Specific contrast ultrasound using sterically stabilized microbubbles for early diagnosis of thromboembolic disease in a rabbit model

Specific contrast ultrasound is widely applied in diagnostic procedures on humans but remains underused in veterinary medicine. The objective of this study was to evaluate the use of microbubble-based contrast for rapid ultrasonographic diagnosis of thrombosis in small animals, using male New Zealand white rabbits (average weight about 3.5 kg) as a model. It was hypothesized that the use of microbubble-based contrast agents will result in a faster and more precise diagnosis in our model of thrombosis. A pro-coagulant environment had been previously established by combining endothelial denudation and external vessel wall damage. Visualization of thrombi was achieved by application of contrast microbubbles [sterically stabilized, phospholipid-based microbubbles filled with sulfur hexafluoride (SF₆) gas] and ultrasonography. As a result, rapid and clear diagnosis of thrombi in aorta abdominalis was achieved within 10 to 30 s (mean: 17.3 s) by applying microbubbles as an ultrasound contrast medium. In the control group, diagnosis was not possible or took 90 to 180 s. Therefore, sterically stabilized microbubbles were found to be a suitable contrast agent for the rapid diagnosis of thrombi in an experimental model in rabbits. This contrast agent could be of practical importance in small animal practice for rapid diagnosis of thrombosis.     

Induction of immune tolerance to caseins and whey proteins by oral intubation in mouse allergy model

This study was designed to evaluate the effect of oral tolerance of caseins (CSN) and whey proteins (WP) in alleviating the allergic response to cow's milk proteins in Swiss albino mice raised on a milk protein–free diet. Oral tolerance was induced by feeding mice with 20 mg of CSN or WP once in a day for 4 days consecutively before immunization with respective protein by intraperitoneal (i.p.) injections (20 μg 200 per μl of PBS) using 2% of alum Al(OH)₃ as adjuvant. Three weeks later, oral tolerance induction was analysed in humoral and cellular compartments of CSN‐ and WP‐fed versus saline‐fed control mice groups by measuring seric and intestinal antibody responses, mRNA abundance in splenic tissue and cytokine secretion patterns. The specific serum immunoglobulin‐E (IgE) levels were significantly suppressed (p < 0.05), while sIgA was enhanced in these groups when compared with their respective saline‐fed mice. Moreover, the mRNA levels of interferon‐γ (IFN‐γ) and interleukin‐4 (IL‐4) in both CSN‐ and WP‐tolerized mice were found to be significantly decreased, while the abundance of interleukin‐10 (IL‐10) and transforming growth factor‐β (TGF‐β) was increased significantly, as compared to respective control groups. Finally, cytokine profiles indicated a reciprocal decrease in IL‐4 and IFN‐γ versus an increase in IL‐10 secretions in supernatants of cultured splenocytes of tolerized mice. Taken together, these results clearly showed that oral administration of cows' milk caseins and whey proteins can induce significant hyposensitization in mice, with the participation of suppressor cytokines.     

Real-time RT-qPCR assay to detect sequences in the Piscine orthoreovirus-1 genome segment S1 associated with heart and skeletal muscle inflammation in Atlantic salmon

During the last decade, Piscine orthoreovirus was identified as the main causative agent of heart and skeletal muscle inflammation (HSMI) in Atlantic Salmon, Norway. A recent study showed that PRV-1 sequences from salmonid collected in North Atlantic Pacific Coast (NAPC) grouped separately from the Norwegian sequences found in Atlantic Salmon diagnosed with HSMI. Currently, the routine assay used to screen for PRV-1 in NAPC water and worldwide cannot differentiate between the two groups of PRV-1. Therefore, this study aimed at developing a real-time polymerase chain reaction (RT-qPCR) assay to target the PRV-1 genome segments specific for variants associated with HSMI. The assay was optimized and tested against 71 tissue samples collected from different regions including Norway, Chile and both coast of Canada and different hosts farmed Atlantic Salmon, wild Coho Salmon and escaped Atlantic Salmon collected in British Columbia, West Coast of Canada. This assay has the potential to be used for screening salmonids and non-salmonids that may carry PRV-1 potentially causing HSMI.     

Relative importance of sorption versus aggregation for organic matter storage in subsoil horizons of two contrasting soils

