Evaluation of a method for assaying sulfonamide antimicrobial residues in cheese: hot-water extraction and liquid chromatography-tandem mass spectrometry

Several sulfonamide antimicrobials (SAAs) are largely used in veterinary medicine. A rapid, specific, and sensitive procedure for determining 12 SAAs in cheese is presented. The method is based on the matrix solid-phase dispersion technique followed by liquid chromatography (LC)-tandem mass spectrometry (MS) equipped with an electrospray ion source. Target compounds were extracted from Mozzarella, Asiago, Parmigiano, Emmenthal, and Camembert cheese samples by 6 mL of water modified with 10% methanol and heated at 120 degrees C. The addition of methanol to hot water served to improve remarkably extraction yields of the most lipophilic SAAs, that is, sulfadimethoxine and sulfaquinoxaline. After acidification and filtration, 100 microL of the aqueous extract was injected in the LC column. MS data acquisition was performed in the multireaction monitoring mode, selecting two precursor-to-product ion transitions for each target compound. Methanol-modified hot water appeared to be an efficient extractant, because absolute recovery ranged between 67 and 88%. Using sulfamoxole as surrogate analyte, recovery of the 12 analytes spiked in the five types of cheese considered at the 50 ng/g level ranged between 75 and 105% with RSD not higher than 11%. Statistical analysis of the mean recovery data showed that the extraction efficiency was not affected by the type of cheese analyzed. This result indicates this method could be applied to other cheese types not considered here. The accuracy of the method was determined at three spike levels, that is, 20, 50, and 100 ng/g, and varied between 73 and 102% with relative standard deviations ranging between 4 and 12%. On the basis of a signal-to-noise ratio of 10, limits of quantification were estimated to be <1 ng/g.     

Effect of high-pressure-moderate-temperature processing on the volatile profile of milk

The effects of high hydrostatic pressure on volatile generation in milk were investigated in this study. Raw milk samples were treated under different pressures (482, 586, and 620 MPa), temperatures (25 and 60 degrees C), and holding times (1, 3, and 5 min). Samples submitted to heat treatments alone (25, 60, and 80 degrees C for 1, 3, and 5 min) were used for comparison. Trace volatile sulfur compounds were analyzed using solid-phase microextraction (SPME) and gas chromatography (GC) with pulsed-flame photometric detection (PFPD), whereas the rest of the volatile compounds were analyzed using SPME-GC with flame ionization detection (FID). Multivariate analysis of variance (MANOVA) and principal component analysis (PCA) were used to study the effect of pressure, temperature, and time on volatile generation. Relative concentration increases of 27 selected volatile compounds were compared to an untreated sample. It was found that pressure, temperature, and time, as well as their interactions, all had significant effects (P < 0.001) on volatile generation in milk. Pressure and time effects were significant at 60 degrees C, whereas their effects were almost negligible at 25 degrees C. The PCA plot indicated that the volatile generation of pressure-heated samples at 60 degrees C was different from that of heated-alone samples. Heat treatment tended to promote the formation of methanethiol, hydrogen sulfide, methyl ketones, and aldehydes, whereas high-pressure treatment favored the formation of hydrogen sulfide and aldehydes.     

Peripheral T lymphocyte changes in neonatal piglets: Relationship with growth hormone (GH), prolactin (PRL) and cortisol changes

