BIOL 2051 - MOCK EXAM #1

Under what environmental conditions would a bacterium prioritize endospore formation over cell division?

Bacteria, such as Bacillus and Clostridium species, initiate sporulation in response to conditions that threaten vegetative survival under: Extreme heat, desiccation, harmful chemicals (antibiotics/disinfectants), radiation, and UV exposure) trigger the need for protection

why does endospore formation not result in a new generation of cells?

Endospore formation is a differentiation process, not reproduction - Instead of symmetric binary fission (one cell split into two identical daughter cells), the bacterium divides asymmetrically, producing a large "mother cell" and a small "forespore" to aid in survival process

Name an archaea that thrives in high-salt environments and is an extremophile.

Halobacterium salinarum

Name the world's largest bacteria.

Thiomargarita magnifica

Name a bacterium discovered by Thomas Brock - found in hot springs!

Thermus aquaticus

A bacterial cell acquires a plasmid containing a gene for tetracycline resistance. For the cell to produce the tetracycline resistance protein, which of its native cellular machinery are essential?

RNA Polymerase (RNAP), Ribosomes (30S and 50S subunits), Transcriptional and Translational Machinery, ATP and GTP

what is the function of RNA Polymerase (RNAP)?

Essential for transcribing the tetracycline resistance gene (tet gene) located on the plasmid into messenger RNA (mRNA).

what is the purpose of Ribosomes (30S and 50S subunits)?

Essential for translating the mRNA into the tetracycline resistance protein. The ribosome must be functional, as tetracycline typically targets the 30S subunit to inhibit this process

what is the function of Transcriptional and Translational Machinery?

This includes sigma factors for RNAP to initiate transcription, initiation factors, elongation factors, and tRNA for translation

A student claims that sporulation in Bacillus is a type of mitosis. This claim is incorrect primarily because:

The student's claim that sporulation in Bacillus is a type of mitosis is incorrect primarily because it involves asymmetric cell division (rather than symmetric), resulting in two distinct compartments—a smaller forespore and a larger mother cell—that possess different cell fates and distinct gene expression profiles, whereas mitosis produces two genetically identical, identical-sized daughter cells. Additionally, sporulation is a survival response to environmental stress, not a mechanism for reproduction/cellular proliferation.

Name the smallest bacteria that lack a cell wall.

Mycoplasma genitalium

Name a bacterium that is extremely resistant to radiation damage.

Deinococcus radiodurans

Primary fate of NADH in fermentation

Regenerates of NAD+

Unicellular eukaryotic organism (Saccharomyces)

Yeast

"Practical use" for fermentation

Yogurt, Bread, Beer

Polysaccharide produced by Streptococcus mutans

Dextran

Initial stage to both fermentation and cellular respiration

Glycolysis

True or False: The oxidase assay specifically tests for cytochrome c oxidase, not all terminal oxidases.

TRUE. The oxidase assay (using reagent: tetramethyl-p-phenylenediamine dihydrochloride) specifically tests the presence of cytochrome c oxidase - bacterial colony (that synthesizes cytochrome c oxidase) turns dark blue/purple when the reagent is added, but it is not oxidized by other terminal oxidases

True or False: Denitrification depletes soil nitrogen by converting usable nitrates into nitrogen gas (N2), which escapes into the atmosphere and is unavailable to most plants

TRUE. Denitrification is a microbial process that reduces nitrate (NO3) to nitrogen gas (N2) and other gaseous compounds (such as N2O). This process converts soluble, plant-usable nitrate into inert, insoluble nitrogen gas that escapes into the atmosphere, making it unavailable for most plants.

True or False: In a Peptone Iron Agar deep (containing peptone), an animal digest protein source rich in the substrate, cysteine to demonstrate the production of hydrogen sulfide.

TRUE. Peptone Iron Agar is used to detect hydrogen sulfide (H2S) production by microorganisms. Peptone Iron Agar contains peptone (specifically peptic digest of animal tissue), which are rich sources of cysteine. Certain bacteria possess the enzyme cysteine desulfhydrase, which acts on the cysteine amino acid to release H2S gas. The medium contains ferric citrate, which react with the H2S produced to form ferrous sulfide (FeS), a black precipitate, confirming a positive test.

True or False: Glycosidic bonds in cellulose, starch, and glycogen contain a specific covalent bond that is cleaved by proteases.

