Volume 65 (2015): Issue 4 (December 2015)

Blood smears stained with Diff Quick are the initial tool for cytological diagnosis of Haemogregarina spp. However, the development of sensitive and specific molecular methods enabled the detection and identification of parasites in the sample and to clarify the evolutionary relationships of adeleorinid parasites within the Apicomplexa.

The current study was attempted in order to perform cytological investigation and molecular identification of the hemoparasites in thirty European pond turtles (Emys orbicularis) from the quarantine section at Belgrade Zoo, which have been found in poor health condition with massive skin hemorrhages, based on intraerythrocytic parasitic forms on hematological smears and 18S rDNA sequence, respectively.

Different life cycle stages of the Haemogregarina sp. were noticed within the erythrocytes in the peripheral blood. Biochemical analysis indicated lower values of AST and iron in most of the infected turtles while hematological analysis showed a changed hematocrit value, a decrease in the number of red blood cells and low hemoglobin levels. Amplifications of the 18S rDNA sequence of Haemogregarina were detected in 30/30 (100%) turtles with clinical symptoms. The identity of PCR products was confirmed by direct DNA sequencing. Future research concerning H. stepanowi in Serbia should be applied to its definitive host-the leech.

The aim of this study was to evaluate the serum cardiac troponin I (cTnI) concentrations in weaned calves with moderate and severe bovine respiratory disease. Eighteen healthy weaned calves (group I), 18 weaned calves with moderate bovine respiratory disease (group II) and 18 weaned calves with severe bovine respiratory disease (group III) were included in the study. Clinical signs and severity of respiratory disease according to clinical index scores were determined. Blood cell counts and cardiac biomarkers, such as serum cTnI concentrations and creatine kinase-myocardial band (CK-MB), and lactate dehydrogenase (LDH) activities were determined in all groups. Temperature and respiratory and pulse rates were significantly increased in calves with moderate and severe bovine respiratory disease compared to healthy calves (P < 0.05). Serum cTnI concentrations and LDH activities were significantly increased in the calves with severe bovine respiratory disease compared to healthy calves and calves with moderate bovine respiratory disease (P < 0.05). A significant increase in white blood cell counts in calves with moderate and severe bovine respiratory disease and lymphopenia and neutrophilia in calves with severe bovine respiratory disease were also found. The study suggests that severe bovine respiratory disease causes increased serum cTnI concentrations in weaned calves.

The objective of the present study was to identify the Prolactin Receptor (PRLR) gene polymorphism related to litter size traits. The study included 101 Topigs 20 line of sows with 426 litters. The traits studied were: Total Number of Born (TNB), Number of Born Alive (NBA), Number of Still Born (NSB), and Number of MUMmified (NMUM) piglets. Polymorphism was identified with the polymerase chain reaction-restriction fragment length polymorphism method. Allelic and genotype frequencies and deviation from Hardy-Weinberg equilibrium were verified with the chi-square test. Analysis of litter size traits was performed using the General Linear Model, which included the potential environmental effects. Additive and dominant allele variances were observed by the regression procedure. In the studied population of sows, the frequency of heterozygotes (0.5149) for PRLR gene exceeded the total number of AA (0.0198) and BB (0.4653) homozygotes, which resulted in a high proportion of B allele (0.7228). The results for PRLR showed statistically significant (P<0.05) differences in first parity sows between BB and AB genotypes for TNB and NBA. Significant differences (P<0.05) were recorded in third parity sows between BB and AB genotypes for NBA, and in AA genotype versus AB and BB genotypes for NMUM. The fourth and subsequent parity sows of AA genotype had a significantly higher (P<0.05) rate of NBA as compared with those of AB and BB genotypes. In all parities analysed, the difference between the BB and AB genotypes for NBA was statistically significant (P<0.05). Interpretation of the results at the levels of phenotypes and either additive or dominant variance was quite difficult due to the small number of AA homozygous sows. The calculation model yielded a significant effect (P<0.05) as well as tendency (P<0.1) for the mentioned effects except for age at first farrowing.

The aim of this research was to determine the influence of 56-days (from day 4 to day 60 post partum) of simultaneous supplementation of selenium (Se) and iodine (I) enriched Chlorella algae on blood and urine Se concentrations and thyroid hormone levels in ewes and lambs. The study included 18 Šumava sheep ewes and their lambs divided into three groups (CON, HSe, HSeI). Selenium and I content in the diet of ewes measured 0.2 and 0.7 mg (CON), 0.4 and 0.7 mg (HSe), 0.4 and 1.3 mg (HSeI) per kg of dry matter. During the experimental period a significantly higher blood serum Se concentration was determined in ewes of the HSeI group (P< 0.001) and urine (P< 0.01). A significantly higher (P< 0.001) blood serum Se concentration was found also in lambs of the HSeI group. The highest Se concentration in the blood serum was recorded in ewes (128.0 μg/L) and lambs (74.2 μg/L) on the 26th day of the experiment. The results demonstrate a higher utilization of Se administered in the organic form accompanied by a higher intake of I applied also in the organic form. Plasma thyroid hormone levels in ewes were not explicitly affected by higher Se and I supplementation. A higher Se concentration in the lambs’ blood serum led to significantly higher (P< 0.05) T3 levels up to 30 days post birth.

