Serum Protein Concentration and Serum Protein Fractions in Bottlenose Dolphins (Tursiops truncatus) under Human Care Using Agarose Gel Electrophoresis.

Serum protein electrophoresis (SPE) is the most used and reliable method to determine the percentage of serum protein subfractions. The interpretation of the kinetics of total proteins and albumin and globulin fractions is receiving increased attention in wild animals, as well as in domestic animals, due to the possibility of identifying typical pathologic patterns. However, the interpretation of these data had to be performed in light of an appropriate method-and species- specific reference intervals (RIs). In marine mammals, as well as other non-domestic species, specific attention should also be given to the different environment (free ranging vs. human managed) and the associated different exposure to environmental stimuli. The aim of this report was to establish RIs for the serum protein fractions evaluated using agarose gel electrophoresis (AGE) in bottlenose dolphins under human care. Peripheral blood samples were collected from 40 bottlenose dolphins during standard veterinary procedures to evaluate their health status. Total protein concentration was determined using the biuret method while AGE was performed using an automated system. A pooled dolphin's serum sample was used to determine the intra-assay and inter-assay imprecision of AGE. The RIs were calculated using an Excel spreadsheet with the Reference Value Advisor set of macroinstructions. The intra and inter-assay imprecisions were 1.2% and 2.5%, respectively, for albumin; 2.9% and 5.7%, respectively, for α-globulins; 3.8% and 4.0%, respectively, for ß-globulins; and 3.4% and 4.8%, respectively, for γ-globulins. The total protein, albumin, α-globulin, ß-globulin, and γ-globulin concentrations were 65.5 ± 5.4 g/L, 45.5 ± 4.9 g/L, 8.0 ± 1.0 g/L, 5.0 ± 2.0 g/L, and 7.0 ± 2.0 g/L, respectively. We established the RIs for the total protein and serum protein fractions using AGE in bottlenose dolphins under human care.

Anti-Müllerian Hormone Concentrations for Determining Resumption of Sertoli Cell Function following Removal of a 4.7 mg Deslorelin Implant in Tomcats.

Background: Deslorelin implant use in cats is a medical alternative to surgical sterilization, and due to its prolonged efficacy, its use has shown growing interest in the veterinary community. In the case of breeding facilities, its removal is often requested for the early restoration of testicular function. As anti-Müllerian hormones (AMH) in males is dependent of testosterone secretion, its assay may determine the restoration of testicular steroid secretion. An average of 3 weeks has been already described for tomcats' testicular function resumption after implant removal, but information about AMH concentrations in deslorelin-treated tomcats is lacking. Methods: Fourteen tomcats were treated for temporary suppression of fertility with a 4.7 mg deslorelin implant, which was surgically removed after 3, 6 or 9 months (n = 6, 4 and 4 tomcats, respectively). A general clinical and reproductive check with a gonadorelin stimulation test for testosterone determination was performed before deslorelin implant administration. After implant removal, tomcats' testicles were ultrasonographically checked for volume determination every 1-2 weeks with observation of the glans penis (presence or absence of spikes) and blood collection to assay both testosterone and AMH concentrations. Results: AMH concentrations increased significantly during the deslorelin treatment from 20.95 ± 4.97 ng/mL to 82.41 ± 14.59 ng/mL (p < 0.05). Following implant removal, AMH concentrations progressively decreased to pre-treatment levels, with a value of 28.42 ± 7.98 ng/mL on the third week post-removal where testosterone secretion was again detected. Conclusions: Even if a big variability of AMH concentrations exists between male individuals, resumption of tomcats' testicular function following a deslorelin treatment can be determined by AMH assay.

Biological Variation and Reference Change Value of Routine Hematology Measurands in a Population of Managed Bottlenose Dolphins (Tursiops truncatus).

