Grantee Research Project Results
2002 Progress Report: Understanding the Sources and Fate of Conventional and Alternative Indicator Organisms in Tropical Waters
EPA Grant Number: R828829Title: Understanding the Sources and Fate of Conventional and Alternative Indicator Organisms in Tropical Waters
Investigators: Harwood, Valerie J. , Rose, Joan B.
Institution: University of South Florida
EPA Project Officer: Hahn, Intaek
Project Period: September 1, 2001 through August 31, 2003
Project Period Covered by this Report: September 1, 2001 through August 31, 2002
Project Amount: $388,335
RFA: Recreational Water Quality: Indicators and Interstitial Zones (2000) RFA Text | Recipients Lists
Research Category: Aquatic Ecosystems , Water , Ecological Indicators/Assessment/Restoration
Objective:
The objective of this research project is to investigate the fate (survival, persistence, and potential for regrowth) of various water quality indicator organisms in subtropical waters. The subjects include fecal coliforms, enterococci, Clostridium perfringens, coliphage MS2, and bacteriophages of Bacteroides fragilis. In addition, differential survival of certain Escherichia colistrains are being investigated by genotypic (ribotyping) and phenotypic (antibiotic resistance) subtyping, which will provide insight about the ecology of these organisms in aquatic environments, as well as information that will aid in refining bacterial source tracking methodologies.Progress Summary:
Phase 1 of this research project was conducted in aqueous mesocosms with associated sediments, which were established in a roofed and unsided greenhouse. Mesocosms contained natural waters of varying salinity (fresh, brackish, and saltwater), and the corresponding sediment associated with each water type. Three replicate mesocosms representing each water type were inoculated with one natural, unamended source of indicator organisms, i.e., dog feces, wastewater, or soil (27 mesocosms total). Over time, we monitored culturable counts of indicator bacteria and bacteriophages in the water column and in sediment.Indicator organisms persisted longer in fresh water than in salt or brackish water. Fecal coliform concentrations began and remained higher than Enterococcus concentrations in the water column and sediments of freshwater mesocosms. In contrast, Enterococcus species persisted longer than fecal coliforms in the sediments of brackish and saltwater mesocosms.
C. perfringens was not detectable in mesocosms inoculated with dog feces, due to very low levels in the inoculum. In mesocosms inoculated with wastewater and soil, C. perfringens declined to undetectable levels in the water column of the fresh and brackish water mesocosms; after 72 hours, C. perfringens numbers increased in the sediments from the time of inoculation until 72 hours, but declined to undetectable levels by the second week. These data suggest that C. perfringens endospores accumulate in the sediments over time, and that they maintain culturability under the conditions examined for approximately the same length of time as the fecal coliforms and enterococci.
We detected coliphages in canine feces and wastewater, but not in the soil inoculum. They declined rapidly over time, surviving 48 hours or less in the water column and sediments of all water types. Bacteriophages of B. fragilis RYC2056 (found in humans and animals) also were generally undetectable after greater than 48 hours or more, and were only isolated from wastewater. Bacteriophages specific for B. fragilis HSP40, the human-specific strain, were not detected in any of the mesocosms or the material with which they were inoculated.
Over time, we noticed a partitioning of bacteria between the water column and sediments. Fecal coliforms and enterococci persisted longer in the sediment of all three water types than in the water column. The initial density of indicator bacteria was consistently lower in sediments than in the water column; however, sediment concentrations generally rose over time as water column concentrations fell. Several possible explanations exist for this phenomenon: (1) bacteria in the water column were deposited to the sediments; (2) some growth of bacteria occurred in the sediments; and (3) the "resuscitation" of stressed, viable but nonculturable bacteria occurred in the sediment, but not in the water column.
The source of inoculum influenced the persistence of indicator bacteria in the mesocosms. In the sediments of all three water types, we consistently noted that fecal coliforms and enterococci in mesocosms with naturally contaminated soil survived for at least 1 week. Persistence of bacteria in mesocosms inoculated with dog feces and wastewater was consistently shorter, particularly in the case of fecal coliforms.
We examined the distribution of E. coli strains over time in the sediments and water column of freshwater mesocosms. This series of experiments was designed to determine whether some subtypes of E. coli display differential survival in aquatic environments. Dominant E. coli subtypes in the mesocosms varied over a 4-week incubation period, suggesting that certain strains survive longer than others in the environment. This phenomenon was most clearly illustrated in mesocosms inoculated with dog feces, as the E. coli population contained in the dog feces used to inoculate the mesocosms was nearly clonal. Five to eight E. coli ribotypes from the mesocosm water column and from the sediments were typed each day. Initial samples and 24-hour samples from mesocosms inoculated with dog feces contained E. coli with identical ribotypes (RT), which matched those obtained directly from dog feces (designated dog-RT). After 48 hours, we obtained E. coli with other ribotypes (nondog-RT) from water column samples. The nondog-RTs matched those obtained from E. coli in the uninoculated fresh water. After 2 weeks of incubation, the dog-RTs remained present in the water column (62.5 percent of all isolates typed), but other E. coli strains also were present. Similar results were noted in the sediment samples; though a longer persistence of E. coli allowed sampling to continue for 4 weeks. Some dog-RTs (62.5 percent) were obtained from samples taken up to 3 weeks incubation time; however, at week 4, only nondog-RTs were obtained from sediment samples.
Future Activities:
Survival experiments in dialysis bags will be conducted in a manmade pond, beginning in September. We anticipate that this study will be completed by December; at this time, we will undertake a seeded pond study.Journal Articles:
No journal articles submitted with this report: View all 12 publications for this projectSupplemental Keywords:
fecal coliform, Escherichia coli, E. coli, Enterococcus, bacteria, bacteriophage, coliphage, bacterial source tracking, water quality, indicator bacteria, tropical, subtropical, source tracking, ribotype, microbial survival., RFA, Scientific Discipline, Water, Geographic Area, Waste, Ecosystem Protection/Environmental Exposure & Risk, Hydrology, Health Risk Assessment, State, Fate & Transport, Environmental Monitoring, Ecological Risk Assessment, Recreational Water, Biology, fishing, recreational water monitoring, risk assessment, fate and transport, bacteria, fate and sources, E. coli, nutrient rich conditions, microbes, swimming-associated gastroenteritis, bacterial source typing, fecal coliform, water quality criteria, water quality, enterococci, indicator organisms, Florida, organisms, human health risk, tropical climate, FLAProgress and Final Reports:
Original AbstractThe 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.