Evolution Of Chikungunya Virus Research Articles

Importation of a novel Indian Ocean lineage carrying E1-K211E and E2-V264A of Chikungunya Virus in Zhejiang Province, China, in 2019.

The chikungunya virus (CHIKV) is widespread. In Zhejiang province, China, CHIKV infection is often associated with travelers from tropical and subtropical countries. In the present study, three CHIKV isolates from serum samples of travelers in Zhejiang province in 2019 were sequenced, and phylogenetically analyzed to study their molecular characteristics. Sequence analysis showed that the non-structural protein and the structural protein had 37 and 28 amino acid mutations, respectively; no mutation site was found at the E1-A226 residue, which could increase the adaptability of CHIKV to Aedes albopictus. All three samples carried two mutations, namely, E1-K211E and E2-V264A, which were introduced to Bangladesh around late 2015 and Thailand in early 2017. Phylogenetic analysis revealed that these three CHIKVs were Indian Ocean lineage of the East Africa/Central/South Africa genotype (ECSA) and that the MF773566 strain from Bangladesh (Australia/Bangladesh 2017) had the closest evolutionary relationship. The three CHICKs imported into Zhejiang province in 2019 belonged to the ECSA genotype and had multiple amino acid variation sites. The variation in the three samples provides a certain reference for the subsequent research on CHIKV evolution.

The evolution of chikungunya virus circulating in Indonesia: Sequence analysis of the orf2 gene encoding the viral structural proteins.

Chikungunya virus (CHIKV) is an arthropod-borne virus that has caused several major epidemics globally, including in Indonesia. Although significant progress has been achieved in understanding the epidemiology and genotype circulation of CHIKV in Indonesia, the evolution of Indonesian CHIKV isolates is poorly understood. Thus, our study aimed to perform phylogenetic and mutation analyses of the orf2 gene encoding its viral structural protein to improve our understanding of CHIKV evolution in Indonesia. Complete orf2 gene sequences encoding the viral structural proteins of Indonesian-derived CHIKV were downloaded from GenBank until August 31, 2022. Various bioinformatics tools were employed to perform phylogenetic and mutation analyses of the orf2 gene. We identified 76 complete sequences of orf2 gene of CHIKV isolates originally derived from Indonesia. Maximum likelihood trees demonstrated that the majority (69/76, 90.8%) of Indonesian-derived CHIKV isolates belonged to the Asian genotype, while seven isolates (9.2%) belonged to the East/Central/South African (ECSA)genotype. The Indonesian-derived CHIKV isolates were calculated to be originated in Indonesia around 95 years ago (1927), with 95% highest posterior density (HPD) ranging from 1910 to 1942 and a nucleotide substitution rate of 5.07 × 10-4 (95% HPD: 3.59 × 10-4 to 6.67 × 10-4). Various synonymous and non-synonymous substitutions were identified in the C, E3, E2, 6K, and E1 genes. Most importantly, the E1-A226V mutation, which has been reported to increase viral adaptation in Aedes albopictus mosquitoes, was present in all ECSA isolates. To our knowledge, our study is the first comprehensive research analyzing the mutation and evolution of Indonesian-derived CHIKV based on complete sequences of the orf2 genes encoding its viral structural proteins. Our results clearly showed a dynamic evolution of CHIKV circulating in Indonesia.

The origin and continuing adaptive evolution of chikungunya virus.

Chikungunya virus (CHIKV) is the responsible agent of chikungunya fever, a debilitating arthritic disease in humans. CHIKV is endemic in Africa and Asia, although transmission cycles are considerably different on these continents. Before 2004, CHIKV had received little attention, since it was only known to cause localised outbreaks in a limited region with no fatalities. However, the recent global reemergence of CHIKV has caused serious global health problems and shown its potential to become a significant viral threat in the future. Unexpectedly, the reemergence is more rapid and is geographically more extensive, especially due to increased intensity of global travel systems or failure to contain mosquito populations. Another important factor is the successful adaptation of CHIKV to a new vector, the Aedes albopictus mosquito. Ae. albopictus survives in both temperate and tropical climates, thus facilitating CHIKV expansion to non-endemic regions. The continuous spread and transmission of CHIKV pose challenges for the development of effective vaccines and specific antiviral therapies. In this review, we discuss the biology and origin of CHIKV in Africa as well as its subsequent expansion to other parts of the world. We also review the transmission cycle of CHIKV and its continuing adaptation to its mosquito vectors and vertebrate hosts. More-complete understanding of the continuous evolution of CHIKV may help in predicting the emergence of CHIKV strains with possibly greater transmission efficiency in the future.

