Molecular tracing of the transmission routes of bois noir in Mediterranean vineyards of Montenegro and experimental evidence for the epidemiological role of Vitex agnus‐castus (Lamiaceae) and associated Hyalesthes obsoletus (Cixiidae)

Epidemiological aspects and transmission routes of bois noir (BN), a grapevine yellows disease induced by ‘Candidatus Phytoplasma solani’, have been exhaustively studied in the affected vineyards of continental Europe but not in the Mediterranean coastal zone. Because ‘Ca. Phytoplasma solani’ and its principal vector Hyalesthes obsoletus presumably originate from the Mediterranean, gaining knowledge of the epidemiological peculiarities of the disease in this area is essential for understanding its global spread and diversification, as well as for designing local management strategies. In this study, molecular epidemiology was applied to trace transmission pathways of ‘Ca. Phytoplasma solani’ in the Mediterranean vineyards of Montenegro, using multilocus sequence typing of tuf, vmp1 and stamp genes of the isolates associated with various hosts. Thus, ‘Ca. Phytoplasma solani’ was tracked from a tentative reservoir plant (inoculum source) through an associated vector population to the infected grapevine. Three pathways of transmission were documented, originating from Urtica dioica, Convolvulus arvensis and Vitex agnus‐castus; however, only the route originating from U. dioica was direct, whereas the latter two were overlapping and could be intermixed. Vitex agnus‐castus is a natural source of ‘Ca. Phytoplasma solani’, representing an important link in disease epidemiology in the Mediterranean and a possible origin of several genotypes occurring in central Europe. Experimental confirmation of the role of Vitex‐associated H. obsoletus in BN transmission in Montenegrin vineyards indicates its tentative role as a vector in the wide area of the Mediterranean, where some of the major wine‐producing regions are located.     

Gemmamyces bud blight of Picea pungens: a sudden disease outbreak in Central Europe

Due to its high tolerance to acid rain, the North American Colorado blue spruce (Picea pungens) has been among the most frequent substitute tree species used in reforestation programmes in spruce mountain forests in the Ore and Sudeten Mountains in the Czech Republic after destruction by sulphur dioxide pollution in the second half of the last century. This species was planted on more than 8800 ha of destroyed mountain forest stands in the Ore Mountains alone. New plantations prospered until a massive outbreak of the gemmamyces bud blight, first identified in 2009. The causal agent, Gemmamyces piceae, induced massive bud blight in infected trees – bud loss progressively increased to 70–80% or more, and because the trees were not able to replace the natural loss of older needles, they quickly withered and died. Many plantations disrupted by the pathogen had to be cut down. Although several resistant genotypes have been identified, the cultivation of this species in forest stands has no future in this area. Because this disease was also identified in several ornamental plantations in the area, the bud blight epidemic haunts public green space administrators and nursery and Christmas tree plantation owners. Other pathogens and pests, such as Lophodermium piceae, Sirococcus conigenus and Elatobium abietinum, have also been identified as involved in the damage to P. pungens.     

Integrative approach of histopathology and histomorphometry of common carp (Cyprinus carpio L.) organs as a marker of general fish health state in pond culture

The semi‐intensive system of common carp monoculture in earthen ponds is currently the most practiced fish rearing system in several countries in Eastern Europe. This system is based on the addition of supplemented feed to ponds to achieve faster growth. In this study, the general state of the fish health was monitored when three different types of feeds were used: grain cereals (CF), commercial pelleted feed and extruded feed. The markers used were: (i) the histopathological index (HP) of the main organs: gills, liver and kidney; and (ii) the histomorphometric parameters of the hepatocytes and enterocytes to assess the state of the digestive system in fish fed different types of supplemented feed. At the end of the experiment, the fish were in good health, since irreversible lesions were not noticed to a greater extent in any of the fish organs. There were also no other severe lesions present. However, the results showed differences in the HP indices of the liver and gills, as well as the total HP index, indicating a better health status of the fish in the CF group. The histomorphometric results showed larger cytoplasmic areas of hepatocytes in fish fed CF, which is attributed to glycogen accumulation and higher numbers of goblet cells in the distal intestines of fish fed compound feeds. This can be attributed to slight inflammation.     

