Identification of major immunogenic proteins of Mycoplasma synoviae isolates

Mycoplasma synoviae isolates differ in patterns of immunogenic proteins, but most of them have not been identified yet. The main aim of this study was their identification in two closely related M. synoviae isolates, ULB 02/P4 and ULB 02/OV6, recovered recently from chickens in Slovenia. N-terminal sequencing identified 17 M. synoviae proteins. Amongst them were 14 major, highly expressed but previously unidentified proteins, including enzymes, chaperones and putative lipoproteins. ULB 02/P4 proteins with increasing molecular weight (M(w)) in the region above the lipoprotein MSPB (approximately 40 kDa) were elongation factor EF-Tu, enolase, NADH oxidase, haemagglutinin MSPA, ATP synthase beta chain, trigger factor, pyruvate kinase and chaperone DnaK. Enolase (approximately 47 kDa) seemed to be immunogenic for chickens infected with M. synoviae, whereas EF-Tu, which might cross-react with antibodies to the P1 adhesin of Mycoplasma pneumoniae, was not. ULB 02/OV6 synthesized several immunogenic proteins and those with M(w) of approximately 70, 78, 82, 90, 110 and 160 kDa, cross-reacted with antibodies to Mycoplasma gallisepticum. They remain to be identified, because besides putative lipoproteins, protein bands of 78, 82, 85 and 110 kDa contained also dehydrogenase PdhD, elongation factor EF-G, enzyme PtsG and putative neuraminidase, respectively.     

Effects of cover crops on soil mineral nitrogen and on the yield and nitrogen content of maize

Current agricultural practice favours winter cover crops, which can not only optimize N management in field crop rotation; but also affect subsequent crops. Three field experiments were carried out in Eastern Slovenia to examine the effects of Italian ryegrass (Lolium multiflorum Lam.), winter rape (Brassica napus ssp.oleifera (Metzg.) Sinsk), subclover (Trifolium subterraneum L.), and crimson clover (Trifolium incarnatum L.) as winter cover crops on the mineral N (N_min) content of soil and on the yield and N content of subsequent maize (Zea mays L.), fertilized with 120 kg N ha⁻¹. Italian ryegrass and winter rape decreased soil N_min contents before winter and in spring more than both clovers. In contrast, clovers accumulated significantly higher amounts of N in organic matter and had lower C/N ratios than winter rape and especially Italian ryegrass. In comparison to the control (bare fallow without cover crop), clovers increased the whole above ground maize dry matter yield, maize grain yield and N contents in whole above ground plants and in grain. The yields and N contents of maize following winter rape were on the same level as the control, while yields and N contents of maize following Italian ryegrass were, in two of the experiments, at the same level as the control. The effects of Italian ryegrass on the maize as subsequent crop in the third experiment were markedly negative. Maize in the control treatment exploited N much more efficiently than in treatments with cover crops. Therefore, cover crop N management should be improved, especially with a view to optimizing the timing of net N mineralization in accordance with the N demands of the subsequent crop.     

Field efficacy of different vaccines against infectious bursal disease in broiler flocks

A field study was performed to determine the efficacy of three commercially available vaccines against infectious bursal disease (IBD) in commercial broilers raised in a high IBD virus (IBDV) risk area. Live attenuated intermediate and intermediate plus vaccines were used in four flocks. Birds were vaccinated orally at the estimated vaccination time. Three broiler flocks were vaccinated subcutaneously with a turkey herpesvirus (HVT)-IBD vector vaccine at one day old. Evaluation of the efficacy of different vaccines was focused on humoral immune response, bursa/body weight (B/Bw) ratio, molecular detection of IBDV in ileocaecal tonsils and bursa of Fabricius, and production parameters. The serological results showed that although the uptake of all three vaccine strains was confirmed in the lymphoid organs, no significant antibody response to vaccination was detected in flocks vaccinated with intermediate and intermediate plus vaccines. A significant increase in antibody titres detected in flocks vaccinated with the vector vaccine indicated its ability to induce an immune response in birds with a high level of maternally derived antibodies. Observations obtained in this field trial did not confirm the expected reduction of the B/Bw ratio in flocks vaccinated with less attenuated vaccines. No significant differences were observed between birds vaccinated with the vector vaccine and those immunised with the intermediate plus vaccine. Very virulent IBDV was confirmed in the flock vaccinated with the intermediate vaccine. The infection induced reduced B/Bw and moderate mortality but did not affect the production parameters. Field infection was not detected in broilers vaccinated with the intermediate plus vaccine and the vector vaccine.     

