Direct examination of soil for sporangia of Synchytrium endobioticum using chloroform, calcium chloride and zinc sulphate as extraction reagents

Fields infested with Synchytrium endobioticum can be descheduled when the soil is found free from sporangia of S. endobioticum . For direct examination, EPPO Standard PM 3/59 describes a soil extraction technique based on the use of a sieve shaker with six sieves. We compared recovery of sporangia between this (modified) method and an extraction method employed by the Dutch Plant Protection Service (PPS method). Recovery was determined using an inoculum dilution series: 125, 25, 5, 1, 0.2 or 0.04 sporangia per g soil. Extraction reagents used were chloroform and calcium chloride in the method described by EPPO, calcium chloride and zinc sulphate in the PPS method. At 125 sporangia per g soil, the mean density determined for the modified EPPO method was 228 sporangia per g soil when chloroform was used. Using calcium chloride, recovery percentage was higher for the modified EPPO method than for the PPS method (286, 136%, n.s. P  < 0.05). The advantage of the modified EPPO method was the larger soil volume to be processed; its disadvantages were use of complex equipment and noxious reagents (chloroform). Both extraction methods showed high variation in recovery between samples, making accurate estimation of sporangial densities in soil awkward.     

CABI/EPPO distribution maps of plant pests and plant diseases and their important role in plant quarantine

Never before has the need for accurate distribution data for plant pests been so important. CAB International (CABI) and EPPO are international organizations with a long history and strong involvement in collating and disseminating information on the global distribution of plant pests. Distribution Maps of Plant Pests and Distribution Maps of Plant Diseases, first published in 1951 and 1942, respectively, are respected, referenced sources of such data, expertly compiled and validated and used by plant health organizations around the world. They have been joint CABI/EPPO publications since 1997, and provide an essential complement to expanding knowledge on plant pest distribution. The Distribution Maps continue to be the most authoritative sources of information on the presence and extent of specific plant pests, sourced from the 4.5 million records in CAB Abstracts as a basis and numerous other sources. They also feed directly into CABI's Crop Protection Compendium (CPC) and EPPO's Plant Quarantine Data Retrieval System (PQR) databases. Their history, compilation and value are discussed.     

Investigation of Senescence in Canine Fibroblasts

Introduction and Aims:  Primary and secondary cell cultures of canine dermal fibroblasts were used to test the suitability of several senescence-linked cellular markers to characterize the chronological age of dogs. In further studies these markers will be used to test the hypothesis that senescence-linked changes at the cellular level are representative for the observed inverse relationship of body mass and life expectancy in different dog breeds.
Methods:  In this study standardized skin samples of Beagles were used to analyse the cell morphology and several senescence-linked parameters as Cumulative Population Doubling Level, Highest Population Doubling Level, Culture Doubling Time and Life Span Completed to determine the replicative potential of fibroblasts. The samples were divided by the age of the donors into three groups ('young', 'middle' and 'old').
Results:  There are notable differences between the replicative potential of young and old Beagles at the cellular level. Preliminary results show that in fibroblast cultures obtained from the young group the highest population doubling level is higher and the culture doubling time is shorter then in cultures of the old group. Cultures from the young and middle-aged group cannot be distinguished by the measured parameters. For final results the complete statistical analysis of sampled data has to be carried out.
Conclusions:  Our results demonstrate a relationship between the chronological age of the skin donor and the replicative potential of the dermal fibroblasts. Therefore cellular senescence-linked markers could serve as suitable tools to compare the age of individual dogs of different breeds.     

