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Microbiology Study Guide from Lectures (summary), Study Guides, Projects, Research of Microbiology

Rijksuniversiteit Groningen Microbiology

Microbiology Study Guide from Lectures (summary) BME year 1 period 2b

Typology: Study Guides, Projects, Research

2021/2022

Available from 08/15/2022

elena.morandini 🇳🇱

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Introduction to Microbes

Chapter 1 - Microbial Life: Origin and Discovery

Microbe = single celled organism derived from single ancestor

- Prokaryotes → bacteria / archaea

- Eukaryotes → algae, fungi, protozoa

- Biofilms and viruses (not living but classified as)

- Range from millimeters to 0.2 micrometers

Historical events → big time gap between first discoveries and next events

- Microscopic world first observed in 1665 by Robert Hooke

- Antonie van Leeuwenhoek observes bacteria with a single lens in 1676

- Semmelweis observes CL reduces pathogens on doctors’ hands in 1847

- Microbes defined in a different class than plants and animals (1866 - Haeckel)

- First antibiotic made by fungus discovered by Flemming in 1929

- Biofilms major form of existence of microbes in 1978 by Costerton

Influenzae → first sequence genome, circular bacterial DNA

Robert Koch → 4 steps to link infection and disease

- Microbe could be inactive in healthy individual, is isolated and grown in pre culture, then

introduced in a healthy susceptible host to see if same disease is shown

- Same strain of microbe obtained by new host

Pasteur → immunization and smallpox virus

- vaccine injects damaged or dead virus into the host → immune system recognizes it and

creates antibodies

- For covid → mRNA vaccine skips recognition, helps the host product the protective proteins

Vaccines / Antiseptics and Antibiotics →

- Vaccines → utilize the virus to produce antibodies, not effective against bacteria

- Antiseptics → can be physical (include heat - bunsen burner or autoclave) or chemical

(molecules that kill bacteria and the host - bleach)

- Antibiotics → kill > 99.9% of microbes but leave host unharmed

Support Ecosystem → wide range for bacteria

- Aerobic environment (a lot of O2) / Anaerobic environment (lack of O2)

- Microaerophilic environment (reduced O2, many forms of metabolism)

- Capnophilic environment (CO2loving)

- Strict anaerobic environment (void of oxygen, lethal)

- In biofilm → more environments can exist simultaneously

Nitrogen Cycle → stages: fixation, mineralization, nitrification, immobilization, and denitrification

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Introduction to Microbes Chapter 1 - Microbial Life: Origin and Discovery Microbe = single celled organism derived from single ancestor

Prokaryotes → bacteria / archaea
Eukaryotes → algae, fungi, protozoa
Biofilms and viruses (not living but classified as)
Range from millimeters to 0.2 micrometers Historical events → big time gap between first discoveries and next events
Microscopic world first observed in 1665 by Robert Hooke
Antonie van Leeuwenhoek observes bacteria with a single lens in 1676
Semmelweis observes CL reduces pathogens on doctors’ hands in 1847
Microbes defined in a different class than plants and animals (1866 - Haeckel)
First antibiotic made by fungus discovered by Flemming in 1929
Biofilms major form of existence of microbes in 1978 by Costerton Influenzae → first sequence genome, circular bacterial DNA Robert Koch → 4 steps to link infection and disease
Microbe could be inactive in healthy individual, is isolated and grown in pre culture, then introduced in a healthy susceptible host to see if same disease is shown
Same strain of microbe obtained by new host Pasteur → immunization and smallpox virus
vaccine injects damaged or dead virus into the host → immune system recognizes it and creates antibodies
For covid → mRNA vaccine skips recognition, helps the host product the protective proteins Vaccines / Antiseptics and Antibiotics →
Vaccines → utilize the virus to produce antibodies, not effective against bacteria
Antiseptics → can be physical (include heat - bunsen burner or autoclave) or chemical (molecules that kill bacteria and the host - bleach)
Antibiotics → kill > 99.9% of microbes but leave host unharmed Support Ecosystem → wide range for bacteria
Aerobic environment (a lot of O 2 ) / Anaerobic environment (lack of O 2 )
Microaerophilic environment (reduced O 2 , many forms of metabolism)
Capnophilic environment (CO 2 loving)
Strict anaerobic environment (void of oxygen, lethal)
In biofilm → more environments can exist simultaneously Nitrogen Cycle → stages: fixation, mineralization, nitrification, immobilization, and denitrification

