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Fulltext Genetic diversity of Japanese encephalitis virus isolates obtained from the Indonesian archipelago between 1974 and 1987

Schuh AJ, Guzman H, Tesh RB, Barrett AD

Vector Borne Zoonotic Dis, 2013 Jul;13(7):479-88.
PMID: 23590316 DOI: 10.1089/vbz.2011.0870

Five genotypes (GI-V) of Japanese encephalitis virus (JEV) have been identified, all of which have distinct geographical distributions and epidemiologies. It is thought that JEV originated in the Indonesia-Malaysia region from an ancestral virus. From that ancestral virus GV diverged, followed by GIV, GIII, GII, and GI. Genotype IV appears to be confined to the Indonesia-Malaysia region, as GIV has been isolated in Indonesia from mosquitoes only, while GV has been isolated on three occasions only from a human in Malaysia and mosquitoes in China and South Korea. In contrast, GI-III viruses have been isolated throughout Asia and Australasia from a variety of hosts. Prior to this study only 13 JEV isolates collected from the Indonesian archipelago had been studied genetically. Therefore the sequences of the envelope (E) gene of 24 additional Indonesian JEV isolates, collected throughout the archipelago between 1974 and 1987, were determined and a series of molecular adaptation analyses were performed. Phylogenetic analysis indicated that over a 14-year time span three genotypes of JEV circulated throughout Indonesia, and a statistically significant association between the year of virus collection and genotype was revealed: isolates collected between 1974 and 1980 belonged to GII, isolates collected between 1980 and 1981 belonged to GIV, and isolates collected in 1987 belonged to GIII. Interestingly, three of the GII Indonesian isolates grouped with an isolate that was collected during the JE outbreak that occurred in Australia in 1995, two of the GIII Indonesian isolates were closely related to a Japanese isolate collected 40 years previously, and two Javanese GIV isolates possessed six amino acid substitutions within the E protein when compared to a previously sequenced GIV isolate collected in Flores. Several amino acids within the E protein of the Indonesian isolates were found to be under directional evolution and/or co-evolution. Conceivably, the tropical climate of the Indonesia/Malaysia region, together with its plethora of distinct fauna and flora, may have driven the emergence and evolution of JEV. This is consistent with the extensive genetic diversity seen among the JEV isolates observed in this study, and further substantiates the hypothesis that JEV originated in the Indonesia-Malaysia region.

Matched MeSH terms: RNA, Viral/genetics; RNA, Viral/chemistry
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: RNA, Viral/analysis; RNA, Viral/genetics
Genome sequence of a novel HIV-1 circulating recombinant form 54_01B from Malaysia

Ng KT, Ong LY, Takebe Y, Kamarulzaman A, Tee KK

J Virol, 2012 Oct;86(20):11405-6.
PMID: 22997423

We report here the first novel HIV-1 circulating recombinant form (CRF) 54_01B (CRF54_01B) isolated from three epidemiologically unlinked subjects of different risk groups in Malaysia. These recently sampled recombinants showed a complex genome organization composed of parental subtype B' and CRF01_AE, with identical recombination breakpoints observed in the gag, pol, and vif genes. Such a discovery highlights the ongoing active generation and spread of intersubtype recombinants involving the subtype B' and CRF01_AE lineages and indicates the potential of the new CRF in bridging HIV-1 transmission among different risk groups in Southeast Asia.

Matched MeSH terms: RNA, Viral/analysis; RNA, Viral/genetics
Development of SYBR green I based one-step real-time RT-PCR assay for the detection and differentiation of very virulent and classical strains of infectious bursal disease virus

Kong LL, Omar AR, Hair Bejo M, Ideris A, Tan SW

J Virol Methods, 2009 Nov;161(2):271-9.
PMID: 19591873 DOI: 10.1016/j.jviromet.2009.06.023

