Tennessee Valley Authority
Toxicity Testing
Invertebrate and Fish Toxicity and Bioaccumulation of Metals
TVA initiated two-phase laboratory sediment and surface water toxicity studies in the spring and summer of 2009. The objective of this testing was to determine whether constituents associated with fly ash found in site sediment and surface water are harmful to or are bio-accumulated by benthic invertebrates. The primary focus of the initial studies was on the potential for short-term effects associated with removing ash from the Emory River. Longer-term toxicity and bioaccumulation studies are either underway or are being planned in conjunction with the CERCLA non-time critical removal actions. The toxic effects endpoints measured include survival, growth and reproduction, and metals bioaccumulation elicited by exposure of benthic and aquatic species to whole ash, ash elutriates, dredge plume water, and ash stilling pond effluent.
The two phases of testing (described as Phase I and Phase II) were linked to TVA’s incremental approach to dredging ash from the Emory River. Phase I testing was conducted in spring and summer (April through June) 2009 with Vibracore®, dredge plume, and plant Stilling Pond (Outfall 001) effluent samples during the pilot phase of dredging. The methods for toxicity tests are discussed in TVA (2009) Kingston Fossil Plant Fly Ash Recovery Project Sampling Plan for Phase I Dredging Operations.
Phase II testing with dredge plume and Stilling Pond effluent samples began in August 2009 to correspond with increased dredging rates and continues to present. This testing was described in TVA (2009) Surface Water Monitoring Plan for the Emory, Clinch, and Tennessee Rivers.
Phase I and Phase II testing are summarized below. Relevant links are provided to laboratory reports and/or data compilations.
Phase I
Phase I Toxicity and Associated Studies for Ash Deposits
Multiple 3.1-m Vibracore® samples were collected immediately upstream (VB.1) and downstream (VB.2) of the main portion of Emory River ash on March 17, composited into 2 discrete samples, individually homogenized, placed in 19-L plastic buckets and kept in cold storage (4ºC) until use. On June 11 and 12 and following 3 months of Phase I dredging operations, two additional Vibracore® samples (VB.3 & VB.4) were collected and processed in the same manner from approximately the same locations as the March 17 samples. June samples were also treated with ion-exchange resins in an effort to provide a suitable reference control that retains the physical properties of ash, but without levels of contaminants that could potentially elicit toxicity. Laboratory control sediment was collected from Clinch River mile 189.0. Various acute and chronic test protocols were used in the sediment toxicity testing, including:
- Corbicula fluminea bioaccumulation for sediments
- Ceriodaphnia dubia 96-hr survival test with whole ash elutriates
- Pimephales promelas 96-hr survival test with whole ash elutriates
- Freshwater Juvenile Mussel (Lampsilis siliquoidea – VB.1 & VB.2; L. cardium – VB.3 & VB.4) 10-d survival for whole ash elutriates
- Freshwater Juvenile Mussel (L. siliquoidea – VB.1 & VB.2; L. cardium – VB.3 & VB.4) 5-d survival for sediments
- Freshwater Juvenile Mussel (L. siliquoidea – VB.1 & VB.2; L. cardium – VB.3 & VB.4) 10-d survival for sediments
- Hyalella azteca 10-d survival and growth for sediments
- Lumbriculus variegatus 4-d toxicity test (pre-bioaccumulation) for sediments
Hydrosphere Phase I Report - Vb.1 & Vb.2
Hydrosphere Phase I Report - Vb.3 & Vb.4
Hydrosphere Plume and Outfall Results April - December 2009
Elutriate Toxicity Evaluation: Ceriodaphnia dubia and Pimephales promelas - University of Maryland
Results from the sampling in March (ash composite samples VB.1 and VB.2) indicated no appreciable bioaccumulation of metals in C. fluminea exposures (28-d) to whole ash nor any toxic effects in C. dubia (96-h), P. promelas (96-h), or L. siliquoidea (10-d) exposures to ash elutriates. L. variegatus exposures (4-d) to whole ash showed no effects on survival, but worms did not burrow so bioaccumulation was not assessed. No effects on survival were noted for L. siliquoidea (5-d) exposures to whole ash, but (10-d) exposures to one of the whole ash samples did result in significant effects on survival relative to laboratory control sediment. H. azteca exposures (10-d) to whole ash samples indicated adverse effects on survival, growth, and biomass.
