Development of green onion and cabbage certified reference materials for quantification of organophosphorus and pyrethroid pesticides

Green onion and cabbage certified reference materials for the analysis of pesticide residues were issued by the National Metrology Institute of Japan, part of the National Institute of Advanced Industrial Science and Technology. Green onion and cabbage samples were grown so as to contain several kinds of organophosphorus and pyrethroid pesticides, and those were collected from a field in the Kochi Prefecture in Japan. The certification was carried out by using multiple analytical methods to ensure the reliability of analytical results; the values of target pesticides (diazinon, fenitrothion, cypermethrin, etofenprox, and permethrin for green onion and chlorpyrifos, fenitrothion, and permethrin for cabbage) were obtained by isotope dilution mass spectrometry. Certified values of target pesticides were 0.96-13.9 and 2.41-6.9 mg/kg for green onion and cabbage, respectively. These are the first green onion and cabbage powder certified reference materials in which organophosphorus and pyrethroid pesticides are determined.     

Genetic characterization and susceptibility on poultry and mammal of H7N6 subtype avian influenza virus isolated in Japan in 2009

From February to March 2009, six strains of H7N6 subtype avian influenza virus were isolated from quails in three farms in Aichi prefecture in Japan. The isolates were shown to be low pathogenic for chicken by the examination performed using the "Manual of Standards for Diagnostic Tests and Vaccines" by World organisation for Animal Health (OIE). The deduced amino acid sequence at the cleavage site was PE (I/Q/L) PKRR (nucleotide sequences were cct gaa (a/c) (t/a) a cc (a/g) aaa aga aga), suggesting persistence in domestic poultry for some time. The direct putative ancestor strain could not be elucidated by phylogenetic analysis of all genome segments of the quail isolates. Diverged date from a putative common ancestor in a non-rooted phylogenetic tree among quail viruses was estimated between March 2002 and July 2004. Three putative N-linked glycosylation sites resided in the vicinity of the receptor binding pocket of HA1 region. They are considered to decrease the reactivity of neutralizing antibody against the virus. Experiments for the infectivity and pathogenicity of a quail strain to poultry indicated that the quail isolate had higher infectivity to quails than chickens and ducks. Direct and dust-borne and/or droplet-borne transmissions among quail were proven in quails with and without direct contact with experimentally infected quails. The virus is seldom transmitted among chickens either directly or indirectly, and indirect transmission from infected quails to chickens was not observed. The pathogenicity of the quail strain for mammalian, pig and mouse was low, although it could replicate in those animals.     

Prevalence of hemoplasma infection among cattle in the western part of Japan

We have examined for hemoplasma infection among cattle in the Hiroshima and Miyazaki prefectures by using a sensitive real-time PCR, with SYBR Green I and with melting curve analysis, which allow to distinguish the two bovine hemoplasma species, Mycoplasma wenyonii and 'Candidatus M. haemobos'. We found 69.4% of 36 cattle in Hiroshima and 93.8% of 32 cattle in Miyazaki infected with either of these two hemoplasma species. High morbidity in western part of Japan may reflect the activity of arthropod vectors for hemoplasma transmission. We also demonstrated neonatal calves less than three months old affected with hemoplasmas without grazing in summer, suggesting a possibility of vertical transmission.     

Comparison of serum lipid compositions, lipid peroxide, alpha-tocopherol and lipoproteins in captive marine mammals (bottlenose dolphins, spotted seals and West Indian manatees) and terrestrial mammals

Concentrations of serum lipid components, lipid peroxide (LPO) and α-tocopherol and electrophoretic patterns of lipoproteins in serum samples obtained from captive marine mammals and terrestrial mammals were compared. Serum concentrations of total cholesterol, free fatty acid, and phospholipid in fish-eating animals were significantly higher than those in manatees and cows. Serum LPO and α-tocopherol concentrations in the fish-eating animals were also significantly higher than those in manatees, cows and dogs. Different patterns of densitometric scans of low density lipoprotein (LDL) and a significantly lower percentage of LDL were demonstrated in the dolphins compared with the seals, cow and dogs. The concentration of LPO was significantly correlated with triglyceride and phospholipid concentrations in serum from the dolphins. These results suggest that triglyceride and phospholipid are susceptible to oxidative reaction in fish-eating animals. Evaluation of serum lipids, LPO and α-tocopherol concentrations is needed for nutritional husbandry for fish-eating animals.     

