Volume 59 (2015): Issue 3 (September 2015)

The aim of this study was to determine the ceruloplasmin (Cp) and vitamin C concentrations, the total antioxidant status (TAS), and selected biochemical parameters in dairy cows spontaneously infected with bovine leukaemia virus (BLV). Of the 27 cows included in the study, 18 animals were seropositive for enzootic bovine leukosis (EBL), whereas nine cows were seronegative and were used as controls. The serum aspartate aminotransferase (AST) (P = 0.003) and Cp concentrations (P = 0.03) decreased (65.17 ± 5.03 and 7.70 ± 0.72 respectively) in BLV-infected cows, as compared to healthy animals (100.67 ± 11.50 and 10.40 ± 0.70 respectively). A slight insignificant increase in alkaline phosphatase activity and unchanged levels of alanine aminotransferase, lactate dehydrogenase, calcium, magnesium, and TAS were demonstrated in EBL cows. As the TAS and vitamin C levels remained unchanged in EBL cows, it may be suggested that ruminants may compensate for the impaired oxidative/antioxidative balance. The results obtained also indicate that BLV may suppress AST and Cp synthesis or secretion in the liver through an unknown mechanism. The mechanism of action of BLV in hepatocytes, especially on AST and Cp, requires further investigation to elucidate the immune suppression caused by oncogenic retroviruses.

The aim of the study was to evaluate the presence of Mycoplasma bovis infection and co-infections with other Mycoplasma spp. infections in cattle. The tested population was one in the eastern region of Poland containing 66 dairy cows and 23 calves showing different clinical signs and suffering from pneumonia, mastitis, and arthritis. The incidence of M. bovis in co-infections with other Mycoplasma spp. was examined using serological traditional mycoplasma culture methods, and the molecular methods - PCR and polymerase chain reaction/denaturing gradient gel electrophoresis (PCR/DGGE). The PCR/DGGE method for detecting Mycoplasma spp. in cattle was used for the first time in Poland. The seroprevalence of M. bovis in the affected cattle herds in the eastern region of Poland was 47.8% in calves and 19.7% in dairy cows. The direct detection and identification of M. bovis from nasopharyngeal swabs by PCR revealed that 56.5% of calves were positive, but all of the dairy cows were negative. The PCR/DGGE identified eight (34.8%) instances of M. arginini and eight (26.1%) instances of M. bovirhinis co-infecting with M. bovis in ten calves. The seroprevalence of M. bovis in the tested population was 33.7%. Any future attempts to control mycoplasma infections require an insight into the current epidemiological situation of M. bovis infection and its relationship to other mycoplasmas in causing clinical disease in cattle. Using these diagnostic methods we have demonstrated that mycoplasmal infections are often caused by multiple species of Mycoplasma and not just the primary M. bovis pathogen.

A 3-year-old female fallow deer was subjected to the necropsy and virological testing, due to a suspected infectious disease in the herd of farmed deer in the Southeastern region of Poland. The animal was found negative for the presence of BVDV, BoHV-1, BTV, and EHDV antibodies and BVDV antigen. The toxicological examination did not reveal any coccidiostats, mycotoxins, rodenticides, carbamate pesticides, and organophosphorus pesticides. The flukes found during postmortem examination were first characterised microscopically as Fascioloides magna and later their identity was confirmed by PCR and sequencing. The autopsy revealed lesions characteristic for F. magna infection, including different size cystic spaces in the liver, filled with brownish mucous fluid and flukes, and black pigment covering the surface of parietal and visceral peritoneum with the highest concentrations localised next to the liver. The changes observed in the liver tissue were typical of liver cirrhosis. The results demonstrated that in Poland, where the cervid farming is developing dynamically, the problem of fascioloidosis is present and may probably exert a significantly negative influence on the productivity of such farms if no antiparasitic treatment is performed.

A liquid chromatography – tandem mass spectrometry (LC-MS/MS) method for the determination of oxytetracycline (OTC), 4-epi oxytetracycline (4-epi OTC), tetracycline (TC), 4-epi tetracycline (4-epi TC), chlortetracycline (CTC), 4-epi chlortetracycline (4-epi CTC), doxycycline (DC), minocycline (MINO), methacycline (META) and rolitetracycline (ROLI) residues in muscles was developed. The procedure consisted of an oxalic acid extraction followed by protein removal with trichloroacetic acid. Further solid phase clean-up on polymeric (Strata X) reversed phase columns was performed to obtain an extract suitable for LC-MS/MS analysis. The tetracyclines were separated on a C 18 analytical column with mobile phase consisting of 0.01% formic acid in acetonitrile and 0.01% formic acid in water in gradient mode. The method was validated according to the Commission Decision 2002/657/EC. The recoveries of all target compounds were 91.8% – 103.6%. The decision limits were from 109.0 to 119.8 μg/kg and detection capability varied within the range of 122.2 to 137.6 μg/kg, depending on the analyte.

