Micro 2

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iRON for bacteria

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considered a trace element but most organisms needed for surviva l

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The primary halide required for bacteria

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Cl is the primary halide required

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trace elements are used as

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enzymes or cofactors in facilitating rxn or as structural elements as well

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Photoautotrophs
energy source
Carbon source

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Light
CO2

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Photoautotrophs example

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Oxygenic: Cyanobacteria, plants. Anoxygenic: Green and purple bacteria.

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Photoheterotrophs
energy source
Carbon source

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Light
Organic compounds

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Photoheterotrophs example

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Green and purple nonsulfur bacteria.

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Chemoautotrophs (Lithoautotrophs)
energy source
Carbon source

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Reduced inorganics
CO2

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Chemoautotrophs (Lithoautotrophs) example

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Fe-oxidizing bacteria, most Archaea.

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Chemoheterotrophs
energy source
Carbon source

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Reduced organics
Organic compounds

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Chemoheterotrophs example

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Fermentative bacteria. Bacteria, protozoa,

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All pathogens are part of

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Chemoheterotrophs

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Facultative Anaerobes*

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O2 not required but utilized when available
Cornybacterium, Escherichia, Listeria, Staphylococus, Y easts

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Obligate Anaerobes

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O2 toxic
Bacteroides, Clostridium, Propionibacterium

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Aerotolerant Anaerobes

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O2 insensitive

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Micro- aerophiles

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O2 required but at levels below 0.2 atm

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In terms of oxygen requirements, most pathogens are

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Facultative anaerobes
Anaerobes

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Essential substances that the organism is unable to synthesize from available nutrients.

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1. Purines and pyrimidines (for DNA and RNA) 2. Amino acids (for proteins)
3. Vitamins (for coenzymes/cofactors)
Growth factors are not metabolized as sources of carbon or energy,
but are directly assimilated by cells to fulfill their specific role in metabolism.

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Microoganisms that are usually pathogenic thrive at a water activity (Aw) of about

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0.7-1. Lowering water activity
by adding salts or sugars is an effective measure for prevention of bacterial growth

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Catabolism of proteins, polysaccharides, and lipids provides

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glucose, pyruvate, tricarboxylic acid (TCA) cycle intermediates, and energy
in the form of ATP and NADH.

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Catabolic process is breaking down

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highly riched and reduced carbonated compounds to get energy out of them, so it would couple enzymatic rxns to oxydize the reduced compounds and sabe the thermodynamic equivalent into energy compounds such as NADH and ATP.

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The glycolytic pathway couples the oxidation of glucose to

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pyruvate with generation of two ATPs and two NADHs:

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Net rxn of glycolysis

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Glucose + 2NAD+ + 2ADP + 2Pi →
2 pyruvate + 2NADH + 2H+ + 2ATP + 2H2O

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Priming stage of glycolysis:

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glucose (reduced carbon compound) comes into organisms as source of energy. Before breaking it down, it has to be primed. Glucose is phosphorylated to commit glucose to the process of glycolysis. The second stage is breaking down glucose to recover the energy from glucose.

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The free energy decrease in this conversion is very large, making glycolysis

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essentially irreversible.

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Different microorganisms yield different end products when fermenting pyruvate. Clinical laboratories use these variations for

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bacterial identification.

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The tricarboxylic acid (TCA) cycle occurs in

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aerobic conditions and is
an amphibolic cycle.
Precursors for the synthesis of amino acids and nucleotides are also shown.

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he TCA cycle
1) is the major mechanism for

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the
ultimate generation of ATP.

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The TCA cycle serves as

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the final pathway for the
complete oxidation of carbohydrates,
amino acids, and fatty acids

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The TCA cycle supplies the key intermediates
(i.e., pyruvate, α-ketoglutarate, oxaloacetate) for

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the ultimate synthesis of amino acids, lipids, purines, and pyrimidines

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NADHs are oxidize and produce energy in the form of ATP. In bacteria, all the components involved witih oxidative phosphorulation are embedded in

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the membrane and thhey are intrinsic.

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Electron carriers are organized into three complexes In the cell membrane. Protons are pumped across the membrane at three points, from

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he inside to the outside. ATP generation is coupled to proton reentry.

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The combination of glycolysis, the TCA cycle, and the electron transport chain
in aerobic glucose metabolism provides

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an efficient method of energy generation: 38 ATP per glucose...
~20X more efficient than fermentation.

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Some bacteria can perform anaerobic respiration, where the terminal electron acceptor is

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a compound other than O2 (e.g., NO3-, SO42-, or CO2).

