Development of a quantitative NS1-capture enzyme-linked immunosorbent assay for early detection of yellow fever virus infection

Jorge, Taissa Ricciardi; Bordignon, Juliano; Koishi, Andrea Cristine; Zanluca, Camila; Mosimann, Ana Luiza Pamplona; Santos, Claudia Nunes Duarte dos

doi:10.1038/s41598-017-16231-6

Cited by 23 publications

(25 citation statements)

References 29 publications

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“…Although frequently used for the diagnosis of DENV in countries where it is endemic, few reports have described the specific detection of NS1 antigen as a diagnostic test for YFV (previously identified in YFV studies as gp48). Ricciardi-Jorge et al described the development of a quantitative YFV NS1-capture ELISA with 80% sensitivity (12/15 cases) and 100% specificity compared to those of RT-PCR (85). NS1 concentrations ranged from 177.6 to 4,597.6 ng/ml of serum and the 3 negative samples came from patients on days 4 (n ϭ 2) and 7.…”

Section: Ns1mentioning

confidence: 99%

Yellow Fever Virus: Diagnostics for a Persistent Arboviral Threat

2018

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Yellow fever (YF) is the prototypical hemorrhagic fever and results from infection with yellow fever virus (YFV), which is endemic to regions of Africa and South America. Despite the availability of an effective vaccine, YFV continues to cause disease throughout regions where it is endemic, including intermittent large outbreaks among undervaccinated populations. A number of diagnostic methods and assays have been described for the detection of YFV infection, including viral culture, molecular testing, serology, and antigen detection. Commercial diagnostics are not widely available, and testing is generally performed at a small number of reference laboratories. The goal of this article, therefore, is to review available clinical diagnostics for YFV, which may not be familiar to many practitioners outside areas where it is endemic. Additionally, we identify gaps in our current knowledge about YF that pertain to diagnosis and describe interventions that may improve YFV detection.

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Section: Ns1mentioning

confidence: 99%

Yellow Fever Virus: Diagnostics for a Persistent Arboviral Threat

2018

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“…Zika virus (ZIKV) secretes NS1 efficiently from infected cells. Other Flaviviruses like Yellow Fever virus (YFV) also secrete NS1 albeit in very low amounts compared with DENV or ZIKV [24]. In order to compare with DENV, we tested the secretion properties of ZIKV and YFV NS1 in infected C6/36 and Vero-E6 cells.…”

Section: Resultsmentioning

confidence: 99%

The dengue virus non-structural protein 1 (NS1) is secreted from mosquito cells in association with the intracellular cholesterol transporter chaperone caveolin complex

Ramirez

Ludert

2018

Preprint

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11 12 Keywords: 13 dengue virus; zika virus; yellow fever virus; flavivirus; NS1; caveolin-1; chaperone caveolin 14 complex; viral protein trafficking; unconventional secretion; mosquito cells 15 16 17 ABSTRACT 18 Dengue virus (DENV) is a mosquito-borne virus of the family Flaviviridae. The RNA viral genome 19 encodes for a polyprotein that is co-translationally processed into three structural proteins and . CC-BY 4.0 International license is made available under aThe copyright holder for this preprint (which was not peer-reviewed) is the author/funder. It . https://doi.org/10.1101/370932 doi: bioRxiv preprint 2 20 seven non-structural proteins. The non-structural protein 1 (NS1) is a multifunctional viral protein 21 actively secreted in vertebrate and mosquito cells during DENV infection. In mosquito cells, NS1 22 is secreted in a caveolin-1 (CAV-1) dependent manner by an unconventional pathway. The 23 caveolin chaperone complex (CCC) is a cytoplasmic complex formed by caveolin-1 and the 24 chaperones FKBP52, Cy40 and CyA which is responsible for cholesterol traffic inside the cell. 25 In this work, we demonstrate that in infected mosquito cells, DENV NS1 is secreted by an early 26 and unconventional route that bypasses the Golgi apparatus in close association with the CCC.27 Treatment of mosquito cells with classic secretion inhibitors such as brefeldin A, golgicide A and 28 Fli-06 showed no effect on NS1 secretion, but significant reductions in recombinant luciferase 29 secretion and virion release. Silencing the expression of CAV1, FKBP52 with siRNAs or the 30 inhibition of CyA by cyclosporine A resulted in significant decrease in NS1 secretion without 31 affecting virion release. Using co-localization, co-inmunoprecipitation and proximity ligation 32 assays, NS1 was found to co-localize and interact with all the protein components of the CCC 33 in mosquito infected cells. In addition, CAV-1 and FKBP52 expression was found augmented in 34 DENV infected cells. Finally, the treatment of ZIKV infected mosquito cells with brefeldin A and 35 golgicide A showed no effect on NS1 secretion, while affecting virion release. ZIKV NS1 was 36 also found to co-localize with CAV-1 in infected mosquito cells. These results suggest that in 37 mosquito cells, ZIKV NS1 follows the same secretory pathway observed for DENV NS1. The 38 association of NS1 with the cholesterol transporter CCC agrees with the lipoprotein nature of 39 secreted hexameric NS1. 40 41 AUTHOR SUMMARY . CC-BY 4.0 International license is made available under a The copyright holder for this preprint (which was not peer-reviewed) is the author/funder. It . https://doi.org/10.1101/370932 doi: bioRxiv preprint 3 42 Dengue protein NS1 is secreted in infected mosquito and vertebrate cells. In humans, secreted 43 NS1 have been associated with pathogenesis. In mosquito cells, NS1 follows an unconventional 44 secretion pathway that is dependent on Caveolin-1. This work shows that in mosquito cells, NS1 45 secretion is associated to the chaperone caveolin complex, a...

