transmitted through blood transfusion or percutaneous exposure to blood  Can occur as a result of an accidental needlestick injury to an employee in a dental office, through the sharing of contaminated needles among users of injection drugs, or through use of contaminated tattoo needles  The carrier rate for HCV is higher than that of HBV  No vaccine against hepatitis C at this time, nor is there a cure for the disease  Treatments available to control effects of the disease 44  HDV is a defective virus that cannot replicate itself without the presence of HBV.  Therefore, infection with HDV may occur simultaneously as a coinfection with HBV or may occur in an HBV carrier.  Persons with a coinfection of HBV and HDV often have more severe acute disease and a higher risk of death than do those infected with HBV alone.  Vaccination against HBV also prevents infection with HDV. 45  Not transmitted through bloodborne contact  Most frequently transmitted by way of the fecal- oral routes through contaminated food or water  HEV is most frequently seen in the form of an epidemic in developing countries  Transmission is not a major concern in a standard dental setting 46  HIV is a bloodborne viral disease  An infection in which the body’s immune system breaks down  AIDS is caused by HIV  When HIV enters the body, it infects special T cells and slowly kills them  As more and more of the T cells die, the body’s ability to fight the infection weakens  A person with HIV infection may remain healthy for many years  AIDS is considered to have developed in an HIV-positive person when he or she becomes sick with serious illnesses and infections that can occur with HIV 47  HIV is spread by sexual contact with an infected person and through needle sharing among drug users  Now that blood is screened for HIV antibodies, the blood supply in this country is safe  Babies born to HIV-infected mothers may become infected before or during birth or, after birth, while breastfeeding  In (nondental) healthcare settings, workers have been infected with HIV after being stuck with needles containing HIV-infected blood 48  Viruses are easily destroyed in the external environment.  Chemicals such as chlorine (bleach), iodine, phenol, and formaldehyde easily and effectively destroy viruses on surfaces and objects.  These agents, however, are too toxic to be used internally. 49 50  A viral infection that causes recurrent sores on the lips  Because these sores frequently develop when the patient has a cold or fever of other origin, the lesions have become commonly known as fever blisters or cold sores 51  This disease, which is highly contagious, makes its first appearance in very young children (1–3 years of age) and is known as primary herpes  Child may have a slight fever, pain in the mouth, increased salivation, bad breath, and a general feeling of illness  Inside of the mouth becomes swollen, and the gingivae are inflamed  Healing begins naturally within 3 days, and the illness is usually over in 7 to 14 days  Supportive measures can be taken to make the child more comfortable, relieve the pain, and prevent secondary infection 52  After the initial childhood infection, the HSV lies dormant and reappears later in life as the familiar recurring fever blister or cold sore  Recurrences tend to take place when the patient’s general resistance is lowered as a result of stress, fever, illness, injury, or exposure to the sun  The use of sunscreen with a sun protective factor of 15 helps to prevent sun-induced recurrences of herpes  Attacks may recur as infrequently as once a year or as often as weekly or even daily 53 From Ibsen OAC, Phelan JA: Oral pathology for the dental hygienist, ed 6, Philadelphia, 2014, Saunders. 54  Genital herpes is one of the most common sexually transmitted diseases in the United States  Initial symptoms: (generally appear 2 to 10 days after infection) tingling, itching, and a burning sensation during urination  Once a person is infected with the virus, outbreaks will recur  The disease can be transmitted only during these recurrences  A mother with active vaginal or cervical herpetic lesions at the time of delivery can pass the virus to her newborn 55  Herpes zoster virus (HHV3)  Causes both varicella (chickenpox) and herpes zoster (shingles)  Chickenpox is the primary infection, and zoster represents reactivation of the illness  Epstein-Barr virus (HHV4)  Responsible for infectious mononucleosis, nasopharyngeal carcinoma, lymphoma, and oral hairy leukoplakia  Cytomegalovirus (HHV5)  Can affect the fetus during pregnancy  Roseola (exanthema subitum) (HHV6)  Roseola occurs as a high fever and a skin rash in infants.  This virus also may cause infectious mononucleosis symptoms.  Human herpesvirus type 7  is isolated from saliva in as many as 70% to 80% of adults and children but has not yet been associated clearly with any particular disease state.  Human herpesvirus type 8  has been associated recently with Kaposi sarcoma  a condition seen in many AIDS patients. 