Microbiology Exam 2

Why is microbial structure important?

* Bacteria control the expression of virulence mechanisms
* We need to understand what an organism is and how it functions to understand how it causes disease
* To use antibiotics we need to understand what it is targeting

Virulence mechanisms

any molecule that contributes to an organism’s virulence and usually aids in entry portal, evasion of defenses, cell damage, and exit portal

Cell envelope

* Partially responsible for structure and shape
* Toxic and immunological properties of cell are often associated with it
* May be responsible for signs/symptoms of disease

Coccus

Sphere

Coccobacillus

Oval/round

Vibrio

Curved rod

Bacilius

Pill/rod

Spirillum

Spiral/worm shape

Spirochete

Corkscrew

Diplo

Two

Tetra

Four

Sarcinae

Cuboidal

Strepto

Chains

Staphylo

Clusters

Cell envelope protection

* protection from osmotic imbalances
* protect cell from elimination by phagocytosis and infection by viruses
* can protect from chemo

Cell envelope metabolic functions

* side of envelope is associated in enzymatic activities
* location of transport proteins
* origin of many signal transduction pathways
* attachment and colonization of environmental surfaces

Gaining entry to host

1. Entry through portal in human body either a common portal or opportunistic (nose vs cut)
2. Pathogens need to attach to host cells otherwise they will be flushed out

Cell membrane

Semipermeable barrier that keeps things in/out despite gradients and osmotic pressure and has a fluid structure because of cholesterol

Cell membrane functions

1. Fast cell division
2. Proteins for responses and reactions
3. Transportation of materials

Fast cell division

* Septum formed from the cell membrane and separates the two new daughter cells
* Ftsz

Ftsz

assembles at site of septum formation, analogous to actin ring in eukaryotic cytokinesis; establish cell polarity

Proteins for responses and reactions

* membrane receptors initiate responses to stimuli
* Movement so they don’t die
* Chemotaxis
* Phototaxis
* membranes are crucial for ETC
* assist in making energy
* Photosynthetic microbes do photosynthesis in cell envelope membrane
* Have carboxysome:
* Endosymbiosis theory
* transportation of materials
* Active transport
* Proton motive force
* Resistance-nodule cell division
* Group translocation
* ATP Binding Cassette

Carboxysome

Protein shell containing RuBisCo for carbon function, the same enzyme found in chloroplasts

Endosymbiosis theory

Complex eukaryotes are a result of symbiotic combinations of prokaryotic cells

Proton motive force

bacteria use the energy of moving protons with gradient to transport molecules against gradient, same way ATP synthase uses proton movement to provide energy for ATP

Resistance-nodule cell division

bacteria use PMF to transport molecules out of cell; drug resistant bacteria pump antibiotic out of cell before it is effective

Group translocation

chemical modification of the transported substance driven by

phosphoenolpyruvate; substrate phosphorylated as a means to transport it against the gradient

ATP Binding Cassette

transporters use the energy from ATP to transport a molecule against its gradient; usually involves a substrate-binding protein, can also be used to transport out of cell (virulence factors)

ABC Type 1

used to secrete factors such as RTX toxins commonly found in E. Coli; vibrio cholera and bordetella pertussis

Cell wall functions

* provides structure and stability for the cell
* protection from osmotic pressure changes
* partially responsible for morphology and arrangement of bacterial cells
* provides points for attachment (colonization) and receptors for viruses (bacteriophage)
* responsible for some disease signs/symptoms
* may contain toxic molecules
* target of antibiotics/chemo
* important for classification

Gram positive

cell wall with thick peptidoglycan (purple in stain)

Gram negative

cell wall with thin peptidoglycan with outer layer of lipopolysaccharides (membrane layer loaded with sugar); pink stain

Composition of peptidoglycan

1. Glycan Backbone
2. Side chain
3. Interbridge (or cross link)

Glycan backbone

Made of NAG (n-acetylglucoamine) and NAM (n-acetylmuranic acid) that repeat to make glycan chain

Side chain

* Consistant from genus to genus
* Made of (in order of attachment):
* D-alanine
* L-lysine (+) or Diaminopimeic acid (-)
* D-glutamic
* L-alanine
* NAM-NAG (glycan backbone)
* Percentage of NAM varies between 35-40% for G- to 80% for G+

Interbridge

* typically a pentaglycine (S glycine) in G+
* Direct link in G-
* 1 to 2 linkage in both that run antiparallel

Teichoic acid

* In G+
* adds structural integrity (scaffolding) holding thick layer of peptidoglycan together and anchors to membrane
* important to cell growth as it holds together through stretching
* important for cell adhesion (to host cells and other bacteria)

Bacterial porins

* In G-
* allow for passive transport of material in and out but are very selective

