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80 Cards in this Set

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Characteristics of Eukaryotic (as compared to Prokaryotic) cells?
- nuclear membrane and nucleoli
- usually have more than one chromosome
- Membrane-bound organelles
- 80S ribosomes
- Cell-surface proteins and extracellular secreted proteins are glycosylated
What are 3 nique features of Gram negative
cell walls
- more complex than Gram-positive cell walls
- peptidoglycan layer is thin and sparsely cross-linked (more easily broken)
- outer membrane is a very important component (LPS, & other proteins & liporoteins embedded)
Characteristics of Prokaryotic (as compared to Eukaryotic) cells?
- No nuclear membrane
- usually one chromosome
- 70S ribosomes
- Proteins are not normally glycosylated
- Almost all bacteria of medical importance have a chemically complex cell wall containing peptidoglycan (murein)
What are LPS?
Lipopolysaccharide (LPS)
makes up most of the outer leaflet of the outer membrane lipid bilayer classical endotoxin
3 parts: Lipid A, embedded in the membrane (toxic)
Core oligosaccharide – serves as the connection
O antigen - repeating polysaccharide which extends into the environment, anteigenic
What are 4 unique features of Gram-postitive cell walls?
-b They are relatively thick
- They consist mainly of multiple layers of densely cross-linked peptidoglycan
- have added peptide bridges
- commonly contain teichoic acids
What is LOS?
lipo-oligosaccharide
LPS with the o antigen missing, so it’s:
Lipid A & Core Oligosaccharide only
Describe teichoic acids?
- composed mainly of ribitol or mainly of glycerol connected by phosphate bonds
- - polymers which form long filaments
- linked to the cytoplasmic membrane and span the peptidoglycan layer
- have a net negative charge
- mediate adherence to mucosal cells
- provide antigenic specificity
- possess a weak endotoxin-like activity
What are the 3 functions of the Outer Membrane?
Its strong negative charge is an important factor in evading both phagocytosis and the action of complement.

It is a barrier to certain antibiotics, some hydrolytic enzymes, and other environmental chemicals.

It prevents the loss of metabolite-binding proteins and hydrolytic enzymes found in the periplasmic space.
Name 2 Gram-positive bacteria that have polysaccharides
(Streptococcus), and even in some cases proteins in the outer part of their cell walls. These may also contribute to the antigenic specificity of the cell
Mycobacterium have waxy lipids in their cell walls which makes them more resistant to environmental insults
What is the periplasmic space?
Between the outer membrane and the inner (cytoplasmic) membrane of Gram-negative organisms, an aqueous space which contains the peptidoglycan, someproteins, oligosaccharides, and lipoprotein

-- periplasmic proteins include degradative enzymes, detoxifying enzymes, and binding proteins involved in membrane transport.
-- oligosaccharides = osmoregulatory.
-- lipoprotein connects the peptidoglycan with the outer membrane
What’s the 1st step for Peptidogylcan synthesis (for cell walls construction)?
Autolysins create sites for insertion of newly synthesized peptidoglycan precursors.

Glycosidases break the glycoside backbone, Amidase releases the tetrapeptide from NAM, & Endopeptidases attack the various peptide bonds of the side chains and cross links.
What are the steps in biosynthesis of peptidoglycan?
-- Soluble monomeric precursors made in the cytoplasm as UDP-NAM and UDP-NAG.
-- Particular amino acids are added to UDP-NAM to build up a UDP-NAM-pentapeptide.
-- The NAM-pentapeptide transferred to a membrane carrier lipid called undecaprenol.
-- NAG is added to form NAG-NAM-pentapeptide, still attached to the membrane carrier lipid.
-- Membrane carrier lipid, with its attached disaccharide pentapeptide, flips from the inner surface of the cytoplasmic membrane to the outer
-- The disaccharide pentapeptide is transferred from the carrier lipid to the cell wall glycan backbone and connected to it via glycosidic bonds.
-- Peptide cross links are formed via transpeptidation to complete wall structure
What are the step in biosynthesis of the outer membrane?
The outer membrane precursors are made in the cytoplasm of the cell, then transferred to the membrane carrier lipid.

In Gram-negative bacteria such as Escherichia coli, the inner and outer membranes are fused to each other at several hundred sites. The lipopolysaccharide molecules are assembled at the inner membrane and then inserted into the inner-outer membrane adhesion sites
What is a special characteristeric of mycoplasma bacteria?
-No cell walls
- Have sterols in cytoplasmic membrane
- Sterols must be acquired from host or environment
plasmids
small circular double- stranded DNA molecules attached to the membrane

- * Replicate independently of chromosomal DNA
* They usually contain from 5 to 100 genes which are not crucial , (ie antibiotic resistance, tolerance to toxic metals, production of toxins, or synthesis of enzymes)
* May be transferred from one bacterium to another- via conjugation
Describe Flagella
Composed of flagellin (a globular protein) molecules arranged in several chains that coil together to form a helix around a hollow core.

