Chikungunya is a re-emerging mosquito-borne viral infection that has spread from East Africa to Indian Ocean islands and re-emerged in India since 2004. In Malaysia, chikungunya re-emerged after a hiatus of seven years, causing a localised outbreak in a north-western coastal town in 2006 and subsequently widespread outbreaks in 2008. Since the first local outbreak of chikungunya in Singapore in January 2008, chikungunya infections have been increasingly reported in Singapore. In this case series, five patients aged 37-62 years, with chikungunya infection confirmed in August 2008, were reported. Three of the five were male, and only one had medical comorbidities. Two had a travel history to Johor, Malaysia, where local outbreaks of chikungunya had been reported. Fever, arthralgia and rash were the most common symptoms. Fever lasted four to five days while viraemia lasted four to 11 days, persisting two to three days after defervescence in three patients. A biphasic pattern of fever was observed in two patients. Leucopenia was noted in all patients, while mild thrombocytopenia and transaminitis occurred in three of five patients. Two patients had persistent polyarthralgia at two to three weeks after the onset of symptoms. Fever, arthralgia and rash should prompt consideration of acute chikungunya in Singapore. While taking the travel history, doctors should be mindful that indigenous chikungunya cases can occur.
In this study, we isolated a virus strain (YN12031) from specimens of Armigeres subalbatus collected in the China-Laos border. BHK-21 cells infected with YN12031 exhibited an evident cytopathic effect (CPE) 32 h post-infection. The virus particles were spherical, 70 nm in diameter, and enveloped; they also featured surface fibers. Molecular genetic analysis revealed that YN12031 was closely related to alpha viruses such as Chikungunya virus and Sindbis virus, and located in the same clade as MM2021, the prototype of Getahvirus (GETV) isolated in Malaysia in 1955. Phylogenetic analysis of the E2 and capsid genes further revealed that YN12031 was located in the same clade as the Russian isolate LEIV/16275/Mag. Analysis of the homology of nucleotides and amino acids in the coding area and E2 gene demonstrated that the YN12031 isolated from the China-Laos border (tropical region) was related closest to the LEIV/16275/Mag isolate obtained in Russia (North frigid zone area) among other isolates studied. These results suggest that GETV can adapt to different geographical environments to propagate and evolve. Thus, strengthening the detection and monitoring of GETV and its related diseases is very crucial.
Getah virus (GETV) was first isolated in Malaysia in 1955. Since then, epidemics in horses and pigs caused by GETV have resulted in huge economic losses. At present, GETV has spread across Eurasia and Southeast Asia, including mainland China, Korea, Japan, Mongolia, and Russia. Data show that the Most Recent Common Ancestor (MRCA) of GETV existed about 145years ago (95% HPD: 75-244) and gradually evolved into four distinct evolutionary populations: Groups I-IV. The MRCA of GETVs in Group III, which includes all GETVs isolated from mosquitoes, pigs, horses, and other animals since the 1960s (from latitude 19°N to 60°N), existed about 51years ago (95% HPD: 51-72). Group III is responsible for most viral epidemics among domestic animals. An analysis of the GETV E2 protein sequence and structure revealed seven common amino acid mutation sites. These sites are responsible for the structural and electrostatic differences detected between widespread Group III isolates and the prototype strain MM2021. These differences may account for the recent geographical radiation of the virus. Considering the economic significance of GETV infection in pigs and horses, we recommend the implementation of strict viral screening and monitoring programs.
In the last four years, Malaysia has had three outbreaks of chikungunya virus infection. The first two occurred in Perak in 2006.The third began in Johor in early 2008. The genome of the viruses suggests that on each occasion a different virus was introduced into the population. The first outbreak in Bagan Panchor was due to an Asian genotype virus. The second in the Kinta district of Perak in late 2006 was due to a Central/East African genotype virus. Contact tracing was even able to discover the patient who was the source of the virus from the Indian subcontinent. The third outbreak in Johor was also of a Central/East African strain of virus, but introduced independently. The epidemiology of that outbreak is described in this issue of the MJM.
