Extensive Screening for Plant Foodstuffs in Okinawa,Japan with Anti-Obese Activity on Adipocytes <Emphasis Type="Italic">In Vitro</Emphasis>

To search for plant foodstuffs with potent anti-obese activity, we conducted a large scale screening based on the inhibitory activity on adipogenesis and the facilitating activity on adipolysis in vitro. That is, inhibition of intracellular lipid accumulation and facilitation of lipid degradation in 3T3-L1 adipocytes were extensively screened from ethanol and hexane extracts of approximately 100 kinds of plant foodstuffs marketed in Okinawa prefecture, which has been famous for the highest prevalence of exceptionally long-lived individuals in the world. Among them thirty one foodstuffs showed potent inhibitory activity on intracellular lipid accumulation in 3T3-L1 adipocytes, whereas only four foodstuffs showed clear facilitating effect on lipid degradation in 3T3-L1 adipocytes. Although further study to examine the in vivo effects on adipogenesis and adipolysis is required, this is the first study to investigate anti-obese characteristics of wide range of traditional Okinawa foodstuffs so that the results give useful information to take another look at Okinawa food culture.     

Construction and evaluation of genetically engineered replication-defective porcine reproductive and respiratory syndrome virus vaccine candidates

Porcine reproductive and respiratory syndrome virus (PRRSV) is an emerging pathogen causing significant economic losses in the swine industry worldwide. Two novel gene-deleted viruses were constructed and evaluated as vaccine candidates. Using the full-length infectious cDNA clone of North American PRRS isolate P129, the ORF2 and ORF4 genes (which encoded minor structural glycoproteins GP2a/2b and GP4, respectively) were individually deleted from the viral genome. Both deletion mutants were non-viable in MARC-145 cells and porcine alveolar macrophages, indicating that both genes are essential for virus replication. To rescue the replication-defective PRRSV, two complementing cell lines, MARC-2000 and MARC-400, were established to stably express the PRRSV GP2 and GP4 proteins, respectively. These cells were able to complement the deleted gene function of PRRSV in trans and supported production of the replication-defective DeltaORF2-PRRSV and DeltaORF4-PRRSV viruses. Both DeltaORF2-PRRSV and DeltaORF4-PRRSV viruses were propagated for 40-50 generations in the corresponding complementing cells and remained replication-defective in MARC-145 cells. To examine the immunogenic potential of the replication-defective PRRSV as vaccine candidates, four groups of pigs, 20 pigs per group, were immunized twice with DeltaORF2-PRRSV or DeltaORF4-PRRSV and challenged with the homologous virulent virus at 3 weeks post-immunization. In spite of the fact one group showed significant reduction in virus load, we could not demonstrate improvement from clinical diseases in this vaccination/challenge study. However, we did show that the cDNA clone of PRRSV can be a useful tool to genetically engineer PRRSV vaccine candidates and to study pathogenesis and viral gene functions.     

The effect of sodium butyrate supplementation on ruminal and fecal pH and serum lipopolysaccharide-binding protein after ruminal acidosis challenge in nonlactating cows

The objective of this study was to evaluate effects of sodium-butyrate supplementation on gastrointestinal function and the inflammatory response to ruminal acidosis (RA) challenge in cows. Four nonlactating cows with a rumen cannula were assigned to two treatments in a crossover design. Treatments were ruminal administration of sodium-butyrate (BUT) or control (CON). Sodium-butyrate was provided as Gustor BP70 and administered at a butyrate dose of 0.04% per kg body weight. The CON premix was made by replacing sodium-butyrate with wheat bran. Experimental periods were 28 days long with 21-day washout period separating the treatments. On Day 25 of each period, corn starch was ruminally administered at 0.7% per kg body weight as RA challenge. After RA challenge, ruminal pH was lower, and endotoxin concentration was higher for cows provided with BUT than those with CON, but the increase in fecal starch and the decrease in fecal pH were attenuated by BUT. The effect of butyrate supplementation on serum lipopolysaccharide-binding protein after RA challenge was not found. From these findings, butyrate supplementation mitigated rectal acidosis by reducing the flux of fermentable carbohydrate into the large intestine. An anti-inflammatory effect of butyrate was not observed, possibly due to lower pH and higher endotoxin concentration in the rumen.     

Leaf blast disease reduction by rice-phyllosphere actinomycetes producing bioactive compounds

Fungal leaf blast, caused by Pyricularia oryzae, is a devastating disease of rice plants that annually causes severe production losses worldwide and is one of the top 10 fungal diseases that threaten global food security. Thus, a reliable control strategy against this disease is essential. In this study, the antagonistic activity of indigenous phyllosphere actinomycetes was elucidated against P. oryzae in vitro and in planta to develop an efficient, effective and environmental friendly approach to protect rice plants against P. oryzae. Of 75 isolates of actinomycetes isolated from the rice phyllosphere, 18 isolates inhibited P. oryzae by >45%. According to analysis of their 16 S rRNA gene sequences, the majority of the 18 isolates belonged to Streptomyces genera; others were identified as belonging to Saccharothrix, Gordonia, or Lentzea. Isolates that potentially produced a bioactive compound(s) were identified among the 18 isolates: 17 isolates (94.44%) had a domain marker for nonribosomal peptide synthetase (NRPS) gene and 12 (66.67%) had type-I polyketide synthase (PKS) gene in their corresponding genome. Interestingly, isolates JSN1.9, SKB2.14, and SKB2.3 suppressed disease suppression by approximately 88%. To our knowledge, this is the first report on the application of rice-phyllosphere actinomycetes producing bioactive compounds to control leaf blast disease in Indonesia. Thus, these findings have escalated the potential application of phyllosphere actinomycetes as a supreme biocontrol agent against fungal leaf blast disease.     

Different cross-reactivity of human and rodent sera to Tula virus and Puumala virus

Tula virus (TULV) and Puumala virus (PUUV) are hantaviruses carried by the bank vole (Myodes glareolus) and European common vole (Microtus arvalis), respectively. PUUV is a causative agent of hemorrhagic fever with renal syndrome (HFRS), while TULV is thought to be apathogenic to humans. The N-terminal regions of the N proteins from TULV and PUUV were expressed and applied as enzyme-linked immunosorbent assay (ELISA) antigens. Colonized Japanese grass voles (Microtus montebelli) and BALB/c mice were used for experimental inoculation of the vole-borne hantaviruses TULV and PUUV. Voles and mice showed significant antibody production toward both viruses, but these antisera showed little cross-reactivity between TULV and PUUV in the immunofluorescence antibody assay and ELISA. In contrast, sera from patients with HFRS caused by PUUV exhibited high cross-reactivity against the TULV antigen, and sera from a natural rodent reservoir showed moderate cross-reactivity against the heterologous antigen, indicating that the antigenic cross-reactivity between TULV and PUUV differs in sera from rodents and humans.