FALSE. Proteases are enzymes that specifically hydrolyze peptide bonds in proteins. They are not responsible for breaking the carbohydrate-based glycosidic linkages found in glucose polymers like starch and cellulose

Amylase producers that are Gram-positive aerobic species

Bacillus subtilis

Gram-negative, motile, Nitrate reductase producers causing UTI infection

Escherichia coli (E. coli); {But there are reports on other gram negative uropathogens species like Proteus mirabilis}

Chymosin, used in cheesemaking, is given as an example of which type of enzyme?

Its primary function is that it’s a Proteolytic enzyme (Protease) – caseinase

A clinical microbiologist needs to differentiate between Staphylococcus and Streptococcus. Which test is most appropriate and why?

Catalase Test; Staphylococcus species produce the enzyme catalase, while Streptococcus species do not. When a small amount of bacterial colony is mixed with 3% hydrogen peroxide - Staphylococcus immediately bubbles (catalase-positive) due to the release of oxygen, whereas Streptococcus does not bubble (catalase-negative).

How does the Cry toxin produced by Bacillus thuringiensis (Bt) kill targeted insect pests like the cotton bollworm?

When an insect pest, (cotton bollworm larvae), consumes plant tissue containing inactive Bt crystals (Cry protein), the alkaline environment of its midgut activates the crystals into Cry toxins. These activated toxins bind specific receptors on the larvae’s gut lining and create pores or holes in the gut wall. This leads to cell lysis, gut leakage, and ultimately resulting in the death of the pest.

How do Small Acid-Soluble Proteins (SASPs) physically change the structure of DNA to protect it from environmental damage?

During sporulation, SASPs saturate and bind non-specifically to the bacterial DNA in minor grooves, causing it to condense and become more rigid. This binding alters the DNA's geometry, twisting it from its standard B-form helix into an A-like conformation. The A-form DNA transformation is shorter, wider and more compact structural form, combined with the core's dehydrated state, protects the genetic material from UV radiation, heat denaturation, and chemical or enzymatic damage.

what is the role of the sigma factor in bacterial transcription at the promoter recognition point?

They directly identify and bind specific -35 bp and – 10 bp DNA promoter consensus sequences upstream of the mRNA transcription start site

what is the role of the sigma factor in bacterial transcription at the transcription initiation point?

By binding the core RNA polymerase, they ensure transcription starts at the correct location.

What is the primary role of sigma factors in bacterial transcription?

Sigma factors facilitate the separation of double-stranded DNA to form an "open complex" (transcription bubble), allowing the template strand to be accessed and Prokaryotes use multiple sigma factors (e.g., housekeeping, heat shock, starvation) to regulate specific regulons, allowing rapid adaptation to new environments.

what is the role of sigma factors in bacterial transcription at dissociation?

After initiation, the sigma factor usually dissociates from the RNA polymerase, allowing elongation to proceed.

How does the enzyme Chloramphenicol Acetyltransferase (CAT) render the antibiotic chloramphenicol incapable of inhibiting bacterial protein synthesis?

This CAT enzyme inactivates the antibiotic by transferring an acetyl group from acetyl-CoA directly onto the chloramphenicol molecule. This enzymatic modification changes the drug's structure so it can no longer bind to its target on the 50S ribosomal subunit (or large ribosomal subunit in prokaryotes), thereby preventing it from inhibiting bacterial protein synthesis.

How does penicillin weaken the bacterial cell wall to cause lysis?

Penicillin binds to and inactivates Penicillin-Binding Proteins (PBPs), such as transpeptidase enzymes, which are responsible for cross-linking the peptidoglycan layer of the cell wall. By inhibiting this transpeptidation reaction, the antibiotic prevents proper cell wall synthesis and weakens the existing bacterial cell wall structure. The compromised cell wall cannot withstand the bacterium's high internal osmotic pressure, leading to osmotic lysis.

What is the target enzyme of the oxidase test?

the presence of the endoenzyme cytochrome c oxidase

what is cytochrome c oxidase role in metabolism?

It is part of the membrane electron transport chain used by organisms that produce energy through aerobic respiration. Its specific role is to reduce molecular oxygen (the final electron acceptor) by transferring electrons to it.

Why must the oxidase test results be read immediately (within seconds)?

Results must be read immediately (within seconds) because the oxidase reagent (tetramethyl-pphenylenediamine dihydrochloride) can be oxidized by the air if exposed for too long. This leads to a color change regardless of the presence of the enzyme, resulting in false positives.

Name one genus that is oxidase-positive

Oxidase-Positive Genera: Neisseria, Moraxella, Pseudomonas, or Vibrio

Name one genus that is oxidase-negative

Oxidase-Negative Genera: Acinetobacter, Escherichia, or Enterobacter

What is the chemical reaction catalyzed by the enzyme catalase?

