Differential diagnosis of dog hookworms based on PCR-RFLP from the ITS region of their rDNA

Species of Ancylostoma infecting dogs and sometimes humans are sympatric in many parts of the world. The establishment of a specific molecular diagnostic tool is important, not only to refine information for epidemiological studies, but also to evaluate the efficacy of vaccine programmes and assist in the development of specific drug treatments. The ITS region from 20 specimens of A. braziliense, collected from three separate geographical areas of Brazil, and from 10 specimens of A. caninum, collected from the same area in Brazil were sequenced and analyzed. Alignment of sequences showed that this gene is highly conserved. The intraspecific polymorphism for both species was less then 1%, whereas the interspecific polymorphism was 6.2, 7.3 and 9.4% between A. ceylanicum and A. braziliense; A. caninum and A. ceylanicum and A. ceylanicum and A. braziliense, respectively. Among the three species it was 12.3%. This revealed the ITS region as highly conserved and consequently a good molecular marker for diagnostic studies. In this work, four restriction enzymes were used in a PCR-RFLP using the ITS region of rDNA, to establish a differential diagnosis which discriminates between three Ancylostoma species, A. braziliense, A. caninum and A. ceylanicum. The best pattern was given by the HinfI enzyme, which produced different fragment sizes for each of the three species. Furthermore, the diagnostic tool differentiates DNA extracted directly from faeces of Ancylostoma-infected dogs.     

Clinical observations of cattle and buffalos with experimentally induced chronic copper poisoning

The susceptibility of cattle and buffalos to chronic copper poisoning (CCP) was compared by using cattle (n = 10) and buffalo (n = 10) steers distributed into two copper supplemented (n = 6) and two control (n = 4) groups. Supplemented animals received 2 mg copper (Cu)/kg body weight daily for one week, with an additional 2 mg weekly until the end of the experiment (day 105). Three liver biopsies (day 0, 45, and 105) were obtained for mineral analyses; clinical examinations and blood samples were obtained every 15 days. Three supplemented cattle and two buffalos with typical manifestations of CCP died. There were no differences in the frequency of mortality between cattle and buffalos; hepatic copper concentration was higher in cattle than buffalos. These findings suggest that buffalos and cattle might be equally susceptible to CCP. However, buffalos accumulate less liver copper than cattle and have a lower threshold of hepatic Cu accumulation, which leads to clinical manifestation of CCP.     

Immunogenicity in dogs of three recombinant antigens (TSA, LeIF and LmSTI1) potential vaccine candidates for canine visceral leishmaniasis

Control of canine visceral leishmaniasis (VL) remains a difficult and serious problem mostly because there is no reliable and effective vaccine available to prevent this disease. A mixture of three recombinant leishmanial antigens (TSA, LeIF and LmSTI1) encoded by three genes highly conserved in the Leishmania genus have been shown to induce excellent protection against infection in both murine and simian models of cutaneous leishmaniasis. A human clinical trial with these antigens is currently underway. Because of the high degree of conservation, these antigens might be useful vaccine candidates for VL as well. In the present study, using the dog model of the visceral disease, we evaluated the immunogenicity of these three antigens formulated with two different adjuvants, MPL-SE and AdjuPrime. The results were compared with a whole parasite vaccine formulated with BCG as the adjuvant. In order to investigate if sensitization with the recombinant antigens would result in recognition of the corresponding native parasite antigens upon infection, the animals were exposed for four weeks after the termination of the immunization protocol with the recombinant antigens to a low number of L. chagasi promastigotes, an etiological agent of VL. Immune response was evaluated by quantitative ELISA in the animal sera before and after exposure to the viable parasites. Both antigen specific IgG1 and IgG2 antibody levels were measured. Immunization of dogs with the recombinant antigens formulated in either MPL-SE or AdjuPrime resulted in high antibody levels particularly to LmSTI1. In addition, this immunization although to low levels, resulted in the development of antibody response to the whole parasite lysate. Importantly, experimental exposure with low numbers of culture forms of L. chagasi promastigotes caused a clear boost in the immune response to both the recombinant antigens and the corresponding native molecules. The boost response was predominantly of the IgG2 isotype in animals primed with the recombinant antigens plus MPL-SE. In contrast, animals primed with the recombinant antigens formulated in AdjuPrime as well as animals vaccinated with crude antigen preparation responded with mixed IgG1/IgG2 isotypes. These results point to the possible use of this antigen cocktail formulated with the adjuvant MPL-SE in efficacy field trials against canine VL.     

