MICROBIOLOGY AND PARASITOLOGY 4.
Parasitology
5. Phycology
by: RV De la Cruz
6. Immunology
CHAPTER 1 Microbiology is important because:
THE SCIENCE OF MICROBIOLOGY 1. It has an impact in our daily lives;
What is microbiology? 2. Some microorganisms are vital in biotechnology and various
- “Mikros” small industries;
- “Bios” life, “logia”, or logos study of 3. Some microorganisms are key sources of antimicrobialagents.
a.) Normal or Indigenous Flora
- dwell within our body
b.) Pathogens (non-pathogenic and paghogenic)
- disease-causing agents
4. Some microorganisms function as saprophytes (decomposers);
5. It advanced understanding of disease-causing microorganisms and
Prokaryotes improved disease control; and
- bacteria 6. The return of diseases, biological warfare risks, and antibiotic
- cyanobacteria (primitive, stromatolites, accumulation – rock like resistance highlight microbiology's importance.
structure.)
- archaeans EVOLUTION OF MICROBIOLOGY
a.) Pyrolobos Fumarii ● 3180 BC
- acid tolerant (0pH) - “plague” in Egypt (Epidemic)
b.) Picrophilus ● 1122 BC
- hyperthermophytes (115*C) - small-pox like disease outbreak
c.) Methanogens ● Mid 1600s
- chemotherapy - discovery of microscope
- CO2 ● Robert Hooke - cell theory
1670s
FIELDS OF STUDY ● Anton Van Leeuwenhoek
. Bacteriology - single lense microscope
2. Virology -“animalcules”
3. Mycology -“Father of Microbiology”
Middle and Late 1800s -> All cocci - gram-positive, exc. Neisseria, Veilonella, and
● Louis Pasteur Branhamella
-Germ Theory, -> All bacilli - gram-negative, exc.
- Pastuerization, Corynebacterium, Clostridium, Bacillus, and Mycobacterium
- Aerobes and Anaerobes
Golden Age of Microbiology (late 1800s and 1st decade of 1900s)
● Edward Jenner
- small pox vaccine
● Joseph Lister
- medical procedures (aseptic surgery)
After World War II (Intro of Antibiotics)
● Paul Ehrlich
- salvarson (chemotherapy) 2. Acid-fast Stain
● Alexander Fleming ● For high lipid content in the cell wall (Gram stain x)
- Penicillin a.) Ziehl-Neelsen stain
Early 20th Century - "hot method"
- study of bacteria - steam bathing the smear after the primary dye
1930s b.) Kinyoun stain
- development of electron icroscope - "cold method"
- no heat after the primary stain
STAINING
1. SIMPLE STAINS
- single dye (aquaseos, alcohol-based)
- Basic dyes: safranin, methylene blue, or crystal violet
2. DIFFERENTIAL STAINS
- differentiate bacterial groups based on cell wall properties
Two (2) Common Differential Staining Procedures
1. Gram Stain
● Gram-positive and gram-negative bacteria 3. Special Stain
● General Rule: ● Demonstrate specific structures in a bacterial cell
Example: - Liquefiable and non-liquefiable forms
● Metachromatic granules - LAMB (Loeffler AlkalineMethylene Blue) 2. Chemical Composition
● Capsule/slime layer - Hiss stain a.) Synthetic Media
● Cell wall - Dyer stain - Chemically defined
● Flagella - Fischer-Conn stain - Known composition
● Spores - Dorner and Shaeffer-Fulton stain - Simple or complex
● Capsule of fungus Cryptococcus neoformans - India ink or b.) Non-synthetic Media
nigrosine - Undefined components like animal, plant, or
yeast extracts
- Support fastidious organisms.
