Lecture 2

Bacterial Cell Structure and Bacterial Growth

Eka Jaiani
2024
 Morphology of Prokaryotic Cells
Prokaryotes exhibit a variety of shape. Most bacteria are
Monomorphic
Some can be Pleomorphic
Cocci
- Round (spherical)
• Bacilli
-Rod shaped
• Coccobacillus
– Short round rod
• Vibrio
– Curved rod
• Spirillum
– Spiral shaped
• Spirochete
– Helical shape
• Pleomorphic
– Bacteria able to vary shape
 Morphology of Prokaryotic Cells
• Prokaryotic cells may form groupings after cell
division - adhere together for characteristic
arrangements Arrangements of cocci.
• Arrangement depends on plane of division
– Especially in the cocci
• Division along a single plane
may result in pairs or chains
of cells
– Pairs = diplococci
Example: Neisseria gonorrhoeae
– Chains = streptococci
Example: species of Streptococcus
 Morphology of Prokaryotic Cells
Tetrads
Example- Micrococcus

• Division along two or three perpendicular planes form

cubical packets
– Example: Sarcina genus
• Division along several random planes form clusters
– Example: species of Staphylococcus
Bacilli

(a) Vibrios.

Diplobacilli

(b) Spirillum.

Streptobacilli

(c) Spirochete.
Prokaryote vs Eukaryote
• Paired chromosomes,
• One circular chromosome, not in in nuclear membrane
a membrane • Histones
• No histones • Organelles
• No organelles • Polysaccharide cell walls
• Bacteria: peptidoglycan cell walls • Mitotic spindle
• Archaea: pseudomurein cell
walls
• Binary fission
 Structures External to Cell Wall
• Glycocalyx: Capsules and Slime Layer
Glycocalyx- meaning sugar coat - Glyco = sugar
calyx = shell (substance is synthesized inside of the
cell and secreted outside of the cell)

Chemical composition - Polysaccharides,

Polypeptides or both.
General function -
Protection
- Protects bacteria from host defenses
Attachment
Enables bacteria to adhere to specific
surfaces
- Capsule is a distinct gelatinous layer
- Slime layer-Biofilm is irregular diffuse
layer
 Flagella, Fimbriae and Pili
• Some bacteria have protein appendages
- Aid in survival in certain environments
• Flagella
– Long protein structure
– Responsible for motility
- Use propeller like movements to push bacteria
One advantage of the motility is ability to
move toward favorable conditions or away
from adverse one.
This movement is called Taxis
Chemotaxis (receptors on the bacterial cell pick up the
chemical stimuli) and Phototaxis the movement
in response to light, either towards the source of light
(positive phototaxis ) or away from it (negative phototaxis )
 Arrangements of Bacterial Flagella
• Monotrichous: Bacteria with a single polar flagellum located at
one end (pole)
• Amphitrichous: Bacteria with two flagella
• Lophotrichous: a tuft of flagella coming from one pole
• Peritrichous: Bacteria with flagella all over the surface
• Atrichous: Bacteria without flagella
• Cocci shaped bacteria rarely have flagella
 Flagella
• Flagella structure has three basic parts
Filament: Extends to exterior, Made of proteins called flagellin
Hook: Connects filament to cell
Basal body: Anchors flagellum into cell wall
- Gram negatives have two paris of rings (L and P rings and S and M rings)

rod

S M rings
Fimbriae and Pili

– Considerably shorter and

thinner than flagella
– Similar in structure
– Protein subunits

• Function
- Attachment
- Movement
- Conjugation
Mechanism of DNA transfer
 Cell Wall
• Bacterial cell wall
– Rigid structure
– Surrounds cytoplasmic membrane
– Determines shape of bacteria
– Holds cell together
– Prevents cell from bursting
– Unique chemical structure
– Distinguishes Gram positive from Gram-negative
 Four Groups Based on Cell Wall
Composition

1. Gram-positive cells
2. Gram-negative cells
3. Bacteria without cell walls
4. Bacteria with chemically unique cell walls
 Differential Staining
 The Gram staining method, named after the Danish
bacteriologist Hans Christian Gram who originally devised it
in 1882 (published 1884).

 Gram staining, is one of the most

important staining techniques in microbiology. It is almost
always the first test performed for the identification of
bacteria.

