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Microbiology > Lab Practical 2 > Flashcards

Lab Practical 2 Flashcards

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1
Q

Antimicrobials, usually of a low molecular weight, produced by microorganisms that inhibit or kill other microbes.

A

Antibiotics

Penicillin and streptomycin

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2
Q

Antibiotics that are chemically altered to make them more effective

A

Semi-synthetics

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3
Q

Antimicrobials that are chemically synthesized in the laboratory and are not produced my microbial biosynthesis

A

Synthetics

Sulfa drugs used to treat some bacterial diseases before the discovery of penicillin. Sulfa drugs target vitamin synthesis, more narrow in spectrum

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4
Q

Antibiotic resistant strain of bacteria?

A

methicillin-resistant S.aureus (MRSA)

Doesn’t respond to penicillin. Vancomycin must be used. Nosocomial infections.

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5
Q

Broad spectrum vs narrow spectrum

A

Some antibiotics are more effective against gram-positive bacteria, some against gram-negative. Broad-spectrum are effective against both kinds of organisms (can target 70S ribosomes which all bacteria have)

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6
Q

Antimicrobial that targets cell wall synthesis

A

Penicillin

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7
Q

Antimicrobial that targets DNA and RNA synthesis

A

Cipro, rifampin

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8
Q

Antimicrobial that targets protein synthesis

A

Tetracyclines, streptomycin

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9
Q

Method used to determine sensitivity or resistance of a bacterium to an antimicrobial.

A

Kirby-Bauer Method
Uniformly lawning a standardized inoculum on a medium and then paper disks with specific concentrations of antimicrobial or antibiotic are placed on the surface. The stuff diffuses out on the agar, creating a concentration gradient.
S. aureus, E. coli, P. aeruginosa

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10
Q

In the Kirby-Bauer Method, when the agents inhibits or kills the test organism?

A

Zone of inhibition where there is no growth shown

It can vary with the diffusibility of the agent, the size of the inoculum, the type of medium, etc.

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11
Q

Medium in Kirby-Bauer method

A

Mueller-Hinton II agar. pH between 7.2-7.4, 4mm thick.
For fastidious organisms, sheep’s blood is added to the medium. 48hrs incubation

Cultures are designated resistant, sensitive, or intermediate

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12
Q

Compounds that kill or inhibit microorganisms.

A

Antimicrobials

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13
Q

Nonionizing short wavelength radiation that falls between 4nm and 400 mm in the visible spectrum.

A

Ultraviolet light

The shorter the wavelength in electromagnetic radiation, the more damaging it is to cells. Most bacteria are killed by it and it’s used sterilize surfaces

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14
Q

Lethality of UV light?

A

Primary lethal effects are due to its mutagenic properties. UV radiation at 260nm is the most germicidal because it’s the specific wavelength at which DNA maximally absorbed UV.
Depends on time of exposure, what’s being exposed, and the blocking of light

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15
Q

These form when UV light causes a covalent bond to form between two adjacent thymine or cytosine molecules in a DNA strand.

A

Pyrimidine dimers
Cause the DNA molecule to be deformed to that the DNA polymerase can’t replicate DNA strands past the site of dimer formation, nor can genes past this point be transcribed.

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16
Q

System that enzymatically removes the dimers and inserts in their place new pyrimidine molecules. The enzymes can move past the point where dimers occur, unlike DNA polymerase.

A

SOS system
Can be overwhelmed and begin making errors by inserting incorrect bases for the damaged bases if there are too many dimers. Results in cell death.

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17
Q

Why are endospores more resistant to UV light than vegetative cells?

A

The DNA of endospores is protected by acid-soluble proteins that bind to it and alter its confirmation. Also photolyase enzyme is generated by UV light in them that functions in repair during germination.
B. megaterium is the endospore
S. aureus isn’t.

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18
Q

A technique used in research for many things. Based on the enzymatic amplification of a desired fragment of DNA. Three-step process of denaturation, annealing, and extension that is repeated several times. After each cycle, the number of copies is doubled.

