Exam 1

Antony van Leeuwenhoek (1632-1723)

First person to accurately observe and describe single-celled micro-organisms.

Sulfate and nitrate ions accumulate in oceans

Through oxygenation of the atmosphere, microbes drastically changed the chemistry of the oceans by allowing oxidation of various chemical species, increasing the diversity of chemicals on earth that could be metabolized my organisms to obtain energy

Transduction

Transduction is DNA transfer mediated by viruses that infect bacteria (i.e. bacteriophages). Conjugation is direct cell-to-cell transfer of DNA. When a bacteriophage attacks a bacterial cell, it first attaches to a receptor on the bacterial surface. The surface attachment is taxon (or species)-specific. Phage attachment proteins are specific to protein receptors on the surface of its host cell. Then it injects its DNA in its capsid into the bacterial cell. Once the DNA of the phage is injected, it starts to reproduce itself. This process is called a lytic infection, because the cell that hosts the phage eventually lyses. During this process, parts of the DNA from the host can be inadvertently picked up and moved. This happens because a phage that is replicating chops up the chromosome of the bacterium it infects. In the frenzy to assemble new phage particles, a piece of bacterial chromosomal DNA instead of the phage's own genome may be packaged into a phage head. The phage particle carrying bacterial DNA can later attach to another bacterium and inject the bacterial DNA into it.

After bacteria are transferred from one medium to another, they often enter a _____ phase immediately after the transfer while they are adapting to the new medium

lag

A newly discovered microbe in the Hudson River was found to obtain its reducing electrons from hopanoid lipids within cyanobacterial cell membranes. This microbe would be considered a(n)

organotroph

rRNA Gene Structure

(3) Good combination of con-served and variable regions.

Staphylococcus aureus

-30% noses -top 5 bacteria causing hospital infections (90k deaths/yr) -MRSA- Methicillin-resistant S.aureus

Morphology of microbial cells

-bacteria, archaea (no nuclear membrane, avg size: 1-5um) -fungi (has nuclear membrane, 4-25um) -protozoa (has nuclear membrane, 10-50 um) -algae (has nuclear membrane, size: 10-50um)

Prokaryotic cell morphology

-cell wall -ribosomes (protein synthesis) -tightly coiled DNA genome (nucleoid), cytoplasmic membrane (energy production)

rooting the tree of life

-knowing whether bacteria or archaea are more closely related to eukaryotes is important -unfortunately, we don't have an outgroup for life on Earth

eukaryotic cell morphology

-nuclear membrane -mitochondrion (energy production) -cytoplasmic membrane -Golgi body (protein localization) -endoplasmic reticulum studded with ribosomes (protein synthesis)

The rRNA Tree of Life

-shape of tree has been now supported by studies of other genes and genomes -DNA-based metric of relatedness among all living organisms

Why worry about phylogeny?

-taxon-based methods assume a "star phylogeny" where all taxa are equally related -phylogenetically related taxa are more likely to be phenotypically similar -community a and b share no exact taxa, but for every lineage in a there is a closely related lineage in b.

other prokaryotic properties

-thicker outer envelope with few exceptions -morphologically simple, metabolically complex (produce and consume methane, nitrogen gas, sulfides) -typically only 1 or 2 chromosomes (humans have 23 & 1000x DNA) -asexual reproduction by binary fission (rapid generation time=high evolvability)

What are the major morphological factors that distinguish bacteria from archaea?

1) Different cell wall biopolymer: bacteria have peptidoglycan (which is targeted by most of our antibiotics), while archaea do not have peptidoglycan in their cell wall. 2) Different cell membrane lipids: archaeal membranes are made up of terpenoid lipids (derived from isoprene), in which every several carbons extends a methyl branch. The branches strengthen the membrane because the methyl groups allow the lipid chains to physically lock into each other, thereby limiting movement of the lipid chains. In addition, archaeal lipids replace the ester link between glycerol and fatty acid with an ether link, C-O-C. Ethers are much more stable than bacterial esters, which hydrolyze easily in water. Finally, archaeal lipids can fuse to create a monolayer (tetraether), which is hypothesized to give archaeal cells greater stability under extreme conditions.

What are the major metabolic factors that distinguish bacteria from archaea?

1) Temperature tolerance: While bacteria can withstand extremely high temperatures (up to 90oC), archaea can persist under even higher temperatures (up to 120oC). 2) Complex carbon utilization: archaea seem to be limited to the use of simple molecules as sources of carbon and energy (glucose). 3) Pathogenesis: whereas bacteria cause many diseases of animals and plants, no disease-causing archaea have been identified.

Bacterial genomes are unique from eukaryotic genomes in several key ways. What are they?

1. Bacterial chromosomes can be circular, rather than linear (as in eukaryotes) 2. Bacteria and archaea are more prone to horizontal gene transfer than eukaryotes 3. Bacterial and archaeal genomes are on average much smaller than eukaryotic genomes 4. Bacterial genomes tend to be way more packed with genes. Bacteria have on average 15% of their genome as non-coding. By contrast, humans have over 90% noncoding DNA. 5. Bacterial genes tend to be organized in units called operon, where multiple genes are under the control of the same promoter.

Which specific environmental conditions on early Earth allowed the first life forms to emerge (around 3.7 GYA)?

1. Earth's crust cooled: The earth's surface had to cool down a bit before liquid water could form 2. Appearance of water: soon after that, life on Earth began to appear 3. Volcanic activity: volcanic activity was higher during this period than it is today and that was actually a good thing for life on Earth. The reason why is that volcanoes spew out lots of carbon dioxide, which is an important greenhouse gas. This helped keep the Earth's surface from freezing by trapping some of the solar radiation in our atmosphere. 4. Water vapor from volcanoes formed the first oceans. During this period, life really began to multiply and many new environments were colonized.

Describe how the bacterial cell wall is synthesized.

1. Precursors are made in the cytoplasm. The glycan (or sugar) part of the peptidoglycan cell wall is composed of alternating units of N-acetylglucosamine (NAM) and N-acetylmuramic acid (NAG). The process of synthesis starts in the cytoplasm, where precursors are made. The precursor molecules consist of NAG and NAM attached to uridine diphosphate (UDP) to make UDP-NAG and UDP-NAM-peptide. 2. These precursor molecules then interact with a lipid carrier (bactoprenol) embedded in the cell membrane. This lipid does two things, 1) serves as the matrix on which a unit of NAG is added and 2) transports the disaccharide across the cell membrane. Uridine monophosphate is released. All of this happens in the cytoplasm because the energy available to conduct those reactions occurs within the cell. 3. Once across the cell membrane, the precursors are polymerized to the existing cell wall structure by transglycosylases. Assembling the peptidoglycan itself is driven by transglycosylases located on the cytoplasmic membrane. As the subunits pass through the membrane, they are attached to a growing sugar backbone. 4. The peptide side chains are cross-linked by transpeptidases. Transpeptidases attach peptides on adjacent sugar backbones together to form peptidoglycan. If this cross-linking doesn't happen, the peptidoglycan layer has no strength and the bacteria ultimately lyse.

What are some basic structural features of bacterial cells?

