Comparison of Salmonella enterica serovar Abortusequi isolates of equine origin by pulsed-field gel electrophoresis and fluorescent amplified-fragment length polymorphism fingerprinting

Equine paratyphoid is caused by Salmonella enterica serovar Abortusequi, and manifests mainly as abortion in the mare. We compared S. Abortusequi strains isolated in Japan and other countries using pulsed-field gel electrophoresis (PFGE) and fluorescent amplified-fragment length polymorphism (FAFLP) analysis. PFGE analysis of S. Abortusequi strains gave 21-27 fragments ranging in size from 33 to 602kb. Although two PFGE profiles were observed among the 20 S. Abortusequi isolates in Japan, the restriction fragments originating from the chromosome were common between the two profiles. The similarity index of the two profiles was 90.9%, while those between Japanese and five other S. Abortusequi strains were 29.8-37.5%. On the other hand, FAFLP analysis of S. Abortusequi strains generated 64-67 amplified fragments ranging in size from 100 to 400bp. One polymorphic fragment was observed among the 20 S. Abortusequi isolates in Japan. These data indicate the close relation of this agent in Japan. S. Abortusequi strains sharing a common ancestry might have been conserved in Japan.     

The regional economic impacts on the development of wood chip utilization in Maniwa city

This paper reports on an empirical investigation about regional economic impacts by the promotion of wood chip utilization in Maniwa city, Japan. We clarify the estimated potential of regional economy development by using input–output analysis for Maniwa city. The result indicates that 315 million JPY of woody chip products can be estimated to generate roughly 448 million JPY of direct, indirect, and induced economic effects. The value of 448 million JPY was equivalent to about 0.2 % of total production value in Maniwa. It is also expected to create approximately 21 jobs by promotion of wood chip utilization. We examined the regional economic impacts on the woody biomass utilization, and evaluated to estimate direct, indirect and induced economic effects, and job creation to clarify the economic impact on Maniwa. We will leave the negative economic effects and the environmental impacts in the process of woody biomass utilization as a future task.     

Synthesis and antifungal activity of the proposed structure of a volatile compound isolated from the edible mushroom Hypsizygus marmoreus

We synthesized the proposed structure of an antifungal compound detected in the culture broth of the edible mushroom Hypsizygus marmoreus. Using the Evans aldol and Abiko–Masamune aldol reactions as the key steps, we synthesized all of the stereoisomers of the compound with high stereoselectivity. The GC retention times and the fragmentation patterns in the mass spectra of the synthesized isomers did not match those of the natural product. Therefore, this result may imply that it is necessary to reisolate the natural product and reconsider its structure. All of the synthesized isomers were found to exhibit antifungal activity against the phytopathogenic fungus Alternaria brassicicola. Due to their simple structures, the obtained isomers could be lead compounds for new pesticides.     

Molecular analysis of bovine leukemia virus in early epidemic phase in Japan using archived formalin fixed paraffin embedded histopathological specimens

Bovine leukemia virus (BLV) is an important pathogen associated with enzootic bovine leukosis. In this study, we performed PCR and sequencing analysis to characterize BLVgp51 sequences from formalin-fixed paraffin-embedded (FFPE) specimens made from 1974 to 2000 and successfully obtained BLV proviral genome sequences from 94% of the analyzed samples. Furthermore, from these samples, we reconstructed eight full-length and nearly full-length BLVgp51 sequences. These sequences were classified as BLV genotype 1, implying that genotype1 has already been circulating in Japan since the 1970s. In our results, the proviral DNA was detected in the 1970s, 1980s, and 1990s in the same manner, indicating that the detection of BLV proviral genome depends on storage conditions rather than storage period. The sequences obtained in this study provide direct insights into BLV sequences before 2000, which serves as a good calibrator for inferring ancient BLV diversity.     

Detection of vaccine-like infectious bursal disease (IBD) virus in IBD vaccine-free chickens in Japan

The prevalence of infectious bursal disease virus (IBDV) was studied in chickens, which had not been vaccinated against IBD. Fifty sera and forty-six bursae of Fabricius from chickens showing impaired growth, collected from 7 IBD vaccination-free farms in Japan were used for virus neutralization (VN) tests and RT-PCR for detection of IBDV genome corresponding to the VP2 hypervariable region. Of the fifty sera, 39 sera (78%) from 6 farms were VN antibodies positive. Of the forty-six bursae, 37 bursae (80.4%) from 6 farms were positive in the RT-PCR assay. The sequences of all the RT-PCR products detected in this study were closely related or identical to those of the vaccine strains. These results show that vaccine-like IBDV is prevalent even in IBD vaccine-free chicken farms in Japan.     

