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Fulltext Isolation and molecular detection of dengue and chikungunya virus from field-collected adult mosquitoes in Kelantan, Malaysia

Jusoh TNAM, Jaafar IS, Shueb RH

J Vector Borne Dis, 2024 Jan 01;61(1):61-71.
PMID: 38648407 DOI: 10.4103/0972-9062.392269

BACKGROUND OBJECTIVES: Dengue and chikungunya infections are one of the major health problems that have plagued the human population globally. All dengue virus (DENV) serotypes circulate within Malaysia with particular serotypes dominating in different years/outbreaks. In the state of Kelantan, an increasing number of DENV and chikungunya virus (CHIKV) new cases have been reported, including several deaths. This study aimed to isolate and detect these arboviruses from adult mosquitoes in Kelantan.
METHODS: Adult mo squito samples were collected from January to August 2019 and were identified according to gender, species and locality. The isolation of the virus was done in C6/36 cells. Dengue NS1 antigen was carried out using direct mosquito lysate and mosquito culture supernatant. Detection and serotyping of the DENV was performed using multiplex RT-PCR and CHIKV detection using a one-step RT-PCR assay.
RESULTS: Of 91 mosquito pools, four were positive for NS1 antigen comprising two pools (2.2%) of male Ae. albopictus (Pulau Melaka and Kubang Siput) and two pools (2.2%) of Ae. aegypti (Kampung Demit Sungai). DENV 1 was detected in one pool (0.9%) of female Ae. albopictus among 114 tested Aedes pools. Two pools of 114 pools (1.7%) from both male Aedes species were positive with double serotypes, DENV 1 and DENV 2 (Pulau Melaka). However, no pool was positive for CHIKV.
INTERPRETATION CONCLUSION: The presence of DENV and the main vectors of arboviruses in Kelantan are pertinent indicators of the need to improve vector controls to reduce arbovirus infections among people in the localities.

Matched MeSH terms: Chikungunya Fever/virology
Fulltext Assessing the threat of chikungunya virus emergence in Australia

Viennet E, Knope K, Faddy HM, Williams CR, Harley D

Commun Dis Intell Q Rep, 2013 Jun;37(2):E136-43.
PMID: 24168087

Chikungunya virus (CHIKV) is a major threat to Australia given the distribution of competent vectors, and the large number of travellers returning from endemic regions. We describe current knowledge of CHIKV importations into Australia, and quantify reported viraemic cases, with the aim of facilitating the formulation of public health policy and ensuring maintenance of blood safety.

Matched MeSH terms: Chikungunya Fever/virology
Fulltext Detection of east/central/south African genotype of chikungunya virus in Myanmar, 2010

Tun MM, Thant KZ, Inoue S, Nabeshima T, Aoki K, Kyaw AK, et al.

Emerg Infect Dis, 2014 Aug;20(8):1378-81.
PMID: 25062511 DOI: 10.3201/eid2008.131431

In 2010, chikungunya virus of the East Central South African genotype was isolated from 4 children in Myanmyar who had dengue-like symptoms. Phylogenetic analysis of the E1 gene revealed that the isolates were closely related to isolates from China, Thailand, and Malaysia that harbor the A226V mutation in this gene.

Matched MeSH terms: Chikungunya Fever/virology*
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 Fever/virology*
Fulltext Independent Emergence of the Cosmopolitan Asian Chikungunya Virus, Philippines 2012

Tan KK, Sy AK, Tandoc AO, Khoo JJ, Sulaiman S, Chang LY, et al.

Sci Rep, 2015 Jul 23;5:12279.
PMID: 26201250 DOI: 10.1038/srep12279

Outbreaks involving the Asian genotype Chikungunya virus (CHIKV) caused over one million infections in the Americas recently. The outbreak was preceded by a major nationwide outbreak in the Philippines. We examined the phylogenetic and phylogeographic relationships of representative CHIKV isolates obtained from the 2012 Philippines outbreak with other CHIKV isolates collected globally. Asian CHIKV isolated from the Philippines, China, Micronesia and Caribbean regions were found closely related, herein denoted as Cosmopolitan Asian CHIKV (CACV). Three adaptive amino acid substitutions in nsP3 (D483N), E1 (P397L) and E3 (Q19R) were identified among CACV. Acquisition of the nsP3-483N mutation in Compostela Valley followed by E1-397L/E3-19R in Laguna preceded the nationwide spread in the Philippines. The China isolates possessed two of the amino acid substitutions, nsP3-D483N and E1-P397L whereas the Micronesian and Caribbean CHIKV inherited all the three amino acid substitutions. The unique amino acid substitutions observed among the isolates suggest multiple independent virus dissemination events. The possible biological importance of the specific genetic signatures associated with the rapid global of the virus is not known and warrant future in-depth study and epidemiological follow-up. Molecular evidence, however, supports the Philippines outbreak as the possible origin of the CACV.

Matched MeSH terms: Chikungunya Fever/virology*
Zika Virus Infection: Current Concerns and Perspectives

Maharajan MK, Ranjan A, Chu JF, Foo WL, Chai ZX, Lau EY, et al.