Taking into account the role played by the neuroendocrine network in affecting the early development of the immune response, the present study aims to assess neonatal immunity in piglets by testing peripheral lymphocyte age-related changes in relationship to plasma levels of some relevant immunoregulatory hormones, such as growth hormone (GH), prolactin (PRL) and cortisol. For this purpose, we studied the peripheral lymphocyte age-related changes in relationship to plasma levels of GH, PRL and cortisol in conventional piglets from birth (day 0) to 41 days of age. A significant decrease was observed in the total number of lymphocytes at day 0, with a subsequent constant increment up to 41 days of age. Concomitantly, the number of T cell subsets (mainly CD8(+) cells and double positive CD4(+)CD8(+)) was low at birth, with strong increments between the 19th and 41st days of life. The CD4(+) T cell number subset was less diminished at birth than that of CD8(+), albeit with significant increments in the post-weaning period. Of interest, gammadelta T cells, which are more involved in innate immune efficiency, displayed the same trend as CD8(+) T cells from birth to the 41st day of life. From day 0 up to the 19th day, significant inverse correlations were found between T cell subsets and GH or PRL or cortisol, albeit with more significant inverse correlations with cortisol. The high levels of GH and PRL in the pre-weaning period may be due to the fact that they have to counteract the cortisol-mediated negative effect on lymphocyte production and development. These findings suggest that stress condition occurs at birth with decreases in the immune parameters, in the same way as in human newborns, with a subsequent gradual normalisation and immune development, as shown by decreased cortisol, GH and PRL normalisation and concomitant increments in T cell subsets.     

Inhibition of hemoglobin- and iron-promoted oxidation in fish microsomes by natural phenolics

Natural phenolic antioxidants have been tested in hake (Merluccious merluccious) microsomes as inhibitors of lipid oxidation promoted by fish muscle prooxidants: hemoglobin (Hb), enzymatic NADH-iron and nonenzymatic ascorbate-iron. The phenolics selected were as follows: (a) a grape phenolic extract (OW), (b) a fraction (IV) with isolated grape procyanidins with a medium-low degree of polymerization and galloylation percentage, (c) hydroxytyrosol obtained from olive oil byproducts, and (d) a synthetic phenolic antioxidant, propyl gallate. All compounds delayed lipid oxidation activated by Hb, enzymatic NADH-iron, and nonenzymatic ascorbate-iron, excluding hydroxytyrosol that was not an effective antioxidant on oxidation promoted by nonenzymatic iron. The relative antioxidant efficiency was independent of the prooxidant system, IV > propyl gallate > OW > hydroxytyrosol, and showed a positive correlation with their incorporation into microsomes (p < 0.05). The reducing capacity or ability for donating electrons and the chelating properties may also contribute to the antioxidant activity of phenolics, although these factors were less decisive than their affinity for incorporating into the microsomes. Conversely, the inhibition of Hb oxidation by phenolics and their polarity did not seem to play an important role on antioxidant mechanism. These results stressed the importance of incorporating the exogenous antioxidants into the membranes where are located key substances for fish lipid oxidation (highly unsaturated phospholipids, iron-reducing enzymes, and endogenous alpha-tocopherol).     

Hemodynamic effects of methylprednisolone acetate administration in cats

OBJECTIVE: To investigate the mechanisms by which corticosteroid administration may predispose cats to congestive heart failure (CHF). ANIMALS: 12 cats receiving methylprednisolone acetate (MPA) for the treatment of dermatologic disorders. PROCEDURE: The study was conducted as a repeated-measures design. Various baseline variables were measured, after which MPA (5 mg/kg, IM) was administered. The same variables were then measured at 3 to 6 days and at 16 to 24 days after MPA administration. Evaluations included physical examination, systolic blood pressure measurement, hematologic analysis, serum biochemical analysis, thoracic radiography, echocardiography, and total body water and plasma volume determination. RESULTS: MPA resulted in a substantial increase in serum glucose concentration at 3 to 6 days after administration. Concurrently, RBC count, Hct, and hemoglobin concentration as well as serum concentrations of the major extracellular electrolytes, sodium and chloride, decreased. Plasma volume increased by 13.4% (> 40% in 3 cats), whereas total body water and body weight slightly decreased. All variables returned to baseline by 16 to 24 days after MPA administration. CONCLUSIONS AND CLINICAL RELEVANCE: These data suggest that MPA administration in cats causes plasma volume expansion as a result of an intra to extracellular fluid shift secondary to glucocorticoid-mediated extracellular hyperglycemia. This mechanism is analogous to the plasma volume expansion that accompanies uncontrolled diabetes mellitus in humans. Any cardiovascular disorders that impair the normal compensatory mechanisms for increased plasma volume may predispose cats to CHF following MPA administration.     