– Bacteria separated into two

major groups
• Gram positive-Stained purple
• Gram negative-Stained red or
pink
Gram staining
 Bacterial Cell Wall
Rigidity of cell wall is due to
peptidoglycan (PTG) or murein.
Peptidoglycan consists of a
repeating disaccharide made up
of monosaccharides called
N-acetylglucosamine(NAG)
and N-acetylmuramic acid (NAM)

Peptide cross-bridge (link)

- Joined subunits form glycan chain
Side chain
- Glycan chains held together by string of four amino acids
Tetrapeptide chain
Cell Wall
• Gram positive cell wall
– Relatively thick layer of PTG
• 20 to 80 layers of PTG
– Regardless of thickness,
PTG is permeable to
numerous substances
– Teichoic acid component of
PTG
- Gives cell negative charge
Gram Positive Bacterial Cell Wall
 Cell Wall
Gram-negative cell wall
– More complex than G+
– Contains thin layer
of PTG (2-8 layers of PTG)
• PTG sandwiched between
outer membrane and plasma
membrane
Region between outer membrane
and cytoplasmic membrane is
called periplasm
-Most secreted
proteins contained
here
-Proteins of
transport system
located here
 Atypical Cell Walls
 Some bacteria naturally lack cell wall
• Mycoplasma
- Bacterium causes mild pneumonia
- Mycoplasma species are the smallest bacterial cells
- Have no cell wall
- Pleomorphic because of absence of cell wall
– Antimicrobials directed towards cell wall ineffective
Sterols in membrane account for strength of membrane
-
• Acid-Fast cell walls
-60% Mycolic acid and thin Peptidoglycan layer
• Bacteria in Domain Archaea
– Have a wide variety of cell wall types
– None contain peptidoglycan but rather pseudopeptidoglycan
 Cytoplasmic (or Plasma) Membrane

• Cytoplasmic membrane

– Delicate thin fluid structure

– Surrounds cytoplasm of cell
– Contains primarily of
Phospholypides
– Does not contain sterols (except Mycoplasma)
– Serves as a semi permeable barrier (has selective permeability)
• Barrier between cell and external environment
 Internal Structures
Cytoplasm –The substance inside the Plasma Membrane
• 80%-Water
• 20%- proteins (enzymes), carbohydrates, lipids, inorganic ions
• Thick, Aqueous, Semitransparent and Elastic
• Nucleoid, Ribosomes, Inclusions

Bacterial cells have variety of internal structures

• Some structures are essential for life
– Chromosome
– Ribosome

• Others are optional and can confer selective advantage

– Plasmid
– Storage granules
– Endospores
 Internal Structures
• Chromosome
– Resides in cytoplasm
• In nucleoid space
– Typically single chromosome
- Circular double-stranded molecule
- Contains all genetic information

• Plasmid
– Circular DNA molecule
• Generally 0.1% to 10% size of
chromosome
– Extrachromosomal
• Independently replicating
– Encode characteristic
• Potentially enhances survival
– Antimicrobial resistance
 Internal Structure
• Ribosome
– Involved in protein synthesis
– Composed of large and small subunits
- Units made of riboprotein and ribosomal
RNA

• Prokaryotic ribosomal subunits

• Large = 50S(contains two rRNA
molecules)
• Small = 30S (contains one rRNA
molecule)
• Total = 70S

– Difference from eukaryotic ribosomes

(40S, 60S, 80S)
• often used as target for antimicrobials
 Internal Structures
• Endospores
– Dormant cell types
• Produced through sporulation
• Theoretically remain dormant for
100 years
• Resistant to damaging conditions
- Heat, desiccation, chemicals and UV
light
• Vegetative cell produced through
germination
- Germination occurs after exposure to
heat or chemicals
• Germination not a source of
reproduction Common bacteria genus that
produce endospores include
Clostridium and Bacillus
Endospore
 Internal Structures
• Storage granules (Inclusions)

Polysaccharide Granules, Lipid Granules, Sulfur Granules…

– Accumulation of polymers
• Synthesized from excess nutrient
– Example : glycogen
– Excess glucose in cell is stored in glycogen granules

Gas vesicles-hollow cavities

Small protein compartments
Provides buoyancy to cell
Regulating vesicles allows organisms
to reach ideal position
in environment
Bacterial nutrition, Growth, Principles of Laboratory
Diagnosis of Bacteria

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 Microbial Growth
Microbial growth: Increase in cell number, not cell size!
Physical Requirements for Growth:
Temperature
pH
Osmotic Pressure
Chemical Requirements for Growth:
Macro & Micro Elements
Carbon, N, O, S, P, etc.
 Temperature
Physical Requirements for Growth: Temperature

