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Chikungunya virus (CHIKV) is a mosquito-borne alphavirus causing a debilitating febrile illness with rheumatic disease symptoms of arthralgia and arthritis. Since its spread outside of Africa in 2005, it continues to cause outbreaks and disseminates into new territories. Intervention strategies are urgently required, including vaccination and antiviral approaches. To test efficacy, the use of small animal models is required. Two mouse strains, A129, with a deficiency in their type-I interferon (IFN) receptor, and C57BL/6 are widely used. A direct comparison of these strains alongside the wild-type parental strain of the A129 mice, 129Sv/Ev, was undertaken to assess clinical disease progression, viral loads in key tissues, histological changes and levels of sera biomarkers. Our results confirm the severe disease course in A129 mice which was not observed in the parental 129Sv/Ev strain. Of the two wild-type strains, viral loads were higher in 129Sv/Ev mice compared to C57BL/6 counterparts. Our results have established these models and parameters for the future testing of vaccines and antiviral approaches., Competing Interests: The authors declare no conflicts of interest. The funders had no role in the design of the study; in the collection, analyses, or interpretation of data; in the writing of the manuscript; or in the decision to publish the results.

Chikungunya (CHIKV) and Mayaro (MAYV) viruses are arthritogenic alphaviruses that promote an incapacitating and long-lasting inflammatory muscle-articular disease. Despite studies pointing out the importance of skeletal muscle (SkM) in viral pathogenesis, the long-term consequences on its physiology and the mechanism of persistence of symptoms are still poorly understood. Combining molecular, morphological, nuclear magnetic resonance imaging, and histological analysis, we conduct a temporal investigation of CHIKV and MAYV replication in a wild-type mice model, focusing on the impact on SkM composition, structure, and repair in the acute and late phases of infection. We found that viral replication and induced inflammation promote a rapid loss of muscle mass and reduction in fiber cross-sectional area by upregulation of muscle-specific E3 ubiquitin ligases MuRF1 and Atrogin-1 expression, both key regulators of SkM fibers atrophy. Despite a reduction in inflammation and clearance of infectious viral particles, SkM atrophy persists until 30 days post-infection. The genomic CHIKV and MAYV RNAs were still detected in SkM in the late phase, along with the upregulation of chemokines and anti-inflammatory cytokine expression. In agreement with the involvement of inflammatory mediators on induced atrophy, the neutralization of TNF and a reduction in oxidative stress using monomethyl fumarate, an agonist of Nrf2, decreases atrogen expression and atrophic fibers while increasing weight gain in treated mice. These data indicate that arthritogenic alphavirus infection could chronically impact body SkM composition and also harm repair machinery, contributing to a better understanding of mechanisms of arthritogenic alphavirus pathogenesis and with a description of potentially new targets of therapeutic intervention.

Alphaviruses are mosquito-transmitted pathogens that induce high levels of viremia, which facilitates dissemination and vector transmission. One prevailing paradigm is that, after skin inoculation, alphavirus-infected resident dendritic cells migrate to the draining lymph node (DLN), facilitating further rounds of infection and dissemination. Here, we assess the contribution of infiltrating myeloid cells to alphavirus spread. We observe two phases of virus transport to the DLN, one that occurs starting at 1 h post infection and precedes viral replication, and a second that requires replication in the skin, enabling transit to the bloodstream. Depletion of Ly6C + monocytes reduces local chikungunya (CHIKV) or Ross River virus (RRV) infection in the skin, diminishes the second phase of virus transport to the DLN, and delays spread to distal sites. Our data suggest that infiltrating monocytes facilitate alphavirus infection at the initial infection site, which promotes more rapid spread into circulation., Competing Interests: Declaration of interests M.S.D. is a consultant or member of a Scientific Advisory Board for Inbios, Vir Biotechnology, IntegerBio, Merck, GlaxoSmithKline, Akagera Medicines, and Moderna. The Diamond laboratory has received unrelated funding support in sponsored research agreements from Emergent BioSolutions, Moderna, IntegerBio, and Vir Biotechnology., (Copyright © 2024 The Author(s). Published by Elsevier Inc. All rights reserved.)

Arthralgia is a hallmark of chikungunya virus (CHIKV) infection and can be very debilitating and associated with a robust local inflammatory response. Many pathophysiological aspects associated with the disease remain to be elucidated. Here, we describe a novel model of CHIKV infection in immunocompetent mice and evaluate the role of tumour necrosis factor in the pathogenesis of the disease. C57BL/6 wild type (WT) or TNF receptor 1 deficient (TNFR1 -/- ) mice were inoculated with 1 × 10 6 PFU of CHIKV in the paw. Alternatively, etanercept was used to inhibit TNF in infected WT mice. Hypernociception, inflammatory and virological analysis were performed. Inoculation of CHIKV into WT mice induced persistent hypernociception. There was significant viral replication in target organs and local production of inflammatory mediators in early time-points after infection. CHIKV infection was associated with specific humoral IgM and IgG responses. In TNFR1 -/- mice, there was a decrease in the hypernociception threshold, which was associated with a milder local inflammatory response in the paw but delayed viral clearance. Local or systemic treatment with etanercept reduced CHIKV-induced hypernociception. This is the first study to describe hypernociception, a clinical correlation of arthralgia, in immunocompetent mice infected with CHIKV. It also demonstrates the dual role of TNF in contributing to viral clearance but driving tissue damage and hypernociception. Inhibition of TNF may have therapeutic benefits but its role in viral clearance suggests that viral levels must be monitored in CHIKV-infected patients and that TNF inhibitors should ideally be used in combination with anti-viral drugs., (© 2022 John Wiley & Sons Ltd.)

Ubiquitin regulatory X domain-containing proteins (UBXN) might be involved in diverse cellular processes. However, their in vivo physiological functions remain largely elusive. We recently showed that UBXN3B positively regulated stimulator-of-interferon-genes (STING)-mediated innate immune responses to DNA viruses. Herein, we reported the essential role of UBXN3B in the control of infection and immunopathogenesis of two arthritogenic RNA viruses, Chikungunya (CHIKV) and O'nyong'nyong (ONNV) viruses. Ubxn3b deficient ( Ubxn3b -/- ) mice presented higher viral loads, more severe foot swelling and immune infiltrates, and slower clearance of viruses and resolution of inflammation than the Ubxn3b +/+ littermates. While the serum cytokine levels were intact, the virus-specific immunoglobulin G and neutralizing antibody levels were lower in the Ubxn3b -/- mice. The Ubxn3b -/- mice had more neutrophils and macrophages, but much fewer B cells in the ipsilateral feet. Of note, this immune dysregulation was also observed in the spleens and blood of uninfected Ubxn3b -/- mice. UBXN3B restricted CHIKV replication in a cell-intrinsic manner but independent of type I IFN signaling. These results demonstrated a dual role of UBXN3B in the maintenance of immune homeostasis and control of RNA virus replication. IMPORTANCE The human genome encodes 13 ubiquitin regulatory X (UBX) domain-containing proteins (UBXN) that might participate in diverse cellular processes. However, their in vivo physiological functions remain largely elusive. Herein, we reported an essential role of UBXN3B in the control of infection and immunopathogenesis of arthritogenic alphaviruses, including Chikungunya virus (CHIKV), which causes acute and chronic crippling arthralgia, long-term neurological disorders, and poses a significant public health problem in the tropical and subtropical regions worldwide. However, there are no approved vaccines or specific antiviral drugs. This was partly due to a poor understanding of the protective and detrimental immune responses elicited by CHIKV. We showed that UBXN3B was critical for the control of CHIKV replication in a cell-intrinsic manner in the acute phase and persistent immunopathogenesis in the post-viremic stage. Mechanistically, UBXN3B was essential for the maintenance of hematopoietic homeostasis during viral infection and in steady-state.

