Clinical Bacteriology Lecture - Microbiology PDF

CLINICAL BACTERIOLOGY LECTURE - MICRO 1 Ms. Giane Paola T. Tanjuaquio, RMT *:･ﾟ✧*:･ﾟ INTRODUCTION TO BACTERIOLOGY DEFINITION OF TERMS PATHOGENS:...

CLINICAL BACTERIOLOGY LECTURE - MICRO 1 Ms. Giane Paola T. Tanjuaquio, RMT *:･ﾟ✧*:･ﾟ INTRODUCTION TO BACTERIOLOGY DEFINITION OF TERMS PATHOGENS: disease-causing MICROBIOLOGY microorganism such as bacteria, fungi, protozoans, and viruses - includes the study of certain non-living entities as well as OPPORTUNISTIC TRUE PATHOGEN PATHOGEN certain living organisms. Collectively, these non-living organism that will organisms that will entities and living organisms are cause disease in a cause disease in an called microbes. healthy host immunocompromised - a branch of biology which deals host with the study of living organisms PATHOGENICITY: ability of an organism that are too small to be seen by to cause disease in a host organism the naked eye VIRULENCE: the degree of pathogenicity; the power by which a BRANCHES OF MICROBIOLOGY pathogen can cause a severe disease PATHOGENIC BASIC DETERMINANTS/VIRULENCE FACTORS: By organism By process Disease related any genetic, biochemical or structural features that enable a pathogen to cause Bacteriology Microbial Immunology disease in a host organism Phycology metabolism Epidemiology INFECTION: the entry, invasion and Mycology Microbial Etiology multiplication of pathogens in or on to Virology genetics the host body system which results to Parasitology subsequent tissue injury and progress to Protozoology overt disease APPLIED A. Based on Source of Pathogen Disease Environmentally ENDOGENOUS EXOGENOUS Industrial related related INFECTION: infection INFECTION: infection Infection arising from colonizing arising from invading Environmental Food and control beverage tech flora pathogen from the microbiology Chemotherapy Pharmaceutical external environment microbiology B. Based on Clinical Onset of Signs and Genetic engineering Symptoms ACUTE INFECTION: CHRONIC INFECTION: rapid/sudden onset of gradual onset of signs 4 GROUPS OF MICROBES signs and symptoms and symptoms that VIRUS: Very simple microbes, consisting which are usually are usually mild to of nucleic acid, a few proteins, and (in severe to fatal that moderate that may some) a lipid envelope may lead to death progress to long FUNGI: Heterotrophic eukaryotes that standing infection obtain nutrients through absorption C. Based on Etiologic/Causative Agent PARASITES: Eukaryotic organisms that live at the expense of their hosts exist as NOSOCOMIAL ZOONOTIC INFECTION unicellular organisms of microscopic size INFECTION: acquired (ZOONOSIS): animal whereas others are multicellular during hospitalization disease which can organisms spread to humans; BACTERIA: Prokaryotic, unicellular animal-acquired organisms that lack a nuclear membrane infection and true nucleus PAGE 1 Viray, M.S.Y. - 3B ˚ ༘♡ ⋆｡˚ ❀  CLINICAL BACTERIOLOGY LECTURE - MICRO 1 Ms. Giane Paola T. Tanjuaquio, RMT *:･ﾟ✧*:･ﾟ D. Based on Clinical Manifestations rabies, anthrax, dysentery, smallpox, SUBCLINICAL/ CLINICAL/ ergotism, botulism, ASYMPTOMATIC/ NON SYMPTOMATIC/ measles, typhoid APPARENT: no APPARENT: associated fever, typhus fever, obvious appearance of with presence of overt diphtheria, and signs and symptoms signs and symptoms of syphilis and the person is the disease - First appeared in unaware of the Europe infection - Was carried by the Native Americans DISEASE: an altered health state in an who were brought infected host in Portugal by → INFECTIOUS DISEASE: caused by Christopher a pathogen which invades body SYPHILIS Columbus tissues and causes damage 1493 (Treponema - Neapolitan Disease → COMMUNICABLE DISEASE: pallidum) (French) capable of spreading from person - French or Spanish to person Disease (Italian) → SYMPTOMS: any subjective - French pox evidence of disease (English) - Spanish, German, ○ E.g. headache, dizziness, Polish and Turkish etc. pocks → SIGNS: readily observable evidence of disease The infection was 1530 NORMAL FLORA: Bacteria that are in or