Notes from 09/06 & 09/08

Which medical treatments involve microbiota?

1.0History of Microbiology cont’d.

Medical Treatments Involving Microbiota

• Fecal microbiota transplant (FMT) 

- life threatening colitis

• Treatment  

- surgery to remove infected intestine  

- transplant of clean fecal matter from family member

Video: Meet your microbes

* Questions on exam for this 

• Summary of video 

- Believed he got diabetes from drinking water from a river while hiking - Microbes do good most of the time instead of hurting us (the ones that cover us) - 10x as many cell so microbes on us than human cells

more mass in microbes than mass of brain

What eveidenec support chemical evolution as origin of life?

We also discuss several other topics like How many tribes consist the early native americans?

- DNA sequencing is the best way to look and learn about microbes

- 1,000 upon 1,000 of types of microbes covering us (microbial diversity differs  between people)

- the variation of microbes may be the cause of diseases in humans

- trigger of type I diabetes: miss communicating with microbial community in the  body Don't forget about the age old question of What do you call the site of respiration?

- Ilia microbes: experiment Don't forget about the age old question of Which aspects of financial accounting fall under liabilities?

• Donor ilium, transplanted into a recipient with microbial community to study the  effects. (ex. Crohn’s disease patients) We also discuss several other topics like What does natural selection consist of?

- Mentioned children born via C-section being swabbed with vaginal microflora - Germaphobia leads to excessive use of antibiotics can lead to the killing of --- the  microbes around us and in us that protect us.  

- What to do to restore microbial community: probiotics

What does astrobiology study?

- Animals eat poop, copraphagia, to help with restoring health

- In humans: probiotic microbe community through fecal transplants to help cure  system infections (FMT)

- View microbes as a functioning organ that is part of our cells

- DNA sequencing allows detailed studies of large amount of patients  

compared to those who are healthy and study the community of genes between the  two and see the differences  

- Later realized that he had diabetic symptoms since he was younger

- Can see microbial community through the affects they have on people and by their  DNA

1.1/1.2 First Life & Three Domain Tree

Origin-of-Life

• 3 ideas Don't forget about the age old question of What are the costs associated with hospitality in hospitality industries?
If you want to learn more check out What supports the cell membrane to become more rigid as cholesterol?

1. Divine creator

• A supernatural being created earth and the 1st cellular organisms 

- anaerobic prokaryotic organisms

- Earth is 4.6 byo

- 1st cells appeared ~3.5 bya

• Unknown where the original organisms come from

2. Pansmeria 

• Popular idea of how life originated

• Life originated from microorganisms or chemical precursors of life from outer space  and were able to initiate life after reaching earth

• Current understanding  

- life can’t withstand heat of meteorites

• causes idea to be less likely possible

3. Chemical Evolution or Abiotic Synthesis 

• The formation of complex organic molecules from simpler inorganic molecules  through chemical reactions in oceans during the early history of earth • Formed protocells

- unknown how it happens

• Carbs, lipids, proteins formed by carbon, oxygen, nitrogen, and hydrogen  - present in earth  

• Sources of energy

- volcanic activity, meteorites, & radioactivity

- lightening, heat, & UV light

• Sources of energy used for simple inorganic molecules to come together - simple macromolecules

• sugars

• amino acids  

• nucleotides

• lipids

• Absence of oxygen is critical to the process

- chemical bonds form without oxygen present

Building Organic Molecules Requires Electrons (e-)

• Oxygen is electron selfish

• Highly electronegative

- grabs all the electrons it can get

- oxygen present: bonds less likely to occur  

• The lack of oxygen in the atmosphere of the early earth’s atmosphere favored the  synthesis of organic molecules 

• Reducing atmosphere 

- there is no oxygen so bonds form

- monomers to polymers

Timeline

• 1.1 billion years between formation of the earth and first prokaryotes • even in a reducing environment an energy source is required

- lightning, UV radiation, thermal and kinetic energy from meteor impacts

• building of biological molecules from inorganic precursors

Experimental Results Supporting Chemical Evolution/ Abiotic Synthesis 1. Primordial Soup Experiment 

• 1953

• Replicate environmental conditions of prebiotic times

• Atmosphere  

- H2O

- H2

- CH4

- NH3

• Experiment has found 

- organic compounds

- amino acids

• Supports chemical evolution

• Experiment has been modified over the last 50 years

- yielded all amino acids and simple organic molecules

• Various gases at different concentrations passed through a source of energy through  a condenser. The collection of molecules are collected for examination  2. Murchison Meteorite 

• 1969

• Found in Murchison, Australia  

• Rock dated to be ~5 byo

• Analysis identified 

- amino acids

- organic compounds

• Organic compounds found in cells can form under the appropriate conditions in  nature 