56  Major transmission route for herpesvirus is through direct contact with lesions  Even when there are no active lesions, there is still the possibility of transmission of the virus through saliva or the aerosol spray from the dental handpiece  Because there is no preventive vaccine against herpes, it is essential that precautions be taken to prevent exposure  Protective eyewear is particularly important because a herpes infection in the eye may cause blindness  Gloves protect against infection through lesions or abrasions on the hands 57  Herpetic whitlow—also called digital herpes simplex  finger herpes, or hand herpes—is a painful viral infection occurring     on the fingers or around the fingernails. Herpetic whitlow is caused by infection with the herpes simplex virus (HHSV). DHCW must always wear proper PPE especially on hands to protect against exposure to HHSV 1. Don’t treat patients when they have active cold sores; reschedule them for at least 2 weeks Keep hands in good shape; no nail biting or skin biting 58  Enteroviruses  It is a type of enterovirus; it is a virus that enters the body through the gastrointestinal tract and thrives there, often moving on to attack the nervous system. The polioviruses are enteroviruses.  Enteroviruses can be found in the respiratory secretions (for example,     saliva, sputum, or nasal mucus) and stool of an infected person. Other people may become infected by direct contact with secretions from an infected person or by contact with contaminated surfaces or objects, such as a drinking glass or telephone. Infections caused by enteroviruses are most likely to occur during the summer and fall. Most people who are infected with an enterovirus have no disease at all. Infected people who become ill usually develop either mild upper respiratory symptoms (a "cold"), a flu-like illness with fever, and muscle aches, or an illness with rash. In early fall 2014, an outbreak of infection with a non-polio enterovirus known as enterovirus D68, or EV-D68, sickened many children across multiple U.S. states, many of whom required care in a hospital intensivecare unit. Enterovirus infections have been suspected to play some role in the development of type 1 diabetes, although they do not directly cause the condition. Newborns who become infected with an enterovirus may rarely develop an overwhelming infection of many organs, including the liver and heart, and die from the infection. 59  Herpangina: caused by the coxsackie virus  Breakout of ulcers in the posterior part of mouth and last about 1 week  Herpangina is associated with fever, sore throat, and blisters in the back of the mouth.  It is caused by a number of viruses, all part of the enterovirus family, coxsackie virus being the most common.  Most children develop a high fever and complain of a sore throat. 60  West Nile virus (WNV) is commonly found in Africa, West Asia, and the Middle East  Has likely been in the United States since the early summer of 1999  Virus is carried by mosquitoes and can infect human beings, birds, horses, and some other mammals  Affects a person’s nervous system, causing inflammation of the brain and spinal cord  Symptoms include fever, headache, tiredness, aches, and sometimes rash  Occurs primarily in the late summer or early fall 61  Not related to previous or current human seasonal influenza viruses  The new H1N1 virus spreads very easily from person to person  The most effective way to protect yourself is to receive the vaccine and practice meticulous hand hygiene  For additional resources, contact: www.osap.org 62  Short, nonmovable rods that normally live in the intestinal tract of insects such as lice, fleas, ticks, and mosquitoes  Very small and require host cells to reproduce  Diseases caused by rickettsiae include typhus and Rocky Mountain spotted fever  Transmitted to humans by way of the bite of an infected insect 63  Algae range from microscopic single-cell organisms to larger multiple-cell organisms such as seaweed and kelp  All algae contain chlorophyll, as well as pigments that cause them to appear yellow-green, brown, or red  Algae are found in abundance in both freshwater and marine habitats  Most algae do not produce human disease 64  A large group of single-cell organisms  Some can remain viable as cysts for long periods outside their hosts  Most do not cause disease, but some live in hosts and do cause damage  A small number of protozoa are responsible for intestinal infections in human beings; others invade the blood, lungs, liver, or brain 65  Tuberculosis  Legionnaires' disease  Tetanus  Syphilis  Methicillin-resistant Staphylococcus aureus (MRSA) 66  Caused by the bacterium Mycobacterium tuberculosis  Leading cause of death resulting from infectious disease worldwide  HIV and tuberculosis are often present together  Of the two diseases, tuberculosis is a greater health risk for healthcare workers  “Tuberculosis kill time” is the benchmark for the effectiveness of a surface disinfectant 67  Legionella pneumophila causes two acute bacterial diseases:  Pontiac fever  Legionnaires' disease  Bacteria are transmitted through aerosolization and aspiration of contaminated water  No person-to-person transmission  Dental personnel have higher titers of antibodies against L. pneumophila than do members of the general public 68 Courtesy Dr. Shannon Mills, New Hampshire. 