Eight-Year Study of Haemogregarina stepanowi Infection in Poached European Pond Turtles (Emys orbicularis) Held in Belgrade Zoo Quarantine.

The eight-year study (2015-2023) was performed on a large sample of poached European pond turtles infected with Haemogregarina stepanowi and held in a pond that belongs to a quarantine section of Belgrade Zoo. The protected species of European pond turtles have been found in poor health, with general weakness, anorexia, and low motility. Comprehensive cytological, hematological, molecular, and postmortem evaluations have been performed. Initially, Diff Quick staining of the blood smears revealed rounded or elongated erythrocytes, often bearing premeront or U-shaped gamont of the hemogregarines inside. The reduced erythrocyte numbers, hemoglobin, and hematocrit values found in the examined population of infected turtles indicated anemia. Macroscopically, shell necrosis and massive skin hemorrhages were the most prominent findings observed in diseased turtles. Microscopically, the lungs, liver, kidneys, and spleen revealed hyperemia, hemorrhages, and the presence of parasitic stages in tissue samples in 31 of 40 necropsied turtles. Cytological and microscopic examination of the samples proved to be sufficient for establishing the infection, but molecular analyses of the 18S sequence were used for phylogenetic studies. Over the years, the number of diseased and dead turtles has decreased, which could be hypothetically attributed to the elimination of leeches as the definitive host.

A case of nerve sheath tumor followed by multicentric high-grade T-cell lymphoma in an African pygmy hedgehog (Atelerix albiventris).

A 3-year-old intact male African pygmy hedgehog was presented at the Teaching Hospital of the Faculty of Veterinary Medicine, University of Belgrade, with a growth on the left side of its abdomen. After clinical examination, the mass was surgically removed, and histopathological findings indicated a nerve sheath tumor. The hedgehog fully recovered after surgery and was euthanized eight months later due to the appearance of multicentric changes in the internal organs. Further necropsy and macroscopic, cytologic, histopathologic, and immunohistochemical findings revealed that the tumor was a multicentric high-grade T-cell lymphoma. This is an unusual case of an African pygmy hedgehog with two different neoplasms-a nerve sheath tumor followed by lymphoma.

The COVID-19 pandemic affects owners walking with their dogs.

The COVID-19 pandemic imposed a strict state of emergency on Belgrade residents with a curfew and restriction on movement. Therefore, the aim of this study was to determine how the measures introduced as a consequence of this disease in the capital of Serbia affected the duration of walking time of owners and their dogs. The study allowed for sociodemographics of owners, characteristics of their homes and breeds of dog. The research was conducted in the period from March 22 to April 4, 2020. In general, the COVID-19 pandemic restrictions reduced the duration of dog walks and owners' walking time. Of the 216 adult dog owners, 59% walked their dogs successfully (≥150 min/week) before the state of emergency. The number of owners who walked their dogs for ≥150 min/week decreased to only 44% (N = 96) during the state of emergency. The pandemic also decreased the number of owners who achieved total walking times of ≥150 min/week, so only 56% of them were successful in total walking time during the state of emergency (100% were successful before the pandemic, as this was one of inclusion criteria). This difference was statistically significant (P < 0.01). A Wilcoxon signed-rank test found significant differences in the total walking time before and during the state of emergency, with regard to characteristics of the owner, home and dog, with strong effect sizes for these differences (P < 0.01). McNemar's test showed that female owners, owners aged 45-64 years and owners with high income who were previously successful in walking their dogs (≥150 min/week) were more affected by the pandemic in regard to walking time with dogs (P < 0.01). Significant correlations (P < 0.01) were observed between dog breed and walking time before the state of emergency; age and walking time with the dog during the state of emergency and; age and total walking time during the state of emergency. Comparisons between different categories within the same characteristics (owner demographics, home characteristics and dog breed) by Fisher's exact test found significant differences only between younger and older dog owners in total walking time during the state of emergency (P < 0.01). Younger owners were significantly more successful in achieving ≥150 min/week total walking time than owners aged 45-64. The COVID-19 pandemic has left some owners jobless and allowed them to spend more time walking dogs. The occurrence of this disease has led to changes in the social structure of households and in the daily habits of household members. These factors have affected on the length of walking time of owners and their dogs in Belgrade.

Kinds of pet chosen and manner of pet acquisition during COVID 19 in Serbia.