Background: Re-emerging viral attacks are catastrophic for health and economy. It is crucial to grasp the viral life cycle, replication and mutation policies and attack strategies. It is also absolute to fathom the cost-efficient antiviral remedies earliest possible., Methods: We propose to use a lab-grown organ (re-cellularized scaffold of sheep kidney) for viral culture and understand its interaction with extra-cellular matrices of the host tissue., Results: Our findings showed that the chikungunya virus (CHIKV) could be better replicated in tissue-engineered bio models than cell culture. A decrease in ds-DNA levels emphasized that CHIKV propagates within the re-cellularized and cell culture models. There was an increase in the viral titres (pfu/ml) in re-cellularized scaffolds and control groups. The lipid peroxidation levels were increased as the infection was progressed in cell culture as well as re-cellularized and control groups. The onset and progress of the CHIKV attacks (cellular infection) lead to transmembrane domain fatty acid peroxidation and DNA breakdown, landing in cellular apoptosis. Simultaneously cell viability was inversely proportional to non-viability, and it decreased as the infection progressed in all infected groups. Histological findings and extracellular matrix evaluation showed the impairment in medullary, cortex regions due to propagation of CHIKV and plaques generations., Conclusion: This method will be a breakthrough for future virus culture, drug interaction and to study its effect on extracellular matrix alterations. This study will also allow us to investigate the correct role of any vaccine or antiviral drugs and their effects on re-engineered organ matrices before moving towards the animal models., (© 2022. Korean Tissue Engineering and Regenerative Medicine Society.)

Chikungunya is a fast-mutating virus causing Chikungunya virus disease (ChikvD) with a significant load of disability-adjusted life years (DALY) around the world. The outbreak of this virus is significantly higher in the tropical countries. Several experiments have identified crucial viral-host protein-protein interactions (PPIs) between Chikungunya Virus (Chikv) and the human host. However, no standard database that catalogs this PPI information exists. Here we develop a Chikv-Human PPI database, ChikvInt, to facilitate understanding ChikvD disease pathogenesis and the progress of vaccine studies. ChikvInt consists of 109 interactions and is available at www.chikvint.com., (© 2022 The Society for Applied Microbiology.)

Chikungunya virus (CHIKV) is a mosquito-borne alphavirus that recently re-emerged in many parts of the world causing large-scale outbreaks. CHIKV infection presents as a febrile illness known as chikungunya fever (CHIKF). Infection is self-limited and characterized mainly by severe joint pain and myalgia that can last for weeks or months; however, severe disease presentation can also occur in a minor proportion of infections. Among the atypical CHIKV manifestations that have been described, severe arthralgia and neurological complications, such as encephalitis, meningitis, and Guillain-Barré Syndrome, are now reported in many outbreaks. Moreover, death cases were also reported, placing CHIKV as a relevant public health disease. Virus evolution, globalization, and climate change may have contributed to CHIKV spread. In addition to this, the lack of preventive vaccines and approved antiviral treatments is turning CHIKV into a major global health threat. In this review, we discuss the current knowledge about CHIKV pathogenesis, with a focus on atypical disease manifestations, such as persistent arthralgia and neurologic disease presentation. We also bring an up-to-date review of the current CHIKV vaccine development. Altogether, these topics highlight some of the most recent advances in our understanding of CHIKV pathogenesis and also provide important insights into the current development and clinical trials of CHIKV potential vaccine candidates.

Chikungunya virus (CHIKV) is one of the emerging viruses around the globe. It belongs to the family Togaviridae and genus Alphavirus and is an arthropod borne virus that transmits by the bite of an infected mosquito, mainly through Aedes aegypti and Aedes albopcitus. It is a spherical, enveloped virus with positive single stranded RNA genome. It was first discovered during 1952-53 in Tanganyika, after which outbreaks were documented in many regions of the world. CHIKV has two transmission cycles; an enzootic sylvatic cycle and an urban cycle. CHIKV genome contains 11,900 nucleotides and two open reading frames and shows great sequence variability. Molecular mechanisms of virus host-cell interactions and the pathogenesis of disease are not fully understood. The disease involves three phases; acute, post-acute and chronic with symptoms including high-grade fever, arthralgia, macupapular rashes and headache. There is no licensed vaccine or specific treatment for CHIKV infection. This lack of specific interventions combined with difficulties in making a precise diagnosis together make the disease difficult to manage. In this review we aim to present the current knowledge of global epidemiology, transmission, structure, various aspects of diagnosis as well as highlight potential antiviral drugs and vaccines against CHIKV., (© 2021 John Wiley & Sons Ltd.)

Chikungunya virus (CHIKV) is a reemerging arthropod-borne alphavirus and a serious threat to human health. Therefore, efforts toward elucidating how this virus causes disease and the molecular mechanisms underlying steps of the viral replication cycle are crucial. Using an in vivo transmission system that allows intrahost evolution, we identified an emerging CHIKV variant carrying a mutation in the E1 glycoprotein (V156A) in the serum of mice and saliva of mosquitoes. E1 V156A has since emerged in humans during an outbreak in Brazil, cooccurring with a second mutation, E1 K211T, suggesting an important role for these residues in CHIKV biology. Given the emergence of these variants, we hypothesized that they function to promote CHIKV infectivity and subsequent disease. Here, we show that E1 V156A and E1 K211T modulate virus attachment and fusion and impact binding to heparin, a homolog of heparan sulfate, a key entry factor on host cells. These variants also exhibit differential neutralization by antiglycoprotein monoclonal antibodies, suggesting structural impacts on the particle that may be responsible for altered interactions at the host membrane. Finally, E1 V156A and E1 K211T exhibit increased titers in an adult arthritic mouse model and induce increased foot-swelling at the site of injection. Taken together, this work has revealed new roles for E1 where discrete regions of the glycoprotein are able to modulate cell attachment and swelling within the host. IMPORTANCE Alphaviruses represent a growing threat to human health worldwide. The reemerging alphavirus chikungunya virus (CHIKV) has rapidly spread to new geographic regions in the last several decades, causing overwhelming outbreaks of disease, yet there are no approved vaccines or therapeutics. The CHIKV glycoproteins are key determinants of CHIKV adaptation and virulence. In this study, we identify and characterize the emerging E1 glycoprotein variants, V156A and K211T, that have since emerged in nature. We demonstrate that E1 V156A and K211T function in virus attachment to cells, a role that until now has only been attributed to specific residues of the CHIKV E2 glycoprotein. We also demonstrate E1 V156A and K211T increase foot-swelling of the ipsilateral foot in mice infected with these variants. Observing that these variants and other pathogenic variants occur at the E1-E1 interspike interface, we highlight this structurally important region as critical for multiple steps during CHIKV infection. Together, these studies further define the function of E1 in CHIKV infection and can inform the development of therapeutic or preventative strategies.

Chikungunya virus (CHIKV) is an emerging mosquito-transmitted alphavirus that leads to acute fever and chronic debilitating polyarthralgia. To date, the mechanism underlying chronic recurrent arthralgia is unknown. In the present study, newborn wild-type C57BL/6 mice were infected with CHIKV, and the virological and pathological features of CHIKV infection were analyzed over a period of 50 days. Acute viral infection was readily established by footpad inoculation of CHIKV at doses ranging from 10 plaque forming unit (PFU) to 106 PFU, during which inoculation dose-dependent viral RNA and skeletal muscle damage were detected in the foot tissues. However, persistent CHIKV was observed only when the mice were infected with a high dose of 106 PFU of CHIKV, in which low copy numbers (103-104) of viral positive strand RNA were continuously detectable in the feet from 29 to 50 dpi, along with a low level and progressive reduction in virus-specific CD8+ T cell responses. In contrast, viral negative strand RNA was detected at 50 dpi but not at 29 dpi and was accompanied by significant local skeletal muscle damage at 50 dpi when mild synovial hyperplasia appeared in the foot joints, although the damage was briefly repaired at 29 dpi. These results demonstrated that a high viral inoculation dose leads to viral persistence and progression to chronic tissue damage after recovery from acute infection. Taken together, these results provide a useful tool for elucidating the pathogenesis of persistent CHIKV infection and viral relapse-associated chronic arthritis., Competing Interests: The authors have declared that no competing interests exist.

