Bacterial Metabolism & Growth Flashcards
Biochemical reactions bacteria use to break down organic compounds and reactions they use to synthesize new bacterial parts from the resulting carbon skeleton
Bacterial metabolism
Biochemical processes of metabolism by which molecules are broken down.
Catabolism
Synthesis of complex molecules in living organisms from simpler ones together with the storage of energy; constructive metabolism.
Anabolism
Diagnostic schemes analyze each unknown microorganism for:
- Utilization of various substrates as a carbon source
- Production of specific end products from various substrates
- Production of an acid or alkaline pH in the test medium
Breakdown of chemical substrate through the degradative process of catabolism coupled with oxidation reduction reactions
Energy Production
Bacteria use biochemical pathways to catabolize (break down) carbohydrates and produce energy by two mechanisms:
- Fermentation
- Respiration (oxidation)
• Anaerobic process carried out by both obligate and facultative anaerobes
• Electron acceptor is an organic compound less efficient in energy generation beginning substrate is not completely reduced
• A mixture of end products lactate, butyrate, ethanol, and acetoin accumulates in the medium identification of anaerobic bacteria
FERMENTATION
• Efficient energy generating process
• Molecular oxygen is the final electron acceptor
• Obligate aerobes and facultative anaerobes
RESPIRATION (OXIDATION)
True or false:
Certain anaerobes can carry out anaerobic respiration, in which inorganic forms of oxygen, such as nitrate and sulfate act as the final electron acceptors
True
Three major biochemical pathways bacteria use to break down glucose to pyruvic acid:
Embden Meyerhof Parnas (EMP) Glycolytic pathway
Pentose Phosphate Pathway
Entner-Doudoroff Pathway
• Major pathway in conversion of glucose to pyruvate
• Generates reducing power in the form of NADH 2
• Generates energy in the form of ATP
• Anaerobic does not require oxygen
o Example:
▪ Enterobacteriaceae
Embden Meyerhof Parnas (EMP) Glycolytic pathway
• Alternative to EMP pathway
• Glucose to ribulose 5 phosphate, which is rearranged into other 3 4 5 6 and 7 carbon sugars
• Provides pentoses for nucleotide synthesis
• Produces glyceraldehyde 3 phosphate, which can be converted to pyruvate
• Generates NADPH, which provides reducing power for biosynthetic reactions
• May be used to generate ATP
Pentose Phosphate Pathway
• Glucose 6 phosphate (rather than glucose) to pyruvate and glyceraldehyde phosphate
• Generates one NADPH per molecule of glucose but uses one ATP
Entner-Doudoroff Pathway
Anaerobic Utilization of Pyruvic Acid (Fermentation):
• Major end product is ethanol
• Example: yeasts
Alcoholic Fermentation
Anaerobic Utilization of Pyruvic Acid (Fermentation):
• End product is almost exclusively lactic acid
• Example
o All members of the Streptococcus genus
o Members of the Lactobacillus genus
Homolactic Fermentation
Anaerobic Utilization of Pyruvic Acid (Fermentation):
• In addition to lactic acid, the end products include carbon dioxide, alcohols, formic acid, and acetic acid
• Example: Some lactobacilli
Heterolactic Fermentation
Anaerobic Utilization of Pyruvic Acid (Fermentation):
Anaerobic Utilization of Pyruvic Acid (Fermentation):
• Propionic acid is the major end product of fermentations
• Example:
o Propionibacterium acnes
o Some anaerobic non spore forming, gram positive bacilli
Propionic Acid Fermentation
Anaerobic Utilization of Pyruvic Acid (Fermentation):
• Produce a number of acids as end products lactic, acetic, succinic, and formic acids
• Strong acid produced is the basis for the positive reaction on the methyl red test
o Example: Members of:
▪ Escherichia
▪ Salmonella
▪ Shigella
Mixed Acid Fermentation
Anaerobic Utilization of Pyruvic Acid (Fermentation):
• End products are acetoin (acetyl methyl carbinol) and 2,3-butanediol
• Detection of acetoin is the basis for the positive VP reaction
• Little acid is produced by this pathway
Butanediol Fermentation
True or false:
Organisms that have a negative VP reaction usually have a negative reaction on the methyl red test, and vice versa
False. Organisms that have a POSITIVE VP reaction usually have a negative reaction on the
methyl red test, and vice versa
Anaerobic Utilization of Pyruvic Acid (Fermentation):
• Involves the conversion of pyruvate to butyric acid along with acetic acid, CO2 and Hydrogen
• Example:
o Clostridium
o Fusobacterium
o Eubacterium
Butyric Acid Fermentation
- Most important pathway for the complete oxidation of a substrate under aerobic conditions
- Pyruvate is oxidized, carbon skeletons for biosynthetic reactions are created, and the electrons donated by pyruvate are passed through an electron transport chain and used to generate energy in the form of ATP
- Results in the production of acid and the evolution of carbon dioxide
Krebs or Tricarboxylic acid (TCA) Cycle
• Use of various sugars (carbohydrates)
• Fermentation is usually detected by acid production and a concomitant change of color resulting from a pH indicator present in the culture medium
Carbohydrate Utilization and Lactose Fermentation
True or false:
Glucose must not be present if the ability to ferment another sugar is being tested.