1.Disease 2.Illustrates how microbes can survive and thrive 3.Detection and identification (i.e. gram stain) 4.Antibiotic resistance

Briefly describe the species concept of "operational taxonomic units" AND its limitation in microbiology:

An operational taxonomic unit (OTU) is a group of DNA sequences (often of the ribosome (16s region)) that have high similarity (>97%). It is possible that bacteria with this DNA similarity are not the same unique type of organism (ie. taxon). The sequence similarity cutoff is arbitrary.

Which groups of organisms are considered microorganisms?

Bacteria and archaea are typically referred to as "microorganisms". However, many people also consider fungi, algae, and protists to be microorganisms. This is because many fungi are small (some smaller than bacteria!), and most algae, protists, and fungi are simple in construction. Many consider viruses to be microorganisms, although whether or not a virus is an organism is a matter of debate.

Describe the difference between bacterial run and tumble. What is the role of the flagellum, and how do the bacterium use run and tumble states to generate a random walk in liquid?

Bacteria move in liquid currents primarily by run and tumble movements. The bacterial flagella acts basically like a propeller on a boat. The direction of rotation determines the nature of bacterial movement. If it rotates one way (counter clockwise, CCW), the bacteria move forward, in what is called a run. If it rotates the other way (clockwise, CW), individual flagella don't rotate in synchrony. This switch in the direction of rotation disrupts the bundle of flagella and you get the bacteria moving in a tumbling motion. This tumbling phenomenon is important because this reorients the bacteria, allowing it to swim towards or away from a certain environment. This is known as taxis, the ability to swim toward favorable environments (attractant signals, such as nutrients) and away from inferior environments (repellent signals, such as waste products). Taxis to specific chemicals is called chemotaxis. When flagellar rotation is reversed again, the cell sets off in a new direction. By periodically reversing the direction of flagellar rotation, the bacterium follows what is known as a "random walk". This walk, however, allows the bacterium to move up a gradient of a desired nutrient (so not really random...).

Describe how plasmid addiction systems work. Why would these systems have evolved?

Because plasmids require host cell DNA polymerase to replicate, they need to make sure to get into the daughter cells to persist. For high copy number plasmids, this is not really a problem because they are so many that enough will randomly diffuse into each daughter cell. For low copy number plasmids, however, they have to coordinate their replication with that of chromosome replication to make sure they are partitioned into both daughter cells. One of the more interesting example of the ways plasmids make sure they get into daughter cells is what are referred to as plasmid addiction systems. Plasmid addiction systems are designed to kill cells that lose the plasmid. These types of plasmids produce a toxin protein and also produce an antidote protein. They ensure their survival because if one of the daughter cells in division doesn't receive the plasmid, it still gets some of the toxin and the antidote in the cytoplasm. The catch is that the antidote protein has a shorter life than the toxic protein, so the absence of the plasmid makes it so the antidote is not being produced. This then kills the daughter cell if the plasmid making the antidote is not in it. Plasmid addiction systems are non-beneficial to the host. Nevertheless, plasmid addiction systems persist in bacteria because if a cell division produces a cell with no plasmid, the cell without the plasmid may be at a competitive advantage for growth because maintaining a plasmid drains energy. Thus, the cells without plasmids could overgrow the plasmid-containing cells, eventually eliminating them from the population.

Describe the process of binary fission, and how it differs from endospore formation.

Binary fission happens when bacterial cells split in two: DNA replicates when DNA synthesis machinery separates the parental strands in the chromosomes while simultaneously synthesizing the new, growing strands. DNA replication is semi-conservative, in that each daughter cell gets one parental strand and one newly synthesized strand of DNA. A septum forms in the center of the cell (from both membranes) and eventually splits the cell into two equal daughter cells. Endospore formation is a type of cell differentiation technique that allows bacteria to survive stressful conditions. DNA replicates, and a septum forms from the cytoplasmic membrane near one end of the cell. Once the endospore is formed inside the mother cell, the mother cell engulfs the endospore, effectively wrapping the spore in a second membrane. The mother cell's chromosomes disintegrate, the spore gets covered in proteins, organic acids, and calcium. Then, the mother cell releases the spore to the environment, where it can germinate when conditions are right.

What are the major features of bacterial biofilms?

Biofilms are matrix-enclosed microbial consortia that adhere to biological or non-biological surfaces. They are often resistant to stressors including biocides, such as human cells designed to kill pathogens (neutrophils). They are also structurally complex, including an exopolysaccharide (EPS) matrix. Biofilms can include cells in both exponential growth phase and stationary phase. Cells deep in the colony, at the agar surface, are in stationary phase because they can't get oxygen. Nutrients from the agar are not consumed by these cells, and instead, diffuse toward the oxygen-rich colony surface, where they feed the actively growing cells.

Prokaryotic Inner (Cell) Membranes

Both types have proteins interspersed throughout. Archaea have: 1.Terpenoid lipids: consistent methyl groups 2.Different (tetra-ether) glycerol-lipid link 3.Partial monolayer (bacteria have phospholipid bilayer)

If you were to create a molecule to serve as the perfect food source (i.e. resource) for a microorganism, which elements would you include and in what ratios?

Carbon, hydrogen, oxygen, nitrogen, phosphorus, sulfur are needed in high concentrations, with trace elements like manganese, copper, zinc, nickel, and molybdenum required in low concentrations. Other elements like iron, magnesium, and calcium are required in high abundance as well. So, I would construct a molecule that included elements perhaps in these ratios: CNSFeMgCa : MnCuZnMoNi = 95:1 Alternatively, you might construct a food source that has 100% major macronutrients (carbon, hydrogen, oxygen, nitrogen, phosphorus, sulfur, iron, magnesium, calcium), as the trace elements can usually be obtained from water.

Knowing about the role of che proteins in chemotaxis, describe the dominant direction of flagellar rotation in presence of attractant for the following E. coli mutant strains: 1) CheR deficient and 2) CheB deficient. In your answer, explain the molecular basis for the direction chosen.

CheR deficient: no methylation= constant de-activation of CheA=no poshphorylation of CheY=CCW (run) CheB deficient: No de-methylation= activation of CheA=phosphorylation of CheY=CW (tumble)

Conjugation

Conjugation is the term used to describe direct cell-to-cell transfer of DNA. During conjugation, two bacteria (a donor and a recipient) make a tight connection with each other. This contact is mediated by a specialized pilus (sex pilus), which attaches to another (recipient) cell and draws the two cells together. On the plasmid that is transferred, there are genes coding for the sex pilus, a channel between the donorand recipient cell. (i.e. mating bridge), and DNA transfer proteins. The plasmid uses one of its proteins called TraI (transferI) in a protein complex called relaxosome to nick the plasmid at its transfer origin (oriT). is followed by attachment of the single strand to the mating bridge, which then transfers the single strand into the cytoplasm of the recipient. DNA polymerase makes both the strand left in the donor and the strand introduced into the recipient double-stranded. So both donor and recipient end up with a double-stranded copy of the plasmid.

What does current and future research in microbiology focus on?

