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Abstract: Background: Virus isolation is the most reliable evidence of dengue virus (DENV) infection. However, conventional virus isolation methods generally posses lower sensitivity and are time consuming as compared to other diagnostic methods such as detection of viral genome by RT-PCR, and determination of NS1 antigen and anti-DENV antibody by ELISA. Objectives: A virus isolation method relying on the antibody-dependent enhancement mechanism was established and the assay''s efficacy in DENV isolation was confirmed. Study design: FcγR-expressing BHK cells were used for DENV isolation from patient serum samples in the presence of a flavivirus-group reactive monoclonal antibody, mAb4G2, which possesses DENV infection-enhancement activity. DENV genome copy numbers in the culture supernatant fluids of FcγR-expressing BHK cells were assessed and compared to those of parent BHK cells and C6/36 mosquito cells, a cell line commonly used for DENV isolation. Results: The virus titer levels were higher in the culture supernatant fluid of FcγR-expressing BHK cells in the presence of enhancing antibody in comparison with other cell lines using laboratory-established strains and some clinical samples. DENV was isolated from 7 of 16 serum samples by using FcγR-expressing BHK cells in the presence of mAb4G2, but not by using cell lines commonly employed in conventional isolation assays, the FcγR-negative BHK cells and C6/36 cell lines. Conclusions: The results demonstrate that FcγR-expressing BHK cell line in the presence of antibodies, which possess antibody dependent enhancement (ADE) activity, is a useful tool for DENV isolation. [Copyright &y& Elsevier]

• Pre-existing immunity is crucial in the dengue virus (DENV)-2 genotype shift in the Philippines. • Cross-reactive antibodies favored the enhancement of the Cosmopolitan virus in vitro. • Testing antibody-dependent enhancement will help understand DENV cross-protection. The invasion of dengue virus (DENV)-2 Cosmopolitan genotype into the Philippines, where the Asian II genotype previously circulated challenges the principle of dengue serotype-specific immunity. Assessment of antibodies in this population may provide a mechanistic basis for how new genotypes emerge in dengue-endemic areas. We evaluated the neutralizing antibody (nAb) and antibody-dependent enhancement (ADE) responses against the two genotypes using archived serum samples collected from 333 patients with confirmed dengue in Metro Manila, Philippines, before, during, and after the introduction of the Cosmopolitan genotype. We quantified nAb titers in baby hamster kidney (BHK-21) cells with or without the Fc γ receptor IIA (Fc γ RIIA) to detect the capacity of virus-antibody complexes to neutralize or enhance DENV. The nAb potency of the archived serum samples against the two genotypes was greatly affected by the presence of Fc γ RIIA. We found significant differences in nAb titers between the two genotypes in BHK-21 cells with Fc γ RIIA (P <0.0001). The archived serum samples were incapable of fully neutralizing the Cosmopolitan genotype, but instead strongly promoted its ADE compared to the Asian II genotype (P <0.0001). These results reinforce the role of pre-existing immunity in driving genotype shifts. Our finding that specific genotypes exhibit differing susceptibilities to ADE by cross-reactive antibodies may have implications for dengue vaccine development. [Display omitted] [ABSTRACT FROM AUTHOR]

• Pre-existing immunity against recent SARS-CoV-2 strains in Vietnam was determined. • Pre-COVID-19 samples neutralize Wuhan and Alpha strains but not newer variants. • Limited cellular immunity to SARS-CoV-2 suggests a lack of cross-immunity among newer variants of concern. • Pre-COVID-19 immunity may influence low transmission in the early phase of a pandemic. SARS-CoV-2 transmission and epidemic potential is related to the population's immunity levels. As such, assessing different regions' preexisting immune responses to SARS-CoV-2 is important to understand the transmission potential of emerging SARS-CoV-2 variants. In 975 serum samples from Vietnam (2014 to 2019), anti-SARS-CoV-2 Immunoglobulin G levels were determined by enzyme-linked immunosorbent assay. Plaque reduction neutralization test (PRNT) was performed using Wuhan strain and variants of concern (VOCs). Cross-reactivity was confirmed by analyzing B-cell receptor (BCR) repertoire sequences and identifying BCR repertoire sequences-derived T-cell epitopes. Overall, 20.9% (n = 76/364) and 9.2% (n = 7) demonstrated SARS-CoV-2 neutralizing activity (PRNT 50) against the Wuhan and Alpha strain, respectively. Neutralizing activity against Beta, Gamma, and Delta strains was absent (PRNT 50 <5) in all samples. Cross-reactive epitopes against SARS-CoV-2 and other coronavirus spike proteins were detected in the N-terminal domain, S2, and receptor-binding domain regions. Following BCR and major histocompatibility complex analysis, T-cell receptor-recognized epitope motif (TREM) among pathogenic coronaviruses and coronaviruses spike proteins were the top TREM peptide, suggesting that pre-existing immunity against SARS-CoV-2 in Vietnam was due to exposure to common cold coronaviruses. With limited immunity against emerging VOCs, further monitoring, and control of the epidemic, along with COVID-19 vaccine programs against VOCs, are necessary. [ABSTRACT FROM AUTHOR]

