A COMPARISON OF DIATRIZOATE* AND IOXAGLATE† FOR POSITIVE CONTRAST SHOULDER ARTHROGRAPHY IN DOGS

In six experimental dogs, arthrographic quality and synovial inflammatory response with shoulder arthrography comparing meglumine-sodium diatrizoate (Urovison) and the monoacidic dimer, meglumine-sodium ioxaglate (Hexabrix) was evaluated. In our study initial films were of equally high diagnostic quality for both contrast media, but delayed films significantly favored ioxaglate for diagnostic quality. The rise in white blood cells in synovial fluid samples collected 24 hours after the arthrographic procedure was significantly lower after the use of ioxaglate. Histologic examination performed 14 days after the intra-articular injection revealed no drug related lesions.     

Development of a gas-liquid chromatographic method for the analysis of fatty acid tryptamides in cocoa products

The determination of the occurrence and level of cocoa shells in cocoa products and chocolate is an important analytical issue. The recent European Union directive on cocoa and chocolate products (2000/36/EC) has not retained the former limit of a maximum amount of 5% of cocoa shells in cocoa nibs (based on fat-free dry matter), previously authorized for the elaboration of cocoa products such as cocoa mass. In the present study, we report a reliable gas-liquid chromatography procedure suitable for the determination of the occurrence of cocoa shells in cocoa products by detection of fatty acid tryptamides (FATs). The precision of the method was evaluated by analyzing nine different samples (cocoa liquors with different ranges of shells) six times (replicate repeatability). The variations of the robust coefficient of variation of the repeatability demonstrated that FAT(C22), FAT(C24), and total FATs are good markers for the detection of shells in cocoa products. The trueness of the method was evaluated by determining the FAT content in two spiked matrices (cocoa liquors and cocoa shells) at different levels (from 1 to 50 mg/100 g). A good relation was found between the results obtained and the spiking (recovery varied between 90 and 130%), and the linearity range was established between 1 and 50 mg/100 g in cocoa products. For total FAT contents of cocoa liquor containing 5% shells, the measurement uncertainty allows us to conclude that FAT is equal to 4.01 +/- 0.8 mg/100 g. This validated method is perfectly suitable to determine shell contents in cocoa products using FAT(C22), FAT(C24), and total FATs as markers. The results also confirmed that cocoa shells contain FAT(C24) and FAT(C22) in a constant ratio of nearly 2:1.     

Indices of soil nitrogen availability for an ecological site classification of British forests

An adequate supply of nitrogen (N) is essential for the successful establishment and sustainable productivity of forest stands. N deficits may necessitate the use of artificial fertilisers. Availability of N in the inorganic forms, and the relative abundance of the NO₃-N and NH₄-N components, influences the species composition of natural forest vegetation. Hence it is essential to use reliable measures of soil N supply that fully reflect its ecological significance. The new Ecological Site Classification (ESC) used in British forestry employs a multi-factorial definition of soil nutrient regime (SNR), including soil N. To develop this, a soil and vegetation study was made at 89 forest sites throughout Great Britain covering the major soil types used for forestry. “Total N” levels were compared with separate pre- and post-incubation measures of the two inorganic N components as potential indices of soil N supply. Multivariate statistical analysis showed that the major discriminant chemical variables for the sampled soils were pH, calcium and NO₃-N and that these were also the main variables influencing the species composition of the ground vegetation. Total N and NH₄-N were less effective discriminant variables for these sites. In some infertile soils the levels of NH₄-N or total N may be of greater importance, as NO₃-N is usually in very limited supply. A multivariate gradient of SNR, which incorporates the NO₃-N measures, has been adopted for use within the ESC system. The position of a site on this gradient can be estimated quantitatively from soil type, ground vegetation species composition and humus type. This enables soil N supply and overall SNR to be assessed in a simple but effective way that guides the operational management of British forest soils for sustainable productivity. It will also be possible to use these techniques to monitor the nutritional status of forest sites over time.     

A proposal for enhanced EU herbage VCU and DUS testing procedures

Herbage production is regarded as having environment-friendly credentials. However, as the ruminant production it supports is facing major challenges on sustainability, environmental footprint and human health concerns, EU herbage cultivar testing must contribute to the solutions. Before new cultivars can be sold in a member state (MS) and gain EU-wide marketing, they must pass official tests to prove they are both novel (distinct, uniform and stable, DUS) with improved value for cultivation and use (VCU). Herbage species present specific challenges, as their allogamy imposes a wide within-cultivar variation that adds complexity to DUS tests and their “value” is only realized in ruminant produce. Current VCU systems measure production, chemical composition and disease/stress tolerances, often on large numbers of candidate cultivars, but prohibitive labour costs and logistics mean that animal intake, ruminant output or environmental benefits cannot be measured directly. Furthermore, some candidate cultivars with proven superior VCU fail DUS even though the non-distinct comparison is with a significantly lower performing registered cultivar. To resolve these problem cases, a “vmDUS” distinctness tool is proposed, which uses molecular markers but conforms to UPOV-declared principles. A short overview of current grassland research shows smart proxy measures of animal value can easily and quickly be adopted into an integrated pan-European (EU-VCU) test network. The proposed EU-VCU scheme will reallocate test resources to conduct these additional tests by placing MS in data sharing collaborations, while retaining their national listing authority. The benefits to all stakeholders from adopting these new testing procedures are considered.     

