Chikungunya Virus (CHIKV): A Comprehensive Overview

Table of Contents

• Introduction
• CHIKV Genome, Structure, and Replication
• Epidemiology and Geographic Distribution of Chikungunya Virus (CHIKV)
• Transmission of Chikungunya Virus (CHIKV)
• Clinical Aspects of Chikungunya Virus (CHIKV)
• Pathogenesis of Chikungunya Virus (CHIKV)
• Clinical Manifestations of Chikungunya Virus (CHIKV)
• Diagnosis of Chikungunya Virus (CHIKV) Infection
• Vaccines and Treatments for Chikungunya Virus (CHIKV)
• Animal Models for Studying Chikungunya Virus (CHIKV) Infection

Introduction

• Chikungunya fever is a viral illness transmitted by Aedes mosquitoes.
• The term "chikungunya" comes from the Makonde language of Africa, meaning "to walk bent over," reflecting the severe joint pain.
• The causative agent is Chikungunya virus (CHIKV), a member of the Alphavirus genus in the Togaviridae family.
• Historically, CHIKV was confined to Africa, Asia, and the Indian subcontinent.
• In December 2013, France reported the first autochthonous cases in the Caribbean island of St. Martin, marking the beginning of CHIKV’s spread to over 43 countries in the Americas.
• CHIKV causes recurring epidemics, particularly in tropical and subtropical regions.
• Transmission occurs mainly through Aedes aegypti and Aedes albopictus mosquitoes.
• Symptoms of Chikungunya fever (CHIKF) include high fever, rash, myalgia, headache, and severe polyarthralgia (joint pain).
• The intense joint pain can persist for weeks, months, or years, leading to significant economic and social impact.
• Although CHIKF is typically non-lethal, severe cases can result in multiple organ failure and death, with mortality rates ranging from 0.024% to 0.7%.
• There is currently no vaccine or effective antiviral treatment for CHIKF.
• Research on CHIKV is challenging due to the lack of reliable experimental models that fully replicate the human disease.
• Understanding CHIKV's epidemiology, replication, and pathogenesis is crucial for advancing prevention and treatment efforts.

CHIKV genome, structure and replication

Chikungunya virus (CHIKV) Structure:

• CHIKV is an enveloped plus-strand RNA virus with icosahedral symmetry.
• The virion has a diameter of 70 nm and consists of:
• Repeating units of E1 and E2 transmembrane glycoproteins (240 heterodimers of E2/E1 arranged as trimeric spikes on its surface).
• A capsid (C).
• A host-derived lipid bilayer.
• A single molecule of genome RNA.

Chikungunya virus (CHIKV) Genome:

• The genome is approximately 12 kb in length and encodes:
• Nonstructural proteins (nsPs) at the 5' end.
• Structural proteins at the 3' end.
• Nonstructural proteins are translated from the genomic RNA.
• Structural proteins are translated from a subgenomic RNA.

Chikungunya virus (CHIKV) Replication:

• Alphaviruses enter target cells via endocytosis.
• Receptors involved include DC-SIGN, L-SIGN, heparin sulfate, laminin, and integrins, though their exact roles are not fully established.
• Prohibitin has recently been identified as a CHIKV receptor protein.
• The acidic environment of the endosome triggers conformational changes in the viral envelope, exposing the E1 peptide which mediates virus-host cell membrane fusion.
• This fusion allows delivery of the core and release of the viral genome into the cytoplasm.
• Two precursors of nonstructural proteins are translated from the viral mRNA.
• Cleavage of these precursors produces nsP1-nsP4.
• Nonstructural proteins form the viral replication complex, which synthesizes a full-length negative-strand RNA intermediate.
• This negative-strand RNA serves as a template for the synthesis of subgenomic (26S) and genomic (49S) RNAs.
• The subgenomic RNA drives the expression of the C-pE2-6K-E1 polyprotein precursor.
• This precursor is processed by autoproteolysis to release the capsid and generate pE2 and E1 glycoproteins.
• PreE2 and E1 associate in the Golgi and are exported to the plasma membrane.
• pE2 is cleaved into E2 and E3.
• The viral nucleocapsid binds to viral RNA, and membrane-associated envelope glycoproteins are recruited for viral assembly.
• The assembled alphavirus particle, with an icosahedral core, buds from the cell membrane.

