Development of a simple method for sperm cryopreservation of Scombridae fishes in outdoor environments

We developed a simple dry shipper method for cryopreserving the sperm of Scombridae fish in outdoor environments. First, we undertook a preliminary study to optimize the sperm cryopreservation conditions using bullet tuna, Auxis rochei (Risso, 1810) sperm. We found that the optimum cryomedium contained 90% foetal bovine serum (FBS) or 300 mM trehalose as an external cryoprotectant and 10% dimethyl sulfoxide (DMSO) as an internal cryoprotectant. Under these optimized conditions, the post‐thaw sperm had a duration of motility of 500 s and a motility rate of >70%. We then performed practical trials of the optimized protocol in various outdoor environments (e.g., fishing boats and ports) using the sperm of five Scombridae species: chub mackerel, Scomber japonicus (Houttuyn, 1782); blue mackerel, S. australasicus (Cuvier, 1832); skipjack tuna, Katsuwonus pelamis (Linnaeus, 1758); longtail tuna Thunnus tonggol, (Bleeker, 1851) and Pacific bluefin tuna, T. orientalis (Temminck & Schlegel, 1844). The post‐thaw sperm of all five of these species had a duration of motility of 650 s and a motility rate of >70%, indicating that this simple method can be used to obtain high‐quality cryopreserved sperm of various Scombridae species in outdoor environments.     

Properties of extracts from defatted rice bran by its subcritical water treatment

Defatted rice bran was extracted with water and subcritical water at 50-250 degrees C for 5 min. The highest extract yield was achieved at 200 degrees C, at which the maximum amounts of protein and carbohydrate were also obtained. The total phenolic and furfural contents, radical scavenging activity, and antioxidative activity for the autoxidation of linoleic acid increased with increasing treatment temperature. The bran extracts exhibited emulsifying activity except for the extract prepared at 250 degrees C, which was concomitant with the disappearance of its high-molecular-mass substances. The extract prepared at 200 degrees C also had the highest emulsion-stabilizing activity.     

Formation of Enzyme‐Resistant Starch in Bread as Affected by High‐Amylose Wheat Flour Substitutions

High‐amylose wheat flour was used to substitute for normal wheat flour in breadmaking and formation of resistant starch (RS) in bread during storage was determined. Substitution with high‐amylose wheat flour (HAF) decreased peak and final viscosities, breakdown, and setback. Doughs with HAF substitutions were weaker and less elastic, and absorbed more water than those of the normal wheat flour. After baking, RS contents in breads with 10, 30, and 50% HAF substitutions were 1.6, 2.6, and 3.0% (db), respectively, higher than that of the control (0.9%, db). The levels of RS increased gradually during storage for one, three, and five days. With substitutions of 30 and 50% HAF, the total levels of dietary fiber (DF) and RS in bread after five days of storage were 15.5 and 16.8% (db), respectively, as compared to 13.0% (db) in bread from the normal wheat flour. The loaf volumes and appearances of bread crumbs made from HAF substitutions of 10 and 30% were not significantly different from those of the control, whereas the substitution with 50% HAF decreased loaf volume and resulted in inferior appearance of breadcrumbs. The firmness of breadcrumbs increased along with increase in the level of HAF substitutions after baking. During storage, the firmness of breadcrumb with 10% HAF substitutions was higher than that of the control, whereas breads with 30 and 50% HAF substitutions had similar firmness to the control. As a result, HAF might be used to substitute for up to 50% normal wheat flour to make bread with acceptable bread quality and significantly high amount of RS.     