Chloroquine (CQ) and hydroxychloroquine (HCQ) have been used as antiviral agents for the treatment of severe acute respiratory syndrome coronavirus 2 (SARS-CoV2) infection. We performed a systematic review to examine whether prior clinical studies that compared the effects of CQ and HCQ to a control for the treatment of non-SARS-CoV2 infection supported the use of these agents in the present SARS-CoV2 outbreak. PubMed, EMBASE, Scopus and Web of Science (PROSPERO CRD42020183429) were searched from inception through 2 April 2020 without language restrictions. Of 1766 retrieved reports, 18 studies met our inclusion criteria, including 17 prospective controlled studies and one retrospective study. CQ or HCQ were compared to control for the treatment of infectious mononucleosis (EBV, n = 4), warts (human papillomavirus, n = 2), chronic HIV infection (n = 6), acute chikungunya infection (n = 1), acute dengue virus infection (n = 2), chronic HCV (n = 2), and as preventive measures for influenza infection (n = 1). Survival was not evaluated in any study. For HIV, the virus that was most investigated, while two early studies suggested HCQ reduced viral levels, four subsequent ones did not, and in two of these CQ or HCQ increased viral levels and reduced CD4 counts. Overall, three studies concluded CQ or HCQ were effective; four concluded further research was needed to assess the treatments' effectiveness; and 11 concluded that treatment was ineffective or potentially harmful. Prior controlled clinical trials with CQ and HCQ for non-SARS-CoV2 viral infections do not support these agents' use for the SARS-CoV2 outbreak., (Published 2021. This article is a U.S. Government work and is in the public domain in the USA.)

Chronic rheumatological manifestations similar to those of rheumatoid arthritis (RA) are described after chikungunya virus infection. We aimed to compare the relevance of joint counts and symptoms to clinical outcomes in RA and chronic chikungunya disease. Forty patients with chronic chikungunya arthralgia and 40 patients with RA were enrolled in a cross-sectional study. The association of tenderness and swelling, clinically assessed in 28 joints, and patient evaluations of pain and musculoskeletal stiffness with modified Health Assessment Questionnaire (HAQ) and quality of life (QoL) assessments were investigated. Tender and swollen joint counts, pain and stiffness scores were all associated with the HAQ disability index in RA (all r > 0.55, p ≤ 0.0002), but only stiffness was significantly associated with disability in chikungunya (r = 0.38, p = 0.02). Joint counts, pain and stiffness were also associated with most QoL domains in RA patients. In contrast, in chikungunya disease, tender joint counts were associated only with one QoL domain and swollen joints for none, while pain and stiffness were associated with several domains. Our results confirm the relevance of joint counts in RA, but suggest that in chronic chikungunya disease, joint counts have more limited value. Stiffness and pain score may be more important to quantify chikungunya arthritis impact., (© 2021. The Author(s).)

Chikungunya virus (CHIKV) is an arthropod-borne virus that causes large outbreaks world-wide leaving millions of people with severe and debilitating arthritis. Interestingly, clinical presentation of CHIKV arthritides have many overlapping features with rheumatoid arthritis including cellular and cytokine pathways that lead to disease development and progression. Currently, there are no specific treatments or vaccines available to treat CHIKV infections therefore advocating the need for the development of novel therapeutic strategies to treat CHIKV rheumatic disease. Herein, we provide an in-depth analysis of an efficacious new treatment for CHIKV arthritis with a semi-synthetic sulphated polysaccharide, Pentosan Polysulfate Sodium (PPS). Mice treated with PPS showed significant functional improvement as measured by grip strength and a reduction in hind limb foot swelling. Histological analysis of the affected joint showed local inflammation was reduced as seen by a decreased number of infiltrating immune cells. Additionally, joint cartilage was protected as demonstrated by increased proteoglycan staining. Using a multiplex-immunoassay system, we also showed that at peak disease, PPS treatment led to a systemic reduction of the chemokines CXCL1, CCL2 (MCP-1), CCL7 (MCP-3) and CCL12 (MCP-5) which may be associated with the reduction in cellular infiltrates. Further characterisation of the local effect of PPS in its action to reduce joint and muscle inflammation was performed using NanoString™ technology. Results showed that PPS altered the local expression of key functional genes characterised for their involvement in growth factor signalling and lymphocyte activation. Overall, this study shows that PPS is a promising treatment for alphaviral arthritis by reducing inflammation and protecting joint integrity., Competing Interests: This study was funded by a joint research grant funded from Paradigm Biopharmaceuticals Ltd (https://paradigmbiopharma.com/), Advance Queensland mid-career fellowship awarded to PAR and Griffith University. RK and CJMS are paid employees of Paradigm Biopharmaceuticals Ltd. As staff members of Paradigm Biopharmaceuticals Ltd, RK and CJMS are entitled to long term and short-term incentives. LJH was a paid part-time consultant for Paradigm Biopharmaceuticals Ltd. The specific roles of these authors are articulated in the ‘author contributions’ section. The funders did not have any role in the study design, data collection and analysis, decision to publish, or preparation of the manuscript beyond those specified. This research may lead to the use of PPS towards the treatment of CHIKV disease. Paradigm Biopharmaceuticals Ltd. hold the following patents PCT/AU2016/050408 “Treatment of Alphavirus-Induced Inflammation” (published as WO2017/201563) and the Australian patent number is 2016407635 “Treatment of Alphavirus-Induced Inflammation”. Furthermore, iPPS will be available in Australia via Australian infectious disease physicians through the TGA Special Access Scheme (SAS) for patients with chronic RRV-induced arthritis where there are no other treatment options. Access will be available in Q2 CY 2021 as a pay-for-use program, providing a path to first revenues for Paradigm. This does not alter our adherence to PLOS ONE policies on sharing data and materials.

Alphaviruses are positive sense, RNA viruses commonly transmitted by an arthropod vector to a mammalian or avian host. In recent years, a number of the Alphavirus members have reemerged as public health concerns. Transmission from mosquito vector to vertebrate hosts requires an understanding of the interaction between the virus and both vertebrate and insect hosts to develop rational intervention strategies. The current study uncovers a novel role for capsid protein during Chikungunya virus replication whereby the interaction with viral RNA in the E1 coding region regulates protein synthesis processes early in infection. Studies done in both the mammalian and mosquito cells indicate that interactions between viral RNA and capsid protein have functional consequences that are host species specific. Our data support a vertebrate-specific role for capsid:vRNA interaction in temporally regulating viral translation in a manner dependent on the PI3K-AKT-mTOR pathway., (Copyright © 2021 Elsevier Inc. All rights reserved.)

Background: Chikungunya fever is considered an abrupt onset arbovirus transmitted by mosquitoes, mainly Aedes aegypti and Aedes albopictus. The disease has a significant impact on the quality of life of affected persons, and many of its numerous symptoms have not yet been properly clarified, such as the manifestations that can occur in the oral cavity. The aim of this study was to identify the main oral manifestations related to chikungunya fever, as well as describe the demographic characteristics of patients, by conducting a systematic review of the literature., Methods and Findings: Searches were performed in MEDLINE (PubMed), Embase (Elsevier), LILACS (VHL), Cochrane Library, Scopus, and CAPES electronic databases for theses and dissertations published up to January 16, 2021 without language and date restrictions. Additional manual searches of gray literature, reference list, and Google Scholar were carried out. We included 27 studies highlighting mainly oral manifestations that cause masticatory discomfort such as ulcers and oral thrush, gingival bleeding, pain and burning of the oral mucous membranes, temporomandibular joint (TMJ) arthralgia, opportunistic infections, and changes in taste., Conclusions: There seems to be a predominance of oral manifestations that cause discomfort when chewing, such as ulcerations in the acute phase of the disease, with complete remission within 3 to 10 days after the onset, apparently mostly affecting women and older persons. These oral manifestations can be compatible with basic viral infections related to inflammatory response and transitory immunosuppression., Competing Interests: The authors have declared that no competing interests exist.

Chikungunya virus (CHIKV) is an emerging, mosquito-borne alphavirus responsible for acute to chronic arthralgias and neuropathies. Although it originated in central Africa, recent reports of disease have come from many parts of the world, including the Americas. While limiting human CHIKV cases through mosquito control has been used, it has not been entirely successful. There are currently no licensed vaccines or treatments specific for CHIKV disease, thus more work is needed to develop effective countermeasures. Current animal research on CHIKV is often not representative of human disease. Most models use CHIKV needle inoculation via unnatural routes to create immediate viremia and localized clinical signs; these methods neglect the natural route of transmission (the mosquito vector bite) and the associated human immune response. Since mosquito saliva has been shown to have a profound effect on viral pathogenesis, we evaluated a novel model of infection that included the natural vector, Aedes species mosquitoes, transmitting CHIKV to mice containing components of the human immune system. Humanized mice infected by 3-6 mosquito bites showed signs of systemic infection, with demonstrable viremia (by qRT-PCR and immunofluorescent antibody assay), mild to moderate clinical signs (by observation, histology, and immunohistochemistry), and immune responses consistent with human infection (by flow cytometry and IgM ELISA). This model should give a better understanding of human CHIKV disease and allow for more realistic evaluations of mechanisms of pathogenesis, prophylaxis, and treatments., Competing Interests: The authors have declared that no competing interests exist.

