Effects of the dioxin-like PCB 126 on larval summer flounder (Paralichthys dentatus).

Little is known about the sensitivity of teleost post-embryonic developmental stages (larval and metamorphic) to dioxin-like compounds. Larval and metamorphosing summer flounder (Paralichthys dentatus) were exposed to the dioxin-like polychlorinated biphenyl congener PCB 126, to compare their sensitivity to other fish species early life stages, and to document effects on metamorphic development, including degree of eye migration and gastric maturation. Median lethal doses (LD 50s) ranged between 30 and 220 ng/g wet mass, indicating that pre- and early-metamorphic stages of summer flounder are equally sensitive to the embryos of some of the most vulnerable fish species tested. Consistent with the presence of a functional aryl hydrocarbon receptor pathway, dose-dependent induction of cytochrome P-4501A (CYP1A) at four days post-exposure was observed in liver, stomach, intestine, and kidney of metamorphosing larvae. Stage-dependent differences in the epithelial distribution of CYP1A immunoreactivity were observed in the developing stomach of fish exposed to relatively high PCB 126 doses. A single sublethal dose (15 ng/g) delayed metamorphic progress (determined by the degree of eye migration), and resulted in abnormally high levels of cell proliferation and abnormal gastric gland morphology in late metamorphic stages. These results suggest that the post-embryonic larval and metamorphic stages of summer flounder, and potentially other fish species with complex life histories, are vulnerable to the effects of dioxin-like compounds, including lethality, developmental delay, and malformations.

Microbial communities in subpermafrost saline fracture water at the Lupin Au mine, Nunavut, Canada.

We report the first investigation of a deep subpermafrost microbial ecosystem, a terrestrial analog for the Martian subsurface. Our multidisciplinary team analyzed fracture water collected at 890 and 1,130 m depths beneath a 540-m-thick permafrost layer at the Lupin Au mine (Nunavut, Canada). 14C, 3H, and noble gas isotope analyses suggest that the Na-Ca-Cl, suboxic, fracture water represents a mixture of geologically ancient brine, approximately25-kyr-old, meteoric water and a minor modern talik-water component. Microbial planktonic concentrations were approximately10(3) cells mL(-1). Analysis of the 16S rRNA gene from extracted DNA and enrichment cultures revealed 42 unique operational taxonomic units in 11 genera with Desulfosporosinus, Halothiobacillus, and Pseudomonas representing the most prominent phylotypes and failed to detect Archaea. The abundance of terminally branched and midchain-branched saturated fatty acids (5 to 15 mol%) was consistent with the abundance of Gram-positive bacteria in the clone libraries. Geochemical data, the ubiquinone (UQ) abundance (3 to 11 mol%), and the presence of both aerobic and anaerobic bacteria indicated that the environment was suboxic, not anoxic. Stable sulfur isotope analyses of the fracture water detected the presence of microbial sulfate reduction, and analyses of the vein-filling pyrite indicated that it was in isotopic equilibrium with the dissolved sulfide. Free energy calculations revealed that sulfate reduction and sulfide oxidation via denitrification and not methanogenesis were the most thermodynamically viable consistent with the principal metabolisms inferred from the 16S rRNA community composition and with CH4 isotopic compositions. The sulfate-reducing bacteria most likely colonized the subsurface during the Pleistocene or earlier, whereas aerobic bacteria may have entered the fracture water networks either during deglaciation prior to permafrost formation 9,000 years ago or from the nearby talik through the hydrologic gradient created during mine dewatering. Although the absence of methanogens from this subsurface ecosystem is somewhat surprising, it may be attributable to an energy bottleneck that restricts their migration from surface permafrost deposits where they are frequently reported. These results have implications for the biological origin of CH4 on Mars.

Subseafloor microbial communities associated with rapid turbidite deposition in the Gulf of Mexico continental slope (IODP Expedition 308).

