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Neurovirulence comparison of chikungunya virus isolates of the Asian and East/Central/South African genotypes from Malaysia

Wei Chiam C, Fun Chan Y, Chai Ong K, Thong Wong K, Sam IC

J Gen Virol, 2015 Nov;96(11):3243-3254.
PMID: 26276497 DOI: 10.1099/jgv.0.000263

Chikungunya virus (CHIKV), an alphavirus of the family Togaviridae, causes fever, polyarthritis and rash. There are three genotypes: West African, Asian and East/Central/South African (ECSA). The latter two genotypes have caused global outbreaks in recent years. Recent ECSA CHIKV outbreaks have been associated with severe neurological disease, but it is not known if different CHIKV genotypes are associated with different neurovirulence. In this study, the neurovirulence of Asian (MY/06/37348) and ECSA (MY/08/065) strains of CHIKV isolated in Malaysia were compared. Intracerebral inoculation of either virus into suckling mice was followed by virus titration, histopathology and gene expression analysis of the harvested brains. Both strains of CHIKV replicated similarly, yet mice infected with MY/06/37348 showed higher mortality. Histopathology findings showed that both CHIKV strains spread within the brain (where CHIKV antigen was localized to astrocytes and neurons) and beyond to skeletal muscle. In MY/06/37348-infected mice, apoptosis, which is associated with neurovirulence in alphaviruses, was observed earlier in brains. Comparison of gene expression showed that a pro-apoptotic gene (eIF2αK2) was upregulated at higher levels in MY/06/37348-infected mice, while genes involved in anti-apoptosis (BIRC3), antiviral responses and central nervous system protection (including CD40, IL-10RA, MyD88 and PYCARD) were upregulated more highly in MY/08/065-infected mice. In conclusion, the higher mortality observed following MY/06/37348 infection in mice is due not to higher viral replication in the brain, but to differentially expressed genes involved in host immune responses. These findings may help to identify therapeutic strategies and biomarkers for neurological CHIKV infections.

Matched MeSH terms: Chikungunya virus/genetics
Fulltext Outbreak of chikungunya due to virus of Central/East African genotype in Malaysia

Noridah O, Paranthaman V, Nayar SK, Masliza M, Ranjit K, Norizah I, et al.

Med J Malaysia, 2007 Oct;62(4):323-8.
PMID: 18551938 MyJurnal

Chikungunya is an acute febrile illness caused by an alphavirus which is transmitted by infective Aedes mosquitoes. Two previous outbreaks of chikungunya in Malaysia were due to chikungunya virus of Asian genotype. The present outbreak involved two adjoining areas in the suburb of Ipoh city within the Kinta district of Perak, a state in the northern part of Peninsular Malaysia. Thirty seven residents in the main outbreak area and two patients in the secondary area were laboratory confirmed to be infected with the virus. The index case was a 44-year Indian man who visited Paramakudi, Tamil Naidu, India on 21st November 2006 and returned home on 30th of November 2006, and subsequently developed high fever and joint pain on the 3rd of December 2006. A number of chikungunya virus isolates were isolated from both patients and Aedes albopictus mosquitoes in the affected areas. Molecular study showed that the chikungunya virus causing the Kinta outbreak was of the Central/East African genotype which occurred for the first time in Malaysia.

Matched MeSH terms: Chikungunya virus/genetics*
Fulltext Rapid detection of chikungunya virus in laboratory infected Aedes aegypti by Reverse-Transcriptase- Polymerase Chain Reaction (RT-PCR)

Rohani A, Yulfi H, Zamree I, Lee HL

Trop Biomed, 2005 Dec;22(2):149-54.
PMID: 16883281 MyJurnal

A study of chikungunya virus was carried out to establish Reverse Transcriptase- Polymerase Chain Reaction (RT-PCR) as a rapid detection technique of the virus. The susceptibility of lab-colonized Aedes aegypti to chikungunya virus was also determined. Artificial membrane feeding technique was used to orally feed the mosquitoes with a human isolate of chikungunya virus. A total of 100 fully engorged female Ae. aegypti were obtained and maintained for 7 days. Seventy of them survived and then pooled at 10 individuals per pool. Total RNA was extracted from the samples and RT-PCR amplifications were carried out. Five out of 7 pools showed positive PCR band at 350-bp, indicating Ae. aegypti is a potential vector of chikungunya virus. The minimum infection rate (MIR) was 71% within these laboratory colonies. RT-PCR is a sensitive technique that is useful in detecting infected mosquitoes in epidemic areas. This technique can de used as a rapid detection method and provide an early virologic surveillance systems of chikungunya virus infected mosquitoes.

