Effects of oocyte-derived growth factors on the growth of porcine oocytes and oocyte–cumulus cell complexes in vitro

During oocyte growth and follicle development, oocytes closely communicate with cumulus cells. We examined the effects of oocyte-derived growth factors, growth differentiation factor 9 (GDF9) and bone morphogenetic protein 15 (BMP15), on the growth and acquisition of meiotic competence of porcine oocytes collected from early antral follicles (1.2–1.5 mm). First, we confirmed that GDF9 and BMP15 mRNAs were expressed almost exclusively in the oocytes. Oocyte–cumulus cell complexes (OCCs) collected from early antral follicles were cultured in growth medium supplemented with 0–100 ng/ml of GDF9 or BMP15 for 5 days. GDF9 dose-dependently increased the OCC diameter, while BMP15 did not. GDF9 and BMP15 had no significant effects on oocyte growth (P > 0.05). When OCCs that had been cultured with 50 and 100 ng/ml BMP15 were subjected to a subsequent maturation culture, they expanded fully by gonadotropic stimulation and 49% and 61% of oocytes matured to metaphase II (MII), respectively. In contrast, GDF9 did not promote cumulus expansion, and < 10% of oocytes matured to MII. Based on the difference in cumulus expansion, we compared the expression of luteinizing hormone/choriogonadotropin receptor (LHCGR) and follicle stimulating hormone receptor (FSHR) mRNAs in cumulus cells. The level of LHCGR mRNA was increased in cumulus cells of the BMP15 group, although there were no significant differences in FSHR mRNA levels among the groups. These results suggest that GDF9 promotes the growth of OCCs and that BMP15 promotes LHCGR mRNA expression in cumulus cells during oocyte growth culture, which may contribute to cumulus expansion and oocyte maturation.     

Genetic diversity and differentiation of sea trout (<Emphasis Type="Italic">Salmo trutta</Emphasis>) populations in Lithuanian rivers assessed by microsatellite DNA variation

The genetic diversity and differentiation of sea trout were studied in three river basins in Lithuania: Akmena-Dane, Bartuva, and Nemunas. A total of 282 individuals were genotyped at eight microsatellite loci. A similar level of genetic diversity was found in all of the populations studied: mean allelic richness ranged from 3.64 to 5.03, and average expected heterozygosity ranged from 0.588 to 0.721. Significant genetic divergence was observed among the different river basins as well as between populations within the drainages. All pairwise F _ST values were highly significant, ranging from 0.027 to 0.197. The analysis of molecular variance showed rather weak hierarchical population structuring within the Nemunas basin, which may be explained by extensive gene flow among different river basins or, alternatively, reflect the influence of artificial breeding. Information on genetic diversity and differentiation of the Lithuanian sea trout populations will be useful for future management decisions.     

Genetic Introgression Between Arctic Charr (Salvelinus Alpinus) and Brook Trout (Salvelinus Fontinalis) in Bavarian Hatchery Stocks Inferred from Nuclear and Mitochondrial DNA Markers

Nuclear insulin-like growth factor 2 gene (IGF-2), growth hormone 1 gene (GH-1) and internal transcribed spacer 1 (ITS-1) of the ribosomal DNA as well as the mitochondrial NADH-3 and NADH-4 dehydrogenase genes (ND-3/4) exhibited species-specific restriction fragment patterns and three microsatellite loci (Sfo18, Ssa85 and Ssa197) had non-overlapping allele size ranges in Arctic charr and brook trout and were used as diagnostic markers for testing genetic purity of hatchery stocks and wild populations of Arctic charr and brook trout in Bavaria, Germany. Screening of four wild populations (three in Arctic charr and one in brook trout) revealed only a single hybrid (back-cross to brook trout) individual in L. Starnberg. In contrast, in three (out of five) hatchery stocks of Arctic charr and in both hatchery stocks of brook trout hybrids were detected with the frequency from 3 to 100%. Three hatchery stocks (SS2, SA and BS1) represent a hybrid swarm because they contained a very high proportion of hybrids (from 83 to 100%) and most or all hybrid individuals had alien alleles at only one or a few of six unlinked diagnostic loci, indicating that post-F₁ hybrids represent the majority of individuals in these stocks and introgression has taken place. Release or escape of introgressed individuals from hatcheries into natural water bodies should be avoided in order to protect the biological diversity and genetic integrity of native fish populations.     

Changes in the pericellular connective tissue and breaking strength of the three types of muscles of the cultured carp Cyprinus carpio during storage in ice

To elucidate the structural changes in pink (P), white (W), and red (R) muscles during storage in ice, we measured the breaking strength and changes in pericellular connective tissues of cultured carp. The breaking strength just after killing was highest in R muscle (1.00 ± 0.20 N), lowest in W muscle (0.37 ± 0.07 N), and intermediate (0.84 ± 0.12 N) in P muscle. During the storage period, the breaking strength decreased first in R muscle, then in P muscle, followed by W muscle. The diameter of muscle fibers was greater in W muscle (113 ± 15 μm) than in P muscle (72 ± 3 μm) and R muscle (48 ± 2 μm). Destruction of the honeycomb structure of the pericellular connective tissue occurred most rapidly in W muscle and most slowly in R muscle. These results suggest that the interposing of P muscle fibers in the dorsal ordinary muscle contributes to the acceleration of post-mortem tenderization in fish.     