Hematological analyses are particularly useful in assessing a dolphin's health status. However, the creation of appropriate reference intervals for this species is difficult due to the low number of reference individuals. The implementation of individual reference intervals (iRIs) allows researchers to overcome this limitation and, moreover, also consider the within-individual variability. The aims of this study were (1) to evaluate the biological variations in some hematological measurands, including erythrocytes (RBC), hematocrit (Hct), mean cellular volume and hemoglobin content (MCV and MCHC, respectively), RBC distribution width (RDW), leukocytes (WBC), and platelets (PLT); and (2) to calculate the index of individuality (IoI) and reference change value (RCV), which enable the production of iRIs, in healthy managed bottlenose dolphins. Seven dolphins were included, and the results of six hematological exams were analyzed for each animal. Analytical imprecision (CVa), within-dolphin variation (CVi), and between-dolphins variations (CVg) were calculated, and the IoI and RCV were derived for each measurand. All the hematological measurands had intermediate IoI except WBC, for which Iol was low. The calculated RCV ranged from 10.33% (MCV) to 186.51% (WBC). The results reveal that the majority of hematological measurands have an intermediate level of individuality in dolphins, and thus the application of iRIs is appropriate. The calculated RCV can also be applied to other managed dolphins and could be useful in interpreting serial CBC exams.

Prostatic fluid composition and semen quality in dogs with benign prostatic hyperplasia undergoing treatment with osaterone acetate.

Benign prostatic hyperplasia (BPH) may alter prostatic fluid biochemical composition causing reduced fertility. Osaterone acetate (OA) is an androgen receptor antagonist marketed for treatment of canine BPH. Little information exists on effects of OA administration on biochemical composition of canine prostatic fluid and its role on fertility. The aim of this research was to study biochemical composition of prostatic fluid and its role on semen quality in dogs with BPH undergoing treatment with OA. Eight intact, 5-11-year-old dogs with benign prostatic hyperplasia were treated orally with OA at a dose of 0.25-0.5 mg/kg once daily for seven days. Prostatic volume, semen evaluation and a biochemical analysis of prostatic fluid were performed on the day before treatment (D0), D60, D120, D180 and D240. A significant reduction (57% and 61%) of prostatic volume was observed at D60 and D120, respectively, and a significant reduction (20%) of normal spermatozoa was observed at D60 coincident with a significant increase of sperm tail defects, which disappeared during the course of the treatment. Prostatic fluid composition did not vary during the OA treatment except for zinc (Zn2+ ) with a significant increase at D120 and D180 correlated with the return to normal sperm values. In conclusion, canine Zn2+ prostatic fluid concentrations decrease during development of BPH and return to normal during treatment with OA. Zn2+ is an important electrolyte for semen quality, suggesting that oral Zn2+ supplementation might be considered a treatment to improve semen quality.

Autologous Platelet-Rich Plasma Enhances the Healing of Large Cutaneous Wounds in Dogs.

Platelet-rich plasma (PRP) is known to play a crucial role in skin wound healing, in both Human and Veterinary Medicine. Remarkably, until now, no studies have reported PRP treatment in subacute full-thickness skin wounds of the dog. The aim of this study was to evaluate the effects of two consecutive applications of autologous PRP, with the second application after 15 days, in 6 dogs showing large subacute skin wounds. The percentage of contraction, re-epithelialization and healing in all treated patients indicated that no complications or side effects, associated with consecutive PRP treatments, occurred in any patient and all wounds achieved complete closure and re-epithelialization. Our results suggest a positive effect of repeated autologous topical PRP treatments in large cutaneous subacute wounds of different etiology. Therefore, this PRP treatment could represent a simple, cost-effective, and valid alternative to promote healing processes in subacute large wounds cases in dogs.

Evaluation of an automated immunoturbidimetric assay for detecting canine C-reactive protein.

C-reactive protein (CRP) is a major acute-phase protein, and it is produced by the liver in response to a pro-inflammatory stimulus. Given that human and canine CRP have a similar molecular structure, the assays used for human CRP detection have been used to measure CRP concentrations in dogs. We evaluated the use of a human CRP assay (Biotecnica CRP assay) and validated its application in dogs. We analyzed 91 canine serum samples with a fully automated analyzer. Our validation was based on the evaluation of imprecision, limits of linearity, limits of quantification, and an evaluation of interferences. The new assay was also compared with the Randox CRP assay, a validated assay for the measurement of CRP. Intra- and inter-assay repeatability were <8% and <11%, respectively. The tested assay proportionally measured canine CRP in an analytical range up to 60 mg/L; however, hemoglobin, triglycerides, and bilirubin interfered with the determination. Good agreement, with the presence of proportional systematic bias, was observed between Biotecnica and Randox assays. The Biotecnica CRP assay provides reliable measurement of CRP in canine serum, provided that samples are free of interferents.

