A 52-year-old man, 3 months after an allogeneic stem cell transplant for leukemia, was not vaccinated this season and has influenza A(H1N1)pdm09 pneumonia confirmed by RT-PCR on a nasopharyngeal swab, and oseltamivir is started. Repeat nasopharyngeal swabs are tested with the same assay and collection method: day 0, Ct 29; day 5, Ct 27; day 10, Ct 24, each with a valid internal control. He remains hypoxic on day 10. Which laboratory test is most useful now?

Neuraminidase gene testing for the H275Y mutation

Paired serology for influenza A antibodies

Hemagglutinin subtyping to confirm H1N1pdm09

A rapid antigen test to confirm the PCR

A laboratory confirms MERS-CoV by RT-PCR on two separate genomic targets, as WHO recommends. A sputum from a camel herder is positive on the upE screening assay (Ct 36) but negative on the ORF1a confirmation assay. What is the best next step?

Report inconclusive; test another target and a new sample

Report MERS-CoV not detected, since ORF1a is negative

Repeat the upE assay alone and report that result

Report MERS-CoV confirmed, since upE is positive

Which swab is suitable for collecting a nasopharyngeal specimen for respiratory virus NAAT?

Flocked swab on a plastic shaft

Charcoal-coated swab in Amies gel

Calcium alginate swab on an aluminium shaft

Virology and respiratory viruses

10 DOH Abu Dhabi (HAAD) lab licensing practice questions on Virology and respiratory viruses, part of Microbiology. The first 10 are below; the first 5 with answers and explanations (tap Show answer after you have chosen), the rest in Learn with a free account.

