Bacteriology- Lesson 1-9 PDF

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LESSON 1 AND 2 : HISTORY AND MICROBIAL TAXONOMY 2 Five kingdoms a.) Kingdom Animalia b.) Kingdom Plantae c.) Kingdom Protista d.) Kingdom Monera e.) Kingdom Bacteria Microbial Taxonomy Giving names, classification of organism Allows to use a common label for every organism studied within the multitu...

Description

2.1 Five kingdoms a.) Kingdom Animalia b.) Kingdom Plantae c.) Kingdom Protista d.) Kingdom Monera e.) Kingdom Bacteria

LESSON 1 AND 2 : HISTORY AND MICROBIAL TAXONOMY

Microbial Taxonomy -

Giving names, classification of organism Allows to use a common label for every organism studied within the multitude of biologic disciplines

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Three Distinct but Highly Interrelated Disciplines I.

Classification Organization of microorganisms into groups or taxa based on similar morphologic, physiologic, and genetic traits

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1.

Hierarchal Levels

Kingdom Phylum / Division Class Order Family Genus Species (Basic Unit) Strain / Variant / Subspecies

2.

Monera / Eubacteria Proteobacteria Gamma-proteobacteria Proteobacteriales Enterobacteriaceae Escherichia Coli Strain 1, 2, 3

Robert Whittaker - Scientist that classify living things into five kingdoms

3.

4.

Classification according to Cellular Organization a.) Unicellular vs Multicellular b.) Prokaryotic vs Eukaryotic Classification according to Nutrition a.) Ingestion b.) Absorption c.) Photosynthesis

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Because genus and species are the groups commonly used by the microbiologist, the discussion of rules governing microbial nomenclature is limited to these two taxa - Genus and species both derived from Greek and Latin words - Species abbreviated as “sp.” (singular) or “spp.” (plural)

III. -

Identification Process by which microorganism’s key features are delineated or described 1. -

5.

Carl Woese - Classify microorganism in three domains 5.1 Domain Bacteria / Prokaryotea a.) Eubacteria b.) Cyanobacteria (bluegreen algae) 5.2 Domain Eukaryota a.) Protozoa b.) Blue / brown algae 5.3 Domain Archaea II. Nomenclature - Naming microorganisms according to established rules and guidelines

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Two General Categories 1.1 Phenotypic Features of microorganisms that are readily observable a.) Size b.) Shape c.) Arrangement d.) Biochemical characteristics: catalase, oxidase, fermenter, etc. 1.2 Genotypic Based on features beyond the genetic level and include both readily observable characteristics and characteristics that may require extensive analytic procedures to be detected Relates to the organism’s genetic makeup

a.) DNA based composition ratio b.) Nucleic acid-base sequence Phylogeny - Study of historical origin or evolutionary relatedness of microorganisms International Journal of Systematic and Evolutionary Microbiology (IJSEM)

1862

1867 1876

1881 1882

1882 1884

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Formerly International Journal Systematic Bacteriology (IJSB), is the official journal of record for novel prokaryotic taxa Authenticates the newly found microorganisms

Anton Van Leeuwenhoek - Father of microbiology Selected Significant Agents and Dates of Discovery Perio d 1665 1667 1796 1850

1861

Developmental Notes Publication of the first description of microbes Observation of “little animals” Smallpox vaccination – first scientific validation Advocating hand washing in prevention of the spread of disease Spontaneous generation

Key Scientist/s Robert Hooke Anton Van Leeuwenhoek Edward Jenner Ignaz Semmelweis Louis Pasteur

disproved Publication of the paper supporting the germ theory of disease Practice of antiseptic surgery Discovery of Bacillus anthracis which became the first proof of germ theory Utilization of solid culture media for bacterial growth Outlined Koch’s postulate Discovered Mycobacterium tuberculosis Development of acid-fast stain Gram stain developed

