Microbiology Final Exam Review

What are Koch's postulates? How do we use these to determine the causal agent of a disease?

(1) The microorganism must be found in diseased but not healthy individuals; (2) The microorganism must be cultured from the diseased individual; (3) Inoculation of a healthy individual with the cultured microorganism must recapitulate the disease; (4) The microorganism must be re-isolated from the inoculated, diseased individual and matched to the original microorganism.

In positive control, how do the inducers activate transcription? Know what activator proteins and activator binding sites do.

Activator proteins bind to specific activator-binding sites on DNA sequence (not the operator). Regulator protein activates the binding of RNA polymerase to DNA

Given a description of different microbes, which domain do they fall under?

Archaea, Bacteria, and Eukarya

What is the difference between cilia and flagella?

Cilia and flagella are cell organelles that are structurally similar but different in length and functions. Cilia are present in organisms such as paramecium while flagella can be found in bacteria and sperm cells. Cilia are shorter and more numerous than flagella. Cilia function for locomotion like flagella but are also used for feeding and circulation.

· What is the difference between a prokaryotic and eukaryotic genome?

Eukaryotic: Linear chromosomes within the nucleus. Much larger, more DNA. Prokaryotic: Generally single circular chromosome that aggregates to form the nucleoid region. It may have plasmids (extrachromosomal DNA) that confer special properties (anti-biotic resistance). Small and compact.

How are enzymes affected by temperature, pH, substrate concentration, and inhibitors?

Extreme high temperatures can cause an enzyme to lose its shape (denature) and stop working. pH: Each enzyme has an optimum pH range. Changing the pH outside of this range will slow enzyme activity. Extreme pH values can cause enzymes to denature.

Negative control

a regulatory mechanism that stops (suppresses) transcription

Regulons

a regulatory protein that controls multiple operons

Lysogenized cell

altered host phenotype because of phage conversion

What does it mean to have a recombinant organism?

an organism that contains a different combination of alleles from either of its parents.

Fusion

is seen in enveloped viruses. When the envelope surrounding the cell fuses with the membrane of the cell. This releases the contents of the enveloped virus into the cell.

Repressible genes

a gene that is regulated by a corepressor or inhibitor, which are small effector molecules that cause transcription to decrease

Transduction

a process of genetic recombination in bacteria in which genes from a host cell (a bacterium) are incorporated into the genome of a bacterial virus (bacteriophage) and then carried to another host cell when the bacteriophage initiates another cycle of infection.

Effectors

term for inducers and repressors

· Decimal reduction time

the D-value or decimal reduction time is the time required, at a given condition or set of conditions, to achieve a log reduction, that is, to kill 90% of relevant microorganisms. (10 fold)

Transformation

the process by which cells take up free ("naked") DNA from the environment.

Enveloped virus

(many animal viruses) have an outer phospholipid bilayer (from host cell membrane) and viral proteins (glycoproteins that form spikes)

Naked virus

(most bacterial viruses) have no additional layers

Specialized transduction

- Infection by bacteriophage - DNA from host chromosome is integrated into virus genome during lysogeny or homologous recombination - lysogenic phage DNA is excised incorrectly when moving into lytic cycle - part of host DNA excised (short fragment)

Generalized transduction

- Infection by bacteriophage - Phage has random piece of previous host DNA - Injected DNA recombines with recipient cell

How can you make bacteria competent and how does this relate to the pGLO transformation lab?

1) Ca2+ and chilling OR 2) electroporation

Operon

A transcriptional unit. May contain one or more different genes.

What are the three ways in which horizontal gene transfer occurs?

1. Transformation 2. Transduction 3. Conjugation

· What are different types of organisms that we study in microbiology? What is their taxonomy: domain and kingdom (if applicable)?

1.) Bacteria & Archaea 2.) Fungi 3.) Protozoa & Algae (protists) 4.) Viruses & Prions 5.) Multicellular animal parasites (Heartworm)

what are chromosomes? Plasmids? Transposable elements?

1.) Chromosome - main genetic element - Most Bacteria and Archaea have single circular chromosome carrying all/most genes - Eukarya have 2 or more linear chromosomes 2.) Plasmids - small (1 kbp to > 1 Mbp) double-stranded DNA that replicate separately from chromosome - Usually circular; beneficial for cell (e.g., antibiotic resistance) - Not essential for function under all conditions - May have a selective growth advantage under some conditions 3.) Transposable elements - segments of DNA that can move from one site to another on the same/different DNA molecule

· In translation, what is the E site, P site, and the A site?