Source-tracking of the Chinese Chikungunya viruses suggests that Indian subcontinent and Southeast Asia act as major hubs for the recent global spread of Chikungunya virus

BackgroundChikungunya fever, caused by the Chikungunya virus (CHIKV), has become a major global health concern, causing unexpected large outbreaks in Africa, Asia, Europe, and the Americas. CHIKV is not indigenous to China, and its origin in the country is poorly understood. In particular, there is limited understanding of the recent global spread of CHIKV in the context of the CHIKV epidemic.MethodsHere we investigated a novel Chikungunya patient who came from Myanmar to China in August, 2019. Direct genome sequencing was performed via combined MinION sequencing and BGISEQ-500 sequencing. A complete CHIKV genome dataset, including 727 CHIKV genomes retrieved from GenBank and the genome sequenced in this study, was constructed. An updated and comprehensive phylogenetic analysis was conducted to understand the virus’s origin, evolution, transmission routes and genetic adaptation.ResultsAll globally distributed CHIKV genomes were divided into West Africa, East/Central/South African and Asian genotypes. The genome sequenced in this study was located in the Indian Ocean lineage, and was closely related to a strain isolated from an Australian patient who returned from Bangladesh in 2017. A comprehensive phylogenetic analysis showed that the Chinese strains mainly originated from the Indian subcontinent and Southeast Asia. Further analyses indicated that the Indian subcontinent and Southeast Asia may act as major hubs for the recent global spread of CHIKV, leading to multiple outbreaks and epidemics. Moreover, we identified 179 distinct sites, including some undescribed sites in the structural and non-structural proteins, which exhibited apparent genetic variations associated with different CHIKV lineages.ConclusionsHere we report a novel CHIKV isolate from a chikungunya patient who came from Myanmar to China in 2019, and summarize the source and evolution of Chinese CHIKV strains. Our present findings provide a better understanding of the recent global evolution of CHIKV, highlighting the urgent need for strengthened surveillance against viral diversity.

Molecular characterization of chikungunya virus isolates from two localized outbreaks during 2014-2019 in Kerala, India.

After the 2005-2009 chikungunya epidemic, intermittent outbreaks were reported in many parts of India. The outbreaks were caused by either locally circulating strains or imported viruses. Virus transmission routes can be traced by complete genome sequencing studies. We investigated two outbreaks in 2014 and 2019 in Kerala, India. Chikungunya virus (CHIKV) was isolated from the samples, and whole genomes were sequenced for a 2014 isolate and a 2019 isolate. Phylogenetic analysis revealed that the isolates formed a separate group with a 2019 isolate from Pune, Maharashtra, and belonged to the East/Central/South African (ECSA) genotype, Indian subcontinent sublineage of the Indian Ocean Lineage (IOL). A novel mutation at amino acid position 76 of the E2 gene was observed in the group. The phylogenetic results suggest that the outbreaks might have been caused by a virus that had been circulating in India since 2014. A detailed study is needed to investigate the evolution of CHIKV in India.

Phylogenetic and sequencing analysis of Chikungunya virus strains from local Pakistani population.

To outline the genotype of Chikungunya virus strains and to form their phylogenetic tree. The cross-sectional study was conducted at the Armed Forces Institute of Pathology, Rawalpindi, Pakistan, from August 2016 to September 2017, and comprised suspected Chikungunya samples. The viral ribonucleic acid was extracted and amplified. The purified product was sequenced using Sanger dideoxy method. Phylogenetic tree was constructed. Sequences generated were checked for alignment with sequences reported globally. Structural prediction and analysis was performed using Phyre2 and the models were visualised in PyMol2.2. In the 11 suspected samples, 7(63.6%) were found to be positive for Chikungunya. Of them, 5(71.4%) sequences generated were found to be aligned with other full structural polyprotein sequences, having 99% similarity with amino acid sequences. The Chikungunya strains in the study belonged to the East/Central/South African genotype.

