Irene Muñoz, Jevrosima Stevanovic, Zoran Stanimirovic and Pilar De la Rúa
Genetic variation of Apis mellifera from Serbia inferred from mitochondrial analysis
Two honeybee subspecies inhabit Serbia; Apis mellifera carnica and A. m. macedonica. Both belong to eastern Mediterranean (C) evolutionary lineage. Furthermore three Serbian honeybee ecotypes restricted to particular regions, were defined through morphometry and cytogenetic analyses. In this study, mitochondrial data have been used to analyze the molecular diversity of the honeybee population from Serbia. Seven haplotypes of the C evolutionary lineage have been found, two of them are newly described (C2o and C2p) and restricted to two regions, which ultimately increased the number of haplotypes found in this lineage. Comparisons with surrounding honeybee populations suggest a hybrid situation between A. m. carnica and A. m. macedonica and also introgression from A. m. ligustica. The results should be considered when dealing with future conservation strategies, and for pathogen-parasite-tolerant breeding programs.
The aim of the study was to establish morphometric, biochemical and hematological values for the endangered Balkan donkey breed (Serbia) and to explore the possible age dependence of the parameters tested. Inter-breed similarity of morphometric parameters was assessed by comparing the data obtained for the Balkan donkey with morphometric measurements of several previously characterized domestic donkey breeds. The study population included 74 donkeys, divided in two age groups (group A ≤ 3 years; group B > 3 years). In total, 18 morphometric, 13 hematological and 14 biochemical parameters were assessed. Significant morphometric differences (p<0.05) in body length, head length, chest circumference and body weight were found between the two age groups. Significant differences in morphological parameters were revealed among the Balkan donkey and other donkey breeds (Catalonian, Croatian and Albanian), but results of cluster analysis demonstrated the smallest distance between the Balkan donkey and Albanian donkeys. The results of morphometric analyses showed consistency of the obtained values within the breed, and diversity as compared to other donkey breeds, and, thus, could be taken as referent for the Balkan donkey. Hematological and biochemical profiles obtained for the Balkan donkey were consistent with previous reports and within the recommended reference ranges. White blood cell, mid cell and granulocyte counts, showed significantly higher (p<0.05) values in donkeys under 3 years of age, while the only biochemical parameter affected by age was alkaline phosphatase. The information gained through characterization of the Balkan donkey breed provides a basis for conservation and development of the breed standard.
Thelazia callipaeda is a parasitic nematode causing ocular infections in different mammalian species and humans, clinically manifested as lacrimation, conjunctivitis, keratitis, corneal opacity or corneal ulcer. In this paper, we reported six cases of autochthonous canine and feline thelaziosis on different localities in Serbia. Total of 285 parasites (85 males and 200 females) were collected from the eyes of infected animals (n=6) suffering from uni- or bilateral conjunctivitis, with the number of parasites ranging from 7 to 150 per animal. All parasites were morphologically identified as T. callipaeda, while molecular analyses of cytochrome oxidase 1 (cox1) gene revealed the presence of h1 haplotype, as the unique previously reported in other studies in Europe. Since T. callipaeda is a newly detected parasite in Serbia with the infective potential for humans, there is a necessity for animal owners education and cooperation among professional services in order to control this zoonosis.
Jòzsef Özvegy, Darko Marinković, Miloš Vučićević, Bojan Gajić, Jevrosima Stevanović, Dejan Krnjaić and Sanja Aleksić-Kovačević
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.
Zoran Stanimirović, Uroš Glavinić, Nada Lakić, Dejan Radović, Marko Ristanić, Elmin Tarić and Jevrosima Stevanović
Varroa destructor is the most important honey bee parasite. There are various methods used in the control of this mite, but none of them meets all requested criteria, to be safe, effective and easy to apply. The objective of this study was to evaluate the varroacidal efficacy of newly created plant-derived formulation Argus Ras (mixture of extracts of Sophora flavescens, Ginkgo biloba, Gleditsia chinensis and Teucrium chamaedrys) in a field trial. The investigation was conducted on 240 Apis mellifera colonies equalized in respect of brood amount, adult bee population and food reserves. Efficiency was evaluated by applying Argus Ras consecutively with two other acaricides, amitraz and oxalic acid. Average acaricidal efficacy of Argus Ras was 80.89%, being higher of other previously tested essential oils. Besides, it showed a potential in knocking down the mites resistant to other acaricides. It should not be neglected that Argus Ras requires a smaller number of treatments and financial investments than other formulations used for the control of Varroa mites.
Bluetongue, a vector-born disease caused by the Bluetongue virus (BTV) and transmitted by Culicoides biting midges, is considered to be one of the most important diseases of domestic ruminants. The first outbreak of bluetongue in Serbia was reported in 2001, when BTV serotype 9 was identified in sampled materials. In 2014, outbreak of BTV-4 in Serbia caused considerable economic losses affecting sheep, cattle and goats. During this outbreak, BTV-4 was recorded in 644 outbreaks within 49 municipalities, part of 17 administrative regions. From the total number of sheep kept in areas affected by bluetongue (n=1 748 110), 2 083 cases (0.2%) were proven to be BTV-4 infected. Total of 206 infected cattle and 24 infected goats were reported during this investigation period, which represents 0.06% and 0.03% of the total number of cattle and goats kept in affected areas, respectively. The highest incidence of infected sheep, cattle and goats was recorded on the territory covered by veterinary institute of Nis. Recorded lethality in cattle, sheep and goats was 18.45% (n=38), 48.10% (n=1002) and 54.17% (n=13), respectively. The peak of the outbreak was in September and October when 94.43% of the confirmed positive cases, regardless of the species, was recorded. Monitoring of bluetongue disease in Serbia relies on active surveillance programmes aimed at: (i) identification and tracing of susceptible and potentially infected animals and (ii) detection, distribution and prevalence of insect vectors. Vaccination of sheep is planned to be implemented as a control measure against bluetongue in Serbia.
