[Less is more in infectious diseases]. / Weniger ist mehr in der Infektiologie.

BACKGROUND: The increasing development of antimicrobial resistance has been identified as one of the greatest threats to public health and is caused to a relevant extent by falsely indicated antibiotic treatment. OBJECTIVE: The main aim of this article is to identify areas in infectious disease diagnostics and treatment where overuse occurs and to provide recommendations on how to avoid it. MATERIAL AND METHODS: The authors identified current and relevant studies on the topic of medical overuse in infectious diseases via a literature search. In particular, contributions from international "less is more" initiatives were included. The focus was on areas in which a reduction of diagnostic and therapeutic measures leads to an optimization of patient outcomes. RESULTS: In many cases overuse in the context of diagnostics and treatment of infectious diseases not only leads to an unnecessary financial burden on the healthcare system and is not beneficial but can also increase the risk of development of antimicrobial resistance and have adverse consequences for patients. CONCLUSION: Correct indications as well as focused selection and adequate application of antimicrobial agents is crucial to provide the best possible medical care. Diagnostic and antibiotic stewardship measures, which should be implemented in collaboration with infectious disease specialists, can help to identify and reduce areas of overuse and misuse.

[Recurrent vomiting and headaches in a 26-year-old male patient]. / Rezidivierendes Erbrechen und Kopfschmerzen bei einem 26-jährigen Patienten.

This article reports the case of a 26-year-old male patient with recurrent emesis and headache due to central nervous system tuberculosis. The thoracic computed tomography showed bilateral disseminated pulmonary micronodular infiltrates and a cavern connecting to the bronchial system. The cranial magnetic resonance imaging showed multiple supratentorial and infratentorial microabscesses with concomitant meningitis. Mycobacterium tuberculosis was detected in sputum, bronchoalveolar lavage and cerebrospinal fluid. The patient received first-line antituberculous drug treatment, including streptomycin (instead of ethambutol) and adjuvant prednisolone.

High vaccination coverage and infection rate result in a robust SARS-CoV-2-specific immunity in the majority of liver cirrhosis and transplant patients: A single-center cross-sectional study.

BACKGROUND: In the third year of the SARS-CoV-2 pandemic, little is known about the vaccine- and infection-induced immune response in liver transplant recipients (LTR) and liver cirrhosis patients (LCP). OBJECTIVE: This cross-sectional study assessed the vaccination coverage, infection rate, and the resulting humoral and cellular SARS-CoV-2-specific immune responses in a cohort of LTR and LCP at the University Medical Center Hamburg-Eppendorf, Germany between March and May 2023. METHODS: Clinical and laboratory data from 244 consecutive patients (160 LTR and 84 LCP) were collected via chart review and a patient survey. Immune responses were determined via standard spike(S)- and nucleocapsid-protein serology and a spike-specific Interferon-gamma release assay (IGRA). RESULTS: On average, LTR and LCP were vaccinated 3.7 and 3.3 times, respectively and 59.4% of patients received ≥4 vaccinations. Altogether, 68.1% (109/160) of LTR and 70.2% (59/84) of LCP experienced a SARS-CoV-2 infection. Most infections occurred during the Omicron wave in 2022 after an average of 3.0 vaccinations. Overall, the hospitalization rate was low (<6%) in both groups. An average of 4.3 antigen contacts by vaccination and/or infection resulted in a seroconversion rate of 98.4%. However, 17.5% (28/160) of LTR and 8.3% (7/84) of LCP demonstrated only low anti-S titers (<1000 AU/ml), and 24.6% (16/65) of LTR and 20.4% (10/59) of LCP had negative or low IGRA responses. Patients with hybrid immunity (vaccination plus infection) elicited significantly higher anti-S titers compared with uninfected patients with the same number of spike antigen contacts. A total of 22.2% of patients refused additional booster vaccinations. CONCLUSION: By spring 2023, high vaccination coverage and infection rate have resulted in a robust, mostly hybrid, humoral and cellular immune response in most LTR and LCP. However, booster vaccinations with vaccines covering new variants seem advisable, especially in patients with low immune responses and risk factors for severe disease.

