Effects of fractions from biodegraded Alaska North Slope crude oil on embryonic inland silversides, Menidia beryllina.

Embryonic inland silversides, Meinida beryllina, were exposed to neutral, water-soluble fractions (WSFs) resulting from microbial degradation of artificially weathered Alaska North Slope (ANS) crude oil. Three individual microbes obtained from Prince William Sound, Alaska, and designated Phe#6 (enriched on phenanthrene), Hexaco#2 (enriched on the straight-chain alkane, hexacosane), and EI2V (grown by enrichment on Bushnell-Haas medium containing 0.2% pristane, a branched alkane) were used to individually biodegrade weathered ANS crude oil for 14 days in darkness in 20-L glass carboys containing nutrient enriched, sterilized 20% salinity sea water at 20 +/- 1 degrees C. Neutral WSFs resulting from biodegradation of ANS (lot 521) by each microbe were recovered and weighted. Neutral WSFs recovered were: 1.76 mg/L for Phe#6, 1.85 mg/L for Hexaco#2, and 13.02 mg/L for the EI2V microbe. Embryo toxicity and teratogenicity tests revealed that exposure of embryos to the WSFs from the EI2V incubation (with a total recovered neutral fraction approximately seven times greater than the Phe#6 and Hexaco#2 incubations) resulted in the most severe responses in craniofacial, cardiovascular, and skeletal organ systems. The total neutral WSFs recovered from the EI2V biodegradation of weathered ANS 521 were subfractionated into saturated (eluted with hexane), aromatic (eluted with CH2Cl2), polar (eluted with ethyl ether), and recombined (saturated + aromatic + polar) fractions. Developing fish embryos were then exposed to each subfraction and the recombined subfractions. The polar subfraction and recombined subfractions proved to be the most embryo toxic and teratogenic. They resulted in statistically significant (p < or = 0.05) responses (compared to controls) for craniofacial, cardiovascular, skeletal, and total severity effects in one or both tests with these subfractions.

Effects of aerially applied fenthion on survival and reproduction of the panacea sand fiddler, Uca panacea, in laboratory habitats.

Sand fiddler crabs, Uca panacea, were exposed in laboratory habitats to measured concentrations of ULV-grade fenthion via simulated aerial spray at 5% and 50% of field rate application of 6-12 mg fenthion/m(2) (0.05-0.10 lbs fenthion/acre). Two habitats served as controls, and two habitats served as replicates for each of the test exposures. The aerial application was repeated 12 times between July 7, 1997, and August 15, 1997, during the period of the most active larval production of the crabs. The resulting measured concentrations of fenthion at the surface and in the water were consistently lower than what the application rate would have suggested. Statistical analysis of total, daily larval counts showed a periodicity of approximately 14 days that did not appear to be affected by fenthion, at least at the 5% application rate, where a 15% reduction in larval production was noted by the end of the second hatching cycle and a 25% reduction by the end of the third. Larval production in the habitats exposed to the 50% application rate was reduced by 18% at the end of the first hatching cycle, 77% at the end of the second, and 100% at the end of the third hatching cycle. At the end of the third hatching cycle, adult crab mortality observed at the surface was 0%, 3%, and 20% for the control, 5%-, and 50%-exposure habitats, respectively. Three weeks after the final fenthion application, survival of adult crabs was 100% in the control habitats, 75% in the 5%-exposure habitats, and 3% in the 50%-exposure habitats. Survival of unfed first-stage zoeae released during the night following the sprays averaged 4.6 +/- 0.9, 3.7 +/- 0.9 and 1.7 +/- 0.6 days for control, 5%-, and 50%-exposure habitats, respectively. Forty-eight-hour LC-50s (nominal) for fenthion exposure of adult crabs submerged in water and for first-stage zoeae were 215 microg fenthion/L and 0.8 microg fenthion/L, respectively.

Preliminary observations on responses of embryonic and larval Pacific herring, Clupea pallasi, to neutral fraction biodegradation products of weathered Alaska North Slope oil.

