Volume 58 (2014): Issue 3 (September 2014)

This article reviews the present state of the science concerning the polybrominated dibenzo-p-dioxins (PBDDs) and dibenzofurans (PBDFs). Everywhere in the world people are exposed to anthropogenic origin chemicals. Some of them are long-lived organic compounds, which persist over the years in the environment. Persistent organic pollutants, such as organohalogen compounds, accumulate in environmental and biological compartments and have adverse effects on the health of humans and animals. Little is known about the brominated and mixed chloro/bromo dioxin and furans. Existing literature suggests that brominated dioxins and furans have similar toxicity profiles to their chlorinated analogues. The exposure data are extremely limited, showing a major data gap in estimating the potential environmental and health risk of these chemicals. The rapid increase in the use of brominated flame retardants (the main source of these pollutants) has raised the level of concern over environmental and health damage from brominated dioxins and furans. It is likely that human as well as wildlife exposure to these contaminants will increase with their greater use. The findings reported here present strong evidence of the PBDDs and PBDFs as an emerging new class of contaminants.

Q fever is a zoonotic disease caused by Coxiella burnetii. The main source of infection are ruminants (cattle, sheep, and goats). C. burnetii is excreted via birth products, vaginal mucus, milk, and faeces. Raw milk is considered useful for epidemiological examinations of animals and evaluation of infection dynamics at the herd level. This article summarises data on prevalence studies on C. burnetii in bulk-tank milk in different European countries with the means of serological tests and PCR. It also summarises the results of studies to evaluate the actual risk of disease transmission to humans through consumption of raw milk. Moreover, the available diagnostic tools for detection C. burnetii infection are presented.

The aim of the study was to identify the genotype of Polish isolates of salmonid alphaviruses (SAV) and to find the origin of the virus. Samples for virus isolation included the kidneys, spleen, and liver pooled from 10 fish. A typical cytopathic effect was observed after inoculation of samples on cell lines. Total RNA was extracted from cell culture supernatant and submitted to RT-PCR with primers amplifying two informative regions of the genome: a conserved region in the E2 gene and a variable region in the nsP3 gene. The sequences revealed that the strain from Poland belonged to subtype SAV 2, indicating a very strong genetic identity with isolates from Italy and France.

The aim of the study was to develop an in situ PCR (IS-PCR) method for detection of bovine leukemia virus (BLV) in cell cultures. Samples from five BLV positive and five BLV negative cows were collected and dendritic cells (DCs) from blood, bone marrow, spleen, and lymph node were cultured. Cultures prepared from healthy animals were infected with BLV. After two weeks, the cells were tested by nested PCR and IS-PCR for the presence of proviral DNA. As a positive control adherent cell line permanently infected with BLV was used. BLV was successfully detected by IS-PCR in DCs from naturally infected cattle and DCs infected in vitro. In control, non-infected DCs, the results of the reaction were negative. The results of provirus detection by IS-PCR were similar with these performed with nested PCR. Additionally, IS-PCR provides many advantages, like specific localisation of infection and smaller number of cells needed as template for PCR.

The study was performed on lambs experimentally infected with bovine leukaemia virus (BLV). The presence of BLV antibodies in sera of infected animals was detected by agar gel immunodiffusion test and ELISA. Proviral DNA was detected by PCR and nested PCR. Dual-colour flow cytometry analysis was performed with the use of specific monoclonal antibodies against lymphocyte CD markers and gp51 viral envelope protein, followed by incubation with fluorescent-labelled secondary antibodies conjugated with FITC or PE. Gp51 viral envelope protein was detected in tumours caused by BLV infection. The BLV infection resulted in depletion of CD4+ lymphocytes, increase in CD8+ lymphocytes, and decrease in CD4+ to CD8+ ratio in infected sheep. Proliferation of IgM+ CD19+ cells was also detected. These cells had an immature character without tendency to differentiate, and their vitality was prolonged. Flow cytometry enabled detection of gp51 expression in sheep blood lymphocytes at the early stages of the infection, before detection of serum antibodies using ELISA.

In total, 14 608 pig sera, collected between 2008 and 2011, were tested with ELISA using antibodies specific for bovine viral diarrhoea virus (BVDV). All doubtful and positive samples were retested by virus neutralisation test (neutralising peroxidase-linked assay). The BVDV seroreagents were detected in 11 (68.75%) out of 16 provinces, the seroprevalence varied from 0.1% to 1.04% (average 0.31%). The obtained results indicate that the prevalence of BVDV infection in pig population in Poland is low.

