Multifactorial Analysis Identifies Conditions for Robust HCoV-OC43 Replication in Primary Human Bronchial Epithelial...

Area of Science:

• Virology
• Cell Biology
• Respiratory Medicine

Background:

• Air-liquid interface (ALI) cultures mimic human airway epithelium, enabling study of respiratory pathogens.
• Human coronavirus OC43 (HCoV-OC43) infection models are crucial for understanding respiratory diseases.

Purpose of the Study:

• To investigate factors influencing HCoV-OC43 replication in ALI-cultured primary human bronchial epithelial cells (pHBECs).
• To identify optimal conditions for supporting HCoV-OC43 infection and replication in vitro.

Main Methods:

• Utilized a Design of Experiments (DoE) approach to test seven variables affecting HCoV-OC43 replication.
• Monitored epithelial differentiation via transepithelial electrical resistance and gene expression (RT-qPCR).
• Quantified viral RNA (genomic and subgenomic) using RT-qPCR to assess replication.

Main Results:

• Epithelial differentiation, completed after 21 days, was the primary factor influencing viral replication.
• Differentiated pHBECs exhibited significantly lower HCoV-OC43 RNA levels than undifferentiated cultures, especially after apical infection.
• Repeated mucus washes before infection increased HCoV-OC43 replication in mature cultures.

Conclusions:

• Epithelial differentiation negatively impacts HCoV-OC43 replication in ALI cultures.
• Conditions maximizing viral replication in differentiated pHBECs were identified, including mucus washing protocols.