Lab Practical 2 Flashcards
What is transformation?
Transformation is the uptake of naked DNA by a bacterial cell
What promoter controls the GFP gene?
Transcription of the GFP gene is controlled by the promoer for arabinose metabolism.
What factors effect transformation of bacteria?
Methods of transformation
- The method of preparation of competent cells
- the length of time of heat shock
- temperature of heat shock
- incubation time after heat shock
- growth medium used and various additives
- Bacterial colonies’ phase of growth
- Size and state of the foreign DNA
What is meant by competency?
Competency is the relative ability of a bacterial cell to incorporate naked DNA (plasmid) and undergo transformation.
Type of medium that contains reagant that will change color or react due to the growth of metabolism of bacteria.
Differential Media
What type of medium is this?
Ex: The reagants in MaConkey Agar (MAC) will turn bacterial colonies pink if they are able to ferment lactose.
Differential Media
What is the purpose of a selective marker?
Selectable markers show the success of a transfection or other procedure meant to introduce foreign DNA into a cell. It is a technique in gene targeting and gene knockout.
Ex: Beta-lactamase which confers ampicillin resistance to bacterial hosts.
What is the purpose of the heat shock step?
Heat shock forms transient pores in the bacteria’s cell wall, providing a path for the plasmid (foreign DNA) to approach the bacterial cell more readily.
Why are cells held on ice prior to heat shock?
What structure is commonly used by bacteria for motility?
Flagella are a common physiological feature of motile bacteria
Are all bacteria motile?
No but whether a bacterium is motile or non-motile can be a useful characteristic for differentiation
You have a suspected gram-positve culture. Would it be more appropriate to grow it on MacConkey agar or PEA agar to confirm these results?
PEA agar, because PEA agar is selective for Gram- positive bacteria. Because the growth of Gram-negative bacteria is suppressed by the interaction of the alcohol with the outer membrane, suppressing growth.
MAC is selective for Gram negative bacteria
In addition to selecting for halophilic bacteria, MSA can detect the fermentation of mannitol. What color will the plate turn if mannitol is fermented?
Yellow
If the halotolerant organism is able to ferment mannitol, the acids produced will cause the phenol red in the medium to turn yellow.
You disover that an unidentified bacterium you have been given is Gram-negative. Would you use MAC, PEA, or MSA to determine if the bacterium ferments lactose?
MAC, because MacConkey Aga is selective for Gram-negative bacteria and differential in lactose fermentation.
- Not:
- PEA is selective only for Gram- postive bacteria
- MSA is selective for Gram-negative but differential for mannitol fermentation
If an organism does not grow on MacConkey, what conclusions can you draw regarding its ability to ferment lactose.
No conclusions on the bacteria’s ability to ferment lactose can be drawn, since MAC is a selective and differential test.
- If an organism does not grow on the MAC agar, no conclusions can be drawn regarding lactose fermentation because selective assay takes precedence over the differential assay.
When using selective media, why must a culture also be grown on TSA?
The TSA plate acts as a positve control in order to compare and differentiate among the enzymatic processes of bacteria, different media (such as selective and differentail assays) to detect metabolic and physiological differences
What are the selective reagants in PEA?
Phenylethyl Alcohol
- The Gram-negative bacteria is suppressed by the interaction of alcohol with the outer membrane
- The Gram-positive bacteria are typically unaffected by the phenylethyl alcohol
What are the selective reagants in MAC?
Bile salts and crystal violet
- The bile salts and crystal violet inhibit the growth of Gram-positive bacteria when compared to the TSA control plate
What are the selective reagants in MSA?
7.5% NaCl
- MSA selects for halotolerant organisms, bacteria that can survive in a relatively high salt environment
What are the differential reagents in MAC?
Sugar and neutral red (a pH indicator)
- If a Gram-negative organism is able to ferment lactose, the colonies will turn pink and may have a haze around them
- Gram-negative organisms unable to ferment lactose will be colorless
What are the differential reagents in MSA?
Sugar mannitol and phenol red (a pH indicator)
- If a halotolerant organism is able to ferment mannitol, the acids produced will cause the phenol red in the medium to turn yellow
- Bacterial growth with no color change indicates halotolerance, but no ability to ferment mannitol
What type of media contains reagents that will change color or react due to the growth or metabolism of bacteria?