Throughout the last decade, chikungunya virus (CHIKV) and Zika virus (ZIKV) infections have spread globally, causing a spectrum of disease that ranges from self-limited febrile illness to permanent severe disability, congenital anomalies, and early death. Nevertheless, estimates of their aggregate health impact are absent from the literature and are currently omitted from the Global Burden of Disease (GBD) reports. We systematically reviewed published literature and surveillance records to evaluate the global burden caused by CHIKV and ZIKV between 2010 and 2019, to calculate estimates of their disability-adjusted life year (DALY) impact. Extracted data on acute, chronic, and perinatal outcomes were used to create annualized DALY estimates, following techniques outlined in the GBD framework. This study is registered with PROSPERO (CRD42020192502). Of 7,877 studies identified, 916 were screened in detail, and 21 were selected for inclusion. Available data indicate that CHIKV and ZIKV caused the average yearly loss of over 106,000 and 44,000 DALYs, respectively, between 2010 and 2019. Both viruses caused substantially more burden in the Americas than in any other World Health Organization (WHO) region. This unequal distribution is likely due to a combination of limited active surveillance reporting in other regions and the lack of immunity that left the previously unexposed populations of the Americas susceptible to severe outbreaks during the last decade. Long-term rheumatic sequelae provided the largest DALY component for CHIKV, whereas congenital Zika syndrome (CZS) contributed most significantly for ZIKV. Acute symptoms and early mortality accounted for relatively less of the overall burden. Suboptimal reporting and inconsistent diagnostics limit precision when determining arbovirus incidence and frequency of complications. Despite these limitations, it is clear from our assessment that CHIKV and ZIKV represent a significant cause of morbidity that is not included in current disease burden reports. These results suggest that transmission-blocking strategies, including vector control and vaccine development, remain crucial priorities in reducing global disease burden through prevention of potentially devastating arboviral outbreaks., Competing Interests: The authors have declared that no competing interests exist.

Chikungunya, a mosquito-borne disease that causes high fever and severe joint pain in humans, is a profound global threat because of its high rate of contagion and lack of antiviral interventions or vaccines for controlling the infection. The present study was aimed to investigate the antiviral activity of Stearylamine (SA) against Chikungunya virus (CHIKV) in both in vitro and in vivo. The antiviral activity of SA was determined by foci forming unit (FFU) assay, quantitative RT-PCR and cell-based immune-fluorescence assay (IFA). Further in vivo studies were carried out to see the effect of SA treatment in CHIKV infected C57BL/6 mice. The anti-CHIKV activity was evaluated using qRT-PCR in serum and muscle tissues at different time points and by histopathology. In vitro treatment with SA at a concentration of 50 μM showed a reduction of 1.23 ± 0.19 log10 FFU/mL at 16 h and 1.56 ± 0.12 log10 FFU/mL at 24 h posttreatment by FFU assay. qRT-PCR studies indicated that SA treatment at 50μM concentration showed a singnificant reduction of 1.6 ± 0.1 log10 and 1.27 ± 0.12 log10 RNA copies when compared with that of virus control at 16 and 24 h post incubation. Treatments in the C57BL/6 mice model revealed that SA at 20 mg/kg dose per day up to 3, 5 and 7 days, produced stronger inhibition against CHIKV indicating substantially decrease viral loads and inflammatory cell migration in comparison to a dose of 10 mg/kg. This first in vivo study clearly indicates that SA is effective by significantly reducing virus replication in serum and muscles. As a next-generation antiviral therapeutic, these promising results can be translated for the use of SA to rationalize and develop an ideal delivery system alone or in combination against CHIKV., (Copyright © 2021 Elsevier B.V. All rights reserved.)

Patients following infection by chikungunya virus (CHIKV) can suffer for months to years from arthralgia and arthritis. Interestingly, methotrexate (MTX) a major immune-regulatory drug has proved to be of clinical benefit. We have previously shown that CHIKV can persist in the joint of one patient 18 months post-infection and plausibly driving chronic joint inflammation but through ill-characterized mechanisms. We have pursued our investigations and report novel histological and in vitro data arguing for a plausible role of a COX-2-mediated inflammatory response post-CHIKV. In the joint, we found a robust COX-2 staining on endothelial cells, synovial fibroblasts and more prominently on multinucleated giant cells identified as CD11c+ osteoclasts known to be involved in bone destruction. The joint tissue was also strongly stained for CD3, CD8, CD45, CD14, CD68, CD31, CD34, MMP2, and VEGF (but not for NO synthase and two B cell markers). Dendritic cells were rarely detected. Primary human synovial fibroblasts were infected with CHIKV or stimulated either by the synthetic molecule polyriboinosinic:polyribocytidylic acid (PIC) to mimic chronic viral infection or cytokines. First, we found that PIC and CHIKV enhanced mRNA expression of COX-2. We further found that PIC but not CHIKV increased the mRNA levels of cPLA2α and of mPGES-1, two other central enzymes in PGE2 production. IFNβ upregulated cPLA2α and COX-2 transcription levels but failed to modulated mPGES-1 mRNA expression. Moreover, PIC, CHIKV and IFNβ decreased mRNA expression of the PGE2 degrading enzyme 15-PGDH. Interestingly, MTX failed to control the expression of all these enzymes. In sharp contrast, dexamethasone was able to control the capacity of pro-inflammatory cytokines, IL-1β as well as TNFα, to stimulate mRNA levels of cPLA2α, COX-2 and mPGES-1. These original data argue for a concerted action of CHIKV (including viral RNA) and cytokines plausibly released from recruited leukocytes to drive a major COX-2-mediated PGE2 proinflammatory responses to induce viral arthritis., Competing Interests: The authors have declared that no competing interest exist.

Background: Perinatally chikungunya infected neonates have been reported to have high rates of post-infection neurologic sequelae, mainly cognitive problems. In older children and adults chikungunya does not appear to have sequelae, but data on postnatally infected infants are lacking., Methods: We performed a prospective, non-controlled, observational study of infants infected before the age of 6 months with a severe chikungunya infection during the 2014-2015 epidemic in Curaçao, Dutch Antilles. Two years post-infection cognitive and motor - (BSID-III) and social emotional assessments (ITSEA) were performed., Results: Of twenty-two infected infants, two died and two were lost to follow up. Eighteen children were seen at follow-up and included in the current study. Of these, 13 (72%) had abnormal scores on the BSID-III (cognitive/motor) or ITSEA., Conclusion: In the first study aimed at postnatally infected infants, using an uncontrolled design, we observed a very high percentage of developmental problems. Further studies are needed to assess causality, however until these data are available preventive measure during outbreaks should also include young infants. Those that have been infected in early infancy should receive follow up.

The WHO defined three clinical forms for chikungunya virus infection (CHIKV, namely, acute, atypical, and severe cases) and a chronic form. These definitions seemed inappropriate for the elderly. So, we propose an adapted definition for elderly people. A cross-sectional analysis was performed including patients aged ≥ 65 years, who attended the emergency department with a positive biological diagnosis of CHIKV in 2014. A total of 267 elderly patients (80 ± 8 years) were included. When using the 2015 WHO definitions, 114 patients could not be classified (42.7%) in any of the category, of whom 43 (37.7%) reported absence of fever, 85 (74.6%) reported absence of joint pain, and 14 (12.3%) reported absence of both fever and joint pain. After adaptation of the WHO definitions, the 114 unclassifiable patients were reclassified as follows: eight as typical cases, 50 as atypical cases, 42 as severe cases, and 14 remained unclassifiable. The atypical clinical form was the most common form. The 2015 WHO definitions of the clinical forms at the acute phase of CHIKV are ill suited to the elderly. The adapted definition we propose here appears to be more appropriate and could help improved management of older patients with CHIKV.