The aim of this study was to evaluate the parameters of intraoperative and postoperative pain in bitches undergoing laparoscopic or conventional ovariectomy. The study was conducted on 20 healthy, adult bitches of medium and large breeds. All the bitches were subjected to the same anesthetic protocol. Carprofen was applied preoperatively, and general anaesthesia was carried out with a combination of medetomidine, propofol and sevoflurane. The bitches were divided into two groups of 10 individuals each. The first group underwent conventional ovariectomy while the second one was subjected to the laparoscopic procedure. The intensity of intraoperative pain was estimated at different time points by measuring changes in heart and respiratory rate, arterial blood pressure, and body temperature. A multifactor pain scale was used to assess the intensity of postoperative pain. Dogs in the group which was scheduled for conventional ovariectomy (COV group) responded to painful intra-operative stimuli with increased heart rate and respiratory rate. Arterial blood pressure and body temperature decrease was detected in COV group. Laparoscopic ovariectomy induced less intra- and postoperative pain when compared to conventional ovariectomy. Beside that, the total anaesthesia time and postoperative recovery were shorter in dogs that underwent laparascopic ovariectomy.

Diflubezuron is increasingly used in areas where mosquito larvae developed resistance to other insecticides. In our community diflubenzuron is not used to control mosquito larvae. Two formulations of 1% diflubenzuron (on corn-cob EF-1, and zeolite EF-2) were tested on Culex pipiens L (larvae) on one canal in the Belgrade suburb area. The effect was followed for seven weeks after application of the formulations. Formulation EF1 achieved a reduction in mosquito L₁L₂ larvae between 23.9% and 89.4%. The change was statistically significant the 21^st and 28^th day (p<0.001), 35^th and 42^nd day (p<0.01) and 49^th day (p<0.05). The maximal reduction obtained by formulation EF2 was 69.1%. The accomplished reduction was significant on the 28^th and 42^nd day (p<0.001), 35^th day (p<0.01) and 21^st (p<0.05). Both formulations have maintained a good residual effect on the lower developmental larval stages. Maximum reduction achieved by EF1 on L₃L₄ larvae was 97.4%. Reduction of larvae was high between the 7^th and 42^nd day (66.4 - 97%). Statistically significant values were recorded on the 21^st, 28^th and 35^th day. Formulation EF2 achieved a reduction of 99.5%. A statistically significant reduction in the value of mosquito larvae was obtained on the 14^th, 21^st, 28^th, 35^th and 42^nd day.

Between the two used formulations there was no significant difference in the number reduction of lower larval stages, but for the higher larval stages EF1 proved to be more efficient.

The bursa of Fabricius (BF) is a lymphoepithelial organ found only in birds. Differences in morphology of BF could play an important role in immune response. The objective of this study was to investigate the histological and histochemical characteristics of the bursa of Fabricius in the long-legged buzzard (Buteo rufinus). The material for the study comprised bursa samples obtained from three long-legged buzzards with permission of the General Directorate of Nature Protection and National Parks (Ankara, Turkey). Briefly, interfollicular epithelium (IFE) was shown to be columnar in shape and not to contain goblet cells. Reticular fibers were located in interfollicular septae. Each lymphoid follicle in the bursa of Fabricius in the long-legged buzzard was remarkably linked to the follicle associated epithelium (FAE). Namely, FAE has been reported to stimulate antibody production by transferring antigens to the medulla and have a leading role in developing of local immune response. Among the others, the species-specific differences in bursa of Fabricius morphology of long-legged buzzard (Buteo rufinus) also might support the continuity of this species in nature.

Sjenica cheese is an artisanal cheese stored in brine, traditionally produced from raw sheep’s milk in the southern part of Serbia - Sjenica Pester plateau.

The aim of this study was to perform the safety evaluation of Sjenica cheese. As one of the safety criteria we considered the number of coagulase positive staphylococci and their enterotoxigenic potential. Antibiotic susceptibility/resistance patterns of autochthonous lactic acid bacteria and coagulase-positive staphylococci isolated from Sjenica cheese was also investigated.

During the monitoring period of the cheese-making process, coagulase positive staphylococci did not reach the value of 10⁵ cfu/g. Among coagulase positive staphylococci, 12 (46,15%) isolates showed enterotoxigenic potential and were identified as Staphylococcus intermedius (11 isolates) and Staphylococcus aureus (1 isolate). Vancomycin resistance was the most prevalent phenotypic resistance profile in coagulase positive staphylococci.