1885 1887

First rabies vaccination Invention of petri dish

1892

Discovery of viruses

1893

Zoonosis – first described

1899

Viral dependence on living host cells for reproduction recognize Proof the mosquitoes carry the agent of yellow fever Discovered the cure for syphilis Discovery of penicillin

1900 1910 1928 1953 1977 1983 1995

Proposed and built the DNA model Development of the DNA sequencing method Invention of the polymerase chain reaction (PCR) Publication of the first microbial genomic structure

Discovery of Bacteria

Louis Pasteur

Period 1876 1879 1880

1892

Bacteria Bacillus anthracis Neisseria gonorrhoeae Staphylococcus, Streptococcus, Pneumococcus Mycobacterium tuberculosis Corynebacterium diphtheria Clostridium perfringes

1894

Yersinia pestis

1900

Coccidiodes immitis

1903

Leishmania donovani

1905

Treponema pallidum

1918 1977

Brucella abortus Leigonella pneumophilla Prions

Joseph Lister Robert Koch

1882 1883

Robert Koch Robert Koch

Paul Erlich Hans Christian Gram Louis Pasteur Richard J. Petri Dmitri Iosifovich Ivanoski T. Smith, F.I. Kilbourne Martinus Bijerinck Walter Reed Paul Erlich Alexander Fleming J. Watson, F. Crick W. Gilbert, F. Sanger Kary Mulis The Institute for Genomic Research

1982

Discoverer Robert Koch Albert Neisser Louis Pasteur

Robert Koch Edward Klebs, Fredrick Loeffler William Welch, George Nutal Emile John Yersin, S. Kitasato W. Ophuls, H.C. Moffett William Leishmann Fritz R. Schandinn, Erich Hoffman Alice Evans Joseph McDade, Charles Shepard Stanely Prusiner

LESSON 3: CELL STRUCTURE General Differences of Prokaryotic from Eukaryotic Nucleus

Cell Wall

Prokaryotic Absent (Counterpart is Nucleoid, found in mesosomes) Present (except Mycoplasma spp and Ureaplasma spp)

Eukaryotic Present

Absent (except fungi with composition of cellulose, chitin, mannan, glucan)

Cytoplasmic Membrane

Present (made up of phospholipids)

Present (made up of steroids)

Absent

Present

Absent

Present

Absent

Present

Absent

Present

70s: 50s, 30s Electron transport in cell membrane Free Ribosome

80s: 60s, 40s Mitochondria

Asexual (Mitosis)

Asexual (Mitosis), Sexual (Meiosis)

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Organelles

4.

Endoplasmic Reticulum Mitochondria Golgi Complex Lysosome Ribosome Size Site of Energy Production Site of Protein Synthesis Means of Reproduction

Rough ER

Three General Part of Bacteria I. II. III. Cell Wall I. 1.

Cell Wall Internal Parts External Parts

Functions

For Protection, Rigidity, and Give Shape to the Cell 2. Responsible for Pathogenicity of the Cell - Ability to carry or cause diseases 3. Endotoxin A (Pathogenic Property) - Component of bacteria that causes diseases

5. -

Stimulate production of fever Commonly on gram negative May cause shock syndrome Responsible for the Antigenic Property of the Cell - Any foreign substance that once introduce in our body, can stimulate the production of antibody - Binding site of antibody is cell wall Staining Properties of the Cell Artificial procedure 5.1 Two general types

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Doesn’t need to put colour / dye to the bacterial structure, but give a dark background against it  Negative / Relief stain - Examples: India ink Borris ink Nigrossin stain II.

1.

a.) Direct Staining - Gives colour directly to the bacterial structure  Simple stain - Colours bacterial structure using one type of dye - Examples: Methylene blue Crystal violet  Differential stain - Use to differentiate two groups of bacteria using two types of dye - Examples: Acid-Fast Bacilli staining Gram staining  Special stain - It colours the special structure of the bacteria b.) Indirect Staining 2.

