Identification of a potent immunostimulatory oligodeoxynucleotide from Streptococcus thermophilus lacZ

Immunostimulatory sequences of oligodeoxynucleotides (ODNs), such as CpG ODNs, are potent stimulators of innate immunity. Here, we identified a strong immunostimulatory CpG ODN, which we named MsST, from the lac Z gene of Streptococcus (S.) thermophilus ATCC19258, and we evaluated its immune functions. In in vitro studies, MsST had a similar ability as the murine prototype CpG ODN 1555 to induce inflammatory cytokine production and cell proliferation. In mouse splenocytes, MsST increased the number of CD80+CD11c+and CD86+CD11c+ dendritic cells and CD4+CD25+ regulatory T cells. We also analyzed the effects of MsST on the expression of regulatory cytokines by real-time quantitative PCR. MsST was more potent at inducing interleukin-10 expression than the ODN control 1612, indicating that MsST can augment the regulatory T cell response via Toll-like receptor 9, which plays an important role in suppressing T helper type 2 responses. These results suggest that S. thermophilus , whose genes include a strong Immunostimulatory sequence-ODN, is a good candidate for a starter culture to develop new physiologically functional foods and feeds.     

Microbiota analysis of Caspian Sea yogurt, a ropy fermented milk circulated in Japan

We analyzed the microbiota of domestic ropy fermented milk, Caspian Sea yogurt (or 'kasupikai yohguruto' in Japanese), circulated in Japan. We collected six varieties from five localities. Lactococcus ( L) lactis ssp. cremoris was isolated from all samples as the dominant strain at levels of 10⁸−10⁹ CFU/g. We show this strain produces an extracellular polysaccharide (EPS) that causes the unique characteristic viscosity of the product. From analysis of the RAPD pattern of 60 bacterial isolates from the six samples, we found that 59 strains from a total of 60 isolates were identical and produced this viscosity. Furthermore, PFGE analysis of representative strains from each sample indicated that the isolates could be classified into four subgroups. This suggests these L. lactis ssp. cremoris strains found in Caspian Sea yogurt may have been slightly mutated during subculture in Japan. In addition, Lactobacillus (L.) sakei ssp. sakei were isolated from three samples; L. plantarum , Gluconoacetobacter sacchari and Acetobacter aceti were isolated from two samples; and L. paracasei , L. kefiri , Leuconostoc ( Leu. ) mesenteroides were isolated from one sample.     

Effect of ensiled hop (Humulus lupulus L.) residues on plasma acetate turnover rate in sheep

An isotope dilution method using [1‐¹³C]sodium acetate was applied to determine the effect of feeding ensiled hop (Humulus lupulus L.) residues on plasma acetate turnover rate in six adult crossbred sheep. The sheep were fed 63 g/kg body weight (BW)^0.75/day of either mixed hay of orchardgrass (Dactylis glomerata L.) and reed canarygrass (Phalaris arundinacea L.) and round bale silage at 3:1 ratio (Hay‐diet), or another where round bale silage was replaced by ensiled hop residues (Hop‐diet) with a crossover design each of a 3‐week period. The isotope dilution method was performed on day 21 of each dietary treatment. Dry matter digestibility was similar between diets, and nitrogen (N) digestibility was lower (P = 0.001) for Hop‐diet than Hay‐diet. However, N retention did not differ between diets. Plasma acetate concentration was lower (P = 0.04) for Hop‐diet than Hay‐diet, and the turnover rate of plasma acetate did not differ between diets. Plasma concentration of lactate and non‐esterified fatty acids were similar between diets. Hop‐diet was found almost comparable to Hay‐diet on plasma acetate turnover rate in the present experimental conditions. Therefore, it could be concluded that hop residues partially could be used as an alternative to traditionally used round bale silage for rearing sheep.     

DNA methylation analysis on satellite I region in blastocysts obtained from somatic cell cloned cattle

Many observations have been made on cloned embryos and on adult clones by somatic cell nuclear transfer (SCNT), but it is still unclear whether the progeny of cloned animals is presenting normal epigenetic status. Here, in order to accumulate the information for evaluating the normality of cloned cattle, we analyzed the DNA methylation status on satellite I region in blastocysts obtained from cloned cattle. Embryos were produced by artificial insemination (AI) to non‐cloned or cloned dams using semen from non‐cloned or cloned sires. After 7 days of AI, embryos at blastocyst stage were collected by uterine flushing. The DNA methylation levels in embryos obtained by using semen and/or oocytes from cloned cattle were similar to those in in vivo embryos from non‐cloned cattle. In contrast, the DNA methylation levels in SCNT embryos were significantly higher (P < 0.01) than those in in vivo embryos from non‐cloned and cloned cattle, approximately similar to those in somatic cells used as donor cells. Thus, this study provides useful information that epigenetic status may be normal in the progeny of cloned cattle, suggesting the normality of germline cells in cloned cattle.     

中国和日本农业活动对流域氮素收支的影响

To analyze the effect of agricultural activity on nitrogen(N) budget at the watershed scale,a comparative study was conducted at two Japanese watersheds,the Shibetsu River watershed(SRW) and Upper-Naka River watershed(UNRW),and one Chinese watershed,the Jurong Reservoir watershed(JRW).The total area and the proportion of agricultural area(in parentheses) of the watersheds were 685(51%),1 299(21%),and 46 km 2(55%) for SRW,UNRW,and JRW,respectively.The main agricultural land use in SRW was forage grassland,while paddy fields occupied the highest proportion of cropland in UNRW(11% of total area) and JRW(31% of total area).The farmland surplus N was 61,48,and 205 kg N ha 1 year 1 for SRW,UNRW,and JRW,respectively.The total input and output for the whole watershed were 89 and 76,83 and 61,and 353 and 176 kg N ha 1 year 1 for SRW,UNRW,and JRW,respectively.The proportion of discharged N to net anthropogenic N input was 31%,37%,and 1.7% for SRW,UNRW,and JRW,respectively.The two watersheds in Japan showed similar proportions of discharged N to those of previous reports,while the watershed in China(JRW) showed a totally different characteristic compared to previous studies.The high N input in JRW did not increase the amount of discharged N at the outlet of the watershed due to high proportions of paddy fields and water bodies,which was an underestimated N sink at the landscape scale.     

