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All Journal Journal of Tropical Life Science : International Journal of Theoretical, Experimental, and Applied Life Sciences Microbiology Indonesia Biotropika Journal of Biomedicine and Translational Research Bioma : Jurnal Biologi Indonesia Global Health Management Journal Acta Biochimica Indonesiana Majalah Kedokteran Andalas Medika Kartika : Jurnal Kedokteran dan Kesehatan Indonesian Journal of Clinical Pathology and Medical Laboratory (IJCPML) Tunas Medika Jurnal Kedokteran & Kesehatan JKKI : Jurnal Kedokteran dan Kesehatan Indonesia JIMKI: Jurnal Ilmiah Mahasiswa Kedokteran Indonesia

Beti Ernawati Dewi

Departemen Mikrobiologi Fakultas Kedokteran Universitas Indonesia

Author-ID : 242224

Agriculture, Biological Sciences & Forestry Biochemistry, Genetics & Molecular Biology Dentistry Education Environmental Science Health Professions Immunology & microbiology Medicine & Pharmacology Neuroscience Nursing Public Health Veterinary

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Cloning and Expression of Nonstructural Protein NS1 of Dengue Virus Serotype 2 BETI ERNAWATI DEWI; FITHRIYAH FITHRIYAH; ANDRIANSJAH RUKMANA; PAISAL PAISAL; DEKA LARASATI; TJAHJANI MIRAWATI SUDIRO
Microbiology Indonesia Vol. 6 No. 1 (2012): March 2012
Publisher : Indonesian Society for microbiology

Early diagnosis of dengue virus (DENV) infection is affirmative for patient management and control of the disease. Detection of nonstructural-1 (NS1) antigen has been proven to provide early detection of DENV infection. Commercial NS1 antigen assays are available in Indonesia with variable sensitivity. In an attempt to develop an NS1-based diagnostic test, we successfully cloned NS1 gene of DENV2 to a glutathione Stransferase- based vector pGEX6P-1 in Escherichia coli system. The recombinant protein (pG2NS12) was expressed in E. coli BL21. After induction with isopropyl-β-D-thiogalactoside 0.1 mM for 4 h at 25 °C a recombinant protein GST-NS1 with molecular size of approximately 75 kDa was obtained. The fusion protein was insoluble and found in the pellet fraction of the cell lysate. Addition of lysozyme (10 mg mL-1) and DNase-I (7.2 mg mL-1) in the lysis buffer was necessary to collect proteins from the pellet fraction. The proteins in the cell pellet were fractionated through Sephadex-G100 column, and GST-NS1 was further purified with Glutathione-Sepharose 4B beads. To obtain pure recombinant NS1 protein to be used in the immunization of mice, the fusion protein was cut with PreScission Protease® by addition of 0.075% Triton-X 100 was necessary to cut the fusion protein. We found that antibodies that recognized the recombinant NS1 protein and DENV2 virus were produced in mice immunized with purified NS1 protein. Therefore, our recombinant NS1 could be used to produce antibody that is potentially useful for developing diagnostic assay to determine the presence of dengue virus NS1 antigen in patient sera.

Five Unique Amino Acid Residues of Hemagglutinin (HA) Proteins of Swine Influenza A (H1N1) Detected in 2009 in Jakarta, Indonesia ANDI YASMON; YULIANTY MUHAYAR; VIVI SETIAWATY; BETI ERNAWATI DEWI; BUDIMAN BELA; FERA IBRAHIM
Microbiology Indonesia Vol. 6 No. 2 (2012): June 2012
Publisher : Indonesian Society for microbiology

Nine HA genes of influenza A (H1N1) viruses originating from swine which were detected in 2009 in Jakarta, Indonesia, were characterized in this study. Nasopharyngeal and/or pharyngeal samples were extracted to obtain viral RNA genomes. Amplification of the HA segment was performed by using the reverse transcription-polymerase chain reaction (RT-PCR), and followed by nested PCR in cases of RT-PCR negative. DNA sequencing was performed by using eight overlapping primers. All the Jakarta strains were closely related to vaccine strain A/California/07/2009. Nine amino acid changes were found in the Jakarta strains, and 5 (P100S, S220T, G239D, R240Q, and I338V) of those were unique to all Jakarta strains with respect to strain A/California/07/2009 used to produce vaccine. An I338V substitution was detected in a cleavage site of HA and no amino acid changes were detected in potential sites for N-linked glycosylation. For seven sites (positions 131, 158, 160, 183, 187, 222, and 227) playing an important role in viral attachment to host receptor, none of the expected amino acid changes was detected; however, a S220T substitution close to amino acid 222 was found in all the Jakarta strains. All amino acid changes potentially affect the pathogenicity of the viruses and the efficacy of strain A/California/07/2009 in neutralizing the Jakarta strains.

Levels of TNF-Î± in PBMC (Peripheral Blood Mononuclear Cells) Induced by Recombinant Non Structural 1 Protein of Dengue Virus Serotype-2 in vitro FITHRIYAH SJATHA; OKTIVIA CHANDRA MUSTIKA; BETI ERNAWATI DEWI; TJAHJANI MIRAWATI SUDIRO
Microbiology Indonesia Vol. 13 No. 2 (2019): June 2019
Publisher : Indonesian Society for microbiology

Dengue infection is a global health problem with an increasing incidence every year and now endemic in more than 100 WHO countries. Dengue infection is caused by dengue virus (DENV) which is an RNA virus with positive single strand, with Â±11kb genome size encoding 3 structural proteins, 7 non-structural proteins, and two Untranslated Region (UTR). NS1 protein is known to have a very important role in the development of severe DENV infection, by the direct effect causing host cells damage and indirect effect by activating immune response to induce the secretion of excess cytokines. This study aims to evaluate whether recombinant pcNS1 plasmids which have been proven expressing recombinant NS1 proteins in previous studies is able to induce cytokine secretion from Peripheral Blood Mononuclear Cells (PBMC). Transfected Chinese Hamster Ovary-K1 (CHO-K1) cells with recombinant pcNS1 plasmid was co-cultured with PBMC from healthy donor. After 48 h post co-cultured, cell supernatant was collected and TNF-Î± levels and NS1 recombinant were measured by ELISA. The results showed that recombinant NS1 protein was expressed in CHO-K1 mammalian cell line and able to induce TNF-Î± with higher levels compared to control.

Levels of CXCL10 Chemokine in Dengue Infected Hepatocyte Huh 7 it-1 Cell Line Co-cultured with Peripheral Blood Mononuclear Cells BETI ERNAWATI DEWI; EVA DAMAYANTI; TJAHJANI MIRAWATI SUDIRO; AGUS SYAHRURACHMAN
Microbiology Indonesia Vol. 13 No. 3 (2019): September 2019
Publisher : Indonesian Society for microbiology

Dengue is a mosquito borne virus that spreads rapidly in the world. At present, it is estimated that more than 3.9 billion people are at risk of being infected with dengue virus (DENV) and there are 96 million clinical cases that have been reported annually in 128 countries worldwide. In DENV infected patients often associated with liver dysfunction which hepatocyte and kuppfer cells as the main target of viral infections. DENV infection induced the expression of several chemokines, which might play an important role during the inflammatory response and pathogenesis of a disease. CXCL10 is known as a chemokine that activates lymphocytes for innate and adaptive immunity, induces tissue damage, and modulates tumor formation. Therefore, we conducted an in vitro study using Huh 7it-1 cells co-cultured with peripheral blood mononuclear cells (PBMCs) to investigate CXCL10 chemokine induction during DENV infection. Huh 7it-1 cells were grown on 96 micro well plate until a monolayer was formed. The cells were infected with DENV-2 at an MOI of 0.5 FFU/cell and 1 FFU/cell in the presence of PBMCs. Heat inactivated DENV-2 and Huh7 cell medium were used as control. After 2 hours of infection, cells were co-cultured with PBMCs and incubated at 37 ÂºC with 5% CO 2 for 48 h. Cell supernatant was collected and CXCL10 chemokine levels were measured using CXCL10 Quantikine ELISA Kit. Statistical analysis was performed by SPSS 23. In the presence of PBMCs, CXCL10 levels from DENV infected Huh 7it-1 at an MOI of 0,5 FFU/cell and MOI of 1 FFU/cell were 552,653 Â± 22,779 pg Â mL-1 and 576,787 Â± 16,901 pg Â mL-1 . Those levels were higher when compared with supernatan from heat inactivated DENV-2 and control cells. Without PBMCs, all of treatments showed lower level of CXCL10. DENV-2 infection in Huh 7it-1 cells co-cultured with PBMCs was able to induce CXCL10 secretion. Furthermore, heat inactivated DENV-2 also still capable to inducen the secretion of CXCL10 chemokine in Huh 7it-1 cells.

