Generation of monoclonal antibodies specific for ORF68 of koi herpesvirus

Outbreaks of koi herpesvirus (KHV) infection in carp are still a serious problem worldwide. KHV is closely related to other two cyprinid herpesviruses, pox herpesvirus (CHV) and haematopoietic necrosis herpesvirus (CyHV-2) in goldfish. In this study, two major KHV antigenic proteins (ORF62 and ORF68) were identified by immunoscreening using a KHV-specific polyclonal antibody, and then monoclonal antibodies were generated for immunodiagnostic studies. After screening hybridoma cells, one mAb against ORF68 (mAb-7C6) was obtained but no mAbs against ORF62. mAb-7C6 specifically reacted with a lysate of KHV-infected koi fin cells (KF-1 cells) but not with lysates of CHV- or CyHV-2-infected KF-1 cells in an immuno-blotting analysis. Similar results were shown in the following tests: (1) a indirect fluorescent antibody test using infected KF-1 cells and (2) an immunohistochemical investigation by fast red stain (infected liver) or FITC detection (infected spleen). These results suggested that mAb-7C6 specifically reacts with KHV ORF68 protein.     

Full genome sequence and virulence analyses of the recent equine isolate of Japanese encephalitis virus

In the past 25 years, there has been only one case of Japanese encephalitis in horses in Japan. We determined the full genome sequence of the Japanese encephalitis virus (JEV) strain JEV/eq/Tottori/2003 isolated from an afflicted horse and also analyzed its virulence in mice. The sequence analysis showed that the genome of JEV/eq/Tottori/2003 is similar to that of genotype I, a dominant genotype of JEV presently circulating in Japan. Its neurovirulence, but not neuroinvasiveness, was still as high as it was for genotype III, thus indicating the necessity for continuation of a vaccination program of horses against JEV.     

Silage quality as influenced by concentration and type of tannins present in the material ensiled: A meta‐analysis

Protein degradation during ensiling is a major problem. Tannins are known to prevent or decelerate protein degradation in the rumen and may be able to do so in silages as well. Therefore, the present evaluation aimed to analyse the influence of tannins on silage quality. This was done by integrating from all suitable experiments found in literature on the topic in a meta‐analysis approach. A total of 122 datasets originating from 28 experiments obtained from 16 published articles and one own unpublished experiment were included in the database. Tannins in the silages originated either from the plants ensiled or from supplementations of tanniferous plants or tannins extracted from such plants. Tannin concentrations ranged from 0 to 57.8 g/kg dry matter, and the ensiling period varied from 30 to 130 days. The analysis was based on the linear mixed model methodology in which the different studies were considered as random effects and tannin‐related properties (either concentration or type of tannins) were treated as fixed effects. Results revealed that greater concentrations of tannins were associated with a decrease of butyrate concentration in the silages (p < 0.05). An increasing tannin concentration was also accompanied with smaller proportions of soluble N, free amino acid N, non‐protein nitrogen and NH₃‐N in total silage N (p < 0.05). The relationships between hydrolysable and condensed tannins and the decline in butyrate and NH₃‐N concentrations in the silages were of different magnitude (p < 0.05). A higher tannin concentration was associated with a decline in in vitro dry matter digestibility. It was concluded that tannins apparently have the ability to limit extensive proteolysis which may occur during ensiling and thus may improve the fermentative quality of silages. A desired side effect seems to be given by the tannins' apparent property to limit the activity of the butyrate‐producing microbes.     

Effects of livestock grazing on the spatial heterogeneity of net soil nitrogen mineralization in three types of Mongolian grasslands

Purpose

Small-scale soil heterogeneity relates to productivity and biodiversity and is crucial to understand. Soil heterogeneity could be affected by vegetation structure, and large mammal grazers could modify it through herbivory and excretion. The objective is to clarify the effects of livestock grazing on the small-scale (～3 m) soil heterogeneity in three types of Mongolian grasslands.

Materials and methods

We sampled soils from inside (ungrazed) and outside (grazed) exclosures in three vegetation types: forest-steppe, shrub-steppe, and desert-steppe. We measured laboratory rates of soil net nitrogen (N) mineralization and net nitrification and geostatistically analyzed heterogeneity.