Differential Media
What is this an example of?
The reagants in MacConkey Agar (MAC) will turn bacterial colonies pink if they are able to ferment lactose?
Differential media
__________ ingredients do not suppress growth, they react in response to growth.
Differential
__________ contain ingredients that suppress the growth of some type of organism and allow a different type of organism to grow.
Selective Media
Selective media are interpreted with regard to the ___________ or _________ of growth
presence or absence
Differential media are interpreted by the _______ or ___________of the growth
color or lack of color
What is this an example of?
The ingredients of the MacConkey agar (MAC) suppress the growth of Gram-positve cells, while selecting the growth of Gram-negative cells.
Selective Media
If an organism does not grow in a selective media what conclusions can be drawn pretaining to the differential media?
No conclusions can be drawn. When using media that are selective and differential, it is important to remember that the selective assay takes precedence over the differential assay
What type of test is the Mannitol Salt Agar (MSA)? What does it test for?
- MSA is a selective and differential media
- MSA selects for halotolerant organisms
- Bacterial growth = halotolerant
- No growth = not halotolerant
- MSA is differential for the ability of bacteria to ferment mannitol
- Growth & Yellow = halotolerant organism that ferments mannitol
- Growth & Colorless = halotolerant organism that can’t ferment mannitol
- No growth = no conclusion can be draw about fermentation
What type of test is PEA? What does it test for?
- PEA is selective for Gram-positive bacteria
- Not a differential test
- The presence of alcohol suppresses the growth of Gram-negative bacteria
- The presence of alcohol doesn’t suppress the growth for Gram-postive bacteria
- Growth = Gram positve
- No growth = Gram negative
What type of test is the MacConkey Agar (MAC)? WHat does the MAC test for?
MAC is a selective and differential medium
- MAC is selective for Gram-negative bacteria
- Growth = Gram negative
- No growth = Gram positive
- MAC is differential for lactose fermentation
- Growth & pink = Gram negative bacteria that can ferment lactose
- Growth & colorless = Gram negative bacteria that cannot ferment lactose
- No growth = Gram positve but no conclusions can be drawn about fermentation
What is the purpose of the KOH test?
The KOH test is a differential test used to confirm the results of the Gram stain.
KOH causes lysis of Gram-negative bacteria (gooey)
KOH will not cause lysis of Gram-positive bacteria (watery)
Why does KOH become gooey in the presence of Gram-negative cells?
KOH causes lysis of Gram negative bacteria cells. After lysis, proteins and other macromolecules from the lysed cells denature, and form a thick, gooey, sticky liquid.
What is denaturation?
Denaturation is the break down of protein molecules.
As a beginning microiologist, which do you think is more reliable, a Gram stain or the KOH test?
THe KOH, because there is less steps to cause error and the results are instanteous.
What happens to Gram positve bacteria cell after the KOH test?
The Gram-positve cultures do not lyses, and the KOH remains watery.
What happens to Gram negative bacteria after the KOH test?
KOH causes lysis of the Gram-negative bacterial cells, causing the proteins and other macromolecules to denature and form a thick, gooey, sticky liquid.
What oxygen indicator is used in thioglycollate broth?
Resazurin acts as an oxygen indicator
(Thio-gly-coll-ate)
(Re-saz-ur-in)
Why is resazurin used as an oxygen indicator for thioglycollate?
Oxygen diffuses into the top portion of the thioglycollate media, which turns the resazurin pink, indicating that the top area of the thioglycollate broth is aerobic.
What component of the thioglycollate broths maintains the anaerobic conditions in the lower portion of the broth?
Sodium thioglycollate acts as a oxguen scrubber, maintaining an anaerobic environment in the thioglycollate medium below the pink band of resazurin.
Where in a thioglycollate broth tube would you expect to find growth for the following types of bacteria?
Facultative anaerobes
Throughout the tube (top and bottom)
Facultative anaerobes can use either aerobic respiration or fermentation, so can use oxygen but does not require it.
Where in a thioglycollate broth tube would you expect to find growth for the following types of bacteria?