Epidemiology and geographic distribution of Chikungunya virus (CHIKV):

• First Cases: Initial cases of a chikungunya-like illness recorded in 1823 in Zanzibar, Africa, followed by similar reports in St. Thomas Island, Caribbean, in 1827 and 1828.
• 1952: CHIKV isolated, identified, and characterized as an arbovirus in Tanzania.
• 2004 Onward: CHIKV spread to approximately 60 countries globally, causing significant outbreaks.
• 2005-2006 Outbreak: Major outbreak on La Réunion Island in the Indian Ocean, infecting over one-third of the population (260,000 people) with 284 deaths. The Aedes albopictus mosquito, a new vector, contributed to virus spread.
• 2007: First autochthonous outbreak in Europe reported in Italy.
• 2013: First CHIKV outbreak in the Americas, starting in Saint Martin with 658 confirmed cases.
• Global Impact: Local transmission identified in around 45 countries and territories in the Americas, resulting in over 3 million confirmed cases.

Genotypes:

• Asian
• East Indian Ocean (IOL)
• West African (WA)
• East/Central/South African (ECSA)

Geographic Distribution:

• Africa: Includes West Africa, Central Africa, and other regions like Democratic Republic of Congo, Nigeria, Angola, and more.
• Asia: Includes Burma, Thailand, Cambodia, Vietnam, India, Sri Lanka, Timor, Indonesia, and the Philippines.
• Indian Ocean Region: Significant outbreaks in Kenya, Comoros, Seychelles, Mauritius, and La Réunion.
• Europe: Autochthonous outbreaks in Italy, re-importation cases from La Réunion, and ongoing transmission.
• Recent Outbreaks: Significant cases in Republic of Congo (2011), India (2012), Cambodia, Papua New Guinea, and the Philippines (2012-2013).
• Situation update: In July 2024, approximately 350 000 CHIKVD cases and over 140 deaths have been reported worldwide. A total of 21 countries have reported CHIKVD cases from the Americas (13), Asia (6), Africa (1) and Europe (1).

Transmission of Chikungunya Virus (CHIKV)

Transmission Cycles:

Sylvatic Cycle:

• Mechanism: Maintained in forested areas between Aedes mosquitoes (e.g., A. furcifer, A. vittatus) and non-human primates.
• Human Involvement: Humans occasionally infected when they live near or visit forested areas where enzootic transmission occurs.
• Characteristics: Results in sporadic human cases and small outbreaks.

Urban Cycle:

• Mechanism: Involves transmission between infected and non-infected individuals facilitated by urban mosquitoes (A. aegypti and A. albopictus).
• Vectors:
• Aedes aegypti: Prefers to feed on humans, lays eggs in artificial containers, and rests inside houses.
• Aedes albopictus: Both zoophilic and anthropophilic, aggressive, active throughout the day, and has a longer lifespan.
• Mutation: The E1 mutation in the ECSA lineage strain enhances CHIKV infectivity and transmission, particularly by A. albopictus.
• Characteristics: Sustained transmission in urban settings leads to larger outbreaks and higher levels of human exposure.