Prevalence of pigeon circovirus infections in feral pigeons in Ljubljana, Slovenia

Pigeon circovirus (PiCV) was detected by real-time PCR in cloacal swabs, pharyngeal swabs, and serum samples taken from 74 feral pigeons (Columba livia var. domestica) that were caught at various locations in the city of Ljubljana, Slovenia. PiCV infections were detected in the majority of the tested birds. The highest (74.3%) detection rate was observed in the cloacal swabs and the lowest (31.1%) in serum samples. PiCV DNA was more readily detected in the cloacal swabs, pharyngeal swabs, and serum samples of birds younger than 1 yr. Molecular analysis of partial open reading frame V1 sequences showed that PiCV strains detected in feral pigeons share high nucleotide and amino acid sequence identities with PiCV strains detected in ornamental, racing, meat, and feral pigeons.     

Molecular characterization of avian paramyxovirus type 1 (Newcastle disease) viruses isolated from pigeons between 2000 and 2008 in Slovenia

Fourteen avian paramyxovirus type 1 (APMV-1; Newcastle disease) viruses isolated from dead free-living and domestic pigeons in Slovenia between 2000 and 2008 were analyzed by a molecular characterization of a part of the fusion protein gene, which included the region encoding the fusion protein cleavage site. Phylogenetic analysis indicated that the Slovene pigeon paramyxovirus type 1 (PPMV-1) viruses do not cluster together but instead are divided into two groups--4bi and 4bii--of sublineage 4b. Nine Slovenian strains were placed in group 4bii. Five other strains clustered together with PPMV-1 from group 4bi. The sequence of the fusion protein cleavage site of all Slovenian strains was typical for pathogenic APMV-1. The 112RRQKRF117 motif was present in the strains from group 4bii, whereas strains from group 4bi displayed the 112GRQKRF117 motif.     

Molecular characterisation of infectious bursal disease viruses isolated in recent acute outbreaks in Slovenia

In 2004 and then in 2006 several outbreaks of infectious bursal disease (IBD) were reported in broiler and broiler breeder flocks in Slovenia. In this report ten recently emerged IBD viruses (IBDV) were characterised by sequence analysis of the VP2 hypervariable region and compared to previous Slovene IBDV strains from 1995/1996 and to some representative serotype 1 IBDV strains of different pathotypes. On the basis of nucleotide and amino acid identities, phylogenetic analyses and the presence of very virulent IBDV (vvIBDV) conserved amino acid substitutions, all Slovene isolates from recent outbreaks were identified as vvIBDV. Although some unique nucleotide exchanges and amino acid substitutions have been observed, the results of this study indicated that recent vvIBDV isolates are closely related with those from outbreaks in the 1990s. However, acute IBD has not been reported in commercial flocks in Slovenia for some years. This could lead to the conclusion that poor biosecurity and relaxed vaccination could be responsible for the re-emergence of vvIBDV.     