• Minimum growth temperature

• Optimum growth temperature

• Maximum growth temperature

 Microbial Growth
Five groups based on optimum growth temperature

Psychrophiles
range -10C- 20C
Psychrotrophs
(cold loving microbes )
range 0C-30C
Mesophiles
(moderate temp. loving microbes)
range 10 C - 50 C
Thermophiles
(heat loving microbes)
range 40 C - 70C
Hyperthermophiles
range 70C -100C
 Physical Requirements for Growth:

pH
Most bacteria grow best between pH 6.5 and pH7.5 Neutrophils
Staph aureus
Some bacteria are very tolerant of acidity or thrive in it:
Acidophiles (preferred pH range 1 to 5) Acidithiobacillus spp
Alkaliniphiles (preferred pH range 9 to 12) Alcaligenes faecalis
Molds and yeasts grow best between pH 5 and pH6
 Osmotic Pressure
Microbes obtain almost all their nutrients in solution
from surrounding water
(cell composition 80%-90% water)
Tonicity
Isotonic ( is one in which its effective osmole concentration is the same as the solute
concentration of another solution with which it is compared)

Hypertonic (higher concentration of solutes on the outside of the cell)

Hypotonic (lower concentration of solutes on the outside of the cell) than its
surroundings, so in an attempt to balance concentrations, water will rush into the cell.
 Hypertonic
cause plasmolysis

Hypotonic
causing swelling (Osmolysis)
Salt loving microorganisms:
In artificial solid media
Obligate halophiles (30% salt)
agar concentration is
Facultative Halophiles (2-15%salt)
1,5%
 Metabolic Diversity Among Organisms

Nutritional Pattern: source of energy and

source of carbon

Energy source
Phototrophs – use light as their primary energy
source
Chemotrophs – depend on oxidation-reduction
reactions of inorganic or organic compounds
for energy
 Metabolic Diversity Among Organisms

Carbone Source
Autotrophs (Lithotrophs) – use carbondioxide

Heterotrophs (Organotrophs ) – use organic carbon source

Nutritional Classification
(Energy Source + Carbon Source)
Photoautotrophs
Photoheterotrophs
Chemoautotrophs
Chemoheterotrophs (most of human infections are
caused by this group)
Chemical Requirements for Growth: Carbon, N, S, P, etc.
Carbon (Structural Backbone)
–  Half of dry weight Vit B1
– Chemoheterotrophs use organic carbon sources
– Chemoautotrophs use CO2

• Nitrogen 14%, Sulfur , Phosphorus 4%

– Organic - Found in amino acids and proteins, DNA, RNA
(most bacteria decompose proteins)
– Inorganic – Ammonium Ions (NH4+),Nitrates and N2 ( Nitrogen fixation)
– S in amino acids, vitamins ( thiamine and biotin), Vit B7
– Phosphate ions (PO43–) Nucleic Acid, Phospholipids
ATP

• Also needed K, Mg, Ca, Fe, Cu, Zn trace elements (as cofactors), and
organic growth factors (vitamins, purines , pyrimidines and amino
acids)
 Chemical Requirements for Growth: Oxygen

O2 requirements vary greatly

The Effects of Oxygen on the Growth of Various Types of Bacteria

• Obligate Aerobes – require oxygen to live,
extracts more energy from nutrients
(disadvantage – oxygen is poorly soluble in
water)
• Facultative Anaerobes – can use oxygen when it
is presented are able to continue on absence of
oxygen by using fermentation or anaerobic
respiration
• Obligate Anaerobes – unable to use molecular
oxygen, most are harmed by oxygen, do use
oxygen atoms presented in cellular materials
 SOURCES OF METABOLIC ENERGY
• Fermentation – glycolise,
substrate
phosphorylation
• Respiration
• Photosynthesis

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CRITERIA FOR IDENTIFICATION OF BACTERIA

42
Gram staining and microscopy
The Gram stain was
developed by Christian
Gram in 1884 and can
differentiate between the
two types of cell walls
Gram positive and Gram-
negative.

Christian
Gram 1884
43
 Acid-fast stain
• Used to stain organisms
that resist conventional
staining.
• Used to stain
Mycobacterium
tuberculosis
• High lipid concentration
in cell wall prevents
uptake of dye
• Once stained difficult to
decolorize.
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45
 Immunologic Tests—Serotypes, Serogroups, and
Serovars
• The designation “sero” simply indicates the Cholera spp. subtyping
use of antibodies (polyclonal or monoclonal)
that react with bacterial cell surface structures
such as lipopolysaccharide (LPS), flagella, or
capsular antigens.
• The terms “serotype,” “serogroups,” and
“serovars” are, for all practical purposes,
identical—they all use the specificity of these
antibodies to subdivide strains of a specific
bacterial species.
• In certain circumstances (eg, an epidemic), it is
important to distinguish among strains of a
given species or to identify a specific strain.
This is called subtyping and is accomplished by
examining bacterial isolates for characteristics
that allow discrimination below the species
level.