HIV evolution: CTL escape mutation and reversion after transmission.

Within-patient HIV evolution reflects the strong selection pressure driving viral escape from cytotoxic T-lymphocyte (CTL) recognition. Whether this intrapatient accumulation of escape mutations translates into HIV evolution at the population level has not been evaluated. We studied over 300 patients drawn from the B- and C-clade epidemics, focusing on human leukocyte antigen (HLA) alleles HLA-B57 and HLA-B5801, which are associated with long-term HIV control and are therefore likely to exert strong selection pressure on the virus. The CTL response dominating acute infection in HLA-B57/5801-positive subjects drove positive selection of an escape mutation that reverted to wild-type after transmission to HLA-B57/5801-negative individuals. A second escape mutation within the epitope, by contrast, was maintained after transmission. These data show that the process of accumulation of escape mutations within HIV is not inevitable. Complex epitope- and residue-specific selection forces, including CTL-mediated positive selection pressure and virus-mediated purifying selection, operate in tandem to shape HIV evolution at the population level.

Infrequent recovery of HIV from but robust exogenous infection of activated CD4(+) T cells in HIV elite controllers.

BACKGROUND. Human immunodeficiency virus (HIV) elite controllers are able to control infection with HIV-1 spontaneously to undetectable levels in the absence of antiretroviral therapy, but the mechanisms leading to this phenotype are poorly understood. Although low frequencies of HIV-infected peripheral CD4(+) T cells have been reported in this group, it remains unclear to what extent these are due to viral attenuation, active immune containment, or intracellular host factors that restrict virus replication. METHODS. We assessed proviral DNA levels, autologous viral growth from and infectability of in vitro activated, CD8(+) T cell-depleted CD4(+) T cells from HIV elite controllers (mean viral load, <50 copies/mL), viremic controllers (mean viral load, <2000 copies/mL), chronic progressors, and individuals receiving highly active antiretroviral therapy. RESULTS. Although we successfully detected autologous virus production in ex vivo activated CD4(+) T cells from all chronic progressors and from most of the viremic controllers, we were able to measure robust autologous viral replication in only 2 of 14 elite controllers subjected to the same protocol. In vitro activated autologous CD4(+) T cells from elite controllers, however, supported infection with both X4 and R5 tropic HIV strains at comparable levels to those in CD4(+) T cells from HIV-uninfected subjects. Proviral DNA levels were the lowest in elite controllers, suggesting that extremely low frequencies of infected cells contribute to difficulty in isolation of virus. CONCLUSIONS. These data indicate that elite control is not due to inability of activated CD4(+) T cells to support HIV infection, but the relative contributions of host and viral factors that account for maintenance of low-level infection remain to be determined.

Substantial differences in specificity of HIV-specific cytotoxic T cells in acute and chronic HIV infection.

Cytotoxic T lymphocytes (CTLs) play a vital part in controlling viral replication during human viral infections. Most studies in human infections have focused on CTL specificities in chronic infection and few data exist regarding the specificity of the initial CTL response induced in acute infection. In this study, HIV-1 infection in persons expressing human histocompatibility leukocyte antigen (HLA)-A*0201 was used as a means of addressing this issue. In chronic infection, the dominant HLA-A*0201-restricted CTL response is directed towards the epitope SLYNTVATL ("SL9") in p17 Gag (residues 77-85). This epitope is targeted by 75% of HLA-A*0201-positive adults, and the magnitude of this A*0201-SL9 response shows a strong negative association with viral load in progressive infection. Despite using the highly sensitive peptide-major histocompatibility complex tetramer and intracellular cytokine assays, responses to the SL9 epitope were not detectable in any of 11 HLA-A*0201-positive subjects with acute HIV-1 infection (P = 2 x 10(-6)), even when assays were repeated using the SL9 peptide variant that was encoded by their autologous virus. In contrast, multiple responses (median 3) to other epitopes were evident in 7 of the 11 A*0201-positive subjects. Longitudinal study of two subjects confirmed that the A*0201-SL9 response emerged later than other CTL responses, and after viral set point had been reached. Together, these data show that the CTL responses that are present and that even may dominate in chronic infection may differ substantially from those that constitute the initial antiviral CTL response. This finding is an important consideration in vaccine design and in the evaluation of vaccine candidates.