Obligate aerobes
Top of the test tube only (where the resazurin is)
Obligate aerobes require oxygen for their metabolism, and are incapable of growing in an anaerobic environment, as they cannot use fermentation or anaerobic respiration.
Where in a thioglycollate broth tube would you expect to find growth for the following types of bacteria?
Obligate anaerobes
Obligate anaerobes are poisoned by oxygen, so they gather at the bottom of the tube where the oxygen concentration is lowest.
Where in a thioglycollate broth tube would you expect to find growth for the following types of bacteria?
Aerotolerant anaerobes
Aerotolerant anaerobes typically use fermentation to make ATP, so they do not require nor utilize oxygen.
Aerotolerant anaerobes will grow within the lower, anaerobic portion of the broth
What type of metabolism is used by obligate aerobes?
Obligate aerobes use (oxygen) aerobic respiration
What type of metabolsim is used by facultative anaerobes?
Facultative anaerobes can use both (oxygen) respiration and (anaerobic/without oxygen respiration) fermentation
What type of metabolism is used by obligate anaerobes?
Obligate anaerobes can only use fermentation
What type of metabolism is used by aerotolerant anaerobes?
Aerotolerant anaerobes use fermentation
Would you expect an obligate aerobe to be capable of fermentation?
No, because obligate aerobes can only use oxygenic/ aerobic respiration, not fermentation to survive.
What is the substrate of gelatinase?
Gelatin (collagen)
Do all types of bacteria have the same oxygen requirements?
No, different types of bacteria have different oxygen requirements.
_______________ require oxygen for their metabolism and are incapable of growing in an anaerobic environment, as they cannot use fermentation or anaerobic respiration.
Obligate aerobes
Both ____________ and __________ will grow in an anaerobic environment.
Facultative anaerobes
Aerotolerant anaerobes
____________ can use either aerobic respiration or fermentation, so they can use oxygen but do not require it
Facultative anaerobes
_______________typically use fermentation to make ATP, so they do not require nor utilize oxygen.
Aerotolerant anaerobes
____________ require low levels of oxygen.
Microaerophiles
What type of medium is used to determine the requirements of bacteria?
Thioglycollate broth
What does thioglycollate both contain?
Sodium thioglycollate, resuzurin, and 0.05% agar
What in thioglycollate both acts as an oxygen indicator?
Resazurin
__________ is a differential nutrient broth used to determine the oxygen requirements of bacteria
Thioglycollate
Organisms that cannot grow in anaerobic conditions, ___________, will grow only in the ________ portion of the tube
- Obligate aerobes
- Upper/ top
Organisms that can grow anaerobically, ___________and ________, will grow within the _______, anaerobic portion of the thioglycollate broth.
- Aerotolerant anaerobes and facultative anaerobes
- Lower/ bottom
What type of enzyme is secreted by bacteria (and other organisms) into their environment?
Exoenzymes
(Exo-enzymes)
What is another name for gelatinase?
Gelatinase is sometimes called collagenase
Gelatinase/collagenase is a type of what?
Protease
What enzyme catalyzes the hydrolysis of gelatin/collagen?
Gelatinase/ collagenase which is a type of protease
Why type of enzyme is used by a wide variety of bacteria to break down proteins for nutrient acquisition?
Exoenzymes
Gelatinase/ collagenase is what type of enzyme?
Exoenzyme
What type of differential medium is used to determine if an organism produces gelatinase?
Nutrient gelatin
What are the ingredients for nutrient gelatin?
Gelatin (collagen)
Peptone
Beef extract
If an organism produces ______, the exoenzyme will break down gelatin to its component amino acids and the solid gelatin will be liquid at room temperature
Gelatinase/ collagenase
How is gelatinase used as a virulence mechanism?
Pathogens may use enzymes like gelatinase to break down connective tissues as part of their invasion process, an important virulence mechanism
Why do you think agar is used to solidify media?
Agar has a higher melting point than gelatin. Gelatin is liquid at 28oC and the incubator’s temperature is at 35oC, so gelatin media would not be good to track growth, like agar would.
How do you interpret the nutrient gelatin assay?
