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PHA112 > Micro TBL Pre-Reading > Flashcards

Micro TBL Pre-Reading Flashcards

1
Q

what do many microorganisms require?

A

B group vitamins

this requirement satisfied by yeast extract

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2
Q

at what temperature does agar form a firm gel?

A

37 degrees C

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3
Q

at what temperature is agar used as a liquid and how this useful?

A

45 degrees C

this is at sufficiently low temp without killing microorganisms

this = imp in pour plate counting methods

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4
Q

solid media designed for the growth of anaerobic organisms usually contain non-toxic reducing agents.

give examples of these?

A
  • sodium thioglycollate

- sulphur containing amino acids

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5
Q

give example of redox indicators?

A
  • methylene blue

- resazurin

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6
Q

why may redox indicators be incorporated into anaerobic media?

A

to confirm the sufficiently low redox potential has been achieved

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7
Q

media for yeasts and moulds have … pH than bacterial culture media?

A

lower

pH of yeasts & moulds: 5.5-6.0

pH of bacterial culture media: 7.0-7.4

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8
Q

why is lactic acid used to impart a low pH?

A

it is not inhibitory to fungi at concentrations used

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9
Q

in bacteria, the patterns of binary fission take place how often?

A

every 25-30 mins under optimal conditions of laboratory cultivation

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10
Q

how do moulds grow?

A

through extension & branching of hyphae to produce mycelium

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11
Q

how may anaerobic organisms be grown?

A

on petri dishes given incubated in anaerobic jar

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12
Q

what are anaerobic jars made of?

A

rigid plastic with airtight lids

  • petri dishes placed in them together with low temp catalyst
  • catalyst made of palladium-coated pellets/wire
  • this causes oxygen in jar to = combined with hydrogen generated by additions of water to NaBH4
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13
Q

what is the periphery of a colony?

A

part which = actively growing and usually non-pigmented

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14
Q

what is the term planktonic used to describe?

A

freely suspended cells

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15
Q

bacteria attached to a substrate (surface in the body) are known as what?

A

sessile

they = said to exhibit biofilm/micro colony mode growth

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16
Q

what are planktonic cells routinely used for?

A

testing procedures designed to assess activity of antimicrobial chemical & processes

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17
Q

there are several situations where it is necessary to measure the number of microbial cells in culture, sample or specimen.

give examples of these?

A
  • when measuring levels of microbial contamination in raw material/manufactured medicine
  • when using microorganisms in making of therapeutic agents
  • when evaluating effects of antimicrobial chemical/decontamination process
  • when assessing nutrient capability of growth medium
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18
Q

in what case is it necessary to know the total number of microbial cells present?

A
  • in vaccine making (dead & living cells may both cause immune response)
  • pyrogen testing (dead & living cells induce fever when injected into body)
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19
Q

what is a viable count?

A

counting living cells only

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20
Q

when might MPN (most probable number) counts be used?

A

when anticipated count is relatively low

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21
Q

what do MPN counts involve?

A
  • inoculating multiple tubes of culture medium (3/5)
  • with 3 different volumes of sample
  • there should be proportion of tubes receiving inocula where no microorganism present
  • these remain sterile after incubation
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22
Q

where are MPN counts used?

A

water, food, dairy industries

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23
Q

MPN counts are highly…

A

innacurate

24
Q

…. measurements are the most common means of estimating the total numbers of bacteria present in sample

A

turbidity

25
Q

how do you take turbidity measurements?

A
  • use spectrophotometer/colorimeter

- reading conc from calibration plot

26
Q

why cannot fungi undergo turbidity measurements?

A

cannot be readily handled in this way because suspension may not = uniform / may sediment in spectrophotometer cuvette

27
Q

what might direct microscopic counting be an appropriate method for?

A
  • bacteria
  • yeasts
  • fungal spores
28
Q

the traditional methods of viable counting all suffer from the same limitations.

what are they?

A
  • labour intensive
  • not easy to automate
  • slow, because need incubation period for colonies to develop/liquid cultures = turbid
  • need large volumes of culture media
29
Q

give examples of the operating principles most common in detection of living cells?

A

epifluorescent techniques:

  • dyes exhibit diff colours in living & dead cells / appear colourless outside cell
  • become fluorescent when absorbed

living cells generate ATP:
- readily detected by enzyme assays e.g. luciferin emits light when exposed to firefly luciferase

resistance, capacitance/impedence of culture medium changes as result of bacteria/yeast growth & metabolism

manometric techniques = appropriate for monitoring growth of organisms that consume/produce large amounts of gas during metabolism

30
Q

give examples of viable count methods?