Molecular typing by pulsed-field gel electrophoresis of Flavobacterium psychrophilum isolates derived from Japanese fish

Sixty-four isolates of Flavobacterium psychrophilum from ayu, Plecoglossus altivelis altivelis (Temminck & Schlegel), and other fish (n=16) in Japan and the type strain (NCIMB 1947(T)) were typed using pulsed-field gel electrophoresis (PFGE) with endonuclease BlnI and XhoI. These isolates were classified into 20 clusters and 42 genotypes by PFGE analysis. The most predominant cluster of isolates from ayu was cluster XII (n=20), followed by clusters XVII, XVI, XX, XI, IX, X, XIII and XV; the remaining 17 isolates from other fish were divided into clusters I, II, III, IV, V, VI, VII, VIII, XIV, XVIII and XIX. The PFGE genotype of isolates from ayu clearly differed from those of other fish. The isolates from ayu in Gunma Prefecture belonged to clusters XII, XVI, XVII and XX, and the strains of three of these clusters (XII, XVII and XX) were isolated from ayu in 15 of 19 prefectures. PFGE typing enabled more accurate classification of isolates into clusters than previously achieved by analysing the restriction fragment length polymorphism of PCR products. These results suggest that F. psychrophilum isolated from ayu and other fish are genetically different and strains with several PFGE types have spread within Japan.     

DRIS and geostatistics indices for nutritional diagnosis and enhanced yield of fertirrigated acai palm

Abstract

Information about Açaí palm (Euterpe oleracea Mart.) nutrition, that gives support for yield increase is sparse. The aim of this study was to assess the nutritional status of fertigated Açaí palm by the Index called Diagnosis and Recommendation Integrated System (DRIS), as well as the spatial variability of this Index and its productivity. We achieved a sampling of 80 geo-referenced points in an Açaí palm commercial crop area. Then we assessed the yield and contents of N, P, K, Ca, Mg, S, B, Cu, Fe, Mn and Zn. The DRIS evaluation indicated that the frequency of nutrients in suitable status was N?>?S?>?Zn?>?B>Fe?>?K>Ca?>?Mg?>?P>Mn?>?Cu, in deficiency was Mn?>?Ca >?B>Cu?>?Mg?>?Fe?>?K?>?P?>?S>Zn?>?N, and in excess was P?>?Cu?>?Mg?> K?>?N?=?Zn?>?Fe?>?Ca?>?S?=?B?>?Mn. The nutrients N and S were well balanced, whereas Mn, Ca and B were the nutrients with the highest frequency of deficiency. The sampling points were close enough to detect the spatial variability of DRIS Index. Thus, it was possible to observe the patterns for the nutritional deficiencies, occurring at the final part of the irrigation, as well as the variability of the Açaí palm yield. The spatial variability of the DRIS Index was efficient to indicate the points in which fittings in the fertilization doses are required.     

A case of equine motor neuron disease (EMND)

We report a case of EMND in a heavy horse that was bred and trained in Hokkaido, Japan. Clinical symptoms included severe ataxia of all four limbs, tilted head, lethargy, and flaccid lips. Numerous axonal degenerations and swellings were observed in nuclei, mostly in the cerebellar dentate nucleus and the nucleus of the hypoglossal nerve, and in the ventral horn of the spinal cord. In the ventral horn of the spinal cord, neuronal degeneration, swelling, and/or necrosis were observed sporadically. The case was diagnosed as EMND from the clinical symptoms and pathological findings.     

GH-producing mammary tumors in two dogs with acromegaly

Two intact female dogs were admitted for growing mammary tumors. They had symptoms of acromegaly including weight gain, enlargement of the head, excessive skin folds, and inspiratory stridor. Serum concentrations of growth hormone (GH), insulin-like growth factor-I (IGF-I), and insulin were elevated in the two cases. From these findings, both dogs were diagnosed with acromegaly. In case 1, the GH, IGF-I, and insulin levels subsided after removal of the focal benign mammary tumors and ovariohysterectomy. In case 2, those levels subsided after removal of only focal mammary carcinoma. In both cases, immunohistochemical investigations for GH were positive in the mammary tumor cells but not in the normal mammary glands. We concluded that GH-producing mammary tumors caused the present acromegaly.     

