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HIV-1 superinfection with a triple-class drug-resistant strain in a patient successfully controlled with antiretroviral treatment
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AIDS July 2014
Castro, Erikaa,c; Zhao, Hongb; Cavassini, Matthiasa; Mullins, James I.b; Pantaleo, Giuseppec; Bart, Pierre-Alexandrec,d aService of Infectious Diseases, Centre Hospitalier Universitaire Vaudois, Lausanne, Switzerland bDepartments of Microbiology, Medicine and Laboratory Medicine, University of Washington School of Medicine, Seattle, Washington, USA cLaboratory of AIDS Immunopathogenesis, Service of Immunology and Allergy dService of Internal Medicine, Centre Hospitalier Universitaire Vaudois, Lausanne, Switzerland.
We report a case of HIV-1 superinfection (HSI) with a clade B, triple-class resistant virus in a patient successfully controlling viremia with continuous combination antiretroviral therapy started 8 years earlier during primary HIV infection. The course of HIV infection prior to HSI was monitored in both the source partner and recipient (8 and 11 years, respectively) and 4 years following HSI. This case report demonstrates re-infection with HIV-1 despite effective combination antiretroviral therapy.
HIV-1 superinfection (HSI), that is, infection with a second strain after the first has been established, has been reported since 2002, mainly involving HIV group M clades [1-6], and in a smaller proportion as inter-group recombinant forms [7,8]. Overall, HSI has been most often observed in untreated patients, during treatment interruption, and in seroconcordant couples with poor viral suppression [9]. Here, we report the onset and 4-year follow-up of HSI with a triple-class resistant clade B virus in a man on effective combination antiretroviral therapy (cART).
Briefly, blood samples were obtained from two men (M1 and M2) chronically infected with HIV-1 and sexual partners since 2006. M2 was diagnosed with HIV infection in 1994 at stage A2 with full Western blot seroconversion, and developed triple-class antiretroviral drug resistance as a consequence of weak adherence to ART, leading to virologic failure. M2 experienced uncontrolled viremia (range 3-4 logs of viral RNA) until September 2008 when a salvage treatment regimen reduced his viral load to undetectable limit for the first time (Fig. 1 a). M1 was diagnosed in 2000 with primary HIV infection, initiated cART in 2000, and remained on cART with undetectable viremia and no drug resistance mutations through the end of 2007 (Fig. 1 b). cART first regimen consisted of zidovudine (ZDV) + lamivudine (3TC) + efavirenz (EFV). In 2001, EFV was replaced by boosted lopinavir (LPV/r) due to EFV neurologic intolerance. The therapy was simplified in 2002 with the association of ZDV/3TC/abacavir (ABC) (Trizivir). M1 was also a vaccine-recipient in the therapeutic HIV vaccine trial TheraVac01 [12]. Briefly, the trial was an open-label one-arm study that took place in Lausanne, Switzerland. All patients (n = 10) were immunized with New York vaccinia virus expressing HIV-1 clade B antigens (NYVAC-B) (Gag/Pol/Nef polygene of HIV IIIB, and Env clade B of HIV BX08) intramuscularly (10E7.4 cell culture infectious dose 50%/ml) at weeks (W)0 and W4. M1 was assigned the code TH#04 and was immunized on May 17 (W0) and on June 14 (W4) 2006, respectively.
In February 2008, M1 presented with a plasma viral load of 280 copies/ml, which increased over the following year. Genotypic analysis from 2008 onward revealed 25 new drug-resistance mutations to nucleoside reverse transcriptase inhibitor (NRTI) (6), non NRTI (NNRTI) (5) and protease inhibitor (14). Of note, this resistance profile shared 22 of 23 mutations (96%) found contemporaneously in M2. The shared mutations included NRTI resistance mutations 41L, 74I, 184V, and 215Y; NNRTI mutations 98G, 103N, and 108I; and protease inhibitor mutations 10V, 13V, 20R, 32I, 33I, 46I, 47V, 50V, 71I, and 77I. M1 also developed 75T (NRTI), 115/Y (NNRTI) and 82A (protease inhibitor) polymorphisms.
