Microbiology - 2 - Bacteria

Question 1

What are the basic shapes of bacterial cells?

Answer 1

1. Rod
   
   • ​Single = Bacillus
   • Chain = Steptobacillus
     
     • can be straigh or curved
2. Sphere
   
   • singular = coccus
   • plural = cocci
     
     • clusters = staphylococcus
     • chain = streptococcus
3. Spiral
   
   • ​can be rigid or flexible & undulating
     
     • Vibrio = comma shaped
     • Spirillum = wavy
     • Spirochete = spiral
4. Other
   
   • square
   • star
   • filamentous
   • appendaged

Question 2

What are the structures & functions of bacterial cells?

Answer 2

Envelope

• Capsule - polysaccharide
  
  • hydrophilic gel = discrete layer = capsule
  • slime layer = amorphous
    
    • provides general protection from immune system
• Cell Wall - Gram (-) / (+)
  
  • prevents cell from taking in too much water
  • made of teichoic acid & lipoteichoic acid
    
    • promote adhesion & anchor wall to membrane
  • Gram (-) = thin cell wall
    
    • little peptidoglycan + LPS + lipoprotein
  • Gram (+) = thick cell wall
    
    • lots peptidoglycan + teichoic acid
• Outer Membrane - Gram (-) ONLY!
  
  • Gram (-) = biphospholipid membrane
    
    • inner leaf = ordinary phospholipids
    • outer leaf = lipopolysaccharides (LPS)
      
      • extremely toxic to humans (endotoxin)
• Peptidoglycan layer - BOTH (+) and (-)
  
  • makes surface hydrophilic
    
    • Gram (+) = thick layer
      
      • highly polar
    • Gram (-) = thin layer
      
      • less polar
• Periplasm
  
  • enzymes w/ hydrolytic functions; antibiotic-inactivating enzymes
    
    • Gram (-) = contains peptidoglycan
    • Gram (+) = small, no peptidoglycan
      
      • has proteins & oligosaccharides
• Cell Membrane
  
  • similar to familiar bi-leaflet membrane of most cells
    
    • made of phospholipids + proteins
  • ​Different than EUK b/c:
    
    • NO STEROLS
    • bacterial chromosome attached
    • site for DNA synthesis
  • Functionally ~ mitochondria
  • Vital for growth & maintenance of cell

Appendages

• Pili (Fimbriae) - protein
  
  • hair-like projections
    
    • found on both gram (+) and (-) species
    • specialize for adherence to certain cell types
  • made from pilin protein
    
    • tube w/ hollow core
  • 2 kinds:
    
    • common
      
      • adhesions = allow for colonization on surfaces
    • sex
      
      • allow for exchange of genetic material btw gram (-) bacteria
• Flagella -
  
  • rotating helical protein structures –> locomotion
    
    • rings on center rod
      
      • independent of ATP!
  • can be found:
    
    • 1 singular pole (polar)
    • tufts (lophotrichous)
    • all over surface (peritrichous)
  • built from flagellin proteins
    
    • Antigenic = H Ags = targets for Ab response

Core

• Cytosol - polyribosomes, proteins carbs, glycogen
  
  • granular b/c packed w/ lots of ribosomes
  • cytoskeleton elements
• Nucleoid - DNA & assoc RNA & protein
  
  • contains the genome
  • single circular chromosome
  • nuclear bodies = 1 when resting; 4 is reproducing a lot
• Plasmid - DNA
  
  • small, circular, covalently closed DNA

Question 3

What is the process & function of Gram-staining?

Answer 3

Gram staining:

• initially stained purple via crystal violet + iodine
• after decolorization + counterstain:
  
  • gram (+) = keep stain –> purple
  • gram (-) = unstained –> red

Function:

• allows us to ID + treat for organism
  
  • Gram (+) = thick peptidoglycan layer
    
    • resists activity of bile in intestitine
    • digested by lysozymes
    • disrupted by antibiotcs
  • Gram (-) = thin peptidoglycan layer + outer membrane
    
    • (LPS) =
      
      1. toxic lipid A
      2. core polysaccharide
      3. O Ag polysaccharide side chain = major surface Ag

Question 4

What are the differences in structure & function of Gram (+) & Gram (-) bacterial cell walls?

