The effect of ovary storage and in vitro maturation on mRNA levels in bovine oocytes; a possible impact of maternal ATP1A1 on blastocyst development in slaughterhouse-derived oocytes

Since BSE testing of slaughtered cattle is obligatory in Japan, storage of ovaries at 15-20 C overnight in phosphate buffered saline has become a routine protocol in in vitro production (IVP) of cattle embryos. Ovary storage is known to reduce developmental competence of oocytes; however, its effects on oocyte gene expression have not been clarified yet. This study compared oocytes collected from stored slaughterhouse-derived ovaries with those collected by Ovum Pick-Up (OPU) in terms of the expression of 20 selected genes to determine if ovary storage affects cellular processes at the molecular level. Expression of mRNA in oocytes was assayed before and after in vitro maturation (IVM) by real-time quantitative PCR. Maternal mRNA levels of genes were investigated in 2-cell stage embryos obtained from slaughterhouse oocytes to assess their roles for blastocyst formation. In immature OPU oocytes, genes related to metabolism (GAPDH), transporters (GLUT8, ATP1A1) and stress resistance protein (HSP70) showed significantly higher expression compared with oocytes derived from stored ovaries. During IVM, the expression of GDF9, GLUT8, CTNNB1 and PMSB1 was significantly decreased irrespective of oocyte source. Two-cell stage embryos cleaving at 22-25 h after in vitro fertilization (IVF) showed a significantly higher blastocyst formation rate and ATP1A1 gene expression level compared with those cleaving at 27-30 h after IVF. Our results reveal that storage of ovaries alters mRNA levels in oocytes. Correlation of Na/K ATPase ATP1A1 expression in IVP embryos at the 2-cell and 8-cell stages with their developmental ability to the blastocyst stage may suggest the importance of maternal mRNA of this gene during blastulation in embryos derived from slaughterhouse oocytes.     

Fine mapping the number of corpora lutea quantitative trait loci on SSC3: Analysis of the porcine follicle‐stimulating hormone receptor gene

In females, follicle‐stimulating hormone (FSH) targets a FSH receptor (FSHR) expressed only on granulose cells, inducing maturation of the ovarian follicles. We hypothesized that genetic variants in the FSHR gene influence litter size by affecting the number of corpora lutea. We fine‐mapped a region of Sus Scrofa chromosome 3 that contains quantitative trait loci for corpora lutea. Polymorphisms were detected in the exons and 5′ flanking region of the porcine FSHR gene, a positional candidate for the statistically most significant of the quantitative trait loci. Finally, 248 F₂ animals from a Duroc and Meishan cross were genotyped for three FSHR SNPs at positions 74, 532 and 1166, and these were correlated with the phenotypes of litter size and corpus luteum number. Three haplotypes were identified: M1 (G/G/C), M2 (C/A/T) and D (C/A/C). In the F₂ population, the M1 haplotype was associated with a greater number of corpora lutea (P < 0.01) and also seemed to be associated with increased litter size, although the association was not significant (P = 0.2571). Some polymorphisms resulting in amino acid substitutions in these genes were excluded from the polymorphisms possibly responsible for the number of corpora lutea.     

Effect of ovary lipid from Skipjack tuna (Katsuwonus pelamis) on brain monoamines in rats

In our previous experiments with rats, ovary lipid from Skipjack Tuna (Katsuwonus pelamis) (OLS) was shown to have a mitigating effect on anxiety and/or fear in elevated T-maze tests. This suggests that OLS has some effect on the central nervous system (CNS) of rats. Thus, we performed experiments to examine the status of CNS in rats given OLS. The effect of OLS on chronic stress was also examined at the same time. The feed for control rats used oil and fat that have the same energy percentages for n-6 and n-3 fatty acids and the same n-6/n-3 ratio as OLS. As a result, rats given OLS for 28 days had lower serotonin levels in various brain areas regardless of stress application, showing that OLS affected the serotonin nervous system. From this, it was inferred that the ability of OLS to mitigate anxiety and/or fear resulted from its action on CNS, especially the serotonin nervous system. Substances other than the essential fatty acids may have been responsible for the action of OLS on monoamines and the metabolites. The effect of OLS on CNS, especially the serotonin nervous system, suggests that OLS may suppress anxiety.     

Allele‐specific polymerase chain reaction typing and sequencing of mitochondrial D‐loop region in broiler chickens in Japan

This study aimed to comprehend a feature of single nucleotide polymorphism (SNP) in mitochondrial DNA (mtDNA) mainly of general broiler chickens in Japan. We typed two SNP sites (199C/T and 792A/G) of the D‐loop region in mtDNA by allele‐specific PCR (AS‐PCR) in 359 broiler (182 chunky and 177 cobb) and 506 layer (233 White Leghorn, 140 Barred Plymouth Rock and 133 Rhode Island Red) chickens. The SNP of 199C or 792A by AS‐PCR was observed in the chunky and cobb chickens, and not in the layers. The haplotype 199T/792G was observed in a part of cobb and all layers. By the result of AS‐PCR haplotyping and the broiler brands, the D‐loop region was sequenced in 44 broiler chickens (20 chunky and 24 cobb) and compared with the layers’ sequence data. Among the broiler and layer chickens, 21 SNP sites (including one insertion) and 11 sequence haplotypes were observed. Haplotype variation or correspondence was observed in and between the broiler brands. This study provides important information to establish a chicken meat traceability system by SNP haplotyping of mtDNA in Japan.     