A SYBR Green I based one-step real-time reverse transcriptase polymerase chain reaction was developed for the detection and differentiation of very virulent (vv) and classical strains of infectious bursal disease virus (IBDV). The assay showed high PCR efficiency >93% and high reproducibility with coefficient of variation less than 0.5%. When tested on characterized IBDV strains, the very virulent and classical-specific primers detected accurately only vvIBDV and classical IBDV strains, respectively. The diagnostic efficacy of the assay was also tested on 140 bursal samples from experimental infection and 37 bursal samples from cases suspected of IBD. The assay was able to detect IBDV from bursal samples collected at days 3 and 5 post-infection with the vvIBDV strain UPM94/273 and the classical IBDV strain D78. The assay was also able to detect bursal samples infected dually with D78 and UPM94/273. The melting temperature values of the amplification products from the classical and very virulent viral infection were statistically significant (P<0.05). The specificity of the assay for detecting IBDV from suspected cases was confirmed by sequence analysis of the VP2 gene. The assay showed high sensitivity since bursal samples which were negative for IBDV were confirmed by virus isolation and PCR amplification. Hence, the new assay offers an attractive method for rapid detection of strains of IBDV.

Matched MeSH terms: RNA, Viral/analysis; RNA, Viral/genetics
Fulltext Molecular analysis of 2009 pandemic influenza A(H1N1) in Malaysia associated with mild and severe infections

Balraj P, Sidek H, Suppiah J, Khoo AS, Saat Z

Malays J Pathol, 2011 Jun;33(1):7-12.
PMID: 21874745 MyJurnal

The 2009 pandemic influenza A(H1N1) was first detected in Malaysia in May 2009. It quickly spread in the general population and contributed to a number of influenza-like illness. The objective of the study is to characterize genetic changes in early Malaysian isolates of mild and severe illness of the novel influenza, and to compare sequences of viruses circulating in Malaysia to those in other countries between May to September 2009. Viral isolates of 56 mild cases and 10 severe (intensive care unit or fatal) cases were sequenced for haemagglutinin (HA) and neuraminidase (NA). Genome sequencing of the viral RNA was conducted on 5 isolates (3 were from fatal cases). Highly conserved sequences with few sporadic variations were identified in HA and NA. E374K and D222N were identified in 2 viral isolates from patients with severe illness. Phylogenetic analysis showed close genetic relatedness to the vaccine strain A/California/07/09 and other isolates circulating worldwide during the same period. Sporadic variations were identified in the viral isolates, however a larger sample size is required to make associations with disease severity.

Matched MeSH terms: RNA, Viral/analysis; RNA, Viral/genetics
Comparative analysis of viral RNA and apoptotic cells in bursae following infection with infectious bursal disease virus

Kong LL, Omar AR, Hair-Bejo M, Aini I, Seow HF

Comp Immunol Microbiol Infect Dis, 2004 Nov;27(6):433-43.
PMID: 15325516

Specific-pathogen-free (SPF) chickens infected with very virulent (vv) infectious bursal disease virus (IBDV) UPM94/273 developed lower pathogenicity compared to UPM97/61. Sequence analysis indicated that UPM94/273 is an exceptional vvIBDV. In this study, a SYBR Green I based real-time reverse transcriptase reaction assay was developed to measure viral RNA in the bursae of SPF chickens infected with IBDV. Specificity of the amplified products was confirmed by melting temperature analysis. A linear relationship was observed between the amount of input viral RNA and the threshold values for IBDV-specific product over five log10 dilutions. The viral RNA level following infection with UPM94/273 was significantly higher at day 1 and 2 post-inoculation (p.i.) compared to UPM97/61 infected chickens. However, chickens infected with UPM97/61 had significantly higher numbers of bursal cells undergoing apoptosis compared to UPM94/273 infected chickens. In both groups, the number of apoptotic cells and viral RNA levels peak at day 3 p.i. This study indicates that UPM97/61 and UPM94/273 have different efficiency of replication and percentage of apoptotic cells in bursae during the acute phase of IBDV infection.

Matched MeSH terms: RNA, Viral/analysis*; RNA, Viral/genetics
Molecular characterization of serotype A foot-and-mouth disease viruses circulating in Vietnam in 2009

Le VP, Nguyen T, Lee KN, Ko YJ, Lee HS, Nguyen VC, et al.

