Microbial Classification PDF

Microbial Classiﬁcation Microorganisms divided either to:  Eukaryotes that contain a membrane bound nucleus.  Prokaryotes that contain no nuclear membrane. Di erences between eukaryotic and prokaryotic cells are shown in the following table: Table (1): Comparison between eukaryotic and prokaryotic cells Eukaryotes Prokaryotes Cells with true nucleus , Contain nucleolus, Cells with premature nucleus , No Has nuclear membrane nucleolus , No nuclear membrane Chromosome is more than one, Chromosome is a one ball of double twisted DNA threads The cytoplasmic membrane contains sterol The cytoplasmic membrane does not contain sterol except mycoplasma There is no mesosomes There is mesosomes They have 80S ribosome They have 70S ribosome The respiratory system is localized in The respiratory system is localized in mitochondrion cytoplasmic membrane Multiply by mitosis Mitosis is absent, Multiply by binary ﬁssion e.g. fungi e.g. bacteria & rickettsia PM 0 Morphology Of The Bacterial Cell Bacteria are small unicellular prokaryotic organisms with rigid cell wall that multiply by simple binary ﬁssion. Bacterial Morphology: Staining properties (Gram stain): According to Gram stain, they are divided into 2 groups: 1. Gram positive: bacteria that resist decolorization by alcohol after application of the primary stain and appear violet in colour under the microscope 2. Gram negative: bacteria that decolorization by alcohol after application of the primary stain and take the counter stain (carbol fuchsin) appeared in colour under the microscope (red in colour) Structure of the Bacterial Cell 1. Cell wall Structure: a rigid structure due to peptidoglycan layer. There is a di erences in component between Gram positive and Gram negative bacteria. o Cell wall component in Gram positive bacteria: 1- Peptidoglycan layer: 50-60% of the thickness of the cell wall. 2- Teichoic acid. Antigenic structure o Cell wall component in Gram negative bacteria. 1- Peptidoglycan layer: 5-10% of the thickness of the cell wall 2- Lipoprotein layer OLPLP 3- Outer membrane: found outside lipoprotein layer 4- The lipopolysaccharide layer: endotoxin (lipid A) + polysaccharide O antigen. 5- Periplasmic space Component Gram + Gram - 1. Peptidoglycan Thick Thin 2. Teichoic acid Present Not present 3. Lipopolysaccharide (LPS) (endotoxin) Not present Present 4. Periplasmic space Not Present Present 5. Outer membrane Not present Present 1 Function of the bacterial cell wall: 1. Preservation of the shape of the cell (rigidity). 2. Osmotic in sensitive 3. Di erentiation of Gram positive & Gram negative staining reaction 4. Antigenicity: - Teichoic acid in Gram +ve is major surface Ag. - Lipopolysaccharides in Gram-ve is major surface Ag called "O" Ag. 5. Toxicity: Lipid A of lipopolysaccharides of G-ve is the endotoxin 6. Target for action of antibiotics: as penicillin and cephalosporins. Cell wall defective bacteria Mycoplasma: only bacterial with deﬁcient in the cell wall. So they are polymorphic and not destroyed by penicillin and not stained by Gram stain. Protoplasts, Spheroplasts and L form: bacterial cell wall may be lost under the e ect of certain environmental conditions e.g. treatment with lysosomes and penicillin. if such treated cell placed in osmotically protective media they liberate protoplasts from Gram positive cells and spheroplasts from Gram negative cells. if they are able to grow and divide they are called L forms (can be converted into normal form upon removal of cell wall inhibitor L form may be found during active infection under the e ect of antibiotics (since resist to antibiotic inhibiting cell wall; cause chronic infection). both are involved in secretion of 2. Cytoplasmic membrane (plasma membrane) lays immediatley interior to peptidoglycan layer of cell wall and forms outer boundary of cytoplasm proteins and active transport and are site of respiration. Structure: It is semi-permeable double layered structure, composed of phospholipid and protein. Function: 4. excretion of hydrolytic enzymes 1. Selective permeability & Active transport. of ions as K, NA and nutrients to degrade nutrients to subunits. 5. biosynthetic function (site if synthesis of cell wall 2. Energy production, site of respiration. Transport of electroms for oxidative phosphorylation precursors amd certain proteins). 6. chemotactic function (contain 3. Excretion of pathogenicity proteins and toxins e.g. IgA protease. receptors pf binding and repellents). Mesosome: irregular convoluted invagination of the cytoplasmic membrane into the cytoplasmic Function: 1. Septal mesosomes are attached to chromosome and involved in cell division 2. Involved in secretion of proteins and active transport 3. The sites of the respiratory enzymes 2 4. Lateral mesosomes: increase the total surface of the membrane II. Cytoplasmic components: 1. Nuclear body: DNA is concentrated in the cytoplasm as a nucleoid, no nuclear membrane or one double stranded circular DNA molecule, nucleolus. carrying the genetic characters of the cell. 2. Plasmid: they are extra chromosomal DNA molecule capable of replication independently of the bacterial chromosome hold non essential DNA 3. Ribosome: (70 S), 30S and 50S subunit. Function: i. protein synthesis ii. target of some antibiotics as tetracycline & chloramphenicol III. Extracellular structures and appendages ALL 3 ARE ANTIGENIC 1. Capsule: it is formed in-vivo. Composed of polysaccharides except bacillus anthracis composed of It is a geletinous layer of high molecular polypeptide.weight surround many bacteria. It is produced by the cell wall and adgeres to its surface. Function: 1. Virulence factor protect against phagocytosis. 