The aim of this study was to assess the impact of asymptomatic infections of mink with Aleutian mink disease virus (AMD on reproductive performance and the morphology of the internal organs. The study drew its subjects from two farms where the disease was endemic and included 450 clinically healthy female mink divided into three equal groups: two AMD positive groups and one AMD negative control group. The mink were subjected to clinical observations throughout the period of reproduction, and then for the first seven weeks after birth. Data collected during this period were evaluated using selected breeding and reproductive indices. All reproductive indices including conception rate, litter size, and litter weight at birth as well as neonatal mortality were substantially poorer. Histopathological examination revealed changes typical for AMD, particularly in the spleen, kidneys, liver, and lymph nodes. The results highlighted and proved the frequently underestimated extent of economic losses on farms where AMD is endemic yet the mink stock is apparently healthy.

The presence of Mycoplasma haemomuris (haemoplasma) in blood samples collected from specific pathogen-free (SPF) laboratory rats bred in Japan was reported. Its presence was examined in Fischer 344, Sprague-Dawley (SD), and Wistar rat strains of both sexes by real-time PCR. All strains were positive for M. haemomuris infection. The 16S rRNA gene of M. haemomuris strain detected in the animals was amplified using end-point PCR. Only the entire nucleotide sequence of 16S rRNA gene of a mycoplasma strain detected in SD rats was determined and compared to those of other haemoplasmas. Our investigations suggest a wide M. haemomuris infection among the SPF rats purchased from commercial breeders in Japan.

Serological study of porcine reproductive and respiratory syndrome virus (PRRSV) infection in pigs and wild boars was conducted in Lithuania between 2009 and 2013. Antibody level was measured using a commercial ELISA. The 4.32% (95% CI 3.92-4.72) out of 9856 examined porcine sera were positive for the PRRSV antibodies. The antibodies were detected in 11.82% (95% CI 10.28-13.36) of all investigated serum samples of sows and gilts. As much as 8.2% of serologically positive samples (95% CI 6.83-9.57) were determined in the piglets under three months of age. Considerably smaller (P < 0.05) seroprevalence was detected in boars (0.62%) and fattening pigs (1.84%). From 1357 examined sera of wild boar, collected between 2009 and 2013 hunting seasons, 5.38% (95% CI 4.52-8.2) of samples were positive for PRRSV antibodies in 23 locations out of 50 investigated. The analysis of seroprevalence in different age groups of wild boars showed that PRRSV antibodies were detected in all age groups; however, it was significantly higher in adults than in juveniles or subadults and reached up to 10.02% (95% CI 7.39-12.65).

The proteomics of inflammatory response in whey from cows with subclinical mastitis were analysed. Whey protein lysates were separated on 24 cm dry IPG strips (pH 3-10 linear) and 24 cm dry IPG strips (pH 4-7) using two-dimensional electrophoresis. The results indicated that the whey proteins in milk from cows with subclinical mastitis are different from those in milk from healthy cows. All protein spots were found to have biologically relevant changes in relative abundance during subclinical mastitis using matrix-assisted laser desorption/ionisation time-of-flight mass spectrometry analysis, including ß-1,4 galactosyltransferase, ß-2 microglobulin, complement 3, a-1-acid glycoprotein, ß-lactoglobulin A, a-S1 casein precursor, ß-casein B, and serotransferrin precursor. The mRNA expression of these genes was verified by quantitative real-time PCR. These proteins are involved in signal transduction, binding, transport, and immune defence activity. The results suggest that the markers may be used for the diagnosis of subclinical mastitis.

A PCR-based procedure for detection of C. botulinum C and D in corn silage samples was validated. During the validation, method specificity, sensitivity, and accuracy were determined according to PN - EN ISO 16140:2004. Additionally, the specificity of the validated methods was proved by sequence analysis of PCR products obtained from examination of samples connected with botulism cases in cattle and mallard ducks. Limit of detection was estimated according to the Spearman - Kärber formula and expressed as LOD50. The obtained results showed that a 100% specificity was achieved. The sequencing of PCR products revealed 99% identity with sequences of bont/C and bont/D genes deposited in the GenBank. The sensitivity value ranged from 63.3% for C. botulinum type C to 75% for type D. The accuracy value varied from 72% for type C to 81.3% for type D. LOD50 was estimated at the levels of 0.272 (0. D 188-0395) spore/g for type C and 0.17 (0.1-0.289) spore/g for type D. The described PCR-based procedure enabled detection of C. botulinum C and D at the stage of liquid culture. This makes examination of feed samples possible without isolation process. The presented procedure could support the diagnosis of botulism by faster and specific laboratory examination process.