Cellular immune responses and viral diversity in individuals treated during acute and early HIV-1 infection.

Immune responses induced during the early stages of chronic viral infections are thought to influence disease outcome. Using HIV as a model, we examined virus-specific cytotoxic T lymphocytes (CTLs), T helper cells, and viral genetic diversity in relation to duration of infection and subsequent response to antiviral therapy. Individuals with acute HIV-1 infection treated before seroconversion had weaker CTL responses directed at fewer epitopes than persons who were treated after seroconversion. However, treatment-induced control of viremia was associated with the development of strong T helper cell responses in both groups. After 1 yr of antiviral treatment initiated in acute or early infection, all epitope-specific CTL responses persisted despite undetectable viral loads. The breadth and magnitude of CTL responses remained significantly less in treated acute infection than in treated chronic infection, but viral diversity was also significantly less with immediate therapy. We conclude that early treatment of acute HIV infection leads to a more narrowly directed CTL response, stronger T helper cell responses, and a less diverse virus population. Given the need for T helper cells to maintain effective CTL responses and the ability of virus diversification to accommodate immune escape, we hypothesize that early therapy of primary infection may be beneficial despite induction of less robust CTL responses. These data also provide rationale for therapeutic immunization aimed at broadening CTL responses in treated primary HIV infection.

Dynamic changes of circulating miRNAs induced by the Ebola virus vaccine VSV-EBOV.

VSV-EBOV is a replication-competent Ebola virus (EBOV) vaccine, which was tested in clinical trials as response to the Ebola virus disease (EVD) outbreak 2013-2016. It is the most advanced EBOV candidate currently in the licensure process. The experimental vaccine was again administered as response to outbreaks in the Democratic Republic of Congo. However, underlying molecular mechanisms that convey protection remain incompletely understood. MicroRNAs (miRNAs) are known key regulators that influence gene expression on a post-transcriptional level. The miRNA-mediated control has emerged as a critical regulatory principle in the immune system, which strongly influences the balance of innate and adaptive immune responses by modulation of signaling pathways critical for differentiation of immune cells. We investigated expression levels of circulating miRNAs (c-miRNAs) in plasma from healthy vaccinees, as they may reflect cellular dynamics following VSV-EBOV immunization and additionally may serve as potential biomarkers for vaccine efficacy. As part of the WHO-led VEBCON consortium, we investigated safety and immunogenicity of VSV-EBOV in a phase I trial. A comprehensive analysis of expression levels on c-miRNAs from plasma samples following VSV-EBOV immunization (day 0, 1, 3 post vaccination) was conducted using RT-qPCR assays. Potential biological relevance was assessed using in silico analyses. Additionally, we correlated dynamics of miRNA expressions with our previously reported data on vaccine-induced antibody and cytokine responses and finally evaluated the prognostic power by generating ROC curves. We identified four promising miRNAs (hsa-miR-146a, hsa-miR-126, hsa-miR-199a, hsa-miR-484), showing a strong association with adaptive immune responses, exhibited favourable prognostic performance and are implicated in immunology-related functions. Our results provide evidence that miRNAs may serve as useful biomarkers for prediction of vaccine-induced immunogenicity. Furthermore, our unique data set provides insight into molecular mechanisms that underlie VSV-EBOV-mediated protective immune responses, which may help to decipher VSV-EBOV immune signature and accelerate strategic vaccine design or personalized approaches.

