Volume 56 (2012): Issue 4 (December 2012)

Phylogenetic analysis of the genes determining influenza virus subtype - haemagglutinin (HA) and neuraminidase (NA), was performed. The results showed that the Polish H1N2 isolate (A/Swine/Poland15817/2011) was reassortant of human-like swine H1N1 and human-like swine H3N2 origin. The novel isolate was presented to have a close phylogenic relationship with one of the latest European isolates of H1N2 (A/SW/Gent/102/2007 and A/SW/Hungary/13509/2007). Our evolutionary analyses also suggested that the HA and NA genes evolved in a significantly higher rate of synonymous substitutions after they were introduced from human to swine and established the European H1N2 swine lineage.

The aim of the study was to estimate the current epidemiological situation concerning swine influenza (SI) in Poland. The study was based on an annual passive survey of 11,770 fatteners’ sera from 584 herds, taken at slaughterhouses within the last 30 months (from January 2010 till June 2012), as well as, an active monitoring conducted in 2011 and 2012, in 25 farms, using 388 sera taken from life pigs of different age/technological groups. The analysis of simultaneous circulation of different swine influenza virus (SIV) subtypes was taken into a deep consideration. The wide spread of SIV in Poland, including the occurrence of multiple SIV infections was demonstrated. In 2010 and 2011, the domination of H1N1 subtype and the most frequently co-circulation of H1N1 and H1N2 viruses was evidenced, while in the first 6 months of 2012, the co-circulation of H1N1 and H3N2 viruses was detected more often. Based on the obtained results, it can be stated that the epidemiological situation concerning SI in Poland is dynamic and similar to that observed in other European regions with high pigs’ density; however, the prevalence of antibodies and the occurrence of mixed SIV infections is lower than in Western Europe.

In the study, a 122 bp fragment of gag gene encoding immunodominant epitope on capsid protein of small ruminant lentiviruses (SRLVs) found in sheep was amplified by PCR and analysed by SSCP and sequencing. Out of 30 DNA samples, five showed different migration patterns, demonstrating the individual variations within gag sequences, which were confirmed afterwards by sequence analysis. In two samples nucleotide changes yielded amino acid substitutions highlighting the conservative nature of gag encoded immunoreactive epitope but also potencial insensitivity of a single-strain-based immunoassay.

The strains of adenoviruses were isolated from 356 birds with clinical form of Marek’s disease and coinfection with adenoviruses. A hexon gene fragment coding loop L1 of adenovirus strains was sequenced and obtained data were analysed with BLAST, Geneious 5.3, and MEGA5 software by comparison with nucleotide sequences of reference strains of fowl adenoviruses (FAdV-1 - FadV-12), two turkey adenoviruses, and two goose adenovirus strains. On this basis, serotypes of adenovirus strains were determined. Sequences of all adenovirus strains isolated from birds infected with Marek’s disease virus were classified into six serotypes representing four species. Mostly FAdV-7, FAdV2/11, and FAdV-8a serotypes were found.

The aim of the study was to determine the influence of simultaneous infection of chicken embryo fibroblasts (CEF) with different doses of adenovirus field strain serotype 7 (FAdV-7 JN-5/10j) and turkey herpesvirus strain FC126 (FC126 HVT) on replication of the herpesvirus in in vitro cultures. Three experiments were performed: simultaneous infection of CEF with adenovirus and HVT; inoculation of CEF culture with adenovirus, followed by infection with HVT after 24 h; and inoculation of CEF with HVT, followed by the infection with adenovirus 24 h later. In order to detect the presence of HVT and adenovirus strains in CEF culture, SORF 1 and hexon genes were determined, respectively. The infection with adenovirus lowered replication of FC126 HVT in chicken embryo fibroblast.

The aim of the study was to determine the prevalence of Mycoplasma bovis infection in clinical cases of bovine respiratory disease (BRD) in 2011. There were examined 841 serum samples and 41 nasal swabs and lungs of cattle from different farms of six regions of Poland (13 provinces). The obtained results indicated that the mean prevalence of M. bovis infection in Polish cattle population suffering from BRD was 64.3% and among them 8.4% of the animals were highly positive. On the other hand, in particular regions, it altered - 72.0% in east region, 52.7% in central region, 66.4% in south region, 64.1% in north-west region, 56.5% in north region, and 58.8% in south-west region. It should be added that within the presented regions, the highest values (82.4%) were reported in south region (Malopolskie province), whereas the lowest ones (42.9%) were in east region (Lubelskie province). However, field strains of M. bovis were isolated from the lungs of affected cattle only in the north-west region of Poland (Zachodniopomorskie province).

The aim of the study was to determine Mycoplasma mycoides subsp. mycoides small colony variant (MmmSC) and Mycoplasma agalactiae antibodies in ruminants from different provinces/regions of Poland. Eight hundred and ten bovine serum samples were examined for MmmSC antibodies by the use of competitive ELISA (c-ELISA) and complement fixation test (CFT). ELISA was also used for M. agalactiae antibody detection in 951 serum samples of sheep and goats. The first screening serological examination of MmmSC antibodies using c-ELISA revealed two (0.25%) positive and 135 (16.92%) doubtful results. The second examination revealed only 52 doubtful results, whereas the rest samples were negative. To compare, the final confirmatory examination by CFT gave 100% of seronegative results. The examination performed in small ruminants demonstrated only one doubtful result, which was finally defined as negative following the second ELISA, whereas the remaining samples were negative. To conclude, the present serological study showed the lack of infections in Polish domestic ruminant caused by two mycoplasmas.