• Second COVID-19 wave in Malaysia doubled every 3.8 days and decayed after 1 month. • Instantaneous reproduction number R t peaked at 3.1 and case fatality rate is 1.9. • Movement control measures began 3 days after peaked R t and lasted for 8 weeks. • Combining multiple control measures could provide effective exit strategy. • Disproportionately impacted subpopulations should be identified and supported. The first wave of COVID-19 epidemic began in late January in Malaysia and ended with a very small final size. The second wave of infections broke out in late February and grew rapidly in the first 3 weeks. Authorities in the country responded quickly with a series of control strategies collectively known as the Movement Control Order (MCO) with different levels of intensity matching the progression of the epidemic. We examined the characteristics of the second wave and discussed the key control strategies implemented in the country. In the second wave, the epidemic doubled in size every 3.8 days (95% confidence interval [CI]: 3.3, 4.5) in the first month and decayed slowly after that with a halving time of approximately 3 weeks. The time-varying reproduction number R t peaked at 3.1 (95% credible interval: 2.7, 3.5) in the 3rd week, declined sharply thereafter and stayed below 1 in the last 3 weeks of April, indicating low transmissibility approximately 3 weeks after the MCO. Experience of the country suggests that adaptive triggering of distancing policies combined with a population-wide movement control measure can be effective in suppressing transmission and preventing a rebound. [ABSTRACT FROM AUTHOR]

In 2013 in Myanmar, dengue epidemic occurred with 20,255 cases including 84 deaths. This study aimed to determine the serological and molecular characteristics of dengue virus (DENV) infection among children with clinical diagnosis of dengue hemorrhagic fever (DHF) or dengue shock syndrome (DSS) during this period. Single acute serum samples were collected from 300 children in Mandalay Children Hospital, Mandalay, Myanmar. Out of the 300 children, 175 (58.3%) and 183 (61%) were positive for anti-dengue IgM and anti-dengue IgG, respectively. Among the IgM positives, 41 (23.4%) had primary DENV infection. Thirty-nine DENV strains (23 DENV-1, 10 DENV-2 and 6 DENV-4) were successfully isolated after inoculation of the patient serum samples onto C6/36 cells. DENV 1 was the dominant serotype in the 2013 epidemic. There was no correlation between the infecting serotypes and clinical severities. The DENV-1 strains belonged to three lineages of the genotype 1; the DENV-2 strains were of the Asian I genotype and were separated into two lineages; and DENV-4 strains belonged to the same lineage of genotype I. It is of interest to note the diversity of DENV-1 and -2 circulating in the same location during June–August 2013. These DENV isolates were genetically close (98%–100%) to the other previously reported isolates from Myanmar and its neighboring countries, namely China, Thailand, Sri Lanka, Cambodia and Vietnam. Primary DENV infection was still high among the severe dengue cases. Different serotypes of DENV were co-circulating in 2013, however, genotype shift was not observed. Additionally, amino acid mutations were detected in the study strains not seen in the previously reported strains from other countries and Myanmar. This paper provided information on the circulating serotypes for the last 15 years and the recent dengue situation in Mandalay, Myanmar after 2006. [ABSTRACT FROM AUTHOR]

Zika virus (ZIKV) is a re-emerging mosquito-borne flavivirus of African origin that is transmitted by Aedes mosquitoes. ZIKV was historically limited to Africa and Asia, where mild cases were reported. However, ZIKV has recently been responsible for major global outbreaks associated with a wide range of neurological complications. Since no antiviral therapy exists for ZIKV, drug discovery research for ZIKV is crucial. Intracellular lipids regulated by sterol regulatory element-binding proteins (SREBPs) are important in flavivirus pathogenesis. PF-429242 has been reported to inhibit the activity of site-1 protease (S1P), which regulates the expression of SREBP target genes. Our primary objective in this study is to elucidate the mechanism of the antiviral activity of PF-429242 against the African genotype (ZIKV MR-766) and Asian genotypes (ZIKV H/PF 2013 and ZIKV PRVABC59) using several primate-derived cell lines. The virus titer was determined via a focus-forming assay; we used flow cytometry to quantify intracellular lipids in ZIKV-infected and mock-treated cells. The PF-429242 molecule effectively suppressed ZIKV infection in neuronal cell lines; T98G, U-87MG, SK-N-SH and primary monocytes cell, indicating that PF-429242 molecule can be used therapeutically. A strong reduction in ZIKV replication was observed at 12 μM and 30 μM in in neuronal cell lines and primary monocytes, respectively. Interestingly, the inhibitory effects of the PF-429242 molecule were observed when it was tested on various ZIKV-lineage infections. Lipid quantification reveals that ZIKV increases lipogenesis in infected cells, while the exogenous addition of cholesterol effectively blocks ZIKV replication. Furthermore, the supplementation of oleic acid increases the ZIKV titer. Fenofibrate, an inhibitor of lipid droplet formation, reduces the ZIKV titer. Collectively, our results demonstrate that the development of antiviral drugs against ZIKV could be based on key regulators of lipid metabolism. In addition, this study reveals that the mechanism of the PF-429242-mediated suppression among flavivirus infections is not entirely identical. Our results warrant further evaluation of PF-429242 as a prospective antiviral drug, given the multiple advantageous properties of this compound, such as its limited toxicity, neuroprotective properties, and broad spectrum of capabilities. [Display omitted] [ABSTRACT FROM AUTHOR]