2. Antigenic: identiﬁcation and typing of some bacteria 3. Adherence of bacteria to human tissues 4. Capsular polysaccharides are used in some vaccines as pneumococcal, meningococcal and H. inﬂuenza vaccine. 2. Flagella: Filamentous appendages that move the bacteria toward nutrients and other attractants (organ of motility). formed of protein ﬂagellin which is antigenic. attached to cell wall and cytoplasmic membrane by basal bodies Types of ﬂagella: 4 types according to arrangement: 1. mono-trichate, single, at one pole 2. amphi-trichate, two, one at each pole 3. lopho-trichate, group of ﬂagella, at one or both poles 4. peri-trichate, all around the surface Function: It responsible for motility of the organism. 1. Important in pathogenesis, by moving the bacteria 2. They are antigenic ( H Ag ), useful in bacterial identiﬁcation. 3- Pilli (ﬁmberiae): Short hair like ﬁne surface ﬁlamentous appendages, they are Shorter and thinner than ﬂagellae. They are formed of protein, found mainly in Gram negative. they originate from the cell membrane pilin 3 There are 2 types : 1- Ordinary or common pili, they are antigenic, have a role in adhesion and are virulence factors. 2- Sex pili (fertility pili), longer thicker than ordinary pili. Has a role in conjugation. Bacterial Endospore Deﬁnition: highly resistant resting forms developed by certain gram +ve bacilli as bacillus and clostridium when there are unfavorable environmental conditions for their growth as depletion of nutrients, heat, dryness..etc DNA+part of the cytoplasm bacterial membrane invaginates inwards Sporulation: (mechanism of spore formation): The nuclear material + core+ essential enzymes + thick cortex (thick peptidoglycan) + the spore coat (tough keratin like protein) Spore has no metabolic activity and can remain dormant for years. Germination (vegetation): the conversion of spore into vegetative cell when the environmental conditions become favorable for growth. Medical importance of spores: 1- They are resistant to heating (killed at 121oc) so -------- autoclaved. 2- Highly resistant to chemical and disinfectant due to thick coat of spore (need sporecidal) 3- Can survive for many years in soil. Wound contamination can be infected with tetanus. Marked resistant due to: 1. Thick spore cortex and taught spore coat. 2. Low water content so it resists dryness 3- The spore has a rigid impermeable wall rich in dipicolinic acid and calcium. 4- Low metabolic and enzymatic activity. Bacterial physiology Bacterial growth requirements bacterial growth = Increase in the cell mass Nutrition: 1. Autotrophic bacteria: utilize inorganic sources of carbon (CO2) and nitrogen (ammonium). These are usually free living, non parasitic organisms of no medical importance. independent, only depend on themselves 2. Heterotrophic bacteria: require organic sources of carbon and nitrogen as sugar and protein. i.e. pathogenic bacteria Gaseous requirements I. Oxygen: 8 1. Strict aerobic: grow only in the presence of oxygen; O2 is the only H2 acceptor ex. TB *contain catalase enzyme, hydrolysis of H2O2. Aerobic bacteria breakdown H2O2 by catalase enzyme and O3 superoxide dismutase enzyme. 2. Obligate Anaerobic: growing only in the absence of O2 e.g. clostridium In presence of O2 two toxic molecules are formed; hydrogen peroxide H2O2 and superoxide radical O3 they are toxic to bacteria. Anaerobic bacteria have no catalase or superoxide dismutase enzyme i.e the presence of O2 ® kills the organism 3. Facultative anaerobic: growing in the presence as well as in absence of O2 *they contain catalase and other H2 acceptors e.g most of pathogenic bacteria 4. Microaerophilic: need very small concentration of O2, contain small amount of catalase enzyme e.g. P acne II. Carbon dioxide: Normal atmospheric conc of CO2 (0.03%) ® is su icient for growth of many bacteria. High CO2 conc (up to 10%) may needed Factors a ect metabolic activity of bacteria. Temperature: *Minimum temperature (10 °C). *Maximum temperature (42°C). *Optimal temperature (37°C). Bacterial Products 1. Bacterial pigments: Endopigment & Exopigment Endopigment Exopigment localized in the bacteria Di uses outside the bacteria Colour the bacterial colonies Colour the bacterial colonies & the surrounding medium e.g. staphylococcus aureus golden e.g. pseudomonas aeruginosa greenish blue yellow colonies Best developed at room temperature They have a role in bacterial respiration 9 2. Bacterial toxins: Types: exotoxins and endotoxins Table 3: Comparison between bacterial exotoxins and endotoxins Exotoxins Endotoxins cell wall 1. Di usibility Di usible toxins Bound to the body and released only bacteria will when the organism disintegrate explode & stuﬀ will be released by cell wall 2. Nature Protein Lipopolysaccharide lipid A 4. Organism producing some Gram +ve &-ve Gram-ve bacteria bcz toxin is part of the cell wall structure in gram negative bacteria only 5. Toxicity Highly toxic Less toxic 6. Antigenicity Strong antigenic stimulates immune system, so can be used in vaccines