microbiology (8) (Kevin Purdy) Flashcards
number of domains
what are they?
3
bacteria
archaea
eukaryotes
bacteria size
1-6 μm
largest are visible (almost mm)
lots cell space, storage - makes big
archaea
once thought to be bacteria
lots are extremophiles - 1st non extremophile found in 2004
no known pathogenic ones
found everywhere
fungi
what is it and size?
eukaryote
size varies enormously
largest is 10,000kg because spreads
protists
what is it and size?
eukaryote most eukaryotes are protists, very diverse usually microscopic and unicellular 1-150 μm e.g. Paramecium
how many cells of microbes on Earth?
4.6 x 10^30
more than the number of stars
what gases are mostly inside microbes?
nitrogen and phosphorous
phototroph
E from light
organotroph
use organic compounds like sugars as electron donors
e.g. humans
lithotroph
use inorganic compounds like water as electron donors
autotroph
use CO2 as carbon source
fixes own carbon
heterotroph
use organic carbon as a carbon source
chemotroph
energy from chemical bonds
cyanobacteria
maybe original chloroplasts
photolithoautotrophs (plants are also this)
E.coli and animals are classed as what in terms of how they use electrons to pass energy around?
chemoorganoheterotroph
bacteria are what kind of trophs (can be 2 types)?
and what does this mean
prototroph - synthesise all their own cellular components like AAs nucleotides and vitamins
fastidious - need to give them some components in media
some microbes have to be grown in eukaryotic cells if require complex media
complex media
like blood, milk, or yeast, of biological origin
exact chemical composition is unknown so complex media is undefined
bacterial growth: asexual reproduction
binary fission or budding
double in size then split
exponential growth
sometimes fail to separate so pairs and long chains
all organisms limited by carrying capacity of medium (space/nutrients)
phases of bacterial growth
lag phase
log phase
stationary
death
measurements of bacterial growth
turbidmetry - light scattered so becomes cloudy (turbid)
weigh biomass
total viable count, colony forming unit - dilute so can count colonies, assume colonies from single cell
identify MOs - microscopy+staining, selective media, test substrates/enzymes for growth, cell characteristics of chemical constituents, sequence genes
selective media
allow growth of only some organisms
to identify pathogens from clinical sample
differential media
identification based on growth and appearance on medium
colour differences from pH can identify if pathogenic
enzyme activity
Apizym test - for pathogens, grow and put cells into wells, compare colour against database
surface origin hypothesis
warm little pond
primordial soup
high UV
meteor strikes, volcanic activity (evidence that organic molecules can form spontaneously)
subsurface origin hypothesis
hydrothermal vents at ocean floor from volcanic activity
gases out of floor
more stable because surrounded by water
constant sources of energy - reduced inorganic compounds from ocean crust and vents
more likely for life to survive
time of domains
eukaryote was 2 billion years ago - more recent than bacteria/archaea
formation of RNA
self-replicating catalyse cheical reactions produce proteins make ribozymes (enzymes) unstable - too many functions
LUCA
last universal common ancestor
from this evolution splits to archaea and bacteria
what do bacteria and archaea make from H and CO2
B - acetate
A - methane
anoxygenic photosynthesis
without oxygen
produce sulfur as waste
by purple and green sulfur bacteria
phylogenetics
and how to measure
relationships of organisms
can’t allow gene transfers or will carry info of where it’s been
must be homologous
look at production of protein via ribosomal RNA
other markers: ATPase, EF-Tu, RecA
how closely related are the 3 domains?