Vet Microbiol, 2010 Jul 29;144(1-2):58-66.
PMID: 20097490 DOI: 10.1016/j.vetmic.2009.12.033

Foot-and-mouth disease (FMD) is a major cause of endemic outbreaks in Vietnam in recent years. In this work, six serotype A foot-and-mouth disease viruses (FMDV), collected from endemic outbreaks during January and February of 2009 in four different provinces in Vietnam, were genetically characterized for their complete genome sequences. Genetic analysis based on the complete viral genome sequence indicated that they were closely related to each other and shared 99.0-99.8% amino acid (aa) identity. Genetic and deduced aa analysis of the capsid coding gene VP1 showed that the six Vietnamese strains were all classified into the genotype IX from a total of 10 major genotypes worldwide, sharing 98.1-100% aa identity each other. They were most closely related to the type A strains recently isolated in Laos (A/LAO/36/2003, A/LAO/1/2006, A/LAO/6/2006, A/LAO/7/2006, and A/LAO/8/2006), Thailand (A/TAI/2/1997 and A/TAI/118/1987), and Malaysia (A/MAY/2/2002), sharing 88.3-95.5% nucleotide (nt) identities. In contrast, Vietnamese type A strains showed low nt identities with the two old type A FMDVs, isolated in 1960 in Thailand (a15thailand iso43) and in 1975 in the Philippines (aphilippines iso50), ranging from 77.3 to 80.9% nt identity. A multiple alignment based on the deduced amino acid sequences of the capsid VP1 coding gene of type A FMDV revealed three amino acid substitutions between Vietnamese strains and the strains of other Southeast Asian countries (Laos, Thailand, Malaysia, and the Philippines). Alanine was replaced by valine at residue 24, asparagine by arginine at residue 85, and serine by threonine at residue 196. Furthermore, type A FMDV strains recently isolated in Vietnam, Laos, Thailand, and Malaysia all have one amino acid deletion at residue 140 of the capsid VP1 protein compared with the two old type A FMDV strains from Thailand and the Philippines as well as most other type A representatives worldwide. This article is the first to report on the comprehensive genetic characterization of type A FMDV circulating in Vietnam.

Matched MeSH terms: RNA, Viral/genetics; RNA, Viral/isolation & purification
The utilization of SiNWs/AuNPs-modified indium tin oxide (ITO) in fabrication of electrochemical DNA sensor

Rashid JI, Yusof NA, Abdullah J, Hashim U, Hajian R

Mater Sci Eng C Mater Biol Appl, 2014 Dec;45:270-6.
PMID: 25491829 DOI: 10.1016/j.msec.2014.09.010

This work describes the incorporation of SiNWs/AuNPs composite as a sensing material for DNA detection on indium tin-oxide (ITO) coated glass slide. The morphology of SiNWs/AuNPs composite as the modifier layer on ITO was studied by scanning electron microscopy (SEM) and energy dispersive X-ray spectroscopy (EDX). The morphological studies clearly showed that SiNWs were successfully decorated with 20 nm-AuNPs using self-assembly monolayer (SAM) technique. The effective surface area for SiNWs/AuNPs-modified ITO enhanced about 10 times compared with bare ITO electrode. SiNWs/AuNPs nanocomposite was further explored as a matrix for DNA probe immobilization in detection of dengue virus as a bio-sensing model to evaluate its performance in electrochemical sensors. The hybridization of complementary DNA was monitored by differential pulse voltammetry (DPV) using methylene blue (MB) as the redox indicator. The fabricated biosensor was able to discriminate significantly complementary, non-complementary and single-base mismatch oligonucleotides. The electrochemical biosensor was sensitive to target DNA related to dengue virus in the range of 9.0-178.0 ng/ml with detection limit of 3.5 ng/ml. In addition, SiNWs/AuNPs-modified ITO, regenerated up to 8 times and its stability was up to 10 weeks at 4°C in silica gel.

Matched MeSH terms: RNA, Viral/genetics; RNA, Viral/metabolism
Fulltext Antiviral activity of silymarin against chikungunya virus

Lani R, Hassandarvish P, Chiam CW, Moghaddam E, Chu JJ, Rausalu K, et al.