Differential induction of pathogenesis-related proteins in banana in response to Mycosphaerella fijiensis infection

Black leaf streak or “black Sigatoka” is one of the most important diseases affecting bananas and plantains worldwide. Very few studies have been published on the host-pathogen interaction of this pathosystem, particularly at the molecular level. The aim of this work was to analyze, under controlled conditions, the enzyme activity of peroxidase (POX), phenylalanine ammonia lyase (PAL), β-1, 3-glucanase (GLU) and chitinase (CHI) as well as the production of H₂O₂ in banana plants infected with Mycosphaerella fijiensis. Defence responses were examined and compared in a resistant (Calcutta 4) and a susceptible (Williams) cultivar. Plants were inoculated and tested for enzyme activity at 0, 6, 12, 18, 24, 48 and 72?h after infection (HAI) and 6, 9, 12, 15 and 18?days after inoculation (DAI). A rapid induction of PAL, POX and GLU was observed in the resistant cultivar at 6–18 HAI as well as H₂O₂ production at 72 HAI. In contrast, in the susceptible cultivar, induction of these enzymes was only observed from 6 DAI. These results suggest that the first 72 HAI are important in determining the response of the host to the disease. Further studies characterizing banana responses to M. fijiensis at the early stages of the infection are necessary in order to better understand this host-pathogen interaction.     

Increasing energy and lysine in diets for growing-finishing pigs in hot environmental conditions: consequences on performance, digestibility, slurry composition, and gas emission

The influence of dietary nutrient concentration on growth performance, manure composition, and gas emission was studied in pigs in hot environmental conditions. A total of 64 intact males and 64 females [(Landrace × Large White) × Pietrain] weighing 63.1 ± 9.7 kg were divided into 2 dietary treatments: high (HD: 14.39 MJ of DE/kg and 1.11% Lys) and low (LD: 13.97 MJ of DE/kg and 1.01% Lys) in energy and Lys contents. Pigs were allocated to 32 split-sex pens with 4 pigs/pen and 16 pens/treatment. Average productive performance was recorded for 41 d (phase 1). After phase 1, 12 females of 103.3 ± 3.15 kg (6 per treatment) were selected and housed individually, and feces and slurry were collected during 3 and 4 consecutive days, respectively, to calculate nutrient digestibility and measure gas emissions (phase 2). For gas emission measurements, slurry was pooled by treatment and stored for 76 d. Initial composition of slurry and pH were analyzed. Maximum and minimum temperatures registered in the barn throughout the growing period were 35.1 and 18.1°C, respectively. Animals fed the HD diet grew more efficiently than pigs fed the LD diet (G:F, 0.43 vs. 0.40; SEM = 0.01; P < 0.05). Fat digestibility was greater in HD compared with LD pigs (88.0 vs. 84.9%; SEM = 0.9; P < 0.05). Slurry from pigs fed the LD diet showed greater DM, OM, total N, and VFA contents than slurry from pigs fed the HD diet. Cumulative NH(3), CO(2), and especially CH(4) emissions were greater in the HD slurry compared with the LD slurry (192.4 vs. 210.g of NH(3)/m(3); 2,712 vs. 3,210 g of CO(2)/m(3); 1,502 vs. 2,647 mL of CH(4)/kg of OM). Increasing feed density in the present study led to a more efficient growth, a decreased nutrient concentration in the slurry, and a greater gas emission.     

