A new sediment contact assay to assess particle-bound pollutants using zebrafish (<Emphasis Type="Italic">danio rerio</Emphasis>) embryos

Goal, Scope and Background. Based on a bioassay battery covering only primary producers and consumers as well as degraders, the potential ecological hazard of sediments to vertebrates cannot be estimated comprehensively. Therefore, there is an urgent need to develop and standardize integrated vertebrate-based test systems for sediment investigation strategies. Whereas vertebratebased in vitro systems have frequently been used for the investigation of aqueous samples, there is a significant lack of whole sediment assays. Thus, the purpose of the present study was: (1) to develop a rapid and reliable, but comprehensive method to investigate native sediments and particulate matters without preceding extraction procedures; (2) to compare the hazard potential of solid phase sediments to the effects of corresponding pore waters and organic extracts in order to characterize the bioavailability of the particle-bound pollutants; and (3) to relatively evaluate the embryotoxic effects of sediments from the catchment areas of the rivers Rhine, Neckar and Danube. Methods (or Main Features).  To investigate the toxicity of sediment samples on vertebrates, the standard embryo toxicity test with the zebrafish (Danio rerio; Hamilton-Buchanan 1922) according to DIN 38415-6 was modified with respect to exposure scheme and toxicological endpoints. Sediments from the catchment area of the Neckar River were assessed using pore waters, acetonic extracts and native sediments in order to get inside into the potential bioavailability of particle-bound pollutants. A comprehensive test protocol for the investigation of native sediments in the embryo toxicity test with the zebrafish is presented. Results and Discussion.  The fish embryo assay with Danio rerio can be carried out with both aqueous and organic sediment extracts as well as native (whole, solid phase) sediment samples. Elongation of exposure time from 48 to up to 196 h significantly increased the mortality. Using the fish egg assay with native sediments, a broad range of embryotoxic effects could be elucidated, including clear-cut dose-response curves for the embryotoxic effects of contaminated sediments; in contrast, absence of embryotoxic effects could be demonstrated even for the highest test concentrations of unpolluted sediments. With native sediments, embryotoxicity was clearly higher than with corresponding pore waters, thus corroborating the view that — at least for fish eggs — the bioavailability of particle-bound lipophilic substances in native sediments is higher than generally assumed. The relative ranking of sediment toxicity was identical using both native sediments and sediment extracts, EC₂₀ values of the latter, however, being eight time lower higher than with the native sediments. A comparison of the embryo toxic effects of samples from the Neckar area with locations along the Rhine and Danube rivers elucidated a broad range of results, thus indicating different levels of contamination. Conclusions.  A modified protocol of the zebrafish embryo test allows the assessment of sediment toxicity in both aqueous extracts and native sediments. The isolated investigation of pore waters may result in a clear-cut underestimation of the bioavailability of lipophilic particle-bound substances (as determined by native sediments). Recommendations and Perspectives.  The zebrafish embryo test with native (whole, solid phase) sediments appears very promising for the evaluation of the bioavailable fraction of lipophilic particle-bound substances and can therefore be recommended for the evaluation of vertebrate toxicity in tiered sediment test strategies and dredging directives such as the HABAB-WSV. Whereas acetone extracts may be tested as a rough estimation of embryotoxicity, native sediment samples will provide a more comprehensive and realistic insight into the bioavailable hazard potential     

Comparison of glucose concentrations in canine whole blood,plasma, and serum measured with a veterinary point-of-care glucometer

