Brainscape's Knowledge GenomeTM

Browse over 1 million classes created by top students, professors, publishers, and experts.


See full index

4.1 Medical Microbiology > Laboratory Diagnosis of Meningitis > Flashcards

Laboratory Diagnosis of Meningitis Flashcards

You may prefer our related Brainscape-certified flashcards:
USMLE® Step 1 flashcards Biology 101 flashcards
1
Q

What is the specimen of choice for meningitis investigation, what is the gold standard?

A

Cerebrospinal fluid (CSF)

The specimen should be obtained before patient is put on antimicrobial therapy

The specimen should be transported rapidly to the lab

CSF should NOT be refrigerated

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
2
Q

What are some specimens other than CSF for investigation of meningitis?
(5)

A

Blood cultures -> these tend to also be positive -> think of where the meningitis originates

Skin scrapings -> only used in meningococcal meningitis -> rapid, helpful, often positive

Wound swabs ->only helpful in trauma/chronic cases

Urine or sputum for tuberculosis meningitis

Blood -> looking for antiodies, antigens, raised CRP etc etc

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
3
Q

How should CSF be collected?
(3)

A

CSF is normally collected sequentially into three or more separate containers -> and should be labelled in accordance of draw

The first and last samples are sent for micro and the second one is sent to bio for glucose and protein quantification

At least 1 ml per tube is necessary

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
4
Q

What is different about the collection of a CSF for Mycobacterium investigation?

A

We need to take as much CSF as posible -> as large a volume as possible

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
5
Q

What counts are done on CSF?

A

A white cell count and a red cell count

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
6
Q

Why do we carry out a serial red cell count on our CSFs?

A

To determine whether a haemorrhagic CSF is due to a traumatic tap or a subarachnoid haemorrhage

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
7
Q

How can you tell between a haemorrhagic sample and a traumatic CSF?

A

The uniform bloodstaining of all samples suggests previous haemorrhage into the subarachnoid space

Reducing bloodstaining in sequentially obtained samples suggest bleeding induced by the tap procedure

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
8
Q

What are some safety considerations to handling CSF?

A

Biological Safety hood required due to the possibiltiy of N. meningitidis

A cat 3 lab is required for any M. tuberculosis meningitis

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
9
Q

How should a CSF specimen be collected?

A

Collect sample before antimicrobial therapy (if possible)
Send sample immediately to the lab and process urgently
Minimal volume of 1ml required but more required for tuberculosis
Insufficient volumes must be prioritised
Collect sequentially into 3 separate containers

Other samples such as a pharyngeal swab, blood for PCR and FBC should be considered

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
10
Q

Why do we always take a blood sample for query meningitis?

A

As we often query sepsis as the cause of meningitis

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
11
Q

What is the first thing that a scientist does when receiving a CSF?

A

Specimen should be signed in
Describe the appearance of the CSF

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
12
Q

How should you describe the appearance of a CSF?

A

Comment on turbidity -> indicater of wbcs
Comment on degree of haemorrhage -> rbcs
Any sign of xanthochromia/straw-coloured
Any cobwebs indicative of TB?

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
13
Q

Why do we send a CSF sample to biochemistry?

A

Quantification of glucose and protein
-> used to differentiate between viral and bacterial meningitis

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
14
Q

What do we do after describing the appearance of a CSF?

A

We carry out a complete cell count:
- rbcs
- cell count for wbcs (neutrophils vs lymphocytes)

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
15
Q

What do we do after carrying out a cell count on a CSF sample?

A

We centrifuge the sample

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
16
Q

Why do we centrifuge our CSF samples?

A

We centrifuge the sample as a means of concentrating any bacterial cells which would be present -> increases likelihood of us being able to cuture an organism

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
17
Q

What do we do after we centrifuge a CSF sample?

A

Gram (can do further confirmatory testing based on what we see microscopically)
Culture on blood and chocolate agar (nothing every grew on these in the Mater)
We do molecular detection using the film array
We can send a sample out to a reference lab for antigen testing etc to confirm ID

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
18
Q

What molecular detection methods do we do on the CSF?