Background: Ras-GTPase activating protein SH3-domain-binding proteins (G3BP) are a small family of RNA-binding proteins implicated in regulating gene expression. Changes in expression of G3BPs are correlated to several cancers including thyroid, colon, pancreatic and breast cancer. G3BPs are important regulators of stress granule (SG) formation and function. SG are ribonucleoprotein (RNP) particles that respond to cellular stresses to triage mRNA resulting in transcripts being selectively degraded, stored or translated resulting in a change of gene expression which confers a survival response to the cell. These changes in gene expression contribute to the development of drug resistance. Many RNA viruses, including Chikungunya (and potentially Coronavirus), dismantle SG so that the cell cannot respond to the viral infection. Non-structural protein 3 (nsP3), from the Chikungunya virus, has been shown to translocate G3BP away from SG. Interestingly in cancer cells, the formation of SG is correlated to drug-resistance and blocking SG formation has been shown to reestablish the efficacy of the anticancer drug bortezomib., Methods: Chikungunya nsP3 was transfected into breast cancer cell lines T47D and MCF7 to disrupt SG formation. Changes in the cytotoxicity of bortezomib were measured., Results: Bortezomib cytotoxicity in breast cancer cell lines changed with a 22 fold decrease in its IC 50 for T47D and a 7 fold decrease for MCF7 cells., Conclusions: Chikungunya nsP3 disrupts SG formation. As a result, it increases the cytotoxicity of the FDA approved drug, bortezomib. In addition, the increased cytotoxicity appears to correlate to improved bortezomib selectivity when compared to control cell lines., (Copyright © 2020 IMSS. All rights reserved.)

Background: Chikungunya virus (CHIKV) is an arthropod-borne RNA virus which induces host Endoplasmic Reticulum (ER) stress by accumulating unfolded or misfolded proteins. ER stress activates the unfolded protein response (UPR) pathway to enable proper protein folding and maintain cellular homeostasis. There is no approved drug or vaccine available for CHIKV treatment, therefore, a pharmacological countermeasure is warranted for preventing CHIKV infection., Objective: With a view to find a treatment modality for chikungunya infection, "andrographolide", a plant-derived diterpenoid with reported antiviral, anti-inflammatory and immunomodulatory effects, was used to investigate its role in chikungunya induced unfolded protein stress and apoptosis., Methods: Cells and supernatant collected on andrographolide and VER-155008, a GRP78 inhibitor, treatment in CHIKV infected and mock-infected THP-1 cells were tested for differential expression of UPR pathway proteins including GRP78, PERK, EIF-2α, IRE-1α, XBP-1 and ATF6. Furthermore, the inflammasome and apoptosis pathway proteins, i.e., caspase-1, caspase-3 and PARP, were tested by immunoblotting, and cytokines, i.e., IL-1β, IL-6 and IFN-γ were tested by ELISA., Results: Andrographolide treatment in CHIKV infected THP-1 cells significantly reduced IRE1α and downstream spliced XBP1 protein expression. Furthermore, CHIKV induced apoptosis and viral protein expression were also reduced on andrographolide treatment. A comparative analysis of andrographolide versus VER-155008, confirmed that andrographolide surpasses the effects of VER-155008 in suppressing the CHIKV induced ER stress., Conclusion: The study, therefore, confirms that andrographolide is a potential remedy for chikungunya infection and suppresses CHIKV induced ER stress and apoptosis., (Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.net.)

Competing Interests: Competing interests: None declared.

We report the case of a 38-year-old woman who initially consulted for an undifferentiated fever. Although her clinical condition evolved with signs and symptoms compatible with dengue with alarm signs and that the anti-dengue IgM detection in a single sample indicated it was a probable case that could have happened during the previous three months, the patient kept consulting due to little improvement. On the tenth day after the onset of symptoms, she presented with painful polyarticular symmetric edema, as well as hyperpigmented lesions in the nasolabial fold. Chikungunya diagnosis was confirmed by the presence of IgM antibodies. In endemic countries for dengue and chikungunya, the possibility of co-infection exists, but it may go unnoticed. On the other hand, the co-infection may worsen the clinical course of these diseases. Therefore, physicians should evaluate the clinical and laboratory characteristics of both infections to be able to diagnose the coinfection for adequate management and to minimize complications.

Chikungunya virus (CHIKV), an emerging alphavirus, has infected millions of people. However, the factors modulating disease outcome remain poorly understood. Here, we show in germ-free mice or in oral antibiotic-treated conventionally housed mice with depleted intestinal microbiomes that greater CHIKV infection and spread occurs within 1 day of virus inoculation. Alteration of the microbiome alters TLR7-MyD88 signaling in plasmacytoid dendritic cells (pDCs) and blunts systemic production of type I interferon (IFN). Consequently, circulating monocytes express fewer IFN-stimulated genes and become permissive for CHIKV infection. Reconstitution with a single bacterial species, Clostridium scindens, or its derived metabolite, the secondary bile acid deoxycholic acid, can restore pDC- and MyD88-dependent type I IFN responses to restrict systemic CHIKV infection and transmission back to vector mosquitoes. Thus, symbiotic intestinal bacteria modulate antiviral immunity and levels of circulating alphaviruses within hours of infection through a bile acid-pDC-IFN signaling axis, which affects viremia, dissemination, and potentially transmission., Competing Interests: Declaration of Interests M.S.D. is a consultant for Inbios, Eli Lilly, Vir Biotechnology, NGM Biopharmaceuticals, and on the Scientific Advisory Board of Moderna. The Diamond laboratory has received unrelated funding under sponsored research agreements from Moderna and Emergent BioSolutions., (Copyright © 2020 Elsevier Inc. All rights reserved.)

Chikungunya virus (CHIKV) is an enveloped virus that enters host cells and transits within the endosomes before starting its replication cycle, the precise mechanism of which is yet to be elucidated. Endocytosis and endosome acidification inhibitors inhibit infection by CHIKV, murine leukemia virus (MLV), or SARS-coronavirus, indicating that these viral entries into host cells occur through endosomes and require endosome acidification. Although endosomal cathepsin B protease is necessary for MLV, Ebola virus, and SARS-CoV infections, its role in CHIKV infection is unknown. Our results revealed that endocytosis inhibitors attenuated CHIKV-pseudotyped MLV vector infection in 293T cells but not in TE671 cells. In contrast, macropinocytosis inhibitors attenuated CHIKV-pseudotyped MLV vector infection in TE671 cells but not in 293T cells, suggesting that CHIKV host cell entry occurs via endocytosis or macropinocytosis, depending on the cell lines used. Cathepsin B inhibitor and knockdown by an shRNA suppressed CHIKV-pseudotyped MLV vector infection both in 293T and TE671 cells. These results show that cathepsin B facilitates CHIKV infection regardless of the entry pathway.

Key Results: Both outbreaks started in small towns, but cases were also detected in nearby larger cities where transmission was limited to small clusters. The time spans between the first and the last symptom onsets were similar between the 2 outbreaks, and the delay from the symptom onset of the index case and the first case notified was considerable. Comparable infection and transmission rates were observed in laboratory. The basic reproductive number (R0) was estimated in the range of 1.8-6 (2007) and 1.5-2.6 (2017). Clinical characteristics were similar between outbreaks, and no acute complications were reported, though a higher frequency of ocular symptoms, myalgia, and rash was observed in 2017. Very little is known about the immune mediator profile of CHIKV-infected patients during the 2 outbreaks. Regarding public health responses, after the 2007 outbreak, the Italian Ministry of Health developed national guidelines to implement surveillance and good practices to prevent and control autochthonous transmission. However, only a few regional authorities implemented it, and the perception of outbreak risk and knowledge of clinical symptoms and transmission dynamics by general practitioners remained low., Major Conclusions: Efforts should be devoted to developing suitable procedures for early detection of virus circulation in the population, possibly through the analysis of medical records in near real time. Increasing the awareness of CHIKV of general practitioners and public health officials through tailored education may be effective, especially in small coastal towns where the outbreak risk may be higher. A key element is also the shift of citizen awareness from considering Aedes mosquitoes not only as a nuisance problem but also as a public health one. We advocate the need of strengthening the surveillance and of promoting the active participation of the communities to prevent and contain future outbreaks., Competing Interests: The authors have declared that no competing interests exist.

