MICRO Block 3

what features do all cells have in common

cytoplasm; cytoplasmic cell membrane; ribosomes; cell wall (some microbes- strength)

properties of all cells

structure, metabolism, growth (proteins convert nutrients into new cells) , evolution (chance mutations in the DNA)

metabolism in all cells

all cells use information in DNA to produce RNA and protein.
via catabolism and anabolism

catabolism

transforming molecules to produce energy

anabolism

synthesising macromolecules

properties of certain cells

communication; motility, horizontal gene transfer; differentiation

size range of prokaryotes

0.2 µm to \>600 µm in diameter and between 0.5 µm and 10 µm long

size range of eukaryotes

5-100 micrometers

how is surface area to volume ratio an advantage in smaller cells

small cells have a higher SA:V ratio and support greater nutrient and waste product exchange per unit cell volume, making them more efficient than larger cells.

coenocytic fungi

aseptate fungi- no septa; continuous cell mass with 100 000s nuclei

Four Kingdoms of Eukarya

Protista, Fungi, Plantae, Animalia

characteristics of algae

- cell walls with cellulose
- can photosynthesize
- many conformations

fungi

Unicelllular/ multicelllular
- cell walls contain chitin
-derive nutrients from environment
- non motile usually
- asexual (budding/fission)/ sexual (meiosis derived spores) reproduction

protozoa

unicellular
asexual/sexual reproduction
nutrition from organic substances
movement through cilia, flagella, pseudopods

viruses

obligate parasites; only replicate in host cell
do not carry out metabolism
small genomes of DNA /RNA

extremophiles

live in extreme environments

beginning of microscopy

Robert hooke- first to describe microbes
Antoni van leeuwenhook- first to see bacteria

light microscopy

passing visible light transmitted or reflected from the sample through one or multiple lenses to magnify the sample.

bright field microscopy

simplest of all light microscopy which uses white light to illuminate the object

three main parts of compound microscope

mechanical- base, arm stage
magnifying- objective and ocular lens
illuminating - substage condenser, iris, diaphragm, light source

total maginification formula

ocular x objective lens

spherical abberation

found in systems that use spherical lenses;
light rays striking off centre are ore or less refracted than those at the centre
blurring of the image

chromatic aberrations

fringes of colour around the image
caused by dispersion of lens material
- variation of refractive index with wavelength of light

resolving power

ability to measure separation of images close together

numerical aperture

the ability of a lens to gather light

how does oil change the RI of transmitted light

-increases resolving power of a microscope
-oils have high RI

Resolution limit

minimum angular separation between two points that can be perceived
- d\=0.5 x λ/ NA

NA for dry and oil immersion

Dry-0.95
Oil- 1.5

basic dyes

stain cation

acidic dyes

stain anions

simple stains

single basic dye
-highlights entire microorganism
-crystal violet, safranin, methylene blue

mordant

additive that increases affinity of stain to sample
- eg grams iodine stain, forms crystal violet-iodine complex which clumps and is contained in layers of peptidoglycan

differential stains

react differently with different kinds of microorganism
eg gram stain, gram+ purple, gram - pink

four steps of gram staining

1. apply basic dye eg crystal violet to stain gram + bacteria
2. apply mordant- iodine
3. decolourize with 95% ethanol to destain gram - bacteria
4. apply counterstain of safranin to stain decolourized cells pink

why does gram + bacteria retain crystal violet stain

the thick peptidoglycan layer retains the primary stain

capsule stain

india ink

flagella staining

leifson stain

phase contrast mircroscopy

improves contrast of unstained life cells
phase ring- amplifies contrast between RI of cell and surroundings

dark field microscopy

used to observe flagella
light enters from the side, does not go through the slide

fluorescence microscopy

cells glow on black background due to filter
naturally fluorescent or dyed cells

electron microscopy

uses electrons instead of light, the shorter wavelength of electrons gives greater resolution
- operates in vacuum
produces electron micrograph