A

Polymerase chain reaction (PCR)
Catalyzed by heat-stable Taq polymerase that is controlled by a thermocycler.
About 1 trillion copies are made in the end, enough to detect the targeted DNA by gel electrophoresis.

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19
Q

What is PCR used for?

A

Identification of infectious agents and detection of antibiotic resistant strains. Sequencing, diagnosis of hereditary disease, identification of genetic fingerprints.

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20
Q

What is the primary mechanism of methicillin resistance in staphylococci?

A

The production of a penicillin-binding protein, PBP2, which is encoded by the mecA gene.

Identification of resistance is accomplished by amplifying the mecA gene using PCR.

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21
Q

Gram-positive spherical bacteria that divide in more than one plan to form irregular clusters of cells. Non-motile, non-spore forming, halophilic, mannitol fermenter. Most are facultative anaerobes.

A

Staphylococci

Nasal membrane, hair follicles, skin, and the perineum

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22
Q

How do infections by staph happen?

A

When a breach of skin or mucosa occurs, when a host’s ability to resist infection occurs, or when a staph toxin is ingested.

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23
Q

How much of the US population carries S. aureus? How much carries MRSA?

A

30%

1%

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24
Q

Symptoms of S aureus?

A

The most clinically significant staph pathogen. Skin infections, wound infections, bone tissue infections, scalded skin syndrome, toxic shock syndrome, food poisoning.

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25
Q

The most notable virulence factor possessed by S. aureus. 97% of the strains S. aureus are positive for this and will cause plasma to form a clot. The other two staph species are always negative for this.

A

Coagulase. May cause a clot to form around the staph infection thus protecting it from host defenses.

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26
Q

A nuclease enzyme that digests DNA.

A

DNase

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27
Q

A pigment produced by S. aureus that can act as a virulence factor. Has antioxidant properties that prevent reactive oxygen being produced by the host immune system from killing the bacteria.

A

Staphyloxanthin

Makes the golden color of S. aureus when grown on blood agar and S110.

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28
Q

Coagulase-negative staph (CNS) that do not produce DNase or alpha-toxin. Clinical significance has increased, especially for those with compromise immune systems or prosthetic or in dwelling devices.

A

Staph epidermidis and Staph saprophyticus

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29
Q

The most common cause of hospital acquired urinary tract infections. Also been involved with catheters, heart valves, and other prosthetic devices.

A

Staph epidermidis

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30
Q

The second most common cause of urinary tract infections in sexually active young women. The only clinically important staph selfies that is resistant to novobiocin. Some strains are able to ferment mannitol to acid.

A

Staph. saprophyticus

Unpigmented and appear opaque when grown on blood agar and S110 plates.

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31
Q

Contains 7.5% NaCl and phenol red indicator. The NaCl inhibits organisms other than staph. If the substance is fermented to produce acid, the medium changes from red to yellow. Of fermented, may be S. aureus or S. saprophyticus

A

Mannitol salt agar

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32
Q

Contains NaCl and mannitol, but lacks phenol red. Advantageous over MSA because it favors colony pigmentation by different species of staph. S. aureus colonies are yellow/orange. CNS colonies are colorless.

A

Staph medium 110 (SM110)

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33
Q

Involves inoculating a small tube of plasma with loopfuls of the organism and incubating. If the plasma turns solid, it’s positive. If there is a loose clot suspended in liquid or the plasma is solid, it’s positive. Valid only for gram+ staph like bacteria.

A

Coagulase test

Liquid fibrogenin to solid fibrin

Pseudomonas, gram- rods, can cause false positives.

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34
Q

Hemolysin that causes a wide, clear zone of beta-hemolysis on blood agar. Lyses red blood cells, damages leucocytes, cardiac muscle, and renal tissue.

A

Alpha toxin

A characteristic of S. aureus.

35
Q

These form part of the normal microbiota of the nose and throat, because it’s warm and moist.

