Francisca Valdivieso

Highly Differentiated, Resting Gn-Specific Memory CD8 + T Cells Persist Years after Infection by Andes Hantavirus

In man, infection with South American Andes virus (ANDV) causes hantavirus cardiopulmonary syndrome (HCPS). HCPS due to ANDV is endemic in Southern Chile and much of Argentina and increasing numbers of cases are reported all over South America. A case-fatality rate of about 36% together with the absence of successful antiviral therapies urge the development of a vaccine. Although T-cell responses were shown to be critically involved in immunity to hantaviruses in mouse models, no data are available on the magnitude, specificity and longevity of ANDV-specific memory T-cell responses in patients. Using sets of overlapping peptides in IFN-γ ELISPOT assays, we herein show in 78 Chilean convalescent patients that Gn-derived epitopes were immunodominant as compared to those from the N- and Gc-proteins. Furthermore, while the relative contribution of the N-specific response significantly declined over time, Gn-specific responses remained readily detectable ex vivo up to 13 years after the acute infection. Tetramer analysis further showed that up to 16.8% of all circulating CD3+CD8+ T cells were specific for the single HLA-B*3501-restricted epitope Gn465–473 years after the acute infection. Remarkably, Gn465–473–specific cells readily secreted IFN-γ, granzyme B and TNF-α but not IL-2 upon stimulation and showed a ‘revertant’ CD45RA+CD27−CD28−CCR7−CD127− effector memory phenotype, thereby resembling a phenotype seen in other latent virus infections. Most intriguingly, titers of neutralizing antibodies increased over time in 10/17 individuals months to years after the acute infection and independently of whether they were residents of endemic areas or not. Thus, our data suggest intrinsic, latent antigenic stimulation of Gn-specific T-cells. However, it remains a major task for future studies to proof this hypothesis by determination of viral antigen in convalescent patients. Furthermore, it remains to be seen whether Gn-specific T cells are critical for viral control and protective immunity. If so, Gn-derived immunodominant epitopes could be of high value for future ANDV vaccines.

Neutralizing Antibodies in Survivors of Sin Nombre and Andes Hantavirus Infection

We evaluated titers of homotypic and heterotypic neutralizing antibodies (NAbs) to Andes and Sin Nombre hantaviruses in plasma samples from 20 patients from Chile and the United States. All but 1 patient had high titers of NAb. None of the plasma samples showed high titers against the heterologous virus.

Person-to-person household and nosocomial transmission of andes hantavirus, Southern Chile, 2011.

Four persons became ill after exposure to a patient infected with the virus; 2 cases involved hospital transmission.

High-Dose Intravenous Methylprednisolone for Hantavirus Cardiopulmonary Syndrome in Chile: A Double-Blind, Randomized Controlled Clinical Trial

BACKGROUND Andes virus (ANDV)-related hantavirus cardiopulmonary syndrome (HCPS) has a 35% case fatality rate in Chile and no specific treatment. In an immunomodulatory approach, we evaluated the efficacy of intravenous methylprednisolone for HCPS treatment, through a parallel-group, placebo-controlled clinical trial. METHODS Patients aged >2 years, with confirmed or suspected HCPS in cardiopulmonary stage, admitted to any of 13 study sites in Chile, were randomized by study center in blocks of 4 with a 1:1 allocation and assigned through sequentially numbered envelopes to receive placebo or methylprednisolone 16 mg/kg/day (≤1000 mg) for 3 days. All personnel remained blinded except the local pharmacist. Infection was confirmed by immunoglobulin M antibodies or ANDV RNA in blood. The composite primary endpoint was death, partial pressure of arterial oxygen/fraction of inspired oxygen ratio ≤55, cardiac index ≤2.2, or ventricular tachycardia or fibrillation within 28 days. Safety endpoints included the number of serious adverse events (SAEs) and quantification of viral RNA in blood. Analysis was by intention to treat. RESULTS Infection was confirmed in 60 of 66 (91%) enrollees. Fifteen of 30 placebo-treated patients and 11 of 30 methylprednisolone-treated patients progressed to the primary endpoint (P = .43). We observed no significant difference in mortality between treatment groups (P = .41). There was a trend toward more severe disease in placebo recipients at entry. More subjects in the placebo group experienced SAEs (P = .02). There were no SAEs clearly related to methylprednisolone administration, and methylprednisolone did not increase viral load. CONCLUSIONS Although methylprednisolone appears to be safe, it did not provide significant clinical benefit to patients. Our results do not support the use of methylprednisolone for HCPS. CLINICAL TRIALS REGISTRATION NCT00128180.