Current research in microbiology focuses on: Genomics: for example, decoding our own genetic make-up will to a large extent involve microorganisms. Much of the information about how to use genome sequences to understand the function of genes in a living organism will come from studies of microbes and genome sequences. Emerging Infectious Diseases: to find ways controlling the spread of microorganisms generating novel diseases in humans, domesticated animals, and crops. Industry: to find ways of understanding and exploiting the genetic and metabolic diversity of the microbial world. Diversity and ecology are additional research areas, because whereas over 50% of mammalian species and 20% of arthropod species have been identified, it has been estimated that fewer than 0.2% of microbial species are known to science. These microbes are the life support system of the Earth - yet we understand little about their fundamental interactions with other organisms and with the environment.

What are some key ways in which microbes changed the Earth system?

Cyanobacteria and other oxygen-producing microbes radically changed the environmental conditions on our planet. During the Great Oxygenation Event (GOE). One major effect was to expand the simple cycles of carbon, nitrogen, and other compounds to include more oxidized forms. For example, sulfides could be turned into sulfate or iron turned into iron oxide. Through oxygenation of the atmosphere, microbes drastically changed the chemistry of the oceans by allowing oxidation of various chemical species, increasing the diversity of chemicals on Earth that could be metabolized my organisms to obtain energy. The second major effect was the formation of the ozone layer. Once the ozone layer formed, UV-sensitive microorganisms could begin to move out of the ocean, where water provided some protection, and into terrestrial habitats. A third major effect occurred somewhat later, when the cyanobacteria were incorporated by eukaryotic cells to from the chloroplasts of algae and green plants. A third major effect was the evolution of symbiosis, when cyanobacteria were incorporated by eukaryotic cells to from the chloroplasts of algae and green plants. Endosymbiosis enabled the evolution of the eukaryotes.

Describe how the gene coding for the "Shiga toxin" (from Shigella bacteria) moved into "killer" E. coli strains O157:H7 and O26.

E. coli O157:H7 causes bloody diarrhea and can also cause kidney failure and death in some cases, particularly among children. So does E. coli O26, the strain that was the culprit in the Chipotle outbreak in 2015/2016. These killer strains appear to have been created by horizontal gene transfer, where a less virulent strain of E. coli acquired the genes to make a new toxin. In this case, the process of acquiring these genes appears to have been mediated by a phage, or a virus. It has been shown by looking at the composition of the toxin that the virus originally entered a Shigella species, which naturally produces the toxin, and inserted itself into its genome. When a new virus particle (virion) broke out of the Shigella cell, it took part of the Shigella genes with it, including the toxin gene. It then infected an E. coli cell and was incorporated into its genome. With this new gene, E. coli not only causes diarrhea, but the toxin it acquired makes that diarrhea bloody, leading to its more lethal effect.

Current Focus: Quantifying & Understanding Microbial Diversity

Estimated 1 Trillion (1012) species of microorganism on Earth. The vast majority of diversity of life on Earth is microbial!

Describe the structural components of the flagella.

Flagellum: a spiral filament of proteins called flagellin (protein FliC). Proteins MotA and MotB form part of the ion channel whose flux of hydrogen ions powers rotation, similarly to the proton flux that drives ATP synthase. Another protein, FliG, forms part of the device that generates torque (rotary force). To generate torque, the MotA/MotB stator unit changes its conformation in response to the ion influx and interacts with the rotor protein FliG.

How do prokaryotes and eukaryotes differ in their approximate average generation times and optimal growth temperatures?

Fungi and protists generally grow more slowly than prokaryotes, and at lower optimal temperatures.

Describe how trophic categories are distributed across taxonomic lineages of microbes and morphological groups of microbes

Generally, trophic groups match up with taxonomic and morphological groups of microbes. The prokaryotes (bacteria and archaea) generally have more diversity of nutrition types (i.e. trophic groups) compared to the eukaryotes. For example, most fungi and protists are chemoorganoheterotrophs. There are some exceptions, however. Algae are photoorganoheterotrophs, like some bacteria, and bacterial pathogens can be chemoorganoheterotrophs as well.

Not All Microbes are Microscopic

In addition to usually being small, microorganisms are relatively simple in construction and typically lack highly differentiated cells and distinct tissues.

Illumina sequencing

In Illumina sequencing, the genomic material to be sequenced is sonically fragmented into 100-300 bp segments and different linker oligonucleotides are ligated to each end. Strands of each fragment are separated and the mixture added and fixed to an optical flowcell. The flowcell has a dense lawn of oligonucelotides that are complementary to the ends of the adapters on each sequence. The fragments bend over and attach to the oligos, which serve as primers for amplification. Once amplification is complete, the double-stranded bridge is denatured and both strands remain attached to the surface. This repeats until 1000 identical copies of a gene fragment are generated = a cluster. Sequencing is then done in a series of single-step, reversible chain termination reactions. A fluorescently labeled nucleotide is added one at a time to the clusters. After a base is added, a snapshot is taken of the plates, illustrating the colors. Next, the fluor is removed, which reverses the chain termination, and the slide is flooded again with tagged bases. Illumina sequencing can sequence ~ 18 Gbp per run. However, it generates the shortest DNA reads (~ 300 bp) of all the sequencing technologies.

Describe why anemia (low iron in blood) could help you stave off infections.

In mammals, iron is tightly bound to proteins (glycoproteins) like hemoglobin and transferrin, leading to a very low concentration of free serum iron (10-24M). This is a challenging condition for microbes to be in, so many pathogens have evolved siderophores to capture these low concentrations of iron. The consequence of microbes having a good iron-scavenging strategy is that when iron concentrations rise in the human body, microbial pathogens can capture that iron very quickly and use it to grow. This idea was put forth by French physician, Trousseau, who in the late 1800s (1872) alerted his medical students in Paris in a seminal lecture series to the dangers of giving iron to infected patients. His evidence was that patients to whom iron was administered in their diet as a form of 'tonic' invariably had a poorer outcome than those patients who received no additional iron and were, in fact, somewhat anemic. Tuberculosis cases amongst the wealthy people, where there was a good chance of a nutritious diet with iron supplementation to correct any anemia, often fared worse than poorer people who had a diet that was inadequate in its iron content. Giving 'tonics' of unprocessed animal blood to tuberculosis sufferers was seemingly widely practiced in the 19th century and such a practice could not in fact have been worse for the outcome of the disease.

Isotopes & Dating Early Life

Isotopic evidence based on 34S/32S ratios show sulfate reduction by microbes around 3 bya.

The Future of Microbiology

Key features of microbes present lots of new challenges and opportunities! •Human, plant, and animal genomics •Emerging infectious diseases (e.g. COVID-19, Ebola, Zika). •New and improved industrial processes. •Microbial diversity and ecology. - Less than 1% of all microbial species are culturable.

Describe the phases of bacterial growth in batch culture. Why is it advantageous to use bacterial cultures in the exponential phase of growth for experiments?

Lag phase occurs as the cell is acclimating to the new conditions of the culture and is repairing any cellular damage that occurred during the transfer process. Exponential phase occurs when the organism is actively growing in the new culture environment. By contrast, stationary phase is when the number of viable cells remains constant: this could be because metabolically active cells stop reproducing or because the reproductive rate is balanced by the death rate of cell. Finally, the death phase occurs in batch cultures because nutrients are not added, so the cultures can only complete a limited number of life cycles before nutrients are consumed and growth stops. During the exponential (log) phase of growth, the number of cells produced is exponential, yet the rate of growth of a single cell is constant- all cellular components are synthesized at constant rates relative to each other. Therefore, the population is most uniform in terms of chemical and physical properties during this phase. This is why it is advantageous to use cultures that are in their exponential phase of growth for laboratory experiments.