The molecular epidemiological relationship among isolates of Salmonella enterica serovar (S.) Saintpaul, which was identified in animals, food, and humans in several EU countries, was investigated. Pulsed-field gel electrophoresis (PFGE) revealed a high degree of genetic diversity (82 XbaI PFGE profiles with 42.6% similarity) among 159 S. Saintpaul isolates from animals (n=91), food of animal origin (n=29), and humans (n=36) in 12 European countries during 2005 to 2009. Most frequent profiles (n=12) comprised almost 50% of the tested isolates. Profiles obtained in isolates from a single source within a particular geographical region or particular period of time were indistinguishable or closely related. Turkeys were confirmed as the major reservoir for S. Saintpaul. Indistinguishable PFGE profiles were identified in up to 19 isolates from turkey breeding and fattening flocks and food over the study period. Other animals, including food and pets, may also contribute to S. Saintpaul spread. International trade of animals and food, as well as travelling contributes to the spread of a specific clone to different geographical areas. Although control programmes in breeding turkey flocks, together with improved biosecurity, may interrupt the major transmission routes, it was concluded that S. Saintpaul will continue to represent a potential threat to human health.

The aim of this study was the evaluation of fluorescence polarisation assay (FPA) in the diagnosis of porcine brucellosis in comparison with Rose Bengal test (RBT), serum agglutination test (SAT), complement fixation test (CFT), 2-mercaptoethanol test, and ELISA. Eight hundred seventeen sera from pigs, including 612 sera from healthy animals, seven sera from Brucella suis bv2 culture positive animals, and 198 sera classified as false positive, originated from confirmatory investigations, were used. All sera from healthy animals, negative in RBT, SAT, CFT, and ELISA were also negative in FPA. All sera positive in serological examination, originated from Brucella infected animals, were also positive in FPA. Among sera classified as false positive almost half of the samples tested (49.49%) reacted positively in FPA. The examinations confirmed the usefulness of FPA in diagnosis of porcine brucellosis, but the method, like the other tests, does not allow resolving the problem of discrimination cross-reacting from specific antibodies.

The paper presents the results of bacteriological and molecular investigations on the presence of Y. enterocolitica O:9 in the head, mammary and genital lymph nodes, spleen, liver, and uterus samples originating from 58 cows slaughtered due to the positive results of serological examinations for brucellosis. All samples were cultured for Brucella and Y. enterocolitica and examined in multiplex PCR assay (mPCR), in order to identify the universal 16S rRNA Brucella sp. marker and amplify the perosamine synthetase (per) gene, specific for Y. enterocolitica O:9 only. Out of 58 examined animals, in 23 cases the presence of Y.enterocolitica was demonstrated. Typical Yersinia-suspected colonies were seen after 24-48 h incubation on Cefsulodin-Irgasan- Novobiocin (CIN) Yersinia selective solid medium. The mPCR analysis confirmed the presence of predicted amplicon of 312 bp, typical for Y. enterocolitica O:9 in 20 out of 58 lymph node samples tested and similarly, as in bacteriological examination, other samples were negative. The presence of the 16S rRNA gene of Brucella, generating the amplicon of 905 bp, was not observed in any of the samples tested.

The aim of the study was to perform a molecular investigations for the presence of pathogenicity genotypic markers of Y.enterocolitica O:9 isolated from cattle, in which initially positive serological reactions for brucellosis were observed. Almost all isolates were ail-, ystA- and myfA-positive (n=19). On the other hand, one isolate, which harboured plasmid encoding gene yadA was ail- , ystA- and myfA-negative. The plasmid encoding yadA marker was present in half of the isolates tested. None of the examined isolates was ystB-positive. The results of the investigations revealed that the Y. enterocolitica O:9 isolates, related to false positive serological results for brucellosis, may be also potentially pathogenic for humans, due to the presence of chromosomal and plasmidencoded molecular markers.

The objective of the study was to determine genotypic profiles of Bordetella bronchiseptica (Bbr) strains, based on the occurrence of genes encoding virulence factors, such as flagella (fla), dermonecrotoxin (dnt), and exogenous ferric siderophore receptor (bfrZ), using PCR. 209 tested Bbr strains were obtained from Polish swine herds with different health status (with progressive atrophic rhinitis - PAR, suspected for PAR, and unknown). In total, seven different Bbr genotypes were determined. In 39.2% of Bbr isolates all three genes were present. In 41.1% of the isolates only two genes were detected. The most common genotype dnt+bfrZ-fla+ was present in 60 (28.5%) Bbr strains, 65% of them were obtained from farms with PAR. Twenty five (12%) Bbr isolates were identified as dnt-bfrZ+fla+ genotype and, as above, they were more frequently isolated from clinical cases of disease (84%). Among 31 (14.8%) strains only fla gene was evident, and in nine (4.3%) only dnt gene was present. There were no Bbr strains with bfrZ gene only. These results confirm the heterogenicity among Bbr strains.

