Genetic succession and spatial genetic structure in a natural old growth Cryptomeria japonica forest revealed by nuclear and chloroplast microsatellite markers

Spatial genetic structure and diversity of Cryptomeria japonica trees in old growth forest but selectively logged in approximately 300 years ago were analyzed using seven nuclear microsatellite markers and one chloroplast microsatellite marker. The individuals were sampled from a 4-ha natural forest plot (200 m × 200 m), which are consisted of three size classes including newly regenerated individuals such as regenerated individuals (REG class; stem diameter at breast height (DBH), 5–110 cm), remaining individuals (REM class; DBH ≥ 110 cm), and old stumps. Genetic diversity was similar between the three groups. These results suggest that genes from the old growth forest may have been passed on to the next generation (i.e., the current forest). DNA extraction and analysis from old stumps was partly successful and we could compare the genetic diversity between old stumps and living trees using four microsatellite loci. Patterns of spatial genetic structure detected by analyses of the nuclear and chloroplast markers differed substantially; all nuclear markers showed significant positive autocorrelation (P < 0.05) over short distances, but the chloroplast marker showed no significant autocorrelation over any distance. We concluded that Moran's I spatial autocorrelation pattern for the chloroplast microsatellite may have been influenced by the extensive pollen flow characteristic of this species, because chloroplast DNA of C. japonica is paternally inherited through pollen and is a wind-pollinated species that produces abundant pollen every few years. In contrast, Moran's I spatial autocorrelation pattern for nuclear microsatellites may reflect limited seed dispersal in gaps where the forest has regenerated and low levels of seed shadow overlap due to past logging. The average seed dispersal distance was 86.0 m at the study site and about 34.0% of the seeds were from the outside of the site, which revealed by parentage analysis. The individuals remaining after logging, and the extensive pollen flow from outside our study plot, may play important roles in maintaining the forests on this island. Gaps created by the selective logging activities appear to have been important for the regeneration of this species in the past but in the present time, natural disturbances such as typhoon and landslide are also important for gap creation.     

Software development for estimating the concentration of radioactive cesium in the skeletal muscles of cattle from blood samples

The 2011 earthquake severely damaged the Fukushima Daiichi Nuclear Power Plant (FNPP), resulting in the release of large quantities of radioactive material into the environment. The deposition of these radionuclides in rice straw as livestock feed led to the circulation of contaminated beef in the market. Based on the safety concern of the consumers, a reliable method for estimating concentrations of radioactive cesium in muscle tissue is needed. In this study, we analyzed the concentrations of radioactive cesium in the blood and skeletal muscle of 88 cattle, and detected a linear correlation between them. We then developed software that can be used to estimate radioactive cesium concentrations in muscle tissue from blood samples. Distribution of this software to the livestock production field would allow us to easily identify high‐risk cattle, which would be beyond the safety regulation, before shipping out to the market. This software is planned to be released as freeware. This software would contribute to food safety, and aid the recovery of the livestock industry from the damage creacted by the 2011 Tohoku earthquake and tsunami.     

Genetic and antigenic characterization of bovine viral diarrhea viruses isolated from cattle in Hokkaido,Japan

In our previous study, we genetically analyzed bovine viral diarrhea viruses (BVDVs) isolated from 2000 to 2006 in Japan and reported that subgenotype 1b viruses were predominant. In the present study, 766 BVDVs isolated from 2006 to 2014 in Hokkaido, Japan, were genetically analyzed to understand recent epidemics. Phylogenetic analysis based on nucleotide sequences of the 5′-untranslated region of viral genome revealed that 766 isolates were classified as genotype 1 (BVDV-1; 544 isolates) and genotype 2 (BVDV-2; 222). BVDV-1 isolates were further divided into BVDV-1a (93), 1b (371) and 1c (80) subgenotypes, and all BVDV-2 isolates were grouped into BVDV-2a subgenotype (222). Further comparative analysis was performed with BVDV-1a, 1b and 2a viruses isolated from 2001 to 2014. Phylogenetic analysis based on nucleotide sequences of the viral glycoprotein E2 gene, a major target of neutralizing antibodies, revealed that BVDV-1a, 1b and 2a isolates were further classified into several clusters. Cross-neutralization tests showed that BVDV-1b isolates were antigenically different from BVDV-1a isolates, and almost BVDV-1a, 1b and 2a isolates were antigenically similar among each subgenotype and each E2 cluster. Taken together, BVDV-1b viruses are still predominant, and BVDV-2a viruses have increased recently in Hokkaido, Japan. Field isolates of BVDV-1a, 1b and 2a show genetic diversity on the E2 gene with antigenic conservation among each subgenotype during the last 14 years.     