If an organism produces gelatinase, the exoenzyme will break down gelatin to its component amino acids and the solid gelatin will be liquid at room temperature
Why must observations for nutrient gelatin be done at temperatures lower that 28oC?
Gelatin is a liquid at 28oC or above
What type of biological molecule is collagen?
Amino acids
Organisms that live in an oxygen containing environment must have _________ to deak with harmful ______ ________.
- Mechanisms
- Oxygen radicals
__________ is an enzyme found in bacteria and other organisms that catalyzes the conversion of peroxide anions into water and oxygen, protecting the organism from oxidative damage.
Catalase
___________ is a differential test used to determine if a culture produces the catalase enzyme.
The catalase test
When hydrogen peroxide is added to a sample of bacteria on a microscope slide, if bubbles form this indicates that the organism possess what type of enzyme?
Bubbles = catalase
When hydrogen peroxide is added to a sample of bacteria on a microscope slide and no bubble form, this indicates that the organism…?
No bubbles = no catalase
If you have a positve catalase test, does this indicate that the organism is an obligate aerobe or facultative anaerobe?
No, a positve catalase test only indicates that the organism possess catalase.
Having a positive catalase test should not be interpreted as the organism being an obligate aerobe or a facultative anaerobe.
What is the last enzyme in the electron transport system of many organisms?
Cytochrome oxidase
What enzyme is responsible for reducing the oxygen in the final step of aerobic respiration, forming H2O as an end product?
Cytochrome oxidase
(Cyto-chrome oxi-dase)
What is the function of catalase?
To protect the organism from oxidative damage
Why is bubbing used to determine if catalase is present?
The catalase enzyme will bubble when hydrogen peroxide is added to is, because it converts the peroxide anions into water and oxygen thus forming bubbles
What type of enzyme is secreted by bacteria (and other organisms) into their environment?
Exoenzymes
(exo-enzymes)
What type of enzyme is used as toxins (like hemolysins and phospholipase) or can function to provide nutrients to the cell?
Exoenzymes
Exoenzymes such as __________ are used to break down large polymeric molecules into their monomers so that they may be taken into the cell.
Amylase
What exoenzymes are used in the brewing industry to catalyze the hydrolysis of polysaccharide amylose, starch, into smaller sugars maltose and glucose, which are then transported into the cell.
Amlyase
Glycosidase
Why is starch hydrolysis a useful differential assay for the characterization of unknown organisms?
Not all bacteria produce the (exo)enzymes amlyase (and glycosidase)
(am-ly-ase)
(gly-co-i-dase)
What differential medium is used to detect the hydrolysis of starch by a bacterial culture?
Starch agar
If an organism can hydrolyze starch in the starch agar, what will be seen on the plate after iodine is added?
A clear/tan area called the zone of clearing will be seen around the bacterial culture due to the absence of starch in the area immediately surrounding the bacterial growth.
What enzymes are used by bacteria to hydrolyze starch?
The exoenzymes:
- Amylase (amy-lase)
- Glucosidase (gluco-si-dase)
Why is iodine added to the starch plate?
To better see the zone of clearing if the bacteria hydrolyzes starch (produces amylase) because iodine turns starch blue/purple
Why does a zone of clearing form around organisms that can hydrolyze starch?
Amylase and glucosidase will “digest” starch, thus there will be a zone of clearing around the culture due to the absence of starch in the area immediately surrounding the bacterial growth.
Is starch agar a differential or a selective medium?
Starch agar is a differential medium because it doesn’t inhibit the growth of the culture, but it does determine the culture’s metabolism
What is a common metabolic strategy among bacteria?
Fermentation
In fermentation, ___________ __________ are used as the final electron acceptors of the metabolic pathway.
Organic molecules (contains C & H)
What metabolic process allows an organism to continue to oxidize NADH back into NAD+ and produce ATPs in the absence of oxygen?
Fermentation
What two areas is fermentation dividied into?
- Alcohol fermentation
- Acid fermentation
What type of fermentation consists of a carbohydrate ultimately being converted into organic acids?
Acid fermentation
What type of fermentation consists of a pathway in which: the pyruvate produced by glycolysis is decarboxylated to produce carbon dioxide and acetaldehyde, the acetaldehyde is then reduced to ethanol by alcohol dehydrogenase, and NADH is oxidized to NAD+.