Recovery from chikungunya is previously considered universal and mortality due to the virus is rare and unusual. Findings from recent chikungunya outbreaks occurred in Reunion Island and India have since challenged the conventional view on the benign nature of the illness. Malaysia has experienced at least of 4 outbreaks of chikungunya since 1998. In the present on-going large outbreak due to chikungunya virus of Central/East African genotype, a previous healthy sixty six years gentleman without co-morbidity was noted to have severe systemic infection by the virus and involvement of his liver. He subsequently passed away due to cardiovascular collapse after 5 days of illness.
Getah virus (GETV), a mosquito-borne alphavirus, is an emerging animal pathogen causing outbreaks among racehorses and pigs. Early detection of the GETV infection is essential for timely implementation of disease prevention and control interventions. Thus, a rapid and accurate nucleic acid detection method for GETV is highly needed. Here, two TaqMan minor groove binding (MGB) probe-based quantitative reverse transcription-polymerase chain reaction (qRT-PCR) assays were developed. The qRT-PCR primers and TaqMan MGB probe were designed based on the conserved region of nsP1 and nsP2 genes of 23 GETV genome sequences retrieved from GenBank. Only the qRT-PCR assay using nsP2-specific primers and probe detected all two Malaysia GETV strains (MM2021 and B254) without cross-reacting with other closely related arboviruses. The qRT-PCR assay detected as few as 10 copies of GETV RNA, but its detection limit at the 95% probability level was 63.25 GETV genome copies (probit analysis, P ≤ 0.05). Further validation of the qRT-PCR assay using 16 spiked simulated clinical specimens showed 100% for both sensitivity and specificity. In conclusion, the qRT-PCR assay developed in this study is useful for rapid, sensitive and specific detection and quantification of GETV.
In the last few years, chikungunya has become a major problem in Southeast Asia, with large numbers of cases being reported in Singapore, Malaysia, and Thailand. Much of the current epidemic of chikungunya in Southeast Asia is being driven by the emergence of a strain of chikungunya virus that originated in Africa and spread to islands in the Indian Ocean, as well as to India and Sri Lanka, and then onwards to Southeast Asia. There is currently no specific treatment for chikungunya and no vaccine is available for this disease. This review seeks to provide a short update on the reemergence of chikungunya in Southeast Asia and the prospects for control of this disease.
Chikungunya virus, a mosquito-borne alphavirus, is endemic in Africa and Southeast Asia but is rarely reported in Taiwan. We report the case of a Taiwanese woman who developed Chikungunya fever, which was first diagnosed by a clinician rather than by fever screening at an airport. The woman presented with fever, maculopapular rash, and arthralgia, the triad for the disease, on the day she returned home after a trip to Malaysia. These symptoms are very similar to those of dengue fever, which is endemic in Southern Taiwan. Chikungunya infection was confirmed by reverse transcriptase-polymerase chain reaction and seroconversion on paired serum specimens. For approximately 40 years until 2006, no cases of Chikungunya fever had been found in Taiwan. Clinicians in Taiwan should consider Chikungunya fever as a possible diagnosis for a febrile patient with arthralgia, rash, and a history of travel to an endemic area, such as Africa or Southeast Asia.
In 2008, an outbreak of chikungunya infection occurred in Johor. We performed a retrospective review of all laboratory confirmed adult chikungunya cases admitted to Hospital Sultanah Aminah, Johor Bahru from April to August 2008, looking into clinical and laboratory features. A total of 18 laboratory confirmed cases of chikungunya were identified with patients presenting with fever, joint pain, rash and vomiting. Haemorrhagic signs were not seen. Lymphopenia, neutropenia, thrombocytopenia, raised liver enzymes and deranged coagulation profile were the prominent laboratory findings. We hope this study can help guide physician making a diagnosis of chikungunya against other arborviruses infection.
Chikungunya infections were detected in Singapore among returning travelers who had visited friends and relatives (VFR) in India and Malaysia. These sporadic imported cases occurred over a year before the 2008 chikungunya outbreaks in Singapore, demonstrating the potential for introducing this emerging viral infection into new areas via VFR travel.