69  Also known as lockjaw  An extremely dangerous and often fatal disease caused by a spore-forming bacillus found in soil, dust, or animal or human feces  This microbe is usually introduced into the body through a wound or break in the skin (e.g., a puncture wound caused by a soiled instrument)  The disease can be prevented by the administration of a vaccine; however, immunity must be kept current through booster doses 70  A sexually transmitted disease (STD) caused by Treponema pallidum pirochetes  Bacteria are quite fragile outside the body, but can cross- infect the dental operator through contact with oral lesions  First stage of syphilis is a painless ulcerating sore, known as a chancre, which is infectious on contact  The second stage is also infectious, and immediate infection may occur as a result of contact with an open sore  The third stage, known as latent syphilis, is usually fatal, and it may occur after the disease has been dormant for 20 years 71 From Ibsen OAC, Phelan JA: Oral pathology for the dental hygienist, ed 6, Philadelphia, 2014, Saunders. A, Courtesy Dr. Norman Trieger; B, Courtesy Dr. Edward V. Zegarelli. 72  A bacteria that is resistant to some antibiotics  This type of Staphylococcus aureus has evolved due to the excessive use of penicillin antibiotics over the years  MRSA infections can be on the surface of the skin or can go into the soft tissue and form a boil or an abscess 73  Always maintain good hand hygiene practices  Never squeeze or try to drain any sore  Keep any wound covered until it has healed  Do not share personal items such as towels, razors, sheets, clothes 74  Small proteinaceous infectious particles  They are composed entirely of proteins that lack nucleic acids (DNA or RNA)  Until the discovery of prions, it was believed that any agent capable of transmitting disease had to be made up of genetic material composed of nucleic acids  Prions convert normal protein molecules into dangerous ones simply by causing the normal ones to change their shape  Prions are a new and separate class, unlike bacteria, fungi, viruses, and all other known pathogens 75  "Mad cow” disease  Linked to human diseases such as Creutzfeldt-Jacob disease (a rare form of dementia) and possibly Alzheimer’s disease  Prion-caused diseases have been found in human beings and animals  Highly resistant to heat, chemical agents, and irradiation  No treatment or vaccine against prion diseases, and the only preventive measure is not eating suspect food 76  Ongoing research to determine whether prions that consist of other proteins play a part in more common neurodegenerative conditions such as Alzheimer’s disease, Parkinson’s disease, and amyotrophic lateral sclerosis  These three disorders have marked similarities  As with all prion diseases, these neuropathologic diseases occur sporadically but sometimes run in families 77  A global disease outbreak  A pandemic occurs when a new virus or disease emerges for which people have little or no immunity, and for which no vaccine is available  In addition to illness and death, an especially severe influenza pandemic may cause severe disruption resulting from school and business closings and interruption of public transportation and food services 78  The amount of healthcare exposure has dropped drastically since improvement in infection control protocols  The greatest dental occupational exposures are:  Injuries from contaminated sharps  Needles, instrument punctures, cuts, bur lacerations  Blood and saliva contamination of cuts and cracks on skin on ungloved hands or hands with torn gloves  Spraying of blood or saliva onto open lesions on the skin or onto mucosa membranes 79
 
									 
									 
									 
									 
									 
									 
									 
									 
									 
									 
									 
									 
									 
									 
									 
									 
									 
									 
									 
									 
									 
									 
									 
									 
									 
									 
									 
									 
									 
									 
									 
									 
									 
									 
									 
									 
									 
									 
									 
									 
									 
									 
									 
									 
									 
									 
									 
									 
									 
									 
									 
									 
									 
									 
									 
									 
									 
									 
									 
									 
									 
									 
									 
									 
									 
									 
									 
									 
									 
									 
									 
									 
									 
									 
									 
									 
									 
									 
									 
                                             
                                             
                                             
                                             
                                             
                                             
                                             
                                             
                                             
                                             
                                            