Chapter 4 - Microbial Growth PDF

Chapter 4 Microbial Growth Bacterial Growth Refers to increase in bacterial cell numbers not an increase in size of individual cells Most bacteria reproduce by binary fission The bacterial cell: Elongates and makes a copy of its DNA Divides into two identical cell...

Chapter 4 Microbial Growth Bacterial Growth Refers to increase in bacterial cell numbers not an increase in size of individual cells Most bacteria reproduce by binary fission The bacterial cell: Elongates and makes a copy of its DNA Divides into two identical cells. Exponential growth Because bacteria divide by binary fission, the population of cells will double every generation Time required for population to double = generation time Varies greatly between different bacteria E. coli has generation time = 20 minutes Mycobacterium tuberculosis generation time = 24 hours. Bacterial Growth in the Lab Culture: microbes growing in a medium Inoculation: introducing microbes into a medium to start a culture Batch culture Closed system Once started, no other nutrients are added When nutrients are used up – bacteria stop growing Continuous culture Open system Nutrients are continuously added, wastes are continuously removed Supports indefinite growth. The Growth Curve in Batch Culture Lag phase A period of adaptation Cells adjust to new media and get ready to grow Exponential phase (log phase) Period of maximal reproduction - Cells numbers increase exponentially Used to calculate generation time. Stationary phase Cells have reached maximum population density Nutrients have been used up, or wastes have accumulated No increase in cell number Death phase Toxic waste products have accumulated Cells die at a uniform rate. Phase of prolonged decline Sometimes a small fraction of population survives the death phase May consume nutrients released from dying cells Selects for the strongest cells in the population. Environmental factors that influence bacterial growth Temperature requirements Each species of microbe has it’s own specific temperature range This range usually spans about 30°C minimum – lowest temp supporting growth optimum – temperature that supports best growth maximum – highest temp supporting growth. Bacteria can be grouped based on temperature range Psychrophiles – cold loving Grow between – 5°C and about 15°C Killed at 20°C Psychrotrophs – have a very broad temp range Min: about – 5°C Max: about 30 – 45°C Optimum: 15 – 30°C These are the microbes that cause food to spoil in your fridge! Mesophiles – moderate temperature loving Min: about 10°C Max: about 45°C Optimum: 25 - 45°C Most bacteria are mesophiles Most pathogens (diseases causing microbes) have temp optimum of 37°C. Thermophiles – heat loving Min: about 40°C Max: about 80°C Optimum: about 65°C Hyperthermophiles Min: about 75°C Max: up to 121°C Restricted to very few places on the Earth where water reaches these temperatures ex. Deep ocean vents. Food Safety Involves the use of both hot and cold temperatures Heat is used to kill mesophilic and psychrotrophic microbes ie. Cooking Cold temp is used to slow growth ie. Only psychrotrophs will grow in a refrigerator – and slowly. Oxygen (O2) requirements Only required by some organisms, can be extremely toxic to others! Obligate aerobes – require O2 for respiration (energy generation) Facultative anaerobes – Can use O2 for respiration but can also grow in its absence. Oxygen (O2) requirements Obligate anaerobes – cannot use O2, and are killed by it Microaerophiles – require O2 in low amounts, but killed by high concentrations Aerotolerant anaerobes – Cannot use O2, but are not killed by it. pH Measurement of acidity or alkalinity pH below 7 = acidic pH above 7 = alkaline pH of 7 = neutral Most bacteria grow at or near neutral pH ie. pH 6.5 – 7.5 Bacteria that grow at very low pH: Acidophiles Bacteria that grow at high pH: Alkaliphiles. Osmotic pressure Osmosis is the movement of solvent molecules across a semi- permeable barrier ex. Movement of water through the cytoplasmic membrane H2O will move from area of high concentration to area of low concentration Hypertonic solution Hypotonic solution High solute concentration Low solute concentration ex. Salt or sugar Water flows into cell Water flows out of the cell Cell bursts – osmotic lysis Cell dries up – plasmolysis Isotonic solution Condition where solute concentration on outside of cell is equal to that inside the cell. Osmotic pressure is important in food preservation ex. Salted fish, honey Some bacteria have adapted to life in high salt concentrations – requiring up to 30% NaCl These bacteria are termed: extreme halophiles ex. Bacteria that live in the Dead Sea Blood has a salt concentration of about 0.9% Does not inhibit the growth of most microorganisms. Nutritional Factors that Influence Growth 1. Carbon Required for all organic molecules – backbone of living matter Heterotrophs – Take carbon from organic matter (ex. sugars) Autotrophs – Use inorganic carbon (ie. CO2) 2. Nitrogen (N), Sulfur (S) and Phosphorus (P) Required in smaller amounts for synthesis of cellular material ex. Protein, nucleic acids, phospholipids, ATP 3. Trace elements Required in very small amounts ex. Iron, zinc, molybdenum Essential to the function of certain enzymes. 4. Energy Organisms need energy to build cell material and drive cellular processes Chemotrophs – Acquire energy from chemical compounds May be organic or inorganic ex. Sugars Phototrophs – Harvest energy from sunlight. Nutritional Diversity Organisms are classified based on how they obtain their carbon and energy Photoautotrophs Use sunlight for energy CO2 as carbon source } Process called: Photosynthesis includes – some bacteria – Algae – Plants Photoheterotrophs Use sunlight for energy Obtain organic carbon from food Some bacteria. Chemoautotrophs Obtain energy from inorganic chemicals ex. H2, H2S Use CO2 as carbon source Only done by some bacteria Chemoheterotrophs Obtain energy from organic chemicals Use the same organic chemicals as their source of carbon All animals, fungi and protozoa Most bacteria All medically relevant bacteria are chemoheterotrophs! Culture media Solid media (Agar Petri plates) Made by adding agar (solidifying agent) to liquid media Cannot be degraded by most bacteria Allows growth of colonies: A genetically identical population of cells Allow the isolation of pure cultures. Culture media can be: Chemically defined – the exact chemical composition of the medium is known ex. Media made from known quantities of salts and sugars Also known as minimal media Chemically undefined - contains rich organic ingredients (so the chemical composition is not known) ex. Media containing yeast extract (all of the soluble components from crushed yeast cells) Also known as complex media. Selective and Differential media Selective media Prevent the growth of unwanted organisms, allowing only the desired microbes to grow ex. Bismuth sulfite agar – used to culture Salmonella typhi Inhibits growth of all Gram positive and most Gram negative bacteria Differential media Used to distinguish different bacteria All can grow – but colonies of certain bacteria look different on the plate Ex. Blood agar plates Used to distinguish bacteria that can lyse (and eat) red blood cells Streptococcus pyogenes. Counting bacteria Direct count Cells are counted using a light microscope Usually employs a special counting chamber Inaccurate because it counts both live and dead cells. Viable counts Only live cells are counted A liquid culture is diluted and plated onto agar plates to grow colonies Each colony on a plate represents a single cell from the original culture Colonies are counted, and used to calculate the number of bacteria in the original culture Counts are always expressed as cfu per ml: colony forming units assumption is that 1 cfu = 1 live bacterial cell.