III. TOXICITY: Based on the scientific literature and ecological context, is there evidence to suggest that effluent from the new outfall may negatively impact lobster?

The wastewater effluent will contain various potentially toxic substances including chlorine and chloramine, other halogenated hydrocarbons, ions and heavy metals. Of these, only DDT's and PCB's pose a potential threat to lobsters because they bioaccumulate. The consequences of bioaccumulating these compounds are poorly understood for lobster. Other potentially toxic substances will apparently be present at sufficiently low concentrations that they should not pose a serious threat.

Chlorine & Chloramine: The toxicity of chlorine to marine organisms is dependent on the available form of chlorine, the temperature of the water (higher temperatures are required for toxic reactions), and time of exposure to the chlorinated waters. The available form of chlorine will depend on the concentrations of ammonia, bromide, organic matter, and other nitrogenous compounds. Generally there are several forms of free chlorine: hypochlorous acid (HOCl), hypochlorite ion (OCl^-), hypobromous acid (HOBr), hypobromous ion (OBr^-), and broamines (EPA, 1984). When nitrogen-containing compounds are present, chloramine (monochloramine and dichloramine) will be formed and it is the compound that has the greatest potential to cause toxic reactions (note that there is a ten-fold difference in the amount of free chlorine vs. chloramine needed to produce a toxic reaction in lobsters, with chloramine being far more toxic), although all of the compounds formed are toxic to aquatic organisms. Of course, in wastewater effluents, there is a high concentration of nitrogenous compounds; thus, chloramine will be formed, and this is why concerns have been raised about the effects on lobster larvae.

While chlorine and chloramine compounds can have serious lethal effects on stage I lobster larvae, they must be in high concentration (16.3 mg/liter for free chlorine and 2.02 mg/l for chloramine) at elevated temperatures (25-30°C). At lower temperatures (20°C), free chlorine resulted in ~20% mortality at levels ranging from 0.08 mg/l to 8 mg/l; mortality increased at 10 mg/l (Fig. 1). However, at 20°C chloramine had dramatic mortality effects at concentrations greater than 1 mg/l (Fig. 2). Thus, temperature and concentration work together to produce greater effects than either would on their own -- they are synergistic. Part of this synergistic effect is due to the fact that higher temperatures are extremely stressful to larvae, as evidenced by the fact that a temperature of 30°C caused more than a 10% increase in mortality to control lobsters exposed to it for 30 minutes and more than a 20% increase in mortality to control lobsters exposed for 60 minutes (Capuzzo et al., 1976).

Lower concentrations of both compounds can also cause respiratory distress in larvae at a constant temperature of 25°C (Fig. 3). Exposure to free chlorine in concentrations of 5.0 mg/l results in a doubling of the respiratory rate of stage I larvae. Concentrations of 10 mg/l result in a significant decrease in the respiratory rate. However, at concentrations of 0.1 mg/l and 1.0 mg/l free chlorine, no respiratory distress is noted. Again, the effects of chloramine are more dramatic with concentrations of 0.05 mg/l and 0.5 mg/l causing initial increases in O₂ uptake, followed by significant decreases after 48 h and increased mortality (Capuzzo et al., 1976).

Exposure to both free chlorine and chloramine (at 1 mg/l) at 25°C can result in increased mortality (compared to control groups), but the mortality value is significantly less than the LC₅₀ values of 16.3 mg/l free chlorine and 2.02 mg/l chloramine, and it stabilizes after 48 h of exposure to between 15-25%. However, these lower concentrations (1 mg/l) reduce the growth of the larvae as measured by both length and weight; this growth reduction is particularly noticeable in the molt from stage I to stage II (Figs. 4 & 5). For free chlorine exposed animals, no further growth rate reduction is noted for length in subsequent molts, but weights are reduced, implying that the intermolt interval may be longer for these larvae. For chloramine exposed animals, growth rate is reduced for each subsequent molt with postlarvae (stage IV) being considerably smaller and lighter than either control or free chlorine exposed animals. Furthermore, respiratory rates are reduced for exposed larvae and postlarvae compared to control animals. These sublethal effects indicate that long-lasting metabolic disturbances may result from acute exposure to chlorinated seawater, but only at elevated temperatures (25°C). At lower temperatures, these effects will not occur (Capuzzo, 1977).

