Post-transplant dynamics and clinical significance of CMV-specific neutralizing antibodies in kidney transplant recipients treated with T-cell-depleting agents.

We measured cytomegalovirus (CMV)-specific antibodies that neutralize epithelial cell infection (CMV-AbNEIs) in 101 CMV-seropositive kidney transplant recipients (KTRs) at baseline and post-transplant months 3 and 6. All the patients received antithymocyte globulin and 3-month valganciclovir prophylaxis. There were no significant differences in pre-transplant AbNEIs titers between KTRs that developed or did not develop any-level CMV infection or the composite of high-level infection and/or disease. One-year CMV infection-free survival was comparable between KTRs with or without pre-transplant CMV-AbNEIs. No differences were observed by months 3 and 6 either. We observed no protective role for CMV-AbNEIs among CMV-seropositive KTRs undergoing T-cell-depleting induction.

Circulatory follicular helper T lymphocytes associate with lower incidence of CMV infection in kidney transplant recipients.

Primary infection and/or reactivation of cytomegalovirus (CMV) in kidney transplant recipients (KTR) favor rejection and mortality. T follicular helper cells (TFH) could contribute to protection against CMV. Circulatory TFH (cTFH) were studied pretransplant and early posttransplant in 90 CMV seropositive KTR not receiving antithymocyte globulin or antiviral prophylaxis, followed-up for 1 year. Patients who presented CMV infection had significantly lower cTFH and activated cTFH pretransplant and early posttransplant. Pretransplant activated cTFH were also lower within patients who developed CMV disease. Pre- and 14 days posttransplant activated cTFH were an independent protective factor for CMV infection (HR 0.41, p = .01; and 0.52, p = .02, respectively). KTR with low cTFH 7 days posttransplant (<11.9%) had lower CMV infection-free survival than patients with high cTFH (28.2% vs. 67.6%, p = .002). cTFH were associated with CMV-specific neutralizing antibodies (Nabs). In addition, IL-21 increased interferon-γ secretion by CMV-specific CD8+ T cells in healthy controls. Thus, we show an association between cTFH and lower incidence of CMV infection, probably through their cooperation in CMV-specific Nab production and IL-21-mediated enhancement of CD8+ T cell activity. Moreover, monitoring cTFH pre- and early posttransplant could improve CMV risk stratification and help select KTR catalogued at low/intermediate risk who could benefit from prophylaxis.

Going beyond serology for stratifying the risk of CMV infection in transplant recipients.

Knowledge of donor and recipient (D/R) cytomegalovirus (CMV) serostatus is critical for risk stratification of CMV infection and disease in transplant recipients, particularly in the solid organ transplantation (SOT) setting. Despite its broad availability and the success of it use, the risk stratification based on the D/R serostatus is not free of limitations since there are a nondepreciable number of patients that are not accurately categorized by this approach. In fact, up to 20% of seropositive SOT recipients, classically considered at intermediate risk, develop episodes of CMV infection and disease after transplantation. Here, we provide an overview of additional donor and recipient factors that may have utility in identifying patients at risk for post-transplant CMV infection. Specifically, we summarize our current understanding regarding the potential use of use CMV-specific T-cell-mediated immunity, neutralizing antibodies and host genetics that may influence the risk of CMV infection and disease. We provide an overview of the benefits and limitations associated with using these immunological factors in risk stratification and propose specific variables that could be analyzed at the pretransplant evaluation to improve the identification of patients with increased individual susceptibility.

COVID-19 vaccine effectiveness against hospitalization due to SARS-CoV-2: A test-negative design study based on Severe Acute Respiratory Infection (SARI) sentinel surveillance in Spain.

BACKGROUND: With the emergence of SARS-CoV-2, influenza surveillance systems in Spain were transformed into a new syndromic sentinel surveillance system. The Acute Respiratory Infection Surveillance System (SiVIRA in Spanish) is based on a sentinel network for acute respiratory infection (ARI) surveillance in primary care and a network of sentinel hospitals for severe ARI (SARI) surveillance in hospitals. METHODS: Using a test-negative design and data from SARI admissions notified to SiVIRA between January 1 and October 3, 2021, we estimated COVID-19 vaccine effectiveness (VE) against hospitalization, by age group, vaccine type, time since vaccination, and SARS-CoV-2 variant. RESULTS: VE was 89% (95% CI: 83-93) against COVID-19 hospitalization overall in persons aged 20 years and older. VE was higher for mRNA vaccines, and lower for those aged 80 years and older, with a decrease in protection beyond 3 months of completing vaccination, and a further decrease after 5 months. We found no differences between periods with circulation of Alpha or Delta SARS-CoV-2 variants, although variant-specific VE was slightly higher against Alpha. CONCLUSIONS: The SiVIRA sentinel hospital surveillance network in Spain was able to describe clinical and epidemiological characteristics of SARI hospitalizations and provide estimates of COVID-19 VE in the population under surveillance. Our estimates add to evidence of high effectiveness of mRNA vaccines against severe COVID-19 and waning of protection with time since vaccination in those aged 80 or older. No substantial differences were observed between SARS-CoV-2 variants (Alpha vs. Delta).