3. Functional Type
General Purpose Media
- Broadly support microbial growth
Ex. Peptone water, nutrient broth, & nutrient agar
Enrichment Media
CULTURE MEDIA - Complex substances (blood, serum, or SGF)
- An aqueous solution containing essential nutrients for organism - Enhance growth of specific microbes (nutritionally exacting)
growth. Commonly used EM
Blood Agar
Three (3) Primary Levels - General nutrients
1. Physical State - 5%-10% blood
a.) Liquid media - Gram-positive bacteria - "exotoxins" - hemolyze RBC
- Nutrient solutions that remain liquid above Hemolysis Types:
freezing i. Beta hemolysis - complete hemolysis
- Growth & fermentation ii.. Alpha hemolysis - partial hemolysis
b.) Semi-solid media ili. Gamma hemolysis - no hemolysis
- ≤0.5% agar Chocolate Agar
- microaerophilic motility studies of bacteria - Culture of fastidious organisms (hemophilis)
c.) Solid media
- 1.5%-2% agar agar
- Isolating organisms and colony studies
Selective Media Eukaryotes
- Encourage specific microbes while inhibiting others. - With true nucleus
a. Thayer Martin agar
- Isolates Neisseria - trimethoprim, nystatin, vancomycin,
colistin
b. Mannitol Salt agar
- Isolates Staphylococcus aureus - 10% NaCl
c. MacConkey's agar
- Selective: gram-negative Enterobacteriacee using bile salts;
also differential.
d. Löwenstein Jensen medium
- recovers Mycobacterium tuberculosis using malachite green.
e. Sabouraud's dextrose agar
- Isolates fungi
Differential Media
- Differences among microorganisms (e.g., colony size or color)
Examples: MacConkey's agar, Triple Sugar Iron agar
MEDICALLY IMPORTANT MICROORGANISMS
1. Viruses
Transport Media
- Acellular
- Preserve specimens during transport and prevent contamination
- Capsid - outer surface (capsomeres)
Examples: Cary-Blair medium, Pike's medium
- DNA or RNA
- Obligate intracellular parasites
Anaerobic Media
- Support oxygen-sensitive organisms with nutrients and reducing
agents
Examples: Chopped cooked meat broth, thioglycolate broth
PROKARYOTIC AND EUKARYOTIC CELLS
Prokaryotes
- No true nucleus
2. Bacteriophages 6. Algae
- Special virus - infect bacteria - Eukaryotic
Roles: - Outer surface - cellulose
-Acquisition of bacterial virulence factors - Plant-like
Transfer genetic material between bacteria - DNA and RNA
3. Bacteria - No true roots, stems, and leaves
- Outer covering - "cell wall" - peptidoglycan
- DNA and RNA
- Nucleoid, smaller ribosomes, no mitochondria
4. Fungi
- Eukaryotic
- Outer surface - "chitin"
- Cell membrane - ergosterol
- DNA and RNA
- True nucleus, mitochondria a.) Diatoms
- Unicellular algae
- Cell wall - SiO2
b.)Dinoflagellates
-Unicellular algae
- Phytoplankton group - Contributes oxygen "Red
tide"
BACTERIA MORPHOLOGY
5. Protozoa
- Eukaryotic
- Outer surface - pellicle
- Unicellular - "binary fission"
- DNA and RNA
Morphologic forms:
1. Cysts
2. Trophozoites
SPECIAL COMPONENTS OF GRAM-POSITIVE CELL WALL
1. TEICHOIC ACIDS
- major surface antigens of gram-positive organisms
- elicit antibody response attachment of the organum to the
host cell
- provide tensil utrength: to gram-positive b. cell wall
2. POLYSACCHARIDES
- include neutral sugars such as mannose, arabinose,
rhamnose, and glucosamine
- includes some acidie sugars such as gluctronic acid and
mannuronic acid.