Matched MeSH terms: Chikungunya virus/genetics
Fulltext Re-emergence of Chikungunya virus in Malaysia

Kumarasamy V, Prathapa S, Zuridah H, Chem YK, Norizah I, Chua KB

Med J Malaysia, 2006 Jun;61(2):221-5.
PMID: 16898316 MyJurnal

An outbreak of Chikugunya (CHIK) fever occurred among the fishing community in Bagan Pancor, Perak. The outbreak was laboratory confirmed within 48 hours after the receipt of the specimens. Fifty-three patients' serum samples were submitted for laboratory investigation and 47 (88.7%) were confirmed to be positive for CHIK infection by RT-PCR, and/or virus isolation, and/or in-house immunoflourescent test. RT-PCR and virus isolation were the tests of choice for patients with illness of four days or less and detection of CHIK specific IgM for those with more than four days of fever. The nucleic acid sequence based on the 354- and 294-bp of the nsP1 and E1 genes of the CHIK virus detected from pools of adults Aedes aegypti mosquitoes were identical to those CHIKV virus isolated from humans in the same locality. Phylogenetic analysis of the CHIK virus based on the 257 nts partial E1 gene indicates that Bagan Panchor's strain was closely related to the first CHIK virus isolated during the outbreak in Klang in 1998.

Matched MeSH terms: Chikungunya virus/genetics
Re-emergence of Chikungunya virus in South-east Asia: virological evidence from Sri Lanka and Singapore

Hapuarachchi HC, Bandara KB, Sumanadasa SD, Hapugoda MD, Lai YL, Lee KS, et al.

J Gen Virol, 2010 Apr;91(Pt 4):1067-76.
PMID: 19955565 DOI: 10.1099/vir.0.015743-0

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.

Matched MeSH terms: Chikungunya virus/genetics
Real-time polymerase chain reaction for diagnosis and quantitation of negative strand of chikungunya virus

Chiam CW, Chan YF, Loong SK, Yong SS, Hooi PS, Sam IC

Diagn Microbiol Infect Dis, 2013 Oct;77(2):133-7.
PMID: 23886793 DOI: 10.1016/j.diagmicrobio.2013.06.018

Quantitative real-time polymerase chain reaction (qRT-PCR) is useful for diagnosis and studying virus replication. We developed positive- and negative-strand qRT-PCR assays to detect nsP3 of chikungunya virus (CHIKV), a positive-strand RNA alphavirus that causes epidemic fever, rash, and arthritis. The positive- and negative-strand qRT-PCR assays had limits of quantification of 1 and 3 log10 RNA copies/reaction, respectively. Compared to a published E1 diagnostic assay using 30 laboratory-confirmed clinical samples, the positive-strand nsP3 qRT-PCR assay had higher R(2) and efficiency and detected more positive samples. Peak viral load of 12.9 log(10) RNA copies/mL was reached on day 2 of illness, and RNA was detectable up to day 9, even in the presence of anti-CHIKV IgM. There was no correlation between viral load and persistent arthralgia. The positive-strand nsP3 assay is suitable for diagnosis, while the negative-strand nsP3 assay, which uses tagged primers to increase specificity, is useful for study of active viral replication kinetics.

Matched MeSH terms: Chikungunya virus/genetics
Fulltext Reemergence of endemic Chikungunya, Malaysia

AbuBakar S, Sam IC, Wong PF, MatRahim N, Hooi PS, Roslan N

Emerg Infect Dis, 2007 Jan;13(1):147-9.
PMID: 17370532

Chikungunya virus infection recently reemerged in Malaysia after 7 years of nondetection. Genomic sequences of recovered isolates were highly similar to those of Malaysian isolates from the 1998 outbreak. The reemergence of the infection is not part of the epidemics in other Indian Ocean countries but raises the possibility that chikungunya virus is endemic in Malaysia.

Matched MeSH terms: Chikungunya virus/genetics
Fulltext Refractoriness of Aedes aegypti (Linnaeus) to dual infection with dengue and chikungunya virus

Rohani A, Potiwat R, Zamree I, Lee HL

Southeast Asian J. Trop. Med. Public Health, 2009 May;40(3):443-8.
PMID: 19842428

In this study, artificial membrane feeding technique was used to orally feed Aedes aegypti with dengue and chikungunya viruses. Virus detection was carried out by reverse transcriptase polymerase chain reaction. The study did not detect dual infection of Ae. aegypti with dengue and chikungunya virus from the same pool or from individual mosquitoes. Oral receptivity of Ae. aegypti to chikungunya virus was higher than that of dengue virus.

Matched MeSH terms: Chikungunya virus/genetics
Fulltext Seroprevalence survey of Chikungunya virus in Bagan Panchor, Malaysia

Ayu SM, Lai LR, Chan YF, Hatim A, Hairi NN, Ayob A, et al.

