Past Papers 7 Flashcards
RT PCR virus RNA PCR run graph
label the x axis of the chart
x-axis - Cycle number
not the cycle threshold (Ct value)
RT PCR virus RNA PCR run graph
label the y axis of the chart
y-axis - fluorescence
RT PCR virus RNA PCR run graph
what is the red line marked Z
red line marked Z - threshold line
set to be in the linear portion of the PCR curves
RT PCR virus RNA PCR run graph
What is the cycle threshold?
A positive result should have fluorescence sufficiently above the threshold value, in order for it to be called positive
Threshold usually set at the limit of detection for the system
Ct value - cycle threshold value tells us at what point the fluorescence crosses the threshold value, and becomes positive
RT PCR virus RNA PCR run graph
What is the effect of moving line Z upward - i.e higher baseline fluorescence?
decrease sensitivity, as low level positives may not be called as such
Increase PPV- all pos will be strongly pos
increase specificity, as all positive results should be strongly positive
Decrease NPV - as some negative results might be weak pos, but called neg
RT PCR virus RNA PCR run graph
Please assess results ABDE with your reasoning
https://www.rcpath.org/static/80d54920-b6e2-45aa-bdb6c34026e99179/Part-2-Virology-sample-questions.pdf
A - good take off, good curve - positive
B - ok take off, poor curve just above threshold . Do sine wave up to Ct 40, so this is negative result
D - ok take off, good curve, positive
E - late take off, ok curve. Would like to see more of curve. Could be a weak positive result, or could be contamination from a strong positive result such as D sample
RT PCR virus RNA PCR run graph
It is revealed:
A positive control
E negative control
BCD were taken from patients with suspected measles
What is your opinion of this run?
https://www.rcpath.org/static/80d54920-b6e2-45aa-bdb6c34026e99179/Part-2-Virology-sample-questions.pdf
A - Positive control worked - but would want to compare to known previous values of control
E - Negative control is flagging weak positive - suggests contamination. This is likely due to a strongly positive sample D
B - almost certainly negative. Low fluorescence, might want to repeat testing?
C - definitely negative
D- more strongly positive than the contaminated E sample. So can accept this result. Strength of this result means it is not due to contamination
A 56-year-old man presented with progressive renal insufficiency. He had received a cadaveric renal transplant 6 months ago for chronic renal failure secondary to hypertension and diabetes mellitus.
CMV IgG status was D- / R-. He was on prednisolone, tacrolimus and mycophenolate mofetil. Progressive deterioration in allograft function was noted .
5a Name two most likely differential diagnoses for the possible deterioration in renal function
graft rejection - could be spontaneous due to poor match, or insufficient immunosuppression
CMV
BK virus nephropathy -also known as Polyomavirus 1
drug related nephropathy - tacrolimus/ MMF
A 56-year-old man presented with progressive renal insufficiency. He had received a cadaveric renal transplant 6 months ago for chronic renal failure secondary to hypertension and diabetes mellitus.
CMV IgG status was D- / R-. He was on prednisolone, tacrolimus and mycophenolate mofetil. Progressive deterioration in allograft function was noted .
Shown H&E biopsy.
Name the virus which is likely responsible
Describe the features shown in H&E stain biopsy
CMV?
Cells infected with CMV are larger, with a thickened nuclear membrane and granular intracytoplasmic inclusions.
Typical “owl’s eyes” or basophilic intranuclear inclusion bodies are representative of active CMV replicating nucleoprotein cores
Histological examination with H&E stain has lower sensitivity than IHC stain
A 56-year-old man presented with progressive renal insufficiency. He had received a cadaveric renal transplant 6 months ago for chronic renal failure secondary to hypertension and diabetes mellitus.
CMV IgG status was D- / R-. He was on prednisolone, tacrolimus and mycophenolate mofetil. Progressive deterioration in allograft function was noted .
Shown H&E biopsy - CMV features
Name 3 other viruses that can also cause these features
Histological examination with H&E stain has lower sensitivity than IHC stain for CMV - as other viruses can appear similar
unclear answer?
HSV1/2
VZV
Adenovirus
All have infected cells which are larger
A 56-year-old man presented with progressive renal insufficiency. He had received a cadaveric renal transplant 6 months ago for chronic renal failure secondary to hypertension and diabetes mellitus.
CMV IgG status was D- / R-. He was on prednisolone, tacrolimus and mycophenolate mofetil. Progressive deterioration in allograft function was noted .
IHC testing was performed
In this immunostained slide, what monoclonal antibody is most commonly used to confirm the main diagnosis?
What is its limitation?
