Bacterial Classification, Anatomy, Nutrition, Growth, Metabolism and Genetics

Classification Systems in the Prokaryotes

1.

2. 3. 4. 5. 6.

Macroscopic morphology colony appearance Microscopic morphology Physiological / biochemical characteristics Chemical analysis Serological analysis Genetic and molecular analysis
G + C base composition DNA analysis using genetic probes Nucleic acid sequencing and rRNA analysis

G + C base composition

Low G+C Gram-Positive Bacteria

Clostridia Mycoplasmas

High G+C Gram-Positive Bacteria

Corynebacterium Mycobacterium

Bacterial Taxonomy Based on Bergeys Manual

Bergeys Manual of Determinative Bacteriology five volume resource covering all known procaryotes
based on genetic information phylogenetic two domains: Archaea and Bacteria five major subgroups with 25 different phyla
classification

Major Taxonomic Groups of Bacteria

Vol 1A: Domain Archaea primitive, adapted to extreme habitats and modes of nutrition Vol 1B: Domain Bacteria Vol 2-5: 2 - Phylum Proteobacteria Gram-negative cell walls 3 - Phylum Firmicutes mainly Gram-positive with low G + C content 4 - Phylum Actinobacteria Gram-positive with high G + C content 5 Loose assemblage of phyla All gram negative

Diagnostic Scheme for Medical Use

Uses phenotypic qualities in identification

restricted

to bacterial disease agents divides based on cell wall structure, shape, arrangement, and physiological traits

Species and Subspecies

Species
bacterial

cells which share overall similar pattern of

traits

Subspecies
Strain

or variety culture derived from a single parent that differs in structure or metabolism from other cultures of that species

Type
subspecies

that can show differences

Bacterial Shapes, Arrangements, and Sizes

Typically described by one of three basic shapes:

coccus

Spherical Rod

bacillus

coccobacillus vibrio

spirillum

Helical, twisted rod,

Spirochete

Bacterial Shapes, Arrangements, and Sizes

Arrangement of cells dependent on pattern of division and how cells remain attached after division:
cocci: singles diplococci tetrads chains irregular clusters cubical packets bacilli: chains palisades

Cocci

Bacilli

Bacterial anatomy

Generalized structure of a prokaryotic cell

Appendages: Cell Extensions Flagella

3 parts

filament long, thin, helical structure composed of proteins Hook curved sheath basal body stack of rings firmly anchored in cell wall rotates 360o

1-2 or many distributed over entire cell

Flagellar Arrangements

monotrichous
single flagellum at one end

lophotrichous
small bunches arising from one end of cell

amphitrichous
flagella at both ends of cell

peritrichous
flagella dispersed over surface of cell, slowest

Fig. 4.4

Movement by flagella

Polar

Rotates counterclockwise Cell swims forward in runs Reverse will stop it All flagella sweep towards one end

Peritrichous

Chemotaxis

Internal Flagella Axial Filaments

aka Periplasmic Endoflagella

Spirochetes

enclosed between cell wall and cell membrane of spirochetes

Appendages for Attachment Fimbrae

fine hairlike bristles from the cell surface function in adhesion to other cells and surfaces

Appendages for Mating Pili

rigid tubular structure made of pilin protein found only in Gram negative cells Functions

joins bacterial cells for DNA transfer (conjugation) Adhesion to form biofilms and microcolonies

The Cell Envelope

External covering outside the cytoplasm Composed of few basic layers:

glycocalyx cell wall cell membrane

Maintains cell integrity

The Cell Membrane

fluid layer of phospholipid and protein phospholipid molecules are arranged in a bilayer Hydrophobic fatty acid chains in the phospholipids form a permeability barrier

The Bacterial Surface Coating Glycocalyx

Coating of molecules external to the cell wall Made of sugars and/or proteins functions

attachment inhibits killing by white blood cells receptor

The Bacterial Surface Coating Glycocalyx

2 types: 1. slime layer loosely organized and attached 2. capsule - highly organized, tightly attached