Am J Trop Med Hyg, 2010 Dec;83(6):1245-8.
PMID: 21118929 DOI: 10.4269/ajtmh.2010.10-0279

In 2006, an outbreak of Chikungunya virus (CHIKV) of the Asian genotype affected over 200 people in Bagan Panchor village in Malaysia. One year later, a post-outbreak survey was performed to determine attack rate, asymptomatic rate, and post-infection sequelae. Findings were compared with recent CHIKV outbreaks of the Central/East African genotype. A total of 180 residents were interviewed for acute symptoms and post-infection physical quality of life and depressive symptoms. Sera from 72 residents were tested for CHIKV neutralizing antibodies. The estimated attack rate was 55.6%, and 17.5% of infected residents were asymptomatic. Arthralgia was reported up to 3 months after infection, but there were no reports of long-term functional dependence or depression. Symptomatic and seropositive residents were significantly more likely to live in the area with the most dense housing and commercial activities. CHIKV had a high attack rate and considerable clinical impact during the Bagan Panchor outbreak.

Matched MeSH terms: Chikungunya virus/genetics
Fulltext The assessment of risk factors for the Central/East African Genotype of chikungunya virus infections in the state of Kelantan: a case control study in Malaysia

Yusoff AF, Mustafa AN, Husaain HM, Hamzah WM, Yusof AM, Harun R, et al.

BMC Infect Dis, 2013 May 08;13:211.
PMID: 23656634 DOI: 10.1186/1471-2334-13-211

BACKGROUND: The aims of the study were to assess the risk factors in relation to cross border activities, exposure to mosquito bite and preventive measures taken.An outbreak of chikungunya virus (CHIKV) infection in Malaysia has been reported in Klang, Selangor (1998) and Bagan Panchor, Perak (2006). In 2009, CHIKV infection re-emerged in some states in Malaysia. It raises the possibilities that re-emergence is part of the epidemics in neighbouring countries or the disease is endemic in Malaysia. For this reason, A community-based case control study was carried out in the state of Kelantan.
METHODS: Prospective case finding was performed from June to December 2009. Those who presented with signs and symptoms of CHIKV infection were investigated. We designed a case control study to assess the risk factors. Assessment consisted of answering questions, undergoing a medical examination, and being tested for the presence of IgM antibodies to CHIKV. Descriptive epidemiological studies were conducted by reviewing both the national surveillance and laboratory data. Multivariable logistic regression analysis was performed to determine risk factors contributing to the illness. Cases were determined by positive to RT-PCR or serological for antibodies by IgM. CHIKV specificity was confirmed by DNA sequencing.
RESULTS: There were 129 suspected cases and 176 controls. Among suspected cases, 54.4% were diagnosed to have CHIKV infection. Among the controls, 30.1% were found to be positive to serology for antibodies [IgM, 14.2% and IgG, 15.9%]. For analytic study and based on laboratory case definition, 95 were considered as cases and 123 as controls. Those who were positive to IgG were excluded. CHIKV infection affected all ages and mostly between 50-59 years old. Staying together in the same house with infected patients and working as rubber tappers were at a higher risk of infection. The usage of Mosquito coil insecticide had shown to be a significant protective factor. Most cases were treated as outpatient, only 7.5% needed hospitalization. The CHIKV infection was attributable to central/east African genotype CHIKV.
CONCLUSIONS: In this study, cross border activity was not a significant risk factor although Thailand and Malaysia shared the same CHIKV genotype during the episode of infections.

Matched MeSH terms: Chikungunya virus/genetics*
Updates on chikungunya epidemiology, clinical disease, and diagnostics

Sam IC, Kümmerer BM, Chan YF, Roques P, Drosten C, AbuBakar S

Vector Borne Zoonotic Dis, 2015 Apr;15(4):223-30.
PMID: 25897809 DOI: 10.1089/vbz.2014.1680

Chikungunya virus (CHIKV) is an Aedes-borne alphavirus, historically found in Africa and Asia, where it caused sporadic outbreaks. In 2004, CHIKV reemerged in East Africa and spread globally to cause epidemics, including, for the first time, autochthonous transmission in Europe, the Middle East, and Oceania. The epidemic strains were of the East/Central/South African genotype. Strains of the Asian genotype of CHIKV continued to cause outbreaks in Asia and spread to Oceania and, in 2013, to the Americas. Acute disease, mainly comprising fever, rash, and arthralgia, was previously regarded as self-limiting; however, there is growing evidence of severe but rare manifestations, such as neurological disease. Furthermore, CHIKV appears to cause a significant burden of long-term morbidity due to persistent arthralgia. Diagnostic assays have advanced greatly in recent years, although there remains a need for simple, accurate, and affordable tests for the developing countries where CHIKV is most prevalent. This review focuses on recent important work on the epidemiology, clinical disease and diagnostics of CHIKV.

Matched MeSH terms: Chikungunya virus/genetics