Development and validation of the first HBV qRT-PCR assay in the Mediterranean area targeting the X region

Madihi, Salma; Laassili, Chaimaa; Boukaira, Samia; Baha, Warda; Khyatti, Meriem; Zyad, Abdelmajid; Ben Mkaddem, Sanae; Benani, Abdelouaheb

doi:10.1016/j.jviromet.2024.114913

Journal of Virological Methods

2024

DOI: 10.1016/j.jviromet.2024.114913

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Development and validation of the first HBV qRT-PCR assay in the Mediterranean area targeting the X region

Salma Madihi,

Chaimaa Laassili,

Samia Boukaira

et al.

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2024

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Sensitive and Accurate Quantification of Enterovirus-D68 (EV-D68) Viral Loads Using Droplet Digital PCR (ddPCR)

Grizer,

Li,

Mattapallil

2024

Microorganisms

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Enterovirus-D68 (EV-D68) is a reemerging virus that has been associated with numerous outbreaks in children in the past 10 years. Most assays examining viral infection kinetics have relied on the use of quantitative RT-PCR (qRT-PCR) assays as an assay of choice. Though valuable, there are inherent limitations that introduce variability, thereby reducing its value when comparing results across the field. Unlike the qRT-PCR assay that uses a standard curve to determine the copy number of viral RNA, the droplet digital PCR assay (ddPCR) directly quantifies the absolute number of copies within a given sample, which in turn makes the assay highly sensitive and accurate. Here, we have developed an EV-D68-specific ddPCR assay that effectively quantifies EV-D68 RNA copies in both cells and supernatants within a dynamic range of 6.7 × 10−3 copies/μL to 1.2 × 104 copies/μL of the sample. The assay was highly specific for a broad range of EV-D68 isolates (Fermon, US/MO/14-18947, US/MO/14-18949, US/KY/14-18953, USA/2018-23088, USA/2020-23336 and EV-D68-infected human nasal turbinate samples from the 2022 outbreak) without cross-reactivity to other viruses such as Enterovirus-A71 (EV-A71), Human Parechovirus (HPeV)-1 and -2, Coxsackievirus (CV)-B1, Human Coronavirus (HCoV)-NL63, SARS-CoV-2, Influenza-A and B, Rhinovirus, and Respiratory Syncytial Virus (RSV)-A2, which are known to cause infection in children. The assay was able to readily quantify EV-D68 in infected cells and supernatants along with nasal turbinate samples collected from children during the 2022 outbreak. Our results suggest that the assay can be readily translated to accurately quantify viral loads in tissues and body fluids such as plasma and lung or nasal aspirates.

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Sensitive and Accurate Quantification of Enterovirus-D68 (EV-D68) Viral Loads Using Droplet Digital PCR (ddPCR)

Grizer,

Li,

Mattapallil

2024

Microorganisms

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Development and validation of the first HBV qRT-PCR assay in the Mediterranean area targeting the X region

Cited by 1 publication

References 12 publications

Sensitive and Accurate Quantification of Enterovirus-D68 (EV-D68) Viral Loads Using Droplet Digital PCR (ddPCR)

Sensitive and Accurate Quantification of Enterovirus-D68 (EV-D68) Viral Loads Using Droplet Digital PCR (ddPCR)

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