True
True or false:
All organisms that can ferment lactose can’t ferment glucose
False. All organisms that can ferment lactose can also ferment glucose
Classifying members of the Enterobacteriaceae family → ability to ferment lactose
- β galactoside permease → transport of lactose across the cell wall into the bacterial cytoplasm
- β galactosidase → break the galactoside bond, releasing glucose, which can be fermented
Energy Utilization:
- Biosynthesis of new cell components
- Maintenance of the physical and chemical integrity of the cell
- Activity of the locomotor organelles
- Transport of solutes across membranes
- Heat production
Time required for one cell to divide into two cells
Generation Time / Doubling Time
Generation time of a fast-growing bacterium such as E. coli
20 minutes
Generation time of a slow-growing bacterium such as Mycobacterium tuberculosis
24 hours
Growth curve:
Little to no multiplication but enzymes are
very active. A period of adjustment and adaptation
Lag phase
Growth curve:
- Organisms grow at maximum rate (exponential rate)
- Most sensitive to antibiotics
Logarithmic/exponential phase
Growth curve:
Plateau-growth ceases because nutrients are exhausted or toxic metabolic products have accumulated
Stationary phase
Growth curve:
Direct microscopic count may remain constant but viable (alive bacteria) count slowly decreases
Decline and death
DETERMINATION OF CELL NUMBERS
Direct counting under the microscope
Direct plate count
Density measurement
o Estimate the number of bacteria present in a specimen
o Does not distinguish between live and dead cells
Direct counting under the microscope
o Dilutions of broth cultures on agar plates—colony forming units per milliliter (CFU/ml)
o Provides a count of viable cells only
o Determining the bacterial cell count in urine cultures
Direct plate count
o Correlated to CFU/ml of the culture
o Prepare a standard inoculum for antimicrobial susceptibility testing
Density measurement
DIRECT MEASUREMENT OF MICROBIAL GROWTH:
Plate counts
Filtration
Most Probable Number (MPN) Method
2 types of plate counts
Serial dilutions
Pour plates and spread plates
o Measures the number of viable cells
o Often reported as colony-forming units
Plate counts
1.0 mL or 0.1 mL of dilutions of the bacterial suspension is introduced into a Petri dish
Pour plates
0.1 mL inoculum is added to the surface of a pre-poured, solidified agar medium; spread uniformly over the surface of the medium with a specially shaped, sterilized glass or metal rod
Spread plates
o Water is passed through a thin membrane filter whose pores are too small to allow bacteria to pass
o Detection and enumeration of coliform bacteria
Filtration
o Statistical estimating technique
o Based on the fact that the greater the number of bacteria in a sample, the more dilution is needed to reduce the density to the point at which no bacteria are left to grow in the tubes in a dilution series
Most Probable Number (MPN) Method
• Measured volume of a bacterial suspension is placed within a defined area on a microscope slide
• Used to count the number of bacteria in milk
DIRECT MICROSCOPIC COUNT
Used in direct microscopic counts
Petroff-Hausser cell counter
ESTIMATING BACTERIAL NUMBERS BY INDIRECT METHODS
Turbidity
Metabolic activity
Dry weight