Comparison of the growth kinetics of neuropathogenic and nonneuropathogenic equid herpesvirus type 1 (EHV-1) strains in cultured murine neuronal cells and the relevance of the D/N(752) coding change in DNA polymerase gene (ORF30)

We compared the growth kinetics of neuropathogenic and nonneuropathogenic equine herpesvirus type 1 (EHV-1) strains in mouse cerebral cortex cells and investigated the relevance of the D/N amino acid change at position 752 of ORF30 in Japanese isolates. Neuropathogenic electropherotype P strains 01c1 and 89c25 exhibited similar growth kinetics to nonneuropathogenic P strain 90c16 in cultured neurons; however, the growth ability of type B strain 97c7 was lower than those of the other strains tested. The amino acid encoded at 752 of ORF30 in 01c1 was asparagic acid; asparagine was encoded in the other EHV-1 strains isolated from Japanese horses. The D/N(752) difference in ORF30 may not be related to replication ability in neurons.     

Chicken B lymphoma DT40 cells as a useful tool for in vitro analysis of pathogenic infectious bursal disease virus

Susceptibility of DT40 cells to pathogenic field strains of infectious bursal disease virus (IBDV) including very virulent and classical virulent strains were studied. After the first and second passage of the virus in DT40 cells, IBDV-specific antigen was readily detected in DT40 cells inoculated with the pathogenic field strain infected bursal homogenates. Nucleotide sequence analysis in the VP2 hypervariable domain, which is critical for the virulence of IBDV, revealed no common amino acid substitutions among the pathogenic IBDVs in accordance with the propagation in DT40 cells. These results indicate that DT40 cells are a useful tool for rapid isolation of pathogenic field strains and successive in vitro analysis of IBDV.     

The effects of migration on the immunity of Black-Headed Gulls (Chroicocephalus ridibundus: Laridae)

In order to elucidate the relationship between migration period and immunity related to susceptibility, we conducted research on Black-headed gulls (Chroicocephalus ridibundus). We captured 260 gulls and collected their peripheral blood. Their leukocyte (WBC) count, percentages of heterophils (Het) and lymphocytes (Lym), heterophil and lymphocyte ratio (H/L ratio), and CD4 and CD8α expression levels (CD4 and CD8α, respectively) were quantitatively analyzed over three migration periods (Autumn migration, Wintering, Spring migration). In Adult gulls, WBC counts and CD4 levels significantly increased. Moreover, the Het and H/L ratio decreased from the Autumn migration to Wintering. Conversely, only WBC counts and CD4 levels measurements significantly decreased from Wintering to Spring migration (P<0.05). The tested parameters of the Tokyo-bay population show a greater significant difference than the measurements of immunity of the Mikawa-bay population. This study suggests that the migratory period has a negative effect on an aspect of the immune system. Including the period-difference in the immune systems in the local population, it is necessary to investigate the relationship between the ecology of migratory birds and their immunity.     

Molecular characterization of avian polyomavirus isolated from psittacine birds based on the whole genome sequence analysis

Seven avian polyomaviruses (APVs) were isolated from seven psittacine birds of four species. Their whole genome sequences were genetically analyzed. Comparing with the sequence of BFDV1 strain, nucleotide substitutions in the sequences of seven APV isolates were found at 63 loci and a high level of conservation of amino acid sequence in each viral protein (VP1, VP2, VP3, VP4, and t/T antigen) was predicted. An A-to-T nucleotide substitution was observed in non-control region of all seven APV sequences in comparison with BFDV1 strain. Two C-to-T nucleotide substitutions were also detected in non-coding regions of one isolate. A phylogenetic analysis of the whole genome sequences indicated that the sequences from the same species of bird were closely related. APV has been reported to have distinct tropism for cell cultures of various avian species. The present study indicated that a single amino acid substitution at position 221 in VP2 was essential for propagating in chicken embryonic fibroblast culture and this substitution was promoted by propagation on budgerigar embryonic fibroblast culture. For two isolates, three serial amino acids appeared to be deleted in VP4. However, this deletion had little effect on virus propagation.     

Polo-like kinase Cdc5 controls the local activation of Rho1 to promote cytokinesis

The links between the cell cycle machinery and the cytoskeletal proteins controlling cytokinesis are poorly understood. The small guanine nucleotide triphosphate (GTP)-binding protein RhoA stimulates type II myosin contractility and formin-dependent assembly of the cytokinetic actin contractile ring. We found that budding yeast Polo-like kinase Cdc5 controls the targeting and activation of Rho1 (RhoA) at the division site via Rho1 guanine nucleotide exchange factors. This role of Cdc5 (Polo-like kinase) in regulating Rho1 is likely to be relevant to cytokinesis and asymmetric cell division in other organisms.