Spatiotemporal distribution of Flavobacterium psychrophilum and ayu Plecoglossus altivelis in rivers revealed by environmental DNA analysis

Outbreaks of bacterial cold-water disease (BCWD), caused by Flavobacterium psychrophilum, are widespread in Japan, especially among ayu Plecoglossus altivelis. There are few investigations of F. psychrophilum in river water, and its seasonal distribution has not been clarified. We aimed to identify the spatiotemporal dynamics of F. psychrophilum and ayu to provide information that is useful for establishing a countermeasure for BCWD. Quantitative analysis of environmental DNA (eDNA) was used to clarify the year-round dynamics of ayu and F. psychrophilum. We sampled river water from the Nagara and Ibi rivers in Japan, and conducted monthly water sampling and eDNA quantification. Changes in the eDNA concentration of ayu were consistent with the known life histories of the fish. There was a strong negative correlation between the eDNA concentration of F. psychrophilum and water temperature, suggesting a strong dependence of F. psychrophilum dynamics in the river on water temperature. Furthermore, relatively high eDNA concentrations were recorded for both organisms in early summer and fall, suggesting that ayu is infected with F. psychrophilum during these seasons when experiencing up- and downmigration, respectively.

     

Growth Rate, Maturation Level and Flesh Quality of Three Strains of Large Rainbow Trout (Oncorhynchus Mykiss) Reared in Estonia

Three all-female strains of large rainbow trout imported to Estonia (Danish, Finnish and Donaldson strain) were evaluated in a communal rearing experiment in tanks. The fish had been reared on different fish farms before the experiment started and they had an initial weight of 400–500 g. There were no significant genetic differences between the investigated strains on the basis of three allozyme loci. The total weight gain of the strains during the trial was similar but differences were revealed between seasons. The growth rate of the Finnish trout was high during the first autumn but decreased during the second summer of the trial, probably due to infestation by eye fluke Diplostomum sp. The strains differed in the percentage of immature fish and in the level of gonad development at the age of 2.5–3 years. The Donaldson strain had the highest number of immature fish of the studied strains. Mature females of the Finnish strain had the smallest gonads but also the lowest percentage of immature fish. The level of maturation was the main factor determining the quality – slaughter yield and flesh color.     

Incorporation of winter grasses suppresses summer weed germination and affects inorganic nitrogen in flooded paddy soil

ABSTRACT

Laboratory experiments were conducted to investigate how incorporation of the winter grasses foxtail (Alopecurus aequalis) and milk vetch (Astragalus sinicus) into flooded paddy soil affects the germination of summer weeds and soil inorganic nitrogen (N) levels. Polyethylene terephthalate (PET) vessels (10.0 × 8.0 × 15.0 cm) were filled with 0.7 kg air-dried soil that had been collected from organic paddy fields located in Tochigi, Japan. Foxtail and milk vetch were then independently incorporated into the soil at rates of 0% (untreated control), 0.25%, 0.5%, 1%, and 1.5%, with four repetitions per treatment. The PET vessels were placed in a growth chamber (Biotron) set to day/night temperatures and humidities of 27/18°C and 70%/65%, respectively, and with an illuminance of 40,000 lx (12 h) for 28 days, and the total number of weeds and changes in the electrical conductivity (EC) of the surface water were investigated throughout this period, while changes in soil inorganic N were determined at the ends of the experiments. The incorporation of both foxtail and milk vetch significantly suppressed the germination of summer weeds by 56% and 50%, respectively, at incorporation rates of 1% and by 78% and 77%, respectively, at incorporation rates of 1.5% compared with their respective controls. There was a high and significant positive correlation between both the rate of winter grass incorporation and the degree of growth suppression and EC. Incorporation of foxtail at rates of 0.25% and 0.5% significantly increased the inorganic N contents of the sub-layer by an average of 68% compared with the control but had no significant effect in the top layer, whereas the incorporation of milk vetch at a rate of 1% significantly increased the inorganic N contents of both the top layer and sub-layer by 200% and 316%, respectively, compared with the control. Furthermore, the effect of milk vetch on inorganic N significantly increased with an increase in the rate of incorporation. These findings demonstrate that incorporation of the winter grasses foxtail and milk vetch into flooded organic paddy soil can suppress weed growth and act as a source of inorganic N.     

A convenient and nondestructive method using bio-impedance analysis to determine fish freshness during ice storage

Fish freshness can be assessed through K values, but this method has a number of limitations, including a complex procedure and destructive sampling. With the aim to develop a convenient method of assessing fish freshness, we measured the changes in K values (up to 40%) and bio-impedance (Z value; frequency 2, 5, 20, 50, 100 kHz) of ordinary muscle in fish of eight marine species, all caught in the East China Sea, during ice storage and examined their relationships. The results indicated that the K value in all fishes increased linearly with storage time, while their Z value decreased only after 24 h of storage. Moreover, after 24 h of storage and at K values of < 40%, impedance ratios at 2–100 kHz (C value, C?=?Z_{2 kHz}/Z_{100 kHz}) were significantly correlated (p ?<?0.05) with both storage time and K values in all fishes. These findings suggest that the bio-impedance ratio effectively reflects the change in ATP-related compounds of fish and that a convenient, nondestructive method using the C value can be used instead of the complicated K value measurement to assess the freshness of marine fishes after 24 h of ice storage.