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“…Another interesting observation is that the CBD is fully conserved in DENV and ZIKV NS1, while incomplete in other flaviviruses such as YFV, JEV and WNV. Secretion of YFV NS1 have been observed in mosquito cells albeit a higher concentration was maintained as cell-associated rather than secreted into the extracellular milieu (28); moreover, no NS1 secretion is observed in WNV or JEV infected mosquito cells (29, 30). Thus, the presence of an active CBD and the recognition of NS1 by the mosquito caveolin may be crucial for NS1 secretion in the mosquito, and suggest unique roles for the DENV and ZIKV soluble NS1 in the mosquito (11, 21).…”

Section: Discussionmentioning

confidence: 99%

The secretory fate of flavivirus NS1 in mosquito cells is influenced by the caveolin binding domain

Ramirez

Ludert

2019

Preprint

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14Flaviviruses of major medical importance worldwide such as dengue (DENV), Zika (ZIKV), and 15 yellow fever (YFV) viruses are transmitted by mosquitoes Aedes sp. The non-structural protein 16 1 (NS1) of these flaviviruses is secreted from the infected cells using different secretion routes 17 depending on the cell and virus nature. The NS1 of DENV and ZIKV contain in the hydrophobic 18 region a conserved caveolin binding domain (CBD) (XXXXXX), which is not conserved in 19 YFV NS1. To ascertain the role of the CBD in the secretory route followed by flavivirus NS1, 20 expression vectors for the NS1 of DENV2, ZIKV and YFV were constructed. Using site-directed 21 mutagenesis, substitutions were made in the aromatic residues within CBD; in addition, the full 22 domain was replaced by those of other flaviviruses, creating chimeras in the CBD of NS1. 23 Substitutions of the aromatic residues to Ala or Thr, or CBD chimeras, results in increased 24 sensitivity of NS1 secretion to brefeldin A treatment, indicating a change to a classical secretion 25 pathway. Likewise, the insertion of the DENV/ZIKV CBD into the recombinant Gaussia-26 130 Gaussia Luc in DENV2-NS1. This insertion was introduced only in the DENV2-NS1 construct. 5.131 General experimental strategy with constructs. All mutations, insertions and chimera constructions 132 were transfected in C6/36 cells and NS1 secretion to the supernatants in cells treated or not with BFA 133 determined by ELISA. 134 135 136 441 2015-1 254461 to JEL. Authors declare no conflict of interest. 442 443 444

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Development of a quantitative NS1-capture enzyme-linked immunosorbent assay for early detection of yellow fever virus infection

Cited by 23 publications

References 29 publications

Yellow Fever Virus: Diagnostics for a Persistent Arboviral Threat

Yellow Fever Virus: Diagnostics for a Persistent Arboviral Threat

The dengue virus non-structural protein 1 (NS1) is secreted from mosquito cells in association with the intracellular cholesterol transporter chaperone caveolin complex

The secretory fate of flavivirus NS1 in mosquito cells is influenced by the caveolin binding domain

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