A survey of avian Mycoplasma species for neuraminidase enzymatic activity

Among 23 currently recognized avian Mycoplasma (AM) species only Mycoplasma gallisepticum, Mycoplasma synoviae, Mycoplasma meleagridis and Mycoplasma iowae cause disease and loss of production in chickens and/or turkeys. Because neuraminidases are considered virulence factors in many pathogenic microorganisms the aim of our study was to determine which AM species possess neuraminidase enzymatic activity (NEAC). Small samples of AM cells were assayed for NEAC using the chromogenic substrate 5-bromo-4-chloro-3-indolyl-alpha-d-N-acetylneuraminic acid. In the case of positive NEAC reaction the substrate gave the insoluble indigoblue product what enabled simple test and easy estimation of NEAC. M. gallisepticum and M. synoviae which share sequences of the gene encoding neuraminidase (sialidase NanH) exhibited considerable levels of NEAC. However, NEAC levels differed among their strains, as well as among cultures of different strains. Only certain cultures of the type strain of M. meleagridis showed NEAC, whereas among six serovars of M. iowae only serovar I (type strain 695) showed NEAC. Weak NEAC was detectable in M. anseris, M. cloacale and M. pullorum, whereas the type strain of M. corogypsi (BV1) showed strong NEAC. Our study provides novel informations about NEAC in AM species and suggests that higher invasiveness and possibly, the pathological processes might be associated with their NEAC.     

Influence of oxygen plasma pre-treatment on the water repellency of cotton fibers coated with perfluoroalkyl-functionalized polysilsesquioxane

Oxygen plasma pre-treatment was applied to cotton fabric with the aim of improving the water repellency performance of an inorganic-organic hybrid sol-gel perfluoroalkyl-functionalized polysilsesquioxane coating. Cotton fabric was pre-treated with low-pressure oxygen plasma for different treatment times and operating powers. Afterward, 1H,1H,2H,2H-perfluorooctyltriethoxysilane (SiF) was applied to the cotton fabric samples using the pad-dry-cure method. The surfaces of the untreated and modified cotton fibers were characterised using Fourier transform infrared spectroscopy, Xray photoelectron spectroscopy, scanning electron microscopy, and atomic force microscopy. The water repellency of the SiF-coated fabric samples was evaluated using static and sliding contact angle measurements with water. The results show that the plasma treatment with the shortest treatment time (10 s) and the lowest operating current (0.3 A) increased the atomic oxygen/carbon ratio of the cotton fiber surface from 0.6 to 0.8 and induced the formation of a nano-sized grainy surface. Increasing the plasma treatment time and/or operating current did not intensify the surface changes of the cotton fibers. Such saturation effects were explained by the large influence of reactive oxygen atoms during the plasma treatment. The measured static water contact angles on the surface of the untreated and plasma pre-treated and SiF-coated cotton fabrics showed that the oxygen plasma pre-treatment enabled the increase of the water contact angle from 135° to ≈150°, regardless of the applied plasma treatment time and discharge power. This improvement in the hydrophobicity of the SiF coating was followed by a decrease in the sliding angle of water droplets by more than 10° compared to the plasma untreated and SiF-coated sample characterized by a water sliding angle of 45°. Additionally, measurements of the water sliding angle revealed that the increase of the static contact angle from 149° to 150° corresponded to a drop of the water sliding angle from 33 to 24°, which suggests that the plasma pre-treatment of 20 s at an operating current of 0.3 A produced the best water-repellent performance of the SiF-coated cotton fabric.     

Biofortification of wheat cultivars with selenium

ABSTRACT

Biofortification experiments with three winter wheat cultivars treated with sodium selenate through foliar- and soil-fertilisation were conducted at two locations in Croatia and Serbia in two consecutive years to increase the selenium (Se) concentration in bread-making wheat grain. The treatments were: (a) 5?g?ha^?1 Se foliar-, (b) 10?g?ha^?1 Se foliar- and (c) 10?g?ha^?1 Se in soil surface-application and they were compared with (d) control. Both Se foliar- and soil-fertilisation increased the Se concentration in grains from 2.6- to 4.6-fold. The concentration in grain was highest with Se foliar-fertilisation of 10?g?ha^?1 and it was increased by 29–32?µg Se kg^?1 dry weight for each gram of Se applied per ha. The wheat cultivars differed in grain yield and Se uptake (g?ha^?1 Se). However, on average, there were no differences between wheat cultivars with respect to Se grain concentrations. Agronomic use efficiency (by grain) was significantly higher for Se foliar- (19%) than for soil-fertilisation (13%). It can be concluded that agronomic biofortification of winter wheat can be effective in increasing Se grain concentration, where the efficiency depends on the rate of Se applied, application method and local environmental conditions rather than on cultivar differences.