Foundations In Microbiology 10Th Edition By Kathleen Park Talaro – Test Bank

 

To Purchase this Complete Test Bank with Answers Click the link Below

 

https://tbzuiqe.com/product/foundations-in-microbiology-10th-edition-by-kathleen-park-talaro-test-bank/

 

If face any problem or Further information contact us At tbzuiqe@gmail.com

 

 

 

Sample Test

Answers to Writing to Learn Questions/Writing Challenge Questions

 

CHAPTER 1

 

1.   What does it mean to say microbes are ubiquitous?

From the Latin ubique, meaning   “everywhere”, it is a succinct way of saying that microbes exist everywhere throughout the natural world, even areas with extreme conditions.  Other ways to express this idea are the terms universally widespread and constantly present. The only environments that are microbe-free have probably been artificially sterilized by humans.

2.   What is meant by diversity?

The term diversity is used to denote the immense variety in different types of organisms, with regard to such characteristics as appearance, life style, and distribution.  Although about 1.2 million different species of organisms have been discovered and named, this is only a small fraction of the true diversity present over the entire planet and its millions of habitats.  The case study emphasizes that we are still at the early stages in exploring the richness of life (especially of the microbial type) that is hidden and largely unknown because of small size and inaccessiblity.

3.   Most important events and discoveries:

Many hundreds of separate scientists and labs contributed to the rise of microbiology during its early history.  Tools such as the microscope allowed direct observation of samples and their microbial contents.  Microbes were subsequently seen as discrete entities that could be observed, described, and documented much like larger organisms.  Development of laboratory techniques for culturing microbes using sterile techniques allowed macroscopic handling and control of microbes so they could be studied and understood in greater depth.

Application of the scientific method and experiments to standardize the requirements for fact-based inquiries were very important. The abandonment of the spontaneous generation theory was especially significant because it departed from superstition and prejudice in favor of the scientific method. The institution of the germ theory of disease and the development of aseptic techniques were an essential contribution to medical aspects of microbiology.  The knowledge that microbes cause food spoilage and disease led to early attempts to control microbes using heat and other methods.

4.   Use of the scientific method:

A hypothesis is a statement put forth by an investigator that purports to explain a phenomenon based upon a collection of observations, tests, and other objective criteria. It can be tested experimentally.  A theory is a statement of confidence that scientifically-based observations provide a factual explanation for some natural phenomenon.  It is supported by measurable data collected from numerous experiments used to test the hypothesis.

Examples:  For years, it was not really known what was the exact cause of tooth decay.  Various hypotheses were proposed that acid, sugar, tooth hygiene, and other factors were involved.  Finally, after having the correct conditions for experimentation (on animals that lacked normal resident microbes), it was determined that a combination of excess dietary sugar, lack of cleaning, and certain streptococci living in the mouth were the primary factors.

Likewise, the germ theory of disease started out as a hypothesis, but after thorough verification, it became not only a theory, but is now considered a law because it has held true over several centuries of investigation.

 

5.   Classification of microbes

Evolution is a process by which organisms gradually change (evolve) over long periods of time through inheritance of modified characteristics from ancestors.  It asserts that all organisms arise from preexisting forms and that this relatedness shows itself by similarities in structure, physiology, and genetics.  Biologists and microbiologists use various means to show the pattern of evolution, including trees, taxonomy, classification, and nomenclature.

Taxonomy is a hierarchical system, from general to specific, for assembling organisms into a scheme that emphasizes their origins and relatedness. Classification is the process of collecting organisms into distinct taxonomic groups according to defined characteristics.  Nomenclature is the naming of these categories in collaboration with their level of classification.  Most classification schemes are based on evolutionary relatedness, with organisms that are more closely related placed in the same taxonomic groups.

The correct order of taxa, from broadest and most inclusive to most specific, is: Domain, Kingdom, Phylum, Class, Order, Family, Genus, and Species.  The five Kingdom system groups the archaea with the bacteria in the prokaryotic category.  In comparison the domain system emphasizes the evolutionary separation of the Archaea from the Bacteria and their early relatedness with the Eukarya.

The binomial system of nomenclature assigns a genus and species (scientific) name to each organism; the first letter of the genus is capitalized, the species is lowercase, and the two names are underlined or italicized.  This system standardizes naming and ensures consistency and universality.  Names may also provide an indication of noteworthy characteristics of the organisms or its discoverer.

 

6.   Sources for new infections

(a)  A large number of new infectious diseases arise from animals living in environments where they have crossed paths with humans.  Since 1969, at least 20 new viral diseases have been reported in humans (see Making Connections 25.1).  Most of these have not spread uncontrollably through the population, but a few, such as HIV and Zika virus infections, developed into pandemics. Some of the new diseases are caused by well-known microbes that have become drug resistant (MRSA) and others have mutated to become more infectious for human beings (some influenza strains).

(b) Most of the time outbreaks of newer diseases receive more negative attention from the media than is warranted, often leading to unfounded fear of diseases and germ phobia rather than helping to educate and inform the public.  Understanding the facts about disease outbreaks and learning methods of dealing with them would be a much more valuable contribution to sensible coverage.

 

 

Chapter 2

 

1.   Explain why all compounds are molecules but not visa versa.

A molecule is any combination of two or more bonded atoms, regardless of the types of atoms present. Compounds are molecules that contain at least two different atoms.  For example, O₂ and N₂ are molecules but not compounds, whereas CO₂ and NaCl are both  molecules and compounds.

 

2.   What causes bonding?

Atoms form chemical bonds because they have a structure that allows them to lose, gain, or share the electrons in their outer orbitals.  Covalent bonds form when atoms share electrons. Elements that tend to make covalent bonds have valences that are more suitable for sharing electrons rather than donating or receiving.

Ionic bonds form when an atom donates electrons to another atom, which leaves the participating atoms now oppositely charged. Atoms with unfilled outer electrons shells can easily lose or gain electrons to create stability and usually form ionic bonds.

Hydrogen bonds from between atoms that have polar covalent bonds that are shared unequally.  This creates partial charges on atoms within molecules and creates attraction between oppositely charged atoms on the same or adjacent molecules.

3.   Why are some covalent molecules polar and others nonpolar?

Polarity of covalent molecules arises when the electron pairs of atoms in the molecule are being shared unequally.  This is usually because the atoms differ greatly in the sizes of their atomic nuclei and number of protons.  The atom exerting greater electronegativity (pull on the electrons) keeps the electrons in its sphere.  Examples of molecules that show polarity are H₂0 and phospholipids.  Since water is a polar molecule, it allows intramolecular hydrogen bonding, which makes water cohesive.

 

4.   What causes diatomic molecules?

A few elements exist in their natural elemental state as two identical atoms bonded by a covalent bond.  Examples include oxygen (O₂), hydrogen (H₂), and chlorine (Cl₂).  This state arises from the electron numbers in their outer orbitals.  Diatomic elements are most stable when they share these electrons with a partner having the same number of electrons.  Chlorine, for example, has a single unpaired electron that can be shared with another chlorine, which allows both atoms to essentially complete their outer shell of 8 electrons.

5.   What causes charges to form?

Remember that an unionized atom is neutral and has the same number of electrons (negatively charged) and protons (positively charged), which balance each other out.  An atom that has become charged (ionized) has an imbalance in the number of electrons and protons because it has either lost or gained an electron.  This means that the atom: 1) has less protons than electrons and is now a negatively charged ion or 2) has more protons than electrons and is a positively charged ion.

6.   Why are hydrogen bonds weak?

Hydrogen bonds are relatively weak because they are based on a slight distribution of positive charges on an H atom and negative charges on molecules that have covalent bonds. When the H atom is near another atom or molecule of opposite charge, the two will be attracted and pulled together.  H bonds are easily disrupted as compared with covalent bonds.  They are also different from ions and ionic bonds, which result from fully charged atoms being strongly attracted.

 

7.   What causes hydrophilic and -phobic compounds?

“Hydro” here refers to water, “philic” means to attract, and “phobic” means to repel.  Recall that opposite charges attract and same charges repel.  Water is highly polar, with positive and negative poles.  In the case of hydrophilic reactions, positively charged ions and polar compounds will be attracted to the negative poles of water molecules, and negatively-charged ions and polar compounds will be attracted to the positive pole of water molecules.  This is the basis for solubility of ions in water and for one action of lipids in the presence of water.  Hydrophobic compounds are not attracted to either pole of water water molecules, and in fact will repel it as is seen in the “tails” of lipids.

 

8.   How is a neutral salt formed?

Mixing an acid with a base in a neutralization reaction can form a neutral salt.  An acid like HCl when combined with a base like NaOH will exchange ions to form water (H₂O) and a neutral salt (NaCl).

 

 

9.   Phospholipids in membranes; saturated vs unsaturated fatty acids.

(a)  Phospholipids have a charged or polar head group and two hydrophobic tails.  The dual nature of phospholipids allows them to form bilayers in water that creates a fluid semipermeable membrane, which is an essential component of cells.

(b)  Unsaturated fatty acids have one or more double covalent bonds between the carbon atoms of the fatty acid chains. Saturated fatty acids contain all single covalent bonds between carbon atoms.  Triglycerides composed of unsaturated fatty acids are more fluid than those composed of saturated chains.