An analytical validation of a screening ELISA for detection of chloramphenicol (CAP) in honey was conducted according to the Commission Decision 2002/657/EC and Guidelines for the Validation of Screening Methods for Residues of Veterinary Medicines. The analyte was extracted from honey with a water and ethyl acetate mixture, and CAP concentrations were measured photometrically at 450 nm. The recovery rate of the analyte from spiked samples was 79%. The cut-off level of CAP in honey as the minimum recovery (0.17 units) was established. Detection capability (CCβ) was fixed at 0.25 μg kg⁻¹. No relevant interferences between matrix effects and structurally related substances including florfenicol and thiamphenicol were observed. The ELISA method should be useful for determination of CAP residues in honey monitoring.

A total of 85 mussel samples of eight species were examined. Analysis of mercury in the freeze-dried samples was carried out by atomic absorption spectrometry method using direct mercury analyser AMA 254. The analytical procedure for determination of mercury was covered by the quality assurance programme of research and participation in national and international proficiency tests. Concentrations of total mercury in all investigated samples were found to be generally low, in the range of 0.033-0.577 mg/kg of dry weight and of 0.003-0.045 mg/kg of wet weight. The results indicate that obtained levels of mercury in bivalve molluscs are not likely to pose a risk to the health of consumers.

The aim of the study was the evaluation of the effect of ageing on the extent of myofibrillar proteins degradation and tenderness of beef in different crossbreeds, BB × HF and SM × HF, from which the musculus semitendinosus was obtained. The pH value, basic composition of meat, and colour parameters were determined on the 3^rd d post mortem. The Warner Bratzler shear force and the extent of protein degradation were evaluated in regard to the effect of ageing time. Meat of BB × HF crossbreed had a lower amount of intramuscular fat and higher protein content (P ≤ 0.05). The shear force decreased with ageing time in the case of both crossbreeds. However, the highest values were noted in SM × HF crossbreed on days 3 and 7 of ageing. The differences in proteolysis of myofibrillar proteins and polypeptides, determined by SDS-PAGE electrophoresis, were observed between crossbreeds and the ageing time. A significant decrease in desmin and increased levels of 49-46 kDa and 32-27 kDa polypeptides (products of proteolytic degradation) were observed with an increasing ageing time. In addition, the rate of increase in the amount of 32-27 kDa polypeptides was more significant in BB × HF crossbreed. The data obtained showed that tenderness and the extent of protein degradation are associated with ageing process and animals’ genotype.

The influence of acidic electrolysed water (AEW) treatment on inactivation of pure bacterial cultures inoculated onto the surface of agarised media and surface microbiota of pork meat were examined. Low-concentrated AEW (low concentration of sodium chloride and low current electrolysis) was generated by electrolysis (5 or 10 min) of 0.001% or 0.01% NaCl solution. The number of viable microorganisms was determined using a plate count method. The effect of AEW on bacterial cell morphology were investigated using scanning electron microscopy (SEM). After treatment with AEW, a significant, about 3.00 log reduction of Pseudomonas fluorescens, Yersinia enterocolitica, Escherichia coli, Staphylococcus aureus, Bacillus subtilis, Listeria monocytogenes, and Micrococcus luteus populations was observed. In the AEW treatment of pork, the highest reduction of total number of microorganisms (2.1 log reduction), yeast and moulds (2.5-2.6 log reduction), and psychrotrophs (more than 1 log reduction) was observed after spraying with 0.001% NaCl subjected to 10 min electrolysis. SEM revealed disruption and lysis of E. coli and S. aureus cells treated with AEW, suggesting a bactericidal effect. Higher available chlorine concentration (0.37-8.45 mg/L), redox potential (863.1-1049.8 mV), and lower pH (2.73-3.70) had an influence on the shape of bacteria and the number of breaks in the bacterial membrane.