Comparison of methods to detect gastrointestinal parasites in llamas and alpacas

OBJECTIVE: To compare relative sensitivity and overall yields of various methods of fecal examination for gastrointestinal parasites in llamas and alpacas. DESIGN: Prospective study. SAMPLE POPULATION: Fecal samples from 42 alpacas and 62 llamas. PROCEDURES: Fecal samples were analyzed via direct smear, a modified McMaster technique with sucrose solution or saturated saline (approx 36% NaCl) solution, and a centrifugation-flotation procedure. McMaster flotation chambers were examined 15 and 60 minutes after loading. Centrifugation-flotation samples were examined after 10 and 60 minutes of flotation. The proportions of samples with positive results and concentrations of parasites were compared among methods. RESULTS: The centrifugation-flotation technique yielded more positive results than other methods for all parasites except small coccidia. Longer flotation time increased the proportion of positive results and parasite concentrations for all parasites except Nematodirus spp. Longer time in the McMaster chamber made little difference. By use of the modified McMaster technique, sucrose solution yielded more positive results for Trichuris spp, Eimeria macusaniensis, and strongyles, whereas saline solution yielded more positive results for Nematodirus spp and small coccidia. The saline solution McMaster test yielded more positive results for small coccidia than did most other methods, and the sucrose McMaster technique yielded more positive results for Trichuris spp. CONCLUSIONS AND CLINICAL RELEVANCE: The centrifugation-flotation technique appeared to offer clear advantages in detecting infection with E macusaniensis, Trichuris spp, Nematodirus spp, and capillarids. The saline McMaster technique appeared to offer an advantage in detecting small coccidia.     

Micro-environmental conditions modulate protein secretion and infectivity of the Trichinella spiralis L1 larva

After digestion of infected meat the free L1 of Trichinella spp. penetrate the intestinal mucosa where they moult to the mature adult stage. We have used proteomics to identify changes in protein secretion during in vitro culture of free T. spiralis muscle larvae under different environmental conditions, and to correlate these changes with their infectivity in mice. Muscle larvae were cultured in different media (RPMI-1640, C-199 and HBSS) under conditions of anaerobiosis, microaerobiosis and in 5% CO₂ at 37 °C. Following incubation the larval excretory/secretory proteins were analysed by two-dimensional gel electrophoresis and the larvae were used to orally infect naïve CD1 mice. For all culture media tested, infectivity of the L1 was preserved following incubation in anaerobic conditions. In contrast, the infectivity of worms cultured in nutrient-rich media was almost completely abolished in both microaerobiosis and in the presence of 5% CO₂. Some infectivity was retained in poor or reduced culture media. Comparative analysis of larval infectivity and protein secretion showed that loss of infectivity correlated with the appearance of non-tyvelosylated proteins that in turn may be related to the onset of moulting.     

The Role of Saprotrophy and Virulence in the Population Dynamics of Botrytis cinerea in Vineyards

ABSTRACT Change in relative frequencies of the three main genetic types of Botrytis cinerea (Group I, Group II vacuma, and Group II transposa) were monitored over time from 1998 to 2000 in three Bordeaux vineyards not treated with fungicides. During 2000, Group I isolates, detected mostly at flowering comprised only 2.5% of the entire population. Within Group II, the complementary frequencies of vacuma and transposa isolates differed significantly depending on grapevine phenological stages and organs. Every year and at all sites, including one noble rot site, transposa isolates dominated at every stage, particularly on overwintering canes and at harvest (greater than 86.7% on berries). The complementary frequency of vacuma isolates reached a maximum on senescing floral caps (between 23.5 and 71.4%) and then decreased significantly until harvest on leaves and berries. In pathogenicity tests on grape berries, transposa isolates were significantly more virulent than were vacuma isolates. Mycelial growth rate was negatively correlated with virulence, notably on leaves in transposa and with double resistance to the fungicides carbendazim and vinclozolin. In vacuma, this double resistance was positively correlated with virulence on leaves. Change in the vacuma and transposa frequencies was most likely caused by differences in saprotrophic and pathogenic fitness. Possible interactions between fungicide resistance profiles and fitness are discussed.     

