Final Report: Automated Sample Collection and Concentration System for Multiple Pathogens in Water

EPA Contract Number: EPD06055
Title: Automated Sample Collection and Concentration System for Multiple Pathogens in Water
Investigators: Hsu, Fu-Chih
Small Business: Scientific Methods, Inc.
EPA Contact: Manager, SBIR Program
Phase: I
Project Period: March 1, 2006 through August 31, 2006
Project Amount: $69,999
RFA: Small Business Innovation Research (SBIR) - Phase I (2006) RFA Text |  Recipients Lists
Research Category: Drinking Water , SBIR - Water and Wastewater , Small Business Innovation Research (SBIR)


The goal of this research project was to develop a simple, rapid, and automated sample collection/concentration system that will concentrate a broad range of pathogens simultaneously. The system will integrate continuous flow centrifugation (CFC) with an innovative, positively charged filter so that large (i.e., protozoan parasites and bacteria) and small (i.e., enteric viruses) biologically active particles can be concentrated using a single procedure. This innovative sample collection and concentration device will provide sample concentrates that can be subjected directly to assays us ing molecular techniques, or to biosensors for routine water quality monitoring. The system also can serve as an emergency response platform that will aid in the rapid collection of microbiological samples during bioterror attacks and post-attack monitoring and remediation efforts.

Summary/Accomplishments (Outputs/Outcomes):

The optimized flow and velocity conditions for CFC concentration of an array of model microorganisms have been determined using seeded water samples. With flow rates u nder 500 mL/min and rotational velocities of 10,000 rpm, more than 50% of seeded E. coli cells could be recovered. When velocities were increased to 12,000 rpm, about 70% of seeded E. coli cells could be recovered. An alternative neutral pH elution buffer, OptimaRE, was developed by the research team for eluting viruses from the electropositive filters; it was capable of recovering more than 80% of the model viruses seeded into the water samples. The average recoveries for E. coli O157:H7, V. cholerae, B. subtilis spores, Cryptosporidium, Giardia, and bacteriophage MS2 were 64%, 42%, 64%, 70%, 85%, and 42%, respectively, using the integrated CFC/filtration system with different water matrices (surface, ground, and tap water samples). An automated recirculation procedure also was evaluated as an alternative to the cumbersome traditional mechanical shaking procedure; no significant differences were observed across the range of test microorganisms evaluated.


The feasibility of the integrated CFC and positively charged capsule filtration was demonstrated for the efficient recovery of multiple pathogens in different water matrices. The advantages of this system for sample collection and concentration of a broad range of pathogenic and indicator organisms include: (1) it is easy to use; (2) it can accommodate a broad range of water qualities, including high turbidity samples; (3) it produces discrete concentrates of viruses and bacteria/protozoa that do not require further concentration or separation prior to analysis; and (4) the innovative virus capsule filter provides 66- to 666-fold concentration of viruses from sample volumes ranging from 10 to 100+ liter water samples, with final concentrates of less than 150 mL. Overall, the results indicate that the integrated CFC/filtration system simultaneously can recover a broad range of pathogens efficiently and readily adapts to complete automation.

Supplemental Keywords:

small business, SBIR, continuous flow centrifugation, pathogen concentration, positively charged capsule filter, waterborne pathogens, filtration, drinking water, groundwater, surface water, drinking water monitoring, microbial agents, biosensors, biological weapon detection, wastewater, water protection, homeland security,, RFA, Scientific Discipline, Water, Environmental Chemistry, Analytical Chemistry, Environmental Monitoring, Drinking Water, Environmental Engineering, pathogens, homeland security, bioterrorism, biopollution, waterborne pathogens, community water system, polymerase chain reaction, drinking water monitoring, analytical methods, continuous monitoring sensors, biosensor, drinking water system

SBIR Phase II:

Automated Sample Collection and Concentration System for Multiple Pathogens in Water