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Microbiology > Cultivation, Microscopy and Immuniology > Flashcards

Cultivation, Microscopy and Immuniology Flashcards

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1
Q

what is clinical microbiology

A

-Very simple strategy
-Collect specimen from patient
-Examine specimen for evidence of pathogen
-Find pathogen in specimen by e.g
Direct microscopy
-Find evidence of pathogen by e.g
Immune response or Biomarker signature (protein, lipid, etc.)
-Arrive at a diagnosis

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2
Q

what are the factors of clinical specimens

A

-Material has to be appropriate to the clinical condition= Diarrhoeal symptoms suggest that faeces be collected and Blood, urine, CSF, biopsy tissue etc. as appropriate
-Aseptic collection
-Collect an appropriate quantity
-Collect specimen before treatment is initiated
-Fast transport from collection to laboratory analysis

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3
Q

what are some clinical collection materials

A

-Swabs =skin, wounds, nose, ear etc.
-Needles= Blood, tissue fluids, Cerebrospinal Fluid
-Sterile cups= mucus, stool
-Catheter = urine, blood
-Intubation= extraction of fluids from hollow organs

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4
Q

what do we use to determine if a pathogen is present in a sample

A

-Microscopy
-Laboratory Culture of microorganisms
-Direct detection= Immunology, nucleic acid technology and analytical chemistry

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5
Q

what traditional methods do microbiology labs rely on

A

culture, phenotypic, immunological and biochemical tests

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6
Q

what samples are anaylsed in a clincal lab

A

blood, faeces, sputum, skin swabs, wound swabs etc

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7
Q

what are the best diagnostic tests to use

A

sensitivity and specificity

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8
Q

what is sensitivity

A

-Sensitivity refers to a test’s ability to designate an individual with disease as positive.
-A highly sensitive test means that there are few false negative results, and thus fewer cases of disease are missed.
-A sensitive test for an infectious disease should also be able to work with relatively low numbers of pathogens present in the specimen.

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9
Q

what is the equation for sensitivity

A

-ability of a test to detect a true positive.

-Sensitivity = True positive / True positive + false negative x100

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10
Q

what is specificity

A

-The specificity of a test is its ability to designate an individual who does not have a disease as negative.
-A highly specific test means that there are few false positive results.

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11
Q

what is the equation for specificity

A

-Specificity = ability of a test to exclude a true negative.

-Specificity = True negative / True negative + false positive x100

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12
Q

what is desirable in a test

A

a test that is both highly sensitive and highly specific. This is frequently not possible

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13
Q

what is looked at in diagnosis using microscopy (direct…)

A

-Direct examination of sample
-Variety of formats

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14
Q

what are the advantages of microscopy

A

-Rapid
-Cheap
-adaptable according to sample
-Specificity possible
(antibody/nucleic acid staining)

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15
Q

what are the disadvantages of microscopy

A

-Limited specificity
-No recovery of organism for further analysis.
-Poor sensitivity
-Expensive for viruses

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16
Q

what occurs in diagnosis using a laboratory culture

A

-Inoculation of specialised medium with specimen
-Incubate for specified time
-Recover individual colonies
(possibly grow a new culture from a single colony to ensure working with a pure culture of the organism of interest)
-Look at morphology and Gram reaction
-Undertake additional biochemical tests to obtain an ID

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17
Q

what mediums are used in laboratory culture strategy

A

selective, differential and chromogenic mediums

18
Q

what tests are used in laboratory culture strategy

A

gram stain, biochemical, immunological and molecular tests

19
Q

what is general purpose media examples

A

Tryptone Soy Agar
Brain-Heart Infusion Agar

20
Q

what is an example of selctive media

A

Vogel and Johnston Agar

21
Q

what is examples of differential media

A

MacConkey Agar
Blood agar

22
Q

what is chromogenic media

A

-Most recent development
-Species-specific chromogens developed to take advantage of what we understand about bacterial metabolism.
-Beta-galactosidase
-Beta glucuronidase

23
Q

what is chromogen

A

“a chemical compound, itself without colour, that can be transformed into a coloured compound, or can react with another material to form a coloured compound”

it is transformed by a specific enzyme and this enzyme is only found in one particular species

24
Q

why do we use Chromogens?

