Microbiology Unit 2-3: Bacteria

1. Microbiology and Parasitology
UNIT 2/3: BACTERIA

2. Microbiology
Virus: Ebola
Algae: Algal bloom
Fungi: Mushroom
Bacteria: Salmonella
ENEMY OR… …FRIEND

3. Test your memory
A. True or False
1. Microbiology is the study of disease-causing microorganisms
2. Microbes are involved in the production of food and drinks
3. Fungi are a type of acellular microorganisms
4. The majority of our microflora are “microbial allies”
5. Microbial intoxication is a type of infectious disease
B. Multiple choice
1. Who invented the microscope
a) Pasteur; b) van Leeuwenhoek; c) Fleming
2. Koch’s Postulates proves that
a) Microbes can be isolated from an infected animal; b) Microbes are always found
in the same animal species; c) Specific microbes cause particular diseases
3. Ignaz Semmelweis is reffered to as
a) Father of Microbiology; b) Father of Handwashing; c) Father of Hygiene

4. Variety of Microbes
• Categories
– Acellar: Viroids, Virus, Prion
– Prokaryotes (cellular without nucleus): Bacteria, Archaea, Cyanobacteria
– Eukaryotes (cellular with nucleus): Algae, Fungi, Protozoa
Microbes
Acellular
Viroids
Prions
Virusus
Cellular
Prokaryotes
Archaea
Bacteria
Cyanobacteria
Eukaryotes
Algae
Fungi
Protozoa

5. Today
• Classification
– Taxonomy
– Morphology
• Structure of cells
• (Endo)spores
• Cell reproduction
– Binary fission
– Generation time
• Bacterial growth
– Encouraging growth
– Inhibiting growth
• Staining
• Other characteristics
• Unique bacteria
– Rickettsia

6. Microbial classification
• Millions of species on Earth
– Bring order in the chaos: system to classify organisms
– Taxonomy = science of classification of species
– Classification, nomenclature and identification (complicated system)
– Scientist still debate about the system
Burton’s Microbiology: Chapter 3,4
23
32
77
182
871
5007
Only 1% known?
3 or 5?

7. Classification by shape
• Categories based on shape of their cells (simpler system)
– Spherical: cocci
– Rod-shaped: bacilli
– Short rods/elongated spheres: coccobacilli
– Curved/Spiral-shaped: spiralla (spirochetes)
– Various shapes: Pleomorphic (cell wall-deficient bacteria)
BacilliCocci Curved/spiral
Burton’s Microbiology: Chapter 4

8. Morphology of cocci
Burton’s Microbiology: Chapter 4
Chains
Clusters
• Bacteria aggregate: morphological arrangement

9. Morphology of bacilli
Chains: streptobacilli
Side-by-side: DiphtheroidsPairs: Diplobacilli
Short rods: Coccobacilli
Burton’s Microbiology: Chapter 4

10. Morphology of spirochetes
Less tightly coiledTightly coiled
highly flexible cell wall
able to move through tissue
Treponema pallidum:
Syphilis
Borrelia:
Lyme disease
Burton’s Microbiology: Chapter 4

11. Size of bacteria
• Spherical bacteria: Coccus
– Typical diameter: 1 μm (1 million micrometers = 1 meter)
– Seven cocci fit the diameter of human red blood cell
• Rod-shaped bacteria: Bacillus
– Typical length: 3 μm
– Typical width: 1 μm
Burton’s Microbiology: Chapter 2

12. Structure: Advanced microscopes
(Optical) Light Microscope
x1000
Scanning Electron
Microscope (SEM)
x10.000
Transmission Electron
Microscope (TEM)
x100.000
1 μm
“STAPHYLOCOCCCUS AUREUS”
Burton’s Microbiology: Chapter 2,3
“Study the structure of cells in detail”

13. Cell structure
Burton’s Microbiology: Chapter 3
• Cell
– Fundamental unit of a living organism
– Basic characteristics of life
• Cell = tiny functional factory
– Metabolism: all chemical reactions in a
cell
– Use food (nutrients) to produce energy
– Grow and reproduce
– Responds to environment
– Can mutate (change genetically)
• Study of cell structure
– Cytology

14. Procaryotic cell
• Bacteria: Procaryotic cell
– Cell envelope
• Cell membrane
• Cell wall
• Slime layers and Capsules
– Chromosome
– Cytoplasm
• Cytoplasmic Particles
– Flagella and Pili
– (Endo)spores
Burton’s Microbiology: Chapter 3
Difference with eucaryotes:
10x smaller than eucaryotic cell
Less complex cell structure: no nucleus

15. Cell envelope
• Cell membrane
– Flexible thin skin that encloses the cells
interior
– Controls in/out transport of substances
(via proteins in membrane)
• Cell wall
– Provide rigidity, strength and protection
– Different structure for Gram+ and Gram-
bacteria
– Cell wall-deficient (CWD) bacteria: cause
chronic diseases, e.g. Lyme
• Glycocalyx (external layer)
– Slime layer: slide along solid surfaces
– Capsule: protection (white blood cells)
Burton’s Microbiology: Chapter 3,4

