Plant viruses and viroids in Japan

Journal of General Plant Pathology - An increasing number of plant viruses and viroids have been reported from all over the world due largely to metavirogenomics approaches with technological...     

Bilateral cataract in a cynomolgus monkey

Severe bilateral cataract was found in a 7 year-old naïve female cynomolgus monkey (Macaca fascicularis) 3 months before necropsy. During macroscopic examination, severe opacity and thinning of the lens were observed in both eyes. Histopathology revealed that the lens nuclei and majority of cortex lens fibers had disappeared and become excavated, while the lens fibers in the subcapsular area were swollen and distorted. Other observations included atrophy and vacuolation in the lens epithelial cells and proliferation of spindle cells and collagen fiber beneath the anterior capsule of the right eye. Immunohistochemical staining of these spindle cells revealed the presence of vimentin, cytokeratin and α-smooth muscle actin (α-SMA), which were considered to be derived from lens epithelial cells. This is a rare case of spontaneous, bilateral, hypermature cataract in a cynomolgus monkey.     

Efficient virus-induced gene silencing in apple, pear and Japanese pear using Apple latent spherical virus vectors

Background

Virus-induced gene silencing (VIGS) is an effective technology for the analysis of gene functions in plants. Though there are many reports on virus vectors for VIGS in plants, no VIGS vectors available for Rosaceae fruit trees were reported so far. We present an effective VIGS system in apple, pear, and Japanese pear using Apple latent spherical virus (ALSV) vectors.

Results

Inoculation of ALSV vectors carrying a partial sequence of endogenous genes from apple [ribulose-1, 5-bisphosphate carboxylase small subunit (rbcS), alpha subunit of chloroplast chaperonin (CPN60a), elongation factor 1 alpha (EF-1a), or actin] to the cotyledons of seeds by a particle bombardment induced highly uniform knock-down phenotypes of each gene on the true leaves of seedlings from 2~3 weeks after inoculation. These silencing phenotypes continued for several months. Northern blot and RT-PCR analyses of leaves infected with ALSV containing a fragment of rbcS gene showed that the levels of rbcS-mRNA drastically decreased in the infected apple and pear leaves, and, in reverse, rbcS-siRNAs were generated in the infected leaves. In addition, some of apple seedlings inoculated with ALSV vector carrying a partial sequence of a TERMINAL FLOWER 1 gene of apple (MdTFL1) showed precocious flowering which is expected as a knock-down phenotype of the silencing of MdTFL1 gene.

Conclusions

The ALSV-based VIGS system developed have provides a valuable new addition to the tool box for functional genomics in apple, pear, and Japanese pear.     

Limb and skin abnormalities in mice lacking IKKalpha

The gene encoding inhibitor of kappa B (IkappaB) kinase alpha (IKKalpha; also called IKK1) was disrupted by gene targeting. IKKalpha-deficient mice died perinatally. In IKKalpha-deficient fetuses, limb outgrowth was severely impaired despite unaffected skeletal development. The epidermal cells in IKKalpha-deficient fetuses were highly proliferative with dysregulated epidermal differentiation. In the basal layer, degradation of IkappaB and nuclear localization of nuclear factor kappa B (NF-kappaB) were not observed. Thus, IKKalpha is essential for NF-kappaB activation in the limb and skin during embryogenesis. In contrast, there was no impairment of NF-kappaB activation induced by either interleukin-1 or tumor necrosis factor-alpha in IKKalpha-deficient embryonic fibroblasts and thymocytes, indicating that IKKalpha is not essential for cytokine-induced activation of NF-kappaB.     

A survey of Blastocystis sp. in livestock,pets, and zoo animals in Japan

The prevalence of Blastocystis sp. was examined in fecal samples collected from cattle, pigs, dogs, and a variety of zoo animals (primates, carnivores, herbivores, pheasants, and ducks) by direct observation of fresh fecal suspensions or cultured materials, using light microscopy. The cattle and pigs were randomly sampled from 11 and 12 commercial farms, respectively, located in the western region of Japan. The dog material used in this study was obtained from pets housed in an animal shelter in the city of Osaka. Zoo animals were chosen based on housing conditions that minimized the possibility of intra-zoo transmission of the organism. The prevalence rate among the groups varied greatly. A high prevalence of infection was observed in the farm animal group, ranging from 95% (58/61) in pigs to 71% (39/55) in cattle, whereas the dog fecal samples were completely free of the organism. Prevalence of the organism in the zoo animal were 85% (29/34) in primates, 80% (8/10) in pheasants, 56% (9/16) in ducks, and 0% (0/58) in various carnivores and herbivores. Among the zoo animals infected with Blastocystis, eight species of primates, eight species of pheasants, and four species of ducks were confirmed as new hosts. Since Blastocystis organisms isolated from various animals were morphologically indistinguishable from Blastocystis hominis by light microscopy, further genomic studies are required for analysis of the zoonotic potential or etiological significance of these isolates.     

Measurement of ferritin and anti‐ferritin autoantibodies in serum and colostrum of Holstein and Japanese Black cows

Anti‐ferritin autoantibody is a ferritin‐binding protein commonly found in mammals; it is thought to form an immune complex with ferritin and thereby mediate the rapid clearance of circulating ferritin. The aim of this study is to determine concentrations of ferritin and anti‐ferritin autoantibodies (immunoglobulin (Ig)M, IgG and IgA) in serum and colostrum of Holstein (H) and Japanese Black (JB) cows within 24 h of normal calving. Blood and colostrum samples were collected from cows of various ages (2–11 years) and calving number (1–8 live births). Mean ferritin concentrations were higher in colostrum than in serum for both breeds, and higher colostrum ferritin concentrations were found in H than JB cows. IgA antibodies in serum and colostrum from both breeds had negligible ferritin‐binding activity. For both breeds, IgM and IgG antibodies had higher ferritin‐binding activity in colostrum than in serum. There was a significant correlation between IgM and IgG ferritin‐binding activities in serum and colostrum of H and JB cows. These results suggest that ferritin and IgM and IgG autoantibodies are actively transferred from the blood stream to the colostrum at prepartum or early lactation.     