Sci Rep, 2015;5:11421.
PMID: 26078201 DOI: 10.1038/srep11421

The mosquito-borne chikungunya virus (CHIKV) causes chikungunya fever, with clinical presentations such as severe back and small joint pain, and debilitating arthritis associated with crippling pains that persist for weeks and even years. Although there are several studies to evaluate the efficacy of drugs against CHIKV, the treatment for chikungunya fever is mainly symptom-based and no effective licensed vaccine or antiviral are available. Here, we investigated the antiviral activity of three types of flavonoids against CHIKV in vitro replication. Three compounds: silymarin, quercetin and kaempferol were evaluated for their in vitro antiviral activities against CHIKV using a CHIKV replicon cell line and clinical isolate of CHIKV of Central/East African genotype. A cytopathic effect inhibition assay was used to determine their activities on CHIKV viral replication and quantitative reverse transcription PCR was used to calculate virus yield. Antiviral activity of effective compound was further investigated by evaluation of CHIKV protein expression using western blotting for CHIKV nsP1, nsP3, and E2E1 proteins. Briefly, silymarin exhibited significant antiviral activity against CHIKV, reducing both CHIKV replication efficiency and down-regulating production of viral proteins involved in replication. This study may have important consequence for broaden the chance of getting the effective antiviral for CHIKV infection.

Matched MeSH terms: RNA, Viral/antagonists & inhibitors*; RNA, Viral/genetics
Fulltext Early detection of dengue virus by use of reverse transcription-recombinase polymerase amplification

Teoh BT, Sam SS, Tan KK, Danlami MB, Shu MH, Johari J, et al.

J Clin Microbiol, 2015 Mar;53(3):830-7.
PMID: 25568438 DOI: 10.1128/JCM.02648-14

A method for the rapid diagnosis of early dengue virus (DENV) infection is highly needed. Here, a prototype reverse transcription-recombinase polymerase amplification (RT-RPA) assay was developed. The assay detected DENV RNA in <20 min without the need for thermocycling amplification. The assay enabled the detection of as few as 10 copies of DENV RNA. The designed RT-RPA primers and exo probe detected the DENV genome of at least 12 genotypes of DENV circulating globally without cross-reacting with other arboviruses. We assessed the diagnostic performance of the RT-RPA assay for the detection of DENV RNA in 203 serum samples of patients with clinically suspected dengue. The sera were simultaneously tested for DENV using a reverse transcription-loop-mediated isothermal amplification (RT-LAMP) assay, quantitative RT-PCR (qRT-PCR), and IgM- and IgG-capture enzyme-linked immunosorbent assays (ELISA). Acute DENV infection was confirmed in 130 samples and 61 of the samples (46.9%) were classified as viremic with qRT-PCR. The RT-RPA assay showed good concordance (κ of ≥0.723) with the RT-LAMP and qRT-PCR assays in detecting the dengue viremic samples. When used in combination with ELISA, both the RT-RPA and RT-LAMP assays increased the detection of acute DENV infection to ≥95.7% (≥45/47) in samples obtained within 5 days of illness. The results from the study suggest that the RT-RPA assay is the most rapid molecular diagnostic tool available for the detection of DENV. Hence, it is possible to use the RT-RPA assay in a laboratory to complement routine serology testing for dengue.

Matched MeSH terms: RNA, Viral/analysis; RNA, Viral/genetics
Fulltext The Nature of Exposure Drives Transmission of Nipah Viruses from Malaysia and Bangladesh in Ferrets

Clayton BA, Middleton D, Arkinstall R, Frazer L, Wang LF, Marsh GA

PLoS Negl Trop Dis, 2016 06;10(6):e0004775.
PMID: 27341030 DOI: 10.1371/journal.pntd.0004775

