Preparative isolation of anthocyanins by high-speed countercurrent chromatography and application of the color activity concept to red wine

Red pigments were isolated from wine and grape-skin extracts using preparative high-speed countercurrent chromatography (HSCCC) and identified by NMR and MS techniques. Four solvent systems were developed in order to separate anthocyanins with different polarities. Malvidin-3-glucoside was the major component present in young red wines, and up to 500 mg of pure malvidin-3-glucoside could be obtained from a single bottle of a red wine. Other isolated pigments were the malvidin- and peonidin-3,5-diglucosides, as well as acetyl-, coumaroyl-, and caffeoyl-derivatives of anthocyanins. Furthermore, condensed red wine pigments formed from malvidin-3-glucoside (vitisin A and acetylvitisin A) were isolated on a preparative scale. Isolated compounds were used as standards for quantification of anthocyanins in a range of red wines. The "color activity concept" was applied to red wine, and visual detection thresholds were determined for some of the isolated anthocyanins. Mono-glucosides were found to exhibit lower visual detection thresholds than di-glucosides and acylated anthocyanins.     

Comprehensive sensomics analysis of hop-derived bitter compounds during storage of beer

For the first time, quantitative LC-MS/MS profiling of 56 hop-derived sensometabolites contributing to the bitter taste of beer revealed a comprehensive insight into the transformation of individual bitter compounds during storage of beer. The proton-catalyzed cyclization of trans-iso-α-acids was identified to be the quantitatively predominant reaction leading to lingering, harsh bitter tasting tri- and tetracyclic compounds such as, e.g. the cocongeners tricyclocohumol, tricyclocohumene, isotricyclocohumene, tetracyclocohumol, and epitetracyclocohumol, accumulating in beer during storage with increasing time and temperature. The key role of these transformation products in storage-induced trans-iso-α-acid degradation was verified for the first time by multivariate statistics and hierarchical cluster analysis of the sensomics data obtained for a series of commercial beer samples stored under controlled conditions. The present study offers the scientific basis for a knowledge-based extension of the shelf life of the desirable beer's bitter taste and the delay of the onset of the less preferred harsh bitter aftertaste by controlling the initial pH value of the beer and by keeping the temperature as low as possible during storage of the final beverage.     

Bluetongue disease reaches Switzerland

Since 2006 bluetongue disease is rapidly spreading across Europe and reached Switzerland in October 2007. In the present article a short overview about the disease and the virus is given, and the first three clinical bluetongue disease cases in cattle, and the respective laboratory findings are presented.     

Two newly developed Erns-based ELISAs allow the differentiation of Classical Swine Fever virus-infected from marker-vaccinated animals and the discrimination of pestivirus antibodies

New generations of Classical Swine Fever virus (CSFV) marker vaccines have recently been developed in order to make emergency vaccination in case of a CSF outbreak more feasible. However, the application of a marker vaccine is dependent on the availability of an accompanying discriminatory test allowing differentiation of infected from vaccinated animals (DIVA). CP7_E2alf, the most promising live marker vaccine candidate currently available, is a genetically modified Bovine Viral Diarrhea virus expressing the E2 glycoprotein of CSFV strain Alfort/187. The DIVA principle going along with CP7_E2alf is based on the detection of CSFV E^rns-specific antibodies that are raised in the host upon CSFV infection but not after vaccination with the marker vaccine. The aim of this study was to develop novel DIVA tests to be used in combination with CP7_E2alf. Two indirect ELISAs (one for screening, the other one for confirmation purposes) using bacterially expressed recombinant E^rns proteins were designed and evaluated. Both ELISAs detected CSFV-specific antibodies against a broad range of strains and genotypes, and as early as 10 days after infection. They were able to distinguish CSFV-infected pigs from pigs vaccinated with CP7_E2alf and allowed discrimination of antibodies against ruminant pestiviruses in both, sera from domestic pigs and wild boar. Sensitivity and specificity of the screening ELISA was ≥95%. Thus, the ELISAs represent promising DIVA diagnostic tools, as well as an alternative to traditional pestivirus antibody differentiation by serum neutralization test.     

Evaluation of mosquito fish Gambusia holbrooki (Girard) as a partial and complete pellet replacement for juvenile barramundi Lates calcarifer (Bloch)

Barramundi (Lates calcarifer, Bloch) fed live mosquito fish (Gambusia holbrooki, Girard) exhibited faster growth and a better food conversion ratio (on a dry‐weight basis) than those fed an iso‐calorific ration of commercial barramundi pellets and a 50:50 ration of mosquito fish and pellets. The improved performance of barramundi fed mosquito fish was attributed to their higher protein content, higher protein to energy ratio and lower carbohydrate content. The essential amino acid indices and fatty acid profiles of the two diets were similar, suggesting that imbalances or deficiencies in these nutrients were not the limiting factor to the slower growth of barramundi fed on pellets. The use of mosquito fish as barramundi food offers a potential use of this noxious species and may allow barramundi to be cultured without the use of conventional fish meal.     

