This document discusses laboratory diagnosis of COVID-19. It describes how the disease spreads and appropriate sample types and collection methods. Molecular RT-PCR tests targeting SARS-CoV-2 genes like E, RdRp, N and ORF1ab are recommended for diagnosis. Serological and antigen tests can also be used but have limitations. Proper sample handling and accurate test interpretation are important for diagnosis. Laboratory networking is important for managing the COVID-19 pandemic.

1. Laboratory diagnosis of COVID-19
Dr Adil Raza
Associate Professor ,
Department of Microbiology,
JNMC, AMU, Aligarh

2. • Coronavirus disease 2019 (COVID-19) is a
severe acute respiratory infection caused by
the novel coronavirus severe acute
respiratory syndrome coronavirus 2
(SARSCoV-2).
• The disease primarily spreads via close contact
of respiratory droplets generated by infected
individuals

3. Sample collection

4. • Adequate standard operating procedures
(SOPs) should be followed before
collecting any specimen, packaging,
storage and transport.

5. Samples to be collected
• Upper respiratory specimens
– Oropharyngeal swab
– Nasopharyngeal swab
– Anterior nasal swab
• Lower respiratory specimens
– Sputum
– BAL
– Endotracheal aspirate
• Other specimens
– Stool
– Blood
– Paired serum
– Lung

6. Viral Transport Media (VTM)

10. Sample collection
• Nasopharyngeal swab
• Oropharyngeal swab
• Anterior nasal swab

11. Normal Anatomy

12. Nasopharyngeal Swab

14. Anterior nasal swab

18. • NOTE
• Calcium alginate and cotton swabs should not be
used as these are inhibitory to viruses.
• Only dacron and polyester swabs should be used
• Wooden stick should not be used.
• Temperature should be maintained 2 to 80 C during
transport, if sample delivery is assured within 5 days.
• If is more than 5 days it should be -700 C.

19. Laboratory testing for COVID-19 virus
1. Nucleic acid amplification tests (NAAT)
2. Serological testing
3. Antigen detection
4. Viral sequencing
5. Viral culture

20. Molecular (rRT-PCR) tests have generally
targeted a combination of the following
genes:
1) Envelope (E)
2) RNA dependent RNA polymerase (RdRp)
3) Nucleocapsid (N)
4) Open reading frame 1ab (ORF 1ab)
Nucleic acid amplification test (NAAT): rRT PCR

21. Sampling
Nucleic acid
isolation
RT
Real time
PCR
amplification
Swab RNA cDNA PCR
Figure: General work flow

23. Test interpretation (WHO):
1. Screening test (+) and confirmatory test (+): positive
for COVID-19 (SARS-CoV-2 detected).
2. Screening test (+) and confirmatory test (-):
negative for COVID-19 (SARS-CoV-2 not detected).
3. Screening test (-) and confirmatory test (-):
negative for COVID-19 (SARS-CoV-2 not detected).
4. Screening test (-) and confirmatory test (+): retest
or refer to a reference laboratory for additional
testing.

24. • Based on the detection of IgM/IgG antibodies.
• Antibody test results are especially important
for detecting previous infections in people
who had few or no symptoms.
• Cross reactivity to other coronaviruses can be
challenging.
• Not recommended by WHO or CDC for
diagnosis.
Serological testing

25. Figure: Variations in the Levels of SARS-CoV-2 RNA, IgM and IgG after infection.

26. Antigen detection
• COVID-19 Antigen test is a lateral flow
immunochromatographic assay intended for
the qualitative detection of the nucleocapsid
protein antigen from SARS-CoV-2
• This test is authorized for use at the Point of
Care (POC).

28. Antigen detection
• Negative results should be treated as
presumptive, and do not rule out SARS-CoV-2
infection.
• Additional confirmatory testing with a
molecular test for positive results may also be
necessary

29. Viral sequencing
• Can be useful to monitor
– viral genome mutations that might affect the
performance of medical countermeasures,
including diagnostic tests.

30. Viral culture
• Not recommended as a routine
diagnostic procedure.
• Human airway epithelial cell lines were
used for the initial isolation of the
virus.

31. Conclusions
• Early diagnosis is the key for prompt
management of COVID-19.
• Serological and molecular assays together will
further strengthen the diagnosis of SARS-CoV-2.
• Laboratory networking is the need of the hour for
real-time diagnosis of COVID-19