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Micro

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CONTENTS Title Introduction to Microorganisms....... Bacteria: Structure and Organization. Bacterial Growth and Physiology.... Bacterial Viruses Bacterial Genetics and Variation @vonnens Overview of the Immune System... Innate Immunity Antigens Acquired Immunity. Cell Mediated immunity. Cytokines. The Humoral Immune Response. Complement, : Mechanisms of Acquired Immunity Immunity to Microbes. Tumour Immunology. Tissue Injury in Immunological Reactions Immunodeficiency Diseases Bacteriology Staphylococcus Streptococcus Enterococcus Neisseria Corynebacterium Anaerobic bacteria: Non-Spore Forming Anaerobes Clostridium d Infections in Dentistry ensure ie ea 102 110 114 15 v7 120 126 142 147 CHAPTER 1 INTRODUCTION TO MICROORGANISMS Many common diseases in the oral cavity are caused by microorganisms. To understand these diseases (their etiology, progression, diagnosis and treatment), it is important to have some understanding of microbiology. Microorganisms are minute living organisms that are usually too small to be seen with the nakedeye. “Smal ys es inl Ary) Microorganisms are either cellular or aceilular (not formed of a cell). sels a oo ‘* Cellular microorganisms are generally uniceltilar i.e. the whole organism is one call. In such cases, a single microbial cell performs all the functions required to maintain its growth and reproduction. = According to the type of cell, cellular microorganisms are either: ~ \ 4 4u pt 5 1 Eunervetes (eu=true; karyote=nucleus) which are characterized by ae the presence of a distinct nucleus, surrounded by a nuclear membrane that separates it from the other contents of the cell Fungi are an example of eukaryotic microorganisms ,\ouf9e«, rnove Sewelopre) 2. Prokaryotes (pro=before) which represent a simpler form of cellular organization, They do not contain a membrane-bound nucleus, but their heredifa{y material is suspended in the cytoplasm and called fucleoid or nuclear region. They are also dévoid of mitochondria and other membrane bound organelles. Bacteria are an example of prokaryotic microorganisms. Seal). \es Jeuclopect * Viruses are acellular microorganisms; they have no cell structure. They are ont of the smaljest infective agents. Viruses are dbilgate intracellular parasit because se the biological machinery of a host cell for their survival and reproduction. ¥ The smallest Ling mcrory anism —> bac ter ict * elec pricNoQYamem —> Nig chateteis ensure ie ea sabes’, SYS of nomenclature (naming) for organisms, in use today Wir was the moro than two contre age eee name are iatized because Lath guage traditionalty used by scholars. ul ae jus name Scientific nomenclature assigns each organism two names—th@. ame (Plural: genera) is the first name and is always capitalized; the spec! specie the follows and is not capitalized. The organism is referred to by both the ge Specific species. By custom after a scentie name has been mentioned once Ht can Be ‘abbreviated with the initial of the genus followed by the specific species. the Scientite names can, describe an organism, hopgr a researches, or Kentty habitat of @ species, For example, covekier Stal Saal a tac ‘Commonty found on human skin. Staphylo- describes the clustel ngement cells; coccus indicates that they are Soa like spheres. The specific species, aureus, {is Latin for golden, the color of many colonies of this bacterium a CE ie ai CHAPTER 2 BACTERIA: STRUCTURE AND ORGANIZATION BACTERIAL MORPHOLOGY Bacteria are differentiated into major categories, based on their morphological features such as shape, size, arrangement and staining characteristics. Bacterial Size: Bacteria vary in size; however, they all measure in micrometres (10° millimetre). Bacterial Shape and Arrangement (Figure 1): ‘There are a few basic shapes for bacteria: A- Gocei (singular. coccus); are spherical organisms with a wide variety of SB REA: arrangements, Oot) Diplococci (pairs . 2. imegutar grape-tike clusters ( vapri\o Cok ») 3. Chains ( styeproc 0(C*) _o/ 8 Baeilli (singular: bacillus = stick); are rod-shaped organisms. These cells may be s arranged singly, in pairs or in chains. \--,\\) a DiPlobee) te! S VAP Cay ° Figure 1; Basic bacterial shapes and arrangements 3 encarta ep aye to classify Staining CI more than one ‘stains which are composed of i in_ clinical microbiology. It divides bacteria accordi to hel bacteria which appear aimgat and Gram-neds" which appear pink. _ ZiehiNeslsen stain is used to identify members of Mycobactorium. (¢w= ysv-ell » avnecae ) he genus G re Pad Fast baa & BACTERIAL ULTRA-STRUCTURES AND FUNCTIONS {All bacteria have a nucleoid, ribosomes and a cytoplasm Most bacteria have a celLwall (except mycoplasmas). , 4 eee emcoforia are futher enveloped by @ capsule ot slime |2¥er (giycetaly ) {Some bacteria have appendages e.9.. flagella and pil. * fare have Poe - Gram-negative ig membrane (Figure 2). Gram-positive Figure 2: Schematic representation of a Gram-positive Gram- te 1 ‘ * /-negative bacter Sin, Sanus RM pyc, Htc ah anda tive bacterium Le Croton tue * u (LPP Ue pe bp 8} (552 by Od encarta ea Cytoplasm: Few components can be found within the cytoplasm: a oN (bacterial DNA): Essential ee aceee of @ bacterial cell is contained in @ single chromosome formed of supercoiled double-stranded DNA. (i(C4 oN Plasmids (Figure 3): Non esertiad fn many bacteria, additional genetic information is contained on plasmids which re mer parcular ds-DNA molecules that can replicate independently of the chromosome. Bacterial ONA Plasmid nr Figure 3: Plasmids ERY ree backs 4 cattiad SME LEMS (x) They are the site of protein synthesis in the cell. Prokaryotic ribosomes have 8 Sedimentation constant of 70S, smaller than the 80S ribosomes of eukaryotes. 4s S- As This difference makes bacterial ribosomes a selective target for antibiotics. SNe eee ee ade te 9 core iS inclusion granules: Steals oS i srepe, ery grgnules of nutrient materials, e.g., carbohydrates and lipids. = -Mesosomes: 2-52» These are invaginations ofthe cytoplasmic membrane into the cytoplasm. They F are involved in cell division and sporulation. 5.» (>, (oy cvs 3 [4ePrumfotmasion) ee aod 2 Cytoplasmic Membrane: - / a ne aa * (Gtmoubune of hunan -5 sHonyl 29 theress ne cel) wall { : : 4 Hele move ssomp ie cytoplasmic membrane by simple diffusion, facilitated 2, - Excretion of extracellular enzymes: @- Hydrolytic enzymes; which aaneameacemes: whieh digest lar food molecules to ba able to penetrate the b- Enzymes used am to destroy harmful chemicals, such as antibiotics, e.g. penicillinase ensure ea 5 Be Mspiral enzymes are located in the cytoplasmic membrane: coreg te ntocrondia aU ORS» 1 me pike chon ‘aie Date pane contains enzymes eure ol wa INES parates 5. Bente ein “2 cytoplasmic membrane aitaches to the DNA and se ma Hens rom ech awe Im alon, 8 sep cre 2 1 ata pata cpa oo aur colar) ‘ palais bind to speci receptors inthe cytoplasmic membrane oe SCAtrectants and r oe Atronale tothe cols Interior. The cel then responds to the surface message. Cell Wall: oth Malt ceria! coll wall is @ strong relatively gid structure that surrounds the cytoplasmic membrane., Com PCY ~ ‘Stracture of the cell wall (Figure 4): . ‘The coll wall of bacteria fs composed of peptidoglycan, which gives the coll wall its ngth. .n, additional components exist in the cell wall. (PF. &) (VRP! . murine) ‘structure and thickness of the cell wall differ in gram-positive and gram-negative F ll il i. i. ie i. is i a Gram-positive cell wall is composed of: . ‘© Peptidoalycan: Comprises up to 50% of the cell wall material, rice © Teichoic acid \+ acs as andigen Gram-negative cell wall is composed of: ‘e Peptidogiycan: It is much thinner, comprises 5-10% of the cell wall material. Quter membrane: “ - tis ne rotein bilayer present external to the peptidoglycan ta : The outer surface of the bilayer is composed of molecules of lipopolysaccharide (LPS) which consist of a complex lipid called lipid A linked to polysaccharides. - Lipid A of the LPS forms the endotoxin of Gram-negative bacteria. = Its the space between the cytoplasmic membrane and outer membrane. i ~ Itcontains the peptidoglycan layer and gel-like solution of proteins. Functions of the cell wall: 1. It maintains the characteristic shape of the bacterium. 2. It supports the weak cytoplasmic membrane. 3. It plays an important role in cell division. 4. Itis responsible for the staining affinity of the organism. = pea te . Cell wall deficient variants: . a : e {tis the only group of bacteria that exists naturally without cell wall, therefore Mycoplasmas - donot have a defined shape - are naturally resistant to cell wall inhibitors, e.g., penicillins and cephalosporins bebe Forms: . pu * L-forms may develop from cells that normally possess cell wall, when ‘ exposed to hydrolysis by lysozyme or by blocking peptidoglycan 2 biosynthesis with antibiotics, such as penicillin, provided that they are present in an isotonic medium. sha * Some L-forms resynthesize their walls once the inducing stimulus is g\) removed. Others permanently lose the capacity to produce a cell wall. Se 7k ody * \-forms may survive antibiotic therapy; their reversion to the walled state aes can produce relapses of the overt infection. ~ geen ya MP cas conualll Deo S dost ensure ie ea eX y (poly cox aL, 4 Giydocaiye and Related Structures (Figure 5): / taining material lying Glycocalyx is defined as the polysaccharide-0on is only formed inside the host linear i) Itcan exist as a capsule(ja slime layér. outside the cell. It | ‘ d \I-defined alo oP Saat face of the cell and forms a wel { yer that adheres to the sul : ee Is 2 layer that acer aot Fader erabes Hactena. +? Ove sees Ho ceutece layer that is loosely distributed around the cell. capsule Figure 5: Capsule and slime layer it protects the cell wall against various kinds of antibacterial agents, e.g. c , complement and lysozymes. {t protects the bacterial cell from phagocytosis. Hence, the capsule is considered an ‘ important virulence factor. ¥ Some bacteria attach to the target surface by using their glycocalyx in order to \2° establish infection. 