Detection and Quantification of Chikungunya Virus by Real-Time RT-PCR Assay

Chikungunya virus (CHIKV) re-emerged as a globalized health threat fifteen years ago. There are dozens of RT-PCR assays published. An inventory of the latter was made, and after in silico analysis, two assays were selected for their ability to detect strains belonging to the five CHIKV genetic lineages. They were combined in order to provide a robust assay not affected by genetic point mutations and the resulting Duo CHIKV real-time RT-PCR assay was compared to the two parental single-plex tests against five strains belonging to the five genetic lineages. The Duo CHIKV assay performed equally, or better, in terms of sensitivity, specificity, linearity and signal intensity. Dual-target assays are better suited for viruses having the propensity to evolve into new variants via point mutations or major sequence deletions/insertions. Here, we demonstrated that combining two single systems into a dual-target assay did not impair sensitivity and specificity, and proved a potent diagnostic tool to face a potential emergence of CHIKV variants by newly evolving mutations.

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Establishment of Real-Time TaqMan-Fluorescence Quantitative RT-PCR Assay for Detection and Quantification of Porcine Lipoprotein Lipase mRNA

Agricultural Sciences in China ◽

10.1016/s1671-2927(08)60336-3 ◽

2009 ◽

Vol 8 (10) ◽

pp. 1256-1262 ◽

Cited By ~ 2

Author(s):

Hong-xia LIAN ◽

De-xun LU ◽

Min GAO

Keyword(s):

Real Time ◽

Lipoprotein Lipase ◽

Rt Pcr ◽

Pcr Assay ◽

Detection And Quantification

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A35 The first laboratory confirmation of chikungunya outbreak in Ethiopia

Virus Evolution ◽

10.1093/ve/vez002.034 ◽

2019 ◽

Vol 5 (Supplement_1) ◽

Author(s):

Mesfin Mengesha Tsegaye ◽

Aadamu Tayachew ◽

Desalegn Belay ◽

Abebe Alemu ◽

Berhane Beyene

Keyword(s):

Nucleic Acid ◽

Real Time ◽

Laboratory Investigation ◽

Emergency Preparedness ◽

Zika Virus ◽

Chikungunya Virus ◽

Febrile Illness ◽

Rt Pcr ◽

Pcr Assay ◽

Serum Samples

Abstract Chikungunya is a viral disease (genus Alphavirus) which is transmitted to humans by infected mosquitoes—including Aedes aegypti and A. albopictus. An outbreak of febrile illness, suspected to have been caused by chikungunya, was reported in June 2016 from Dolloado district, Suuf Kebele, in the Somalia regional state of Ethiopia that borders the Mandera county of Kenya where a confirmed chikungunya outbreak was ongoing. Laboratory investigation was carried out to confirm if the outbreak in Ethiopia was caused by Chikungunya virus. Ten serum samples were collected from suspected patients visiting a health center in Suuf Kebel, who were then sent to the Nation laboratory in Ethiopian Public Health Institute. RNA was extracted from the serum samples using QIAgene RNA Mini kit, and PCR detection of dengue, chikungunya, and Zika virus nucleic acid was done using Trioplex Real-time RT-PCR Assay following the protocol from the Center for Disease Control (CDC). The Trioplex Real-time RT-PCR assay, for detection and differentiation of RNA from dengue, Chikungunya and Zika, was provided by CDC as part of the zika emergency preparedness effort. Of the nine samples tested, eight (88.88%) were found to be positive for chikungunya virus nucleic acid but negative for dengue and Zika virus nucleic acids. The median age of the affected sampled patients was 40 years, and males appear to be more affected (66.6% of sampled patients). The laboratory investigation confirmed that the outbreak was caused by chikungunya virus. Even though further molecular characterization of the positive isolates will provide more information as to the circulating genotypes and elucidate the origin of the outbreak virus, it is also possible to assume that the outbreak was an extension of the outbreak in neighboring countries in Kenya and, therefore, warrants that cross-border integration efforts to control chikungunya should be implemented by the concerned countries.

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