Current Status of Chikungunya in India.

Chikungunya fever (CHIKF) is an arbovirus disease caused by chikungunya virus (CHIKV), an alphavirus of Togaviridae family. Transmission follows a human-mosquito-human cycle starting with a mosquito bite. Subsequently, symptoms develop after 2–6 days of incubation, including high fever and severe arthralgia. The disease is self-limiting and usually resolve within 2 weeks. However, chronic disease can last up to several years with persistent polyarthralgia. Overlapping symptoms and common vector with dengue and malaria present many challenges for diagnosis and treatment of this disease. CHIKF was reported in India in 1963 for the first time. After a period of quiescence lasting up to 32 years, CHIKV re-emerged in India in 2005. Currently, every part of the country has become endemic for the disease with outbreaks resulting in huge economic and productivity losses. Several mutations have been identified in circulating strains of the virus resulting in better adaptations or increased fitness in the vector(s), effective transmission, and disease severity. CHIKV evolution has been a significant driver of epidemics in India, hence, the need to focus on proper surveillance, and implementation of prevention and control measure in the country. Presently, there are no licensed vaccines or antivirals available; however, India has initiated several efforts in this direction including traditional medicines. In this review, we present the current status of CHIKF in India.

Molecular Epidemiology, Evolution and Reemergence of Chikungunya Virus in South Asia

Chikungunya virus (CHIKV) is a vector (mosquito)-transmitted alphavirus (family Togaviridae). CHIKV can cause fever and febrile illness associated with severe arthralgia and rash. Genotypic and phylogenetic analysis are important to understand the spread of CHIKV during epidemics and the diversity of circulating strains for the prediction of effective control measures. Molecular epidemiologic analysis of CHIKV is necessary to understand the complex interaction of vectors, hosts and environment that influences the genotypic evolution of epidemic strains. In this study, different works published during 1950s to 2020 concerning CHIKV evolution, epidemiology, vectors, phylogeny, and clinical outcomes were analyzed. Outbreaks of CHIKV have been reported from Bangladesh, Bhutan, India, Pakistan, Sri Lanka, Nepal, and Maldives in South Asia during 2007–2020. Three lineages- Asian, East/Central/South African (ECSA), and Indian Ocean Lineage (IOL) are circulating in South Asia. Lineage, ECSA and IOL became predominant over Asian lineage in South Asian countries during 2011–2020 epidemics. Further, the mutant E1-A226V is circulating in abundance with Aedes albopictus in India, Bangladesh, Nepal, and Bhutan. CHIKV is underestimated as clinical symptoms of CHIKV infection merges with the symptoms of dengue fever in South Asia. Failure to inhibit vector mediated transmission and predict epidemics of CHIKV increase the risk of larger global epidemics in future. To understand geographical spread of CHIKV, most of the studies focused on CHIKV outbreak, biology, pathogenesis, infection, transmission, and treatment. This updated study will reveal the collective epidemiology, evolution and phylogenies of CHIKV, supporting the necessity to investigate the circulating strains and vectors in South Asia.

Insight into the origin of chikungunya virus in Malaysian non-human primates via sequence analysis