Role of Neutralizing Antibodies in CMV Infection: Implications for New Therapeutic Approaches.

Cytomegalovirus (CMV) infection elicits a potent immune response that includes the stimulation of antibodies with neutralizing activity. Recent studies have focused on elucidating the role of neutralizing antibodies in protecting against CMV infection and disease and characterizing viral antigens against which neutralizing antibodies are directed. Here, we provide a synthesis of recent data regarding the role of neutralizing antibodies in protection against CMV infection/disease. We consider the role of humoral immunity in the context of the global CMV-specific immune response, and the implications that recent findings have for vaccine and antibody-based therapy design.

Cytotoxic cell populations developed during treatment with tyrosine kinase inhibitors protect autologous CD4+ T cells from HIV-1 infection.

Tyrosine kinase inhibitors (TKIs) are successfully used in clinic to treat chronic myeloid leukemia (CML). Our group previously described that CD4+ T cells from patients with CML on treatment with TKIs such as dasatinib were resistant to HIV-1 infection ex vivo. The main mechanism for this antiviral activity was primarily based on the inhibition of SAMHD1 phosphorylation, which preserves the activity against HIV-1 of this innate immune factor. Approximately 50% CML patients who achieved a deep molecular response (DMR) may safely withdraw TKI treatment without molecular recurrence. Therefore, it has been speculated that TKIs may induce a potent antileukemic response that is maintained in most patients even one year after treatment interruption (TI). Subsequent to in vitro T-cell activation, we observed that SAMHD1 was phosphorylated in CD4+ T cells from CML patients who withdrew TKI treatment more than one year earlier, which indicated that these cells were now susceptible to HIV-1 infection. Importantly, these patients were seronegative for HIV-1 and seropositive for cytomegalovirus (CMV), but without CMV viremia. Although activated CD4+ T cells from CML patients on TI were apparently permissive to HIV-1 infection ex vivo, the frequency of proviral integration was reduced more than 12-fold on average when these cells were infected ex vivo in comparison with cells isolated from untreated, healthy donors. This reduced susceptibility to infection could be related to an enhanced NK-dependent cytotoxic activity, which was increased 8-fold on average when CD4+ T cells were infected ex vivo with HIV-1 in the presence of autologous NK cells. Enhanced cytotoxic activity was also observed in CD8 + T cells from these patients, which showed 8-fold increased expression of TCRγδ and more than 18-fold increased production of IFNγ upon activation with CMV peptides. In conclusion, treatment with TKIs induced a potent antileukemic response that may also have antiviral effects against HIV-1 and CMV, suggesting that transient use of TKIs in HIV-infected patients could develop a sustained antiviral response that would potentially interfere with HIV-1 reservoir dynamics.

First cases of European bat lyssavirus type 1 in Iberian serotine bats: Implications for the molecular epidemiology of bat rabies in Europe.

Previous studies have shown that EBLV-1 strains exclusively hosted by Eptesicus isabellinus bats in the Iberian Peninsula cluster in a specific monophyletic group that is related to the EBLV-1b lineage found in the rest of Europe. More recently, enhanced passive surveillance has allowed the detection of the first EBLV-1 strains associated to Eptesicus serotinus south of the Pyrenees. The aim of this study is the reconstruction of the EBLV-1 phylogeny and phylodynamics in the Iberian Peninsula in the context of the European continent. We have sequenced 23 EBLV-1 strains detected on nine E. serotinus and 14 E. isabellinus. Phylogenetic analyses were performed on the first 400-bp-5' fragment of the Nucleoprotein (N) gene together with other 162 sequences from Europe. Besides, fragments of the variable region of the phosphoprotein (P) gene and the glycoprotein-polymerase (G-L) intergenic region were studied on Spanish samples. Phylogenies show that two of the new EBLV-1a strains from Iberian E. serotinus clustered together with French strains from the North of the Pyrenees, suggesting a recent expansion southwards of this subtype. The remaining seven Iberian strains from E. serotinus grouped, instead, within the cluster linked, so far, to E. isabellinus, indicating that spatial distribution prevails over species specificity in explaining rabies distribution and supporting interspecific transmission. The structure found within the Iberian Peninsula for EBLV-1b is in concordance with that described previously for E. isabellinus. Finally, we have found that the current EBLV-1 European strains could have emerged only 175 years ago according to our evolutionary dynamics analyses.