In summary, a toxic reaction to chloramine is dependent on temperature. If the temperature is 25-30°C, chloramine is very toxic even in very low concentrations (2 mg/l); if the temperature is 20°C, 1 mg/l chloramine is sufficient to cause respiratory distress and to affect growth, but a toxicity effect is not noted until the concentration of chloramine reaches 4.08 mg/l. At lower temperatures, a greater and greater concentration of chloramine will be required to cause a toxicity effect. It is highly unlikely that conditions potentially harmful to lobster larvae would arise because the new outfall pipe is located at a depth ranging from 28-32 meters, in an area where the bottom temperatures rarely exceeds 12°C and where mixing with warmer effluent waters will not result in waters warmer than 12.25°C.

Surface temperatures above the new outfall site may exceed 20°C and it is possible that larval lobsters could occur there if females release hatchlings nearby. However, there is a seasonal thermocline present which would most likely form an effective barrier to compounds within the effluent plume. Thus, those compounds would not be able to reach the upper layer waters where temperatures would be great enough for toxic effects to occur assuming that NO dilution of original emitted concentrations took place between the bottom and the surface -- an event which is unlikely. Similarly, as the food of the pelagic larvae is also planktonic and concentrated at the water's surface, it too should not be affected by chlorine and chloramine compounds.

The basic flaw of these above-mentioned tests is that they have been conducted with pulsed doses -- the kinds typically used in power plants to eliminate fouling species. However, in the case of wastewater effluent, the dose of chlorine is not intermittant, but continuous. Thus the effects of short-term exposures will likely be underestimates of the potential effects during continuous exposure. However, these pulsed experiments provide us with baseline data that we can use to predict the effects of continuous doses. Currently, the chlorine concentrations found in wastewater effluents in Boston Harbor are ~2 mg/l before dilution with seawater occurs (Ken Keay, MWRA, personal communication). These are free chlorine concentrations, which if combined with high water temperatures and organic matter, could cause problems for lobster larvae and other marine organisms only in the event of no dilution. Unfortunately, this ongoing effluent discharge is not what is causing the current concern. It is the proposed new outfall pipe's effluent that is causing the concern -- and the chlorine concentration for the new effluent has been set at 0.456-0.631 mg/l (upon dilution with seawater, it is expected to go no higher than 0.1 mg/l; Ken Keay, MWRA, personal communication). Clearly, these lower concentrations of free chlorine are far better than the present situation -- there is far less chance that any toxic or sublethal effects will occur. Furthermore, according to EPA testing (EPA, 1984), the Atlantic silverside, calanoid copepod (Acartia tonsa), and the eastern oyster are three of the most sensitive genera to chlorine toxicity. MWRA currently uses the silverside as an indicator species and we have recommended above that they continue their use of copepods. Additional information on the relative half-lives of these compounds in seawater and turbulent flow would help determine how likely they are to cause problems.

Other Halogenated Hydrocarbons: Effects of exposure to other halogenated hydrocarbons are summarized in Mercaldo-Allen and Kuropat (1994). Of particular concern is DDT retention in egg masses, since that may alter reproductive performance. Adult lobsters withstand sublethal acute exposures to organics such as DDT or PCB by sequestering the pesticides into their lipophilic tissues (hepatopancreas and egg masses). Egg masses retain more than 1% of a dose of 100 µg/l of DDT one month after an intravascular injection. If the dose is administered through ambient seawater or in food, residue concentrations are highest in eggs masses (1000 ng/g wet weight) and hepatopancreas (400 ng/g wet weight) one week later. The impact on developing embryos has not been studied thus far, so it is currently impossible to assess the effect that the MWRA effluent concentrations reported in Table 3 of Mitchell et al. (1998) will have given that they range between 1.51 to 3.7 ng/l. However, the effluent discharges of DDT are well below the levels used to determine residual concentrations and will be diluted upon exposure to the seawater. They may or may not represent a problem to developing embryos, but this would need to be determined in laboratory studies where embryonic development of chronically exposed females could best be assessed.