Identification of Plasma Fatty Acids in Four Lipid Classes to Understand Energy Metabolism at Different Levels of Ketonemia in Dairy Cows Using Thin Layer Chromatography and Gas Chromatographic Techniques (TLC-GC).

Excessive mobilization of adipose tissue in high milk producing dairy cows predisposes to metabolic diseases. The aim of this research was to identify the plasma fatty acids in four lipid classes as biomarkers for the diagnosis of hyperketonemia in bovines using thin layer chromatography and gas chromatographic techniques (TLC-GC). Sixty multiparous Holstein-Friesian dairy cows were enrolled in the study. Blood samples from the coccygeal vein were collected and ß-hydroxybutyrate (BHB) was evaluated. All animals were divided into three groups on the basis of ketonemia: BHB < 0.50 mmol/L, 0.50 < BHB < 1.0 mmol/L, and BHB > 1.0 mmol/L. Plasma fatty acid concentrations were evaluated in four lipid classes: Free Fatty Acids (FFA), Triglycerides (TG), Cholesterol Esters (CE) And Phospholipids (PL). The concentration of fatty acids was analyzed using TLC-GC. The results showed the following significance in the lipid classes: 19 fatty acids were significant (p < 0.053) in FFA, nine fatty acids were significant (p < 0.050) in TG, eight fatty acids were significant (p < 0.050) in CE and three fatty acids were significant (p < 0.049) in PL. Eleven parameters were considered as predictive fatty acids related to animals in hyperketonemia. The FFA increased simultaneously with blood BHB levels, although the identified predictive fatty acids related to the TG and CE lipid classes decreased, meanwhile the BHB values increased. In the PL lipid class, no fatty acids were predictive.

Serum haptoglobin and protein electrophoretic fraction modifications in buffaloes (Bubalus bubalis) around calving and during early lactation.

Serum protein distribution and concentration can be affected by different physiological and pathological conditions. The aim of this study was to evaluate the changes in the concentration of serum protein fractions and haptoglobin in clinically healthy dairy buffaloes during late pregnancy and early lactation. Blood and milk samples were collected from 30 buffaloes at around 7 d before expected calving (blood only) and 7, 30 and 50 d after calving. In serum samples, the total protein, haptoglobin, albumin, α1-, α2-, ß1-, ß2-, γ-globulins, and albumin/globulin ratio (A/G) values were evaluated. In milk, fat%, protein%, lactose%, somatic cell score (SCS) were assessed, along with milk yield (MY) and daily milk production (DMP). The peripartum period significantly influenced (P < 0.005) total protein, albumin, haptoglobin, α2-, ß2- and γ-globulins (P < 0.005). Milk yield, DMP and fat% changed significantly throughout the monitoring period (P < 0.005). Milk yield and DMP were positively correlated with total protein, albumin, ß2-globulins and A/G ratio, and negatively correlated with haptoglobin and α2-globulins. These results provide new knowledge about the serum protein electrophoretic pattern in Italian Mediterranean Buffaloes during the last phase of pregnancy and early stages of lactation.

Metabolic and hormonal adaptation in Bubalus bubalis around calving and early lactation.