A

-Improve overall specificity of a diagnostic medium.
-Positive colonies are easy to identify.
-Reduce number of sub-cultures required.

25
Q

what are the advantages of culture

A

-Unequivocal demonstration of presence of pathogen
-Pathogen is available for further antimicrobial susceptibility testing
-Established technology and standards
-Relatively inexpensive
-Low technology
-Less to go wrong (hopefully)

26
Q

what are the disadvantages of culture

A

-slow= It can take days to identify a bacterial pathogen and to determine the most appropriate antibiotics
-Technically challenging in certain instances= e.g Chlamydia spp. + Mycobacterium tuberculosis
-Impossible to culture some pathogens= e.g Treponema pallidum A bacterium causing syphillis
-Culturing methods assume that all pathogens are culturable all of the time and does not take into account either= injured organisms and viable but non-culturable organisms.

27
Q

what is the equation for the biochemical testing of bacteria

A

-Catalase test
-2 H2O2 –> Catalase –> 2 H2O + O2
S. aureus +

Streptococcus -

28
Q

explain bacteria and fermenting sugars

A

-Many bacteria have different abilities to ferment sugars
-Specific sugar and pH indicator in the broth e.g glucose, galactose, mannose etc.

29
Q

what are the colours produced when bacteria ferment sugar

A

-Purple: no fermentation
-Yellow: fermentation occurs and acid produced
-Yellow: fermentation occurs and both acid and gas produced

30
Q

what is API

A

-API: Miniaturised biochemical tests on a strip.
-Can easily see that these three species are different
-Different patterns
-ID based on database

31
Q

What is the advanatages of Immunodiagnostics

A

-Recognition of species
-Fast assays
-Very flexible

32
Q

what is Immunodiagnostics used for

A

-Direct analysis of sample to detect presence of pathogen
-Used to confirm identification when pathogen isolated
-Lots of simple systems in use= Agglutination and ELISA

33
Q

what test is used in Immunodiagnostics

A

Agglutination Test

34
Q

what is the Agglutination Test

A

-Simplest test available to see Visible clumps on a slide
-Widely used for confirmation
tests on organisms isolated on an agar plate
-S. aureus on Mannitol Salt Agar
-Remove colony
-Mix with anti-S. aureus serum
-Shake on slide
-Look for agglutination

35
Q

what is ELISA

A

Enzyme-linked immunosorbent assay

36
Q

what does the name of ELISA suggest

A

-Name suggests three components
-Antibody= Allows for specific detection of antigen of interest and If antigen is specific to a bacterium then we can identify the bacterium.
-Solid phase (sorbent)= Allows one to wash away all the material that is not specifically captured
-Enzymatic amplification= Allows you to turn a little capture into a visible color change that can be quantified using an absorbance plate reader

37
Q

what is Alkaline Phosphatase

A

-One of the most widely used reporter systems in ELISA and Histochemistry
-A general purpose Hydrolyase enzyme that removes phosphate groups under alkaline conditions
-Can act on many different substrates

38
Q

what is used in the lateral flow test strip

A

-Conjugation of Particles
-Conjugate Pad
-Test Strip
-Absorbent pad (wicking)

39
Q

what is the advantage of the lateral flow test strip

A

rapid diagnosis

40
Q

what are the main elements in clincal diagnostic microbiology

A

-Specimen subjected to a number of treatments
-Microscopy
-Laboratory Culture of microorganisms
-Direct detection= Immunology, Nucleic acid technology and Analytical chemistry
-In practice more than one set of techniques are used together to give a more accurate result.

Microbiology (29 decks)

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