Summary Tables VB1&2
Full Calcs VB1
Full Calcs VB2
Toxicity results from the untreated samples from June (ash composite samples VB.3 and VB.4) indicated no appreciable bioaccumulation of metals in C. fluminea exposures (28-d) to whole ash. Effects on survival were noted in both of the centrifuged and uncentrifuged elutriate samples for L. cardium (10-d) exposures. Effects on survival were noted in one of the centrifuged elutriate samples for C. dubia (96-h) exposures. No effects on survival were observed in the uncentrifuged elutriate samples for C. dubia (96-h) exposures. Effects on survival were noted in one of the centrifuged and one of the uncentrifuged elutriate samples for P. promelas (96-h) exposures. Similar results were seen with ultraviolet-treated elutriate samples. L. variegatus exposures (4-d) to whole ash showed no effects on survival, but worms did not burrow so bioaccumulation was not assessed. Effects on survival were noted in one of the whole ash samples for L. cardium (5-d) exposures as well as one of the whole ash samples for (10-d) exposures. H. azteca exposures (10-d) to both whole ash samples indicated adverse effects on survival, growth, and biomass.
Unlike the freshwater juvenile mussel (L. siliquoidea) tests conducted with VB.1 and VB.2 samples, large masses of “debris” developed in the L. cardium test exposure solutions with VB.3 and VB.4 samples. The laboratory reported on benchsheets that test animals in several replicates appeared stressed as early as 48 h into the exposure, including those in which laboratory control water (MHSW) was used. Within 96 h, test animals were being entangled in the debris, and this was confirmed by TVA’s toxicologist during a site visit while the test exposures were ongoing. Microscopic examination indicated that the “debris” consisted of fungal hyphae with large colonies of protozoa. It is not known if there is a relation between this apparent pathogenic interference and that commonly observed in larval fathead minnow tests with Emory River water, or if the fungal material originated with the test organism supplier. Test organisms were supplied by Genoa National Fish Hatchery, and since L. siliquoidea were not available for the VB.3 and VB.4 tests, L. cardium were used instead.
The results for VB.3 and VB.4 were significantly different than for VB.1 and VB.2, but showed no discernable patterns of toxicity. Effects were observed in more test exposures with the downstream sample, but were not consistent. Several efforts have been undertaken to try to elucidate the reasons for the VB.3 and VB.4 results, with little success. It is likely that the fungal material observed in the VB.3 and VB.4 tests contributed to the complicated results for those samples, at least for the L. cardium tests.
Summary Tables VB3&4 - Untreated
Full Calcs VB3 - Untreated
Full Calcs VB4 - Untreated
Summary Tables VB3&4 - Treated
Full Calcs VB3 - Treated
Full Calcs VB4 - Treated
Results from the resin-treated samples from June (ash composite samples VB3 and VB4) did not provide conclusive results, probably at least partly because treatment durations differed between tests. Additional investigations of resin treatment to attempt to produce a suitable non-toxic reference material with the same physical characteristics as fly ash are planned.
As a result of the differences between the results for the March and June Vibracore samples and the difficulty in interpreting the June results, TVA initiated several additional activities described in the discussion of Phase II below.
Phase I Toxicity and Associated Studies for Surface Water
Once per month during April, May, and June 2009, samples were collected for testing from Emory River dredge plumes and the Stilling Pond (Outfall 001). Emory River dredge plumes were visually located, followed by determination of the most turbid area using a Hydrolab. At that point, a peristaltic pump was used to collect an aqueous plume sample. Samples were held at 4ºC until use. Reference control and dilution water were collected from unaffected areas of the Emory River. In addition, 24-h composite stilling pond effluent samples were collected with a ISCO sampler, and samples were held at 4ºC until use. The methods for toxicity test are discussed in more detail in TVA (2009) Kingston Fossil Plant Fly Ash Recovery Project Sampling Plan for Phase I Dredging Operations. Various acute and chronic test protocols were used in the Phase I surface water toxicity testing, including:
- C. dubia 7-d survival and reproduction test with Emory River Plume
- C. dubia 7-d survival and reproduction test with Outfall 001
- P. promelas 7-d survival, growth, and biomass test with Emory River Plume
- P. promelas 7-d survival, growth, and biomass test with Outfall 001
No effects (survival, reproduction) were observed in 7-day chronic exposures with C. dubia to plume or stilling pond effluent samples collected from April to June. Results with identical exposures to the April and May samples by P. promelas (survival, growth) were invalidated due to confirmed pathogen interference. P. promelas chronic tests with Ultraviolet-treated plume and stilling pond effluent samples collected in June resulted in no adverse effects.