Blood serum samples collected from randomly selected groups of 32 pigs and 41 cows reared in farms belonging to the rural community “A” located in eastern Poland and exposed to the Vistula river floods, and serum samples from groups of 41 pigs and 40 cows from farms belonging to the rural community “B” located also in eastern Poland but not in the area exposed to floods, were examined by the microscopic agglutination test for the presence of antibodies against 18 Leptospira serovars. The percentage of serum samples presenting positive results with at least one serovar were higher in pigs and cows from community “A” comparing to community “B” (34.4% vs. 4.9% and 26.8% vs. 15.0%, respectively). In the case of pigs, the difference was statistically significant (P=0.0015). The reactions with 12 Leptospira serovars (Australis, Bataviae, Bratislava, Canicola, Hardjo, Hebdomadis, Icterohaemorrhagiae, Pomona, Poi, Cynopteri, Grippotyphosa, Celledoni,), belonging to four species (L. interrogans, L.borgpetersenii, L. kirschneri, L. weili) were found in the examined animals. In community “B”, six reactions with one serovar and two reactions with two serovars were noted whereas in community “A” - 19 reactions with one serovar, one reaction with two serovars and two reactions with six serovars were observed. The titres in animals reared in community “A” were significantly higher (up to 25,600) compared to community “B” (up to 200, P=0.0094). The obtained results suggest that the exposure to flooding may increase the infection rate in pigs and cows from afflicted areas to some extent.

Clostridium perfringens isolates were obtained from pigs of five porcine farms in Poland. The presence of C. perfringens was detected in 92% of faeces samples and its number ranged from 1.0 x 101 cfu/g to 1.2 x 107 cfu/g. All the isolates belonged to type A and 48.7% of them contained cpb2 gene. The qualitative assessment of toxin genes expression by type A subtype β2 isolates showed expression of cpa gene in 100% of strains and cpb2 gene in 71% of the analysed strains. The isolate from one-day-old piglets demonstrated also the expression of cpa and cpb2 genes.

The aim of the work was to find out what is the prevalence and burden of Campylobacter sp. in the poultry exudates, what are the predominant species, how diverse they are, and can poultry exudates be considered as the essential carriers of campylobacters posing a health risk of campylobacteriosis in Poland. Broilers’ and turkeys’ exudates, collected directly from the bulk containers at the day of delivery were subjected to pre-enrichment and direct plate counting according to ISO 10272 (1995) with presumptive C.jejuni strains subjected to confirmatory genetic identification by the nested PCR and RAPD typing. The results indicated that over 90% of the exudates tested carried Campylobacter sp., with their number being significantly higher in broilers’ (10³ to10⁵ cfu mL^-1) compared to turkeys’ (10² to 10⁴ cfu mL^-1) (P≤0.05), and essentially higher (P≤0.05) for the broiler parts’ “skin on” exudates. C.jejuni was the only species present in the turkeys’ and predominant in broilers’ parts’ exudates. RAPD-typing of the isolates revealed 14 distinct RAPD types with the majority of C. jejuni isolates considered to be closely related.

The investigations comprised 100 piglets of crossbreed Polish Landrace x Large White Polish breed. Faeces samples were collected on the 2^nd d of piglets’ life (control). On the 5^th d of life of the piglets, probiotic paste was applied and 7 d later, faecal samples were collected again. The material included 100 isolates of Campylobacter sp. obtained from healthy piglets. All isolates were assigned to the Campylobacter coli species. The occurrence of virulence genes was determined by the PCR method. Drugresistance of the obtained isolates was determined using diffusion tests and E-test strips. All isolates deriving from the control group piglets were found to contain the cadF gene responsible for adhesion, as well as, gene flaA influencing motility of the examined bacteria. In piglets fed diets supplemented with probiotics, the cadF gene occurred in 100% isolates and gene flaA - in 99% isolates. Campylobacter coli isolates obtained from piglets from the control group exhibited the highest resistance with respect to ciprofloxacin and enrofloxacin. The similar results were recorded in the case of isolates obtained after the probiotic application. The majority of the isolates generated α type haemolysis (91%-92%). No significant differences were recorded in the capability of generating haemolysis between isolates obtained before probiotic administration and the isolates obtained after the application of the experimental probiotic.

The aim of the study was to evaluate the effectiveness of Q fever treatment with oxytetracycline based on the level of Coxiella burnetii antibodies in the sera of infected goats and cows, and excretion of Coxiella burnetii in milk. The study was performed in naturally infected goats and cattle. Forty-six goat sera and 35 cows’ sera were investigated three times before, and twice after treatment with oxytetracycline. The percentage of seronegative goats after treatment (the third examination) was 86.96% while the percentage of seronegative cows after treatment was 52.77%. Moreover, the molecular studies (real-time PCR) of cheese from milk of these animals showed that the specific sequences of DNA for Coxiella burnetii were present despite treatment with oxytetracycline.

Blood morphology in dogs infected with intracellular parasites of the genera Babesia and Ehrlichia was examined. The parasites were detected in peripheral blood smears stained by the May-Grünwald-Giemsa method. During the parasitic invasion, a decrease in thrombocyte counts was observed. This was connected with forming platelet aggregates. It was noted that the number of giant platelets simultaneously increased.

The aims of this study were estimation of thyroxin status in dogs infected with B. canis, and determination of the association between azotaemia and concentrations of total thyroxin (TT4) and free thyroxin (FT4) in canine babesiosis. Concentrations of TT4 and FT4 were determined using an immunoassay in 23 dogs infected with B. canis (nine azotaemic dogs and 14 non-azotaemic dogs). Mann-Whitney U test was used to compare the concentrations of TT4 and FT4 in groups of azotaemic and non-azotaemic dogs. Correlations between azotaemic parameters (i.e. serum urea and creatinine) and TT4 and FT4 concentrations were calculated. The obtained results showed high prevalence of dogs infected with B. canis showing decreased TT4 and FT4 concentrations. No difference between concentrations of TT4 and FT4 in azotaemic and non-azotaemic dogs was demonstrated, but statistically significant correlations between the level of TT4 and FT4 and serum levels of urea and creatinine were shown.

