Search

OBJECTIVE: To investigate the mechanism explaining the persistence of human immunodeficiency virus (HIV) type 1 variants with multiple protease inhibitor (PI)-resistance mutations in the absence of PI therapy. METHODS: Longitudinal genotypic analyses were performed on sequential samples obtained from 2 HIV-1-infected patients who had stopped PI therapy for 4 years. Replication capacity (RC) was determined using recombinant viruses. Subsequently, the effect that changing individual protease mutations back to wild type has on RC was analyzed. RESULTS: We observed prolonged persistence (up to 4 years) of viruses with multiple protease mutations after PI therapy was stopped, despite the fact that the RC of the viruses was severely reduced. Forcing the virus to evolve toward wild type by changing individual protease mutations to wild type was unsuccessful, because all variants displayed a decreased RC in comparison with that of their predecessors. CONCLUSIONS: We propose compensatory fixation as a mechanism for the in vivo persistence of variants with multiple PI-resistance mutations in the absence of PI therapy. Viruses with multiple PI mutations have (partially) compensated for the initial loss in RC. Therefore, reversion of a single mutation causes a (further) reduction in RC and, as a consequence, the route to wild type is blocked. Copyright © 2007 Infectious Diseases Society of America [ABSTRACT FROM AUTHOR]

BACKGROUND: Airway infection with influenza virus induces local expression of the tryptophan-catabolizing enzyme indoleamine 2,3-dioxygenase (IDO), which has been shown to enhance inflammatory mediator responses in vitro. Because secondary pneumococcal infections occurring shortly after recovery from influenza are associated with enhanced inflammatory responses, we hypothesized that IDO activity contributes to the enhanced response to bacterial challenges in mice previously infected with influenza virus. METHODS: On day 14 after influenza virus infection (with strain A/PR/8/34), C57Bl/6 mice were intranasally inoculated with 1 x 10(4) colony-forming units of S. pneumoniae (serotype 3). Matrix-driven delivery pellets that contained 70 mg of the IDO inhibitor 1-methyl-DL-tryptophan (MeTrp) released over a period of 7 days were subcutaneously implanted 48 h before pneumococcal infection. RESULTS: MeTrp treatment resulted in a 20-fold reduction in pneumococcal outgrowth 48 h after bacterial inoculation. Remarkably, pulmonary levels of interleukin-10 and tumor necrosis factor-alpha were significantly reduced in mice treated with MeTrp. CONCLUSIONS: Our data suggest that IDO expression during influenza virus infection alters the inflammatory response and facilitates the outgrowth of pneumococci during secondary bacterial pneumonia. Copyright © 2006 Infectious Diseases Society of America [ABSTRACT FROM AUTHOR]

Background: The connection domain mutation N348I confers resistance to zidovudine (AZT) and is associated with the lamivudine (3TC) mutation M184V. We explored the biochemical and virological influence of N348I in the context of M184V., Methods: Genotypic resistance data for patients receiving monotherapy or dual therapy with AZT, lamivudine (3TC), or AZT/3TC were analyzed. Rates of N348I emergence were compared between treatment groups. Mutant reverse transcriptases (RTs) containing M184V and/or N348I were generated to study enzymatic and virological properties., Results: We included 50 AZT-treated, 11 3TC-treated, and 10 AZT/3TC-treated patients. N348I was observed in 3 (6%), 0, and 4 (40%) of these patients, respectively. The rate of N348I emergence was increased by 5-fold in the AZT/3TC group (11.7 instances [95% confidence interval {CI}, 3.2-30.1 instances] per 100 person-years of receipt of AZT), compared with the rate noted for the AZT group (2.3 instances [95% CI, 0.4-6.8 instances] per 100 person-years of receipt of AZT; P = .04). Biochemical data show that N348I can partially compensate for the diminution in processive DNA synthesis and the reduction in AZT excision associated with M184V. Furthermore, virological analyses demonstrate that N348I confers low-level resistance to AZT and partly restores the reduced RT activity of the M184V variant., Conclusion: In vivo selection of N348I is driven by AZT and is further facilitated when 3TC is coadministered. Compensatory interactions between N348I and M184V help to explain these findings.

Background: Virological failure of first-line antiretroviral therapy based on lopinavir boosted with ritonavir (lopinavir/r) has rarely been associated with resistance in protease. We identified a new genotypic resistance pathway in 3 patients who experienced failure of first-line lopinavir/r treatment., Methods: Viral protease and the C-term part of Gag were sequenced. The observed mutations were introduced in a reference strain to investigate impact on protease inhibitor susceptibility and replication capacity., Results: A detailed longitudinal analysis demonstrated the selection of the M46I+L76V protease mutations in all 3 patients. The L76V conferred a solitary 3.5-fold increase in one-half the maximal inhibitory concentration to lopinavir but severely hampered viral replication. Addition of M46I, which did not confer any lopinavir resistance on its own, had a dual effect. It partly compensated for the loss in replication capacity and increased the one-half maximal inhibitory concentration to above the lower clinical cutoff (11-fold). Analysis of a large clinical database (>180,000 human immunodeficiency virus [HIV] sequences) demonstrated a significant association (Spearman rho, 0.93) between the increased presence of L76V in clinical samples (0.5% in 2000 to 3.4% in 2006) and lopinavir prescription over time., Conclusions: The HIV protease substitution L76V, in combination with M46I, confers clinically relevant levels of lopinavir resistance and represents a novel resistance pathway to first-line lopinavir/r therapy.