Ovarian activity in F1 prepubertal ewe lambs under tropical conditions

Seventy crossbred ewe lambs, born from Pelibuey (PB) dams mated with five sire breeds: White Dorper × PB (WD × PB), Black Head Dorper × PB (BHD × PB), Ile de France × PB (Ile × PB) Katahdin × PB (KT × PB) and Pelibuey (PB × PB), were used to determine age and weight to puberty, follicular populations during prepubertal development, ovulation rate at first ovulation and function of the first corpus luteum (FCL). Ewe lambs were maintained under grazing conditions and were supplemented with 300 g head^− 1 day^− 1 of a concentrate with 14% CP and 3.0 Mcal ME kg^− 1 of DM. From 145 days of age, monthly observations were performed by endoscopy to monitor follicular populations and luteal structures. Ovarian follicles were classified according to size into small (≥ 1 and < 3 mm), medium (≥ 3 and ≤ 4 mm) and large (> 4 mm). Additionally, weekly observations by ultrasonography were performed in four ewes per genotype to assess follicular populations. Luteal function from first ovulation was determined by circulating concentrations of progesterone. Ovulation rate was not affected by genotype (P > 0.05). All breed groups had the same pattern of follicular populations, where most of follicles were small, followed by medium and a small proportion of large follicles. Maximum diameter of follicles was similar for all genotypes during prepubertal development (P > 0.05), with an average range of 3.9 ± 0.20 to 4.4 ± 0.20 mm. The BHD × PB reached puberty at the youngest age of 240.9 ± 13.0 days, compared to all other genotypes (P < 0.05), 259.1 ± 11.7 for WD × PB, 279.3 ± 9.6 for KT × PB and 289.0 ± 15.3 days for PB × PB and 308.6 ± 11.9 days for Ile × PB ewe lambs. Ile × PB had greater weight to first ovulation (31.5 ± 1.19 kg) as compared to other breed groups (P < 0.01). The largest percentage of FCL was for KT × PB ewes (100%) (P < 0.05), followed by BHD × PB (88.9%), PB × PB (83.3), WD × PB (78.6%) and Ile × PB (77.8%). There were no differences in ovulation rate attributable to type of lambing from which ewe lambs came from (P > 0.05). BHD × PB ewe lambs reached puberty at an earlier age than other in this study. KT × PB ewe lambs had the highest percentage of FCL in their first ovulation. Breed of sire Ile de France increased weight to puberty and decreased FCL at first ovulation. In conclusion, the BHD × PB ewe showed puberty at an earlier age than other genotypes, while the KT × PB had the highest percentage of functional corpus luteum in their first ovulation. Ewe lambs coming from single lambings reach puberty at a heavier weight than those coming from multiple lambings. The ovulation rate in the first ovulation of ewe lambs is not affected by genotype and type of lambing.     

Development and evaluation of a high-fidelity canine patient simulator for veterinary clinical training

High-fidelity human patient simulators have been used for decades in medical education to provide opportunities for students to practice technical skills, diagnostic and therapeutic planning, and communication skills in a safe environment. A high-fidelity canine patient simulator (CPS) was developed using components from a human patient simulator and a low-fidelity foam core canine mannequin. Ninety-six veterinary students participated in cardiopulmonary arrest scenarios in groups of three to five students. Afterwards, participants were asked to complete an anonymous online survey describing their experiences. A total of 70 students (73%) completed the survey. All of the students (100%) felt that the simulator session expanded their cardiopulmonary resuscitation (CPR) knowledge base, and 97% responded that their skills and abilities had improved. Students also expressed positive opinions about the CPS, with 89% agreeing or strongly agreeing that the CPS was realistic and 73% agreeing or strongly agreeing that the scenarios generated emotions similar to real clinical situations. Most participants (98.5%) agreed or strongly agreed that the simulator was an engaging learning experience. Students commonly commented that the simulations allowed them to practice communication and teamwork skills and were more effective than paper-based, problem-oriented learning opportunities and lecture. Students also commented that they wanted more opportunities to participate in simulation exercises. These results suggest that high-fidelity veterinary simulation is an engaging educational methodology that addresses some limitations of other forms of problem-based learning. More studies are needed to quantitatively determine the effectiveness of this novel veterinary educational technology in comparison with more traditional approaches.