Microbiology: Exam 1 Essay

Essay Question Preparation

Part III. An essay – Dynamics and control of prokaryotic growth. (Up to 20 points available). Each part answered completely and correctly counts as 4 points. Each part must be clearly marked and separated from other part of the essay.

a) Principles and Phases of prokaryotic growth – Describe: • Exponential growth of bacteria: explain specifics of bacterial cell division and growth. Describe equation used to calculate the bacterial population – Lecture 2 – slides 2, 3 and notes • Phases of bacterial growth curve: Describe what occurs at each phase of growth – Lecture 2 – slide 7 and notes

Bacteria grow by means of binary fission:

1.     Chromosome attaches to plasma membrane.

2.     Chromosome duplicates and cell enlarges.

3.     DNA is moved into each future daughter cell, and cell wall is formed.

4.     Cell is divides into two and separates.

Bacteria grow exponentially. Formula: N_n = N₀ * 2ⁿ, where N₀ = initial # of cells and n = # of generations.

Bacteria growth phases:

1.     Lag: Bacteria still adjusting to environment and preparing to grow.

2.     Log - Early (exponential growth): Production of primary metabolites

3.     Log – Late (exponential growth): Production of secondary metabolites

4.     Stationary: Rate of death = rate of growth, using dead cells’ nutrients. Stage of endospore formation for some bacteria.

5.     Death: Rapid decrease in pop. By more than 99%, As cell death > cell growth

6.     Prolonged decline: Remaining live cells undergo gradual decrease on pop.

b) Environmental factors influencing microbial growth – Describe and give species examples of bacterial groups by their requirements of: • Temperature: Describe 5 bacterial groups and their importance to humans – Lecture 2 – slide 8 and notes • Oxygen: Describe 5 bacterial groups and give example for each – Lecture 3 – slide 9 and notes • pH: name 3 bacterial groups and their pH range – Lecture 2 – slide 10 • Water availability: describe two bacterial groups based on their salt tolerance – Lecture 2 – slide 10 and notes

Psychophiles (Optimal 5ºC – 10ºC): Inhibit artic regions, NO significant importance

Psychotrophs (Optimal 20º - 28ºC): Responsible for food spoilage in refrigerator

Mesophiles (Optimal 30º - 37ºC): Majority of pathogens

Thermophile (Optimal 60º - 80ºC): Hotsprings, source of useful enzyme (PCR rxns)

Extreme thermophile (Optimal 85º - 95ºC): Hydrothermal vents.

Obligate aerobes – Need oxygen Mycobacterium tuberculosis, Bordetella pertussis

Facultative anaerobe – Grow better in oxygen, but can live w/o Escherichia coli, S. aureus

Obligate anaerobe – Grows in absence of oxygen, O₂ toxic Clostridium tetani, C. perfringens

Microaerophiles – Grows in small O₂ concentrations (2-10%) Helicobacter pylori

Aerotolerant microbes – Grows w/ or w/o O₂, obligate fermenters Streptococcous pyogenes

pH acidophile – Listeria monocytogenes, Acetobacter aceti

pH neutrophile – Escherichia coli

pH alkaliphiles – Bacillus halodurans, Microbacteriaceae spp.

Vast majority of bacteria prefer isotonic solution

Bacteria that grow in high salt concentration: facultative halophiles, halophiles

Facultative halophiles: Bacillus subtilis, Staphylococcus epidermidis, Staphylococcus aureus

Bacteria sensitive to salt: Escherichia coli, Streptococcus pyogenes

c) Nutritional factors influencing microbial growth & Cultivating prokaryotes in the laboratory – Describe: • Essential elements, Growth factors and fastidious bacteria: give a definition for each and give examples – Lecture 2 – slide 11 • Energy sources used by bacteria – name 3 bacterial groups and source of energy they use – Lecture 2, slide 11 • Define selective media and use a particular example to describe how they work – Lecture 2 – slides 13, 14 and notes • Define differential media and use a particular example to describe how they work – Lecture 2 – slides 14 and notes

Essential elements = all chemical elements required to grow bacteria (carbon, nitrogen, hydrogen

Trace elements only needed in small amounts: zinc, iron, manganese

Growth factors = Complex organic molecules, needed for fastidious bacteria to grow. (ex: beef extract, blood)

Fastidious bacteria = Bacteria with very specific requirements for growth (ex: Neisseria gonorrhoeae)

Photoautotroph – sunlight, CO₂ carbon source (ex: cyanobacteria)
Photoheterotroph – sunlight, Organic C source (ex: purple non-sulfur bacteria, green non-sulfur bacteria, and heliobacteria)

Chemolithoautotroph – inorganic energy, CO₂ C source (ex: sulfur bacteria, nitrogen fixing bacteria)

Chemoorganoheterotroph – organic molecules for both energy and C source (human pathogens)

Selective media:

MacConkey agar → bile salts selects for only gram negative bacteria (in particular enteric bacteria)

EMB agar → dyes select for gram negative bacteria

Thayer-Martin → Selects for N. gonorrhea, but preventing growth of other bacteria and fungi (vancomycin, colistin, treimethoprim, nystatin).

