Microbiology

For Saier's BIMM 120 Course

First Law of Biology

• All living organisms obey the laws of thermodynamics

• Cells exist in open systems due to material exchange

• Genetic variation, sexual reproduction and evolutionary divergence

First Law: Corollaries

• Temporary creation goes against the 2nd Law of Thermodynamics

• Anything at equilibrium is dead, as change must be occurring

• Entropy in a CLOSED system always increases

Second Law of Biology

• All living organisms consist of membrane-encased cells

  • Physical separation, exchange

Second Law: Corollaries

• The cell is the only structure that can grow and divide fully on its own

• Sexual reproduction requires genetic instructions

Third Law of Biology

• Darwin credited for Third Law

• All living organisms arose in an evolutionary process

  • Evolution is the guide through which biology makes sense

• Natural selection occurs at BOTH phenotypic and genotypic levels

• Programmed genetic similarities and differences

Third Law: Corollaries

• All living organisms contain homologous macromolecules (DNA, RNA, and Proteins) that are derived from a common ancestor

  • Homology: Relatedness, or a descent from a common ancestor

• Genetic code is universal

RNA Viruses Evolve Faster than DNA Viruses

• Why?

  • RNA is less stable, used by viruses

  • DNA viruses can access host proofreading mechanisms

Relationship between Eating Animals and Viral Epidemics

• Many animal pathogens have infected humans BUT there has yet to be a plant virus to cause an epidemic

• 75% of emerging infections are zoonotic

• “Arms Race” for us vs. pathogens

MERS (Middle East Respiratory Syndrome)

• Needs an intermediate host (bats are usually the reservoir); most likely a camel

• Transmission was limited but more deadly than Covid

SARS-CoV-2

• SARS: Severe Acute Respiratory Syndrome

• Viral Spike (S) Protein binds to ACE2 cell surface receptors

• The replicase, RNA-dependent RNA Polymerase, does not have good proofreading capabilityies (antigenic drift)

• Bat reservoir and Pangolin intermediate

Antigenic Drift

Gradual change in mutations leading to adaptations, large accumulation of point mutations over time leading to variation

Antigenic Shift

• Crossing of viruses (genetic recombination) between two seemingly unrelated animals generating more deadly and more easily transmitted viruses

• Occurs at animal farms/markets

Ebola Virus

• Fruit bat reservoir

• Contact with green monkeys and other wild animals

• Highly transmissible and deadly hemorrhagic fever virus

  • Transmitted by just touching dead bodies during funerals

Marbug Virus

• Enveloped RNA filovirus (another hemorrhagic fever virus)

• Transmission via consumption of bushmeat or direct contact

HIV (Human Immunodeficiency Virus)

• RNA enveloped lentivirus

  • Retroviral

  • Very high mutation rate due to retrovirus mechanism (faulty RTase)

Viruses and Non-Human Primates (NHPs)

• Includes Zika and HIV

• Bats, primates, and rodents have a higher proportion of zoonotic diseases

Influenza Virus

• Fast Evolution: Multiple viral chromosomes allow recombination when infecting the same cell

• 1918 Spanish Flue Epidemic

  • H1N1: Spanish flu back then, but today it is the swine flu

• H5N1 is the strain we are not immune to from cattle (cows)

  • Currently not human-to-human transmissible

AIV (Avian Influenza Virus): H

• H: Hemagluttinin (viral surface glycoprotein that binds to the host receptor for entry)

• Cleaves sialic acid for viral exit

Swine Influenza Virus

• Pigs have 2 kinds of receptor: One targets bird viruses, the other targets human viruses

• Pigs can thus swap their genetic material and create novel, more virulent strains

• Considered to be “mixing bowls” of influenza viruses

Prion Diseases

• Transmissible spongiform encepalopathies (TSEs)

• Prion proteins also cause other normal proteins to fold incorrectly and so on in a chain reaction