Chikungunya virus (CHIKV) is maintained in the sylvatic cycle in West Africa and is transmitted by Aedes mosquito species to monkeys. In 2006, four verified CHIKV isolates were obtained during a survey of arboviruses in monkeys (Macaca fascicularis) in Pahang state, Peninsular Malaysia. RNA was extracted from the CHIKV isolates and used in reverse transcription polymerase chain reactions (RT-PCR) to amplify PCR fragments for sequencing. Nucleic acid primers were designed to generate overlapping PCR fragments that covered the whole viral sequence. A total of 11,238 base pairs (bp) corresponding to open reading frames (ORFs) from our isolates and 47 other registered isolates in the National Center for Biotechnology Information (NCBI) were used to elucidate sequences, amino acids, and phylogenetic relationships and to estimate divergence times by using MEGA 7.0 and the Bayesian Markov chain Monte Carlo method. Phylogenetic analysis revealed that all CHIKV isolates could be classified into the Asian genotype and clustered with Bagan Panchor clades, which are associated with the chikungunya outbreak reported in 2006, with sequence and amino acid similarities of 99.9% and 99.7%, respectively. Minor amino acid differences were found between human and non-human primate isolates. Amino acid analysis showed a unique amino acid at position 221 in the nsP1region, at which a glycine (G) was found only in monkey isolates, whereas arginine (R) was found at the same position only in human isolates. The time to the most recent common ancestor (MRCA) estimation indicated that CHIKV probably started to diverge from human to non-human primates in approximately 2004 in Malaysia. The results suggested that CHIKV in non-human primates probably resulted from the spillover of the virus from humans. The study will be helpful in understanding the movement and evolution of CHIKV in Malaysia and globally.

Evolution of Chikungunya virus in mosquito cells

It has been observed that replication of Chikungunya virus (CHIKV) in C6/36 Aedes albopictus cells has little effect on virus evolution. To characterize evolutionary patterns associated with CHIKV replication in mosquito cells, we performed serial passages of the LR2006 strain in Ae. albopictus cells (75 and 30 passages in C6/36 and U4.4 respectively) and Ae. aegypti cells (100 passages in AA-A20 and in AE) and studied genotypic changes accompanying adaptation during this evolutionary process. Quantitative analysis revealed cell specific patterns: low mutation rates in C6/36 cells except when a CHIKV strain pre-adapted to mammalian was used and typical features of adaptation to cell culture conditions with a high number of fixed mutations in AE and AA-A20 cells probably due to the weak permissiveness of these latter cell lines. Altogether, these results suggested that both cell line and viral strain influence rates of viral evolution. In contrast, characteristics and distribution of mutations were qualitatively very similar in all mosquito cells with a high level of parallel evolution including 4 deletion mutations. Serial passage in mammalian cells of viruses pre-adapted to mosquito cells revealed disappearance of almost all shared mutations suggesting that many of these mutational patterns are vector-specific.

Differing epidemiological dynamics of Chikungunya virus in the Americas during the 2014-2015 epidemic.

Chikungunya virus (CHIKV) has been detected sporadically since the 1950s and includes three distinct co-circulating genotypes. In late 2013, the Asian genotype of CHIKV was responsible for the Caribbean outbreak (CO) that rapidly became an epidemic throughout the Americas. There is a limited understanding of the molecular evolution of CHIKV in the Americas during this epidemic. We sequenced 185 complete CHIKV genomes collected mainly from Nicaragua in Central America and Florida in the United States during the 2014–2015 Caribbean/Americas epidemic. Our comprehensive phylogenetic analyses estimated the epidemic history of the Asian genotype and the recent Caribbean outbreak (CO) clade, revealed considerable genetic diversity within the CO clade, and described different epidemiological dynamics of CHIKV in the Americas. Specifically, we identified multiple introductions in both Nicaragua and Florida, with rapid local spread of viruses in Nicaragua but limited autochthonous transmission in Florida in the US. Our phylogenetic analysis also showed phylogeographic clustering of the CO clade. In addition, we identified the significant amino acid substitutions that were observed across the entire Asian genotype during its evolution and examined amino acid changes that were specific to the CO clade. Deep sequencing analysis identified specific minor variants present in clinical specimens below-consensus levels. Finally, we investigated the association between viral phylogeny and geographic/clinical metadata in Nicaragua. To date, this study represents the largest single collection of CHIKV complete genomes during the Caribbean/Americas epidemic and significantly expands our understanding of the emergence and evolution of CHIKV CO clade in the Americas.

Chikungunya virus evolution following a large 3'UTR deletion results in host-specific molecular changes in protein-coding regions.