Weak antigenic wont stimulate immune system so wont be able to be converted into vaccine 7. Speciﬁcity Speciﬁc in action on cells & Non speciﬁc tissues 8. E ect of heat Destroyed bcz its protein Stable bcz its lipid (60-80 °C) 9. E ect of formalin Detoxicated : change into formol Not detoxicated bcz loss of its toxic toxoid used as a vaccine factor and preservation of its antigenic structure 10 Bacterial Growth & Reproduction Bacterial Growth : increases in number. Bacteria divide by asexual simple binary ﬁssion i.e Bacterial Growth Curve A. Lag phase: stage of preparation for multiplication during which the organism adapt itself by synthesis of new enzymes speciﬁc for the new medium. This stage is characterized by:  No increase in the number of bacteria and there may be a slight decrease due to death of some inoculated bacteria.  Little or no cell division (the bacteria prepare themselves for active cell division). marked increase in metabolic activity and susceptibility to physical and checmical agents. it varies from few hours in E. coli to several weeks in TB depending on: 1. Type of organism (short in E. Coli and long in T.B). 2. Size of the inoculum (the bigger the inoculum, the shorter the lag phase). 3. Stage from which the bacteria are grown (if taken from logarithmic phase the lag phase will be very bcz it has already divided short). if from death phase its already dead. 4. The more suitable the medium, the shorter the lag phase. This phase corresponds in natural infection (in vivo) to the incubation period of disease. B. Logarithmic phase (Exponential phase): rapid multiplication, regularly increase by time. antibiotics are e ective during this phase, as B lactam. In vivo it corresponds to the invasion period of the disease. it continues till 1 or more nutrients in medium become B lactam drug acts when the cell is making peptidoglycan exhausted(done) or the toxic metabolic products accumilate C. Stationary phase: the rate of division decreases and the rate of death increase, this is due to accumulation of metabolic products and O2 starvation. The number of bacteria divide is equal to the number of bacteria died (the number remains stationary). In vivo it corresponds to the period of clinical signs and symptoms of the disease. D. Decline phase: the death of bacteria gradually increases, at the end the bacteria are completely died; due to accumulation of toxic waste products & release of lytic enzymes and exhaustion of nutrients and does bacterial lysis (breaking) O2. It corresponds to the convalescence stage of the disease. time spent recovering from an illness or medical treatment 11 Pathogenesis of bacterial infection The pathogenicity: is the ability of organism to cause disease. Virulence: Is the degree of pathogenicity of the microbe. According to natural habitat and relation to the host, bacteria are divided into: 1- Saprophytic bacteria that grow on dead tissue. free on nature as soil 2- Parasitic bacteria that live in or on host tissue. They are divided into: a- Commensal organisms that live in a balance with the host, does not cause disease (Non pathogen) e.g: normal ﬂora of human body. b- Opportunistic pathogen under certain conditions microbiota (some of normal ﬂora) cause disease, some of this conditions are: 1- Impaired host defense mechanisms (i.e. when the patient is "immunocompromised"). impaired system immune 2- Alteration of the host tissues, e.g Strept veridance (normal inhabitant in the mouth) caused endocarditis when reach blood stream after tooth extraction in rheumatic heart patients. inflammation of the inside lining of the heart chambers and heart valves 3- Change in the natural habitat of the organisms, e.g if E. coli leave intestine and reach to urinary tract. c- Pathogenic organisms: can cause disease in previously healthy individual with intact immunological defenses. Infection and Diseases Infection is multiplication of an infectious agent within the body. may be inapparent or asymptomatic. pathogenic infectious bacteria Disease the development of signs and symptoms of disease. suﬃcient harm (enough). A cycle of transmission from a source of infection, through a portal of entry, into a susceptible host and on again I- Sources of infection: (1) Human (Patient, Carrier): - Carrier is a apparently healthy person that carry pathogenic organism in his body, secrete it to outside without any signs or symptoms. Carriers are more dangerous than patient because: a. They don`t show manifestation of the disease. b. They contact easily with other persons. c. They are not easily detected, not isolated, not treated. (2) Animals (zoonotic infections). transmission of microbes from animals to humans. (3) Inanimate sources (soil, water, air). not living. II- Root of transmission: (1) Direct transmission: - Direct respiratory spread via large droplets. 