named “Syphilis” on different sites of the body that usually PIONEERS IN MICROBIOLOGY do not harm the host unless the host - First person to see live bacteria defense is compromised and protozoa COLONIZATION: the establishment of - “Father of Microbiology, substantial number of microorganisms Bacteriology, Protozoology” usually in the skin or mucosa but there’s - He ground tiny glass lenses, no penetration of tissues which he mounted in small metal Anton Van frames, thus creating what today EARLIEST KNOWN INFECTIOUS DISEASES Leeuwenhoek are known as single lens - Fatal epidemic (1632-1723) microscopes or simple disease caused by a microscopes bacteria called - He observed various tiny living PESTILENCE 3180 BC Yersinia pestis – creatures, which he called and PLAGUE BLACK DEATH “animalcules” (bacteria and - The first recorded protozoa) epidemic - Demonstrated that different near the end of the types of microbes produce Trojan war, the different fermentation products Around BUBONIC Greek army was - Disproved theory of 900 BC PLAGUE decimated by this spontaneous epidemic Louis Pasteur generation/Abiogenesis EBERS describing epidemic (1822-1895) − Life can arise spontaneously 1500 BC from non-living materials PAPYRUS fevers - He introduced the terms 1122 BC SMALLPOX occurred in China “aerobes” (requires oxygen) and “anaerobes” (does not requires - occurred in Rome oxygen) 790, 710, EPIDEMICS OF and in Greece and 640 BC PLAGUE around 430 BC PASTEURIZATION - Early accounts of - Heating liquids to 63–65°C for 30 minutes or PAGE 2 Viray, M.S.Y. - 3B ˚ ༘♡ ⋆｡˚ ❀  CLINICAL BACTERIOLOGY LECTURE - MICRO 1 Ms. Giane Paola T. Tanjuaquio, RMT *:･ﾟ✧*:･ﾟ 73 75°C for 15 seconds Semmelweis - showed quite clearly that - Type of sterilization. Only kills pathogen women became infected in the maternity ward after GERM THEORY DISEASE examinations by physicians - Specific microbes cause specific infectious diseases coming directly from the autopsy - Developed vaccines to prevent chicken cholera, room anthrax and swine erysipelas - First to introduce aseptic - Made many significant techniques aimed at reducing contributions to the germ theory Joseph Lister microbes in a medical setting and of disease preventing wound infections - Discovered that B. anthracis produces spores, capable of KEY DEVELOPMENTAL resisting adverse conditions PERIOD SCIENTIST/S NOTES - Developed methods of fixing, Robert Koch Publication of the first staining, and photographing (1843-1910) 1665 Robert Hooke description of bacteria, methods of cultivating bacteria on solid media microbes - Discovered the bacterium (M. Anton van Observation of “little tuberculosis) that causes 1667 Leeuwenhoek animals” tuberculosis and the bacterium (Vibrio cholerae) that causes Smallpox Vaccination cholera 1796 Edward Jenner – first scientific validation Koch’s Postulate Advocating Ignaz handwashing in the 1850 Semmelweis prevention of the spread of disease Spontaneous 1861 generation disproved Louis Pasteur Publication of the paper supporting the 1862 germ theory of disease Practice of Antiseptic 1867 Joseph Lister THE DISCOVERY OF SPORES AND STERILIZATION Surgery - Provided the initial evidence Discovery of Bacillus that some of the microbes in dust anthracis which 1876 and air have very high heat became the first proof John Tyndall resistance and that particularly of germ theory vigorous treatment is required to Robert Koch Utilization of solid destroy them 1881 culture media for - Developed Tyndallization bacterial growth - Clarified the reason that heat Outlined Koch’s would sometimes fail to 1882 Ferdinand postulate completely eliminate all Cohn Microorganisms Development of Paul Erlich - “Sterile” was established Acid-fast stain THE DEVELOPMENT OF ASEPTIC TECHNIQUES Hans Christian Gram stain developed 1884 Gram observed that mothers who gave Dr. Oliver birth at home experienced fewer First Rabies Wendell 1885 Louis Pasteur infections than did mothers who Vaccination Holmes gave birth in the hospital Invention of the Petri 1887 Richard J. Petri Dr. Ignaz - Father of Hand Hygiene Dish PAGE 3 Viray, M.S.Y. - 3B ˚ ༘♡ ⋆｡