Culturing Microbes Flashcards
what is a microbial cell made of
-55% protein
-25% nucleic acid
-lipid
-polysaccharide
chemical components of microbial cell
-CHONPS
-HOPS can come from organic molecules
Mg 2+ role in microbial cells
-key role in stabalizing negative charges in membranes
-also used by enzymes
growth media or culture media
-can be highly varibale depending on microbe
defined media
-media prepared by adding precise/known quantities of chemicals to water
-know the exact composition
-know what your working with
complex media
-contains extracts or digested organic material with unknown composition
-cheaper easier and work for a broader arrray of different microbes
What microbes can make most or al of the organic molecules they need from a few basics
-microbes living in nutrient poor environments
-certain flexible microbes that adapt to many different environments
microbes that require a great number of growth factors
-many organisms that have olbigate symbiotic lifestyle
-organiss that live in nutrient rich environemtns such as LAB
auxotrophy
-inability to produce a molecule you need for gorwth
what is ecoli grown on
-simple concotion
-grown on complex media
what is lactic acid bacterium, l.mesentheroides, need to grow
-requires a number of organic building blocks such as AA and vitamins
selective media
-used to isolate a limited assortment of microbes
-combination of positive selection and negative selection
enrichment culture
-similar to selective media but less selective and richer media
-promotes geowth to increase cell numbers of particular microbes from samples
-usually somewhat selective
differential media
-contwan some sort of indicator when particular organisms are present
what are solid plates most useful for
-usefule for isolating single colonies
-ideally single colony oiginates from single cell that grows to large numbers and can be readily seen
what should liquid cultures geenrally be started using
-isolated single colonies from agar plates
great plate count anomoly
-when environmental samples are plated on generic growth plates meant to be non-selective, >99% of the microbes dont grow
why can we not culture the majority of microbes in isolation
-syntrophy`- microbes feeding off one another, metabolismintegrated between multiple different species
direct count
-cells in the sample are counted using a microscope
viable plate counts
-a small sample of cells are spread on agar plates and incubated - colonies counted
turbidimetric
-absorbance of light by a microbe growing in a lqiuid culture is measured in a spectrophotometer
other indirect methods of counting mircobial cells
-O2 consumption, CO2 production, metabolic activity
-Quantitative PCR to determine the number or genome equivalents
how does direct microscope counting work
-known volume added to gridded microscope slide/coverslip and cells are counted under microscope
-doesnt require growth but is laborious and prone to inaccuracies and you likely need to stain microbes
-hard to differentiate between viable and dead cells
how does viable plate counts work
-common and reliable
-have to know how to grow microbes
-assumes colonies emerge from single cell
-assumes all cells will grow to form colony, though some cells can be viable but none culturable under plating conditions
how does turbidity measurements work
-microbes catter light and amount of light scattered proprotional to number microbes in a liquid sample
-very common
-need to find correlation between optical density at chosen wavelength and cell numbers
-only works in pure cultures and liquid cultures
-limited concentration range
-not labour intensive and growth can be continuosly or regularly measured over time
-growth can be visually oserved
-need right amount of cell numbers (not too low and not too high_
-time on X axis and OD on Y axis
microbial growth definition
-refers to increase in population size
-cell division resulting in multiplying in numbers
generation time
-the amount of time it takes for one cell to become two
-varies greatly depending on mcirobe and growth conditions
batch cultures
-cultures in a fixed volume in a closed contained like a flask or test. tube
continous cultures
-cultures within systems where waste product are being removed and new media fed in
lag phase in batch cultures
-period of slow or no growth
-,icrobes adjust to new environment
-length of this phase varies
-bigger adjustments = longer lag phase
exponential phase in batch cultures
-growin population doubles at regular intervals
-nutrients not yet exhausted waste products not slowing growth
stationary phase
-nutrients begin to be exhaustedand or accumulating waste products inhibit growth
-little to no net growth, still some growth and death
decline phase
-cells start to die if left long enough
-net decline in celel numbers
-this phase isnt all relevant
generation time equation
-growth time/number of generations
total number of cells at any given time equation
-Nt =N0 x2^n
-Nt = total number of cells
-N0 = starting number of cells
-2^n = number of generations
biofilm formation
-sessile growth can develop into biofilms
-cells encased in matrix attached to surface
-matrix is often polysacchardies but can also include protein and DNA
-many times it a complex community with complex cellular differentiation
-cells with different properties in different layers of biofilm
sessile growth vs planktonic growth
-sessile - growth on surface
-planktonic growth - free living organisms in liquid
formation of biofilms
-can start with planktonic cells attaching to a surface via appendages such as pili, fimbriae or even flagellum
-colonization begins and cells multiply and produce extracellular polymers such as polysaccharides to hold matrix together
-during decelopment, cells biological programs change
-cells with begin to disperse an resume planktonic state
how does cells change their programing during biofilm formation
-express unqiue combinations and amounts of genes to facilitate this biofilm lifestyle
-different cells on different layers have different programs