12 - Fecal-oral spread. - Sexual transmission. (2) Vector-borne transmission: Is mediated by arthropods or insects. mechanical OR biological (3) Airborne transmission: Mediated by aerosols suspended in the air for long periods. (4) Zoonosis: Any infection spread from a vertebrate animal to a human. III- Portals of entry of pathogenic bacteria into the body: skin, respiratory, gastrointestinal, genital, and urinary tracts. , abnormal areas of mucous membranes and skin (eg. cuts, burns, and other injuries). IV-Multiplication of the parasite within the host: either locally at site of entry or may be spread through tissues, blood, lymphatic to reach other sites. V- Portal of exit from the host: e.g urine, stool, respiratory or genital discharge, or from blood by injections or insects. Host Parasites Relationships The relationship between a host and infectious agents may take one of 4 forms: 1- Colonization: colonizes the host tissue without causing any harmful e ect. e.g commensal bacteria in oral cavity. bacteria just sits in host, doesnt cause disease 2- Infection: invades the host tissues + elicits immune response but causes minor tissue damage so that no clinical signs appears (subclinical infection). 3- Infectious disease: invades the host tissues + elicits immune response and causes marked tissue damage so that clinical signs and symptoms appears (clinical infection). 4- Carrier multiplication but no symptoms show Factors a ecting host parasites relationships 1- Factors related to the host: Natural and acquired immunity. 2- Factors related to the microorganism: Pathogenicity &Virulence. the more pathogenic the more it spreads Virulence factors include: (1) Mechanisms for colonization (adherence and initial multiplication): - Adhesion to cells of tissue surface by speciﬁc surface structures called bacterial adhesins as pilli. (2) Invasion of host cells & tissues:.e.g Collagenase: breaks down collagen. (3) Toxin production: exotoxins and endotoxins. Bacterial toxins may be transported by blood and lymph. (4) Ability to bypass or overcome host defense mechanisms: a – Antiphagocytic Factors: e.g Polysaccharide capsules & Pili of Neisseria gonorrhea. b- Intracellular Pathogenicity: Some bacteria (e.g. M. tuberculosis) live and grow in the phagocytic cells. c- IgA Proteases: that split IgA. 13 Antimicrobial chemotherapy act on microbes without aﬀecting its host, by selective toxicity(toxic to microbes but not toxic to human cells by attacking the bacteria while having a target not present in humans - Bactericidal drugs have a rapid killing action of bacteria, which is irreversible. Examples include penicillins, cephalosporins and aminoglycosides. act on cell wall - Bacteriostatic drugs merely inhibit bacterial multiplication, but do not kill them. The bacteria can grow again when the drug is withdrawn. In this case, host defence mechanisms, such as phagocytosis, are required to kill bacteria. Examples include sulphonamides, tetracyclines and act on ribosomes chloramphenicol. act on ribosome - Spectrum of Action of Chemotherapeutics: - Broad-spectrum antibiotics; active against several types of microorganisms, both gram positive and gram negative e.g. tetracyclines, chloramphenicol and ampicillin. - Narrow-spectrum antibiotics are active against one or very few types, e.g. vancomycin is primarily used against certain gram positive cocci i.e. staphylococci and enterococci. Mechanisms of action of antimicrobials: - An ideal antimicrobial agent should have selective toxicity, i.e. it can kill or inhibit the growth of a microorganism in concentrations that are not harmful to the cells of the host. - Several mechanisms are known: 1-Inhibition of Bacterial Cell Wall Synthesis: Due to its unique structure and function, the bacterial cell wall is an ideal point of attack by selective toxic agents e.g. Penicillin, cephalosporins and vancomycin, interfere with cell wall synthesis by inhibit peptidoglycan synthesis. β-lactams e.g. penicillin and cephalosporins, and Vancomycin 2- Inhibition of Bacterial Cytoplasmic Membrane Functions: cause disruption of the cytoplasmic membrane and leakage of cellular proteins and nucleotides leading to cell death e.g. Polymyxins, amphotericin B, and nystatin (These drugs are highly toxic as they have a narrow margin of selective bcz no specific target toxicity). 3-Inhibition of Bacterial Protein Synthesis: Several drugs inhibit protein synthesis in bacteria without signiﬁcantly interfering with protein synthesis in human cells. This selectivity is due to the di erences between bacterial and human ribosomal proteins, RNA, and associated enzymes. Bacteria have 70S ribosomes (with 50S and 30S subunits), whereas human cells have 80S ribosomes (with 60S and 40S subunits) e.g. Chloramphenicol, erythromycin, linezolid and streptogramins (quinupristin / dalfopristin) act on 50S subunits, while tetracycline and aminoglycosides (gentamicin and arnikin) act on 30S subunits. 4-ln hibition of Bacterial Nucleic Acid Synthesis: These can act on any of the steps of DNA or RNA replication  Quinolones; inhibit DNA synthesis by blocking DNA gyrase,  Nitrofurantoin; Act through damaging bacterial DNA  Rifampicin; inhibits RNA synthesis by binding to RNA polymerase &  Trimethoprim and sulfonamides; inhibit nucleotide synthesis. 14 5-Competitive Inhibition: In which the chemotherapeutic agent competes with an essential metabolite for the same enzyme. Para-aminobenzoic acid (PABA) is an essential metabolite for many organisms. They use it as a precursor in folic acid synthesis which is essential for nucleic acid synthesis. Sulphonamides: similar to PAPA essential for folic acid synthesis (competitive inhibition) Trimethoprim: inhibit dihydropholic acid reductase.. Dihydrofolic acid and tetrahdrofolic acid… (purine synthesis) -prevents infection. -not a treatment. -used as soon after aquisition of microbe and before symptoms appear. -even before contact of infection we take it. Antimicrobial prophylaxis -prophlaxis can also reduce the risk of endogenous infection associated with certain surgical and dental procedures. 