Tropheryma whipplei in children with diarrhoea in rural Ghana.

Tropheryma whipplei has been hypothesized to be able to cause diarrhoea, but data from young children are scarce. In this hospital-based case-control study 534 stool samples of children aged between 2 months and 15 years from rural Ghana were analysed for the presence of T. whipplei. Overall stool prevalence of T. whipplei was high (27.5%). Although there was no difference in T. whipplei carriage overall between cases and controls, cases aged between 0 and 12 months carried T. whipplei in their stool twice as often as controls without diarrhoea. The results from this study may support the hypothesis that T. whipplei can cause diarrhoea in first-time infection.

Regulatory and accessory HIV-1 proteins: potential targets for HIV-1 vaccines?

The HIV-1 regulatory proteins Tat and Rev and the accessory proteins Vpr, Vpu and Vif are essential for efficient viral replication, and their cytoplasmic production suggests that they should be processed for recognition by cytotoxic T lymphocytes. However, only limited data is available, evaluating the role of immune responses directed against these proteins in natural HIV-1 infection. Recent advances in the methods used for the characterization of HIV-1-specific cellular immune responses, including quantification of antigen-specific IFN-gamma production by ELISpot assay and flow-cytometry-based intracellular cytokine quantification, have allowed for a much more comprehensive assessment of virus-specific immune responses. Emerging data show that the regulatory and accessory proteins serve as important targets for HIV-1-specific T cell responses, and multiple CTL epitopes have been identified in functionally important regions of these proteins. Moreover, the use of autologous peptides have allowed for the detection of significantly stronger HIV-1-specific T cell responses in the more variable regulatory and accessory HIV-1 proteins Tat and Vpr. These data indicate that despite the small size of these proteins, regulatory and accessory proteins are targeted by cellular immune responses in natural HIV-1 infection and contribute importantly to the total HIV-1-specific CD8+ T cell response. A multi-component vaccine, with the inclusion of these proteins plus structural proteins remains the most promising choice for an effective AIDS vaccine.

Cytotoxic T-lymphocyte (CTL) responses directed against regulatory and accessory proteins in HIV-1 infection.

The HIV-1 regulatory proteins Tat and Rev and the accessory proteins Vpr, Vpu, and Vif are essential for viral replication, and their cytoplasmic production suggests that they should be processed for recognition by cytotoxic T lymphocytes. However, only limited data is available evaluating to which extent these proteins are targeted in natural infection and optimal cytotoxic T lymphocyte (CTL) epitopes within these proteins have not been defined. In this study, CTL responses against HIV-1 Tat, Rev, Vpr, Vpu, and Vif were analyzed in 70 HIV-1 infected individuals and 10 HIV-1 negative controls using overlapping peptides spanning the entire proteins. Peptide-specific interferon-gamma (IFN-gamma) production was measured by Elispot assay and flow-based intracellular cytokine quantification. HLA class I restriction and cytotoxic activity were confirmed after isolation of peptide-specific CD8+ T-cell lines. All regulatory and accessory proteins served as targets for HIV-1- specific CTL and multiple CTL epitopes were identified in functionally important regions of these proteins. In certain individuals HIV-1-specific CD8+ T-cell responses to these accessory and regulatory proteins contributed up to a third to the magnitude of the total HIV-1-specific CTL response. These data indicate that despite the small size of these proteins regulatory and accessory proteins are targeted by CTL in natural HIV-1 infection, and contribute importantly to the total HIV-1-specific CD8+ T-cell responses. These findings are relevant for the evaluation of the specificity and breadth of immune responses during acute and chronic#10; infection, and will be useful for the design and testing of candidate human immunodeficiency virus (HIV) vaccines.

Cellular immune responses in transplantation-associated chronic viral infections.