Weathered Alaska North Slope crude oil (ANS 521) was subjected to biodegradation in vigorously stirred incubations for 14 days at 15 +/- 1 degrees C in 20/1000 salinity sterilized seawater, amended with nutrients and inoculated with a hydrocarbon-degrading microorganism (EI2V) isolated from an oil-contaminated beach in Prince William Sound, Alaska. A total of 13.7 mg/L water-soluble neutral fraction (WSF) was recovered from the incubation of weathered ANS 521. Toxicity/ teratogenicity tests were conducted with WSF recovered from the biodegradation system using embryonic and larval Pacific herring, Clupea pallasi. Exposures were begun at 4, 48, and 96 h postfertilization of herring eggs. Exposure concentrations were 1, 10, and 100% of the original concentration of WSF recovered from incubations (redissolved in 20/1000 salinity sterile seawater at 15 +/- 1 degrees C). Sterile 20/1000 salinity seawater without the addition of redissolved neutral fraction was used as a control. Significant (p < or = 0.05) embryo mortality or teratogenic responses were observed at WSF concentrations of 10 and 100%. On days 5 through 8 of embryogenesis, counts of heart contraction rates were significantly lower (p < or = 0.05) at the 100% WSF concentration for embryos exposed beginning at 4 and 48 h postfertilization. Grow-out of larvae from selected exposures was conducted. High mortality was noted in larvae exposed to the 10% WSF concentration beginning at 4 and 48 h postfertilization. Most of these larvae died 5 to 8 days after hatching when they elicited vertebral displacements at a time concurrent with the onset of feeding behavior.

Evaluation of bleached kraft mill process water using Microtox(R), Ceriodaphnia dubia, and Menidia beryllina toxicity tests.

To determine whether a 7- to 10-d embryo toxicity/teratogenicity test with the inland silverside fish, Menidia beryllina, is a sensitive indicator for evaluation of bleached kraft mill effluents, we compared this test with the Microtox(R) 15-min acute toxicity test and the Ceriodaphnia dubia 7-d chronic toxicity test. Water samples used in each test were collected from three areas in a bleached kraft pulp and paper mill using a 100% chlorine dioxide bleaching process: 1) river water prior to use in the mill; 2) the combined acid/base waste stream from the pulping process prior to biological treatment in the aerated stabilization basin (ASB); and 3) the effluent from the ASB with a retention time of approximately 11 d. Relative toxicity determined by the three tests for each water sampling location was compared. All three toxicity tests were predictive indicators of toxicity; however, the C. dubia and M. beryllina tests were the more similar and sensitive indicators of toxicity. Process water (ASB influent) prior to biological treatment in the ASB was toxic at all concentrations using the Microtox(R) and C. dubia tests. The fish embryo test showed no toxicity at 1% concentrations, slight toxicity at 10%, and acute toxicity at the 100% ASB influent concentration. Tests with biologically-treated ASB effluent indicated a substantial reduction in observed toxicity to Microtox(R) bacteria, C. dubia, and M. beryllina. No toxic responses were observed in any test at a 1% ASB effluent concentration which was the approximate effluent concentration in the receiving river following mixing. No relationship was found among any toxicological response and effluent levels of adsorbable organic halides, polychlorinated phenolic compounds, 2,3,7,8-tetrachlorodibenzo-p-dioxin, 2,3,7,8-tetrachlorodibenzofuran, total suspended solids, color, chemical oxygen demand, or total organic carbon.

Responses of embryonic and larval inland silversides, Menidia beryllina, to a water-soluble fraction formed during biodegradation of artificially weathered Alaska North Slope crude oil.