The enzyme catalase breaks down hydrogen peroxide (H2O2) into water (H2O) and molecular oxygen (O2).

In the catalase test, what visual sign confirms a positive result?

A positive result is confirmed by the release of oxygen, which is visible as bubbles immediately after adding hydrogen peroxide to a colony or smear.

Which specific microbe is a clinical example of a catalase-positive organism?

Examples of Catalase-Positive Organisms: Specific clinical examples include species of the genera Staphylococcus (e.g., Staphylococcus aureus or Staphylococcus epidermidis) and Micrococcus.

What is the substrate for the gelatinase test?

gelatin

what is the chemical nature of gelatin?

Gelatin is a protein extracted from the connective tissue and bones of animals. It consists of long chains of amino acids joined by peptide bonds.

How does a positive result appear on a nutrient gelatin agar plate after adding a developer?

A positive result appears as a clear, transparent area (zone of hydrolysis) around and under the gelatinase-producing colony. This clearing becomes visible after the addition of the developer (1N HCl) because the enzyme has hydrolytically cleaved the protein into shorter peptides, meaning there is no longer intact gelatin in that area to precipitate.

What reagent is used to precipitate unhydrolyzed gelatin to make a zone of hydrolysis visible?

1 N hydrochloric acid (HCl); HCl reacts with the unhydrolyzed gelatin remaining in the agar, causing it to precipitate and turn the agar opaque (faint white). Wherever around or below the colony producing gelatinase, gelatin has been hydrolyzed by the enzyme, no precipitate forms, leaving the clear zone

What specific medium is used to test for the production of proteases like caseinase?

The specific medium used is milk agar. This is composed of nutrient agar supplemented with sterile skim milk, which provides the substrate casein.

Why is the presence of a colony alone insufficient to confirm casein hydrolysis?

The presence of a colony alone is insufficient because some microorganisms can grow on milk agar without producing caseinase. A positive result requires the presence of a clearing (zone of hydrolysis) around and under the colony. This clearing indicates that the organism has secreted the extracellular protease caseinase to hydrolyze the large, opaque casein proteins into smaller, transparent peptides and amino acids.

Which enzyme is responsible for the hydrolysis of starch?

The enzyme responsible for the hydrolysis of starch is amylase (specifically alpha-amylase). It is an extracellular enzyme that breaks the alpha (1→4) glycosidic bonds between the glucose subunits in the starch polysaccharide

What reagent is flooded over a starch agar plate to visualize the results?

The reagent flooded over the starch agar plate to visualize results is iodine. Iodine reacts with any remaining starch in the agar to produce a blue-black or brown color.

Describe the appearance of a negative result for starch hydrolysis.

A negative result for starch hydrolysis appears as a blue-black or dark brown color in the medium surrounding the bacterial growth after the addition of iodine. This dark coloration indicates that the starch has not been broken down and is still present in the medium.

What is the function of the Durham tube in a nitrate broth culture?

The Durham tube is used to trap gas produced during the process of denitrification. If an organism reduces nitrate (NO3 - ) all the way to nitrogen gas (N2) example by Paracoccus denitrificans, the gas will appear as a bubble inside the small, inverted Durham’s tube.

Why is zinc powder added to nitrate broth tubes for nitrate reduction that do not change color after adding reagents alpha-naphthylamine acetate and sulfanilic acid?

Zinc powder is a reducing agent that is added to determine if unused nitrate is still present in the tube. It chemically reduces any leftover nitrate (NO3 - ) to nitrite (NO2 - ). If the tube turns red after adding zinc, it means the zinc did the work that the bacteria could not, confirming that nitrate was never reduced by the organism and organism is nitrate reductase negative.

A tube turns red only after adding zinc. Is the organism positive or negative for nitrate reduction?

If the tube turns red only after adding zinc, the organism is negative for nitrate reduction; The red color indicates that nitrate was still present in the medium until the zinc was added. This proves the organism did not produce nitrate reductase to reduce the nitrate to nitrite or any other products.

What is the black precipitate formed in a positive Cysteine Desulfhydrase test?

The black precipitate is ferrous sulfide (FeS). It forms when the gas produced by the bacteria reacts with the ferric citrate (the indicator) present in the Peptone Iron Agar medium.

Which gas is released during the degradation of the amino acid cysteine?

The gas released during the degradation of the amino acid cysteine is hydrogen sulfide (H2S). This occurs when the enzyme cysteine desulfhydrase cleaves the sulfur-containing amino acid cysteine into hydrogen sulfide, ammonia, and pyruvic acid.

what is the purpose of phenol red?

