Design and Validation of a Simulator for Feline Cephalic Vein Cannulation-A Pilot Study.

In recent years there has been an increased use of alternative methods for teaching veterinary clinical skills, since ethical considerations preclude the use of live animals for demonstration or practice of many procedures. Skills training on cats (i.e., feline venipuncture) is a particularly challenging area. This study aimed to develop a simulator for cephalic venipuncture in cats and to validate this simulator using questionnaires answered by undergraduate students and experienced veterinarians. The simulator was developed to provide an experience that was close to reality, including an artificial blood system that flows through the catheter when venipuncture is correctly performed, while at the same time using simple methodology and accessible materials so that it could be reproduced in other universities. The experienced vets agreed (44.4%) or strongly agreed (55.6%) that the simulator was good for venipuncture training, and the most useful feature was the experience of catheter manipulation and fixation on the cat's limb. All the students agreed that the practical class with the simulator was important for learning this skill. Both groups (students and experienced veterinarians) unanimously agreed that it is important to train using a simulator before trying the procedure on a live cat. This simulator offers undergraduate students an alternative way to learn and practice venipuncture in cats helping to reduce the use of live animals in practical classes.

Secretory production in Escherichia coli of a GH46 chitosanase from Chromobacterium violaceum, suitable to generate antifungal chitooligosaccharides.

A chitosanase (CvCsn46) from Chromobacterium violaceum ATCC 12472 was produced in Escherichia coli, purified, and partially characterized. When subjected to denaturing polyacrylamide gel electrophoresis, the enzyme migrated as two protein bands (38 and 36 kDa apparent molecular masses), which were both identified as CvCsn46 by mass spectrometry. The enzyme hydrolyzed colloidal chitosan, with optimum catalytic activity at 50 °C, and two optimum pH values (at pH 6.0 and pH 11.0). The chitosanolytic activity of CvCsn46 was enhanced by some ions (Ca2+, Co2+, Cu2+, Sr2+, Mn2+) and DTT, whereas Fe2+, SDS and ß-mercaptoethanol completely inhibited its activity. CvCsn46 showed a non-Michaelis-Menten kinetics, characterized by a sigmoidal velocity curve (R2 = 0.9927) and a Hill coefficient of 3.95. ESI-MS analysis revealed that the hydrolytic action of CvCsn46 on colloidal chitosan generated a mixture of low molecular mass chitooligosaccharides, containing from 2 to 7 hexose residues, as well as D-glucosamine. The chitosan oligomers generated by CvCsn46 inhibited in vitro the mycelial growth of Lasiodiplodia theobromae, significantly reducing mycelium extension and inducing hyphal morphological alterations, as observed by scanning electron microscopy. CvCsn46 was characterized as a versatile biocatalyst that produces well-defined chitooligosaccharides, which have potential to control fungi that cause important crop diseases.

Structural and functional features of a class VI chitinase from cashew (Anacardium occidentale L.) with antifungal properties.

A partial cDNA sequence from Anacardium occidentale CCP 76 was obtained, encoding a GH19 chitinase (AoChi) belonging to class VI. AoChi exhibits distinct structural features in relation to previously characterized plant GH19 chitinases from classes I, II, IV and VII. For example, a conserved Glu residue at the catalytic center of typical GH19 chitinases, which acts as the proton donor during catalysis, is replaced by a Lys residue in AoChi. To verify if AoChi is a genuine chitinase or is a chitinase-like protein that has lost its ability to degrade chitin and inhibit the growth of fungal pathogens, the recombinant protein was expressed in Pichia pastoris, purified and biochemically characterized. Purified AoChi (45 kDa apparent molecular mass) was able to degrade colloidal chitin, with optimum activity at pH 6.0 and at temperatures from 30 °C to 50 °C. AoChi activity was completely lost when the protein was heated at 70 °C for 1 h or incubated at pH values of 2.0 or 10.0. Several cation ions (Al3+, Cd2+, Ca2+, Pb2+, Cu2+, Fe3+, Mn2+, Rb+, Zn2+ and Hg2+), chelating (EDTA) and reducing agents (DTT, ß-mercaptoethanol) and the denaturant SDS, drastically reduced AoChi enzymatic activity. AoChi chitinase activity fitted the classical Michaelis-Menten kinetics, although turnover number and catalytic efficiency were much lower in comparison to typical GH19 plant chitinases. Moreover, AoChi inhibited in vitro the mycelial growth of Lasiodiplodia theobromae, causing several alterations in hyphae morphology. Molecular docking of a chito-oligosaccharide in the substrate-binding cleft of AoChi revealed that the Lys residue (theoretical pKa = 6.01) that replaces the catalytic Glu could act as the proton donor during catalysis.

Humane Education in Brazil: Organisation, Challenges and Opportunities.