1- Prophylaxis in persons of normal susceptibility exposed to speciﬁc pathogen e.g. Prophylaxis from Rheumatic fever by long acting Penicillin. Prophylaxis from meningitis by Rifampicin. 2- Prophylaxis in persons of increased susceptibility e.g.: - Heart diseases. - Respiratory diseases (Chronic). - Recurrent urinary tract infections. - Immunosuppressed host. 3- Surgical prophylaxis In dentistry, prophylactic antibiotics before dental or surgical treatment of patients who: At risk of infective endocarditis Immunocompromised Recently received radiotherapy to the jaws. Prosthetic hip replacements, insertion of implants or bone grafting. Resistance to Antimicrobial Agents The mechanisms by which the organism develops resistance may be one of the following: 1. Inactivating enzyme production: e.g. (penicillinases enzymes and B lactamase destroy the penicillin) made up of B lactame 2. Alteration of permeability cell membrane to the drug e.g. microorganisms change their permeability to the drug. 3. Alteration of target (receptor) for the drug e.g. microorganisms develop altered structural target for the drug. 4. Alteration of metabolic pathway: e.g. microorganisms develop altered metabolic pathway that bypass the reactions inhibited by the drug. e.g. some bacteria not require PABA but utilize performed folic acid. so uses alternative pathway instead of sulphonomides that compete with PABA 5. Alteration of enzyme: e.g. microorganisms develop altered enzyme that can still perform its metabolic function but is much less a ected by the drug. Antimicrobial combinations Advantages: 1. Serious infection e.g. Peritonitis , meningitis 2. Mixed or unknown infection.e.g. Polytraumatized patients 15 3. Chronic infection or prolonged treatment.e.g. T.B 4. Prevention or delay of drug resistance.e.g. T.B Disadvantages: 1- Cost is high. 2. Increased incidence of drug reaction 3. Drug antagonism. 4. Increased incidence of Super infection like fungal infections by killing of normal flora combination of 2 drugs has much greater eﬀect Mechanisms of Drug synergism: than 1 drug on its own 1- Sequential block of a microbial metabolic pathway by the 2 drugs. combining sulfonamides and trimethoprim blocks two steps in the folic acid synthesis 2- One drug may enhance the uptake of the second drug; one drug may affect cell membrane and facilitate the entry of the second drug. Aminoglycoside + penicillin 3- Drug combination may inhibit the bacterial enzymes that destroy the one drug. combination of amoxicillin with clavulanic acid. Clavulanic acid inhibits β-lactamase enzymes produced by some bacteria, preventing them from breaking down amoxicillin. This allows amoxicillin to remain active and effective against β-lactamase-producing bacteria. Microbial Ecology of the Oral Cavity Normal Flora of the Body Definition: Normal flora includes microorganisms (bacteria, fungi, and protozoa) living in harmony on or within animal and plant bodies, generally without causing harm to the host. Symbiosis: o Definition: Symbiosis is the intimate interaction between two organisms. o Types of Symbiotic Interactions: 1. Commensalism: One organism benefits, and the other remains unaffected (e.g., common resident flora on the human body that doesn’t cause disease). 2. Mutualism: Both organisms benefit (e.g., gut bacteria synthesize vitamin K for humans while using available nutrients). 3. Parasitism: One organism benefits at the host’s expense, causing harm (e.g., pathogenic bacteria and viruses). Benefits of Normal flora 1. Synthesize and excrete vitamins:relating Enteric bacteria secrete Vitamin K and Vitamin B12 to intestines 2. Prevent colonization by pathogens: Compete for attachment /Essential nutrients by attaching to receptor 3. Antagonize other bacteria: Production of inhibitory substances which inhibit or kill other species by producing toxins to kill bacteria (Peroxides and Bacteriocins) 4. Stimulate the development of immune tissues. 16 5. Stimulate the production of cross-reactive antibodies. 2 antigens attach to 1 antibody 6. Maintain inhibitory pH in vagina and skin. by acidity Harmful Effects of Normal Flora Harmonious friendly relationship of microflora with the host can be changed and cause disease. 1. Opportunistic Pathogenicity: Normal flora can become pathogenic a- Overgrow of normal flora as in immunocompromised individuals naturally occuring fungus in our body b- Depletion (reduction) of normal bowel flora following antibiotic therapy (e.g., over growth of Candida oral normal intestinal flora thrush). white yellow tongue 2. Displacement (e.g., oral flora causing bacterial endocarditis after tooth extraction). bacterial infection 3. Conversion of Food to Carcinogens: Certain bacteria in the colon can turn ingested substances into carcinogenic compounds can cause cancer Oral Diseases have Impact on General Health spreading far 1. Metastatic Infection: Microbes enter into the blood stream (e.g infective endocarditis). circulatory system 2. Metastatic Injury: Products of bacteria, such as exotoxins and endotoxins. 