Alain Michault

Genome microevolution of chikungunya viruses causing the Indian Ocean outbreak.

Background A chikungunya virus outbreak of unprecedented magnitude is currently ongoing in Indian Ocean territories. In Réunion Island, this alphavirus has already infected about one-third of the human population. The main clinical symptom of the disease is a painful and invalidating poly-arthralgia. Besides the arthralgic form, 123 patients with a confirmed chikungunya infection have developed severe clinical signs, i.e., neurological signs or fulminant hepatitis. Methods and Findings We report the nearly complete genome sequence of six selected viral isolates (isolated from five sera and one cerebrospinal fluid), along with partial sequences of glycoprotein E1 from a total of 127 patients from Réunion, Seychelles, Mauritius, Madagascar, and Mayotte islands. Our results indicate that the outbreak was initiated by a strain related to East-African isolates, from which viral variants have evolved following a traceable microevolution history. Unique molecular features of the outbreak isolates were identified. Notably, in the region coding for the non-structural proteins, ten amino acid changes were found, four of which were located in alphavirus-conserved positions of nsP2 (which contains helicase, protease, and RNA triphosphatase activities) and of the polymerase nsP4. The sole isolate obtained from the cerebrospinal fluid showed unique changes in nsP1 (T301I), nsP2 (Y642N), and nsP3 (E460 deletion), not obtained from isolates from sera. In the structural proteins region, two noteworthy changes (A226V and D284E) were observed in the membrane fusion glycoprotein E1. Homology 3D modelling allowed mapping of these two changes to regions that are important for membrane fusion and virion assembly. Change E1-A226V was absent in the initial strains but was observed in >90% of subsequent viral sequences from Réunion, denoting evolutionary success possibly due to adaptation to the mosquito vector. Conclusions The unique molecular features of the analyzed Indian Ocean isolates of chikungunya virus demonstrate their high evolutionary potential and suggest possible clues for understanding the atypical magnitude and virulence of this outbreak.

A mouse model for Chikungunya: young age and inefficient type-I interferon signaling are risk factors for severe disease.

Chikungunya virus (CHIKV) is a re-emerging arbovirus responsible for a massive outbreak currently afflicting the Indian Ocean region and India. Infection from CHIKV typically induces a mild disease in humans, characterized by fever, myalgia, arthralgia, and rash. Cases of severe CHIKV infection involving the central nervous system (CNS) have recently been described in neonates as well as in adults with underlying conditions. The pathophysiology of CHIKV infection and the basis for disease severity are unknown. To address these critical issues, we have developed an animal model of CHIKV infection. We show here that whereas wild type (WT) adult mice are resistant to CHIKV infection, WT mouse neonates are susceptible and neonatal disease severity is age-dependent. Adult mice with a partially (IFN-α/βR+/−) or totally (IFN-α/βR−/−) abrogated type-I IFN pathway develop a mild or severe infection, respectively. In mice with a mild infection, after a burst of viral replication in the liver, CHIKV primarily targets muscle, joint, and skin fibroblasts, a cell and tissue tropism similar to that observed in biopsy samples of CHIKV-infected humans. In case of severe infections, CHIKV also disseminates to other tissues including the CNS, where it specifically targets the choroid plexuses and the leptomeninges. Together, these data indicate that CHIKV-associated symptoms match viral tissue and cell tropisms, and demonstrate that the fibroblast is a predominant target cell of CHIKV. These data also identify the neonatal phase and inefficient type-I IFN signaling as risk factors for severe CHIKV-associated disease. The development of a permissive small animal model will expedite the testing of future vaccines and therapeutic candidates.