• Peyer Patches are where major prion proteins can be found

• Recently discovered, least understood, low # of treatments

Prion Diseases: Bovine Spongiform Encephalopathy (BSE)

• Aka Mad Cow Disease

• BSE (Bovine Spongiform Encephalopathy) in cattle, a chronic degenerative disease in cows that forms “sponges” in the brain and spine

Prion Diseases: Kuru

Passed between people who practiced cannibalism

Eating Animal Products: A Common Cause of Human Diseases

• Most important pathogens for foodborne illnesses: Bacteria, viruses, prions and protozoans

• Alzheimer’s and Parkinson’s act like prion diseases but not from contaminated food

  • AD from aggregate Tau protein that causes neurofibrillary tangles

  • Cells constitutively secrete Tau at low levels under normal conditions

• “Food poisoning” is not from one single agent

• Gasteroenteritis (Stomach Flu): Long lasting diarrhea

Bacterial Food Poisoning

• Gram (-) protobacteria most common type for food poisoning

  • E. coli is the most studied and best understood organism

  • Broad Specificity Pathogens: A consequence of adhesive fimbriae on the bacteria surfaces grab onto particular glycoproteins and glycolipids on our cell surfaces (many specific options to grab)

• Not everyone is susceptible to food poisoning

• Not always the same symptoms

• Loss of fluids very common

Viral Food Poisoning

• Viral food poisoning is rare

• Costly to detect, hardest to treat

• Bats are often the original hosts because their immune systems allow for viruses to replicate quickly but are not virulent towards them

Virus Recombination

Two related viruses, when inside the same host, mix their genetic material and become more virulent mutants

Viral Food Poisoning: Rotaviruses

Recognized as the primary cause of gastroenteritis in young children

Viral Food Poisoning: Noroviruses

Common in seafood and oysters

Protozoan Food Poisoning

• Most Common: Toxoplasma, Cryptosporidium, and Giardia

• Giardia lamblia aka “Beaver Fever” is a pathogen found in mountain stream water

Genetic Code = Universal Triple Code

• P2 is thus the most important for determining the type of the amino acid based on the codon (most conserved)

  • Determines if P3 is important

  • When P3 is important, it only matters whether the position is a purine (A&G) or a pyrimidine (C&U)

• P1 important only for the specific amino acid

• Negative Selection Principle: What is least important changes the most

• Similar amino acids likely only vary in one position

P2 Amino Acid Types

• T = Hydrophobic

• A = Hydrophilic

Alternate Wheel Based on P2

• 4 quadrants

• 1 hydrophobic (T/U), 1 hydrophilic (A), 2 Semi-Polar (C or G)

Wobble Base Pairing

• P3 allows specific alternate base pairing

• For Start Codons, (fMet/Met), wobble is P1, not P3

Purine + Pyrimidine Binding

• More H-Bonds are formed between C+G than A+U

• Also bonds between tRNA and mRNA are in general stronger when the tRNA is a purine and mRNA is a pyrimidine

• Not all A:T binding or C:G binding are equivalent in energy

  • Depends what is the tRNA and what is the mRNA

  • T:A = mRNA:tRNA is stronger than the other way around

  • C:G = mRNA:tRNA is stronger

Stop Codons (eg UAA)

• Most common chain termination codon

• Lowest # of H-Bonds possible out of all the stop codons

• All of the common nonsense codons use U in P1

  • Pyl = Pyrrolysine (in archaea and bacteria)

  • Sec = Seleno-cysteine (in all domains of life)

• Ochre, Amber and Opal are all stop codons

Primordial Soup

Gave rise to lipids, nucleic acids, etc.

