Volume 62 (2018): Issue 1 (March 2018)

Classical swine fever (CSF) has caused severe economic losses in pig production in many countries. Recent CSF outbreaks in China are mainly associated with sub-genotype 2.1 of CSF virus (CSFV). Although there is abundant information regarding 2.1 isolates, few data are available on whole-genome analysis.

The biological and genome characteristics of three recently emerged Chinese CSFV isolates, i.e. SD2014-1, SD2014-2, and SD2014-3, were fully analysed.

Sequence analysis showed that the isolates shared 83.4%–95.0% nucleotide identity with eight other CSFV isolates. In addition, the 5′ untranslated region (5′UTR) and the non-structural (NS) proteins NS3, NS4A, and NS4B were more conserved than other regions of the genome. Phylogenetic analysis based on the complete genome sequences or full-length structural protein E2 gene sequences revealed that the three isolates belonged to sub-genotype 2.1b. In addition, several unique molecular characteristics of the 5′UTR, 3′UTR, and E2 were identified.

The genomic variations of the three isolates will support further analysis of virulence determinants and the evolutionary trend of CSFV.

Dengue is one of the major emerging viral diseases in the world, with dramatic increases in reported cases in the last few decades and annual worldwide occurrence of approximately 390 million infections. It is a highly important mosquito-vectored disease and is a problem in tropical and subtropical areas of the world. The major aim of this study was to clone and express the dengue NS3 gene, in service to its therapeutic importance for the development of stable cell lines.

Blood samples from dengue fever (DF) patients were collected and subjected to PCR amplification of the NS3 gene of dengue virus serotype-2 (DENV-2). The NS3 gene was amplified using gene specific primers and cloned in the TA cloning vectors.

The gene was successfully expressed in mammalian expression vector pcDNA3.1. The current finding was different from a previously reported DENV-2 strain replicon constructed in different cells, in which the whole genetic material of the virus was used instead of an active protease gene, and which gave a low yield of replicon expressing cells.

Recombinant NS3 could be used to produce an antibody that is possibly helpful for developing a single step diagnostic assay to detect the dengue virus NS3 antigen in sera of dengue patients.

This study consisted in histopathological and immunohistochemical examinations of the central nervous system of 15 sheep suspected of infection with Coenurus cerebralis. The sheep displayed compulsive circling and were submitted for necropsy in 2012–2016.

Species identification was made on the basis of the PCR analysis and parasitological examination of the cysts.

Coenurus cerebralis cysts were detected only in the cerebral tissue of 13 sheep and in the cerebral and cerebellar tissues of 2 animals. Out of the 33 parasite cysts, most (21.21%) were located in the right and left frontal lobes of the cerebrum. The largest cyst measured 6 × 5 cm and the smallest cyst was 2 × 2 cm in size. The highest and lowest numbers of scolices were 55 and 21, and the number of rostellar hooks ranged between 22 and 30. Histopathological examination revealed the presence of typical parasitic granulomatous inflammatory foci. Immunohistochemical staining showed that most common in the periphery of the parasite cysts were, in descending order by cell number, GFAP, CD163, CD3, and CD79α-positive cells.

The study confirms the role of cellular defence mechanisms in the pathogenesis of Coenurus cerebralis infection in sheep.

Phagocytic activity and oxygen metabolism of peripheral blood granulocytes from rabbits with experimental trichophytosis were assessed by flow cytometry.

Virulent species of T. mentagrophytes var. granulosum (Tm-K) isolated from rabbits with natural trichophytosis was used for experimental infection. The phagocytic activity of granulocytes was measured in whole blood by flow cytometry using the commercial Phagotest kit. Oxidative burst was measured in whole blood by flow cytometry using the commercial Bursttest kit.

It was found that rabbits were susceptible to infection with Trichophyton mentagrophytes under experimental conditions. The analysis of the phagocytic activity indices and oxygen metabolism of granulocytes in peripheral blood of infected rabbits showed that changes of the indices were connected with the progression and regression of the disease. A significant decrease in phagocytic activity and oxygen metabolism was observed during development of fungal lesions and it remained similar throughout the progress of the disease. The highest means of the percentage of activated and ingesting phagocytes and a significant increase in the mean fluorescence intensity (representing the number of ingested bacteria) were observed during spontaneous recovery. Therefore, the decrease or increase in the indices of phagocytic activity and oxygen metabolism of granulocytes from rabbits experimentally infected with T. mentagrophytes is somehow related to the progress of infection and suppressive activity of the fungus, whose elimination during recovery caused significant increases in investigated indices of non-specific cellular immunity.