A

We use the film array

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
19
Q

What are some downfalls of molecular methods of detecting meningitis

A

The film array is not quantiative

We need to culture regardless for DST and for epidemiology

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
20
Q

Why do we carry out grams for CSF?

A

A lot of the time nothing will grow on the blood and choocalte agar due to patients already being on antimicrobial treatment

Because of this we can use the gram to confirm ID of organism -> lets us confirm what we get on molecular

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
21
Q

In general what six things do we do on CSF

A

Macroscopic appearance: colour and clarity

Cell count

Centrifugation

Gram stain

Culture on blood and choc

Further tests based on microscopy results

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
22
Q

Talk about the macroscopic examination of CSF

A

CSF should be clear and colourless = normal

Cloudy/turbid sample is abnormal -> relates to wbcs, rbcs and microorganisms

Colour is related to haemorrhage or pigment

NB: a CSF sample may contain abnormal numbers of cells while still appearing “clear”

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
23
Q

How many white cells are required to make a CSF sample cloudy?

A

50 WBCs and the CSF will start to loose clarity

200 WBCs and the CSF will look turbid

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
24
Q

What is considered a normal white cell count in adults

A

<5/ul

Any elevation in this is abnormal

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
25
Q

When carrying out a cell count what might you notice other than wbcs

A

RBCS: these are not diagnostic of meningitis but they should still be reported

Microorganisms may also be seen and their presence recorded e.g. Listeria, pneumococci, candida or cryptococcus

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
26
Q

What is a normal CSF wcc for neonates, 1-4yr olds, 5 yrs-puberty and adults?

A

0-30 cells in neonates
0-20 cells in 1-4 year olds
0-10 cells in 5 year olds to puberty
0-5 cells in adults

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
27
Q

What is a normal erythrocyte count for newborns vs adults?

A

Newborn = 0-675 cells x 10^6/L
Adults = 0-10 cells x 10^6/L

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
28
Q

What is the normal protein in csf of neonates <6days old vs others?

A

0.7 g/L in neonates <6days old
0.2-0.4 g/L in others

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
29
Q

When would you not be concerned with a csf wcc of 35?

A

a wcc of 35 is perfectly normal in any baby less than 1

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
30
Q

In what cases will lymphocytes in CSF predominate over PNMs?

A

Viral meningitis
Fungal meningitis
Tuberculous meningtisi

You might also see more lymphs in early stage listeria meningitis

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
31
Q

How do we centrifuge our CSF samples?

A

Centrifuge 1,200g for 5-10 minutes
if query TB centrifuge at 3,000g for 15-20 minutes

Aseptically remove supernatant
Resuspend deposit

Retain supernatant and store at 40 degrees celsius

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
32
Q

What is the one reasons why we would not centrifuge a csf sample?

A

only done if you have more than half a ml

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
33
Q

What is xanthochromia?

A

The yellow colouration of the supernatant of centrifuged CSF

34
Q

What causes xanthochromia?

A

Metabolism of products of RBC breakdown

Increased CSF protein concentration

Bilirubin staining

35
Q

How should we quantify xanthochromia?

A

It should be determined accurately only by spectrophotometry

36
Q

After centrifugation, why might the supernatant be pink and what does this mean?

A

This may be due to oxyhaemoglobin

This confirms pre-tap intracranial haemorrhage

After 24 hours this degrades to bilirubin and xanthochromia increases to peak at 36-48 hours

37
Q

What is the gold standard for meningitis diagnostics

A

culturre

38
Q

What factors affect meningitis culture

A

Antimicrobial therapy
Very small nnumbers of bacteria
bacteria may be fastidious
low volume of csf => low numbers of bacteria

special culture may be required for Mycobacterium, Candida and crypto

39
Q

What kinds of culture can we put up for meningitis?

A

Always:
- blood agar in 5-10% CO2
- chocolate agar in 5-10% CO2

Blood anaerobically
Broth
Possibly mycobacterial media is query TB
Possibly fungal media (SDA) if query fungus

40
Q

What kinds of culture can we put up for meningitis?

