Ethanol production from sugi pulp under simultaneous saccharification and fermentation using a cocktail enzyme of T. reesei and A. tubingensis produced by solid-state fermentation

The process of solid-state fermentation was used to produce a cocktail enzyme of Trichoderma reesei ATCC 66587 and Aspergillus tubingensis KRCF 700-33. Wheat bran, corncob, and sugi pulp were supplemented with ammonium sulfate as an enzyme-producing medium using T. reesei and A. tubingensis. The corncob blend ratio, duration of incubation, and ammonium sulfate concentration were optimized for enhancing cellulase production from T. reesei using a Box-Behnken design. Filter paper degrading activity more than tripled when T. reesei was grown in the optimized medium, as compared with the initial medium. The highest activity of 4.03 FPU/ml (about 29 FPU/g of dry material) was obtained with a cocktail enzyme having a 25 % content of A. tubingensis and 75 % of T. reesei. The sugi pulp was then fermented to ethanol with the cocktail enzyme and thermotolerant yeast (Saccharomyces cerevisiae BA-11) under simultaneous saccharification and fermentation (SSF) at 40 °C. An ethanol concentration of 4.48 % (w/v) was achieved using the cocktail enzyme (4 FPU/g-pulp) that was produced on-site with a substrate loading level of 12.5 wt %, which achieved an ethanol yield of 76 % after 72 h.     

Contagious equine metritis eradicated from Japan

Contagious equine metritis (CEM), a contagious venereal disease of horses, invaded Japan in 1980 and spread in the Thoroughbred population of the Hidaka-Iburi district of Hokkaido. To eradicate CEM, we ran a program aimed at detecting Taylorella equigenitalis, the causal agent, in carrier horses by using the PCR test, followed by culling or treatment. In 2001, the first year of the program, 12,356 Thoroughbred racing stallions and mares were tested and 11 carriers were found. Four, two, one, and one carrier mares were detected in 2002, 2003, 2004, and 2005, respectively, by application of the program at the same scale as in 2001. No PCR-positive horses were found from 2006 to 2010. These results strongly suggest that CEM was eradicated from Japan by 2010.     

Locally synchronized synaptic inputs

Synaptic inputs on dendrites are nonlinearly converted to action potential outputs, yet the spatiotemporal patterns of dendritic activation remain to be elucidated at single-synapse resolution. In rodents, we optically imaged synaptic activities from hundreds of dendritic spines in hippocampal and neocortical pyramidal neurons ex vivo and in vivo. Adjacent spines were frequently synchronized in spontaneously active networks, thereby forming dendritic foci that received locally convergent inputs from presynaptic cell assemblies. This precise subcellular geometry manifested itself during N-methyl-D-aspartate receptor-dependent circuit remodeling. Thus, clustered synaptic plasticity is innately programmed to compartmentalize correlated inputs along dendrites and may reify nonlinear synaptic integration.     

Relations between plasma IGF-I concentrations during treatment with CIDR-based or Ovsynch protocol for timed AI and conception in early postpartum Japanese black beef cows

We examined the relations between plasma insulin-like growth factor (IGF) -I concentrations during treatment with CIDR-based or Ovsynch protocol for timed AI and conception and plasma steroid concentrations in early postpartum Japanese Black beef cows. Cows in the control group (Ovsynch; n = 21) underwent Ovsynch protocol (GnRH analogue on Day 0, PGF(2alpha) analogue on Day 7, and GnRH analogue on Day 9), with AI on Day 10, approximately 20 h after the second GnRH treatment. Cows in the Ovsynch+CIDR group (n = 22) received Ovsynch protocol plus a CIDR for 7 days (starting on Day 0). Cows in the further treatment group (EB+CIDR+GnRH; n = 22) received 2 mg of estradiol benzoate (EB) on Day 0 in lieu of the first GnRH treatment, followed by the same treatment as in the Ovsynch+CIDR protocol. Plasma IGF-I concentrations were determined on Days -7, 0, 7, 9 and 17. Conception rates were improved in the CIDR-combined groups (both CIDR-treated groups were combined) relative to Ovsynch group (P < 0.05) for cows with low IGF-I concentrations (<1,000 ng/ml) on Days -7, 0, and 7, but improved conception rate produced by the CIDR-based protocols did not occur in cows with a high IGF-I concentration (> or =1,000 ng/ml). Plasma estradiol-17beta concentrations increased from Day 0 to 7 (P < 0.05) and were unchanged from Day 7 to 9 in the Ovsynch group with low IGF-I concentrations on Day 0, while they were unchanged from Day 0 to 7 and increased from Day 7 to 9 (P < 0.05) in the Ovsynch group with high IGF-I concentrations on Day 0 and in the CIDR-combined group. Plasma progesterone concentrations in the Ovsynch group with low IGF-I concentrations on Day 0 were higher on Day 14 than in the Ovsynch group with high IGF-I concentrations on Day 0 and in the CIDR-combined group (P < 0.05). In conclusion, CIDR-based protocols may improve conception relative to Ovsynch in early postpartum beef cows with lower plasma IGF-I concentrations at the start of the protocols. This improvement is probably due to prevention of premature increases of estradiol-17beta and progesterone concentrations, which occurred in cows with low IGF-I concentrations treated with Ovsynch, by the CIDR treatment.     