˚ ❀  CLINICAL BACTERIOLOGY LECTURE - MICRO 1 Ms. Giane Paola T. Tanjuaquio, RMT *:･ﾟ✧*:･ﾟ Dmitri Iosifovich Discovery of Viruses recognized or named and placed in 1892 overall taxonomic scheme (genotypic Ivanovski and phenotypic characteristics) T. Smith and F.I. Zoonosis - first 1893 Kilbourne described CLASSIFICATION Viral dependence on Martinus living host cells for - Provided the initial evidence 1899 Beijerinck reproduction that some of the microbes in dust recognized Carl von Linne and air have very high heat (Linnaeus; resistance and that particularly Proof the mosquitoes vigorous treatment is required to 1701-1778) 1900 Walter Reed carry the agent of destroy them yellow fever - Developed Tyndallization Discovered the cure 1910 Paul Erlich Carl von Linne’s Classification System for syphilis Alexander Discovery of Penicillin 1928 Fleming J. Watson and F. Proposed and built 1953 Crick the DNA model Development of the W. Gilbert, F. 1977 DNA sequencing Sanger method Invention of the Kary Mulis Polymerase Chain Reaction (PCR) TAXONOMY - Whittaker’s tree - based on structural similarities Greek taxes: arrangement; Greek nomos: law and differences, such as - is the orderly classification and grouping of Robert prokaryotic and eukaryotic organisms into taxa (categories) Whittaker cellular organization, and the way - it is based on similarities and differences in these organisms obtain their genotype and phenotype nutrition. GENOTYPE: genetic makeup of an WHITTAKER’S TREE organism, or combinations of forms of one or a few genes under scrutiny in an organism's genome. PHENOTYPE: readily observable physical and functional features of an organism expressed by its genotype. THREE CATEGORIES OF TAXONOMY 1. NOMENCLATURE: is the assignment of scientific names to the various taxonomic categories and individual organisms 2. CLASSIFICATION: Attempts the orderly arrangement of organisms into a hierarchy of taxa (categories) 3. IDENTIFICATION: Is the process of discovering and recording the traits or organisms so that they may be PAGE 4 Viray, M.S.Y. - 3B ˚ ༘♡ ⋆｡˚ ❀  CLINICAL BACTERIOLOGY LECTURE - MICRO 1 Ms. Giane Paola T. Tanjuaquio, RMT *:･ﾟ✧*:･ﾟ - devised a Three Domain System STANDARD RULES FOR DENOTING of Classification BACTERIAL NAMES - there are two domains of prokaryotes – based on cellular Carl R. Woese organization and function Family name is capitalized and has an (Archaea and Bacteria) and one "-aceae" ending (Micrococcaeceae) domain (Eukarya or Eukarya), Genus name is capitalized and followed which includes all eukaryotic by the species epithet, which begins with organisms. a lowercase letter, both the genus and THREE-DOMAIN SYSTEM OF CLASSIFICATION the species should be italicized in print but underlined when written in script (E.g. Staphylococcus aureus or Staphylococcus aureus) Abbreviated by using the first letter (capitalized) of the genus followed by a period and the species epithet (S. aureus). First two letters or the first syllable are used when two or more genera names begin with the same first letter (Staph. And Strept. For Staphylococcus and Streptococcus) Escherichia coli (E. coli or Esch. coli) is a bacterium, but Entamoeba coli (Ent. coli) is an intestinal parasite NOMENCLATURE IDENTIFICATION International Code of Nomenclature of Bacteria (ICNB) or the Bacteriological GENOTYPIC CHARACTERISTICS: relate Code (BC): provides the accepted labels to an organism’s genetic makeup, by which organisms are universally including the nature of the organism’s recognized genes and constituent nucleic acids PHENOTYPIC CHARACTERISTICS: are BINOMIAL SYSTEM OF based on features beyond the genetic NOMENCLATURE level and include both readily observable characteristics and characteristics that - every organism is assigned a genus and a may require extensive analytical species of Latin or Greek derivation Each procedures to be detected organism has a scientific “label” consisting of SUBSPECIES: based on phenotypic two parts: differences (abbreviated "subsp.") Genus – the first letter is always SEROVARIETIES: based on serologic capitalized differences (antigenic properties) Species – first letter is always lower case (abbreviated "serovar") ○ PRINTED: Italicized BIOVARIETIES: based on biochemical or ○ SCRIPT/HANDWRITTEN: physiological test result differences Underlined (separate line for (abbreviated "biovar") genus and species) PHAGE TYPING: based on susceptibility to specific bacterial phages Analysis of ribosomal RNA (rRNA) PAGE 5 Viray, M.S.Y. - 3B ˚ ༘♡ ⋆｡