Question 1 · Hard
A 52-year-old man, 3 months after an allogeneic stem cell transplant for leukemia, was not vaccinated this season and has influenza A(H1N1)pdm09 pneumonia confirmed by RT-PCR on a nasopharyngeal swab, and oseltamivir is started. Repeat nasopharyngeal swabs are tested with the same assay and collection method: day 0, Ct 29; day 5, Ct 27; day 10, Ct 24, each with a valid internal control. He remains hypoxic on day 10. Which laboratory test is most useful now?
1. APaired serology for influenza A antibodies
2. BHemagglutinin subtyping to confirm H1N1pdm09
3. CNeuraminidase gene testing for the H275Y mutation
4. DA rapid antigen test to confirm the PCR
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Answer: C · Neuraminidase gene testing for the H275Y mutation
A falling Ct means a rising amount of viral RNA, so the virus is still replicating after 10 days of oseltamivir. Immunocompromised patients shed influenza for long periods and are the group in whom antiviral resistance most often emerges during treatment. In A(H1N1)pdm09, the commonest oseltamivir-resistance change is H275Y in the neuraminidase (H274Y in N2 numbering), detected by targeted RT-PCR or sequencing. This mutation usually leaves zanamivir active, so the result directly guides treatment. Subtyping, serology and antigen tests cannot detect resistance.
- A: Serology is slow, may fail after transplant and does not detect resistance.
- B: The subtype is already known and does not explain the rising viral load.
- D: A rapid antigen test is less sensitive than PCR and gives no resistance result.
Question 2 · Medium
On the same SARS-CoV-2 RT-PCR assay, patient A's sample has a Ct of 18 and patient B's a Ct of 33. Compared with patient B's sample, patient A's sample contains:
1. AMore PCR inhibitors
2. BA newer virus variant
3. CFar less viral RNA
4. DFar more viral RNA
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Answer: D · Far more viral RNA
The cycle threshold (Ct) is the PCR cycle at which the signal crosses the detection threshold. Each cycle roughly doubles the target, so a 15-cycle difference means about 2^15, or over 30,000-fold, more starting RNA in sample A. Ct values are only semi-quantitative, because sampling, transport and the assay all affect them. A high Ct can reflect early infection, late infection or a poor sample.
- A: Inhibitors delay amplification and raise the Ct, not lower it.
- B: The Ct reflects the amount of target, not the variant.
- C: A higher Ct, not a lower one, means less target RNA.
Question 3 · Hard
A laboratory confirms MERS-CoV by RT-PCR on two separate genomic targets, as WHO recommends. A sputum from a camel herder is positive on the upE screening assay (Ct 36) but negative on the ORF1a confirmation assay. What is the best next step?
1. AReport inconclusive; test another target and a new sample
2. BReport MERS-CoV not detected, since ORF1a is negative
3. CRepeat the upE assay alone and report that result
4. DReport MERS-CoV confirmed, since upE is positive
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Answer: A · Report inconclusive; test another target and a new sample
Under the WHO approach, a confirmed MERS-CoV case needs a positive result on two specific genomic targets, or one positive target plus sequencing of a second. A positive upE screen with a negative ORF1a, especially at a high Ct of 36, is inconclusive: it may be a low viral load or a false positive. The laboratory tests a further target such as the N gene or sequences the product, and asks for a repeat sample. Infection control precautions continue until the result is resolved.
- B: A negative confirmation assay at low viral load does not exclude infection.
- C: Repeating the screening assay alone cannot confirm the result.
- D: One positive target is not enough for confirmation under this rule.
Question 4 · Easy
The main animal source of human MERS-CoV infection in the Arabian Peninsula is:
1. APalm civets
2. BDromedary camels
3. CDomestic pigs
4. DDomestic poultry
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Answer: B · Dromedary camels
Dromedary camels are the main reservoir of MERS-CoV, and many camels in the Arabian Peninsula and Africa carry antibodies to it. People are infected through close contact with camels, possibly also through raw camel milk, and the virus can then spread between people, especially in hospitals. Bats are thought to be the original, ancestral source. Palm civets were linked to the SARS outbreak of 2003, not to MERS.
- A: Palm civets were linked to the original SARS outbreak in 2003.
- C: Pigs are linked to swine influenza, not MERS-CoV.
- D: Poultry are a source of avian influenza, not MERS-CoV.
Question 5 · Easy
Which swab is suitable for collecting a nasopharyngeal specimen for respiratory virus NAAT?
1. AFlocked swab on a plastic shaft
2. BCharcoal-coated swab in Amies gel
3. CCalcium alginate swab on an aluminium shaft
4. DCotton swab on a wooden shaft
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Answer: A · Flocked swab on a plastic shaft
Flocked swabs made of nylon or other synthetic fibres collect and release cells and virus well, and plastic or wire shafts do not interfere with testing. The swab is placed in viral or universal transport medium. Calcium alginate and wooden shafts contain substances that can inhibit PCR or harm viruses. Charcoal swabs in gel are designed for bacterial culture.
- B: Charcoal swabs in Amies gel are for bacterial culture, not viral NAAT.
- C: Calcium alginate can inhibit PCR and inactivate some viruses.
- D: Wooden shafts and cotton can release substances that inhibit NAAT.
Question 6 · Easy
Tubes of respiratory specimen from a suspected MERS-CoV case are opened and aliquoted for PCR, before the virus is inactivated, in a:
1. AChemical fume hood
2. BHorizontal laminar flow clean bench
3. CClass II biological safety cabinet
4. DOpen bench, with gown and mask
Answer and explanation in LearnFree account
Question 7 · Easy
Influenza A viruses are divided into subtypes, such as H1N1 and H3N2, by their surface:
1. ACapsid and polymerase proteins
2. BSpike and envelope proteins
3. CHemagglutinin and neuraminidase
4. DMatrix and nucleoprotein
Answer and explanation in LearnFree account
Question 8 · Medium
A nasopharyngeal swab in viral transport medium for SARS-CoV-2 RT-PCR cannot be tested for 5 days. Following CDC guidance, how should it be stored?
1. AAt room temperature, about 25 °C
2. BAt 2-8 °C in a refrigerator
3. CFrozen at -70 °C or colder
4. DFrozen at -20 °C
Answer and explanation in LearnFree account
Question 9 · Hard
A poultry worker has severe pneumonia. Influenza RT-PCR on a nasopharyngeal swab: influenza A matrix gene detected (Ct 27); H1pdm09 not detected; H3 not detected; influenza B not detected. What is the most appropriate action?
1. AReport it as inconclusive and close the request
2. BReport it as seasonal influenza A(H3N2) infection
3. CRefer it urgently as a possible novel influenza A
4. DReport it as influenza A not detected
Answer and explanation in LearnFree account
Question 10 · Medium
Which specimen is preferred for MERS-CoV RT-PCR in a patient with pneumonia?
1. AA lower respiratory sample, such as sputum or BAL
2. BA stool sample in viral transport medium
3. CA blood film stained with Giemsa
4. DA midstream urine sample collected in a sterile cup
Answer and explanation in LearnFree account

Virology and respiratory viruses

On the same SARS-CoV-2 RT-PCR assay, patient A's sample has a Ct of 18 and patient B's a Ct of 33. Compared with patient B's sample, patient A's sample contains:

A laboratory confirms MERS-CoV by RT-PCR on two separate genomic targets, as WHO recommends. A sputum from a camel herder is positive on the upE screening assay (Ct 36) but negative on the ORF1a confirmation assay. What is the best next step?

The main animal source of human MERS-CoV infection in the Arabian Peninsula is:

Which swab is suitable for collecting a nasopharyngeal specimen for respiratory virus NAAT?

Tubes of respiratory specimen from a suspected MERS-CoV case are opened and aliquoted for PCR, before the virus is inactivated, in a:

Influenza A viruses are divided into subtypes, such as H1N1 and H3N2, by their surface:

A nasopharyngeal swab in viral transport medium for SARS-CoV-2 RT-PCR cannot be tested for 5 days. Following CDC guidance, how should it be stored?

A poultry worker has severe pneumonia. Influenza RT-PCR on a nasopharyngeal swab: influenza A matrix gene detected (Ct 27); H1pdm09 not detected; H3 not detected; influenza B not detected. What is the most appropriate action?

Which specimen is preferred for MERS-CoV RT-PCR in a patient with pneumonia?