Volume 62 (2018): Issue 3 (September 2018)

Simvastatin is a substance which is commonly used as a medicine to reduce cholesterol level. Unfortunately, it shows numerous side effects. Simvastatin affects various internal organs, and among other detriments to health may cause persistent muscle weakness, osteolytic processes, headaches, and rashes. Until now knowledge of the influence of simvastatin on bone marrow cells has been rather scant and fragmentary.

During this experiment the numbers of all types of cells in the leukocytic system of porcine bone marrow were evaluated after 28 and 56 days of oral administration of simvastatin at a dose of 40 mg/day/animal.

Simvastatin caused an increase in the number of all types of cells in the leukocytic system, and the most visible fluctuations concerned promyelocytes.

Observations obtained during the present study indicated that the results of the action of simvastatin on porcine bone marrow differ from those observed in other mammal species, including human. This may be due to various metabolic pathways within the bone marrow in the particular species, but the exact mechanisms of these actions are unknown at the present time.

Mycoplasma bovis is known as a causative agent of many disorders in cattle. In Europe, there is still a lack of commercial vaccines against M. bovis infection. Acute phase response (APR) is a non-specific host reaction to infection, most seen in changes in production of acute phase proteins. The aim of this study was to analyse APR in calves administered with an experimental M. bovis vaccine.

Twelve healthy female calves were divided into two equal groups: experimental and control. The experimental vaccine containing the field M. bovis strain and two adjuvants such as saponin and lysozyme dimer was subcutaneously administered to the experimental group. Phosphate buffered saline was taken as the placebo and given to the control group by the same route as the vaccine. Blood samples were collected prior to the study (day 0), then daily up to day 7, and then each seven days until day 84 post vaccination. The concentrations of serum amyloid A (SAA), haptoglobin (Hp), interferon-γ (IFN-γ), and inteleukin-4 (IL-4) were determined using commercial ELISA kits.

Following the vaccination, a significant increase in SAA, Hp, and IFN-γ concentrations was observed when compared to the unvaccinated calves, whereas the IL-4 concentration was not detectable.

The experimental saponin-based M. bovis vaccine containing lysozyme dimer adjuvant visibly stimulated the APR in the calves, and some specific cytokines (Th1-dependent) directly involved in this response.

Lyme borreliosis/Lyme disease is caused by Borrelia burgdorferi and is one of the most common vector-borne diseases transmitted by ticks.

A total of 136 Ixodes ricinus ticks, collected in the Ternopil (Ukraine) region, including 126 adults (70 females and 56 males), and 10 nymphs were examined. The identification of the species and their developmental form was based on morphological characteristics.

PCR with B5S-Bor and 23S-Bor primers resulted in Borrelia burgdorferi sensu lato DNA amplification among six ticks (4.4%). The detailed analysis based on the DNA sequencing showed the presence of DNA of Borrelia afzelii in four samples; the remaining two represented Borrelia burgdorferi sensu lato complex, although their genospecies were not determined. The research confirmed the dominance of Borrelia afzelii genospecies in the ticks from Ukraine.

It seems reasonable to undertake similar research in ticks from other regions of Ukraine. Knowledge in this field can be useful for public health and planning the prevention of tick-borne diseases.

The aim of this study was examination of honey samples collected from apiaries situated in all Polish provinces for occurrence of Clostridium spp., especially C. perfringens.

The study was carried out on 240 honey samples (15 samples/province). Estimation of Clostridium titre, its cultures and C. perfringens isolate characterisation were performed according to the standard PN-R-64791:1994. A multiplex PCR method for detection of genes coding cpa (α toxin), cpb (β), cpb2 (β2), etx (ε), iap (ι), and cpe (enterotoxin) toxins was used.

Clostridium spp. was noticed in 56% (136/240) of samples, and its titres ranged between 0.1 g and 0.001 g. Clostridium perfringens occurrence was evidenced in 27.5% (66/240) of samples. All isolates were classified to toxinotype A.

Evidence of a high number of positive samples with occurrence of Clostridium spp. indicates a potential risk to consumers’ health. The infective number of Clostridium spp. is unknown; however, the obtained results have shown that a risk assessment on the entire honey harvesting process should be made in order to ensure microbiological safety. Moreover, a detailed study should be undertaken on the antibiotic resistance of C. perfringens isolates from honey samples.