(c)  The hydrophilic end of phospholipids is attracted to water because it forms hydrogen bonds with polar water molecules, while the hydrophobic tails repel water and orient themselves away from a water interface.

 

10.                How are amino acids different from each other?

An amino acid has a central carbon atom, a hydrogen group, an amino group, a carboxyl group, and an R group, which is responsible for the distinctive chemical nature of each of the 20 amino acids.  Each R group varies in size, chemical make-up, acidity or basicity, or being polar or nonpolar.

 

11.                Why is DNA considered a double helix?

DNA is called a double helix because its two complementary chains of DNA are held together with hydrogen bonds and the chains are assembled into a  structure that looks like a twisted ladder or helix.

 

12.                Describe a dehydration synthesis reaction.

Several macromolecules including polysaccharides and proteins are synthesized by bonding the atoms on individual smaller units together by a specialized bond, forming a longer molecule similar to linking units on a chain.  In order for this bond to form, one atom releases an -OH group and the other releases an -H, which combine to make a water molecule.  Since this action involves removal of water, it is called dehydration synthesis.

 

13.                How does chemistry support the study of microbiology?

Because organisms are composed of atoms and molecules like all other matter in the universe, we can develop a much greater understanding of their structures and functions if we know something about the chemicals they are made of.  A knowledge of chemistry forms a solid basis for explaining how enzymes and metabolism work, how cells grow and reproduce, how microbes infect and what their toxins do to the human body, how they contribute to the ecology of environments, how drugs work to control them, the operations of their genetic material, and much more that you will discover in later chapters.  The more we find out about the chemistry of microorganisms, the more we insight we gain about their biology.

 

 

Chapter 3

 

1.   (a) What features are most important in selecting a microscope?

It should have the capacity for high magnification and good resolution, so that the image it makes provides an accurate view of very small things.

Magnification involves the creation of an enlarged image by means of lenses.  Resolution describes these lenses’ ability to create a clear image, or differentiate two adjacent objects as separate, regardless of degree of magnification.  Both are limited by the structure of the microscope lenses, but the resolution is the more critical quality. In general, one would look for a microscope that has a small maximum resolving power, since a small number indicates the size of the objects that can be viewed in detail.

1.   b) It is possible to have “empty” magnification, with the object being enlarged but not resolved. Beware of a cheap microscope that claims to have 1000X magnification, because it probably cannot resolve what it magnifies.

To improve resolution, a microscope can be fitted with magnifying lenses with large numerical apertures, a blue filter can be used to limit long wavelength visible light from passing through the specimen, and oil can be used with the oil immersion lens to decrease light scattering.

 

2.   How does one get 2000X magnification from a 100X objective?

By fitting it with a 20X ocular.

 

 

3.   Differentiate between macroscopic and microscopic methods of inspection, with examples.

Microscopic observations of microbes require a microscope to see individual cells and characterize them based upon their size, shape, arrangements, physical features (e.g. flagella) and color, if using a differential stain (e.g. Gram stain).  Macroscopic observations of microbes require culture media that expand populations to visible masses or colonies.  These growths can be characterized based upon growth patterns, color, texture, size, and odor.  The media may also contain substances (e.g. differential media) that allow for the characterization of species based upon biochemical or physiologic properties of the microbe.

 

4.   The steps of the Gram stain and how it is useful in diagnosis

The Gram stain is a differential stain that reacts differently with two bacterial cell types and stains them different colors (e.g. gram-positive cells are purple and gram-negative cells are pink at the end of the procedure).

In the first step, a slide bearing a fixed smear is stained with crystal violet.  This dye will stain all cells the same color (purple, regardless of cell type).  This is followed by a series of steps that will allow the differences to be readily seen.  First, a mordant–Gram’s iodine– is placed on the smear to chemically change the nature of the crystal-violet in certain cells.  It binds the crystal violet into the cell walls of the gram positive cells in a way that it will not be readily removed.  It does not bind the dye into gram negative cells this way.  This step is critical, but to actually see the differences, it is essential to next decolorize and then recolorize.

During decolorization, ethyl alcohol (or acetone) is added to remove the purple dye from cells that are actually gram negative.  It decolorizes the gram negatives by dissolving the dye out of the cell wall.  It is a solvent that also removes part of the cell wall of gram negative cells.  When this step is performed correctly, gram positive cells will remain purple and gram negative ones will be colorless.  In order to see them more clearly, the smear is treated with a counterstain, safranin, that will stain the colorless cells red.  Purple gram positive cells will not be changed by this second dye.  If the specimen being tested is a pure culture, you should have all cells the same color.  Mixed specimens will generally show both colors, indicating more than one type of cell is present.

The Gram stain is a very effective early test for specimens from potential infections. The Gram reaction in conjunction with the shape of the cells can give clues to the identity of a potential pathogen. Note that chapter 4 has an outline of the major bacterial infectious agents broken down by Gram reaction and shape.   For example,

A Gram stain done on a urine sample can quickly uncover the type of microbe that may be present. Gram negative rods point to Escherichia coli as the infectious agent.  A stain of a throat specimen will also help identify or eliminate the type of pathogen that may be involved. We saw in the case file that a stain of CSF can be important in rapid diagnosis of meningitis.

An initial Gram stain can also guide the selection of early treatments with antibiotics.

 

5.   Steps taken to isolate, cultivate, and identify a pathogen in urine.

You would use many of the 6 “I” techniques: a) inoculation, the introduction of microbes from the specimen into a growth medium; b) incubation, the culturing of microbes under controlled environmental conditions for a period of time; c) isolation, the separation of microbes from a mixed culture into isolated colonies for the purposes of creating a pure culture; d) inspection, the macroscopic and microscopic evaluations of microbial growth and cellular characteristics; e) Information gathering, doing additional tests as required for identification and f) identification, pinpointing and naming the microbe of interest.

A urine sample will undoubtedly contain several types of microbes, so most labs would start isolation with selective, differential media that can block the growth of unwanted microbes and favor the growth of the pathogen.  It could also highlight differences in biochemical reactions that provide additional identification characteristics. For example, since Escherichia coli is the most common cause of urinary tract infections, MacConkey agar or other selective medium that grows only gram negative bacteria would be a good choice.

The plate of media would be inoculated with the urine sample using the streak method and incubated for 24-48 hours to form separate or isolated colonies to work with.  It would be common practice to Gram stain the urine to clarify microscopically what types of bacteria might be present.

The isolated colonies would be examined for their reactions and selected for subculture to make pure cultures for further testing.  This could include a panel of biochemical tests that are useful in identifying urinary tract pathogens and drug sensitivity testing to aid in the selection of correct therapy.

The combination of all test data will point to the correct pathogen.  Other urinary pathogens would require different media, but the same or similar steps would be followed.

 

6.   Trace the pathway of light from source to eye and the areas it passes through.

The light originates from a bulb in the base of the microscope; this light is from the visible spectrum, giving off a range of wavelengths.  The light usually is directed through a blue filter housed above the lamp that can filter out most wavelengths except for the shorter blue ones that facilitate good resolution.

Light leaving the filter shines through the opening of the iris diaphragm, which is the primary device for regulating the amount of light that enters the condenser lens.  This condenser is placed just below the specimen on the stage and functions to collect the light beams into a tight bundle that focuses on the specimen area.

At the stage, light enters the specimen and continues on a pathway through the objective lens where the first of two images is made.  This image, called the real image, is the first level of magnification, and varies according to the magnification of the objective, usually from 4X to 100X.

As the image passes upward through the tubular body of the microscope, it enters the ocular lens, where a second image, the virtual image, is formed. This is an image of the real image, and provides a total magnification from the two lenses operating together  of 40X to 1000X, depending on the objective in place and the ocular power.

This image is captured by the eye and interpreted by the brain.

 

 

 

CHAPTER 4

 

1.   Label the parts of the bacterial cell.

The parts clockwise from top right are: fimbriae, granule, cell wall, cell membrane, ribosomes, flagellum, cytoplasm/actin, outer membrane, pilus, bacterial chromosome, glycocalyx.

 

2.   Properties of life and prokaryotic cell structures involved.

All life forms show a collection of characteristics that, together, we define as life or being alive.  These include having a cellular composition; reproduction, passage of DNA to offspring; growth and development; metabolism; responsiveness, meaning the capacity to interact and communicate; transport of materials into and out of the cell; and processing of nutrients. Even the smallest, simplest cells must demonstrate these properties.  Viruses are not considered to be alive because they lack most of these characteristics and are inactive without the presence of their host cell.

Prokaryotic cells are smaller and appear to be simpler in structure because they lack organelles.  They do have simple reproductive methods such as binary fission or budding. Most have a single chromosome that is copied during cell division and distributed to offspring.  Being single-celled, they have simple developmental phases, although some make specialized structures such as endospores.  The prokaryotic cell membrane is the center for many critical functions that involve metabolism and transport.  They have complete protein synthesis machinery, including ribosomes. The cell surface sprouts appendages such as flagella for movement, pili for communication of genetic material, and fimbriae that serve as hold-fasts to the environment.