The article describes the analysis of Toxoplasma gondii infection among people and animals from the selected rural households in the Lublin province. To identify the potential sources of infection, parasitological and/or PCR examinations of the samples of water, soil, cats’ faeces, and meat eaten by the household members were carried out. The study indicated high percentages of seropositive results among the household members (N=57) and domestic animals (N=94) - 61.4 % and 53.2%, respectively. In two households, the parasite was detected in drinking water. The study results seem to confirm the possibility of family character of toxoplasmosis, and point at the environmental conditions leading to the infection in rural environment.

The aim of the study was to estimate the effectiveness of intestinal scraping technique (IST) in the detection of Echinococcus multilocularis. The experimental assessment of the limit of detection and comparison with “gold standard” (sedimentation and counting technique - SCT) was also performed. Samples of fox small intestines experimentally enriched with known numbers of E. multilocularis tapeworms, were used. Twenty four samples containing 10, 30, 60, and 90 E. multilocularis tapeworms were prepared. Moreover, in order to compare IST with SCT, 127 intestines of foxes were examined using both methods. The limit of detection was estimated at 30 E. multilocularis tapeworms per sample of the intestine. Moreover, mean number of Echinococcus found by IST were several dozen times lower than the real content of these tapeworms in the samples (on average only 2 to 3.2% of worms were recovered). Among 127 intestinal samples examined with the use of two methods, eight samples (8.2%) were positive by SCT and only two (1.6%) when IST was used. A relatively high limit of detection estimated experimentally in the first part of the study, as well as, the results obtained in field investigations showed clearly that IST method could significantly decrease the reliability of the results of investigations, especially carried out in regions where a very low prevalence of E.multilocularis occurs or in countries, which want to demonstrate that they are free from this parasite.

Screening of the prevalence of Paenibacillus larvae spores in honey bee colonies in apiaries from 162 districts, belonging to nine provinces was carried out during 2009-2011. The honey samples were examined by the use of a culture method. Based on the number of CFUs grown on Columbia sheep blood agar medium, the level of infection and probability of American foulbrood outbreak was estimated. Altogether, 6,510 pooled honey samples from 32,550 bee colonies located in 2,294 apiaries were collected. P. larvae was identified in 45% of the surveyed apiaries. The widest distribution of P. larvae was found in the Małopolskie province. Culture-positive honey samples were obtained for 71% of the apiaries and in a half of them, the level of spores was high. In the Warmińsko-Mazurskie province, the presence of the bacterium was detected in 58% of the apiaries. In the remaining provinces, from 26% to 47% of the apiaries were contaminated with P. larvae spores

This article presents the essential stages leading to development of a modified method for Hypoderma bovis protein transfer, and estimation of the usefulness of semi-quantitative densitometric analysis of western-blottig results. The principal of the method was to incubate the gel obtained with the native electrophoresis method in buffer containing SDS prior to transfer of the separated proteins of L1 H. bovis onto nitrocellulose membrane. In two experiments, the authors estimated the efficacy of the new method and the possibility of using this method for semi-quantitative densitometric investigations.

Age-related changes in serum concentrations of C-reactive protein (CRP), haptoglobin (Hp), serum amyloid A (SAA), and pig major acute phase protein (pig-MAP) were investigated in healthy pigs from birth to slaughter under field conditions. Repeated blood samples were obtained from 60 pigs at ages of 1-19 weeks. Concentrations of acute phase proteins (APP) were measured with the use of commercial ELISA kits. Concentrations of all APP increased with age (P<0.05) and positive correlations were evidenced between their concentrations and the age of pigs. Great variations in CRP, Hp, and SAA concentrations were found, as can been seen from standard deviation values. The minimal individual variability was found in regard to pig-MAP. A significant increase in all APP was observed in pigs’ serum after weaning, constituting an important characteristic of this period. The elevation of APP after weaning may be associated with stress induced by mixing animals after weaning or changes in the pattern of feed administration. The peak in APP may be also caused by the initiation of synthesis of these proteins by piglets. Because a significant association between age and APP concentrations exists, further studies are needed to decide whether the age may influence the diagnostic value of APP as a marker of infection. Additionally, studies are needed to estimate whether the APP response in infection is age-dependent to any clinical importance degree.

This study describes preparation of test samples composed of freeze-dried strain of S. aureus and powdered milk as a matrix. In the first part of the study, the number of S. aureus cells freeze-dried in skim milk or horse serum were compared at two levels of contamination (10⁴ and 10⁵ cfu g^-1). The analysis of the samples was performed three times within a week. The preliminary results showed that the samples composed of S. aureus freeze-dried in horse serum were more stable and homogeneous than those prepared with skim milk. These results were further confirmed after analysing a higher number of such samples. Therefore, this procedure was then chosen for preparation of the samples for proficiency tests (PTs). Homogeneity and stability of these samples were checked according to ISO 13528. The results obtained showed that the samples met the criteria of stability and homogeneity required for PTs and were used in PT for enumeration of S. aureus in powdered milk.