Plasma viral load threshold for sustaining intrahost HIV type 1 evolution.

The objective of the present study was to determine if natural suppression of plasma viremia below the detection limit of commercial assays (50-80 copies HIV-1 RNA/ml) can contain the HIV-1 evolution. HIV-1 quasispecies complexity in PBMC DNA was assessed in the env gene at two time points in 14 long-term nonprogressors (LTNPs). Sequence changes consistent with viral evolution was found in all patients with a median plasma RNA viral load >100 copies/ml. Evidence of low-level viral evolution was detected in two of four patients with intermittent viremia and a median plasma HIV-1 RNA load of >80 copies/ml. No significant evolution was observed in the three LTNPs with persistent viral suppression below the detection limit. Overall, a significant positive correlation (p < 0.001) was observed between viral evolution and plasma RNA viral load in the LTNPs analyzed. These results suggest that the detection limit of ultrasensitive viremia assays could represent an important threshold below which intrahost HIV-1 evolution does not occur.

Identification of a cluster of HIV-1 controllers infected with low replicating viruses.

Long term non-progressor patients (LTNPs) are characterized by the natural control of HIV-1 infection. This control is related to host genetic, immunological and virological factors. In this work, phylogenetic analysis of the proviral nucleotide sequences in env gene from a Spanish HIV-1 LTNPs cohort identified a cluster of 6 HIV-1 controllers infected with closely-related viruses. The patients of the cluster showed common clinical and epidemiological features: drug user practices, infection in the same city (Madrid, Spain) and at the same time (late 70's-early 80's). All cluster patients displayed distinct host alleles associated with HIV control. Analysis of the virus envelope nucleotide sequences showed ancestral characteristic, lack of evolution and presence of rare amino-acids. Biological characterization of recombinant viruses with the envelope proteins from the cluster viruses showed very low replicative capacity in TZMbl and U87-CD4/CCR5 cells. The lack of clinical progression in the viral cluster patients with distinct combinations of protective host genotypes, but infected by low replicating viruses, indicate the important role of the virus in the non-progressor phenotype in these patients.

Prevalence of HIV-1 dual infection in long-term nonprogressor-elite controllers.

INTRODUCTION: Human immunodeficiency virus type 1 (HIV-1) dual infection (DI) in long-term nonprogressor-elite controller patients (LTNP-EC) has been described only in sporadic cases and then, consequences in disease progression are not clearly established. To fill-up this limited knowledge, we analyzed, for the first time, the prevalence, host genetic polymorphisms, and clinical consequences of HIV-1 DI in a group of LTNP-EC. METHODS: For DI detection, nucleotide sequences in env gene from viruses from 20 LTNP-EC were analyzed by maximum likelihood. Epidemiological and clinical parameters and host factors of patients were also studied. RESULTS: DI was detected in 4 (20%) of the 20 LTNP-EC, of which 3 maintained the elite controller status. CD4⁺ T-cell counts were not different between single and DI patients although higher CD8⁺ T-cell counts were observed in DI patients, and, consequently, the CD4⁺/CD8⁺ ratios were lower in LTNP-EC DI patients. CONCLUSIONS: Prevalence of HIV-1 DIs in LTNP-EC is similar to other groups of HIV-1 patients; in addition, DI was not associated with loss of disease control in the patients. These DI LTNP-EC patients showed, in comparison with single infected patients, higher numbers of CD8⁺ T cells and lower CD4⁺/CD8⁺ ratios.

Comparison of real-time RT-PCR, shell vial culture, and conventional cell culture for the detection of the pandemic influenza A (H1N1) in hospitalized patients.