Pregnancy and lactation are physiological periods that result in an increased metabolic demand that, if not satisfied, could provoke a threat to homeostasis. In this study changes in the values of Non-Esterified Fatty Acids (NEFA) and ß-hydroxybutyrate (BHB), glucose, insulin, thyroid hormones, milk composition and yield were investigated in buffaloes during the late pregnancy and early lactation. From a total of 50 buffaloes, blood samples were collected -7±5 days before expected calving; +7±5; +30±5 and +50±5 days post-partum; milk samples were collected at the same post-partum time points. On serum samples, the values of Non-Esterified Fatty Acids (NEFA), ß-hydroxybutyrate (BHB), glucose, insulin, Triiodothyronine (T3), Thyroxine (T4) and Thyroid-stimulating hormone (TSH) were evaluated. On milk, fat %, protein %, lactose %, somatic cells score (SCS), milk yield and daily milk production (DMP) were assessed. Peripartum period significantly influenced all studied parameters (P<0.05). Milk constituents and productivity statistically changed throughout monitoring period (P<0.005). Milk yield resulted positively correlated with insulin and TSH values, negatively correlated with NEFA and BHB. Insulin was negatively correlated with lactose % and positively correlated with SCS. The obtained results showed that the peripartum period and lactation are accompanied by marked changes in some biochemical variables and in the thyroid hormones values in Italian Mediterranean Buffaloes. Moreover, the relationship found between TSH values and milk yield seems to suggest a possible role of thyroid gland on the maintenance of lactogenesis. This study underlines the importance of monitoring the hormonal status of buffalo during the transition period in order to understand when adjustments of regulatory mechanisms break through physiological limits predisposing the buffalo to metabolic problems.

Salivary pH, calcium, phosphorus and selected enzymes in healthy dogs: a pilot study.

BACKGROUND: Saliva in dogs, as in humans, is a complex fluid secreted by different salivary glands in the oral cavity to protect the oral mucosa and teeth. The use of saliva as a substitute for blood in diagnosing and prognosticating disease in humans is widely accepted. Salivary biochemistry has also been used as a marker for periodontal disease in humans. No studies have as yet investigated the relation between salivary biochemistry and periodontal disease in dogs, however; neither has the salivary composition of healthy dogs with no oral disease been assessed. The purpose of this study was to obtain an overview on pH distribution and a set of salivary biochemical analytes (calcium, phosphorus, lactate dehydrogenase, lysozyme and amylase) commonly related to oral health in humans in a subset population of healthy young dogs with no periodontal disease or previous oral disease. Data were analyzed to gather salivary reference ranges for pH and each parameter and to assess a possible correlation between salivary and serum analytes. RESULTS: Twenty-nine adult client-owned dogs were recruited for the study. Lactate dehydrogenase and lysozyme showed higher concentrations in saliva than in serum, whereas amylase showed the contrary. Salivary biochemistry values did not differ between males and females or between non-neutered and neutered individuals. No significant correlations between salivary and serum calcium, phosphorus, lactate dehydrogenase, amylase and lysozyme were identified in this study. Data allowed intervals for the salivary pH and other analytes investigated to be obtained from healthy dogs with healthy oral conditions. CONCLUSIONS: These preliminary data can contribute to enlarge our understanding of the functional role of saliva and its relation to oral health in dogs.

Effect of combined acetylmethionine, cyanocobalamin and α-lipoic acid on hepatic metabolism in high-yielding dairy cow.

The aim of the study reported in this Research Communication was to investigate the effect of a combined acetylmethionine, cyanocobalamin and α-lipoic acid treatment, on some metabolic parameters of early lactating high-yielding dairy cows. Thirty cows were randomly divided into two groups: experimental group (EG, n = 20) and control group (CG, n = 10). EG received 20 ml of treatment and CG received 20 ml of placebo. The treatments were administered for seven times every 2 d. Blood samples were collected from all cows at 3 time points: 10 ± 2, 30 ± 2 and 50 ± 2 d postpartum. Glucose, ß-hydroxybutyrate (BHB), nonesterified fatty acids (NEFA), triglycerides, total cholesterol (TC), alanine aminotransferase (ALT), aspartate aminotransferase (AST), γ-glutamiltransferase (GGT), total bilirubin (TB), conjugated bilirubin (CB), total proteins (TP), globulins, albumin and urea concentrations were determined. Two-way repeated measure analysis of variance was applied. Significant differences in the values of glucose, BHB, NEFA, triglycerides, TC, AST and urea were found between EG and CG. Moreover, the increased glucose, TC, ALT, GGT, TP and globulins, and the reduced BHB, NEFA, AST, triglycerides, TB, CB and urea concentrations were evident in both groups, but the changes were more pronounced in EG. Our findings indicate that our treatment positively influenced liver metabolism in high-yielding dairy cows.