Viral pathogens are important causes of morbidity following transplantation. Cytomegalovirus (CMV) and Epstein-Barr virus (EBV) infections represent two major viral complications in transplant recipients. Recent advances in methodology have led to a better understanding of host immune responses directed against chronic viral infections. We review the nature of antiviral immunity involved in control of CMV and EBV. Viral mechanisms of immune evasion and immunotherapeutic strategies in the transplantation setting will also be addressed.

T(H)1 to T(H)2 shift of cytokines in peripheral blood of HIV-infected patients is detectable by reverse transcriptase polymerase chain reaction but not by enzyme-linked immunosorbent assay under nonstimulated conditions.

BACKGROUND: Dysregulation of cytokines has been implicated in the pathogenesis of HIV infection. Therefore, we determined tumor necrosis factor-alpha (TNF-alpha), interleukin-1beta (IL-1beta), IL-4, IL-10, and interferon-gamma (IFN-gamma) mRNA and serum levels in HIV-infected patients under nonstimulated conditions. MATERIAL AND METHODS: Blood samples of 32 HIV-infected patients and 10 healthy HIV-negative controls were analyzed. Cytokine serum levels were quantified by enzyme-linked immunosorbent assay (ELISA). Cytokine mRNA levels were determined semiquantitatively by competitive reverse transcriptase polymerase chain reaction (RT-PCR) and expressed as ratios relative to those of beta-actin. RESULTS: Competitive RT-PCR was shown to be more sensitive than protein ELISA in analyzing cytokine production. We found a significant correlation between steady-state mRNA ratios and serum protein levels for TNF-alpha. Significantly higher cytokine mRNA ratios were found in those patients with IL-10 and IFN-gamma levels detectable by ELISA. Steady-state mRNA ratios of TNF-alpha, IL-4, and IL-10 were significantly increased in patients with highly replicative HIV-infection. Furthermore, elevated IL-4:IFN-gamma ratios were related to both high viral load and loss of CD4 cells. DISCUSSION: Determination of steady-state mRNA ratios by semiquantitative RT-PCR represents a sensitive method to analyze cytokines in peripheral blood of HIV-infected patients under nonstimulated conditions. The data obtained with this technique provide further evidence for a T(H)1 to T(H)2 cytokine shift with progressive HIV disease.

Rapid definition of five novel HLA-A*3002-restricted human immunodeficiency virus-specific cytotoxic T-lymphocyte epitopes by elispot and intracellular cytokine staining assays.

Human immunodeficiency virus (HIV)-specific cytotoxic T lymphocytes (CTL) play a major role in control of viral replication. To understand the contribution of this antiviral response, an initial step is to fully define the specific epitopes targeted by CTL. These studies focused on CTL responses restricted by HLA-A*3002, one of the HLA-A molecules most prominent in African populations. To avoid the time-consuming effort and expense involved in culturing CTL prior to defining epitopes and restricting alleles, we developed a method combining Elispot assays with intracellular gamma interferon staining of peripheral blood mononuclear cells to first map the optimal epitopes targeted and then define the HLA restriction of novel epitopes. In two A*3002-positive subjects whose CTL responses were characterized in detail, the strongest response in both cases was to an epitope in p17 Gag, RSLYNTVATLY (residues 76 to 86). Using this method, CTL epitopes for which there were no motif predictions were optimized and the HLA restriction was established within 48 to 72 h of receipt of blood. This simple and convenient approach should prove useful especially in the characterization of CTL responses specific to HIV and other viruses, particularly in localities where performing cytotoxicity assays would be problematic.

The HIV-1 regulatory proteins Tat and Rev are frequently targeted by cytotoxic T lymphocytes derived from HIV-1-infected individuals.