Person-to-person transmission is a key feature of human Nipah virus outbreaks in Bangladesh. In contrast, in an outbreak of Nipah virus in Malaysia, people acquired infections from pigs. It is not known whether this important epidemiological difference is driven primarily by differences between NiV Bangladesh (NiV-BD) and Malaysia (NiV-MY) at a virus level, or by environmental or host factors. In a time course study, ferrets were oronasally exposed to equivalent doses of NiV-BD or NiV-MY. More rapid onset of productive infection and higher levels of virus replication in respiratory tract tissues were seen for NiV-BD compared to NiV-MY, corroborating our previous report of increased oral shedding of NiV-BD in ferrets and suggesting a contributory mechanism for increased NiV-BD transmission between people compared to NiV-MY. However, we recognize that transmission occurs within a social and environmental framework that may have an important and differentiating role in NiV transmission rates. With this in mind, ferret-to-ferret transmission of NiV-BD and NiV-MY was assessed under differing viral exposure conditions. Transmission was not identified for either virus when naïve ferrets were cohoused with experimentally-infected animals. In contrast, all naïve ferrets developed acute infection following assisted and direct exposure to oronasal fluid from animals that were shedding either NiV-BD or NiV-MY. Our findings for ferrets indicate that, although NiV-BD may be shed at higher levels than NiV-MY, transmission risk may be equivalently low under exposure conditions provided by cohabitation alone. In contrast, active transfer of infected bodily fluids consistently results in transmission, regardless of the virus strain. These observations suggest that the risk of NiV transmission is underpinned by social and environmental factors, and will have practical implications for managing transmission risk during outbreaks of human disease.

Matched MeSH terms: RNA, Viral/analysis; RNA, Viral/blood
Fulltext Advances in the study of nodavirus

Yong CY, Yeap SK, Omar AR, Tan WS

PeerJ, 2017;5:e3841.
PMID: 28970971 DOI: 10.7717/peerj.3841

Nodaviruses are small bipartite RNA viruses which belong to the family of Nodaviridae. They are categorized into alpha-nodavirus, which infects insects, and beta-nodavirus, which infects fishes. Another distinct group of nodavirus infects shrimps and prawns, which has been proposed to be categorized as gamma-nodavirus. Our current review focuses mainly on recent studies performed on nodaviruses. Nodavirus can be transmitted vertically and horizontally. Recent outbreaks have been reported in China, Indonesia, Singapore and India, affecting the aquaculture industry. It also decreased mullet stock in the Caspian Sea. Histopathology and transmission electron microscopy (TEM) are used to examine the presence of nodaviruses in infected fishes and prawns. For classification, virus isolation followed by nucleotide sequencing are required. In contrast to partial sequence identification, profiling the whole transcriptome using next generation sequencing (NGS) offers a more comprehensive comparison and characterization of the virus. For rapid diagnosis of nodavirus, assays targeting the viral RNA based on reverse-transcription PCR (RT-PCR) such as microfluidic chips, reverse-transcription loop-mediated isothermal amplification (RT-LAMP) and RT-LAMP coupled with lateral flow dipstick (RT-LAMP-LFD) have been developed. Besides viral RNA detections, diagnosis based on immunological assays such as enzyme-linked immunosorbent assay (ELISA), immunodot and Western blotting have also been reported. In addition, immune responses of fish and prawn are also discussed. Overall, in fish, innate immunity, cellular type I interferon immunity and humoral immunity cooperatively prevent nodavirus infections, whereas prawns and shrimps adopt different immune mechanisms against nodavirus infections, through upregulation of superoxide anion, prophenoloxidase, superoxide dismutase (SOD), crustin, peroxinectin, anti-lipopolysaccharides and heat shock proteins (HSP). Potential vaccines for fishes and prawns based on inactivated viruses, recombinant proteins or DNA, either delivered through injection, oral feeding or immersion, are also discussed in detail. Lastly, a comprehensive review on nodavirus virus-like particles (VLPs) is presented. In recent years, studies on prawn nodavirus are mainly focused on Macrobrachium rosenbergii nodavirus (MrNV). Recombinant MrNV VLPs have been produced in prokaryotic and eukaryotic expression systems. Their roles as a nucleic acid delivery vehicle, a platform for vaccine development, a molecular tool for mechanism study and in solving the structures of MrNV are intensively discussed.

Matched MeSH terms: RNA, Viral
Fulltext Little Evidence of Zika Virus Infection in Wild Long-Tailed Macaques, Peninsular Malaysia

Chua CL, Chan YF, Andu ESGS, Rovie-Ryan JJ, Sitam FT, Verasahib K, et al.

Emerg Infect Dis, 2019 02;25(2):374-376.
PMID: 30666941 DOI: 10.3201/eid2502.180258

We tested a sample of 234 wild long-tailed macaques (Macaca fascicularis) trapped in Peninsular Malaysia in 2009, 2010, and 2016 for Zika virus RNA and antibodies. None were positive for RNA, and only 1.3% were seropositive for neutralizing antibodies. Long-tailed macaques are unlikely to be reservoirs for Zika virus in Malaysia.