In mid-March 2020, a state of emergency was declared in Serbia in order to prevent the spread of COVID 19. Many activities of Serbian citizens were limited or completely stopped. Citizens had to spend most of their time confined in their homes. However, some still wanted and managed to acquire pets. Therefore, the aim of this study was to examine which kinds of pets (dogs, cats and non-traditional animals) the residents of Serbia acquired during the first year of the COVID 19 pandemic and in which manner (purchase, adoption or gift). If the pets were acquired by adoption, the participants were asked from what sources they adopted them. Finally, how the characteristics of the owner and their household influenced the adoption of pets was examined. A total of 566 pet owners who acquired pets in the pre-COVID 19 and COVID 19 periods participated in the research. The COVID pandemic reduced the number of pet owners 2.7-fold in 2020 compared to 2019. Dogs were the most favourite pets acquired before (43%) and during (43%) the pandemic. During the pandemic, 61% of pets were acquired by adoption and the percentage of dogs, cats and non-traditional pets adopted directly from previous owners increased by 32%, 13% and 12%, respectively. The probability of acquiring pets by adoption was significantly increased by the presence of people older than 65 in the household (OR=2.56; P=0.02), the absence of children up to 5 years of age (OR=0.17; P<0.001) in the household, lower monthly incomes (OR=0.47; P<0.001), the greater available free time of the owner (OR=2.09; P<0.001), and the period of adoption (OR=2.42; P<0.001).

Evaluation of the effects of two anaesthetic protocols on oxidative status and DNA damage in red-eared sliders (Trachemys scripta elegans) undergoing endoscopic coeliotomy.

The aim of this study was to assess how red-eared sliders (Trachemys scripta elegans) respond to anaesthesia itself and coelioscopy. For that purpose, the turtles were anaesthetised with ketamine-medetomidine or propofol, and the activities of superoxide dismutase (SOD), catalase (CAT) and glutathione S-transferase (GST) and the level of malondialdehyde (MDA) were determined by spectrophotometry. The possible genotoxic effects of the anaesthetic agents were estimated by comet assay. A total of 24 turtles were included in this study. The animals were divided into four groups according to the anaesthetic protocol and according to whether endoscopy would be performed. Significantly decreased activities of CAT were found only in the propofol group and in turtles undergoing coelioscopy. Both anaesthetic protocols induced significantly increased MDA levels, while no differences were observed after the intervention. A significant increase in GST activity was detected in turtles after both anaesthetic protocols, but after coelioscopy significant changes in GST activity were found only in the propofol group. However, no differences in SOD activity and no DNA damages were detected in either group. These findings suggest that ketamine-medetomidine may be more suitable anaesthetic agents in red-eared sliders than propofol.

A molecular and haematological study of Theileria equi in Balkan donkeys.

Equine piroplasmosis in donkeys has been recognised as a serious problem of major economic importance. The present molecular study is the first investigation of the presence of Theileria equi and Babesia caballi in Balkan donkeys and of the possible haematological alterations related to it. A total of 70 apparently healthy donkeys from Serbia were included in this study. The overall prevalence of T. equi infection in donkeys tested with multiplex PCR was 50%. There was no B. caballi-positive sample. Infections in donkeys included in this study seem to be associated with decreased red blood cell count, haemoglobin concentration, haematocrit and platelet count, and with increased white blood cell count, mean corpuscular haemoglobin and mean corpuscular haemoglobin concentration. Altered haematological parameters in donkeys can lead to a decrease in working capacity and production performance. Further molecular research and long-term monitoring of equine piroplasmosis is needed in Serbia and throughout Europe.

Molecular detection and prevalence of Theileria equi and Babesia caballi in horses of central Balkan.

Equine piroplasmosis is significant tick-borne disease with wide distribution. The prevalence of equine piroplasmosis in Serbia, Montenegro and Bosnia and Herzegovina is unknown. In aim to obtain a first insight into the prevalence we performed molecular epidemiological study which included 142 horses, on seven locations in these three countries. We first performed PCR for the detection of a 450bp long section of the 18S rRNA of piroplasma-specific region. For all positive samples we have done multiplex PCR for the species detection. Species determination was further confirmed by sequencing PCR products of 10 randomly selected Theileria equi and all Babesia caballi samples. The overall prevalence rates in analysed region for T. equi and B. caballi were 22.5% and 2.1%, respectively. Possible risk factors (such as location, age, sex and activity) associated with PCR positivity were evaluated. Marked differences were found in prevalence between geographic areas. There was no significant association between positivity and age group. T. equi was more prevalent in females and farming horses. This is the first report on the molecular survey of T. equi and B. caballi in central Balkan. Further prevalence studies on definitive host and vectors in this region are necessary.

First report of Polycystic kidney disease occurrence in Persian cats in Serbia.