3. Metastatic Inflammation: Caused by immunological injury due to oral organisms. E.g. Soluble antigens react with circulating specific antibodies. Examples of Normal flora in Mouth Anaerobes: Veillonella Spirochetes: Treponema species Gram-positive Cocci and Bacilli: Streptococci, Actinomycetes, Lactobacilli Gram-negative Cocci and Bacilli (HACEK group): Haemophilus, Actinomycetemcomitans Protozoa: Entamoeba gingivalis Yeasts: Candida spp., especially C. albicans Pathogenic Reservoir: Helicobacter pylori can sometimes be found in dental plaque, associated with gastric issues. Mouth acts as a reservoir for pathogenic organisms: The Mouth as a Microbial Habitat Ecological Terminology Habitat: The specific site where microorganisms grow. Microbial Community: The population of microorganisms in a habitat. Resident Microflora: Organisms regularly found at a specific site. Biofilm: Microbial communities growing on surfaces. Ecosystem: The microbial community and its surroundings, both biotic and abiotic. 17 Four Distinct Features of the Oral Cavity: The oral cavity is a delicate balance between allowing microbial colonization and activating defense mechanisms. Each of the four features—specialized mucosal surfaces, teeth, saliva, and gingival crevicular fluid—provides an environment where microorganisms can thrive, but also plays an essential role in the defense against infection. Effective oral hygiene and a healthy oral environment are vital in maintaining this balance to prevent the overgrowth of harmful microbes while ensuring the proper functioning of the immune defenses in the mouth. 1. Specialized Mucosal Surfaces (Lips, Cheeks, Palate, Tongue):  Allowing Microbial Colonization: The lips, cheeks, palate, and tongue provide a high surface area, creating ideal environments for the colonization of both aerobic and anaerobic microorganisms. These areas, due to constant exposure to food and liquids, facilitate the establishment of microbial communities, including potential pathogens.  Defense Mechanisms: While these surfaces promote colonization, they also possess natural barriers such as mucosal immunity, which includes secretory IgA, to help limit the overgrowth of harmful microbes and prevent infection. 2. Teeth: Allowing Microbial Infection: Teeth are non-shedding surfaces, meaning they provide a stable habitat for biofilms, which can include harmful bacteria. These biofilms are especially prevalent in areas such as fissures, smooth surfaces, and gingival crevices, where food particles and plaque accumulate, creating a niche for microbial growth.  Defense Mechanisms: The saliva and gingival crevicular fluid (GCF) play a key role in controlling microbial colonization on teeth. The antimicrobial agents in saliva, such as lysozyme and sialoperoxidase, contribute to microbial control. antimicrobial eﬀect 3. Saliva Allowing Microbial Infection: Saliva provides a rich medium for microorganisms, offering nutrients that can support the growth of both commensal and potentially pathogenic bacteria. While it is an essential part of oral health, an imbalance (e.g., reduced saliva flow) can create conditions conducive to infections such as candidiasis or caries. antibodies Defense Mechanisms: Saliva contains various antimicrobial factors, including secretory IgA, sialoperoxidase, lysozyme, and antimicrobial peptides, which act to inhibit microbial growth. It also acts as a buffer, maintaining a neutral pH to protect oral tissues and neutralize acids produced by bacteria that could lead to infection or damage, like dental caries or periodontal disease. 1. Gingival Crevicular Fluid (GCF) Allowing Microbial Infection: GCF, particularly in the gingival crevices, can promote microbial growth if the oral hygiene is poor, especially in the presence of inflammation. Pathogenic microorganisms can invade these areas, leading to gingivitis and periodontitis. Defense Mechanisms: GCF acts as an immune defense fluid, containing components such as IgG, neutrophils, complement proteins, and enzymes. These help combat microbial invasion, especially during periods of increased bacterial activity. During infection or inflammation, GCF volume increases, facilitating a stronger immune response to protect periodontal tissues. 18 Factors Affecting Microbial Growth in the Oral Cavity Temperature, pH, nutrients, host defenses, genetics, and antimicrobial agent Temperature: The oral cavity provides a stable, warm environment around 37°C, ideal for microbial growth. However, slight temperature fluctuations can occur, especially with the intake of hot or cold food and beverages, affecting the metabolic activity and growth rate of oral microbes. Ph : The oral cavity typically maintains a slightly acidic to neutral pH range (6.5–7.5), which is favorable for most resident microbes. Saliva plays a critical role in buffering pH, but diet (e.g., acidic foods) and microbial metabolism (e.g., lactic acid production by bacteria) can temporarily alter pH, influencing microbial survival and the risk of dental caries. Nutrients: available in the oral cavity, such as dietary sugars, proteins, and glycoproteins from saliva, support microbial growth. Resident microbes utilize these nutrients for energy and to sustain biofilm communities, while oral hygiene practices and diet influence nutrient availability and microbial composition.  