Case Report : T-Cell Responses During Clearance of Andes Virus From Blood Cells 2 Months After Severe Hantavirus Cardiopulmonary Syndrome

Hantavirus Cardiopulmonary Syndrome (HCPS) due to Andes virus (ANDV) is endemic in Chile and Argentina and currently demonstrates a case‐fatality rate of 37% in humans. By contrast to the chronically infected rodents, it is believed that ANDV in humans is cleared during the acute phase. Moreover, to date, both magnitude and quality of human T‐cell responses during ANDV infection and clearance are unknown. Using IFN‐γ and granzyme B ELISPOT assays as well as flow cytometry, we prospectively studied the ANDV‐specific T‐cell responses in a 56‐year‐old convalescing survivor of severe HCPS, whose blood cells remained PCR‐positive for ANDV‐RNA until day 53 after hospital admission, that is, 67 days after infection and 42 days after discharge. PCR‐negativity was closely related to the increase and function of (Gn46–60)‐specific IFN‐γ+ granzyme B+ CD8+ T‐cells, but not to neutralizing antibody titers. Concurrently, the phenotype of CD45RA+CCR7− Gn46–60‐specific T‐cells shifted from a CD28−CD27+ “intermediate” to a CD28− CD27− “late” effector memory beyond day 53 after hospital admission. This is the first report that shows that ANDV can persist in the human hosts for more than 2 months. Moreover, the kinetics of T‐cell responses during ANDV clearance may indicate a major role of T‐cells for clearance of ANDV and human immunity to this pathogen. J. Med. Virol. 80:1947–1951, 2008.

Hantavirus pulmonary syndrome, Southern Chile, 1995-2012.

Early clinical suspicion should prompt urgent transfer of patients to a hospital with intensive care facilities.

Molecular method for the detection of Andes hantavirus infection: validation for clinical diagnostics

Hantavirus cardiopulmonary syndrome is a severe disease caused by exposure to New World hantaviruses. Early diagnosis is difficult due to the lack of specific initial symptoms. Antihantavirus antibodies are usually negative until late in the febrile prodrome or the beginning of cardiopulmonary phase, while Andes hantavirus (ANDV) RNA genome can be detected before symptoms onset. We analyzed the effectiveness of quantitative reverse transcription polymerase chain reaction (RT-qPCR) as a diagnostic tool detecting ANDV-Sout genome in peripheral blood cells from 78 confirmed hantavirus patients and 166 negative controls. Our results indicate that RT-qPCR had a low detection limit (~10 copies), with a specificity of 100% and a sensitivity of 94.9%. This suggests the potential for establishing RT-qPCR as the assay of choice for early diagnosis, promoting early effective care of patients, and improving other important aspects of ANDV infection management, such as compliance of biosafety recommendations for health personnel in order to avoid nosocomial transmission.

Knowledge, attitudes, and practices regarding hantavirus disease and acceptance of a vaccine trial in rural communities of southern Chile

ABSTRACT Andes hantavirus cardiopulmonary syndrome, transmitted by Oligoryzomys longicaudatus, has no approved treatment, a case fatality rate of 35%, and documented person-to-person transmission. An Andes vaccine, highly needed for prevention, is in development. We aimed to evaluate knowledge, attitudes and practices (KAP) regarding hantavirus disease and willingness to participate in a future Andes vaccine trials through a cross sectional face-to-face oral survey of a randomly selected adult sample from 2 rural communes in southern Chile. Human subjects approval was obtained from our institutional IRBs, and participants signed informed consent. We enrolled 319 subjects from Corral and 321 from Curarrehue; 98% had heard about hantavirus disease and its reservoir but only half knew about transmission, symptoms and prevention. Participants fear the disease but are only partially aware of their own risk. One third of participants reported presence of rodents inside their homes. Despite moderate confidence in their health system, most subjects perceived vaccines as beneficial, and 93% would accept an approved hantavirus vaccine. Half would agree to participate in a vaccine trial and 29% would allow their children to participate. Motivations to participate were mainly altruistic, while risk perception was the main reason for declining. Knowledge about hantavirus disease and prevention practices require reinforcement, and a vaccine trial seems feasible in these populations.