Serial endosymbiosis Theory → how eukaryotes form

Differentiation between plant and animal cells was based on what was ingested (mit/chl)
formation of mit and chl 5 kingdom scheme → prokaryotes and bacteria are isolated in their own group Tree of life Kingdom Scheme (Carl Woese)
Eukaryotes = multi celled, Bacteria (common) = single celled (proteo = mit / cyano = chloro)
Archaea (rare) = thermophiles, sulfur oxidizers (not susceptible to antibiotics) Chapter 2: the Microbial Cell Gram staining technique → gram positive (thick peptidoglycan layer, purple) and gram negative bacteria (thinner layer, red or pink)
Procedure: stain with violet die, iodine, alcohol, safranin (for 1 minute / 30 seconds) Bacteria categorization →

All antibiotics require active metabolism (energy) → bacteria can slow down metabolism if there are not enough nutrients Bacterial Cell division →
Bacteria use energy through mitosis
DNA origin replicates and migrates → replication is bidirectional, septum forms and division Specialized Structures →
Adhesion structures = pili and stalks
Movement structures = rotary flagella (help propel) Chapter 4: Bacterial Culture, Growth and Development Bacteria → purpose = survive and grow
No nutrients → survival mode with shut down metabolism (low energy state)
Abundant nutrients → replication mode Heterotrophy (humans) → use nutrients to produce CO 2 / Autotrophy (plants) → use CO 2 to produce nutrients Life Cycle →
Log phase = exponential
Stationary → bacteria = nutrients
Death → bacteria > nutrients, metabolism slows down Bacteria → grown in culture medium
Pure culture = suspension derived from a single bacterial colony in liquid medium
After growth → cultures are placed on agar plate to confirm purity of culture (dilution streaking → apply culture on edge of plate, flame and streak → isolation) Choosing Medium →
Complex → provides what bacteria need to produce (depending on # of bacteria in sample)
Selective → limits growth of bacteria not wanted (can contain antibiotics, will kill gram - )
Differential → provides cues that differ between colonies for identification Quantifying Bacteria →
Microscope → live + dead bacteria per unit area
Dilution plating → viable colonies from single bacteria (CFU / unit area)
Optical density → used with standard curve of suspension (McFarland standards)
Calibration curve → every bacteria has different size

Biochemical Assay → translate DNA, ATP to counts Biofilms → community of bacteria in extracellular polymers 6 stages of growth → conditioning of film, reversible attachment of cells, colonizers become irreversibly attached, growth and cell division, formation of water channels, attachment of colonizers and dispersion of microbes to new sites Chapter 5: Environmental Influences and Control of Microbial Growth Conditions and Classification of Microbes →

Controlling Growth Rates → optimum growth curve (21-37°)

Higher temp, lower growth rate (enzymes denature)
Lower temp, lower growth rate (decrease in membrane fluidity and enzymatic activity)

Physical, Chemical and Biological Control → Sterilization → all living cells, spores and viruses are destroyed Disinfection → killing or removal of disease producing organisms from inanimate surfaces Antisepsis → killing or removal of pathogens from living tissues Sanitation → reducing the microbial population to safe levels (clean è disinfect) Bacteriostatic → inhibits growth (number of viable cells remains constant) Bactericidal → kills (number of viable cells reduced) Germicidal → kills pathogens but not spores Physical Agents that kill microbes →

Biofilms Biofilms in Medicine Biofilm → group of microorganisms, cells embedded in extracellular polymeric substance (EPS, made of water, polysaccharides, proteins and extracellular DNA)

Hospital setting → contact lenses keratitis, infection of implants or burns
Survival mechanism for bacteria forming communities in a gel like coating (survival method for bacteria → form community) History →
Van Leeuwenhoek (1676): discovered bacteria
Costerton (1978): biofilms first described (3 centuries later)
Estimated that biofilms make part of >60% human infection How do Biofilms Work → 6 stages

Formation of conditioning film (Pre stage, forms favorable condition for bacteria to adhere)