Matched MeSH terms: RNA, Viral
Fulltext Serological evidence and experimental infection of cynomolgus macaques with pteropine orthoreovirus reveal monkeys as potential hosts for transmission to humans

Tan CW, Wittwer K, Lim XF, Uehara A, Mani S, Wang LF, et al.

Emerg Microbes Infect, 2019;8(1):787-795.
PMID: 31132935 DOI: 10.1080/22221751.2019.1621668

Pteropine orthoreoviruses (PRV) are emerging bat-borne viruses with proven zoonotic transmission. We recently demonstrated human exposure to PRV in Singapore, which together with previous reports from Malaysia and Vietnam suggest that human infection of PRV may occur periodically in the region. This raises the question whether bats are the only sources of human infection. In this study, we screened 517 cynomolgus macaques caught in Singapore for evidence of exposure to PRV3M (also known as Melaka virus), which was first isolated from human patients in Melaka, Malaysia. We found that 67 serum samples were PRV3M positive by ELISA and 34 were also positive by virus neutralization assay. To investigate whether monkeys could act as hosts for PRV transmission, we experimentally infected cynomolgus macaques with PRV3M and housed these animals with uninfected monkeys. Although no clinical signs of infection were observed in infected animals, viral RNA was detected in nasal and rectal swabs and all infected macaques seroconverted. Additionally, one of the uninfected animals seroconverted, implying active shedding and transmission of PRV3M. We provide evidence that PRV exposure in the macaque population in Singapore occurs at a relatively high prevalence and this study suggests that cynomolgus macaques may be an intermediate or reservoir host for PRVs.

Matched MeSH terms: RNA, Viral
Fulltext Capsid structure of Leishmania RNA virus 1

Procházková M, Füzik T, Grybchuk D, Falginella F, Podešvová L, Yurchenko V, et al.

J Virol, 2020 Nov 18.
PMID: 33208443 DOI: 10.1128/JVI.01957-20

Leishmania parasites cause a variety of symptoms, including mucocutaneous leishmaniasis, which results in the destruction of the mucous membranes of the nose, mouth, and throat. The species of Leishmania carrying Leishmania RNA virus 1 (LRV1), from the family Totiviridae, are more likely to cause severe disease and are less sensitive to treatment than those that do not contain the virus. Although the importance of LRV1 for the severity of leishmaniasis was discovered a long time ago, the structure of the virus remained unknown. Here, we present a cryo-electron microscopy reconstruction of the virus-like particle of LRV1 determined to a resolution of 3.65 Å. The capsid has icosahedral symmetry and is formed by 120 copies of a capsid protein assembled in asymmetric dimers. RNA genomes of viruses from the family Totiviridae are synthetized, but not capped at the 5' end, by virus RNA-polymerases. To protect viral RNAs from degradation, capsid proteins of totivirus L-A cleave the 5' caps of host mRNAs, creating decoys to overload the cellular RNA quality control system. Capsid proteins of LRV1 form positively charged clefts, which may be the cleavage sites for the 5' cap of Leishmania mRNAs. Capsid proteins of LRV1 contain a putative RNA binding site distinct from that of the related L-A virus. The structure of the LRV1 capsid enables the rational design of compounds targeting the putative de-capping site. Such inhibitors may be developed into a treatment for mucocutaneous leishmaniasis caused by LRV1-positive species of LeishmaniaIMPORTANCE Twelve million people worldwide suffer from leishmaniasis, resulting in more than thirty thousand deaths annually. The disease has several variants that differ in their symptoms. The mucocutaneous form, which leads to disintegration of the nasal septum, lips, and palate, is predominantly caused by Leishmania parasites carrying Leishmania RNA virus 1 (LRV1). Here, we present the structure of the LRV1 capsid determined using cryo-electron microscopy. Capsid proteins of a related totivirus L-A protect viral RNAs from degradation by cleaving the 5' caps of host mRNAs. Capsid proteins of LRV1 may have the same function. We show that the LRV1 capsid contains positively charged clefts that may be sites for the cleavage of mRNAs of Leishmania cells. The structure of the LRV1 capsid enables the rational design of compounds targeting the putative mRNA cleavage site. Such inhibitors may be used as treatments for muco-cutaneous leishmaniasis.