1.) E site - ejection site, where the tRNA that no longer has an amino acid is released back into the cytoplasm. 2.) P site - peptide site, where the tRNA is located that has the peptide chain (the growing protein) attached. 3.) A site - acceptor site, where the next tRNA comes in and decodes the mRNA.

· What are the phases of bacterial growth in a batch culture system? And, what is occurring at each phase?

1.) Lag Phase - the initial period in the life of a bacterial population when cells are adjusting to a new environment before starting exponential growth. 2.) Exponential Phase - The exponential phase of growth is a pattern of balanced growth wherein all the cells are dividing regularly by binary fission, and are growing by geometric progression. 3.) Stationary Phase - the stage when growth ceases but cells remain metabolically active. 4.) Death Phase - As waste builds up and nutrient-rich media is depleted, the death phase is the point where the living cells stop metabolic functions and begin the process of death.

· Factors affecting antimicrobials

1.) Number of microbes - Greater numbers decrease effectiveness. 2.) Concentration of antimicrobial - Higher concentration makes it easier to kill bugs. 3.) Duration of exposure - Longer times leads to killing. 4.) Environmental Factors - Metal ion concentrations, pH, moisture, etc. 5.) Endospore formation - Resistance to agents. 6.) Temperature - Lower temperatures decrease. antimicrobial effectiveness; longer exposure times needed. 7.) Microbial characteristics - Some are more sensitive/resistant.

Decontamination terminology

1.) Sterilization - All vegetative cells and endospores. 2.) Disinfection - Vegetative cells (pathogens, not all); not spores; only for inanimate objects 3.) Sanitation - Removing dirt and dust 4.) Antisepsis - All vegetative cells; not spores; on a biological surface 5.) Degerming - Removing most microbes from a limited area

· What are transcriptional units? Operons? Polycistronic mRNA? ORFs?

1.) Transcriptional Units - DNA segments transcribed into 1 RNA molecule bounded by initiation and termination sites. 2.) Operons - multiple genes (related to a metabolic pathway) transcribed into a single mRNA (polycistronic mRNA) containing multiple open reading frames that will be translated to amino acids. 3.) Polycistronic mRNA - mRNA that encodes several proteins and is characteristic of many bacterial and chloroplast mRNAs. 4.) ORFs - the length of DNA, or RNA, which is transcribed into RNA, through which the ribosome can travel, adding one amino acid after another before it runs into a codon that doesn't code for any amino acid.

What are biofilms and why are they important?

A biofilm is like a well-protected burrow where bacteria hibernate. The matrix creates a barrier against antibiotics. But this defensive barrier also limits the penetration of oxygen and food. Bacteria located in the deepness of the biofilm will start starving. Biofilms have great importance for public health because of their role in certain infectious diseases and importance in a variety of device-related infections.

· What do all cells have in common?

A plasma membrane, cytoplasm, ribosomes, DNA

What is the purpose of a simple stain vs a differential stain?

A simple stain will generally make all of the organisms in a sample appear to be the same color, even if the sample contains more than one type of organism. In contrast, differential staining distinguishes organisms based on their interactions with multiple stains.

· What does the tree of life tell us in terms of the evolution of life?

All life is related by common descent.

What was the Great Oxidation event? How did this contribute to the evolution of life?

An event where O2 rose to one part per million. Atmosphere became oxic, causing new metabolisms (sulfide oxidation, nitrification, other aerobic chemolithotrophy) to evolve. O2 was energetically advantageous because of high reduction potential, allowing aerobes to reproduce much faster than anaerobes.

What type of metabolism do scientists hypothesize as being the first and then what followed after that?

Anaerobic then Authotrophic then chemoauto and chemohetero.

What are the differences/similarities between Archaea, Bacteria and Eukarya?