Chikungunya virus (CHIKV) is a mosquito-borne virus that causes arthralgic fever. Fibroblast-like synoviocytes play a key role in joint damage in inflammatory arthritides and can additionally serve as target cells for CHIKV infection. To gain a better understanding of CHIKV-induced arthralgia, the interaction between CHIKV and synoviocytes was investigated at the protein level. A gel-enhanced liquid chromatography-mass spectrometry (GeLC-MS/MS) approach was used to examine protein expression from primary human fibroblast-like synoviocytes (HFLS) infected with clinical isolates of CHIKV at 12 and 24 hr post infection. Our analysis identified 259 and 241 proteins of known function that were differentially expressed (>1.5 or <-1.5 fold change) following CHIKV infection at 12 and 24 hpi, respectively. These proteins are involved in cellular homeostasis, including cellular trafficking, cytoskeletal organization, immune response, metabolic process, and protein modification. Some of these proteins have previously been reported to participate in arthralgia/arthritis and the death of infected cells. Our results provide information on the CHIKV-induced modulation of cellular proteins of HFLS at an early stage of infection, as well as highlighting biological processes associated with CHIKV infection in the main target cells of the joint., (© 2020 The Societies and John Wiley & Sons Australia, Ltd.)

Chikungunya fever (CHIKF) is a major public health concern and is caused by chikungunya virus (CHIKV). In 2005, the virus was reintroduced into India, resulting in massive outbreaks in several parts of the country. During 2010 and 2016 outbreaks, we recruited 588 patients from a tertiary care hospital in New Delhi, India, during the acute phase of CHIKF; collected their blood and clinical data; and determined their arthralgic status 12 weeks post-onset of fever. We evaluated IgM/IgG CHIKV-binding antibodies and their neutralizing capacity, sequenced complete genomes of 21 CHIKV strains, and correlated mutations with patient sequelae status. We also performed infections in murine models using representative strains from each outbreak to evaluate differences in pathogenesis. Our screening and analysis revealed that patients of the 2016 outbreak developed earlier IgM and neutralizing antibody responses that were negatively correlated with sequelae, compared with 2010 patients. Mutations that correlated with human disease progression were also correlated with enhanced murine virulence and pathogenesis. Overall, our study suggests that the development of early neutralizing antibodies and sequence variation in clinical isolates are predictors of human sequelae.

Background: The chikungunya virus (CHIKV) is a re-emerging alphavirus that can cause chronic and potentially incapacitating rheumatic musculoskeletal disorders known as chronic chikungunya arthritis (CCA). We conducted a prospective cohort study of CHIKV-infected subjects during the 2013 chikungunya outbreak in Martinique. The aim of this study was to assess the prevalence of CCA at 12 months and to search for acute phase factors significantly associated with chronicity., Methodology/principal Findings: A total of 193 patients who tested positive for CHIKV RNA via qRT-PCR underwent clinical investigations in the acute phase (<21 days), and then 3, 6, and 12 months after inclusion. The Asian lineage was identified as the circulating genotype. A total of 167 participants were classified as either with or without CCA, and were analyzed using logistic regression models. The overall prevalence of CCA at 12 months was 52.1% (95%CI: 44.5-59.7). In univariate analysis, age (RD 9.62, 95% CI, 4.87;14.38, p<0.0001), female sex (RD 15.5, 95% CI, 1.03;30.0, p = 0.04), headache (RD 15.42, 95% CI, 0.65;30.18 p = 0.04), vertigo (RD 15.33, 95% CI, 1.47;29.19, p = 0.03), vomiting (RD 12.89, 95% CI, 1.54;24.24, p = 0.03), dyspnea (RD 13.53, 95% CI, 0.73;26.33, p = 0.04), intravenous rehydration (RD -16.12, 95% CI, -31.58; -0.66 p = 0.04) and urea (RD 0.66, 95% CI, 0.12;1.20, p = 0.02) were significantly associated with the development of CCA. For the subpopulation with data on joint involvement in the acute phase, the risk factors significantly associated with CCA were at least one 1 enthesitis (RD 16.7, 95%CI, 2.8; 30.7, p = 0.02) and at least one tenosynovitis (RD 16.8, 95% CI, 1.4-32.2, p = 0.04)., Conclusions: This cohort study conducted in Martinique confirms that CCA is a common complication of acute chikungunya disease. Our analysis emphasized the importance of age and female sex for CCA occurrence, and highlighted the aggravating role of dehydration during the acute phase. Early and adequate hydration were found to reduce the risk chronic chikungunya disorders., Trial Registration: clinicaltrials.gov (NCT01099852)., Competing Interests: The authors have declared that no competing interests exist.

Chikungunya virus (CHIKV) is an arbovirus capable of causing a severe and often debilitating rheumatic syndrome in humans. CHIKV replicates in a wide variety of cell types in mammals, which has made attributing pathologic outcomes to replication at specific sites difficult. To assess the contribution of CHIKV replication in skeletal muscle cells to pathogenesis, we engineered a CHIKV strain exhibiting restricted replication in these cells via incorporation of target sequences for skeletal muscle cell-specific miR-206. This virus, which we term SKE, displayed diminished replication in skeletal muscle cells in a mouse model of CHIKV disease. Mice infected with SKE developed less severe disease signs, including diminished swelling in the inoculated foot and less necrosis and inflammation in the interosseous muscles. SKE infection was associated with diminished infiltration of T cells into the interosseous muscle as well as decreased production of Il1b, Il6, Ip10, and Tnfa transcripts. Importantly, blockade of the IL-6 receptor led to diminished swelling of a control CHIKV strain capable of replication in skeletal muscle, reducing swelling to levels observed in mice infected with SKE. These data implicate replication in skeletal muscle cells and release of IL-6 as important mediators of CHIKV disease.

After the mass campaign of Measles and Rubella vaccination in 2003 in Iran, the cases of measles and rubella infection decreased but still, the cases of rash and fever were reported. It is worth noting that some other viral infections show signs similar to measles and rubella such as some arboviruses. Considering the epidemic outbreak of arbovirus infections in countries neighbouring Iran, we performed this study to estimate the possibility of chikungunya and dengue fever among measles and rubella IgM negative patients presenting with rash and fever from December 2016 to November 2017 in the National Measles Laboratory at Tehran University of Medical Sciences. Serum samples were selected at random from patients from eight provinces. The presence of DENV IgM and CHIKV IgM was examined by enzyme-linked immunosorbent assay. Of the 1306 sera tested, 210 were CHIKV seropositive and 82 were dengue seropositive. Statistical analysis demonstrated a significant increase in the CHIKV IgM antibody seropositivity rate in Kerman (OR = 2.07, 95% CI: 1.10-3.92; P = 0.024) and Fars (OR = 1.77, 95% CI: 1.06-2.93; P = 0.027). The DENV and CHIKV seropositivity rate in summer is higher than in other seasons (P < 0.01). Our seropositive samples suggest possible CHIKV and DENV infection in Iran. It is likely that these viruses are circulating in Iran and there is a need to study vector carriage of these two viruses.

In 2013, cases of chikungunya virus (CHIKV) infection were first detected in the Caribbean. Chikungunya virus rapidly spread through Central and South America, causing explosive outbreaks in naive populations. Since its emergence in 2004, the number of case and series reports describing severe, atypical manifestations seen in chikungunya patients has increased substantially, calling into question whether clinicians and health services are failing to diagnose these atypical cases because of not only insufficient knowledge but also limitations in the case classification. Although this classification based on the duration of the musculoskeletal (acute, subacute, and chronic forms) complaints helped guide therapeutic approaches directed to these manifestations, patients presenting severe or complicated forms, which are less frequent but produce most of the fatal outcomes, were not properly addressed. In Brazil and the Caribbean, a clear temporal and spatial association between excess overall mortality and the occurrence of chikungunya epidemics has been shown, supporting the hypothesis that many of these excess deaths were a consequence of CHIKV infections. Thus, accumulated experience has highlighted that the current chikungunya case classification does not encompass the actual needs presented by certain cases with atypical features nor does it contribute to early detection and management of potentially severe cases. With continued CHIKV circulation in three continents and recent reemergence in Asia and Europe, we need a classification that is prospective and informed both by initial clinical presentation and by progression of signs and symptoms.