Lactococci present the most dominant population among lactic acid bacteria. The most prevalent resistance phenotype in lactococci was resistance to streptomycin (83.33%), ampicillin and penicillin (70.83%); lactobacilli were characterized by resistance to vancomycin (62.5%) and tetracycline (54.17%), while resistance to streptomycin (82.46%) was the most prevalent phenotypic profile in enterococci.

All coagulase positive staphylococci and lactic acid bacteria isolates that showed resistance to tetracycline on disc diffusion and E-test, were tested for the presence of ribosomal protection proteins, tet(M) and tet(K) genes. All isolates were positive for ribosomal protection proteins genes; 14 (60.87%) isolates showed tet(M) gene presence, while 2 lactobacilli isolates revealed the presence of tet(K) gene.

The aim of this study was to describe the prevalence of antibiotic resistance to beta-lactams and to evaluate two resistance genes bla_TEM and bla_OXA-1 in Escherichia coli isolates from faeces on six swine farms in the Republic of Bulgaria. A total of 186 E. coli isolates from 192 faecal swabs were tested by the disk diffusion method to determine resistance patterns to 11 antimicrobial agents. Resistance to beta-lactams was determined by disk diffusion method, E-test, micro-broth dilution method and PCR. About 40.3% of the E. coli isolates from swine were resistant to ampicillin. The highest resistance was observed in E.coli isolates from weaned pigs to ampicillin – 60.0% and to cephalotin – 45.5 %. The E. coli isolates resistant to beta-lactams were examined for the presence of bla_TEM and bla_OXA-1 genes. The most common bla gene identified was bla_TEM, which was found in 92.0% of swine isolates.

Resistance to tetracycline was studied in Salmonella Infantis isolated from 28 poultry farms in the Northern part of Serbia (The Autonomous Province of Vojvodina). A total of 18 isolates were resistant to nalidixic acid (NAL) and tetracycline (TET). The minimal inhibitory concentration (MIC) to TET, ranged from 1-256 mg/L. Namely, 13 isolates exhibited MIC to TET at 256 mg/L, in four of the isolates, the MIC was 128 mg/L and one isolate had MIC 64 mg/L. Ten isolates were exhibiting a MIC of 1mg/L. It was evident that Salmonella Infantis had also spread to breeders and layers. In this work, we detected the tetA gene and the corresponding tetR gene (encoding the repressor protein) as well as the truncated transposon Tn1721, which are responsible for the resistance to TET. The presence of the non conjugative transposons from the conjugative plasmid has facilitated the spread of resistance to TET in Salmonella. It was concluded that higher biosecurity practice in poultry farming presents the best option to eliminate infections caused by Salmonella spp. from poultry flocks in Serbia. A rational use of antimicrobials is necessary to prevent any further spread of Salmonella Infantis resistant clones.

The results on serological testing of blood sera from stallions and mares used for breeding and the presence of the viral genome of Equine Arteritis Virus (EAV) in stallion semen are presented. The blood and semen samples were taken from a horse stable on the territory of the Republic of Serbia during 2012, 2013 and 2014. Detection of anti-EAV specific antibodies in blood sera was performed by the virus neutralization test (VNT), and identification of EAV genome RNA in stallion semen was done by reverse-transcriptase polymerase chain reaction (RT-PCR). In 2012, high seroprevalence of EAV was detected in the investigated stable. In total, 45% and 65 % of stallions and mares reacted positive, respectively, and the antibody titre values ranged between 2 and 10 log₂. High seroprevalence was confirmed in the same animals again in 2013. Out of two stallions tested semen samples in 2013, the viral genome was detected by RT-PCR in 3 examined semen samples from a seropositive stallion, while EAV was not detected in 3 semen samples of a seronegative stallion. During 2014, 11 semen samples were collected from two seropositive stallions. Again, the presence of EAV was confirmed by RT-PCR in all 8 semen samples originating from the same stallion with the EAV genome positive semen result in 2013, whereas the virus was not detected in semen samples originating from the second anti-EAV antibody positive stallion. The presence of EAV-specific antibodies was confirmed in the blood sera of the mares inseminated with the semen of seropositive stallions in 2012 and 2013.

Priapism, defined as a persistent and painful penile erection in the absence of sexual excitation, is an uncommon disorder of cats that usually requires penile amputation and perineal urethrostomy. This report describes a case of priapism in a short-haired Persian male cat aged 3 years. The condition had appeared after attempted mating with a female at the end of oestrus and the time of referral to the authors was 10 days after onset. Previous treatment had consisted of corticosteroids and local lubricants. On physical examination, the cat appeared bright and alert; the penis was erect but the colour was normal; blood analysis excluded viral pathologies. The cat history revealed that he had been treated with 500 IU hCG i.m. because he had to mate with three queens in a short period of time. In consideration of the mild symptoms, conservative treatment was adopted. The condition resolved in about 40 days. This is the first report of priapism following hCG administration in a cat and indicates that the misuse of this gonadotropin might cause potentially harmful side effects.