˚ ❀  CLINICAL BACTERIOLOGY LECTURE - MICRO 1 Ms. Giane Paola T. Tanjuaquio, RMT *:･ﾟ✧*:･ﾟ organelles BY CELLULAR TYPE Golgi bodies Present in some PROKARYOTES: Gr. “before kernel” Present in some; (nucleus) contain Lysosomes - domains Archaea and Bacteria hydrolytic (Eubacteria) enzymes - Simple cells that lacking Present in all; membrane bound nucleus Endoplasmic Absent in all lipid synthesis, - Prokaryotes fall under the domain reticulum transport Archaea and Bacteria. Bacteria fall Mitochondria Present in most in either of these two domains EUKARYOTES: Gr. eu: “well or good” and Nucleus Present in all Gr. Karyon: kernel Present in algae Chloroplasts for - fungi, algae, protozoa, animals, photosynthesis and plants and Plants o With membrane Ribosome: site bound nucleus of protein - Categorized under the domain synthesis Present in all Present in all Eukarya (nonmembrano us) size CHARACT 70S consisting of In the inner PROKARYOTE EUKARYOTE ERISTICS Electron 50S and 30S membrane of transport for 0.4-2 um in 10 - 100 um in energy subunits mitochondria Typical size diameter 0.5-5 diameter >10 um and chloroplasts um in length in length Sterols in Absent except in No nuclear Membrane cytoplasmic Mycoplasmatace Present membrane, bound nucleus membrane ae Nucleus nucleoid region Plasma Peptidoglycan in of the cytosol membrane most bacteria In the nucleoid, In the nucleus Location Peptidoglycan in Cellulose, at the mesosome most bacteria phenolic Circular; Linear; Cell wall, if polymers, lignin complexed with complexed with present (plant), chitin Chromosomal RNA basic histones (fungi), other DNA and other glycans (algae) proteins Present in most Plasmids, small In mitochondria as an organized Present; some circular molecule and chloroplasts Glycocalyx capsule or animal cells of DNA unorganized containing slime layer accessory Cilia Absent Present information; most Extrachromo commonly found Flagella, if Complex cilia or somal circular Simple flagella DNA in gram-negative present flagella bacteria; each Pili and carries genes for Present Absent fimbriae its own replication; can confer resistance to antibiotics Asexual (binary Sexual and Reproduction fission) asexual Membrane Absent All -bound PAGE 6 Viray, M.S.Y. - 3B ˚ ༘♡ ⋆｡˚ ❀  CLINICAL BACTERIOLOGY LECTURE - MICRO 1 Ms. Giane Paola T. Tanjuaquio, RMT *:･ﾟ✧*:･ﾟ BACTERIAL STRUCTURE AND FUNCTION IMPORTANT CHARACTERISTICS OF BACTERIA PROKARYOTIC Have no organelles; No membrane enclosed structure Do not have histones o Have 70S ribosomes and are HAPLOID with a single chromosome. Cell wall: with peptidoglycan except for Mycoplasma and Ureaplasma ARCHAEA (ARCHAEOBACTERIA): more Cytoplasmic membrane: fluid closely related to eukaryotic cells / found phospholipid bilayer with carbohydrate in microorganisms that grow under and sterol extreme environmental peptidoglycan - Site of energy production conditions/lack Free Ribosomes: Site of protein synthesis - Greek: (archaics) ancient, origin Unicellular; contains both DNA and RNA from the earliest cells o Average size of 0.2-5.0 microns - Those that belong in the domain - Smallest living organism: Archaea are some of the most Mycoplasma resistant - Largest: Bacillus - organisms are found in extreme Multiplies by binary fission environmental conditions. Biofilms: property of bacteria to attach on - Has cell wall, plasma membrane, solid surface o Pathogenicity: ability of a ribosomes and flagella (no microbe to produce disease in a nucleus and membrane bound susceptible individual organelles Virulence: relative ability of a - Never contain