- Insert link to October 28 Hydrosphere lab reports (again)
Phase II
Phase II testing was approved by EPA as a component of the TVA (2009) Surface Water Monitoring Plan for the Emory, Clinch, and Tennessee Rivers. In response to preliminary results from the VB.3/4 testing summarized above, this plan included resuming the C. dubia and P. promelas acute toxicity tests on both the stilling pond outfall and the dredge plume samples on a weekly schedule for an unspecified period of time until there was sufficient data to propose elimination of that activity. The reasoning was that because the VB.3 and VB.4 results could not be clearly interpreted, it was important to continue to perform the plume and outfall sampling with acute toxicity tests on as frequent a schedule as was sustainable. The weekly plume and outfall acute toxicity testing resumed on August 26, and after six weeks with no observed adverse effects, was reduced to a biweekly schedule.
Therefore, Phase II testing (96-h C. dubia and P. promelas) involved weekly (August – September) and biweekly (October – present) monitoring of dredge plume and stilling pond effluent samples. Grab samples were collected from observed dredge plumes on the Emory River and 24-hour composite samples were collected from the KIF Stilling Pond Outfall 001 and shipped to Hydrosphere Research laboratory in Alachua, Florida. Unaffected Emory River water (collected at Emory River Mile 12.2) was used as reference control and dilution water. The methods used were modifications of EPA Method 2002.0 and 2000.0.The following toxicity tests were conducted:
- C. dubia 96-hr survival test with Emory River Plume
- P. promelas 96-hr survival test with Emory River Plume
- C. dubia 96-hr survival test with Outfall 001
- P. promelas 96-hr survival test with Outfall 001
A total of 50 tests, including one re-test of Outfall 001, were performed between April and December 31, 2009. Only one test from a sample collected on October 27, 2009 from Outfall 001 exhibited any toxicity for C. dubia. The same sample showed no toxicity for P. promelas. On November 5, 2009 another sample from Outfall 001 was collected and exhibited no toxicity for either C. dubia or P. promelas. Table 1 below summarizes these results.
Table 1
Sample Date |
Results |
August 26, 2009 |
Emory River Dredge Plume and Outfall 001
|
September 1, 2009 |
Emory River Dredge Plume and Outfall 001
|
September 9, 2009 |
Emory River Dredge Plume and Outfall 001
|
September 15, 2009 |
Emory River Dredge Plume and Outfall 001
|
September 22, 2009 |
Emory River Dredge Plume and Outfall 001
|
September 29, 2009 |
Emory River Dredge Plume and Outfall 001
|
Table 1 (continued)
Sample Date |
Results |
October 13, 2009 |
Emory River Dredge Plume and Outfall 001
|
October 27, 2009** |
Emory River Dredge Plume
Outfall 001
|
November 5. 2009** |
Outfall 001
|
November 10, 2009 |
Emory River Dredge Plume and Outfall 001
|
December 1, 2009 |
Emory River Dredge Plume and Outfall 001
|
December 15, 2009 |
Emory River Dredge Plume and Outfall 001
|
December 29, 2009 |
Emory River Dredge Plume and Outfall 001
|
TVA further investigated the VB.3 and VB.4 results by performing a homogenization and metals variability study during November and December 2009. Randomly selected aliquots of VB.3 and VB.4 ash were homogenized and samples collected for total metals analyses. Similarly, randomly selected aliquots of VB.3 and VB.4 ash that had been homogenized and retained at the KIF site were sampled for total metals analyses. In all, 47 samples were analyzed. The objective was to determine the possibility that metals concentration variability might help to explain the toxicity test results. The data show a high degree of analytical precision among all samples (insignificant variations considering the analytical method). However, it could not be determined whether the small degree of analytical variability might have affected the results.