Lipopolysaccharide (LPS)

* On G- outermost outer membrane (inner part is phospholipids)
* Varies in length and composition; repeating

Lipooligosaccharide (LOS)

* no repeating
* Only 1 O antigen
* ex. Neisseria spp and haemophilus spp

Lipid A

* Bacterial endotoxin
* in the outer most portion of cell wall in G-; liberated when the cell dies and cell wall breaks apart
* can trigger immune response
* if it gets into blood can cause systemic response and sepsis

Acid-fast cell wall

* Mycobacteria
* membrane, thin peptidoglycan, arabinogulactan layer and then thin mycolic acid/lipid outer layer
* named due to decolorizing technique
* contains large amounts of unique, high molecular weight lipids
* mycolic acids contain anywhere from 60-90 carbon atoms in long chain fatty acids
* Ex. Mycobacterium spp and nocardia spp

Cell “wall-less” bacteria

* Mycoplasma spp
* Ureaplasma spp (T-strains)
* L forms of bacteria (protoplasts G+ or spheroplasts G- that lost their peptidoglycan)
* No true shape/form; change based on the environment

S-layer proteins

* repeated proteins found on the exterior of some bacterial cell walls
* increases structural integrity of cell wall
* involved in attachment and protection

Glycocalyx

* material surrounding the cell; thick, sticky sugar coat
* gives bacteria its sticky and slimy characteristics
* Slime layer or capsule
* gives protection against toxic compounds, bacteriophage infections, elimination by phagocytosis, and desiccation
* Adherence proteins present and important in pathogenesis of almost all bacteria; cell to cell recognition and interaction depending on attachment
* Biofilm formation
* role in attachment of streptococcus mutens in dental plaque that can only be removed by metal scraping teeth
* Classification of many species
* Carbohydrate reserves possibly?

Slime layer

temporary and less consistent

Capsule

organized and considered a permanent part of the cell

Bacteria appendages

* outside of the cell envelope
* Three main types:
* Flagella
* Pilus
* Fimbriae

Flagella

* primary role is locomotion
* can be a sensory organelle
* anchored into cell envelope
* Basal apparatus and filament

Atrichous

0 flagella

Monotrichous

1 flagella

ex. ampylobacter spp.

Amphitrichous

2 flagella

ex. ampylobacter spp.

Peritrichous

Many flagella all over

ex. E. coli and salmonella

Lophotrichous

many flagella on one end

ex. Helicobacter pylori

Cephalotrichous

many flagella on each end

ex. salmonella spp.

Basal apparatus

flagella that allows bacteria to rotate

Filament

flagella that allows for bacterial movement

Run and tumble motion

bacteria move forward for awhile, pause and spin in a circle to sense where they are and then move forward again all using flagella

Spirochetal endoflagella

* anchored to inner membrane, remains in periplasm and doesn’t extend through outer so it can’t sense the same way


* Diderm, especially G- but no LPS/LOS
* Whole organism moves like a propeller instead of just flagella
* Ex. Treponema pallidum and Borrelia burgdoferi

Specialized flagella: Type III secretion system

* Uses “needle” to inject into plasma of neighbor
* Direct delivery= no dilution from secretion
* Secrete virulence factors (attachment proteins, toxins, immune evasion)
* AKA injectosomes
* Found in G- pathogens
* Chlamydia trachomatis, E. Coli, Pseudomonas aeruginosa, salmonella, shigella, vibrocholera, yersina pestis

Pilus

Specialized for movement, secretion, conjugation, attachment

Type IV pilus

primary function is motility; ulike flagella, movement is more rigid resulting in “twitching” (like climbing ice with a pick)

* ex. Acinetobacter baumannii

Type IV secretion system

* Pilus secretion system
* secrete virulence factors, transfer and uptake DNA, potentially “steal” resources from host (move both ways unlike Type III flagella); needle forms not always out


* ex. bordetella pertussis, helicobacter pyloris, legionella pneumophila, neissaeria, streptococcus, argobacterium tumefuliens

Conjugation pilus

* transfer genetic material from donor to recipient cell, “sex pilus”
* T45S often a major mediator of congugation
* F pilus encoded by F factor (fertility)
* plasmids may contain genes as environmental conditions, host immune systems, and other bacteria
* donor cell doesn’t loose plasmid DNA, just transfers a copy and does not benefit

Type VI secretion system

* secrete virulence factors (toxin, bacteriocins) into target cell, bares a striking resemblance to bacteriophage tail
* not a flagellum, pilus or fimbria just an appendage
* G- pathogens: vibrio cholerae, pseudomonas aeruginosa