Computed Tomography of the Lungs of the Dog by a Six-generation CT Scanner, Intravenous Contrast Medium and Different Windows

Computed tomography (CT) is a modern technique of image diagnosis particularly recommended in human medicine to evaluate the existence of pulmonary pathological changes such as neoplasms, metastasis, interstitial infiltrates, etc. In veterinary medicine, however, few anatomical and clinical CT studies in the dog have used apparatus of the latest generation, including injection of intravenous contrast and application of regional specific CT windows with different window width (WW) and window level (WL) to evaluate the lungs, the pulmonary vessels and the bronchial structures. This methodology allows the obtaining of clear CT images with high capacity of tissue discrimination and different shades of attenuation. In this work we have planned a tomographic study of the lungs of the dog by using a six-generation spiral CT scanner (Toshiba Ex Vision), belonging to the private Medical Institute of Radiology 'Irion' of Porto Alegre, Brazil. Four mixed-breed mature dogs (4–6 years, 15–20 kg) were used, two males and two females. The dogs were anaesthetized and kept in a maximum inspiration when obtaining the images. Dogs were placed in a stretcher in a ventral or sternal recumbency. Previously, the contrast urografin® was injected in the cephalic vein. Different CT windows were applied in order to increase the quality of the images: pulmonary window (WW 928; WL -680), high-resolution pulmonary window (WW 1085; WL -750), and soft tissue window (WW 652; WL -34). The use of intravenous contrast, different CT windows and a modern CT apparatus produced excellent images of the pulmonary parenchyma, the pleural cavity, the pulmonary veins, the lobular rami of the pulmonary artery and the lobular bronchi.     

An improved protocol for the production of AFLPTM markers in complex genomes by means of capillary electrophoresis

The amplified fragment‐length polymorphism (AFLP) technology is a recently introduced method to investigate genomes of different complexity, from microbial to higher organisms. It is applied to purposes as diverse as identification of species, strain and varieties, investigation of genetic diversity within and between populations, simple and complex trait mapping, and construction of linkage and physical maps. This technology has been designed on the use of primers labelled with radioactivity and on AFLP fragment separation on sequencing gel. We show that the original EcoRI/TaqI AFLP protocol does not perform appropriately when transferred to fluorescent labelling and capillary electrophoresis (CE), and propose an improved protocol for the production of high‐quality AFLP markers in fish, rodents and artiodactyles by means of the Beckman‐Coulter CEQ2000 automatic DNA sequencer. In addition, we describe the procedure routinely used in our laboratory to obtain binary matrices from AFLP profiles with the aid of Genographer free‐share software (vers. 1.6.0, J.J. Benham, Montana State University), able to elaborate original fragment data and convert them to standard graphical formats for phylogenetic analyses. Comparison with radioactive AFLPs in goats confirmed the reliability of the protocol developed for CE. In fact, 107 fragments generated by two primer combinations and identified by both techniques were attributed the same scoring. Compared with traditional methods, the use of capillary systems and automated analysis increases data throughput and scoring reliability, decreasing the overall experimental error.     

The coding sequence of the ASIP gene is identical in nine wild-type coloured cattle breeds

The aim of this study was to ascertain the role of the Agouti signaling peptide (ASIP) gene coding region in the Agouti locus variation within wild‐type coat colour in cattle. We determined the Extension genotype in 241 individuals from six Spanish and three French brown cattle breeds representative of wild‐type coat variation. Polymerase chain reaction–single‐strand conformation polymorphism (PCR‐SSCP) analysis was carried out in individuals of each Extension genotypes within the same breed in an attempt to identify variants in the three coding exons of the ASIP gene. No SSCP variants were found. Results were confirmed by sequencing the coding exons of the ASIP gene in 20 individuals. Our results suggest that the ASIP coding region does not play a central role in coat colour variation in cattle.     

ECHOCARDIOGRAPHIC AND DOPPLER DIAGNOSIS: FIRST CASE OF ATRIAL SEPTAL ANEURYSM IN A CAT

A 6-year-old female cat developed pleural and abdominal effusion. Cardiac ultrasound and 2D color tissue Doppler imaging revealed restrictive cardiomyopathy with severe biatrial dilatation and hypertension. This cardiomyopathy was associated with atrial septal aneurysm and a patent foramen ovale. The atrial septal aneurysm involved the entire atrial septum. Necropsy and histological examination confirmed all these findings. ASA is a rare malformation and, as in this cat, it generally occurs concomitantly with congenital or acquired heart disease and may be explained by greater pressure in one atrium, leading to controlateral protrusion of the atrial septum.     