What were the historical obstacles to assessing microbial diversity and how did using rRNA genes to measure diversity overcome them?

Microbes were officially discovered long after people had been studying plants and animals for many years. Because people were used to identifying plants and animals by physical traits such as appendages, shapes, and color, they started identifying microorganisms this way as well. However, this led to some challenges: Problem #1: vastly different kinds of microbes looked more or less alike. Linnaeus thought we would never be able to clearly distinguish one from another. Problem #2: Microbes do not readily fit the classic definition of a species—that is, a group of organisms that interbreed. For example, bacteria usually breed asexually. As a result, they were thought to be a relatively undiverse group of organisms. Problem #3 Another issue with the physically based systems emerged when scientists tried to place microorganisms on the tree of life. To make comparisons among all of the different lineages, you need a set of common traits from which to do so. Unfortunately with macrobes and microbes, they really don't share a lot of physical traits, so there is no one common yardstick against which all organisms can be measured. What the scientific world really needed was a classification system based on some trait possessed by all living organisms. In the 1970s, a scientist named Carl Woese put forth a suggestion that was radical for its time that went straight to the center of the diversity issue. He suggested that DNA sequences of certain common genes be used to determine the relatedness of different organisms. He chose to use the gene that encoded an RNA molecule found in ribosomes. Ribosomes are the RNA/protein complexes on which proteins are synthesized and they are found in all organisms. He could have chosen other common genes, but rRNA provided a couple of advantages: 1. There are lots of these genes in a cell because lots of ribosomes need to be made (10-3 to 10-5 molecules/cell). Thus finding this gene was not really a needle in a haystack. 2. The right size of the gene. There are three rRNA molecules in each ribosome. Woese chose the intermediate sized one because it was large enough to contain enough information for genetic comparisons but also small enough to be sequenced easily. 3. Good combination of conserved and variable regions. Conserved regions and variable regions make the ribosome a good yardstick. That is because you can use the conserved regions to (a) identify and amplify them in the cells of all organisms, and then you can (b) align the genes of different species to calculate divergence (or differences) among organisms.

A Brief History of Microbiology

Microbial metabolism have been informally known to humans for ages. Domestication: fermented foods (since at least 10,000 B.C.) Disease: soil identified as the source in 60 B.C.

What are two major defining features of microorganisms?

Microorganisms are 1) very small (often too small to be seen with the naked eye), and/or 2) simple in morphological construction, lacking highly differentiated cells and distinct tissues They are also: 3) capable of rapid rates of evolution 4) metabolically diverse for their morphological simplicity 5) taxonomically diverse, compared to other domains and kingdoms of organisms

When and where would it be beneficial to engage in mixotrophy?

Mixotrophy is energetically expensive - a lot of energy is needed to maintain all the proteins needed for many different trophic strategies. However, mixotrophy is a benefit in low nutrient (i.e. oligotrophic), unpredictable environments - so where conditions for growth are generally poor. Mixotrophy is also a benefit if you are able to disperse to many different types of habitats (i.e. are a "cosmopolitan" species, like Rhodopseudomonas palustris), and need to survive in those new habitats, which could be quite different (anaerobic lake sediments vs. glaciers in Greenland vs. earthworm droppings).

You have been invited to join the team of biologists studying extremophilic microbial communities in the hot spring ponds of New Zealand. In the pond you are assigned to sample, you take samples and generate a number of pure culture isolates. You check the temperature of the pond and it is exactly 90 degrees C. Knowing that both bacteria and archaea can persist at this temperature, name three ways (one based on morphology, one based on metabolism, and one based on genetic material) that a pure culture isolate could be differentiated these microbial groups.

Morphology: cell membrane lipid structure (archaea have terpenoid lipids, tetra ether linkages, and partial monolayers) Cell wall composition (bacteria have peptidoglycan, and archaea don't) Metabolism: Degradation of complex C compounds (primarily bacteria) Disease (bacteria) Genetic material: 16S rRNA gene composition

Nanopore sequencing

Nanopore sequencing is not synthesis based, but is electric-current based. It is a single-molecule sequencing technology that relies on the changes in electric current that occur when a single strand of DNA passes through a nanopore structure. DNA is ligated to an adapter and the adapter is recognized by an enzyme that ratchets one of the strands through the nanopore. The sequencer cannot recognize an individual nucleotide, but it can recognize short strings of bases (3-6 bases long) because they have signature changes in electrical current that they produce as they pass through the nanopore. Nanopore can generate ~800 kbp read per strand, yet it has a large error rate (92% accuracy only), which means that the same DNA strand needs to be sequenced multiple times to identify (and correct) the errors.

The Cell Wall and Outer Layers (Cell Envelope)

Protects the cell membrane -Surface polysaccharides (capsule) -The cell envelope includes at least one structural supporting layer. -Most commonly: the cell wall

What is a mobilizable plasmid, and how does it differ from a self-transmissible plasmid?

Not all plasmids have the genes that encode the proteins that form the mating bridge and carry out transfer of the DNA segment. In fact, only a small fraction do and because of the large number of genes needed for conjugation, self-transmissible plasmids are usually quite large. Other plasmids, mobilizable plasmids, cannot transfer themselves because they lack the genes needed to create a mating bridge. Mobilizable plasmids can, however, make use of the mating bridge furnished by the self-transmissible plasmid to complete their own transfer. Mobilizable plasmids can exploit the conjugative-plasmid mating pore by encoding a mimic sequence of the conjugative-plasmid oriT that is recognized by the relaxase of the transferable plasmid. The relaxase of. The self-transmissible plasmid can then cleave and covalently attach to DNA at oriT, forming a nucleoprotein complex referred to as the relaxasome. The relaxasome is recruited to the mating-pore, after which it is transferred (along with the attached plasmid DNA) to recipient cells through the mating bridge.

What is/are the major cellular feature(s) that distinguishes prokaryotic and eukaryotic cells?

Nuclear membrane

Given that stromalites seem like good fossil evidence - what else could you look for in the stromalites that would further corroborate the argument that they are products of biological activity? List at least two other pieces of chemical biosignature evidence you could collect.

One could look for many types of chemical biosignature evidence of biological activity in stromatolites. We look for (a) macromolecules produced exclusively by microbial life (e.g. hopanoids produced by cyanobacteria, for example). We could also use (b) isotopes as a measure to see if rocks have delta C that indicates isotopic fractionation of carbon due to biological activity.

What is the benefit of rooting the tree of life?

One of the big questions about the Tree of Life is figuring out which of the three major domains diverged first. This matters because when people look at groups of organisms to study parallel functions, ideally you want to use more closely related organisms. Both bacteria and archaea have many advantages for study as model organisms for similar processes in eukaryotes, but if we don't know which is more closely related to eukaryotes then we don't know which model is best. To figure this out, we need to root the tree of life (i.e. find an outgroup for all lineages of organisms on Earth, or an organism with a sequence from outside modern day organisms on Earth). This is because an unrooted tree does not indicate which of the sequences diverged earliest from the common ancestor they all shared.

Outline two lines of evidence that would support Panspermia theory.