- with the right amount of time

• Supports chemical evolution

3. Astrobiology 

• Study of the origin, evolution, distribution, and future of life in the universe • The Mars orbiters and rovers look for chemical signatures consistent with organic  molecules associated with cells

• The Philae lander conducted chemical analysis to determine, what if any, organic  compounds are present on comet 67P

- Nov 12, 2014

• Looking for biologically relevant molecules in the universe

Evidence for Early Life 

Fossil Timeline

• Oldest fossil: 4.6 byo

- known from oldest rocks dated by geologists

• Microorganisms existed for ~3.2 billion years before multicellular organisms • Multicellular bodies with bilateral symmetry were not present before 600 mya

- multicellular organisms with a body appear in the fossil record 600 million years  ago

• Cambrian Explosion  

- explosion of biodiversity

- started 545 mya

- by 600 mya fossil ancestors appeared…

(all eukaryotes)

• crustaceans

• starfish

• sponges

• mollusks (creatures with shells) (ex. snails)

• worms

• chordates (no backbones)

• algae

Stromalites  

• Microbial fossils in layered sedimentary rock

• Seen in Western Australia

• ~3.5 byo

** Microscopic resemblance to photosynthetic organisms 

• Fossils are not living organisms

• Section them thin, you’ll see objects in them (like cells) but are fossils so can’t be  alive

• Similar to modern microbial mats

• Community of physiologically integrated organisms

When did eukaryotes arise?

• 2.3-2.4 bya

• based on oil droplets within quartz crystals

- sterols, including cholesterol, found inside

• Prokaryotes don’t have cholesterol 

• Sterols produced almost exclusively by eukaryotes  

• Microfossil resembling eukaryotes are just older than 2 bya

Hypothesized Time Line

• Earth formed 4.6 bya

• Methane dominant environment

- anaerobic planet  

• 3.5 bya anaerobic prokaryotes appeared

- seen from stromanocytes  

- no oxygen present

• 2.3-2.4 bya photosynthetic bacteria appeared

- byproduct is oxygen

- oxygen accumulates rapidly in atmosphere

• 1.5 bya eukaryotes appeared

- without bilateral symmetry  

• Cambrian explosion

- 545 mya

- eukaryotes with bilateral symmetry appeared  

• Eukaryotes are fundamentally aerobic  

RNA may have been the 1st nucleic acid  

• Many early life biologists assert this idea

• Nucleic acids of the 1st cells

• Reasoning  

- RNA has catalytic activity

- DNA requires complex proteins for replication

• Pulls apart the double helix

- RNA is single stranded; simpler design

- Some have enzymatic activity and can catalyze the synthesis of new RNA • Auto-catalyze short strands  

→ autocatalytic properties  

- RNA of ribosomes catalyze peptide bond formation during translation • Catalyze other enzymes

- Store genetic information

Single stranded RNA can fold back on itself  

• Can form complex structures

• Ability classifies it as a secondary structure (2°)

• Functions in cells

- tRNA

- ribosomes

- regulatory RNA

• DNA doesn’t have these abilities

Question: Are chemical evolution & abiotic synthesis a possible factor of spontaneous  generation?

• Yes

How did eukaryotes arise from prokaryotes?

• LUCA were anaerobic prokaryotic organisms

• Model of life is a rooted tree

• Model presents modern organisms  

** How did eukaryotes come about?

Endosymbiosis  

- best explanation

• Relationship between two organisms (prokaryotes), in which one is living inside the  other 

• Engulfed but not digested

• Both organisms benefit

• Relationship became obligatory (required)

• Abundant evidence

- eukaryotes developed from endosymbiosis

Constructing the Three Domain Tree of Life  

Domain 

• Largest taxonomic group

**repeated Tree is built from comparing DNA sequences for ribosomal RNA (rRNA)  genes

• All cells have ribosomes

- made up of RNA

• All cells have genes for rRNA

• rRNA genes mutate slowly

- constant/predictable rate

• Closely related species have similar rRNA sequences 

• Distantly related have more differences 

** Ribosomes are composed of protein & RNA

• Composed of large subunits and small subunits

• Relative estimate of size comes from small ribosomal subunit 

• Prokaryotic 

- 16S (RNA)

• Eukaryotic 

- 18S (RNA)

** S – unit used to describe size

Svedberg Unit (S)

• unit describing sedimentation rate in a centrifuge tube 

• size of particles increase the lower into the tube the particles rest  Gene (DNA) for the small subunit ribosomal RNA

• rRNA used for building the tree

• The small subunit (30S) of the prokaryotic ribosome is composed of 16S RNA • The small subunit (40S) of the eukaryotic ribosome is composed of 18S RNA Compare DNA sequences that code for ribosomal RNA (rRNA)