MAGNETIC RESONANCE IMAGING FINDINGS IN THE EQUINE DEEP DIGITAL FLEXOR TENDON AND DISTAL SESAMOID BONE IN ADVANCED NAVICULAR DISEASE—AN EX VIVO STUDY

We describe the abnormal magnetic resonance (MR) imaging findings in the deep digital flexor tendon (DDFT) and distal sesamoid bone in horses with radiographic changes compatible with navicular syndrome. Thirteen postmortem specimens were examined using a 1.5-T magnetic field, with spin echo (SE) T1-weighted, turbo SE (TSE) proton density-weighted (with and without fat saturation), and fat saturation TSE T2-weighted sequences. The limbs were then dissected to compare the MR findings with the gross assessment and histologic examination of the DDFT and distal sesamoid bones. Tendonous abnormalities were detected by MR imaging in 12 DDFTs and confirmed at necropsy. Most tendon lesions were located at the level of the distal sesamoid bone and the proximal recess of the podotrochlear bursa. Tendon lesions were classified based on their MR imaging features as core lesions, dorsal lesions, dorsal abrasions, and parasagittal splits. Areas of increased MR signal in the DDFTs were characterized by tendon fiber disturbance and lack of continuity of the collagen fibers, foci of edema, hemorrhages, and formation of lakes containing eosinophilic plasma-like material or amphophilic material of low density. Bone marrow signal alterations in the distal sesamoid bone were seen in all digits. Two main phenomena were responsible for the abnormal signal, respectively, in T1-weighted (decreased signal) and in T2-weighted fat-suppressed images (increased signal): a decrease in the fat marrow content in the trabecular spaces and an increase in the fluid content. Histologic examination revealed foci of bone marrow edema, hemorrhage, necrosis, and fibrosis. Cyst formation and trabecular abnormalities (disorganization, thinning, remodelling) were also observed in areas of abnormal signal intensity. Increased bone density because of trabecular thickening induced a decrease in signal in all sequences.     

In vitro antimicrobial inhibition profiles of Mycoplasma bovis isolates recovered from various regions of the United States from 2002 to 2003.

Antimicrobial therapy continues to be important in reducing losses due to pneumonic forms of Mycoplasma bovis disease in beef and dairy calves. Although M. bovis diseases have been documented as frequent and economically important in the United States, there are no published reports on the antimicrobial activity of approved compounds against US strains. In this study, the authors report on the activity of 9 different antimicrobials against 223 recently recovered isolates of M. bovis. These isolates represent accessions from 5 geographic regions of the United States and were grouped by 4 tissues of origin (milk, respiratory, joint, or ear and eye). A broth microdilution test was used to determine minimum inhibitory concentration (MIC) values by reading redox changes detected in broth with alamarBlue (resazurin) indicator. For each antimicrobial, the median, MIC50, MIC90, mode, and range were calculated, and the values used for comparisons. In the absence of accepted breakpoint values, published MIC cutoff values for animal mycoplasmas as well as Clinical Laboratory Standards Institute interpretive criteria were used as a reference to define in vitro activity. The MIC values from active antimicrobials were found to distribute independently of region of origin of the isolates or of tissue of origin. Enrofloxacin, florfenicol, and spectinomycin were found to be active compounds in vitro. Oxytetracycline and chlortetracycline were active against more than half of the isolates. Very few isolates were inhibited by tilmicosin and none by erythromycin, ampicillin, or ceftiofur. The antimicrobial profiles determined for these US strains were remarkably similar to those reported for European isolates. However, unlike in Europe, there appears to be no diversity of profiles when US isolates are grouped by region or tissue of origin.