An adult Malaysian woman returned to Japan from Kuala Lumpur and had onset of dengue fever-like symptoms including high fever, malaise and arthritis in early January 2009. Serum obtained on the following day was tested at the National Institute of Infectious Diseases in Tokyo, where it was determined to be positive for chikungunya virus (CHIKV) RNA. IgM antibody against CHIKV was negative on January 6 and sero-converted to be positive on January 14, confirming a recent CHIKV infection. Except for arthralgia, all her symptoms resolved uneventfully within 10 days.
Many countries neighboring Malaysia have reported human infections by chikungunya virus, a mosquito-borne togavirus belonging to the genus Alphavirus. However, although there is serological evidence of its presence in Malaysia, chikungunya virus has not been known to be associated with clinical illness in the country. An outbreak of chikungunya virus occurred in Klang, Malaysia, between December 1998 and February 1999. The majority of the cases were in adults and the clinical presentation was similar to classical chikungunya infections. Malaysia is heavily dependent on migrant workers from countries where chikungunya is endemic. It is speculated that the virus has been re-introduced into the country through the movement of these workers.
Getah virus (GETV), which was first isolated in Malaysia in 1955, and Sagiyama virus (SAGV), isolated in Japan in 1956, are members of the genus Alphavirus in the family Togaviridae. It is a consensus view that SAGV is a variant of GETV. In the present study, we determined the complete sequences of the prototype GETV MM2021 and SAGV M6-Mag132 genomic RNA extracted from plaque-purified viruses. The MM2021 genome was 11,692 nucleotides (nt) in length in the absence of 3' poly(A) tail, and the length of M6-Mag132 genome was 11,698 nt. Through sequence alignment of MM2021 and M6-Mag132, we located all the amino acid differences between these two strains, which were scattered in all the encoded proteins. Subsequently, we validated the close evolutionary relationship between GETV and SAGV by constructing phylogenetic trees based on either complete genomes or structural genomes. We eventually analyzed the growth kinetics of GETV and SAGV as well as other representative alphaviruses in various mammalian and insect cell lines. It was shown that human-oriented cell lines such as HEK-293T and Hela cells were relatively resistant to GETV and SAGV infection due to absence of proviral factors or species-specific barrier. On the other hand, both GETV and SAGV replicated efficiently in non-human cell lines. Our results provide essential genetic information for future epidemiological surveillance on Alphaviruses and lay the foundation for developing effective interventions against GETV and SAGV.
Sagiyama virus (SAG) is a member of the genus Alphavirus in the family Togaviridae, isolated in Japan from mosquitoes in 1956. We determined the complete nucleotide sequence of the SAG genomic RNA from the original stock virus which formed a mixture of plaques with different sizes, and that from a full-length cDNA clone, pSAG2, infectious RNA transcripts from which formed uniform large plaques on BHK-21 cells. The SAG genome was 11698 nt in length exclusive of the 3' poly(A) tail. Between the complete nucleotide sequences of the full-length cDNA clone, pSAG2, and the consensus sequence from the original stock virus, there were nine amino acid differences; two each in nsP1, nsP2 and E1, and three in E2, some of which may be responsible for plaque phenotypic variants in the original virus stock. SAG was most closely related to Ross River virus among other alphaviruses fully sequenced, with amino acid sequence identities of 86% in the nonstructural proteins and of 83% in the structural proteins. The 3' terminal 280 nt region of SAG was 82% identical to that of Barmah Forest virus, which was otherwise not closely related to SAG. Comparison of the nucleotide sequence of SAG with partial nucleotide sequences of Getah virus (GET), which was originally isolated in Malaysia in 1955 and is closely related to SAG in serology and in biology, showed near identity between the two viruses, suggesting that SAG is a strain of GET.
The identification of new virus strains is important for the study of infectious disease, but current (or existing) molecular biology methods are limited since the target sequence must be known to design genome-specific PCR primers. Thus, we developed a new method for the discovery of unknown viruses based on the cDNA--random amplified polymorphic DNA (cDNA-RAPD) technique. Getah virus, belonging to the family Togaviridae in the genus Alphavirus, is a mosquito-borne enveloped RNA virus that was identified using the Virus-Discovery-cDNA RAPD (VIDISCR) method.