Cell Wall

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

Structure of the Cell Wall Peptidoglycan

macromolecule composed of a repeating framework of long glycan chains

cross-linked

by short peptide fragments

provides strong, flexible support

keep

bacteria from bursting or collapsing because of changes in osmotic pressure

The Gram Stain

Important basis of bacterial classification and identification Practical aid in diagnosing infection and guiding drug treatment Differential stain

Gram-negative

lose crystal violet and stain red from safranin counterstain retain crystal violet and stain purple

Gram-positive

Gram Positive Cell Wall (1)

Consists of

a thick, homogenous sheath of peptidoglycan

tightly bound acidic polysaccharides teichoic acid and lipoteichoic acid

Periplasmic space cell membrane

Retain crystal violet and stain purple

Gram Negative Cell Wall (2)

Consists of

an outer membrane containing lipopolysaccharide (LPS) periplasmic space thin shell of peptidoglycan periplasmic space cell membrane

Protective structure while providing some flexibility and sensitivity to lysis Lose crystal violet and stain red from safranin counterstain

Gram Negative Cell Wall

LPS

endotoxin that may become toxic when released during infections may function as receptors and blocking immune response contains porin proteins in upper layer

Regulates molecules entering and leaving cell

Atypical Cell Walls

Some bacterial groups lack typical cell wall structure

Mycobacterium and Nocardia Gram-positive cell wall structure with lipid mycolic pathogenicity high degree of resistance to certain chemicals and dyes basis for acid-fast stain

acid

Some have no cell wall

Mycoplasma cell wall is stabilized pleomorphic

by sterols

Chromosome

single, circular, doublestranded DNA molecule contains all the genetic information required by a cell DNA is tightly coiled around a protein

dense area called the nucleoid central subcompartment in the cytoplasm where DNA aggregates

Plasmids

small circular, doublestranded DNA stable extrachromosomal DNA elements that carry nonessential genetic information duplicated and passed on to offspring

replicate independently from the chromosome

Plasmids

may encode antibiotic resistance, tolerance to toxic metals, enzymes & toxins used in genetic engineering

readily manipulated & transferred from cell to cell

F plasmids allow genetic material to be transferred from a donor cell to a recipient R plasmids carry genes for resistance to antibiotics

Storage Bodies Inclusions & Granules

intracellular storage bodies vary in size, number & content Examples:

Glycogen poly-b-hydroxybutyrate gas vesicles for floating sulfur polyphosphate granules

Endospores

resting, dormant cells produced by some G+ genera

Clostridium, Bacillus & Sporosarcina

resistance linked to high levels of calcium & certain acids longevity verges on immortality 25 to 250 million years pressurized steam at 120oC for 20-30 minutes will destroy

Endospores

have a 2-phase life cycle

vegetative cell endospore formation of endospores return to vegetative growth

sporulation

Germination

withstand extremes in heat, drying, freezing, radiation & chemicals

Endospores
stressed cell
undergoes asymmetrical cell division creating small prespore and larger mother cell prespore contains:

Cytoplasm DNA dipicolinic acid

mother cell matures the prespore into an endospore

then disintegrates

environmental conditions are again favorable

protective layers break down spore germinates into a vegetative cell

Microbial nutrition, growth, and metabolism

Microbial Nutrition
nutrition
process

by which chemical substances (nutrients) are acquired from the environment and used for cellular activities

Two

categories of essential nutrients:

or trace elements

macronutrients

micronutrients

Inorganic nutrients Organic nutrients

Obtaining Carbon

Heterotroph
organism

that obtains carbon in an organic form made by other living organisms proteins, carbohydrates, lipids and nucleic acids

Autotroph

an organism that uses CO2 (an inorganic gas) as its carbon source not dependent on other living things

Growth Factors

organic compounds that cannot be synthesized by an organism & must be provided as a nutrient

Carbon Energy source source

photoautotrophs CO2 sunlight

chemoautotrophs

CO2

Simple inorganic chemicals sunlight

essential amino acids, vitamins Chemo

photoheterotrophs

organic

Nutritional types

chemoheterotrophs organic

Chemical compounds light

Metabolizing organic cmpds

Photo

Types of Heterotrophs

Saprobes Parasites / pathogens

Obligate

Nutritional Movement
Osmosis Facilitated diffusion Active transport Endocytosis

Phagocytosis

Pinocytosis

Extracellular Digestion

digestion of complex nutrient material into simple, absorbable nutrients accomplished through the secretion of enzymes (exoenzymes) into the extracellular environment