Archaea: Cell type - Prokaryotes. Cell wall - composition varies, no peptidoglycan. Membrane lipids - Carbon chains attached to glycerol by ether linkage (branched carbon chains). 1st amino acid in protein synth - Methionine. Antibiotic sensitivity - No. rRNA loop - Lacking. Common Arm of tRNA - Lacking. Eukarya: Cell type - Eukaryotic. Cell wall - Composition varies; contains carbohydrates. Membrane lipids - Composed of straight carbon chains attached to glycerol by ester linkage. 1st amino acid in protein synth - Methionine. Antibiotic sensitivity - No. rRNA loop - Lacking. Common Arm of tRNA - Present. Bacteria: Cell type - Prokaryotic. Cell wall - Contains peptidoglycan. Membrane lipids - Composed of straight carbon chains attached to glycerol by ester linkage. 1st amino acid in protein synth - Formylmethionine. Antibiotic sensitivity - Yes. rRNA loop - Present. Common Arm of tRNA - Present.

What are the differences between Bacteria, Eukarya, and Archaea?

Bacteria: prokaryotes. Eukarya: Kingdom Protista (protozoans, algae), Kingdom Fungi, Kingdom Animalia (helminths); first were unicellular, may have appeared two billion years ago. Archaea: prokaryotes; less morphological diversity than Bacteria; historically associated with extreme environments, but not all extremophiles; lack known parasites or pathogens of plants and animals

Conjugation

Bacterial conjugation is the transfer of genetic material between bacterial cells by direct cell-to-cell contact or by a bridge-like connection between two cells. This takes place through a pilus. It is a parasexual mode of reproduction in bacteria.

Bacteriophage

Bacterial viruses

What is the different mechanism of attraction for basic dyes and acidic dyes?

Basic Dye: Positive Ion Acidic Dye: Negative Ion

· Why was rRNA essential in building the tree of life? What happened when we didn't have to culture bacteria? What are the rRNA genes used for the tree of life?

Because it was universally distributed, functionally constant, highly conserved, and had an adequate sequence length for analysis quality.

Know the tests for dehydrogenation, decarboxylation, desulfurylation, and urease production. What is a positive result for these?

Dehydrogenation- hydrogen is being removed. decarboxylation- the carboxyl group is being removed. desulfurization- sulfur removed with a hydrogen or sulfur attached to oxygen. urease production- ability to hydrolyze urea with the enzyme urease.

Taxonomy of the Domain Eukarya vs Domain Bacteria

Domain: Eukarya Kingdom: Animalia, Fungi, Plantae Phylum: Chordata, Ascomycota, Tracheophyta Class: Mammalia, Hemiascomycetes, Angiospermae Order: Carnivora, Saccharomycetales, Rosales Family: Canidae, Saccharomycetaceae, Rosaleae Genus: Canis, Saccharomyces, Rosa Species: C. familiaris, S. cerevisiae, R. pratincula Domain: Bacteria Kingdom: None Phylum: Proteobacteria Class: Gammaproteobacteria Order: Enterobacteriales Family: Enterobacteriaceae Genus: Escherichia Species: E. coli

What were the first autotrophs and then which evolved next? What was the difference between them?

First autotrophs were anoxygenic. Then Cyanobacteria evolved later. 1.) Oxygenic. 2.) Photoautotrophs. 3.) H2O replaced H2S as the reducing agent for CO2. 4.) O2 then became the waste product.

How did the ozone shield form?

Formed naturally through the interaction of solar ultraviolet (UV) radiation with molecular oxygen (O2).

vertical gene transfer

Occurs during reproduction between generations of cells.

Repression

Gene expression is turned off

Activation

Gene expression is turned on

inducible genes

Genes whose expression is turned on by the presence of some substance. Regulate catabolic pathways.

What is a glycocalyx and what is its function? What is the difference between a slime layer and a capsule?

Glycocalyx layer is considered as an additional layer to the cell wall. The main difference between capsule and slime layer is that capsule is a thick glycocalyx layer that is tightly bound to the cell, defining boundaries of the cell whereas slime layer is a thin glycocalyx layer that is loosely bound to the cell.

· What are the differences between Gram + and Gram - bacteria?

Gram + = Cell wall has a thicker (primarily one layer) of peptidoglycan. Gram - = Cell wall (or cell envelope) has at least two layers: LPS and Peptidoglycan.

When there are two carbon sources (glucose and lactose), what happens? What if glucose isn't available?

If both glucose and lactose are present, lactose binds to the repressor, and thus prevents it from binding to the operator. If glucose is absent, then the cAMP concentration is high. cAMP forms a complex with CAP, which binds to the promoter and stimulates lac gene transcription.

