Molecular characterization of a lipid-modified virulence-associated protein of Rhodococcus equi and its potential in protective immunity.

Virulent strains of Rhodococcus equi produce plasmid-mediated 15- and 17-kDa proteins, which are thermoregulated and apparently surface-expressed. We demonstrated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) that R. equi produce three antigenically-related virulence-associated proteins, a diffuse 18-22-kDa, a 17.5-kDa and a 15-kDa protein. Phase partitioning of whole cells of R. equi strain 103 with Triton X-114 (TX-114) and labelling with [3H]-labelled palmitic acid showed that the two higher molecular weight proteins are hydrophobic and lipid modified. The 15-kDa protein did not partition into TX-114 and was not lipid modified. Cloning and expression of a fragment of the R. equi virulence plasmid in Escherichia coli showed that the three proteins were expressed from a single gene. Sequence analysis of this gene (designated vapA) revealed a 570-bp open reading frame encoding a polypeptide of 189 amino acids with a calculated molecular mass of 19,175 Da. The mature, nonlipid modified protein had a calculated mass of 16,246 Da. The 17.5- and 18-22-kDa forms of the protein are therefore due to lipid modification. No significant sequence homology of the vapA gene with other reported nucleotide sequences were found. Opsonization of virulent R. equi with an IgG1 mouse monoclonal antibody (MAb103) to the VapA protein significantly enhanced uptake in the murine macrophage cell line IC-21. Intraperitoneal injection of mice with Mab103 enhanced initial clearance from the liver of mice challenged intravenously with R. equi. Immunization of mice with the lipid-modified VapA purified by SDS-PAGE fractionation or with acetone precipitated VapA protein following TX-114 extraction resulted in significantly enhanced clearance from the liver and spleen following intravenous challenge. The VapA protein of R. equi appears therefore to be a protective immunogen.     

Hereditary congenital heart defects in dogs

Congenital heart defects is probably the most common class of malformations found in dogs, occurring with a frequency approaching 1 per cent in animals presented to veterinary clinics. The frequency is significantly higher among purebred dogs than in dogs of mixed breeding and specific anatomic malformations occur with highest frequency in certain breeds. Genetic studies of patent ductus arteriosus, pulmonic stenosis, subaortic stenosis, ventricular septal defect, tetralogy of Fallot and persistent aortic arch have confirmed that these are specific heritable defects, the genes for which are concentrated in a number of different breeds. Each of these defects is inherited in a complex manner consistent with a polygenic basis. This paper will describe evidence supporting the view that the common forms of congenital heart disease in the dog are polygenic threshold traits. The general criteria for recognition of polygenic traits and methods for their control will be discussed.     

Inherited phosphofructokinase deficiency in an American cocker spaniel.

A 3-year-old female American Cocker Spaniel with a chronic hemolytic disorder and hemolytic crises was found to have M-type phosphofructokinase deficiency. This inherited erythroenzymopathy and myopathy is commonly diagnosed in English Springer Spaniels, but the family study of this Cocker Spaniel, although supporting an autosomal recessive mode of inheritance, did not reveal any English Springer Spaniel ancestors. Molecular genetic studies did, however, identify the same mutation in this dog as we previously reported in the English Springer Spaniel breed, suggesting that this mutation originated prior to the separation of these 2 breeds.     

Irradiation‐induced off‐odour in chicken and its possible control

1. Volatiles isolated from irradiated raw chicken were analysed by gas chromatography (GC) in conjunction with olfactory assessment of the effluent carrier gas to locate compounds with strong smells.

2. Sixteen odours of differing intensities were registered, some, but not others, coinciding with recognisable GC peaks. Identifications were made on the basis of retention data, mass spectrometric information and odour quality agreement.

3. Dimethyltrisulphide was found to be the most potent and obnoxious compound (foul gas, sulphurous), followed by cis‐3‐ and trans‐6‐nonenals (soapy), oct‐1‐en‐3‐one (mushroom) and bis(methylthio‐)methane (foul). With the exception of oct‐1‐en‐3‐one, none of these compounds has been reported before in irradiated raw chicken.

4. α‐Tocopherol and ascorbic acid induce stability in tissues in vivo and post mortem. Chickens were reared on diets supplemented with high concentrations (800 mg/kg food) of each of these vitamins. Yields of irradiation volatiles from the tissues of these birds were very much reduced, compared to yields from similar tissues from birds fed unsupplemented diets.

5. Concomitantly with the reduced yield of volatiles, less odour was associated with the samples when analysed by GC‐olfactory analysis.

6. The use of enhanced concentrations of the two vitamins in combination in the diet of poultry may provide a means of controlling development of off‐odour in irradiated raw chicken, thus improving acceptability to the consumer.     

Prothrombin time and activated partial thromboplastin time using a point‐of‐care analyser (Abaxis VSpro®) in Bennett's wallabies (Macropus rufogriseus)

There are few reports of coagulation times in marsupial species. Blood samples collected from 14 Bennett's wallabies (Macropus rufogriseus) under anaesthesia during routine health assessments were analysed for prothrombin time (PT) and activated partial thromboplastin time (aPTT) using a point‐of‐care analyser (POC) (Abaxis VSPro®). The wallabies had an aPTT mean of 78.09 s and median of 78.1 s. The PT for all wallabies was greater than 35 s, exceeding the longest time measured on the POC. Although PT was significantly longer, aPTT was similar to the manufacturer's domestic canine reference range.     

Laboratory cultivation of prochloron, a tryptophan auxotroph

Laboratory cultures have been established of the didemnid symbiont Prochloron, a unique prokaryotic alga that synthesizes chlorophylls a and b but no phycobilin pigments. Cell division in Prochloron cultures occurs under acidic conditions (pH 5.5) in the presence of tryptophan. The alga is a naturally occurring tryptophan auxotroph that survives in nature by close association with the host, Diplosoma similis. The metabolic dysfunction that renders Prochloron auxotrophic may involve only the initial step of the tryptophan biosynthetic pathway.     

Osteochondrosis dissecans in a juvenile roan antelope (Hippotragus equinus).

A 4-mo-old, juvenile roan antelope (Hippotragus equinus) presented for an acute and progressive lameness of the left hind limb, which was non-responsive to anti-inflammatory medication. The antelope was anesthetized for examination, radiography, and lab work. The left stifle joint had limited range of motion and was markedly swollen on physical examination. Radiography revealed thin cortices of the left distal femur and proximal tibia, as well as multiple lytic areas in the condyles. Euthanasia was performed due to the severity of the lesions and the animal's overall physical condition. Postmortem findings included a parasitic infection of the gastrointestinal tract. Cartilaginous erosions, associated with fibrotic connective tissue and viscous joint fluid, were identified in all appendicular joints. Perimortem lab work demonstrated anemia and hypoproteinemia. Joint fluid was inflammatory but non-infectious, and aerobic culture of the joint was negative for pathogens. Histologic examination identified focal, transmural necrosis of the articular cartilage with clefting. Osteochondrosis dissecans was diagnosed, which initiated an evaluation of the nutritional status and management of the herd. This is the first documented case of osteochondrosis dissecans in a roan antelope.