General Microbiology for Dentistry Students PDF

General microbiology for dentistry students Dr. Haneya Ali Anani Prof. of Medical Microbiology and Immunology Faculty of Medicine  Lecture 1 Bacterial cell morphology On completion of this lecture, the student will be able...

General microbiology for dentistry students Dr. Haneya Ali Anani Prof. of Medical Microbiology and Immunology Faculty of Medicine  Lecture 1 Bacterial cell morphology On completion of this lecture, the student will be able to: 1. To know the classification and nomenclature of bacteria 2. To know morphology and structure of bacterial cell  Differences between Prokaryotes /Eukaryotes Prokaryotic cell Eukaryotic cell Reproduction They reproduce asexually by They reproduce sexually or a simple binary fission sexually Respiration Through cytoplasmic Mitochondria membrane Nucleolus absent present Nuclear membrane absent Present Chromosome Single chromosome Pairs of chromosomes Circular Linear Location of DNA In cytoplasm In nucleus Cytoplasmic Bacterial cell membrane does Contains sterols membrane not contain sterols except mycoplasma Mitochondria Absent Present Ribosomes Prokaryotic ribosomes 70S eukaryotic ribosomes 80S Examples Bacteria and Rickettsia Human cell, Animal cell, Fungi, Protozoa Differences between prokaryote and eukaryote Classification and Nomenclature of Bacteria Binomial nomenclature The scientific naming of microorganisms. Bacteria are named binomially by incorporating the genus and the species Staphylococcus (Genus) aureus ( species)  Morphological characters of bacteria ▪ Size ▪ Cell shape and arrangement ▪ Gram stain ▪ Growth temperature ▪ Sporulation ▪ Motility ▪ Capsule formation  Cell Morphology Size : bacteria are unicellular microorganisms measured in micrometer Shape and arrangement Arrangement  Structure of the bacterial cell The essential components of a bacterial cell ▪ Cell wall, ▪ Cytoplasmic membrane ▪ Cytoplasm containing (ribosomes, plasmids, metabolic granules) ▪ Nuclear material Surface structures Capsules Flagellae Pili. Endospores External structure of the bacterial cell  Cell wall According to cell wall, bacteria divided into 1. Gram-positive cell wall 2. Gram-negative cell wall 3. Acid-fast bacilli cell wall 4. Cell wall deficient forms Gram positive versus Gram negative bacteria  Lipopolysaccharide The lipopolysaccaride of gram-negative cell walls is a union of lipid with sugar. The lipid portion of LPS is known as lipid A. attached to a hydrophilic linear polysaccharide region, consisting of the conserved core polysaccharide and O- specific polysaccharide or O antigen (highly variable). The lipid A inside Gram negative cells called endotoxin. A dead cell releases lipid when the outer membrane disintegrates, and lipid A may trigger fever, septic shock, inflammation  Function of the bacterial cell wall 1. It maintains the characteristic shape of bacteria. 2. Supports the cytoplasmic membrane against the high effects of osmotic pressure. 3. It is responsible for gram reaction. 4. Play an essential role in cell division 5. Providing attachment sites for bacteriophages 6. Antigenic determinants may be found in cell wall. 7. It is responsible for endotoxin activity of Gram-negative bacteria. 8. They are one of the most important sites for attack by antibiotics. Note: bacteriophage is virus infect the bacteria  Bacteria with deficient cell wall Naturally Induced L –form Mycoplasma Causes of L-forms bacteria 1. Failure of cell wall synthesis by antibiotics 2. High salt concentration. 3. Lysozyme which is found in saliva and tears dissolve rigid cell wall Protoplast is gram-positive organism that is devoid of cell wall Spheroplast is Gram-negative bacteria that is devoid of cell wall  Staining properties According to Gram stain, bacteria are divided into two groups 1. Gram positive bacteria 2. Gram negative bacteria  Cytoplasmic membrane It is semi-permeable double-layered structure, composed of phospholipids and protein. Functions of the Cytoplasmic Membrane; 1. Selective permeability barrier to different molecules that regulates the passage of substances into and out of the cell. 2. Active transport of ions ( H+, Na+, K+ etc...) and nutrients to achieve osmotic balance and a pool of nutrients. 3. It also supplies the cell with energy through electron transport and oxidative phosphorylation, i.e. site of respiration. 4. Excretion of hydrolytic exoenzymes which degrade the different nutrients into subunits small enough to penetrate cytoplasmic membrane. 5. It provides enzymes and lipid carriers for cell wall synthesis. 6. It play a role in DNA replication.  