Pathogenicity of highly pathogenic avian influenza viruses of H5N1 subtype isolated in Thailand for different poultry species

Highly pathogenic avian influenza (HPAI) viruses of the H5N1 subtype have caused several rounds of outbreaks in Thailand. In this study, we used 3 HPAI viruses isolated in Thailand in January 2004 from chicken, quail, and duck for genetic and pathogenetic studies. Sequence analysis of the entire genomes of these isolates revealed that they were genetically similar to each other. Chickens, quails, domestic ducks, and cross-bred ducks were inoculated with these isolates to evaluate their pathogenicity to different host species. A/chicken/Yamaguchi/7/04 (H5N1), an HPAI virus isolated in Japan, was also used in the chicken and quail studies for comparison. All four isolates were shown to be highly pathogenic to chickens and quails, with 100% mortality by 10(6) EID50 inoculants of the viruses. They caused sudden death in chickens and quails within 2-4 days after inoculation. The mean death times (MDT) of quails infected with the Thai isolates were shorter than those of chickens infected with the same isolates. Mortality against domestic and cross-bred ducks ranged from 50 to 75% by intranasal inoculation with the 10(6) EID50 viruses. Neurological symptoms were observed in most of the inoculated domestic ducks and appeared less severe in the cross-bred ducks. The MDTs of the ducks infected with the Thai isolates were 4.8-6 days post-inoculation. Most of the surviving ducks infected with the Thai isolates had sero-converted until 14 dpi. Our study illustrated the pathobiology of the Thai isolates against different poultry species and would provide useful information for improving control strategies against HPAI.     

Effect of alcohol-based organosolv treatment combined with short-time ball milling on the enzymatic hydrolysis of Japanese cypress (Chamaecyparis obtusa)

Japanese cypress (Chamaecyparis obtusa) is a recalcitrance softwood widely used as building material in Japan. A combined technique of alcohol-based organosolv treatment and ball milling (BM) of this softwood for the production of fermentable sugars from lignocellulose was investigated. Ethanol, ethylene glycol, water, and their mixtures were used as solvents, and acetic acid was used as a proton donor. After solvolysis, a residual sample was subjected to short-time BM, and the entire residue was evaluated by saccharification with Acremonium cellulase. Short-time BM after alcohol-based organosolv treatment greatly improved the enzymatic digestibility and decreased the required severity of organosolv treatment. Moreover, alcohol-based organosolv treatment increased the efficiency and reduced the time required for BM despite small quantity of removed lignin. It was found that the combination of alcohol-based organosolv treatment in mild condition and short-time BM had a synergistic effect on the enzymatic digestibility of Japanese cypress.     

Effect of mosapride on prednisolone-induced gastric mucosal injury and gastric-emptying disorder in dog