Chikungunya virus (CHIKV) is an emerging mosquito-borne alphavirus that has caused multiple unprecedented and re-emerging outbreaks in both tropical and temperate countries. Despite ongoing research efforts, the underlying factors involved in facilitating CHIKV replication during early infection remains ill-characterized. The present study serves to identify host proteins modulated in response to early CHIKV infection using a proteomics approach.
This is a retrospective cross-sectional study based on the database of clusters of patients with clinical diagnosis of chikungunya (CHIK) that were referred to the National Public Health Laboratory for diagnostic investigations from January 2006 to December 2009. Of the 13,759 referred patients, a total of 6314 (45.9%) patients were laboratory confirmed to have CHIK and 7445 (54.1) patients were considered as clinical cases of CHIK by epidemiological link. Epidemic curves plotted using date of onset of illness for all referred clusters of cases showed that there were three unrelated outbreaks of CHIK in Malaysia from 2006 to 2009. There were two small outbreaks that occurred within the state of Perak in 2006. The cluster of cases in 2008 and 2009 were of related outbreak which started in Johor state and subsequently spread to various parts of Malaysia. The mean age of the patients was 37.0 years old and those patients in the laboratory confirmed group were significantly younger than those in the epidemiological linked group. The main presenting clinical features recorded in this study were fever, arthralgia, myalgia and rashes. Those patients in the laboratory confirmed group had a significant higher incidence of fever, arthralgia and rash than those in the epidemiological linked group.
During an outbreak of chikungunya in a dengue hyperendemic area within the Kinta district of Perak, two patients with acute febrile illness were laboratory confirmed to have co-infection of both dengue and chikungunya viruses in their blood. The concomitant presence of two types of viruses transmitted by the same vector in a susceptible population contributed to the resultant event. A good understanding of virus vector ecology in association with population dynamics and wider application of improved laboratory techniques by using different cell-lines suited for optimal replication of each type of virus and the correct utilization of powerful molecular techniques will enhance accurate diagnosis of these infectious diseases.
Many triggering factors for onset of emerging infectious diseases are now recognised, such as: globalisation, demographic increase, population movements, international trade, urbanisation, forest destruction, climate changes, loss in biodiversity, and extreme life conditions such as poverty, famine and war. Epidemic burden is often leading to disasters, in terms of human losses, as well as economic, political or social consequences. These outbreaks may jeopardize within a few weeks or months, industry, trade, or tourism. While dengue and its most severe forms (hemorrhagic and shock syndrome) is spreading all over the tropical world, another arbovirosis, chikungunya disease dramatically spread in Indian Ocean islands where 30 to 75% of population were infected in 2005 and 2006, and then extended its progression towards India, Sri Lanka, Indonesia, Malaysia, Maldives islands with more than a million people infected with the East-African strain, replacing the former Asian strain which was known to prevail more than 30 years ago in India. Patients experience sequelae with disability, work loss, and rarely severe outcome recently identified in La Réunion and Mayotte (French overseas territories). No country, no part of the world may consider itself as protected against such events. However, consequences of emerging or re-emerging diseases are more and more unacceptable when they impact the poorest countries of the world. Viruses, bacteria, as well as wild animals, birds, or arthropods are not stopped by borders. It is time now to promote barriers against infectious diseases, including prevention, anticipation, disease surveillance and research. This is not only for humanitarian reasons, but also for contributing to a sustainable development with equity for worldwide population. This report presents comprehensive actions taken in 2006 for tracing the epidemic and mobilise research, as requested to the task force set up by the Prime Minister by March 20, 2006.
Purified preparations of Getah virus strains have been analysed by sodium-dodecyl-sulphate polyacrylamide gel electrophoresis (SDS-PAGE) to reveal their structural proteins. Two envelope proteins (E1 and E2) and core protein (C) were found with the prototype AMM2021 strain both under reducing and nonreducing conditions, while separation of E1 and E2 was observed only under nonreducing conditions for 3 strains isolated in Japan. Limited digestion by Staphylococcus aureus V8 protease revealed difference in the peptide patterns of E1 between AMM2021 and Japanese isolates. Mobility of E1 and E2 was slower for the virus grown in BHK21 cells compared with the virus grown in Aedes albopictus cells, indicating host-controlled modification on the envelope glycoproteins.