The 3′untranslated region (UTR) in alphavirus genomes functions in virus replication and plays a role in determining virus host range. However, the molecular evolution of virus UTRs is understudied compared to the evolution of protein-coding regions. Chikungunya virus (CHIKV) has the longest 3′UTR among the alphaviruses (500–700 nt), and 3′UTR length and sequence structure vary substantially among different CHIKV lineages. Previous studies showed that genomic deletions and insertions are key drivers of CHIKV 3′UTR evolution. Inspired by hypothesized deletion events in the evolutionary history of CHIKV, we used experimental evolution to examine CHIKV adaptation in response to a large 3′UTR deletion. We engineered a CHIKV mutant with a 258 nt deletion in the 3′UTR (ΔDR1/2). This deletion reduced viral replication on mosquito cells, but did not reduce replication on mammalian cells. To examine how selective pressures from vertebrate and invertebrate hosts shape CHIKV evolution after a deletion in the 3′UTR, we passaged ΔDR1/2 virus populations strictly on primate cells, strictly on mosquito cells, or with alternating primate/mosquito cell passages. We found that virus populations passaged on a single host cell line increased in fitness relative to the ancestral deletion mutant on their selected host, and viruses that were alternately passaged improved on both hosts. Surprisingly, whole genome sequencing revealed few changes in the 3′UTR of passaged populations. Rather, virus populations evolved improved fitness through mutations in protein coding regions that were associated with specific hosts.

Extensive evolution analysis of the global chikungunya virus strains revealed the origination of CHIKV epidemics in Pakistan in 2016.

Chikungunya virus (CHIKV) is a mosquito-borne virus that causes epidemics widely in the world especially in the tropical and subtropical regions. Phylogenetic analysis has found that the CHIKV lineages were associated with the spatial and temporal distributions, which were related to the virus adaption to the major mosquito species and their distributions. In this study, we reported the complete genome sequences of eight CHIKV isolates from the outbreak in Pakistan last year. Then we reviewed the evolutionary history using extensive phylogenetic analysis, analyzed lineage-specific substitutions in viral proteins, and characterized the spreading pathway of CHIKV strains including the Pakistani strains. The results showed that the Pakistani stains belonged to the ECSA.IOL sub-lineage and derived from India. The genetic properties of the Pakistani strains including the adaptive substitution to vectors were further characterized, and the potential risks from the occurrence of CHIKV infection in Pakistan were discussed. These results provided better understanding of CHIKV evolution and transmission in the world and revealed the possible origination of the CHIKV outbreak and epidemic in Pakistan, which would promote the disease prevention and control in the identified countries and territories with the history of CHIKV infections as well as new regions with potential risk of CHIKV outbreaks.

Chikungunya virus-vector interactions.

Chikungunya virus (CHIKV) is a mosquito-borne alphavirus that causes chikungunya fever, a severe, debilitating disease that often produces chronic arthralgia. Since 2004, CHIKV has emerged in Africa, Indian Ocean islands, Asia, Europe, and the Americas, causing millions of human infections. Central to understanding CHIKV emergence is knowledge of the natural ecology of transmission and vector infection dynamics. This review presents current understanding of CHIKV infection dynamics in mosquito vectors and its relationship to human disease emergence. The following topics are reviewed: CHIKV infection and vector life history traits including transmission cycles, genetic origins, distribution, emergence and spread, dispersal, vector competence, vector immunity and microbial interactions, and co-infection by CHIKV and other arboviruses. The genetics of vector susceptibility and host range changes, population heterogeneity and selection for the fittest viral genomes, dual host cycling and its impact on CHIKV adaptation, viral bottlenecks and intrahost diversity, and adaptive constraints on CHIKV evolution are also discussed. The potential for CHIKV re-emergence and expansion into new areas and prospects for prevention via vector control are also briefly reviewed.

