microbio midterm 1

phylogeny

evolutionary relatedness btwn organisms

strain

• isolate distinguished within species by genotype/phenotype

variables that make up strain

• antimicrobial resistance

• phenotypic growth patterns

• virulence factors (enzymes, proteins, lipids that aid organism in colonizing host)

definition of LIFE

• carry out metabolism independently

  • chem rxns → continuous energy → sustain life

• 
  

what allows for metabolism

• membrane encolsed compartment

• can replicate (hereditary info, evolves over time)

archaea

• prokaryotes

• extremophiles (acidic, super cold/hot environments

• hard to grow

• outer cell membrane but no peptidoglycan

magnification

ability to enlarge objects

resolving power

ability to show detail

• distinguish two adjacent points

parfocal

maintains focus btwn objectives

difference btwn simple and compound microscope

1 or 2+ lenses

total mag

objective power x ocular power

what does resolution depend on

• wavelength of light

• amount of light captured by lens

RP = wavelength/2 NA = (nm)

numerical aperture

how much light we capture (angles of lens)

nsin(theta)

• n=c/v

• theta = ½ angle of max amt of light that can enter lens (highest: 90 degrees, sin90=1)

  • increase by increasing lens size

refractive index

n=c/v

• what you shoot light thru

• v = medium

want to be as close to 1 as possible

what are light field microscopes used for

live and preserved stained specimens

• specimen darker than surroundings

• we have these

dark field microscopes?

live, thin, unstained specimens

• brightly illuminated specimens

• more internal detail

• expensive

pathonomonic

determine disease by signs and symptoms of patient

phase contrast microscope

bright field with great contrast

• manipulate phase of light

• best observe intracellular structures

• z direction

allows visualization of live cells and organelles

fluorescence microscope

has UV radiation source and filter

• dyes emit visible light when bombarded with shorter UV rays

• fluorescence

green fluorescent protein

can fold spontaneously

has self fluorescent core

expressed in ANY cell type (bacteria, insects, humans)

scanning confocal microscope

not squashed

laser beam of light to scan specimen

• allows focus on multiple depths or planes (integrates images

• z stacking

• live imaging, better resolution, 3D imaging

electron microscopy

accelerated e- as illumination source (instead of light photons)

• used by virologists

• 1000000x +

• view ultrastructure (viruses)

• NOT for live samples

goal of microbial investigation

obtain pure culture (all genetically same)

sample collection must dos

• label it (name, location, time/date, tests)

• know properties of sample

• store it well

inoculation

introduce specimen into growth media

incubation

put into controlled environment

• temp

• humidity

• co2 and o2

• time

isolation

separate single species from other species in sample

• make assumption it’s derived from single cell

pour plate technique

bacterial sample added to petri dish

• pour over molten agar

spread plate technique

hockey stick spreads

inspection

• colony morphology

• unique pigments

• check for contaminants

• CONSTANTLY inspect

sub culture

pick colony off of pure culture → put it in another environment

mixed culture

1 + genetically identical microorganisms

contaminated culture

unexpected microorganism

broth

liquid, does not solidify

advantage: grows large quantities of culture

• fast and efficient

semi solid agar

with solidifying agent

• 0.3-0.5% agar

advantage: better access to nutrients and motility assays

solid agar

firm surface for colony forming

• most common

• not digestible for most microbes

advantage: useful for storage/slant

agar

polysaccharide extract from marine algae/seaweed

• polymerizes

• liquifies 85 C, resolidifies 42 C

synthetic media

aka chemically defined

• used to grow specific organisms w defined requirments

advantage: known amounts of pure compounds in medium

disadvantage: expensive and time consuming

complex media

chemically undefined

• rough idea of what’s in there

• used for many microorganisms

advantage: helpful when exact requirements are unknown

disadvantage: exact components unknown

selective medium

certain microbes grow, some inhibited

• “can it grow or not”

differential medium

certain characteristics show pH or color differences

• “can i tell difference btwn colonies?”

enrichment medium

enriched with compound required for growth

• increases # of desired organisms

positive staining

basic dye, stains specimen

• ex: crystal violet, methylene blue

negative stain

acidic dye, stains background

• ex: congo red

do NOT heat fix

simple stain

single staining agent used

• reveals shape, size, arrangement

differential stain

primary and counterstain

• distinguish cell types/parts

• ex: Gram stain, acid-fast stain, endospore stain

structural stain

reveal cell parts not revealed by conventional methods

• ex: capsule stain, flagellar stain

cell envelope

structures around cytoplasm

• can’t rely on other cells

gram positive

cell wall(thick peptidoglycan) plasma membrane

• teichoic acid

• lipoteichoic acid

gram negative

cell wall(outer membrane, thin peptidoglycan) plasma membrane

• lipopolysaccharides LPS (endotoxin layer

• periplasm

plasma membrane

fluid mosaic model

• contains proteins, lipids, and sugars

• NOT static

phospholipid

head: polar, hydrophilic

tail: nonpolar: hydrophobic

• head and tail linked by ester

• ether linkage found in archaea

what do bacterial membranes generally lack (compared to Eukarya)

sterols

• stabilization is achieved by hopanoids

membrane function

1. permeability barrier: prevents leakage, gateway for nutrients in n out

2. protein anchor: involved in transport

3. energy conservation: generate and use protein motive force

cell wall structure

crosslinking peptidoglycan

• helices of sugars and amino acids

cell wall function

bacterial integrity

• shape and rigidity against external forces

protects from osmotic lysis

• antibiotics that target cell wall and expose it

fimbriae

attach to surfaces

• short, thin, hairlike

• proteinaceous appendages (pilin)

pilus/pili

surface binding (primary func)

• conjugation: share nucleic acids

  • sex pilus: latches to cell wall of another bacteria → draws it in → share nucleic acid

flagella

motility and surface attachment

• basal body, hook, filament

flagellum structure

basal body

• embedded in cell envelope via protein rings

  • gram pos: 2 rings

  • gram neg: 4 ring (2 at outer membrane)

hook

• wider than filament

• array of pro subunits

filament

• hollow, rigid cylinder

• many flagellin (pro subunit)

what is flagellar mvmt dependent on

environmental signals

• attractants

  • chemotaxis: search for nutrients

• repellants

monotrichous flagella

1 tail

lophotrichous flagella

multiple tails at one end

amphitrichous flagella

one tail at each end

peritrichous flagella

tails EVERYWHERE

flagellar run

counterclockwise rotation towards stimulant

flagellar tumble

clockwise rotation due to……

• chemotactic (nutrient) signal lost

• change of gradient

• repellent sensed

glycocalyx

bacterial surface coating\

• neg stains

slime layer

• less organized

• brittle

capsule

• specialized, structured, tightly attached

• layers of polysaccharides

advantages of glycocalyx

• surface binding

• resist phagocytosis

• protects against drying

• resists toxins/antimicrobes

clinically relevant genus of endospores

clostridium, bacillus

advantages of sporulation

dormant (metabolically inactive)

resistant structures

• increase bacterial survival

• doesnt replicate

endospores are resistant to

alcohols, ammonium solutions, most detergents

endospore triggers

lack of water and nutrients

• environmental stressors

endospore advantages

resistance against:

• heat

• uv and gamma radiation

• chemical disinfectants

• desiccation

endospore sporulation

asymmetric division (happens on one pole)