Activity of Ricinus communis (Euphorbiaceae) and ricinine against the leaf-cutting ant Atta sexdens rubropilosa (Hymenoptera: Formicidae) and the symbiotic fungus Leucoagaricus gongylophorus

The focus of this study was the identification of compounds from plant extracts for use in crop protection. This paper reports on the toxic activity of fractions of leaf extracts of Ricinus communis L (Euphorbiaceae) and isolated active compounds in the leaf-cutting ant Atta sexdens rubropilosa Forel and its symbiotic fungus Leucoagaricus gongylophorus (Singer) M?ller. The main compounds responsible for activity against the fungus and ant in leaf extracts of R communis were found to be fatty acids for the former and ricinine for the ants.     

A biomechanical comparison of headless tapered variable pitch and AO cortical bone screws for fixation of a simulated slab fracture in equine third carpal bones

OBJECTIVE: To compare the mechanical shear strengths and stiffnesses obtained from in vitro testing of a simulated complete third carpal bone (C3) frontal plane radial facet slab fracture (osteotomy) stabilized with either a 4/5 Acutrak (AT) compression screw or a 4.5-mm AO cortical bone (AO) screw inserted in lag fashion. Drilling, tapping, and screw insertion torques, forces, and times also were compared between AT and AO implants. STUDY DESIGN: In vitro biomechanical assessment of site preparation, screw insertion, and shear failure test variables of bone screw stabilized simulated C3 slab fracture in paired cadaveric equine carpi. SAMPLE POPULATION: Eight pairs of cadaveric equine C3 without orthopedic abnormalities. METHODS: Standardized simulated C3 slab fractures were repaired with either AO or AT screws (AO/C3 and AT/C3 groups, respectively). Drilling, tapping, and screw insertion torques, forces, and times were measured with a materials testing machine for each screw type. Repaired specimens were tested in axially oriented shear until failure. Paired Students t-tests were used to assess differences between site preparation, screw insertion, and shear testing variables. Significance was set at P <.05. RESULTS: There were no significant differences in bone fragment measurements of the standardized simulated C3 slab fractures created for AO or AT screws. There were no significant differences for mean and maximum drilling torques; however, the tapered AT drill had greater maximum drilling force compared with the 3.2-mm and 4.5-mm AO drill bits. Mean insertion torque and force measured from the self-tapping AT screw were not significantly different compared with the 4.5-mm AO tap. There were no significant differences in maximum screw torque among constructs. Total procedure time was significantly longer for the AT group (5.8 +/- 1.6 minutes) compared with the AO group (2.9 +/- 1.1 minutes; P =.001). AT stabilized specimens had significantly greater mean +/- SD initial shear stiffness (3.64 +/- 1.08 kN/mm) than AO specimens (1.64 +/- 0.73 kN/mm; P =.005). All other shear mechanical testing variables were not statistically different among screw types. CONCLUSION: The 4/5 Acutrak insertion technique was accurate and safe, and the AT screw effectively stabilized simulated equine C3 frontal plane slab fractures. When tested in shear, this screw type was mechanically comparable to the 4.5-mm AO screw; however, AT constructs had greater initial shear stiffness. Initial shear stiffness was likely an indirect measure of interfragmentary compression, and thus may indicate that the AT screw provides a more rigid fixation for frontal plane C3 slab fractures in horses. CLINICAL RELEVANCE: Considering the comparable mechanical behavior, greater initial shear stiffness for AT screw stabilized C3 slab fracture fragments, the ability to accurately insert the screw with the aid of a guide pin, and the potential for less persistent soft tissue irritation with the headless screw design, the 4/5 tapered AT screw is an attractive alternative for repair of C3 slab fractures in horses.     

