Improved SARS-CoV-2 RNA recovery in wastewater matrices using a CTAB-based extraction method

Area of Science:

• Environmental microbiology
• Molecular biology
• Public health

Background:

• Wastewater-based epidemiology (WBE) is vital for tracking infectious diseases like SARS-CoV-2.
• Accurate SARS-CoV-2 detection in wastewater relies heavily on efficient RNA extraction, concentration, and purification.
• Optimizing these steps is critical for reliable public health surveillance data.

Purpose of the Study:

• To compare the efficiency of four common RNA extraction methods for SARS-CoV-2 detection in sewage.
• To identify a protocol that enhances viral RNA recovery from wastewater samples.
• To evaluate the impact of sample physicochemical characteristics on recovery rates.

Main Methods:

• Four RNA extraction methods were tested: CTAB-based, TRIzol-based, guanidinium thiocyanate (GTC)-based with silica spin columns, and an automated paramagnetic particle protocol.
• Sewage samples were artificially inoculated with SARS-CoV-2.
• RNA recovery rates were quantified using quantitative reverse transcription polymerase chain reaction (RT-qPCR).

Main Results:

• The CTAB-based method demonstrated the highest viral RNA recovery rates.
• CTAB consistently provided stable recovery percentages across diverse wastewater matrices.
• TRIzol-based extraction was the second most effective method.
• GTC-based and automated methods showed significantly lower recovery rates compared to CTAB and TRIzol.

Conclusions:

• The CTAB-based RNA extraction protocol is highly effective for recovering SARS-CoV-2 RNA from complex wastewater.
• This method shows promise for improving the accuracy and reliability of wastewater-based epidemiology.
• Optimized RNA recovery is essential for robust infectious disease surveillance.