TEM

transmission electron microscope
-cut bacterium into slices to view internal structure of the cell
-0.2nm resolving power
- specimen stained with high atomic weight substances that scatter electrons and improve contrast

SEM

-specimen coated with thin film of heavy metal
-only visualises surface
scattered electrons collected + projected to produce an image

aseptic technique

A procedure performed under sterile conditions (no living organisms)

enrichment culture technique

isolates microbes having particular metabolic characteristics from nature

Louis pasteur

disproved spontaneous generation theory (that life arose spontaneously from non living material)

Koch's Postulates

1) Pathogen must be present in all disease cases
2) Isolate pathogen, cultivate in pure culture
3) Inoculate into susceptible animal, initiate disease symptoms 4) Re-isolate pathogen, confirm it's the same pathogen

Sergei Winogradsky

demonstrated that specific bacteria are linked to specific biogeochemical transformations
proposed chemolithotrophy

Martinus Beijerinck

enrichment culture technique

Carl Woese

rRNA sequences could be used to infer evolutionary relationships (aligned sequences)

Chemolithotrophy

oxidation of only inorganic compounds to yield energy

the cell envelope

layered structures surrounding cytoplasm
- cell wall
-cytoplasmic membrane
- outer membrane
- s layers

plasma membrane

surrounds cytoplasm
- selective permeability; prevents influx of ions and loss of nutrients

bacterial and eukaryotic cytoplasmic membrane

phospholipid bilayer with embedded proteins
- fatty acid tails ; hydrophobic
- glycerol+ phosphate+ other functional group (sugar; ethanolamine, choline)
ester linkages in phospholipids

membrane proteins

embedded- integral
transmembrane- extend completely across
peripheral- loosely attached/associated with membrane

archael cytoplasmic membrane

ether linkages in phospholipids
isoprenes instead of fatty acids for hydrophobic tails

function of cytoplasmic membrane

permeability barrier (polar and charged molecules must be transported )
protein anchor
energy conservation (proton motor force to drive flagella etc)

eukaryotic plasma membrane

carbohydrates- attachment and receptor sites
glycoproteins (proteins attached to carbs)
sterols- role in membrane fluidity
humans- cholesterol
fungi- ergosterol

active trasport

simple (driven by energy in proton motor force
group translocation (binding proteins and energy from ATP) ABC system (chemical modifications of the substance driven by PEP- phosphoenolpyruvate)

simple transport

symport- solute and H+ cotransported in one direction
antiport- solute and H+ transported in opposite direction

group translocation

substance chemically modified eg glucose phosphorylated
eg phosphotransferase in E.coli
-glucose, fructose, mannose (not as specific as ABC)

ABC transporter system

ATP binding cassette
substrate binding proteins have high substrate affinity

What organisms prefer ABC transporters

extremophiles- as the transporters are very specific, efficient at finding required substances

The cell wall

withstands osmotic and turgor pressure to prevent lysis
maintains shape and rigidity

gram positive and gram negative bacteria

gram positive - thick cell wall make of peptidoglycan and NO outer membrane, purple color
gram-negative - thin cell wall made up of peptidoglycan WITH outer membrane, red-pink color

Bacterial Cell wall

glycan tetrapeptide contains :
-sugar backbone (NAG and NAM joined by B-1,4 linkages)
- peptide attached to NAM
- amino acids, L-alanine, D-alanine, L-Lysine, D-glutamic acid, DAP

Acid fast cell walls

eg TB causing bacteria
- high % mycolic acid (waxy lipid)
- layer outside thin peptidoglycan layer
-cells stick together and stick to surfaces

Archael Cell walls

no peptidoglycan, no outer membrane, S layer protein shell

pseudomurein cell wall

- archaea similar to peptidoglycan
-NAG and NAT
-B1,3 linkages (lysozyme can't hydrolyse)
all AAs are L stereoisomers no D

Eukaryotic cell walls

Plants- cellulose
Fungi (Chitin)
-NAG units
-arthropod exoskeleton
Yeast cells- Glucan and Mannan