The study was carried out on live bivalve molluscs available on Polish market. Microbiological tests were performed for the presence of Salmonella sp., Vibrio parahaemolyticus, spore-forming anaerobe bacteria, and coagulase-positive Staphylococcus sp., and for the enumeration of Escherichia coli. ELISA was used for the determination of marine biotoxins, paralytic shellfish poisoning (PSP), amnesic shellfish poisoning (ASP), and diarrhoeic shellfish poisoning (DSP). Microbiological examinations were performed according to ISO and Polish Standards. Salmonella sp. was not detected in any sample tested. Coagulase-positive staphylococci were identified in 9.0% of the samples. V. parahaemolyticus was isolated from 17.0% of mussels. Shellfish were highly contaminated by anaerobes, which were isolated from 68.0% of the samples. The number of E. coli ranged from <2.0 x 10¹ up to >1.8 x 10⁴ MPN/100 g. The majority of mussels were free from the marine biotoxins tested or contained them bellow the permitted level. The analysis of microbiological and toxicological status of raw bivalve molluscs available on Polish market indicates that they are generally safe for the consumers.

One hundred and ninety seven samples of molluscs representing different species were tested for the presence of antibacterial substances using a microbiological diffusion test - “4-plate” method. It was found that 58 samples (29.4%) were positive. The percentage of positive samples depended on species and varied from 0 (Ostrea edulis, Perna canaliculis, Cardiumedule, Myretrix lyrata, Mercenaria mercenaria) to 41.2 (Mytilus edulis) and 50.0 (Tapes semidecussatus and Ruditapesphillipinarium). The randomly performed confirmatory analyses using HPLC -MS/MS method did not show the presence of any known antibiotics.

Four different beef muscles (PM, SM, LTL, TB) were examined. These muscles were fried or grilled. Each muscle was observed at various internal temperatures: 60°C, 65°C, 70°C, and 75°C. At each temperature level, a measurement of colour was taken using a Minolta Chroma Meter CR-400 in a L*a*b* system. The values of the parameters L*, a*, and b* were measured on a cross-section sample (two areas) after cutting perpendicular to the fibres at the middle of its height. Area I was located at the central part of the steak with a diameter of 2 cm. Area II was located at a distance of 1 cm from the edge of the steak. From the results obtained, the difference in colour values between areas was noted. The results indicated that area I was lighter and redder in comparison to area II. The values of parameters L* and a* recorded in the first area were higher in comparison with the values from the second region, indicating that the sample in the area I was lighter and redder than in the area II at the outside of the sample. There was a large variation in the colour of the two areas. It was observed that for all sampled fried muscles, the highest difference in lightness (ΔL*) between the two test areas was obtained at a heat treatment temperature of T_i= 65°C inside the sample. With grilling the highest value for overall difference in colour between areas was from samples heated to a final temperature of T_i=60 °C.

The cytototoxic potential of metronidazole, tinidazole, ronidazole, and ornidazole, using human and rat hepatoma cell lines (HepG2 and FaO) in culture was assessed. The cells were treated with drugs for 24, 48 and 72 h at 37 °C in 5% CO₂ at concentrations of 0.1 to 200 μg/mL. Following the treatment period, the cells were assayed by four independent assays: MTT reduction, neutral red uptake (NRU), total protein content (TPC), and LDH leakage. The results suggest that nitroimidazoles are of low cytotoxic potential (EC₅₀ >200μg/mL). The exception was ronidazole, which demonstrated a distinct endpoint sensitivity related to the species. EC₅₀ (μg/mL) in human cells were: in MTT assay - 196±5.5 and 122±9.3 at 24 and 48 h, respectively, and in NRU assay - 150±1.25 at 72 h. Based on minimal toxic concentrations (EC₂₀) for ronidazole, determined by all methods used in HepG2 cells, it could be concluded that their sensitivity was as follows: MTT>NRU>LDH>TPC.

Procedure for determination of chromium in biological materials by Zeeman graphite furnace atomic absorption spectrometry method using a Perkin-Elmer spectrometer equipped with hollow-cathode lamp at 357.9 nm was developed. The samples of animal tissues, food, and feed were digested in muffle furnace at 450ºC. The ash was dissolved in 1 N hydrochloric acid and the final solution was diluted in 0.2% nitric acid. Magnesium nitrate (1%) was used as a matrix modifier. The method was validated in terms of basic analytical parameters. The mean recoveries of chromium was 84.4% for muscle, 79.0% for canned meat, and 80.2% for feed, and analytical detection limit was 0.003 μg/g. Certified reference materials were used for analytical quality assurance. The proposed analytical procedure is well adapted for monitoring chromium content in food and feedstuffs. Content of total chromium in the tested samples (animal muscles and liver) was low and was situated in the range of 0.031-0.101 mg/kg (muscles) and 0.047-0.052 mg/kg (liver).

Magnesium chloride was given via drinking water (500 mg Mg/L) to rats exposed intragastrically to cadmium (cadmium 109) at a dose corresponding to 10 mg/kg diet for 7, 14, 21, and 28 d. The results provided the evidence that magnesium decreased cadmium retention in the duodenum, kidneys, and liver after 7 d. Significant reduction in cadmium retention was also found on days 14 d and 21 in kidneys. No significant decreases in cadmium retention in the organs examined were noted after 28 d. The results suggest that beneficial action of magnesium on body cadmium accumulation ceased with time after the two metal administration.