Intracytoplasmic structures 1. Nuclear body a) It is a single, circular packed bundle of double stranded DNA molecule (chromosome) b) There are no nuclear membrane or nucleolus. c) The DNA carries the genetic information to daughter cells d) It appears by the electron microscope 2. Ribosomes a) They are composed of protein and RNA. b) They are the site of protein synthesis. 3. Inclusions; The cytoplasm contains granules which represent accumulation of food reserve, 4. Plasmids; These are extrachromosomal double stranded circular DNA molecules that are capable of replicating independent of bacterial chromosome.  Capsule It is formed outside the cell wall It is chemical structure, usually polysaccharide except in Bacillus anthracis which has a protein capsule. The capsule can be seen by special stains Functions: 1. It protects the bacteria against phagocytosis 2. It is used in identification and typing of bacteria 3. It consider virulence inside host tissue  Flagellae Flagella are long structures that extend beyond the surface of a cell. It is organ of locomotion. They are protein in nature.  Pili ( fimbriae) It is hairy like projections the are shorter and thinner than flagella; they are composed of structured protein subunits termed pilin. They are mainly in gram negative bacteria. Two classes are known: *ordinary pili, which play a role in the adherence to host cells *Sex pili It is responsible for the attachment of donor and recipient cells in bacterial conjugation. It transfer DNA from bacteria to another  Bacterial endospores (spores) 1. It is a special tough resistant form of bacterial cell that can survive in a dormant state for years 2. The active multiplying bacteria called vegetative form 3. Sporulation is response to unfavourable conditions or starvation 4. It formed inside vegetative bacteria , it acquire a thick cortex layer and spore coat which contain calcium and less water 5. Spore released after lysing of remaining vegetative part 6. Bacterial endospores have different shapes, may be central, terminal or subterminal in bacterial cell. 7. Spores is stained by spore stain Sporulation  Spore resistance They are highly resistant to boiling and dryness It is killed by sterilization by autoclaving The marked resistance of spores is due to: 1. The thick spore cortex and tough spore coat. 2. The large amounts of calcium and less water 3. It is low metabolic and enzymatic activity.  Lecture 2 Bacterial physiology On completion of this lecture, the student will be able to: 1. To know the physical requirements , bacterial nutrition, oxygen requirement and bacterial reproduction 2. To describe the bacterial growth curve and it is related clinical application  Bacterial nutrition 1. Autotrophic bacteria 2. Heterotrophic bacteria can use organic sources of carbon (sugars and proteins) Most pathogenic bacteria are heterotrophic. Medical important  Autotrophic bacteria No medical importance According to nutritional requirements, the bacteria are classified into: Autotrophic bacteria can use inorganic substance for source of carbon and nitrogen. The energy needed by these bacteria is obtained from sun or oxidation of inorganic substance. Many autotrophic bacteria are saprophytic. Organic Simple inorganic substances Organic substances substances Carbon…….Co2 Nitrogen….. Ammonium salts proteins and carbohydrates  Oxygen requirements of bacteria aerobes anaerobes facultative aerotolerant microaerophilic Anaerobic bacteria that grow only in complete absence of oxygen. Obligate aerobes : these can grow only in the presence of free O2 Facultative anaerobes : These bacteria can grow in the presence of O2 and also grow when deprive of it Microaerophilic : these organisms grow best in the presence of low amount of O2  Physical requirements Physiology and metabolic activities if bacteria are divergent. The aim of these activities is growth and reproduction. Factors affecting the metabolic activities of bacteria 1. Moister: water is a major component of the bacterial cell, so moisture is absolutely necessary for growth of bacteria and high water content is essential in any medium used for bacterial cultivation. 2. Temperature for growth Mesophilic 25 C - 40 oC. ( pathogenic and opportunists) Psycorophilic ( saprophytic) bacterial grow below the minimum temp. Thermophilic : bacterial grow above the maximum temp. 3. Hydrogen ion concentration pH: Most pathogenic species of bacteria can grow at neutral pH 7.5 Some bacteria tolerate alkaline medium while other are more tolerant to acids  CO2 Requirement Normal CO2 concentration in atmospheric air (0.04%) is sufficient for growth of most bacteria. Some bacteria need high percentage (5 - 10% ) of CO2 e.g Neisseria gonorrhea. CO2 is needed to enhance toxin production and capsule formation.  