Gram - outer membrane

second lipid bilayer external to cell wall
- Lipopolysaccharide layer
- surface recognition, virulence factors, strength
- porins (transmembrane transport proteins)

Lipopolysaccharides

-ionic bonds to divalent cations (MG, Ca)
-Lipid A: endotoxin, when cell dies it is responsible for fever shock and thrombosis
- core polysaccharide: structural stability
-O polysaccharide: extends outwards; function as an antigen

S layer

paracrystalline structure of protein/glycoprotein
- outermost layer
-strength, lysis protection, creates periplasmic space, cell surface interactions, protect cell from host defenses
some archaea without a cell wall depend on s layer for strength.

Glycolax layer( capsule/ slime layer)

polysaccharide coat outside cell envelope
capsule- visible with india ink, tight matrix, tightly attached
slime- loosely attached and easily deformed

function of slime and capsule

-attachment to surfaces
-maintenance of biofilms
-infectivity
-prevent dessication
-macrophage resistant

fimbrae and pili

composed of pillin
-fimbriae: can number in 100s; stick to each other and surfaces
-pili: longer than fimbriae, few per cell
motility and horizontal DNA transfer

Hamus/Hami

archaeal grappling hooks, surface attachment and biofilms

Carbon storage polymers

PHB and PHA
synthesized when C in excess
glycogen

polyphosphate granules

inorganic phosphate
elemental sulfur accumulates in periplasmic granules

magnetosomes

magnetotaxis- migration along magnetic field lines
biomineralized magnetic iron oxides

gas vesicles

confer buoyancy (floating bacteria; remain at surface for sun+oxygen)
conical shaped gas filled structures

Enospores

usually gram +
dormant, tough, non-reproductive
formation triggered by lack of nutrients
not metabolically active
UV, heat, chemical pasteurisation resistant

Exosporium

interacts with environment/host
contains spore antigens that may trigger immune response
triggers germination in favourable environments

spore coat

resistant to toxic molecules
contains enzymes needed for germination

cortex

peptidoglycan- temp resistance

core well

UV and harsh chemical resistance

core

chromosomal DNA encased in SASPs
protect DNA from UV and heat
DPA stabilises the proteins and DNA
ribosomes and other structures

Mechanisms to destroy endospores

-burning/autoclaving
-ionising radiation
-10% sodium hypochlorite
-ethylene oxide (hospital)

flagella

15-20 nm wide
polar, tufts, lophotrichous, amphitrichous etc
filament (flagellin)
hook
Basal body (motor)
gram - : 2 sets of rings
gram + : inner set of rings (no outer membrane so don't require additional rings to attach to membrane)

cell movement

Clockwise - tumbles
counterclockwise- runs

Archaella

smaller than flagella
rotation driven by ATP hydrolysis
proteins unrelated to flagella

surface motility

millions of cells spread over surface to colonise new area
twitching and gliding motility (pili)

twitching motility

type IV pili
extend from one cell pole, attach to surface, retract to pull forward
ATP hydrolysis

gliding motility

smooth, continuous motion along long axis without external structures
only bacteria
helical protein track, corkscrew movement

taxis

Movement toward or away from a stimulus.

phototaxis

phototrophic organisms optimise position for receiving light
scotophobotaxis: entering darkness causes cells to tumble and head back to light
algae, cyanobacteria

magnetotacic bacteria

use magnetic fields to remain upright to swim towards or away from O2
saves time
display aerotaxis

Cytoskeleton (eukaryotes)

microtubules, microfilaments, intermediate filaments

move proteins and chemicals/compounds to the correct area
speed up movement around cell
highly conserved gene sequence

eukaryotic flagella and cilia

whiplike motion do not rotate
dynein protein
cilia are short flagella that beat in synchrony

macronutrients

-carbon
-oxygen
-hydrogen
-nitrogen
-sulphur
-phosphorous
96% dry weight of the cell