0G - For example, Streptococcus mutans forms glycocalyx, with which the bacteria stick to the tooth enamel. More bacteria become entrapped in the glycocalyx, resulting in the formation of the layer known as plaque on the tooth surface. f Then, acidic products excreted by these bacteria start the process of dental A- Flagella: Flagelia are long, whip-like appendages in many i j “They cn : eS fihe cages present ‘ genera of bacteria. A . consist i oe les of aoe type of protein called flagellin, which is highly 2s Three iypes of flagellar distribution ere known, as shown in (Figure 6): oP Monotfichdus (single polar flagellum) | Lophotrichous (muttiple polar flagella) > Perttrichous (flagella distributed over the entire cell). - = 7 Monotrichous Lophotrichous Peritrichous Figure 6: Different distribution of flagella Function: Flagella are responsible for bacterial motility; i.e. they Mave the, bacteria towards attractants (e.g. nutrients) or away from repellants (&:0- fisinfectants) by @ oe called chemotaxis. (movenrercl of bacteria. veapind iJ 40 chenu cal Factor Spirothstes move by using flagellum-Ike structures called axial filaments: cepwle These structures are composed af two groups of Thre that originate === _ within the opposite ends of the cell and overlap in the middle. «When the call moves, it rotates around its longitudinal axis, flexes and bends along its length. B- Pili (fimbriae): Same in Rackeva so net andyer Pil (singular: pilus) are hair-like appendages, shorter and thinner than flagella and are present in many genera of bacteria. «They can only be seen by the electron microscope. * They are composed of structural protein subunits termed pilin. 5 ‘ & & : Functions: 4. Adherence: They mediate the attachment of bacteria to specific receptors on the human cell surface, which is a necessary step in the Initiation of infection (je. virulence factor). CE Baus fatlee 2 PH Is ) 2. Conjugation: A specialized kind of pilus (sex pilus) forms an attachment tube between two bacter leading to the transfer of DNA between them (Figure 7). Figure 7: Sex pilus conjugating two bacterial colls 9 ensure io ea ~ mI cs 4 * Cc we é Mechanism of ter € i * ercoag's seston of te oforaam at Soi at wl uA F rkasames ut to ' be necded 1 penton. N acquires a thick cortex end 2 tin . ‘outer spore coat (Figure 8), s ovat Pray nertrane Sun Pepidogean ze # Cael ee a as a Neriooid ONA - Figure 8: Mechanism of bacterial sporulation - ‘Spores resistant than vegetative bacteria to disinfectants © Se oes "they are also more resistant {0 heating to the extent that e moist heat of 121°C for 15 30 minutes is needed to kill spores while 60°C © is sufficient to kill the vegetative forms. Hatorotrophs: © They require organic sourpas for carbon, organo bor as they can nol synthesize complex + Moat bacteria of medical importance are heterotraphic. u 2, Oxygen requirements: i zymes: st two ef For bacteria to grow in the presence of oxygen, they reauire ‘Superoxide dismutase and catalase enzymes. ‘According to O2 requirements, bacteria are classified into n for growin, e.g. Pseudomonas a- Strict or obligate aerobes: they require xygen to grow of © b- Strict or obligate anaerobes: they require complete absence fies! (because they lack superoxide dismutase and catalase enzymes), Co Bacto c- Facultative anaerobes: they generally grow better in the presence oxyge" still able to grow in its absence, e.g. Staphylococci. cause they have - Micro-aerophilic organisms: they require ret superoxide dismutase and a limited amount of catalase enzyme), 0.9, Helicon e Aerotolerant anaerobes: they require complete absence of oxygen to grow put tolerate the presence of oxygen (because they possess superoxide dismu' e.g. Clostridium perfringens. 3. CO; requirements: The minute amount of CO: present in air is st certain species require higher concentration of COz ‘e.g. Neisseria spp. .ufficient for most bacteria. However, (5-10%) for growth (capnophilic) 4, Temperature: Bacteria vary regarding the optimum temperature needed for growth as follows: + Mesophiles are organisms able to grow within a temperature range of 20-40°C (optimum 37°C). Human pathogens are mesophiles. = Psychrophiles are capable of growth at refrigeration temperature (0-8°C). + Thermophiles (heatieving) ‘grow best at high temperature (>60°C) e.g. Geobacillus stearothermophilus. 5. Hydrogen ion concentration (pH): ‘Most microorganisms of clinical significance grow best at pH 7.2. However, some microorganisms prefer acidic pH (4 or less); such as lactobacilli e iicorpanis po ) 6 lactobacilli. Others grow better at ensure ie ea ii i ace ale TENE CHAPTER 4 BACTERIAL VIRUSES (BACTERIOPHAGES) Bacteriophages (or phages) are viruses that parasilize bacteria. - Morphology of the Bacteriophage (Figure 10): The bacteriophage consists of: Tra head, containing the nucleic acid core (usually DNA, rarely RNA) surrounded by a protein coat (capsid). 2. A tail; consists of a hollow core surrounded by a contractile sheath which ends in a base plate to which tail fibres are attached. HEAD TAIL Figure 10: Structure of a bacteriophage Replication of Bacteriophages: ‘ Seer are viruses, they need to replicate inside living host cells Two cycles for phage replication are known: Ae Lytic cycle (Figure 11): ites oo ae becouse it ends in-lysis of the bacterial host cell and release of the pe eta e 3 Figure 11: The lytic cycle of bacteriophage replication B-Lysogenic cycle (Figure 12): wee Ss SS BARA ee In this cycle, the phage does not replicate or lyse the bacteria but the phage DNA\ ‘is integrated with the bacterial chromosome and divides with it to pass into daughter fells. The integrated phage genome is called “prophage” and the bacteria carrying it! are called “lysogenic” bacteria. The presence of a Prophage in the lysogenic bacterium results in bacterial, acquisition of new properties, e.g. diphtheria bacilli can produce toxin only when Lysogenic conversion (phage conversion): It is the acquisiti 3 charactor inthe bacterial host call coded for by a provhage Dhue Seawskion of 8 ne a ensure ie ea Sac Loe LYSOGENIC CYCLE Host replication Integration of genetic material Figure 12: The lysogenic cycle of bacteriophage replication important remarks: » Bacteriophages may transfer few genes from one bacterial cell to another, giving it new characters (transduction). + To start replication, bacteriophages attach to specific receptors on the surface of bacteria. This specificity means that a bacteriophage can infect only certain bacteria carrying receptors to which they can bind. Accordingly, phages are used to identify and type strains of bacteria. This phage typing is important in epidemiologic studies, e.g. to trace the source of infection in outbreaks of post- ‘operative wound sepsis caused by Staphylococcus aureus. ensure ea CHAPTER 5 BACTERIAL GENETICS and VARIATION The bacterial genome Is the total set of genes present inside the bacterial call. It comprises: 1. The bacterial chromosome ‘Additional genes may be carried on: 2. Bacteriophage DNA (prophage) (Chapter 4) 3. Plasmids (Chapter 2) Functions of plasmids: Plasmids bacteria with additional characters! properties, not essential for bacterial survival; for example: Sex pilus formation: Some plasmids code for the formation of a sex pilus which mediates the process of conjugation. Antibiotic resistance: Some plasmids carry genes for resistance to one or several entimiorobial drugs. They often control the formation of enzymes: capable of destroying the antimicrobial drug(s), @.g. B-lactamase enzyme = which breaks the BHactam ring of penicillins and cephalosporins - can be transferred among bacteria by conjugation = Virulence: Some plasmids may code for exotoxins, adhesins or invasion factors. Bacterial Variation change | vacates as in the phenotype of the bacterium or its genotype is known as stall és cel aba na aia SEs ensure ie ea ‘Table (1): Comparison between phenotypic ‘and genotypic variations Phenotypic variation Genotypic variation “lis a bacterial change which occurs | - Ttis @ bacterial change which occurs as a in response to changes in the| result of a change in the underlying conditions without genetic constitution. change in the ic constitution. ~ Reversible (transient) | [irreversible (permanent) = Not-henttable | Heritable examples: = Examples; It occurs through: 1. L-forms of bacteria 4. Mutation 2. Loss of flagella upon exposure to 2. Gene transfer: phenol a- Transformation b- Transduction c- Conjugation Mutation: It results from a change in the nucleotide sequence of DNA that may occur ‘spontaneously as a replication error, or may be induced by radiation or chemicals. Gene Transfer: ‘There are 3 methods for gene transfer among bacteria: 4. Transformation (Figure 13): Dying bacteria release DNA which can be taken up by other bacteria. Such DNA may be chromosomal or plasmid in origin and may carry genes that “transform” the Integrated DNA Figure 13: Gene transfer by transformation 17 ensure ie ea 2. Transduction: means of 8 it is the transfer of DNA from one bacteria! bacteriophage. 