Complementation

In bacteria, when a copy of a wild type gene is introduced to a mutant and the wild type phenotype is restored

How does conjugation occur? What is the role of the F plasmid?

In conjugation, one bacterium grows a duct, called a pilus, which attaches to the other bacterium. A genetic element known as a plasmid is then passed through the pilus from the donor cell to the recipient. In another case, viruses play a role in genetic exchange between bacteria. Carries genes for sex pili and transfer of the plasmid • tra genes that encode transfer functions (synthesis of sex pilus and type IV secretion system for DNA transfer)

What is the difference between vertical and horizontal gene transfer?

In vertical gene transfer, the transfer of genetic material is from parents to offspring. It may be through sexual or asexual reproduction. In contrast, the horizontal gene transfer is the movement of genetic material from a donor organism to a recipient organism that is not its offspring.

In terms of function/structure, what are the similarities/differences between pili and flagella?

Pili is used for prokaryotic attachment to surfaces and is an appendage while flagella assist the prokaryote in movement. Flagella are not straight but helical while Pili are straight and non-helical. Flagella are whip-like and long while pili are hair-like and short. Also, flagella are seen in gram-negative and gram-positive bacteria and originate from the cell wall, pili are seen in gram-negative bacteria and originate from the cytoplasmic membrane.

· Direct Measurements of Growth: Plate Counts, MPN method, Direct Counts

Plate Counts: (1:10), (1:100), (1:1,000), (1:10,000), (1:100,000) MPN Method: a method for estimating the number of bacteria in a food or water sample. In this technique, replicate portions of the original sample are cultured to determine the presence or absence of microorganisms in each portion (10 fold). Direct Counts: Can count bacteria that are unculturable or not selected/enriched based on medium. Disadvantages: Dead and Live cells counted (unless using a BacLight system). Unrepresentative sample (technical)

· What is different between prokaryotic and eukaryotic cells?

Prokaryotes: Bacteria & Archaea; No membrane-enclosed organelles (membrane-enclosed structures), no nucleus. Eukaryotes: Plants, Animals, Algae, Protozoa, Fungi. Contain organelles, DNA enclosed in a membrane-bound nucleus.

· How did life evolve on Earth?

Prokaryotes; earliest life forms, fed on carbon compounds, over time enough oxygen accumulated in Earth's atmosphere to allow for the evolution of oxygen metabolizing organisms.

· What is the general difference in the ribosome of prokaryotes and eukaryotes?

Prokaryotic ribosomes are bacterial ribosomes that are small (70S) while eukaryotic ribosomes are large ribosomes (80S). Prokaryotic ribosomes occur free in the cytoplasm while most eukaryotic ribosomes are membrane-bound.

· flow of genetic information: replication, transcription, translation

Replication - DNA is duplicated by DNA polymerase. Transcription - Information from DNA is transferred to RNA by RNA polymerase. mRNA (messenger RNA): encodes polypeptides. Translation: Information in RNA is used to build polypeptides. • tRNA (transfer RNA): convert mRNA to amino acid sequence of protein • rRNA (ribosomal RNA): catalytic and structural ribosome components

How do repressors, corepressors and inducers participate in negative control? What are the two ways in which negative control occurs?

Repressor: prevents the synthesis of an enzyme in response to sufficient levels of a product Corepressor: substance that helps repress enzyme synthesis Inducer: substance that induces the production of an enzyme in response to the presence of a substrate 1. Block the binding of RNA polymerase • DNA in a closed complex • Bind RNA polymerase binding site 2. Block the movement of RNA polymerase. • Bind in front of the RNA polymerase and prevent it from moving forward.

· What are types of simple and differential stains and their purpose?

Simple: Basic Stain - Stain negatively charged molecules and structures, such as nucleic acids and proteins Acidic Stain - Stain positively charged molecules and structures, such as proteins Negative stains - Stains background, not specimen Differential: Gram stain - Used to distinguish cells by cell-wall type (gram-positive, gram-negative) Acid-fast stain - Used to distinguish acid-fast bacteria such as M. tuberculosis, from non-acid-fast cells. Endospore stain - Used to distinguish organisms with endospores from those without; used to study the endospore

Specificity vs. Sensitivity

Specificity - the ability to recognize the target pathogen, minimizing false positives. Sensitivity - the minimum amount of a pathogen needed for the test to detect it, minimizing false negatives.