Benefit of Redundancy

Because multiple codons can code for the same AA< a SNP has less of a chance to change the AA identity and cause a problem

Asgard Archaea

• Found on sea floor sediments near hydrothermal vents

• Eukaryotes branched off from Asgard archaea

• Saw fusion of bacterial and archaeal cells

• Many archaea genes are considered Eukaryotic signature proteins

Asgard Genomes Encode Typically Eukaryotic Systems

• Suggest Eukaryotes should be placed in Archaea

  • MVA Lipid Synthesis Pathway

  • Membrane Remodeling/Trafficking Systems

  • Cytoskeletal Proteins

  • Ubiquitin

  • Vesicle Formation Systems

CPR and DPANN are Mostly Symbionts and Episymbionts

• Episymbiont: A symbiont that lives on the surface of another organism for survival

• Slightly different genetic codes

• Have unusual ribosome compositions (missing universal “essential” ribosomal proteins)

• Gaps shared for metabolic capacities

  • Gaps aren’t shared well with non-CPR symbiotes

CPR

• Have small genomes and cell sizes and most have symbiotic lifestyle

• CPR bacteria lack key biosynthetic pathways

• Nano-Archaea look very similar, both are associated with much larger microbes

DPANN

• Small genomes, small cell sizes and limited metabolic abilities (cannot make nucleotides, AAs, and lipids)

• Have an outer membrane and unique surface attaching grappling hooks called hami

Radiation

Increased taxonomic diversity caused by elevated rates of speciation

Metagenomics

• Genome-resolved metagenomics approaches allow for studying metabolic pathways spanning different species in a community

• Metabolic insights

Nanobacteria

Most are obligate anaerobes and cannot tolerate oxygen

Non-CPR Bacteria

• Can mix aerobic and anaerobic pathways, metabolic versatility, and can survive in changing conditions

• Exist in our microbiome

• Carry out fermentation, not photosynthesis

• Secondary metabolites give insight into lineages

Episymbiosis Common for CPR and DPANN

• Associated with cell surfaces, cell-to-cell contact

• Cryo-TEM show pili-like structures extending from CPR cell surfaces

Isoprenoids

Metabolites that are essential in all living organisms in all domains of life

“Lipid Divide”

• Archaea membranes composed of isoprenoid-based lipids, precursors made via MVA Pathway

• Bacteria have nonhomologous MEP pathways

  • MEP pathways not reported in Archaea

  • MVA lost in most bacteria

• Lipid Divide between bacteria and archaea by MVA and MEP distribution is muddied by these results

Metabolic Handoffs

Everything is made, consumed and digested

Rapid Evolution

Reduced genomes tend to increase evolution rates

Early Divergence

Possibly don’t have enough missing links

Convergent Evolution

Mostly ruled out by nonoverlap with non-CPR genome-reduced symbionts like buchnera

Horizontal Gene Transfer

• “Borrow” systems from other lineages

Gut Brain Axis in Autism Disorders

• ASD: Autism Spectrum Disorder

  • Highly multi-factorial (genetic and environmental)

• Many children with autism suffer from various GI issues such as IBD and Crohn’s Disease

• Biochemical signaling between the GI tract and the ENS/CNS via the Vagus Nerve

Gut Microbiota

• Microbiome strengthens intestinal barrier integrity, protects from inflammatory disorders

• Microbiota development begins at birth, stabilizes at ~3 y/o

• Microbiota and CNS metabolism can directly or indirectly affect homeostasis

• Disruption of the mucosal microbiota

  • Dysbiosis: Can cause several diseases and disorders such as ASD

  • “Leaky Gut”: Cytokines from the gut can get into the bloodstream and then pass the blood-brain barrier, wreaking havoc

Il-6

• Inflammatory product in excess can cause abnormal neuron growth

• Inhibition of Il-6 has reversed behavioral abnormalities in offspring prenatally exposed to Maternal Immune Activation (MIA)

• Affects stress levels

Metabolism

• Microbiota produces many metabolites and ligands

• SCFAs: Short Chain Fatty Acids act on the CNS through the microbiota-gut-brain axis

• Tryptophan can be converted into a metabolite that generates neurotoxic products depending on changes to microbome