Anatomy

Gram Positive 1.1 Surface protein 1.2 Peptidoglycan layer / Murein layer - Backbone of the cell wall - Site where antibiotics are being directed - Large in size (Thick) - Penicillin (antibiotic) is effective in gram positive a.) Two sub unit of protein  n-acetyl-d-muramic acid  n-acetyl-d-glucosamine 1.3 Teichoic acid - Regulates magnesium, salts, etc. - Passageway of molecules from outside, going inside - Anchored to peptidoglycan layer - Insoluble to alcohol 1.4 Lipoteichoic acid - Passageway of molecules from outside, going inside - Anchored all the way down to the plasma membrane or cell wall Gram Negative

2.1 Surface protein 2.2 Peptidoclglycan layer / Murein layer - Small in size (Thin) - Penicillin (antibiotic) is not strong in gram negative a.) Two sub unit of protein  n-acetyl-d-muramic acid  n-acetyl-d-glucosamine 2.3 Outer membrane a.) Porins - Counterpart of gram negative - Where molecular pass through inside the cell such as antibiotics, nutrients, etc. b.) Phospholipids c.) Lipopolysaccharide (LPS) - Soluble in alcohol - Largest part of outer membrane - Responsible for antigenic property  Somatic antigen / surface antigen - Bind the antibody  Endotoxin A / Lipid A - Responsible for fever and shock syndrome 2.4 Periplasmic space / periplasm - Exclusive for gram negative - Contains enzymes and protein for metabolic pathway III.

Staining - Procedure for artificially colouring the cell using different dyes / reagent

1. 2.

- With Steam (Mordant)  Pappenheim’s Method - Uses Rosalic Acid instead of Acid Alcohol (Decolourizer) - Used to differentiate Mycobacterium tuberculosis from Mycobacterium smegmatis - M. tuberculosis (+) – red - M. smegmatis (-) – blue  Baumgarten Method - Uses Alcoholic Fuchsin instead of Carbol Fuchsin - Used to differentiate Mycobacterium leprae from Mycobacterium tuberculosis - M. leprae (+) – red - M. tuberculosis (-) – blue

Simple Stain Differential Stain 2.1 Gram stain a.) Crystal Violet - Primary stain - 1 min b.) Gram’s Iodine - Mordant - 2 mins c.) Acetone Alcohol - Decolourizer - 30 secs d.) Safranin - Secondary stain - 1 min

Crystal Violet Gram’s Iodine Acetone Alcohol Safranin

Gram (+) Violet Violet Violet Violet

Gram (-) Violet Violet Colourless Red / Pink

2.2 Acid-Fast Bacilli stain a.) Carbol Fuchsin (5 minutes) - Primary stain - 5 mins b.) Acid Alcohol - Decolourizer - 1 – 2 mins / variable c.) Methylene Blue - Secondary stain - 1 min  Methods of Acid-Fast Bacilli  Kinyoun Method (Cold Method) - With Tergitol # 7 ( Mordant)  Ziehl – Neelsen Method (Hot Method)

3.

Special Stain 3.1 Capsular stain a.) Hiss b.) Tyler c.) Muir d.) Welch e.) Novelli f.) Anthony 3.2 Metachromatic stain a.) Albert b.) Neisser c.) LAMB d.) Lindergren e.) Ljubinsky 3.3 Flagellar stain a.) Leiffson b.) Gray Method

4.

c.) Silver Technique d.) Caesares Gil e.) Fisher & Conn 3.4 Endospore stain a.) Fulton b.) Schaeffer c.) Dorner d.) Wirtz e.) Conklin Indirect 4.1 Negative / Direct / Relief a.) India Ink stain b.) Nigrossin stain

2.

3. Mycolic Acid / Hydroxymethox Acid - Specific part / bacterial structure for acidfast bacilli stain - Hard to stain and difficult to decolourize - Extra part of cell wall that is present in two genera of gram positive - Examples: Mycobacterium spp. Nocardia spp. Three Things to Differentiate Gram Positive from Gram Negative Bacteria I. II. III.