Matched MeSH terms: RNA, Viral
Fulltext Occurrence of Norovirus GI in green and red onion

Noor Hidayah, M.S., Tuan Zainazor, C., Pui, C.F., Noorlis, A., Noor Eliza, M.R., Naziehah, M.D., et al.

International Food Research Journal, 2011;18(2):677-681.
MyJurnal

Several Norovirus cases due to consumption of green onions have been reported during recent years but reports on red onions are not found. Onions are one of the major tastes in Malaysian food which are sometimes consuming raw especially the green onion. Viral contamination in onions can occur due to planting condition and not properly prepared food. This situation can pose the human health risk. A method was developed to detect the Norovirus that might present on different type of onions. In this study, 60 samples were collected from local market. Elution by Tryptose Phosphate Glycine broth and concentration steps using negatively charge filter were applied to enhance the detection of virus in food due to low copies of virus on food surface. The viral RNA was extracted using Qiagen Rneasy Mini kit before further detection using One-step RT-PCR. The total incidence of Norovirus in green onion and red onion was 13.33% (4/30) and 3.33 % (1/30) respectively. This is the first report of the detection of Norovirus in red and green onions in Malaysia. Based on the results, it is concluded that this method is reliable to detect Norovirus on onions and vegetables surface and hence can be applied in the laboratories for routine or food borne outbreak investigation.

Matched MeSH terms: RNA, Viral
Enterovirus A71 and coxsackievirus A16 show different replication kinetics in human neuronal and non-neuronal cell lines

Yogarajah T, Ong KC, Perera D, Wong KT

Arch Virol, 2017 Mar;162(3):727-737.
PMID: 27878462 DOI: 10.1007/s00705-016-3157-4

Enterovirus A71 (EV-A71) and coxsackievirus A16 (CV-A16) are closely related enteroviruses that cause hand, foot and mouth disease (HFMD) in children. Serious neurological complications almost always occur in EV-A71 infection, but are rare in CV-A16 infection. Based on the hypothesis that this may be because EV-A71 infects neuronal cells more easily than CV-A16, we compared virus infection, replication and spread of EV-A71 and CV-A16 in SK-N-SH cells. We found that CV-A16 invariably showed significantly lower replication and caused less necrotic cell death in SK-N-SH cells, compared with EV-A71. This was not due to a lower proportion of CV-A16-infected cells, since both viruses showed similar proportions of infected cells at all time points analyzed. Furthermore, reduced replication of CV-A16 in SK-N-SH cells does not appear to be due to limited viral receptor availability, which might limit viral entry, because experiments with viral RNA-transfected cells showed the same results as for live virus infections. On the other hand, no differences were observed between EV-A71 and CV-A16 in RD cells and results were generally similar in RD cells for both viruses. Taken together, our findings suggest that the poor growth of CV-A16 and EV-A71in SK-N-SH cells, compared with RD cells, may be due to cell type-specific restrictions on viral replication and spread. Furthermore, the lower viral replication and necrotic cell death in CV-A16-infected SK-N-SH cells, compared with EV-A71-infected SK-N-SH cells, is consistent with the lower prevalence of neurotropism observed in CV-A16-associated HFMD outbreaks. Nonetheless, in vivo data and more extensive comparisons of different viral strains are essential to confirm our findings.

Matched MeSH terms: RNA, Viral
Fulltext Overview of anti-viral effects of probiotics via immune cells in pre-, mid- and post-SARS-CoV2 era