 

3.   The basic process of biofilm formation:

Most microbes become part of a biofilm association in their natural habitats.  This involves colonizing the habitat and participating in complex networks that support a rich and diverse interchange.  The initial biofilm is started by primary settler microbes on the moist surfaces of objects such as rock, soil, plants, and even living tissues.  These early colonists secrete a sticky glycocalyx matrix that affixes them to the surface.  There they grow and build up a layered mat of cells.  Other microbes will be drawn to the biofilm, adding thickness and complexity.  Biofilms can remain viable for long periods.  Consider the large stromatolites of cyanobacteria that form in oceans and lakes and can be so long-lived that they eventually fossilize.  Other biofilms are involved in biogeochemical cycles, decomposition, plant associations, and many other ecological actions.  Biofilms also colonize the body as normal residents of the mouth, intestine, and skin and are involved in infection. In most biofilms, the microbes communicate and cooperate almost as a single organism, with the members deriving benefit from the stable community a biofilm provides.

 

4.   What observations place Archaea closer to Eukarya in evolution.

Archaea are prokaryotic and have 70S ribosomes, but the ribosomes have a structure that is more similar to 80S eukaryotic ribosomes,  and archaea do not have peptidoglycan in their cell walls.  Archaea differ from eukaryotes in that they have a single circular chromosome, no nucleus or organelles, and no sterols in the membrane.

Evidence that suggests that Archaea are more closely related to the Eukarya is the similarity in their rRNA sequences and protein synthesis.  Because of these simililarities and differences, the Archaea are considered a completely separate microbial group and cell type that evolved from an ancestral form that produced the Bacteria and Eukarya lines.

 

5.   How are bacterial endospores killed/dated?

Bacterial endospores are capable of withstanding extremes of heat, drying, freezing, radiation, and chemicals that would readily kill ordinary cells.  In order to kill these hardiest of cells, conditions must generally be created that are far more harsh than those found usually found on Earth.  An autoclave for example reliably kills endospores by subjecting them to a combination of moisture, heat, and pressure far greater than is normally encountered.

Generally, the age of an endospore can be inferred from the age of other things in the immediate vicinity.  An endospore found in a fossil may be dated through the use of radiometric (Carbon 13) dating of the fossil, for instance, and the fossil itself may have been discovered buried at a depth that indicates the age of both the fossil and its accompanying endospores.

 

CHAPTER 5

1.   The anatomy and functions of the major eukaryotic organelles:

The nucleus is where the DNA genome is stored and is the site for DNA and RNA synthesis.  A large structure in the nucleus, the nucleolus is the location for ribosome synthesis.  The nuclear membrane contains pore proteins to facilitate transport of nucleic acids to the cell cytoplasm.

The endoplasmic reticulum (ER) is continuous with the nuclear membrane and is a membrane-enclosed organelle connected by a vesicle system.  It becomes either rough (RER), which contains ribosomes and is where proteins are synthesized and stored; or it becomes smooth (SER), where lipids are synthesized and modified.  Both types of ER exist as a continuous internal membrane passageway that transports materials throughout the cell.  The Golgi apparatus is formed from vesicles coming from the ER and acts in sorting, processing, and packaging molecules from the ER for secretion.

Mitochondria are double-membraned organelles where most energy metabolism takes place..  Chloroplasts contain the photosynthetic pigments and are the sites of photosynthesis in algae and plants.  Both mitochondria and chloroplasts contain their own DNA and ribosomes and are capable of self-replication.

Flagella and cilia are both protein appendages of motility that undulate back and forth to create movement.  Flagella are long and usually exist as single appendages whereas cilia are shorter but exist in large numbers over the cell surface.  Both have an outer layer of the cell membrane covering them.

Organisms use lysosomes for digestion of foods or microbes contained in endocytic or phagocytic vesicles.  They are also used to remove cell debris in damaged tissue.

Eukaryotic cells that lack a cell wall for support (e.g. animal cells) use an intracellular skeleton, or cytoskeleton, to maintain their cell shape and support.  The cytoskeleton is also used for anchoring organelles, movement (e.g. pseudopodia), vesicle trafficking, and cell division.

4.   The chart below reviews the major similarities and differences between prokaryotic and eukaryotic cells.

Characteristic	Prokarya (Bacteria)	Eukarya
Cell membrane	Lipid bilayer with embedded proteins One group has sterols	Phospholipid bilayer with embedded proteins Sterols in most cells
Cytoplasm	Yes	Yes, but generally more complex because of organelles and cytoskeleton
DNA genome	Yes usually a single chromosome	Yes, usually 3 or more chromosomes
Nucleus	No	Yes
Organelles (e.g. ER, mitochondria)	No	Yes
ATP synthesis	Located in cytoplasm and cell membrane	Located in cytoplasm and mitochondria
Photosynthesis	Some members (e.g. cyanobacteria); pigment is in thylakoid membranes	Some members (e.g. algae); pigment is in chloroplasts
Motility	Bacterial flagella with filament, hook, and basal body	Eukaryotic flagella with microtubules; cilia, pseudopodia
Cell wall	Peptidoglycan in most	Cellulose, chitin
Cytoskeleton	Actin in some bacteria	Microfilaments and microtubules
Ribosomes	70S	80S (70S in mitochondria and chloroplasts)
Cell size	Generally 0.5-5 µm	Generally 10-100 µm
Reproduction	asexual only with binary fission or budding	asexual (mitosis) and sexual (fusion of gametes)

 

 

Eukaryotic microbes are found in the kingdom Fungi (yeasts and molds), Protista (algae and protozoa), and Animalia (worms).  Unicellular microbes exist on their own, colonial organisms can exist independently but typically exists with a group of other cells, and multicellular organisms contain cells that act together to sustain life but cannot exist independently.

Unicellular fungi are yeasts and multicellular fungi include mushrooms and molds.  Unicellular protists include Paramecium and Euglena, colonial protists include Volvox, and multicellular protists include sea kelp.  The parasitic helminthes are multicellular animals with tissues and organs.

 

 

2.   Trace the synthesis of cell products through the organelle network.

The Messenger RNA that gives the codes for making a protein is transcribed from DNA in the nucleus.  It leaves the nucleus through a pore and is transported to the cytoplasmic ribosomes or to the rough endoplasmic reticulum (RER) ribosomes for translation.  Proteins synthesized in the RER are carried to its outer edge where they are packed into membranous sacs or vesicles.  These transport vesicles are taken into the cisterni of the Golgi apparatus where they may be modified (carbohydrates may be added).  The Golgi repackages the proteins into condensing vesicles for secretion through the cell membrane into the extracellular environment.  Some proteins may be packaged into cytoplasmic vesicles e.g. lysosomes) and remain intracellular.  Lipids (e.g. phospholipids and sterols) produced in the SER may also pass through the Golgi on their way to the cell membrane where they are inserted as the membrane grows.

 

3.   (a) What is the reproductive potential of molds?

A single colony of mold on a moldy lemon may produce 2000 spore-bearing structures, each with the potential to release 2000 single spores.  This means that just one colony could give off 4,000,000 spores, each spore having the potential to germinate and produce another colony if it falls onto a favorable surface.  This explains the relatively high spore counts in habitats with adequate moisture and food substrates.

(b)  How are mold spores different from bacterial endospores?

Mold spores differ from bacterial endospores in that mold spores are produced through asexual or sexual reproduction and bacterial endospores are resistant survival cells and not reproductive.  Most mold spores do not have the dense coatings that protect them from chemicals and heat so they are less resistant. Most mold spores are produced extracellularly on special hyphae and do not arise from inside another cell like endospores.

 

 

 

4.   (a) (b) Chart comparing microscopic eukaryotes

Characteristic	Fungi	Algae	Protozoa	Function in cell
Flagella	A few (chytrids)	Many groups	Many groups	Swimming, feeding
Cilia	No	No	Yes, in the Phylum Ciliophora	Movement, feeding
Glycocalyx	Yes	Yes	Yes	Attachment, communication
Cell wall	Chitin	Cellulose, silicone	None	Structural support
Cell membrane	Contains ergosterol lipids	Contains sterols	Contains sterols	Transport and engulfment
Nucleus	Yes	Yes	Yes	The location of DNA (genes) and ribosome synthesis
Mitochondria	Yes	Yes	Yes	Site of most aerobic (O2-using) metabolism
Chloroplasts	No	Yes	Most do not, some mastiogophorans have them	Site of photosynthesis
Endoplasmic reticulum/	Yes	Yes	Yes	An extensive series of hollow passageways that act to synthesize, transport, and modify materials
Ribosomes	Yes–80S	Yes–80S	Yes–80S	Site of protein synthesis
Cytoskeleton	Yes	No, cell wall maintains shape and support	Yes	Support, locomotion, anchors for organelles
Lysosomes	Yes	Yes	Yes	A vesicle containing enzymes for breaking down food and other materials inside the cell
Microvilli	Not likely, due to cell wall	No, cell wall prevents it	Yes	Fine extensions of the cell membrane that aid in absorption of nutrients
Centrioles	No	No	Yes	Pairs of bodies that release the spindle fibers for mitosis

 

 

 

Chapter 6

 

1.   (a) What characteristics of virus are “lifelike?”

The primary feature of viruses that fulfills one life criterion is that they contain genetic material.with a blueprint for constructing new viruses.  It could also be said that they interact with their host cells and evolve.