The emergence of the pandemic influenza virus A H1N1 has made fast and accurate diagnosis essential. However, few well-validated diagnostic techniques exist. The real-time RT-PCR developed by the Centers for Disease Control and Prevention (CDC) is the recommended technique. Our objective was to compare the CDC real-time RT-PCR assay, shell vial (SV), and conventional cell culture [with Madine-Darby canine kidney (MDCK) cells and A549] for the detection of pandemic influenza A H1N1 in hospitalized patients. We performed a prospective study comparing the efficacy of 5 diagnostic techniques (RTPCR, SV in A549, SV in MDCK, conventional cell culture in A549, and conventional cell culture in MDCK) using nasopharyngeal swabs from patients ≥18 years of age hospitalized with clinical symptoms of influenza at our institution. Detection of the virus by conventional culture was considered the gold standard. An "extended gold standard" was also used to recalculate validity values. The sensitivities, specificities, positive predictive values, and negative predictive values (NPVs) for the detection of influenza A H1N1, determined using conventional culture as the gold standard, were, respectively, as follows: RT-PCR: 95.6, 82.3, 78.3, 96.5%; SVA549: 91.2, 99.01, 98.4, 94.4%; SV-MDCK: 82.3, 100, 100, 89.4%; tube-A549: 94.12, 100, 100, 96.2%; tube-MDCK: 86.7, 100, 100, 91.9%. Sensitivities and NPVs using an extended gold standard were as follows: RT-PCR: 96.5%, 96.6%; SV-A549: 73.3%, 78.5%; SV-MDCK: 65.1%, 73.7%; tube-A549: 74.4%, 79.2%; tube-MDCK: 68.6%, 75.7%. The average time to detect pandemic influenza A H1N1 by RT-PCR, SV culture, and conventional culture was, respectively, 4 h, 48 h, and 7 days. Real-time RT-PCR displayed high sensitivity and specificity for the detection of influenza A H1N1 in adult patients when compared with conventional techniques. In addition, the A549 cell line was not inferior to the MDCK line.

A combination of defective DNA and protective host factors are found in a set of HIV-1 ancestral LTNPs.

We studied viral evolution in three HIV-1 ancestral patients from a group of LTNPs; although some minor sequences showing viral evolution were detected in all patients, the extremely low viral evolution of their viruses was shown by the phylogenetic analysis of the env sequences. Complete nucleotide sequencing of viral DNA showed the major presence of deletions. In two patients, deletions of 1088 and 228 nucleotides mapped to 5' LTR-gag region; in the other, a 247 nucleotide deletion was positioned in pol gene up to the vif ORF. These deleted genomes became dominant during follow up. Patient's viruses displayed 13 common mutations in conserved residues, from the 5' LTR to the nef gene. These mutations provided evidence of a common origin. Regarding host characteristics, one patient had HLA B2705/B5801; another B1402/B5701; whereas a third showed B3901/B4402 and was Delta32-CCR5 heterozygous. These HIV controllers presented a combination of deleted viral genomes and host protective factors.

Coinfection and superinfection in patients with long-term, nonprogressive HIV-1 disease.

Human immunodeficiency virus 1 (HIV-1) dual infections are considered important because they have been related to AIDS progression. We identified dual infections in 2 patients with long-term, nonprogressive HIV-1 disease; the first patient was diagnosed as being already coinfected, on the basis of the first sample analyzed, but a previous superinfection could not be excluded; the second patient was diagnosed as having a superinfection, on the basis of the 9-year difference between the viral dating of the 2 strains. Dual infections occur in patients with long-term, nonprogressive disease, with no immediate clinical manifestations. Such occurrences could indicate a general phenomenon in natural HIV-1 infections.

A subset of human immunodeficiency virus type 1 long-term non-progressors is characterized by the unique presence of ancestral sequences in the viral population.

Within human immunodeficiency virus type 1 (HIV-1)-infected patients, there are those who have been infected for more than 10 years with a CD4+ cell count of >500 cells microl(-1) and who remain asymptomatic without antiretroviral therapy; these patients are designated long-term non-progressors (LTNPs). In a set of 16 LTNPs, viral dating, DNA viral load, quasispecies heterogeneity and antibody (Ab) titres against gp160 and beta2 microglobulin (beta2m) were determined. Plasma viral RNA and CD4+ and CD8+ T-cell numbers were estimated in more than three samples per patient. Host genetic characteristics, such as Delta32-CCR5 genotype and human leukocyte antigen (HLA) genotype and supertypes, and clinical-epidemiological factors were evaluated. Dating of global populations and of DNA and RNA viral quasispecies identified two subsets of patients: one displaying only ancestral sequences and the other displaying predominantly modern sequences. The ancestral patients displayed a significant reduction in RNA and DNA viral loads, quasispecies heterogeneity, CD8+ cell number, anti-gp160 Ab titres and beta2m level, and they were also associated with better use of safe-sex practices and higher presence of the HLA sB58 supertype than the modern subset. Viral dating has therefore permitted the segregation of LTNPs into two subsets that show very different virological, immunological, host and clinical-epidemiological characteristics. Moreover, whereas the modern subset displayed low levels of virus replication, the ancestral group displayed not only a very limited virus replication, often to undetectable levels, but also very slow or arrested viral evolution, maintaining the close relationship of the viral population to the transmitted virus.