Effects of natural (flooding and drought) and anthropogenic (copper and salinity) stressors on the earthworm Aporrectodea caliginosa under field conditions

Laboratory toxicity tests are usually conducted under stable conditions, while exposures in the field occur under variable environmental conditions. Field studies are therefore more appropriate in understanding the effect of single or mixed pollutants in the environment. Short-term interactive effects of natural environmental factors (flooding and drought) and anthropogenic stressors (copper and salinity) on the earthworm Aporrectodea caliginosa were investigated using outdoor microcosm experiments. Specimens of the lumbricid earthworm A. caliginosa were exposed in microcosms loaded with soils with increasing salinity with electrical conductivity (EC) ranging from 0.08 to 1.05 dS m^?1, with or without copper oxychloride spray treatments at recommended and elevated dosages. The experiment was conducted in August/September (end of winter) and repeated in November/December (end of spring) in the Stellenbosch area, South Africa to attain exposure under variable environmental conditions. In the soil, changes in Cu concentrations along the gradient of salinity were monitored using three methods: CaCl₂, DTPA and nitric acid extraction. Survival, weight change, and cocoon production of worms and body Cu concentrations were used as indices of interaction. None of the three extraction methods could reveal interactive effects between salinity and Cu in both seasons either because concentrations of Cu were too low or below detection limits. Copper, on its own, did not have a significant effect on the measured worm parameters during both seasons. During the winter experiment, flooding of microcosms significantly reduced the survival and affected the weight change of worms, and probably caused leaching of chemicals. Interaction between salinity and Cu had no significant effects on the measured worm parameters in winter while salinity only had a significant negative effect on weight change of worms. During spring, significant synergistic interaction between salinity and Cu occurred but only at 0.3 dS m^?1 by day 14 of the exposure period. At this and higher levels, salinity had a significant individual effect on survival and weight change of worms at days 14 and 28. The results indicate that higher toxicity of salinity could be expected during the dry spring periods than during the wet winter periods which are typical for southern temperate zones.     

Eliminating amino acid interference during spectrophotometric NH4 analysis

Amino acids can interfere with NH₄⁺ in spectrophotometric NH₄⁺ determination hampering accurate quantification of the fate of NH₄⁺ and dissolved organic N in soils. Serious interference has been reported for soils rich in organic matter, and for soils that have been fumigated, oven-dried or fertilized where between 5 and 60% of the NH₄⁺ detected could be attributed to amino acid interference. We investigated whether a combination of gas diffusion and the classical Berthelot method can eliminate this interference, increasing the selectivity of the NH₄⁺ analysis. We tested this approach using synthetic amino acid solutions and a large set of terrestrial samples (n = 353), including pore water samples, (fumigated or dried) soil extracts, and extracts of manures, composts and crop residues. The evaluated method produced accurate (recovery > 99.7%) and reproducible (standard error = 2.2%) NH₄⁺ concentrations, eliminating any interference between amino acids and NH₄⁺ (interference < 0.3%). Interference from K₂SO₄ was also eliminated. The method is robust, broadly applicable and will improve our understanding of nitrogen cycling in various ecosystems, in particular those where high levels of amino acids occur.     

Identification and mapping of quantitative resistance to late blight (<Emphasis Type="Italic">Phytophthora infestans</Emphasis>) in <Emphasis Type="Italic">Solanum habrochaites</Emphasis> LA1777

Late blight (Phytophthora infestans) can have devastating effects on tomato production over the whole world. Most of the commercial cultivars of tomato, Solanum lycopersicum, are susceptible. Qualitative and quantitative resistance has been described in wild relatives of tomato. In general qualitative resistance can more easily be overcome by newly evolved isolates. Screening of three S. habrochaites accessions (LA1033, LA2099 and LA1777) through a whole plant assay showed that accession LA1777 had a good level of resistance to several isolates of P. infestans. To explore the potential in this wild species, an introgression line (IL) population of S. habrochaites LA1777 was used to screen individual chromosome regions of the wild species by a detached leaf assay. Two major isolates (T_1,2 and T_1,2,4) were used and two parameters were measured: lesion size (LS), and disease incidence (DI). Substantial variation was observed between the individual lines. QTLs were identified for LS but not for DI. The presence of five QTLs derived from LA1777 (Rlbq4a, Rlbq4b, Rlbq7, Rlbq8 and Rlbq12) results in unambiguous higher levels of resistance. All QTLs co-localized with previously described QTLs from S. habrochaites LA2099 except QTL Rlbq4b, which is therefore a novel QTL.     

Uptake,distribution and metabolic fate of 3H-triforine in plants. II. Long-term experiments

Uptake of ³H-triforine by tomato and barley seedlings from soil with a high organic matter content was much less efficient than from aqueous suspensions, even though the period of exposure was much longer—at least 1 week (“long-term treatment”) vs 1 day (“short-term treatment”). After transplanting to fresh soil, part of the label in the roots was lost probably by desorption. Distribution of label in tomato shoots was as irregular as after short-term treatment; label was virtually confined to the leaves which expanded before about 14 days after cessation of the treatment. In shoots of barley seedlings which were pretreated in an aqueous suspension of ³H-triforine for 1 day before being subjected to a long-term soil treatment, almost all radioactivity present could be ascribed to uptake during the pretreatment phase. The distribution pattern strongly resembled that obtained after short-term treatment, hardly any label being found in leaves which unfolded after the pretreatment phase. Rates of conversion of ³H-triforine in barley shoots depended to some extent on whether or not seedlings were transplanted to fresh soil after 1 week.     