As the test material mink feed with natural microflora was used. The analyses were conducted using Wrzosek and TPGY broth media, and Willis–Hobbs and Zeissler differential agar media. Wrzosek, Willis–Hobbs, and Zeissler media are described in Polish Standards approved by the National Standards Body in Poland and routinely used in detection of anaerobic bacteria in Poland. Detection and identification of C. botulinum was performed with a previously validated real-time PCR method based on ntnh gene detection, which is common in all C. botulinum toxotypes. The use of Wrzosek broth and Zeissler agar in routine analyses for detection and identification of C. botulinum was ineffective and limited. The obtained results showed the highest culturing process effectiveness in TPGY broth with 72 h incubation at 30°C and isolation on Willis–Hobbs agar. The real-time PCR method based on ntnh gene detection used in this study could be utilised as a supplementary tool to the mouse lethality assay.

The aim of this study was to determine the correlation between lipophilicity and maximum residue limit (MRL) value specified for veterinary drugs in the fatty tissue of various animal species. The analysis was performed on a group of 73 compounds with different modes of action and MRL values determined for the fatty tissue of animals. Additionally, the logarithm of water/organic phase partition ratio (LogP) and the ratio of ionised and unionised substance in buffer with pH 7.4 (LogD_7.4) were calculated. The main analysis was performed after the division of the whole group into six fractions. The linear correlation and regression analysis were used to determine the indirect relationship between the mean arithmetic value of LogP or LogD_7.4 in selected fractions and related LogMRL of the drugs tested. The calculations revealed a linear correlation between fractioned lipophilicity and LogMRL values for the analysed compounds. The existence of indirect relationship between lipophilicity and MRL values determined for fatty tissue was confirmed.

The aim of the study was to analyse the effect of the feed additive alfalfa protein concentrate (APC), on pig health. The trial involved 40 crossbred gilts and 40 crossbred castrates (Polish Landrace × Polish Large White) × Duroc of 29.0 ± 0.5 kg initial body weight. Allocation of experimental animals was into four treatment groups: the control group (C) was fed standard mixtures, without APC addition; group E-15 was fed a basal diet supplemented with 1.5% APC; and groups E-30 and E-30P were fed diets with 3.0% APC inclusion. There were two feeding systems. In the first system, animals of groups C, E-15, and E-30 were fed continuously with suitable mixtures. The second feeding system was used in group E-30P where animals received the experimental or control mixture alternating at two-week intervals. The addition of APC supplement to diets significantly increased (P ≤ 0.05) red blood cell indices, i.e. haematocrit (Ht), red blood cell count (RBC), and haemoglobin concentration (Hb) in growing and finishing periods. The analysis of enzyme activity demonstrated a markedly higher activity of aspartate aminotransferase (AST), alanine aminotransferase (ALT), and especially alkaline phosphatase (ALP) in the blood plasma of pigs fed APC supplement. This increase may indicate a negative impact of APC on the animal’s liver. A positive effect of dietary APC on blood lipid parameters was associated with a decreased level of total cholesterol and reduced low-density lipoprotein fraction. Analysis of the haematological and biochemical blood indices demonstrated that APC additive may affect animal health.

The aim of the study was the evaluation of changes in the percentage profile of CD4+, CD8+, and CD25+ T lymphocytes, and their predictive value with respect to the course of experimental skin burns and early necrectomy in pigs. Thirty Large White Landrace pigs of both genders, weighing 50 kg (±2 kg), were used. Burns to their skin were performed with the use of a computer-controlled heating plate, applied to the animal’s body and heated to 2000°C, using 2.5 kg pressure for 10 s. It produced a burn of 30% (±2%) of body surface with a range of damage between II b° and III°. In animals of each experimental group fascial necrectomy was performed, according to the testing module. Blood from experimental and non-treated control animals was collected from the external jugular vein before the beginning of the experiment (hour 0) and at 12, 24, 36, 48, 60, 72, 84, 96, 108, 120, 132, 144, 156, 168, and 180 h of the experiment. An immune response profile was evaluated using flow cytometry analysis of the level and expression dynamics of CD4+, CD8+, and CD25+ particles on the surface of T lymphocytes. The study demonstrated that experimentally-induced burns in pigs caused cell-mediated immune response reflected in the changes in the percentage of CD4+, CD8+, and CD25+ T lymphocytes, and that early necrectomy in burnt pigs acted in a protective manner for the organism, based on the immunological index values. The study also proved that the dynamics of cell-mediated immunological response intensification determined on the basis of the percentage of CD4+, CD8+, and CD25+ T lymphocytes is conditioned by the size of the burnt surface and the time of necrectomy procedure.