Answer 4

1. Peptidoglycan
   
   • gram (+) = thick
     
     • lots of peptidoglycan
   • gram (-) = thin
     
     • little peptidoglycan
2. (lipo)teichoic acids
   
   • more in gram (-)
3. Porins
   
   • gram (-)
4. Lipopolysaccharide (LPS)
   
   • gram (-)
5. Endotoxin
   
   • LPS = gram (-)

Question 5

What happens to bacterial metabolism after glycolysis?

Answer 5

Following glycolysis:

• 1 glucose –> 2 pyruvate (+ 2ATP)
  
  • if O2 present = aerobicrespiration –> lots of ATP
  • if no O2 = anaerobic fermentation –> no ATP

Aerobic Respiration

• with O2, more energy can be made
• pyruvate –> mitochondria –> 36 ATP (18x more energy)
  
  • occurs in cell membrane

Anaerobic Fermenation

• necessary if no O2 available (or if organism is resistant to O2)
• cells cannot stockpile pyruvate
• pyruvate –> lactic acid or other acids

Function in Bacteria & Role in Metabolism:

• O2 gives rise to at least 2 extremely reactive & toxic substances:
  
  • H2O2
  • superoxide anion (O2-)
• O2 –> O2- [superoxide dismutase] –>
• H2O2 [catase] = H20 + O2 ….or
• H2O2 [peroxidase] –> reduces NADH2 –> NAD = H20

Note:

• bacteria that LACK ability to make superoxide dismutase & catalase = grow anaerobically (sensitive to O2)

Question 6

What are the differences btw bacterial aerobes & anaerobes w/ respect to oxygen & mode of respiration?

Answer 6

​Basics:

• Bacteria response depends on:
  
  • ability to ferment or respire
  • ability to protect from deleterious effects of O2
    
    • aka do they have enzymes to break down oxygen

Leads to 4 General Classes:

1. Aerobes
   
   • + O2 / + enzymes / - ferment
   • requires O2, cannot ferment
     
     • ex: fungi, protozoa, many bacteria (bacilis, TB)
2. Anaerobes
   
   • • O2 / - enzymes / + ferment
   • killed by O2; ferments
     
     • ex: oral/intestinal bacteria
3. Facultative anaerobes
   
   • • O2 / + enzymes / + ferment
   • respires w/ O2, but can ferment if no O2 available
     
     • ex: gram (-) intestinal bacteria, staphylcoccci
4. Microaerophiles
   
   • • O2 / + enzymes / + ferment
   • grows best at LOW O2
     
     • ex: bacteria that live in soil/water/host (h. pylori)

Note:

Aerotolerant anaerobes

• do not utilize O2, but can survive in its presence to a limited extent
• NOT harmed by O2
  
  • have alternate mechanisms for breaking down H2O2 & superoxide
• ex: lactobacilli & streptococci & clostridial species

Question 7

How does bacterial DNA replication occur?

Answer 7

Replication important for survival

Role of components:

• OriC
  
  • origin of replication
  • multienzyme replication complex (MeRC) binds to this & initiates unwinding & separation via
    
    • DNA gyrase (topoisomerase) = relaxes coiling
    • DNA helicase = removes H bonding & separates
• DNA polymerase
  
  • replicates DNA via base pairs
  • Proof reading of new strand

Therapeutic targets:

• quinoiones =
  
  • inhibit unwinding of DNA gyrase during replication
• penicillin =
  
  • inhibitors of peptidoglyvcan cell wall synthesis

Question 8

How does bacterial gene expression occur?