1 call to another PY 3. Conjugation (Figure 14): er by plasmids through jugatin (OW "4 tn ne bacterial ct 1 snot sex pilus. Chromosomal DNA Gwomosoms! ON plasmid Donor Lo Recipient DNA Fase F plasmid F plasmid Pilus Pilus Old donor New donor Figure 14: Gene transfer by conjugation nscarer ia ea APE CHAPTER 6 ANTIMICROBIAL CHEMOTHERAPY There ished groups of antimicrobial agents used in the treatment of infectious Ssceost anniotics ‘anich are natural substances produced by certain ‘groups of microorganisms, and chemotherapeutic agents, which are chemically Definitions: * Bacterlostatic agent; is an antimicrobial agent that is capable of inhibiting bacterial multiplication. Multiplication resumes upon removal of the agent. * Bactericidal agent; is an antimicrobial agent that is capable of killing bacteria. * Selective toxicity; is the ability of an antimicrobial agent to harm a pathogen without harming the host. * Spectrum of activity; is the range of microorganisms that are affected by a certain antibiotic. It may be broad spectrum, narrow spectrum or limited spectrum. MECHANISMS OF ACTION OF ANTIMICROBIAL AGENTS A- inhibition of Bacterial Cell Wall Synthesis: * These antibiotics are bactericidal with minimal tissue toxicity. © These include: ' 1. BHactam antibiotics: e.g. Penicillins and cephalosporins. 2. Glycopeptides: e.g. Vancomycin and teicoplanin. 3. Cycloserine and bacitracin. B- Interference with the Cell Membrane Function: ¢ These agents are microbicidal. * They are highly toxic as they have narrow margi i . 0 high margin of selective toxicity. 1. Antibacterial agents: e.g. Polymyxin and colistin, 2. Antifungal agents: e.g. Amphotericin B, nystatin and imidazoles. C- Inhibition of Bacterial Protein Synthesis: * Bacteria have 70S ribosomes (with 30S i i cals have Bos ne met ae bg and 50S subunits) whereas mammalian «These include: unit: @-9-- Aminoglycosides 308 ribosomal sul 1. Agents ns on sta noe ides (erythromycin) (gentamicin, amikacin, on te B08 rosea 2. Agents sxting oni onan le re may occur by: 1 JTinmpiton of RNA synthesis 0.9 2 Rewer anlage Inhibition of folic thesis: . - Oo a orainpdrofolic acid reductase enzyme eg. trimethoprim and - Ota. bition of para-amino benzoic acid (PABA) which is ‘essential for the synthesis of folic acid; €.9.. sulphonamides. rifampin. , quinolones and novobiocin. RESISTANCE TO ANTIBIOTICS ‘Resistance to antibiotics Is either intrinsic or acquired: 1. intrinsic resistance: it is natural resistance to an antibiotic i.e., the organism lacks the target or receptor for the antibiotic. Enterococci are naturally resistant to cephalosporins. 2 Acquired resistance: Itis the resistance to an antibiotic due to the acquisition of resistance genes. Two genetic processes are involved: @- Mutation. b- Gene transfer: by transformation, transduction or conjugation. 8, Reduction of intracellular concentration of the antibiotic due to: -A A decrease in the influx of antibiotic by reduction in the permeability of the . Pesos efflux pump: antibiotic across ~ Pamanee of te ne amie me b oa eee he antibiotic by destroying enzymes F hydrolyse P-lactam ring thus inactivate penicillins and eG. See ne Sop/yfocnoan sues eee (PBPs) results in mecithillin a Bctngjeg sina eae , bY developing winethopnm, SereoPing New metabot: pathway: 9. resistance tc COMPLICATIONS OF ANTIBIOTICS (yellowish discolouration) of teeth in Met -os may cause permanent staining ( ‘Strepton cranial nerve leading to deafness. - Sueno ries may cause nephrotoxicity. can cause bone marrow depression. Allergy (hypersensitivity): @.9., a - Penicilins may cause he anaphylactic shock or serum sickness. i = Local application of sulphonamides may result in contact dermatitis. Emergence Resistant Strains: ae * Ae See ot Retbiotics (low dosage, interrupted course, no real indication and the emergence of resistant mutants. | RP cece wil overgrow and replace the originally susceptible Dacor “Therefore, it is recommended that in vitro susceptibility testing should be performed to guide the selection of antibacterial drugs. 4. Superinfection: It occurs as a result of overgrowth of resistant members of normal flora when the sensitive ones are eradicated during antibiotic therapy; €.9. ~ Oral thrush caused by overgrowth of the yeast Candida. = Bseudomembranous colitis (antibiotic-associated diarrhoea) caused by overgrowth of Clostridium difficile. Empiric Antibiotic Therapy Empiric antibiotic therapy (also known as “best guess therapy") is the use of antibiotic for therapy prior to the availability of antibiotic sensitivity results. - Depending on the site of infection, there will be a short list of bacteria most likely to be causing that infection = Depending on the type of bacteria, there will be an antibiotic most likely successfully treat that infection. an ~ Enipiric antimicrobial therapy Is indicated for severe oral and dental infections becat of the possibility of infection spread via lymph and blood ircuation with, the { a i ymp! circulation with the development of septicemia; for example facial cellulitis, pericoronitis (swollen and infected Lethe) around the wisdom teeth), lateral periodontal abscess and necrotizing NB. Empiric ther ben ee be started without delay but after collecting specimens for er encarta ea joTIC THERAPY COMBINED ANTIBI tiie 19 sch necessitate tne use OF MO than one 30 There are conditions wh _order to achieve @ ‘successful clinical sepsis in Indications, of having serious infections; ©-9- achieve bactericidal : jcillin combined action through synergistic effect; e.g., pen! n co ’ br ‘metronidazole in the treatment of acute necrotizing ulcerative gingivitis. massive trauma (caused by aerobic and 5. Mixed infections ©.9. infections following anaerobic bacteria). ‘ ; OICE OF AN ANTIBIOTIC ‘ be done before giving an antibiotic. ite of infection and achieve ” GUIDELINES FOR THE CH 4. In vitro culture and susceptibility testing should 2. Select an antibiotic that is able to penetrate to the si effective concentration. 3. Identify the nature of infection, whether bacterial, viral or fungal. Antibiotics are only given for bacterial infections. --4. Choose a narrow-spectrum antibiotic rather than a broad-spectrum one. The use of broad-spectrum antibiotics is likely to faster the induction of resistance to antit ibiotics and may be complicated by superinfection. ane 5. on bactericidal rial than bacteriostatic antibiotics. 8. .e appropriate dose of the antibiotic for the proper duration. Inadequate t r . te dosa, , = et prolonged therapy may result in antibiotic resistance or drug tonicity. “ . eS potential of the drug to produce toxicity and use antibiotics that are only certain patients (e.g., pregnant or lactating women and for infants anc children). j ensure ie ea ANTIBIOTIC PROPHYLAXIS ‘Antibiotic prophylaxis is the administration o of an antibiotic to prevent rather than to treat an infection caused by & specific Examples: ~ «Penicillin or erythromycin; ie given to individuals with abnormal heart vaives prior fo dental procedures to prevent endocarditis. © Preoperative in some surgical operations; such as Implant surgery where 2 grams of ‘amoxicillin are given orally 1 hour preoperatively. . Rifampicin: is given to close contacts of meningococcal meningitis cases for 2 days meningitis. CHAPTER 7 STERILIZATION AND DISINFECTION one of the primary responsbities of dental healthcare professionale ie to create © and comfortable environment for their patients. This involves sterilization of all types of dental instruments, each time they are Basic Terms and Definitions: Steritzation; i @ process, which if performed property, kills all microbes, Disinfection: is a process in which all microbes are removed, except bacterial Ersinfection has been categorized into 3 levels; high, intermediate and low as follows (Figure 15): 1. High level disinfection: It destroys all vegetating bacteria (including Mycobacterium tuberculosis), fungi and viruses. [Link] level disinfection: It destroys all vegetating bacteria (including Mycobacterium tuberculosis), majority of fungi and majority of viruses. 