Grantee Research Project Results

2002 Progress Report: Mycobacterium avium Complex in Drinking Water: Detection, Distribution, and Routes of Exposure

EPA Grant Number: R828036
Title: Mycobacterium avium Complex in Drinking Water: Detection, Distribution, and Routes of Exposure
Investigators: Ford, Timothy E.
Institution: Harvard University , Montana State University
EPA Project Officer: Hahn, Intaek
Project Period: (Extended to March 21, 2004)
Project Period Covered by this Report: January 1, 2002 through January 1, 2003
Project Amount: $516,679
RFA: Drinking Water (1999) RFA Text | Recipients Lists
Research Category: Drinking Water , Water

Objective:

The objectives of this research project are to: (1) develop and improve methods for detecting Mycobacterium avium complex (MAC) in drinking water and biofilms; (2) examine presence and distribution of MAC in drinking water distribution systems; and (3) explore routes of exposure to MAC at residential homes.

Progress Summary:

The work in the last project year focused on the following three areas:

Validating techniques and methods for detecting MAC in both drinking water and biofilm samples

Conducting an epidemiological survey on the presence and distribution of MAC in both residential homes and water distribution systems in four eastern Massachusetts towns

Investigating the routes of exposure by detecting MAC in samples collected from both kitchen taps and showerheads of residential homes.

Validating Methods for the Detection of MAC

To detect MAC from both drinking water and biofilm samples, several laboratory protocols were tested and finalized early in the last project year:

1. The filtering technique has been optimized and used in the epidemiological study. This technique has been compared with selective culture on paraffin slides.

2. To improve the detection of the MAC strains, different types of antibiotics have been added to solid media at various concentrations to inhibit non-MAC bacteria.

3. To find a more precise and reliable method for detection and quantification of MAC in drinking water and in biofilms, fluorescence in situ hybridization (FISH) probes have been developed for the MAC 16sRNA gene. Two fluorescent oligo-nucleotide probes have been designed and tested.

4. PCR-restriction enzyme pattern analysis of the gene encoding for the 65-kDa protein (HSP65 gene) has been used as a rapid identification test for Mycobacterium species. Identification of these species was confirmed by comparing the 16sRNA gene sequence to the National Center for Biotechnology Information-GenBank database.

5. Laboratory experiments have continued on biofilm formation using a bypass system that allows recirculation of hot water. Environmental biofilms are generated to assess MAC survival and growth in hot water systems. A parallel system now has been constructed at Montana State University.

An Epidemiological Survey on Presence and Distribution of MAC

After extensive efforts to recruit households, 55 residential homes have been identified from 4 municipal towns (i.e., Duxbury, Lowell, Boston, and Cambridge). A total of 240 samples have been collected from these residential homes for each quarterly sampling period. To date, three quarterly sampling periods have been completed. The final sampling period will be completed early in the fall of this year.

While collecting drinking water samples from residential homes, samples also have been collected from locations used by the local water utilities for their own routine monthly water quality monitoring. These sampling locations/sources include raw water, filter influent and effluent water, and finish water samples from each water utility plant. Collaborating with local water utility teams serving each of the 4 municipal towns, a total of 70-80 samples have been collected from each of the 4 water distribution systems at each quarterly sampling date. The final quarterly sampling will be completed by November 2003.

All collected samples have been analyzed. The chemical analyses performed on these samples included pH, temperature, free and total chlorine residues, alkalinity, ammonia-N, nitrite-N, nitrate-N, total iron, and assimilable organic carbon. The microbial analyses include the detection for MAC and heterotrophic plate counts. These samples also were used to identify and refine the technique and protocol for detecting MAC, as described above. The labor-intensive process of these analyses has consumed much of the research teams’ efforts in the last project year.

Investigating the Route of Exposure

To investigate the possible route of exposure, samples have been collected from two different potential exposure sources (kitchen and shower) in the end-users’ homes. At each visit, at least four water samples are taken from the kitchen faucet (cold and hot water) and from the shower head (cold and hot water). These samples have been analyzed for the presence of MAC.

At the first sample collection, glass and metal slides were installed into the back of the most frequently used toilet tank at each home. This is to allow biofilm formation to assess whether toilet tank biofilms represent a potential reservoir for MAC and could therefore present an exposure route through aerosolization during toilet flushing. The slides have been placed inside the toilet tanks for at least 6 months and will be analyzed for MAC over the coming months.

There was no significant difference in the presence of MAC between the two potential exposure points in the residential homes (i.e., kitchen faucet versus shower head). Numbers of MAC, however, were slightly higher in samples collected from the shower head than from the kitchen tap.

Future Activities:

In the next year, we will: (1) complete the comparative study to investigate the use of paraffin slide cultures; (2) finalize the FISH protocols to hybridize directly to the biofilm samples; (3) improve the M. intracellulare probe and protocols; (4) complete the seasonal sampling procedure and complete the intensive sampling at the sites found to be persistently positive for MAC; (5) perform statistical analyses; and (6) prepare manuscripts of our findings.

Journal Articles:

No journal articles submitted with this report: View all 6 publications for this project

Supplemental Keywords:

pathogens, survival, proliferation, new techniques, mycobacteria, drinking water pathogens, Mycobacterium avium, Mycobacterium avium complex, MAC, Mycobacterium intracellulare, fluorescence in-situ hybridization, FISH, drinking water biofilms, hot water recirculating systems, bypass systems, water distribution systems, routes of exposure, community water system, contaminant candidate list, contaminant removal, detection, drinking water system, drinking water contaminants, drinking water treatment, emerging pathogens, exposure, exposure and effects, microbial exposure, monitoring, treatment, water quality parameters,, RFA, Scientific Discipline, Water, Environmental Chemistry, Health Risk Assessment, Environmental Microbiology, Environmental Monitoring, Ecological Risk Assessment, Drinking Water, alternative disinfection methods, monitoring, CCL, detection, water quality parameters, exposure and effects, mycobacterium avium complex, routes of exposure, exposure, community water system, treatment, emerging pathogens, microbial exposure, water quality, contaminant removal, drinking water contaminants, drinking water treatment, water treatment, contaminant candidate list, DNA microarrays

Progress and Final Reports:

Original Abstract

The 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.