Background: Measurement of adenosine deaminase (ADA) can provide information about cell-mediated immunity. This report’s objective was to study the enzymatic activity of total ADA (tADA) and its isoenzymes ADA1 and ADA2 in canine, equine, porcine, and bovine serum and saliva and their changes in different inflammatory situations in each species. Besides, an automated method for ADA2 measurement was developed and validated.Results: tADA was present in serum and saliva of healthy animals of the four species. Erythro-9-(2-hydroxy-3-nonyl) adenine (EHNA) concentration of 0.47 mM was needed for ADA1 inhibition in canine and porcine samples (serum and saliva) and bovine saliva, whereas for equine saliva 0.94 mM was needed. ADA2 activity was not detected in bovine serum and was very low or absent in equine serum and bovine saliva. An automated procedure to measure ADA2 consisting of adding EHNA to a commercial reagent for tADA measurement provided repetitive (coefficients of variation < 8.8% in serum and < 10% in saliva) and accurate (linearity of serial sample dilutions withR² > 0.90) results, being equivalent to a manual incubation of the sample with EHNA at a similar concentration. Salivary tADA, as well as ADA1 and ADA2, were higher in dogs with leishmaniosis, horses with acute abdominal disease and pigs with lameness than in healthy animals. tADA and isoenzymes in saliva showed a positive significant correlation with serum ferritin in dogs (r = 0.602,P < 0.01;r = 0.555,P < 0.05; andr = 0.632,P < 0.01; respectively for tADA, ADA1 and ADA2) and serum C-reactive protein in pigs (r = 0.700,P < 0.01, for both tADA and ADA1;r = 0.770,P < 0.001, for ADA2), whereas salivary ADA2 significantly correlated with serum amyloid A in horses (r = 0.649,P < 0.01). In cows, salivary tADA and ADA1 significantly increased after calving, correlating with total white blood cell count (r = 0.487,P < 0.05, for both tADA and ADA1).Conclusions: The activity of total ADA and its different isoenzymes, can be measured in serum and saliva of dogs, horses, pigs and cows by a simple and fast procedure described in this report. When measured in saliva, these analytes correlated with other biomarkers of inflammation and it could potentially be used as a biomarkers of inflammation and immune activation in the species of this study.

The use of anesthesia is a common practice in aquaculture to sedate fish and mitigate handling stress. Although the employ of anesthesia is considered beneficial for fish, as it reduces stress and improves welfare, at the same time it may induce hazardous side-effects. The aim of the present study was to investigate the effects of clove oil (CO) and tricaine methanesulfonate (MS222), two of the most used anesthetics, on several oxidative stress related parameters in gilthead sea bream (Sparus aurata), as these types of effects of anesthetics have been seldom investigated. To assess these effects, S. aurata juveniles were placed in a setup of mobile water tanks and were transported during 6 h with either 2.5 mg/L CO or 5 mg/L MS222. After transport, half of the fish were sampled, whereas the remaining fish were transferred to tanks without anesthetics where they were allowed to recover for 18 h before sampling. Changes in the expression levels of several target genes related with the antioxidant response and cell-tissue repair were evaluated in the gills, liver and brain. Those transcripts included glutathione peroxidase 1 (gpx1), catalase (cat), glutathione S-transferase 3 (gst3), glutathione reductase (gr), superoxide dismutase [Zn] (sod2), heat shock protein-70 (hsp70), and metallothionein (mt). Antioxidant enzymatic activities glutathione S-transferase, GST; catalase, CAT; and glutathione reductase, GR, levels of non-enzymatic antioxidants (non-protein thiols – NPT), and pro-oxidative damage, assessed as lipid peroxidation (LPO), were determined in gills, liver and brain. Acetylcholinesterase activity (AChE) was determined in plasma, gills, brain, muscle and heart as an indicator of neuro-muscular alterations. In plasma, the total antioxidant capacity (TAC) and total oxidative status (TOS) were also measured. Results showed that the use of both anesthetic agents, CO and MS222, interferes with fish antioxidant status. All tested biological matrices displayed alterations in antioxidant endpoints, confirming that these substances, although minimizing the effects of transport stress, may have long term effects on fish defenses. This result is of high relevance to aquaculture considering that the oxidative stress, may increase the susceptibility to different environmental or biotic stress and different types of pathologies.

Uric acid (UA) is the most abundant antioxidant compound in saliva and one of the most sensitive biomarkers for detecting changes in the oxidative status of the organism. The aim of this study was to evaluate the effect of: (i) different methods of saliva sampling and (ii) the correction by salivary flow or total protein on UA concentrations in saliva. Paired saliva (collected by two different methods, passive drooling and using Salivette cotton rolls) and serum samples were obtained from 12 healthy men after the performance of two resistance training exercises of different level of effort that can produce different concentrations in UA in saliva. There were no significant differences between values of uric acid in saliva using Salivette and passive drool. Correlations between UA in serum and saliva and increases in UA in saliva after exercise were detected when saliva samples were obtained by passive drool and Salivette and were not corrected by salivary flow or total protein concentration. Therefore for UA measurements in saliva it would not be recommended to normalize the results by salivary flow or protein concentration. This study highlights the importance of choosing an adequate sampling method selection as well as the expression of results when analytes are measured in saliva.