Panspermia theory states the view that life arose on Mars first, then traveled from one planet to another. Evidence that would support Panspermia theory would have to come from what microbial life looks like on Mars and Earth. For example, if life on Mars showed a completely different biochemical or metabolic basis than that of Earth - for example, if it was based on silicon polymers instead of carbon - such a finding would not support the Panspermia theory. Instead, it would suggest that life originated independently on each planet, or that both planets were seeded from somewhere else. Biochemical/metabolic lines of evidence that would support Panspermia theory would be the presence of similar macromolecules in the biomass of the two microbes (i.e. cell membrane lipids like isoprenoid lipids or peptidoglycan). Genetic lines of evidence that would support Panspermia theory would be similar genetics, perhaps the two microbes even showing the same genetic code, this would serve as another line of evidence to support the view both planets were seeded from the same source.

How does Penicillin inhibit bacterial cell wall synthesis?

Penicillin is a beta-lactam antibiotic that binds to the transpeptidase enzymes that cross-link the peptides and inactivates them, so cross-linkage cannot happen. Beta-lactams all share a similar lactam ring, which look enough like one of the peptides on the NAM molecules in 3-d that the cross-linking transpeptidase enzymes try to cleave it. So the transpeptidases end up getting bound by the antibiotic and don't get used in making actual peptidoglycan.

Describe some of the major events in the history of microbiology that led to the research on microbiology being conducted today.

People learned that microorganisms could ferment foods in 10,000 B.C. and that swamps gave rise to "minute animals" that infected the air and caused human disease in 60 B.C. However, microorganisms were not observed until the 1670s. Antony von Luewenhoek was the first person to observe and accurately describe single-celled microorganisms with a microscope. He was proud and paranoid, refusing to share the design of his microscope with anyone. It took another 200 years for someone to develop a microscope with similar magnifying strength (~300x). As a consequence, there a lag in the development of microbiology as a field of research between the mid-1600s until the mid-1800s. The Golden Age of microbiology occurred from the 1850s through the 1940s, when better microscopes and culturing techniques emerged. These advances allowed scientists like Lister and Pasteur to develop the practice of antisepsis in hospitals, and the first vaccines. Pasteur also discovered the products produced by fermentation (e.g. alcohol). This continued through the 1940s with the production of antibiotics like penicillin. The Golden Age gave rise to advances in molecular biology (through the study of gene expression control by lactose in E. coli) and genetic engineering (i.e. the production of human insulin by E. coli).

What are the major metabolic/ecological factors that distinguish kingdoms of eukaryotic microbes?

Protozoa graze on other life forms. Particularly in soils and aquatic systems, protozoa can be important regulators of bacterial populations. Other protozoa are parasites and prey on much larger organisms (Trypanosoma and Giardia). Algae, by contrast, are photosynthesizers: they form the base of many aquatic food chains, particularly in marine ecosystems. Fungi are decomposers, acting as key recyclers of dead biomass, particularly of plant parts that bacteria can't initially breakdown.

Sanger sequencing

Sanger sequencing is a sequencing technology that sequences a single strand of DNA at a time. It relies on dideoxnucleotides, which can form a phosphodiester bond to a growing strand of DNA, but lack the 3' OH acceptor for a new incoming nucleotide. So, after the ddNTP is added, no new phosphodiester bond is formed to an incoming nucleotide (dNTP). Very small amounts of dideoxynucleotides are added to Sanger samples that are actively being synthesized. So, during the PCR reaction, every now and then, a ddNTP will be added to the growing DNA strand and terminate the reaction. Sanger sequencing can generate a ~ one thousand bases per run, but it takes a lot of time to run the reaction.

Compare and contrast Sanger sequencing, Illumina sequencing, and Nanopore sequencing. What are the pros and cons of each method? Pick a method that would you use to sequence the genome of a bacteria you isolate from the human gut and justify it.

Sanger sequencing is a sequencing technology that sequences a single strand of DNA at a time. It relies on dideoxnucleotides, which can form a phosphodiester bond to a growing strand of DNA, but lack the 3' OH acceptor for a new incoming nucleotide. So, after the ddNTP is added, no new phosphodiester bond is formed to an incoming nucleotide (dNTP). Very small amounts of dideoxynucleotides are added to Sanger samples that are actively being synthesized. So, during the PCR reaction, every now and then, a ddNTP will be added to the growing DNA strand and terminate the reaction. Sanger sequencing can generate a ~ one thousand bases per run, but it takes a lot of time to run the reaction. In Illumina sequencing, the genomic material to be sequenced is sonically fragmented into 100-300 bp segments and different linker oligonucleotides are ligated to each end. Strands of each fragment are separated and the mixture added and fixed to an optical flowcell. The flowcell has a dense lawn of oligonucelotides that are complementary to the ends of the adapters on each sequence. The fragments bend over and attach to the oligos, which serve as primers for amplification. Once amplification is complete, the double-stranded bridge is denatured and both strands remain attached to the surface. This repeats until 1000 identical copies of a gene fragment are generated = a cluster. Sequencing is then done in a series of single-step, reversible chain termination reactions. A fluorescently labeled nucleotide is added one at a time to the clusters. After a base is added, a snapshot is taken of the plates, illustrating the colors. Next, the fluor is removed, which reverses the chain termination, and the slide is flooded again with tagged bases. Illumina sequencing can sequence ~ 18 Gbp per run. However, it generates the shortest DNA reads (~ 300 bp) of all the sequencing technologies. Nanopore sequencing is not synthesis based, but is electric-current based. It is a single-molecule sequencing technology that relies on the changes in electric current that occur when a single strand of DNA passes through a nanopore structure. DNA is ligated to an adapter and the adapter is recognized by an enzyme that ratchets one of the strands through the nanopore. The sequencer cannot recognize an individual nucleotide, but it can recognize short strings of bases (3-6 bases long) because they have signature changes in electrical current that they produce as they pass through the nanopore. Nanopore can generate ~800 kbp read per strand, yet it has a large error rate (92% accuracy only), which means that the same DNA strand needs to be sequenced multiple times to identify (and correct) the errors.

What are the major morphological factors that distinguish kingdoms of eukaryotic microbes?

Single cells vs. multicellularity: Among the eukaryotic microbes, both protozoa and algae normally grow as single-celled organisms. On the other hand, most fungi can form large multicellular branches called hyphae. Hyphae are produced in the same way, with new cells dividing by binary fission, but instead of separating they stay together as part of the growing hyphal structure. A mass of hyphae is called mycelia. Some fungi grow as single cells - those are the yeasts and they reproduce by budding (which we'll discuss later in the course). Most fungi grow strictly as either mycelia or yeasts, but some can switch between the two and therefore called dimorphic. Fungi that cause serious human infections are usually dimorphic fungi, living in the mycelia form outside the body and switching the yeast form when they infect.

What is "taxa"?

Taxon = group of very closely related organisms Taxon = group of very closely related organisms (S. aureus vs. S. epidermis) Bacteria, archaea: group that has higher genomic DNA sequence similarity than an arbitrary cutoff (97%) •Usually morphologically and metabolically similar •Very closely related evolutionarily (phylotype) Operational taxonomic unit (OTU) = taxon = phylotype (prokaryotes) ~species (fungi)

Compare and contrast taxonomic diversity measures and phylogenetic diversity measures. What is the benefit of phylogenetic diversity over taxonomic diversity estimates?