Results and discussion

Average rates of net N mineralization and net nitrification were lower at shrub-steppe and desert-steppe and were decreased by grazing. Semivariograms showed vegetation-induced heterogeneity in ungrazed plots, except for net nitrification at forest-steppe. We found linear change with distance under dense and uniform vegetation at forest-steppe, 1.3 m patch under patchy vegetation at shrub-steppe, and linear change, but with much smaller semivariance, under sparse and poor vegetation at desert-steppe. At forest-steppe, grazing randomized the spatial patterns of net N mineralization and net nitrification. At shrub-steppe and desert-steppe, grazing greatly decreased the semivariances of net N mineralization and net nitrification as well as their averages, and the soil heterogeneity was virtually disappeared.

Conclusions

Grazing in Mongolian grasslands homogenized the spatial patterns of net N mineralization and net nitrification, irrespective of their original spatial patterns determined by the differences in vegetation structure.     

Sesbania spp., Aeschynomene indica and Crotalaria spp. are amide-exporters

Abstract

Extract

Leguminous plants consist of two groups, amide-exporting and ureide-exporting plants. The former legumes export a large fraction of fixed-N in the form of amides (asparagine and glutamine), and the latter legumes in the form of ureides (allantoic acid and allantoin). Another characteristic of the nodules is the enrichment in ¹⁵N. There are two types of legumes: one characterized by the enrichment with ¹⁵N in N₂-fixing nodules, in contrast to the other where the enrichment does not occur. The first investigation by Shearer et al. (1982) suggested that the nodules exporting fixed-N in the form of ureides were enriched in ¹⁵N unlike those exporting it in the form of amides. Soybeans, mungbean, and cowpea belong to the former group and groundnut, alfalfa, white clover to the latter. Although pea and faba bean were first classified into the latter group (Shearer et al. 1982), a recent investigation (Yoneyama 1988) showed that these nodules were also enriched in ¹⁵N.     

Occurrence of Guanidino Compounds in Several Plants

The stored nitrogen in perennial plants plays an important role in their regrowth in spring. Free L-arginine is one of the major forms of stored nitrogen (6, 9) .. LArginine is metabolized via the urea cycle in animal and plant tissues. In animal tissues, L-arginine is also metabolized via r-guanidinobutyramide, r-guanidinobutyric acid, and r-aminobutyric acid to the TCA cycle. In plant tissues, few attempts have been made to study arginine-metabolic routes other than the urea cycle. In white spruce tissues (3), L-arginine is metabolized via r-guanidinobutyric acid. Also in citrus tissues (4, 5), the operation of the same metabolic route has been reported. In these experiments, it was observed that various guanidino compounds reacted with the Sakaguchi reagent.

To obtain more information about the guanidino compounds involved in the arginine metabolism, the analytical system was evaluated and the presence of these compounds in several plant species was examined.     

Pathogenetic roles of beet necrotic yellow vein virus RNA5 in the exacerbation of symptoms and yield reduction,development of scab-like symptoms,and Rz1-resistance breaking in sugar beet

Beet necrotic yellow vein virus (BNYVV) generally has a four-segmented positive-sense RNA genome (RNAs 1–4), but some European and most Asian strains have an additional segment, RNA5. This study examined the effect of RNA5 and RNA3 on different sugar beet cultivars using a Polymyxa-mediated inoculation system under field and laboratory conditions. In field tests, the degree of sugar yield served as an index for assessing the virulence of BNYVV strains. Japanese A-II type isolates without RNA5 caused mostly 15%–90% sugar yield reductions, depending on the susceptibility of sugar beet cultivars, whereas the isolates with RNA5 induced more than 90% yield losses in the seven susceptible cultivars, but small yield losses in one Rz1-resistant and Rizor cultivars. However, a laboratory-produced isolate containing RNA5 but lacking RNA3 caused higher yield losses in Rizor than in susceptible plants, and induced scab-like symptoms on the root surface of both susceptible and resistant plants. In laboratory tests, A-II type isolates without RNA5 had low viral RNA accumulation levels in roots of Rizor and Rz1-resistant plants at early stages of infection, but in the presence of RNA5, viral RNA3 accumulation levels increased remarkably. This increased RNA3 accumulation was not observed in roots of the WB42 accession with the Rz2 gene. In contrast, the presence of RNA3 did not affect RNA5 accumulation levels. Collectively, this study demonstrated that RNA5 is involved in the development of scab-like symptoms and the enhancement of RNA3 accumulation, and suggests these characteristics of RNA5 are associated with Rz1-resistance breaking.     