peptidoglycan microorganism to cause disease or the contain protein or glycoprotein degree of pathogenicity wall structure known as S-layer - usually measured by the numbers EXTREMOPHILES: lovers of the extreme of microorganisms necessary to conditions cause infection in the host E.g. Sulfurovum epsilonproteobacteria HALOPHILES: salt-loving cells - in Utah's Great Salt Lake Found in environments high in salinity or salt concentrations E.g. Salinibacter ruber THERMOPHILES: heat-loving cells - In hot springs and deep ocean vents - Thrive in environments with extreme heat E.g. Methanobacterium wolfei PAGE 7 Viray, M.S.Y. - 3B ˚ ༘♡ ⋆｡˚ ❀  CLINICAL BACTERIOLOGY LECTURE - MICRO 1 Ms. Giane Paola T. Tanjuaquio, RMT *:･ﾟ✧*:･ﾟ II. Based on Structure VIRULENCE FACTORS CELL ENVELOPE: Composed of layers ADHERENCE FACTORS: pili/fimbriae (capsule, cell wall, cell membrane) that ANTIPHAGOCYTIC FACTORS: capsule surround the bacterium. and self-component of cell wall CELL WALL: also referred to as the ENZYMES: Coagulase - S. aureus, peptidoglycan or murein layer Fibrinolysin - spreading and clotting - Gives bacterial cell shape and strength to withstand changes in EXOTOXIN ENDOTOXIN the environmental osmotic Released by all major Usually produced by gram pressures gram (+) (-) organisms - Protects against mechanical Exotoxin is excreted by It requires cell death for disruption of the cell living bacteria and it release (Cell wall - Some bacteria use the cell wall for does not require cell disintegration) pathogenicity: death for release; → M PROTEIN: major metabolic product of virulence factor and bacteria prevents phagocytosis for Composition: mainly Composition: S. pyogenes PROTEIN in nature Polysaccharide and lipids → MYCOLIC ACID: responsible for acid Effect: "Local" - one area Effect: “Systemic” all over fastness of Mycobacterium Not associated with fever the body species and prevents Toxicity: High Toxicity: Low digestion during Stability to heating: Stability to heating: phagocytosis UNSTABLE at 60 STABLE → POLY-D-GLUTAMIC ACID: Bacillus species Stimulates antitoxin Stimulates antitoxin → CHITIN: fungi production: b (can be production: NO (cannot converted into toxoid; be converted into toxoid; → L-FORM: organisms that easily neutralized by anti- not easily neutralized by temporarily lost their cell toxin) anti- toxin) wall as a result of environmental conditions Usually bind to specific Specific receptors not → Peptidoglycan consists of receptors on cells found on cells glycan chains of Frequently controlled Synthesis directed by alternating extrachromosomal (e.g. chromosomal genes N-acetyl-d-glucosamine plasmids) by genes (NAG) and N-acetyl-d-muramic acid OTHER CHARACTERISTICS OF (NAM) BACTERIA I. Based on Classification DOMAIN: Bacteria (Prokaryotes) PHYLA: E.g. Proteobacteria, Firmicutes, Actinobacteria, and Bacteroidetes GENUS AND SPECIES: Based on genetic, biochemical, and phenotypic characteristics PAGE 8 Viray, M.S.Y. - 3B ˚ ༘♡ ⋆｡˚ ❀  CLINICAL BACTERIOLOGY LECTURE - MICRO 1 Ms. Giane Paola T. Tanjuaquio, RMT *:･ﾟ✧*:･ﾟ GRAM + GRAM - - Motility is best seen at room temperature – Single-layered – Double Layered – Thick protective – Thin peptidoglycan TYPES OF FLAGELLA peptidoglycan layer (single−layer) → ATRICHOUS: absence of flagella – layer (several layers) – Large periplasmic – Small periplasmic space → MONOTRICHOUS: single flagellum in layer – Presence of outer one end – Absence of outer membrane → LOPHOTRICHOUS: multiple flagella in membrane – No teichoic acid one end – Has teichoic acid – Thickness: 10 → AMPITRICHOUS: single flagella in both – Thickness: 15 80 nanometers ends nanometers – Presence of porins → PERITRICHOUS: flagella around in the – Absence of porins – High Lipid content: bacterium – Very low lipid 15-20% → AMPHILOPHOTRICHOUS: multiple content (2-5%) – Has – No lipopolysaccharides flagella in both ends lipopolysaccharides Gram Staining Procedure: WAYS TO DEMONSTRATE MOTILITY Gram Staining - No retention of Procedure: crystal violet dye a. Flagellar stain - Retains the - Appears pink in b. Hanging