The subseafloor microbial communities in the turbidite depositional basins Brazos-Trinity Basin IV (BT Basin) and the Mars-Ursa Basin (Ursa Basin) on the Gulf of Mexico continental slope (IODP holes U1319A, U1320A, U1322B and U1324B) were investigated by PCR-dependent molecular analyses targeted to the small subunit (SSU) rRNA genes, dsrA and mcrA, and hydrogenase activity measurements. Biomass at both basins was very low, with the maximum cell or the SSU rRNA gene copy number <1 x 10(7) cells mL(-1) or copies g(-1) sediments, respectively. Hydrogenase activity correlated with biomass estimated by SSU rRNA gene copy number when all data sets were combined. We detected differences in the SSU rRNA gene community structures and SSU rRNA gene copy numbers between the basin-fill and basement sediments in the BT Basin. Examination of microbial communities and hydrogenase activity in the context of geochemical and geophysical parameters and sediment depositional environments revealed that differences in microbial community composition between the basin-fill and basement sediments in the BT Basin were associated with sedimentation regimes tied to the sea-level change. This may also explain the distributions of relatively similar archaeal communities in the Ursa Basin sediments and basement sediments in the BT Basin.

A versatile and sensitive tritium-based radioassay for measuring hydrogenase activity in aquatic sediments.

We present a method for the measurement of hydrogenase (H(2)ase) activity in aquatic sediments. The assay is based on the H(2)ase-mediated isotopic exchange between dissolved molecular hydrogen (H(2)) and water. A slurry of sediment material is incubated with a tritiated hydrogen (HT) headspace in a glass syringe on a rotary shaker. The method includes a procedure for preparing HT from radiolabeled sodium borohydride, which is a useful alternative to purchasing HT directly. A method for measuring HT specific activity based on liquid scintillation counting is also presented. Validation tests were run using live and frozen cultures of Clostridium pasteurianum and Desulfovibrio vulgaris, and freshly collected marine sediments. Adherence to Michaelis-Menten kinetics was demonstrated. An interassay coefficient of variation of 15% was determined using frozen C. pasteurianum cultures as reference material. Serial dilutions of cultures and sediments showed that measured H(2)ase activity scales with cell concentration, and indicate that the method can detect C. pasteurianum cell concentrations of between 300 and 3000 cells/ml. This technique allows measurement of H(2)ase activity in a variety of environmental samples, and will be particularly useful in the study of deep marine sediments with low microbial activity.

Age-dependent changes in the response of the stomach to thyroidal signaling in developmentally arrested larval summer flounder.

Thyroid-dependent stomach development occurs between approximately 35 and 50 days post-hatch (dph) in laboratory-reared summer flounder larvae. The process can be blocked by thiourea (TU), and TU effects are reversed by exogenous thyroxine (T4). To establish whether a window of sensitivity exists for T4-dependent gastric development, we arrested development of larvae with 0.39 mM TU given from 26 to 61dph, and measured weekly the developmental response of the stomach to 13 nM T4 for 0, 2, or 7 days. We examined cell proliferation in surface epithelium, gastric glands, and connective tissue, pepsinogen immunoreactivity in gastric glands, and Ulex Europaeus I (UEA I) lectin staining of gastric mucous neck cells, indicative of mucous content. In 26-47dph larvae, cell proliferation was increased 5- to 10-fold in all cell types after 2 days in T4, and returned to pretreatment values by 7 days of treatment. In 54dph fish, however, the proliferative response of gastric gland and surface epithelial cells decreased significantly from that of younger fish, while that of connective tissue was unchanged. For the differentiation markers, T4-induced mucous content increased at 54dph, but not in older fish, while pepsinogen induction was not different at any of the ages tested. The age interval between 47 and 54dph corresponds with the completion of gastric development in spontaneously metamorphosing larvae. The findings suggest that a critical window exists for the mitogenic actions of T4 in epithelial cells, but not for connective tissue cells, whereas no critical period was found for markers of differentiation.