During the past years, human coronaviruses (HCoVs) have been increasingly identified as pathogens associated with more-severe respiratory tract infection (RTI). Diagnostic tests for HCoVs are not frequently used in the routine setting. It is likely that, as a result, the precise role that HCoVs play in RTIs is greatly underestimated. We describe a rapid, sensitive, and highly specific quantitative real-time reverse-transcriptase polymerase chain reaction (RT-PCR) for the detection of HCoV that can easily be implemented in the routine diagnostic setting. HCoV was detected in 28 (11%) of the 261 clinical specimens obtained from patients presenting with symptoms of RTI ranging from common cold to severe pneumonia. Only 1 (0.4%) of the 243 control specimens obtained from patients without symptoms of RTI showed the presence of HCoV. We conclude that HCoVs can be frequently detected in patients presenting with RTI. Real-time RT-PCR provides a tool for large-scale epidemiological studies to further clarify the role that coronavirus infection plays in RTI in humans.

The relationship between clinical changes in stavudine activity and stavudine resistance was investigated in 16 human immunodeficiency virus (HIV)-infected children who received stavudine monotherapy for 18 months. Seven patients responded well to stavudine therapy, 3 experienced transient reductions in virus load, and all others had no detectable virologic response. In both the responders and nonresponders, no changes in stavudine susceptibility or specific baseline/emergent mutations in reverse transcriptase were observed. Only posttherapy HIV isolates from transient responders had elevated IC(50) values for stavudine. In 2 of the 3 transient responders, substitutions at codons 41, 210, and 215 were selected. The significance of these mutations was confirmed in viral competition experiments, site-directed mutagenesis, and in vitro selection. Selection of mutations previously associated with zidovudine resistance can be an important mechanism through which HIV may escape stavudine. The effect of these mutations on phenotypic stavudine susceptibility is relatively small but apparently large enough to be clinically significant.

Use of human immunodeficiency virus (HIV) drug-resistance testing in therapeutic decision making may be aided by understanding the relationship between results of genotypic and drug-susceptibility phenotypic assays. We investigated this relationship by applying 3 different statistical methods-cluster analysis, recursive partitioning, and linear discriminant analysis-to results for 72 patients followed in the Adult AIDS Clinical Trials Group (ACTG) protocol 333. ACTG 333 was a multicenter, randomized trial comparing 2 formulations of saquinavir (SQV) to indinavir (IDV) in patients with extensive hard-gel SQV experience. Data include protease amino acid sequences and 50% inhibitory concentrations for SQV and IDV at baseline. The 3 methods give similar results showing the association of mutations at codons 10, 63, 71, and 90 with in vitro resistance to IDV and SQV. Recursive partitioning is especially useful because it can identify interactions among mutations at different codons and accommodates many types of data as well as missing observations.

Exposure of human immunodeficiency virus to the nucleoside analogue lamivudine (3TC) rapidly selects for resistant variants with a valine at codon 184 (M184V) in the catalytic site of reverse transcriptase. In vitro, 184V demonstrated increased enzyme fidelity and suppressed zidovudine resistance. Clinical trials demonstrated that 3TC-zidovudine combination therapy results in a strong and sustained antiviral response. To investigate the role of 184V on in vivo virus evolution, the effect of zidovudine addition in 3TC-pretreated patients harboring 184V was studied. In vivo, no significant change in fidelity was observed with 184V, shown by generation of the classical pattern of zidovudine mutations. Of interest, in contrast to zidovudine monotherapy, in which just one substitution is sufficient for in vivo development of significant zidovudine resistance, multiple substitutions are required for the same level of zidovudine resistance in strains harboring 184V. This need for multiple substitutions may be one of the mechanisms explaining the sustained antiretroviral response of the 3TC-zidovudine combination.

The effect of the appearance of drug-resistant human immunodeficiency virus type 1 (HIV-1) on viral RNA load was studied in patients treated with the reverse transcriptase inhibitor lamivudine. During the first 12 weeks of treatment, HIV-1 RNA concentrations and amino acid changes in codon 184, causing high-level resistance to lamivudine, were determined in longitudinal serum samples from HIV-1 p24 antigen-positive and -negative patients. A marked decline in the amount of HIV-1 RNA (approximately 95% below baseline) and HIV-1 p24 antigen was observed within 2 weeks, followed by a rise that coincided with the appearance of lamivudine-resistant viruses in serum (isoleucine mutants initially, which were subsequently replaced by valine variants). After 12 weeks, a partial antiviral effect was observed despite the presence of a complete codon 184 mutant virus population in serum. This study shows that the rapid appearance of drug-resistant virus in serum is followed by an increase in viral RNA load.