A reverse transcription loop-mediated isothermal amplification for broad coverage detection of Asian and African Zika virus lineages.

BACKGROUND: Early detection of Zika virus (ZIKV) infection during the viremia and viruria facilitates proper patient management and mosquito control measurement to prevent disease spread. Therefore, a cost-effective nucleic acid detection method for the diagnosis of ZIKV infection, especially in resource-deficient settings, is highly required. METHODS: In the present study, a single-tube reverse transcription loop-mediated isothermal amplification (RT-LAMP) assay was developed for the detection of both the Asian and African-lineage ZIKV. The detection limit, strain coverage and cross-reactivity of the ZIKV RT-LAMP assay was evaluated. The sensitivity and specificity of the RT-LAMP were also evaluated using a total of 24 simulated clinical samples. The ZIKV quantitative reverse transcription-polymerase chain reaction (qRT-PCR) assay was used as the reference assay. RESULTS: The detection limit of the RT-LAMP assay was 3.73 ZIKV RNA copies (probit analysis, P ≤ 0.05). The RT-LAMP assay detected the ZIKV genomes of both the Asian and African lineages without cross-reacting with other arthropod-borne viruses. The sensitivity and specificity of the RT-LAMP assay were 90% (95% CI = 59.6-98.2) and 100% (95% CI = 78.5-100.0), respectively. The RT-LAMP assay detected ZIKV genome in 9 of 24 (37.5%) of the simulated clinical samples compared to 10 of 24 (41.7%) by qRT-PCR assay with a high level of concordance (κ = 0.913, P < 0.001). CONCLUSION: The RT-LAMP assay is applicable for the broad coverage detection of both the Asian and African ZIKV strains in resource-deficient settings

Autoimmunity in the pathogenesis of severe dengue / Nurhafiza Zainal

Dengue, an arboviral disease is transmitted between humans by the bite of infected female Aedes mosquitoes. Annually, about 50 million cases of dengue virus (DENV) infections were reported with approximately 500,000 cases of severe dengue and about 5% fatality rate. Understanding the mechanism leading to dengue or severe dengue is crucial, considering the fact that it is not completely comprehended. Autoimmunity has known to be involved in the pathogenesis of dengue. Here, we investigated the association of dengue and autoimmunity by comparing the genetic variant distribution among severe, non-severe dengue and systemic lupus erythematosus (SLE) patients using Ion Torrent sequencing method. Four significant variants were identified; novel variant at chr3:49679737 of BSN gene was associated with susceptibility to severe dengue, while polymorphisms rs76018112 of ABCF1 gene, rs1557370 of Mx1 gene and rs945635 of FCRL3 were associated with protective effects against severe dengue. Theoretical analysis on the effects of novel variant (chr3:49679737) suggests the importance of calcium release in the prevention of severe manifestation, whereas rs76018112, rs1557370 and rs945635 demonstrated that effective early antiviral responses and faster virus clearance associated with protective effects against severe dengue manifestation. SLE patients showed high distribution of all variants related to protection against severe dengue, suggesting that autoimmune disease patients might possess protective factors against acute dengue. We then analysed the protective factor of autoimmunity against dengue by determining the capability of SLE-sera to neutralize DENV using foci reduction neutralization test (FRNT). A total of 82 dengue serology negative sera of SLE patients were collected and FRNT against DENV was performed. Results revealed that 69%, 61% and 52% of the SLE patients’ sera showed FRNT50 neutralization against DENV1, DENV2 and DENV3, respectively. SLE-sera significantly neutralized DENV iv in comparison to healthy donors, though tested negative for dengue IgG/IgM antibodies, signifying that SLE patient might have an efficient antiviral response against DENV, perhaps via other antibody isotypes or autoreactive antibodies. We further investigated the association of dengue and autoimmunity by observing the role of high mobility group box 1 (HMGB1), a DNA-binding protein commonly linked to the progression of autoimmune diseases, in the pathogenesis of dengue. HMGB1 resides in the nucleus, but translocated out to the cytoplasm and extracellular milieu during DENV infection. HMGB1-knockdown cells showed higher DENV replication and lower interferonstimulated genes (ISGs) production than wild-type cells, proposing the novel role of HMGB1 in the antiviral response against DENV, through the regulation of innate immune response. Resveratrol (RESV) treatment inhibits the translocation of HMGB1 via Sirt1, and the accumulation of nuclear HMGB1 consequently increased production of ISGs and reduced DENV replication, verifying the role of nuclear HMGB1 in regulating ISGs in the antiviral response against DENV. Nuclear HMGB1 was found to bind to the promoter region of ISG and positively regulated the expression of ISGs. Our findings highlighted the importance of efficient early antiviral responses and protective effects of autoimmunity against severe dengue progression. We also introduce the novel role of nuclear HMGB1 in the antiviral response and RESV as an antiviral drug against DENV

Sera of patients with systemic lupus erythematosus cross-neutralizes dengue viruses

Dengue is the most prevalent mosquito-borne disease in Southeast Asia, where the incidence of systemic lupus erythematosus (SLE) is approximately 30 to 53 per 100,000. Severe dengue, however, is rarely reported among individuals with SLE. Here, whether sera of patients with SLE cross-neutralize dengue virus (DENV) was investigated. Serum samples were obtained from individuals with SLE who were dengue IgG and IgM serology negative. Neutralization assays were performed against the three major DENV serotypes. Of the dengue serology negative sera of individuals with SLE, 60%, 61% and 52% of the sera at 1/320 dilution showed more than 50% inhibition against dengue type-1 virus (DENV-1), DENV-2 and DENV-3, respectively. The neutralizing capacity of the sera was significantly greater against DENV-1 (P < 0.001) and DENV-3 (P < 0.01) than against DENV-2 (P < 0.05). Neutralization against the DENV correlated with dengue-specific IgG serum titers below the cut-off point for dengue positivity. Depletion of total IgG from the sera of patients with SLE resulted in significant decreases of up to 80% in DENV inhibition, suggesting that IgG plays an important role. However, some of the SLE sera was still able to neutralize DENV, even with IgG titers <0.1 OD absorbance. Our findings suggest that sera of patients with SLE contain IgG, and possibly other type of antibodies, that can cross-neutralize DENV, which may explain the rarity of severe dengue in individuals with SLE. Further studies, are needed to further substantiate this finding and to elucidate the specific neutralizing epitopes recognized by the sera of individuals with SLE

Draft genome of the emerging pathogen, Kocuria marina, isolated from a wild urban rat

<div><p>Kocuria marina has recently emerged as a cause for catheter-related bloodstream infections in patients with underlying health complications. One K. marina strain was recently isolated from the lung tissues of a wild urban rat (Rattus rattus diardii) caught during rodent surveillance. Here, we present the draft genome of the first K. marina animal isolate, K. marina TRE150902.</p></div