Absorptive root areas of large pedunculate oak trees differing in health status along a road in South Bohemia,Czech Republic

The health status of 200- to 400-year-old trees growing in an area alongside a road was described in detail on the basis of evaluation of hazard trees in urban areas and analyzed using the acoustic scanning and modified earth impedance (MEI) methods. Acoustic analysis was used to characterize tree trunks and coarse roots; MEI was used to characterize the active absorptive root surfaces (the electric impedance of soils and sapwood were also included). Several groups of trees were distinguished according to the extent of damage (slight, serious or extreme), which was easily detectable from the MEI results. It was confirmed that both methods gave similar results, which also corresponds to the detailed observations. Instrumental measurements provided additional information about trunk and root injuries, which cannot be directly observed. A combination of both methods is recommended to achieve the most reliable results. This unique approach is applicable for testing eventual threats by old ill trees for safety reasons over the landscape.     

Examining the Relationships Between the Activities of 3H in Precipitation and 137Cs in Ground-Level Air in Belgrade City Area

Specific activity of tritium (³H) in precipitation and specific activity of ¹³⁷Cs in ground-level air were monitored at three locations in Belgrade (Meteorological Station of Belgrade at Zeleno Brdo (ZB), Meteorological Station Usek (USEK), and Vin?a Institute of Nuclear Sciences (VINS)). Data presented cover the period 1985?C1997 for ¹³⁷Cs for all locations and 1985?C2009 (ZB), 1988?C1997 (USEK), and 1988?C2009 (VINS) for ³H. Concentrations of ³H in precipitation have been determined using electrolytic enrichment and liquid scintillation spectrometer LKB-Wallac 1219 RackBeta. The activity of ¹³⁷Cs in air was determined on an HPGe detector (Canberra, relative efficiency 23?%). ³H concentrations in precipitation ranged from 0.40?±?0.08 to 74.6?±?5.2?Bq?l^?1 decreasing with distance from the nuclear facilities. Significantly higher tritium levels were measured in samples in VINS compared with those from an off-site location. The observed seasonal variations of tritium concentration indicate the stratospheric source of tritium. Increases in activity concentration of ¹³⁷Cs in the atmosphere were observed after the nuclear plant accident at Chernobyl in April 1986. The concentrations obtained for ¹³⁷Cs in 1986 were compared with the integrated air concentrations of ¹³⁷Cs in the region. The increases of ¹³⁷Cs air concentrations in 1987 and 1988 were attributed to local resuspensions from the ground. Since 1989, the activity level before the accident has been obtained. The average monthly concentrations of ¹³⁷Cs in ground-level air were shown spread maximum in spring?Csummer period and pronounced maximum during winter. The obtained results were statistically analyzed, i.e., the following parameters were determined: tritium deposition, monthly activities of ³H and ¹³⁷Cs, seasonal indices, radionuclide loading indices, and linear correlation coefficients.     

On determination of chitin content in mushrooms

Nitrogen compounds constitute the greatest proportion of the dry weight of mushrooms. These nitrogen compounds are partly protein and partly non-protein. A modified Heneberg-Stohman's method was used to separate non-protein from protein nitrogen, as well as to determine raw fiber content in mushrooms, since the methods used previously for this purpose are time-consuming and complicated. Elemental analysis of crude fiber was performed to confirm the uniformity of isolated products. The content of total crude fiber (fungin) and the proportion of chitin and cellulose in it were determined. The chitin content of 15 species of field grown edible mushrooms was in the range of 1.87–6.93% of total dry mass and 34.51–67.72% of isolated fungin.     

Microbiological identification and analysis of swine tonsils collected from carcasses at slaughter

The primary objective of this 7-month study was to determine the prevalence of porcine pathogens of the tonsil of the soft palate of swine at slaughter. Additional objectives were to determine if sampling the carcasses of normal or abnormal hogs provided different microbiological profiles and if the slaughter plant provides a feasible sampling frame and environment for detecting and monitoring important pathogens in tonsils that have health implications for both swine and humans. A total of 395 samples were collected from 264 farms. Of these, 180 tonsils were collected from normal carcasses and 215 tonsils were collected from carcasses that were diverted to the hold rail. Laboratory testing included bacteriological culture and identification as well as real time-polymerase chain reaction (PCR) testing for porcine reproductive and respiratory syndrome virus (PPRSV) and immunohistochemistry (IHC) for porcine circovirus-2 (PCV-2). The most commonly isolated bacteria included: Streptococcus suis (53.7%), Arcanobacterium pyogenes (29.9%), Pasteurella multocida (27.3%), and Streptococcus porcinus (19.5%). Virus screening revealed evidence of PRRSV and PCV-2 in 22.0% and 11.9% of the samples, respectively. Salmonella Typhimurium and Yersinia enterocolitica were isolated in 0.5% and 1.8% of the samples, respectively. Tonsils collected from the hold rail were more likely to be positive for Staphylococcus hyicus [odds ratio (OR) = 7.51, confidence interval (CI) = 2.89 to 19.54], Streptococcus porcinus (OR = 9.93, CI = 4.27 to 23.10), and Streptococcus suis (OR = 2.16, CI = 1.45 to 3.24). Tonsils collected from abnormal carcasses were less likely to be positive for Staphylococcus aureus (OR = 0.05, CI = 0.005 to 0.482).     