Genes related to iron metabolism play an important role in inflammatory response. The objective of this study was to investigate the role of ferritin, transferrin receptors 1a and 1b, and transferrin genes in the response to blood parasite infection in common carp (Cyprinuscarpio L.).

Two genetically distinct carp groups were used: R3 carp, which are established as being sensitive to parasitic infection, and SA carp (Cyprinus carpio haematopterus) of wild origin. An established challenge model with Trypanoplasma borreli was applied. Challenged carp were sampled to determine their expression levels of transferrin receptors 1a and 1b, ferritin, and transferrin mRNA. Mortality and serum iron concentration were also measured.

The study revealed contrasting differences in the expression profiles of all key iron regulatory genes except the transferrin gene. In the case of other parameters, significant differences were also observed.

Our results demonstrate that the level of parasitic infection depends on the blood iron status. This parameter was related to the origin of the fish.

Toxocara canis and Toxocara cati are roundworms of dogs and cats. The purpose of this study was to investigate the infection caused by these ascarids in cats and dogs, using microscopic and molecular analysis methods.

Adult ascarids were gathered from the faeces of dogs and cats in Van province, in 2015–2016. Existing keys and PCR sequencing of the ITS-2 fragment were used to identify the morphological features of the parasite species.

It was observed that out of 20 adult ascarids, 17 and 3 were found to be Toxocara canis and Toxocara cati, respectively. The ITS-2 gene region was amplified by PCR to perform molecular analysis. Genotyping indicated that the dogs and cats were infected with T. canis and T. cati, respectively, and none had Toxascaris leonina.

To the best of our knowledge, this is the first report on the molecular characteristics of adult ascaridoid nematodes from cats and dogs in Turkey. The molecular approaches established in this study enable molecular identification and genetic structure studies of the ascaridoids.

Amitraz is a formamide exhibiting both acaricidal and insecticidal activity and is frequently used by beekeepers to protect honeybee colonies against Varroa destructor mites. The aim of this apiary trial was to evaluate the impact of honeybee colony fumigation with amitraz on the level of contamination of honey stored in combs.

Experimental colonies were fumigated four times every four days with one tablet of Apiwarol per treatment. Honey was sampled from combs of brood chambers and combs of supers one day after each amitraz application and from harvested honey. Amitraz marker residues (as a total of amitraz and metabolites containing parts of molecules with properties specific to the 2,4-DMA group, expressed as amitraz) were evaluated in honey.

All analysed samples were contaminated with amitraz metabolites. 2,4-DMA and DMPF were the most frequently determined compounds. The average concentration of amitraz marker residue in honey from groups where a smouldering tablet was located directly in beehives was significantly higher than that of residue in honey from groups with indirect smoke generation. No significant effect on the honey contamination deriving from the place where it was exposed to smoke (combs of brood chambers and supers) was noted. Amitraz marker residues exceeded the MRL in 10% of honey samples from combs.

Fumigation of beehives with amitraz results in contamination of honey stored in combs.

Salicylic acid is a derivative of benzoic acid and occurs in nature. The main target of this study was to develop the liquid chromatography coupled with tandem mass spectrometry technique as a method for determination of salicylic acid in feed materials and compound feed.

Salicylic acid was extracted from feed with 0.1% hydrochloric acid in methanol. Separation was achieved in 8 min in a gradient elution using 0.1% formic acid and acetonitrile. The analyte was detected using negative electrospray tandem mass spectrometry. The procedure was validated to the specifications of the European Commission Decision No. 2002/657/EC.

The validation results showed the repeatability of the method, which was evaluated at three levels (0.25, 0.5, and 1.0 mg/kg). Calibration curves for the working ranges were linear (R² 0.9911 to 0.9936), and recoveries ranged from 98.3% to 101%. The LOD and LOQ for compound feed were 0.02 and 0.05 mg/kg, respectively. Salicylic acid was found mostly in corn, and its concentrations differed depending on whether it was young or fully grown (5.30–12.8 mg/kg and 0.13–1.01 mg/kg, respectively).

A sensitive and reliable method for the determination of salicylic acid in feed and compound feed using LC-MS/MS was developed.

Epstein–Barr virus (EBV) is a γ-herpesvirus associated with various neoplasms in humans and is a probable aetiological agent in breast cancer; however, a causal relationship has not yet been established. Because of the epidemiological and clinicopathological similarities between breast cancer and canine mammary tumours, dogs have been proposed as a valid model for breast cancer.

