Macrobrachium rosenbergii nodavirus (MrNv) infects giant freshwater prawns and causes white tail disease (WTD). The coding region of the capsid protein of MrNv was amplified with RT-PCR and cloned into the pTrcHis2-TOPO vector. The recombinant plasmid was introduced into Escherichia coli and protein expression was induced with IPTG. SDS-PAGE showed that the recombinant protein containing the His-tag and myc epitope has a molecular mass of about 46 kDa and it was detected by the anti-His antibody in Western blotting. The protein was purified using immobilized metal affinity chromatography (IMAC) and transmission electron microscopic analysis revealed that the recombinant protein assembled into virus-like particles (VLPs) with a diameter of about 30±3 nm. The size of the particles was confirmed by dynamic light scattering. Nucleic acids were extracted from the VLPs and treatment with nucleases showed that they were mainly RNA molecules. This is the first report describing the production of MrNv capsid protein in bacteria and its assembly into VLPs.
From August 2006 to July 2007 a prospective study of out-of-pocket costs incurred by care-givers of children hospitalized for rotavirus gastroenteritis was conducted in a hospital in Malaysia. Data on caretaker out-of-pocket costs were collected from 260 children hospitalized with diarrhoea. A stool sample was collected from 198 of these children of which 46 (23%) were positive for rotavirus by latex agglutination assay. The mean (median; interquartile range) out-of-pocket cost incurred by the care-givers was US$194 (US$169; US$47-738), constituting 26% of average monthly income of the households surveyed. Major components of the cost were hospital expenses (45%) and productivity loss (37%). These findings will allow further assessment of the cost-effectiveness of any future rotavirus immunization program in Malaysia.
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.
A molecular epidemiological investigation was conducted among various risk populations (n = 184) in Kuala Lumpur, Malaysia, in 2003 to 2005, on the basis of nucleotide sequences of protease and reverse transcriptase regions. In addition to circulating HIV-1 strains, including CRF01_AE (57.1%), subtype B (20.1%), and subtype C (0.5%), we detected a candidate with a new circulating recombinant form (CRF). We determined four near-full-length nucleotide sequences with identical subtype structure from epidemiologically unlinked individuals of different risk and ethnic groups. In this chimera, two short subtype B segments were inserted into the gag-RT region in a backbone of CRF01_AE. The recombinant structure was distinct from previously identified CRF15_01B in Thailand. In agreement with the current HIV nomenclature system, this constitutes a novel CRF (CRF33_01B). The overall prevalence of CRF33_01B is 19.0% (35/184). Although the prevalence of CRF33_01B is particularly high among injecting drug users (42.0%, 21/50), it is also detected in a substantial proportion of homo-/bisexual males (18.8%, 3/16) and heterosexuals (9.8%, 9/92). Moreover, unique recombinant forms composed of CRF01_AE and subtype B that have a significant structural relationship with CRF33_01B were detected in 1.6% (3/184) of study subjects, suggesting an ongoing recombination process in Malaysia. This new CRF seems to be bridging viral transmission between different risk populations in this country.
An effective live attenuated rubella vaccine was available since 1969 and congenital rubella syndrome can be prevented with appropriate vaccination. We report a baby with congenital rubella syndrome born in Klang valley to indicate that the Universal Rubella Vaccination Programme adopted by the Ministry of Health Malaysia since 2002 has yet to achieve its effect of eliminating transmission of rubella and preventing congenital rubella infection in the community. To our knowledge, the virus isolate represents the first successful isolation of rubella virus in this country and will serve as the reference strain for future comparison in molecular epidemiological tracking of rubella virus activity this country.
Newcastle disease virus (NDV) strains can be classified as virulent or avirulent based upon the severity of the disease. Differentiation of the virus into virulent and avirulent is necessary for effective control of the disease. Biopanning experiments were performed using a disulfide constrained phage displayed heptapeptide library against three pathotypes of NDV strains: velogenic (highly virulent), mesogenic (moderately virulent) and lentogenic (avirulent). A phage clone bearing the peptide sequence SWGEYDM capable of distinguishing virulent from avirulent NDV strains was isolated. This phage clone was employed as a diagnostic reagent in a dot blot assay and it successfully detected only virulent NDV strains.