Humane education and the debate on alternatives to harmful animal use for training is a relatively recent issue in Brazil. While animal use in secondary education has been illegal since the late 1970s, animal use in higher science education is widespread. However, alternatives to animal experiments in research and testing have recently received attention from the Government, especially after the first legislation on animal experiments was passed, in 2008. This article proposes that higher science education should be based on a critical and humane approach. It outlines the recent establishment of the Brazilian Network for Humane Education (RedEH), as a result of the project, Mapping Animal Use for Undergraduate Education in Brazil, which was recognised by the 2014 Lush Prize. The network aims to create a platform to promote change in science education in Brazil, starting by quantitatively and qualitatively understanding animal use, developing new approaches adapted to the current needs in Brazil and Latin America, and communicating these initiatives nationally. This paper explores the trajectory of alternatives and replacement methods to harmful animal use in training and education, as well as the status of humane education in Brazil, from the point of view of educators and researchers engaged with the network.

Canine Prostate Palpation Simulator as a Teaching Tool in Veterinary Education.

Adult dogs, especially elderly ones, are commonly affected by prostate diseases. Performing rectal palpation during physical examination in dogs is important in small animal clinical diagnosis. Prostate palpation training allows students to learn how to correctly introduce the finger into the rectum and identify the location, size, symmetry, and consistency of the prostate. Alternative methods are needed to teach this technique without using live dogs. Thus, our aim was to develop a canine prostate palpation simulator to provide students with the opportunity to learn the prostate palpation technique in dogs and to assess their opinion of this simulator as a teaching tool. The inner part of the canine mannequin contains a rotation system with three types of prostates that can be exchanged during the exam. Of the 64 participating students, 81% had never used alternative methods and 92.2% had never performed any prostatic palpation. According to the students' opinions, performing a clinical examination on a simulator allowed them to be prepared and familiarized with the palpation technique. They felt satisfied learning a practical method in a harmless way. Both the 3R (replacement, reduction, and refinement) and dog welfare principles were present in most of the students' concerns. We conclude that the simulator can help students to develop clinical skills for prostate palpation in dogs.

Clinicopathologic and electrocardiographic features of Akita dogs with high and low erythrocyte potassium phenotypes.

BACKGROUND: Canine RBCs are expected to have high sodium and low potassium (LK) concentrations. However, some dogs have a low sodium and high potassium (HK) phenotype due to a Na-K pump in the mature RBC membrane. Awareness of this particularity avoids misinterpretation of hyperkalemia in affected dogs. OBJECTIVE: The purpose of this study was to compare the CBCs, serum biochemistry profiles, urinalysis data, and electrocardiography data in HK and LK Akita dogs. METHODS: In this study, the intra-RBC potassium (KRBC) was measured in 48 healthy adult Akita dogs. The HK group included dogs with KRBC concentration at least 5-fold that of plasma, while the LK dogs served as controls. RESULTS: The HK phenotype was determined in 10 dogs (21%). With the exception of higher plasma potassium concentrations in the HK group compared with LK controls (6.6 mmol/L vs 4.4 mmol/L, P < .001), there were no other differences in serum biochemistry variables. In hematology, mean HGB concentration (13.5 vs 15.9, P < .001), PCV (42.9 vs 46.7, P = .009), RBC (6.1 vs 7.4, P < .001), and MCHC (31.3 vs 34.0, P < .001) were significantly lower in the HK group, while MCV (70.0 vs 63.4, P < .001) was higher compared with the LK controls. There were no significant differences in urinalysis or electrocardiography data between groups. CONCLUSIONS: HK Akita dogs showed significant differences in several hematologic variables, as well as higher plasma potassium concentration. Therefore, the HK phenotype should be considered in the interpretation of these variables in this breed.

Laparoscopic versus open splenectomy in dogs

In the last few years, the use of laparoscopy in veterinary medicine has expanded and consequently so was the need for studies that establish the advantages, disadvantages and possible complications of each procedure. The purpose of the current study was to describe a laparoscopic splenectomy technique and the alterations due to this access, and compare it to the open procedure in dogs. A total of 15 healthy female mongrel dogs were used, with mean weight of 17.4±2.5kg. The animals were distributed into three groups: Group IA of open splenectomy (laparotomy) using double ligation of the vessels of the splenic hilum with poliglicolic acid, Group IB of open splenectomy (laparotomy) with bipolar electrocoagulation of the splenic hilum, and Group II of laparoscopic access with bipolar electrocoagulation of the splenic hilum. Operative time, blood loss, size of incisions, complications during and after surgery were evaluated. Other parameters included pain scores, white blood cell (WBC) counts and postoperative serum concentrations of alanine aminotransferase (ALT), alkaline phosphatase (ALP), creatine kinase (CK), C-reactive protein (CRP), glucose and cortisol. No differences were found in the evaluation of parameters between both open splenectomy techniques employed. Laparoscopic access presented significant differences (p<0,05) when compared with open surgery: Longer operative time, smaller abdominal access, decrease in blood loss, lower concentrations of CRP, higher levels of CK and ALP, and lower scores in the pain scale. Laparoscopic surgery showed fewer complications of the surgical wound. No significant differences were observed between groups in the postoperative temperature, WBC, ALT, cortisol and glucose concentrations. In conclusion, the laparoscopic technique is useful for splenectomy in dogs, being advantageous in terms of blood loss, surgical stress and surgical wounds. However, it expends more operative time and causes transitory...