Detects acid production from carbohydrate fermentation. It appears orange-red at neutral pH, turns yellow in acidic conditions (below pH 6.8), and bright pink/fuchsia in alkaline conditions (above pH 8.2).

phenol red is used as an indicator in what assay/mediums?

Fermentation Test (Carbohydrate Broths) & Mannitol Salt Agar (MSA)

what is the purpose of bromocresol green?

Indicates the relative number of Lactobacilli in saliva. It is blue-green at pH 4.8 and changes to yellow as the pH drops due to lactic acid production from glucose fermentation.

bromocresol green is found in what assay/medium?

Snyder's Test Agar

what is the classification of Mannitol Salt Agar (MSA)?

Selective & Differential

what makes Mannitol Salt Agar (MSA) selective?

Contains 7.5% NaCl, which inhibits the growth of most bacteria while favoring salt-tolerant organisms like Staphylococcus .

what makes Mannitol Salt Agar (MSA) differential?

Contains the carbohydrate mannitol and the pH indicator phenol red to distinguish mannitol fermenters (turn agar yellow) from non-fermenters.

what is the classification of blood agar?

Differential & Enrichment

what makes blood agar differential?

Contains 5% sterile sheep blood to differentiate bacteria based on their hemolytic properties (alpha, beta, or gamma hemolysis)

what makes blood agar enrichment?

It is a highly nutritious medium that supports the growth of fastidious organisms like Streptococcus.

whats the classification of Snyder's Test Agar (STA)?

Selective & Differential

what makes Snyder's Test Agar (STA) selective?

Has a low pH (4.8) to inhibit the growth of most bacteria while favoring acid-tolerant Lactobacilli .

what makes Snyder's Test Agar (STA) differential?

Contains glucose and the pH indicator bromocresol green; acid production from fermentation turns the medium from blue-green to yellow

what makes Sabouraud Dextrose Agar (SDA) selective?

Has a low pH (5.6) that favors the growth of fungi (yeasts and molds) over most bacteria.

what is the classification of Sabouraud Dextrose Agar (SDA)?

selective

what is beta hemolysis?

Beta hemolysis is characterized by the complete destruction of red blood cells and the total decolorization of hemoglobin in the medium.

what is the visual appearance of beta hemolysis?

a clear, transparent zone in the agar surrounding and under the bacterial colony. You can typically see through the plate when held up to a light source in these areas

what is an example of beta hemolysis?

Streptococcus pyogenes (the causative agent of strep throat)

What causes the greenish discoloration seen in Alpha (α) hemolysis?

The greenish-brown discoloration is caused by the partial destruction of red blood cells and the oxidation of hemoglobin to methemoglobin.

what is the appearance of alpha hemolysis?

Unlike beta hemolysis, the medium does not become transparent. Instead, it develops a cloudy, greenish zone around the bacterial colonies.

what is an organism example of alpha hemolysis?

Streptococcus mutants and Streptococcus sanguinis - species of the "Viridans" group of streptococci

A Snyder tube changes from blue-green to yellow. What does this indicate?

A color change from blue-green to yellow indicates acid production by oral bacteria (primarily Lactobacilli) through the fermentation of glucose. Clinically, this suggests a susceptibility to dental caries (cavities). The faster the color change occurs (e.g., within 24–48 hours), the higher the relative number of lactobacilli in the saliva and the greater the risk of tooth decay.

Which organism is typically Mannitol Positive (turns the agar yellow) on MSA?

Staphylococcus aureus is the clinical organism typically mannitol-positive. It ferments the mannitol in the medium, producing acid that changes the color of the agar from pink/red to yellow.

In the lab example of commensalism, how does S. epidermidis help C. sporogenes?

Staphylococcus epidermidis (a facultative anaerobe) helps Clostridium sporogenes (an obligate anaerobe) by removing oxygen from the environment; S. epidermidis uses up the available oxygen during its own aerobic respiration. This creates the anaerobic (oxygen-free) conditions necessary for C. sporogenes to grow in an environment that would otherwise be toxic to it

What is a positive result for the coagulase test?

the formation of a clot or solidification of the plasma in the tube

which microbe is the target for the coagulase test?

The primary target for this test is detect pathogenic Staphylococcus aureus. The test is used to distinguish the pathogenic S. aureus (which produces the enzyme coagulase to form protective clots) from other non-pathogenic staphylococci (like S. epidermidis) which are coagulase-negative; Coagulase is an enzyme produced by certain bacteria that functions as a virulence factor. It breaks down fibrinogen (a soluble protein in plasma) into short strands of fibrin. The fibrin strands interact to form a fibrin clot. In a clinical infection, this clot, protects the bacteria from the host's immune defenses.