The occurrence of commonly used veterinary antimicrobial agents was investigated in 159 fresh water, 443 fish, and 150 sediment samples from Polish rivers and lakes. The agents included aminoglycosides, ß-lactams, diaminopyrimidines, fluoroquinolones, lincosamides, macrolides, pleuromutilins, sulfonamides, and tetracyclines. The analysis was performed by three different sample preparation procedures for each matrix and it was performed by liquid chromatography-tandem mass spectrometry with electrospray ionisation source in positive mode, under the same conditions. All analytical methods used were validated and showed good sensitivity, accuracy, and precision. The LOQ was in the range from 5 μg/kg to 125 μg/kg for fish samples, from 0.02 μg/L to 10 μg/L for fresh water samples, and from 1 μg/kg to 8 μg/kg for sediment samples.

The occurrence and concentrations of organochlorine pesticides and polychlorinated biphenyls (PCBs) were determined in 158 muscle samples of bream (Abramis brama) and roach (Rutilus rutilus), and 84 samples of sediments collected from 10 river and lake sampling sites in 2011 and 2012. The concentrations of DDTs (p,p’-DDT, o,p’-DDT, p,p’-DDE, and p,p’-DDD), HCH isomers (a-, ß-, and y-HCH), HCB, and PCBs (six indicator PCB congeners 28, 52, 101, 138, 153 and 180) were determined using the capillary gas chromatography. The mean concentrations of DDTs in bream and roach were in the range of 11.2-654 and 4.5-121 ug/kg wet weight respectively, and PCBs were in the range of 1.3-75.9 and 1.1-112 ug/kg wet weight, respectively. Mean concentrations of DDTs and PCBs in sediments were 0.5-270 ug/kg dry weight and ⋋0.1-2.2 ug/kg dry weight respectively. The study showed clear spatial differences in the levels of organochlorine pesticides and PCBs in fish and sediments from different aquatic ecosystems. The highest levels of contaminants were detected in fish and sediments from the Vistula River in vicinity of Cracow. The possible risk to the fish meat consumers and ecological risk were evaluated.

A rapid liquid chromatography tandem mass spectrometry method was developed and validated to detect and confirm five thyreostatic drugs: tapazole, thiouracil, methylthiouracil, propylthiouracil, and phenylthiouracil in animal feeding stuff samples. Thyreostats were extracted from feed with methanol, and then degreasing of the extract with petroleum ether was performed, followed by the derivatisation of the compounds with 3-iodobenzylbromide in basic medium (pH 8.0). The derivatives were extracted with diethyl ether and analysed by gradient elution on a Poroshell 120-EC C18 column with triple quadrupole MS detection with turbo spray source in positive ionisation mode. The method was validated in accordance with the Commission Decision 2002/657/EC. For validation level of 10 ļig kg^-1, the recovery ranged from 82% to 97.5% for all examined compounds. The repeatability and reproducibility did not exceed the limit of 20% for all analytes. The linearity was good for all thyreostats in the whole range of tested concentrations, as proved by the correlation coefficients greater than 0.99. The decision limits (CCa) ranged from 1.63 ļig kg^-1 to 3.95 ļig kg^-1, whereas the detection capabilities (CCß) ranged from 2.74 ļig kg^-1 to 6.73 ļig kg^-1. The developed analysis is sensitive and robust, and therefore useful for quantification and confirmation of thyreostats in residue control programme.

The study presents the analysis of the findings of 234 post-mortem examinations on free-ranging and captive European bison selectively culled or having fallen between 2008 and 2013 in Białowieża Primeval Forest. Pneumonia, emphysema, nephritis, bodily traumas, and intestinal lesions were observed in 106 (45.3%), 77 (32.9%), 82 (35.0%), 68 (29.1%), and 56 (23.9%) animals respectively and were the most common pathological changes. Almost half of all males (66 out of 140; 47.1%) tested showed some pathological changes of prepuce and penis, described as posthitis or balanoposthitis. Infection with liver fluke (Fasciola hepatica) and lungworm (Dictyocaulus viviparus) was observed macroscopically in 114 (48.7%) and 80 (32.9%) bison respectively. F. hepatica prevalence was associated with the emergence of other liver changes such as hepatitis and cirrhosis (P < 0.001). Similarly, the prevalence of D. viviparus coincided with pneumonia (P = 0.001), changes in the upper respiratory tract (P = 0.04), and emphysema (P < 0.001). Hepatitis, infection with F. hepatica, and pathological lesions in the male and female reproductive tracts were associated with the animals’ age. Mechanical injuries, caused by other bison or less commonly by traffic accidents, were the most common cause of death of bison below six months of age. Most pathological changes were significantly more frequent in the selectively culled animals in comparison with the ones having fallen, which confirms the desirability of elimination as a tool to improve the health and welfare of the bison population and limit the number of reservoirs of invasive and possibly infectious diseases.