A
  • pour plate (counting colonies in agar)
  • surface spread/surface drop methods
  • membrane filter methods
  • MPNs
31
Q

give examples of total count methods?

A
  • direct microscopic counting
  • turbidity methods
  • dry weight determination
  • nitrogen, protein/nucleic acid determination
32
Q

give examples of rapid methods (indirect viable counts)?

A
  • epifluorescence
  • ATP methods
  • impedance
  • manometric methods
33
Q

what are the advantages of the pour plate method?

A
  • doesn’t need pre-drying of agar surface

- will detect lower conc than surface spread methods

34
Q

what are the disadvantages of the pour plate method?

A
  • very small colonies of strict aerobes at base of agar may = missed
  • colonies of diff species in agar appear similar (difficult to detect contaminants)
35
Q

what are the advantages of surface spread/ surface drop methods?

A
  • gives larger colonies than pour plates (easier to count)

- easier to find contaminants by appearance of colonies

36
Q

what are the disadvantages of surface spread/ surface drop methods?

A
  • agar surface needs pre-drying to absorb sample
  • possibility of confluent
    growth, particularly with moulds, making individual colonies
37
Q

what are the advantages of membrane filtration methods?

A
  • will detect lower conc than other methods

- antimicrobial chemicals in sample = physically removed from cells

38
Q

what are the disadvantages of membrane filtration methods?

A
  • viscous samples won’t go through membrane

- particulate samples = block membrane —> restricting filtration capacity

39
Q

apart from adding preservatives, give another factor that can potentially attenuate the preservative activity?

A

oil-water partitioning

40
Q

with tests involving liquid systems, how can the early growth of viable cells be assessed?

A

by some light scattering processes

41
Q

what might quantitative PCR be useful in demonstrating?

A

presence/growth of microorganisms that are slow/difficult to culture under usual lab conditions e.g. viruses

42
Q

why must determination of MIC values be conducted under standardised conditions?

A

deviation from standard test conditions can result in considerable variation in data

43
Q

what have the historical gradient plates, ditch-plate and cup-plate techniques been replaced with?

A

more quantitative techniques:

  • disc diffusion
  • broth & agar dilution
  • e-tests
44
Q

describe briefly what a disc test is?

A

filter-paper discs impregnated with antimicrobial replace antimicrobial-filled cups / wells

45
Q

disc test process?

A
  • standard suspensions of log-phase growth cells prepared and inoculated on to surface of appropriate agar plates to form lawn
  • paper discs with known conc of antimicrobial agent placed on dried lawn
  • plates incubated aerobically at 35 degrees C for 18 hours
46
Q

when do problems arise with the disc test?

A

when inoculum density is e.g. too low —> indistinct edge to inhibition zone after incubation

47
Q

as the distance from the disc …., there is a logarithmic …. in the antimicrobial concentration

A

increases

reduction

48
Q

what is the most convenient and presently accepted method of determining bacterial MICs?

A

E (epsilometer) test

49
Q

give examples of antimicrobial-resistant bacterial pathogens?

A
  • vancomycin-resistant enterococci (VRE)
  • methicillin-resistant staph. aureus (MRSA)
  • vancomycin intermediate staph.aureus (VISA)
50
Q

standard methodology may fail to detect the resistant phenotype.

give the factors that cause this??

A
  • heterogenous expression of resistance e.g. VISA, MRSA
  • poor agar diffusion of antimicrobial
  • slow growth of resistant cells
51
Q

give the conditions where expression is enhanced?

A
  • lower temperatures

- higher salt conc

52
Q

how can you improve the clarity of inhibition zone edges? (tests for fungistatic and fungicidal activity)

A

adding methylene blue

53
Q

what is the MFC?

A

the lowest drug conc showing no growth / fewer than 3 colonies per plate to obtain 99-99.5% killing activity

54
Q

give examples of bacterial species that are cocci and gram pos?

A
  • e.faecium
  • s.aureus
  • s.epidermidis
  • s.pneumoniae
55
Q

give examples of bacterial species that are bacilli and gram pos?

A
  • b.subtilis
  • c.difficile
  • l.monocytogenes
  • c.sporogenes
56
Q

give examples of bacterial species that are cocci and gram neg?

A
  • h.influenzae
  • m.catarrhalis
  • n.gonorrhoeae
  • v.parvula
57
Q

give examples of bacterial species that are bacilli and gram neg?

A
  • p.aeruginosa
  • s.enterica
  • p.mirabilis
  • e.coli

PHA112 (37 decks)

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