Answer 8

1. Transcription:

• DNA is copied by DNA-dependent RNA polymerase –> RNA transcript

Components:

• RNA polymerase
  
  • DNA –> mRNA
• Promoter
  
  • nucleotide sequence in DNA that binds RNA polymerase
• Operon
  
  • results in polycistronic arrangement
  • provide a way of ensuring protein subunits of enzyme complexes are made in correct stoichiometry
• Monocistronic
  
  • 1 promotor = 1 gene => 1 protein
• Polycistronic
  
  • 1 promotor = signals many genes => many proteins

2. Translation:

• exact sequence of AA of a protein = specificed by nucleotides in mRNA

Components:

• Initiation complex
  
  • starts mRNA –> protein via Shine-Dalgarno sequence where ribosomes bind
  • comprises of mRNA, ribosome, initiator tRNA + formyl methionine
• Codon
  
  • tRNA reads and places AA
• START & STOP codons
  
  • Start = AUG
  • Stop = UAA, UGA, UAG

Importance of bacterial coupling of these 2 processes:

• translation is started BEFORE transcription is COMPLETED
  
  • multiple ribosomes can bind to mRNA => polyribosome
  • bind to free 5’ end of mRNA
• protein synthesis = target of greater variety of antimicrobials than any other process

Question 9

How are bacterial genes regulated?

Answer 9

Purpose of gene regulation:

• Bacteria adapt to environment by CONTROLLING gene expression

Important sites:

• Promoter (P)
• Operator site (O)
• Activation & activator
• Repression & repressor
• Regulon
  
  • all genes controlled by SAME REGULATOR

Function of lac operon

• (+) lactose & glucose
  
  • prefers metabolism of glucose
  • cAMP levels = low = CAP not activated
• (+) lactose only
  
  • cAMP levels rise = CAP activated = lac operon on
• (-) lactose & glucose
  
  • LacI binds to operator of lac promoter & BLOCKS transcription

Quorum sensing

• specific gene transcription = activated in response to bacterial concentration
  
  • aka = biofilms (cystic fibrosis)
• at ceratin concentrations = activation of transcriptino of genes to turn on biofilm production

Environmental regulation of Gene Expression

• 2 component regulation:
  
  • signal transduction
    
    • allows cellular function to react in response to changing environment
  • appropriate environmental stimulus results in AUTOPHOSPHORYLOATION of sensor proteins
    
    • activates a response that affects gene regulation

Question 10

What is the function of bacteral endospore formation?

Answer 10

• Endospore
  
  • small, dehydrated forms of bacteria
  • few species produce spores
    
    • anthrax, tetanus, botulism
  • GRAM (+) rods = medical relavent spores
• Sporulation
  
  • 1 cell forms 1 spore under adverse conditions
    
    • spore may persist for a LONGGG time —> give rise to single vegetative bacterial cell
  • Process:
    
    • walling off of nucleoid & surrounding cytosol
    • germination = activation by heat, acid, etc
      
      • initiation –> outgrowth of new vegetative cell that produced the spore

Question 11

Describe & label bacterial growth cycle

Answer 11

4 Major phases:

• Lag phase
  
  • no growth
• Exponential growth phase
  
  • LOTS of growth
• Stationary phase
  
  • some cells start to die
  • no growth
• Death phase
  
  • many cells die
  • some remain viable

Notes:

• Divide via binary fission
  
  • Happens in 20 minutes

Question 12

What are the 2 major bacterial mobile genetic elements?

Answer 12

1. Plasmid
   
   • ​small, independently replicating extrachromosomal nucleic acid molecules
   • have genes for replication & transfer
     
     • phenotypic advantages!

• R plasmid
  
  • carry genes for resistance to antimicrobials
  • carry virulence genes & encode toxins
    
    • increase virulence of microorganism

1. Transposable Elements
   
   • ​​DNA sequences that can jump (transpose) from a site in 1 DNA molecule to another in a cell
     
     • transposase gene
       
       • encodes enzyme necessary
     • IR & DRs
       
       • IR = inverted repeats
       • DR = direct repeats

​​​Notes:

• AKA= horizontal gene transfer

Question 13

What are the major processes for bacterial genetic transfer?

Answer 13

1. Transformation
   
   • involves release of DNA into environment by lysis
   • followed by uptake of that DNA from recipient cells
2. Transduction
   
   • DNA is introduced into recipient cell by a bacteriophage (virus)
     
     • lysogenic cycle = genome of virus = replicated passively as host cells replicate
     • lytic cycle = new virus particles are made & release when host cell lyses
       
       • virulent phages = limited to lytic cycle
3. Conjugation (sex pilus)
   
   • involves actual contact btw donor & recipient cell
   • plasmid is transfered

Question 14

What are ways to classify bacteria?

Answer 14