Authentication of natural vanilla flavorings: isotopic characterization using degradation of vanillin into guaiacol

The isotopic investigation of vanillin has been extended to the new sources of natural precursors of vanillin recently introduced with a view of obtaining natural vanillin by biotechnological processes. To check the consistency of the isotopic composition of vanillin with that of the corresponding aromatic fragment, a selective degradation reaction into guaiacol was carried out. The reaction was shown to proceed without significant isotopic fractionation at the sites of interest, and an optimized procedure was defined from the results of an experimental design involving the quantity of reagent and the temperature and duration of the experiment. Guaiacol, which can be easily obtained in a reasonable time, is an interesting isotopic probe for carbon- and oxygen-isotope ratio mass spectrometry (IRMS) determinations. It provides (13)C information specific to the aromatic fragment and, combined with delta(13)C values measured on vanillin itself, it improves the authentication potential of carbon-IRMS. Thus, the natural status of ferulic acid may be characterized by significant (13)C depletion at the formyl site. Similarly, the oxygen-18 content of guaiacol is a better authentication tool than delta(18)O of vanillin because it does not suffer the drawback of being altered by chemical exchange of the sp(2) oxygen atom with water in industrial or laboratory procedures. Although collaborative studies are still necessary to improve the interlaboratory reproducibility of the delta(18)O parameters, consistent results can be obtained in an intralaboratory context. It is shown in particular that chemical oxidation of ferulic acid is characterized by a relative enrichment of the aromatic moiety of vanillin.     

Shoot biomass growth is related to the vertical leaf nitrogen gradient in Salix canopies

Plant canopy optimization models predict that leaf nitrogen (N) distribution in the canopy will parallel the vertical light gradient, and numerous studies with many species have confirmed this prediction. Further, it is predicted that for a given canopy leaf area, a low vertical light extinction coefficient will promote rapid growth. Therefore, the ideal canopy of fast-growing plants should combine high leaf area index with a low light extinction coefficient; the latter being reflected in a flat vertical leaf N gradient throughout the canopy. Based on data from an experimental Salix stand (six varieties) grown on agricultural land in central Sweden, we tested the hypothesis that shoot growth is correlated with vertical leaf N gradient in canopies of hybrid willows bred for biomass production, which could have implications for Salix breeding. Tree improvement research requires screening of growth-related traits in large numbers of plants, but assessment of canopy leaf N gradients by chemical analysis is expensive, time-consuming and destructive. An alternative to analytical methods is to estimate leaf N gradients nondestructively with an optical chlorophyll meter (SPAD method). Here we provide a specific calibration for interpreting SPAD data measured in hybrid willows grown in biomass plantations on fertile agricultural land. Based on SPAD measurements, a significant and inverse relationship (r(2) = 0.88) was found between shoot biomass growth and vertical leaf N gradient across canopies of six Salix varieties.     

Effect of oat decortication on chemical composition,in vitro digestibility and in situ degradability

The effect of decortication on chemical composition and degradability characteristics of four Danish oat varieties was investigated. Effective degradability (ED) and post-ruminal disappearance (PRD) were measured by in situ and mobile bag techniques respectively. Decorticated oat showed higher (p = .01) concentrations of crude protein (CP; 134 vs. 108 g/kg DM) and crude fat (71.6 vs. 53.1 g/kg DM) and a higher (p = .001) organic matter digestibility (OMD; 888 vs. 703 g/kg OM) than oat. The content of total fatty acids (FA) in DM was higher in decorticated oat. The proportion of linoleic acid (C18:2 n6) increased (p < .05) due to decortication, while the linolenic acid (C18:3 n3) proportion of total FA decreased in decorticated oat. Decortication increased (p = .01) the concentration of amino acids (AA), but the proportion of lysine in total AA decreased (p < .002). Effective degradability (ED) of both DM and CP was (p < .001) higher in decorticated oat. Decortication increased the total tract disappearance (TTD) and PRD of CP (p < .001). In conclusion, decortication can be used as a practical approach to increase the nutritional value of oat.     