Acts as a propeller

~15-20 nm
Describe Ftypes of flagella arrangement
monotrichous (one at end)
polar/lophotrichous (many at one end)
petritrichous (like rays of sun)
amphitrichous (groups at each end)
Describe Pili
Thin filamentous appendages distributed, often in large numbers, over the surface of some bacteria. Present in Gram-negative bacteria but only rarely in Gram-positive.

Too slender to be visible in the light microscope – smaller than flagella – 8-9 nm

Composed of a protein called pilin which forms the shaft of the pilus plus a thin tip called the fibrillum (which is the binding part of the pilus)
What are the the functions of pili
*most often involved in attachment of Gram-negative bacteria - fibrillum confers the specificity of adherence
*antiphagocytic
* special F-pilus permits transfer of DNA from one bacterium to another by conjugation
Slime layer
amorphous, unorganized outer layer that is loosely attached to the bacterial cell
replicates independently of the bacterial chromosome.
Capsules & their function
Organized polysaccharides or sometimes polypeptides firmly attached to the cell wall

-My prevent phagocytosis, promote attachment, protect from dehydration & usually antigenic
Describe EXOTOXIN
Toxic proteins secreted into the environment

*Antigenic and are usually heat-labile
*Often exotoxins are responsible for the particular symptoms of the diseases caused by the organisms
*DNA coding for these exotoxins often located on a plasmid
*Ex.: Tetanus & Cholera
Describe Endospores
Highly resistant, metabolically dormant structures
primarily in certain Gram-positive rods

Core surrounded by peptidoglycan cortex & tight keratin coat seals core
Dehydrated, protective (not reproductive) device Resistance to many stains & disinfectants
What are storage granules?
Some bacteria under certain circumstances produce intracellular granules.
These inclusion granules are usually metabolic reserves used for storage of nutrients (or “energy") in a polymerized form, e.g., glucose as glycogen. (Polymerization minimizes the osmotic effect of the stored material)
What are the requirements for culture media?
sterile and usually must contain (at a minimum) water and sources of energy, carbon, nitrogen, sulfur, phosphorus, as well as salts (K, Na, Cl) and trace elements (Fe, Mg, etc)
How can Bacterial cultures can be quantitated
by plating known volumes of known serial liquid dilutions on solid media, incubating, counting the resulting colonies, and then calculating the viable titer of the original culture. Viable titers are expressed as colony-forming units per milliliter (CFU/ml), which may or may not be equivalent to viable cells/ml
Names for bacteria with different temperature ranges for growth?
Psychrophiles grow best at low temperatures, such as those found in a refrigerator.
Mesophiles grow best at moderate temperatures around the temperature of the human body.
Thermophiles grow best at elevated temperatures, such as those found in hot springs.
Bacteria grow fastest in the upper middle portion of their growth range, and slower at the extremes
Obligate anaerobes
grow only in the absence of oxygen. These can be divided into strict anaerobes, which not only are prevented from growing by oxygen but also are rapidly killed by exposure to oxygen after they have grown, and aerotolerant anaerobes, which are not rapidly killed by exposure to oxygen
Facultative bacteria
capable of growth under both aerobic and anaerobic conditions
Microaerophilic
grow best at low oxygen tensions; high tensions (like that of air) are inhibitory
Capnophilic
increased carbon dioxide tension for growth
the most common method for Quatitaion of baceria in liquid culture?
Plate count or viable titer

A sample of culture is serially diluted in liquid then plated.
The dilution factor must be one which will yield countable numbers of colonies on a Petri dish, and the number of colonies must be sufficient to be statistically meaningful (approximately 100 is good)
The plate is incubated to and colonies are counted.
The viable titer of the original culture is calculated from the number of colonies obtained times the total dilution factor

Measures only viable cells. It is slow, but sensitive
Mordant
“to bite” – help dye to bind
(ie iodine in a grain stain procedure helps bind the crystal violet to gram-positive cells walls)
How does Transmission electron microscopy work?
used to visualize internal structures
electron beam is passed through a dry sample in a vacuum. Contrast results from the differential scattering of electrons by the specimen
How is bacterial growth mathematically expressed
The increase in cell number in a growing bacterial population is exponential (also called logarithmic or geometric).
This is as distinguished from an arithmetic (or linear) increase
Obligate aerobes
require oxygen for growth
Descrie the 4 phases of bacterial growth
Lag phase is a time during which the cells are reorganizing their metabolism in order to prepare to resume exponential growth. Growth begins slowly but then accelerates.

Exponential phase (sometimes called logarithmic phase or log phase) is a period of constant exponential growth rate, i.e. constant generation time.