Chemiluminescent immunoassay in comparison with the indirect ELISA as reference method for detecting Salmonella antibodies in swine meat juice

The efficiency of chemiluminescent immunoassay (CLIA) in detecting Salmonella antibodies in the meat juice of slaughter swine was compared with the indirect ELISA (BgVV method). Based on the screening test results of 987 meat juice samples obtained from different laboratories in Germany, a good level of agreement between the two systems was obtained with a kappa value of 0.824 at 20% cut-off and 0.798 at 40% cut-off. At 20% and 40% cut-off levels, a sensitivity of 96.2% and 97.3%, respectively, and a specificity of 94.6% and 95.1%, respectively, were demonstrated between CLIA and ELISA. The detecting LPS antigen was tested for specificity and a cross-reaction with two E. coli and Yersinia strains was found when tested with ELISA. This reaction was not observed in CLIA, possibly because of the broader measurement spectrum of this test which allows a more distinctive definition of immunologic reactions. The same explanation can be given for the increased number of meat juice samples which were positively detected only in ELISA but not in CLIA. The positively classified samples in screening were further tested for reciprocal titers in both test systems, and a higher correlation between screening and titration results was obtained for CLIA. Towards the end of the study, a preliminary comparison of CLIA with two available commercial ELISA test kits was conducted and the same tendency was observed, namely, wider detection range of CLIA compared to the other tests. Based on the results of this study, CLIA can be used as a reference method in detecting Salmonella antibodies in the meat juice of slaughter pigs.     

Measurement of serum immunoglobulin concentration in killer whales and sea otters by radial immunodiffusion

Killer whales and sea otters maintained in captivity are the subjects of routine health monitoring programs, and interest in immunologic studies in sea otters has been rising recently in response to potential impacts from infectious disease and environmental pollution on the threatened southern sea otter population. Development of species-specific reagents for immunologic studies in these two marine mammals is currently in its infancy. In this study, killer whale and sea otter immunoglobulin-specific polyclonal antibodies were generated, and used to develop tests for serum Ig concentration in the killer whale (Orcinus orca) and the southern (Enhydra lutris nereis) and northern sea otter (Enhydra lutris lutris). Killer whale serum IgG was purified using caprylic acid/ammonium sulfate precipitation. Sea otter plasma IgG was purified using protein-A-agarose. Polyclonal anti-Ig antisera were produced in rabbits, and specificity confirmed by immunoelectrophoresis. Radial immunodiffusion was used to measure Ig concentration in serum or plasma samples derived from 21 captive killer whales, 18 wild and 4 captive southern sea otters and 15 wild and 4 captive northern sea otters grouped by age. Mean killer whale serum Ig concentration (+/-95% confidence interval) ranged from 15.04 +/- 3.97 g/l for animals aged 0-5 years to 26.65 +/- 9.8 g/l for animals aged >10 years. Mean sea otter serum Ig concentration (+/-95% confidence interval) ranged from 28.39 +/- 11.00 g/l for southern sub-adults to 32.76 +/- 11.58 g/l for southern adults. No significant difference in serum Ig concentration was found between southern and northern sea otters. Serum Ig concentrations in two northern sea otter pups were low compared to those of adult sea otters. The two serum Ig quantitation assays produced were highly specific and reproducible and will be useful additions to the limited number of tests available for immune function in these marine mammal species.     

Serological responses against the pathogenic dimorphic fungus Mucor amphibiorum in populations of platypus (Ornithorhynchus anatinus) with and without ulcerative mycotic dermatitis

Mucor amphibiorum, a dimorphic fungus, causes ulcerative dermatitis and systemic infections in the platypus Ornithorhynchus anatinus in some river systems in Tasmania but apparently not in other regions of Australia. As yet there are no suitable tests for population surveys, nor for detection of internal lesions in live animals. Consequently, immunoglobulins were purified from the serum of platypuses and anti-immunoglobulin antisera were prepared in rabbits in order to develop an enzyme-linked immunosorbent assay (ELISA) for anti-M. amphibiorum antibodies. Antigens from plate-grown cultures resulted in greater signal-to-noise ratios in indirect ELISA than those from broth-grown cultures. Platypuses with clinical ulcerative dermatitis had elevated anti-Mucor antibody levels compared to apparently unaffected individuals. Seroconversion was observed in one animal coincident with the development of cutaneous ulcers. The results suggested that platypuses in affected rivers were exposed to M. amphibiorum at a higher frequency than the occurrence of clinical disease. Some platypuses from New South Wales had elevated antibody levels but these increased significantly with age suggesting exposure to cross-reactive antigens, although exposure to M. amphibiorum cannot be excluded. Further studies are warranted to determine factors that result in progression from infection to disease, the occurrence of the fungus in areas where disease has not been observed and the specificity of antigen used in ELISA.