drop crystal violet dye color. c. Semi-solid media - Appears purple i. NON-MOTILE: growth is in the line in color ii. MOTILE: outside the line TYPES OF MOTILITY CAPSULE: slimy area surrounding the → Tumbling, gliding, darting, cork screw, cell wall, responsible for mucoid colonies twitching, shooting star - Usually made of polysaccharide or polypeptide - Neufeld-Quellung capsular Ag (+) capsular swelling due to Ag- Ab METACHROMATIC GRANULES: serves as - Serotyping: energy source and food reserves → Somatic O Ag - heat stable - Composed of glycogen, lipids, → Vi Ag (Salmonella) and K polyphosphates or Ag (E. coli) - heat labile pyrophosphates PLASMA MEMBRANE: surrounds the - stained using Albert, Lamb, cytoplasm Neisser, and Ljubinsky - Site of energy synthesis (ATP site) - Transport of nutrients and out of TYPES OF GRANULES the cell → MUCH GRANULES: Mycobacterium PILI/ FIMBRIAE: hair-like proteinaceous tuberculosis structured made up of “pilin” → BABES-ERNST/METACHROMATIC TWO CLASSES: GRANULES: C. dIptheriae a. Common/ordinary pili → BIPOLAR BODIES: Yersinia pestis b. Sex pili → SULFUR BODIES: Actinomyces ENDOSPORES: composed of Calcium dipicolinate/ Dipicolinic Acid - Found in Clostridium and Bacillus RIBOSOMES: for protein synthesis species - give the bacterial cytoplasm a - highly resistant to desiccation, granular appearance when heat and chemical agents viewed in the electron FLAGELLA: whip-like structure used as microscope an organ for locomotion MESOSOMES: point of attachment of - Made up of “flagellin” chromosomes - Antigenic (H-antigen) PAGE 9 Viray, M.S.Y. - 3B ˚ ༘♡ ⋆｡˚ ❀  CLINICAL BACTERIOLOGY LECTURE - MICRO 1 Ms. Giane Paola T. Tanjuaquio, RMT *:･ﾟ✧*:･ﾟ - convoluted invaginations of cell Cocci in chains membrane - important for DNA Sarcinate Coccus in cubical packets replication and cell division CYTOSOL: fluid portion of the cell Cocci in clusters - Contains ribosomes and various types of nutritional storage Tetrads Packets of four granules NUCLEOID: where the bacterial DNA is found; also known as nuclear body - Contains bacterial chromosome III. Based on Function METABOLISM: enzymatic pathways for energy production (aerobic or anaerobic) REPRODUCTION: binary fission for asexual reproduction GENETIC EXCHANGE: conjugation, transformation, transduction for genetic variation SURVIVAL: endospore formation in some bacteria under harsh conditions B. Bacilli (Bacillus) - Rod-shaped, cylindrical or elongated IV. Based on Morphology Single bacilli Two cocci adjacent to each other Diplobacillus Coccus in cubical packets Streptobacilli Bacillus in chain Bacillus that are small, short, Cocobacilli stout/plump Irregular bacilli C. Spirilla (Spirals) - Helical or twisted bacteria - Spiral with two or more curves, quite rigid - Loosely twisted spiral resembling a stretched A. Cocci (Coccus) spiral - Round, spherical-shaped bacteria - Tightly twisted spiral resembling a cork - May occur singly, in pairs, in chains or in screw clusters - Tightly twisted spiral with one or both ends Diplococcus Two cocci adjacent to each other bent into a hook, sometimes even resembling an interrogative symbol PAGE 10 Viray, M.S.Y. - 3B ˚ ༘♡ ⋆｡˚ ❀  CLINICAL BACTERIOLOGY LECTURE - MICRO 1 Ms. Giane Paola T. Tanjuaquio, RMT *:･ﾟ✧*:･ﾟ V. Antigenic Determination Based on BACTERIAL GENETICS Serological Typing O Antigen: Associated with the cell wall DNA H Antigen: Associated with the flagella K Antigen: Associated with the capsule - Deoxyribonucleic acid Vi Antigen: Specific capsular antigen of - It is a double-helical chain of Salmonella typhi deoxynucleotides. - The helix is a double strand twisted VI. Based on Molecular Techniques together, which many scientists refer to as a spiral PCR: Amplification of specific DNA sequences THE DISCOVERY OF DNA DNA Sequencing: determines the genetic sequence FRIEDRICK Discovered DNA in 1869 RFLP: restriction fragment length MIESCHER polymorphism for DNA profiling FISH: Fluorescent in situ hybridization for PHOEBUS A. T. Discovered the composition of LEVINE DNA in 1920s detecting specific DNA/RNA sequences in cells ROSALIND Discovered the helical structure FRANKLIN by x-ray crystallography JAMES Described the three-dimensional WATSON and (3D) structure of the DNA FRANCIS CRICK molecule in 1950s NUCLEOTIDE COMPOSITION Phosphate group (PO4) Cyclic five-carbon pentose Nitrogen-containing base NITROGENOUS BASES IN NUCLEOTIDES - PURINE: consists of a fused ring of nine carbon atoms and nitrogen PAGE 11 Viray, M.S.Y. - 3B ˚ ༘♡ ⋆｡