A

Always:
- blood agar in 5-10% CO2
- chocolate agar in 5-10% CO2

Blood anaerobically
Broth
Possibly mycobacterial media is query TB
Possibly fungal media (SDA) if query fungus

41
Q

Talk about the gram stain for meningitis

A

It can be invaluable to diagnose meningitis in minutes
It can be technically difficult as organisms are often intracellular organisms - will need to look in wbcs etc
You can concentrate the gram by adding drop on drop of centrifuged concentrate to slide

42
Q

What would gram-positive diplococci indicate?

A

S. pneumoniae

43
Q

What would gram negative coccobacilli mean?

A

Haemophilus

44
Q

What would gram negatve diplococci indicate?

A

N. meningitidis

45
Q

What would gram positive cocci in chains indicate

A

S. agalactiae

46
Q

What would gram positive bacilli indicate

A

Listeria but these are rarely visible - intracellular

47
Q

What are the two most likely yeast in csf?

A

Candida
Cryptococcus

48
Q

What special stains might be used on CSF?

A

Wet prep and india ink for cryptococcus

Ziehl Neelson for tb

49
Q

What is the recommended preparation for the white cell differential?

A

Cytospin + Leishman differential wcc:
- polymorph:lymphocyte ratio

50
Q

What are some exceptions to our wcc limits for meningitis?

A

Neonates, will be higher
Viral and TB meningitis -> polys may predominate in early infection
Neutropenic patients - still work up even if low neuts

51
Q

Why is csf protein levels not diagnostic?

A

Elevated protein is not diagnostic of meningitis

It may indicate the duration of the infection or a poor prognosis

52
Q

What is normal glucose levels?

A

60% of blood glucose level usually

Its between 2.2-3.3 m.mol/L

53
Q

What is the one bacterial meningitis where there is not a decrease in glucose?

A

Listeria meningitis -> its usually 60%

54
Q

What antimicrobial susceptibility should be carried out for meningitis

A

Should use standard EUCAST methods

Choose antimicrobials based on microorganism identified

Should test for B-lactamase production by H. influenzae

55
Q

How should we treat meningitis?

A

Intravenous antimicrobials

56
Q

What are the five methods of diagnosing meningitis

A

Gram stain
Culture
PCR test
Serology
Antgen test

57
Q

How dependent is sensitivity of blood and culture on antimicrobial use

A

If no prior antibiotic use:
- CSF culture is 95% sensitive
- Blood culture is 50% sensitive

If prior antibiotic use:
- Blood sens is only 5%
- CSF -> nothing really grows

58
Q

Give a specific reason for the need for clinical need for molecular detection of samples in CSF

A

In a study of 103 patients in the UK with clinically defined meningitis only 13% of cases were positive on CSF cultures

Our reference lab in 2019 carry out a study of N.meningitidis cases whereby 40% were detected by PCR methods only i.e. culture negative

59
Q

What is the high clinical impact of meningitis and encephalitis

A

High morbidity
High mortality
Significant health care costs
Rapid treatment is associated with improved outcome
Current gap in diagnostics: identification of the causing pathogen
Time-consuming (culture based)
Sue to pre-treatmenet sometimes unsuccessful

60
Q

Talk about the use of multiplex meningitis encephalitis platforms for the detection of meningitis

A

Syndromic testing - multiplex nested PCR
Reverse transcription - cDNA used
Broad coverage without the need to select specific tests
Enhanced ability to detect co-infections
Increased sensitivity
Higher throughput but not really cause the film array can only process one sample at a time
Reduced sample volume requirements

61
Q

What bacteria is the biofire film array capable of detecting?

A

E. Coli K1
H. influenzae
L. monocytogenes
N. meningitidis
S. agalactiae
S. pneumoniae

62
Q

What viruses are detectable by the biofire meningitis/encephalitis panel?
(7)

A

Cytomegalovirus
Enterovirus
Herpes simplex virus 1
Herpes simplex virus 2
Human herpes virus 6
Human paraechovirus
Varicella zoster

63
Q

What fungi are detectable by the filmarray

A

Yeasy
Cryptococcus neoformans
Cryptococcus gattii

64
Q

What are the three main benefits of the biofire?

A

Everything happens in one single pouch, very easy to use

Very small amount of specimen needed ideal for CSF

Very well controlled - multiple on board controls

65
Q

What are the three steps to the film array?