The g.841G>C SNP of FASN gene is associated with fatty acid composition in beef cattle

The objective of the current study is to evaluate the association between fatty acid composition and fatty acid synthase gene polymorphisms as responsible mutations. For this purpose, we selected seven previously reported single nucleotide polymorphisms (SNPs) in FASN gene, including one within promoter region (g.841G>C) and six non‐synonymous SNPs (g.8805C>T, g.13126C>T, g.15532A>C, g.16024A>G, g.16039C>T, g.17924A>G), and genotyped them in Japanese Black cattle. Genotyping results revealed that g.8805 C>T and g.17924 A>G were monomorphic loci. Genome‐wide association analysis including the other five SNPs revealed that only g.841G>C showed significant associations with the percentages of C14:0, C14:1, C16:1 and C18:1 at 5% genome‐wide significance level. In order to further evaluate the effect, we genotyped g.841G>C using additional three populations, including two Japanese Black populations and a Holstein cattle population. g.16024A>G was also genotyped and included in the analysis because it has been reported to be associated with fatty acid composition in Japanese Black cattle. In the result of analysis of variance, g.841G>C showed stronger effects on fatty acid percentage than those of g.16024A>G in all populations. These results suggested that g.841G>C would be a responsible mutation for fatty acid composition and contribute to production of high‐grade beef as a selection marker in beef cattle.     

Easy Preparation of Dietary Fiber with the High Water-Holding Capacity from Food Sources

Dietary fibers were prepared as alkali- and acid-insoluble fractions with chemical phosphorylation from Tossa jute (Corchorus olitorius), defatted soybean (Glycine max), and Shiitake (Lentinula edodes). The dietary fiber fractions treated with alkaline solution containing sodium metaphosphate had the lower protein content and higher total dietary fiber content than those of the preparations without phosphorylation. Alkaline extraction followed by phosphorylation led to a 1.5-fold increase in the water holding capacity of dietary fiber compared with no phosphorylation, whereas the binding capacity to bile acids of dietary fiber was almost the same. The alkali- and acid-insoluble extraction with phosphorylation provided an efficient preparation of water-insoluble dietary fiber with high-water holding capacity from various food sources.     

Can harvest outsourcing services reduce field harvest losses of rice in China?

The purpose of this study is to quantify the rice harvest losses in China and to evaluate the impacts of machinery and harvest outsourcing services on these losses, given the background of high-speed mechanization and outsourcing services. Data were collected from a national survey conducted in 2016 by the research team in conjunction with the Research Center for the Rural Economy of the Ministry of Agriculture and Rural Affairs of China. A non-parametric method was used to test whether combine harvesting and outsourcing services could significantly reduce harvest losses. Next, quantile regression was used to estimate the real effects of machinery and outsourcing services on harvest losses. The analysis yielded four main study outcomes. First, the harvest loss rate of rice in China was 3.65%. Second, mechanical reaping and winnowing caused greater losses than manual methods, while the opposite was true of field transportation. Third, combine harvesting increased the losses. Fourth, the effects of an outsourcing service on losses differed among the different harvesting methods. Outsourcing services increased losses in segmented harvesting but they reduced losses in combine harvesting.     

Clarification of adsorption and movement by predicting ammonia nitrogen concentrations in paddy percolation water

We noted that ammonia nitrogen was not adsorbed by the cultivated layers of highly permeable paddy fields during the initial fertilization period, but reached the lower layers relatively early. In our study, we considered an exponential equation from an aqua-environmental perspective with the goal of obtaining good growth of rice plants in order to estimate the concentrations and integrated volume of ammonia nitrogen accompanying paddy percolation. Using this exponential equation, we were able to derive a relation between time and concentrations of paddy percolation water, and hypothesized that if percolation rates were less than 10 mm/day, percolation would have no effect on rice growth, while simultaneously helping to maintain the good water quality of the extra-paddy environment. We also clarified the differences between the potential ammonia nitrogen adsorption volume derived from the CEC value and the integrated amount of ammonia nitrogen water in soil, and considered the causes from the perspectives of solute movement and water movement. Electronic Publication     

Pathology of cutaneous fowlpox with amyloidosis in layer hens inoculated with fowlpox vaccine

Cutaneous fowlpox occurring in vaccinated layer hens was investigated pathologically and microbiologically. Anorexia, decrease of egg production, increased mortality, yellow scabs on faces, and alopecia of feathered skins with yellow scabs were observed in affected hens. Histologically, proliferative and necrotic dermatitis with eosinophilic ring-shaped cytoplasmic inclusions (Bollinger bodies) and clumps of gram-positive cocci (Staphylococcus hyicus) were noted in the affected birds. Fowlpox lesions were primarily observed in the feathered skins. Proliferation of feather follicle epidermal cells, with cytoplasmic inclusions and degeneration of the feather, and bacterial clumps in the feather follicles were noted in the affected skins. Ultrastructurally, characteristic fowlpox viral particles were observed in the cytoplasmic inclusions of hyperplastic epidermal cells. Amyloid deposition was observed in the Disse space of the liver, splenic sinus, and lamina propria of the bronchiolar, bronchial, and tracheal areas. Amyloidosis could be one factor inducing the fowlpox infection in vaccinated chickens.