Abnormal development of nephrons in claudin-16-defective Japanese black cattle

The kidneys of 37 Japanese Black calves aged 2 to 65 months diagnosed with Claudin 16 (CL-16) defect by the DNA-based test were examined pathologically. The animals exhibited clinical symptoms such as growth impairment, renal failure, overgrowth of hooves, and anemia at a young age. There was no correlation between the time of onset and age. Kidney weights relative to body weight were similar to those in normal animals, but both kidney net weights and size were reduced due to atrophy in animals that showed severe renal dysfunction. Histopathological examination of the kidneys showed reduction in the number of glomeruli, compensatory hypertrophy of glomeruli and tubules, and glomerular and tubular atrophy accompanied by interstitial fibrosis and lymphocytic infiltration. Glomeruli were clearly less in number in the kidneys of CL-16-defective animals than those of normal animals even in the cases with mild lesions. A small number of immature glomeruli and tubules were also detected, suggesting that there were fewer nephrons developed at birth in CL-16-defective animals. It was suggested that a defect of the CL-16 gene is involved in the "abnormal development of nephrons". Immunohistopathological examination of the kidneys showed that the epithelium of thick ascending limb of Henle was stained with anti-CL-16 antibody in the control animals, but not in the affected animals, suggesting a defect of CL-16 in the epithelium of renal tubules in the affected animals.     

Pathological and immunohistochemical studies of subclinical infection of chicken anemia virus in 4-week-old chickens

Subclinical infection of chicken anemia virus (CAV) at 4 to 6 weeks of age, after maternal antibodies have waned, is implicated in several field problems in broiler flocks. In order to understand the pathogenesis of subclinical infection with CAV, an immunopathological study of CAV-inoculated 4-week-old SPF chickens was performed. Sixty 4-week-old SPF chickens were equally divided into CAV and control groups. The CAV group was inoculated intramuscularly with the MSB1-TK5803 strain of CAV. Neither mortality nor anemia was detected in the CAV and control groups. In the CAV group, no signs were observed, except that some chickens were grossly smaller compared with the control group. Sporadic thymus lobes appeared to be reddening and atrophied. Within the first two weeks p.i. of CAV, there was a mild to moderate depletion of lymphocytes in the thymus cortex and spleen in some chickens. Moreover, lymphoid depletion of the bursa of Fabricius, proventriculus and cecal tonsils was observed. Hyperplastic lymphoid foci were observed in the liver, lungs, kidneys and heart at the 4th week p.i. of CAV. Immunohistochemically, a moderate lymphoid depletion of CD4(+)and CD8(+) T cells in the thymus cortex and spleen was observed in some chickens within two weeks p.i. of CAV. CAV inclusions and antigens were detected infrequently in the thymus cortex and spleen. It could be concluded that the immunosuppression in subclinical infection with CAV occurs as a result of reduction of cellular immunity.     

Delayed-type hypersensitivity in sheep induced by synthetic peptides of bovine leukemia virus encapsulated in mannan-coated liposome

Four sheep were immunized with synthesized peptides, derived from bovine leukemia virus (BLV) envelope glycoprotein, encapsulated in mannan coated liposomes as adjuvant. On the seventh day after the immunization, the sheep were intradermally challenged with BLV antigen, or synthesized peptides. The areas challenged with antigen were increased skin thickness and biopsied sequentially for immunohistological examinations. Strong delayed-type hypersensitivity was induced in sheep immunized and challenged with peptides encapsulated in mannan-coated liposomes. The major phenotype of the infiltrating lymphocytes was CD5(+). The ratio of CD4(+) to CD8(+) cells was about 1:1.     