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Aerobic Glucose Metabolism Net reaction:

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Glucose + 6O2 + 38ADP + 38Pi →
6CO2 + 38ATP + 6H2O

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amphibolic importance of the krebs cycle..meaning

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it creates intermediates for the amino acid synthesis

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Bacterial Growth
Definition:

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Cell growth is orderly increase in all components of the organism. Population growth is cell multiplication as a consequence of cell growth.*

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Bacterial Growth measurements

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Biomass density; cell concentration; number of cells per time point.

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Lag Phase

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Adaptation to new environment. Time to switch on new metabolic machinery. Population remains unchanged, but cells are growing in mass and metabolic activity.

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Exponential Growth Phase

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Rate of growth in non-limiting conditions.
Most metabolically active period, and most sensitive to antibiotics.

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Exponential Growth

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Exponential growth is more the exception than the rule.
Growth will continue only if there is ample space and nutrients and no accumulation of waste products.

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ll populations will ultimately be limited by their resources and environment. They all will eventually

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plateau and decline

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Stationary Phase

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Closed system results in exhaustion of nutrients/space and accumulation of waste. Viable cell count stays constant (kdeath = kbirth).
Population is not quiescent; may be adjusting.

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4) Death Phase

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Individual cell death = loss of ability to reproduce (grow and divide). Death is when the viable population declines (kdeath > kbirth).
The death decline is also exponential.

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Different experimental conditions can influence the stages of the growth curve:

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Liquid media...solid media...chemostat...steady state culturing...synchronized growth

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psychrophiles:
mesophiles:
thermophiles

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-10–20 °C
10–50 °C
40–70 °C

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pH optimal conditions

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pH 4.5–8.0)
(bacillus)
pH:
Fermenters and yeast are acid tolerant.

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H2O requirements

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Obviously required for viability, but desiccating environments can be tolerated through sporulation.

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Difficult organisms to culture:

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Treponema pallidum: syphilis
Mycobacterium leprae: leprosy Ricksettiae: typhus

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Plating allows
Selective Liquid Media:
Selective Solid Media:

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Select individual colonies.
Enrich the population of desired organisms
Permit direct isolation

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Colony Identification

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1) Size: Sensitive measure of growth rate. 2) Color.
3) Odor.
4) Surface Texture:
Smooth: Suggests capsulation; compact cellular orientation. Rough: Dry or wrinkled, can vary within species.
Often correlates with virulence (e.g., M. tuberculosis).
Due to no capsulation, or filamentous growth.
Mucoid: Glossy appearance due to polysaccharide exudate. These parameters depend on environmental as well as genetic factors (e.g., P. aeroginosa).
5) Microscopic examination of stained specimens.

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Selective Media:

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Suppress growth of unwanted microbes while encouraging growth of other(s).
For example, bismuth sulfite agar is used to isolate the gram-negative typhoid bacterium Salmonella typhimurium from feces, since bismuth sulfite inhibits gram-positive bacteria as well as most other gram-negative intestinal bacteria.

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Differential Media:

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Reveal specific characteristics without being selective.
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E. coli on eosin-methylene blue medium.
The black-centered colonies are surrounded by a characteristic metallic green sheen. Weaker lactose fermenters have pink/purple- colored colonies, while colonies of non-lactose fermenters remain colorless.
http://microbelibrary.org
For example, fermentation plates with eosin-methylene blue (EMB) mixture can be used to stain acid-producing fermenting organisms.

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Combined Selective and Differential Media

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Contains reagents tolerated only by the organisms of interest, but also includes reagents that aid in differential identification of characteristics. For example, the mannitol-salt agar (MSA) contains a high (7.5%) concentration of NaCl that is tolerated only by
Staphylococcus sp., but also contains phenol red to differentiate between the S. aureus that can ferment the mannitol from the non-fermenting S. epidermidis.

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Sterilization:

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Totally inactivation of all living organisms.

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Disinfection:

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Killing of organisms (not spores) on a surface. Three levels: high, medium, and low.
Not safe for bodily tissues.

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Antisepsis

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Killing or prevention of growth, can be used on bodily surfaces.

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Sterilization Physical

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Moist heat: 120° C for 20 min @ 15 psi (2 atm) Dry Heat: 180° C for 120 min
Incineration
Radiation: UV, X-rays, γ-rays, microwaves Filtration

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Sterilization (cont) Chemical

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Ethylene Oxide Formaldehyde Glutaraldehyde Peracetic Acid Hydrogen Peroxide Ozone

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Disinfection

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High Level
Moist heat, glutaraldehyde, peracetic acid, H2O2, Cl2 Medium Level
Alcohols, iodophores, phenolics Low Level
Quaternary ammonium compounds

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Antisepsis

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Chemical
Alcohol
Iodophores Chlorhexidine Parachlorometaxylenol Trichlosan