Alcohol fermentation
What type of fermentation consists of glucose being broken down to pyruvate by glycolysis, producing 2 ATPs by substrate level phosphorylation. Pyruvate is then reduced into lactic acid by the enzyme lactate dehydrogenase. During this reduction of pyruvate, NADH is oxidized to NAD+.
Homolactic acid fermentation
Why type of fermentation consists of an organism fermenting a carbohydrate into lactic acid, ethanol, and gas?
Heterolactic acid fermentation
Do all bacteria utilize the same types of carbohydrates and use them in the same way?
No. Different species, and strains of bacteria due to their genetic makeup, do not necessarily utilize the same types of carbohydrates nor utilize them in the same way.
What differential test broths contain a sugar (like glucose, lactose, sucrose, or mannitol and phenol red), a pH indicator that turns from red to yellow in the presence of acid?
Phenol red carbohydrate broth
What is a Durham tube?
The Durham tube is a smaller, upside down tube in the Phenol red carbohydrate broth tube, used to capture any gases that may be produced during fermentation.
What is the purpose of the Durham tube in the phenol red carbohydrate broth tube?
To capture any gases that are produced during fermentation
What does it mean when the phenol red carbohydrate broth turns yellow and no bubbles have formed in the Durham tube?
The organism produces acid only
What does it mean when the phenol red carbohydrate broth tube turns yellow and there is a bubble in the Durham tube?
The organism ferments a sugar to produce an acid and gas
What does it mean if the Phenol red broth tube does not turn yellow?
The organism cannot ferment the sugar in the broth
What does it mean when the phenol red turns a faint orange color but not an intense yellow?
The organism may be an obligate aerobe, since obligate aerobes can convert sugars into acid, but do not employ a fermentative pathway.
Some amounts of acids may be been produced, but no fermentation occured.
What happens is the phenol red carbohydrate broth tube is incubated for an extended period of time?
The tue may turn from yellow back to red, this is usually seen at the top of the broth
What can be determined about an organism using the Phenol red carbohydrate broth tubes?
We can only determine if an acid and/or gas is produced via fermentation
What can the Phenol red carbohydrate broth tubes not determine about fermentation?
Does not provide any information regarding the types of acids produced (lactic, acetic) or the gas produced (CO2, H2)
What can be concluded by doing the Phenol red carbohydrate broth tube tests?
Only can conclude whether acid alon or acid or gas were produced
Phenol red turns what color in an acidic solutions?
Yellow
You have an unknown culture and do not have Gram stain results. You are allowed one medium to determine if it can ferment lactose. Would you use phenol red lactose broth or MacConkey agar?
Phenol red lactose broth, because we want to know if the unknown organism can ferment lactose only.
The MacConkey agar test is a selective test for Gram negative and a differential test for lactose fermentation.
We just want to know if the unknown can ferment lactose, not that if its a Gram negative or positive.
You have grown an obligately aerobic culture in phenol red glucose broth, and some of the media has turned yellow. How would you explain this?
Obligate aerobes can convert sugars, like glucose, into acid, but they do not employ a fermentative pathway.
A culture ferments mannitol and glucose to acid, but not lactose and sucrose. Is this possible? Why?
Yes, because different bacteria have different DNA, different proteins, and different enzymes therefore different metabolisms.
Different species, and strains of bacteria due to their genetic makeup, do not necessarily utilize the same types of carbohydrates, nor utilize them in the same way
What type of molecule is used as the final electron acceptor in fermentation?
Organic compounds (have C & H)
You check your fermentation broth and find that the top is red and the bottom is yellow. What can you conclude?
It means that the organism has fermented and the top portion means that the tube has been incubated for an extended period of time, which may have turned from yellow back to red
What is the purpose of the Durham tube?
The purpose of the Durham tube is to capture any gases that are produced during fermentation during the phenol red carbohydrate broth tube test.
What are the end products in each of the following types of fermentation?
Alcohol fermentation by yeast
Ethanol and carbon dioxide
What are the end products in each of the following types of fermentation?
Homolactic fermentation
2 ATPs and lactic acid
What are the end products in each of the following types of fermentation?
Heterolactic fermentation?