The aim of the study was to evaluate the distribution and number of mast cells and eosinophils in rat mammary gland tumours induced by N-Nitroso-N-methylurea. The highest density of mast cells was found in cystic papillary adenocarcinomas of grade II. Eosinophils were detected only in the cystic papillary adenocarcinoma of grades I and II, in non-invasive cribriform adenocarcinoma and comedo-type carcinoma. Mast cell populations were observed perivascularly in the tumour stroma, in the host tumour interface, as well as in necrotic areas of neoplasms. Mast cells were observed to be intact according to their morphological changes, collectively referred to as degranulation. The obtained results indicate that mast cells and eosinophils play an important role in tumour micro-environment formation. The increased density of these cells in experimentally-induced rat mammary gland tumours suggests a poor prognosis in these cancers. Our results also confirmed that rat mammary gland tumours are good models for the study of breast cancers.

The aim of the study was to determine total antioxidative capacity (TAC) and zinc concentration in serum of dogs suffering from perianal tumours just before the start of the antihormonal treatment (AHT) and one and six months later. The study was performed on 45 dogs divided into two groups: control group suffering from non-malignant tumours (N = 24) and a group with malignant neoplastic changes (N = 21). Serum TAC and zinc concentrations were measured using photometric and atomic absorption spectrophotometric methods. Six months after the start of the AHT, TAC was significantly lower by 10.6% in dogs with malignant tumours when compared to controls (P = 0.03). In the non-malignant group, serum zinc concentration was higher before the treatment than in the malignant group, while the opposite results were observed six months later (P < 0.001). In the non-malignant group, gradually decreasing values of serum zinc concentration at each stage of the investigation were observed, while the opposite results were obtained in the malignant group (P < 0.05). The obtained results indicate that malignant neoplastic process is associated with significantly reduced TAC. Determination of serum zinc concentration in dogs with non-malignant and malignant perianal tumours may have practical diagnostic and prognostic values and may serve towards increasing the effectiveness of AHT monitoring.

Non-healing wounds are associated with high morbidity and might greatly impact a patient’s well-being and economic status. For many years, scientific research has focused on developing and testing several natural and synthetic materials that enhance the rate of wound healing or eliminate healing complications. Honey has been used for thousands of years as a traditional remedy for many ailments. Recently, honey has reemerged as a promising wound care product especially for infected wounds and for wounds in diabetic patients. In addition to its proposed potent broad-spectrum antibacterial properties, honey has been claimed to promote wound healing by reducing wound hyperaemia, oedema, and exudate, and by stimulating angiogenesis, granulation tissue formation and epithelialisation. Several animal models, including large animals, dogs and cats, and different species of laboratory animals have been used to investigate the efficacy and safety of various natural and synthetic agents for wound healing enhancement. Interpreting the results obtained by these studies is, however, rather difficult and usually hampered by many limiting factors including great variation in types and origins of honey, the type of animal species used as models, the type of wounds, the number of animals, the number and type of controls, and variation in treatment protocols. In this article, we provide a comprehensive review of the most recent findings and applications of published experimental and clinical trials using honey as an agent for wound healing enhancement in different animal models.

The therapeutic effects of Sidr honey on second-intention healing of contaminated full-thickness skin wounds in dogs were investigated. Povidone-iodine was used as a standard treatment and served as a control. Healthy adult (two-to-four-year-old) mongrels, comprising six dogs and two bitches, were divided into four equal groups in order to obtain multi-aged wounds at the end of the study. Four 2cm × 2cm full-thickness skin wounds were created on both sides of the back area of each dog under general anaesthesia adhering to aseptic technique. Contaminated wounds were then divided into two treatment groups: Group 1, Sidr honey treated (right side wounds) and group 2, povidone-iodine treated (left side wounds). All wounds were evaluated grossly daily at the time of treatment application and digitally photographed once every week. Images were analysed using ImageJ software. The parameters of unhealed wound area and length of advancing epithelium were obtained. The epithelialisation areas, percentage of wound area, and wound contraction rate were then calculated. No significant differences were found between the two treatment groups in any of the parameters studied. Overall, both honey treated and iodine treated wounds healed well within the time period of the study (28 d). However, the study showed a beneficial effect of Sidr honey on second-intention healing of full thickness contaminated wounds in healthy dogs and the effect was comparable to that of Povidone iodine.