Host Defenses: Host defense mechanisms include the innate and adaptive immune components present in saliva and gingival crevicular fluid. Saliva contains antimicrobial factors (e.g., lysozyme, lactoferrin, and secretory IgA) that target and neutralize microbes. Additionally, GCF has leukocytes and antibodies that help combat pathogenic invasion, limiting microbial overgrowth and infection. Genetics: Individual genetic variations influence the composition and resilience of the oral microbiome. Genetic differences affect immune responses, saliva composition, and even structural features of oral surfaces, all of which shape microbial colonization and growth patterns unique to each person.  Antimicrobial Agents: Regular exposure to antimicrobial agents, such as in mouthwashes and toothpastes, can inhibit the growth of certain microbes or shift the microbial balance. Antibiotics can also impact oral microbial communities, reducing the number of susceptible bacteria while potentially allowing resistant organisms or opportunistic pathogens to thrive. 19 General Properties of viruses Viruses are the smallest infectious agents known (20 -300 nm diameter). So pass through bacterial filters, need bcz they are smaller than bacteria high speed of centrifugation for sedimentation and seen by electron microscope. unlike bacteria, they cant be seen by LM high speed bcz viruses are too small are gravity seperation would take too long and the seperation force must be increased - Doesn't contain nucleus, ribosomes, and mitochondria. so wont produce proteins -single stranded DNA not found in bacteria -double stranded DNA - Contains one type of nucleic acid (RNA or DNA). bcz too small -single standed RNA -double stranded RNA - Metabolically inert (no ribosomes which synthesize protein), require host for multiplication and survival. so cant live alone - Obligate intracellular parasites, only replicate inside living cells. - They can infect man, animals, insects, plants and bacteria. - Not susceptible to the action of antibiotics. Viral Structure Virion (virus particle) is composed of: 1. Nucleic Acid (genome or core): DNA or RNA, single or double stranded, linear or circular. The DNA is always a single molecule, while the RNA can exist either in a single molecule or in several pieces. Functions: protected by 1- Nucleic acid caries the genetic information of the virus like replication, antigenicity and virulence. severity or harmfulness 2- It is the infectious part of the virus, coreless particles are noninfectious. of a disease 2. Capsid: Surrounds viral nucleic acid (capsid + its nucleic acid called nucleocapsid), made up of subunits called capsomers, each capsomer consist of one or several proteins. Functions: a. Protection of viral genome against inactivation by nuclease enzymes. protected by b. Gives the virus its antigenicity. can stimulate the immune system c. Gives the virus its geometrical symmetry. /shape d. Adsorption (attachment) of virions to susceptible cells. attachment to organism 3. Envelope: Some viruses have additional coat outer to capsid, others don't have, which composed of lipid or lipoprotein in nature (lipid fraction derived from host cell membranes & protein fraction is virus specific) may be covered by glycoproteins (spike like projections) on the surface. attach to the host cell receptors and antigenic and determine virus specificity Functions: 1- Additional protection of the virus 2- Share in viral symmetry 3- Viral antigenicity 4- Pathogenesis and replication (attach to the host cell receptors and determine virus specificity). 20 4-Viral enzymes: Some viruses carry enzymes like RNA polymerase in orthomyxoviruses and reverse transcriptase in retroviruses. Functions: Important for viral replication and survival. Viral Classification Viruses were classified according to nucleic acid into: 1- DNA viruses 2- RNA viruses Viruses were classified according to envelope into: 1- Enveloped viruses 2- Non Enveloped viruses Viruses were classified according to symmetry into: 1- Icosahedral symmetry: in which the capsomers arranged in 20 triangles that form a symmetric figure (an icosahedron) like crystal. e.g. Herpes viruses and adenoviruses. multisurface 2- Helical symmetry: in which the capsomers are arranged in hollow coil that appears rod-shaped or helix, all human helical viruses are enveloped. e.g. Influenza viruses. capsomeres rap around the DNA 3- Complex symmetry: the virus is complicated in structures. e.g. Bacteriophage. Viral Replication Viruses are obligatory intracellular with no metabolic activity, so they depend on living host cells for providing the viral parts by using the machinery of host cells, the viral genome provides the host with the genetic information needed for its replication. The viruses replicate in the following steps: 1- Adsorption: It is the attachment of the virus to the host cell, it is receptor specific, this explain viral tropism (each virus is specific for certain cells). 2- Penetration a- In non-enveloped viruses penetration occurs by crossing the plasma membrane directly or by receptor mediated endocytosis. b- In enveloped viruses penetration occurs by fusion of viral envelope with cell membrane at the cell surface. only the nucleocapsid part enters 3- Uncoating: It is the release of viral nucleic acid by cellular enzymes. Uncoating may occurs at cell surface removing the capsid so that only the genetic like bacteriophage, at cytoplasm like poliovirus or at the nucleus like herpes viruses. material is left alone. which renders the viral nucleic acid accessible for transcription and translation 4- Eclipse: The virus cannot be detected for certain time in the cell (viral nucleic acid incorporated into cellular bcz the virus genetic material is too small nucleic acid), it is the eclipse phase. 