archaea closer related with eukaryotes than bacteria is to both of them
endosymbiotic theory
defo true - chloroplasts came from phototrophic cyanobacteria into eukaryotic cell
debatable - aerobic chemo-organotrophic bacteria became mitochondria in host cells
some bacteria examples (8)
aquifex - hyperthermophile, chemolithoautotroph
deinococcus - radiation resistant, bio-remediation
cyanobacteria
actinobacteria - heterotroph, antibiotic or pathogen
firmicutes - mostly heterotroph, probiotics or pathogen
chlamydia - parasite, infect eyes, STD
spirochaetes - spin, heterotroph, parasite
proteobacteria - very diverse
systematics
study of diversity of organisms and their relationships
links phylogeny and taxonomy
bacteria taxonomy
phenotypic comparisons - what they did
bacterial names
reflects shape/what they do
staphylococcus epidermis - bunch of group clusters, coccus-shaped, from skin
bacillus thermophilus - rod, at high temp
microbiology species
asexual so viable offspring definition doesn’t make sense, also lateral gene transfer occurs
groups of strains that show a high degree of overall similarity and differ considerably from related strain groups with respect to independent characteristics
issue: single mutation can change definition of species e.g. pleomorphism (exist in diff shapes)
polyphasic bacterial taxonomy
phenotype - what do, morphology, metabolism, physiology, chemicals
and genotype
and history - phylogenetic evolutionary relationships
morphology
motility
nutrition
shape of cells
Gram stain
phase-contrast microscopy, some swim
what grow on
decomposition of simple carbohydrates
acid from glucose turns pH indicator from red to yellow
see if gas produced - info about metabolism
enzymes that decompose large molecules
tested in agar plates
starch used up so stain with iodine
colony characteristics
shape margin surface texture colour odour
acidophile alkaliphile microaerophile barophile halophile facultative anaerobe obligate anaerobe psychrophile mesophile thermophile hyperthermophile
pH < 6 pH > 8 low O2 conc high pressure salty better with O2 but can w/o can't tolerate O2 low temp < 15 normal temp high temp >50 very high > 80
molecular analysis
FAME DNA-DNA hybridisation DNA profiling (PCR, AFLP) MLST GC base ratios
outer layer of microbes
capsule/ S-layer
Gram +ve
Gram -ve
outer cell wall, 90% of cell wall is peptidoglycan
2 outer membranes with periplasmic space between (stores peptidoglycans)
10% peptidoglycan
lipoprotein attaches outer M to peptidoglycan covalently
capsules
outside cell wall - glycocalyx/slime layers (polysaccharide components)
no stain on capsule with phase contrast miscroscopy (negatively staine with india ink so stain things not in cell)
loose network of polymer fibres extend from wall
form biofilms
capsule is organised, tight matrix, not easily removed
slime layer is unorganised, easily removed
function of capsule
carbon store prevent desiccation capture nutrients attach to surfaces biofilm defense difficult for immune system to recognise - resist phagocytosis
S-layer
paracrystalline outer wall layer composed of protein/glycoprotein
external to cell wall
protect
peptidoglycan (murein)
after plasma membrane
alternating NAG & NAM (sugar moieties)
dimers cross link by AA chains creating amide bonds
mesh-like polymer
stained in Gram stain (more in Gram +ve)
joined by cross links between peptides
cross links between carboxyl group of terminal D-alanine connected to amino group of DAPA
porous, elastic, stretchable
lysozyme
antibacterial enzyme in saliva, tears, airways
targets peptidoglycan and degrades 1-4 glycosidic bonds between NAG & NAM
so makes bacteria sensitive to osmotic pressure so burst and die
penicillin’s effect on bacterial walls
inhibits peptidoglycan synthesis so better against Gram +ve
archaeal cell walls
no pathogens no peptidoglycan but some have pseudomurein 1-3 beta not 1-4 no D-amino acids in linker not affected by penicillin/lysozyme
membranes
stiff - hopanoids/sterols sit alongside fatty acids (too flexible) so stabilise
no hopanoids in archaea
archaeal membranes
isoprene hydrocarbons attached to glycerol by ether link not ester (bacteria/eukarya)
branched/rings - function same as hopanoids/sterols
lipopolysaccharide (LPS)
large complex molecules on surface of Gram -ve cells
contain lipid and carbohydrates - called endotoxin when free in host (us)
lipid A, core polysaccharide, O side chain
Lipid A
2 glucosamine residues linked to fatty acid and phosphate/pyrophosphate
induces largest immune response
in outer membrane while the rest of LPS projects from surface
core polysaccharide/R-antigen
sugar residues
side chains of NAG, phosphate, ethanolamine
O side chain
variable region - antigenic make-up
make antibodies to this
different O serotypes linked to diseases
flexible
rough/smooth depending on side chain length
often phosphorylated and anchored into OM via lipid
R = more easily detected so phagocytosis - less pathogenic
functions of LPS on bacteria
Lipid A - inflexible so stabilises outer membrane
core polysaccharide - -ve and hydrophilic so reduces permeability
loss of O-antigen means reduced virulence, diagnostic tool
endotoxin
produced by pathogen when cells are attacked and destroyed or during cell division/lysis
what does LPS of bacteria cause in our body?