Mouse recombinant CMV-p65 protein antibody IgG binds to CD8 specific T-cells
Made using E.coli bacteria
limitations - cannot find anything for this
p65 has been used in trials as a marker of CMV reactivation, as it suggests virions are being formed. E.g viral load 1000, but p65 present - recommend treating
A 56-year-old man presented with progressive renal insufficiency. He had received a cadaveric renal transplant 6 months ago for chronic renal failure secondary to hypertension and diabetes mellitus.
CMV IgG status was D- / R-. He was on prednisolone, tacrolimus and mycophenolate mofetil. Progressive deterioration in allograft function was noted .
Primary CMV infection suspected
List the key features of a post-transplant policy aimed at preventing loss of graft due to this virus infection
Matching donors and recipients for status
CMV negative blood products
solid organ transplant - 100/3 months days of gancliclovir if recipient CMV neg, and donor CMV pos
Do not routinely offer Valganciclovir prophylaxis if donor and recipient are seronegative for CMV (D-/R-)
viral load monitoring in this case if infection occurs
This is an Internal Quality Control chart for positive standard within a quantitative CMV DNA assay.
https://www.rcpath.org/static/80d54920-b6e2-45aa-bdb6c34026e99179/Part-2-Virology-sample-questions.pdf
What is the name of this type of chart?
Statistical process control (SPC)
Levy-Jennings chart is a type of SPC, that uses Westgard rules
This is an Internal Quality Control chart for positive standard within a quantitative CMV DNA assay.
https://www.rcpath.org/static/80d54920-b6e2-45aa-bdb6c34026e99179/Part-2-Virology-sample-questions.pdf
Which Westgard rule was violated in the course of this chart?
13s
one measurement exceeds 3SD above or below the mean
This is an Internal Quality Control chart for positive standard within a quantitative CMV DNA assay.
https://www.rcpath.org/static/80d54920-b6e2-45aa-bdb6c34026e99179/Part-2-Virology-sample-questions.pdf
What actions need to be taken on discovery of this violation?
If a Westgard rule is broken, then the run must be rejected
Investigate -
samples - temperature/ storage/ handling
equipment mainteance
staff errors
new batch of controls - new mean may need calculated
This is an Internal Quality Control chart for positive standard within a quantitative CMV DNA assay.
https://www.rcpath.org/static/80d54920-b6e2-45aa-bdb6c34026e99179/Part-2-Virology-sample-questions.pdf
Name two other Westgard rules
13s - one measurement exceeds 3SD above or below the mean
22s - two consecutive measurements exceed 2SD above or below mean
R4s - two measurements in the run have 4SD difference. For example 1 result 2SD above, and another result 2SD below
41s - four consecutive measurements 1SD above or below mean
10x-bar - ten consecutive measurements are on the same side of the mean
This is an Internal Quality Control chart for positive standard within a quantitative CMV DNA assay.
https://www.rcpath.org/static/80d54920-b6e2-45aa-bdb6c34026e99179/Part-2-Virology-sample-questions.pdf
If the internal quality control is 3 SD below the mean on 4 consecutive days, what would you recommend?
Issue with equipment/ analysis
Issue with IHC - possibly new batch. New mean may need to be calculated, if no other issues found
Candidates will see a virus marker Positive
HBV cirrhosis recipient
Is liver donation permitted?
Unknown which virus marker.
HCV RNA pos?
Can donate
Ideally treat donor first
If no time, transplant, and treat recipient with DAAs
Candidates will see a virus RNA Positive
Sperm donation to wife
Is this donation permitted? State any precautions.
Unclear virus
Sperm donors are screened for HIV1/2, HTLV, HBV, HCV, syphilis, chlamydia/ gonorrhoea
CMV sometimes tested
If CMV pos, then would only recommend donating to recipients who are also CMV pos
Zika? Male wait 3 months before conception
Candidates will see a virus DNA Positive
Kidney donation
Is this donation permitted? State any precautions.
CMV??
Can donate
screen recipient
Consider Valganciclovir prophylaxis for at least 3 months after starting treatment for acute allograft rejection if either donor or recipient are CMV positive (D+/R-, D+/R+ or D-/R+)
Someone wishing to donate sperm
Initial screening infection tests are negative.
What else would exclude patient from donating?
If high risk recent sexual exposures e.g sex worker/ IVDU/ MSM or someone known to have infection.
Need to wait 3 months and re-screen, to ensure they are not incubating
if had tattoos/ piercings, do not need to wait
Candidates will see a virus RNA Positive - Zika?
Sperm donation to wife
Is this donation permitted? State any precautions.
Should wait 3 months from infection before donating.
Can donate sooner if sperm is tested and found to be Zika negative
females should not try and get pregnant for 2 months after having Zika infection