• Alignment 

- create a closer relation

• Closely related 

- similar rRNA sequences

• Distantly related 

- more differences

- further on the tree

Ribosomal RNA

• Genes are present in all organisms

• Sequence is sufficiently conserved to define groups 

- define 3 main groups, and the subgroups

- slow mutation rate

• Adequate variability to determine evolutionary relationships

• Basis of 3 Domain Tree of Life

Tree Consists of Current/ Extant Organisms

• Extinct organisms are absent  

• Bacteria and archaea near the main trunk are used as a model for LUCA • Alive organisms today

Study Modern Day Organisms that are Closest to the Hypothesized LUCA on the Tree

• Hyperthermophiles 

- bacteria and archaea that can live in very warm temperatures

- 100° C

The Tree is complicated by Horizontal Gene Transfer 

• Tree gets messy

• Comparison of complete genomes from all 3 domains shows substantial transfer of  genes during the beginning  

** HGT (Horizontal Gene Transfer)

- 2 cells that bump into each other and exchange DNA

• Humans

- vertical gene transfer

** Horizontal/Lateral Gene Transfer 

• Genetic exchange

• Transfer of genes across species barrier

• 5 mechanisms

1. Transformation 

• Uptake of DNA from the environment

2. Conjugation 

• Physical contact between 2 cells

3. Transduction 

• DNA is moved by viruses/transposable elements  

4. Endosymbiosis 

• DNA transferred from the endosymbiont/organelle to the nucleus

5. Fusion of organisms 

• Two organisms become one

** Cells done have to be in the same species

1.3 Eukaryotes & Endosymbiosis  

Endosymbiosis

• Ex. mitochondria & chloroplast  

- ancestor prokaryotes residing in another

Lynn Marguis

• Cell biologist

• Proposed that eukaryotic organelles were formed by endosymbiosis • Paradigm shift in cell and evolutionary biology

• Used light microscopy and electron microscopy to study metabolism • She argued that the similarity in shape, size, structures and metabolism between  mitochondria/aerobic bacteria and chloroplasts/photosynthetic bacteria was more than  coincidence  

- looked at the metabolism of aerobic bacteria

- can look at DNA, mitochondria of chloroplasts (prokaryotic DNA)

Evidence for Endosymbiosis

• Modern mitochondria share characteristics with prokaryotes

• Approximate size and shape of a prokaryote

- similar internal structure

• Mitochondrial circular chromosome with Rickettsia – like genes

• Rickettsia

- pathogens that invade eukaryotic host cells 

• several species  

• ex. typhus (human pathogen)

• Mitochondria divide by independent division or binary fission 

• Prokaryotic like ribosomes

- different from eukaryotes

• Prokaryotic like membranes

• Mitochondria decedents of prokaryotes

Chloroplast

• Modern chloroplasts share characteristics with prokaryotes 

- approximate size and shape

• Circular chromosome with cyanobacteria-like genes 

- blue/green in color

- photosynthetic bacteria

• Independent division

• Prokaryotic like ribosomes

- different than ribosomes in plants

- cyanobacteria chlorophylla & thylakoid membranes

• composition of membrane are similar to prokaryotes  

Mitochondrion & Chloroplast

• Unique DNA

• Ribosomes

- make their own proteins

• Manipulate electrons in a membrane to generate ATP

• Have internal membrane that are prokaryote like

• Mitochondrial DNA is closely related to Rickettsia DNA

• Chloroplast DNA is closely related to cyanobacteria DNA and not algal DNA Symbiodinium dinoflagellae

• Animals

• Unicellular, marine photosynthetic eukaryote 

• Endosymbionts of coral, sea anemones, and jellyfish

- coral largest structure that look like plant

• Conducts photosynthesis within the host – coral

- symbiosis get carbon & nitrogen from what coral eats

- live within host – derived vacuoles inside the gut cells 

- translocate photosynthetically fixed carbon compounds that support the coral’s  metabolic needs

• byproduct is sugar

• The coral provides nutrients for algal growth  

Hydras

• Common in rivers and lakes

- fresh water

• 1”- 1 ½ “ in length

• Eat whatever they can reach and put into their mouth

• When hydra reproduces sexually, algal cells pass to the next generation via the eggs Azolla (plant)

• Water fern and the leaves that float on the surface of ponds

Anabaenza azollae 

• Not a pathogen

• Cyanobacteria that grows in chains within Azolla

Azolla & Anabaenza azollae

• Two cell types

- Vegetative cells 

• Perform photosynthesis

- Heterocyst 

• Fix atmospheric N2 to NH3

→ Nitrogen fixation

→ Create amino acids & nucleotides (DNA,RNA)

o with the ammonia

• Larger cell

• Thick cell wall

• Thrive together

- can grow separate but doesn’t have chain formation or do nitrogen fixation • Anabaena receives sugar = carbon source