small amounts are fine because trigger immune system
high levels lead to septic shock
trigger cytokines - activates transcription factors
inflammation, fever, coagulation of blood
it’s heat stable
porins
protein channels
protein export to periplasm
sec
TAT
flagella
extracellular helical structure
protein
motor spins
longer than cell
rings and hook of flagella = rigid and attached
shaft = easily removed, signle protein flagellin
motor driven by transfer of protons through rings
base = hook (diff proteins)
single protein connects shaft to motor
Gram -ve: L ring in lipopolysaccharide of OM, P ring in peptidoglycan, S-M ring on membrane, C ring in cytoplasm
how does flagella get longer?
growth at tip, adds at end, channel allows flagellin to pass up to tip to grow
types of flagella (4)
monotrichous (1 end)
amphitrichous (2 ends)
lophotrichous (multiple at 2 ends)
peritrichous (everywhere, like E.coli)
aerotaxis
chemotaxis
magnetotaxis
phototaxis
towards O2
to nutrients and away from toxins
along lines of magnetism
towards lights
how do bacteria sense changes in nutrient conc.
transmembrane proteins
MCP
how many does E.coli have?
Methyl-accepting chemotaxis proteins (transducer protein)
interact with cytoplasmic proteins and change toxic behaviour of cell by interacting with rings of motor
E.coli - 5 transducers to sense different compounds
Che B
methylesterase
what happens to MCP in low conc. attractant?
attractant decreases CheA-P so low conc, means high CheA-P high CheB-P (which removes methyl from MCP) demethylation of MCP increased sensitivity to attractant
what happens to MCP in high conc.attractant?
low CheA-P, low CheB-P, methylated MCP
decreased sensitivity
if harder and harder to sense - know going up conc. gradient
if no sensitivty - short runs and tumble and stay in env.
what happens to MCP in high conc. repellent?
methylated MCP leads to longer runs, less tumblr and leaves env.
which sequences of receptors are conserved and which vary?
cytoplasmic domains of all receptors are highly conserved
periplasmic sensing domains vary
Gliding
pulls along slime extruded on outside
little feet
3 gld proteins - ABC transporters
5 lipoproteins in membranes
dsrupting gld genes means loss of motility, lose ability to digest chitin, increase resistance to bacteriophage infection
Twitching
via type IV pilus
extend then retract so drag cell
powered by ATP
Gas vesicles
in planktonic bacteria/archaea
float to right level in water
Fimbriae/Pili e.g. CFA
adhesion
7 groups
virulence, resist phagocytes, antigenic
adhesion to epithelial cells
type 1 fimbriae
properties
types of proteins involved
how is it made?
thin (7nm), short (1-2μm), only attached to outer membrane
FimA protein stacked in helical cylinder
FimC chaperone
FimD usher protein (catalyse FimA polymerisation at base of pili)
Fim F,G,H - adhesin onto fimbriae
FimH - tip adhesin binds to D-mannose containing structures
don’t have middle channel so grow from base
proteins transported into periplasm via sec translocase then fold to fimbriae
type 4 pili
lots in Gram -ve, some in +ve, long, both ends of cell, twitching motility
aggregate to form bundles which causes virulence
e.g. CFA in E.coli (K antigen), ETEC interacts with mucosal epithelium but no CFA so not pathogenic
F pilus
conjugation?
10μm long, 8nm wide, central 2nm channel, in Gram -ve, plasmid encoded
adhesion like type 4 pili
e.g. E.coli F pilus
conjugation - only attach if other cells have no F-pilus, remember video shown of attaching to other cells and giving them pilus (attachment, retraction, exchange F pilus genes)
endospore
dormant stage/dispersal stage to survive difficult conditions
only Gram +ves
stain with Malachite green
resistant to 150 degrees
human microbiome
ecological community of commensal, symbiotic, and symbiotic, and pathogenic MOs that share our body space
shield against pathogens
virulence factors
anything that allows bacteria to avoid detection and stick to cell
toxins, adhesins, surface capsules, enzymes, LPS
secretion machineries, siderophones, catalases counteract phagocytes
who invented vaccines?
Edward Jenner
Louis Pasteur
infected chicken with old culture and didn’t affect them, then new culture didn’t either
