When to do HIV drug resistance testing

  HIV drug resistance is one of the important reasons for the failure of HIV antiretroviral therapy. The 2020 edition of the National AIDS Testing Technical Specifications will continue to include HIV drug resistance testing. What do we need to know about HIV drug resistance testing?
What is HIV drug resistance

  HIV resistance, also known as HIV resistance, refers to the phenomenon of reduced sensitivity or insensitivity to the inhibitory effects of antiviral drugs caused by genetic variation in HIV.
Where does HIV drug resistance come from?

  (1) Direct infection. Primary drug resistance, also known as disseminated drug resistance, occurs when the infected strain is a drug-resistant strain of HIV.
  (2) Drug selection. HIV is a virus that replicates quickly and is highly mutated. In the absence of antiviral drugs, HIV-resistant mutants will appear randomly, but have no replication advantage and will not accumulate in the body; during antiviral therapy, when the drug concentration When insufficient, HIV replication cannot be completely inhibited, and the replication advantage of HIV drug-resistant mutants will appear, and then the drug-resistant mutants will gradually accumulate and increase, resulting in secondary drug resistance, also known as acquired drug resistance.
How to detect HIV drug resistance

  (1) Genotype detection. Detection of drug resistance-related mutations in the HIV genome by sequencing or gene chip hybridization. Genotype detection usually uses sequencing. The detection process is as follows: using HIV genomic RNA as a template, reverse transcription PCR to amplify the target gene fragment, sequencing to obtain the HIV gene sequence, and compare it with the wild-type sequence to determine whether there are drug resistance-related genes. Mutations, and then associated with the degree of resistance through the algorithm of the drug resistance database. Sample types can be plasma and dried blood spots. The sequencing method usually adopts the Sanger sequencing method. With the development of sequencing technology, the deep sequencing method is gradually used.
  (2) Phenotypic detection. Also known as in vitro drug susceptibility test, it is expressed as 50% inhibitory concentration (IC50), and the IC50 of the tested strain is compared with that of the wild-type strain, and the degree of drug resistance is evaluated by fold change (FC). The virus to be tested can be directly isolated and cultured from the PBMC of the infected person, or it can be a recombinant virus containing the HIV gene of the infected person.
  Generally speaking, genotype testing is simpler in operation and has a short testing period, so it is used more clinically, but phenotype testing has advantages for complex mutations and drug resistance testing of new drugs.
How to read the results of genotype resistance test

  (1) Representation of drug-resistant mutations. The drug resistance mutation is the change in the nucleotide sequence of the HIV gene, but it is expressed by the change of the amino acid. The amino acid was mutated from lysine (K) to asparagine (N).
  (2) Drug resistance interpretation system. Refers to a database with a drug resistance degree algorithm, through which the drug resistance mutation result can be converted into a drug resistance degree. For example, in the widely used Stanford HIVdb resistance interpretation system, the K103N mutation is highly resistant to nevirapine (NVP) and efavirenz (EFV).
Considerations for HIV drug resistance testing

  (1) Detection timing. Since HIV is very easy to mutate, when drug-resistant patients stop or discontinue treatment, and there is no antiviral drug in the body, the replication advantage of the wild strain will appear and gradually accumulate, and eventually no drug resistance will be detected. Therefore, the best time for drug resistance detection is For patients on antiviral therapy, or within one month of interruption of therapy.
  (2) Be alert to cross-contamination between samples. Blank and negative controls should be set during the detection process, and cross-contamination should be excluded through sequence evolutionary tree analysis.
  (3) When no drug-resistant mutation is detected, the existence of a lower proportion of drug-resistant strains cannot be ruled out. This is because when the proportion of drug-resistant strains in individual viral quasispecies is less than 10-20%, its presence is usually undetectable by Sanger sequencing.

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