A sensitive and reliable method has been developed and validated to determine residues of abamectin, doramectin, eprinomectin, ivermectin, and moxidectin in bovine milk. Isolation of the analytes from milk was performed with the use of liquidliquid extraction with acetonitrile in the presence of sodium chloride. The extract was defatted with hexane and cleaned up using solid phase extraction (C8 cartridge) after forming ion pairs with triethylamine. The analytes were derivatized with N, Ndimethylformamide, acetic acid anhydride, and N-methylimidazole (100°C, 90 min). The derivatives were determined by reverse phase liquid chromatography with fluorescence detection (excitation and emission wavelength 365 nm and 475 nm, respectively). Recoveries of the lactones from milk samples fortified at 10-30 μg kg^-1 ranged from 52% to 80% with intra-laboratory reproducibility (CV) of 12.7%-22.8%. The critical concentrations (decision limit, CCα and detection capability, CCβ) were in accordance with target limits. The method has been verified in the proficiency studies by EURL/CVL Berlin (all z-scores in the range of ±2). The method was transferred to routine laboratories, verified in inter-laboratory comparison and successfully applied in the National Residue Control Plan.

An in-house reference material of chloramphenicol (CAP) in pigs muscle was prepared from the chloramphenicol treated animals. The incurred muscle material was diluted by mixing with blank muscle sample. The concentration of CAP at the level 0.33 μg kg⁻¹ was reached. For the homogeneity study, 10 random samples were analysed and the results were interpreted by Cochran’s test and the sufficient homogeneity test. Additionally, the samples were tested for their stability according to the following scheme: 1, 7, 14, 28, 56, 84, and 112 d. It was confirmed that an appropriate homogeneity and stability of the produced in-house reference material was obtained.

The aim of the study was to evaluate the concentration of lead in blood of dogs from Polish urban polluted areas including the Lower Silesia Region (LSR; 102 dogs), KGHM Polish Copper Region (PCR; 102 dogs), and Upper Silesia Industrial Region (USIR; 102 dogs). Moreover, it was investigated whether age, height, gender, and weight influence blood lead concentrations in the dogs. The mean concentrations of lead in dogs from LSR, PCR, and USIR were 16.18 μg/L, 31.82 μg /L, and 32.53 μg /L, respectively. In the same age groups of dogs, the concentrations of lead were the smallest and significantly lower in LSR than those reported in PCR and USIR. Mean blood concentrations of lead demonstrated a decreasing tendency in regard to the height of the examined dogs - 30.00 μg /L (low dogs), 27.37 μg /L (medium dogs), and 25.12 μg /L (high dogs).These findings indicate that blood lead concentrations mainly depend on lead contamination of the dogs’ habitat. In all regions examined, lead concentrations significantly increased with the length of the dogs’ life. Height, weight, and gender had no significant effect on lead content.

Natural occurrence of thiouracil in bovine and swine urine in Poland was investigated. Under the national residue control programme, 537 urine samples were tested. In 77 samples (14.3%) thiouracil was detected above decision limit CCα (0.91 μg L^-1), including eight samples over the recommended concentration of 10 μg L^-1. Of the bovine urine samples, 95 and 99 percentiles have thiouracil concentration below 4.50 and 14.85 μg L^-1 ,and of porcine samples below 2.35 and 6.80 μg L^-1, respectively.

Histopathological examination of liver, kidney, spleen, pancreas, duodenum, jejunum, skeletal muscle, and bursa of Fabricius samples, collected from broiler chickens, laying hens, fattening pigs, and calves fed genetically modified corn MON 810 and soybean meal MON-40-3-2 (Roundup Ready, RR), was performed The examination showed no significant differences between the control animals fed diets containing no genetically modified feeds and animals fed genetically modified feeds. In some cases, congestion of parenchyma and focal lymphoid cell infiltrations were observed in all dietary groups, including controls, and therefore, it was assumed that the lesions were not associated with the feeding transgenic feeds.

The aim of the study was to show the variability of exterior type in Arabian horses bred in three leading stud farms in Poland. A total of 334 adult animals from the basic herd of stud farms in Bialka, Janow Podlaski, and Michalow were studied. Each horse underwent 26 exterior measurements. In the first stage, the method of exploration of multidimensional data - principal component analysis, was used, and subsequently, a stepwise canonical discriminant analysis. It was found that each stud farm breeds horses with a different metrical pattern. Horses bred in Bialka represent clearly different morphotype comparing to horses from the other stud farms. Individuals from Janow Podlaski are more similar to horses originating from Michalow but morphological variation also occurs between them. From among the 26 metric traits, 10 parameters have a meaningful discriminative power. These include in particular such parameters as the length of the metatarsus, pelvis, arm, and neck, and depth of the chest.

The electrocardiographic examination was performed in 33 training horses (2-16 years of age, 11 males and 22 females). Einthoven and precordial leads (I, II, III, aVR, aVL, aVF, CV₁, CV₂, CV₄) were used. The ECG was performed in resting horses and immediately after exercise (10 min walk, 15 min trot, 10 min canter) using a portable Schiller AT-1 3-channel electrocardiograph, with a paper speed of 25 mm sec^-1 and a sensitivity of 10 mm.mV^-1. The heart rate, wave amplitudes, and duration time were estimated manually. All horses presented a significant increase in heart rate after exercise (rest 43.83 ±10.33 vs. exercise 73.2 ±14.8). QT intervals were significantly shortened in most of the leads. In resting horses, all P waves in the lead I were positive and almost all II, III and CV₄ leads were positive. Simple negative P wave dominated in aVR and only simple negative T wave was found in the leads I. The biphasic shape was observed. After exercise, the amplitude of P and T waves rose, however, clear changes were not observed in wave polarisation and form. In the absence of specific racial characteristics of the electrocardiogram in the Polish Anglo- Arabians, electrocardiographic findings can be interpreted according to ECG standards adopted for horses.