Outbreak of Chikungunya on Reunion Island: Early Clinical and Laboratory Features in 157 Adult Patients

BACKGROUND Chikungunya is a reemerging disease. In 2005-2006, a severe outbreak occurred on Reunion Island in the southwestern part of the Indian Ocean. Other islands in this area were affected during the same period. METHODS Adult patients with acute chikungunya (defined as onset of fever and/or polyarthralgia in the 5 days preceding consultation) and laboratory-confirmed chikungunya who were referred to Groupe Hospitalier Sud Reunion during the period from March 2005 through April 2006 were included in this retrospective study. Their clinical and laboratory features are reported. RESULTS Laboratory-confirmed acute chikungunya was documented in 157 patients. The mean age of patients was 57.9 years, and the ratio of male to female patients was 1.24 : 1. Sixty percent of patients had at least 1 comorbidity. Ninety-seven patients (61.8%) were hospitalized, and 60 (38.2%) were treated as outpatients. Five fatalities were reported. One hundred fifty-one patients (96.1%) experienced polyarthralgia, and 129 (89%) experienced fever. Gastrointestinal symptoms were reported by 74 patients (47.1%), and skin rash was reported by 63 (40.1%). Hemorrhagic signs were rare. Lymphopenia and hypocalcemia were the prominent laboratory findings. Severe thrombocytopenia was rarely observed. CONCLUSIONS Chikungunya virus can be responsible for explosive outbreaks of disease. Polyarthralgia and fever are the 2 main clinical features. In this era of travel and globalization, chikungunya should be considered in the differential diagnosis of febrile polyarthralgia with an abrupt onset.

Characterization of Reemerging Chikungunya Virus

An unprecedented epidemic of chikungunya virus (CHIKV) infection recently started in countries of the Indian Ocean area, causing an acute and painful syndrome with strong fever, asthenia, skin rash, polyarthritis, and lethal cases of encephalitis. The basis for chikungunya disease and the tropism of CHIKV remain unknown. Here, we describe the replication characteristics of recent clinical CHIKV strains. Human epithelial and endothelial cells, primary fibroblasts and, to a lesser extent, monocyte-derived macrophages, were susceptible to infection and allowed viral production. In contrast, CHIKV did not replicate in lymphoid and monocytoid cell lines, primary lymphocytes and monocytes, or monocyte-derived dendritic cells. CHIKV replication was cytopathic and associated with an induction of apoptosis in infected cells. Chloroquine, bafilomycin-A1, and short hairpin RNAs against dynamin-2 inhibited viral production, indicating that viral entry occurs through pH-dependent endocytosis. CHIKV was highly sensitive to the antiviral activity of type I and II interferons. These results provide a general insight into the interaction between CHIKV and its mammalian host.

Multidisciplinary Prospective Study of Mother-to-Child Chikungunya Virus Infections on the Island of La Réunion

Background An outbreak of chikungunya virus affected over one-third of the population of La Réunion Island between March 2005 and December 2006. In June 2005, we identified the first case of mother-to-child chikungunya virus transmission at the Groupe Hospitalier Sud-Réunion level-3 maternity department. The goal of this prospective study was to characterize the epidemiological, clinical, biological, and radiological features and outcomes of all the cases of vertically transmitted chikungunya infections recorded at our institution during this outbreak. Methods and Findings Over 22 mo, 7,504 women delivered 7,629 viable neonates; 678 (9.0%) of these parturient women were infected (positive RT-PCR or IgM serology) during antepartum, and 61 (0.8%) in pre- or intrapartum. With the exception of three early fetal deaths, vertical transmission was exclusively observed in near-term deliveries (median duration of gestation: 38 wk, range 35–40 wk) in the context of intrapartum viremia (19 cases of vertical transmission out of 39 women with intrapartum viremia, prevalence rate 0.25%, vertical transmission rate 48.7%). Cesarean section had no protective effect on transmission. All infected neonates were asymptomatic at birth, and median onset of neonatal disease was 4 d (range 3–7 d). Pain, prostration, and fever were present in 100% of cases and thrombocytopenia in 89%. Severe illness was observed in ten cases (52.6%) and mainly consisted of encephalopathy (n = 9; 90%). These nine children had pathologic MRI findings (brain swelling, n = 9; cerebral hemorrhages, n = 2), and four evolved towards persistent disabilities. Conclusions Mother-to-child chikungunya virus transmission is frequent in the context of intrapartum maternal viremia, and often leads to severe neonatal infection. Chikungunya represents a substantial risk for neonates born to viremic parturients that should be taken into account by clinicians and public health authorities in the event of a chikungunya outbreak.