Transmission Pathways:

• Vertical Transmission: CHIKV can be transmitted from mother to fetus during pregnancy. Outcomes vary from asymptomatic to severe conditions such as myocarditis and meningoencephalitis. Risks are higher if the mother is viremic at childbirth.
• Blood Transfusion and Transplants: CHIKV can be transmitted through blood transfusions or organ transplants. This has been observed during outbreaks in various regions, including La Réunion and Puerto Rico. Prevalence of CHIKV RNA in blood donations can range from 0.4% to 2.1% during epidemics.
• Enzootic Cycle:
• Vectors: Arboreal mosquitoes in Africa, like A. furcifer.
• Reservoirs: Non-human primates serve as principal reservoirs.
• Spillover: Virus may spill over to humans living nearby.
• Urban Cycle:
• Vectors: Primarily A. aegypti and A. albopictus.
• Reservoirs: Humans serve as the primary reservoirs during epidemic periods. In non-epidemic times, other animals like monkeys, rodents, and birds can maintain virus circulation.

Clinical Aspects of Chikungunya Virus (CHIKV)

Incubation and Viremia:

• Incubation Period: CHIKV incubation ranges from 1 to 12 days in humans.
• Viremia Levels: Viremia can reach up to 3.3 x 10^9 copies/ml during the first week, significantly higher than levels seen with other arboviruses like DENV and ZIKV.

Symptoms and Disease Spectrum:

• Common Symptoms: High fever, headache, myalgia, and severe polyarthralgia. Most infected individuals show symptoms, with less than 15% being asymptomatic.
• Serological Findings: Possible findings include lymphopenia, moderate thrombocytopenia, elevated levels of ALT, AST, creatinine, and creatinine kinase. Some cases may also involve calcium deficiency related to bone absorption.

Chronic Phase:

• Duration: Acute symptoms typically resolve within 1-2 weeks, but 30–40% of cases may develop chronic arthralgia lasting for months or years.
• Dermatological Manifestations: Skin rashes, facial edema, and oral mucosa bleeding affect 40–50% of infected individuals.

Severe and Atypical Cases:

• Incidence: Severe manifestations, including respiratory issues, hypertension, and cardiac problems, are more common in older adults.
• Newborns: High risk for severe outcomes such as pain, prostration, fever, thrombocytopenia, encephalopathy, and intracranial bleeding.

Neurological Complications:

• Prevalent Symptoms: Include abnormal mental status, headache, focal deficits, seizures, meningoencephalitis, meningoencephalomyeloradiculitis, and papillitis.
• Mortality: Severe neurological involvement can be fatal, particularly with the ECSA lineage of the virus.

Pathogenesis of Chikungunya Virus (CHIKV)

Initial Infection:

• Viral Entry: CHIKV enters the body through subcutaneous capillaries, infecting macrophages, fibroblasts, and endothelial cells. It then spreads to secondary lymphoid organs and the bloodstream, targeting tissues like the liver, muscles, joints, and brain.

Immune Response:

• Systemic Response: CHIKV infection triggers a strong innate immune response, including the production of antiviral IFN-α and pro-inflammatory cytokines. The adaptive response shifts from CD8+ T cells to CD4+ T cells and anti-inflammatory cytokines later in the acute phase.

Immune Response and Viral Load:

• Cytokines and Chemokines: Elevated levels of IL-16, IL-17, MCP-1, IP-10, MIP-1, and IL-6 during acute infection indicate a significant inflammatory response.
• Antibodies: Specific IgM appears within the first week and lasts for 3–4 months, while IgG can persist for over 6 months. The role of these antibodies in chronic arthralgia is not fully understood.

Cellular Entry and Replication:

• Receptors and Entry: CHIKV binds to receptors such as prohibitin (PHB), TIM-1, and DC-SIGN. It enters cells through clathrin-mediated endocytosis or micropinocytosis and can use apoptotic blebs for cell-to-cell transmission.
• Immune Response: The virus targets synovial fibroblasts and osteoblasts, leading to joint pathology and chronic arthritis. The resulting proinflammatory response correlates with disease severity and chronic symptoms.

Clinical Manifestations of Chikungunya Virus (CHIKV)

Initial Presentation:

• Abrupt Onset: CHIKV infection typically starts with a sudden febrile illness.
• Common Symptoms: Includes polyarthralgia and maculopapular rash.
• Incubation Period: Generally lasts 2 to 4 days, but can range from 1 to 12 days. Asymptomatic infections occur in 5-15% of cases.