Nipah virus infection of porcine stable kidney cells (PS), human neuronal cells (SK-N-MC), human lung fibroblasts cells (MRC-5), and human monocytes (THP-1) were examined. Rapid progression of cytopathic effects (CPE) and cell death were noted in PS cell cultures treated with Nipah virus, followed by MRC-5, SK-N-MC, and THP-1 cell cultures, in descending order of rapidity. Significant increase in the intracellular Nipah virus RNA occurred beginning at 24 hr PI in all the infected cells. Whereas, the extracellular release of Nipah virus RNA increased significantly beginning at 48 and 72 hr PI for the infected MRC-5 cells and PS cells, respectively. No significant release of extracellular Nipah virus RNA was detected from the Nipah virus-infected SK-N-MC and THP-1 cells. At its peak, approximately 6.6 log PFU/microl of extracellular Nipah virus RNA was released from the Nipah virus-infected PS cells, with at least a 100-fold less virus RNA was recorded in the Nipah virus-infected SK-N-MC and THP-1. Approximately 15.2% (+/-0.1%) of the released virus from the infected PS cell cultures was infectious in contrast to approximately 5.5% (+/-0.7%) from the infected SK-N-MC cells. The findings suggest that there are no differences in the capacity to support Nipah virus replication between pigs and humans in fully susceptible PS and MRC-5 cells. However, there are differences between these cells and human neuronal cells and monocytes in the ability to support Nipah virus replication and virus release.
The HIV protease-reverse transcriptase (PR-RT) (1047 bp), gp120-env (891 bp) and gp41-env (547 bp) regions from the plasma of 115 HIV-1-infected patients in Kuala Lumpur (KL), Malaysia were sequenced. Detailed phylogenetic and bootscanning analyses were performed to determine the mosaic structure of the HIV-1 strains and their recombination breakpoint(s). Among the 50 patient samples in which all three regions could be amplified, the HIV-1 CRF01_AE subtype (46%) was predominant followed by subtypes B (10%) and B' (6%). A total of 9/50 (18%) patients were infected with a CRF01_AE/B inter-subtype recombinant, displaying a recombinant form (RF)(PR-RT), CRF01_AE(gp120-env) and CRF01_AE(gp41-env). This RF was derived from the Thai variants of CRF01_AE and B' subtype, with two distinct B' subtype segments in the backbone of CRF01_AE, similar to the newly identified CRF33_01B. In addition, one sample demonstrated a close structural relationship with the new CRF33_01B in the PR-RT region but displayed B' segment in part of the env region (RF(PR-RT), CRF01_AE/B'(gp120-env) and B'(gp41-env)) indicating continuing evolution of CRF33_01B. The remaining 18% of samples were identified as unique recombinant forms (URFs).
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.
Variants of Coconut cadang-cadang viroid have been identified in a plantation oil palm growing in Malaysia. Three size classes are described, comprising 297, 293, and 270 nt. Compared with the 296-nt form of coconut cadang-cadang viroid (CCCVd), all variants substituted C31 --> U in the pathogenicity domain and A175 --> C in the right-hand terminus. Other mutations and deletions accounted for the different sizes. These are the first sequences reported for variants of Coconut cadang-cadang viroid in a species other than coconut palm, and this is the first evidence that variants closely related to CCCVd occur outside the Philippines.
Japanese encephalitis (JE) is a global public health issue that has spread widely to more than 20 countries in Asia and has extended its geographic range to the south Pacific region including Australia. JE has become the most important cause of viral encephalitis in the world. Japanese encephalitis viruses (JEV) are divided into five genotypes, based on the nucleotide sequence of the envelope (E) gene. The Muar strain, isolated from patient in Malaya in 1952, is the sole example of genotype V JEV. Here, the XZ0934 strain of JEV was isolated from Culex tritaeniorhynchus, collected in China. The complete nucleotide and amino acid sequence of XZ0934 strain have been determined. The nucleotide divergence ranged from 20.3% to 21.4% and amino acid divergence ranged from 8.4% to 10.0% when compared with the 62 known JEV isolates that belong to genotype I-IV. It reveals low similarity between XZ0934 and genotype I-IV JEVs. Phylogenetic analysis using both complete genome and structural gene nucleotide sequences demonstrates that XZ0934 belongs to genotype V. This, in turn, suggests that genotype V JEV is emerging in JEV endemic areas. Thus, increased surveillance and diagnosis of viral encephalitis caused by genotype V JEV is an issue of great concern to nations in which JEV is endemic.
Dengue virus type 4 (DENV-4) circulates in tropical and subtropical countries from Asia and the Americas. Despite the importance of dengue virus distribution, little is known about the worldwide viral spread. Following a Bayesian phylogenetic approach we inferred the evolutionary history of 310 isolates sampled from 37 countries during the time period 1956-2008 and the spreading dynamics for genotypes I and II. The region (tropical rainforest biome) comprised by Malaysia-Thailand was the most likely ancestral area from which the serotype has originated and spread. Interestingly, cross-correlation analysis on demographic time series with the Asian sequences showed a statistically significant negative correlation that could be suggestive of competition among genotypes within the same serotype.