Kanauchi O, Low ZX, Jounai K, Tsuji R, AbuBakar S

Front Immunol, 2023;14:1280680.
PMID: 38116008 DOI: 10.3389/fimmu.2023.1280680

The COVID-19 outbreak has caused significant global changes and increased public awareness of SARS-CoV-2. Substantial progress in developing vaccines, enhancing sanitation practices, and implementing various measures to combat the virus, including the utilization of probiotics has been made. This comprehensive review examined the medical impact of clinically proven probiotics on infectious diseases, considering three crucial time periods: before (pre-), during (mid-), and after (post-) COVID-19 pandemic era. This review also showed a perspective on the use of probiotics to stimulate the innate immune system and prevent infectious diseases. In pre-COVID-19 era, several probiotic strains were found to be clinically effective in addressing gastrointestinal infectious diseases, the common cold and flu. However, the mechanism by which probiotics exerted their antiviral effects remained relatively unclear during that period. Nevertheless, probiotics, Lactococcus lactis strain Plasma (LC-Plasma), and others have gained attention for their unique ability to modulate the immune system and demonstrate antiviral properties. While some probiotics have shown promise in alleviating gastrointestinal symptoms linked to COVID-19, their direct effectiveness in treating or preventing COVID-19 progression has not yet been conclusively established. As we transition into the post-COVID-19 era, the relationship between COVID-19 and plasmacytoid dendritic cells (pDCs), a vital component of the innate immune system, has been gradually elucidated. These findings are now being applied in developing novel vaccines and treatments involving interferons and in immune activation research using probiotics as adjuvants, comparable to CpG-DNA through TLR9. The role of the local innate immune system, including pDCs, as the first line of defense against viral infections has gained increasing interest. Moving forward, insight of the immune system and the crosstalk between probiotics and the innate immune system is expected to highlight the role of probiotics in adjunctive immunoregulatory therapy. In combination with drug treatments, probiotics may play a more substantial role in enhancing immune responses. The immunoregulatory approach using probiotics such as LC-Plasma, which can induce anti-infectious factors such as interferons, holds promise as a viable therapeutic and prophylactic option against viral infectious diseases due to their good safety profile and protective efficacy.

Matched MeSH terms: RNA, Viral
Fulltext Recent advances in HIV diagnostics: Point-of-care CD4 + T-cell count & viral load assays

Balakrishnan P, Saravanan S, Vignesh R, Shankar EM

Indian J Med Res, 2023 Nov 01;158(5&6):447-450.
PMID: 38063301 DOI: 10.4103/ijmr.ijmr_1616_23
Matched MeSH terms: RNA, Viral
A label-free electrochemical DNA biosensor used a printed circuit board gold electrode (PCBGE) to detect SARS-CoV-2 without amplification

Zambry NS, Awang MS, Beh KK, Hamzah HH, Bustami Y, Obande GA, et al.

Lab Chip, 2023 Mar 14;23(6):1622-1636.
PMID: 36786757 DOI: 10.1039/d2lc01159j

The emergence of coronavirus disease 2019 (COVID-19) motivates continuous efforts to develop robust and accurate diagnostic tests to detect severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2). Detection of viral nucleic acids provides the highest sensitivity and selectivity for diagnosing early and asymptomatic infection because the human immune system may not be active at this stage. Therefore, this work aims to develop a label-free electrochemical DNA biosensor for SARS-CoV-2 detection using a printed circuit board-based gold substrate (PCBGE). The developed sensor used the nucleocapsid phosphoprotein (N) gene as a biomarker. The DNA sensor-based PCBGE was fabricated by self-assembling a thiolated single-stranded DNA (ssDNA) probe onto an Au surface, which performed as the working electrode (WE). The Au surface was then treated with 6-mercapto-1-hexanol (MCH) before detecting the target N gene to produce a well-oriented arrangement of the immobilized ssDNA chains. The successful fabrication of the biosensor was characterized using cyclic voltammetry (CV), electrochemical impedance spectroscopy (EIS), and atomic force microscopy (AFM). The DNA biosensor performances were evaluated using a synthetic SARS-CoV-2 genome and 20 clinical RNA samples from healthy and infected individuals through EIS. The developed DNA biosensor can detect as low as 1 copy per μL of the N gene within 5 minutes with a LOD of 0.50 μM. Interestingly, the proposed DNA sensor could distinguish the expression of SARS-CoV-2 RNA in a patient diagnosed with COVID-19 without any amplification technique. We believe that the proposed DNA sensor platform is a promising point-of-care (POC) device for COVID-19 viral infection since it offers a rapid detection time with a simple design and workflow detection system, as well as an affordable diagnostic assay.

Matched MeSH terms: RNA, Viral

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