The results of the present investigation confirm that the mechanism of oxygen-dependent killing is crucial in infections caused by T. mentagrophytes.

One aim of the study was to evaluate the impact when added to feed of the two potentially probiotic strains of lactic acid bacteria (LAB) Lactobacillus plantarum K KKP 593/p and Lactobacillus rhamnosus KKP 825 on production performance, health, and the composition of gut microbiota. The complementary aim was to assess the safety of these strains in broiler rearing.

A total of 500 one-day-old Ross 308 chicks were divided into four groups. The experimental factor was the admixture of bacterial preparation to the feed at different doses: the recommended maximum dose, a dose ten times higher, the recommended minimum dose, and a zero dose for the control group not receiving bacteria.

Addition of bacteria to the diets did not have a significant effect on the final body weight, final body weight gain, nor total feed intake or feed conversion. However, lactic acid bacteria had a positive effect on chicken health. Mortality among chickens fed with LAB was reduced. Moreover, LAB feeding inhibited the growth of Salmonella spp. and Clostridium perfringens in the intestines. There were no significant differences in chicken performance by dose of bacteria in the feed. The group dosed with LAB ten times higher than the recommended maximum did not demonstrate changes in biochemical or haematological parameters of blood compared to the remaining groups.

Feeding chicken broilers with two potentially probiotic LAB strains is safe and impacts animal health positively.

In recent years, there has been a great interest in biogenic amines such histamine, as they are associated with the quality and safety of some kinds of fermented foods. The aim of this study was to evaluate the effect of temperature and storage time on the content of histamine in cheeses.

Samples of mould and hard cheeses were examined with RP-HPLC with an organic-aqueous mobile phase containing acidic buffer and chaotropic salt. The samples were stored either at 22 ± 2°C for 42 days (mould and hard cheeses) or at 4 ± 2°C for 112 days (mould cheeses) and 133 days (hard cheeses).

The mean total histamine content in cheeses stored at 22°C was higher than the content in those stored at 4°C, with the highest concentrations found in Gorgonzola Piccante cheese (730.47 mg/kg). Histamine concentration in some types of cheeses exceeded the toxic threshold dose, indicating that after long or inadequately cool storage they may not be safe for consumers.

To protect cheeses from contamination with histamine-producing bacteria and to safeguard consumers from poisoning, factors conducive to this amine’s formation should be minimised during cheese processing. Suitable temperature and time during storage of cheeses are recommended to avoid the intoxication. Monitoring of this toxin in food is necessary to ensure safety of consumers.

Reports that the presence of persistent organic pollutants in fat may affect fatty acid metabolism prompted this research aiming to study the relationship between the contents of γ-HCH and DDT, DDE, DDD, and ΣDDT, and fatty acid composition of milk fat.

The material consisted of 50 samples of cow and mare milk, collected in 2015. Ludwicki’s and the Röse-Gottlieb and IDF Standard methods were used to prepare the samples. Statistical analyses were conducted using Statistica 12.0.

There was a negative correlation between the content of γ-HCH and C16:1, C17:1, C18:1c9, C18:1c9c12, and ΣMUFA in cow milk fat and C13:0, C14:0, and C10:1 in mare milk fat. A positive correlation was observed between γ-HCH and C6:0 to C12:0, C14:0, C18:1t16, and ΣSFA in cow milk fat, and between this compound and C14:0iso, C16:1, C17:1, C18:1c9,11, and ΣMUFA in mare milk fat. A negative correlation between the contents of ΣDDT and C16:1, C17:1, C18:1c9,11,13 and ΣMUFA in cow milk fat and C16:0iso, C17:0, and C18:3 in mare milk fat was noted. A positive correlation was found between the contents of ΣDDT and saturated and polyunsaturated fatty acids and ΣSFA and ΣPUFA in cow milk fat, and C18:2c9c12 in mare milk fat.

The correlation between the content of selected organochlorine compounds and the composition of fatty acids in cow and mare milk fat indicates the strong influence of these environmental pollutants on the nutritional value of milk fat.

The experiment evaluated the effects of intravenous administration of polymyxin B on experimental endotoxaemia in sheep.

Twenty clinically healthy fat-tailed sheep were randomly divided into: a group treated with 6,000 U/kg of polymyxin B, a group at 12,000 U/kg, and positive and negative controls. Endotoxaemia was induced by intravenous administration of lipopolysaccharide (LPS) from E. coli serotype O55:B5 at 0.5 μg/kg. polymyxin was infused intravenously along with 2.5 L of isotonic intravenous fluids at 20 mL/kg/h. The positive control group received LPS and 2.5 L of isotonic fluids, the negatives receiving just 2.5 L of isotonic fluids. Clinical signs were evaluated before and at 1.5, 3, 4.5, 6, 24, and 48 h after LPS administration. Blood was also sampled at the denoted hours and serum haptoglobin, tumour necrosis factor-α (TNF-α), and plasma lactate concentrations were assayed.