˚ ❀  CLINICAL BACTERIOLOGY LECTURE - MICRO 1 Ms. Giane Paola T. Tanjuaquio, RMT *:･ﾟ✧*:･ﾟ - PYRIMIDINE: consists of a single ring of CONJUGATION: horizontal transfer of six atoms of carbon and nitrogen genetic material by cell-to-cell contact TRANSPOSON: some fragments of DNA are mobile and can jump from one place in the chromosome to another place - All deoxynucleotides made up of MUTATION: changes that occur in the combinations of four chemical units DNA code and often result in a change in called nucleotides: the coded protein or in the prevention of Adenine (A) its synthesis Thymine (T) GENETIC RECOMBINATION: a method Cytosine (C) by which genes are transferred or Guanine (G) exchanged between homologous (similar) regions on two DNA molecules, forming new combinations of genes on a → Adenine - Thymine (A with T) chromosome → Guanine - Cytosine (G with C) MECHANISM OF GENE TRANSFER TERMINOLOGIES TRANSFORMATION: the uptake and incorporation of free or naked DNA into a bacterial cell GENOTYPE: the genetic potential of the DNA of an organism PHENOTYPE: consists of observed characteristics by the genome REPLICATION: The duplication of chromosomal DNA for insertion into a daughter cell BINARY FISSION: a single bacterial cell divides into two genetically identical daughter cells TRANSDUCTION: the transfer of bacterial TRANSCRIPTION: the synthesis of genes by a bacteriophage from one cell mRNAa by the enzyme RNA polymerase to another using one strand of DNA as template TRANSLATION: the synthesis of a specific protein from the mRNA code PROTEIN EXPRESSION: synthesis of protein CODON: group of three nucleotides in an mRNA molecule that signifies a specific amino acid ANTICODON: the triplet of bases on the tRNA that bind the triplet of bases (codon) on the mRNA BACTERIOPHAGE: consists of genetic material (DNA or RNA) surrounded by a GENETIC ELEMENTS AND protein coat ALTERATIONS CONJUGATION: the transfer of genetic material from a donor bacterial strain to a BACTERIAL GENOME: consists of a recipient strain single, closed, circular piece of dsDNA that is super coiled to fit inside the cell PLASMIDS: small, circular pieces of extrachromosomal dsDNA PAGE 12 Viray, M.S.Y. - 3B ˚ ༘♡ ⋆｡˚ ❀  CLINICAL BACTERIOLOGY LECTURE - MICRO 1 Ms. Giane Paola T. Tanjuaquio, RMT *:･ﾟ✧*:･ﾟ BACTERIAL GROWTH AND NUTRITION BACTERIAL GROWTH - Refers to the increase in the number of bacteria - This growth is affected by various factors such as optimum growth requirements, dynamics of growth, including the use of a medium that can be artificially prepared in the laboratory RESTRICTION ENZYME: produced that cut (restrict) incoming, foreign DNA at NUTRITIONAL REQUIREMENTS specific DNA sequences - Autotroph CARBON - heterotroph ENERGY - Phototrophic SOURCE - chemotrophic WATER/ MOISTURE/ HUMIDITY MINERAL Needed as co-factors in various ELEMENTS metabolic process of the bacteria - Lithotroph, organotroph ELECTRON - NADH reduced from NAD SOURCE - FADH2 reduced from FAD - Needed for the synthesis of proteins - Free nitrogen from the air NITROGEN - Nitrogenous compounds in the culture media (e.g. peptone, yeast, beef extract) - Halophilic bacteria/ halophiles SALT - Organotroph - NADH reduced from NAD - FADH2 reduced from FAD ADDITIONAL/SPECIAL GROWTH REQUIREMENTS FASTIDIOUS: Haemophilus spp. - Requires both X & V factor - Culture Media: Blood Agar Plate - 1st priority: 5% defibrinated sheep’s blood - 2nd priority: horse blood - 3rd priority: rabbit’s blood - Least priority: human blood X Factor (Hemin/ Hematin) V Factor PAGE 13 Viray, M.S.Y. - 3B ˚ ༘♡ ⋆｡