Grantee Research Project Results
Final Report: A Hydrosol Concentrator for Improved Measurement of Microbial Pathogens in Drinking Water
EPA Contract Number: EPD10015Title: A Hydrosol Concentrator for Improved Measurement of Microbial Pathogens in Drinking Water
Investigators: Page, Andrew E.
Small Business: AlburtyLab, Inc.
EPA Contact: Richards, April
Phase: I
Project Period: March 1, 2010 through August 1, 2010
Project Amount: $69,972
RFA: Small Business Innovation Research (SBIR) - Phase I (2010) RFA Text | Recipients Lists
Research Category: SBIR - Homeland Security , Small Business Innovation Research (SBIR)
Description:
The importance of fast, reliable detection of pathogens in drinking water in the United States and worldwide, is rapidly increasing due to rising populations and increasing environmental degradation, as well as the threat of terrorism. Although the technological readiness level of rapid detection methods for pathogens in water has increased dramatically in recent years, rapid, automated sample concentration and preparation have lagged significantly.
The InnovaPrep hydrosol concentrator was developed through a partnership of two Missouri companies, AlburtyLab, Inc., and Page Applied Research LLC, to quickly and efficiently concentrate waterborne bacteria, viruses, toxins, protozoa, or other particles of interest from a large liquid volume into a very small liquid volume. The InnovaPrep system uses a novel elution method to recover organisms that have been captured onto hollow fiber ultrafiltration membranes into very small volumes. InnovaPrep LLC, of Drexel, Missouri, obtained funding through EPA's SBIR program to develop of a two-staged unit for rapidly concentrating volumes of 10 to 100 liters of water into volumes as small as 500 microliters in less than 1 hour. One method was evaluated for removal of inhibitors and further concentration of the InnovaPrep output. This used immunomagnetic beads, prior to lysis, to separate target organisms. A final analysis-ready sample of 50 microliters was to be analyzed by quantitative polymerase chain reaction (qPCR); however, due to technical difficulties and delays in development of qPCR sample preparation methods, these samples were analyzed by staining and microscopic counting.
Summary/Accomplishments (Outputs/Outcomes):
Under this program, InnovaPrep developed a prototype two-stage system that can concentrate waterborne microbial pathogen-sized particles contained in 100 L of water into 0.5 mL at efficiencies up to 80% in less than 45 minutes. Because of the difficulties associated with the lysis and cleanup procedures and technical hurdles expected to be associated with automating these procedures, especially the freeze-thaw lysis methods, InnovaPrep recently has began exploring the potential of coupling the concentration system with a Luminex Corporation system using immuno-based assays. In this way, it is possible to develop multiplexed detection systems that do not require front-end lysis. This is especially important with regard to Cryptosporidium oocysts and Giardia cysts, due to the difficulty associated with their lysis and the apparent need for freeze-thaw cycles as part of this process. Additionally, the newest Luminex system, the MAGPIX® system, uses magnetic beads and a capture substrate that provides IMS-based cleanup of the sample prior to fluorescent imaging on the same substrate. This integrated cleanup step potentially will allow for connection of the InnovaPrep two-stage concentrator directly to the MAGPIX system without requiring development of additional automated portions.
Conclusions:
Despite the difficulties encountered in Phase I, InnovaPrep has successfully demonstrated the ability to concentrate particles of interest from 100 liters of drinking water into volumes approaching 0.5 mL with total run times approaching 30 minutes. Using water line pressure to achieve higher feed pressures should allow total run times of less than 15 minutes to be achieved. Final elution volumes of less than 0.5 mL also are possible with a specially designed stage 2 concentration cell.
Commercialization:
Currently, InnovaPrep is following two paths towards development of two-stage concentration/detection systems for improved detection of pathogens in drinking water. The first path is that InnovaPrep has formed a partnership with ElutraSep, Inc., and has begun development of an automated elution system (InnovaPrep AutoElutor) for use with the 0.45 polysulfone hollow fiber concentration cells licensed to ElutraSep by MinnTech Corporation. ElutraSep plans to offer these systems for use in Cryptosporidium and Giardia laboratories for elution of their Cryptonite and Cryptonite HV concentration cells after field collection and transfer to the laboratory.
Initial renders of the AutoElutor and initial testing of a new plastic extraction fluid reservoir for use in this system have been completed. The unit will accept prefilled IV-type plastic fluid bags and will auto fill the extraction reservoir. Elution of the concentration cells will be performed with a single button push. ElutraSep currently is selling concentration cell and syringe kits and offering a novel manual elution method for use with these kits. They are interested in offering the AutoElutor to laboratories that perform significant numbers of analyses.
The second path is toward development of a fully automated online detection system that will couple the InnovaPrep two-stage concentrator with the Luminex MAGPIX system. InnovaPrep and Luminex have had several discussions in the past towards development of integrated systems for Biodefense detection applications. More recently, the discussions have centered on integration of the two-stage concentration system and the MAGPIX system.
A fully automated system using the InnovaPrep two-stage concentrator and the Luminex MAGPIX system would potentially provide online detection of up to 50 targets approximately every hour. Using immuno-based assays will result in slightly higher detection limits, but will significantly reduce the time to development and the final system cost. Using ultrafiltration membranes in the two-stage InnovaPrep system will provide the ability to concentrate and subsequently detect Cryptosporidium, Giardia, and other parasites, as well as pathogenic bacteria and viruses, and toxins. Based on the cost of the current InnovaPrep HSC-40 and MAGPIX system, it is possible to achieve a system cost of less than $100K, which would make the system potentially available to large and small municipal water supplies.
Supplemental Keywords:
small business, SBIR, EPA, homeland security, drinking water pathogens, hydrosol concentrator, quantitative polymerase chain reaction, qPCR, drinking water, microbiological methods, water security monitoring, automated microorganism detection systems, microbial pathogens measurement, rapid detection methodsThe perspectives, information and conclusions conveyed in research project abstracts, progress reports, final reports, journal abstracts and journal publications convey the viewpoints of the principal investigator and may not represent the views and policies of ORD and EPA. Conclusions drawn by the principal investigators have not been reviewed by the Agency.