Molecular Detection of Drug Resistant Mutants

Molecular detection of nucleoside/nucleotide analog resistance by line probe hybridization assay or by sequence analysis.

Serum or plasma sample. Minimum volume required for serum or plasma – 1.0 mL.

Collect blood in serum separator tubes (SST) or EDTA tubes.

Store samples frozen until shipped for testing. Ship frozen 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.

Patient on previous or current antiviral therapy for Hepatitis B infection. For patients on current treatment, HBV DNA viral load levels suggest antiviral failure (breakthrough) or nonresponse (i.e., confirmed rise on 2 occasions at least 1 month apart in HBV DNA by > 1 log10 IU/ml from treatment initiation, or HBV DNA reduction < 1 log10 IU/ml, respectively). Previous antiviral treatment warrants drug resistance testing to determine subsequent therapies.

Completed Molecular and Immuno Diagnostics on Hepatitis requisition including sender name, address and telephone number. Patient name or identifier (referring specimen lab #), date of birth, suspected exposure, test(s) requested and all current or previous antiviral medications. Type of specimen and date collected. If possible, include the clinical history and lab results that have already been done at local or provincial laboratories.

Conventional PCR followed by line probe hybridization assay. Reactivity to drug resistance or compensatory mutation specific probes indicates resistance or intermediate sensitivity to certain antiviral therapies dependent on the mutation site (Lamivudine resistance: rtL80V/I, rtV/G173L, rtL180M, rtA181T/V, rtM204V/I/S; Adefovir resistance: rtA181T/V, rtN236T; Entecavir resistance: rtL180M, rtT184S/C/G/A/I/L/F/M, rtS202G/C/I, rtM204I/V, rtM250V/I/L; Tenofovir resistance: rtA181T/V, rtA194T, rtN236T; Telbivudine resistance: rtL180M, rtA181T/V, rtM204I). LiPA analysis will be followed up by sequence analysis in cases of probe site negativity.

19 calendar days.

1. Osiowy C, Villeneuve J, Heathcote EJ, et al. Detection of rtN236T and rtA181V/T mutations associated with resistance to adefovir dipivoxil in samples from patients with chronic hepatitis B virus infection by the INNO-LIPA HBV DR line probe assay (version 2). J Clin Microbiol 2006. 44:1994-1997.
2. Degertekin B, Hussain M, Tan J, et al. Sensitivity and accuracy of an updated line probe assay (HBV DR v.3) in detecting mutations associated with hepatitis B antiviral resistance. J Hepatol 2009. 50:42-48.