We obtained viral gene sequences from M1 from July 2000 [n = 1,1 ~9 kb near full-length genomes (NFLG)], February 2008 (n = 12 gag and env genes) and May of 2008 (n = 14 gag and n = 16 env genes), and from M2 in January (n = 10 NFLG) and March of 2008 (n = 7 NFLG). All 86 viral sequences were assigned to clade B. Additionally, all sequences from M1 obtained from 2008 clustered with M2 sequences from the same year and were phylogentically unrelated to M1 sequences from 2000 (data not shown). This indicated HSI of M1 with substantial or complete replacement with virus from M2. No recombination between the M1 and M2 strains was observed.
The viral load of patient M1 increased from undetectable to above 3 logs after HSI, decreasing progressively from 2009 to 2012 without cART modification. A continuous drop of CD4+ cells of at least 10% was observed from time of HSI detection in February 2008 (30.8%) through April 2011 (18%). However, in April 2012, the CD4+ cell count for M1 recovered to 25.4% (757 cell/μl) (Fig. 1b). The follow-up for patient M2 was taken over by a general practitioner in 2009 after three consecutive undetectable viral loads following start of salvage antiretroviral treatment (Fig. 1a).
We detected superinfection and replacement by HIV-1 clade B triple-class resistant virus in a patient on long-term cART controlled infection, with the initial indicator of HSI being a detectable and increasing viral load. Other resistance mutations not found in the superinfecting strain also emerged following HSI, though their origin is unclear. They could have been present as minority populations prior to superinfection or transmitted with the superinfecting strain but below the detection level of our assessment of M2's quasispecies (no additional specimen was available for massively parallel sequencing). Despite HSI with a triple-class resistant virus, the ART for M1 remained unchanged. During the 4 years of additional follow-up, a continuous drop of viral load occurred, followed by CD4+ recovery in the past year (Fig. 1b). The impact of HSI in CD4+ decline and disease progression following subtype B coinfection was initially suggested by Gottlieb et al.[13]. Evidence of CD4+ T-cell decline as the initial indicator of HSI was also reported in untreated patients during primary HIV infection as well as in untreated elite controllers [14-16].
In contrast to previous studies of superinfecton among long-term known seroconcordant couples undergoing ART [9], this study underscores the fragile chemoprophylactic barrier exerted by ART despite excellent adherence. Apart from the complex resistance profile of the superinfecting strain present in patientM2, we cannot rule out the role of viral escape to pre-existing cytotoxic T-lymphocyte or antibody responses in the establishment of the second infection, which involves the same clade, and therefore, to some extent challenged similar immune signatures in the superinfected individual, as reported elsewhere [17,18]. Of interest, the HIV-specific T-cell responses of M1 were enriched following NYVAC-B immunizations (W0 and W4) and 21 months prior to superinfection [12]. Briefly, two novel Env/Pol vaccine-induced responses emerged at W2 and remained present throughout the study with a clear decline by W48 [12], greater than 1 year prior to HSI. The protective role of these vaccine-induced responses to prevent a second infection with a resistant virus, as well as their longevity, remains unproven. Haplotypes such as human leukocyte antigen (HLA)-B3503 have been shown to be associated with HIV-1 superinfection susceptibility as a consequence of late or weak immune response priming [16]. In this regard, patient M1 carries HLA haplotype A*30/23, B*35/44, DRB1*14/04 (performed by PCR-SSO using LabType kit on the Luminex System). He also had no CCR5-delta 32 mutations (in-house PCR modified from Wilkinson et al.[19]). In summary, patient M1's superinfection onset agrees with the current knowledge on host factors and re-exposure to a resistant strain, and demonstrates the previously unexpected scenario of re-infection during well established chronic infection, despite continuously suppressed viremia with cART started during primary HIV infection.

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