PCB's are also retained in egg masses and have a half life of 40 days. Lobsters can take up PCB's directly from the water via their gills and via trophic exchange by feeding on animals containing PCB's. Again, we know nothing of the effect of PCB's on developing lobster embryos. MWRA effluent concentrations will range from 11.52 to 20.4 ng/l for total PCB's. Whether this will represent a problem for lobsters cannot be determined without laboratory testing. PAH LC₅₀ concentrations have been determined for both larvae and adults. Creosote at a concentration of 0.02 µg/g is lethal after 96 hours for lobster larvae at 20°C (McLeese and Metcalfe, 1979). It is not known if this concentration is more or less or equally lethal at other temperatures. However, at 10°C, 1.76 µg/g of creosote administered to adult lobsters for 96 hours is lethal. At elevated temperatures, PAH's are known to accumulate in lobster tissues in greater concentrations and lobsters are capable of metabolizing and flushing these compounds out of their system if held in clean water (Uthe et al., 1984; James, 1989). The MWRA effluent concentrations will range from 3090 to 6813 ng/l of total PAH's, which would seem to represent no problem for the lobster larvae or the adults given the differences in concentration magnitudes.

Other Compounds (Ag, Cd, Cr, Cu, Mo, Ni, Pb, Zn, Hg, Fe, Se, Sn): According to Table 3 in Mitchell et al. (1998), all of these metals would be released in accordance with EPA ambient water quality criteria. Again the effects of these compounds are summarized in Mercaldo-Allen and Kuropat (1994). Cadmium, copper, and mercury effects have been assessed for stage I larvae (Johnson and Gentile, 1979) reared in temperatures of 20°C. A dose administered over 96 hours of 78 µg/l Cd results in 50% mortality, while that of 48 µg/l Cu results in 50% mortality and that of 20-33 µg/l Hg results in 50-73% mortality. Higher doses over shorter times, also result in considerable mortality: Twenty-four hour exposure to 1000 µg/l Cd results in 50% mortality while 48 hours of exposure results in 100% mortality. A dose of 330 µg/l of Cu results in 100% mortality after 24 hours, whereas dosages ranging from 100-330 µg/l of Hg result in 97-100% mortality. MWRA effluent levels of Cd range from 0.05-0.22 µg/l; those of Cu range from 29.79-48.63 µg/l; and those of Hg range from 0.101-0.226 µg/l, and will be released to the surrounding waters at temperatures under 20°C (concentrations taken from Table 3 of Mitchell et al., 1998). With the exception of copper, all of these concentrations are 80 to 350 times lower than the LC₅₀ levels determined for 24, 48, and 96 hours and thus do not represent a problem for larval lobsters. Copper is of concern, however, given the behavior of the larvae (described above in Section I), we do not anticipate that they will come into contact with this metal.

The lethal and sublethal effects of silver, iron, nickel, lead, selenium, tin, and zinc have not been determined for larval lobsters. In juveniles and adults however, doses of 500 µg/g Zn can cause sluggish behavior. Again, Zn levels at the outfall will range from 30.02-36.33 µg/l (concentration taken from Table 3 of Mitchell et al., 1998), which is under the value at which juvenile and adult lobsters altered their behaviors.

Salinity: Larvae raised to the postlarval stage at various temperatures and salinities exhibit the following survival patterns: at 15-17°C in artificial light with a day:night cycle, survival rates are 83% at 30.9-31.8 ppt, 67% at 27.6-28.4 ppt, 58% at 21.8-22.5 ppt, and 8.3% at 19-19.8 ppt. Survival of the larvae from stage I to stage II and III is higher than that during the metamorphic molt into the postlarval stage. At 15-17.5°C in artificial light with a day:night cycle, survivals of animals passing through all three molts to the postlarval stage are 83% at 30.1-31.8 ppt, and 67% for both 26.3-27.2 ppt and 21.1 to 22.1 ppt. In the absence of light and at higher temperatures (17.5-20.2°C), survival ranges from 95% at 30.8-31.2 ppt to 85% for 25.4-25.9 ppt and 20.7-21.5 ppt (Templeman, 1936). Clearly salinity drops to 19-20 ppt can prove quite harmful to larvae exposed for a long duration (>20 days), but lethal limits are 13.6 ppt. However, given the fact that larvae will avoid swimming into low salinity waters or will remove themselves from them by swimming elsewhere (Scarratt and Raine, 1967), the low salinities at the immediate location of the effluent are not expected to cause a problem with the larvae.