Secretion

a critical component of bacterial survival and pathogenicity

Fimbriae

* unique, specialized pili but own category because they have a single unique function
* Specialized attachment or adherence pili important for virulence, often called colonization factor antigens
* neissera gonorrhea, klebstella pneumonia, e coli (UTIs and intestinal)

Enterotoxigenic (ETEC)

noninvasive, produces toxins, ex. travelers diarrhea

Enteroaggressive (EAEC)

noninvasive, forms aggregates, produces toxins, ex. acute and chronic diarrhea

Enteropathogenic (EPEC)

moderately invasive, does not produce toxins, ex. diarrhea especially in newborns

Enteroinvasive (EIEC)

highly invasive, produces toxins, ex. dysentery, fever

Enterohemorrhagic (EHEC)

moderately invasive, produces toxins, ex. dysentery, hemorrhagic fever, hemolytic-anemic syndrome (RBC being destroyed)

Stages of fimbriae attachment

1. Pili bind to receptors (initial attachment)
2. Attachment proteins bind to other receptors (intimate)
3. Colonization and inflammation

Afimbrial adhesion

attachment without fimbriae, adhesins found on cell wall, glycocalyx attachments are classified as afimbrial

Cytosol structures

* ribosomes
* granules
* endospores
* nucleoid
* plasmids

Ribosomes

* composed of RNA and proteins and made of large and small subunits
* Key player in translation to make proteins
* Structural differences exist between eukaryotes and prokaryotes allow for specific antibiotic targeting

Subunit sizes in prokaryotes

5s, 23s, 50s; 16s, 30s

Granules

* storage bodies/inclusion bodies
* stores synthesized or obtained nutrients/metabolites as well as products of metabolic activity
* Includes specialized for sugars, fats and phosphates

Endospores

* produced under threatening environmental conditions known as sporulation
* purpose is to protect
* resistant to heating, freezing, desiccation (drying), some chemicals, and radiation
* no reproductive capacity when in endospore form but can exist for many years
* produced by only a few pathogenic genre of bacteria
* Bacillus spp. and Clostridium spp.

Endospore structure

* Core
* Cell wall
* Cortex
* Spore coat
* Exosporium

Endospore core

DNA, ribosomes, granules

Endospore cell wall

normal gram-positive peptidoglycan

Endospore cortex

thick layer of loosely cross linked peptidoglycan

Spore coat

highly cross-linked keratin proteins as well as other structural proteins

Exosporium

not always present, varies in composition, primarily proteins and carbs

Endospore formation

1. recognizes resources are being depleted and environment is no longer optimal
2. chromosome duplicated and separated
3. cell separated into sporangium and forespore
4. sporangium engulfs forespore for further development
5. sporangeium synthesizes spore layers around forespore
6. cortex and outer core layer deposited
7. mature endospore in sporengium
8. free spore released, loss of sporangium, in vegetative state
9. germination: spore swells and releases vegetative cell when conditions become optimal again

Nucleiod

* region of dense genomic DNA (single circular chromosome)
* Origin of replication

How do prokaryote genomes compare to eukaryote genomes?

Prokaryote genomes are considerably smaller but there still is a lot DNA that is about 1000x the length of the cell

How is a prokaryote chromosome condensed?

* DNA gyrase and topoisomerase supercoiling
* Scaffolding proteins assist as well

CRISPR/Cas

* Bacterial immune system within its genome
* Stores genome of virus previously encountered so it can fight it if infected again
* Artificially can be used in gene editing

CRISPR

* clustered regularly interspaced short palindromic repeats
* Family DNA sequences within prokaryotic genomes
* Flanked by viral DNA from previous infections

Cas

* CRISPR-associated genes/proteins
* DNA endonuclease
* Uses crRNA to target specific complementary DNA sequences

How does the CRISPR-Cas system work?

* Bacteria infected with virus receives a copy of its viral DNA
* Cell creates CRISPR sequences with the DNA and spacers
* Spacers allow for DNA to create a hairpin structure that can match up with viral DNA should they find it again
* When it binds to viral DNA, Cas genes are activated so they can cut up the viral DNA
* This is an inheritable system so each division passes this on to new cells

Plasmids

* Small circular pieces of DNA
* Considered more “fluid” than the chromosome
* Contains “non-essential genes”
* will only be kept if advantageous
* can combine with other DNA to form new plasmids
* Classification based on characteristics like copy number and organism specification

Engineered plasmids

synthetically made or modified by scientists

Colicin (Col) plasmids

code for bacteriocins

Degraditive plasmids

code for digestive enzymes

Fertility (F) plasmids

code for conjugation genes

Virulence plasmids

code for genes important for pathogenicity; contain pathogenicity islands

Pathogenicity islands (PAI)

series of genes that code for virulence factors