Novel codon T215C/D/N/S mutations have been described in antiretroviral-naive patients infected with zidovudine-resistant HIV-1-carrying 215F/Y. Viruses with codon 215-revertant genotypes are sensitive to zidovudine and stavudine, but rapidly develop resistance under exposure to these drugs. In pre-treated HIV-infected patients who discontinue zidovudine or stavudine there is no evidence of reversion to these unusual genotypes, because of an overgrowth of pre-existing wild-type viruses. Therefore, these novel codon 215 mutations almost always reflect a past primary infection with zidovudine-resistant viruses.
Resistance to zidovudine generally results from the selection and accumulation of several mutations within the HIV reverse transcriptase gene, involving codons 41, 67, 70, 210, 215, and 219, which tend to emerge in an ordered fashion [1,2]. Studies of HIV-1 seroconverters infected with viruses carrying the T215F/Y resistance mutation have shown that in the absence of drug therapy in the new host the resistant virus tends to shift to 215C, 215D, 215N, or 215S [3–5]. These new genotypes represent revertants from 215F/Y, but exert improved fitness .
In a recent report form the United States, codon 215 revertants were recognized in 3.3% of 603 untreated individuals , reflecting indirectly the rate of transmission of zidovudine-resistant viruses in that community. Moreover, the same authors pointed out that viruses with codon 215-revertant mutations were fully sensitive to zidovudine, but readily gave rise to zidovudine-resistant 215Y mutants in vitro in the presence of zidovudine  or stavudine .
Given that most currently available drug-resistance assays interpret as sensitive the sequences from viruses harbouring codon 215-revertant mutations, we were interested to know the prevalence of these novel genotypes among antiretroviral-experienced patients. A substantial proportion of individuals on antiretroviral therapy currently discontinue zidovudine or stavudine for virological failure or toxicity issues, and to our knowledge no data exist on the rate of these novel codon 215 mutations in treatment-experienced patients. Given that they may be prone to the rapid selection of 215F/Y under zidovudine or stavudine, it could be misleading to ignore them when planning salvage therapies.
Plasma samples collected from a total of 530 HIV-positive individuals experiencing virological failure and on regular follow-up at different hospitals across Spain, and for whom drug-resistance testing had been requested during 2001, were examined. Genetic sequencing was performed using an ABI Prism 3100 automatic sequencer (Applied Biosystems, Foster City, CA, USA). At the time of the analysis, the mean plasma viral load and the mean CD4 cell count was 43 490 HIV-RNA copies/ml, and 392 cells/μl, respectively. More than half of the individuals (n = 271; 51%) had wild-type codon 215 viruses, whereas codon 215F/Y-resistant genotypes were found in 249 individuals (47%). Only 10 patients (2%) harboured HIV-1 with unusual codon 215-revertant forms. These substitutions were Ile (n=4), Val (n = 3), Cys (n = 2), and Ser (n = 1) (see Table 1). In all instances, the revertant forms at codon 215 were present in viruses harbouring a median of three classic zidovudine resistance mutations (41L, 67N, 70R, 210W, 219E/Q) (range 1–5), K70R being recognized in up to eight of them. It should be kept in mind that when at least two of these changes are present, including the 70R substitution, a significant loss of susceptibility to zidovudine is produced [7,8]. Therefore, all those patients should have been considered zidovudine resistant, even ignoring the unusual polymorphisms at codon 215.
Although one quarter of the study population (n = 134; 25.3%) was not taking zidovudine or stavudine when the analysis was performed, all subjects with codon 215 revertants but one were taking either zidovudine (n = 1), stavudine (n = 4), or stavudine plus didanosine (n = 4), as part of their antiretroviral regimen. Therefore, codon 215-revertant viruses were not particularly prevalent among patients not receiving zidovudine or stavudine. This observation argues against their evolution from viruses in which 215Y/F was selected in the past under regimens containing zidovudine or stavudine, or both.
The novel codon 215I/V/S/C/N/D variants arise as result of single nucleotide changes from 215Y or 215F (Fig. 1). Their recognition provides a kind of fossil record of the transmission of zidovudine-resistant strains in the past. Return to the wild-type codon from 215Y/F requires two nucleotide transversions, which makes it very unlikely in patients who were primarily infected with T215Y/F-resistant viruses (see Fig. 1). Our data suggest that the rate of codon 215-revertant viruses is very low (2%) among HIV-infected patients failing antiretroviral therapy. As long as these subjects were infected with codon 215 wild-type viruses and selected 215F/Y after failing zidovudine or stavudine, an overgrowth of the originally acquired wild-type virus almost always takes place when discontinuing zidovudine or stavudine, without an opportunity for the selection of codon 215 revertants.
In summary, in pre-treated HIV-infected patients who discontinue zidovudine or stavudine, there is no evidence of the selection of the novel codon 215-revertant forms, probably because of an overgrowth of pre-existing wild-type viruses. Therefore, these genotypes should almost always be considered a marker of past primary infection with a zidovudine-resistant virus.
The authors would like to thank Professor González-Lahoz for his continuous support.
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