A total of 47 canine mammary gland tumour tissues were processed by routine histopathological technique with haematoxylin-eosin staining and classified according to the type of neoplasm. DNA was extracted from paraffin-embedded tissues and the EBNA-1 gene and the BamHI-W region specific for EBV were evaluated by nested PCR.

The histopathological evaluation revealed 2 benign neoplasms, and many carcinomas: 2 in situ, 9 simple, 3 solid, 10 complex, and 21 mixed. One sample was positive for the EBNA-1 gene, while all were negative for the BamHI-W region.

No association was found between EBV and mammary tumours in dogs. However, here we report for the first time the presence of an EBV gene sequence in a canine mammary tumour. It is likely that detection of EBV might be affected by the quality and quantity of DNA extracted from paraffin-embedded tissues. Additional studies are necessary to establish any association of EBV with mammary gland cancer in humans and in dogs, which could eventually lead to better public health prevention and control.

The aim of the study was to determine the effects of supplementation of sows’ and growing pigs’ diets with three newly developed synbiotic and two extant commercial probiotic products on selected immune parameters under field conditions.

The study was performed on 30 sows and 48 piglets of the Danbred breed. Immune parameters such as concentration and proportion of white blood cells and their subpopulations, immunoglobulins amount in serum, and serum concentration of cytokines and acute phase proteins were recorded with the use of a haematology analyser and ELISA kits.

No significant differences between treatment groups and controls were found with regard to the immune parameters evaluated except for serum immunoglobulin concentration, which was significantly increased by synbiotic products B and C and probiotic product D.

The results of the study indicate that the synbiotic products B and C and probiotic product D are worthy of further investigation as promising candidates to improve the immune status of healthy sows and their offspring.

The study aimed to observe TNF-α serum concentration as well as changes in respiration rate, body temperature, and pulse rate in burn victims during 84 h post burn.

A total of 30 healthy pigs were divided into two groups: A, the test group and N, the control group. The experimental group suffered burns to 30% of the body surface, and after infliction of the burns both groups were closely monitored.

The biggest increase in TNF-α serum concentration in the test subjects occurred around the 6^th h of the study, and the second biggest increase took place between 12^th and 36^th h. In the 36^th h, TNF-α was 2.5 times more concentrated in serum in the test group than in the control group. In the test group, the biggest increase in respiration rate occurred up to the 6^th h post burn, on average up to 29/min. In the 12^th h post burn, the mean pulse rate in the test group was 133/min and dropped to the lowest value in the 72^nd h of the experiment. A gradual increase in body temperature up to 41.72°C was observed up to the 30^th h post burn and decreased to a significant value of 40.74°C by the 84^th h of the study.

In a period of a pronounced rise in TNF-α serum concentration, this parameter, pulse rate, and respiration rate are highly correlated and are also influenced by multiple inflammation forming factors.

The study was designed to investigate the effects of repeated lipopolysaccharide (LPS) treatment on growth performance, lymphoid organ indexes, and blood cells in Sprague-Dawley rats.

Forty healthy weaned Sprague-Dawley rats were randomly equally divided into LPS and control groups. Each rat in the LPS group was injected via the caudal vein with LPS (100 μg/kg b.w.) for 10 days, and the control group was treated with an equal volume of normal saline. On the 1^st, 4^th, 7^th, and 10^th days, growth performance, lymphoid organ indexes, and blood cells were evaluated in five necropsied rats.

When rats were treated 3–10 times with LPS, their body weight and average daily gains increased more slowly than in the control group (P < 0.05). Repeated LPS treatment significantly increased spleen weight and the ratio of spleen to body weight (P < 0.05). White blood cells, neutrophils, and neutrophil percentage increased (P < 0.05) remarkably, but lymphocyte percentage, haemoglobin, and blood platelet counts decreased significantly (P < 0.05).

LPS treatment obviously suppresses growth and promotes peripheral immune organ proliferation. It is indicated that host protective mechanism can be activated by multiple small doses of LPS and prevents organs from further damage during stress status.

Methimazole-induced hypothyroidism is a clinical problem in the treatment of hyperthyroidism in people and animals and is an example of metabolic disease that can lead to fertility disorders and can give elastographic testicular changes.

Ultrasound elastography using the Esaote MyLab Twice ultrasound system and a morphological examination of testes were performed in seven methimazole-administered (group E) and seven healthy rats (group C).