3. Low level disinfection: It destroys vegetating bacteria, some fungi and some viruses. It is not effective against resistant bacteria such as Mycobacterium tuberculosis, which is used as an indicator because it is relatively resistant to Crearetent spores syeonnetere Small non-envetoped virumes Srem-negative bacteria Pune Large non-envitoped vieueen Srem-poative pacterm Lind anvetonen v. Figure 15: Descending order of ‘ crocrganisms to sterilization and Giaibecticy ensure ie ea +: are special { chemical disinfectants (e.g. ethyl and isopropyl Peri ianentcanlbs! safely aiid to the skin but are not suitable for systemic administration Cleaning; is an essential process which Is performed before sterilization or disinfection of dental instruments. This involves cleaning all surfaces of dental instruments to remove blood and organic material immediately after use to prevent their drying. It is usually done with water and soap, detergents or enzymatic products. Decontamination; is a general term that is applied to any procedure by which pathogenic microorganisms are reduced to a level where items are safe to handle (Figure 16). Decontamination includes sterilization and all disinfection levels. Figure 16: Decontamination steps Selection of an Adequate Level of Decontamination: Dental instruments classification - into and patient care items are classified - i epending on the ik of wane gees (etic, cembeiileal ang mana {© Spaulding of transmitting infection (Table 2), and non-critical items) 25 ensure ie ea mended ification of instruments and patient care items with the recom Table (2): Classi methods of decontamination $ Recommended 7 Characteristics Examples decontamination method(s) i Critical used to forceps, scalpels, Sterilization after each use 4 items ‘softtissue | bone chisels, or bone, or enter Into | scalers and or contact the surgical burs bloodstream or other normally sterile tissue ‘Semi- Spec that come in| mirors, reusable | Sterilization after ‘each use. critical. | contact with mucous impression trays | In some cases, however, items | membranes or non- | and amalgam sterilization is not feasible and, intact skin condensers therefore, high-level disinfection is appropriate. A high-level disinfectant must be : labeled as “sterilanUdisinfectant” to be used. ‘Such devices have a relatively Non- Objects that contact external critical | intact skin only but not | components of low risk of transmitting items: mucous membranes | x-ray heads and _| infection; and, therefore, may pulse oximeters be reprocessed between patients by intermediate-level or low-level disinfection. NB. Sterilization is recommended for all high-speed dental hand-pieces, low-s| hand-piece components used intra-orally and reusable prophylaxis angles. ‘ MAIN METHODS OF STERILIZATION : . 1. Heat: 1. Moist heat or steam sterilization [Link] heat sterilization Il. Low temperature ("Cold") sterilization methods: A- Chemicals: Liquid sterilization process Commonly used liquid sterilants include: - Gluteraldehyde 2% (contact time 10 hrs) - Hydrogen peroxide 6 - 7.5% (contact time 6 hrs) 26 ensure ie ea the single important variable distinguishing the © Contact, time Is rom high-level disinfection. a I should be finse ith_sterile in this manner should be rinsed with sterile . insteamarts ie ace a stan conte t used immediately). B. Hydrogen peroxide gas plasma C- Ethylene oxide gas IW. tonizing radiation rilizatior There are two forms: moist heat (or steam sterilization) and dry heat. Steam Sterilization: The steam sterilizer (autoclave) is the most effective and most commonly used method of sterilization jin dental practices for items that can withstand high -al dressings, cotton and gauze. temperatures; such as surgical instruments, surgicé » Principle: nC irvhen water is heated in a closed vessel at @ pressure abo boiling point of water rises above 100°C. Thus, pressure serves aS @ means to obtain the high temperatures f necessary to quickly Kill microorganisms. The autoclave uses moist heat in the form of saturated steam under pressure for a specified exposure time and at a specified temperature. Saturated steam means free of air because air acts as an insulator and hinders penetration. - The autoclave can atmospheric pressure ve atmospheric, the be used at 121°C for 15 - 30 minutes (at double = 2 bars) or at 134°C for 3-6 minutes (at 3 bars). © There are two basic types of autoclaves: ee Tho gravity displacement autoclave where incoming steam displaces residual air through a drain in the bottom of the sterilizer chamber. - The prevacuum autoclave is similar the gravity displacement autoclaves except that it is fitted with pump to ensure air removal from the sterilizing chamber befo dmitted. The advantage of using a vacuur s steam penetration. é © This method results in shorter cycle removal of air and the usually higher ter 27 ensure ie ea ae ain in ize and the packs rama” oclave must depressurize a” The autoclay Aftor the storiization cycte, Ine eer rase may take 20 - igure “n). the sterivpe opened ater completion of the drying cycle tored in a clean, dry area. tbe si ‘ utoclave, sterile packs must * Upon removal from the ai a & ee Figure 17: Removal of sterile dental packs from the autoclave «Advantages of the autoclave: ; | 4. Agood ability of hot saturated steam {o penetrate into loaded porous items. 2. Liberation of latent heat of vaporization after cooling due to condensation of the However, the main disadvantages include: 1. Instruments must be dried at completion of sterilization cycle. 2. Some items cannot withstand steam at high temperatures, e.g., rubber and plastic. 3. Steam sterilization is not suitable for sterilizing powders and oils. 4. Autoclaving tends to rust carbon steel instruments and burs. “Monitoring of steam sterilizers (autoclaves): 1. Mechanical indicators; jen physical measurement of the time, temperature and ( 2. Chemical indicators; chemically impregnated : at certain temperature (0.9. 121°C oF 132°C) ave ning cals their colour assess the level of sterilization, : each areas, to § N.B. Mechanical and chemi help detect proced ensure ie ea strips containing the spores of Geobacillus 3. Biological Indicators; are paper indicators are placed in ‘Giout to-reach areas. stearothermophilus. Ldap pees asl ion, spore strips are incubated in a fluid medium for viability. Absence of bacterial growth indicates an ‘spore should be used on each sterilizer al least weekly. It should also be aoa eotey toad wth an implantable device. Ideally, implantable items should not be used until the spore test result reads negative. Dry Heat Sterilization: 4. Incineration: is particularly applicable for infectious hospital waste such as used surgical dressings, needies.....etc. 2. Hot air sterilizer (hot air oven): In this method, the sterilizing agent is dry hot air with the following schedules: Table (3): Sterilization temperatures and exposure times in hot air sterilizers ‘Sterilization temperature Exposure time 60 minutes 120 minutes ‘Major advantages of the hot air sterilizer: 4. Absence of a corroding effect 2. Used for sterilizing metallic instruments, powders, oils and glassware 3. Low cost Major disadvantages of the hot air sterilizer: 1. Slow and uneven penetration of heat into the Toatorials to be sterilized 2. Prolonged exposure time 3. Damage to the rubber items and some fabrics * Monitoring of hot air sterilizers: Bacillus atrop ; a biological indicator. 29 nscarer ie ea Hydrogen peroxide gas plasma: ‘* Gas plasmas have been referred to as the fou solids, gases and gas plasmas). Gas plasmas chamber under deep vacuum using radio freq excite the gas molecules and produce charged particles, many of the form of free radicals. «The free radicals interact with essential cell components (2 acids) and thereby disrupt the metabolism of microorganism: direct inactivation by hydrogen peroxide. Total time of sterilization cycle is about 50 minutes. Medical materials and devices that cannot tolerate high temperatures rth state of matter (i.e. are generated it metal alloys, can be sterilized by this method. * Geobacillus stearothermophilus spores are used as a biological indicator to monitor efficiency of the sterilization process. Ethylene oxide gas: Exposure time is long (3 to 6 hours). The method is expensive, explosive and toxic. Bacillus atrophaeus spores are used as a biological indicator. lonizing radiatio: Two types of ionizing radiation are commonly used for sterilization: 1. Gamma rays; which are emitted by radioactive elements such as cobalt-60. 2. High-energy electrons (B-rays): which are produced by electron accelerators. lonizing radiation has a high penetrating power and is, theref i i . ore, a means for sterilization of pre-packed heat-sensitive items such as bone grafts, ps liquids, in an enclosed r microwave energy to Mericies f which are in enzymes, nucleic in addition to the humidity, such as some plastics, electrical devices and corrosion-susceptible It can be used for devices that cannot be subjected to steam (rubber and plastic). ln i il, i. ‘sutures, dis i sures lisposable plastic syringes, gloves, catheters and intravenous (IV) Infusion 4 * Monitoring of radiation steril biological indicator. 30 ensure ea ization: Bacillus pumilus ‘Spore strips are used as a | MAIN METHODS OF DISINFECTION 1. Boiling water: Boiling at 1 00°C for 20 minutes achieves high disinfection. It can be useful in emergencies if no sterilizer is available. rization (temper ire lower than 100°C): IL Pasteurization ak 63°C for 30 minutes or at 72°C for 20 Pasteurization of milk by heating at ~ seconds, followed by rapid cooling, destroys e.g. Mycobacterium bovis. Ill, Ultraviolet radiation: important pathogenic organisms in milk; Utraviolet imadiation (UV) is a low-energy, non-ionizing radiation present in sun rays oF in operating rooms. attificially produced by mercury lamp’ power. Therefore, UV rays can be u! s. UV radiations have extremely weak ised only for air and surface disinfection IV. Chemical disinfectants (Table 4): Table (4): Commonly used chemical disinfectants Disinfectant Use ‘Alcohol 70% (ethyl alcohol | Used as a skin antiseptic (alone or cor isopropyl alcohol) combined with iodine or chlorhexidine) ‘Biguanides Used as an antiseptic for the skin and (chlorhexidine 2%) mucous membranes (as a mouth wash). It F is often combined with alcohol as a 8 handrub. Hydrogen peroxide ‘Antiseptic for open wounds or as a mouth wash (6%)_ Todophores e.g. povidone | 1% used as an oral antiseptic solution iodine 7.5% used as a scrub solution 10% used as a skin antiseptic Gluteraldehyde 2%* High level disinfection or sterilization of instruments | Chlorine-active compounds | Decontamination of blood spills (1:10 of e.g. hypochlorite or household | 5.25% for a contact time of at least 20 bleach (5.25%) minutes) } Phenol-containing } preparations (phenolics) Cleaning of the floors, walls, furniture ‘Quaternary ammonium f compounds Exposure to gluteraldehyde can cause severe irritation to the skin, mucot system. Its vapors have been associated with cases of occupational ast ‘outside a closed container. 