The HIV-1 regulatory proteins Rev and Tat are expressed early in the virus life cycle and thus may be important targets for the immune control of HIV-1-infection and for effective vaccines. However, the extent to which these proteins are targeted in natural HIV-1 infection as well as precise epitopes targeted by human cytotoxic T lymphocytes (CTL) remain to be defined. In the present study, 57 HIV-1-infected individuals were screened for responses against Tat and Rev by using overlapping peptides spanning the entire Tat and Rev proteins. CD8+ T cell responses against Tat and Rev were found in up to 19 and 37% of HIV-1-infected individuals, respectively, indicating that these regulatory proteins are important targets for HIV-1-specific CTL. Despite the small size of these proteins, multiple CTL epitopes were identified in each. These data indicate that Tat and Rev are frequently targeted by CTL in natural HIV-1 infection and may be important targets for HIV vaccines.

Identification of dominant optimal HLA-B60- and HLA-B61-restricted cytotoxic T-lymphocyte (CTL) epitopes: rapid characterization of CTL responses by enzyme-linked immunospot assay.

Human immunodeficiency virus type 1 (HIV-1)-specific cytotoxic T-lymphocyte (CTL) responses play a major role in the antiviral immune response, but the relative contribution of CTL responses restricted by different HLA class I molecules is less well defined. HLA-B60 or the related allele B61 is expressed in 10 to 20% of Caucasoid populations and is even more highly prevalent in Asian populations, but yet no CTL epitopes restricted by these alleles have been defined. Here we report the definition of five novel HLA-B60-restricted HIV-1-specific CTL epitopes, using peripheral blood mononuclear cells in enzyme-linked immunospot (Elispot) assays and using CTL clones and lines in cytolytic assays. The dominant HLA-B60-restricted epitope, Nef peptide KEKGGLEGL, was targeted by all eight subjects with B60 and also by both subjects with B61 studied. This study additionally establishes the utility of the Elispot assay as a more rapid and efficient method of defining novel CTL epitopes. This approach will help to define new CTL epitopes that may play an important role in the immune control of HIV-1.

Persistent recognition of autologous virus by high-avidity CD8 T cells in chronic, progressive human immunodeficiency virus type 1 infection.

CD8 T-cell responses are thought to be crucial for control of viremia in human immunodeficiency virus (HIV) infection but ultimately fail to control viremia in most infected persons. Studies in acute infection have demonstrated strong CD8-mediated selection pressure and evolution of mutations conferring escape from recognition, but the ability of CD8 T-cell responses that persist in late-stage infection to recognize viruses present in vivo has not been determined. Therefore, we studied 24 subjects with advanced HIV disease (median viral load = 142,000 copies/ml; median CD4 count = 71/ micro l) and determined HIV-1-specific CD8 T-cell responses to all expressed viral proteins using overlapping peptides by gamma interferon Elispot assay. Chronic-stage virus was sequenced to evaluate autologous sequences within Gag epitopes, and functional avidity of detected responses was determined. In these subjects, the median number of epitopic regions targeted was 13 (range, 2 to 39) and the median cumulative magnitude of CD8 T-cell responses was 5,760 spot-forming cells/10(6) peripheral blood mononuclear cells (range, 185 to 24,700). On average six (range, one to 8) proteins were targeted. For 89% of evaluated CD8 T-cell responses, the autologous viral sequence was predicted to be well recognized by these responses and the majority of analyzed optimal epitopes were recognized with medium to high functional avidity by the contemporary CD8 T cells. Withdrawal of antigen by highly active antiretroviral therapy led to a significant decline both in breadth (P = 0.032) and magnitude (P = 0.0098) of these CD8 T-cell responses, providing further evidence that these responses had been driven by recognition of autologous virus. These results indicate that strong, broadly directed, and high-avidity gamma-interferon-positive CD8 T-cells directed at autologous virus persist in late disease stages, and the absence of mutations within viral epitopes indicates a lack of strong selection pressure mediated by these responses. These data imply functional impairment of CD8 T-cell responses in late-stage infection that may not be reflected by gamma interferon-based screening techniques.

Comprehensive epitope analysis of human immunodeficiency virus type 1 (HIV-1)-specific T-cell responses directed against the entire expressed HIV-1 genome demonstrate broadly directed responses, but no correlation to viral load.