Weathered Alaska North Slope crude oil (ANS 521) was stirred for 2 and 14 days in 20 per thousand salinity sterile seawater or for 14 days in seawater with nutrients and a group of three (GO3) microorganisms from Prince William Sound, Alaska, that were capable of biodegrading hydrocarbons. A total of 0.65 and 0.69 mg/L water soluble fraction (WSF) of neutral fraction hydrocarbons was recovered from the 2- and 14-day stirred sterile systems, respectively. In comparison, a total of 7.5 mg/L WSF neutral fraction hydrocarbons was recovered from systems containing ANS 521 that were stirred and biodegraded by the GO3 microbes for 14 days. Toxicity/teratogenicity tests were conducted with neutral fraction hydrocarbons recovered from the sterile and biodegraded systems using embryonic inland silversides, Menidia beryllina. Hydrocarbons from the sterile systems did not cause statistically significant teratogenic responses at concentrations of 1%, 10%, and 100% (w/v) of recovered fractions (redissolved in 20 per thousand salinity sterile seawater). Counts of heart contraction rates were significantly lower (alpha

Infectivity and teratogenicity of Beauveria bassiana in Menidia beryllina embryos.

Developing embryos of the inland silverside fish, Menidia beryllina, were exposed to conidiospores of the insect pathogenic fungus, Beauveria bassiana, that possessed activity against the migratory grasshopper, Melanoplus sanguinipes. Various adverse effects were observed in Menidia beryllina embryos and larvae. They included rupture of the chorion, embryo death, developmental defects (vertebral abnormalities) in the embryo or hatched larvae, and fungal infections on the mandibles of larvae. Although there was little evidence of a definitive dose-response trend based on densities of viable conidiospores, statistically significant (p < or = 0.01) responses were observed in tests in which conidiospore densities were as low as 7.1 x 10(3)/ml and as high as 1.3 x 10(6)/ml. Viable spores were required for adverse effects to occur; heat-killed spores failed to cause significant adverse effects.

Toxicological assessment of biodegraded pentachlorophenol: Microtox and fish embryos.

A Gram-negative bacterium, Pseudomonas sp. strain SR3, was isolated from soil at a former wood treatment plant in north central Florida. The ability of this bacterium to degrade pentachlorophenol (PCP) was confirmed by growing cells in a basal salts medium in which PCP was the only source of carbon and energy. Degradation from a measured concentration of 39-40 micrograms PCP/ml to 0.0006 micrograms PCP/ml was observed within 120 h of incubation in the presence of PCP-induced cells of Pseudomonas sp. strain SR3. The initial cell density in these cultures was 6 x 10(6) cfu/ml. Microtox 5 min EC50 toxicity tests revealed that aqueous solutions of PCP, measured concentrations 39-40 micrograms/ml were toxic but that final biodegraded samples, 0.0006 micrograms PCP/ml were nontoxic. However, bioassays with embryonic inland silversides, Menidia beryllina, showed that the biodegraded samples were embryotoxic or teratogenic. Water containing added PCP at concentrations up to 30 times higher than measured in the final biodegraded samples was less toxic/teratogenic. These results indicate that while biodegradation of PCP was nearly complete, intermediate metabolites of the degradation process or undegraded impurities in PCP were toxic or teratogenic. Thus, the M. beryllina bioassay allows extremely sensitive assessment of toxicity associated with biodegraded environmental pollutants and may be a useful criterion for determining whether bioremediated water or soil is safe for discharge back into the environment.

Sick building syndrome. Medical evaluation of two work forces.

Sick building syndrome, characterized by upper respiratory irritative and central nervous system symptoms, is poorly understood. Building ventilation problems are frequent, although causative agent(s) are unknown. Few studies have addressed clinical characterization of symptomatic building occupants. Employees from two sites underwent standardized evaluation including medical history, physical examination and screening neurologic and neuropsychologic testing while acutely symptomatic. Both symptomatic and asymptomatic individuals were evaluated when one of the sites was evacuated. Baseline evaluation results for this group were available for comparison. Symptoms of both work forces mirrored those reported in the literature. General medical examination abnormalities were few and minor, while neurologic and neuropsychologic examinations documented mental status, cerebellar, and neurobehavioral deficits. There were statistically significant changes from baseline. Abnormalities were self-limited. Controlled evaluations of symptomatic sick building occupants should be performed to verify these findings.

Effects of Beauveria bassiana on embryos of the inland silverside fish (Menidia beryllina).