31 cantare pe ep Filtration i jutions, A process used to remove bacteria from thermolabile fluids {antibiotic so! hormones, vitamins) that cannot be purified by any other means. Se can Tendored free of bacteria by passage through bacterial membrane filters pore size as small as 0.22 pm. - The endopigment producing Serratia. marcescens is used to efficiency of bacterial membrane filters. test the Filters can also be used to remove microorganisms from air supplied to critical areas such as operating rooms. ‘Such filters are known as high efficiency particulate air (HEPA) filters. - Spores of the fungus Aspergillus is used {0 test the efficiency of HEPA filters. CHAPTER 8 BACTERIAL PATHOGENESIS Infection: Entrance and multiplication of microorganisms on/in the host. Disease: Damage of the host tissue due to invasion or toxin production by microorganisms. ‘Most infections end without ‘occurrence of pathological changes and thus are not manifested as clinical disease. These infections are termed subclinical, silent or abortive infections. The outcome of bacterial infections depends on the mutual relationship between bacteria and host. Accordingly, bacteria could be classified into: 1. Saprophytic bacteria: are those which live freely in nature and they do not require a living host. 2. Parasitic bacteria: are those which live on or in a living host. They are classified according to their relation to the host into: a Pathogenic: Bacteria capable of causing disease, BL Nompathogenic (commensals): Bacteria that do not cause disease and are part of the normal flora’. c Opportunistic pathogens: These are potentially pathogenic bacteria that do not Qppertiiisease under normal conditions but can cause disease in famunocompromised patients, or when they find their way to another site other than their normal habitat. Many of these opportunistic pathogens are originally commensals. * The normal flora denotes the population of microorganisms that inhabit the skin and mucous membranes of healthy normal people (discussed in detail in Chapter 34). Stages of the Infectious Proces 2. Mode of transmission e.g. airborne, di contact and transplacental. 3. Portal of entry e.g. respiratory tract, gastroi then starts to multiply within the host causing tissu 4, Portal of exist e.g. urine, stools, blood, respiratory or the organism is transmitted to a new host. 33 ensure ie ea Modes of transmission include: G4 L._ Airborne (aerosol) transmission (small- ‘irbome transmission occurs by dissemination of either airborne droplet nuctel (tr bene particles; $5 ym in size) produced from evaporated droplets oF containing the infectious agent. ‘ be «Microorganisms remain suspended inthe air for long periods of time and can dispersed widely by air currents. «Tro saiclos may Become inhaled by a susceptible host within the same room oF over a longer distance from the source. it fl, Droplet transmission nctivae or the mucous membranes of Droplet transmission involves contact of the conjut um in size) the nose or mouth of a susceptible person with large droplets (>5 oS containing microorganisms generated during coughing, sneezing or talking. «Transmission via large droplets requires close contact between the source and the recipients, because droplets do not remain suspended in the air and generally travel only short distances, usually one metre or less, through the air. Ml. Contact transmission Contact transmission is the most common form of transmitting diseases. «There are two types of contact transmission: direct and indirect. ~ Direct contact transmission occurs when there is physical contact between an infected person and a susceptible person. Examples of direct contact are touching, kissing, contact with oral secretions, i ore tu , oF contact with body { - Indirect contact transmission occurs when there i i iS no. direct human-to- human contact. Contact occurs from a reservoir to contaminated aries { or objects (such as door knobs, hand-pic i tu mengitioon fies thd Orn ind-piece and chairs) or to vectors (such 4 i 34 cancers ep Carriers: rently healthy individual harbouring © pathogenic organism, oP eatations ry an transmit this organism to others. jer state, carriers may be: Transient carriers e.g. during the ¥ cabation period and early convalescence, o Ghronie carers ‘sg. hepatis-B carriers: The organism may be discharged from the carrier in an intermittent or @ continuous manner. A cartier is an without having clinical ‘According to the duration of the cart is than cases aS @ source of infection. This is Carriers are more dangerou because: 1. They are not known to public. 2. They are not easily detected. 3. They are not restricted to bed. They carry the organism in the inter-epidemic periods. BACTERIAL VIRULENCE of thé organism to cause disease (qualitative) Pathogenicity is the ability Virulence is the degree of pathogenicity (quantitative). Virulence Factors of Bacteria: ‘A virulence factor is either a structure or 2 P cause disease. oduct that enables the organism to A- Adherence factors: « They enable bacteria to adhere to th establishment of the infection. e host surfaces, thus contributing to the For example: ror erammbriae of Neisseria gonorrhoeae help their attachment to mucosal epithelial cells. Streptococcus mutans forms glycocalyx, with which the bacteria stick to the tooth enamel. B- Invasion factors: in mechanisms by lowed by inflammation is one of the mair Invasion of tissue fol isease. This invasion is helped by: which bacteria can cause di 4, Enzymes: They include: * Collagenase . © Lipase < Immunoglobulin A1 protease: This degrades IgA. 35 2. Antiphagocytic factors: They include * Capsule: prevents phagocytes from adhering to bacteria. ‘+ Cell wall proteins: e.g., M protein of Streptococcus pyogenes Pao + Coagulase: It forms fibrin clot from fibrinogen, This clot can protect bacte! Phagocytosis, e.g. Staphylococcus aureus. €- Toxin production: Toxin production is another mechanism by which bacteria can produce disease. Bacterial toxins are either exotoxins or endotoxins (Table 5). Table (5): Comparison of the main features of exotoxin and endotoxin Exotoxin Endotoxin Secreted by living organisms [Integral part of the cell wall of both Gram-positive (mainly) | Gram-negative bacteria, therefore and Gram-negative bacteria | liberated only upon cell death and lysis Encoded “by genes on|Encoded by genes on the chromosomes, plasmids or | chromosome bacteriophages Protein Lipopolysaccharide Heat stability | Unstable at temperatures | Stable at temperatures above above 60°C 60°C for several hours Toxicity igh Low Antigenicity | Highly antigenic Poorly antigenic Detoxification | Can be converted into toxoid® | Can not Specificity Every toxin has a specific | Non-specific action, all give fever action and shock Examples Corynebacterium diphtheriae | Meningococcal endotoxins (phage) Clostridium tetani (plasmid) * Treatment of exotoxin with formalin removes its toxicity and of retains its antigenicity converting it into toxoid, which can be used for immunization. Dental unit watertines are usually lined by a biofim bacteria. These bacteria are rolea: composed predominantly of Gram-negative then delivered through the distal 204 n high numbers info the dental unit coolant and irigant waters) with the potential for endotoxin Inrvasclrcongulaton. Taste iis eee a intravascular coagul werefore, itis Pyrogen-‘re0 sterile water forall surgical dental procedures. : pace Recent studies suggest a Song correlation between endotoxins and clinical canal infection. ensure ie ea IMMUNOLOGY Ml. Acquired Immunity (Adaptive Immunity): Table (6): Comparison between innate (natural) and acquired (adaptive) immunity | \ { CHAPTER 9 OVERVIEW OF THE IMMUNE SYSTEM inly involved in combating infections of most tumours. caused by The immune system is mai different microbes and preventing development The immune response to infections includes two parts: 1. Innate Immunity (Natural Immunity): ivi i II times. It is very This is present in all normal individuals since birth and at al n important at the beginning of an infection. However, it is not always successful in Siminating infectious organisms on its own. It does not increase with repeated ‘exposure ‘and cannot tell the difference between different pathogens. It includes many different resistance mechanisms: such as barriers, chemicals ‘and cells present in the body; mainly granulocytes (neutrophils, eosinophils and basophils), as well as monocytes/macrophages and natural killer (NK) cells. { This is a specific immune response directed against a particular pathogen. It is { associated with the development of immunological memory, so that in many cases it | gives life-long protection against the same pathogen. It is very effective in combating infection, but there is a delay of a few days until it is effective. | It depends mainly on T and B lymphocytes and is divided into cell-mediated and | humoral (antibody-mediated) immunity. Innate Acquired Presence ‘Since birth Following exposure to athogens ‘Onset of action | immediately after infection Relatively delayed Main cells Granulocytes, monocytes/ B and T lymphocytes macrophages and NK cells ee i Memory ‘Absent Efficiency Less efficient and does not increase on repeated exposure Specificity Non-specific 37 ncaa ea ’ CHAPTER 10 INNATE IMMUNITY microorganisms in the protected entially harmful Any individual from Polen nit since birth, They are * gqviconment by @ number of very effective mechanisms prese « ton-specific and can act against any ‘microorganism. Mechanisms of Innate Immunity: 1, Mechanical barriers and surface secretions: 4. Intact skin and mucous membranes constitute a barrier that cannot be penetrated by most microorganisms. ; 2. The sticky mucus covering mucous membranes traps any foreign material. F Gita of the respiratory tract epithelium sweep foreign material out. Blinking, sneezing and coughing reflexes expel foreign particles. $. Sweat and sebaceous secretions contain substances that inhibit microorganisms. © Saliva, tears and mucous secretions of respiratory, alimentary and genito-urinary tracts contain lysozyme whieh is bactericidal. 