Stationary phase represents a cessation of exponential growth due to the exhaustion of nutrients and/or the accumulation of toxic metabolites

Death phase is a decline in the viable population. It also follows exponential kinetics, but it is an exponential decrease, not an exponential increase. (exp decrease, similar to a “half life” graph)
How does dark field microscopy work?
used to visualize living bacteria. This requires a special condenser with a stop to block light from the center of the field. Then the objective picks up only the light scattered by the specimen
Used to visualize somewhat smaller objects than can be seen with ordinary bright-field illumination
classically been used in the diagnosis of syphilis
How does Scanning electron microscopy work?
It electronically analyzes signals generated by sequentially irradiating points on the surface of a specimen with a very narrow beam of electrons
visualizes surface structures
How does Electronic countingof bacterial cells work?
A defined volume of culture is drawn through a tiny orifice across which an electrical field exists. Each cell that passes through the hole causes a transient increase in the impedance (resistance) and is counted electronically. This method is technically difficult and counts both viable and dead cells
Germicide
Chemical agent capable of killing microbes
Bactericida
kills bacteria
Bacteriostatic
inhibits growth of bacteria, but may not kill them
Sterilization
The use of physical or chemical means to destroy all microbial life including spores
Disinfection
A physical or chemical procedure that destroys most pathogenic microorganisms. High level destroys all microorganisms except high numbers of bacterial endospores. Use w/ items involved in invasive procedures that cannot withstand sterilization. Intermediate use with items not thought to be contaminated w/ spores or resistant bacteria (mycobacteria). Low level treats non critical devices.
Antiseptic
Germicide on skin or living tissue to inhibit or destroy microorganisms. Not used on inanimate objects
Decontamination
Process or treatment that renders a medical device, instrument, or environmental surface safe to handle. Does not necessarily ensure item is safe for patient reuse
Sanitation
Reduction of microbial populations to levels that are considered safe for public health by cleaning using a sanitizer. This usually takes place on an inanimate object
Microorganisms control mechanism of: Alcohol
Disrupts membranes and denatures protein
Ethanol and isopropanol are used
Microorganisms control mechanism of: Detergents
Soaps and detergents help to remove organisms from surfaces; however they are not very effective disinfectants. Quaternary ammonium compounds such as benzalkonium chloride are used for skin antisepsis. They disrupt cell membranes due to a positively charged quaternary nitrogen and long hydrophobic chain
Microorganisms control mechanism of: Phenols
Damage membranes and denature proteins. Phenol derivative is active ingredient in Lysol
Microorganisms control mechanism of: Iodine-
an oxidant inactivates sulfhydryl groups in microbial proteins
Microorganisms control mechanism of: Hydrogen peroxide
An oxidizing agent that attacks sulhydrl groups. Microorganisms that can produce catalase that inactivates H2O2 are resistant to its effects (Staphylococcus aureus)
Microorganisms control mechanism of: Ethylene Oxide
Alkylates proteins and nucleic acids. This gas is used extensively in hospitals for the sterilization of heat sensitive equipment (see picture of gas sterilization unit in power point)
Microorganisms control mechanism of: Dyes
Can inhibit the growth of microorganisms by binding to nucleic acids
Microorganisms control mechanism of: Chlorine
hypochlorite (bleach) is a strong oxidizing agent that kills by cross linking sulhydryl groups thus inactivation microbial proteins
Microorganisms control mechanism of: Heavy Metals
Act at sulfhydrl groups
Microorganisms control mechanism of: Formaldehyde & Gluteraldehyde
denatures proteins and nucleic acids as well as alkylates proteins. Used to sterilize bronchscopes etc
Microorganisms control mechanism of: Acids and Alkalis
- May kill by denaturing proteins as well as altering the pH homeostasis within the microorganism
Microorganisms control mechanism of: Heat
Kills by denaturation of proteins and membrane damage. Moist heat aids in the denaturation process of the proteins by allowing water molecules to bind to the protein and stabilize the denatured structure. Boiling bacteria may kill vegetative cells but will not inactivate spores. Pressure must be increased to allow the water to achieve higher temperatures necessary to inactivate spores. Autoclaves work in this fashion. Dry heat requires more time to achieve sterilization and acts on proteins by oxidizing them.
Microorganisms control mechanism of:Radiation
250-260 nm ultraviolet light is microbicidal. It causes formation of thymine dimers in DNA. UV light is used in hospitals to kill airborne infectious agents. Mycobacterium tuberculosis is exquisitely sensitive to UV light, and containment rooms will often incorporate a UV light source, however distance from the microorganism to the source must be relatively short
Microorganisms control mechanism of: Filtration
separation of microbial contamination from liquid. A small pore size will retain bacteria and spores on the filter surface with sterile liquid passing through to the holding vessel. The benefit over autoclaving is that immunogenic lysed bacterial components present in autoclaved solutions are absent in filtered solutions
What are the stops of the phage life cycle?
-adherence to the bacterium;
-injection of the phage genome (never the capsid);
-expression of phage genes (early genes, then late genes) to produce the gene products (capsid proteins, etc.);
-self-assembly of the progeny virions; and
-lysis of the host cell to release the progeny phage. This is called the lytic cycle.
Phage which can only perform the lytic cycle are called ________?
virulent phage
How are temperate phage different than virulent phage?
In addition to the lytic cycle, they also can undergo lysogenic cycle, turning off their gene expression by producing a specific repressor
Prevents the cell from being killed and prevents the production of progeny phage
Basically lies dormant in host for up to decades
Often contain genes for toxins
Transduction
Involves transfer of DNA from a donor cell to a recipient cell inside a bacteriophage capsid. Phage grows inside of the donor bacterium and accidentally packages some bacterial DNA into a phage coat, instead of just phage DNA. After the phage infection kills that cell the phage capsid containing a fragment of bacterial DNA is released and goes on to infect another cell. The bacterial DNA is injected into that cell (the recipient cell). There are two kinds of transduction, generalized and specialized
Transformation
(sometimes called transfection)
involves the uptake of naked DNA by a recipient cell. Cells must be in a competent state to take up DNA. In some species of bacteria competence is induced by a natural physiological mechanism, while other species must be artificially manipulated chemically or electrically to become competent
lysogenic conversion
Non-toxigenic bacteria can become toxigenic if they are lysogenized by a phage genome bearing a toxin gene
the genetics of a bacterium is entwined with that of the bacteriophage parasites which infect it
Phenol coefficient
Phenol not used much any more, but used to compare how well germicide sterilizes. Phenol coefficient of 1 means the germicide works just as well as phenol. Greater than 1 means more active and less than 1 means less active. It is not very accurate as phenol is not active at all conditions (temp and pH) that other sterilizers are
Efficacy of Killing of Microorganisms Depends on:
Nature of Item being disinfected (more porous = harder to disinfect)
Type of germicide used
Concentration of germicide (each has its optimum) - In general a higher concentration results in more killing. There are exceptions: ethanol is actually more effective at 70% than at 100%.
Duration, temp and pH of exposure-
Nature & Concentration of organisms
Physiological state of organism (vegetative or dormant)
Presence of extraneous organic matter inhibits sterilization and disinfection!
Conjugation
involves transfer of DNA from donor cells to recipient cells by direct cell-to-cell contact. Donor cells contain a fertility factor, such as the F factor, which gives them the ability to transfer DNA
Describe transposable elements
genetic elements found in bacteria including insertion sequences, transposons, and certain temperate phages which integrate at random into the chromosome