46
 Genetic Diversity
• Developments in DNA sequencing
now make it possible to investigate
the relatedness of genes or genomes
by comparing sequences among
different bacteria.
• It should be noted that genetic
instability can cause some traits to
be highly variable within a biologic
group or even within a specific
taxonomic group.
• For example, antibiotic resistance
genes or genes encoding toxins may
be carried on plasmids or
bacteriophages.
• Many organisms are difficult to
cultivate, and in these instances,
techniques that reveal relatedness by
measurement of DNA sequence
analysis may be of value. 47
 Ribosomal RNA sequencing
• Ribosomes have an essential role in
protein synthesis for all organisms and
as such are indispensable.
• Genetic sequence encodings both
ribosomal RNAs (rRNA) and ribosomal
proteins (both of which are required to
comprise a functional ribosome) are
highly conserved throughout
evolution and have diverged more
slowly than other chromosomal genes.
• Comparison of the nucleotide
sequence of 16S rRNA from a range of
prokaryotic sources revealed
evolutionary relationships among
widely divergent organisms and has led
to the elucidation of a new kingdom,
the archaebacteria.
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 Culture Media
• Culture medium: Nutrients prepared for microbial growth
• Different microorganisms require different Culture media
(some of them can’t grow on any of them)
• Inoculum: Microbes introduced into medium
• Culture: Microbes growing in/on culture medium
What criteria mus culture media meet?
• Right nutrients
• properly adjusted pH
• Have to be sterile (not contain living microbes)
 Culture Media
Wide variety of media are commercially available
1) Add water
2) Sterilize
Liquide Media Solid Media (agar is added)
1,6%, 0,7%, 0,4%
Prepearing petry plates and Slants:
1)Melting Solid media (100ºC)
2) Cooling (50ºC)
3) pouring into petri dishes or reaction tubes

Petry Dishes Slants deeps

 Selective Media and Differential Media
Selective medium: Additives
suppress unwanted and
encourage desired microbes –
e.g. Bismuth Sulfite agar
MacConkey agar

Differential medium: changed

in recognizable manner by
some bacteria  Make it easy
to distinguish colonies of
different microbes – e.g.  and
 hemolysis on blood agar;
Bacterial Division Binary fission
 The Growth of Bacterial Cultures
Binary fission – exponential growth

Generation time – time required for cell to divide (also

known as doubling time)
Ranges from 20 min (E. coli) to > 24h (M. tuberculosis)
Consider reproductive potential of E. coli
 Bacterial Growth Curve
Illustrates the dynamics of growth
Phases of growth
– Lag phase
– Exponential or
logarithmic (log) phase
– Stationary phase
– Death phase (decline phase)
 Control of bacterial growth
• Sterilization - killing all the organisms, including
spores
• Disinfection – elimination of the vegetative cells
but not the spores
• Pasteurization- the process of applying heat,
usually to milk or cheese, for a specified period for
the purpose of killing or retarding the growth of
pathogenic bacteria.
• Aseptic - free of, or using methods to keep free of,
microorganisms.
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Physical and chemical agents affecting bacterial growth

• Heat
 Application of heat is the simplest means of sterilizing
materials, provided the material is itself resistant to heat
damage.
 A temperature of 100°C will kill all but the spores of
eubacteria within 2–3 minutes in laboratory-scale cultures;
 A temperature of 121°C for 15 minutes is used to kill spores
• Radiation
 Ultraviolet (UV) radiation that has a wavelength of about 260
nm causes thymidine dimers resulting in the inability of
bacterial DNA to be replicated. This is generally bactericidal,
but not against spores.
• Filtration – for fluids 56
Physical and chemical agents affecting bacterial growth

• Alcohols
• These agents effectively remove water from biologic systems.
Thus, they functionally act as “liquid desiccants.”
• alcohol, isopropyl alcohol, and n-propanol exhibit rapid,
broad-spectrum antimicrobial activity against vegetative
bacteria, viruses, and fungi but are not against spores.
• Aldehydes
• Compounds like glutaraldehyde or formaldehyde are used for
low-temperature disinfection and sterilization of instruments,
endoscopes, and surgical tools.
• They are normally used as a 2% solution to achieve sporicidal
activity. These compounds are generally bactericidal and
sporicidal. 57
Physical and chemical agents affecting bacterial growth
Peroxygens
• Hydrogen peroxide (H2O2 ) has broad-spectrum
activity against viruses, bacteria, yeasts, and
bacterial spores.
• Sporicidal activity requires higher concentrations
(10–30%) of H2O2 and longer contact times.
Phenols
• Phenol and many phenolic compounds have
antiseptic, disinfectant, or preservative
properties. In general, these are not sporicidal
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 Reading

Tortora
Chapters 4, 6