The serum concentration of TNF-α in the positive control group increased significantly up to 48 h after LPS administration. The concentration of TNF-α was significantly different from those of the polymyxin B and positive control groups from 3 to 48 h; also, the concentrations of haptoglobin at different times in the polymyxin groups were lower than those of the positive control group and were significant at hours 3 to 48 (P < 0.05). Following the LPS administration, haptoglobin and TNF-α concentrations changed without significant difference between the two polymyxin B groups.

Polymyxin B (6,000 U/kg) restrained blood lactate concentrations. Furthermore, it significantly improved the clinical signs in endotoxaemic animals, including rectal temperature and heart and respiratory rates. Polymyxin B may be an antiendotoxic in fat-tailed sheep.

Veterinarians use flumequine (FLU) widely but its toxicological effects are still unclear.

FLU doses of 53, 200, or 750 mg/kg were administered orally for six weeks to pubertal male rats for evaluation of their toxicity.

Weight gain was poorer after seven days of exposure to FLU 750, but relative weights of the brain, adrenal and thyroid glands, and testes were notably higher. Haematological and lipid profile parameters, cardiac markers, and inorganic phosphate significantly increased in the FLU 750 group. Blood glucose, oestradiol and serum concentrations of immunoglobulins G (IgG) and E (IgE) significantly decreased after treatment. The levels of interleukins 10 (IL-10) and 6 (IL-6) fell significantly in the FLU 200 and FLU 750 groups. Cytochrome P450, family 1, subfamily A, polypeptide 1 (CYP1A1) and cyclooxygenase-2 (Cox-2) expression amplified after treatment. Serum levels of free triiodothyronine (fT3) and free thyroxine (fT4) reduced in the FLU 200 and FLU 750 groups without changes in total T3 or T4 level. All doses of FLU significantly depressed concentrations of thyroid-stimulating hormone (TSH) and testosterone. Histopathology of thyroid glands from rats treated with FLU 750 showed degeneration and depletion of thyroid follicular epithelial cells. Expression of 8-hydroxydeoxyguanosine (8-OHdG) was increased in a dose-dependent manner in the brain, but decreased in the testes. Expression of CYP1A1 increased in the adrenal and pituitary glands.

The results of this study suggest that the toxicity of FLU in rats is an effect of its disruptive influence on the pituitary-thyroid hormonal system and on the dysfunction of the immune system.

Nerium oleander is a plant of the Apocynaceae family toxic to humans, animals, and insects. This study was performed to determine the cardiac and neurotoxicity of the plant extract by oral administration in Wistar rats and Balb/c mice and to compare the susceptibility of these animal models to oleander toxicity.

Four groups of eight mice and eight rats received N. oleander extract orally while a fifth group was the control. Serum concentrations of the biochemical toxicity indicators, namely troponin and creatine kinase (CK), were determined and histopathological evaluation of the heart and brain was performed.

In mice, CK and troponin concentrations were respectively 1.5 and 7 times higher than in the control group (P < 0.05), while in rats, they were 6–7 and 11 times higher. Hyperaemia, haemorrhage, and myofibrolysis, without infiltration of inflammatory cells, were observed in the heart. In the brain the authors observed hyperaemia associated with perivascular and perineuronal oedema, and in higher-dosed rats multifocal haemorrhagic and liquefactive necrotic lesions.

Oleander can affect serum levels of CK and troponin due to nervous and cardiac injuries. Rats showed more severe changes in the biochemical indicators and histopathological lesions than mice. Therefore, biochemical and pathological findings indicate that Wistar rats are more susceptible to the cardiac toxicity and neurotoxicity effects of N. oleander poisoning than Balb/c mice.

The efficiency of five natural antioxidants (curcumin, cranberry, pomegranate, grape seed extract (GSE), and açai berry) in reducing lipid oxidation in dog food was compared to that of the synthetic antioxidant butylated hydroxyanisole (BHA).

In two different experiments content parameters were measured after 12 days of storage at 55°C. In experiment one, the natural antioxidants were added at 0.2% and BHA at 0.02% of the food (DM basis), and samples were analysed for thiobarbituric acid-reactive substances (TBARS). In experiment two, the effects of GSE and curcumin at two admixture proportions (0.1% and 0.2% of food DM) on omega-3 fatty acid (FA) content were evaluated.

