Dataset Information

Efficient Plant Production of Recombinant NS1 Protein for Diagnosis of Dengue.

ABSTRACT: Dengue fever is endemic in more than 120 countries, which account for 3.9 billion people at risk of infection worldwide. The absence of a vaccine with effective protection against the four serotypes of this virus makes differential molecular diagnosis the key step for the correct treatment of the disease. Rapid and efficient diagnosis prevents progression to a more severe stage of this disease. Currently, the limiting factor in the manufacture of dengue (DENV) diagnostic kits is the lack of large-scale production of the non-structural 1 (NS1) protein (antigen) to be used in the capture of antibodies from the blood serum of infected patients. In this work, we use plant biotechnology and genetic engineering as tools for the study of protein production for research and commercial purposes. Gene transfer, integration and expression in plants is a valid strategy for obtaining large-scale and low-cost heterologous protein production. The authors produced NS1 protein of the dengue virus serotype 2 (NS1DENV2) in the Arabidopsis thaliana plant. Transgenic plants obtained by genetic transformation expressed the recombinant protein that was purified and characterized for diagnostic use. The yield was 203 ?g of the recombinant protein per gram of fresh leaf. By in situ immunolocalization, transgenic protein was observed within the plant tissue, located in aggregates bodies. These antigens showed high sensitivity and specificity to both IgM (84.29% and 91.43%, respectively) and IgG (83.08% and 87.69%, respectively). The study goes a step further to validate the use of plants as a strategy for obtaining large-scale and efficient protein production to be used in dengue virus diagnostic tests.

SUBMITTER: Xisto MF

PROVIDER: S-EPMC7649140 | biostudies-literature | 2020

REPOSITORIES: biostudies-literature

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Efficient Plant Production of Recombinant NS1 Protein for Diagnosis of Dengue.

Xisto Mariana Fonseca MF Dias Roberto Sousa RS Feitosa-Araujo Elias E Prates John Willians Oliveira JWO da Silva Cynthia Canedo CC de Paula Sérgio Oliveira SO

Frontiers in plant science 20201026

Dengue fever is endemic in more than 120 countries, which account for 3.9 billion people at risk of infection worldwide. The absence of a vaccine with effective protection against the four serotypes of this virus makes differential molecular diagnosis the key step for the correct treatment of the disease. Rapid and efficient diagnosis prevents progression to a more severe stage of this disease. Currently, the limiting factor in the manufacture of dengue (DENV) diagnostic kits is the lack of larg ...[more]

PMID: 33193526

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Project description:UNLABELLED: / BACKGROUND: Dengue is the most important arthropod borne viral disease worldwide in terms of morbidity and mortality and is caused by any of the four serotypes of dengue virus (DENV-1 to 4). Brazil is responsible for approximately 80% of dengue cases in the Americas, and since the introduction of dengue in 1986, a total of 5,944,270 cases have been reported including 21,596 dengue hemorrhagic fever and 874 fatal cases. DENV can infect many cell types and cause diverse clinical and pathological effects. The goal of the study was to investigate the usefulness of NS1 capture tests as an alternative tool to detect DENV in tissue specimens from previously confirmed dengue fatal cases (n = 23) that occurred in 2002 in Brazil. METHODOLOGY/PRINCIPAL FINDINGS: A total of 74 tissue specimens were available: liver (n = 23), lung (n = 14), kidney (n = 04), brain (n = 10), heart (n = 02), skin (n = 01), spleen (n = 15), thymus (n = 03) and lymph nodes (n = 02). We evaluated three tests for NS1 antigen capture: first generation Dengue Early ELISA (PanBio Diagnostics), Platelia NS1 (BioRad Laboratories) and the rapid test NS1 Ag Strip (BioRad Laboratories). The overall dengue fatal case diagnosis based on the tissues analyzed by Dengue Early ELISA, Platelia NS1 and the NS1 Ag Strip was 34.7% (08/23), 60.8% (14/23) and 91.3% (21/23), respectively. The Dengue Early ELISA detected NS1 in 22.9% (17/74) of the specimens analyzed and the Platelia NS1 in 45.9% (34/74). The highest sensitivity (78.3%; 58/74) was achieved by the NS1 Ag Strip, and the differences in the sensitivities were statistically significant (p<0.05). The NS1 Ag Strip was the most sensitive in liver (91.3%; 21/23), lung (71.4%; 10/14), kidney (100%; 4/4), brain (80%; 8/10), spleen (66.6%, 10/15) and thymus (100%, 3/3) when compared to the other two ELISA assays. CONCLUSIONS/SIGNIFICANCE: This study shows the DENV NS1 capture assay as a rapid and valuable approach to postmortem dengue confirmation. With an increasing number of DHF and fatal cases, the availability of new approaches useful for cases confirmation plays an important tool for the disease surveillance.

Project description:Purpose of Review:The requirement for large quantities of therapeutic proteins has fueled a great interest in the production of recombinant proteins in plant bioreactors. The vaccines and bio-therapeutic protein production in plants hold the promise of significantly lowering the cost of manufacturing life-saving drugs. This review will reflect the current status and challenges that the molecular farming platform faces becoming a strategic solution for the development of low-cost bio-therapeutics for developing countries. Recent Findings:Different plant parts have been successfully identified as suitable expression systems for the commercial production of therapeutic proteins for some human and animal diseases ranging from common cold to AIDS. The processed therapeutics from such sources are devoid of any toxic components. The large-scale cultivation of these transgenic plants would be possible anywhere in the world including developing countries, which lack sophisticated drug manufacturing units. A couple of such commercially generated products have already hit the market with success. Newer methods using suitable plant viruses and recombinant gene expression systems have already been devised for producing therapeutic proteins and peptides. Summary:Plants are promising bio-factories for therapeutic protein production because of their several advantages over the other expression systems especially the advanced mechanisms for protein synthesis and post-translational modification which are very much similar to animal cells. Plant biotechnologists are much attracted to the bio-farming because of its flexibility, scalability, low manufacturing cost, as well as the lack of risk of toxic or pathogenic contamination. A number of projects on bio-farming are designed and are at various developmental stages but have not yet become available to the pharmaceutical industry. Therefore, we need further advancement in the optimization of lab protocols for up-scaling the production of such therapeutics at commercial level with a promise to offer their best clinical use.

Dataset Information

Efficient Plant Production of Recombinant NS1 Protein for Diagnosis of Dengue.

Publications

Efficient Plant Production of Recombinant NS1 Protein for Diagnosis of Dengue.

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