16. Bacterial genome
Burton’s Microbiology: Chapter 3
• Genome carries genetic
information (3000 genes)
– Chromosome
• Duplicating itself
• Guiding cell division
• Directing cellular activity
• Embedded in cytoplasm
– Plasmid

17. Nutrient matrix
• Cytoplasm
– Semifluid nutrient matrix of cell
• Cytoplasmic particles
– Complex mixture of all materials required for metabolic functions of cell
– Mostly ribosomes, sites for protein synthesis
Burton’s Microbiology: Chapter 3

18. Flagella and Pili
• Flagella
– Threads of protein
– Movement: flagellated bacteria
are motile
– Number and arrangment vary
Burton’s Microbiology: Chapter 3
• Pili (fimbriae)
– Hair-like structures (gram-)
– Rigid, not associated with
movement
– Adhere/attach to surfaces
(e.g. human body tissues)
– Sex pilus: transfer of genetic
material (plasmid) from one
bacteria to another
Salmonella cell

19. Spores
Burton’s Microbiology: Chapter 3
• Endospores (some bacteria)
– Thick protein coat
– In order to survive when moisture
or nutrient supply is low
– Resistant to
heat/cold/drying/chemicals
• Sporulation
– Process to form endospores
– Copy of chromosome (most
valuable) and cytoplasm enclosed
• Germination
– Dried spore lands on moist
nutrient-rich surface
– New bacterial cell emerges (a seed)
Terminal spores
Subterminal spores

20. Cell reproduction
• Binary fission (reproduction)
– Chromosome is duplicated
(DNA replication)
– Parent cell splits in half to
become two daughter cells
– Daughter cells contain same
genetic info as parent cell
Burton’s Microbiology: Chapter 3
Binary fission

21. Generation
• Generation time
– Time it takes to splitt cells
– Depends on species and conditions (pH, T, nutrients)
• Slow and rapid growers
– E. coli: 20 minutes (ideal conditions)
• Mathematics
– What time does E. coli need to reach a population of 1.000.000, starting
from just two bacterial cells?
Burton’s Microbiology: Chapter 3

22. Bacterial growth
Burton’s Microbiology: Chapter 8
Encouraging/Inhibting growth
MICROBIOLOGY
LABORATORY
In vitro: outside the body
Artificial culture media
Growth requirements
Inoculate & Incubate
Population growth curve

23. Factors affecting growth
• Fundamental needs of microbes
– Nutrients: energy sources (C, H, O, S, P, N)
– Moisture: Water
– Temperature: thermophiles or pshychrophiles
– pH: acidity or alkalinity
– Pressure: Osmotic (cell membrane) and barometric pressure (atmosphere)
– Atmosphere: O2 and CO2
Burton’s Microbiology: Chapter 8

24. Encouraging growth
• Bacterial growth
– No increase in size, but increase in number (multiplication)
– Binary fission results in bacterial colony
– Control the (ideal) conditions for optimum growth
• Culture media in the lab, artificial
– Content: chemically defined / complex
– State: solid / liquid
– Growth: selective / differential
• Culturing bacteria in the lab, in vitro
– Clinical lab: Enable identification of pathogens
– Research lab: Study, harvest antibiotics, produce vaccines
– Industry lab: Produce microbes for food/beverage companies
Burton’s Microbiology: Chapter 8
Selective: only Gram-
Differential: lactose-
fermenters turn pink

25. Culture bacteria
1. Inoculate
– Add bacteria (specimen) to medium
– Sterile: prevent contamination
2. Incubate
– Chamber with appropriate conditions
– Monitor temperature and atmosphere
Burton’s Microbiology: Chapter 8
3. Count population
– Determine bacterial growth
– Bacterial population growth curve
• Lag: absorb nutrients
• Log: binary fission (growth)
• Stationary: lack of nutrients
• Death: toxic waste
Innoculation of solid agar

26. Growth
EXPERIMENT 001
Title: Potato germs
Topic: Growth of bacteria
1. Peel and slice potato (with gloves) in 4
pieces of equal size
2. Put one slice (with gloves) in box
labeled “control”
3. Put second slice (with gloves) in box
labeled “oral” and cough/blow on it
4. Take third slice and rub it on the table
before putting it in box labeled “table”
5. Take out your gloves and rub fourth
slice with your hands before putting it
in box labeled “hands”

27. Inhibiting growth
• Physical methods
– Heat / Cold
– Heat + pressure
– Desiccation (freeze-drying)
– Radiation / Ultrasonic waves
– Filtration
– Gaseous atmosphere
• Chemical methods
– Disinfection (soap/alcohol)
• Chemical agents on materials
(nonliving)
– Antisepsis (prevent infection)
• Chemical agents on body
(living)
“Inhibiting: from reducing microbial growth to killing pathogens”
Burton’s Microbiology: Chapter 8

28. Classification by staining
• Observing microbes
– Bacteria are colorless, transparent: difficult to see
• Examine bacteria via staining
– Smear
– Dry
– Fix
• Kills organisms
• Preserves morphology
• Anchors to slide
– Observe cell morphology
• Size
• Shape
• Arrangement
• Cell walls, capsules, flagella, endospores
Burton’s Microbiology: Chapter 4

