Ultrafiltration-based techniques for rapid and simultaneous concentration of multiple microbe classes from 100-L tap water samples

This study focused on ultrafiltration as a technique for simultaneously concentrating and recovering viruses, bacteria and parasites in 100-L drinking water samples. A chemical dispersant, sodium polyphosphate, and Tween 80 were used to increase microbial recovery efficiencies. Secondary concentration was performed to reduce sample volumes to 3–5 mL for analysis using tissue culture, microscopy, and real-time PCR and RT-PCR. At seeding levels of 100–1000 (CFU, PFU, oocysts, or particles), a “high-flux” ultrafiltration procedure was found to achieve mean recoveries of 51–94% of simultaneously seeded MS2 bacteriophage, echovirus 1, Salmonella enterica subsp. enterica serovar Typhimurium, Bacillus atrophaeus subsp. globigii endospores, Cryptosporidium parvum oocysts, and 4.5-μm microspheres. When 4–7% of the final sample concentrate volume was assayed using real-time PCR and RT-PCR, overall method sensitivities were < 100 C. parvum oocysts, < 240 PFU echovirus 1, < 100 CFU Salmonella and ∼ 160 CFU B. atrophaeus spores in 100-L drinking water samples. The “high-flux” ultrafiltration procedure required approximately 2 h, including time required for backflushing. Secondary concentration procedures required an additional 1–3 h, while nucleic acid extraction and real-time PCR procedures required an additional 2–2.5 h. Thus, this study demonstrated that efficient recovery and sensitive detection of diverse microbes in 100-L drinking water samples could be achieved within 5–8 h using ultrafiltration, rapid secondary processing techniques, and real-time PCR.