Differential media: Separate growing bacteria into groups. Maybe overlap with selective.

MacConkey agar → Differentiates between fermenting (pink) and non-fermenting bacteria (no change).

EMB agar → Differentiates between non-lactose fermenters (colorless) and lactose fermenters (produce lactic acid making colony stained dark, esp. E. coli = green metallic sheen).

Blood agar → alpha-hemolytic (green ring = limited pathogenicity) , beta-hemolytic (white halo = extreme pathogenicity), gamma-hemolytic (No effect - Less likely to be pathogenic).

d) Selection of antimicrobial procedure – Describe how selection of antimicrobial

procedure is affected by:

• Type of microbes present (use the graphs reviewed in lecture): give

examples of the most and the least resistant microbes. Define decimal reduction

time and explain how it affects duration of antimicrobial treatment – Lecture 2 –

slide 20 and notes • Number of microbes (use the graphs reviewed in lecture): explain how the

level of microbial contamination affects duration of treatment – Lecture 2 – slide

20 and notes • Risk of infection & medical instruments: describe how medical instruments

are treated depending on its use – Lecture 2, Slide 21 –

• Composition of the item: describe how the composition of an item to be

treated affects the selection of the antimicrobial treatment – Lecture 2, Slide 21

Most resistant = Prions

Endospores

Mycobacteria → acid-fast, hydrophobic waxy cell wall

Protozoan cysts → disinfectant resistant, but heat sensitive

Vegetative protozoan

Gram negative bacteria

Fungi

Naked viruses

Gram positive bacteria

Least resistant = enveloped viruses

Decimal reduction time (D) – Time required to reduce microbial population by 90%.

t = D * (lgN+1)

Longer decimal reduction time = longer treatment

If sample with 10⁴ microbes D = 1min, sample is sterilized in 4 mins → D = 2mins, sample is sterilized in 8 mins.

More microbes present in sample = longer treatment (Direct relationship)

If sample with 10⁴ microbes D = 1min, sample sterilized in 4mins → sample with 10² microbes, sample sterilized in 2mins.

Risk of infection:

Critical instruments - contacts with open wound (sterilized)

Semi-critical – mucous contact (disinfected)

Non-critical instruments – skin contact (sanitized)

                                                                             

Composition of item affects choice of treatment:

- Moisture sensitive → NO water/steam → Dry heat/(g) steriliants

- Heat sensitive → NO heat → (g) steriliants, irridation, chemical solutions

e) Use of Heat, Alcohol and Ethylene oxide to destroy microbes – Describe: • Moist heat: compare conditions (time, temp, pressure etc) and outcomes (what is killed, what survives) of pasteurization, boiling and autoclaving – Lecture 2 – slide 22 and notes • Dry heat: compare conditions and outcomes of flaming and hot air sterilization – Lecture 2 – slide 23 and notes • Alcohol: describe conditions of treatment. Compare the outcomes of alcohol treatment to outcomes of autoclaving – Lecture 2 – slide 27 • Ethylene oxide: describe conditions of treatment. Compare the outcomes of ethylene oxide treatment to that of autoclaving – Lecture 2 – slide 27

Boiling – 100ºC for at least 5mins

Pasteurization (3 settings):

Classical 63ºC for 30mins

High temp. Short Time 72ºC for 15sec

Ultra high 140ºC for 2 sec

Autoclaving 121ºC at 15psi for 20mins → Kills all except prions

Dry heat 170ºC for 2hrs / 200ºC for 1.5hrs

Flaming – burns microbes instantly 600 – 1600ºC

Alcohol requires at least 60% alcohol for 10 sec → kills bacteria and enveloped viruses, but NOT non-enveloped viruses and endospores

Ethylene oxide – Very strong oxidant that damages proteins, nucleic acids, lipids → Kills everything (except maybe prions). Treatment 3-5hrs → 8-12 heated forced air to remove possible carcinogenic chemicals.