TBARS values were lower than the control (P < 0.01) for curcumin, cranberry, pomegranate, and GSE but not for the açai berry (P > 0.05). By day 12, although there were no significant differences (P > 0.05) between the two curcumin treatments, they preserved higher concentrations of eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) (P < 0.05) than the BHA and control treatments. The addition of GSE or BHA to dog food held (P < 0.05) the concentrations of EPA higher than the control. The concentrations of EPA and DHA for the 0.2% GSE treatment were greater (P < 0.05) than the 0.1% GSE treatment. Grape seed extract at 0.2% lost less (P < 0.05) EPA concentration than BHA.

The present results showed that, except for açai berry, the tested natural antioxidants could be used as a substitute for BHA in dog food.

Serological diagnosis of brucellosis is still a great challenge due to the infeasibility of discriminating infected animals from vaccinated ones, so it is necessary to search for diagnostic biomarkers for differential diagnosis of brucellosis.

Cell division cycle 42 (Cdc42) from sheep (Ovis aries) (OaCdc42) was cloned by rapid amplification of cDNA ends (RACE), and then tissue distribution and differential expression levels of OaCdc42 mRNA between infected and vaccinated sheep were analysed by RT-qPCR.

The full-length cDNA of OaCdc42 was 1,609 bp containing an open reading frame (ORF) of 576 bp. OaCdc42 mRNAs were detected in the heart, liver, spleen, lung, kidneys, rumen, small intestine, skeletal muscles, and buffy coat, and the highest expression was detected in the small intestine. Compared to the control, the levels of OaCdc42 mRNA from sheep infected with Brucella melitensis or sheep vaccinated with Brucella suis S2 was significantly different (P < 0.01) after 40 and 30 days post-inoculation, respectively. However, the expression of OaCdc42 mRNA was significantly different between vaccinated and infected sheep (P < 0.05 or P < 0.01) on days: 14, 30, and 60 post-inoculation, whereas no significant difference (P > 0.05) was noted 40 days post-inoculation. Moreover, the expression of OaCdc42 from both infected and vaccinated sheep showed irregularity.

OaCdc42 is not a good potential diagnostic biomarker for differential diagnosis of brucellosis in sheep.

Xylazine, a type of α₂-adrenoceptors, is a commonly used drug in veterinary medicine. Xylazine-induced changes in the content of amino acid neurotransmitters – glycine (Gly) and aspartic acid (Asp), in different brain regions and neurons were studied.

Wistar rats were administered 50 mg/kg or 70 mg/kg of xylazine by intraperitoneal injection. In addition, in vitro experiments were conducted, in which neurons were treated with 15 μg/mL, 25 μg/mL, 35μg/mL, and 45 μg/mL of xylazine. Test methods were based on the enzyme-linked immunosorbent assays (ELISA).

During anaesthesia, Asp levels in each brain area were significantly lower compared to the control group. Except for the cerebrum, levels of Gly in other brain areas were significantly increased during the anaesthesia period. In vitro, xylazine-related neuron secretion of Gly increased significantly compared to the control group at 60 min and 90 min. Moreover, xylazine caused a significant decrease in the levels of Asp secreted by neurons at 20 min, but gradually returned to the level of the control group.

The data showed that during anaesthesia the overall levels of Asp decreased and overall levels of Gly increased. In addition, the inhibitory effect of xylazine on Asp and the promotion of Gly were dose-dependent. Our data showed that different effects of xylazine on excitatory and inhibitory neurotransmitters provided a theoretical basis for the mechanism of xylazine activity in clinical anaesthesia.

Electrical cardioversion is a therapeutic procedure used to convert various types of arrhythmias back to sinus rhythm. It is used to restore the sinus rhythm in dogs with atrial fibrillation. The effect of the electrical energy used during cardioversion on red blood cells (RBC) is not fully understood. Studies on humans reported lysis of RBC following electrical cardioversion. Similar studies have not been carried out on dogs. The aim of the study was to assess the effect of electrical cardioversion on chosen RBC parameters.

The study was carried out on 14 large and giant breed dogs weighing from 30 to 84 kg with lone atrial fibrillation (lone AF). Electrical cardioversion was carried out under general anaesthesia by biphasic shock with 70–360 J of energy. Blood was collected at T0 – during atrial fibrillation, prior to cardioversion, and at T1 – 30 min after electrical cardioversion. Complete blood counts as well as total and direct bilirubin concentrations were evaluated. A maximum output of 360 J was used.

In all cases, electrical cardioversion was effective, and no significant changes in the number of RBC and RBC indices were noted. Similarly, there were no statistically significant differences in the levels of total and direct bilirubin.

Electrical cardioversion in dogs led neither to statistically nor clinically significant RBC lysis.