The effects of potato pulp and feeding level of supplements on digestibility, in situ forage degradation and ruminal fermentation in beef steers

Six ruminally cannulated Wagyu (Japanese Black) steers (average initial bodyweight (BW) 387 ± 29 kg) were used in a split‐plot design experiment, comprising a 3 × 3 Latin square design (whole plot) and a randomized block design (subplot). The whole plot treatments were three different feeding levels of supplemental diet, fed at 0.2, 0.4 and 0.6% of BW, on a dry matter (DM) basis. Subplot treatments were two different supplemental diets: a potato pulp silage‐based diet (PPS) and a grain‐based diet (GRAIN). Chopped, medium‐quality cool‐season grass hay (predominately Timothy, Phleum pratense L) was fed daily at 0.7% BW (on a DM basis) as the basal diet. Each period consisted of 21 d, which included 11 d of adaptation to the diets and 10 d of the collection period. Chromium oxide was used as an indigestible marker. In situ forage degradation was measured using the nylon bag technique. The dry matter intake increased (linear; P < 0.01) as the feeding level increased and was not affected by the diet. Digestibility was not affected by any treatments. The GRAIN diet tended to decrease the rate of in situ forage degradation as the feeding level increased, but this trend was not found in the steers fed the PPS diet. Steers fed the GRAIN diet had a lower (P < 0.1) ruminal pH compared with steers fed the PPS diet. Ruminal pH was not significantly affected by feeding level; however, it was numerically higher for steers supplemented at 0.2% per BW than that for the steers supplemented above 0.4% per BW due probably to the higher starch intake. The total volatile fatty acids concentration numerically increased as the feeding level increased and was not affected by the diet. Increasing the feeding level decreased (linear, P < 0.01) the proportion of acetate. Neither diet nor the feeding level had any effects on the proportion of ruminal propionate. The results suggested that, for steers fed the PPS diet, there are not adverse effects on forage digestion in the rumen that occur as the feeding level is increased.     

Estimation of the carcass composition from a cross-section of the rib-loin from crossbred Japanese Black × Limousin F2 cattle by computer image analysis

The carcass composition of crossbred Japanese Black × Limousin F2 cattle was examined in order to find an accurate carcass composition equation. The test animals included 17 steers and 17 heifers. The 28 image measurements from the area encircling the vertical line to the thoracic vertebra and the line from the thoracic vertebra between the sixth and seventh rib‐bones were measured by computer image analysis. The relationships between the 29 parameters that added the carcass left side weight of the animal and the carcass composition were suggested. The carcass composition included muscle weight, muscle ratio, fat weight and fat ratio. The carcass composition from steers was estimated by an equation composed of these three or four parameters (R² = 90.80%, 79.30%, 90.75% and 73.70%, respectively). The selected parameters were measured without cutting the thoracic vertebra. The carcass composition from heifers was estimated by an equation composed of two to four parameters (R² = 96.15%, 90.98%, 93.60% and 88.22%, respectively). The parameters for the estimation of the muscle and fat weight, and muscle and fat ratio are very similar. Furthermore, the equations using the parameters could estimate the carcass composition from the Japanese Black × Limousin cattle resource population.     

Physiological Races and Vegetative Compatibility Groups of Fusarium oxysporum f. sp. lactucae Isolated from Crisphead Lettuce in Japan

One hundred and sixteen isolates of Fusarium oxysporum f. sp. lactucae obtained from 85 fields in three crisphead lettuce-producing areas in Nagano Prefecture, Japan were typed for races using differential cultivars Patriot, Banchu Red Fire and Costa Rica No. 4. They were also grouped into vegetative compatibility groups (VCGs) using complementation tests with nitrate non-utilizing (nit) mutants. Two California strains reported as F. oxysporum f. sp. lactucum, a type culture of F. oxysporum f. sp. lactucae, and 28 avirulent isolates of F. oxysporum obtained from crisphead lettuce were included for comparison. Among Nagano isolates, 66 isolates were identified as race 1, and 50 as race 2. Race 1 strains derived from Shiojiri and Komoro cities and race 2 from Kawakami village and Komoro city. All isolates of race 2 were biotin auxotrophs, and the race could be distinguished based on its requirement for biotin on minimal nitrate agar medium (MM). Pathogenic isolates were classified into two VCGs and three heterokaryon self-incompatible isolates. Strong correlations were found between race and VCG. All the race 1 strains were assigned to VCG 1 except self-incompatible isolates, and all the race 2 strains to VCG 2. The 28 avirulent isolates of F. oxysporum were incompatible with VCG 1 and VCG 2. California strains was vegetatively compatible with VCG 1, and they were assigned to race 1. Based on vegetative compatibility, these two races of F. oxysporum f. sp. lactucae may be genetically distinct, and F. oxysporum f. sp. lactucae race 1 is identical to F. oxysporum f. sp. lactucum. Received 7 May 2002/ Accepted in revised form 6 September 2002