Vladimir Dimitrijevic, Jevrosima Stevanovic, Mila Savic, Branko Petrujkic, Predrag Simeunovic, Ivan Milosevic and Zoran Stanimirovic
The aim of the study was to assess a commercially available microsatellite panel for use in paternity and identification analyses in the Yugoslavian Shepherd Dog Sharplanina, an ancient livestock guarding breed. Allele frequencies for 10 microsatellite loci (PEZ01, FHC2054, FHC2010, PEZ05, PEZ20, PEZ12, PEZ03, PEZ06, PEZ08 and FHC2079) were determined in 103 unrelated Yugoslavian Shepherd Dog individuals. The loci revealed varied levels of polymorphism (five to 12 alleles), with an average of 7.83 per locus. Average values of observed heterozygosity and polymorphic information content (PIC) were 0.64 and 0.66, respectively. Nine out of 10 microsatellite markers were highly informative with PIC values higher than 0.5. The obtained value of combined power of exclusion (0.9989) confirms usefulness of this panel of microsatellites for parentage verification, while the value of combined power of discrimination of 0.9999 clearly shows that the panel can conclusively identify individual dogs. In conclusion, the results obtained suggest that the selected set of commercially available microsatellite markers may be used as a routine tool for parentage verification and individual identification in the Yugoslavian Shepherd Dog. Although analysis of genetic variability of the Yugoslavian Shepherd Dog was not the primary focus of the study, the established values of major diversity indices disclose a highly variable gene pool in the breed.
In this study 55 honey bee colonies from different Serbian regions were monitored for the presence of Deformed Wing Virus (DWV) and Acute Bee Paralysis Virus (ABPV) using TaqMan-based real-time RT-PCR assay. The results revealed the presence of DWV in each sampling location, and ABPV in 10 out of 11 apiaries. High frequency of DWV (76.4%) and ABPV (61.8%) positive samples in asymptomatic colonies can be the consequence of inefficient and postponed Varroa treatment concerning the role of this mite in the transmission and activation of honey bee viruses. The real-time RTPCR technique described in this paper is proved to be the most reliable method for this kind of investigation.
Control of swine dysentery with antibiotics is often ineffective due to the resistance of Brachyspira hyodysenteriae. The potential of some herbal-based components against B. hyodysenteriae was previously studied in vitro. This study aims at the evaluation of in vivo efficacy of phytogenic feed additives in the control of swine dysentery
The study involved 64 seven-week old weaned pigs allotted to 4 groups: two were fed on feed supplemented with either Patente Herba® or Patente Herba® Plus, the third received tiamulin (positive control), while the negative control was not given antibiotics or additives. Fecal consistency was recorded daily. The presence of B. hyodysenteriae in the feces was investigated weekly using microbiological assays and the PCR test. Weight gain and feed conversion ratio were calculated for each week, and for the whole experiment.
B. hyodysenteriae was detected in all samples by both methods. The additives showed efficacy in the prevention and control of swine dysentery as only normal and soft stool was observed in the treated groups. By contrast, in the negative control all feces categories were detected. Frequencies of feces categories significantly differed (p<0.001) between feed-supplemented groups and the negative control. Efficacy of both additives in the prevention of SD is comparable to tiamulin, based on insignificant differences in the frequency of the various feces categories.
Beneficial effects of both additives resulted in significantly (p≤0.05) higher weight gain and lower feed conversion ratio in comparison to the negative control. The average weight gains between additive-fed groups and tiamulin-treated group did not differ significantly.
Honey bee mite Varroa destructor and microsporidium Nosema ceranae are currently considered the most important threats to honey bees and beekeeping. It has been believed that both N. apis and N. ceranae invade exclusively epithelial cells of the honey bee ventriculus. However, some fi ndings suggest that these microsporidia may infect other tissues of honey bees. There are indications that these pathogens could be found in honey bee haemolymph, as the medium for its distribution to anatomically distant tissues. Knowing that V. destructor being an ectoparasitic mite feeds on the honey bee’s haemolymph, the aim of this study was to investigate if DNA of Nosema spp. microsporidia could be found in honey bee haemolymph and in V. destructor.
The study was conducted on bee haemolymph and V. destructor mites from 44 Apis mellifera colonies. From each hive five mite individuals and 10 μL of haemolymph (from 4-5 bees) were used as samples for DNA isolation and PCR detection of Nosema spp.
The DNA of N. ceranae was confi rmed in 61.36% of V. destructor mites and 68.18% of haemolymph samples. This is the first report of N. ceranae DNA in honey bee haemolymph and in V. destructor mites. The finding of DNA of N. ceranae in V. destructor could be interpreted as the result of mite feeding on N. ceranae infected bee haemolymph. However, for a full confi rmation of the vector role of V. destructor in spreading of nosemosis, further microscopy investigations are required for the detection of spores in both investigated matrices (haemolymph and V. destructor internal tissues).