Anna Zaghini

Aflatoxin B1 and Clinoptilolite in Feed for Laying Hens: Effects on Egg Quality, Mycotoxin Residues in Livers, and Hepatic Mixed-Function Oxygenase Activities

Ninety-six laying hens were allocated to four groups administered different diets (group 0-0 received a complete diet, group 0-AF received a diet supplemented with 2.5 ppm of aflatoxin B1 [AFB1], group 2-0 received a diet supplemented with 2% clinoptilolite [CPL], and group 2-AF received a diet supplemented with 2% CPL and 2.5 ppm of AFB1) for 4 weeks to evaluate the effect of AFBI and/or CPL on egg quality and the ability of CPL to interact with the oral administration of AFB1. The possible effects of AFB1 on cytochrome P450-dependent hepatic mixed-function oxygenase (MFO) activities were also evaluated. Mycotoxin reduced yolk weight, while CPL influenced albumen percentage relative to that of eggs laid by chickens in group 0-AF Eggs laid by chickens in groups 0-AF and 2-AF had stronger shells and weighed less than the eggs of other groups. The eggs of treated groups were lighter in color than those of the control group (P < 0.01), and the tendency to yellowness in eggs was increased by CPL, probably through the affinity of red pigments for adsorbents and a consequent prevalence of yellow tonality. Color parameters might be connected with AFB1s interference with lipid metabolism and pigment deposition. The livers of hens in groups 0-AF and 2-AF showed very low mycotoxin concentrations that were significantly different (P < 0.01). The highest levels observed were those in the livers of the hens receiving the diet supplemented with the mycotoxin alone. AFB1 did not exert any significant effects on the hepatic MFO activities examined.

Long Depletion Time of Enrofloxacin in Rainbow Trout (Oncorhynchus mykiss)

ABSTRACT The international production of farmed fish has been growing continuously over recent years. Until now few veterinary drugs have been approved by the European Union for use in aquaculture, and this has favored the off-label use of products authorized for use in food-producing animal species different from fishes among fish farmers. Adequate field studies are lacking, especially for those species called minor species which are consumed extensively only in some European countries. In the present investigation we studied the depletion of the fluoroquinolone antibacterial enrofloxacin over time in a minor species, the rainbow trout (Oncorhynchus mykiss), reared on a real fish farm and treated with medicated feed (10 mg kg of trout body weight−1 day−1). Edible tissue samples (muscle plus skin in natural proportions) and fish bone samples were analyzed for enrofloxacin and for its major metabolite, ciprofloxacin, by high-performance liquid chromatography with fluorescence detection at different times after the end of treatment. Our results show that at 500°C-day (in which degree-days are calculated by multiplying the mean daily water temperature by the total number of days on which the temperature was measured), which is the minimum withdrawal period established by European Economic Commission Directive No. 82/2001 for any type of product administered off-label, edible trout tissues might still contain about 170 μg of enrofloxacin kg−1, whereas the maximum residue level for enrofloxacin plus ciprofloxacin is set at 100 μg kg−1. To our knowledge, no studies of the depletion of enrofloxacin in rainbow trout have been performed. On the basis of the data obtained in the present study, we suggest a more appropriate withdrawal time of 816°C-day for the sum of enrofloxacin plus ciprofloxacin levels in rainbow trout muscle plus skin tissues.

Seabream (Sparus aurata L.): disposition of amoxicillin after single intravenous or oral administration and multiple dose depletion studies

Abstract Single dose administration (trial 1): Serum and tissue concentrations of amoxicillin (AMX) were investigated in seabream ( Sparus aurata L.) kept in seawater at 22 °C and 32‰ of salinity. Amoxicillin was given intravenously (i.v.) at 40 mg/kg b.w. or orally (p.o.) at 80 mg/kg b.w. The serum concentrations were examined by using both radioimmunoassay-microbial receptor technology (Charm II test for β-lactams) and microbiological assay (spores of Bacillus stearothermophilus ATCC 10149); the tissue levels were determined by Charm II test. A slow clearance of AMX from serum was observed after i.v. administration, the concentrations at 72 h ranging from 0.94 to 0.66 μg/ml. Despite using the trihydrate form of amoxicillin, the apparent oral bioavailability was only 0.33%. Low levels were determined in muscle, skin and liver. Multiple dose administration (trial 2): A depletion kinetic study was conducted at water temperature of 22–26 °C after in feed administration (10 days) of AMX at the dose of 80 mg/kg b.w./day. AMX was only occasionally detected at very low concentrations in muscle, liver, skin, and vertebrae both during and after treatment cessation. Different formulations (conventional, micronized and microencapsulated AMX) were assayed in seabream at 24–26 °C after a 5-day period on medicated diet at the dose of 80 mg/kg b.w./day (trial 3) to verify if the nonconventional forms could improve the tissue distribution of AMX after in-feed administration. The results achieved in muscle and adherent skin were below the LOQ at each scheduled sampling time regardless of the formulation administered.