Soil organic matter (OM) stabilization by the mineral phase can take place through sorption and aggregation. In this study we examined both of these processes, (i) organic carbon (OC) sorption onto clay‐sized particles and (ii) OC occlusion in silt‐size aggregates, with the objective of evaluating their relative importance in OM storage and stabilization in soil. We studied two loamy soil profiles (Haplic Luvisol and Plinthic Cambisol) currently under agricultural use down to a depth of 2 m. Our approach was based on two parallel fractionation methods using different dispersion intensities; these methods isolated a free clay fraction (non‐occluded) and a clay fraction occluded within water‐stable silt‐size aggregates. The two clay fractions were analysed for their C content and ¹⁴C activity. The proportion of sorbed OC was estimated as OC loss after hydrofluoric acid (HF) demineralization. Our results showed an important contribution to SOM stabilization by occlusion of OC into silt‐size aggregates with depth through both soil profiles. In the Haplic Luvisol, OC associated with clay and located in silt‐size aggregates accounted for 34–64% of the total soil OC, whereas in the Plinthic Cambisol this occluded material represented 34–40% of total OC. In the Haplic Luvisol, more OC was located in silt‐size aggregates than was sorbed onto clay‐size minerals, suggesting that silt‐size aggregation plays a dominant role in OC storage in this soil. In the Plinthic Cambisol, the abundance of sorbed OC increased with depth and contributed more to the stored C than that associated with silt‐size aggregates. Radiocarbon dating of both clay fractions (either occluded within silt‐size aggregates or not) suggests, in the case of the Plinthic Cambisol, a preferential stabilization of OC within silt‐size aggregates.     

Comparison of anal sac cytological findings and behaviour in clinically normal dogs and those affected with anal sac disease

No previous study has explored the relationship between cytology and the frequency of behaviours associated with anal sac disease (ASD). The goals of the study were: (i) to compare the cytological findings between anal sac secretions from normal dogs with no history of ASD to those with non-neoplastic ASD; (ii) to determine whether anal sac cytological findings can be used to differentiate between normal dogs and dogs with ASD; (iii) to explore the correlation of anal sac cytology and behaviour between normal dogs and dogs with ASD; and (iv) to describe behaviours typical of ASD as reported by owners. Thirty dogs were selected for this study, based on their behavioural history as detailed in a questionnaire completed by their owners. Of the thirty dogs, ten were considered normal insofar as they had no history of ASD clinical signs. The remaining 20 dogs were characterized as having ASD, with a chronic history of perianal pruritus, but no other pruritus. All dogs had their anal sacs manually expressed, and the discharge was examined microscopically in a blinded manner. A total of 171 oil immersion fields (OIFs) were examined from normal dogs and 333 OIFs from dogs with ASD. The behavioural results for dogs with ASD revealed that scooting recurred with a median frequency of 3 weeks post-anal sac expression. There were no clinically statistically significant cytological differences between normal dogs and those with ASD, thereby leading to the conclusion that cytology is an ineffective tool for diagnosing ASD.     

Development and validation of the Canine Atopic Dermatitis Lesion Index,a scale for the rapid scoring of lesion severity in canine atopic dermatitis

Background – The third iteration of the Canine Atopic Dermatitis Extent and Severity Index (CADESI‐03) is the only tool rigorously validated for canine atopic dermatitis (CAD) lesion scoring. The CADESI‐03 requires 248 evaluations, limiting its widespread use. Hypothesis/Objectives – The goal of the study was to develop and validate a practical method of grading CAD lesions that requires scoring only the frequently affected body regions. Animals – Fifty‐seven privately owned atopic dogs were used in the study. Methods – The Canine Atopic Dermatitis Lesion Index (CADLI) was evaluated in an open, multicentre reliability study. Validity was assessed with expert opinion (content validity) and comparison of CADLI with existing disease severity measures (construct and criterion validity). Reliability was evaluated by analysing repeated observations of each dog. Convenience was assessed in terms of the time required to complete the scale. Results – The CADLI scores correlated with overall assessment scores (r = 0.60, P < 0.001, linear mixed model) and pruritus severity scores (r = 0.53, P < 0.001, linear mixed model), establishing construct validity. The CADLI was strongly correlated with CADESI‐03 (r = 0.84, P < 0.001, linear mixed model), establishing criterion validity. The CADLI values obtained by two observers correlated very strongly (r = 0.91, P < 0.001), as did the repeat values for the same observer (r = 0.98, P < 0.001). The mean time to complete the CADLI was less than that required for CADESI‐03 (1.9 and 12.6 min, respectively), a highly significant difference (P < 0.001). Conclusion and clinical importance – The CADLI was found to be an effective measure of CAD lesion severity, strongly correlating with CADESI‐03. The convenience of CADLI makes it suitable for use in both clinical research and practice.     

Crystal growth inhibitors for the prevention of L-cystine kidney stones through molecular design

Crystallization of L-cystine is a critical step in the pathogenesis of cystine kidney stones. Treatments for this disease are somewhat effective but often lead to adverse side effects. Real-time in situ atomic force microscopy (AFM) reveals that L-cystine dimethylester (L-CDME) and L-cystine methylester (L-CME) dramatically reduce the growth velocity of the six symmetry-equivalent {100} steps because of specific binding at the crystal surface, which frustrates the attachment of L-cystine molecules. L-CDME and L-CME produce l-cystine crystals with different habits that reveal distinct binding modes at the crystal surfaces. The AFM observations are mirrored by reduced crystal yield and crystal size in the presence of L-CDME and L-CME, collectively suggesting a new pathway to the prevention of L-cystine stones by rational design of crystal growth inhibitors.