A

Staphylococcus
Streptococcus
Neisseria
Haemophilus

36
Q

Gram+ cocci that occur in pairs and chains rather than clusters. Lack the enzyme catalase which dehydrates peroxide to form water and oxygen.

A

Streptococci

37
Q

This works in favor of the normal bacterial inhabitants, which suppress the growth of pathogens by competition for nutrients and production of inhibitory substances.

A

Microbial antagonism

38
Q

The major cause of the bacterial sore throat or acute pharyngitis, as well as necrotizing fasciitis.

A

Streptococcus pyogenes

39
Q

Explain blood agar plates.

A

5% defibrinated sheep blood, .5% NaCl, and nutrient agar.
Beta: complete, clear zone with a clean edge around colony
Alpha: incomplete, cloudy zone of browning due to methemoglobin
Gamma: no hemolysis or change in agar

40
Q

Which streptococci are normally part of the normal throat microbiota? Which are pathogens?
Which are what hemolysis?

A

Alpha and gamma hemolytic are normal, whereas beta is pathogenic.
S. aureus is beta
S. saprophyticus is gamma
S. epidermidis is gamma

41
Q

Definitive groupings of strep may be achieved by using this, which is based on antigenic differences of cell wall polysaccharides.

A

Lancefield Serogrouping Method.

42
Q

How are Streptococci grouped by using the Lancefield Serogrouping Method?

A

From A to O. Human isolates have been found to belong to groups A to G only.

43
Q

Microbes that have been implicated in cases of subacute bacterial endocarditis (SBE) and bacteremia after dental work.

A

Viridans Streptococci

44
Q

Microbe that can be screened using the Bile Esculin Agar and 6.5% Salt Agar Tolerance Test. Has been associated with UTIs, bacteremia, SBE, and septicemia. Vancomycin resistant variety is a thing.

A

Enterococcus faecalis

45
Q

A zone of inhibition 10mm or greater around the disk is indicative of bacitracin sensitivity. What can the organism be identified as?

A

Beta Streptococcus Group A
Streptococcus pyogenes

Resistant are Non-Group A Streptococcus

46
Q

A zone of inhibition 14mm or greater around the disk is indicative of optochin sensitivity. What can the organism be identified as?

A

Streptococcus pneumoniae

Resistant isolates are Viridans Streptococcus

47
Q

A Bile Esculin slant that is more than half darkened is considered to be a positive test. No color change indicates a negative test. What organism is a positive test presumptive of?

A

Enterococci

Requires further testing using salt tolerance

48
Q

A color change from purple to yellow in the salt tolerance test indicates a positive test, which means what?

A

The presence of Enterococci

49
Q

Associated with neonatal sepsis and meningitis, as well as post part infections (puerpueral fever)

A

Streptococcus agalactiae

50
Q

Grouping of tests used to differentiate organisms. Valuable for testing water for sewage contamination because they rule out E. coli which is not always associated with sewage.

A

Indole test

51
Q

A grouping of tests used to differentiate organisms. Can be valuable when testing water for sewage contamination because they can rule out E. aerogenes which is not always associated with sewage. Used for gram- rods and facultative anaerobes.

A 
IMViC
Indole
Methyl red
Voges-Proskaur
Citrate
52
Q

This test contains 0.4% agar and TTC. Bacteria will be able to swim through the medium and will reduce TTC to form a red precipitate called formazan.

A

Motility test
Motility medium with TTC

Motile, the tube will appear red and cloudy and the organism will spread
Nonmotile, the organism will grow along the streak line only.

53
Q

This test uses a medium that contains glucose, cystine, and sodium thioglycollate to lower the oxidation-reduction potential. The oxygen tension is high at the surface of the medium. Resazurin causes it to turn pink when near oxygen.

A

Oxygen requirement test

Fluid thioglycollate medium

54
Q

Test used to determine the ability to produce the enzyme gelatinase, which hydrolyses a large protein into smaller components that can then enter the organism and be metabolized.