29. Simple stain
Dye: methylene blue
Burton’s Microbiology: Chapter 4
• Determine shape and morphological arrangement

30. Gram staining (1)
• Gram staining procedure
– By Dr. H.C. Gram (1883)
– Most important staining method
• Differentiates Gram- / Gram+
• Color depends on cell wall structure
– Gram+ = blue-to-purple
– Gram- = pink-to-red
Burton’s Microbiology: Chapter 4

31. Gram staining (2)
Gram negative
Gram positive
Burton’s Microbiology: Chapter 4
Result of Gram staining

32. Overview of staining
• Acid-fast staining
– To stain gram-variable bacteria (Mycobacterium)
• M. tuberculosis  in sputum of TB patient
• M. leprae
Burton’s Microbiology: Chapter 4
Acid-fast TB bacilli

33. Characteristics of bacteria
• Characteristics used to identify bacteria
• Already discussed…
– Cell shape (cocci, bacilli, spiral-shaped, etc.)
– Morphological arrangement (chains, clusters, pairs, coiled, etc.)
– Staining reactions (gram+, gram-, no cell wall)
• But there is more…
– Motility
– Colony morphology
– Atmospheric requirements
– Nutrional requirements
– Biochemical and metabolic activities
– Pathogenicity
– Genetic composition
Burton’s Microbiology: Chapter 4

34. Motility
• Motility of bacteria
– Bacterium is able to “swim”
– Associated with presence of flagella
– Determined with semisolid agar method
• Pattern of growth of nonmotile organism
– Turbidity only seen along stab line
– Most cocci are unmotile
• Pattern of growth of motile organism
– Turbidity throughout the medium
– Most spiral-shaped and half of bacilli
Burton’s Microbiology: Chapter 4
Semisolid agar method

35. Colony morphology
• When bacteria divides over and over…
– … the result is a colony
– Colony contains millions of microbes
– Morphology varies in:
• Size (indication for growth rate)
• Color
• Shape
• Elevation
• Appearance at edge
Burton’s Microbiology: Chapter 4

36. Atmosphere
• Classify bacteria on relationship to O2 or CO2
Burton’s Microbiology: Chapter 4
Require ~20% O2
Prefer ~5% O2
Require 0% O2
Prefer 0% O2
No preference
Majority of
clinical specimens

37. Nutrition and Activities
• Biochemical and metabolic activities
– Characterize by waste products
• Secretion of enzymes or gases
• Nutritional requirements
– All: C, H, O, S, P, N
– Some: K, Ca, Fe, Mn, Mg, Co, Cu,
Zn, U, vitamins
– Demanding: Fastidious (fussy)
Burton’s Microbiology: Chapter 4

38. Pathogenicity
Burton’s Microbiology: Chapter 4
• Ability to cause disease
– Many pathogens are
able to cause disease
due to…
• Capsules
• Pili
• Endotoxins
• Exotoxins
• Enzymes
– …which damage cells
and tissues

39. Genetic composition
• Molecular diagnostic procedures
– Modern labs: analyze DNA or RNA of bacteria
– Identify organism by composition of genetic material
– Determine relatedness between two species
Burton’s Microbiology: Chapter 4

40. Unique bacteria
• Rudimentary (unique) bacteria: different from the others…
– Rickettsias & Chlamydias
• Pathogen must live within host cell (like a virus)
– Mycoplasmas: lack cell walls
• Occur in many shapes: pleomorphic
• No Gram staining possible
• Resistant to treatment with penicillin
• Can be free-living or parasitic pathogens
– Megabacteria (750 μm) & Nanobacteria (20 nm)
• Photosynthetic bacteria
– Use light as energy source
– e.g. Cyanobacteria
• Produce oxygen, oxygenation of the atmosphere
• Like blue green algae that overgrow lakes (toxic)
Burton’s Microbiology: Chapter 4

41. Rickettsias (1)
• Unique bacteria: Rickettsias
– Named after Howard T. Ricketts
– Gram- cell wall
– Obligate intracellular pathogen
• Must live within host cell
• Cannot grow on artificial (synthetic)
culture media
• Grow inside eggs/animals/cell cultures
– Disease (transmitted by lice, fleas, ticks)
• Rickettsialpox
• Epidemic and Endemic typhus
• Rocky Mountain spotted fever
– Medicine: Tetracyclines
• Attack bacterial ribosomes
(site for protein synthesis)
Burton’s Microbiology: Chapter 4,8,9

42. Rickettsias (2)
• Obligate intracellular pathogen
– Invade and live in host cells
– Produce their own proteins and RNA/DNA
– Still require intracellular environment:
“leaky membranes”
• Intracellular survival mechanisms
– Attack: Phagocytes (like white blood cells)
protect body, destroy pathogens
– Protection: Rickettsias produces enzymes that
destroy the interior of the phagocytes
Burton’s Microbiology: Chapter 4,14

43. The End