Symptoms:

• Arthralgia: Affects 78.4% of patients, usually bilateral and symmetrical. Commonly involves ankles, knees, hands, wrists, feet, shoulders, and elbows.
• Rash: Present in 54% of patients, primarily on the trunk and arms.
• Periarticular Edema: Noted in 45% of patients, especially around the ankles.
• Myalgia and Headache: Affect 72% and 63% of patients, respectively.
• Hemorrhagic Signs: Gingivorrhagia and epistaxis occur in 10.6% of patients.

Disease Progression and Duration:

• Acute Phase: Signs and symptoms generally resolve within 2 weeks.
• Chronic Phase: Arthralgia may persist for weeks, months, or years. This distinguishes CHIKV from dengue virus infection.
• Gastrointestinal Symptoms: Less common in CHIKV compared to dengue, with a higher proportion of myalgia and arthralgia.

Severe Cases:

• Risk Factors: Severe forms are more common in older patients and those with comorbidities such as cardiovascular, neurological, respiratory disorders, or diabetes.
• Severe Manifestations: Can include encephalopathy, encephalitis, myocarditis, hepatitis, and multiorgan failure. These rare cases can be fatal and are more likely in patients with underlying medical conditions.
• Neonatal Risk: Neonates born to viremic mothers can have a high infection rate (up to 50%), leading to severe disease and neurological complications.

Prognosis and Long-Term Effects:

• General Prognosis: Typically good, but older patients (<45 years) may develop chronic rheumatic musculoskeletal pain.
• Chronic Pain: Severe initial rheumatic involvement correlates with a higher likelihood of chronic pain. High titers of CHIKV-specific IgG may be associated with long-lasting arthralgia.
• Mechanistic Hypothesis: Chronic pain may result from an imbalance in B cell expansion and differentiation, influenced by IL-6 secretion and viral persistence.

Diagnosis of Chikungunya Virus (CHIKV) Infection

Laboratory Diagnosis:

1. Virus Isolation:

• Specimen Collection: Can be performed on mosquitoes or acute serum specimens collected within 8 days of symptom onset.
• Transport and Processing: Serum should be transported cold (2°C to 8°C or on dry ice) and processed as soon as possible, ideally within 48 hours.
• Methods: Inoculate serum into a susceptible cell line or suckling mouse in a reference laboratory.

2. Reverse Transcriptase-Polymerase Chain Reaction (RT-PCR):

• Assays: Several RT-PCR assays are available to detect CHIKV RNA. Real-time, closed-system assays are preferred for their higher sensitivity and lower contamination risk.
• Recommendations: Use CHIKV RT-PCR protocols from the Centers for Disease Control and Prevention (CDC) and the Institut Pasteur.
• Specimens: Serum from whole blood is used for RT-PCR testing and virus isolation.

3. Serological Diagnosis:

• Specimen Handling: Serum obtained from whole blood should be transported at 2°C to 8°C and should not be frozen.
• ELISA Testing: Enzyme-linked immunosorbent assay (ELISA) is used to detect IgM antibodies specific to CHIKV or a four-fold rise in IgG titer between acute and convalescent samples.
• Sensitivity: Techniques using the complete virus as antigen are preferred over those using recombinant proteins. In-house IgM/IgG ELISA with purified viral antigen following CDC protocols is recommended.
• Timing: The second serum sample for serological testing should be collected 1 to 2 weeks after the first sample. Seroconversion can be detected as a four-fold increase in IgG between acute and convalescent-phase samples.

Additional Considerations:

• Rapid Tests: Generally not recommended due to lower sensitivity and specificity compared to well-established ELISA and RT-PCR methods.
• Follow-Up: A confirmatory second sample may be necessary for accurate diagnosis, especially if the initial tests are inconclusive.