Nos últimos anos, a utilização da laparoscopia em Medicina Veterinária vem expandindo e, conseqüentemente, a necessidade de pesquisas que determinem as vantagens, desvantagens e possíveis complicações de cada procedimento. Este estudo teve como objetivo descrever uma técnica de esplenectomia laparoscópica, assim como as alterações decorrentes deste acesso, e compará-la ao procedimento convencional em cães. Foram utilizadas 15 cadelas hígidas, sem raça definida, com peso médio de 17,4 ±2,5kg. Os animais foram distribuídos em três grupos: Grupo IA de acesso convencional (por laparotomia) utilizando ligadura com ácido poliglicólico no selamento vascular do hilo esplênico, Grupo IB de acesso convencional (por laparotomia) com eletrocoagulador bipolar do hilo esplênico, e Grupo II de acesso laparoscópico com eletrocoagulador bipolar para selamento vascular dos ramos esplênicos. Estes grupos foram avaliados em relação ao tempo cirúrgico, à perda de sangue, ao tamanho das incisões e às complicações durante e após a cirurgia. Também foram comparadas as avaliações da escala de dor e as alterações no leucograma e nas concentrações séricas da alanina aminotransferase (ALT), da fosfatase alcalina (FA), da creatina quinase (CK), da proteína C-reativa (CRP), da glicose e do cortisol no pós-operatório. Os acessos convencionais não diferiram entre si nos parâmetros avaliados. O acesso laparoscópico apresentou diferenças significativas (p<0,05) quando comparado ao convencional: maior tempo cirúrgico, menor acesso abdominal, diminuição na perda de sangue, menores concentrações de CRP, maiores níveis de CK e FA, além de pontuação menor na escala de dor. A cirurgia laparoscópica apresentou menor número de complicações das feridas cirúrgicas. A ALT, o cortisol, a glicemia, o leucograma e a temperatura retal pós-operatórias não diferiram significativamente entre os acessos convencional e laparoscópico. Conclui-se que a cirurgia laparoscópica é viável para...

Design, optimization, and application of a conventional PCR assay with an internal control for detection of 'Candidatus Mycoplasma turicensis' 16S rDNA in domestic cats from Brazil.

BACKGROUND: 'Candidatus Mycoplasma turicensis' (CMtc) is a hemotrophic bacterial species that can, alone or in combination, induce anemia in cats. The diagnostic test of choice for hemoplasma infections is PCR. Conventional PCR assays have been developed for the detection of Mycoplasma haemofelis (Mhf) and 'Candidatus M. haemominutum' (CMhm) but not for CMtc. Although real-time PCR assays have been reported for all of the feline hemoplasmas, the expense of necessary instrumentation precludes its use in Brazil and many other countries. OBJECTIVES: The goals of this study were to develop and optimize a conventional PCR assay to diagnose CMtc using an internal control to detect false-negative results, and to evaluate the occurrence of CMtc infection in domestic cats from Brazil. METHODS: Species-specific primers were designed and a PCR assay was developed for the detection of CMtc 16S rDNA in cat blood. Sensitivity was determined by serial 10-fold dilutions of plasmid and DNA extracted from blood from an experimentally infected cat. EDTA blood samples from 373 cats were collected. DNA was extracted using a silica-based protocol and tested using the PCR assay. RESULTS: Primer concentration, annealing temperature, and MgCl(2) concentration were optimized in the presence and absence of the internal control. Two samples negative for the internal control were excluded. Of the remaining 371 samples (117 healthy and 254 unhealthy cats), 17 (4.6%) were positive for CMtc. CONCLUSION: These results demonstrate the utility of an optimized PCR assay to detect CMtc in feline blood samples. We also report for the first time the prevalence of CMtc infection in domestic cats in Brazil.

Association of canine obesity with reduced serum levels of C-reactive protein.

The prevalence of obesity is increasing in dogs as well as in humans. C-reactive protein (CRP) is an important tool for the detection of inflammation and/or early tissue damage and is linked to obesity in humans. The objective of the present study was to determine if serum CRP levels are altered in obese dogs. Fifteen lean (control group) and 16 overweight (obese group) dogs were examined. Blood samples were collected under fasted conditions for serum determination of CRP, glucose, insulin, cholesterol, triglyceride, and fructosamine. Results indicated that obese dogs were insulin resistant because serum insulin and insulin/glucose ratios were higher than in lean dogs (P < or = 0.05). Serum CRP concentrations were lower in obese dogs than in controls (P < or = 0.001). C-reactive protein was negatively correlated with insulin/glucose ratio (R = -0.42) and cholesterol (R = -0.39; P < or = 0.05). Furthermore, levels of cholesterol, triglycerides, and fructosamine were increased in the obese group compared with the control group. Based on these results, it can be postulated that CRP production is inhibited by obesity and insulin resistance in dogs.