Herpes Simplex Virus (HSV-1 & HSV-2) anti-viral resistance genotyping

Herpes Simplex Virus (HSV-1 & HSV-2) anti-viral resistance genotyping.

• Herpes simplex 1
• Herpes simplex 2

HSV-1 or HSV-2 positive lesion swab 1 mL (0.5 mL minimum), CSF 1 mL (0.5 mL minimum), or plasma 1 mL (0.5 mL minimum). Viral cultures 1 mL (0.5 mL minimum) may also be sent. Minimum volume will not allow repeat testing of samples with equivocal results.

For lesion swabs, place swabs in 2-3 mL of viral transport medium (VTM) for a minimum of 1 hour and then remove the swab.  For plasma, collect blood in plasma preparation tube (PPT) or K2EDTA tube (plastic; lavender or pink top). Do not use heparin as an anti-coagulant. Centrifuge the tube at room temperature at 1,100 RCF for 10-15 min to separate the blood. Aliquot plasma into separate tube. Collect CSF in a dry tube subject to classic conditions for carrying out lumbar punctures. For viral cultures, collect infected cells and media. 

Plasma and CSF may be stored and shipped refrigerated to NML within 48h of sample collection or must be stored and shipped frozen.  Viral cultures must be stored frozen at all times and shipped on dry ice. 

Shipping of specimens shall be done by a TDG certified individual in accordance with TDG regulations. For additional information regarding classification of specimens for the purposes of shipping, consult either Part 2 Appendix 3 of the TDG Regulations or section 3.6.2 of the IATA Dangerous Goods Regulations as applicable.

Suspected anti-viral resistant HSV-1 or HSV-2, characterized by no improvement (or relapse) in HSV-1 or HSV-2 lesions.  Drug resistant HSV is more commonly isolated from immunocompromised patients. (3).

Results of HSV-1 or HSV-2 type-specific PCR including copy number. Completed “Viral STI, Polyoma and Herpesvirus Testing” requisition form including sender lab name, address and telephone number, patient identifier, sex, specimen reference #, type of specimen, date collected, test requested and any other relevant clinical information.

The specimen should be positive for HSV-1 or HSV-2 DNA and results included with the requisition form. 

Regions of the thymidine kinase ( UL23) and polymerase (UL30) genes that include known acyclovir and derivative anti-viral resistant mutations are amplified by PCR and sequenced  by Sanger sequencing (1,2). The HSV-1 or HSV-2 sequence data are then analysed and compared to wild-type reference sequences and mutations and/or frameshifts are identified. An online tool is used for HSV-1 analysis (4); the HSV-2 database is limited and based on published results (5). Please note that no phenotypic assays are performed. Interpret results with caution as this assay is for research purposes only.

10 calendar days.  STAT samples may be considered under special circumstances.  Please contact the lab to inquire about shorter turn around time.

1. Doris Chibo, Julian Druce, Joe Sasadeusz, Chris Birch, Molecular analysis of clinical isolates of acyclovir resistant herpes simplex virus, Antiviral Research,Volume 61, Issue 2, 2004, Pages 83-91, ISSN 0166-3542, https://doi.org/10.1016/j.antiviral.2003.08.018
2. Emilie Frobert, Jean-Claude Cortay, Tadamasa Ooka, Fatiha Najioullah, Danielle Thouvenot, Bruno Lina, Florence Morfin, Genotypic detection of acyclovir-resistant HSV-1: Characterization of 67 ACV-sensitive and 14 ACV-resistant viruses,Antiviral Research, Volume 79, Issue 1, 2008, Pages 28-36, ISSN 0166-3542, https://doi.org/10.1016/j.antiviral.2008.01.153.
3. Jiang, YC., Feng, H., Lin, YC. et al. New strategies against drug resistance to herpes simplex virus. Int J Oral Sci 8, 1–6 (2016). https://doi.org/10.1038/ijos.2016.3
4. https://dna.informatik.uni-ulm.de/software/mra/app/index.php?plugin=form
5. Andreas Sauerbrei, Kathrin Bohn-Wippert, Marisa Kaspar, Andi Krumbholz, Matthias Karrasch, Roland Zell, Database on natural polymorphisms and resistance-related non-synonymous mutations in thymidine kinase and DNA polymerase genes of herpes simplex virus types 1 and 2, Journal of Antimicrobial Chemotherapy, Volume 71, Issue 1, Pages 6-16. (2016) https://doi.org/10.1093/jac/dkv285

Pathogen Safety Data Sheets: Infectious Substances – Herpes simplex virus