(b)  How are they more similar to lifeless molecules?

Other than these properties, they lack all other life criteria:  they do not have cells, they do not grow, they do not metabolize or release energy to drive reactions, and they do not have an independent nutrition. All of these activities can only occur in association with a host cell they invade.  Other ways they seem like lifeless molecules is their crystalline structure and the fact that they are inert and inactive by themselves.  It seems that they exist in a transitional zone between fully-functioning live organisms and non-living molecules.

 

2.   Why is it that a virus like HIV can only infect certain human cells?

One of the major adaptations of viruses is how they hook up with their host cells.  In being inert and inactive, they cannot chase and attack host cells, so they need to be brought into close contact with them. One important factor in this contact is that the virus must be able to adsorb or bind specifically to a type of surface receptor on the host cell as the first step in invasion.  It is striking that when this binding takes place, the host cell becomes a willing accomplice in bringing the virus into the cell in some form.  So, in the case of HIV, the virus first is transmitted into the body through blood or sexual fluids and is carried to locations where only certain white blood cells have the correct receptors to complete the attachment and facilitate penetration.  HIV cannot bind onto all types of human cells because most cells lack the receptors that HIV needs for binding.  This same explanation fits for hepatitis viruses that attack only liver cells or influenza viruses that attack respiratory epithelial cells.

 

3.   (a) How can viruses synthesize the components they need to make new viruses?

Viruses contain no synthetic machinery such as ribosomes and they lack most enzymes required for producing virus parts.  They become master regulators that take over the natural cell processes for synthesis and packaging that already exist in the host cell. Depending on the virus, this includes using host’s enzymes and energy to make new virus capsids, nucleic acids, and receptors using the virus DNA or RNA as a guide.  Enveloped viruses also pick up their envelope from the host cell.

(b)  What enzymes are found in viruses?

Some viruses come equipped with their own specialized enzymes for synthesizing components such as nucleic acids.  This would include polymerases for synthesizing DNA and reverse transcriptase (in HIV) for converting their RNA genetic material into DNA.

 

4.   What dictates host range of animal viruses?

Most viruses are limited by the types of cells and organisms they can use as a host.  An animal virus can multiply only in animal cells that have a specific receptor for them.  And a virus cannot display a wide variety of receptors to fit many animals.  So, most viruses will only infect a single species or possibly close relatives.  For instance, the human hepatitis virus will not infect cats or horses, and the feline distemper virus infects felines, including domestic and wild cats, but not humans, chickens, or any other animal.  An exception is some viruses that have a wider range because the animal hosts all possess a universal receptor that is compatible with the virus.  Rabies virus can infect almost any mammal, because it uses a receptor common to their nerve cells.

 

(b)  How is influenza virus different in its host range from most viruses?

The influenza viruses are of the type discussed in question 4 in having a wider host range than most other viruses.  Its hemagglutinin (H) spikes and neuraminidase (N) spikes are subject to alterations in shape through genetic mutations.   This means that viruses from swine and many varieties of birds could potentially develop a spike configuration that would be compatible with attaching to human cells.  This is why there is great concern when a new virus arises, since it could jump hosts and infect large segments of the population that would lack immune protection.

 

CHAPTER 7

 

1.   Predict the direction of osmosis; is one example a halophile and why?

(a)  In this one, the external solution is more concentrated (is hypertonic) than the internal solution (hypotonic).  Since the internal solution has more water to lose, the water will osmose in the direct of the external solution.   (b)  In this one, the inner solution is hypertonic to the external solution, so the water will flow into the cell.  (c) In this one, the solutions are the same tonicity (isotonic), so there will be the same rate of flow both directions, and no net movement of water.

Yes, (c) could be a halophile because it lives in a high salt environment and maintains a high salt internal solution, thereby becoming isotonic with and stable in this environment.

 

2.   Basic metabolism of methanogens.

Methanogens are a type of archaeon that lives on inorganic chemicals for both its nutrient and energy needs (chemoautotroph).  Their metabolism is totally anaerobic, and they live in a variety of low oxygen habitats such as swamps, deep ocean, soil, and intestines.   What is unusual about their biochemistry is that they can use H₂ gas as an electron donor and combine it in a chemical reaction with CO₂ to produce methane gas (CH₄), the simplest organic compound.  Another product from this reaction is water.  Because they generate methane, this group has been termed methanogens.  Ancient beds of these microbes appear to underlie the ocean, where they constantly release methane into the upper regions.

 

3.   Discuss extreme environments and extremophiles.

The earth contains many extreme zones and habitats, ranging from the hottest to the coldest, from salty to acidic, from toxic with heavy metals to high in pressure.  And in all of these places, scientists have isolated forms of hardy archaea, bacteria, eukaryotic microbes, or even viruses.   Many of these habitats are remnants of the early earth as it existed billions of years ago. So, it only makes sense that there would be microbes that evolved in the presence of these early conditions that still occupy these habitats.  These “lovers of extreme conditions” or extremophiles have developed specialized strategies in their structure and enzyme function that allow them to survive what would be deadly conditions to most other organisms.  To extremophiles this environment is “normal”.

Other extremophiles may be recent colonists in an extreme environment that have made rapid adaptations for surviving in this environment.  This is what often happens in human-made sites like polluted waters, toxic waste dumps, and oil spills.  Many of these facultative extremophiles can be used in bioremediation because of their natural capacity to break down toxic substances.

 

4.   Differentiate between mutualism and syntrophy, with examples.

In both of these associations, there is a cooperative relationship between microbes and other organisms that favors the survival of all participants. Both types of partnerships  usually have a nutritional basis of some sort.   Some of the major differences between them are that 1)  mutualism is an obligatory and co-dependent type of symbiosis.  It requires the participants to live in intimate contact and interact for survival.   Examples include the relationship between termites and the microbes in their guts, cattle and their rumen microbes, and deep sea worms and their bacterial symbionts.  2) syntrophy is a cooperative exchange among the members that is not symbiotic.  It involves a close association between the partners, but it is not obligatory.  The members can live separate existences but cooperate in cross feeding or breaking down a complex substrate.  Examples include soil microbes that give off products that can be used by the partners or bacteria that fertilize plants by their actions.

 

5.   (a) What events in quorum sensing help a biofilm function in unison?

A biofilm is a living network of microbes that function together as a unit to colonize an environment and establish favorable conditions.  The members of a biofilm are known to communicate through a type of chemical signaling called quorum sensing.  To appreciate the impact of quorum sensing, consider the problem of a single free-swimming microbe attempting to digest a large food particle.  Its actions will be greatly diluted out by its surroundings, and digestion will be very slow and inefficient.  But in the case of a biofilm, the microbe would settle down and establish a flourishing network of thousands of similar microbes on this food particle, thereby making it possible to amplify and coordinate a response.  When the biofilm organisms reach a significant size called the quorum, they synthesize and release special inducer molecules in unison.  These inducers ensure that a large aggregate of cells will be stimulated simultaneously to produce digestive enzymes to break down the mass of food, with all microbes participating and sharing the products.

(b)  During infections, the biofilm could coordinate the production of enzymes for invading the host and produce toxins in unison.  It could also favor development of drug resistance, making it easier to avoid the effects of drugs given to treat the infection.

 

6.   What is happening to the population at points A, B, C, D?

This graph depicts a growth curve in a closed system where no additional nutrients are added and space is limited by the culture vessel.  (A) This represents the early stage in growth called the lag period.  It tends to show a flat or very slow rate of division, because the number of new cells being added is very low.  The cells are active metabolically and synthesizing molecules for cell division, but they are not yet dividing at the high rates yet to come.  (B)  This is the exponential or log phase, during which the population is doubling during every generation.  Each new cell will divide during its generation time, increasing the number of cells geometrically until the limiting factors start to come into play. (C)   During the stationary phase, the rate of cell division slows due to a lack of nutrients, space, and factors such as lack of oxygen. Now, there are as many cells dying as being added to the population, so the number of live cells plateaus out during this period.  During this time many cells are synthesizing but not dividing.  (D)  When the adverse conditions of the culture vessel intensify, the cells begin to die more rapidly and growth rates are at a minimum.  During this death phase the viable count goes down dramatically as more and more cells are destroyed by a build up of toxic metabolic wastes.

 

7.

	Source of Carbon	Usual Source of Energy	Examples
Photoautotroph	CO2	Sunlight	Algae, cyanobacteria
Photoheterotroph	Organic substrates	Sunlight, respiration	Purple, green sulfur bacteria
Chemoautotroph	CO2	Oxidation of simple inorganic substrates	Methanogens, thermal vent bacteria
Chemoheterotroph	Organic substrates	Respiration, fermentation	Fungi, protozoa, many types of bacteria, animals
Saprobe	Matter from dead organisms	Respiration, fermentation	Fungi, bacterial agents of decomposition
Parasite	Tissues and fluids from living organisms	Respiration, some use host’s sources	Pathogens, infectious agents

 

 

 

Chapter 8

 

1.   (a) Relationship of:  anabolism and catabolism

 

Both are types of metabolism associated with the biochemical reactions in cells, although they are actually opposing types of reactions that complement and support each other.  Catabolism refers to reactions that break bonds and degrade molecules into smaller compounds, usually with the release of energy.  Anabolism refers to reactions in the cell that make bonds and build cell structures, such as those of synthesis and growth.  The two forms of metabolism create a continuous cycle in the cell, whereby nutrients are catabolized to release energy that is used to power anabolic activities.  They maintain an essential balance in the physiology of the cell.

 

(b)  ATP to ADP:

 

Adenosine triphosphate is a type of chemical currency that is expended to do work in the cell.  It is composed of an adenine and a ribose molecule bonded to a chain of 3 phosphates.  The bonds between the last two phosphates release free energy when they are broken.  This energy can be used in a variety of anabolic activities.  ATP is produced during several energy-yielding cell processes, including respiration, photosynthesis, and fermentation. Adenosine diphosphate is produced when the first phosphate bond on ATP is broken to release energy.  ADP can then be used again to capture the energy from another catabolic reaction and, combined with a phosphate group, recreate the high energy bond of an ATP.  This cycle of

ATP<—->ADP is continuous in an active cell.

 

(c ) Glycolysis to fermentation

 

Glycolysis is the first series of biochemical reactions that catabolize glucose.  It is an anaerobic phase shared by the three pathways of aerobic respiration, anaerobic respiration, and fermentation.   Its primary function is to initiate the breakdown of glucose, with the release of a small amount of ATP, NADH, and pyruvic acid.

 

Fermentation is the end stage in the metabolism of some bacteria and yeasts that converts the pyruvic acid and NADH from glycolysis into alcohol, organic acids, gases, and numerous other products.  Like glycolysis, it occurs anaerobically and does not, by itself, generate any ATP.  It is a common metabolic strategy for facultative anaerobes and other microbes that encounter or live in oxygen-free habitats.

 

 

(d)  Electron transport to oxidative phosphorylation

 

Both electron transport (ET) and oxidative phosphorylation (OP) represent the final stages in the metabolism of glucose.  They are located in the same place (cristae of the mitochondria) and do not operate totally independently from one another.  The ET chain transports electrons provided by NADH from other pathways and OP powers synthesis of ATP using energy released by the transport process.  The transfer of electrons by ET occurs along a series of cytochrome carriers.  This is accompanied by creation of a proton (H+) gradient that powers the synthesis of ATP through phosphorylation.  In its final step, an oxygen based acceptor binds the H+s generated.  Oxidative phosphorylation differs between aerobic and anaerobic respiration. In aerobic systems, ATP synthesis is linked to the use of oxygen gas (0₂) as the final electron acceptor and in anaerobes, the final electron acceptor is some oxygen-containing compound such as nitrate, sulfate, or carbon dioxide.

 

 

2.   Give the general name of the enzyme that

3.   converts citrate to isocitrate

isomerase  (specific name is aconitate)

1.   reduces pyruvic acid to lactic acid

lactate dehydrogenase

1.   reduces nitrate to nitrite

nitrate reductase

 

3.   Explain what is unique about the actions of ATP synthase?

 

ATP synthase is an enzyme stationed in the cristae of the mitochondrion in eukaryotes and in the cell membrane of prokaryotes.  It performs the extremely important role in final synthesis of ATP in microbes that use the electron transport system.  The significant functions of this enzyme are 1) to pump H+ ions from the outer to the inner compartments of the mitochondrion and 2) to couple the release of energy (from the proton motive force) with the synthesis of ATP.  This enzyme is unique in the biological world by having one unit ((f₁) that rotates in the membrane to pull in ADP and P and create the high energy bond, yielding ATP.

 

4.   Compare the equation for aerobic metabolism with the summary of aerobic metabolism (figure 8.22), verify the final totals of reactants and products, and indicate where they occur in the pathways.

 

The equation reads:  glucose  +  6 O2 + 38ADP +38Pi–à 6 CO2 + 6 H2O + 38 ATP

 

If you add up the ATPs (net #) formed by oxidation of one glucose molecule in glycolysis (2), the Krebs cycle (2), and the electron transport of NADHs and FADH2s (34), it comes out to 38 ATP, which is a theoretical estimate.

 

If you also look under the heading “Summary of Aerobic Respiration”, you will find that, from one glucose, 6 oxygens are consumed in oxidative phosphylation, 6 carbon dioxides are produced between acetyl CoA and Krebs cycle, and 6 waters are produced as the final step in electron transport. The ADPs and Pis do not appear in the summaries, in that it is assumed they must be present to synthesize ATP.

 

5.   Describe four requirements for fermentation to take place.

 

Fermentation is an anaerobic process that is a way to further oxidize pyruvic acid at the end of the glycolytic pathway.  So, it proceeds without oxygen and uses pyruvic acid as the starting reactant.  A fermentative microorganism will require a constant supply of NADH for reducing various substrates they use to produce products such as alcohols and acids; and it also releases NAD+ to be fed back into the glycolysis pathway.  Fermentation is different from aerobic respiration because it requires organic compounds to serve as the final electron acceptor molecules.

 

 

6.   Is fermentation catabolic or anabolic and why?

 

It actually has components of both.  Part of the process of fermentation includes glycolysis, which is clearly catabolic since it breaks glucose into two molecules of pyruvic acid, and releases ATP through substrate level phosphorylation and NADH⁺ through oxidation-reduction. But by using a broader definition, fermentation is also a means by which various organic compounds such as vitamins, hormones, and antibiotics can be synthesized  Because this involves the assembly of new molecules rather than their breakdown, in this context, it would be considered a type of anabolism.

 

7.   Name three electron carriers and the vitamins that are essential to their function. Explain the actions of the electron carriers and their vitamins in metabolic reactions.

 

Three important electron carriers are NAD, FAD, and coenzyme A.

NAD is composed of adenine, ribose, and the vitamin nicotinamide (niacin). The niacin portion of the molecule serves to remove electrons from substrates in glycolysis and Krebs cycle (forming NADH⁺) and to transfer the electrons to complex I of the electron transport chain.

 

FAD is similar in composition to NAD, but it contains the vitamin riboflavin as the electron carrier part of the molecule.  It is active mainly in the Krebs cycle, where it oxidizes (removes electrons from) succinate, converting it to fumarate.  The FADH₂ formed then shuttles the electrons to complex II of the ETS.

 

Coenzyme A contains the vitamin pantothenic acid and is part of the pyruvate dehydrogenase enzyme complex that converts pyruvate into a 2- carbon acetyl group. The primary function of coenzyme A is to combine the acetyl group with the oxaloacetate at the end of the Krebs cycle, forming the citrate that will begin the cycle.

 

 

8.   Explain the similarities and differences between anaerobic respiration and fermentation.

 

They are similar in that they both take place under anaerobic conditions.  But in most other ways, they are quite different.  The differences include 1)  anaerobic respiration includes glycolysis, Krebs, and the electron transport system and fermentation is based on the glycolytic pathway alone.   2)  Anaerobic respiration uses mostly oxygen-containing inorganic molecules (nitrates, sulfates) as the final electron acceptors, and fermentation uses organic molecules such as acids to accept electrons.  3) Anaerobic respiration yields more ATP per glucose than fermentation, which is limited to 2 ATPs.  4)  Fermentative microbes are often aerobic and can operate with oxygen (respiration) or without oxygen (fermentation), depending on the conditions. Obligate anaerobes are unable to use oxygen gas, and lack enzymes to handle it in this same way.  So, they have an alternate enzyme system for processing the H+ ions, usually by reducing some oxygen-containing salt such as NO₃, CO₂, CO₃, SO₄.  Metabolic water will not be a byproduct of these types of reactions.

 

Chapter 9

 

1.   Explain the diagram.

 

This is a shorthand way of representing the origin of DNA from DNA and the relationships of DNA, RNA, and proteins. This is sometimes called the “central dogma of biology”, since it provides a universal summary of the flow of genetic information.  In general, it shows that that 1) DNA replicates itself to form identical DNA to permit inheritance of the parents’genetic material by offspring, 2) DNA conveys its message to RNA through a special copying process called transcription, and 3) proteins are the products that come from the translation of the message contained in the RNA.

 

2.   (a) Replicate the segment of DNA

 

You would separate the two strands into templates to serve as a guide for making new matching strands. This would involve your bringing in the corresponding bases (actually nucleotides) along the single template strands according to the rules of pairing, thereby forming double strands.  At the end, you would have two new strands that are identical to each other and to the original parent strand.

 

(b) Show the direction of the strands; explain leading and lagging strands.

 

The strands do not orient the same direction, using the order of bonding of the phosphate-deoxyribose backbone as a point of reference.  It is based on the numbering system on the carbons on deoxyribose where the phosphate bonds occur.  On one strand, a phosphate attaches first to the #5 carbon and then to the #3 carbon.  This reads as the 5’ to 3’ direction (‘ means prime).  On the complementary strand, the order is just opposite:  The phosphate attaches first to the #3 carbon and then to the #5 carbon, reading in the 3’ to 5’ direction.  This is called an antiparallel arrangement.