Antioxidant activity in astringent and non-astringent persimmons

Summary

Catechins, ascorbic acid and -cryptoxanthin concentrations during fruit development and the antioxidant activity in skin and flesh were investigated in astringent ‘Saijyo’ and non-astringent ‘Fuyu’ persimmons (Diospyros kaki Thunb.). The IC₅₀ values (a midpoint of 50% between zero and full inhibition of diazo dye formation) of superoxide (O₂^–)-scavenging activity remained low in the skin throughout fruit development.The IC₅₀ values of the activity increased with d after full bloom (DAFB) in the flesh of the non-astringent type, whereas values remained low until harvest in the astringent type. However, IC₅₀ values increased sharply after the removal of astringency with ethanol spray. Catechin concentrations in the flesh also decreased after the removal of astringency, whereas the concentrations in the skin did not decrease. Ascorbic acid concentrations in the skin and flesh were high at the beginning and middle of fruit development, but -cryptoxanthin in the skin and flesh increased toward harvest. However, -cryptoxanthin concentrations in the skin and flesh were lower than catechins and ascorbic acid. In addition, IC₅₀ in the flesh of the non-astringnet type was high despite the increase of -cryptoxanthin at harvest.These results suggest that catechins and ascorbic acid influence O₂-scavenging activity at the beginning and middle of fruit development, and catechins are associated with the O₂-scavenging activity at ripening. Changes in 1-diphenyl-2-picrylhydrazyl (DPPH)-radicalscavenging activity with fruit developmental stage were similar to that for O₂^–. That is, IC₅₀ values of DPPH-radical-scavenging activity decreased rapidly after the removal of astringency. -cryptoxanthin did not eliminate the DPPH-radicals. Therefore, the DPPH-radicals in persimmons may be eliminated primarily by catechins and ascorbic acid. The utilization of tannins before the removal of astringency is discussed.     

Epizootology and experimental infection of Yokose virus in bats

To reveal whether bats serve as an amplifying host for Yokose virus (YOKV), we conducted a serological survey and experimentally infected fruit bats with YOKV isolated from microbats in Japan. YOKV belongs to the Entebbe bat virus group of vector unknown group within the genus Flavivirus and family Flaviviridae. To detect antibodies against YOKV, we developed an enzyme-linked immunosorbent assay (ELISA) using biotinylated anti-bat IgG rabbit sera. Serological surveillance was conducted with samples collected in the Philippines and the sera supplied from Malaysia. One of the 36 samples from the Philippines (2.7%) and 5 of the 26 samples from Malaysia (19%) had detectable ELISA antibodies. In the experimental infections, no clinical signs of disease were observed. Moreover, no significant viral genome amplification was detected. These findings revealed that YOKV replicates poorly in the fruit bat, suggesting that fruit bats do not seem to serve as an amplifying host for YOKV.     

Identification of a host 14-3-3 Protein that Interacts with Xanthomonas effector AvrRxv

AvrRxv is a member of a family of pathogen effectors present in pathogens of both plant and mammalian species. Xanthomonas campestris pv. vesicatoria strains carrying AvrRxv induce a hypersensitive response (HR) in the tomato cultivar Hawaii 7998. Using a yeast two-hybrid screen, we identified a 14-3-3 protein from tomato that interacts with AvrRxv called AvrRxv Interactor 1 (ARI1). The interaction was confirmed in vitro with affinity chromatography. Using mutagenesis, we identified a 14-3-3-binding domain in AvrRxv and demonstrated that a mutant in that domain showed concomitant loss of interaction with ARI1 and HR-inducing activity in tomato. These results demonstrate that the AvrRxv bacterial effector recruits 14-3-3 proteins for its function within host cells. AvrRxv homologues YopP and YopJ from Yersinia do not have AvrRxv-specific HR-inducing activity when delivered into tomato host cells by Agrobacterium. Although YopP itself cannot induce HR, its C-terminal domain containing the catalytic residues can replace that of AvrRxv in an AvrRxv-YopP chimera for HR-inducing activity. Phylogenetic analysis indicates that the sequences encoding the C-termini of family members are evolving independently from those encoding the N-termini. Our results support a model in which there are three functional domains in proteins of the family, translocation, interaction, and catalytic.     

Microbial mineralization of organic matter in sinking particles,bottom sediments and seawater in a coastal fish culturing area

Microbial mineralization rates in sinking particles, bottom sediments and seawater were determined in a coastal fish (red sea bream Pagrus major) culturing area to clarify the mineralization process of organic matter (OM) in the entire water column. The mineralization rates of ¹⁴C‐labelled glutamate and glucose per unit volume were highest in the sinking particles and were 131–572 and 7–49 times higher than those of the seawater and bottom sediments respectively. The turnover time of glutamate tended to be shorter than that of glucose at all three sites of the water column. Bacteria appeared to prefer amino acids to monosaccharides because amino acids could be utilized as both energy and nitrogen sources. The sedimentation rate of particulate organic carbon (POC) derived from phytoplankton accounted for 9–61% of the total POC, and it was particularly high in early summer (61% and 50% at fish cage and non‐cage stations respectively). The present study clearly shows that sinking particles serve as an important site of microbial mineralization process of OM within the water column, and that phytoplankton production was another serious cause of organic pollution of the seafloor in addition to the organic wastes directly discharged from fish farms.