· How did Pasteur's study show evidence against the idea of spontaneous generation?

The broth in the broken flasks quickly became cloudy—a sign that it teemed with microbial life. However, the broth in the unbroken flasks remained clear. Without the introduction of dust—on which microbes can travel—no life arose. Pasteur thus refuted the notion of spontaneous generation.

What are the differences between lytic and lysogenic pathways of viruses?

The main difference between lytic cycle and lysogenic cycle is that lytic cycle destroys the host cell whereas lysogenic cycle does not destroy the host cell. Viral DNA destroys the host cell DNA and arrests the cell functions in the lytic cycle. However, in the lysogenic cycle, viral DNA may merge with the host DNA.

Promoter

The region for RNA polymerase to bind.

Operator

The region of the regulatory protein to bind.

horizontal gene transfer

The transfer of genes between cells of the same generation.

What are the roles of DNA binding proteins? How do they interact with DNA? How do they perform negative/positive regulation?

Their functions include control of transcription and translation, DNA repair, splicing, apoptosis and mediating stress responses. • DNA-binding protein may catalyze a specific reaction on the DNA molecule (ex: RNA polymerase binds for transcription) • Negative regulation occurs: binding event blocks transcription • Positive regulation: binding event can activate transcription

· Disk-diffusion method: what is it? Be able to interpret results.

This method is based on the principle that an antibiotic-impregnated disk, placed on agar previously inoculated with the test bacterium, pick-up moisture, and the antibiotic diffuses radially outward through the agar medium producing an antibiotic concentration gradient.

· How do we name bacteria?

Through the use of the Linnaean system: Genus + Species. Escherichia coli or E. coli

· Indirect methods of estimating numbers: turbidity, metabolic activity, dry weight

Turbidity: Microbiologists use machines called photometers and spectrophotometers that shine different types of light through culture samples to determine turbidity. Advantages:• quick and easy to perform. • typically do not require destruction or significant disturbance of the sample. • the Same sample can be checked repeatedly. Disadvantage: sometimes problematic (e.g., microbes that form clumps or biofilms in liquid medium) Metabolic activity: •Production of metabolic products: CO2 or acids •Enzyme activity Dry weight: •For filamentous organisms, molds, and actinomycetes. •Microbe removed, filtered, and dried in a desiccator.

· What was key in decreasing the number of deaths by infectious diseases in the U.S.?

Vaccines, Sanitation, Medical Treatments

In which taxonomic groups do we find microbes?

archaea, bacteria, fungi, and viruses.

constituitive genes

are constantly expressed (transcribed and translated into functional products) (60-80% of genes are in this category)

Prophage

bacteriophage DNA that is integrated into host genome

What are prions? What is Prnp, PrPC , PrPSc?

cause Transmissable Spongiform Encephalopathies • Infectious protein; no nucleic acid • Diseases include scrapie, BSE (bovine spongiform encephalopathy), kuru, chronic wasting disease, CruetzfeldtJakob disease • Prnp - produces native prion protein • PrPC - prion protein cellular; alpha helical • PrPSc - prion protein scrapie; Betapleated sheets

Competent Cells

cells that can be transformed

What are viroids?

infectious RNA molecules that lack a protein component • small, circular, ssRNA molecules • smallest known pathogens (246-399 bp) • cause a number of important plant diseases (TMV) • do not infect animals or microorganisms

Virus

genetic elements that cannot replicate independently of a living (host) cell

catabolite repression

inhibits cells from using carbon sources other than glucose

Tegument/matrix

layer of protein between the envelope and the capsid(s) of a virus

Induction

production of an enzyme in response to presence of substrate

Capsid

protein that surrounds the genome of a virus particle (virion)

Activator

proteins bind to specific activator-binding sites on DNA sequence (not the operator)

Global control systems

regulate expression of many different genes simultaneously

Positive control

regulator protein activates the binding of RNA polymerase to DNA

Thermal death time (TDT)

time to kill all cells at a given temperature; affected by population size.

diauxic growth

two exponential growth phases if two energy sources available

Uncoating

when the nucleic acid is released from the capsid.

What is the difference between negative control and positive control?

• Positive Control: regulator protein activates the binding of RNA polymerase to DNA • Negative control: a regulatory mechanism that stops (suppresses) transcription