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Assay Sheet

Test ID

8700 2009 H1N1 Influenza A Antiviral Resistance

CPT Code

83891, 83894, 83898, 83904 (x2), 83912

Clinical Utility

The emergence of the 2009 H1N1 influenza A virus has lead to increased vigilance in monitoring these infections. Upon proper diagnosis, patients infected with 2009 H1N1 have been effectively treated with oseltamivir.  However, oseltamivir-resistant 2009 H1N1 strains have emerged, and incidence is likely to rise due to  increased drug exposure and horizontal transmission.  Studies have indicated resistance is conferred by a single base mutation (H275Y) in the neuraminidase gene. Utilizing sequencing technology to identify this mutation has proven effective in confirming infections with oseltamivir-resistant 2009 H1N1, allowing physicians to reassess treatment strategies.

Procedure

Extraction of Influenza A nucleic acid from respiratory specimen. Reverse transcription of viral RNA, then amplification of cDNA using conventional PCR followed by gene sequencing. Sequencing analysis provides information on a selected location in the neuraminidase gene implicated in 2009 H1N1 Influenza A antiviral resistance to oseltamivir.

Specimens

Bronchial Lavage/Bronchial Wash: 1 to 3 ml, collected in sterile, screw-cap tube; ship ambient.

Sputum: 1-2 ml submitted in a sterile, screw-top tube; ship ambient.

Throat Gargle:  Instruct patient to gargle with 2-3ml sterile saline.  Expectorate into sterile cup, then transfer contents to sterile, screw-cap tube for shipment; ship ambient.

Upper respiratory aspirate (NP aspirate, nasal aspirate, tracheal aspirate, etc.): Instill 1 to 2 ml sterile saline into desired location and gently aspirate contents. Place collected fluid into sterile, screw-cap tube; ship ambient. 

Upper respiratory swab (NP swab, throat swab): Swab desired location with sterile, flexible shaft swab, preferably a flocked swab. Place swab into 1 to 2 ml sterile saline, M4, or viral transport media in sterile, screw-cap tube. Do not use calcium alginate swab or wood shafted swab; ship ambient.

All suction-type collection devices are inappropriate for specimen transport. Transfer specimen into sterile, leakproof tube for transport.

Causes for rejection

Wood shafted swab, calcium alginate swab. Call ViraCor at 800-305-5198 if specimen is greater than 96 hrs old.

Specificity

The primers utilized for conventional PCR and sequencing in this assay are specific for 2009 H1N1 Influenza A virus based on similarity search algorithms.  Additionally, no cross reactivity was detected when tested against seasonal influenza A subtypes H1N1 and H3N2, influenza B,  RSV A, RSV B, parainfluenza 1, parainfluenza 2, parainfluenza 3, hMPV, rhinovirus, and adenovirus.

Assay Range

Qualitative results (Mutation detected at H275Y/None Detected) for antiviral resistance in 2009 H1N1 Influenza A. 

Turnaround Time

3 days (Monday through Friday)

Shipping

Ship Monday through Friday. Friday shipments must be labeled for Saturday delivery. All specimens must be labeled with patient's name and collection date. Multiple tests can be run on one specimen. Ship specimens FedEx Priority Overnight to: