Sains Malaysiana 50(10)(2021): 3035-3043

http://doi.org/10.17576/jsm-2021-5010-16

Dengue Virus Non-structural (NS) 1 Gene as A Molecular Marker for Early Detection of in vitro Dengue Virus Infection

(Gen Virus Denggi Bukan Struktur (NS) 1 sebagai Penanda Molekul untuk Pengesanan Awal Jangkitan Virus Denggi secara in vitro)

NUR AZIZAH A RAHMAN^1,2, FADHILAH MOH DJAMIL^1,4, VINOD RMT BALASUBRAMANIAM³, SHARIFAH SYED HASSAN³ & WEI BOON YAP^1,2*

¹Biomedical Science Programme, Faculty of Health Sciences, Universiti Kebangsaan Malaysia, Jalan Raja Muda Abdul Aziz, 50300 Kuala Lumpur, Federal Territory, Malaysia

²Center of Toxicology and Health Risk Studies, Faculty of Health Sciences, Universiti Kebangsaan Malaysia, Jalan Raja Muda Abdul Aziz, 50300 Kuala Lumpur, Federal Territory, Malaysia

³Jeffrey Cheah School of Medicine & Health Sciences, Monash University Malaysia, Jalan Lagoon Selatan, 46150 Bandar Sunway, Selangor Darul Ehsan, Malaysia

⁴Microbiology Unit, Department of Pathology, Tuanku Ja’afar Hospital, Jalan Rasah, Bukit Rasah, 70300 Seremban, Negeri Sembilan Darul Khusus, Malaysia

Diserahkan: 26 Oktober 2020/Diterima: 19 Februari 2021

ABSTRACT

Serology-based dengue assays at times produce inaccurate results especially in the early phase of disease onset. A more precise diagnostic approach detecting dengue infections in the early phase enables better management of the disease. This helps reduce dengue-associated morbidity and mortality. Besides, an early diagnosis of dengue is also very beneficial in a dengue outbreak and in endemic regions. In this light, this study aimed to determine the potential of the dengue virus (DENV) non-structural 1 (NS1) gene as an early detection biomarker. The cytopathic effects (CPE) were monitored and the cell death of DENV serotype-2 (DENV2)-infected Vero cells was evaluated for fourteen consecutive days. Only Lemos and in-house NS1-specific primer pairs showed positive amplifications in the preliminary primer validation. Thus, both of the primer pairs were then used to amplify the NS1 gene from the infected cells. The NS1 gene was detected as early as day-2 post-infection using the in-house primers. There was no amplicon produced using the Lemos primers. This is speculated to be attributable to the relatively lower complementarity of the primer sequences with that of the template and low amount of viral mRNA in the DENV2-infected cells. Conclusively, the DENV NS1 gene is a potential early detection marker, however, the NS1-specific primers should be pre-validated to ensure a reliable dengue diagnosis.

Keywords: Dengue; dengue virus (DENV); early diagnosis; non-structural (NS)1; polymerase chain reaction (PCR)

ABSTRAK

Ujian denggi berdasarkan serologi kadang-kadang memberikan hasil yang tidak tepat terutama pada fasa awal permulaan penyakit. Pendekatan diagnostik yang lebih tepat untuk mengesan jangkitan denggi pada fasa awal membolehkan pengurusan penyakit ini dengan lebih baik. Ini membantu mengurangkan morbiditi dan kematian yang berkaitan dengan denggi. Selain itu, diagnosis denggi awal juga sangat bermanfaat dalam wabak denggi dan di daerah endemik. Kajian ini bertujuan untuk menentukan potensi gen bukan struktur 1 (NS1) virus denggi sebagai biopenanda pengesanan awal. Kesan sitopatik (CPE) dipantau dan kematian sel Vero yang dijangkiti serotip-2 (DENV2) dinilai selama empat belas hari berturut-turut. Hanya pasangan primer khusus Lemos dan NS1 dalaman yang menunjukkan peningkatan positif dalam pengesahan primer awal. Oleh itu, kedua-dua pasangan primer kemudian digunakan untuk memperkuat gen NS1 daripada sel yang dijangkiti. Gen NS1 dikesan pada awal hari ke-2 selepas jangkitan menggunakan primer dalaman. Tidak ada amplikon yang dihasilkan menggunakan primer Lemos. Ini diduga disebabkan oleh pelengkap urutan primer yang lebih rendah dengan templat dan jumlah mRNA virus yang rendah pada sel yang dijangkiti DENV2. Kesimpulannya, gen DENV NS1 adalah penanda pengesanan awal yang berpotensi, namun, primer khusus NS1 harus disahkan terlebih dahulu untuk memastikan diagnosis denggi yang boleh dipercayai.

Kata kunci: Bukan struktur (NS)1; diagnosis awal; denggi; tindak balas rantai polimerase (PCR); virus denggi (DENV)

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*Pengarang untuk surat-menyurat; email: yapweiboon@ukm.edu.my