SPECIAL COMPONENTS OF GRAM- NEGATIVE CELL WALL
1. OUTER MEMBRANE
- bi-layered trocture where the inner leaflet is composed of a
lipopolysaccharide (LPS)
- has a special protein channels that allow the passage of small
or low-molecular weight hydrophillic substances (sugar, a.
acid)
ENVELOPE STRUCTURES
- LPS has a complex glycolipid called Lipid A (endotoxin
1. GLYCOCALYX
activity)
- outermost covering
- LIPID A - located in the outer leaflet of the outer membrane
- gelatinous
- inner core is a polysaccharide made up of units (O antigen)
- capsule - strongly attached
2. LIPOPROTEIN
- slime layer - loosely attached
- anchor the outer membrane to the peptidoglycan layer and
2. CELL WALL
stabilizes the outer membrane
- “Murein sacculus”
3. PERIPLASMIC SPACE
- peptidoglycan (murien or mucopeptide)
- fluid-filled space between outer membrane and inner plasma
- shape and support
membrane
- protects the bacteria from osmotoc damage on important role
- contains enzymes for the breakdown, nontransferable
in cell division
molecules into transferable
- detoxify and inactivate antibiotics
- function for adherence the cell surface (common pili) or attachment
ACID-FAST CELL WALL another bacterium during a form of bacterial gene exchange called
- possess an outer layer that is lipid rich conjugation (sex pili)
- composed of large amounts of waxes that are known as 3. AXIAL FILAMENTS
myolic acid - also called as endoflagella, found in spirochetes (e.g. Treponema
- inner layer of cell wall is made up of peptidoglycan pallidum causing Syphilis)
- hydrophobic, cannot stain using the regents used in gram - composed of bundles of fibrils
staining (protect from harsh chemicals, such as strong acids - arise from the end of bacterial cells and spiral around the cell
and detergents.) - rotate producing movement of the outer sheath of the spirochetes,
propelling them forward
PROTECTING STRUCTURES
1. FLAGELLA
- thread like structures made up of protein sub unit called
flagellin
- project from the capsule and our organs for motility
CLASSIFIED INTO 4 TYPES
a. MONOTRICHOUS
- single polar flagellum
b. LEPHOTRICHOUS
- a tuft of flagella at one end of the bacteria
c. AMPHITRICHOUS
- flagella of both ends of the bacterium
d. PERITRICHOUS BACTERIAL GROWTH REQUIREMENTS
- flagella all around the bacteria - orderly and organized increase in the sum of all components of the
e. ANTRICHOUS organism
- bacteria without flagella
2. PILI or FIMBRIAE NUTRITIONAL REQUIREMENTS
- rigid surface appendages found in many gram-negative bacteria 1. Carbon
- fine and short in comparison with flagella - structural backbone
- Pilins, structural protein subunits Based on carbon source:
- Pili may also function for motility ● Autotrophs (lithotrophs)
● Heterotrophs (Organotrophs)