21 5- Viral nucleic acid synthesis: Uncoating renders viral nucleic acid accessible for transcription and replication (viral gene expression known as transcription and protein synthesis is the translation) by using a strand of the nucleic acid as a template for the production of progeny DNA or RNA molecules. 6- Assembly: Assembly of viral nucleic acid and protein coats to form mature virus particles. 7- Release: Virus particles are released from the cell either by: all the things produced should be gathered together inside host to a- Enveloped viruses, budding through the outer cell membrane assemble , EXCEPT the envelope is left at the door to be taken by the virus while the virus is budding out of the host cell b- Unenveloped viruses, rupture of the cell membrane and release of the mature particles. Atypical virus-like agents 1- Defective virus particles: Definition: Virus particles that cannot multiply by themselves but can multiply in cells simultaneously infected with infectious helper virus. Structure: They contain normal structure capsid proteins and only part of a viral genome so; helper virus provides the missing function. 2- Pseudovirions: the shape from the outside is like a virus but from the inside it isnt a virus Contain host cell DNA instead of viral DNA within the capsid, it occurs with certain viruses when the host DNA is fragmented, and pieces are incorporated within the capsid. Pseudovirions can infect cells but cannot replicate. 3- Prions Are infectious particles that are composed of protein only without nucleic acid, they cause slow diseases. Pathogenesis of viral infections The virus needs a susceptible cell (cells contain receptors for the virus). Each virus has a chain of infection which consist of the following: 1- Source of infection 2- Portal of exit from the source 3- Mode of transmission 4- Portal of entry to the recipient 5- Pathogenesis inside the recipient 6- Portal of exit from the recipient (who become source again) Incubation period: It is the period between the infection and the appearance of symptoms which range from few hours (influenza virus) to years (HIV virus). Viruses may produce local or systemic infections 22 - Local infections: Which occurs at the portal of entry virus without viraemia e.g. influenza virus at the mucous not present in blood membrane of the respiratory tract and rotavirus at GIT causing diarrhea. - Systemic infections: After multiplication at the portal of entry the virus spread through blood, lymphatics or nerves to cause systemic infections e.g. polio, mumps, hepatitis and rabies. Viral infections may be transient or persistent - Transient viral infections: Virus was removed after infection with remaining the immunological memory - Persistent viral infections: The virus persists for long time in the host in one of the following forms: 1- Chronic infection: The virus can be continuously detected with mild or no symptoms e.g. HCV and HBV. when we take a blood sample the virus will be detected, its contagous 2- Latent infection: The virus persists hidden (latent) most of the time, viral genome present inside host cell with cant be detected no production of progeny virus. Reactivation of latent viruses can occur after months or years leads to production when immunity decreases of progeny virus andsigns initiate the disease e.g. herpes virus and symptoms appear 3- Slow virus infections: These have a very long incubation period, months or years without clinical symptoms e.g. subacute sclerosing panencephalitis with incubation period 2-20 years and is caused by variants of measles can be detected after 2-20years virus and AIDS, other infections caused by prions like mad cow disease in cattle. HIV Reaction To Physical And Chemical Agents Heat and Cold Heat Treatment: 50c / 30 minutes disrupt viral structures and stop infectivity. Exceptions hepatitis B virus need higher temperatures. Cold Storage and Virus Preservation: Cold low) subfreezing (conditions don’t harm most viruses and help preserve them for longer periods.thats why vaccines should be in cold containers so that the virus can be transmitted Lyophilization (Freeze-Drying): dried out and stored in a stable form at 4°C or even at room temperature. These viruses, when in their dry, lyophilized form, more heat- so 50C wont aﬀect it resistant than they would be in their normal, wet state. Ether Susceptibility: Enveloped ethyl alcohol viruses “inactivating” the virus. Non-enveloped (or denatures the virus which is imp for attachment of virus so killing of virus "naked") not loss infectivity Detergents: Nonionic and anionic detergents solubilize viral envelopes; in addition, they disrupt capsids into separated polypeptides. Formaldehyde: Formaldehyde destroys viral infectivity by reacting with nucleic acid. Formaldehyde has minimal adverse e ects on the antigenicity of proteins and therefore has been used frequently in the production of inactivated viral vaccines. 23

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