The aim of the study was to find out the relationship between the progression of endometrosis and the appearance of the structural microscopic changes in mares’ ovaries. The investigation was performed on slaughtered mares of various age and breed. Four groups each received a portion of the specimens, and designation to group was according to the stage of endometrosis determined on the basis of Kenney’s classification. Uterine and ovarian sections were stained with haematoxylin and eosin. The results of the microscopic evaluation were compared between mares classified into specific Kenney’s categories. It was shown that an increase in ovarian follicular cysts was related to mares’ ages and correlated with significant progression of the endometrosis. These observations suggest that the same aetiological factors may take part both in triggering disorders of ovarian oo/folliculogenesis and in spurring uterine endometrosis. Further more detailed methods of investigation are needed to elucidate the mechanisms of both disease processes.

Samples of tumours collected during necropsies or received from veterinary clinics, as well as biopsies were analysed. The analysis revealed 2259 tumours, including 1814 (80.30%) cases from dogs, followed by 295 (13.06%) cases from cats, 79 (3.50%) cases from horses, and 71 (3.14%) cases from exotic animals. Skin and subcutaneous tissue (45% of dogs and 48% of cats), and mammary gland (23% of dogs and 29% of cats) were the most common location of tumours. Malignant forms prevailed in dogs (59%), cats (66%), and horses (89%). The analysis revealed interesting facts concerning the middle age of dogs and cats with tumours present with a peak in nine-year-old dogs and 10-12-year-old cats.

The expression of miRNA-21, miRNA-10b, and miRNA-34a in malignant and benign tumours and non-neoplastic lesions in canine mammary gland, using real-time PCR with TaqMan probes was determined. The expression in normal tissues was compared to neoplastic and non-neoplastic lesions using one-way ANOVA test. Significant changes in miRNA expression in neoplastic tissues, as compared to normal ones, were demonstrated. In all neoplastic tissues, the miRNA-21 expression increased while in non-neoplastic lesions slightly decreased in comparison to normal ones. MiRNA-10b expression in malignant and benign tumours increased in comparison to normal tissues and non-neoplastic lesions. MiRNA-34a expression profile in neoplastic and non-neoplastic tissues differed from other examined miRNAs (miRNA-21 and miRNA-10b). In all samples miRNA-34a expression level decreased in comparison to normal tissues.

The aim of the study was to evaluate the effect of an inulin source (aqueous or aqueous-alcoholic extract, dried chicory root, or dried Jerusalem artichoke tubers) on piglet performance, plasma lipid profile, and immunoglobulin concentration. The study was conducted on 534 piglets (44 litters) allocated to five nutritional groups: group I (control) - fed basal diet, groups II and III receiving basal diet supplemented with 2% of inulin (aqueous and aqueous-alcoholic extract respectively), and groups IV and V -4% additive of dried artichoke or dried chicory respectively. During the trial, piglets’ body weight, feed intake, diarrhoea incidence, and mortality were controlled. Blood samples were collected twice from six animals of each group. In blood plasma, indices of lipid profile and concentrations of IgA, IgG and IgM were measured. The addition of inulin, regardless of its form (extracts or dried plants), significantly improved the rearing indices. In piglets of groups III, IV and V a significant improvement of daily weight gains and feed efficiency was noted. Inulin showed hypolipidemic activity (lowered total cholesterol level) and stimulated piglet immune system manifested by elevated IgA and IgG concentrations. Irrespective of the inulin source, a lower mortality rate resulting from the improvement of animal health was noted in all experimental groups.

Hepatoprotective effect of plant drugs against hepatic tissue injury induced by non-steroidal anti-inflammatory drugs (NSAIDs) was assessed on Beagle dogs. The adverse effects of carprofen and robenacoxib on the hepatic tissue were evaluated on the basis of histopathological examination of liver sections. It was demonstrated that the use of NSAIDs with liquorice and composed plant remedy Pectosol¯ caused a reduction of hepatic adverse effects induced by the administration of NSAIDs. This fact indicates a hepatoprotective effect of the tested plant remedies during the treatment with NSAIDs. However, the results require further studies on a larger group of animals. Liquorice and Pectosol¯ reduce the hepatic side effects, which develop after the treatment with carprofen and, to a lesser extent, robenacoxib in young Beagles. Such studies allow to investigate the negative and positive effects of using robenacoxib and carprofen in dogs and, therefore, help to limit the NSAID-induced side effects on the liver in these animals.