Gram Reaction Shape Arrangement

I. 1.

Gram Staining Rules To Identify Bacteria All Cocci are Gram Positive Except

a.) Neisseria spp. b.) Monaxella spp. c.) Viellonella spp. All Bacilli are Gram Negative Except a.) Bacillus spp. b.) Corynebacterium spp. c.) Mycobacterium spp. d.) Clostridium spp. e.) Nocardia spp. f.) Erysipelothrix spp. g.) Actinomyces spp. h.) Listeria spp. i.) Lactobacillus spp. All Spiral are Difficult to Stain but are Gram Negative a.) Examples: Treponema spp. Leptospira spp.

Internal Structure I. Function 1. Responsible for the Permeability of the Organism 2. Responsible for the Viability of the Organism 3. Site of Energy Production (Metabolic Reaction) II. Parts 1. Cell Wall - Gives protection to the cell 2. Plasma Membrane - Part lying next to the cell wall inward

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Semi-permeable membrane that is made up of lipoproteins 2.1 Functions - Site of enzymatic activity of the cell - Site of attachment of naked chromosomes (DNA and RNA) - Site of septum formation during cell division 3. Free Ribosomes - Responsible for protein synthesis 3.1 Protein formation 4. Metachromatic Granules - Food reserve - Source of energy - Examples: Corynebacterium diphtheriae - Called as Babes-Ernst Granules Mycobacterium tuberculosis - Contains Much Granules Nocardia braciliensis - Contains Sulfur Granules Yersinia pestis - Bipolar bodies 5. Endospores - Used to resist extreme condition such as when it is exposed to high temperature, it is not easily destroyed - Limited to gram positive bacilli - Examples: Clostridium spp. Bacillus spp. 5.1 Three types of spores

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6.

Based on location a.) Terminal Spore - Located at the end of the cell - Example: Clostridium tetani b.) Sub-terminal Spore - Located near the end of the cell - Example: Clostridium botulinum (lives in the absence of oxygen / anaerobic) c.) Central Spore - Located at the center or equatorial spore - Examples: Bacillus anthracis Bacillus subtilis

Plasmid Extra chromosomal, circular pieces of DNA - Often carries virulence genes and antibiotic resistance genes - Product of disintegration of DNA -

External Structure I. Parts 1. Capsule - Not all bacteria are capsulated - Thick, shinny, gelatinous layer that surround some of bacteria

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Serves as organ of attachment on the tissue cell or host cell - Colonies of capsulated bacteria is Mucoid 1.1 Functions a.) Virulence factor / inhibit phagocytosis - Degree of pathogenicity b.) Serologic specificity / antigenic property - Binding site of antibody (in vitro) 1.2 Several mechanism of immune system for us to be protected a.) Neutrophil - Acts of phagocytes 1.3 Examples: Klebsiella pneumoniae Streptococcus pnuemoniae Neisseria meningitides Haemophilus influenzae 2. Flagella / Flagellum - Slender whip like structure made up of protein called Flagellin - Refers to trichous 2.1 Functions - For locomotion or movement

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For antigenic property (binding site of antibody) 2.2 Several type of flagellated bacteria base on the absence or number of flagella a.) Atrichous - Absence of flagella or flagellum - Examples: Shigella spp. (gram negative bacilli) Pneumoniae spp. (gram negative bacilli) b.) Monotrichous - One flagellum - Examples: Vibrio cholerae (gram negative bacilli) Pseudomonas aeruginosa c.) Lopotrichous - Several, group or tuft of flagella - Example: Spirillum minus (gram negative bacilli) d.) Amphitrichous - Has flagellum at both ends - Example: Alkaligenes faecalis (gram negative bacilli)

e.) Peritrichous - Numerous flagella surround the cell - Example: Escherichia coli 3. Pili / Pilus / Fimbrae - Tiny hair like structure that is present among flagellated organisms - Except Neisseria gonorrhoeae (has no pili flagella) 3.1 Two types of pili a.) Sex pili - Utilize for genetic transfer during bacterial conjugation b.) Common pili - Serves as virulence factor, inhibits phagocytosis - Serves as organ of attachment in the tissue cell or host cell 4. Axial Fibril / Filaments - Cork-screw like movement - Majority are present for spiral bacteria - Made up of protein - Examples: Leptosira spp. Treponema spp.