 

This arrangement of the strands impacts how DNA is replicated.  The DNA polymerase III is configured to add bases only in the 5’ to 3’ direction on the new strand, and this so-called “leading” strand is the only one that can be replicated straight through.  The other strand is called the “lagging strand” because it cannot be replicated the same way–its synthesis is discontinuous.  The polymerase will replicate it in small bits, going backwards from 5’ to 3’ to make short fragments that will later be filled in and made complete by another polymerase.  It is important to emphasize here that it is the direction of the new strand being synthesized that is 5’ to 3’.  This means that the direction of the template strand is 3’ to 5’ for the leading strand and the direction of the template strand is 5’ to 3’ for the lagging strand.

 

(c ) How is this semiconservative replication?.

 

The basic way that DNA is replicated is called semiconservative, because it retains or conserves the original template that is half of the parent strand as part of the finished new DNA.  This is one way to ensure that the language of the DNA will remain intact for many generations. Yes the strands are identical to the original.

 

 

3.   (a) Give the mRNA, tRNA, and amino acids that go with this sequence.

 

We need to start by transcribing the DNA triplets shown into complementary codons of RNA, and in this case, it will be mRNA.

 

For instance, TAC will be transcribed into AUG.  Remember that in practice, transcription is very similar to replication, except only one template strand is copied and U will be the mate of A, rather than T.

 

The mRNA codons for this sequence will read

 

AUG GUC UAU GUG AGG GGA CGC UGA

 

The corresponding tRNAs will be anticodons that match the codons:

 

UAC CAG AUA CAC UCC CCU GCG ACU

 

(You will notice that the tRNAs come out similar to the original template strand of

DNA, except that the Ts on DNA are replaced by Us)

 

To determine the correct amino acids, you need to refer to the master code provided in figure 9.13.  Look up each mRNA codon to find which amino acid it codes for.  The resultant peptide will contain:

 

f-methionine, valine, tyrosine, valine, arginine, glycine, arginine, STOP codon, so no amino acid

 

(b)  A different mRNA strand that would give the same sequence:.

 

AUG GUU UAC GUA AGA GGG CGG UAA

 

Because of redundancy, a given amino acid can be specified by different codons.  So, for example, this mRNA sequence will encode the same amino acids as the original one: (note that there is only one start codon–AUG).  For a group of four codons that specify the same amino acid there is a “wobble” effect, in which only the third base is different.  This makes it possible for a mistake in that base to retain the correct amino acid.

 

(c )  Give the amino acid structure for figure 9.15:

 

f-methionine, leucine, proline, glycine, isoleucine, STOP

 

 

 

4.   Describe the actions of all enzymes involved in replication of DNA.

 

During the early phase of DNA replication, the DNA molecule is tightly wound into a bundle of chromatin, and this would be a barrier to replication, so the cell has specialized gyrase that unwinds the coils in the chromatin to convert it to a free strand of DNA.  At this stage, however, DNA is still double stranded and can be replicated only if its nitrogen bases are exposed.  A type of enzyme called a helicase does the job of unwinding the DNA by lysing the hydrogen bonds and creating two separate strands.

 

5.   Because DNA cannot be replicated without a specific starting point, a primase adds a short RNA primer at the origin of replication to initiate the synthesis of a new DNA strand.

 

The synthesis itself (adding of the correct matching nucleotide to pair with the template strand) is carried out by DNA polymerase III.  As it moves along the template it makes the bonds on the new strand as it adds the nucleotides in the 5’ to 3’ direction. It also serves as a proofreading enzyme for locating errors in the replication process.

 

The lagging strand will require additional enzymes to finish up the DNA molecule.  First, DNA polymerase I removes the primers that attach to each Okazaki fragment.  This enzyme also repairs mismatches and gaps.  Then, a ligase that can fill in the missing nucleotides between the fragments adds the correct matching nucleotides.

 

5.   Compare and contrast the actions of DNA and RNA polymerase.

The similarity is that they both synthesize (replicate) a strand of nucleic acid using template strands from an original DNA molecule.  But DNA polymerase must replicate two strands and RNA replicates only one strand (a process called transcription). DNA polymerase also comes in several forms, each of which has a specific function.  DNA poly III’s major function is to add nucleotides to both strands of DNA during replication. Part of replication requires the actions of other enzymes such as a helicase to separate the two strands, DNA poly I to remove the RNA primers and ensure that the correct replacement nucleotides are added, and ligases to fill in missing bonds in the lagging strand.  RNA polymerase does not come in different functional forms and does not require a helicase or ligase.  Because it only transcribes the single template strand that runs in the 5’3’ direction, it does not have to process a lagging strand.

 

6.   Examine the following series of words and identify what type of mutation they represent, starting with the first word in non-mutated form.

Beast–original

Feast–substitution

Breast–insertion

Best–deletion

Beats–inversion

 

 

 

CHAPTER 10.

 

1.   Why do bacteria make restriction endonucleases?

 

Bacteria are constantly being invaded by viruses called bacteriophages in their natural environment.  As a way of reducing harmful effects of DNA from viruses and other sources, they have evolved specialized enzymes that chop up this “foreign” DNA and render it harmless.  The cuts made by the enzyme (a deoxyribonuclease, or nuclease for short) are along the strand of DNA, so it is internal or “endo”.

 

2.   Why do these enzymes not work on their own DNA?

 

They are called restriction enzymes because their effect is restricted only to DNA from other sources and cannot cut the bacterium’s own DNA.  This means that the entire genome of the bacterium will lack the palindrome that goes with that particular restriction enzyme.  (This makes sense if you consider what would happen to them if they randomly chopped up their own DNA).

 

Show the circularization of DNA.

 

To circularize a linear piece of DNA, it would be necessary to locate

identical palindromes that flank the desired piece of DNA at each end.

When the DNA at the sites of this palindrome is exposed to the correct restriction endonuclease, it will cut across the DNA in a way that produces short single DNA strands that hang off the end of the molecule called “sticky ends”.  They are sticky because they will attract another sticky end that has a complementary series of bases.  When these are located at each end of a segment of DNA, they will come together, bind, and form a circle of DNA. This is one way to make a plasmid.

 

 

3.   What advantages are there in using reverse transcriptases for making cDNA?

 

The answer involves the differences in prokaryotic and eukaryotic gene transcription.  Most cloning hosts are prokaryotic (bacteria), and they lack the mechanisms (a spliceosome) that modify the initial mRNA transcript by removing the introns and splicing the exons together.  Often, a DNA version of a processed mRNA transcript is needed for genetic engineering, studying RNA relationships, or sequencing.  It is very difficult to pull the original gene that was transcribed out of a large genome.  A relatively simple answer is first to isolate RNA from an active cell using a microarray and then to expose it to a reverse transcriptase.  This enzyme has the remarkable ability to synthesize a DNA molecule using an RNA as a guide.  Because it is a copy of DNA made from RNA it is termed cDNA.

 

4.   (a) Explain hybridization and Southern blotting.

 

Just as hybrids in biology are the products of two different parents that produce uniquely different offspring, hybrid molecules are two different strands of DNA, RNA, or a strand of DNA and RNA that are similar enough in the order and type of bases, that they unite and form a double strand at complementary sites.  This process is called hybridization.

 

(b)  How do gene probes work?

 

Gene probes are based on the concept of nucleic acid hybridization.  One can construct or isolate oligonucleotides–specific segments of DNA whose exact order of bases is known.  These probes carry some form of reporter or visualizer for locating the sites where hybridization has occurred.  Probes can be used to analyze unknown DNA or even RNA that may contain regions that match.  In fact, a probe is a very fine tool for finding genetic loci on a long segment of DNA.  It is used to identify infectious agents and to locate specific genes.

 

Both concepts of hybridization and gene probes are used in making a Southern blot, named for its discoverer.  A DNA test sample is cut by endonucleases and electrophoresed on a gel.  The gel is transferred to a special filter that can affix the DNA fragments on the gel like a piece of blotting paper (hence the name).  The filter is exposed to specific probes that can hybridize with and identify any complementary sequences. The bands that develop on the filter will be visualized by a photographic film or by fluorescence.

 

 

5.   Use figure 10.7 to predict the template strand of the sequence.

 

Keeping in mind that the shortest fragment will be at the bottom of the electrophoretic gel and also begins the DNA sequence, you should start at the bottom and read up.  So, the template sequence will be TCGGCTAGG.  I know that you can read this from the illustration, but you would not know it from the actual procedure except by reading the nontemplate strand.

 

 

6.   (a) Why is PCR unlikely to amplify contaminating DNA in a sample of human DNA.?

 

The main reason is that the primers used to initiate the synthesis of DNA during the priming phase can be chosen to specifically affix only to a known sequence of human DNA.  We now know a great deal about the sequences of human and bacterial genomes, so this selectivity makes it possible to rule out amplification of non-human DNA. This does not solve the problem of contamination with other human DNA during the procedure, and this problem is solved by ultra clean techniques to prevent this type of lab contamination.

 

 

(b)  How can PCR pick out a specific gene out of a complex genome and amplify it?

 

It takes advantage of the way that DNA can be annealed or denatured with heat.  PCR is a heat-driven process. One the strands are separated, the specific primers can be added to guide DNA synthesis at a specific site on the DNA.  Then, special DNA polymerases that function at high temperatures carry out the replication of the new strand.  The system for amplification involves repeated cycles of annealing, priming, and replication, until a desired number of strands has been produced.  The gene that is amplified will be the one that the primer has tagged.