Dilated cardiomyopathy in juvenile Portuguese Water Dogs

Dilated cardiomyopathy recently has been recognized in juvenile Portuguese Water Dogs. The purpose of this study was to evaluate unaffected and affected puppies by physical examination, electrocardiogram (ECG), echocardiogram, specific biochemical assays, and ultrastructure to document disease progression and to develop a method of early detection. Results of segregation analysis were consistent with autosomal recessive inheritance. Of 124 puppies evaluated clinically and echocardiographically, 10 were affected. No significant differences were found between unaffected and affected puppies for blood and myocardial carnitine or taurine concentrations, serum chemical variables, results of ophthalmological examinations, ECGs, or measurement of urine metabolites. Ultrastructural examination of myocardium from affected dogs revealed myofibrillar atrophy and small regions of myofibrillar degeneration, most prominently at the region of the intercalated discs. Only echocardiography allowed detection of affected puppies before clinical signs became evident. Echocardiography revealed a significant difference in the shortening fraction, E point to septal separation, and the end systolic and diastolic left ventricular internal diameters. Affected puppies were detected 1-4 weeks before the development of acute congestive heart failure.     

Comparison of two indirect techniques for local delivery of a high dose of an antimicrobial in the distal portion of forelimbs of horses

OBJECTIVE: To compare isolated limb retrograde venous injection (ILRVI) and isolated limb infusion (ILI) for delivery of amikacin to the synovial fluid of the distal interphalangeal and metacarpophalangeal joints and to evaluate the efficacy of use of an Esmarch tourniquet in standing horses. ANIMALS: 6 healthy adult horses. PROCEDURES: Horses were randomly assigned in a crossover design. In ILRVI, the injection consisted of 1 g of amikacin diluted to a total volume of 60 mL administered during a 3-minute period. In ILI, the infusion consisted of 1 g of amikacin diluted to 40 mL administered during a 3-minute period followed by administration of boluses of diluent (82 mL total) to maintain vascular pressure. During ILI, the infusate and blood were circulated from the venous to the arterial circulation in 5-mL aliquots. Synovial fluid and serum samples were obtained to determine maximum amikacin concentrations and tourniquet leakage, respectively. RESULTS: Both techniques yielded synovial concentrations of amikacin > 10 times the minimum inhibitory concentration (MIC) for 90% of isolates (80 microg/mL) and > 10 times the MIC breakpoint (160 microg/mL) of amikacin-susceptible bacteria reported to cause septic arthritis in horses. These values were attained for both joints for both techniques. Esmarch tourniquets prevented detectable loss of amikacin to the systemic circulation for both techniques. CONCLUSIONS AND CLINICAL RELEVANCE: Both techniques reliably achieved synovial fluid concentrations of amikacin consistent with concentration-dependent killing for bacteria commonly encountered in horses with septic arthritis. Esmarch tourniquets were effective for both delivery techniques in standing horses.     

Effect of larval stocking density on growth,survival and whole body cortisol of the Mediterranean mussel Mytilus galloprovincialis (Lamarck, 1819) larvae reared under laboratory conditions

Interest of mussel hatchery development has increased during the last years as natural seed collection does not always satisfy the demand of producers. Success on larval bivalve culture depends on several factors such as diet, temperature or stocking density. In this study, the effect of larval density in mussel culture success has been assessed testing three larval densities: high (50 larvae mL^?1), medium (25 larvae mL^?1) and low (5 larvae mL^?1). Results showed higher growth and survival at low density but the amount of postlarvae was low. The highest number of settled larvae was observed at high density but mortality was also high and a delay on the larval development was detected producing a prolongation on the culture length. However, medium density resulted in a good survival and growth and also in a considerable amount of postlarvae, as this density could be adequate for massive mussel seed production. Cortisol content on the larvae and postlarvae was also analysed and significant differences were obtained among groups probably due to the stress associated to settlement and metamorphosis.