Environmental Influences on Microbial Growth

1. 2. 3. 4. 5. 6.

temperature oxygen requirements pH Osmotic pressure UV light pressure

1. Temperatures

Minimum temperature
lowest

temperature that permits a microbes growth and metabolism temperature that permits a microbes growth and metabolism the fastest rate of growth and metabolism

Maximum temperature
highest

Optimum temperature
promotes

Temperature Adaptation Groups

Psychrophiles
optimum temperature 15oC capable of growth at 0 - 20oC

Mesophiles
optimum temperature 10o - 40oC most human pathogens

Thermophiles
optimum temperature 60oC capable of growth at 40 - 70oC

Hyperthermophiles

Archaea that grow optimally above 80C found in seafloor hot-water vents

2. Oxygen Requirements

Aerobe

requires oxygen
grow without oxygen not require oxygen

Obligate aerobe
cannot

Anaerobe
does

Facultative anaerobe
capable

of growth in the absence OR presence of oxygen

Thioglycollate broth can be used to test an organisms oxygen sensitivity Gas chamber

3. pH

The pH Scale
Ranges

from 0 - 14 pH below 7 is acidic

[H+] > [OH-]
pH

above 7 is alkaline
[OH-] > [H+]

pH

of 7 is neutral
[H+] = [OH-]

3. pH

Acidophiles

optimum pH is relatively to highly acidic optimum pH ranges about pH 7 (plus or minus) optimum pH is relatively to highly basic

Neutrophiles

Alkaphiles

4. Osmotic Pressure

Bacteria 80% water

Require

water to grow

Sufficiently hypertonic media at concentrations greater than those inside the cell cause water loss from the cell
Osmosis Fluid leaves the bacteria causing the Causes the cell membrane to separate

cell to contract

Plasmolysis
Cell

shrinkage to and require high salt concentrations

extreme or obligate halophiles

Adapted

5. UV Light

Great for killing bacteria Damages the DNA (making little breaks)

in sufficient quantity can kill the organisms in a lower range causes mutagenisis

Spores tend to be resistant

can survive much longer exposures

6. Barophiles

Bacteria that grow at moderately high hydrostatic pressures

Oceans membranes and enzymes depend on pressure to maintain their threedimensional, functional shape Grows at pressures from 100500 Atm 400-500 Higher than 500

Barotolerants

Barophilic

Extreme barophilic

Microbial Associations

Symbiotic
organisms

live in close nutritional relationships;

Mutualism
Obligatory Dependent Both members benefit

Commensalism
One member benefits Other member not harmed

Parasitism
Parasite is dependent and benefits Host is harmed

Microbial Associations

Non-symbiotic
organisms

are free-living relationships not required for survival

Synergism

members cooperate and share nutrients some member are inhibited or destroyed by others

Antagonism

Microbial Associations

Biofilms

Complex relationships among numerous microorganisms Develop an extracellular matrix

Adheres cells to one another Allows attachment to a substrate Sequesters nutrients May protect individuals in the biofilm

Microbial Growth in Bacteria

Binary fission:
Prokaryotes

reproduce

asexually one cell becomes two

basis for population growth parent cell enlarges duplicates its chromosome forms a central septum

Process:

divides the cell into two daughter cells

Population Growth

Generation / doubling time time required for a complete fission cycle Length of the generation time is a measure of the growth rate of an organism Some populations can grow from a small number of cells to several million in only a few hours!!