7. Gastric and vaginal acidity inhibit growth of microorganisms. 8. The flushing action of saliva, tears and urine helps in washing microbes from the body. “4. Normal flora: Produce bacteriocins and acids that destroy microorganism compete with pathogens for essential nutrients. s and Soluble defence mechanisms: They are a number of microbicidal substances present in tissues and body fluids. They include: 4. Lysozyme: it destroys peptidoglycan of the bacterial cell wall. 2. Complement: It is a group of plasma proteins that are present in an inactive form al § m ind can be activated by certain pathogens leading to their lysis (chapter 16). 3, Acute phase proteins: These are present at very low levels in normal serum, but their concentration rises dramatically shortly after the onset of an infecti to destroy or inhibit growth of microbes; io fl inoge rand C- ee eat 9s; examples include fibri in and C- 4 Sea a raeia (a and f interferons): They are secreted by virus-infected replication in uninfected neighbouring cells. ena 38 ‘interfere’ with viral cantare ep 1V. Cellular defence mechanism: The process of phage 1. Migration (chemotaxis): lv. Intracellular Killing: (phagocytes, natural killer cell ind eosinophils) 1. Phagocytes: These are cells that engulf and destroy particulate matter including bacteria by a hagocytosis. process called pt The main phagocytes are: Neutrophils * Monocytes/macrophages (monocytes in the blood and macrophages in tissues) + Dendritic cells Phagocytosis: }ocytosis occurs in subsequent steps (Figure 18): * Microorganisms and injured tissues produce chemotactic factors that attract Phagocytes to the site of infection. fi, Attachment: * Attachment occurs between receptors on the surface of phagocytes and non- Specific molecules common to many pathogens. * Some microorganisms may lack these molecules or cover themselves with a thick capsule that is not recogni: ; €.g. complement or other molecules (e.g. CRP). This process is thus called opsonization”, {,Opsonization is the process by which bacteria are rendered more easily engulfed by phagocytes, lil. Engulfment: * The cytoplasmic membrane of the pha encloses it in a vacuole termed . Which qazymes, then fuse with the phagosome forming a phage ani the engulfed material is killed and digested, * Once ingested, most microorganisms are dig (e.g., lysozyme, elastase and hydrolase). 39 cantare eat TELAT NGL RT © Figure 18: Stages of phagocytosis killer cells: These are large lymphocytes capable of non-specific killing of tumour cells and cells by an extracellular killing mechanism. 5 3. Eosinophils: Eosinophils have cytoplasmic granules which contain enzymes and toxic molecules that are active against parasites. The release of these molecules allows the extracellular killing of parasites that are too large to be phagocytosed, e. helminths. V. Inflammation: ES Inflammation is a process by which the body's white blood cells and substances Q produce protect the body against infection with foreign microorganisms, such as and viruses. Chemical mediators released at the site of infection trigger an response in the form of vasodilatation, increased vascular permeability ind migration of white blood cells from the blood stream across the vascular ‘endothelium into the inflamed tissues to combat the invading microbe. l l encarta ea ‘ CHAPTER 11 ; ANTIGENS antigen stem An (or immunogen) is a substance that can stimulate the immune syst to produce an acquired immune response and reacts specifically with the product of this response. Epitopes (Antigenic Determinants): = The immune system does not recognize the antigen molecule as a whole but reacts to structurally limited parts of the molecule called epitopes. - They are very small, composed of just four to five amino acids or monosaccharide residues. ~ They determine the specificity of the antigen. . The same antigen may possess different epitopes. that share one or more similar epitopes are known as cross-reactive ‘similar a _-— epitopes ~_ | CQ )) Cy s at different (3% epitopes o { | Figure 19: Heterophil antigens Hapten: * This is a low molecular weight substance whi t Ms alone (nonentoata ee mien is not able to induce an immune coe np eat an antigen vhen coupled with a . ae of haptens include bacterial capsular polysacchi 4t encarta eat rs e 5 Factors Affecting Immunogenicity: 1. Forelannes®: acule to be antigenic, it must be foreign to the host in which it is troduced. The immune system can normally distinguish between body components (call) and foreign substances (non-self) and |s normally tolerant (non-reactive) to self- antigens (autotolerance). : 2. Molecular Size: . Usually, the larger the molecule is, the stronger the antigenicity. 3. Chemical Nature: ; ; The chemical complexity of a molecule contributes significantly to its immunogenicity. The molecule is more immunogenic if it is more complex, therefore, the most potent immunogens are proteins. 4, Route of Administration: a The route of administration of an antigen may affect the type and intensity of the response. As a general rule, the subcutaneous and intramuscular routes are e best in provoking an immune response. ‘5. Dose: There is an optimum dose at which any antigen is most immunogenic. Very low and very high doses may result in a state of unresponsiveness (tolerance). 6. Adjuvants: ‘Adjuvants are non-specific potentiators of the immune response. The administration of an adjuvant together with an antigen enhances the immune response to that antigen. : (One of the commonly used adjuvants is aluminium hydroxide (alum) which is added to diphtheria and tetanus toxoids that are used for human immunization. Aluminium hydroxide delays the absorption of toxoids and prolongs the period of their exposure to the immune system. 7. Host Factors: _ The generation of an immune response is genetically controlled. Thus, some antigens stimulate an immune response in humans but not in animals. Also, the immune system may not respond to an antigen at the extremes of age (due to immaturity or aging of the immune system). ’ 42 CHAPTER 12 ' ACQUIRED IMMUNITY | main arms; cell- Acquired (or adaptive immunity) immunity is classified into two mediated and humoral (antibody-mediated). The main cells involved in the adaptive immune response are: "lymphocytes classified into T-helper and T-cytotoxic cells | | | + Blymphocytes | Both cells originate and mature in lymphoid organs. \ \ | e Lymphoid Organs: Lymphoid organs are divided into: A. Primary (central) lymphoid organs: ‘e The bone marrow: It is the site of origin of T and B cells. It is also the site where B cells complete their maturation. «The thymus gland: It is the site where T cells complete their maturation. B. Secondary (peripheral) lymphoid organs: «These are the sites where mature Iymphooytes meet antigens leading to their activation. Secondary lymphoid organs include the spleen, lymph nodes and various mucosal associated lymphoid tissues (MALT) such as the tonsils, adenoids appendix and Peyer's patches. " Circulation of Lymphocytes between Blood and Lymph (Figure 20): 1. Small B and T lymphocytes that have matured, but have not 1 called naive lymphocytes. They leave the bone marrow and and circulate continually from the blood into the secondar the lymph nodes, until they meet an antigen or die. et met an antigen are 2. Microbial antigens are drained from the site of infect tion Lymphocytes recognize the antigen and then undergo a se make them ready to start working against the antigen to which the 43 ensure ea ‘The changes which occur are: a. Activation, Lymphocytes become lymphoblasts. : b. Proliferation; Lymphoblasts undergo rapid multiplication. ¢. Differentiation; Lymphoblasts change into effector cells, capable of being effective against the invading agent: © Bcells change into plasma cells, capable of Cytotoxic T cells and helper T cells become @' effector helper T cells, respectively. secreting antibodies. ffector cytotoxic T cells and e efferent lymphatic vessels and lymph nodes through thi lood, they reach the 3. Effector cells now leave the retum to the blood through the thoracic duct. From the bl peripheral tissues where they start functioning. Some of the antigen-specific lymphocytes produced by these events remain after the antigen has been eliminated. These are ‘called memory cells and are the basis of immunological memory which ensures a more rapid and effective response on a second encounter with the same pathogen and, therefore, gives long lasting immunity. {3} Lymphocytes return via thoracic duct to [blood and from there} to tissues 44 cantare pe ep Recognition and Effector Mechanisms of Acquired Immunity: Different pathogens need different mechanisms for recognition: « Effector B lymphocytes recognize extracellular pathogens; such as most bacteria. © Effector T lymphocytes recognize intracellular pathogens; such as viruses and intracellular bacteria. CED: Effector mechanisms: 1. Effector Mechanisms of B cells: { Effector B cells (plasma cells) produce antibodies which eliminate extracellular pathogens and their toxins. Antibodies are found in plasma and extracellular fluids. | _Immunity mediated by antibodies is called humoral immunity. Mechanisms of T cells: All viruses, some bacteria and parasites replicate inside cells, where they cannot be detected by antibodies. The destruction of these invaders is the function of the T lymphocytes. This is called cell-mediated immunity. 