Effects of bacterial strains on the development of the ripening flavor of cured pork loins

To determine the effect of bacteria on the development of the ripening flavor of cured meat, pork loins were cured with pickles containing 8 or 16% sodium chloride at 0 and 8°C. The bacterial flora, total free amino acid content and total free fatty acid content in the cured loins, sensory properties of cooked pork loins and the relationships between the proteolytic and lipolytic activities of isolates and curing conditions were investigated. Desirable bacteria, including salt‐tolerant bacteria and lactic acid bacteria, were predominant under the condition of 8% sodium chloride in the pickles and a curing temperature of 8°C. The total free amino acid content and total free fatty acid content at a curing temperature of 8°C were higher than those at 0°C. Cooked pork loins cured in pickles containing 8% sodium chloride at a temperature of 8°C for more than 7 days were preferred in terms of color, flavor and taste. Before the curing procedure, gram‐negative bacteria (Vibrio, Acinetobacter, Pseudomonas and Enterobacteriaceae) were predominant in the pork loins. During the curing period, the numbers of viable gram‐positive bacteria (Micrococcus, Staphylococcus and Pediococcus) increased and the numbers of viable gram‐negative bacteria decreased. Strains of Micrococcus and Staphylococcus with proteolytic and/or lipolytic activities in cured meat also increased during the curing period and were more predominant in the pork loins cured in pickles containing 8% sodium chloride and at a curing temperature of 8°C than in pork loins cured in pickles containing 16% sodium chloride and at a curing temperature of 0°C. The actions of these bacteria were thought to be important factors affecting the flavor of the cured pork. Micrococcus and Staphylococcus with proteolytic activity might contribute to the development of the ripening flavor of ham. The results of the present study together with the results of further investigations on the relationships of the enzyme activities of Micrococcus and Staphylococcus in cured meat with a preferable flavor would be useful for establishing a novel effective method for using bacteria to produce ham of a high quality.     

Ovsynch plus CIDR protocol for timed embryo transfer in suckled postpartum Japanese Black beef cows

We compared synchronization and pregnancy rates, and the increase in blood progesterone concentrations during luteal development, between (1) Ovsynch plus an intravaginal controlled internal drug release (CIDR) device protocol followed by timed embryo transfer (timed ET), and (2) a conventional estrus synchronization method using PGF(2 alpha) and ET in suckled postpartum Japanese Black beef cows. Cows in the PGF group (n=18) received a PGF(2 alpha) analogue when a CL was first palpated per rectum at 10-d intervals after 1 to 2 month postpartum. Cows (n=11), which showed estrus (Day 0) within 5 d of the PGF(2 alpha), and had a CL on Day 7, received ET. Cows in the Ovsynch+CIDR group (n=19) underwent the Ovsynch protocol plus a CIDR for 7 d (GnRH analogue and CIDR on Day-9, PGF(2alpha) analogue with CIDR removal on Day-2, and GnRH analogue on Day 0), with ET on Day 7. The ovulation synchronization (100%) and embryo transfer (100%) rates in the Ovsynch+CIDR group were greater (P<0.01) than the estrus synchronization (66.7%) and the embryo transfer (61.1%) rates in the PGF group. The postpartum interval at ET in the Ovsynch+CIDR group (62.5 +/- 2.5 d) was shorter (P<0.01) than in the PGF group (74.9 +/- 3.9 d). The pregnancy rate in the Ovsynch+CIDR group (57.9%) did not differ significantly from that in the PGF group (50.0%). Plasma progesterone concentrations were not significantly different in the two groups on Days 0, 1, 2, 5, 7, 14 and 21. In summary, higher synchronization and transfer rates, and shorter postpartum interval to ET, can be achieved with timed ET following the Ovsynch plus CIDR protocol than after estrus with the single PGF(2 alpha) treatment followed by ET in suckled postpartum recipient beef cows. Pregnancy rates were similar. Also, the increase in blood progesterone concentrations during luteal development following ovulation synchronized by the Ovsynch plus CIDR protocol was similar to that after estrus induced by the PGF(2 alpha) treatment.