MICR221 Module 2

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growth

increase no. population
through binary fission
1 parent--> 2 daughter cells --> cetc
rod and cocci
mean generation time- time taken for one cell to divide e.g. E.coli= 20 mins
plane of division depends as cell arrangement

classification

energy: phototrophs, chemoautotrophs
carbon: autotrophs, heterotrophs

phototrophs

sunlight

chemoautotroph

oxidation of chemical compounds e.g. sugar

autotrophs

inorganic carbon of CO2

hetertrophs

organic carbon-protein, lipids

physical factors for growth

gaseous atmosphere
temperature
pH
osmotic pressue

chemical factors for growth

water 80-90%
energy- phototrophs, chemoautotrophs,
carbon- autotrophs, heterotrophs
nitrogen, phosphate, sulfur

obligate and facilitate aerobe

use superoxide dismatase, catalase

most aerotolerant anaerobes

use superoxide dismatatse, peroxidase

obligate anaerobes

use none

gaseous atmosphere

aerobe require O2= aerobic respiration, electron transport chain, final electron acceptor O2
anaerobes prefer absense of O2= anaerbic respiration-->use electron transport chain--> final electron acceptor exogenous, O2 toxic, or fermentation-->use no electron transport chain-->use ATP synthase by substrate level phosphorylation-->final electron acceptor= endogenous

anaerobic respiration

use electron transport chain
final electron acceptor, exogenous, O2 toxic

fermentation

use no electron transport chain
use ATP synthase by substrate level phosphorylation
final electron acceptor= exogenous

obligate aerobe

needs O2

facultative anaerobe

prefer O2
grows with or without O2

aerotolerant anaerobe

tolerant O2

strict anaerobe

O2 toxic
super oxide radicals toxic

microaerophile

little O2
2-10% O2

caphophile

increase CO2
need CO2

temperature

minimum-slow down
optimum-at peak
maiximum-denature, damage, cell death

psychrophile

-10 to -17 degrees

pyschrotrophs

2 to 35 degrees

mesophiles

14 to 43 degrees

thermophiles

42 to 80 degrees

hyperthermophiles

68 to 105 degrees

pH

optimum pH for growth
netral 6-9 pH
acidophillic bacteria 1-3 pH

osmotic pressure

no of molecules the soln
isotonic
hypertonic- increase conc-sugar h2o plasmolysis
hypotonic- decrease conc-increase cell size

death

loss of ability to multiply

sterilisation

complete killing or removal of all microorganisms- physical and chemcial

disinfection

destruction of microorganism on inanimate objects by chemical means

sanitation

reducing microbial load, inanimate objects to an acceptable level

thermal death pt

lowest temp required to kill all cells in 10 mins

thermal death time

length of time required to kill at a given temp

decimal reduction time

length of time taken to obtain ten-fold reduction

control methods using heat

boiling water
steam-autoclave
dry heat
batch pasteurisation
flash pasteurisation
ultra-heat treatment

boiling water

100 degrees
15 mins
kills cells
doesn't kill spores

steam-autoclave

121 degrees
15 mins
kills cells
kills spores

dry heat

160 degrees
2hrs
kills cells
kills spores mostly

batch pasteurisation

63 degrees
30 mins
kills some cells
doesn't kill spores

flash pasteurisation

71 degrees
15 sec
kills some cells
doesn't kill spores

ultra-heat treatment

140 degrees
1 sec
kills cells
kills spores

methods of measuring growth

microscopy- glass slide, totvboial counts
Spectrometry- density, curve, total counts
Dilution series- Plate count, Viable
Growth is exponential and rapid

measuring growth

population-final no.= initial no x 2(power of n)

primary metabolite

essential for growth, logarithmic phase

secondary metabolite

not essential
accumulate during stationary phase

medias for growth

complex
defined
specialised

complex media

not know exact composition
contains energy and carbon
powder
most microbes grow

defined media

know exact composition
know every component and amount
specific sugars, amino acids, vitamins
specific microbe

specialised media types

transport
enrichment
specialised enriched
selective
differential

transport media

temporary, storage, being transported
no microbial growth, no microbial death
buffers and salts- protective
lack carbon, nitrogen, organic factors

enrichment broth media

specifically encouraging
low no.
give competitive edge
clinical labs- faecal specimen

specialised enriched media

general nutrients
harvests many microbes
encouraging

selective media

encourages growth of same organisms and inhibiting others

differential media

containing indicator to distinguish which organism

blood agar

enrichment and differential
alpha haemolysis- partial lysis RBC- green
beta haemolysis- complete- clear zone
gamma- no haemolysis

hektoen agar

selective
lactose, sucrose, salicin, acid fuchsin, bromo blue
carbohydrate fermented-->acid (yellow/pink)= E.coli
non-fermenters--> blue/green and black colonies from H2S production= Shigella/Salmonella