Previous studies have determined that, compared to whole blood, serum or plasma used in a portable blood glucometer (PBG) may provide more accurate results. We investigated the accuracy of a veterinary PBG (AlphaTRAK 2; Zoetis) for the measurement of glucose concentrations in serum, plasma, and whole blood compared to plasma glucose concentration measured by a biochemical analyzer. Blood samples from 53 client-owned dogs were collected. Lin concordance correlation coefficient (ρ_c) and Bland–Altman plots were used to determine correlation and agreement between the results obtained for the different sample types. Glucose concentration in whole blood measured by the veterinary PBG was more strongly correlated with the glucose concentration measured by the biochemical analyzer (ρ_c = 0.92) compared to plasma and serum glucose concentrations (ρ_c = 0.59 and 0.57, respectively). The mean differences between the glucose concentrations in whole blood, plasma, and serum measured by the veterinary PBG and the glucose concentration determined by the biochemical analyzer were 1.0, 6.3, and 6.7 mmol/L (18, 113, and 121 mg/dL), respectively. Our findings suggest that, when using this veterinary PBG, the accuracy of a glucose measurement obtained is higher when using whole blood compared to plasma or serum. Use of whole blood allows for more correct assessment and diagnosis, which are necessary for appropriate therapeutic intervention.     

Computed tomography of granulomatous pneumonia with oxalosis in an American alligator (Alligator mississippiensis) associated with Metarhizium anisopliae var anisopliae

An 18-yr-old, male, albino, American alligator (Alligator mississippiensis) was evaluated for decreased appetite and abnormal buoyancy. Computed tomography (CT) of the coelomic cavity showed multifocal mineral and soft tissue attenuating pulmonary masses consistent with pulmonary fungal granulomas. Additionally, multifocal areas of generalized, severe emphysema and pulmonary and pleural thickening were identified. The alligator was euthanized and necropsy revealed severe fungal pneumonia associated with oxalosis. Metarhizium anisopliae var. anisopliae was cultured from lung tissue and exhibited oxalate crystal formation in vitro. Crystals were identified as calcium oxalate monohydrate by X-ray powder defractometry. Fungal identification was based on morphology, including tissue sporulation, and DNA sequence analysis. This organism is typically thought of as an entomopathogen. Clinical signs of fungal pneumonia in nonavian reptiles are often inapparent until the disease is at an advanced stage, making antemortem diagnosis challenging. This case demonstrates the value of CT for pulmonary assessment and diagnosis of fungal pneumonia in the American alligator. Fungal infection with associated oxalosis should not be presumed to be aspergillosis.     

Surgical resolution of an avulsion fracture of the peroneus tertius origin in a giraffe (Giraffa camelopardalis reticulata)

A 4-mo-old, 185-kg male giraffe (Giraffa camelopardalis reticulata) was presented due to stifle effusion and lameness of 3-wk duration. Radiographs revealed a fracture of the extensor fossa of the lateral trochlear ridge of the femur at the origin of the peroneus tertius. Under anesthesia, dysfunction of the reciprocal apparatus was documented by flexing the stifle while the tarsus remained extended. An avulsion fracture of the origin of the peroneus tertius and extensor digitorum longus muscle was diagnosed. An exploratory arthroscopy of the femorotibial joint was followed by arthrotomy to excise the large bone fragment from its soft tissue attachments. Because of the fractious temperament of the animal, postoperative care was restricted to stall rest for 3 mo, and no postoperative complications arose. Only a mild residual lameness remained by 6 mo after surgery.     

Evaluation of medetomidine, ketamine and buprenorphine for neutering feral cats

A combination of medetomidine (M, 100 μg/kg), ketamine (K, 10 mg/kg) and buprenorphine (B, 10 μg/kg), administered by intramuscular injection, was evaluated for spaying and castration (neutering) of feral cats (n = 101). Eleven animals (11%) required supplemental anesthesia (isoflurane by mask) to maintain an adequate plane of surgical anesthesia. Atipamezole (A, 125 μg/kg) was administered subcutaneously at the completion of surgery. All cats recovered from surgery and were released the following day. A hemoglobin saturation (SpO(2)) value of < 95% was recorded at least once during anesthesia in all cats. This MKB combination can be used in a feral cat sterilization clinic, but isoflurane supplementation may be necessary. Further research is indicated to determine the clinical significance of the low SpO(2) values associated with this anesthetic regimen.     

