Molecular and phylogenetic analysis of the H5N1 avian influenza virus caused the first highly pathogenic avian influenza outbreak in poultry in the Czech Republic in 2007

On 19th July 2007 re-occurrence of the H5N1 highly pathogenic avian influenza (HPAI) virus was noticed in Europe. The index strain of this novel H5N1 lineage was identified in the Czech Republic where it caused historically the first HPAI outbreak in commercial poultry. In the present study we performed molecular and phylogenetic analysis of the index strain of the re-emerging H5N1 virus lineage along with the Czech and the Slovak H5N1 strains collected in 2006 and established the evolutionary relationships to additional viruses circulated in Europe in 2005-2006. Our analysis revealed that the Czech and the Slovak H5N1 viruses collected during 2006 were separated into two sub-clades 2.2.1 and 2.2.2, which predominated in Europe during 2005-2006. On the contrary the newly emerged H5N1 viruses belonged to a clearly distinguishable sub-clade 2.2.3. Within the sub-clade 2.2.3 the Czech H5N1 strains showed the closest relationships to the simultaneously circulated viruses from Germany, Romania and Russia (Krasnodar) in 2007 and were further clustered with the viruses from Afghanistan and Mongolia circulated in 2006. The origin of the Czech 2007 H5N1 HPAI strains was also discussed.     

Suppressive effect of pectic polysaccharides from Cucurbita pepo L. var. Styriaca on citric acid-induced cough reflex in guinea pigs

Several water-soluble pectic polysaccharides were isolated from the pumpkin fruit biomass and characterized by composition, structural features and molecular properties. The pectic polysaccharides were tested for antitussive activity by studying the effects of citric acid-induced cough reflex in guinea pigs and reactivity of the airway smooth muscle in vivo conditions in comparison to the narcotic drug codeine. Oral administration of all pectic polysaccharides from pumpkin inhibited the number of coughs induced by citric acid in guinea pigs, but to various extents. The results indicated that the antitussive activity of the pectic polysaccharides is affected by their molecular and structural properties, whereby a synergistic action between the polysaccharide and non-carbohydrate components on the biological response has been suggested as well. The cough depressive efficacy of most of the tested polysaccharides was comparable and even higher than that of codeine. Moreover, the application of these polysaccharides provoked any side effects what is their advantage towards the conventional opioid-derived antitussive agents.     

In sacco evaluation of ruminal degradability of isoflavones from full-fat soybean and extracted soybean meal—A pilot study

The aim of the study was to determine the ruminal degradability of dry matter (DM), daidzein, genistein, glycitein and total isoflavones in ground full-fat soybean (GFFS) and solvent-extracted soybean meal (SSBM) using the in sacco method. The experiment was carried out in three replications on ruminally cannulated sheep that were fed twice a day with a diet consisted of hay and supplemental mixture (6:4, DM basis). The nylon bags with 2 g feed samples ground to 2 mm were incubated in the rumen for 0, 2, 4, 8, 16 and 24 h. The effective degradability (ED) of DM, daidzein, genistein, glycitein and total isoflavones was calculated at outflow rate of 0.06 h. The ED of DM in GFFS was 77.8% and was higher than in SSBM being 71.8% (p < 0.001). The ED of daidzein (96.8%) and genistein (93.6%) was higher for SSBM compared with GFFS (93.9% and 92.8%, p < 0.001 and p = 0.003, respectively) while ED of glycitein was lower for SSBM than for GFFS (75.5 and 81.7%, respectively, p < 0.001). All isoflavones in the incubations were extensively degraded in the rumen, and regardless of dietary source, they were almost completely degraded after 16 h of incubation. Further, the disappearance patterns, that is the functions describing the time courses of the analyte disappearance, were assessed. The disappearance patterns of daidzein, genistein, glycitein and total isoflavones were similar and showed greater disappearance of mentioned isoflavones from SSBM compared to GFFS (p < 0.001 for daidzein, genistein and total isoflavones and p = 0.002 for glycitein). The study provides knowledge on the effect of processing on degradability of isoflavones in rumen that can be used to clarify the interrelationship between isoflavones and rumen microbiota.     

Amino acid profile of escaped feed protein after rumen incubation and their intestinal digestibility.

The crude protein content and amino acid profile of seven feedstuffs (linseed meal, maize gluten meal, rapeseed meal, rapeseed meal protected, soybean meal, fullfat soybean extruded and sunflower meal) were determined before and after ruminal incubation for 16 h in three bulls with large rumen cannulas. The intestinal disappearance of amino acids was measured using mobile bag technique. Ruminal incubation affected amino acid profile of all experimental feedstuffs. Crude protein degradation varied from 29.3% for maize gluten meal to 86.4% for rapeseed meal. A tendency towards increased disappearance was observed for glutamic acid, histidine, lysine and proline and decreased disappearance for branched-chain amino acids. The intestinal crude protein digestibility was higher than > 80%, except rapeseed meal (66.4%) and sunflower meal (77.8%). The least digestible individual amino acids were methionine and isoleucine in rapeseed meal, histidine and methionine in rapeseed meal protected and arginine in sunflower meal. In general, the lowest amino acid digestibilities were found in feedstuffs with the highest fibre content. The feedstuffs show that they have different potential for supplying of limiting amino acids. Of particular value are the feedstuffs with low crude protein degradability in the rumen and high intestinal digestibility of amino acids.     

