CICEET Progress Report for the period 3/15/05 Through 9/15/05

Project Title: A New Autonomous Technology for Monitoring Microbial Indicators of Fecal Con-tamination in Coastal Waters
Principal Investigator(s): Alfred K. Hanson
Additional Investigator(s): David C. Smith, Heather L. Saffert
Project Start Date:1 September 2004

Figures

Figure 1

Figure 2

Figure 3

Tasks to meet objectives

1. The testing of the bench-top prototype involved the following tasks:
   1. Test in the lab for detection of known concentrations of ATCC strains of E. coli and enterococci.
   2. Evaluate a disinfection step between samples with solutions containing bromine.
   3. Test with field samples from marine waters.
   4. Assess whether a filtration step is necessary.
   5. Develop quantification software program and test further.
2. Prepare a specification document for the instrument manufacturer (SubChem Systems, Inc.) to use as a guide for designing and fabricating the submersible prototype.
3. Laboratory testing of the submersible prototype will be conducted, once it becomes available (expected late October, 2005).
   1. Test for detection of known concentrations of ATCC strains of E. coli and enterococci.
   2. Evaluate the bromine solution disinfection technique.
   3. Test with field samples from marine waters.
   4. Refine quantification software program.
4. Prepare a standard operating procedure for using and calibrating the field instrument.
5. Conduct some preliminary tests in situ in Narragansett Bay.

Progress on Tasks

1. Testing with the bench-top prototype;
   1. Controls and samples with known concentrations of E. coli and enterococci were tested in the instrument at room temperature (20° -22° C) and at elevated temperatures (32° -44° C).
   2. Disinfection with a bromine solution was tested. The pH of the bromine solution was observed to be critical for effective disinfection. Tests were also completed using E. coli and enterococci collected on filters, to demonstrate that they could be effectively disinfected with the bromine solution.
   3. Additional calibrations of Milli-Q and sterile seawater were conducted and shown to be consistent over time. Increases in temperature slightly decreased the fluorescent signals.
   4. It was demonstrated that repeated exposure to the optical detectors does not appear to be affecting the growth of bacteria.
2. Transitioning the bench-top into a submersible prototype;
   1. A revised specification document, for the submersible prototype, has been submitted to the SubChem engineers for their review. They are in the process of fabricating a submersible prototype, based upon these specifications and requirements, which resulted from testing with the bench-top prototype.
   2. The first generation submersible prototype for the SubBioAnalyzer is scheduled to be completed during October. At that time it will become available for testing, evaluation, and optimization.
3. In other related activities, Heather Saffert teamed with other URI CICEET project investigators, David Smith and Marek Kirs, who are working on Microbial Source Tracking using F-specific coliphage and Real-time QPCR.
   1. Field samples collected from the Providence River were analyzed for the EPA using IDEXX’s QuantiTrays to quantify E. coli and enterococci levels. Kirs also analyzed sub-samples using his newly developed method.
   2. This collaboration was also extended to study water quality in the Narrow River and Narragansett Beach and to identify potential pollution sources in the watershed.

Difficulties
Temperature variances of the incubation fluidic circuit of bench-top system were greater than the ideal +0.5° C. The SubChem engineers are working on an improved design for heaters in the incubation fluidic circuit. The time to detection was found to be somewhat longer (2-3 hours) than anticipated, based upon our earlier results. The reason for this is not yet understood.

We have had some electronic problems with bench-top prototype that have delayed testing.

Project Objectives for Next Reporting Period

Objectives
For the next reporting period, we plan to continue working to accomplish the previously stated objectives. We expect to finish laboratory testing with the bench-top prototype and then begin working with the submersible prototype this fall and winter.

Tasks to Meet Objectives

1. To complete the bench-top prototype testing, the following tasks will be performed:
   1. Further optimize the bromine disinfectant solution
   2. Test with field samples from marine waters
   3. Assess whether a filtration step is necessary
   4. Further develop quantification software and test
2. The submersible prototype is scheduled to be completed in October. At that time we will commence laboratory testing of the submersible prototype.
   1. Test for detection of known concentrations of ATCC strains of E. coli and enterococci.
   2. Test with field samples from marine waters.
   3. Refine quantification software program.
3. Prepare a standard operating procedure for using and calibrating the field instrument.
4. Conduct some field tests, in situ in Narragansett Bay.

Work Plan for Next Reporting Period
We have revised our project timeline.

Anticipated Success in Meeting Project Objectives
We expect that we will meet all the project objectives during the following year.

Overall Project Timeline Update
Task 1 ­ Bench-top Tests - October-November 05
Task 2 ­ Submersible Tests ­ December 05 - March 06
Task 3 ­ Develop Protocol - January-March 06
Task 4 ­ Field Tests ­ April-August 06

Preliminary Data
Engineering test data have been collected demonstrating the capabilities of the bench-top prototype for temperature controlled incubation and the optical detection of bacterial indicators (see Figure 1, Figure 2, and Figure 3). Data is also available on the effectiveness of a bromine solution for between measurement disinfection.

Dissemination
Contact with End Users:
David Burnett (RIDOH) was contacted and sent data on Aug. 31 from a survey the previous day because high levels above EPA recreational guidelines of both E. coli and enterococci were detected at Narragansett Beach. David Burnett had follow-up testing conducted. Testing was also conducted for the EPA (Narragansett Laboratory) on samples from the Providence River.

We have also been working with Annette DeSilva who is involved in the Narrow River Preservation Association and leads the URI Watershed Watch sampling of the river. Working with DeSilva, we are hoping to obtain additional funding to further investigate the Mumford Brook area, which has a history of very high indicator levels.

Outreach
Heather Saffert participated in the Metcalf Institute Annual Program for Journalists at URI. She discussed water quality issues, collected samples, analyzed results, and explained our work funded by CICEET with journalists. We also plan to present results from the joint Narrow River/Narragansett Beach Study for a Friend’s of Oceanography Seminar at URI and to the local SurfRider Organization this fall.

Expenditures
The expenditures are in the range anticipated for the work accomplished to date.

End User Advisor Feedback
Name: David Burnett
Organization: Rhode Island Dept. of Health
Location: 3 Capitol Hill, Room 203 Providence, RI 02908
Phone number: 401-222-2749
E-mail: davidb@doh.state.ri.us

1) At this stage, what are the potential applications for this research? Please discuss how you and others could potentially use the technology.

2) What, if anything, has changed about this project's potential applicability since the last reporting period (not applicable to the first Progress Report)?

3) Do you see any key challenges that the researchers may want to address or keep in mind?

4) Does this report offer you enough information to adequately address the above questions?

5) Other feedback?