Development of an in vitro model of feline cartilage degradation

Osteoarthritis is the most common arthropathy of mammalian species including cats. Cartilage degradation is central to the disorder and here we present, for the first time, an in vitro model of feline cartilage degradation which will be useful for further studies in this target species. Feline articular cartilage explant cultures were maintained for 28 days and in the presence of oncostatin M with and without interleukin (IL)-17, tumour necrosis factor (TNF), IL-1α, or IL-1β. Media samples and digested cartilage explants were analysed for glycosaminoglycan (GAG) and collagen content. The combination of IL-1β and OSM, both at 20 ng/ml, was able to promote GAG release to the greatest extent at 14 days. At 28 days, all groups showed relatively high release of GAG. At 14 days, only IL-1β and OSM in combination were associated with a statistically significant increase in collagen release over and above control tissue. IL-1β dose-response studies showed that an IL-1β dose of 10 ng/ml promotes a statistically significant increase in GAG breakdown when used with OSM, and higher doses of IL-1β did not result in significantly greater response. The model demonstrated both GAG and collagen degradation and will be of use for further understanding of feline cartilage metabolism and for screening of potential structure-modifying agents to be used in cats.     

Sportfisheries,conservation and sustainable livelihoods: a multidisciplinary guide to developing best practice

Ecotourism ventures in developing countries are often among the few alternatives for enhancing sustainable livelihoods without altering traditional ways of life. The best way forward is to continually develop and implement best practice guidelines and, in particular, to flexibly develop them to suit individual cases. We conduct a multidisciplinary assessment of best practice guidelines required to develop and sustain sportfishing tourism in developing countries, while enhancing local livelihoods and promoting environmental stewardship. In general, best practice guidelines should be developed around a sustainable livelihood framework that includes short‐term coping mechanisms and longer‐term capacity building. Sportfishing development that conforms to ecological and socially orientated criteria, founded on site‐specific research that captures local environmental and social complexities, has the potential to provide mutual benefits to tourists and local people, fuelling community development and enhancing the cultural experience of tourists. Best practice guidelines for sportfishing that do not address these dimensions are unlikely to result in a viable industry. Given the current interest and growth of sportfishing in developing countries, the proposed guidelines can help a range of end users manage, conserve and maximize livelihood benefits from their fishery.     

Characterisation of midgut digestive proteases from the maize stem borer Busseola fusca

BACKGROUND: Insect damage is a major constraint on maize production. Control of Busseola fusca (Fuller) in sub‐Saharan Africa is relatively ineffective; the major larval digestive enzymes were characterised with a view to developing future control strategies. RESULTS: Using BODIPY‐FL Casein, maximal activity was at pH 9.5, with six protease forms visualised by gelatin‐PAGE. Synthetic substrates and diagnostic inhibitors demonstrated the presence of serine proteases. Chymostatin was a potent inhibitor of general proteolysis (90%), providing strong evidence for the presence of chymotrypsin; it also caused significant inhibition (>95%) with SA₂PFpNA as substrate. The I₅₀ values for chymostatin with casein and SA₂PFpNA were 0.0075 µM and 0.06 µM respectively. Z‐Phe‐Arg‐pNA activity was inhibited by chymostatin and TLCK (50 and 30% respectively), suggesting the presence of trypsin‐like activity. BApNA hydrolysis was also strongly inhibited by chymostatin and TLCK (92 and 75%), suggesting trypsin activity, while SBBI, PMSF, pepstatin and E‐64 had no significant effect. Interestingly, SBBI (I₅₀ = 0.39 µM ) and SBTI both inhibited general proteolysis by approximately 70%, suggesting that SBBI's dual inhibitory role makes this inhibitor a potentially useful candidate for expression in maize for control of B. fusca. CONCLUSION: These results provide a basis for the rational design of insect‐resistant transgenic maize expressing protease inhibitors. Copyright © 2008 Society of Chemical Industry