cause of mutation
have short (20 to 40 base pair) sequences in inverted repeat on each end
transposase, which recognizes the inverted repeats at the two ends of the element and catalyzes its transposition to a new site on the DNA
Insertion sequences
the smallest type of transposable element , and are about the size of one gene

If the insertion sequence hops into the middle of a gene, it disrupts the sequence of that gene and causes a mutation. Insertion sequences can also cause deletion and inversion of DNA sequences adjacent to their ends
Describe cloning vectors
A mechanism for getting DNA into a bacteria for replication of that DNA

Cloning vectors are usually derived from plasmids, although some vectors are based on bacteriophages. The essential requirement for a cloning vector is that it must be a replicon (it must possess an origin of DNA replication).
DNA ligase
In gene cloning, it is used to attach the ends of a DNA fragment to the two free ends of the linearized vector, thus forming a larger circular DNA molecule
R factors
resistance factors

genes on a plasmid for antibiotic resistance, often multiple antibiotic resistance, which they can easily transfer to recipient bacteria
restriction endonucleases
enzymes called are used to cut DNA into fragments to be cloned and also to convert the circular cloning vector to a linear form so that it can accept the DNA fragment to be cloned
Gene cloning
a fragment(s) of DNA from any organism can be incorporated into a cloning vector which is then put inside a bacterium to be replicated, so that the cloned DNA fragment will be replicated as well
transposon
a segment of DNA containing an antibiotic resistance gene and having an insertion sequence on each end

When it transposes to a new site (e.g. onto a plasmid) it carries the antibiotic resistance gene with it. This is how R factors come to exist
homologous recombination
Because of a lack of a replication orgin, the incoming fragment of bacterial DNA aligns with the region of the resident chromosome with which it has a nucleotide sequence similarity, and cellular enzymes cause the incoming piece of DNA to be substituted for the corresponding resident DNA segment. In this way (recombination) the piece of DNA transferred by transformation, transduction, or conjugation is permanently incorporated into the chromosome of the recipient cell.