Environmental mycobacteria closely related to the pathogenic species evidenced in an acidic forest wetland

The diversity of environmental mycobacteria was studied in water-logged acidic forest soil. Mycobacteria were assessed in upper and lower soil horizons and summer and winter seasons using T-RFLP and sequencing of 16S rRNA gene produced with Mycobacterium-specific primers. Mycobacteria diversity differed between both the two seasons and soil horizons. Cloning revealed the presence of mycobacteria belonging to three major clusters recognized within the genus, i.e. fast-growing, intermediate, and slow-growing species, with unprecedented abundance and diversity of the latter. Two novel clusters of sequences unrelated to the known mycobacteria were identified. This study raises the possibility that forest wetlands could serve as environmental reservoirs for an unexplored diversity of mycobacteria including those related to pathogenic species.     

Improved strategy for phylogenetic analysis of classical swine fever virus based on full-length E2 encoding sequences

ABSTRACT: Molecular epidemiology has proven to be an essential tool in the control of classical swine fever (CSF) and its use has significantly increased during the past two decades. Phylogenetic analysis is a prerequisite for virus tracing and thus allows implementing more effective control measures. So far, fragments of the 5′NTR (150 nucleotides, nt) and the E2 gene (190 nt) have frequently been used for phylogenetic analyses. The short sequence lengths represent a limiting factor for differentiation of closely related isolates and also for confidence levels of proposed CSFV groups and subgroups. In this study, we used a set of 33 CSFV isolates in order to determine the nucleotide sequences of a 3508-3510 nt region within the 5′ terminal third of the viral genome. Including 22 additional sequences from GenBank database different regions of the genome, comprising the formerly used short 5′NTR and E2 fragments as well as the genomic regions encoding the individual viral proteins Npro, C, Erns, E1, and E2, were compared with respect to variability and suitability for phylogenetic analysis. Full-length E2 encoding sequences (1119 nt) proved to be most suitable for reliable and statistically significant phylogeny and analyses revealed results as good as obtained with the much longer entire 5′NTR-E2 sequences. This strategy is therefore recommended by the EU and OIE Reference Laboratory for CSF as it provides a solid and improved basis for CSFV molecular epidemiology. Finally, the power of this method is illustrated by the phylogenetic analysis of closely related CSFV isolates from a recent outbreak in Lithuania.     

The effects of sage extract feed supplementation on biochemical parameters, weight of internal organs and Salmonella counts in chickens

We investigated the effects of dietary addition of sage extract on the biochemical parameters, weight of some body organs and changes in the counts of Salmonella Enteritidis PT4 (SE) in experimentally infected chickens. The following diets were used: basal diet, basal diet with addition of an extract of Salvia officinalis L. (S), basal diet and SE, and basal diet and S and SE (SSE). Compared to the SE group, sage extract in the SSE group decreased activities of ALP and ALT and concentrations of glucose and bilirubin on the 4th day post inoculation (p.i.). However, on the 18th day p.i., lower levels of bilirubin and ALT activity only were detected. Addition of sage extract to the diets decreased the counts of Salmonella in the liver, spleen and caecum at both sampling times, along with lower production of mucus in the chickens' intestines. Our results suggest that the addition of sage extract to the diet could be effective in protecting SE-infected chickens.     

Colonization of central European abandoned fields by dry grassland species depends on the species richness of the source habitats: a new approach for measuring habitat isolation

Abandoned fields are perceived as potential habitats for species of threatened semi-natural dry grasslands. However, information is lacking regarding how the spontaneous colonization of abandoned fields depends on the broader spatial context. We recorded the occurrence of 87 target species in 46 abandoned fields and 339 dry grasslands. We tested the effect of the isolation of abandoned fields from source grasslands on the number of dry grassland species occurring in abandoned fields either with or without habitat characteristics being used as covariates. The isolation of the fields was calculated using the distance and area (I _A ) or distance and species richness (I _S ) of source habitats. I _S always explained the number of grassland species in the abandoned fields better than I _A . The effect of isolation became smaller or even non-significant with the inclusion of covariates; it also changed with the method used for measuring distance (edge-to-edge or center-to-center), and it was lower when other abandoned fields were considered as additional source habitats. The different performance of the two isolation measures can be explained by the weak species–area relationship in the grasslands, indicating differences in their habitat quality. Species richness is a better proxy of habitat importance in terms of propagule source than habitat area, and the new isolation measure is therefore suitable for studying the effects of landscape structure on species richness in landscapes presenting a weak species–area relationship, such as areas exhibiting pronounced effects of land-use history. Inclusion of habitat characteristics as covariates may considerably alter conclusions regarding the effect of isolation, which might actually be overestimated when assessed separately.     

Soil moisture content as a predictor of soil disturbance caused by wheeled forest harvesting machines on soils of the Western Carpathians

Limiting surface soil disturbance caused by forest harvesting machines is an important task and is influenced by the selection of efficient and reliable predictors of such disturbance. Our objective was to determine whether soil moisture content affects soil load bearing capacity and the formation of ruts. Measurements were conducted in six forest stands where various machines operated. We measured the formation of ruts along skid trails in connection with varying soil moisture content. Soil moisture content was determined through the gravimetric sampling method. Our results showed that severe(rut depth16–25 cm) to very severe disturbance(rut depth [26 cm)occurred in forest stands where the instantaneous soil moisture exceeded its plasticity limits defined through Atterberg limits. Atterberg limits of soil plasticity ranged from 26 to 32 % in individual stands. Regression and correlation analysis confirmed a moderately strong relationship(R = 0.52; p \ 0.05) between soil moisture content and average rut depth. This confirmed that soil moisture is a suitable and effective predictor of soil disturbance.