Nervous System

• ANS: Autonomic Nervous System

• ENS: Enteric Nervous System

• HPA: Hypothalamic-Pituitary-Adrenal Axis

  • Communicate via vagus nerve to mediate gut-brain axis

Neurotransmitters

• Metabolites cross BBB and change NT levels

• GABA is not able to cross the blood-brain barrier and only acts locally

Maternal Microbiota

• Maternal gut microbiota modulates growth and fetal brain development

• Maternal gut microbiota influences fetal BBB, where it can upregulate the expression of tight junction proteins

• Vertical transmission of microbiomes, birth method can have significant differences in microbiome diversity

Epigenetics

• Change acetylation and deacetylation

• Deacetylation: Gene inhibition by condensing the chromatin

• HDACs: Histone Deacetylases are targets of microbiota-derived metabolites

• ASD postmortem brain samples show abnormal alternative splicing of mRNA

BBB

• Blood Brain Barrier keeps brain free of pathogens

• Has tight junctions reinforced by Zonulin

• Glial cells also surround blood vessels in the BBB

  • Altered Blood-Brain Barrier allows in inflammatory cytokines and causes neural inflammation

• Some microbes produce metabolites like butyrate which can alter activity of BBB cells

Food Selectivity

• Picky eating and severe food selectivity (FS) in ASD patients

• Consumption of particular diets alters gut microbiota to favor specific bacterial genera

• Less-diverse diet could reduce microbiome diversity

ASD: Social Interactions

Deficits in social behavior which can be reversed by probiotics or microbiota recolonization

Germ-Free (GF) Animals

GF mice have social behavioral problems compared to normal mice

Probiotics, Prebiotics, and Synbiotics

• Probiotics: Species of live microorganisms that provide health benefits to the host upon ingestion

• Prebiotics: Non-digestible fiber compounds that act as a substrate for the growth of beneficial microbes

• Synbiotics: Contain a combination of specific probiotics and suitable prebiotics may produce similar beneficial effects in ASD

Fecal Microbiota Transplantation (FMT)

Significantly reduced GI problems

Biomarkers

• Butyric Acid: SCFA with proinflammatory function of intenstinal macrophages, regulates BBB permeability

• Zonulin Levels: Gut tight junction-stabilizing protein upregulated in ASD children, zonulin could be helpful with intestinal permeability

Therapeutics

• No universal treatment as of yet

• Significant heterogeneity of ASD is not helpful in truly understanding the significance of one variable

Cooperation Not Competition

• Cooperation may be the true driving force for survival and evolution

• Plastids in plants and mitochondria in most major Eukaryotic subdivisions arose via initial endosymbiotic events from cyanobacteria and proteobacteria

Bacterial Parasites

• Bba: B. bacteriovirus

• Bex: B. exovorus

• Two Parasitic Modes:

  • Bba grows in the periplasm of the prey cell

  • Bex grows externally, often attached to outer surface of the host bacterium

Two Parasitic Modes: Similarities and Differences for Bba and Bex

• Differences:

  • Growth

    • Bba grows in the periplasm of the prey cell

    • Bex grows externally, often attached to outer surface of the host bacterium

  • Different transport systems

  • Different iron acquisiton systems

  • Bex has smaller genome than Bba

• Similarities:

  • Have large numbers of pseudogenes and incomplete systems, suggesting they are undergoing genome size reduction with a rapid loss of function

Bba Life Cycle

• Attack phase

• Outer membrane penetration

• Periplasmic growth

• Final synchronous cell division

  • Bdelloplast: Single snake-like multi-nucleoid cell

Bex Life Cycle

• Bex grows completely outside the cell

• Grow and divide binary fission

Symbiotic Archaeal Species

• Nanoarchaeum equitans: Novel small type of archaeon

• Ignicoccus

Metagenomics and CPR

• CPR has self-splicing introns and proteins encoded within their rRNA genes

• Metagenomics restated: The DNA of entire communities of bacteria simultaneously sequenced without separation of the different species from each other