A

1st stage PCR
2nd stage PCR
DNA melting analysis

66
Q

What are the steps of the 1st stage PCR

A

Reverse transcription of viral RNA into cDNA
Multiplex nested PCR
Enrich for the target nucleic acid in the sample
Diluted

67
Q

Wha are the steps to the 2nd stage of pcr on the film array

A

Primers are nested or internal to the specific products of the 1st stage multiplex PCR

68
Q

What is DNA melting analysis on the biofire

A

Fluoresence in each well of the array is monitored and analysed to generate a melt curve

This curve tells is if the organism is detected or not

69
Q

What are the controls on the biofire

A

RNA process control:
- RNA transcript from Schizosaccharomyces pombe
- Lysis purification RT PCR 1st stage 2nd stage melt analysis
- A positive control indicates that all steps carried out in the film array ME pouch were successful
- this controls the PCR amplification steps

A PCR2 control
- DNA target
- a positive result indicates that the 2nd stage PCR was successful

Both controls must work or we will get an invalid result

70
Q

What organisms are we missing from the biofire?

A

Were missing organisms that occur in shunt ingections such as S. aures, CNS, P acnes etc

71
Q

What are the advantages of the Biofire?

A

High sensitivity with a low limit of detection for pathogens
Short hands-on-time in sample preparation
Short assay turn around time, overall about 3 hours
Assessment of the most common community acquired pathogens

72
Q

What are the disadvantages of the Biofire

A

Diagnostic gaps for apthogens e.g. staphylococci for shunt infections

No information on antibiotic resistance

No quantification

Costs

73
Q

What organisms does the BIOFIRE strugle to detect?

A

Group B streps
-> culutre should be used to confirm these as they are often missed on molecular platforms

74
Q

What is the main issue with the BIOFIRE?

A

Too many false positives: for viral, fungal and bacterial agents -> S. pneumoniae false positives frequent, as well as E. Coli K1 and S agalactiae -> Viral false positives in herpes frequent

Issues with detection of Group B streps

75
Q

What bacteria does the BIOFIRE struggle to detect?

A

It can miss Group B streps

S. pneumoniae False Positive

E. Coli K1 false positives

S. agalactiae false positive

-> i.e streps and E. Coli K1

76
Q

What viruses does the BIOFIRE struggle to detect and why?

A

Herpes viruses

These can be tricky to diagnose as some patients might have an initial infection while others migh have a latent infection

False positives are associated with latent infection

77
Q

What might cause so many S. pneumoniae false positives?

A

It was suggested that lab staff should wear masks etc to prevent S. pneumoniae contamination

Suggested that a degree of these positives were jut to lab staff however this is unlikely as it was seen across a lot of labs

78
Q

What are the limitations of the FilmArray?

A

Lack of clinical details on all patients -> cannot tell if latent or active infection for Herpes etc

High clinical suspicion is required

Positive bacterial and cryptococcus results should be correlate with Gram stain results and cultures as well as clinial findings -> discrepant molecular results should be witheld pending further investigation

Confirmatory testing should be carried out e.g. PCR for HSV

79
Q

Why should we confirm any organisms IDd on the BIOFIRE, give an example of why?

A

There was a case where a patient was diagnosed with HSV-1 on the biofire but was later diagnosed with MTB 7 days later

The HSV-1 diagnosis was not confirmed by PCR methods -> this should have been done as antimicobrial therapy was 7 days late for patient etc etc

80
Q

For th HSV/TB case what should have been done?

A

The gram stain should have been checked for correlation

Clinical symptoms should have been looked at - they wouldnt have matched

Confirmatory testing should have been carried out

81
Q

What is the sensitivity of CSF culture, why so low?

A

Sensitivty of 13%

Low due to antimicrobials

82
Q

Compare the BIOFIRE to Reference lab, what is the main advantage of both

A

BIOFIRE only takes abour 70mins for a result while the reference lab takes up to 5.6 days

The reference lab has a lower detection rate

4.1 Medical Microbiology (30 decks)

Brainscape helps you reach your goals faster, through stronger study habits.
© 2024 Bold Learning Solutions. Terms and Conditions