Taxonomy-based metrics of diversity (i.e. names of organisms) are useful because distinct species or taxa of microbes may be metabolically distinct as well. However, taxonomy-based metrics of diversity assume that there is no different evolutionary relationships between species - i.e., all species are equally related. This may not be an appropriate assumption, because 1) we know that species are evolutionarily related and 2) closely related species may be more similar in their phenotypes than distantly related species. If this is the case, then taxonomic diversity will mean something very different than phylogenetic diversity for the diversity of microbial phenotypes in a community. A sample that has high taxonomic diversity could have very low phenotypic diversity, if all taxa/species are closely related. By contrast, a sample that has high phylogenetic diversity is likely to have high phenotypic diversity, because it is likely that species that are distantly related have very different traits.

What are the specific terms used to identify taxa using DNA sequences, and what are the differences between these terms?

Terms include: a. Phylotype: a group of organisms classified by overall similarity in phylogenetic relatedness. A single phylotype is defined by DNA sequences of the organisms sharing more than an arbitrarily chosen level of similarity (e.g. 97% similarity) of a particular gene marker (e.g. the 16S rDNA). Phylotypes are often used to describe prokaryotes since prokaryotes are difficult to classify by mating experiments (i.e. they reproduce asexually and can also exchange DNA with distantly related organisms). b. Operational taxonomic unit (OTU): a group of organisms that have high sequence similarity of their ribosomal DNA sequences (e.g. 16S rDNA). Very similar (often the same as) phylotype, although this term is typically used to refer to prokaryotes (rather than fungi). For fungi, OTUs are a proxy for species. c. Taxon: a unique microorganism. This can be a single microbial species (if there are good species concepts for that type of microorganism, like the fungi). When we don't know what constitutes a "species" of microbe (e.g. many bacterial lineages), the term taxon can refer to an OTU or phylotype.

What are some differences between viruses and other microbes?

The major difference between viruses and other microbes is that viruses are more simple in nearly every way. They are much smaller than bacteria and archaea (to image them, it is necessary to use an electron microscope, which magnifies samples 1000X or more) and they are symbiotic with live organisms (such that they don't need the machinery to conduct independent metabolism). Viruses basically consist of a genome and set of closely associated proteins that stabilize the genome and perform essential functions during replication. Both of those parts are wrapped in a capsid that protects the genome during the times the virus is outside the cell it invades. Some viruses have an additional coating called an envelope, consisting of a protein-coated phosholipid bilayer.

What is the major factor that allows us to distinguish prokaryotes and eukaryotes? What are some of the minor factors?

The major factor that distinguishes prokaryotes and eukaryotes is: 1) Presence of a nuclear membrane*: prokaryotes lack a nuclear membrane that encloses the organism's genome, whereas eukaryotes do possess a nuclear membrane *This is the major factor that distinguishes prokaryotes and eukaryotes. Other more minor factors are listed below: 2) Morphological diversity: eukaryotes have generally higher morphological diversity (shapes and sizes) than prokaryotes 3) Average cell size: prokaryotes are generally 10x smaller in diameter and 1000x smaller in volume than eukaryotes 4) Prokaryotes have a thick outer envelope that makes them physically/chemically resistant to environmental stress 5) The diversity of prokaryotes lies in their metabolisms or ways of getting energy: prokaryotes guys produce and consume compounds like methane, nitrogen gas, and sulfides that are neither produced or utilized by plant or animal cells. 6) Few chromosomes: eukaryotes generally have high numbers of chromosomes, while prokaryotes generally have only 1 or 2 7) Reproduction by binary fission: prokaryotes reproduce by binary fission, as opposed to mitosis like most eukaryotes (although some fungi can reproduce by binary fission)

What are the major morphological groups of microbes?

The major morphological groups of microbes include the prokaryotes, eukaryotes, and viruses.

What are the major taxonomic groups of microbes?

The major taxonomic groups of microbes include the bacteria, archaea, fungi, protists, algae, and viruses.

What is a plasmid and how is it unique from bacterial chromosomes? Describe an example of a plasmid that is beneficial to bacteria and one that is detrimental to bacteria.

The primary importance of plasmids is that they are one of the main ways bacteria share genes. For example, many of the genes involved in nitrogen fixation (N2 à NH4+) for the bacteria associated with legume plants are plasmid born. Plasmids are also a major way that disease-causing bacteria become resistant to an antibiotic. Rather than mutate, they can pick the up the genes needed to resist antibiotics. Plasmids are also the way that good bacteria can go bad, by picking up virulence genes that allow them to do damage, such as genes encoding toxins that aid in pathogenesis. Plasmid addiction systems, on the other hand, are designed to kill cells that lose the plasmid. These types of plasmids produce a toxin protein and also produce an antidote protein. They ensure their survival because if one of the daughter cells in division doesn't receive the plasmid, it still gets some of the toxin and the antidote in the cytoplasm. The catch is that the antidote protein has a shorter life than the toxic protein, so the absence of the plasmid makes it so the antidote is not being produced. This then kills the daughter cell if the plasmid making the antidote is not in it.

Compare and contrast the conditions of early Earth, under which microbes evolved, and the conditions on Earth today.

The relatively stable environmental conditions that we experience today are by no means typical of Earth's past, which has been marked by: (a) meteorite bombardment. Early Earth's history was punctuated by a succession of catastrophic impacts of large chunks of space rock. The first period of Earth's existence, ranging from 4.5 to 3.8 Gyr ago, is designated the Hadean eon. It is thought that, during this time, Earth actually sustained more impacts than the moon. Some of the impacts were so big and powerful that they could vaporize oceans, creating clouds of steam that would have sterilized the Earth's surface. (b) very cold air, which may have been the dominant environmental condition before we gained an atmosphere to trap heat. While this would have been bad for organisms living on the surface, life could have survived deep underground, perhaps as spores. (c) periods of heavy volcanic activity (d) Instability: periods when entire oceans went anoxic. (e) strong UV radiation, because there was no ozone layer like we have today to filter out UV radiation.

What is Microbiology?

The study of biological entities too small (typically) to be clearly seen by unaided eye (i.e. microorganisms).

If you found an old microbial specimen in a museum that was characterized as a chemolithoautotroph that lived on rusty water pipes underground, what sorts of materials would it likely use as a carbon source, energy source, and electron source?

This organism would likely use iron as an electron source (i.e. be iron-oxidizing), maybe other inorganic molecules found in water, like sulfate or nitrate, as well. Changes in the oxidation state of iron (i.e. redox chemistry) would be used as an energy source, and CO2 would be used as a carbon source.