The aim of the study was to compare densitometric and geometric parameters of the proximal phalanges in forelimbs of Warmblood and Coldblood horses, at 15%, 50%, and 85% of the bone proximal length. Peripheral quantitative computed tomography was used to analyse the structure of phalanges. The following parameters were determined: bone mineral content, volumetric bone mineral density, total bone area, trabecular area, cortical area, cortical thickness, periosteal circumference, endocortical circumference, and strength strain index. The most significant differences between pastern bones in thoracic limbs of the horses were visible in the proximal and distal metaphyseal parts, for instance. at 15% and 85% of the bone length, respectively. Statistical analysis showed that at 15% of the bone length, total bone area, trabecular area, periosteal circumference, and endocortical circumference were significantly higher in Warmblood horses. At the mid-shaft, volumetric bone mineral density and endocortical circumference were significantly higher in Coldblood horses. At 85% of the bone length, volumetric bone mineral density, total bone area, trabecular area, periosteal circumference, and endocortical circumference were significantly higher in Coldblood horses.

The aim of the study was to analyse the hair coat of the three dachshund varieties, i.e. shorthaired, longhaired, and wirehaired, with respect to its histological structure and content of selected chemical elements. The study was conducted on female dachshunds originating from individual breeders. The study included the assessment of the contribution of particular hair fractions, measurements of hair thickness and length, and also the analysis of histological structure of particular hair fractions. A great differentiation in the parameters and structure of particular kinds of hair from the examined dogs was demonstrated. Thickness and length of hair of particular varieties showed statistically significant differentiation, as well as percentage contribution of particular hair fractions. As regard the content of chemical elements in particular hair fractions and dachshund variety, it appeared that the elements forming hair keratin were the most abundant. The findings presented in the paper may be a useful tool for the recognition of dachshund varieties based on their hair coat structure. They may be also a valuable contribution to the expanding breed standards, which lack histological hair description.

The research was performed on 16 one-year-old domestic turkeys of Beltsville race and 16 domestic ducks of the Pekinese race, of both sexes. Standard histological technique was used and morphometric analysis was performed using histological samples. Ciliary ganglion morphometric analysis revealed the presence of two cell populations of ganglionic neurocytes: choroid and ciliary cells, clearly differing in diameter. Ciliary cells were predominant in turkeys, whereas choroid cells in ducks. The statistical analysis showed that the diameter and surface area of ganglionic neurocytes, the diameter and cross sectional area through the ciliary ganglion, as well as, the number of ganglionic cells on the cross-section through the ganglion were significantly larger (P<0.001) in turkeys than in ducks. The difference in the number of choroid and ciliary cells in domestic turkey and domestic duck is probably connected with eye accommodation, which seems to be greater in turkeys.

Clinically healthy Polish Holstein-Friesian calves were divided into two groups: a control group (12 individuals) and an experimental group (12 individuals). The experimental group calves received a mixture of 20% water herbal extracts prepared from Urtica dioica, Hb., Hypericum perforatum, Hb., Melissa officinalis, Fol., Chamomillae recutita Anth., Calendula officinalis, Flos, and Plantago lanceolata, Fol. The mixture was given from the 2^nd to 14^th week of their life. Control calves received no addition of herbal extracts to their diet. Red blood cell count, white blood cell count, haemoglobin concentration, haematocrit value, and mean corpuscular volume were determined in the 4^th, 9^th, and 14^th week of the experiment. The beneficial effect of the herbal mixture on the level of the examined haematological parameters was demonstrated in all tested weeks. In the calves from experimental group, a significantly higher red blood cell count, haemoglobin concentration, and corpuscular volume, and lower white blood cell count were found. There were no statistically significant differences in the haematocrit values between both groups.

The study aimed at defining the immunity risks connected with high iodine supplementation in ewes and their lambs. The feed ration for control group (C) of ewes (n=6) contained 3.1 mg of iodine per kilogram of dietary dry matter and for experimental group (E) of ewes (n=6) 5.1 mg of iodine per kilogram of dietary dry matter. The animals were fed the diets for 8 months. Parallelly with different iodine supplementation, some immunological parameters (percentage of γ-globulins, concentration of immunoglobulin G, and white blood cell count) were examined in ewes and their lambs. The difference in the average concentration of immunoglobulin G between the ewes of group C and E was not significant during the experiment. A significant decrease (P<0.01) in the concentration of immunoglobulin G from 19.6 mg/mL before the experiment to 9.8 mg/mL on day 60 of the experiment was demonstrated in the ewes of group E. The average percentage of γ-globulins in the whole period after parturition was lower in the ewes of group E than in group C. The concentration of immunoglobulin G and the percentage of γ-globulins in lambs from dams in group E was continually lower (P<0.01) for the entire period of 1-30 d and demonstrated immunological risks of the excessive intake of iodine. The obtained results document a dangerous influence of high iodine intake on immunity in the prenatal period in ewes and mainly in the postnatal period in their lambs.