Persistent arthralgia associated with chikungunya virus: a study of 88 adult patients on reunion island.

BACKGROUND An outbreak of chikungunya virus infection occurred on Reunion Island during the period 2005-2006. Persistent arthralgia after chikungunya virus infection has been reported, but few studies have treated this aspect of the disease. METHODS Adult patients with laboratory-confirmed acute chikungunya virus infection who were referred to Groupe Hospitalier Sud Reunion during the period 2005-2006 were asked to participate in the study. Patients were assessed a mean of 18 months after acute disease occurred. Assessment consisted of answering questions on a standard form, undergoing a medical examination, and being tested for the presence of IgM antibodies to chikungunya virus. RESULTS Eighty-eight patients (mean age, 58.3 years; male-to-female ratio, 1.1:1.0) were included in this study. Fifty-eight patients (65.9%) had been hospitalized for acute chikungunya virus infection, and a history of arthralgia before chikungunya virus infection was reported by 39 patients (44%). Fifty-six patients (63.6%) reported persistent arthralgia related to chikungunya virus infection, and in almost one-half of the patients, the joint pain had a negative impact on everyday activities. Arthralgia was polyarticular in all cases, and pain was continuous in 31 patients (55.4%). Overall, 35 patients (39.7%) had test results positive for IgM antibodies to chikungunya virus. CONCLUSIONS Persistent and disabling arthralgia was a frequent concern in this cohort of patients who had experienced severe chikungunya virus infection approximately 18 months earlier. Further studies are needed to evaluate the prevalence of persistent arthralgia in the general population to determine the real burden of the disease.

Type I IFN controls chikungunya virus via its action on nonhematopoietic cells.

Chikungunya virus (CHIKV) is the causative agent of an outbreak that began in La Réunion in 2005 and remains a major public health concern in India, Southeast Asia, and southern Europe. CHIKV is transmitted to humans by mosquitoes and the associated disease is characterized by fever, myalgia, arthralgia, and rash. As viral load in infected patients declines before the appearance of neutralizing antibodies, we studied the role of type I interferon (IFN) in CHIKV pathogenesis. Based on human studies and mouse experimentation, we show that CHIKV does not directly stimulate type I IFN production in immune cells. Instead, infected nonhematopoietic cells sense viral RNA in a Cardif-dependent manner and participate in the control of infection through their production of type I IFNs. Although the Cardif signaling pathway contributes to the immune response, we also find evidence for a MyD88-dependent sensor that is critical for preventing viral dissemination. Moreover, we demonstrate that IFN-α/β receptor (IFNAR) expression is required in the periphery but not on immune cells, as IFNAR−/−→WT bone marrow chimeras are capable of clearing the infection, whereas WT→IFNAR−/− chimeras succumb. This study defines an essential role for type I IFN, produced via cooperation between multiple host sensors and acting directly on nonhematopoietic cells, in the control of CHIKV.

Chikungunya Virus-associated Long-term Arthralgia: A 36-month Prospective Longitudinal Study

Background Arthritogenic alphaviruses, including Chikungunya virus (CHIKV), are responsible for acute fever and arthralgia, but can also lead to chronic symptoms. In 2006, a Chikungunya outbreak occurred in La Réunion Island, during which we constituted a prospective cohort of viremic patients (n = 180) and defined the clinical and biological features of acute infection. Individuals were followed as part of a longitudinal study to investigate in details the long-term outcome of Chikungunya. Methodology/Principal Findings Patients were submitted to clinical investigations 4, 6, 14 and 36 months after presentation with acute CHIKV infection. At 36 months, 22 patients with arthralgia and 20 patients without arthralgia were randomly selected from the cohort and consented for blood sampling. During the 3 years following acute infection, 60% of patients had experienced symptoms of arthralgia, with most reporting episodic relapse and recovery periods. Long-term arthralgias were typically polyarthralgia (70%), that were usually symmetrical (90%) and highly incapacitating (77%). They were often associated with local swelling (63%), asthenia (77%) or depression (56%). The age over 35 years and the presence of arthralgia 4 months after the disease onset are risk factors of long-term arthralgia. Patients with long-term arthralgia did not display biological markers typically found in autoimmune or rheumatoid diseases. These data helped define the features of CHIKV-associated chronic arthralgia and permitted an estimation of the economic burden associated with arthralgia. Conclusions/Significance This study demonstrates that chronic arthralgia is a frequent complication of acute Chikungunya disease and suggests that it results from a local rather than systemic inflammation.

