-temperature

-pH

-osmotic pressure

• a cold-loving microbe
• an organism that grows best at about 15^oC & does not grow above 20^oC

an organism that is capable of growth between about 0^oC & 30^oC

• a moderate-temperature-loving microbe
• an organism that grows between about 10^oC & 50^oC

• a heat loving microbe
• an organism whose optimum growth temperature is between 50^oC & 60^oC

• an organism whose optimum growth temperature is at least 80^oC
• also called extreme thermophile

- minimum, optimum, maximum

* most bacteria: optimum= 6.5-7.5

* molds and yeasts: optimum= 5-6

* human pathogens: optimum= 7.4 (pH of blood)

- acidophiles: acid-tolerant microbes

* stomach pH: 1 (Helicobacter pylori)

* vagina pH: 3-5 (Lactobacillus)

- acids from fermentation preserve foods

- microbial wastes are acidic

* use buffers in media

• a solution that has a lower concentration of solutes than an isotonic solution
• water moves inside the cell & the cell will lyse
• gram-negative bacteria are more susceptible to this
• most microbes is in this type of environment

• a solution that has a higher concentration of solutes than an isotonic solution
• water moves out the cell & the cell shrink
• extreme & facultative halophiles

• energy
• carbon
• hydrogen
• nitrogen
• oxygen
• phosphorus
• sulfur
• trace elements
• organic factors

• 50% of dry weight of a typical bacterial cell
• obtain through energy sources (carbs,lipids,proteins) or CO₂
• autotroph: CO₂ is carbon source
• heterotroph: organics as carbon source

needed for protein, nucleic acid & ATP synthesis

needed for protein (in Cys & Met) & vitamin synthesis (Biotin & thiamine)

• DNA & RNA
• phosopholipids
• ATP

• potassium, magnesium, calcium
• trace elements: iron, copper, molybdenum, zinc
• organic molecules obtained from the environment

• molecular oxygen (o₂) yields toxic products
• aerobes require O₂ for aerobic respiration
• anaerobes don't use O₂ for energy production
• anaerobic respiration & fermentation

- radicals are formed

* when cells grow in O₂ environment

* ionizing radiation

- O₂ ^-: superoxide OH^-: hydroxyl ion H₂O₂: hydrogen peroxide O₂ ^2-: peroxide ion

- unstable; disrupt electron arrangements around other molecules > DNA or cell damage

- naturally occurring antioxidants

* tocopherol (vitamin E)

* ascorbic acid (vitamin C)

* melatonin

- detoxifying enzymes

* superoxide dismutase

* catalase

* peroxidase

- superoxide dismutase (SOD)

O ₂ ^- + O₂ ^- + 2H⁺ > H₂O₂ + O₂

- catalase

2H₂O₂ > 2H₂O + O₂

-Peroxidase

H₂O₂ + 2H ⁺ > 2H₂O

• reducing medium
• take the oxygen & reduce it into water O₂ > H₂O

organisms that require oxygen to live

an organism that can grow with or without oxygen

organisms that does not use oxygen to live & is killed in the presence of oxygen

an organism that does not use oxygen to live but is not affected by its presence

an organism that grows best in an environment with less oxygen than is normally found in air

- bacteria are usually found in communities

* rarely as isolated species

- matrix of polysaccharide, DNA & proteins

- quorum sensing

* communication via chemical cues

- benefits

* share nutrients

* shelter

* increase opportunity for genetic exchange

* cooperative activity

- impact human health

* 70% of human infections involve biofilms

~ responsible for most nosocomial disease

* form on catheters & medical devices

* more resistant to antimicrobial agents

- medium: nutrient preparation used to grow microbes

* H₂O, carbon, required elements, O₂ (if needed)

* correct pH and temp

* broth vs agar

*steril

- chemically defined vs. complex medium

selective & differential medium

- anaerobic growth

- special culture techniques

* selective & differential medium

- supports growth of specific organisms while inhibiting others

- eosin methylene blue (EMB)

* selects for gram-negative

~ enterobacteria- enteric/gut bacteria

~ facultative anaerobes

* dyes inhibit growth for gram-positive

• selects for gram-positive species
• little/no growth of gram-negative

• selects for facultative halophiles (7.5% salt)
• staphylococcus

selects for fungi (pH 5.6)

- distinguishes between microbes based on identifiable reactions with the medium

- differential media

- contains 5% sheep's blood

- hemolysins: exotoxins that destroy RBCs

* alpha-hemolysis > partial lysis

* beta-hemolysis > total lysis

* gamma-hemolysis > no lysis

• greenish-brown halo
• Streptococcus pneumoniae

• clear halo around colonies
• Streptococcus pyogenes
• Staphylococcus aureus

most bacteria

• selects for gram-negative species
• differentiates between lactose/sucrose fermenters and non-fermenters

• selects for Staphylococcus (high salt 7.5%)
• differentiates between mannitol fermenters and non-fermenters

-reducing media

* contains ingredients that deplete O₂

* example: Thioglycollate medium (liquid)

- anaerobic jars

* used for plate cultures

* colony growth

- anaerobic chamber

- animals

* some microbes don't grow on artificial media

* Mycobacterium leprae

~ mouse foot pads and armadillos (low temp)

* Treponema pallidum

~ rabbit testes

- cell culture (intracellular bacteria: example: Chlamydia)

- growth of capnophiles

* require elevated CO₂ but are aerobic

~ intestines, respiratory tract, body tissues

* CO₂ incubator

* candle jar

- most reproduce by binary fission

- budding

* outgrowth of cell that grows and separates from parent

- external spores (conidiospores)

* filamentous bacteria

* spores develop into new cells

* more common for fungi

-fragmentation

* new cells grow from fragments

- time required for cell number to double

- factors affecting generation time

* species of microbe

* environmental: temp, pH, nutrients, type of media

- exponential growth

- cell # increases by a constant factor for each unit of time

- Log: the power to which a base number is raised to produce a given number (Log₁₀1=10 & Log₁₀6=1000000)

* bacterial growth is linear on log graph

little or no change in the number of cells, but metabolic activity is high

bacteria multiply at the fastest rate possible under the condition provided

there is an equilibrium between cell division & death

the number of deaths exceeds the number of new cells formed

• population numbers (cells/ml or cells/g)
• direct measurement
• indirect measurement

• technique that involves some counting
• count small sample & determine total population

• estimation of cell numbers without counting cells
• turbidity, mass, spectrophotometry

- most frequently used

- advantage: measures only living cells

- disadvantage: takes time

- cell vs colony forming unit

* colony can form from 1 cell or a clump of cells

* plate counts are in CFU/mL

- serial dilutions to get good # colonies/plates

* 30-300 CFU/mL

- pour plate

- spread plate

- volume passed through membrane (bacteria trapped)

- transfer to plate & grow

- count colonies & divide by number of mL filtered (CFU/mL)

- good when bacteria # is small

* can't count 1 CFU/mL

*example: water samples (coliform bacteria)

• count number of cells spread over grid using a microscope
• average # microbes in several squares
• average X conversion factor X dilution factor
• advantages: quick (no incubation required) & counts all cells (live & dead)
• disadvantage: difficult to count motile bacteria

- indirect

- uses spectrophotometer

* measures amount of light passing through a sample

* % transmittance

* absorbance

- # cells is directly proportional to turbidity of culture

* increase # cells/ml (turbidity), increase absorbance

- advantage: quick & easy

- disadvantages: must be read during log phase & needs to be correlated to direct measurement