The elasticity ratio of strains in the scrotal wall of the near-field test area to testicular tissue (ELX-T-RAT) and hardness percentage of strained tissues in the defined area of a testicle (ELX-T%HRD) in group E were statistically significantly lower than in group C. The degree of spermatogenesis was statistically significantly higher in group E than in group C and similarly seminiferous tubule diameters in group E were statistically significantly higher than in group C. Body weight and testicular weight in group E were statistically significantly lower than in group C.

Changes in the elastographical parameters of testes may result from disorders secondary to hypothyroidism. The usefulness of elastography is noteworthy in the case of evaluation of testis function in patients with some metabolic disorders.

Determination of sperm concentration and morphology in ejaculate is essential in evaluating fertility. Sperm shape and dimensions may depend on their concentration in ejaculate. The aim of the study was to evaluate the dependence of Hypor boar morphological and morphometric sperm characteristics on concentration in ejaculate.

The study was conducted on 120 ejaculates from 12 Hypor boars. Depending on sperm concentration, they were put into three groups: low, medium, and high. Ejaculate sperm concentration was determined with the photometric method. Slides were prepared from semen samples which were evaluated for the morphology of spermatozoa. The preparations for morphological analyses were by the eosin-gentian staining method.

As the sperm concentration in the ejaculate increased, the length, perimeter, and area of the sperm heads also did. In the ejaculates with higher sperm concentration the sperm heads were rounder. The ratios of head dimensions to tail length or total length were the highest in the sperm from ejaculates with the highest concentrations. The highest percentage of morphologically abnormal sperm was noted in ejaculates with low concentrations.

The dimensions and shape of sperm depend on the sperm concentration in the ejaculate. In ejaculates with the highest concentrations, the sperm have larger dimensions. Sperm concentration affects the frequency of morphological anomalies in the spermatozoa.

In recent years, the high sensitivity and specificity of novel miRNA biomarkers have been utilised for early diagnosis and treatment monitoring of various diseases. Previous reports showed that abnormal expression of miR-208 in mice resulted in the development of an aberrant cardiac conduction system and consecutive arrhythmias. On the other hand, a study on infarcted human heart tissue showed upregulation of miR-208a in subjects with ventricular tachyarrhythmias compared to healthy controls. We prospectively investigated the expression of miR-208a and -208b in the serum of dogs presenting different cardiac arrhythmias.

A total of 28 dogs with atrial fibrillation (n = 8), ventricular premature contractions (n=6), conduction system disturbances (n = 7), and free of heart conditions (as controls) (n = 7) were enrolled in the study. Total RNA was extracted from serum samples and miR-208a and -b, miR-16 as well as a cel-miR-39-5p spike-in were analysed with qPCR and ddPCR.

miR-208a and miR-208b were not expressed in any of the samples. The calculated ddPCR miR-16 relative expression (normalised with cel-miR-39 spike-in) showed a good correlation (r = 0.82; P < 0.001) with the qPCR results.

This outcome warrants further investigation, possibly focusing on tissue expression of miR-208 in the canine heart.

Ventricular rhythm disturbances are a common pathology in human and veterinary medicine. In humans, the algorithmic approach is used to differentiate wide QRS complex tachycardia. The most commonly used are the aVR and Brugada algorithms as well as the ventricular tachycardia (VT) score developed by Jastrzębski and coworkers. In veterinary medicine, no such algorithms are available and the only parameter used to describe VT abnormalities is the duration of the QRS complexes. The aim of this analysis was determining whether human medicine algorithms for VT are applicable in veterinary medicine to differentiate wide QRS complex tachycardia in dogs.

A retrospective analysis was performed on 11 dogs of both sexes and various breeds and age diagnosed with VT. The diagnosis was based on ambulatory ECG, further established based on the reaction to lidocaine or adenosine or an invasive electrophysiological study.

Of the 11 tracings passed through the aVR algorithm, 10 met the VT criteria. The most common criterion was the Vi/Vt ratio (8 out of 11 tracings). Based on the VT score, seven out of eight dogs had a high probability of VT.

Retrospective analysis of ECGs by aVR and VT score indicates that the applied algorithms may be useful in differentiating wide QRS complex tachycardia as a quick, easy, and non-invasive alternative to cardiac electrophysiology.

The study evaluated the ameliorative effects of Helianthus annuus leaf extract on nephrotoxicity, cardiac, and haematologic disorders in alloxan-induced hyperglycaemic rats.