Estimating Chikungunya prevalence in La Réunion Island outbreak by serosurveys: Two methods for two critical times of the epidemic

BackgroundChikungunya virus (CHIKV) caused a major two-wave seventeen-month-long outbreak in La Réunion Island in 2005–2006. The aim of this study was to refine clinical estimates provided by a regional surveillance-system using a two-stage serological assessment as gold standard.MethodsTwo serosurveys were implemented: first, a rapid survey using stored sera of pregnant women, in order to assess the attack rate at the epidemic upsurge (s1, February 2006; n = 888); second, a population-based survey among a random sample of the community, to assess the herd immunity in the post-epidemic era (s2, October 2006; n = 2442). Sera were screened for anti-CHIKV specific antibodies (IgM and IgG in s1, IgG only in s2) using enzyme-linked immunosorbent assays. Seroprevalence rates were compared to clinical estimates of attack rates.ResultsIn s1, 18.2% of the pregnant women were tested positive for CHIKV specific antibodies (13.8% for both IgM and IgG, 4.3% for IgM, 0.1% for IgG only) which provided a congruent estimate with the 16.5% attack rate calculated from the surveillance-system. In s2, the seroprevalence in community was estimated to 38.2% (95% CI, 35.9 to 40.6%). Extrapolations of seroprevalence rates led to estimate, at 143,000 and at 300,000 (95% CI, 283,000 to 320,000), the number of people infected in s1 and in s2, respectively. In comparison, the surveillance-system estimated at 130,000 and 266,000 the number of people infected for the same periods.ConclusionA rapid serosurvey in pregnant women can be helpful to assess the attack rate when large seroprevalence studies cannot be done. On the other hand, a population-based serosurvey is useful to refine the estimate when clinical diagnosis underestimates it. Our findings give valuable insights to assess the herd immunity along the course of epidemics.

Serious acute chikungunya virus infection requiring intensive care during the reunion island outbreak in 2005-2006

Objective:To report the clinical and laboratory findings of adults with serious chikungunya virus acute infection hospitalized in an intensive care unit. Design:Case series study from August 2005 to May 2006. Setting:Medical intensive care unit, South Reunion Hospital. Patients:We observed 33 episodes of confirmed acute chikungunya virus infection (chikungunya virus-IgM or reverse transcription-polymerase chain reaction positive in the serum) admitted to the intensive care unit. Interventions:We collected cerebrospinal fluid, serum, and sometimes tissue samples from patients with suspected chikungunya fever in our intensive care unit. These samples underwent viral testing for evidence of acute chikungunya virus infection. Measurements and Main Results:Of the 33 patients, 19 (58%) had chikungunya virus specific manifestations, 8 (24%) had associated acute infectious disease and 6 (18%) exacerbations of previous complaints. Among the chikungunya virus specific manifestations, we identified 14 cases of encephalopathy, one case each of myocarditis, hepatitis and Guillain Barré syndrome. Eighty-five percent of patients had a McCabe score = 1 (for nonfatal or no underlying disease). Mortality was 48%. Conclusions:Chikungunya virus infection may be responsible for very severe clinical presentation, including young patients with unremarkable medical histories. Chikungunya virus infection is strongly suspected to have neurologic, hepatic, and myocardial tropism leading to dramatic complications and high mortality rate.