Comparison of bacterial culture, polymerase chain reaction, and a mix-enzyme-linked immunosorbent assay for the detection of Salmonella status in grow-to-finish pigs in western Canada with a Bayesian approach

Among grow-to-finish pigs from 10 herds in Alberta and Saskatchewan, 23 (16%) of 144 fecal samples were culture-positive and 40 (28%) of 144 pigs were seropositive for Salmonella. With a Bayesian model specifying dependence between the 2 tests, the sensitivity (Se) of culture and real-time polymerase chain reaction (RT-PCR) was 79% to 86%, depending on the cut-off value for the enzyme-linked immunosorbent assay (ELISA). Culture specificity (Sp) was assumed to be 100%; RT-PCR Sp was found to be 94%. The ELISA Se was 76% and 51% at optical density cut-off values ≥ 20% and ≥ 40%, respectively; the Sp was 94% at each cut-off value. The model showed some sensitivity to ELISA prior information, the ELISA Se being approximately 8% lower when informative prior information was specified in the model. When there was no adjustment for dependence between culture and RT-PCR, the posterior estimates for both culture and RT-PCR Se were 11% higher than with the conditional-dependence model and had considerably narrower probability intervals, which suggests that correlation between culture and PCR is important and should be adjusted for in future studies.     

Toxoplasma gondii infection in slaughter pigs in Serbia: seroprevalence and demonstration of parasites in blood

ABSTRACT: A seroepizootiological study of Toxoplasma gondii infection involving a total of 488 slaughter pigs (468 market-weight pigs and 20 sows) in the Belgrade area, also included examination of the presence of T. gondii in the blood. Blood sampled at the slaughter line was examined for specific antibodies by modified direct agglutination, and blood clots of those seropositive at titres of 1:50-1:12800 were bioassayed in mice. The overall seroprevalence was 9.2%, significantly higher (p = 0.0063) in sows (30.0%) than in market-weight pigs (8.3%). Amongst the 22 bioassays performed, a total of 16 (72.7%) were positive, by observation of T. gondii cysts (12), seropositivity (7, including 3 in which cysts were not detected), and/or detection of T. gondii DNA by real-time PCR (12, including one otherwise negative). The positive bioassays originated from the blood of 12 market-weight pigs and 4 sows. Despite a general increase in the rate of demonstration of T. gondii with the increase in the specific antibody level, the association was not significant (p = 0.101). The risk of infection was 41-fold increased in sows vs market-weight pigs, and 15-fold in pigs from smallholders' finishing type farms vs those from large farrow-to-finish farms. The presence of viable T. gondii in a proportion of the samples indicates that some of the pigs had an active parasitaemia at the time of slaughter, which, along with the seroprevalence established, points to a potential source of human infection in Serbia. This is the first report on parasitaemia in naturally infected swine.     

Ivermectin effects on motor coordination and contractions of isolated rat diaphragm

Ivermectin, the antiparasitic drug from the macrocyclic lactones class raises attention due to its high efficiency against nematodes and arthropods and very specific toxic and side effects that it may produce in host. Dominant clinical symptoms of adverse effects and toxicity of ivermectin in animals are tremor, ataxia, CNS depression and coma which often results in mortality. In our study increasing intravenous doses of ivermectin, (6 or more times higher than therapeutic dose: 1.25, 2.5, 3.75, 5.0, 6.25 and 7.5 mg/kg), caused dose-dependent disturbance of motor coordination in treated rats. The median effective dose (ED50) that was able to impair the rota-rod performance in rats treated 3 min before testing was 2.52 mg/kg. This effect weakens over time, while in the rats treated 60 min before the rota-rod test, ED50 of ivermectin was 4.21 mg/kg. Whereas, all tested doses of ivermectin did not cause any other clinical symptoms of toxicity. Ivermectin has no effect on the contractions of isolated diaphragm caused by the EFS, which effectively blocked mecamylamine (100 μM) and pancuronium (1 and 2 μM). Effect on motor coordination is the first detectable clinical symptom of ivermectin toxicity and apparently is a result of its central effects.