Cellular immune responses play a critical role in the control of human immunodeficiency virus type 1 (HIV-1); however, the breadth of these responses at the single-epitope level has not been comprehensively assessed. We therefore screened peripheral blood mononuclear cells (PBMC) from 57 individuals at different stages of HIV-1 infection for virus-specific T-cell responses using a matrix of 504 overlapping peptides spanning all expressed HIV-1 proteins in a gamma interferon-enzyme-linked immunospot (Elispot) assay. HIV-1-specific T-cell responses were detectable in all study subjects, with a median of 14 individual epitopic regions targeted per person (range, 2 to 42), and all 14 HIV-1 protein subunits were recognized. HIV-1 p24-Gag and Nef contained the highest epitope density and were also the most frequently recognized HIV-1 proteins. The total magnitude of the HIV-1-specific response ranged from 280 to 25,860 spot-forming cells (SFC)/10(6) PBMC (median, 4,245) among all study participants. However, the number of epitopic regions targeted, the protein subunits recognized, and the total magnitude of HIV-1-specific responses varied significantly among the tested individuals, with the strongest and broadest responses detectable in individuals with untreated chronic HIV-1 infection. Neither the breadth nor the magnitude of the total HIV-1-specific CD8+-T-cell responses correlated with plasma viral load. We conclude that a peptide matrix-based Elispot assay allows for rapid, sensitive, specific, and efficient assessment of cellular immune responses directed against the entire expressed HIV-1 genome. These data also suggest that the impact of T-cell responses on control of viral replication cannot be explained by the mere quantification of the magnitude and breadth of the CD8+-T-cell response, even if a comprehensive pan-genome screening approach is applied.

Identification of novel HLA-A2-restricted human immunodeficiency virus type 1-specific cytotoxic T-lymphocyte epitopes predicted by the HLA-A2 supertype peptide-binding motif.

Virus-specific cytotoxic T-lymphocyte (CTL) responses are critical in the control of human immunodeficiency virus type 1 (HIV-1) infection and will play an important part in therapeutic and prophylactic HIV-1 vaccines. The identification of virus-specific epitopes that are efficiently recognized by CTL is the first step in the development of future vaccines. Here we describe the immunological characterization of a number of novel HIV-1-specific, HLA-A2-restricted CTL epitopes that share a high degree of conservation within HIV-1 and a strong binding to different alleles of the HLA-A2 superfamily. These novel epitopes include the first reported CTL epitope in the Vpr protein. Two of the novel epitopes were immunodominant among the HLA-A2-restricted CTL responses of individuals with acute and chronic HIV-1 infection. The novel CTL epitopes identified here should be included in future vaccines designed to induce HIV-1-specific CTL responses restricted by the HLA-A2 superfamily and will be important to assess in immunogenicity studies in infected persons and in uninfected recipients of candidate HIV-1 vaccines.

Evolution and transmission of stable CTL escape mutations in HIV infection.

Increasing evidence indicates that potent anti-HIV-1 activity is mediated by cytotoxic T lymphocytes (CTLs); however, the effects of this immune pressure on viral transmission and evolution have not been determined. Here we investigate mother-child transmission in the setting of human leukocyte antigen (HLA)-B27 expression, selected for analysis because it is associated with prolonged immune containment in adult infection. In adults, mutations in a dominant and highly conserved B27-restricted Gag CTL epitope lead to loss of recognition and disease progression. In mothers expressing HLA-B27 who transmit HIV-1 perinatally, we document transmission of viruses encoding CTL escape variants in this dominant Gag epitope that no longer bind to B27. Their infected infants target an otherwise subdominant B27-restricted epitope and fail to contain HIV replication. These CTL escape variants remain stable without reversion in the absence of the evolutionary pressure that originally selected the mutation. These data suggest that CTL escape mutations in epitopes associated with suppression of viraemia will accumulate as the epidemic progresses, and therefore have important implications for vaccine design.