 

 

7.   If a complete PCR cycle takes 3 minutes, how many strands of DNA would theoretically be present after 10 minutes; after 1 hour?

 

It works on an exponential scale similar to population growth, with each cycle doubling the number of copies.

Eight copies will be present after 10 minutes (3 cycles)

After one hour, there will be 1,048,576 copies (20 cycles)

 

8.   Explain what is involved in genomics, proteomics, and bioinformatics.

 

These are all sciences developed around the topics of molecular genetics and DNA technology. Once biotechnologists started sequencing the genomes of organisms and viruses, there was an enormous explosion in sequence data (the base sequence of the human genome alone would fill a book of 1000 pages).  This kind of data is far too complex and massive to be handled by the usual methods.  It required supercomputers to manage and analyze.  From this need emerged bioinformatics, the marriage of modern genetics, computers, and statistics.

 

Genomics is a subscience of bioinformatics that analyzes the sequence data to determine the locations and functions of various genes and attempts to show similarities and differences among the genomes of organisms.  We can thank genomics for newer identification techniques like that used in the Case Study and a greater knowledge of human diseases.

 

Proteomics is another subscience of bioinformatics that concentrates on the end product of DNA expression–proteins.  Since proteins are largely responsible for the traits that we see in organisms, proteomics can be an important tool in predicting protein structure (amino acid composition and folding properties, for instance) and function (what structure or enzymatic activity a protein actually performs in a cell or virus.

 

10.                For what reasons would gene therapy be more effective performed on embryos than fully developed animals?

 

Gene therapy would be more effective in an embryo than a fully-developed animal because genetic expression is more effective if all of the cells of the organism contain the newly implanted normal gene.  Delivering the genes into all of the cells of an embryo is far less complicated than attempting to get the genes into just the cells and tissues that are most affected by the defective gene.  For example, implanting the cystic fibrosis gene into the lungs or the muscular dystrophy gene into muscle cells has not been particularly successful.  However, experiments with animals have shown us that embryos can easily be induced to accept and express these genes as mature adults.

A large problem in using embryonic therapy with humans is the moral and ethical concerns with the manipulations of human zygotes or embryos (consider the enormous controversy over cloning humans or stem cell research).   Many groups are in total opposition to using human embryos for any scientific purposes, yet germ line engineering (altering the genes of embryos) will require research with hundreds of thousands of embryos before it could ever become a routine process.  After all, there is no “artificial uterus” that can carry these experimental embryos, so it involves much more than just the embryo.   Even if a way to use human embryos becomes available or occurs in countries with more relaxed laws, the success of introducing the gene and its effectiveness are not guaranteed.  In addition, the gene must go into the nucleus of the cell and become inserted into a precise position on its chromosome.   If it fails to become inserted or inserts in the wrong place, any number of problems will arise. Right now the genetics of development is still in the earlier phases of understanding, and all of the possible effects of “foreign” genes on development are not yet known.

 

11.                Explain the basics of DNA STR profiling for identification of unknown DNA.

 

Single tandem repeats (STRs) are short DNA segments (2 to 8 nucleotides long) interspersed throughout the human genome.  They occur in a repeating pattern that is inherited from parents like other genetic material.  Because the number, distribution, and types of STRS in a given person’s DNA are unique to them, STRs are useful for DNA profiling and identification.

 

The first step in this profiling technique is to release the nuclear DNA from a sample of cells. The isolated DNA molecules must next be amplified with PCR to produce millions of amplified fragments.  During PCR the fragments are tagged with specific fluorescent primers that hybridize with the STRs on the fragments.  This is the key step that allows for tracing the pattern of STRs by scanning for areas of fluorescence. Next the DNA fragments are separated by electrophoresis into tiny fluorescent bands that are detected by a laser and displayed as a series of colored peaks that constitute the profile. This profile may be entered into a data base and used for matching with possible suspects and forensic evidence.

 

CHAPTER 11

 

1.   What is wrong with this statement and what is a more correct way to say it.

 

It is a mistake in terminology to use the term “sterilize” when the process being used could not possibly kill all microbes present in a location such as the skin.  The term sterilization means that all microbes are destroyed or removed and it is usually for describing a technique such as an autoclave that is used on inanimate objects.  Any method that could kill all of the microbes on the skin would severely damage it.  Another problem with the statement is that alcohol is not a sterilizing agent.  The correct terms to use are degerm or antisepticize the patient’s skin.

 

2.   Would it be proper to say a person has been disinfected? Explain.

 

Not really.  The term disinfection denotes the use of a method to kill vegetative cells on inanimate objects.  It involves heat or strong chemicals that, again, would be too detrimental for use on the human body.

 

 

3.   Why are antimicrobials inhibited in the presence of organic matter?

 

Biologic matter such as blood, sputum, feces, and tissue fluids can form a

barrier around microbes, especially if dried.  This can prevent the chemicals

from having the close contact with microbes required to kill them.  In some   cases the organic material may cause chemical changes in the antimicrobial compound that render it less effective.  It also is known to inhibit the mode of action of disinfectants and even heat.  It is for this reason that instruments are cleaned or sanitized before treatment.

 

 

4.   Give three situations in which the same microbe would be considered a serious

contaminant or a harmless one in microbial control.

 

Because we are constantly in contact with microbes, it is essential to understand circumstances that require stringent controls and those that do not.

 

Microbe # 1 is Escherichia coli.  If present in the toilet bowl it is probably just a harmless contaminant.  If present in drinking water or salad, it can cause serious infections and disease.

 

Microbe #2 is Clostridium perfringens spores.   If present in garden soil and carried on the bottom of shoes, it causes little harm.  If it is present in a syringe that is injecting drugs, it can cause serious disease.

 

Microbe # 3  is the fungus Fusarium, a common mold on plants.   It is widely carried by the air in human-frequented habitats, where it is basically harmless.  But if it gets onto contact lenses or into contact lens solutions and is inserted into your eye, it can cause a severe corneal infection.

 

 

5.   Explain what features of endospores make them so resistant to microbial control and which sterilizing techniques would destroy them.

 

Endospores are compact survival units made by certain bacteria that are considered the most resistant cells on earth. They can withstand extremes of heat, drying, freezing, radiation, and chemicals.  During development, they form several protective coats and a central cortex that are difficult to penetrate.  Within this tight packet, the water is removed and a salt is laid down that blocks the effects of heat. Being metabolically inactive and in a state of dormancy also decreases their sensitivity to antimicrobial agents.

6.   Describe some problems in sterilizing delicate instruments.

 

A number of instruments have to be reused on patients because of their high replacement cost. These include endoscopes, dental handpieces, special syringes and needles for bone marrow sampling, respirators, and heart-lung machines.  Most of these types of instruments enter the patient’s body in a way that they can become contaminated with secretions, excretions, or blood and so will require sterilization after use.  Many of them have components that are made of plastic or other heat-sensitive materials.  They also have surface features which can harbor contaminants.

 

The goal is to kill the most resistant bacteria, spores, and viruses without damaging the instrument.  For most applications, these instruments are too delicate for heat to be used, so autoclaving is out of the question.  Radiation could be used, however, it requires very high tech machinery and does not provide a rapid turnaround time. The sterilization techniques that are most practical for these instruments is based on chemicals called sterilants.  These include hydrogen peroxide, glutaraldehyde, and ethylene oxide gas circulated within closed cabinets or chambers.  Usually the items need to be precleaned to remove any biologic wastes and then exposed from 1-3 hours, depending on the particular sterilant.

 

 

7.   Personal reactions to radiation sterilization and chemical sterilization of food.

 

This answer is going to vary with the person, however, if one looks at the scientific evidence, irradiated food has been shown through long years of research and usage to be harmless, and it is certainly safer to eat from the microbiological standpoint.  It is an indispensable method to preserve food that must be stored for long periods and it serves as the primary food source for the military and astronauts on assignment.  Objections usually include changes in nutrition, taste, and texture that make the food unappealing.

 

 

CHAPTER 12

 

 

1.   Use the diagram to explain the three interacting factors in drug therapy.

 

When a drug is given to treat or prevent an infection, there are always three factors to consider:  the patient (host) who has the infection, the infectious agent or microbe, and the nature of the drug itself.  The point in the center where all three circles come together represents a successful treatment outcome.  However, due to complex interactions between the host, drug, and microbe, there can be other consequences besides a cure.

 

Host/drug interactions

The drug can interact with the human in adverse ways.  This violates a basic tenet of drug therapy–the drug needs to be selectively toxic against the microbe.  Adverse reactions are a common side effect of drug therapy.  In addition, a patient can respond to the drug because it is a foreign molecule that causes hypersensitivity.  Humans might also break the drug down or excrete it before it has a chance to act.  The drug may not be soluble in host fluids and fail to penetrate into organs where the infection has traveled.

 

Drug/Microbe interactions

The drug has a mode of action that works on a structure or function of the microbe.  It is preferable for it to kill rather than just inhibit the microbe. Though it is necessary for the microbe to be sensitive to the drug, sometimes the microbe may have developed resistance by breaking the drug down, pumping it out, or changing structure or physiology.  Some drugs destroy the normal microbiota and cause superinfections

 

Microbe / host interactions

The microbe can release toxins and other substances that interfere with the immune system or that are unaffected by the drugs.  Microbes can grow in biofilms that prevent the drug from penetrating and acting on the infectious agents. The host’s immune system plays an important role in the removal of the infectious agent and final cure.