45 cantare ep CHAPTER 13 CELL MEDIATED IMMUNITY The main functions of T cells are the destruction of intracellular pathogens as ~) well as helping other cells of the immune system. As mentioned, for T cells to start ful _ response, they must first change from naive T cells into they meet their specific antigen. nctioning in the acquired immune ‘effector T cells, the first time _ Recognition of antigens by T cells requires the following (Figure 21): -"e Antigens to be carried on special molecules called major histocompatibility e complex (MHC) molecules present on the surface of antigen presenting cells (APCs). © T cell receptors (TCR), with the help of a number of other molecules, present on the T cell. receptor MHC antigen ———> Figure 21: T cell recognition of an antigen Antigen Presenting Cells: These are the only cells capable of activating naive T cells. Une) include: |. Dendritic cells are the most important APCs. Thi eo . They are so-called because they have am mal ue projections called aiiee They are present in nearly all tissues of the sells and are essentially cells of the innate ws:them to contribute to the acquired ral immunity, but they can also act encarta a MHC Molecules: MHC molecules are cell-surface glycoproteins. . * There are two classes of MHC molecules which are important in T cell activation; known as class | and class !| MHC molecules (MHC | and MHC Il). - All nucleated cells of the body have MHC | molecules on their surfaces. ~ APCs have both MHC | and MHC II molecules on their surfaces. T Cell Surface Molecules (Figure 22): These are molecules important in antigen recognition, interaction with other cells and serve as markers for identifying T cells as well as dividing them into subsets. The most important T cell surface molecules are: 4. T Cell Receptor (TCR): * The T cell receptor can recognize certain antigens. + AI TCRs ona single T cell are identical and recognize the same antigen. TCR —— ‘achain Pehain' Figure 22: T-cell surface molecules 2. D3 Molecule: * This is found close to the TCR on all T cells. + Itis involved in transmitting signals from the TCR to the inside of the cell, 3. CD4 and CD8 Molecules: T cells carry either CD4 or CD8 molecules on their surface and classified into two major classes: Se tus * CD4 T cells; also called T helper (Th) cells, and their main function i other cells of the immune system by eecreting cytokines UHR Is to help + CDB T cells; also called T cytotoxic (Te) cells, and their main function is to i infected colls and tumour cells. in function isto kill citi i “. cine i ll ce Al tc 4. CD28 Molecule: This molecule * During the process of ant! present on the APCs, as part of present on all T cells. igen recognition, it binds to a molecule called B7 f T cell activation. 5. CD40 Ligand (CD40L): se This molecule is present on activated T helper cells. . It is involved in activation of B cells by T cells. Sequence of Events in Activation of Naive T Cells: 1. Antigen Presentation to T cells (Figure 23) * For antigens to be presented to T cells they must first enter inside the APC; undergo degradation (antigen processing) producing peptides. « These peptides are carried inside clefts on the MHC molecules to the cell surface, to be presented to T cells (antigen presentation). Figure 23: Antigen processing and presentation Il. T cells Recognition of Antigens (Figure 24) «+ Peptides carried on MHC I molecules presented on the surface of APCs will wager COB (cytotoxic) T cells with TCRs specific for the MHC I-peptide complex. «Similarly, peptides carried on MHC Il molecules presented on the surface of APCs will trigger CD4 (helper) T cells with TCRs specific for the MHC II- 5 peptide complex. Antigen n "| vy “ la ; D4 receptor # | bg CD8 receptor Figure 24: CD4 T cell and CD8 T cell recognition of antigens 48 iu. Teells Activation (Figure 25) «The binding of TCR first signal needed fc Then, CD28 (presen second signal neces: ‘As a result, the T call becomes a lymphobl repeatedly (proliferation) and cells (differentiation). This sequence of events is known as addition to the production of effector T cells, which are long-lived T cells tuth a rapid response and provide protection against s memory T cells, same pathogen. Functions of effector T with MHG (1 or ti)-peptide complex is considered the (or T cell activation. i ton T calls) binds to B7 (present on APCs) delivering sary for T cell activation. cp4T cell Figure 25: Signals for T cell activation last (activation), divides its progeny (daughters) finally become effector the primary immune response. In it also results in the development of that respond to antigen stimulation ubsequent exposure to the cells: Functions of effector CD4 T cells (Helper T cells): The main function of e' According to the types of se ffector CD4 T cells is to secrete cytokines. creted cytokines, they are classified as either Th1 or Th2 calls. "Tht cells: produce cytokines which predominantly activate macrophages making them more capable of inflammation. f killing the bacteria they harbour. They also promote 2. Th2 cells: produce cytokines which predominantly help B cells and promote humoral immunity. 11, Functions of effector’ in funct - virus-infected cells otoxic T Cell: is to oliminate abnormal cells, such as cantare ep | { CHAPTER 14 CYTOKINES : f the “ Cytokir ie or glycoprotein mediators that are produced by cells of = immune ral Ser ‘an effect on the behaviour and properties of many cells. T cells are the major source of cytokines, many ‘other cells can produce them. They have a wide range of functions extending beyond the immune system (e.g. wound healing). Many different and overlapping names have been given to the various cytokines: . produced by lymphocytes are often called lymphokines. © Many cytokines are given the name interleukin (IL), followed by a number (¢.9., IL- 1, 2.3....). They were named so because many of these proteins are produced by leukocytes and act on leukocytes. Chemokines are cytokines that are involved in the migration and activation of cells, especially phagocytes. : «Interferons are cytokines capable of inducing but they have other important functions as well. _ More than 100 cytokines have been identified so far. Classifying cytokines is because many are produced by more than one cell type and many have ‘actions. Figure (26) shows a schematic overview of the most important cytokines. They can be classified into two main groups: body cells to resist viral replication, . Cytokines Cytokines that mediate and i Cytokines that mediate and ‘Tegulate innate immunity regulate acquired immunity IFN-a and IFN-B (Type | interferon) ™ The a 2 4 Z IFN-y (type Ht interferon) IL-5, 6, 10 Figure 26: Schematic overview of important cytokines cantare ep 1. Cytokines that Mediate and Regulate Innate Immunity: it it tem and their These cytokines are produced by cells of the innate immune syst actions serve to mediate innate immunity. However, some of them are produ other calls as well and their actions can also affect cells of the adaptive immune system. Type | Interferon (IFN): » Types: — Itincludes IFN-a and IFN-B. Source: o IFN-d is produced mainly by monocytes/macrophages. — IFN-B is produced mainly by fibroblasts. — Many other cell types also produce both. * Stimulus: sete most important stimulus for production of type | IFN is viral infection. lowing actions: ° Actions: 4 Type | IFN is very important in combating viral infections by the fol ‘a. Inhibition of viral replication (Figure 27): Virus infected cells produce type | IFN which inhibits intracellular replication of viruses in neighbouring cells. + Type | IFN induces the neighbouring cells to synthesize a number of enzymes that interfere with the translation of viral mRNA. = A call that has responded to IFN is resistant to any further viral infection (i.e. an interferon induced by one virus can inhibit replication of any other virus). b. Activation of NK cells: This is very important early in the course of infection, before the onset of the adaptive immune response. c. Increased expression of MHC | molecules and consequently there is more efficient killing of virally-infected cells by CD8 Tc cells. 2. Type I IFN also inhibits cell proliferation and tumour growth. MiSetc mca Antiviral Sisking cantare pe ea Interleukin-12 (IL-12): Source: — It is produced mainly by monocytes/macrophages. e Stimulus: — Bacterial constituents « Actions (Figure 28): . 4, IL-12 activates NK cells. 