Multiplex real–time RT–PCR for detecting chikungunya virus and dengue virus

Pongsiri, Piyathida; Praianantathavorn, Kesmanee; Theamboonlers, Apiradee; Payungporn, Sunchai; Poovorawan, Yong

doi:10.1016/s1995-7645(12)60055-8

Cited by 48 publications

(45 citation statements)

References 19 publications

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“…According to this author, a level of larval infestation comprised between 2% and 5% is enough to keep dengue endemic and to trigger small-scale outbreaks. In the present study, BI was 1.66 during DS and as high as 4.83 for the wet period, resulting in an average of 3.24. Referring to Vanzie [74][75][76][77][78][79] , it appears clear that the annual and WS's BI obtained in Makkah City are within the range believed to require attention, as they indicate increased population maintenance and are adequate to cause outbreaks.…”

Section: Discussionmentioning

confidence: 99%

Household survey of container–breeding mosquitoes and climatic factors influencing the prevalence of Aedes aegypti (Diptera: Culicidae) in Makkah City, Saudi Arabia

Aziz

Dieng

Ahmad

et al. 2012

Asian Pacific Journal of Tropical Biomedicine

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Section: Discussionmentioning

confidence: 99%

Household survey of container–breeding mosquitoes and climatic factors influencing the prevalence of Aedes aegypti (Diptera: Culicidae) in Makkah City, Saudi Arabia

Aziz

Dieng

Ahmad

et al. 2012

Asian Pacific Journal of Tropical Biomedicine

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“…According to the manufacturer, the possibility of cross-reactivity of the Altona assay with Usutu virus, another flavivirus, cannot be ruled out due to sequence homology with the target region used for the detection of ZIKV RNA (18); even when the Altona PCR was performed on only 4 serum specimens, it did not show any cross-reactivity with specimens confirmed to be positive for DENV RNA by a verified PCR assay in use in PHOL, which provides additional data on the specificity of the assay (19). Although the number of urine specimens available for this study was small, the Altona PCR performed as well as the reference PCR with this specimen type.…”

Section: Discussionmentioning

confidence: 99%

Evaluation of Altona Diagnostics RealStar Zika Virus Reverse Transcription-PCR Test Kit for Zika Virus PCR Testing

et al. 2017

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With the emerging Zika virus (ZIKV) epidemic, accessible real-time reverse transcription-PCR (rRT-PCR) assays are needed to streamline testing. The commercial Altona Diagnostics RealStar ZIKV rRT-PCR test kit (Altona PCR) has been approved for emergency use authorization by the U.S. FDA. Our aim was to verify the Altona PCR by comparing it to the CDC-designed dual-target ZIKV rRT-PCR reference assay (reference PCR) and describe the demographics of patients tested for ZIKV by rRT-PCR in Ontario, Canada. A large set of clinical specimens was tested for ZIKV by the Altona PCR and the reference PCR. Positive or equivocal specimens underwent PCR and Sanger sequencing targeting the ZIKV NS5 gene. A total of 671 serum specimens were tested by the reference PCR: 58 (8.6%) were positive, 193 (28.8%) were equivocal, and 420 (62.6%) were negative. Ninety percent of the reference PCR-positive patients were tested in the first 5 days after symptom onset. The Altona PCR was performed on 284/671 specimens tested by the reference PCR. The Altona PCR was positive for 53/58 (91%) reference PCR-positive specimens and 16/193 (8%) reference PCR-equivocal specimens; the ZIKV NS5 PCR was positive for all 68 Altona PCR-positive specimens and negative for all 181 Altona PCR-negative specimens that underwent the NS5 PCR. The Altona PCR has very good sensitivity (91%) and specificity (97%) compared to the reference PCR. The Altona PCR can be used for ZIKV diagnostic testing and has less extensive verification requirements than a laboratory-developed test.

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“…However, only three groups have described single-reaction real-time assays for DENV and CHIKV [23,26,27], whereas others reported modified conventional RT-PCRs that require gel electrophoresis for detection [24,25,28]. All multiplex rRT-PCRs appear to perform well, as reported, though most primer and probe sequences contain mismatches when aligned to consensus DENV and CHIKV sequences that include contemporary viruses obtained globally (see materials and methods).…”

Section: Discussionmentioning

confidence: 99%

Clinical evaluation of a single-reaction real-time RT-PCR for pan-dengue and chikungunya virus detection

Waggoner

Ballesteros²,

Gresh

et al. 2016

Journal of Clinical Virology

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Background Dengue virus (DENV) and chikungunya virus (CHIKV) now co-circulate throughout tropical regions of the world, with billions of people living at risk of infection. The differentiation of these infections is important for epidemiologic surveillance as well as clinical care, though widely-used molecular diagnostics for DENV and CHIKV require the performance of two to four separate PCR reactions for detection. Objectives In the current study, we sought to develop and evaluate a single-reaction, multiplex real-time RT-PCR (rRT-PCR) for the detection and differentiation of DENV and CHIKV (the pan-DENV–CHIKV rRT-PCR). Study design From an alignment of all available CHIKV complete genome sequences in GenBank, a new CHIKV rRT-PCR was designed for use in multiplex with a previously described assay for pan-DENV detection. Analytical evaluation was performed in accordance with published recommendations, and the pan-DENV–CHIKV rRT-PCR was clinically compared to reference molecular diagnostics for DENV and CHIKV using 182 serum samples from suspected cases in Managua, Nicaragua. Results The pan-DENV–CHIKV rRT-PCR had a dynamic range extending from 7.0 to 2.0 log10 copies/μL for each DENV serotype and CHIKV, and the lower limits of 95% detection were 7.9–37.4 copies/μL. The pan-DENV–CHIKV rRT-PCR detected DENV in 81 patients compared to 75 using a reference, hemi-nested DENV RT-PCR, and it demonstrated perfect agreement with a reference CHIKV rRT-PCR (54 positive samples). Conclusions The single-reaction, multiplex format of the pan-DENV–CHIKV rRT-PCR, combined with sensitive detection of both viruses, has the potential to improve detection while decreasing testing costs and streamlining molecular workflow.

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Multiplex real–time RT–PCR for detecting chikungunya virus and dengue virus

Abstract: This rapid and reliable assay provides a means for simultaneous early diagnosis of CHIKV and DENV in a single-step reaction.

Cited by 48 publications

References 19 publications

Household survey of container–breeding mosquitoes and climatic factors influencing the prevalence of Aedes aegypti (Diptera: Culicidae) in Makkah City, Saudi Arabia

Household survey of container–breeding mosquitoes and climatic factors influencing the prevalence of Aedes aegypti (Diptera: Culicidae) in Makkah City, Saudi Arabia

Evaluation of Altona Diagnostics RealStar Zika Virus Reverse Transcription-PCR Test Kit for Zika Virus PCR Testing

Clinical evaluation of a single-reaction real-time RT-PCR for pan-dengue and chikungunya virus detection

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