Lactic acid, ethanol, and gas (carbon dioxide)
What type of microbial control can be accomplished by using heat treatment, cold storage, disinfectants and radiation
Environmental control
Control in or on the a host employs the use of ________ and ____________ drugs.
- Antiseptics
- Chemotherapeutic
This category of microbial control includes oral and topical antimicrobias and antibiotics.
Chemotherapeutic agents
(chemo-therap-eu-tic)
What is a general term for any compound that negatively affects the growth of microbes?
Antimicrobial
How can antimicrobials be antibacterial, antifungal, antiviral, antihelminthic, etc?
Depends on the target organism
What term has been historically reserved for those antimicrobials produced by organisms such as fungi and bacteria?
Antibiotic
(anti-bio-tic)
What terms is used interchangeably with antibiotic?
Antimicrobial
(anti-microb-ial)
What is the term used to describe some antibacterial drugs that are effective against a variety of bacteria?
Broad spectrum drugs
What do broad spectrum drugs usually target?
A common aspect of the bacterial cells
Ex: the drugs that attack bacterial ribosomes tend to be broad spectrum drugs
How do antimicrobial drugs work?
Antimicrobial drugs work because they are selectively toxic
What is selective toxicity?
An aspect of antimicrobial drugs in which they attack some aspect of the target cells’ physiology that is not found in the host
Why do most antibacterial drugs not interfere with the functions within the human cells?
Because bacterial physiology differs from eukaryotic physiology
What kind of drug is only effective against only one or a few types of bacteria?
Narrow spectrum drugs
How do narrow spectrum drugs usually work?
Narrow spectrum drugs usually attack an aspect of cell physiology not found in all bacterial cells
The drug isoniazid affects the production of mycolic acid, and its use is limited primarily to the Mycobacteria.
What kind of drug is this?
Narrow spectrum drug
Class of drug that includes:
- streptomycin (strept-o-mycin)
- kanamycin (kana-mycin)
- neomycin (neo-mycin)
Aminoglycosides
(amino-glyco-sides)
What is an ingredient in Neosporin, an over the counter antibacterial cream?
Neomycin (neo-mycin)
From the class aminoglucosides (amino-gluco-sides)
What is the mode of action of aminoglucosides?
(amino-gluco-sides)
To bind to the bacterial ribosomes, preventing protein synthesis
What antibacterials are included in the drug class aminoglucosides?
“New Kittens Sleep”
- Neomycin (neo-mycin)
- Kanamycin (kana-mycin)
- Streptomycin (strepto-mycin)
What class of antibacterial drugs is an antibacterial polypeptide produced by strains of Bacillus subtilis.
Bacitracin (bac-i-tracin)
What is the mode of action for bacitracin?
Bacitracin works by preventing cell wall synthesis
What class of antibacterial drugs is also an ingredient in Neosporin, along with neomycin and polymyxin B?
Bacitracin
What class of antibacterial drugs includes:
- penicillin (pen-i-cillin)
- amoxicillin (amox-i-cillin)
Beta-Lactams
Beta-lactams include which antibacterials?
“A&P”
- Amoxicillin (amox-i-cillin)
- Penicillin (pen-i-cillin)
How is the mode of action for beta lactams?
Beta-lactams prevents the formation of peptidoglycan molecules in the bacterial cell wall.
(pepti-do-gly-can)
Amoxicillin is used in conjectin with what Beta-lactamase inhibitor?
Clavulanic acid
(cla-vul-anic)
What Beta-lactam combination drug therapy can circumvent some resistance mechanisms?
Amoxicillin with clavulanic acid (a Beta-lactamase inhibitor)
What is the mode of action for chloramphenicol?
(chlor-am-phen-icol)
Chloramphenicol (chlor-am-phen-icol) disrupts protein synthesis by binding to the large ribosomal subunit
What is the mode of action for fluoroquinolones?
(fluoro-quin-olones)
Fluoroquinolones works by binding to the enzyme DNA Gyrase, preventing replication and killing bacterial cells
Ciprofloxacin is commonly used with which class of antibacterial drug?
(ci-pro-flox-acin)
Fluoroquinolones
(fluoro-quin-olones)
Erythromycin (ery-thro-mycin) is under which class of antibacterial drug?