Bacterial products 1. Bacterial enzymes 2. Bacterial toxins 3. Bacterial pigments Endopigments : it is non-diffusible pigment which remain to the body of the organisms. They are best demonstrated when the organism is grown on solid media e.g. golden pigment (Staph.aureus) Exopigments are diffusible pigments, which diffuse into the surrounding medium, e.g. pyoverdine and pyocyanin pigments of pseudomonas aeroginosa  Bacterial reproduction Bacterial reproduction is the process in which bacterial population increases in number under optimal conditions. Most bacteria reproduce by binary fission: One cell divided into 2 equal daughter cells similar in genetic character to the mother cell through generation time. Growth on solid media produce cell aggregates or colonies Growth on fluid media may give uniform turbidity, granular precipitate or surface pellicles  Bacterial growth curve on a bacterial cell culture A curve produced when a small number of a certain organism is placed in a suitable fluid medium and the viable cell number/ ml is determined periodically  Bacterial growth curve Lag Phase: This is the stage of preparation for bacterial multiplication during which the organism adapts itself by synthesis of new enzymes specific for the new medium. No cell division or initial number of bacteria remains constant. This process refers to the incubation time of the disease in vivo. Exponential (log) Phase. During this phase, rapid cell division occurs and the number of bacterial cells increases steadily by time. Antibiotics are effective during this phase. This phase corresponds to the acute phase of disease. Stationary Phase. Exponential growth cannot be continued Population growth is limited by exhaustion of food, accumulation of inhibitory metabolites. The number of dying cells equals that of newly formed cells. The number of viable bacteria remains constant. During this phase the sign and symptoms of the disease are still present. Decline Phase: exhaustion of nutrients and accumulation of toxic products continue, the death rate exceeds the multiplication rate and the number of living bacteria decreases steadily. This phase corresponds in vivo to the convalescent phase.  Growth phases stages Number of viable bacteria Lag phase The initial number of bacterial cells remains constant Log phase Markedly increased Stationary phase The number of viable bacteria remains constant Decline phase Markedly reduced  Lecture 3 Host-parasite relationship Intended Learning Outcomes (ILOs) On completion of this lecture, the student will be able to: Classification of antimicrobials 1. To know the relation between the host and bacteria 2. To know the factors affecting to host- parasite relationship 3. List the virulence factors of bacteria 4. To compare between the exotoxin and endotoxin  Types of bacteria According to natural habitat and relation to the host, bacteria are divided into 1--Saprophytic bacteria: – They found in nature as in soil, water, air and dust – Does not require a living host – Organism lives on dead animal 2-Parasitic bacteria, they are divided into A. Commensal organisms that live in a balance with the host. This group are microbiota of the human body B. Opportunistic bacteria: they produce disease in patients with suppressed immunity C. Pathogenic bacteria: they cause disease in healthy persons with intact immunological defense.  Commensals Test yourself?? Not cause disease (Normal flora) Function of commensals Microorganisms that live in or on To Inhibits pathogenic bacteria through the human body without causing – Competition damage include: – Bactericidal A. Microbiota – Decrease pH B. Normal flora C. Colonizers D. Human microbiome E. All of the above  The relationship between the host and bacteria 1-Colonization The parasite colonizes the host at different site without causing any harmful effect ( commensal bacteria in oral cavity) 2- infection The parasite invades the host tissues, elicits immune response but cause minor damage so that no sign and symptoms( subclinical infection) 3- infectious disease The parasite invades the host tissues, elicits immune response and cause tissue damage so that clinical sign and symptoms are appeared. 3- Carrier A human or animal who harbors the microorganisms responsible for causing infection but show no apparent sign or symptoms of infection or disease.  