The cold maceration method with 80% methanol was used in the preparation of H. annuus extract. Thirty alloxan-induced hyperglycaemic rats were randomly assigned to five equal groups (A–E). Groups A and B received 5% tween-20 solution in water (5 mL/kg) and glibenclamide (2 mg/kg), respectively; while groups C, D, and E received 150, 300, and 600 mg/kg of the extract, respectively, per os once daily for 21 consecutive days. The levels of serum urea, creatinine, haematological indices, and histopathological changes in the kidneys and heart were evaluated 24 h after the last treatment on day 21.

The extract and glibenclamide significantly (P < 0.05) reduced the levels of serum urea and urea : creatinine ratio in diabetic rats when compared with the vehicle treated group. The extract and glibenclamide also ameliorated haematological disorders and kidney and cardiac damage induced by alloxan.

H. annuus extract produced nephroprotective, cardioprotective, and haematoprotective effects and might prevent the advancement of diabetic complications such as diabetic nephropathy and cardiovascular diseases in diabetic patients.

Intestinal obstruction such as atresia coli causes pathophysiological changes in gastrointestinal tissue due to the rise of intra-abdominal pressure. The aim of this study is to determine the intestinal damage with intestinal biomarkers in calves with atresia coli.

The study was conducted on 40 Holstein calves diagnosed with atresia coli with mild to moderate abdominal distention and 10 healthy Holstein calves which served as the control. Blood samples were collected from all calves, and then serum concentrations of intestinal biomarkers were estimated, namely intestinal fatty acid binding protein (IFABP), liver fatty acid binding protein (LFABP), trefoil factor 3 (TFF3), and intestinal alkaline phosphatase (IAP), using commercially available specific bovine ELISA kits. An automatic blood gas analyser was employed for determining the lactate concentration.

The concentrations of serum LFABP (P < 0.01), IFABP, TFF3, IAP, and blood lactate (P < 0.001) were significantly higher in calves with atresia coli than in healthy calves.

The calves affected with atresia coli exhibited severe intestinal damage, and IFABP, LFABP, and TFF3 have significant diagnostic importance and play a useful role in determining the intestinal damage due to intestinal obstruction. High levels of IAP and lactate may serve as a signal for the development of intestinal injury.

In recent years, the use of bone scaffolds as bone tissue substitutes, especially the use of such as hydroxyapatite and tricalcium phosphate, has been very popular. Today, the use of modern engineering techniques and advances in nanotechnology have expanded the use of nanomaterials as bone scaffolds for bone tissue applications.

This study was performed on 60 adult male New Zealand rabbits divided into four experimental groups: the control group without any treatment, the second group receiving hydroxyapatite, the third group treated with β-tricalcium phosphate, and the fourth group receiving nanocomposite polycaprolactone (PCL) scaffold. In a surgical procedure, a defect 6 mm in diameter was made in a hind limb femur. Four indexes were used to assess histopathology, which were union index, spongiosa index, cortex index, and bone marrow.

The results showed that nanocomposite PCL and control groups always had the respective highest and lowest values among all the groups at all time intervals. The histopathological assessment demonstrated that the quantity of newly formed lamellar bone in the nanocomposite PCL group was higher than in other groups.

All these data suggest that PCL had positive effects on the bone healing process, which could have great potential in tissue engineering and clinical applications.

The purpose of this study was to investigate the protective effect of hydrogen-rich saline (HRS) against liver ischaemia-reperfusion combined resection injury.

Eighteen miniature pigs were randomly divided into three groups: a sham operated group (sham group, laparoscopic liver ischaemia-reperfusion combined resection injury group (IRI group), and a hydrogen-rich saline intervention group (IRI + HRS group). Samples of hepatic tissue and serum were collected at the time of reperfusion and then 3 h, 1 d, and 3 d post reperfusion. Liver function, oxidative stress, autophagy-related mRNA genes, and protein expression were evaluated. Changes in cell and tissue ultrastructure were examined by transmission electron microscopy.

Compared with the sham group, the level of autophagy of hepatocytes increased in the IRI and IRI + HRS groups, corresponding to high oxidative stress and severe liver function injury. Liver function, antioxidant content, autophagy levels, and liver injury were improved after intervention with HRS in the IRI + HRS group compared with the IRI group.

Intervention with hydrogen-rich saline could exert a protective effect against liver ischaemia-reperfusion combined resection injury through the reduction of oxidative stress and hepatocyte autophagy.