The aim of the study was to determine the suitability of basic haematological, biochemical, and gasometric tests in checking the effectiveness of transfusion therapy in foals during isoerythrolysis. The number of red blood cells, haemoglobin, haematocrit, and partial pressure of carbon dioxide, oxygen, and blood pH was determined immediately before and several times after blood transfusion. The concentration of serum free bilirubin was also measured to confirm haemolysis. Fluids (0.9% NaCl, multielectrolytic fluid, 5% glucose) and antibiotics (penicillin, amikacin) were provided to the foals. The lowest values of haematological parameters were observed before transfusion. This was accompanied by decreased partial pressure of oxygen, low pH, and increased arterial carbon dioxide tension. Transfusion of whole blood led to a gradual normalisation of the haematological parameters, also accompanied by the normalisation of gasometric indicators (decrease in pCO2 and pO2 and pH increase). Monitoring of selected haematological and gasometric parameters allows to evaluate the efficacy of blood transfusion during treatment of haemolytic disease of foals.

The studies, conducted between 2010 and 2012, involved 102 dogs and 505 people from Lower Silesia (LS), 104 dogs and 578 people from the Legnica - Głogów Copper Mining Region (LGCMR), and 101 dogs and 897 people from the Upper Silesian Industrial Region (USIR). A significant positive correlation between blood lead concentration (BLC) in dogs and people living in the same environment was found. Moreover, the data revealed an increase in BLC in dogs and people with the progressive aging of the body. The highest average BLC in dogs and humans were reported in the LGCMR followed by USIR and LS.

The aim of this study was to define the effects of diet containing the same mineral content of mineral salt or amino acid chelate, and diet containing various levels of Cu amino acid chelate on liver histomorphometry in growing rats. Male Wistar rats were used in the 12th week experiment. The control group (n = 12) was fed standard diet, which provided Cu in an inorganic form at the level required for rats. The experimental animals were divided into four groups (each n = 12) depending on different levels (100%, 75%, 50%, 25% covered daily demand) of Cu supplementation in chelated form. Cu content was determined in the liver tissue and blood plasma. Immunohistochemical staining with caspase-3 antibody was performed. Microscopic assessment of the liver structure indicated that Cu supplementation did not change the liver architecture. However, histomorphometric analysis revealed a significant increase in the number of nuclei, total cell number, and multinucleated hepatocytes in rats supplemented with the organic form of Cu at the level of 25% compared with the control group. There was a considerable increase in the number of apoptotic cells and ballooning degeneration of hepatocytes, especially in groups supplemented with organic form of Cu covering the daily demand in 100% and 75%, in comparison to control group. Moreover, there was no Cu deposition in the liver and changes in Cu content in blood. Cu provided in the diet in organic form covering an amount of its minimum daily demand in 25% appears to be the least harmful with regard to the liver. It indicates that there is a need to establish the level of diet supplementation with Cu amino acid chelates.

The study was conducted on spermatozoa of selected farm and free-living animal species, isolated post mortem from the tail of the epididymis, and stained with silver nitrate - AgNO3. The material was collected from pigs, goats, wild boar, and European roe deer. Twenty morphologically normal spermatozoa randomly selected from each animal and well visible under the microscope, were analysed. The following measurements were considered: head length, width, perimeter and area, acrosome area, mid-piece length, tail length, and overall sperm length. AgNO3 staining differentiated the acrosomal (light hue) and distal (dark hue) part of the sperm head, and a light-hued mid-piece was visible within the sperm tail. Silver nitrate staining revealed species and variety-related differences, particularly in reference to the sperm head. Clear-cut differentiation within the head and tail area made it possible to perform detailed morphometric measurements of the spermatozoa.

The aim of the study was to evaluate the possibility of using heart rate (HR) as a metric parameter that can be used for the characterisation of behaviour of primitive horses and their relatives, related to reactions to the stress resulting from the contact with humans and adaptation to living in various conditions, including natural environment. This characterisation served the authors to expand the knowledge of such behaviour of primitive horses, and to assess the impact of the environmental and genetic factors. Studies were conducted in three populations of horses: two herds of Polish Konik and one herd of Biłgoraj horses. The studies were performed between 1993 and 2010. They concerned the behaviour of horses during grooming - breeding procedures (hooves clearing, body measurements) performed cyclically and the daily observations when HR was monitored continuously. HR results for the respective age categories, during particular grooming - breeding procedures and reserve observations indicate that Polish Konik horses, closely related to the primitive Tarpan breed, are genetically better adapted to living in conditions similar to the natural (reserve) than the Biłgoraj horses. They show less stress symptoms, which are evidenced by HR values noted during inhabiting the natural environment.