IgG subclasses identification of immunized mice sera with Dengue tetravalent DNA vaccine based on prM-E genes: Identifikasi Subkelas IgG Dari Mencit yang Diimunisasi dengan Kandidat Vaksin DNA Dengue Tetravalent Beti Ernawati Dewi; Rizka Andhitia Mentari Putri; Tjahjani Mirawati Sudiro; Fitriyah Sjahta
Microbiology Indonesia Vol. 16 No. 2 (2022): December
Publisher : Indonesian Society for microbiology

Background: Dengue fever is still a serious health problem in the world. DENV consists of 11 kb of single positive-stranded RNA encoding three structural proteins and seven non-structural proteins. PrM and E proteins are the main targets of the antibody response that rich of epitopes and able to induce protective immunity. There are four DENV serotypes that have similar antigenic structures in the amino acid sequence of protein E. In our previous study, we successfully constructed a recombinant tetravalent DNA vaccine candidate consisting pUMVC4a-based expression plasmid for prM-E protein of all DENV serotypes (pUMD1, pUMD2, pUMD3 and pUMD4). It has been proved that the vaccine candidate was able to induced anti-dengue IgG as well as neutralization antibody to all DENV serotypes. This study aims to determine IgG subclasses of immunized mice with recombinant tetravalent DNA vaccine candidates based on prM-E genes of all serotypes. Methods: Mice (Balb/c) were immunized with a dose of 100 Î¼g 100 uL/mouse in triplicate, at three weeks interval. Blood was drawn two weeks post immunization as well as termination blood. IgG subclasses titre were measured using in-house indirect ELISA. Results: The titer of IgG2a subclass was the highest levels with optical density of 1.004Â±0.154 followed by IgG1,IgG2b, and IgG3 to DENV-2, respectively. Conclusion: The data demonstrate the humoral immune response IgG subclasses of this recombinant tetravalent DNA vaccine candidates based on prM-E genes of all serotypes, supporting further translational studies to advance the development of this candidate in response to DENV infection. Keywords: dengue vaccine, DNA vaccine, recombinant, IgG subclass, Tetravalent

Co-Authors . Andriansjah Agus Syahrurachman Agustin Iskandar Aldo Ferly Andi Yasmon Apriyanto, Dadan Ramadhan Aryati Aryati Budiman Bela Chie Aoki-Utsubo Cita Christine Mayorita DEKA LARASATI Desti, Hidayati Dewi Wulandari Dewi Wulandari Dewijanti, Indah Dwiatmi Eduardus Gilang Putra Eleanor Louana Urfa Elisabeth D. Harahap EVA DAMAYANTI Fera Ibrahim Fithriyah Fitriyah Sjahta Hak Hotta Hardi Darmawan Heni Pujiastuti Herdiman T. Pohan Kartika, Aisya Alma Asmiranti Leonard Nainggolan Lia Meilawati Linlin Haeni Marissa Angelina Mirawati Sudiro Nadia Tita Indriasti Nikki Aldi Massardi Nugraheni, Enny Oktania Sandra Puspita OKTIVIA CHANDRA MUSTIKA Pramita G. Dwipoerwantoro Putra, Eduardus Gilang Rahajuningsih Dharma Rama Samara Brajawikalpa Rayasari, Husnaya Rianto Setiabudy Rizka Andhitia Mentari Putri Safrullah, Muhamad Iqbal Santi, Mei Ria Settrin Chenderawasi SJATHA, FITHRIYAH Soemanadi Soemanadi Sri Hartati Sri Hartati Suhendro Suhendro Syamsiar, Syamsiar T. Mirawati Sudiro Tjahjani Mirawati Sudiro TJAHJANI MIRAWATI SUDIRO Tjahjani Mirawati Sudiro VIVI SETIAWATY Yulianti, Selsa YULIANTY MUHAYAR Zanjabila, Sabighoh

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