LESSON 4: BACTERIAL GROWTH -

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Grows in numbers, unlike human that grows by size and length

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organic

d.) Hydrogen – 8%

Undergo cell division

I.

Ammonia, nitrides, compound, free nitrogen

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Requirements

Water, organic compound, hydrogen in the air

free

e.) Phosphorus – 3%

1. Nutritional Requirements 1.1 Macronutrients (larger amount) a.) Carbon – 50% 

Types of bacteria according to the sources of carbon

f.)

Inorganic phosphates, sulfates, hydrogen sulfite, organic sulfur Sulfur – 1%

1.2 Micronutrients (smaller amount) a.) Potassium – 1%

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Autotrophic / litotrophic b.) Magnesium – 0.5% -

Microorganism derived their carbon in carbon dioxide

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Heterotrophic / organotrophic -

Microorganism derived carbon from organic substances (decaying plants, animal, human)

b.) Oxygen – 20% -

Derived from water, organic compound, carbon dioxide, free oxygen from the air

c.) Nitrogen – 14%

c.) Iron – 0.5% d.) Calcium – 0.5% (salts) e.) Manganese – less than 0.5% (smaller amount) f.)

Zinc – less than 0.5% (smaller amount)

g.) Molybdenum – less than 0.5% (smaller amount)

2. Environmental Requirements 2.1 Gas

a.) Obligate aerobe / strict aerobe -

Propionobacterium spp. (Propionobacterium acnes)

f.)

Lives in the presence of oxygen only 

Two enzymes neutralize oxidative toxic product

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Aerobe but they are enhance to grow in the presence of small amount of carbon dioxide (5-10%)

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Examples:

c.) Microaerophilic -

 Catalase

Requires small amount of oxygen (210%), use catalase of the foselate infect to neutralize toxic substances

HACEK group (common cause endocarditis)

 Superoxide Dismutase -

Capnophilic

Examples:

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Haemophilus

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Aggregatibacter

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Cardiobacterium

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Eikenella

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Kingella

Examples: Mycobacterium tuberculosis Brucella abortus Bordatella pertussis Leptospira interogans Neiserria gonorrhoeae Neisseria meningitides Pseudomonas aeruginosa

Campylobacter spp. Helicobacter spp. d.) Facultative anaerobe -

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Aerobe but in the absence of oxygen they can survive

3. Temperature Requirement

Examples:

b.) Obligate anaerobe / strict anaerobe -

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Do not require oxygen (lethal to oxygen), do not have catalase and superoxide dismutase to neutralize oxygen Examples: Clostridium tetani Clostridium perfringens Clostridium botulinum Clostridium difficile Bacteroides spp. (gram negative bacilli) Bacteroides fragilis (pre-dominant normal flora in large intestine)

Enterobacteriaceae Micrococcaceae Streptococcaceae e.) Aerotolerant anaerobe -

Aerobe but tolerates the presence of oxygen but does not require it for growth

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Instead, use fermentation to survive

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Example:

Temperature in order to grow 3.1 Cryophiles - Also called as “psychrophiles” - Requires low temperature, temperature below 20 ˚C

optimum

- Majority of organism that cause food spoilage 3.2 Mesophiles Enterococcus faecalis

Majority of clinically significant pathogen that cause disease to human

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Optimum temperature requirement is 35 – 37 ˚C

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Example:

High concentration outside than inside (hypertonic)

1 – 42 ˚C 3.3 Thermophiles -

4. pH Requirement

Group of microorganism that grow at high temperature

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