Enamel surface covered by salivary conditioning film

Initial (reversible) adhesion

Maintains biofilm on a surface → first cells adhere to the surface through reversible (van der Waals and electrostatic - smaller = both interactions)
Cells can anchor themselves more permanently with cell adhesion

Anchoring (irreversible) adhesion (Specific and non specific adhesion are non interchangeable)

Single layer, permanent attachment → create small cluster

EPS production (Happens together with 3)
Growth and maturation

Biofilm growth → cell division + recruitment → biofilm established
If environment is unfavorable → bacteria sacrifice themselves for protection
Maturation → cells = 15% of volume, water channels deliver nutrients

Cell release or detachment

Release of free floating microorganisms → maintain properties for short time
Will start looking for new locations, leave behind a auto maintained biofilm
After being removed → back to planktonic form, free floating Characteristics →
Highly dynamic structure → may detach in clumps or alone, can deform or revert back to position

Detachment → enables biofilm to spread and colonize new surfaces (overpopulation)
Different genetic expression in biofilm bacteria and planktonic bacteria (phenotype shift in behavior, genes regulated differently)
Behavior coordinated via intercellular communication with signaling molecules (talk and listen signals → cell cell communication / quorum sensing) - Quorum sensing → density dependent → biofilms held together in dense population, secreted molecules reach higher concentrations → signaling molecules can trigger changes in genetic activity but not expression (genotype) - Causes bacteria to act in groups → community with open water channels
Bacteria in biofilm are less susceptible to antimicrobial agents → higher resistance, but sensitive to them in planktonic conditions (different from antibiotic resistance)
Bacteria less susceptible to antimicrobial resistance → no penetration of antimicrobial in biofilm, altered microenvironments (nongrowing bacteria) Metabolic Activity → further away from exchange = lower activity
Inside → heterogeneous environment, best chance for bacteria to survive and reproduce → needs to be attacked to kill biofilm
Reduced activity = less susceptibility to growth dependent antibiotic killing Persister cells → resistant variants → survive, cause infection to relapse
Antibiotic treatment kills planktonic cells and biofilm cells Host immune system → biofilm protects cells against host immune responses Invasion by Leukocytes → invade biofilms through water channels, inactive in killing cells Efficient Genetic exchange → extracellular DNA component of EPS → released from bacteria → gives ideal conditions for exchange of genetic material, multidrug resistance is promoted Controlling Biofilms on Implants Biofilms → responsible for implant related infections (76%)
All materials react to different types of bacteria differently
Metals (titanium, steel), polymers (silicone, PE, PMMA), ceramics
Ex. tooth implant → cuts is where bacteria can invade (not all internal) vs hip implant (total tissue integration)

positive cocci, haemophilus influenzae, E.coli and gram negative bacteria Infections tissue related → Infections without implant → ex. Endocarditis (inner surface of heart) or respiratory infection

Bacterial Genetics Chapter 7: Genomes and Chromosomes Bacterial Genomes → genome = all genetic information, for bacteria and archaea = chromosomes and plasmids

Range in size from 106 kbp to 16000 kbp (eukaryotic = 2900-100,000,000, human = over 3,000,000)
Chromosomes → circular or linear, carry hereditary information
Plasmids → circular, small extrachromosomal DNA molecules that replicate independently, easily transferred, smaller
Not a lot of wanted space in DNA (human genome is 98% noncoding) Replication → picture Nucleotide → different from nucleoside (sugar + base), nucleotide also has phosphate group
ATP = specific nucleotide with 3 phosphates
Nitrogen base → hexagon (pyrimidine → cytosine, uracil, thymine) / hexagon + pentagon (purine → larger guanine, adenine)
Pairs = UA / TA / CG (pyr + pur) RNA → single stranded, forms loops to have interacting complementary bases
At high temperature H bonds that connect base pairs break → separate strands (method: polymerase chain reaction PCR) DNA vs RNA →
Different nucleotide (T/U), RNA single stranded
Two different missions → DNA archives information, long term stability and replication, makes temporary copies of genes to make proteins / RNA modulates DNA expression, 2’ position in 5 carbon sugar (OH in ribose, H in deoxyribose → structural difference), lack of oxygen (more flexibility), prevents enzymes from acting on RNA and DNA DNA → in bacteria → supercoiled, topoisomerases type I relieves supercoiling, type II induces it) Replication → semi conservative (one parent strain is conserved, compared for accuracy)

