Development of a Poly-γ-Glutamic Acid Flocculation Method for Enhanced Virus Recovery from Wastewater

Abstract

Environmental surveillance serves as a monitoring tool for detecting the prevalence of viruses and assessing the burden of disease within a population. It is particularly effective in screening for both waterborne and non-waterborne pathogens. Fecal contamination of water resources poses a significant threat to public health, primarily through the transmission of enteric viruses. Therefore, environmental surveillance is vital for evaluating target areas and investigating viral presence. Traditional methods for virus concentration, such as the Bag-Mediated Filtration System (BMFS), ultrafiltration, ultracentrifugation, polyethylene glycol (PEG) precipitation, and skimmed milk flocculation (SMF), are commonly employed. However, these methods face limitations, including being time-consuming, costly, or having low collection efficiency, which can compromise the accuracy of environmental surveillance and lead to underestimation of the prevalence of viruses. There is a need for virus concentration methods that are straightforward, quick, cost-effective, and reproducible. In response, the development of novel concentration methods that enhance recovery efficiency from aquatic environments is imperative. One such promising candidate is Poly-γ-Glutamic Acid (PGA), known for its robust flocculation activity and applications in many fields, such as cosmetic production, medicines, and wastewater treatment. Nevertheless, there is currently no research exploring its potential for virus recovery from aquatic environments.This study evaluated the efficiency of Poly-γ-Glutamic Acid (PGA) in recovering viruses from aquatic environments, using nonenveloped bacteriophage MS2 as a surrogate for human viruses to test various parameters of PGA, including different processed volumes (5 mL and 10 mL), sample types (pellet and supernatant), groups (silica powder only vs. silica with PGL), and water sources (lake water vs. wastewater). Recovery efficiency (RE) was calculated to assess the effectiveness of the PolyGlu (PGL) method, which contains PGA. Results indicated that different processed volumes did not significantly affect MS2 concentration or RE, although sediment accumulation in spin columns suggested potential sample loss. There were significant differences in MS2 concentration and RE between pellet and supernatant samples, with supernatant samples generally showing higher values of MS2 concentration and RE. PGL significantly improved virus capture in lake water samples compared to silica powder alone but was less effective in wastewater samples. The Generalized Linear Model (GLM) analysis underscored the significant impact of groups and sample types on MS2 concentration and RE. Despite PGL’s potential to enhance virus capture, the overall recovery efficiency remained low (13.06%). Further optimization of parameters is necessary, including the presence of cations, pH levels, stirring conditions, temperature, and the dosage of PGA. Additionally, it is essential to consider the presence of inhibitors in water samples and to comprehend the impact of silica powder and PGL on the results of qPCR in order to enhance the accuracy of detection in environmental surveillance. This study emphasizes the necessity for further research to optimize and validate PGL-based methods for environmental virus concentration. The objective is to provide a reliable, rapid, and cost-effective tool for public health surveillance.