Genetic characterization of E2 region of Chikungunya virus circulating in Odisha, Eastern India from 2010 to 2011

Chikungunya virus (CHIKV) infection has caught attention yet again as it rages around the globe affecting millions of people. The virus caused epidemic outbreaks affecting more than 15,000 people in Odisha, Eastern India since 2010. In this study, complete genetic characterization of E2 gene of CHIKV circulating in Odisha from 2010 to 2011 was performed by virus isolation, RT-PCR, molecular phylogenetics and bioinformatics methods. Phylogenetic analyses revealed the circulation of Indian Ocean Lineage (IOL) strains of ECSA genotype of CHIKV in Odisha. Several mutations were detected in the E2 gene, viz. E2-R82G, E2-L210Q, E2-I211T, E2-V229I and E2-S375T which had various adaptive roles during the evolution of CHIKV. The CHIKV E2 peptide 57KTDDSHD63 was predicted to be the most probable T-cell epitope and peptide 84FVRTSAPCT92 predicted to be the common T and B cell epitope having high antigenicity. The amino acid positions 356–379 and 365–385 were predicted to be transmembrane helical domains and indicated E2 protein anchorage in intracellular membranes for effective interaction with the host receptors. Positive selection pressure was observed in five specific sites, 210, 211, 318, 375, and 377 which were observed to be fixed advantageously in most viral isolates. Structural modeling revealed that E2 gene of CHIKV was composed of 3 domains and the major adaptive mutations were detected in domain B, which can modulate binding of CHIKV to host cells, while the transmembrane domain in domain C and the epitopes were located in domain A, which was found to be most conserved. This is the first report from Eastern India demonstrating a predictive approach to the genetic variations, epitopic regions and the transmembrane helices of the E2 region. The results of this study, combined with other published observations, will expand our knowledge about the E2 region of CHIKV which can be exploited to develop control measures against CHIKV.

Sequential Adaptive Mutations Enhance Efficient Vector Switching by Chikungunya Virus and Its Epidemic Emergence

The adaptation of Chikungunya virus (CHIKV) to a new vector, the Aedes albopictus mosquito, is a major factor contributing to its ongoing re-emergence in a series of large-scale epidemics of arthritic disease in many parts of the world since 2004. Although the initial step of CHIKV adaptation to A. albopictus was determined to involve an A226V amino acid substitution in the E1 envelope glycoprotein that first arose in 2005, little attention has been paid to subsequent CHIKV evolution after this adaptive mutation was convergently selected in several geographic locations. To determine whether selection of second-step adaptive mutations in CHIKV or other arthropod-borne viruses occurs in nature, we tested the effect of an additional envelope glycoprotein amino acid change identified in Kerala, India in 2009. This substitution, E2-L210Q, caused a significant increase in the ability of CHIKV to develop a disseminated infection in A. albopictus, but had no effect on CHIKV fitness in the alternative mosquito vector, A. aegypti, or in vertebrate cell lines. Using infectious viruses or virus-like replicon particles expressing the E2-210Q and E2-210L residues, we determined that E2-L210Q acts primarily at the level of infection of A. albopictus midgut epithelial cells. In addition, we observed that the initial adaptive substitution, E1-A226V, had a significantly stronger effect on CHIKV fitness in A. albopictus than E2-L210Q, thus explaining the observed time differences required for selective sweeps of these mutations in nature. These results indicate that the continuous CHIKV circulation in an A. albopictus-human cycle since 2005 has resulted in the selection of an additional, second-step mutation that may facilitate even more efficient virus circulation and persistence in endemic areas, further increasing the risk of more severe and expanded CHIK epidemics.

Molecular genome tracking of East, Central and South African genotype of Chikungunya virus in South–east Asia between 2006 and 2009

Objective To understand the epidemiology of the East, Central and South African (ECSA) genotype of Chikungunya virus (CHIKV) in terms of emerging and re-emerging infections, this study has been aimed at investigating the evolutionary parameters, genomic signatures and molecular tracking of the CHIKV ECSA genotype in South-east Asia and coastal areas of the Indian Ocean between 2006 and 2009 by using phylogenetic analysis and the Bayesian Markov Chain Monte Carlo (BMCMC) evolutionary estimation. Methods Nearly complete genome sequences of 53 CHIKV isolates from all genotypes were subjected to phylogenetic analysis and evolutionary parameter estimation. The amino acids of 67 of ECSA genotype during 2006 to 2009 were compared for finding molecular signature tracking. The ECSA genotype signatures were visualized to find the possible transmission root was projected onto a geographic map. Results Phylogenetic analysis showed the ECSA genotype was divided into 2 groups. The first group comprises viruses from India and Southeast Asian countries. The second group consists of strains typically circulating in Sri Lanka in 2008. The evolutionary parameters of these groups depicted the time of the most recent common ancestor at approximately 7.5 years ago. The genomic signatures revealed the positions of amino acid variation in each group. Conclusions The molecular evolution projected onto a geographical map showed the routes of CHIKV transmission from 2006 to 2009. Molecular tracking will assist in understanding transmission routes, epidemiology and molecular evolution of CHIKV.