˚ ❀  CLINICAL BACTERIOLOGY LECTURE - MICRO 1 Ms. Giane Paola T. Tanjuaquio, RMT *:･ﾟ✧*:･ﾟ ENVIRONMENTAL REQUIREMENTS BACTERIAL METABOLISM TEMPERATURE - biochemical reactions bacteria use to break down organic compounds and the Incubation temperature for most reactions they use to synthesize new bacteria and viruses: 35° - 37°C bacterial molecules from the resulting Incubation temperature and time for carbon skeletons AEROBES: 35° - 37°C for 18-24 hours - Carbohydrates 🡪 Glucose 🡪 Pyruvic acid 🡪 Incubation temperature and time for energy + By-products ANAEROBES: 35° - 37°C for 2-48 hours Diagnostic Laboratory usually incubate cultures for bacterial growth at 35°C FERMENTATION Pseudomonas and Campylobacter can grow at 35°C and 42°C - anaerobic process carried out by Mesophilic obligate, facultative and aerotolerant Psychrophilic/ Cryophilic anaerobes Thermophilic - Also important because it generates Hyperthermophilic/ Extremely nicotinamide adenine dinucleotide (NAD Thermophilic - molecule necessary for maintaining the Eurithermophilic Krebs cycle) Sternothermophilic - mixture of end products (e.g. lactate, butyrate, ethanol, and acetoin) pH accumulates in the medium Acidophilic - used loosely in the diagnostic Alkaliphilic microbiology laboratory to indicate any Osmophilic type of utilization—fermentative or oxidative—of a carbohydrate—sugar—with the resulting BACTERIAL GROWTH PHASE production of an acid pH GENERATION TIME Time to replicate AEROBIC RESPIRATION (OXIDATION) Refers to the stages of bacterial growth Fast Growing Bacteria: 20 minutes - an efficient energy-generating process Slow Growing Bacteria: 24 hours molecular oxygen (O2) is the final electron acceptor - Obligate aerobes and facultative anaerobes undergo aerobic respiration - Certain anaerobes can carry out anaerobic respiration, in which molecules other than molecular oxygen, such as nitrate and sulfate, act as the final electron acceptors. - Anaerobic respiration is less energy yielding than aerobic respiration THREE MAJOR METABOLIC PATHWAYS - Pyruvate: can be further catabolized either fermentatively or oxidatively. 1. Embden-Meyerhof-Parnas (EMP) glycolytic pathway 2. Pentose phosphate pathway PAGE 14 Viray, M.S.Y. - 3B ˚ ༘♡ ⋆｡˚ ❀  CLINICAL BACTERIOLOGY LECTURE - MICRO 1 Ms. Giane Paola T. Tanjuaquio, RMT *:･ﾟ✧*:･ﾟ 3. Entner-Doudoroff pathway glucose must not be present if the ability to ferment another sugar is being tested GLUCOSE 🡪 PYRUVIC ACID/PYRUVATE – ability to ferment lactose is an important step in classifying members of the family Embden-Meyerhof-Parnas Glycolytic Enterobacteriaceae Pathway ○ classified as either lactose – Major source (90% glycolysis) fermenters or lactose – Yields 2 ATOs (Anaerobic) non-fermenters Pentose-Phosphate (Phosphogluconate) ○ utilization of lactose by a Pathway bacterium requires two steps: – 10% glycolysis 1. requires an enzyme, β-galactoside permease, – Yields 38 molecules of ATPs for the transport of lactose across the cell wall (Anaerobic) into the bacterial cytoplasm 2. occurs inside the cell and requires the enzyme PYRUVATE 🡪 ENERGY + β-galactosidase to break the galactoside bond, BY-PRODUCTS releasing glucose, which then can be fermented. Respiration (Aerobic Process) – Kreb’s Cycle – Electron Transport Chain – Glucose 🡪 CO2 and H2O Oxidation – Glucose - Acid Fermentation – Anaerobic process carried out by both obligate and facultative anaerobes – Organic molecule is the final electron transporter CLASSIFICATION OF FERMENTATION BASED ON END PRODUCTS Alcoholic Fermentation Homolactic Fermentation Heterolactic Fermentation Propionic acid Fermentation Mixed acid Fermentation Butylene Glycol Pathway Butyric Acid Fermentation CARBOHYDRATE UTILIZATION AND LACTOSE FERMENTATION – ability of microorganisms to use various sugars (carbohydrates) for growth is an integral part of many diagnostic identification – fermentation of the sugar is usually detected by acid production and a concomitant change of color resulting from a pH indicator present in the culture medium – Bacteria generally ferment glucose preferentially over other sugars, so PAGE 15 Viray, M.S.Y. - 3B ˚ ༘♡ ⋆｡˚ ❀