Malaysia experienced its first outbreak of chikungunya virus (CHIKV) infection in late 1998 in Klang District in Selangor; six years later the virus re-emerged in the state of Perak. All the CHIKV isolates in 1988 and 2006 shared high sequence similarities and belonged to the Asian genotype. In 2007 and 2008 CHIKV infection again reemerged but the genotype was the Central/East African genotype. This strain was found to be similar to the strains causing outbreaks in the India Ocean. In 2009, the strains circulating in Malaysia, including the state of Kelantan, based on the partial E1 gene, also belong to the Central/East African genotype.
From 2005 to 2009, the Institute for Medical Research (IMR), Kuala Lumpur received a total of 7,117 respiratory specimens from patients with influenza-like illness (ILI) for influenza screening. Seasonal influenza virus was isolated from 17.3% of patients with ILI in 2005, 31.6% in 2006, 12.8% in 2007, 10.2% in 2008 and 13.5% in 2009. There were one or more influenza A and B virus strains circulating in Malaysia throughout the year, with distinctly a peak in May to August. The predominant circulating strains of seasonal influenza A were A/California/7/2004-like (H3N2) in 2005, A/New Caledonia/20/99-like (H1N1) in 2006, A/ Brisbane/10/2007-like (H3N2) in 2007 and 2008, and A/Perth/16/2009-like (H3N2) virus in 2009. The predominant circulating strains of influenza B were B/Hong Kong/330/2001-like in 2005, B/Malaysia/2506/2004-like in 2006, B/Florida/4/2006-like in 2007 and 2008, and B/Brisbane/60/2008-like in 2009.
From 2005 to 2009, the Institute for Medical Research (IMR), Kuala Lumpur, Malaysia received 488 serum and blood samples from hospitalized patients on the East Coast of Peninsular Malaysia, suspected of having dengue infection. In this study we determined the prevailing dengue serotypes using a real time polymerase chain reaction assay (RT-PCR). All 4 dengue virus serotypes were found circulating during the study period; however the predominant serotype varied. In 2005 and 2006, the predominant serotypes circulating were DENV-1 and DENV-3, in 2007, DENV-1 and DENV-2 were predominant, and in 2008 and 2009, DENV-3 was the predominant serotype.
We previously described three new Malaysian orthoreoviruses designated Pulau virus, Melaka virus and Kampar virus. Melaka and Kampar viruses were shown to cause respiratory disease in humans. These viruses, together with Nelson Bay virus, isolated from Australian bats, are tentatively classified as different strains within the species Pteropine orthoreovirus (PRV), formerly known as Nelson Bay orthoreovirus, based on the small (S) genome segments. Here we report the sequences of the large (L) and medium (M) segments, thus completing the whole-genome characterization of the four PRVs. All L and M segments were highly conserved in size and sequence. Conserved functional motifs previously identified in other orthoreovirus gene products were also found in the deduced proteins encoded by the cognate segments of these viruses. Detailed sequence analysis identified two genetic lineages divided into the Australian and Malaysian PRVs, and potential genetic reassortment among the M and S segments of the three Malaysian viruses.
A total of 246 stool samples were collected from the public who participated in a Medical Fair held at the University Malaya Medical Center. The stools were examined for intestinal parasites using the formalin-ether concentration technique. The overall infection rate was 6.9% (17 out of 246), with Trichuris trichiura being the most common parasite (4.5%), followed by Ascaris lumbricoides (0.8%), Clonorchis sinensis (0.8%), hookworm (0.4%), and Entamoeba histolytica (0.4%). None of these participants showed any clinical symptoms. The highest infection rate was among the Chinese 7.7% (13 out of 169), followed by the Malays 7.0% (3 out of 43) and Indians 3.3% (1 out of 30). The highest infection rate was in the age group 16-30 years, which was 9% (6 out of 67). The two cases of clonorchiasis were from two Chinese women aged 28 and 66 years. The 28-year-old Chinese woman was born in Malaysia and had never left the country, while the older woman was also born in Malaysia but had visited Hong Kong as a tourist on two occasions. Both enjoyed eating raw fresh water fish with porridge.
A Hepatitis A outbreak occurred in Mukim Hulu Langat, Hulu Langat district from April 2002 to October 2002. Of the 51 cases notified, most were among students and the ethnic groups involved were Malays and the Orang Asli (local indigenous community). Epidemiological investigations revealed that the cases were localized along rivers used for recreational activities in this area. River water analysis indicated human faecal contamination and it was believed that the contamination was due to the Orang Asli community living upstream. This occurred due to lack of toilet facilities, water at point of use and the existing traditional practices of the Orang Asli community. Control measures instituted were intense health education to the Orang Asli to avoid using the rivers for defecation, multi agency efforts to provide sanitary toilets and adequate water to the villages affected. Future measures include conducting a sero- prevalence survey to determine the feasibility of Hepatitis A immunization to the susceptible population in this area. The outbreak that began in April 2002 was controlled by October 2002.