Susceptibility of sheep to European bat lyssavirus type-1 and -2 infection: a clinical pathogenesis study

European bat lyssaviruses (EBLVs) have been known to cross the species barrier from their native bat host to other terrestrial mammals. In this study, we have confirmed EBLV-1 and EBLV-2 susceptibility in sheep (Ovis ammon) following intracranial and peripheral (intramuscular) inoculation. Notably, mild clinical disease was observed in those exposed to virus via the intramuscular route. Following the intramuscular challenge, 75% of the animals infected with EBLV-1 and 100% of those that were challenged with EBLV-2 developed clinical signs of rabies and then recovered during the 94-day observation period. Disease pathogenesis also varied substantially between the two viruses. Infection with EBLV-1 resulted in peracute clinical signs, which are suggestive of motor neuron involvement. Antibody induction was observed and substantial inflammatrory infiltrate in the brain. In contrast, more antigen was detected in the EBLV-2-infected sheep brains but less inflammatory infiltrate and no virus neutralising antibody was evident. The latter involved a more protracted disease that was behaviour orientated. A high infectious dose was required to establish EBLV infection under experimental conditions (> or =5.0 logs/ml) but the infectious dose in field cases remains unknown. These data confirm that sheep are susceptible to infection with EBLV but that there is variability in pathogenesis including neuroinvasiveness that varies with the route of infection. This study suggests that inter-species animal-to-animal transmission of a bat variant of rabies virus to a terrestrial mammal host may be limited, and may not always result in fatal encephalitis.     

Photometric application of the MTT- and NRR-assays as biomarkers for the evaluation of cytotoxicity ex vivo in Eisenia andrei

In soils, the toxicity of noxious substances such as metals is determined by a number of different factors of chemical, physical, biological, and environmental origin. Addressing these factors as a sum of different parameters, we studied these effects in soil-dwelling indicator species by using ‘sub-cellular’ biomarkers, to see if they can provide an early stage response to environmental disturbances. ‘Sub-cellular’ biomarkers such as the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) and the Neutral Red Retention (NRR) assays are well established for the assessment of cytotoxicity of toxicants on cell cultures. Both assays can be carried out in 96-well microplates and evaluated photometrically. To be able to assess the risk potential of contaminated soils using these assays, without changing the soils chemically or physically to accommodate cell cultures, the assays were adapted for ex vivo use. The assays were performed on coelomic liquid extracted from exposed earthworms. Eisenia andrei was exposed in a standardized laboratory soil substrate (OECD soil) spiked with cadmium at a range of concentrations known to be cytotoxic to earthworms. Earthworms were also exposed to ultramafic soils as field-relevant soil substrates, containing high concentrations of a mixture of metals of natural origin such as chromium, cobalt, manganese, and nickel. A significant response was measured with both assays in the coelomocytes of the earthworms exposed to the Cd-spiked OECD soil as well as to the ultramafic soils. It was shown that as biomarkers for the assessment of ‘sub-cellular’ toxicity, the photometrical application of the MTT and the NRR assay promised to be rapid, objective, and easily conductible tools.     

Impacts of climate change on the complex life cycles of fish

To anticipate the response of fish populations to climate change, we developed a framework that integrates requirements in all life stages to assess impacts across the entire life cycle. The framework was applied on plaice (Pleuronectes platessa) and Atlantic herring (Clupea harengus) in the North Sea, Atlantic cod (Gadus morhua) in the Norwegian/Barents Seas and European anchovy (Engraulis encrasicolus) in the Bay of Biscay. In each case study, we reviewed habitats required by each life stage, habitat availability, and connectivity between habitats. We then explored how these could be altered by climate change. We documented environmental processes impacting habitat availability and connectivity, providing an integrated view at the population level and in a spatial context of potential climate impacts. A key result was that climate‐driven changes in larval dispersion seem to be the major unknown. Our summary suggested that species with specific habitat requirements for spawning (herring) or nursery grounds (plaice) display bottlenecks in their life cycle. Among the species examined, anchovy could cope best with environmental variability. Plaice was considered to be least resilient to climate‐driven changes due to its strict connectivity between spawning and nursery grounds. For plaice in the North Sea, habitat availability was expected to reduce with climate change. For North Sea herring, Norwegian cod and Biscay anchovy, climate‐driven changes were expected to have contrasting impacts depending on the life stage. Our review highlights the need to integrate physiological and behavioural processes across the life cycle to project the response of specific populations to climate change.     

Toxicity of 2,3,7,8-TCDD and short-term bioaccumulation by earthworms (oligochaeta)

The earthworms Allolobophora catiginosa and Lumbricus rubellus were used to study the toxicity of 2,3,7,8-TCDD (dioxin) for earthworms. The earthworms were exposed to soil containing concentrations ranging from 0.05 to 5.0 μgg^?1. No worms were killed or showed any other observable toxicological effects when exposed to concentrations up to 5 μg g^?1 for 85 days in soil. The lethal threshold concentration for TCDD to earthworms falls between 5 and 10 μg g^?1 in this study. In soils containing 0.05 μg g^?1 earthworms accumulated TCDD up to 5 times the original soil concentration within 7 days. Worms were also exposed to TCDD on filter paper to study the behaviour of earthworms and the uptake of TCDD after surface contact. The earthworms did not avoid TCDD in their environment, indicating an indifference to it. No active penetration of TCDD into the body occurred where earthworms were exposed to surface concentrations. No indication was found of possible biological breakdown of TCDD on passing through the earthworm gut, although the search for metabolites was limited to the mono-, bi- and trichlorinated dioxins. There was a steady decrease (a T_{$\text{1}\text{2}$}-value of 80–400 days) in the amount of TCDD recovered from worm-worked soil compared to soil without worms.     

Coral reef organisms as bioregion indicators off Halmahera,Moluccas, Indonesia

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