In addition to the negative taxis of larvae towards lower salinities, non-reproductive female lobsters are also very selective about salinities and exhibit much higher levels of activities in response to them than do male lobsters. When given choices between moving into salinities of 20-25 ppt or 10-15 ppt, both males and females preferred to enter the higher salinities. Lobsters left their shelters but remained in their vicinity when salinities dropped to 18.4 ppt ± 1.42 (SE); when the salinity dropped to 12.62 ppt ± 1.59 (SE), they left the vicinity of their shelters altogether (Jury et al., 1994). Lethal salinities vary with temperature: as temperature increases above 20°C, tolerance for low salinity decreases to 16.4 ppt, whereas at lower temperatures (5°C), lethal salinity may be not be reached until 11 ppt (McLeese, 1956). While it is not understood why non-reproductive females would be more active than males in lower salinity (and would move away from it), it is suggested that this may be a physiological response having to do with potential reproductive events at a later date.

Conclusion: From all the available evidence, the current outfall is not expected to have any effect upon lobster larvae. Toxicity testing has already been conducted for both adult, juveniles, postlarvae, and larvae for a number of the metals, halogenated hydrocarbons, and other organic compounds (see Renee Mercaldo-Allen and Catherine Kuropat's 1994 NOAA Technical Memorandum NMFS-NE-105 for review) and, in the cases for which we have available data, the projected MWRA outfall emission concentrations are well below levels of concern. The EPA has developed ambient water quality criteria which require that toxic compounds be released in concentrations that will have only a 5% probability of toxicity to test organisms (as opposed to the 50% mortality or LC₅₀'s). This is conservative and prevents MWRA from releasing any compound in excess of these criteria. Thus lobster larvae will not be at risk. Furthermore, there is already a monitoring program underway for these compounds, given that MWRA must monitor their concentrations and keep to the ambient water quality criteria set by the EPA, or be found in violation of EPA standards. Given that the lobster's ranking in the toxicity testing varies depending on the compound tested (i.e., they are more sensitive to some things and less sensitive to others) and they are generally 2 to 20 times less sensitive than the most sensitive test organism, use of lobster as an indicator species is not warranted.

The only area which might present a concern is that involving the accumulation of PCB's and DDT's in egg masses and the subsequent effect these accumulations may have on developing embryos. To determine accumulation effects, we would need to know:

1. the movements and residence periods of ovigerous females in the area of the outfall;
2. if ovigerous females are found to be residential near the outfall, would they take up these compounds at greater rates than ovigerous females in outlying areas (such as Cape Cod or further up in the Gulf of Maine); and
3. the effect of dosage concentrations similar to those at the outfall on embryonic development (if any).

Movements and residence times of egg-bearing females could be determined by MADMF or NMFS agents tagging ovigerous females in the abdomen (with sphyrion tags) and recapturing them via the commercial fishery (again coordinated efforts would be required for this study to be successful, as previously discussed). This study would be best designed by MADMF officials and NMFS scientists to determine the appropriate number of ovigerous females to be tagged in order to provide the best possible recapture rate, since tag-recapture studies typically result in less than a 30% return rate (Estrella and Morrissey, 1997).

If a tag-recapture study indicates that females are indeed residential, then a study to determine bioaccumulation of DDT's and PCB's would be warranted. This study could be effected by MADMF or EPA agents collecting females within Boston Harbor and at some outlying area (matched for depth and temperature) and running comparative concentration analyses of egg masses.

Dose responses could be assessed by holding a large number ovigerous females at various concentrations of PCB's and DDT's (representative of those at the outfall) and determining chronic accumulation concentrations and effects on developing embryos (as compared to embryos of ovigerous females held in clean water). A facility that has the space, water quality, and resources to conduct this kind of study is the EPA Environmental Laboratory in Narragansett, RI or the Woods Hole Oceanographic Institution. Given that many sources other than the MWRA outfall contribute to the presence of DDT's and PCB's in our coastal waters, these kinds of studies could be viewed as a public service and would be within the purview of EPA.