Macrolides
(macro-lides)
What is the mode of action for macrolides?
(macro-lides)
Macrolides disrupt protein synthesis by binding to bacterial ribosomes
What group/class of antibacterials includes tetracycline?
(tetra-cycl-ine)
Tetracyclines
(tetra-cycl-ines)
What is the mode of action for tetracyclines?
(tetra-cycl-ines)
Tetracyclines bind to the bacterial ribosomes and interfere with protein production
What group of antibacterial drugs act as competitive inhibitors of the enzymes used in the biosynthesis of folate by bacteria?
Sulfonamides/Trimethoprim
(sulf-on-amides)/ (trime-tho-prim)
What type of bacteria is Bacitracin used against?
Gram positive
Sulfamethoxazole with trimethoprim (SxT) is an example of a combination drug. What class of antibiotic does it belong to?
(sulf-a-meth-ox-azole)
(trime-tho-prim)
Sulfonamides/ trimethoprim
(sulf-on-amids)
(trime-tho-prim)
What drug class is often used in combination therapy, as each antibacterial targets a different enzyme in the metabolic pathway?
Sulfonamides/ trimethoprim
(sulf-on-amides)
(trime-tho-prim)
What test was used in lab to test the antibacterial drug susceptbility of an organism, using paper discs impregnated with a known concentration of a specific antibiotic?
Kirby-Bauer Disk Diffusion Assay
If a culture is susceptible to the antimicrobial disk in the Kirby Bauer Disk Diffusion assay what will be observed?
Why?
A zone of clearing
The zone of clearing is produced because the bacterium cannot grow in the presence of the drug.
How is the zone of clearing measured in the Kirby Bauer Diffusion Disk assay?
The diameter of th zone of clearing is measured (in mm)
What information can be obtained from measuring the zone of clearing from the Kirby Bauer Diffusion Disk assay?
The diameter’s length can determine the relative susceptibilties of each culture in relation to the antimicrobial used.
_________ __________ is a potent mutagen and microbial control agent.
Ultraviolet (UV) light
What causes the formation of thymine dimers in DNA molecules?
UV light
What happens if the thymine dimers in a bacteria’s DNA is not repaired?
Thymine dimers that are not repaired can block DNA replication and transcription
Can UV light kill bacteria?
Yes, thymine dimers which left unrepaired can block DNA replication and transcriptase
What factors of UV light affect its microbial properties?
- Intensity of light used
- Duration of UV exposure
What are the limited uses of UV light?
- Surfaces
- Clear substrates
Why does UV light have limited applications to surfaces and clear substrates?
UV light does not penetrate into the items being treated,
So UV light is most effective for disinfecting surfaces
What is UV light most effective at disinfecting?
Surfaces
What does disinfection with UV light require?
Required an adequate amount of exposure time so that the amount of DNA damage is such that the organism will die before the damage can be repaired
Explain why UV light exposure may not be effective?
Many bacteria possess enzymes to repair UV light. If the amount of damage is minimal, the bacteria can repair the thymine dimers
How do you interpret the disk diffusion assay for antibacterial efficacy?
The diameter of the zone of clearing
What would you hypothesize to explain why there may be differences in the usefulness of antibiotics on different species of bacteria?
The specific features that antibiotics target can be different between different types of bacteria cells. This makes some antibiotics less or more effective against targeting and killing the pathogenic bacteria.
What is meant by a broad-spectrum drug?
Drug spectrum drugs usually target a common aspect of bacterial cells.
Is UV effective control of all bacteria? Explain.
No, many bacteria possess enzymes to repair UV light damage. So if the exposure is minimal, the bacteria can repair the thymine dimers and the UV exposure will not be an effective disinfectant treatment.
How does UV light affect bacteria?
Thymine dimers
___________ is the uptake of naked DNA by a bacterial cell.
Transformation
The relative ability of a bacterial cell to incorporate our plasmid is referred to as __________.
Competence
____________ cells have the innate ability to take in DNA from the environment.
Competent
Can all bacteria undergo natural transformation?
No
Where are bacterical cells manipulated to induce artificial competency?
In the lab
What is transformation?