Pharmacokinetics of cefovecin sodium after subcutaneous administration to Hermann's tortoises (Testudo hermanni)

OBJECTIVE To determine the pharmacokinetics of cefovecin sodium after SC administration to Hermanns tortoises (Testudo hermanni). ANIMALS 23 healthy adult Hermanns tortoises (15 males and 8 females). PROCEDURES Cefovecin (8.0 mg/kg) was injected once in the subcutis of the neck region of Hermanns tortoises, and blood samples were obtained at predetermined time points. Plasma cefovecin concentrations were measured via ultraperformance liquid chromatography coupled to tandem mass spectrometry, and pharmacokinetic parameters were calculated with a noncompartmental model. Plasma protein concentration was quantified, and the percentage of cefovecin bound to protein was estimated with a centrifugation technique. RESULTS Cefovecin was absorbed rapidly, reaching maximum plasma concentrations between 35 minutes and 2 hours after administration, with the exception of 1 group, in which it was reached after 4 hours. The mean ± SD time to maximum concentration was 1.22 ± 1.14 hours; area under the concentration-time curve was 220.35 ± 36.18 h•μg/mL The mean protein-bound fraction of cefovecin ranged from 41.3% to 47.5%. No adverse effects were observed. CONCLUSIONS AND CLINICAL RELEVANCE Administration of a single dose of cefovecin SC appeared to be well-tolerated in this population of tortoises. Results of pharmacokinetic analysis indicated that the 2-week dosing interval suggested for dogs and cats cannot be considered effective in tortoises; however, further research is needed to determine therapeutic concentrations of the drug and appropriate dose ranges.

Ochratoxin A in artisan salami produced in Veneto (Italy)

ABSTRACT Fifty samples of artisan salami purchased in Veneto (Italy) were analysed for the determination of ochratoxin A (OTA). The analytical method, based on a sample preparation procedure with immunoaffinity columns (IACs), together with analysis by high-performance liquid chromatography with fluorescence detection (HPLC-FD), has guaranteed a high rate of recovery (about 97%), limit of detection (LOD) and limit of quantification (LOQ), respectively, of 0.06 µg kg−1 and 0.20 µg kg−1. OTA was detected in five samples, but only one exceeded the guideline value (1 µg kg−1) established by the Italian Ministry of Health for pork meat and derived products. The results would seem to suggest that salami made with the traditional, non-industrial production method can be considered safe as regards contamination by OTA. However, the very high concentration observed in one sample proves that a high OTA contamination is also possible in this type of product. Thus, the controls of mycotoxin contamination must consider also salami.

Rapid depletion of marbofloxacin residues in rabbit after therapeutic treatment.

Although rabbit meat production represents a very small percentage of the world meat market, this percentage has been growing continuously during the last 30 years. Rabbit is considered a minor food species, and therefore no drugs are specifically registered for this animal. This situation encourages rabbit farmers to make off-label use of antibacterial drugs authorized for food-producing animal species other than rabbits. In the present study, the distribution and elimination of the fluoroquinolone antibacterial agent marbofloxacin in rabbit muscle, liver, and kidney was investigated. Marbofloxacin was chosen as a representative of a new generation of antibacterial drugs active against most gram-positive and gram-negative bacteria and mycoplasms; it is well tolerated and has short elimination times in bovine and swine species. Rabbits were treated with marbofloxacin at 2 mg kg of body weight(-1) for 5 days. Residual concentrations in liver, kidney, and muscle tissues were determined posttreatment with high-performance liquid chromatography and fluorescence detection. Marbofloxacin was rapidly distributed and eliminated from rabbit tissues. Concentrations were higher in the liver and kidney than in muscle. However, 48 h after the end of treatment, marbofloxacin concentrations dropped below the maximum residue level fixed for this antibacterial drug in cattle and pigs. Considering the efficacy of marbofloxacin for the treatment of the most common rabbit diseases, its tolerability, and its short elimination time as verified in the present study, use of this antibacterial drug could be extended to therapeutic treatment of rabbits.

Dual ligand insertion in gB and in gD of oncolytic HSVs for the retargeting to a producer Vero cell line and to cancer cells.