A chemical toxicity and teratogenicity test was adapted to assess potential adverse effects of a microbial pest control agent on a nontarget fish. Developing embryos of the inland silverside, Menidia beryllina, were exposed to conidiospores of the insect-pathogenic fungus Beauveria bassiana. Embryo rupture and death were observed. Embryo rupture did not always result in death, nor was death always associated with embryo rupture. Adherence of spores to the chorion, followed by germination and penetration by the germ tube, probably caused the embryos to rupture. Statistically significant (P less than or equal to 0.05) responses were observed in tests in which conidiospore concentrations were greater than or equal to 8.3 x 10(4) or less than or equal to 1.5 x 10(6)/ml. Conidiospores treated with a dispersant (biological detergent) showed significantly less binding (P less than or equal to 0.01) to embryos than did untreated spores. Both detergent-treated and heat-killed spores failed to cause significant adverse effects.

Detoxification of pentachlorophenol and creosote contaminated groundwater by physical extraction: chemical and biological assessment.

Chemical analyses revealed that polycyclic aromatic hydrocarbons (PAHs) and other organic compounds were present in a perennial freshwater stream that flowed through the abandoned American Creosote Works and into Pensacola Bay, Florida. Moreover, groundwater pumped from a well depth of 21 m at a location adjacent to the site was heavily contaminated with PAHs and other organics. A study was conducted to determine the efficacy of ultrafiltration for removal of organics from groundwater at this U.S. Environmental Protection Agency, Super Fund site. Ultrafiltration reduced the concentration of total identified organics from 210.0 mg/L in groundwater to 1.5 mg/L in the post-filtration permeate. Tests for toxicity/teratogenicity in embryonic inland silversides, Menidia beryllina; and Microtox 15 min EC50's were conducted with: 1) streamwater, 2) untreated groundwater, 3) feedwater used in the ultrafiltration system and 4) permeate water that passed through the ultrafiltration system. A concentration of 100% streamwater caused significant (alpha less than or equal to 0.05) teratogenic responses in fish embryos and larvae; the Microtox EC50 was 3.7% streamwater. Groundwater and feedwater caused significant embryo toxic or teratogenic responses at concentrations of 100, 10, and 1%; the Microtox EC50's were 0.85 and 0.48%, respectively. In contrast, only 100% permeate water caused significant increases in terata, compared to the control response; at 10 and 1% concentrations greater than 90% of hatched larvae appeared normal. The Microtox EC50 was 30% permeate water.

Biodegradation of creosote and pentachlorophenol in contaminated groundwater: chemical and biological assessment.

Shake flask studies examined the rate and extent of biodegradation of pentachlorophenol (PCP) and 42 components of coal-tar creosote present in contaminated groundwater recovered from the American Creosote Works Superfund site, Pensacola, Fla. The ability of indigenous soil microorganisms to remove these contaminants from aqueous solutions was determined by gas chromatographic analysis of organic extracts of biotreated groundwater. Changes in potential environmental and human health hazards associated with the biodegradation of this material were determined at intervals by Microtox assays and fish toxicity and teratogenicity tests. After 14 days of incubation at 30 degrees C, indigenous microorganisms effectively removed 100, 99, 94, 88, and 87% of measured phenolic and lower-molecular-weight polycyclic aromatic hydrocarbons (PAHs) and S-heterocyclic, N-heterocyclic, and O-heterocyclic constituents of creosote, respectively. However, only 53% of the higher-molecular-weight PAHs were degraded; PCP was not removed. Despite the removal of a majority of the organic contaminants through biotreatment, only a slight decrease in the toxicity and teratogenicity of biotreated groundwater was observed. Data suggest that toxicity and teratogenicity are associated with compounds difficult to treat biologically and that one may not necessarily rely on indigenous microorganisms to effectively remove these compounds in a reasonable time span; to this end, alternative or supplemental approaches may be necessary. Similar measures of the toxicity and teratogenicity of treated material may offer a simple, yet important, guide to bioremediation effectiveness.

Cognitive function in hypertensives treated with atenolol or propranolol.