• Azolla received NH3  = a usable source of nitrogen

Pea aphid & Buchnera aphidicola

(insect)   (bacteria)

• Puncture plants & feed on the product of photosynthesis  

• Pea aphids feed on plant sap

- sap is carbohydrate rich, but amino acid poor 

• Buchnera lives inside special cells of the aphid and produces essential amino acids - 106 cells/aphid

- from photosynthate

• Buchnera is an obligate intracellular organisms  

- cannot survive outside its aphid host

• Has 600 genes, a remarkably small number for a bacterium  

- because it gets a lot of what it needs from aphid 

• Aphids reproduce parthenogenetically and Buchnera are placed in all eggs  - lay eggs without mating

- so next generation has the bacteria  

• The two organisms are dependent upon one another

- one dies without the other

Elysia chlorotica

• Green sea slug  

- marine animals

• Feeds on algae

- can grow on and suck out contents of cytoplasm

- digests cytoplasm & nucleus

- removes whole chloroplasts which are sequestered in slug vacuoles • do not digest

• Algo chloroplasts can source photosynthesis  

** Slug has algal genes within its nucleus that codes for proteins needed for photosynthesis  and chloroplast it gets from algae 

• Horizontal gene transfer between an alga and an animal 

- this is endosymbiosis 

Cyanophora paradoxa

• Found in fresh water

• Unicellular, aquatic photosynthetic eukaryote

• Chloroplast contains small amounts of peptidoglycan between its inner and outer  membrane 

- usually only found in cell wall of bacteria except in this case

- a carbohydrate

• Chloroplast are of prokaryotic origin

Hydrogenosomes

• Associated with anaerobic eukaryotic organisms

• Some microbial eukaryotes are obligate anaerobes or aerotolerant  - don’t have mitochondria

• Obligate anaerobes 

- do not use O2 and O2 is deadly

• Aerotolerant 

- tolerate O2, but do not use it

• Examples

- Trichomonas

• Causes STD

- Ciliated protists

• Inhabit the animal rumens

- Anoxic muds

- Sediments

• Resemble mitochondria  

- double membrane

- most do not have any DNA

- those with DNA are clearly related to mitochondria by sequence comparison How Hydrogenosome’s Work

• Enzymes within the hydrogenosomes

- derived from prokaryotes

• Acetyl-CoA is converted to acetate 

- produce ATP 

• Lack the citric acid cycle and the electron transport chain

• Microbial eukaryotes have endosymbiotic bacteria living within their cytoplasm Endosymbiotic Bacteria 

• Consume CO2 & H2 (waste product)

• Produce CH4

1.4 Evolution

Evolution

• Change in the frequency of alleles within a population over time 

• Alleles : different versions of genes

• Involves inherited traits

• All populations have genetic variation/diversity

• Beneficial adaptations are identified during differential reproduction  • Change allele frequency

- mutations produce new alleles 

• The new allele must be inherited to be useful

- true in all organism

• include

→ bacteria

→ archaea

→ viruses

• Mutations are random

- sometimes useful

Genetic Changes

• Occurs in a population over time (evolution)

• Microevolution 

- stable phenotypes

• Macroevolution 

- new species

• Microevolutionary events

- new stable phenotypes

• If the sub-populations become reproductively isolated & microevolutionary events  accumulate a speciation event may occur

- macroevolution

• Three Domain Tree of Life

- describes evolutionary relationships

- focus on small subunits of RNA

Microbial Evolution

• Random mutations 

- allele frequencies

• Natural selection 

- stabilized through

• Horizontal gene transfer 

- prokaryotes

- bacteria & archaea

- species barrier isn’t a factor

Bacteria & Archaea

• Divide by binary fission 

- copy chromosomes, stick to membrane, and pinch in half

• Division proceeded by chromosome replication from single origin  • E. coli cells can divide every 20 min

• DNA molecules are identical except for mutations  

Mutations

• Rate ~1 mutation/chromosome/generation

• Short generation time = lots of mutations

- 107-108 mutations/12 hours

• High rate of mutation for eukaryotes 

Viruses

• Especially high mutation rates 

- ~1 in 10,000 nucleotides

- human rate: ~1 in billion nucleotides

• Low fidelity DNA and or RNA polymerases 

- lack proof-reading capability

- sloppy

- mutations accumulate

• Reassortment is common 

- exchange nucleic acid segments with different strains of the same virus • mixing & matching

** Mutation & reassortment generate a genetically diverse virus population Influenza virus

• great example of viral evolution

• strains move from host to host

• ability to reproduce in a new host = natural selection 

** New human flu strains appear each year through mutation & reassortment