Endogenous development of Hemolivia mauritanica (Apicomplexa: Adeleina: Haemogregarinidae) in the marginated tortoise Testudo marginata (Reptilia: Testudinidae): evidence from experimental infection

Six young tortoises Testudo marginata Schoepff, 1792 were experimentally infected with Hemolivia mauritanica (Sergent et Sergent, 1904). The prepatent period ranged from 6 to 8 weeks. Young, smaller, club-like forms (6-9 x 3-6 Am) of gametocytes appeared in the peripheral blood first, whereas mature, elongated, cylindrical forms (9-12 x 5-7 Am) were detected after 1-2 weeks and predominated during later patency. Three of the infected tortoises were euthanized and dissected to study the endogenous stages. Meronts occurred in the cells of the reticulo-endothelial system and in the erythrocytes; these were observed mostly in parenchymatous organs. Mature forms measured 14.2 x 9.3 microm and contained 7-12 merozoites. Cysts with two (exceptionally one) cystozoites were also found predominantly in parenchymatous organs and measured 14.8 x 7.9 microm. Pathological changes attributable to Hemolivia were mild and limited to liver and kidneys. The role of individual developmental stages of haemogregarines is discussed with respect to evolution of heteroxenous life cycle and long-term persistence of parasites in their intermediate hosts.     

Viral and bacterial agents associated with experimental transmission of infectious proventriculitis of broiler chickens.

Proventriculitis of broilers can be reproduced by oral inoculation of day-old chicks with a proventricular homogenate from affected 3-wk-old broilers. The objective of the following studies was to isolate from this homogenate viral and bacterial isolates that could produce proventriculitis. A monoclonal antibody to infectious bursal disease virus (IBDV) was used to precipitate virus from the homogenate. A primary chicken digestive tract cell culture system was also used to isolate virus from a 0.2-microm filtrate of the homogenate, and a bacterium was also isolated from the homogenate. In trial 1, day-old birds were orally inoculated with either proventriculus homogenate or monoclonal antibody immunoprecipitated IBDV (MAB-IBDV). At 4, 7, 14, and 21 days postinfection (PI), 12 birds from each treatment group were subjected to necropsy. In trial 2, day-old birds were orally inoculated with either infectious proventriculus homogenate, suspect virus isolated in cell culture and propagated in embryo livers and spleens, or a bacterial isolate. Twelve birds from each treatment were subjected to necropsy at days 7, 14, 21, and 28 PI. In trial 3, treatments were maintained in negative pressure isolation chambers, and an additional treatment included virus plus bacterial isolate. Twenty-four birds from each treatment were subjected to necropsy at day 21 PI. In trial 1, infectious homogenate decreased body weight and relative gizzard weights at 4, 7, 14, and 21 days PI. Proventriculus relative weight was increased at days 7, 14, and 21 PI, and proventriculus lesion scores were increased at days 14 and 21 PI. Bursa/spleen weight ratios were decreased at day 14, and feed conversion was increased at days 4 and 21. The MAB-IBDV treatment decreased proventriculus and gizzard relative weights at day 4 PI, increased proventriculus lesion scores and bursa/spleen weight ratios at day 14, and decreased heterophil/lymphocyte ratios at day 21. In trial 2, all infected birds had significantly higher mean relative proventriculus weights at 21 days PI and had higher 4-wk mean proventriculus scores as compared with both control groups. In trial 3, birds treated with homogenate and birds treated with both suspect virus and the bacterial isolate had significantly higher proventriculus lesion scores; higher relative weights of proventriculus, gizzard, liver, and heart; lower body weights; and lower relative bursa weights compared with the saline control group. These studies suggest that infectious proventriculitis has a complex etiology involving both viral and bacterial infection.     

Appraisal and standardization of curvilinear velocity (VCL) cut‐off values for CASA analysis of Japanese quail (Coturnix japonica) sperm

One of the basic steps in objective analysis of sperm motility is the subdivision of a motile sperm population into slow, medium and rapid categories based on their velocity. However, for CASA analysis of quail sperm, the velocity values for categorization of slow, medium and rapid sperm have not yet been standardized. To identify the cut‐off values of “velocity curvilinear” (VCL) for quail sperm categorization, we captured and analysed 22,300 tracks of quail sperm using SCA^®‐CASA. The median and mean VCL values were 85 and 97 μm/s. To define the VCL cut‐off values, we used two methods. In the first, we identified the upper (rapid sperm) and lower (slow sperm) cut‐off values using: (i) median VCL ± 25% or ± 50% or ± 75% of median VCL value; (ii) first and third quartile values of VCL data (i.e. 25% cut‐off setting); and (iii) 33% and 66% of VCL data. Among these settings, sperm categories and their corresponding motility characteristics recorded using the “25%” setting (i.e. slow ≤36 ≤ medium ≤154 ≤ rapid) were found the most realistic and coherent with male ranking by fertility. In the second method, we calculated heteroscedasticity in the total VCL data using PCA and the two‐step clustering method. With this approach, the mean of the high and low clusters was 165 and 51 μm/s, respectively. Together, the mean from two methods suggested that, for SCA^®‐CASA categorization of quail sperm, sperm should be classed as “rapid” at VCL ≥160 μm/s and “slow” at VCL ≤45 μm/s.     