Transformation

Transformation is the uptake of free DNA from the environment. There are two types of transformation: natural transformation and artificial transformation. In natural transformation, DNA from the environment first attaches to the surface of the bacterial cell. Then, one of the strands is nicked and digested away by bacterial enzymes to make the DNA single-stranded. The single-stranded form is taken up by an active transport system (meaning it takes energy) and enters the cytoplasm. In the cytoplasm, if the incoming DNA has regions of sequence identical with the chromosome of the recipient, the incoming DNA can enter the chromosome by homologous recombination. most bacteria associated with humans, such as E. coli, are not naturally transformable, it is necessary to use artificial conditions to force them to take up DNA. The heat or chemical or electric shock basically allow us to bypass the need for specialized protein complexes that introduce DNA into the cell. As a result, the DNA that enters the cell is double stranded. That DNA is then replicated either by insertion via homologous recombination or if a plasmid has been introduced, it replicates autonomously. Unlike human and animal microbes, soil and water microbes often have natural transformation systems. They can account for as much as 5% of the cultivable bacterial population. The reason that soil and water microbes have these natural transformation systems is not totally clear, but one explanation has to do with nutrient availabilities in the different environments. DNA released from dying microbes may be an important food source for many soil bacteria.

Describe transformation, transduction, and conjugation. What are the differences between these three methods of HGT?

Transformation is the uptake of free DNA from the environment. There are two types of transformation: natural transformation and artificial transformation. In natural transformation, DNA from the environment first attaches to the surface of the bacterial cell. Then, one of the strands is nicked and digested away by bacterial enzymes to make the DNA single-stranded. The single-stranded form is taken up by an active transport system (meaning it takes energy) and enters the cytoplasm. In the cytoplasm, if the incoming DNA has regions of sequence identical with the chromosome of the recipient, the incoming DNA can enter the chromosome by homologous recombination. most bacteria associated with humans, such as E. coli, are not naturally transformable, it is necessary to use artificial conditions to force them to take up DNA. The heat or chemical or electric shock basically allow us to bypass the need for specialized protein complexes that introduce DNA into the cell. As a result, the DNA that enters the cell is double stranded. That DNA is then replicated either by insertion via homologous recombination or if a plasmid has been introduced, it replicates autonomously. Unlike human and animal microbes, soil and water microbes often have natural transformation systems. They can account for as much as 5% of the cultivable bacterial population. The reason that soil and water microbes have these natural transformation systems is not totally clear, but one explanation has to do with nutrient availabilities in the different environments. DNA released from dying microbes may be an important food source for many soil bacteria. Transduction is DNA transfer mediated by viruses that infect bacteria (i.e. bacteriophages). Conjugation is direct cell-to-cell transfer of DNA. When a bacteriophage attacks a bacterial cell, it first attaches to a receptor on the bacterial surface. The surface attachment is taxon (or species)-specific. Phage attachment proteins are specific to protein receptors on the surface of its host cell. Then it injects its DNA in its capsid into the bacterial cell. Once the DNA of the phage is injected, it starts to reproduce itself. This process is called a lytic infection, because the cell that hosts the phage eventually lyses. During this process, parts of the DNA from the host can be inadvertently picked up and moved. This happens because a phage that is replicating chops up the chromosome of the bacterium it infects. In the frenzy to assemble new phage particles, a piece of bacterial chromosomal DNA instead of the phage's own genome may be packaged into a phage head. The phage particle carrying bacterial DNA can later attach to another bacterium and inject the bacterial DNA into it. Conjugation is the term used to describe direct cell-to-cell transfer of DNA. During conjugation, two bacteria (a donor and a recipient) make a tight connection with each other. This contact is mediated by a specialized pilus (sex pilus), which attaches to another (recipient) cell and draws the two cells together. On the plasmid that is transferred, there are genes coding for the sex pilus, a channel between the donorand recipient cell. (i.e. mating bridge), and DNA transfer proteins. The plasmid uses one of its proteins called TraI (transferI) in a protein complex called relaxosome to nick the plasmid at its transfer origin (oriT). is followed by attachment of the single strand to the mating bridge, which then transfers the single strand into the cytoplasm of the recipient. DNA polymerase makes both the strand left in the donor and the strand introduced into the recipient double-stranded. So both donor and recipient end up with a double-stranded copy of the plasmid. Although bacteria can acquire new genes by either transduction or transformation, these types of transfer tend to occur mainly between members of the same species or between members of very closely related species. For transduction, the phage will only be able to affect cells that it can stick to. The receptors on individuals of the same species or closely related species are similar enough that the phage can stick to them, but for very unrelated species the surface receptors are not similar, so the phage won't be able to bind to it. For both of these mechanisms of gene transfer they involve the introduction of linear DNA. For that DNA to survive in the cell, it needs to get incorporated into the chromosome, otherwise it will get chopped up by restriction enzymes and nucleases. The integration of DNA into the chromosome is typically done by homologous recombination. For HR to work, the regions have to share enough affinity that the enzymes involved in HR bring the strands into close proximity. These will generally happen in comparisons among the same species or closely related species, but not among ones distantly related. Conjugation, which is the third way HGT happens, does not have these constraints because what is transferred are typically plasmids or circular DNA elements that can survive in the new cell outside of the chromosome.

Dating early life on the planet Earth often takes multiple lines of evidence to distinguish biotic from abiotic processes. Imagine you find a stromatolite fossil off the coast of Mexico that dates to 3.2 billion years before present. To bolster your argument the fossil was not generated by an abiotic process, describe two other lines of evidence that could be assessed to confirm the stromatolite fossil was in fact once a living organism. In your answer, explain exactly what kind of data from each line of evidence would support your argument.

Two lines of evidence besides the presence of fossils themselves would be biosignature molecules and isotope ratios. In the case of cyanobacteria, a unique biosignature molecules would be a hopanoid, which is a lipid that is part of their cell membrane. Therefore, the presence of hopanoids in the stromatolite fossil would be suggestive that it was not simply the product of abiotic (non-biological) processes. The presence of isotope fractionation in the stromatolite would be further evidence of the fossil resulting from a biotic process. In the case of isotope fractionation, it is known that photosynthetic cyanobacteria discriminate against 13C during photosynthesis, so their resulting tissue (which becomes fossilized in stromatolites) is significantly depleted in 13C relative to materials generated through abiotic processes. Therefore, a highly negative delta 13C signature would be consistent with a cellular process, like photosynthesis in the cyanobacteria.

Early Earth (4-1 GYA): Not Really "Eden"-Challenging Conditions

a)Constant bombardment by large chunks of space rock •Hadean eon (4.5-3.8 GYA): more impacts than the moon • •Some impacts were powerful enough to vaporize oceans, sterilizing Earth's surface. b)Environment was cold! c)Much greater volcanic activity than today. d)Unstable: periods when entire ocean went anoxic e)No ozone layer, so the earth's surface was bathed in strong UV radiation. •Microbes lived underground

Which structural features of bacterial cells allow the Gram stain to differentiate bacteria? Make sure to discuss the differences in cell wall between Gram positive and Gram negative bacteria.

What is happening in the Gram stain is you are adding a dye to visualize the bacteria. In this case the dye is called crystal violet, which is a big purple molecule. After the dye is added, iodine is added next which binds with the crystal violet into a complex that is harder to remove from the cell. The cells are then washed and, depending on the amount of cell wall, either keep the dye or lose it. For the ones that have lost it, to see them, we need to use another stain, in this case, safranin, so in the end we have two different colored cells. Gram positive are dark purple and Gram negative are pink. Gram positive retain the crystal violet stain because they have more peptidoglycan that can trap the large crystal violet stain complex. The way this works mechanistically is that crystal violet dissociates into CV+ and Cl- ions. The CV+ ion associates with Iodide (I-), creating a large complex (CV-I) that gets stuck within the multiple peptidoglycan layers of Gram positive cell walls. Because there are fewer layers of peptidoglycan in Gram negative cells, it is released following the washing.