Biological characteristics of fish germ cells and their application to developmental biotechnology

We have revealed several unique characteristics of germ cell development using rainbow trout, including the fact that spermatogonia transplanted into the peritoneal cavity of newly hatched embryos migrate toward recipient gonads, that spermatogonia transplanted into female recipients start oogenesis and produce functional eggs and that diploid germ cells transplanted into triploid trout can complete gametogenesis. By combining these unique features of fish germ cells, we established allogeneic and xenogeneic transplantation systems for spermatogonia in several fish species. Spermatogonia isolated from the mature testes of vasa-green fluorescent protein (Gfp) transgenic rainbow trout were transplanted into the peritoneal cavity of triploid masu salmon newly hatched embryos. These spermatogonia migrated toward recipient salmon genital ridges with extending pseudopodia and were subsequently incorporated into them. We further confirmed that the donor-derived spermatogonia resumed gametogenesis and produced sperm and eggs in male and female salmon recipients, respectively. By inseminating the resulting eggs and sperm, we obtained only rainbow trout offspring in the F1 generation, suggesting that the triploid salmon recipients produced functional gametes derived only from donor trout. We further confirmed that this intra-peritoneal transplantation of germ cells is applicable to several marine fishes, which could be of benefit in the production of bluefin tuna that has a large broodstock (>100 kg) and is difficult to maintain in captivity. Gamete production of bluefin tuna could be more easily achieved by generating a surrogate species, such as mackerel, that can produce tuna gametes.     

Effects of storage conditions and pH on chlorine loss in electrolyzed oxidizing (EO) water.

The chlorine loss of electrolyzed oxidizing (EO) water was examined during storage under different light, agitation, and packaging conditions. The chlorine loss of pH-adjusted EO water was also examined. Under open conditions, the chlorine loss through evaporation followed first-order kinetics. The rate of chlorine loss was increased about 5-fold with agitation, but it was not significantly affected by diffused light. Under closed conditions, the chlorine loss did not follow first-order kinetics, because the primary mechanism of chlorine loss may be self-decomposition of chlorine species rather than chlorine evaporation. The effect of diffused light was more significant compared to agitation after two months of storage under closed conditions. The chlorine loss of EO water and commercial chlorinated water decreased dramatically with the increase of pH from the acidic (pH 2.5) to the alkaline (pH 9.0) region.     

Japanese star anise ringspot-associated virus is a distinct emaravirus transmitted by the eriophyid mite (the family Diptilomiopidae)

Japanese star anise (Illicium anisatum L., JSA) is seriously damaged by a ringspot disease in Japan. Herein, to determine the causal agent using high-throughput sequencing, we discovered viral RNAs associated with JSA ringspot disease. We then determined the complete or near-complete nucleotide sequences of these RNAs using Sanger sequencing and RACE. The complementary strand of viral RNAs 1, 2, 3, 4, and 5 encoded a single protein, which shared sequence identity with P1 (RNA-dependent RNA polymerase), P2 (glycoprotein precursor), P3 (nucleocapsid protein), P4 (movement protein), and a protein with unknown function of emaraviruses (genus Emaravirus), respectively; however, the highest amino acid sequence identity for the P1–P5 proteins between JSARaV and known emaraviruses was 41.9%, 30.0%, 30.1%, 52.2%, and 38.0%, respectively, all of which were lower than the species demarcation criterion. Furthermore, RNA segments harbored conserved 12-nt terminal sequences at the 5′- and 3′-termini, and a high complementarity of approximately 20 nt in 5′- and 3′-terminal sequences. Transmission electron microscopy confirmed the presence of virus-like particles. JSA ringspot disease was found to be transmitted by an eriophyid mite (subclass Acari, superfamily Eriophyoidea) that belongs to the family Diptilomiopidae. Taken together, these results identified the virus responsible for the ringspot disease of JSA as a new member of the genus, Emaravirus, which we named as the Japanese star anise ringspot-associated virus (JSARaV). Moreover, this is the first report noting that eriophyid mites of the family Diptilomiopidae are capable of transmitting emaravirus.