Effect of Bacillus subtilis C-3102 supplementation in milk replacer on growth and rumen microbiota in preweaned calves

We aimed to assess the effect of feeding Bacillus subtilis C-3102 on the growth and rumen microbiota in the preweaned calves. Twelve newborn Japanese Black calves were randomly allocated to either the control (n = 6) or the treatment (n = 6) groups in the present study. Calves in the treatment group were offered B. subtilis C-3102 supplemented milk replacer throughout the preweaning period. Rumen fermentation during the first 21 days of life seemed to be slightly suppressed by feeding B. subtilis C-3102. This fermentation shift was probably attributed to the lower abundance of the core members of rumen microbiota until 21 days of age in the calves fed B. subtilis C-3102. However, feeding B. subtilis C-3102 did not influence the abundance of the core members of rumen microbiota at 90 days of age. Distribution of Sharpea spp. and Megasphaera spp., which potentially contribute to low methane production and are regarded as beneficial rumen bacteria, was higher in the rumen of calves fed B. subtilis C-3102 at 90 days of age. These results suggest that B. subtilis C-3102 supplementation in milk replacer could potentially contribute to the improvement of feed efficiency after weaning via the establishment of beneficial rumen bacteria.     

Estimation of concentration of radionuclides in skeletal muscle from blood,based on the data from abandoned animals in Fukushima

The damage caused by the earthquake on 11 March, 2011 resulted in a serious nuclear accident in Japan. Due to the damage to the Fukushima Daiichi Nuclear Power Plant (FNPP), large amounts of radioactive substances were released into the environment. In particular, one of the largest safety concerns is radioactive cesium (¹³⁴Cs and ¹³⁷Cs). Due to the FNPP nuclear accident, a 20 km area was restricted from human activity, and various types of domestic animals were left in the zone. We collected the organs and tissues from sacrificed animals to obtain scientific data to evaluate the internal deposition of radioactive compounds. At first, we found there is a strong correlation between blood ¹³⁷Cs and organ ¹³⁷Cs with data from 44 cattle, indicating that skeletal muscle is the target organ of deposition of radioactive cesium. Second, we analyzed the relationship between blood ¹³⁷Cs and muscle ¹³⁷Cs within relatively lower radioactive concentration, suggesting that estimation of concentration of ¹³⁷Cs is possible from blood concentration of ¹³⁷Cs. Finally, we developed computer software to estimate the muscle ¹³⁷Cs concentration from blood samples. Our study contributes to the food safety of livestock products.     

Leptospiral lipopolysaccharide stimulates the expression of toll‐like receptor 2 and cytokines in pig fibroblasts

Pigs throughout the world are afflicted with leptospirosis, causing serious economic losses and potential hazards to human health. Although it has been known that leptospiral lipopolysaccharide (L‐LPS) is involved in an immunological reaction between an antigen and a host cell, little is known about how the immune system of pigs can respond to L‐LPS. Here, we stimulated pig fibroblasts by L‐LPS and then quantitatively measured gene and protein expression levels of two toll‐like receptors (TLRs), TLR2 and TLR4, by real‐time PCR and Western blotting. As a result, expression of TLR2 was found to be significantly up‐regulated within 24 h after L‐LPS stimulation whereas induction of TLR4 expression was relatively weak. We also revealed that of myeloid differentiation primary response gene 88 (MyD88), interleukin 6 (IL‐6) and IL‐8 gene expressions were markedly up‐regulated by L‐LPS stimulation. These results may suggest that the pig cell can activate TLR2 rather than TLR4 by L‐LPS stimulation, thereby inducing expression of cytokines.     