Protein synthesis → 3 stages

Initiation of translation → mRNA gains access to subunit as ribosome moves along, tRNA gains access to 3 binding sites (A acceptor site where anticodon binds to codon in mRNA, P peptidyl site which binds tRNA to growing peptide, E exit site where tRNA exits after giving up the amino acid)
Stop codon reaches A ride → remaining tRNA ejected
Bacteria have no organelles → coupled transcription and translation in ribosomes → efficient, making protein before mRNA is finished Folding and Degradation → peptide folds in lowest energy state using bonds and complementary groups (to stabilize)
3d conformation = specific → enzymes target sites and shapes
Degradation → occurs naturally when protein is no longer needed (or induced by antibiotics) Chapter 9: Genetic Change and Genome Evolution Types of Mutations → codes Point mutation Change in single nucleotide (transition = swap similar base ring / transversion = switch purine and pyrimidine - no more H bond) Insertion / deletion Addition or subtraction, changes length Inversion DNA fragment flipped (5-3/3-5) Duplication Second copy of DNA fragment Transposition Movement of a fragment Reversion Restores mutated sequence to original (proofreading) Types of Mutations → effect Silent Does not change sequence Missense Changes sequence of protein Loss of function Missense that decreases or eliminates function Gain of function Missense that enhances function Knockout Eliminates function, can include Nonsense Changes amino acid codon

multiple insertions, deletions, nonsense mutations into a termination codon Bacteria and Mutations →

Exponential phase of growth → bacteria produce many generations quickly (increases change of random mutation)
Environmental stress (ex. antibiotic) promotes survival → those that survive will reproduce and change characteristics of the population (generate resistance) DNA repair → error proof and error prone pathways
Methyl mismatch repair, photoreactivation, base excision repair, recombinational repair, SOS response, nonhomologous end joining Gene Transfer → DNA function in bacteria = food, repair, evolution
Mechanisms of gene exchange = transformation, transduction and conjugation
Horizontal gene transfer → transfer info from donor to recipient of same generation
Vertical gene transfer → genetic tree, parent to daughter cell Chapter 10: Molecular Regulation Transcriptional Repressors and Activators →
Control → use regulatory proteins to stimulate or prevent the binding of RNA polymerase to the promoter, sense the external environment (sensor kinase and response regulator) Alternative Sigma Factors or Anti Sigma Factors → ex. Regulon (collection of coregulated operons) → controlled by activators and repressors but also sigma factors → help located promoted sequence RNA regulation →
Attenuation → transcriptional regulatory mechanism, translation affects mRNA structure
Riboswitches → secondary structure at 5’ end of mRNA that obscure access to ribosomes
Regulatory RNA → regulate transcription and stability of mRNA
Cis-antisense RNA → affect expression of single gene Quorum sensing → on when signaling molecule concentration is above threshold, requires signal to be sensed by autoinducers to trigger a response

Techniques → cloning by PCR, random mutagenesis, site directed mutagenesis Gene Expression Analysis →
RNA and transcription / Gene cloning → genetic code can be inserted or removed from bacterial species to test the effect
Help mass production → vaccine antigens produced in plants → sequence sticked into bacteria and mass produce the vaccine Synthetic Biology → engineering a life form from scratch
Like an engineering circuit (NOR → neither on to have expression, AND → both have to be on to have expression)
IMPLY → best one, more options to express gene Switches → toggle = on/ off position, oscillator = production only when concentration is low
system noise = unexpected factors not controlled
Riboswitches and switchboards = sense metabolites to control gene expression
Kill switches = suicide gene (prevent out of control sprede), permanent off Chapter 13: Energetics and Catabolism Sources of energy →
Chemotrophs → use of chemicals for electron transfer (or organic chemicals)
Heterotrophs → use preformed organic compounds for biosynthesis
Autotrophy → CO 2 fixed and assembled in organic molecules
Phototrophs → yield energy from light absorption Enthalpy, Entropy, Gibbs Free Energy →
Entropy (S) = amount of disorder (release heat = increase)
Enthalpy (H) = amount of heat energy
Gibbs (G) = direction of thermodynamic equation (negative is favorable)
∆𝐺= ∆𝐻−𝑇∆𝑆
Negative ∆𝐺 drives reaction forward
H → + = endothermic, - = exothermic, molecular stability increases
S → increases when large molecule is broken down (favorable -G)
STP = 298K, 1 atm, 1M