STUDY OBJECTIVE: To test reports that beta blockers, particularly lipophilic forms, impair cognitive function and cause psychiatric disturbances. DESIGN: Randomized, double-blind, controlled crossover trial with eight-week treatment periods. PATIENTS: Sequential sample of 42 male veterans, with untreated diastolic blood pressures (DBP) between 90 and 110 mmHg, aged 35-64 years. INTERVENTIONS: Propranolol-LA, 80-mg tablets, or atenolol, 50-mg tablets, were given daily, incremented by one tablet at weekly intervals until DBP less than or equal to 90 mmHg. Hydrochlorothiazide was added, if necessary. MAIN RESULTS: Repeated-measures ANOVA was performed on all cognitive tests. Cognitive test performance was not affected by beta blocker therapy in seven of nine tests and was enhanced on Trail Making Test. Performance was impaired only on Digit Cancellation. Neither Speilberger's State Trait Anxiety Inventory nor the Beck Depression Inventory was affected by either beta blocker. CONCLUSIONS: Atenolol or propranolol therapy does not impair cognitive function or contribute significantly to psychiatric side effects.

Relative sensitivity of six estuarine fishes to carbophenothion, chlorpyrifos, and fenvalerate.

The acute toxicity (96-hr LC50) of carbophenothion, chlorpyrifos, and fenvalerate to six estuarine fishes was determined in flow-through laboratory tests. The atherinid fishes (Menidia menidia, M. peninsulae, M. beryllina, and Leuresthes tenuis) consistently were among the most sensitive species tested and were similar to each other in their sensitivity to pesticides. The sensitivity of sheepshead minnows (Cyprinodon variegatus) to carbophenothion was the same as that of the atherinids. For fenvalerate, the sheepshead minnow LC50 was an order of magnitude greater than that of the most sensitive atherinid, whereas the LC50 for chlorpyrifos and sheepshead minnows was two orders of magnitude greater. Gulf toadfish (Opsanus beta) were the least sensitive fish tested with carbophenothion and chlorpyrifos and their 96-hr LC50 for fenvalerate ranked between the LC50 for sheepshead minnows and atherinids. Test results were compared to acute toxicity data for other estuarine fishes and invertebrates.

Preliminary data on use of the inland silverside, Menidia beryllina, to control mosquito larvae.

A study of procedures for spawning and culture of the inland silverside, Menidia beryllina, was conducted. The efficacy of young Menidia, 20 to 22 and 31 to 33 days old, to control mosquito larvae was determined in the laboratory with first and second larval instars of the saltmarsh mosquito Aedes taeniorhynchus. Feeding trials were run at salinities of 1, 5, 15 and 25%. Field trials were also conducted to determine if Menidia would effectively control Culex quinquefasciatus in brackish water impoundments.

A new early life-stage toxicity test using the California grunion (Leuresthes tenuis) and results with chlorpyrifos.

California grunion were continuously exposed as embryos and fry to technical chlorpyrifos in two toxicity tests conducted in the same exposure apparatus. The first test, a 35-day early life-stage (ELS) test, began with approximately 2.5-day-old embryos that were exposed in flow-through aquaria in darkness until hatching was stimulated on Day 9 of exposure. The second toxicity test, a fry test, began with newly hatched fry and lasted 26 days. Test water temperature ranged from 23 to 26 degrees C and salinity from 24.5 to 34.0%. Results of the two tests were similar, indicating that exposure of embryos added little to the overall toxicity of chlorpyrifos to grunions. Percentage hatch of embryos was unaffected by the chlorpyrifos concentrations tested. Fry survival was apparently reduced in nominal concentrations greater than or equal to 1.0 micrograms/liter in both tests, but significantly so in concentrations greater than or equal to 1.0 micrograms/liter in the ELS test and greater than or equal to 2.0 micrograms/liter in the fry test. When compared with carrier controls, mean fish weights were significantly reduced in nominal chlorpyrifos concentrations greater than or equal to 0.5 micrograms/liter in the ELS test and greater than or equal to 1.0 micrograms/liter in the fry exposure. Mean bioconcentration factors were 770X for fish that survived the ELS test and 190X for those that survived the fry test. Results demonstrate the practicality of conducting ELS tests for the first time with a marine fish from the Pacific coastal waters of the United States.