VASA (DDX4) is a Putative Marker for Spermatogonia,Spermatocytes and Round Spermatids in Stallions

Expression of the protein DDX4/MVH, or VASA, has been reported in germ cells of several species. The main objectives of this study were to (i) investigate VASA expression patterns in testicular cells of stallions at two different reproductive stages (pre‐pubertal and post‐pubertal) and (ii) evaluate the use of VASA antibody as a molecular marker for single germ cells from stallions. Testicular tissues were obtained from stallions and categorized as pre‐pubertal and post‐pubertal based on the formation of lumen and status of spermatogenesis on the cross section of seminiferous tubules. The results of Western blot showed a VASA protein band located at 76 kDa, indicating that the rabbit antibody has a cross‐reactivity with horse testicular tissues. The result of immunolabelling showed that VASA was expressed in the cytoplasm of spermatogonia at both reproductive stages and in spermatocytes and round spermatids at the post‐pubertal stage. GATA4‐positive Sertoli cells and Leydig cells located in the interstitial space were not immunolabelled with VASA. These results suggest that VASA can be utilized as a molecular marker for germ cells of stallions at pre‐pubertal and post‐pubertal stages. Interestingly, immunolabelling intensity was significantly higher in pachytene spermatocytes compared to spermatogonia and round spermatid. VASA antibody was also effective for staining of single germ cell preparations. In conclusion, VASA protein expression can be used as a marker for identification of spermatogonia, spermatocytes and round spermatids in testicular tissues of stallions.     

A new population of very high energy gamma-ray sources in the Milky Way

Very high energy gamma-rays probe the long-standing mystery of the origin of cosmic rays. Produced in the interactions of accelerated particles in astrophysical objects, they can be used to image cosmic particle accelerators. A first sensitive survey of the inner part of the Milky Way with the High Energy Stereoscopic System (HESS) reveals a population of eight previously unknown firmly detected sources of very high energy gamma-rays. At least two have no known radio or x-ray counterpart and may be representative of a new class of "dark" nucleonic cosmic ray sources.     

Altered Serum Thyrotropin Concentrations in Dogs with Primary Hypoadrenocorticism before and during Treatment

Background

Thyrotropin (TSH) can be increased in humans with primary hypoadrenocorticism (HA) before glucocorticoid treatment. Increase in TSH is a typical finding of primary hypothyroidism and both diseases can occur concurrently (Schmidt's syndrome); therefore, care must be taken in assessing thyroid function in untreated human patients with HA.

Objective

Evaluate whether alterations in cTSH can be observed in dogs with HA in absence of primary hypothyroidism.

Animals

Thirty dogs with newly diagnosed HA, and 30 dogs in which HA was suspected but excluded based on a normal ACTH stimulation test (controls) were prospectively enrolled.

Methods

cTSH and T4 concentrations were determined in all dogs and at selected time points during treatment (prednisolone, fludrocortisone, or DOCP) in dogs with HA.

Results

cTSH concentrations ranged from 0.01 to 2.6 ng/mL (median 0.29) and were increased in 11/30 dogs with HA; values in controls were all within the reference interval (range: 0.01–0.2 ng/dL; median 0.06). There was no difference in T4 between dogs with increased cTSH (T4 range 1.0‐2.1; median 1.3 μg/dL) compared to those with normal cTSH (T4 range 0.5‐3.4, median 1.4 μg/dL; P=0.69) and controls (T4 range 0.3‐3.8, median 1.8 μg/dL; P=0.35). After starting treatment, cTSH normalized after 2–4 weeks in 9 dogs and after 3 and 4 months in 2 without thyroxine supplementation.

Conclusions and Clinical Relevance

Evaluation of thyroid function in untreated dogs with HA can lead to misdiagnosis of hypothyroidism; treatment with glucocorticoids for up to 4 months can be necessary to normalize cTSH.     

Optimizing Electrical Activation of Porcine Oocytes by Adjusting Pre‐ and Post‐Activation Mannitol Exposure Times

Modifying electrical activation conditions have been used to improve in vitro embryo production and development in pigs. However, there is insufficient information about correlations of porcine embryo development with oocyte pre‐ and post‐activation conditions. The purpose of this study was to compare the developmental rates of porcine oocytes subjected to different mannitol exposure times, either pre‐ or post‐electrical activation, and to elucidate the reason for the optimal mannitol exposure time. Mannitol exposure times around activation were adjusted as 0, 1, 2 or 3 min. Blastocyst development were checked on day 7. Exposure of oocytes to mannitol for 1 or 2 min before electrical activation produced significantly higher blastocyst rates than exposure for 0 or 3 min. There was no significant difference in blastocyst rates when activated oocytes were exposed to mannitol for 0, 1, 2 or 3 min after electrical activation. While exposure of oocytes to mannitol for 1 min pre‐ and 3 min post‐activation showed significantly higher blastocyst development than 0 min pre‐ and 0 min post‐activation. It also showed higher maintenance of normal oocyte morphology than exposure for 0 min pre‐ and 0 min post‐activation. In conclusion, exposure of oocytes to mannitol for 1 min pre‐ and 3 min post‐activation seems to be optimal for producing higher in vitro blastocyst development of porcine parthenogenetic embryos. The higher blastocyst development is correlated with higher maintenance of normal morphology in oocytes exposed to mannitol for 1 min pre‐ and 3 min post‐activation.