 

 

 

2.   Refer to Table 12.4 and explain the modes of drug action and how this affects the specificity of the drug and its spectrum.

 

The table is separated into microbial groups, drug groups, and the primary targets and modes of action. In general, a mode of action directed towards a specific target will often reduce the spectrum, and a mode of action directed towards a target found in more microbes will produce a broader spectrum.

 

For antibacterial drugs, the primary drug actions are to:

 

1) Block cell wall synthesis, with the result that the cells will lyse.  These drugs tend to act on the formation of peptidoglycan, which will make them quite specific in their effects, since only bacteria have this molecule in their cell walls.  The spectrum varies from narrow (bacitracin) to broad according to how well the drug penetrates the outer membrane of gram negative bacteria.  For example, the original penicillin is narrow spectrum and works primarily on gram positive bacteria, but newer forms have been synthesized to have a broader spectrum and include gram negative bacteria.

 

2) Stop prokaryotic ribosomes from working properly, which will prevent cell synthesis and division.  This action has the potential to affect most bacteria, regardless of gram reaction, so these drugs tend to be broader spectrum.  And indeed most of the drugs with this mode of action (tetracyclines, chloramphenical, erythromycin)  are effective in both gram positive and negative infections.

 

3) Inhibition of DNA gyrase, which stops DNA replication and cell division.  This also has the potential to affect most bacteria, since all of them require a functioning gyrase.  Examples of these drugs include the fluoroquinolones such as ciprofloxacin, which have broad spectrum actions.

 

Most antifungal drugs break down cell membranes and lyse the cells, so they are fungicidal, and they tend to be effective on a broad spectrum of fungal pathogens.

 

Antiviral drugs have several modes of action, and each one is relatively specific to just a single virus group such as influenza, herpes, or human immunodeficiency viruses.  None of the drugs act on a universal structure found in all viruses, so there are no broad spectrum antiviral drugs.

 

3.   (a) Word used to describe drug therapy given pre- or post-exposure to prevent an infection.

 

The term used for this practice is prophylaxis.

 

(b)  What is the purpose of this treatment?

 

It is a form of preventive therapy given to people who have been or could possibly be exposed to an infectious agent, and are more vulnerable to infection as a result.  The thought behind this “pretreatment” is that there will be an antimicrobic drug already in the patient’s system before the pathogen can establish an infection.  Examples would be surgical patients and dental patients who could develop an infection due to medical procedures, and healthy people given medications during outbreaks of infections or epidemics such as meningitis or influenza.

 

(c ) What are some undesirable side effects?

 

Like any drug therapy, it could damage the patient’s organs.  The side effects of some prophylactic drugs can be so disabling as to not be worth it, as we saw with the nurse in the case file.  This type of therapy can also disrupt normal microbiota leading to superinfection.

 

 

(d)  Define probiotics and explain their use.

 

Probiotics are foods or supplements that contain actual pure or mixed cultures of microorganisms thought to have beneficial effects in various body sites.  Most probiotics are bacteria normally found in the intestinal microbiota.  The idea behind these preparations is that the ingested microbes will establish themselves as a mixed population of residents that will stabilize and balance the intestinal environment and displace pathogens.   Studies have shown that probiotics can also boost immunities.

 

 

4.   Summarize the concerns in drug therapy, including resistance, allergies, superinfections, and other adverse effects.

 

Besides the intended beneficial effects of antimicrobial drugs, the complexity of therapy means that unintended or undesirable effects can also occur.

 

Many drugs have adverse toxic effects.  These often occur because their mode of action harms parts of the human cell as well as the microbe.  Nearly any organ or system can be damaged by drugs, but some of the most common effects are on the liver, kidney, nervous system, intestinal tract, bone marrow, and skin.  For example, amphotericin B used to treat fungal infections is very damaging to the kidney and isoniazid used to treat tuberculosis is damaging to the liver.

 

Superinfections occur when broad spectrum antimicrobials are administered for infections.  These tend to affect a much wider range of microbes, and are not specific just for the pathogen being treated.  As a result, beneficial microbes in the normal microbiota can be destroyed.  This in itself is a problem, but what may also happen is an overgrowth by hidden pathogens that survive the treatment and create another, often more severe infection called a superinfection.  This is what happens with many yeast infections and some types of colitis.

 

About 5% of people have an allergic reaction or hypersensitivity to antimicrobial drugs.  This is caused by them having been sensitized to a drug when they were first given it, and then upon a later exposure, they react allergically by developing a rash, respiratory attack, or gastrointestinal reaction.

 

We will probably always have to administer drugs for infections, but one of the down sides of this is that eventually, microbes adapt to them and become drug resistant.  In fact, the combination of drugs as an environmental selective agent and the microbe’s abilities for genetic change and evolution, drug resistance is an expected outcome.  This is a continuing problem that continues to plague the medical establishment and will probably do so for many years.  A push is on to improve drug usage, drug research, and a worldwide misuse of drugs.

 

5.   (a) What is the basis for combined therapy?

 

Combined or combination therapy is treatment with 2 or more drugs simultaneously.  The rationale behind this therapy is the reduced likelihood that a microbe will have simultaneous resistance to more than one drug.  So we assume that if one of the drugs doesn’t work, then the other drug will work.  This means that the microbe will die one way or another, and even better, any resistance it had will be gone as well. Occasionally, several drugs will be necessary to make sure that no drug resistant forms persist.  Examples are tuberculosis and HIV infections, both of which show extreme tendencies for drug resistant.

 

(b)  Why is it useful in treating HIV infection?

 

Because HIV can rapidly mutate to a drug resistant strain right in the patient, a cocktail of at least 3 drugs is commonly given, and each drug interferes with a step in the viral cycle.  Most of them act on enzymes that HIV needs to complete DNA synthesis, integration, and viral assembly.

 

Another reason to give combined therapy is that drugs can have a synergistic effect, meaning that together they are more effective than either drug alone. An example is sulfa-trimethoprim, a two-drug combination that works on two different steps in the same metabolic cycle.

 

 

 

6.   Explain the kinds of tests that would differentiate between a broad and narrow spectrum antibiotic.

 

One experiment that could be done in the lab is to take a wide variety of gram-negative and gram-positive species of bacteria and perform disc diffusion tests with a range of drug choices from several drug classes.  Analysis would involve measuring results for all combinations of bacteria and drug.  A drug with broad spectrum effects would show effectiveness (have a large zone of inhibition) for the majority of test bacteria, both gram-positive and gram-negative.  If the test shows drugs that work on some gram positive and negative bacteria but not all, these would be considered medium or moderate spectrum.  The narrow spectrum drugs would be effective only on gram-positive species or gram-negative species, but not both.   If it is important to determine a more detailed spectrum that includes obligate parasitic bacteria such as rickettsia and chlamydia, disc diffusion tests would have to be done in experimental animals or cell culture.

 

 

7.   Summarize the causes of the medical dilemma we have in using antimicrobial drugs.

There has been an ongoing problem with these drugs since the very first one (penicillin G) was introduced.  Their use as a cure-all for any and all medical conditions has become a worldwide problem.  First, they are overprescribed, often given for viral infections and respiratory illnesses that have not been diagnosed.  Remember that most antimicrobials and all antibiotics are useless against viruses.  Many infections are treated by guesswork or by using a broad spectrum drug, rather than by identifying the infections agent and doing antimicrobial sensitivity testing.  This is responsible for an increase in unnecessary toxic effects and superinfections.  Drugs are misused in many countries, being added to animal feed or being sold over the counter without medical supervision.

 

Much of the overprescription and overuse of drugs leads to resistance, so that a number of drugs that were once effective no longer are.  We are seeing more and more episodes in medical settings in which there are few drug choices left for treatment.  Drug companies are in a race to develop new antimicrobial drugs, but the microbes have the lead, and we are racing to keep up.

 

 

CHAPTER 13

 

 

1.   What are the important clinical implications of positive blood or cerebrospinal fluid?

 

There are locations in the human body that are maintained in a sterile state for normal function.  Two of these places are the blood circulation and the fluid that bathes the brain and spinal cord.  They do not harbor microbiota.  It is a given that if a culture of these substances comes out showing positive growth in culture media, this is a sign of infection and must be dealt with.

 

 

2.   The expected result if a compromised patient is exposed to a true pathogen?

 

This scenario brings together the most serious possible combination. Since a patient who is predisposed to infection because of a compromised immune system is highly vulnerable even to weakly pathogenic microbes, then being in contact with a primary pathogen could be deadly.   Such infections become widely systemic and reach the period of invasion very rapidly.  A good example of this combination would be AIDS patients (without treatment).  Because HIV destroys part of the immune system, they are already attacked by a wide variety of opportunists.  But when they acquire infections such as tuberculosis or toxoplasmosis, the pathogen invades very rapidly and the patient becomes gravely ill and may die (discussed in case file 12).

 

 

3.   Explain how endotoxin enters the blood of a patient with endotoxic shock.