mannitol salt agar

differential
mainitol, phenol red
mannitol fermented-->acid yellow, pathogens e.g. Saiureus
non-mannitol fermentors-->no change e.g. S. epi

endospore

modified cell wall
pathogenic or non-pathogenic
vegetative-->binary= 8 hrs SPORALATION
indefinite survival time
endospore--> cell = 15 mins

endospore structure

core- DNA and ribosomes but metabolically inactive
cortex- peptidoglycan less cross-linked
spore coat- large several protein layers, impermeable, physical resistant, UV radiation and dessiccation PROTECTION
exosporidium- thin outer most layer, lipid, carbohydrate coat surrounds endospore `

resistance of endospore

1. physical spore coat
2. chemical
- low water content- 15%- gel-like substance= heat resistant, protective
-small acid-soluble proteins- bind DNA altering confirmation, protect from damage
-high content of dipicolinic acid and Ca2+ in core- lattice protein makes resistant

germination of endospore

1. activation- chemicals or heat
2. germination- breaking spore state, swell rupture, loss protection, increase metabolic activity
3. out growth- spore emerges out of coat, divides into active bacterial cell

filtration- control microbial growth

filter with pores too small for microorganisms, but large enough for liquies to pass through
pore size= 0.45um or 0.20um
use e.g. antibiotic, tissue culture mieda, serum, gases, filtered beer

depth filter

asbestos or glass
random array of fibers
particles trapped in fibers
no definite pore size
thick filter- 5mm high
retain some liquid- disadvantage
high dirt handling capacity
cope with alot of microbes
used as pre-filter- containment

membrane filter

cellulose acetate or nitrate
thin piece
pore size regular
like sieve, trap microbe at top
disadvantages: easily blocked by contaminants
therefore depth and membrane filter used

nucleopore filter

absolute pore
low flow rate
low no. of pores
not used often
used in prep for electron microscopy

HEPA

(High Efficiency Particulate Air) filters
laminar flow hood (basic specimen protection)
class II biological safety cabinets (protection of specimen, you and environment from contamination)

radiation- control microbial growth

non-ionising radiation e.g. UV radiation
ionising radiation e.g. xrays

non-ionising radiation

wavelength of 260 nm damages DNA in cell forming pyrimidine diamers or direct protein damage
sterilise benches and air
not used on large volumes of liquids penetrate these in significant depth
gram -ve more insensitive-->+ve-->endospore

ionising radiation

xrays kills indirectly by inducing reactive chemical radicals (free radicals) by breaking individual moelcules into ions
0.3-0.4 millirads-generally kills microbes
>50 rads- human becomes ill from radiation
hydroxyl free radicals= strong oxidising agents
+ve= more penetrating-sterlise products for packaging
-ve= but expensive and need elaborate safety precautions
used to sterilise lab products (petri dishes), treatment of sewerage and industrial sludges, food preservation

SOS Repair System

bacteria possess repair enzymes which can repair damaged pyrimidine diamers
so UV will cause death of bacteria only if the damage incurred is greater then that which can be repaired

Antiseptics

are chemical agents that kill or inhibit growth of microorganisms and that are sufficiently non-toxic to be applied to living tissues

Preservations

included in pharmaceutical preparations to prevent microbial spoilage of product

conditions influencing effectiveness of antimicrobial agents

population size
properties of chemical agents
population composition
environmental factors
toxicity of agent

population size

only fraction of microorganisms die during certain time interval
one log decrease=90% of population killed
if rate of killing is same it will take longer to kill all members of a larger population than a smaller one

properties of chemical agents

pH
dilution

population composition

- Phase of growth- exponential quickly die with chemicals
- Polymer or capsule production- tuberculosis waxy cell wall take a long time to kill
- Altered cell wall or membrane- reduce permeability to chemicals
- Modified sensitive sites- enzymes more resistant
- Cellular aggregation/biofilms- degree of protection
- Resistant structures- endospores- much more difficult to destroy

environmental factors

- Neutralisation by organic material- absorb or chemically inactivate chemical material- milk proteins blood fetal material
- Temperature- higher temperatures than lower temp

toxicity of agent

correct agent to correct microorganism
use and exposurer and environment

ideal characteristics of chemical control agents

high antimicrobial activity
broad spectrum
stability
homogeneity
adequate soluability
minimum toxicity
detergent activity
minimum material effects
minimum inactivation by organic material
activity at ordinary temperatures
deoderising ability
low cost

high microbial activity

be able to kill microorganisms

broad spectrum

of antimicrobial activity

stability

to be able to store for a long period of time, no loss of activity

homogeneity

do not want to settle out or sediment out

minimum toxicity

shouldn't harm environment, animals or humans

detergent activity

remove dirt action

minimum material effects

different surfaces shouldn't harm

activity at ordinary temperatures

do not need to heat up or cooled down at room temperature

deodourising ability

microbes smell a little bit

low cost

within market range

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