HLA Class I Restriction as a Possible Driving Force for Chikungunya Evolution

After two decades of quiescence, epidemic resurgence of Chikungunya fever (CHIKF) was reported in Africa, several islands in the Indian Ocean, South-East Asia and the Pacific causing unprecedented morbidity with some cases of fatality. Early phylogenetic analyses based on partial sequences of Chikungunya virus (CHIKV) have led to speculation that the virus behind recent epidemics may result in greater pathogenicity. To understand the reasons for these new epidemics, we first performed extensive analyses of existing CHIKV sequences from its introduction in 1952 to 2009. Our results revealed the existence of a continuous genotypic lineage, suggesting selective pressure is active in CHIKV evolution. We further showed that CHIKV is undergoing mild positive selection, and that site-specific mutations may be driven by cell-mediated immune pressure, with occasional changes that resulted in the loss of human leukocyte antigen (HLA) class I-restricting elements. These findings provide a basis to understand Chikungunya virus evolution and reveal the power of post-genomic analyses to understand CHIKV and other viral epidemiology. Such an approach is useful for studying the impact of host immunity on pathogen evolution, and may help identify appropriate antigens suitable for subunit vaccine formulations.

Re-emergence of Chikungunya virus in South-east Asia: virological evidence from Sri Lanka and Singapore

Chikungunya fever swept across many South and South-east Asian countries, following extensive outbreaks in the Indian Ocean Islands in 2005. However, molecular epidemiological data to explain the recent spread and evolution of Chikungunya virus (CHIKV) in the Asian region are still limited. This study describes the genetic Characteristics and evolutionary relationships of CHIKV strains that emerged in Sri Lanka and Singapore during 2006-2008. The viruses isolated in Singapore also included those imported from the Maldives (n=1), India (n=2) and Malaysia (n=31). All analysed strains belonged to the East, Central and South African (ECSA) lineage and were evolutionarily more related to Indian than to Indian Ocean Islands strains. Unique genetic characteristics revealed five genetically distinct subpopulations of CHIKV in Sri Lanka and Singapore, which were likely to have emerged through multiple, independent introductions. The evolutionary network based on E1 gene sequences indicated the acquisition of an alanine to valine 226 substitution (E1-A226V) by virus strains of the Indian sublineage as a key evolutionary event that contributed to the transmission and spatial distribution of CHIKV in the region. The E1-A226V substitution was found in 95.7 % (133/139) of analysed isolates in 2008, highlighting the widespread establishment of mutated CHIKV strains in Sri Lanka, Singapore and Malaysia. As the E1-A226V substitution is known to enhance the transmissibility of CHIKV by Aedes albopictus mosquitoes, this observation has important implications for the design of vector control strategies to fight the virus in regions at risk of chikungunya fever.

A226V mutation in virus during the 2007 chikungunya outbreak in Kerala, India

Kerala State in India was gripped by a renewed and widespread outbreak of Chikungunya virus (CHIKV) infection during 2007. Here, we report the A226V mutation in the glycoprotein envelope 1 (E1) gene of the virus among isolates collected from the three worst-affected districts of the state during this outbreak. This mutation had already been suggested to be directly responsible for a significant increase in CHIKV infectivity in Aedes albopictus. The badly affected districts in Kerala State during 2007 have abundant rubber plantations, which supported prolific breeding of Ae. albopictus mosquitoes. The abundance of Ae. albopictus in the region and molecular evolution of CHIKV may be contributing factors for the renewed epidemic of chikungunya fever during 2007.