The aim of the study was to evaluate the effect of high rape cake content and microbial phytase added to low-phosphorus diet on deposition of nutrients and energy in body of broiler chickens and quality of their bones. At the beginning, seven 1-day-old Ross 308 chickens were weighed, slaughtered, dried, and their bodies were analysed for gross energy and nutrient content. Next, 90 one-day-old chickens were randomly divided into three groups. The group I was fed control diet (RC), which contained 15% (starter) and 20% (grower) of rape cake of Lirajet cultivar with P (7.5 g kg^-1) and Ca (10 g.kg^-1). The group I I was fed diet denoted as RC LP, which contained less P and Ca (5.8 g.kg^-1 and 6.8 g.kg^-1, respectively) than the control diet. The diet for group III, denoted RC LP+ Phy, contained 5.8 g.kg^-1 of P and 6.8 g.kg^-1of Ca, and was supplemented with enzyme preparation containing 0.35g.kg^-1 of phytase. On the 21^st d of life, the chickens were weighed and six birds from each group were slaughtered, dried, and analysed. The rest of the chickens were fed grower mixtures to 49 d of age and then six chickens were slaughtered and the quality of their bones was evaluated. The reduction of P and Ca contents in diets and phytase application did not influence body weight and feed consumption, but significantly improved the deposition of total phosphorus in chicken body at 1-21 d of age. The highest content of total phosphorus was stated in the femur of the chickens fed diet with normal P and Ca contents. The application of phytase to low-phosphorus diet did not influence the quality of bones.

The study aimed at the determination of dynamic relationship between mineralisation processes and antioxidative/oxidative status during the development of osteopenia. One hundred and two healthy female Wistar rats at the age of 2 months and initial body weight of 200 g were used in the experiment. The rats were divided into control (CON, n=6), sham operated (SHO, n=48), and ovariectomised (OVX, n=48) groups. Animals from SHO (n=6) and OVX (n=6) groups were sacrificed every week during 8 weeks of the experiment in order to detect dynamic changes in examined parameters. The samples were collected weekly from day 7 to day 56. The femora were examined with the use of DXA (bone mineral density) and pQCT (area, mineral content, volumetric density of trabecular and cortical part of distal femora). The pQCT scans were performed 5 mm from distal end of the tibia. The determination of activity of glutathione peroxidase (GSH-Px) and superoxide dismutase (SOD) in haemolysates of erythrocytes were performed spectrophotometrically. Obtained data showed wave like changes in both enzyme activities and bone parameters and indicated the importance of the 2^nd-3^rd and 5^th-6^th week after surgery as a key moment for bone metabolism and activity of enzymatic antioxidative defence during the development of osteopenia induced by bilateral ovariectomy. The obtained results proved that alterations in activity of GSH-Px and SOD, and pQCT ahead the changes registered by DXA by 7 d.

The aim of the study was to assess the prepubic minilaparotomy approach in the surgical treatment of prostatic disorders in dogs. The technique of a limited approach to the prostate was developed on the fresh cadavers of 14 intact adult male dogs, diversified in regard to size and body weight. The abdominal cavity was opened in midline, from the right-sided prepubic parapenile incision. The cut was limited to 7 cm. Afterwards, it was expanded by self-retaining retractors: Gelpi, Weislander, Finnochetto Baby, and Adson. Finnochetto Baby and Gelpi retractors used in pairs enabled an adequate exposition of the prostate in small dogs. The Finnochetto Baby retractor enabled an adequate exposition of the prostate in medium dogs and the Adson retractor - in large and giant dogs. The Weislander and Gelpi retractors used individually did not allow for sufficient access to the prostate in any group of the cadavers. Prepubic minilaparotomy can be a useful surgical approach in the treatment of prostate disorders in dogs. It can be used especially for minor surgical procedures, e.g. sampling of large tissue specimens for histopathological examination, partial prostatectomy, or treatment of intraparenchymal cavitary lesions.

Morphological changes developed in rat bucal mucosa, parotid glands, and lymphoid nodes following 6 weeks exposure to dental acetal resin implants were compared to metal/titanium, acrylic implants, and sham-operated rats. Morphological evaluation of bucal mucosa of rats exposed to acetal resin showed an increased accumulation of lymphoid and macrophage-like cells compared to sham-operated rats. Eosinophils were found in bucal mucosa and in parotid glands of rats exposed to acetal resin, although this change did not differ significantly from sham-operated rats. Morphological findings with respect to granular reaction and proliferation were similar in acetal resin exposed and sham-operated rats. Acetal resin implants may induce inflammatory and potentially allergic reactions at the site of implantation.

The aim of the study was to compare erupted permanent bovine teeth of two types involved in the process of chewing in respect to the localisation and histomorphometric characteristics of enamel tufts. The research material comprised 240 fully erupted premolars and molars from the maxillae and mandibles of 27 heads of cattle from the Polish Black-and-White breed. Overall, 1,986 specimens of bovine teeth were analysed using a Nikon Eclipse E600 microscope with Micro Image v4.0 software. Enamel tufts were relatively numerous in the enamel of smooth surfaces - on average from 5.6 to 6.4 per cross-section of tooth crown. The average length of the enamel tufts expressed by means of a median was smaller for premolars (89.3 μm) than for molars (123.9 μm). The analysis of the value of Spearman's rank correlation coefficient revealed that no relationship existed between the length and width of tufts in molars (r_s =0.26), and a low-level relationship was noted (r_s =0.45) in premolars. On the basis of the Mann-Whitney U test for the metric measurements, no significant differences were discovered for the width of enamel tufts (P=0.427), but significant differences were observed for the length of enamel tufts (P=0.032). The observed differences in the histological structure of the enamel of posterior bovine teeth in comparison to human teeth indicate that a certain degree of caution ought to be considered when using bovine teeth as a substitute for human teeth in in vitro trials.