Type I interferons (IFNs) are key mediators of the innate immune response. Although members of this family of cytokines signal through a single shared receptor, biochemical and functional variation exists in response to different IFN subtypes. While previous work has demonstrated that type I IFNs are essential to control infection by chikungunya virus (CHIKV), a globally emerging alphavirus, the contributions of individual IFN subtypes remain undefined. To address this question, we evaluated CHIKV pathogenesis in mice lacking IFN-β (IFN-β knockout [IFN-β-KO] mice or mice treated with an IFN-β-blocking antibody) or IFN-α (IFN regulatory factor 7 knockout [IRF7-KO] mice or mice treated with a pan-IFN-α-blocking antibody). Mice lacking either IFN-α or IFN-β developed severe clinical disease following infection with CHIKV, with a marked increase in foot swelling compared to wild-type mice. Virological analysis revealed that mice lacking IFN-α sustained elevated infection in the infected ankle and in distant tissues. In contrast, IFN-β-KO mice displayed minimal differences in viral burdens within the ankle or at distal sites and instead had an altered cellular immune response. Mice lacking IFN-β had increased neutrophil infiltration into musculoskeletal tissues, and depletion of neutrophils in IFN-β-KO but not IRF7-KO mice mitigated musculoskeletal disease caused by CHIKV. Our findings suggest disparate roles for the IFN subtypes during CHIKV infection, with IFN-α limiting early viral replication and dissemination and IFN-β modulating neutrophil-mediated inflammation. IMPORTANCE Type I interferons (IFNs) possess a range of biological activity and protect against a number of viruses, including alphaviruses. Despite signaling through a shared receptor, there are established biochemical and functional differences among the IFN subtypes. The significance of our research is in demonstrating that IFN-α and IFN-β both have protective roles during acute chikungunya virus (CHIKV) infection but do so by distinct mechanisms. IFN-α limits CHIKV replication and dissemination, whereas IFN-β protects from CHIKV pathogenesis by limiting inflammation mediated by neutrophils. Our findings support the premise that the IFN subtypes have distinct biological activities in the antiviral response., (Copyright © 2019 American Society for Microbiology.)

Mxra8 is a recently described receptor for multiple alphaviruses, including Chikungunya (CHIKV), Mayaro (MAYV), Ross River (RRV), and O'nyong nyong (ONNV) viruses. To determine its role in pathogenesis, we generated mice with mutant Mxra8 alleles: an 8-nucleotide deletion that produces a truncated, soluble form (Mxra8 Δ8/Δ8 ) and a 97-nucleotide deletion that abolishes Mxra8 expression (Mxra8 Δ97/Δ97 ). Mxra8 Δ8/Δ8 and Mxra8 Δ97/Δ97 fibroblasts show reduced CHIKV infection in culture, and Mxra8 Δ8/Δ8 and Mxra8 Δ97/Δ97 mice have decreased infection of musculoskeletal tissues with CHIKV, MAYV, RRV, or ONNV. Less foot swelling is observed in CHIKV-infected Mxra8 mutant mice, which correlated with fewer infiltrating neutrophils and cytokines. A recombinant E2-D71A CHIKV with diminished binding to Mxra8 is attenuated in vivo in wild-type mice. Ectopic Mxra8 expression is sufficient to enhance CHIKV infection and lethality in transgenic flies. These studies establish a role for Mxra8 in the pathogenesis of multiple alphaviruses and suggest that targeting this protein may mitigate disease in humans., (Copyright © 2019 The Author(s). Published by Elsevier Inc. All rights reserved.)

Objective: Arthritogenic alphaviruses, such as Ross River virus (RRV) and chikungunya virus (CHIKV), particularly affect joints of the extremities and can lead to debilitating and potentially chronic polyarthritis/polyarthralgia. The innate immune response of the host plays a crucial role in inducing proinflammatory host factors, leading to tissue destruction and bone loss in the joints. This study was performed to assess how the inhibition of interleukin-1β (IL-1β) signaling using the clinical rheumatoid arthritis drug anakinra influences bone loss in mice with arthritogenic alphavirus infections., Methods: Mice (n = 5 per group) were infected with RRV or CHIKV and then treated with anakinra. Weight gain and disease severity were measured, tissue viral titers were determined, and histologic changes in joint tissues were assessed., Results: Anakinra therapy reduced RRV- and CHIKV-induced bone loss in this murine model (P < 0.001 and P < 0.05, respectively). Histologic analysis of the knee joint showed that treatment with anakinra decreased epiphyseal growth plate thinning, loss of epiphyseal bone volume, and osteoclastogenesis in the tibia. Importantly, pharmacologic IL-1 receptor (IL-1R) blockade did not improve other clinical features, including disease score, weight loss, or viremia., Conclusion: The present findings suggest that anakinra therapy may reduce bone loss in experimental murine models of RRV and CHIKV. Further investigations are needed to assess the potential therapeutic benefits of anakinra in patients with arthritogenic alphavirus disease., (© 2019, American College of Rheumatology.)

Chikungunya (CHIK) is a febrile arboviral illness caused by chikungunya virus (CHIKV) and has been identified in more than 60 countries across the globe. A major public health concern, the infection occurs as an acute febrile phase and a chronic arthralgic phase. The disease manifests differently in different age groups that can range from asymptomatic infection in the younger age group to a prolonged chronic phase in the elderly population. The present study was undertaken to evaluate strain-specific pathogenesis of ECSA genotype of CHIKV strains derived from clinical isolates in adult C57BL/6J mice model. The strain that was pathogenic and developed distinct acute and post-acute phase of CHIK infection was further evaluated for dose-dependent pathogenesis. Upon arriving on the optimal dose to induce clinical symptoms in the mice, the disease progression was evaluated across the acute and the post-acute phase of infection for a period of 15 days post-infection in two age groups of mice, namely eight weeks old and 20 weeks old mice groups. Biochemical, hematological, and virology attributes were measured and correlated to morbidity and linked neurotropism and limb thickness in the two age groups. Our results show that CHIKV exhibit strain-specific pathogenesis in C57BL/6J mice. Distinct dissimilarities were observed between the two age groups in terms of pathogenesis, viral clearance and host response to CHIKV infection.

The largest ever recorded epidemic of the Chikungunya virus (CHIKV) broke out in 2004 and affected four continents. Acute symptomatic infections are typically associated with the onset of fever and often debilitating polyarthralgia/polyarthritis. In this study, a systems biology approach was adopted to analyze the blood transcriptomes of adults acutely infected with the CHIKV. Gene signatures that were associated with viral RNA levels and the onset of symptoms were identified. Among these genes, the putative role of the Eukaryotic Initiation Factor (eIF) family genes and apolipoprotein B mRNA editing catalytic polypeptide-like (APOBEC3A) in the CHIKV replication process were displayed. We further compared these signatures with signatures induced by the Dengue virus infection and rheumatoid arthritis. Finally, we demonstrated that the CHIKV in vitro infection of murine bone marrow-derived macrophages induced IL-1 beta production in a mechanism that is significantly dependent on the inflammasome NLRP3 activation. The observations provided valuable insights into virus-host interactions during the acute phase and can be instrumental in the investigation of new and effective therapeutic interventions., Competing Interests: The authors have declared that no competing interests exist.