Early phylogenies used morphology + nutrition

Whittaker and Margulis' early five-kingdom system • Three modes of nutrition •Distinction between unicellular and multicellular body plans. Problem #3: no one common yardstick against which all organisms can be measured.

Discuss three ways in which bacterial surface polysaccharides (i.e. a capsule) might benefit the growth of a pathogenic bacterium inside human tissues.

a. One of the most recognized benefits to surface polysaccharides is physical protection from phagocytic cells (phagocytes, like your white blood cells). They do this by basically helping disguise the bacteria so they are not ingested. The capsules are uniform and slippery, making it difficult for phagocytes to lock onto the bacterial cell b. Capsules contain a great deal of water and can protect against desiccation. c. They exclude viruses and hydrophobic toxic materials such as detergents. d. They can also aid in attachment to solid surfaces - biofilms.

studying microbiology applied aspects:

are concerned with practical problems-disease, water, food and industrial microbiology

Major Taxonomic Groups of microbes

bacteria (prokaryotes), archaea (prokaryotes), eukarya (algae and plants, fungi and animals, protists)

Studying Microbiology basic aspects:

concerned with individual groups of microbes: taxonomy, physiology, genetics, and ecology

Surface Polysaccharides (Survival + Disease)

•Capsules: thin covering layer of poly-saccharides (rarely proteins). •Gives colonies a shiny and wet appearance. •One role: protection from human phagocytic cells. -Protect against desiccation -Exclude viruses and hydrophobic toxic materials such as detergents -Aid in attachment to solid surfaces - biofilms

(2a) Biomolecules & (2b) Metal Oxidation States

•Certain molecules are unique product of biological activity. -2-Methylhopane (hopanoid): 2.5 Gyr -Banded Iron Formations •chemolithotrophs using Fe2+ as e- donor and O as e- acceptor. •photolithotrophs using using Fe2+ as e- donor •Cycles of SiO2 (gray) & Fe2O3 (red)

Traditional Measures of Diversity

•Defining organismal diversity was long based on morphology. -Emphasis on physical traits such as appendages, shape, and color. •Problem #1: vastly different kinds of microbes looked more or less alike. •As a result, microbial diversity was for years thought to relatively low. •Problem #2: Microbes do not readily fit the classic definition of a species

Over the Past 1Billion Years

•Environmental conditions have, generally speaking, been relatively stable. -Temperature: warmer -Meteorites: fewer -Volcanism: less -Oceans: less fluctuation •"benign" conditions + GOE + endosymbiosis helped promote the immense diversity of life currently present on Earth.

Biosignature (1a) Microfossils: Mixed Evidence

•Fossils of microbes can be formed when cells die and their form is filled by minerals. •Many have very similar form to modern microbes. •Earliest dated to ~2 Gyr. •Martin Brasier: similar forms are created by abiotic processes

The Oxygen Revolution (GOE)

•Geological evidence shows O2 levels in oceans rose around 2 Gyr (Fe2O3) •Resulted in a number of major changes to life on Earth. 1.Greatly expanded cycling of major elements. 2.Resulted in the formation of an ozone layer. 3.Endosymbiosis and evolution of "higher" life.

The Oxygen Revolution (GOE)

•Geological evidence shows O2 levels in oceans rose around 2 Gyr (Fe2O3) •Resulted in a number of major changes to life on Earth. 1.Greatly expanded cycling of major elements: S: H2S -> SO42- N: NH4+ -> NO3-

Modern Measure of Diversity

•In the later 1970s, Carl Woese was the first to propose using DNA-based data. •All living organisms share a common set of genes - "housekeeping genes" •rRNA genes encode the scaffolding in ribosomes - all living organisms have •Other genes would have worked, but 16s rRNA gene was (1) high copy # and (2) right size

The Panspermia Theory

•Life on planet Earth came from elsewhere. •Rationale based on some planets cooling faster than others. •Mars, being smaller and farther from the sun, cooled more quickly and used to have water. -Collisions of space rocks with Mars'surface could have knocked fragments towards Earth.

Differentiating Bacteria & Archaea: morphology

•Major difference in cell wall biopolymer - Bacteria have peptidoglycan •Major difference in cell membrane lipids

Viruses: More Simple in (Nearly) Every Way

•Much smaller than bacteria and archaea, can't be seen with ordinary light microscope. •Unique genomes •Genomes not necessarily composed of DNA, quite a few use RNA. •Genomes not always double-stranded or circular (like many free-living prokaryotes). •Not free-living: mutualistic symbionts or pathogens •Cause many human diseases including AIDS, hepatitis, polio, and influenza.

Evidence for Early Life on Earth

•Often hard to interpret evidence because biological and non-biological processes can create the same products. -E.g. Metals can be oxidized by microbial activity but also by UV light. •Need to use multiple lines of evidence and see where they overlap. •Biosignatures: (1) cell-like formations, (2) chemical signatures

Morphology of Eukaryotic Microbes:Protists, Algae, and Fungi

•Protozoa and algae normally grow as single cells. •Fungi can be single cells or multiple cells •Filamentous fungi = create large multi-cellular branches (hyphae/mycelium) •Yeasts = single cells that reproduce by budding (daughter buds off mom). •Dimorphic fungi: can grow as yeasts and hyphae •More impressive diversity of shapes than prokaryotes

Ecological Roles of Eukaryotic Microbes

•Protozoa are grazers - important regulators of prokaryotic populations (phagocytosis). •Algae are photosynthesizers- form food chain bases in both aquatic and marine systems (w/CO2+light). •Fungi are decomposers - recycle nutrients (C, N, P, Mg, Ca) from dead biomass •Release enzymes outside of the cell (extracellular enzymes) that break down large plant biopolymers (lignin and cellulose) •All three groups contain members that are mutualistic symbionts (+) or pathogens (-)

(1b) Earliest Confirmed Fossils: Stromatolites

•Stromatolites are large fossilized mounds of cyanobacteria. •Occur in the fossil record starting about 3 Gyr. •Living fossils present in both freshwater and marine systems. •Played a key role in the generation of an oxygenic atmosphere.

Differentiating Bacteria & Archaea: metabolism

•Temperature growth limit much higher in archaea (120°C vs. 90°C). •Degradation of complex carbon polymers: only bacteria (human gut) •Pathogens: only bacteria (no archaea are known to cause disease) -Eckerg et al. (2006) Infection and Immunity. 71: 591-596. •Both are mutualistic symbionts of many macrobes, including humans.

"Golden Age" of Microbiology

•The 1850s through the 1940s (•Antisepsis/disinfection (Joseph Lister) •Fermentation (Louis Pasteur) •Vaccines (Pasteur) and antibiotics) After that: A New Era of Microbiology! •Prevention/cure of infectious diseases •Control of gene expression - E. coli •Genetic engineering - Insulin

Major features of microorganisms

•Usually are smaller than can be seen clearly with naked eye •Are relatively simple in construction -RAPID EVOLUTION (SA->MRSA 1yr, MRSA->VRSA several months, 35 mutations) -metabolically diverse -taxonomically diverse