Cesium radioactivity in peripheral blood is linearly correlated to that in skeletal muscle: Analyses of cattle within the evacuation zone of the Fukushima Daiichi Nuclear Power Plant

The accident at the Fukushima Daiichi Nuclear Power Plant (FNPP) released a large amount of radioactive substances into the environment. Furthermore, beef contaminated with radioactive cesium above the 500 Bq/kg safety standard was circulated in the food chain in 2011. Japanese consumers remain concerned about the safety of radioactively contaminated food. In our previous study, we detected a linear correlation between radioactive cesium (¹³⁷Cs) activity in blood and muscle around 500 to 2500 Bq/kg in cattle. However, it was unclear whether the correlation was maintained at a lower radioactivity close to the current safety standard of 100 Bq/kg. In this study, we evaluated 17 cattle in the FNPP evacuation zone that had a ¹³⁷Cs blood level less than 10 Bq/kg. The results showed a linear correlation between blood ¹³⁷Cs and muscle ¹³⁷Cs (Y = 28.0X, R² = 0.590) at low radioactivity concentration, indicating that cesium radioactivity in the muscle can be estimated from blood radioactivity. This technique would be useful in detecting high‐risk cattle before they enter the market, and will contribute to food safety.     

Effect of sorbitol on moisture transportation and textural change of fish and squid meats during curing and drying processes

SUMMARY: Atka mackerel (Am) and Japanese common squid (Sq) meats were cured in 0.5–1.5 M sorbitol solutions (pH 7.0) and dried at 30°C (relative humidity, 60%), and the effect of sorbitol on the moisture transportation and textural change during the curing and drying processes was investigated. With an increase in sorbitol permeated through samples, the moisture contents decreased by 52% (Am) and 42% (Sq) by curing in 1.5 M sorbitol solution. When the cured meats were dried, slow moisture vaporization occurred at the initial drying period, and the critical moisture content significantly decreased with an increase in the sorbitol content of the cured meats. Further, the hardening of the dried products was effectively suppressed by sorbitol curing. These effects of sorbitol would contribute to the reduction of drying time and particularly the elimination of the excess hardening of dried fish products.     

Optimum Dietary Level of L-ascorbic Acid for Japanese Eel, Anguilla japonica

A feeding experiment was conducted to determine the optimum dietary level of ascorbic acid (AsA) for Japanese eel juveniles using L-ascorbic acid Ca based on growth, AsA content in tissues, hematology, and bactericidal activity of serum with Escherichia coli . Test diets with six levels of AsA (3, 10, 27,126, 645, and 3,135-mg/kg diet) as Ca ascorbate were fed to juvenile Japanese eels (11.0 ± 0.2 g) once a day for 8 wk. High survival rates (> 80%) were observed among all dietary treatment groups. The specific growth rates of the fish fed diets containing 3 and 10-mg AsA/kg were significantly (P < 0.05) lower than those of other groups. Liver and brain AsA contents of the fish fed diets containing 3, 10, and 27-mg AsA/kg were significantly (P < 0.05) lower than those of the fish fed diets containing 645 and 3,135-mg AsA/kg. Hemoglobin content tended to be higher in the fish fed diets containing 645 and 3,135-mg AsA/kg than those of the fish fed other diets. Hematocrit value and total serum protein content of the fish fed diets containing 645 and 3135 mg AsA/kg were significantly (P < 0.05) higher than those of the other groups. The fish fed diets containing more than 27-mg AsAlkg showed a higher bactericidal activity of serum than the fish fed the diets containing 3-mg and 10-mg AsA/kg. The optimum dietary level of AsA for the Japanese eel juveniles growth was estimated to be more than 27-mg AsA/kg. Furthermore, the inclusion of 645-mg AsA/kg or more also increased the hematocrit, hemoglobin, total serum protein value, and liver and brain vitamin C concentrations.     

Biometric analysis on diversity of coconut palm: cultivar classification by botanical and agronomical traits

Botanical and agronomical traits were observed using 39 cultivars of coconut palms which mainly collected in the PhilipApines, and statistically analyzed to clarify the variation between and within cultivar groups (typica, nana and javanica). Although there were broad variations in all the traits except for several male flower characters, significant differences among three cultivars were found in a dozen of traits. The variation within a cultivar group was higher in typica and javanica. Nana was noted as an aggregate group, which was far distance from typica. Javanica was characterized as the intermediate group having overlapping boundaries with other groups. This revised version was published online in July 2006 with corrections to the Cover Date.