● Both: Either light or IS oxidation Pasteurization - 62.8*C
2. Nitrogen, Sulfur, Phosphorus Human body - 37*C
- synthesis of cellular materials Refrigerator - 4*C
3. Inorganic lons
- Mg, K, Ca, Fe, & trace elements (e.g., Mn, Zn, Cu, Co) pH
1. Alkalophiles: Thrive at pH 8.4-9.0.
PHYSICAL REQUIREMENTS 2. Neutrophiles: Optimal at pH 6.5-7.5
1. Moisture/Water 3. Acidophiles: Grow below pH 6.0.
● Biochemical reactions
● Nutrient dissolution and transport OSMOTIC PRESSURE
● Maintaining cell structure - salt concentration
● Hydration of cellular components - normal cytoplasmic salt concentration - 1%
● Metabolism and energy production - Tonicity - Lysis, tysis, & iso-
● Thermal regulation 1. Halophiles - high salt
● Adaptation to growth conditions 2. Osmophiles - high osmotic pressure
2. Oxygen
● Aerobic bacteria - cellular respiration NORMAL FLORA OF THE HUMAN BODY
● Final electron acceptor - ETC Microbial Ecology
- relationship: microorganisms & environment
CLASSIFICATION BASED ON OXYGEN REQUIREMENTS Types of Flora
1. Aerobes: Require oxygen for growth. (M. tuberculosis) 1. Resident - permanent
2. Obligate Anaerobes: Die in oxygen due to lack of detoxifying 2. Transient- temporary; environment-derived
enzymes. (Clostrudium botulinum) - Inhibit pathogenic organisms
3. Facultative: Grow in both oxygen and no oxygen. (E. coli) - Protect external-contact organs from invasions
4. Microaerophiles: Prefer low oxygen levels. (Helicobacter pylori) - Synthesize essential vitamins
5. Capnophiles: Thrive with extra carbon dioxide. (Neisseria Other Benefits:
gonorrhoeae) - Prevent pathogens from attaching to tissues
- Digest food through enzymes
TEMPERATURE - Metabolize steroids
1. Thermophiles. Above 40 °C
2. Mesophiles. Optimal at 20 °C-40 °C
3. Psychrophiles. Optimal at 10 °C-20 °C
NORMAL FLORA OF THE SKIN Mouth & Respiratory Tract
Factors eliminating non-resident flora of the skin: ● Tongue & buccal mucosa
1. Lysozyme in skin - Streptococcus viridans (S. mutans, S. milleri, S. salivarius, & S.
2. Acidic pH from sweat sanguis)
3. Fatty acids in sebaceous secretions
4. Skin sloughing off Pharynx & Trachea
- Haemophilus influenzae
Three Regions of the Skin - Streptococcus pneumoniae
1. Axilla, perineum, and toe webs - Neisseria meningitidis
Higher: - Mycoplasma
- Moisture levels
- Temperature Upper Respiratory Tract
- Surface Lipids - Neisseria meningitidis
- More microorganisms - Corynebacterium diphtheriae
- Predominant inhabitants: gram- negative bacilli - Bordetella pertussis
2. Hand, face, and trunk
3. Upper arms and legs Conjunctiva
● Lysozyme (tears) - interferes with cell wall synthesis
Dry Sites - Neisseria, Moraxella, & Corynebacterium
● Diverse flora - environmental exposure - Staphylococci & Streptococci
● Staphylococcus epidermidis
● Staphylococcus hominis Digestive Tract
- Esophagus: transient mouth flora
- Stomach: acidic - scanty - Helicobacter pylori
- Small intestine: scanty flora - constant peristaltic movement -
Streptococci, Lactobacilli, & Bacteroides
- Large intestine: greater flora
- Colon: 95 - 99% anaerobes: Bacteroides fragilis (most common),
Bifidobacterium/Lactobacillus bifidum (breastfed infants),
Eubacterium, Peptostreptococcus, and Clostridium
- Bottle-fed infants: Lactobacillus acidophilus
- Colon: 1 - 4% flora-facultative (E. coli & Enterobacteriaceae)
Importance of Intestinal Flora
● Synthesize vitamins B and K
● Convert bile into bile acids
● Compete with transient flora for nutrients
● Prevent colonization by transient flora
● Produce toxic byproducts harmful to transient flora
Genitourinary Tract
- UT (sterile) - above distal 1 cm of the urethra
- Mycobacterium smegmatis - secretions
- Penile urethra - Gardnerella vaginalis, Bacteroides, and alpha
streptococci
- Female urethra - sterile or with Staphylococcus epidermidis
Vaginal Flora
- Infants: Lactobacillus spp.
- 1 month - puberty: Glycogen cessation (higher pH ~ 7.0)
Staphylococcus epidermidis, diphtheroids, and E. coli.
- Puberty onset: Glycogen resumption (acidic) Lactobacillus
acidophilus, peptostreptococci, streptococci, staphylococci.
streptococci, corynebacteria, Bacteroides,
- After menopause: Higher pH
Flora - the same as pre-puberty
Fungi - Toulopsis & Candida (10-30%)