SONOGRAPHY OF THE MUSCULOSKELETAL SYSTEM IN DOGS AND CATS

Sonography of the musculoskeletal system in dogs and cats was undertaken to evaluate the application of this imaging procedure in orthopedics. In most of the patients a 7,5 MHz linear transducer was used because of its flat application surface and its resolving power. The evaluation of bone by sonography is limited, but sonography can provide addition information regarding the bone surface and surrounding soft tissue. Ultrasound is valuable for assessing joint disease. Joint effusion, thickening of the joint capsule and cartilage defects can be identified sonographically. It is also possible to detect bone destruction. Instabilities are often identified with the help of a dynamic examination. Soft tissue abnormalities of the musculoskeletal system lend themselves to sonographic evaluation. Partial or complete muscles or tendon tears are able to be differentiated and the healing process can be monitored. Most of the diseases that are in the area of the biceps or the achilles tendon, such as dislocation of the tendon, old injuries with scarification, free dissecates in the tendonsheath, tendinitis and/or tendosynovitis can be differentiated by sonography. In addition, with clinical and laboratory findings, it is often possible to make a correct diagnosis with ultrasound in patients with abscesses, foreign bodies, hematomas, soft tissue tumors and lipomas.     

Immunohistochemical differentiation of reactive from malignant mesothelium as a diagnostic aid in canine pericardial disease

Objectives

To develop a provisional immunohistochemistry panel for distinguishing reactive pericardium, atypical mesothelial proliferation and mesothelioma in dogs.

Materials and Methods

Archived pericardial biopsies were subject to haematoxylin and eosin staining, immunohistochemistry for cytokeratin, vimentin, insulin‐like growth factor II mRNA‐binding protein 3, glucose transporter 1 and desmin. Samples were scored for intensity and number of cells stained.

Results

Ten biopsies of reactive mesothelium, 17 of atypical mesothelial proliferation, 26 of mesothelioma and five of normal pericardium were identified on the basis of haematoxylin and eosin staining. Cytokeratin and vimentin were expressed in all biopsies, confirming mesothelial origin. Normal pericardial samples had the lowest scores for insulin‐like growth factor II mRNA‐binding protein 3, glucose transporter 1 and desmin. Mesothelioma and atypical proliferative samples were similar to each other, with higher scores for insulin‐like growth factor II mRNA‐binding protein 3 and glucose transporter 1 than the reactive samples. Desmin staining was variable. Insulin‐like growth factor II mRNA‐binding protein 3 was the best to distinguish between disease groups.

Clinical Significance

An immunohistochemistry panel of cytokeratin, vimentin, insulin‐like growth factor II mRNA‐binding protein 3 and glucose transporter 1 could provide superior information compared with haematoxylin and eosin staining alone in the diagnosis of cases of mesothelial proliferation in canine pericardium, but further validation is warranted.     

Age of Neoproterozoic bilatarian body and trace fossils, White Sea, Russia: implications for metazoan evolution

A uranium-lead zircon age for a volcanic ash interstratified with fossil-bearing, shallow marine siliciclastic rocks in the Zimnie Gory section of the White Sea region indicates that a diverse assemblage of body and trace fossils occurred before 555.3 +/- 0.3 million years ago. This age is a minimum for the oldest well-documented triploblastic bilaterian Kimberella. It also makes co-occurring trace fossils the oldest that are reliably dated. This determination of age implies that there is no simple relation between Ediacaran diversity and the carbon isotopic composition of Neoproterozoic seawater.     

Mapping QTL controlling soybean seed sucrose and oligosaccharides in a single family of soybean nested association mapping (SoyNAM) population

The meal value of Soybean for monogastric animals is determined partly by sucrose, raffinose and stachyose. Of these, sucrose is desirable, while raffinose and stachyose are indigestible, causing flatulence and abdominal discomfort. The objective of this study was to identify quantitative trait loci (QTL) controlling seed sucrose, raffinose, and stachyose in a set of 140 SoyNAM (Nested Association Mapping) recombinant inbred lines (RILs), developed from the cross between lines IA3023 and LD02‐4485. A total of 3,038 SNP markers from the Illumina SoyNAM BeadChip SNP were used to map the QTLs for sucrose and the RFOs, raffinose, and stachyose. Significant genotypic differences (p < .001) among RILs were observed for sucrose, raffinose and stachyose contents across years. A 3038 Illumina SoyNAM BeadChip SNPs identified three QTLs for sucrose, one on chromosome 1, explaining 10% variance and two on chromosome 3 each explaining 22%. Raffinose QTL was detected on chromosome 6, explaining 6% variance. The mapped QTLs were novel and spanned regions harbouring candidate genes with roles in plant growth including seed development.