Symbiotic Archaeal Species: Asgard

• Small cells depend on cooperative symbiotic relationships, host provide one or more essential metabolite

• “Reverse Flow Model”: Updated symbio-genetic model for the origin of eukaryotes that involves electron or Hydrogen flow from an organoheterotrophic archaeal host to a bacterial symbiont

• Much greater diversity of the bacterial domain suggesting bacteria were the first inhabitants of the Earth

Cooperation Rather Than Competition: Main Takeaways

• No primordial life form could have a full complement of metabolic and biosynthetic catalytic proteins that would allow living organisms to reproduce in the presence of only abiotic sources

• Genetic recombination and interspecies gene transfer, mediated by transposons and plasmid-like elements facilitated events that bring the DNA together

• Since evolution tends towards complexity, suggested symbiotic relationships provide a potential pathway for creation of more complex organisms

Obligate

Must be a certain way

Facultative

• Has a preference but can survive in either

• Flips the preferred condition

  • Ex. Facultative Anaerobe: Can grow without oxygen but prefers having oxygen

Aerobe

Oxygen-dependent

Anaerobe

Oxygen-independent

Lamarck

• Lamarckism (Soft Inheritance): First theory of inheritance of acquired characteristics

• Theory of Use and Disuse for phenotypic characteristics

Bioinformatics

• Every detail of an organism is encoded

• Decipher this information

• And then render this information

Molecular Phylogeny

Most reliable guide for function and mechanisms and phenotype

Microbiome

• Multiple microbiomes, one in each tissue

• Influences health and activity

Immune System

• Two Types:

  • Innate: Fast and general

  • Adaptive: Slow and specific

• May accidentally kill commensal bacteria

  • Commensal: Beneficial (mutualistic)

  • Virulent: Pathogenic

• Reinforces barrier immunity of host, limit inflammation

Lines of Defense

• First Defense: Mucosal Firewall (Mucus line)

• Second Defense: Antimicrobial Peptides produced from epithelial cells

• Third Defense: Macrophages (adaptive immune cells)

Interdependence Triangle

• Diet, Microbiota and Immunity

  • Dietary and Environmental stress alters GI microbiota

  • 80% of serotonin is made in the gut while only 20% is made in the brain

Dynamic Instability

• “Stabilizing Cap”: GTP-bound subunits

• Under the cap, tubulin subunits are in the GTP-bound state

• GDP-bound microtubules (Catastrophe) depolymerization

FtsZ

• Prokaryotic tubulin homolog

• Forms Single Strand Polymers

• Main component of the cell division machinery

  • Assembled at mid-cell in a discontinuous ring-like structure

• Exhibit Treadmilling: Polar filaments grow on one end and disassemble at the other

Fts stands for..

Filament temperature sensitive

Different Domains or Regions of Bacterial Tubulins

• A poorly conserved N-Terminal region

• The main GTPase domain that has a tubulin fold

• Highly diverse C-Terminal linker

• C-Terminal conserved (CTC) peptide

Bacterial Actin: MreB

• Conserved in many rod-shaped bacteria

• Filaments are non-polar so they do not exhibit the treadmilling activity that is a hallmark of eukaryotic actin

• Uses an amphipathic alpha helix to bind negatively curved membranes, guiding peptidoglycan synthesis and enforcing the rod shape

Bacterial Actin: FtsA

• Acts in cell division, anchoring the Z-Ring to the membrane

ParM

Plasmid separation

Intermediate Filaments: Crescentin

• Aligns along the inner curvature of the bacterium’s crescent-shaped body

• Crescentin filaments are elastic and solid-like

• Does not physically bend the cell

• Instead, it imposes mechanical strain on peptidoglycan biosynthesis machinery, leading to uneven longitudinal growth, results in curved shape

• Membrane tethering via the positively charged N-Terminus

Pbp

Penicillin Binding Proteins, involved in cell wall synthesis

Rod Shape Development

• FtsZ is a ring in the center

• MreB helps keep shape