Prokaryotic Growth

Bacterial Growth Curve

lag phase

no cell division occurs while bacteria adapt to their new environment

Exponential growth of the population occurs Human disease symptoms usually develop When reproductive and death rates equalize accumulation of waste products and scarcity of resources

logarithmic (log) phase

stationary phase

decline (exponential death) phase

Other Methods of Analyzing Population Growth

Turbidity Direct microscopic count Coulter counting

Turbidity

Direct Microscopic Count

Electronic Counting

Microbial genetics

Genomes

Prokaryotic Genomes

Prokaryotic chromosomes

Main portion of DNA, along with associated proteins and RNA Prokaryotic cells are haploid (single chromosome copy) Typical chromosome is circular molecule of DNA in nucleoid

DNA Replication in Prokaryotes

Genetic Recombination in Prokaryotes

Genetic recombination
occurs when an organism acquires and expresses genes that originated in another organism

Genetic information in prokaryotes can be transferred vertically and horizontally

Vertical gene transfer (VGT)

transfer of genetic material from parent cell to daughter cell transfer of DNA from a donor cell to a recipient cell Three types

Horizontal gene transfer (HGT)

Transformation Transduction Bacterial conjugation

DNA Recombination Events

3 means for genetic recombination in bacteria:
Conjugation 2. Transformation 3. Transduction
1.

Transmission of Genetic Material in Bacteria

conjugation requires the attachment of two related species & formation of a bridge that can transport DNA
transfer of naked DNA

transformation

transduction

DNA transfer mediated by bacterial virus

1. Conjugation

transfer of a plasmid or chromosomal fragment from a donor cell to a recipient cell via a direct connection Gram positive and gram negative
Gram-negative

cell donor has a fertility plasmid

(F plasmid, F factor) allows the synthesis of a conjugation (sex) pilus

recipient cell is a related species or genus without a fertility plasmid donor transfers fertility plasmid to recipient through pilus F+ and F-

Physical Conjugation

2. Transformation

chromosome fragments from a lysed cell are accepted by a recipient cell

genetic

code of DNA fragment is acquired by recipient

Donor and recipient cells can be unrelated Useful tool in recombinant DNA technology

Transformation of Insulin Gene

human insulin gene isolated and cut from its location on the human chromosome using a restriction enzyme plasmid is cut using the same restriction enzyme desired DNA (insulin gene) and plasmid DNA can be joined using DNA ligase plasmid now contains the genetic instructions on how to produce the protein insulin Bacteria can be artificially induced to take up the recombinant DNA plasmids and be transformed successfully transformed bacteria will contain the desired insulin gene transformed bacteria containing the insulin gene can be isolated and grown As transformed bacteria grow they will produce the insulin proteins coded for the recombinant DNA Insulin harvested and used to treat diabetes

3. Transduction

DNA is transferred from one bacterium to another by a virus Bacteriophages

Virus

that infects bacteria consist of an outer protein capsid enclosing genetic material serves as a carrier of DNA from a donor cell to a recipient cell

3. Transduction

Two types:
generalized

transduction

random fragments of disintegrating host DNA are picked up by the phage during assembly any gene can be transmitted this way
specialized

transduction

a highly specific part of the host genome is regularly incorporated into the virus

Generalized Transduction

Specialized Transduction

Transposons

Special DNA segments that have the capability of moving from one location in the genome to another jumping genes Can move from

one chromosome site to anotherr chromosome to a plasmid plasmid to a chromosome

May be beneficial or harmful Changes in traits Replacement of damaged DNA Transfer of drug resistance

Mutations

Result of natural processes or induced

Spontaneous

mutations

heritable changes to the base sequence in DNA result from natural phenomena such as radiation or uncorrected errors in replication

UV

light is a physical mutagen that creates a dimer that cannot be transcribed properly

Nitrous acid is a chemical mutagen that converts adenine bases to hypoxanthine

Hypoxanthine

base pairs with cytosine instead of thymine

Base analogs bear a close resemblance to nitrogenous bases and can cause replication errors

Point Mutation

Result of spontaneous or induced mutations affects just one base pair in a gene Base-pair substitutions

result in an incorrect base in transcribed mRNA codons

Base-pair deletion or insertion

results in an incorrect number of bases

Repair Mechanisms

Attempt to correct mistakes or damage in the DNA Mismatch repair involves DNA polymerase

proofreading the new strand removing mismatched nucleotides

Excision repair
involves

cutting out damaged DNA replacing it with correct nucleotides