Clin Rev Allergy Immunol, 2016 Dec;51(3):383-394.
PMID: 27236440

The Zika virus outbreaks highlight the growing importance need for a reliable, specific and rapid diagnostic device to detect Zika virus, as it is often recognized as a mild disease without being identified. Many Zika virus infection cases have been misdiagnosed or underreported because of the non-specific clinical presentation. The aim of this review was to provide a critical and comprehensive overview of the published peer-reviewed evidence related to clinical presentations, various diagnostic methods and modes of transmission of Zika virus infection, as well as potential therapeutic targets to combat microcephaly. Zika virus is mainly transmitted through bites from Aedes aegypti mosquito. It can also be transmitted through blood, perinatally and sexually. Pregnant women are advised to postpone or avoid travelling to areas where active Zika virus transmission is reported, as this infection is directly linked to foetal microcephaly. Due to the high prevalence of Guillain-Barre syndrome and microcephaly in the endemic area, it is vital to confirm the diagnosis of Zika virus. Zika virus infection had been declared as a public health emergency and of international concern by the World Health Organisation. Governments and agencies should play an important role in terms of investing time and resources to fundamentally understand this infection so that a vaccine can be developed besides raising awareness.

Matched MeSH terms: Chikungunya Fever/virology
Fulltext A combination of doxycycline and ribavirin alleviated chikungunya infection

Rothan HA, Bahrani H, Mohamed Z, Teoh TC, Shankar EM, Rahman NA, et al.

PLoS One, 2015;10(5):e0126360.
PMID: 25970853 DOI: 10.1371/journal.pone.0126360

Lack of vaccine and effective antiviral drugs against chikungunya virus (CHIKV) outbreaks have led to significant impact on health care in the developing world. Here, we evaluated the antiviral effects of tetracycline (TETRA) derivatives and other common antiviral agents against CHIKV. Our results showed that within the TETRA derivatives group, Doxycycline (DOXY) exhibited the highest inhibitory effect against CHIKV replication in Vero cells. On the other hand, in the antiviral group Ribavirin (RIBA) showed higher inhibitory effects against CHIKV replication compared to Aciclovir (ACIC). Interestingly, RIBA inhibitory effects were also higher than all but DOXY within the TETRA derivatives group. Docking studies of DOXY to viral cysteine protease and E2 envelope protein showed non-competitive interaction with docking energy of -6.6±0.1 and -6.4±0.1 kcal/mol respectively. The 50% effective concentration (EC50) of DOXY and RIBA was determined to be 10.95±2.12 μM and 15.51±1.62 μM respectively, while DOXY+RIBA (1:1 combination) showed an EC50 of 4.52±1.42 μM. When compared, DOXY showed higher inhibition of viral infectivity and entry than RIBA. In contrast however, RIBA showed higher inhibition against viral replication in target cells compared to DOXY. Assays using mice as animal models revealed that DOXY+RIBA effectively inhibited CHIKV replication and attenuated its infectivity in vivo. Further experimental and clinical studies are warranted to investigate their potential application for clinical intervention of CHIKV disease.

Matched MeSH terms: Chikungunya Fever/virology
Deciphering the potential of baicalin as an antiviral agent for Chikungunya virus infection

Oo A, Rausalu K, Merits A, Higgs S, Vanlandingham D, Bakar SA, et al.

Antiviral Res, 2018 02;150:101-111.
PMID: 29269135 DOI: 10.1016/j.antiviral.2017.12.012

The past decade has seen the re-emergence of Chikungunya virus (CHIKV) as a major global health threat, affecting millions around the world. Although fatal infections are rare among infected patients, the occurrence of long-lasting polyarthralgia has a significant impact on patients' quality of lives and ability to work. These issues were the stimuli for this study to determine the potential of baicalin, a bioflavonoid, as the novel antiviral compound against CHIKV. It was found that baicalin was well tolerated by Vero, BHK-21 and HEK 293T cells with maximal nontoxic doses >600 μM, ≈ 350 μM and ≈110 μM, respectively. Antiviral assays indicated that baicalin was the most effective inhibitor when tested for its direct virucidal activity with EC50 ≈ 7 μM, followed by inhibition of virus entry into the host cell, attachment of virus particle to cellular receptors and finally intracellular replication of viral RNA genome. In silico analysis using molecular docking demonstrated close interactions between baicalin and CHIKV envelope protein with considerably strong binding affinity of -9.7 kcal/mol. qRT-PCR analysis revealed that baicalin had the greatest effect on the synthesis of viral negative stand RNA with EC50 ≈ 0.4 μM followed by the inhibition of synthesis of positive-strand genomic (EC50 ≈ 13 μM) and subgenomic RNAs (EC50 ≈ 14 μM). These readings indicate that the compound efficiently inhibits replicase complexes formation but is a less potent inhibitor of existing replicase complexes. Coherent with this hypothesis, the use of recombinant CHIKV replicons harboring Renilla luciferase marker showed that replication of corresponding replicon RNAs was only slightly downregulated at higher doses of baicalin, with EC50 > 100 μM. Immunofluorescence and western blotting experiments demonstrated dose-dependent inhibition of expression of different viral proteins. It was also observed that levels of important protein markers for cellular autophagy (LC3) and apoptosis (Bax) were reduced in baicalin treatment groups as compared with untreated virus infected controls. In summary, given its low toxicity and high efficacy against CHIKV, baicalin has great potential to be developed as the novel antiviral compound for CHIKV. In vivo studies to evaluate its activity in a more complexed system represent a necessary step for future analysis.

Matched MeSH terms: Chikungunya Fever/virology*