Novel Influenza A (H1N1) Laboratory Testing

Abhay Chowdhary
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Testing for Novel Influenza A (H1N1) Virus
Clinicians should consider testing suspected cases of novel influenza A (H1N1), especially those with severe illness, by obtaining an upper respiratory specimen to test for novel influenza A (H1N1) virus.

The following should be collected as soon as possible after illness onset: nasopharyngeal swab, nasal aspirate or a combined nasopharyngeal swab with oropharyngeal swab. If these specimens cannot be collected, a nasal swab or oropharyngeal swab is acceptable. For patients who are intubated, an endotracheal aspirate should also be collected.

Bronchoalveolar lavage (BAL) and sputum specimens are also acceptable. Specimens should be placed into sterile viral transport media (VTM) and immediately placed on ice or cold packs or at 4°C (refrigerator) for transport to the laboratory. Recommended infection control should be followed.

Ideally, swab specimens should be collected using swabs with a synthetic tip (e.g. polyester or Dacron®) and an aluminum or plastic shaft. Swabs with cotton tips and wooden shafts are not recommended. Specimens collected with swabs made of calcium alginate are not acceptable. The swab specimen collection vials should contain 1-3ml of viral transport medium (e.g. containing, protein stabilizer, antibiotics to discourage bacterial and fungal growth, and buffer solution).

All respiratory specimens should be kept at 4°C for no longer than 4 days.

Clinical specimens should be shipped on wet ice or cold packs in appropriate packaging. All specimens should be labeled clearly and include information requested by your state public health laboratory. Suspected case specimens shipped from the state public health laboratory to CDC should include all information required for seasonal influenza surveillance isolate or specimen submission.

Real-time RT-PCR is the recommended test for confirmation of novel influenza A (H1N)1 cases. Currently, novel influenza A (H1N1) virus will test positive for influenza A and negative for H1 and H3 by real-time RT-PCR. If reactivity of real-time RT-PCR for influenza A is strong (e.g. Ct <30) it is more suggestive of a novel influenza A (H1N1) virus. Confirmation as novel influenza A (H1N1) virus by real-time RT-PCR was originally performed only at CDC, but at this time it is available in Haffkine Institute, Mumbai, NIV, Pune, NICD, Delhi and few other laboratories.

Some commercially available rapid tests can distinguish between influenza A and B viruses. A patient with a positive rapid test for influenza A may meet criteria for a suspected case of novel influenza A (H1N1) virus infection. However, it is not possible to differentiate from seasonal influenza A viruses. Also, these tests have unknown sensitivity and specificity to detect human infection with novel influenza A (H1N1) virus in clinical specimens, and have suboptimal sensitivity to detect seasonal influenza viruses. Therefore, a negative rapid test could be a false negative and should not be assumed a final diagnostic test for novel influenza A (H1N1) virus infection.

These tests can distinguish between influenza A and B viruses. A patient with a positive for influenza A by Immunofluorescence may meet criteria for a suspected case. However, it is not possible to differentiate from seasonal influenza A viruses. Immunofluorescence depends upon the quality of a clinical specimen, operator skills, and has unknown sensitivity and specificity to detect human infection with novel influenza A (H1N1) virus in clinical specimens. Therefore, a negative Immunofluorescence could be a false negative and should not be assumed a final diagnostic test for novel influenza A (H1N1) virus infection.

Isolation of novel influenza A (H1N1) virus is diagnostic of infection, but may not yield timely results for clinical management hence not recommended. A negative viral culture does not exclude infection with novel influenza A (H1N1) virus. It is requires Bio Safety Level 3 facility.

CDC recommends that people with influenza-like illness remain at home until at least 24 hours after they are free of fever (100° F [37.8°C]), or signs of a fever without the use of fever-reducing medications. This is a change from the previous recommendation that ill persons stay home for 7 days after illness onset or until 24 hours after the resolution of symptoms, whichever was longer. The new recommendation applies to camps, schools, businesses, mass gatherings, and other community settings where the majority of people are not at increased risk for influenza complications.
In India the behaviour of virus is very mild and is not of major concern. However due care must be ensured for children and elderly. The panic button need not be pressed yet.
Please remember, saying namaste with folded hands from a distance is a good age old Indian way to greet and avoid unnecessary contact with others.


Novel Influenza A (H1N1) Laboratory Testing Novel Influenza A (H1N1) Laboratory Testing 01/06/2009
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