A

Gelatin liquefaction test
Nutrient gelatin
A positive test is liquified, while a negative is solid. Continue incubation of negative tubes another 4-5 days to make sure.

55
Q

Test used to determine the ability to ferment a specific carb with or without the production of gas. Fermentation causes acid production, changing the phenol color. Durham tube collects gas.

A

Carbohydrate fermentation
Glucose, lactose, mannitol, maltose, sucrose broths
Positive for acid if phenol red turns yellow
Positive for gas if there’s a bubble

56
Q

Test used to determine the presence of an enzyme. H2O2 is an end product of breakdown of sugars, becoming toxic if there’s too much. The enzyme decomposes it. Only obligate anaerobes and aero tolerant bacteria.

A

Catalase test
Organism on slant, add H2O2
Positive test is bubbling as the O2 gas is liberated from the H2O2.

57
Q

Test used to determine if it is capable of using this as the sole source of carbon with production of the enzyme citratase. Medium contains sodium citrate as C source, ammonia salts for N, and brom thymol blue.

A

Citrate utilization
Simmons citrate agar
Positive means that alkaline pH causes the medium to change from green to Prussian blue.

58
Q

Test used to determine the ability to reduce nitrate to nitrite or nitrogen gas by production of nitratase. Takes place under anaerobic conditions in which an organism derives its O from nitrate.

A

Nitrate reduction test
Nitrate broth, with 5 drops nitrate reagent A and B added after incubation.
Positive is a red color
Negative needs zinc confirmation,
Positive is no color change
Negative is red color as nitrate is reduced by zinc

59
Q

Test used to determine the ability of an organism to produce mixed acid end products from gluc fermentation. Lactic, acetic, etc. Also produces gas due to the enzyme for mic hydrogen lyase. The acids lower the pH to less than 5.

A 
Methyl red test 
MR-VP broth 
Add methyl red reagent to tubes. 
Positive is red color
Negative is yellow color
60
Q

Test used to determine ability to produce acetoin, 2,3 butanediol, and ethanol, which cause less lowering of pH than methyl red positive organisms. Detects the presence of acetoin, which precedes 2,3 butanediol.

A 
Voges-proskauer test
MR-VP broth
Put culture in clean test tube, add Barritt's solution A and B. 
Positive is wine red color
Negative is brown
61
Q

Test used to determine the ability to hydrolyze starch. Amylase breaks the starch down.

A 
Starch hydrolysis
Starch agar plate 
Gram's iodine on the plate. 
Positive is a zone of clearing adjacent to the streak line. 
Negative is a blue-black area.
62
Q

Used to determine ability to produce caseinase which hydrolyzes casein to more soluble products.

A

Casein hydrolysis
Skim milk agar
Positive is a streak of clearing along the streak line

63
Q

Test used to determine ability to split indole from the amino acid tryptophan using tryptophanase.

A 
Tryptophan hydrolysis/Indole test 
Tryptone broth 
Add 10-12 of Kovac's reagent. 
Positive is a pink layer on the surface of the medium. 
Negative is yellow.
64
Q

Test used to determine the ability to split urea to form ammonia by the action of urease. Contains phenol red.

A 
Urea hydrolysis 
Urea broth 
Positive is an intense pink color
Negative is no color change 
7 additional days to make sure if negative.
65
Q

Test used to determine the ability to hydrolyze the sulfur containing amino acid cysteine to produce pyruvic acid, ammonia, and hydrogen sulfide. Removal of sulfide from the cysteine which is catalyzed by cysteine desulfurase. Contains ferris salts that react with H2S to produce black ferrous sulfate.

A

Hydrogen sulfide reaction
Triple sugar iron agar
Positive is a black precipitate along the stab line

66
Q

Test to determine ability to deaminate the amino acid phenylalanine resulting in production of phenylpyruvic acid and ammonia. Catalyzed by phenylalanase.