G3BP-1 and -2 (hereafter referred to as G3BP) are multifunctional RNA-binding proteins involved in stress granule (SG) assembly. Viruses from diverse families target G3BP for recruitment to replication or transcription complexes in order to block SG assembly but also to acquire pro-viral effects via other unknown functions of G3BP. The Old World alphaviruses, including Semliki Forest virus (SFV) and chikungunya virus (CHIKV) recruit G3BP into viral replication complexes, via an interaction between FGDF motifs in the C-terminus of the viral non-structural protein 3 (nsP3) and the NTF2-like domain of G3BP. To study potential proviral roles of G3BP, we used human osteosarcoma (U2OS) cell lines lacking endogenous G3BP generated using CRISPR-Cas9 and reconstituted with a panel of G3BP1 mutants and truncation variants. While SFV replicated with varying efficiency in all cell lines, CHIKV could only replicate in cells expressing G3BP1 variants containing both the NTF2-like and the RGG domains. The ability of SFV to replicate in the absence of G3BP allowed us to study effects of different domains of the protein. We used immunoprecipitation to demonstrate that that both NTF2-like and RGG domains are necessary for the formation a complex between nsP3, G3BP1 and the 40S ribosomal subunit. Electron microscopy of SFV-infected cells revealed that formation of nsP3:G3BP1 complexes via the NTF2-like domain was necessary for clustering of cytopathic vacuoles (CPVs) and that the presence of the RGG domain was necessary for accumulation of electron dense material containing G3BP1 and nsP3 surrounding the CPV clusters. Clustered CPVs also exhibited localised high levels of translation of viral mRNAs as detected by ribopuromycylation staining. These data confirm that G3BP is a ribosomal binding protein and reveal that alphaviral nsP3 uses G3BP to concentrate viral replication complexes and to recruit the translation initiation machinery, promoting the efficient translation of viral mRNAs., Competing Interests: The authors have declared that no competing interests exist.

Chikungunya (CHIK) has emerged as a major public health concern worldwide. Recently, atypical manifestations are drawing special attention because these might be associated with various complications. Information on atypical manifestations of CHIK is still limited. Here, we analyzed a dataset of 1,326 cases from our recent Dhaka outbreak study to explore the demographics and distributions of atypical manifestations. About 80% of cases reported at least one atypical symptom. Among all atypical symptoms, the most common and unique atypical symptom was joint pain before fever (90.2%), occurred predominantly in female respondents. Other common symptoms included red eye (68.2%), oral ulcer (37.7%), and dermatological manifestations (27.1%). More than two-thirds of patients reported multiple atypical symptoms. Atypical manifestations were not significantly different across age groups, except ocular complications. This study would be an important resource for clinicians and epidemiologists to understand the diversity of Chikungunya infection and, thus, help in better patient management.

Chikungunya (CHIK) has re-emerged as a potential neurotropic virus, with outbreaks recently being reported from many parts of India. The present study was conducted to study the spectrum and outcome of neurological complications in patients of CHIK during the 2016 outbreak in Delhi. A total of 42 cases seropositive for IgM CHIK antibodies by MAC-ELISA and developing neurological complications were enrolled. The male:female ratio was 1:2 (age range = 18-90 years). The neurological manifestations observed were encephalitis (n = 12), bulbar palsy (n = 3), acute disseminated encephalomyelitis (n = 1), cerebellitis (n = 1), myelopathy (n = 1), radiculoneuropathy (n = 3), carpal tunnel syndrome (n = 9) and tremors (n = 1). Ten patients reported worsening of pre-existing neuropathic symptoms of diabetic peripheral neuropathy (n = 4) and carpal tunnel syndrome (n = 6). One patient had aggravation of myasthenia gravis leading to respiratory failure. The majority of patients (n = 32) showed a good outcome; ten had a poor prognosis, out of which four died, all from the encephalitis group, particularly the elderly with co-morbidities.

Background: Chikungunya is a mosquito-borne virus transmitted by mosquitoes from the Aedes genus. The virus, endemic to parts of Asia and Africa, has recently undergone an emergence in other parts of the world where it was previously not found including Indian Ocean Islands, Europe, the Western Pacific and the Americas. There is no vaccine against chikungunya virus, which means that prevention and mitigation rely on personal protective measures and community level interventions including vector control., Methodology/principal Findings: A systematic review (SR) was conducted to summarize the literature on individual and community mitigation and control measures and their effectiveness. From a scoping review of the global literature on chikungunya, there were 91 articles that investigated mitigation or control strategies identified at the individual or community level. Of these, 81 were confirmed as relevant and included in this SR. The majority of the research was published since 2010 (76.5%) and was conducted in Asia (39.5%). Cross sectional studies were the most common study design (36.6%). Mitigation measures were placed into six categories: behavioural protective measures, insecticide use, public education, control of blood and blood products, biological vector control and quarantine of infected individuals. The effectiveness of various mitigation measures was rarely evaluated and outcomes were rarely quantitative, making it difficult to summarize results across studies and between mitigation strategies. Meta-analysis of the proportion of individuals engaging in various mitigation measures indicates habitat removal is the most common measure used, which may demonstrate the effectiveness of public education campaigns aimed at reducing standing water., Conclusions/significance: Further research with appropriate and consistent outcome measurements are required in order to determine which mitigation measures, or combination of mitigation measures, are the most effective at protecting against exposure to chikungunya virus., Competing Interests: The authors have declared that no competing interests exist.

Chikungunya virus (CHIKV) is a re-emerging mosquito-borne virus that displays a large cell and organ tropism, and causes a broad range of clinical symptoms in humans. It is maintained in nature through both urban and sylvatic cycles, involving mosquito vectors and human or vertebrate animal hosts. Although CHIKV was first isolated in 1953, its pathogenesis was only more extensively studied after its re-emergence in 2004. The unexpected spread of CHIKV to novel tropical and non-tropical areas, in some instances driven by newly competent vectors, evidenced the vulnerability of new territories to this infectious agent and its associated diseases. The comprehension of the exact CHIKV target cells and organs, mechanisms of pathogenesis, and spectrum of both competitive vectors and animal hosts is pivotal for the design of effective therapeutic strategies, vector control measures, and eradication actions.

Background: Hospitalization of patients during outbreaks of chikungunya virus has been reported to be uncommon (0.5-8.7%), but more frequent among infants and the elderly. CHIKV was first detected in Puerto Rico in May 2014. We enrolled patients with acute febrile illness (AFI) presenting to two hospital emergency departments in Puerto Rico and tested them for CHIKV infection to describe the frequency of detection of CHIKV-infected patients, identify risk factors for hospitalization, and describe patients with severe manifestations., Methodology/principal Findings: Serum specimens were collected from patients with AFI and tested by rRT-PCR. During May-December 2014, a total of 3,035 patients were enrolled, and 1,469 (48.4%) had CHIKV infection. A total of 157 (10.7%) CHIKV-infected patients were hospitalized, six (0.4%) were admitted to the intensive care unit, and two died (0.1%). Common symptoms among all CHIKV-infected patients were arthralgia (82.6%), lethargy (80.6%), and myalgia (80.5%). Compared to patients aged 1-69 years (7.3%), infant (67.2%) and elderly (17.3%) patients were nine and two times more likely to be hospitalized, respectively (relative risk [RR] and 95% confidence interval [CI] = 9.16 [7.05-11.90] and 2.36 [1.54-3.62]). Multiple symptoms of AFI were associated with decreased risk of hospitalization, including arthralgia (RR = 0.31 [0.23-0.41]) and myalgia (RR = 0.29 [0.22-0.39]). Respiratory symptoms were associated with increased risk of hospitalization, including rhinorrhea (RR = 1.68 [1.24-2.27) and cough (RR = 1.77 [1.31-2.39]). Manifestations present among <5% of patients but associated with patient hospitalization included cyanosis (RR = 2.20 [1.17-4.12) and seizures (RR = 3.23 [1.80-5.81)., Discussion: Among this cohort of CHIKV-infected patients, hospitalization was uncommon, admission to the ICU was infrequent, and death was rare. Risk of hospitalization was higher in patients with symptoms of respiratory illness and other manifestations that may not have been the result of CHIKV infection., Competing Interests: The authors have declared that no competing interests exist.

There is a need for more detailed elucidation of T-cell immunity in chikungunya infection. CD8 T cells are one of main actors against viruses. Here, we analysed CD8 + T lymphocytes from patients in the acute and chronic phases of chikungunya disease (CHIKD). Our results demonstrate that CD8 + T cells expressed higher ex vivo granzyme B, perforin and CD107A expression in patients in the acute phase of CHIKD compared with healthy individuals and higher ex vivo expression of CD69, interleukin-17A, interleukin-10 and CD95 ligand, and co-expression of CD95/CD95 ligand. These results elucidate the importance of these lymphocytes, demonstrating immune mechanisms mediated in human chikungunya infection., (© 2018 John Wiley & Sons Ltd.)