Cell wall metabolism of pear fruit on the tree after 2,4-DP treatment

Summary

The effect of 2,4-dichlorophenoxy-propionic acid (2,4-DP) on fruit ripening on the tree was investigated in `La France' pears (Pyrus communis L.). Forty-five ppm, 2,4-DP solution was applied at 159 d after full bloom (DAFB), which was optimum harvest date for `La France' pears. They were compared with fruit harvested at 159 DAFB and placed in a controlled room with a 208C and 90% r.h. After harvest, the fruit firmness decreased dramatically 13 d after harvest (DAH). In 2,4±DP treatment, the levels decreased gradually 13 d after treatment (172 DAFB). In the untreated control, the levels also showed a slight decrease with time. Final ethylene concentration was almost the same in harvested and 2,4-DP treated fruit, but lower in the untreated control. In each treatment, water-soluble polyuronide (WSP) concentrations increased with time and hexametaphosphate-soluble polyuronide (HMP) and HCl-soluble polyuronide (HP) concentrations decreased. WSP concentration increased rapidly 20 DAH in harvested fruit and 194 DAFB in 2,4-DP treatment. Harvested and 2,4-DP treated fruit reached the edible condition on 17 DAH and 194 DAFB, respectively. Pectinesterase (PE) activity increased slightly with time for all treatments, showing no significant difference between treatments. Polygalacturonase (PG) activity increased gradually in all cases, being higher in 2,4-DP treated fruit than in control fruit left on the tree. These results suggest that 2,4-DP treatment before harvest induces pear fruit softening. This implies that 2,4-DP may be used as a replacement for low-temperature storage to promote the ripening, and good quality ripe fruit with buttery and juicy texture may be harvested on the tree.     

Changes of abscisic acid and its metabolite during development of apple fruit

Summary

Free and conjugated abscisic acid (ABA) and phaseic acid (PA) concentrations in the seed and pulp of apples (Malus pumila Mill. cv. Tsugaru) and ABA metabolism in the fruit after cis-(+)-ABA application were investigated. In the seed, the levels of cis- and trans-ABA and conjugated cis- and trans-ABA showed a general increase towards harvest, except for the decrease of cis-ABA at 131 d after full bloom (DAFB) (harvest). PA levels reached a peak at 41 DAFB, then decreased until harvest. The levels of free ABA in the seed were higher than those in the pulp. In the pulp, the levels of cis-ABA increased gradually from 20 DAFB and rapidly from 111 DAFB towards harvest. The levels of trans-ABA and conjugated trans-ABA also showed similar changes to cis-ABA, i.e. increased steeply from 111 DAFB. In contrast, the level of conjugated cis-ABA was the highest at 20 DAFB, then fell rapidly until 111 DAFB and then increased again towards harvest. PA showed the highest level at 20 DAFB, then decreased gradually until 111 DAFB and then increased again towards harvest. It is therefore assumed that not only cis-ABA but also its metabolite may be associated with the ripening of apple fruit. Cis- and trans-ABA and conjugated cis- and trans-ABA concentrations in cis-(+)-ABA treated fruit increased significantly on 11 d after ABA application (122 DAFB). However, at 20 d after cis-(+)-ABA application, only trans-ABA and conjugated trans-ABA showed higher levels than those of the untreated control, while the levels of cis-ABA and conjugated cis-ABA fell to the same levels as the untreated control. PA levels did not show significant increase on 11 and 20 d after cis-(+)-ABA application. These results suggest that cis-(+)-ABA treated exogenously to apple fruit may eventually metabolize to trans-ABA, conjugated trans-ABA and further metabolites.