A 
Phenylalanine demination
Phenylalanine agar
Place 5 drops of 10% ferric chloride on slant culture. Wait 5 mins. 
Positive is green color
Negative is yellow color
67
Q

Test to determine ability to decarboxylate the amino acid lysine, resulting in the production of an alkaline end product cadaverine by producing decarboxylase. Medium has lysine, gluc, and brom creosol purple. Requires pH below 5 for activation.

A

Lysine decarboxylase test
Lysine decarboxylase broth
Microbe ferments gluc, indicator turns yellow cause low pH. Lysine decarboxylase is activated. Cadaverine is formed, pH rides, and turns original purple.
Positive is purple. Neg is yellow.

68
Q

Test used to determine ability to decarboxylate the amino acid ornithine, resulting in production of alkaline putrescine by producing enzyme ornithine decarboxylase. Medium has ornithine, gluc, and brom cresol purple. Requires pH below 5.

A

Ornithine decarboxylate test
Ornithine decarboxylate broth
Microbe ferments gluc, turns yellow cause low pH. Ornithine decarboxylase is activated. Putrescine is formed, pH rises, and it turns original purple.
Positive is purple, neg is yellow

69
Q

What specialized field of microbiology involves anti-body mediated immunity in the detection of causative agents of human diseases?

A

Immunoserology

70
Q

What does ELISA stand for? What is it based on?

A

Enzyme-Linked Immunosorbent Assay

Based on the fact that antibodies produced in response to pathogenic microbes attach to their antigenic sites called epitopes. This results in the formation of antigen-antibody complexes.

71
Q

Tests for the presence of microbial antigens in the patient’s blood.

A

Direct ELISA

72
Q

Used to detect infection by testing a patient’s blood for the presence or absence of antibodies against a particular microbe.

A

Indirect ELISA

73
Q

In indirect ELISA, if the serum samples contain antibodies against the bound antigen, what will happen?

A

The antibodies will attach to the antigens, forming tight complexes.

74
Q

What dark repair enzymes function in repairing UV light damage?

A

DNA polymerase
DNA ligase
Endonuclease

75
Q

Why is PCR more advantageous?

What is Taq polymerase?

A

It’s more rapid. It’s more specific because the polymerase follows the base-pairing rule.
Bad because it’s really expensive.

Thermos aquaticus

76
Q

What are the steps of the process of PCR? Temperatures and times?

A

Denaturation at 94C for 30 sec
Annealing at 55C for 30 sec
Extension at 72C for 1 min
Final extension at 72C for 5 min

77
Q

How does one tell the difference between strep and staph?

A

Catalase test, staph is positive

78
Q

Why is there additional screening after ELISA?

A

There can be false positives, so you have to confirm using nucleic acid amplification, PCR

79
Q

In the indirect ELISA, how does it work?

A

The antigen, a protein, is added to the wells of a plastic polystyrene microtiter plate. It binds to the bottom of the well by forming hydrophobic associations with the surface. The patient’s serum is then added. If it contains antibodies against bound antigen, they form tight complexes.

80
Q

In order to visualize antigen-antibody complexes in indirect ELISA, what is used?
In the final step, what is added?
Results of ELISA?

A

An enzyme-linked anti-human globulin, a conjugate.
In the final step, a chromogen substrate is added. The enzyme portion of the conjugate catalyzes a chemical reaction that changes the color of the chromogen. Any color change indicates infection.

81
Q

How does HIV work?

A

It attacks and destroys a type of T lymphocyte of the immune system called CD4+ cells, thereby severely weakening the immune defense. The body then produces antibodies, flu-like symptoms. Virus enters latency, where the cells continue to attack each other.

82
Q

Why is each sample tested in triplicate in the simulated indirect ELISA?

A

Statistical significance and to ensure reproducibility. Known positive and negative samples serve as controls.

83
Q

In interpreting TSI agar during hydrogen sulfide production, what are the symbols?

A 
A=acid production, yellow slant
K=alkaline reaction, red slant
G=gas production, gas in butt
S=H2S production, black butt
NC=no change

Microbiology (8 decks)

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