Previous report demonstrated that prokinetic agent mosapride has anti-ulcerogenic action in rat-indomethacin gastric mucosal injury model. Here, we assessed the prophylactic effect of mosapride on gastric mucosal injury and emptying disorder induced by prednisolone in dogs. Crossover study design was employed. Six healthy beagles were administered prednisolone alone (2 mg/kg, twice a day [BID] subcutaneously) and prednisolone with mosapride (1 mg/kg, BID, orally), followed by an interval of at least 6 weeks. In each treatment, gastric mucosal injury was scored endoscopically according to the modified Lanza scale, and gastric emptying was assessed with (13)C-octanoic acid breath test. The incidence of gastrointestinal adverse events was also investigated. Coadministration of mosapride with prednisolone significantly (P<0.05) reduced the gastric mucosal injury score (mean ± SD, 17.67 ± 6.96), compared with that of prednisolone treatment alone (25.50 ± 13.03). Prednisolone treatment delayed the half-emptying time (184 ± 45 min) compared with that of controls (137 ± 19 min), and coadministration of mosapride improved this gastric-emptying delay (143 ± 29 min). Furthermore, the incidence of the gastrointestinal adverse event vomiting became less frequent upon coadministration with mosapride. In addition to its prokinetic action, our study suggests that mosapride has an anti-ulcerogenic action in dogs. The use of mosapride in combination with prednisolone is effective for attenuating prednisolone-induced gastrointestinal adverse events.     

Effects of pre-slaughter nutritional condition on intramuscular collagen solubility, pyridinoline cross-links and meat tenderness in aged goats

Effects of preslaughter nutritional condition on intramuscular collagen characteristics were studied, in order to clarify the potential of intensive feeding to aged animals in improving meat tenderness. Ten castrated male goats were assigned into one of two groups: one group was allowed ad libitum access to a concentrate diet (total digestible nutrients 70%, crude protein 15%) and Italian ryegrass hay (ADLIB‐group), and the other group was fed a restricted amount of their diet (concentrate diet 0.5% of bodyweight/day; hay 1.5% of bodyweight/day) to maintain their bodyweight (MAIN‐group). After 3 months of the experimental period, goats were slaughtered and meat samples were obtained immediately. Goats in ADLIB‐group had lower total and insoluble collagen concentrations, higher fat concentrations and collagen solubility than those in MAIN‐group, but soluble collagen concentrations of muscles were similar for both groups. Goats in ADLIB‐group had lower pyridinoline concentrations than those in MAIN‐group, in all muscles, but the differences of pyridinoline concentration between the groups were not statistically significant. Warner‐Bratzler shear force values of Longissimus and Biceps femoris muscles were lower in ADLIB‐group than in MAIN‐group. The increase of meat tenderness by preslaughter intensive feeding seemed to be associated with the increase in intramuscular fat deposition and high collagen solubility, and pyridinoline cross‐link appeared to be one of the factors related to collagen solubility.     

Characterization of mutants of the chestnut blight fungus (<Emphasis Type="Italic">Cryphonectria parasitica</Emphasis>) with unusual hypovirus symptoms

The type virus of the family Hypoviridae, Cryphonectria hypovirus 1 strain EP713 (CHV1-EP713), infects Cryphonectria parasitica, the filamentous causal fungus of chestnut blight, and reduces its virulence. This pathosystem serves as a model to study fungus-mycovirus interactions. We previously developed a genetic screening protocol for host factors associated with symptom induction by CHV1-EP713 and its mutants. In the procedure the standard field fungal isolate EP155 was transformed by cDNA from a mild hypovirus mutant Cys(72), launching virus infection, and mutagenized by random plasmid insertion with pHygR conferring hygromycin resistance. We now report an extension of the study to characterize different mutant strains, with different phenotypes than their parental strain TCys(72)-1. TCys(72)-1 is moderately reduced in pigmentation and sporulation compared to the uninfected wild-type strain EP155. Mutants sfb1, sfb2 and k202 were characterized biologically and molecularly in comparison to the previously isolated mutant (namA) and the parental strain. These mutants harbored one (sfb1) or more copies (sfb2 and k202) of the mutagenic plasmid, pHygR. The three mutants had similar biological attributes; that is, vegetative growth rate, conidiation and virulence (assay on apples) was reduced on potato dextrose agar media, relative to the parental strain and pigmentation was the same or slightly increased. Interestingly, viral dsRNA accumulation levels were apparently unaltered in these mutants. The screening method was efficient for mining fungal mutants with unusual hypovirus symptoms. Further, characterization of the mutants provides interesting insights into symptom induction by the hypovirus.