Vaccines and Treatments for Chikungunya Virus (CHIKV)

Current Status of CHIKV Vaccines

Vaccine Development and Challenges

Currently, there are no widely available vaccines for Chikungunya virus (CHIKV). The need for an effective CHIKV vaccine is pressing due to the virus's endemic status in many regions. A viable vaccine should be economical, easy to handle, and suitable for mass distribution, particularly in developing countries.

Several vaccines are in various stages of development:

• Preclinical Vaccines: These include whole-virus inactivated vaccines, VEE/CHIKV chimeric vaccines, recombinant adenovirus vectored vaccines, DNA-based vaccines, virus-like particle (VLP) vaccines, and live-attenuated vaccines. Some promising candidates are currently undergoing preclinical trials.
• mRNA Vaccines: Leveraging the success of mRNA vaccines for SARS-CoV-2, pharmaceutical companies are exploring mRNA-based vaccines for CHIKV. These vaccines show promise but face challenges regarding their cost, accessibility, and the need for intravenous administration, which limits their use in resource-limited settings.

Specific Vaccine Candidates

1. TSI-GSD-218

• Type: Live attenuated vaccine
• Strain: CHIK strain 15561 from the 1962 Thailand outbreak
• Status: Phase II trial
• Results: Induced high levels of neutralizing antibodies in 98% of vaccinees, with 85% maintaining seropositivity one year after immunization. The vaccine showed safety but has not progressed beyond phase II trials and is considered abandoned.

2. VLA1553

• Type: Live attenuated vaccine
• Strain: CHIKVLR2006-OPY1, genetically modified to enhance safety
• Developer: Valneva
• Status: Phase I completed, Phase III underway in Brazil
• Results: Showed high titers of neutralizing antibodies after a single dose, with 100% seroconversion rate. Safe with common mild side effects.

3. PXVX0317

• Type: Virus-like particle (VLP) vaccine
• Developer: PaxVax, in collaboration with the National Institutes of Health
• Status: Phase III trial
• Results: Evaluates safety and immunogenicity with a two-dose regimen. Common side effects include nausea, injection site pain, and fatigue, with no serious adverse effects reported.

4. ChAdOx1-Chik

• Type: Recombinant chimpanzee adenovirus-vectored vaccine
• Developer: Jenner Institute, University of Oxford
• Status: Phase I completed
• Results: Induced broadly neutralizing antibodies against all CHIKV lineages. Safe with no serious adverse reactions, although adenovirus vectors have raised concerns due to rare but serious adverse effects seen in other vaccines.

5. MV-CHIKV

• Type: Measles-vectored vaccine
• Developer: Themis Bioscience
• Status: Phase II trial
• Results: Promising safety and immunogenicity with high seroconversion rates. Uses a platform with a long history of safety in over 1 billion children.

6. BBV87

• Type: Inactivated whole virus vaccine
• Strain: East/Central/South African genotype
• Developer: Bharat Biotech International Ltd.
• Status: Phase II and III trials in Costa Rica
• Results: Induces high levels of neutralizing antibodies with satisfactory immune responses in Phase I. Phase II and III trials will further evaluate efficacy and safety.

Treatment Options

• Anti-inflammatory Drugs: Currently, the treatment for CHIKV infection is symptomatic, primarily using anti-inflammatory drugs to alleviate symptoms such as joint pain and fever. Some disease-modifying antirheumatic drugs (DMARDs) like methotrexate and hydroxychloroquine have been tested but with limited success.
• Antiviral Research: Various antiviral drugs have been investigated for their effectiveness against CHIKV, but none are currently in clinical use. Research continues to explore potential antivirals and repurposed drugs to inhibit CHIKV replication.

Animal Models for Studying Chikungunya Virus (CHIKV) Infection

Understanding CHIKV infection requires diverse animal models to accurately replicate the virus's impact and to aid in the development of treatments and vaccines. 