Energy Carriers → ATP carries energy load, NAD transfers electrons, NADH responsible for redox

Enzymes → catalyze metabolic reactions Glucose Fermentation and Respiration → catabolic (break down) and anabolic
Carbohydrates and sugars converted to glucose → fats and proteins broken down

Chapter 14: Electron Flow in Organotrophy, LIthotropy, Phototrophy ElectronTransport Systems (ETS) and Proton Motive Force →

Redox → electron donor = reduced / electron acceptor = oxidized
Proton motive force → combination of H+^ concentration gradient + charge difference across membrane → drives cellular processes (ATP synthesis) Phototrophy →
Retinal based proton pump (similar form of chlorophyll) → photoexcitation and photolysis
Chlorophylls contain light absorbing electron carriers, differ based on their absorbance Photosystems I and II → 1 = organic material and sulfides, 2 = purple bacteria Chapter 15: Biosynthesis Biosynthesis requires substrates →
Essential elements (C,O,H,N) for metabolism and energy
Cell components such as lipids and amino acids are reduced Biosynthesis spends energy → only when necessary

Microbial Ecology and Evolution Chapter 17: Origins and Evolution Early Life → essential elements = H,C,O,N,S,P

Main source of energy → solar radiation (temperature permits metabolism) Elements of Life → formed within stars, Earth’s atmosphere was made of CO 2
Organisms consumed CO 2 and replaces atmosphere with N 2 and O 2 → temperature dropped but too far (ice age → reversed by methanogens → cyclic balance) Life → Archaean eon → earliest geological evidence
Carbon dating → dating of fossils and microfossils known through 12 C and 13 C Oxygen → entrance in atmosphere grows and valleys before stabilizing at current level (cycles of aerobic and anaerobic microbial metabolism prevailed)
First life → kicks off CO 2 , first eukaryotes, geological processes move continents First cells → prebiotic soup = if energy is added to the gasses that made up Earth's early atmosphere, the building blocks of life would be created
Water, methane, hydrogen → sonic book
Redox → oceans oxidized by solar radiation and no ozone layer
Theories → Cyanobacteria (3.7 billion years ago)
Unanswered questions → how did life start in such hard conditions RNA → performed all duties that DNA and proteins now perform
RNA replication = earliest form Mutation and Natural Selection →
Random mutation = DNA mistake sequences (1 in 10 bacteria could see a mutation, most are neutral)
Natural selection and adaptation = caused by environmental stresses, enables population to adapt to a changing environment by increasing mutation rate
Reductive evolution = deletion of a gene that is eventually removed (ex. Gene that is not used will be deleted instead of wasting energy) Molecular Clock → time in generations for macromolecule (tracks common ancestors and new mutations) Phylogenetic tree →
Maximum parsimony – the tree that requires the fewest mutations to fit the data

Maximum likelihood – requires statistical calculations and large computational power, but results in a single tree, or a small subset of possible trees
Distance = similarity between species (point in center = most common ancestor to 3 domains)
3 domains → bacteria, archaea, eukarya Natural Selection and Adaptation → Genomic Analysis
Gene duplication = one of the two can gain new function → paralog genes (major source of evolution, the lesser gene is subject to reductive evolution to save energy) Antibiotic Selection → horizontal gene transfer
Antibiotics kill bad and good bacteria protecting body from infection so that drug resistant bacteria can grow, bacteria give resistance to other bacteria (may cause problems) Evolution → all coexist in natural environments *bacteria life cycle and biofilm stages are not the same thing Microbial Species →
Closer genetic code = more similar bacteria → subspecies and strain number = smallest taxonomic classification for bacteria
Same genus = 2 organisms that have >95% orthologous genes within the genome
Same species if they have >95% identity and share common habitat / metabolism Unclassified and Uncultured Bacteria →
Emerging = organism or pathogen that is recently discovered or described

Microbiology Study Guide from Lectures (summary), Study Guides, Projects, Research of Microbiology

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