2 Italso stimulates NK cells to secrete IFN-y. 3. It promotes the differentiation ‘of Th cells into Th1 cells. These, in turn, produce IFN-y which activates macrophages. ‘Thus, IL-12 provides an important link between macrophages and NK cells, and also between innate immunity and adaptive immunity. Figure 28: Actions of IL-12 "Il. Cytokines that Mediate and Regulate Acquired Immunity: These cytokines are produced mainly by Th cells. IL-2 and IFN-y are produced by Th1 cells, while IL-4 is produced by Th2 cells (Table 7). Table (7): Comparison between Tht and Th2 cells Thi Th2 5 12 14 ytokines produced IFN-y IL-5,6,10 | [evelopment is promoted IFNy 4 Large doses of antigen _| Small doses of antigen Development is inhibited | !L-4 IFN-y by IL-10 . Promote Cell-mediated immunity | Humoral immunity Interleukin-2 (IL-2): « Source: otis produced by activated Tht cells. . aie proliferation of T cells (T helper and T cytoto? i Is. cytokine production by activated Th cell 3. . ponaed : activates NK cells so that their killing ability is enhanced. an Ne cells activated by IL-2 become lymphokine-activated killer cells (LAK cells). xic) and increases Interferon-y (IFN-y): * [tis called type II interferon or immune interferon. * Source: — Itis produced by Th1 cells and NK cells. e Actions: : \ 4. IFN-y is mainly known as a macrophage activator and is, thus, very important in dealing with intracellular bacteria through the following mechanisms: ‘a. It promotes fusion of phagosomes containing the bacteria to lysosomes containing anti-bacterial substances. b. It induces synthesis of nitric oxide and other bactericidal substances. c. Itinduces macrophages to secrete their cytokines. 2. IFN-y also increases the expression of MHC | molecules, leading to better killing of target cells by Tc cells. 3. IFN-y increases the expression of MHC II molecules on APCs, leading to better presentation of antigens to Th cells. 4. IFNy promotes the development of Tht cells and inhibits the development of Th2 Sie? 5. Italso activates NK cells. Interleukin-4 (IL-4): © Source: ~ Itis produced by Th2, NK cells and mast cells. * Actions: 4; [Khelps activation and growth of B-cells. Promotes production of IgE and growth of mast cell 3. It promotes the development of Th2 cells and init and eosinophils. its the development of Th1 cantare pe ep Therapeutic Uses of Cytokines: : ‘There is considerable interest in the possible use of me cents Mtwever the earanisraton of tokines In therapeutic doses may lead 0 Examples of Therapeutic Uses: ‘Treatment of viral hepatitis. © Treatment for many malignancies; especially lymphomas and leukaemias. a) Interleukin-2: tion to normal or immunodeficient mice enhances ited by severe toxic side effects. ), its administrat immune responses, but its use in humans is lim CHAPTER 15 THE HUMORAL IMMUNE RESPONSE are the destruction of lular pathogens as they The main functions of the humoral immune response by antibodies or extracellular pathogens and the prevention of spread of intracell move from ato all through the extracellular fluids. This is achieved immunoglobulins (Igs). . B Cell Surface Molecules: . | i Certain molecules are expressed on the surface of B cells and are essential for their be activation (Figure 29). |The most important B cell surface molecules are: ‘a+ B cell receptor (BCR): * This is a membrane-bound antibody molecule, also called immunoglobulin that acts as an antigen receptor. * AIlBCRs ona single B cell are of identical specificity and a B cell that binds an antigen through this molecule will then secrete antibodies of the same specificity. b- CD40: * These molecules are essential for the interaction between B and T cells. surface RF c- MHC Il: * These molecules are essential for antigen presentation to T helper cells. Sequence of Events in Activation of Naive B Cells: The mature naive B cell leaves the lymphoid organ (e.9., lymph node). Upon entry, iin the absence of its target antigen, the B cell traverses this region rapidly and re-enters the circulation. | | crear “ot its target antigen. the B cell binds its specific antigen via the surface Igs (BCR). This is considered the first signal needed for B cell activation. To become fully activated, naive B cells must receive a second signal which is delivered by activated helper T cells (mainly Th2) through: ; ‘3 CD40 ligand (CD40L), which is a T cell surface molecule that binds to the B cell surface molecule CD40. | b-IL-4, IL-5 and IL-6, which are B cell stimulatory cytokines. blood stream and enters a secondary the B cell migrates into a region rich in As a result, the B cell becomes a lymphoblast (activation) and divides repeatedly (proliferation). Its progeny (daughter cells) become effector cells, which ro antbedy-secreting plasma cells (differentiation). Some B-lymphocytes do not undergo this terminal differentiation, instead, they become memory cells which are tong-lived. This sequence of events is known as the primary immune response. IgM is first secreted, and then the B cell can switch to the production of another class of the same immunologic specificity. IMMUNOGLOBULINS Antibodies or immunoglobulins (Igs) are glycoproteins which bind speci - 0 \ specifically to p re antigen Set | induced oa a In the blood, most of the inecanesbaiina ore resent e gamma globulin fraction of plasma proteins. i re yh ord teu fds pl a proteins. They are also present in There are 5 classes of Igs, namely, IgG, IgA, IgM, IgE and IgD. Antibody Structure: The basic structure is common to all class es of Igs. An antibody molecule i: ate etes and consists of 2 identical light (L) and 2 identical neavy “e) lypeptide chains linked together by disulphide (S-S) bonds (Figure 30). The light chains: . There are two 5 antibody molecule. types; kappa (k) or lambda (A). Only one type is found in one : nd epsilon (), The heavy chains: dette (8) 8 ‘There are five main types; gamma (y), alpha (a), mu (y)), del ( n coresponding to the 5 classes of Igs; IgG, IA, IgM, IgD and IgE, respectively, The tre heavy chains are joined by a number of S-S bonds in the region known as region. Immunoglobulin Domains (regions): Each light and heavy chain is subdivi « These regions are: 4. Variable regions or domains, which show a wide vari domains, which show a much more uniform (constant) ai ded into domains or regions. jation in amino acid composition. 2. Constant regions or imino acid sequence. «Importance of the immunoglobulin regions: - The variable regions of both the light and heavy chains are responsible for antigen-binding and are called the antigen-binding fragment (Fab). Since the antibody has two identical light chains and two identical heavy chains, each antibody has two identical antigen-binding fragments. ~ The constant region of the heavy chain is responsible for the various biologic functions (e.g., complement activation and binding to cell surface receptors) and is called the Fc fragment. = The constant region of the light chain has no known function. Light chain \ Fe Hinge region {crystallizable fragment) on NNN c —=-\ coon! coon! Constant heavy chain Variable domain JConstant light chain me (antigen-binding fragment) Figure 30: Basic structure of immunoglobull cantare pa ; { { | : (ict eee Functions of Antibodies: There are several ways in which antibodies contribute to immunity: dies to a particulate antigen (e.g. bacteria) results in clumping of the pathogen which prevents its dissemination and stimulates its removal by other mechanisms (@.9- phagocytosis). 2. Neutralization: Antibodies can inhibit the infectivity of a pathogen (viruses or bacteria) or the toxicity of a toxin molecule by binding to them, thereby preventing their attachment to their specific receptors on their target cells. 4. Agglutination: Binding of antibo 3. Opsonization: Phagocytic cells have Fe receptors on their surface that can recognize and bind Fc portion of antibody molecules coating a pathogen. This facilitates the engulfment and subsequent intracellular killing of the pathogen by the phagocytes (Figure 31). Figure 31: Antibody-mediated opsonization 4. Complement activation: Antibodies bound 4 3 to the activate proteins of the complement system (Chapter ca ah aan 5. este cased cai cell-mediated cytotoxicity (ADC): It is the destruction of Fe portion of aii anaes (NK) cells. NK cells possess receptors for the bind to the NK cell rie antibody bound to an antigen on a target cell can also target cell and pee hy fearien > facitating adhesion of the NK cell to the receptors, e.g., macrophages may also ey (rleute 32). Other cells possessing Fc Figure 32: Antibody-dependent cell-mediated cytotoxicity (ADCC) IMMUNOGLOBULIN CLASSES _ Immunoglobulin G (IgG): ‘IgG is composed of a single basic unit (monomer). It is the principal class found in blood (75% of circulating Igs) and extracellular fluids. « it is the only Ig that can pass the placental barrier to the foetal circulation. This — provides passive protection to the newborn during the first few months of life. * Anti-Rh antibodies are of the IgG class. « Biological activities: 1. Neutralization 2. Opsonization 3. Complement activation 4, ADCC I. Immunoglobulin M (IgM): * IgM is composed of 5 basic units (pentamer) held together by di bonds and a single J (joining) chain (Figure 33). y ceeene ee * Because of its large size, IgM is mainly confined to the blood (8-10% of circulating \gs). * Anti-A and Anti-B to ABO blood group antigens of human RBCs are || IgM cannot cross the placenta; ey Daisies inratnine nfecton, es eae ton ma. newbom blood Biological activities: 1, Agglutination 2. Complement activation 3. IgM found on the surface of B-cells forms BCI ensure ea

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