Type of carriers According to the duration of carriage can be classified into Transient carrier ( during incubation period) Chronic carrier ( hepatitis B virus carrier) According to the sites of carriers Urinary carriers ( salmonella carriers) Faecal carriers Throat carriers as in diphtheria Nasal carriers ( staphylococcal carriers) Nasopharyngeal carriers ( in meningococcal infection)  Virulence factors 1.Adherence factors : – Fimbrial adhesins protein that are located on the end of fimbriae that attach to host cell. – Non-fimbrial adhesins The bacteria have protein of cell wall components such as M protein of Streptococcus pyogenes, lipopolysacchride side chains in Gram negative bacteria and teichoic acid of coagulase negative Staphylococci 2. Invasion factors such as ability to survive and grow intracellular ( TB) 3. Capsule : antiphagocytic factor that prevent phagocytosis 4.Bacterial toxins 5. Toxic products or enzymes 6. Bacterial biofilm 7. Peptidoglycan of Gram-positive bacteria is the same biological effect of endotoxin but less potent.  Bacterial biofilm Bacterial biofilm: it is an aggregation of bacteria to each other and to solid surface and encased in exopolysaccharide matrix. e.g. biofilm in dental plaques Bacteria in biofilm It is protected from host immune mechanisms. It is out of reach of the antibiotics  Comparison between exotoxins and endotoxins Exotoxins Endotoxins Exotoxins are extracellular diffusible are integral part of the cell wall toxins lipopolysaccharide Protein in nature Heat stable at 60-80C Destroyed by heat ( heat liable) 60-80C poorly antigenic strongly antigenic.Weakly toxic highly toxic Have non-specific action( fever , shock) have specific action on cells cannot be converted to toxoids (neurotoxins, enterotoxins, cytotoxins) Produce fever can be converted into toxoids Produced by Gram-negative bacteria do not produce fever (E.coli and Salmonella) Produced by Gram-positive bacteria (Corynebacterium diphtheria) Secreted when cell dies Secreted by living cells  Lecture 4 antimicrobial drugs Intended Learning Outcomes (ILOs) On completion of this lecture, the student will be able to: 1. Classification of antimicrobials 2. To describe the mechanism of action 3. To know the spectrum of activity 4. List the complication of antibiotics 5. To know the indication of antibiotics combination and chemoprophylaxis  Classification of antibiotics Bacteriostatic : inhibition of bacterial growth Bactericidal: Kill of the microorganisms The minimal inhibitory concentration MIC is the lowest concentration of an antibiotic that prevents or inhibits bacterial growth The spectrum of antibiotics mean variety of bacteria that are affected by these antibiotic Antibiotics act on gram positive bacteria ( narrow spectrum) Antibiotics act on both gram positive and gram negative ( broad spectrum) Mechanisms of action of antibiotics  Mechanism of action of antimicrobial agents 1. Inhibition of cell wall synthesis: β-lactam antibiotics include penicillins and cephalosporins 2. Inhibition of Cell membrane function:. Polymyxin, amphotericin B and nystatin are examples. 3. Inhibition of Protein synthesis: They inhibit protein synthesis by binding to 30S subunit of microbial ribosomes. e.g. Aminoglycosides (streptomycin, gentamycin) and tetracycline They inhibit protein synthesis by binding to 50S subunit of microbial ribosomes. e.g. Erythromycin macrolide, clindamycin and linzolid chloramphenicol 4. Inhibition of nucleic acids synthesis: The antibiotics inhibit DNA synthesis by blocking DNA gyrase e.g. quinolones. And the antibiotics inhibit the synthesis of RNA by binding to RNA polymerase enzyme e.g. rifampicin. 5-Competitive Inhibition: Some antibiotics are competitive inhibitors of essential metabolites are needed in bacterial metabolism. Such a drug will compete with metabolite for the enzymes that act on this metabolite. This will be result in formation of non-functional products e.g. Sulfonamides Para amino-benzoic acid (PABA) is an essential metabolite for many organisms. |They use it as a precursor in folic acid synthesis which is essential for nucleic acids synthesis. Sulfonamides are structurally similar to Para-amino-benzoic acid so they enter into the reaction in place of PABA and compete for the active center of the enzymes thus inhibiting folic acid synthesis. Principles of Antimicrobial Use Patient with Symptoms & Sign Pathogen Directed Narrowest Spectrum Best Outcome  Mechanism of antibiotics resistance 1. Production of enzymes that inactivate the antimicrobial agent ( penicillin resistance) 2. Alteration of the target site: (tetracyclines and chloramphenicol resistance) 3. Alteration of permeability to antimicrobial agent (aminoglycosides resistance). 4. Development of new metabolic pathways: e.g. sulfonamides resistant 5. Efflux- pump mediated antimicrobial resistance: Bacteria actively pump the drug out across the cytoplasmic membrane using efflux or multi-drug resistance pump.  Complications of antimicrobials Prolonged use – it will be killed commensal organisms – Resistance Early use of antibiotics Mask of symptoms Inhibition of the immunity Toxicity Superinfections Hypersensitivity penicillin anaphylactic shock Thank you