Rodent Models

1. Wild-Type Rodents

C57BL/6J Mice:

• Age Variations: Studies use mice from newborns to 48 weeks old.
• Virus Dose Variations: CHIKV inoculation ranges from 10² to 10⁸ plaque-forming units (pfu)/ml.
• Observations: Models display diverse outcomes depending on age and virus dose:
• Young Mice (14-day-old): Develop gross swelling and severe tenosynovitis when inoculated in the foot.
• 3-Week-Old Mice: Maintain high levels of viral RNA for up to 98 days post-infection.
• 8-Week-Old Mice: Replicate virus particles only until 9 days post-infection.
• 18-Month-Old Mice: Show prolonged viremia and severe joint pathology.
• Resistance Variation: Older mice may show increased resistance or different disease progression compared to younger ones.

Historic Models:

• Ross’s 1950s Study: Observed lethality in 6-day-old Albino Swiss mice after intracerebral virus administration.

2. Genetically Modified Rodent Models

Single-Gene Knockout Mice:

• C1q−/− or FcRγ−/− Mice: Used to study CHIKV-induced arthritis.
• ISG15−/− Mice: Exhibit increased susceptibility to CHIKV infection.
• Double Knockout Mice (UbE1L−/− and ISG15−/−): Show increased cytokines and lethality.
• Sting-deficient Mice (Stinggt/gt): Exhibit increased immune cells in muscle and synovial cavities.
• IFN-α/βR−/− Mice: More susceptible to CHIKV infections despite preserved immune responses.

Special Strains:

• Rag1 KO Mice: Exhibit persistent virus in joint-associated tissues.
• Irf3−/− Irf7−/− dKO Mice: Develop disease independent of Irf3, Irf7, and IFNAR1 pathways.
• TLR and IFNAR Knockouts: Studies on TLR3−/−, TLR3/7/9−/−, TLR9−/−, and IFNAR−/− mice show varying roles of innate immune responses.

Non-Human Primate (NHP) Models

1. Rhesus Macaques (Macaca mulatta)

• Early Studies (1960s): Showed neutralizing antibodies and clinical symptoms after infection.
• Infection Studies:
• Viremia Duration: Detected for at least 6 days with peak levels 1-2 days post-infection.
• Clinical Symptoms: Include fever and rash, with a peak immune response around 21 days post-infection.
• Aging Effects: Older macaques showed significant differences in viremia and immune responses compared to younger ones, highlighting immune senescence.

2. Other Macaque Species

• Bonnet Macaques (Macaca radiata) and Cynomolgus Macaques (Macaca fascicularis): Used to study age-related effects and pregnancy outcomes.
• Pregnancy Studies: Infected pregnant rhesus macaques showed similar viremia dynamics but no detectable virus in fetal tissues or placenta.

3. Vaccine and Immunotherapy Testing:

• Vaccine Candidates: All CHIKV vaccine technologies tested in NHP models, including live-attenuated and virus-like particle vaccines, with results showing similar endpoints to human trials.
• Monoclonal Antibodies: Tested in rhesus macaques, demonstrating protection and reduced viremia.

Prevention of CHIKV Infection

1. Vector Control:

• Mosquito Control: Since there is no vaccine available, controlling mosquito populations remains the most effective preventive measure. This involves:
• Eliminating Breeding Sites: Regularly remove or destroy mosquito breeding sites such as stagnant water in containers, tires, and other receptacles.
• Treating Water Sources: Use larvicides in water sources that cannot be removed.
• Cleaning and Maintenance: Frequently clean and empty containers that hold water.

2. Personal Protection:

• Clothing: Wear long-sleeved shirts and long pants to reduce skin exposure to mosquitoes.
• Repellents: Apply repellents to exposed skin and clothing. Effective repellents include:
• DEET (N,N-diethyl-3-methylbenzamide)
• IR3535 (3-[N-acetyl-N-butyl]-aminopropionic acid ethyl ester)
• Icaridin (1-piperidinecarboxylic acid, 2-(2-hydroxyethyl)-1-methylpropylester)
• Indoor Protection: Use mosquito coils or insecticide vaporizers to reduce mosquito bites indoors.

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