Polycystic kidney disease (PKD) is an inherited autosomal disorder in cats, mostly diagnosed in Persian cats. Renal cysts can be diagnosed by ultrasound, but cats must be at least 16 weeks old. The goals of this study were to assess the occurrence of PKD in Serbia using a randomly selected group of Persian cats, to compare the diagnostic efficacy of ultrasound and genetic tests, and to measure haematological and selected biochemical parameters. We examined 70 cats of Persian breed, between 4 months and 8 years of age. Complete blood count and selected biochemical parameters were measured, renal ultrasound was performed. Swabs of the oral cavity were obtained for genetic testing. Percentage of PKD positive cats identified by genetic testing was 48.6%, whilst only 18.6% were detected through ultrasound. Animals that were polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) positive and ultrasound negative ranged from 4 months to 3.5 years. All haematological and biochemical parameters were within the the normal range values in all examined cats. Genetic methods proved to be the most effective for reliable and early diagnosis of PKD in Persian cats. DNA analysis can be used right after birth, and excludes the need for other diagnostic procedures, such as ultrasound.

Clinical babesiosis and molecular identification of Babesia canis and Babesia gibsoni infections in dogs from Serbia.

Canine babesiosis is a frequent and clinically significant tick-borne disease. Sixty symptomatic dogs with clinical findings compatible with babesiosis were included in this study conducted in Serbia. After clinical examination, blood samples were taken for microscopic examination, complete blood count (CBC), Canine SNAP 4Dx Test, DNA analyses and sequencing. The main clinical signs included apathy, anorexia, fever, brown/red discoloration of urine, pale mucous membranes, icterus, splenomegaly, and vomiting. The main clinicopathological findings in Babesia infections were a slight to severe thrombocytopenia and a mild to very severe normocytic normochromic anaemia. Microscopic evaluation revealed 58 positive samples with the presence of large and small intraerythrocytic piroplasms in 57 and 1 sample(s), respectively. No co-infections were found using SNAP test. Two Babesia species, B. canis (58/60) and B. gibsoni (2/60), were differentiated by polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP). Species identification was further confirmed by sequencing PCR products of B. gibsoni samples and six randomly selected B. canis samples. All dogs were treated with imidocarb dipropionate (6.6 mg/kg of body weight), given intramuscularly twice at an interval of 14 days. This report presents the first molecular evidence of the occurrence of B. gibsoni and B. canis, confirmed by DNA sequencing, in sick dogs from Serbia.

Variability of the honey bee mite Varroa destructor in Serbia, based on mtDNA analysis.

Only two mitochondrial haplotypes (Korea and Japan) of Varroa destructor, the ectoparasitic honey bee mite, are known to be capable of infesting and successfully reproducing in Apis mellifera colonies worldwide. Varroa destructor (then called Varroa jacobsoni) was observed in Serbia for the first time in 1976. In order to obtain insight into the genetic variability of the mites parasitizing A. mellifera we analyzed 45 adult female mites sampled from nine localities dispersed throughout Serbia. Four fragments within cox1, atp6, cox3 and cytb mtDNA genes were sequenced. The Korea haplotype of V. destructor was found to be present at all localities, but also two new haplotypes (Serbia 1 and Peshter 1) were revealed, based on cox1 and cytb sequence variability. The simultaneous occurrence of Korea and Serbia 1 haplotypes was observed at five localities, whereas Peshter 1 haplotype was identifed at only one place.

Sex determination in 58 bird species and evaluation of CHD gene as a universal molecular marker in bird sexing.

The aim of this research was to test the CHD gene (Chromo Helicase DNA-binding gene) as a universal molecular marker for sexing birds of relatively distant species. The CHD gene corresponds to the aim because of its high degree of conservation and different lengths in Z and W chromosomes due to different intron sizes. DNA was isolated from feathers and the amplification of the CHD gene was performed with the following sets of polymerase chain reaction (PCR) primers: 2550F/2718R and P2/P8. Sex determination was attempted in 284 samples of 58 bird species. It was successful in 50 bird species; in 16 of those (Alopochen aegyptiacus, Ara severus, Aratinga acuticaudata, Bucorvus leadbeateri, Cereopsis novaehollandiae, Columba arquatrix, Corvus corax, C. frugilegus, Cyanoliseus patagonus, Guttera plumifera, Lamprotornis superbus, Milvus milvus, Neophron percnopterus, Ocyphaps lophotes, Podiceps cristatus, and Poicephalus senegalus), it was carried out for the first time using molecular markers and PCR. It is reasonable to assume that extensive research is necessary to define the CHD gene as a universal molecular marker for successful sex determination in all bird species (with exception of ratites). The results of this study may largely contribute to the aim.