ABSTRACT Oncolytic viruses gain cancer specificity in several ways. Like the majority of viruses, they grow better in cancer cells that are defective in mounting the host response to viruses. Often, they are attenuated by deletion or mutation of virulence genes that counteract the host response or are naturally occurring oncolytic mutants. In contrast, retargeted viruses are not attenuated or deleted; their cancer specificity rests on a modified, specific tropism for cancer receptors. For herpes simplex virus (HSV)-based oncolytics, the detargeting-retargeting strategies employed so far were based on genetic modifications of gD. Recently, we showed that even gH or gB can serve as retargeting tools. To enable the growth of retargeted HSVs in cells that can be used for clinical-grade virus production, a double-retargeting strategy has been developed. Here we show that several sites in the N terminus of gB are suitable to harbor the 20-amino-acid (aa)-long GCN4 peptide, which readdresses HSV tropism to Vero cells expressing the artificial GCN4 receptor and thus enables virus cultivation in the producer noncancer Vero-GCN4R cell line. The gB modifications can be combined with a minimal detargeting modification in gD, consisting in the deletion of two residues, aa 30 and 38, and replacement of aa 38 with the scFv to human epidermal growth factor receptor 2 (HER2), for retargeting to the cancer receptor. The panel of recombinants was analyzed comparatively in terms of virus growth, cell-to-cell spread, cytotoxicity, and in vivo antitumor efficacy to define the best double-retargeting strategy. IMPORTANCE There is increasing interest in oncolytic viruses, following FDA and the European Medicines Agency (EMA) approval of HSV OncovexGM-CSF, and, mainly, because they greatly boost the immune response to the tumor and can be combined with immunotherapeutic agents, particularly checkpoint inhibitors. A strategy to gain cancer specificity and avoid virus attenuation is to retarget the virus tropism to cancer-specific receptors of choice. Cultivation of fully retargeted viruses is challenging, since they require cells that express the cancer receptor. We devised a strategy for their cultivation in producer noncancer Vero cell derivatives. Here, we developed a double-retargeting strategy, based on insertion of one ligand in gB for retargeting to a Vero cell derivative and of anti-HER2 ligand in gD for cancer retargeting. These modifications were combined with a minimally destructive detargeting strategy. This study and its companion paper explain the clinical-grade cultivation of retargeted oncolytic HSVs and promote their translation to the clinic.

Biliary ochratoxin A as a biomarker of ochratoxin exposure in laying hens: An experimental study after administration of contaminated diets

To evaluate the levels of ochratoxin (OTA) in kidney, liver and bile of laying hens, forty-five laying hens were enrolled in this study and divided into three equal groups: a control group D₀, and two experimental groups, D₁ fed with 10 µg/kg OTA diet and D₂ fed with 200 µg/kg OTA diet for 6 weeks. Kidneys, livers, and bile from all hens were collected and analyzed by HPLC method for the presence of OTA. Eggs collected 2 days before the start of the experiment and 2 days after its end were also analyzed for the presence of OTA. Results show a relevant biliary excretion of the mycotoxin, with high levels of OTA in the bile after administration of the toxin. OTA level in eggs was below the limit of detection (LOD). These results suggest the suitability of using bile as a matrix for screening measurements of OTA in laying hens.

Kinetics of Ketamine Plasma and Urine Metabolite Levels Following Intravenous Administration in the Dog

P. Roncada1*, A. Zaghini1, C. Riciputi1, N. Romagnoli2 and A. Spadari2 1Dipartimento di Sanità Pubblica Veterinaria e Patologia Animale; 2Dipartimento Clinico Veterinario, Sezione Chirurgica, Università di Bologna, Bologna, Italy *Correspondence: Dipartimento di Sanità Pubblica Veterinaria e Patologia Animale, V ia T olara di Sopra, 50, 40064 Ozzano Emilia (Bologna), Italy E-mail: roncada@vet.unibo.it

Evaluation of liver parenchyma and perfusion using dynamic contrast-enhanced computed tomography and contrast-enhanced ultrasonography in captive green iguanas (Iguana iguana) under general anesthesia

BackgroundContrast-enhanced diagnostic imaging techniques are considered useful in veterinary and human medicine to evaluate liver perfusion and focal hepatic lesions. Although hepatic diseases are a common occurrence in reptile medicine, there is no reference to the use of contrast-enhanced ultrasound (CEUS) and contrast-enhanced computed tomography (CECT) to evaluate the liver in lizards. Therefore, the aim of this study was to evaluate the pattern of change in echogenicity and attenuation of the liver in green iguanas (Iguana iguana) after administration of specific contrast media.ResultsAn increase in liver echogenicity and density was evident during CEUS and CECT, respectively. In CEUS, the mean ± SD (median; range) peak enhancement was 19.9% ± 7.5 (18.3; 11.7-34.6). Time to peak enhancement was 134.0 ± 125.1 (68.4; 59.6-364.5) seconds. During CECT, first visualization of the contrast medium was at 3.6 ± 0.5 (4; 3-4) seconds in the aorta, 10.7 ± 2.2 (10.5; 7-14) seconds in the hepatic arteries, and 15 ± 4.5 (14.5; 10-24) seconds in the liver parenchyma. Time to peak was 14.1 ± 3.4 (13; 11-21) and 31 ± 9.6 (29; 23-45) seconds in the aorta and the liver parenchyma, respectively.ConclusionCEUS and dynamic CECT are practical means to determine liver hemodynamics in green iguanas. Distribution of contrast medium in iguana differed from mammals. Specific reference ranges of hepatic perfusion for diagnostic evaluation of the liver in iguanas are necessary since the use of mammalian references may lead the clinician to formulate incorrect diagnostic suspicions.