Loop-mediated isothermal amplification for the rapid, sensitive, and specific detection of the O9 group of Salmonella in chickens

In the present study, the loop-mediated isothermal amplification (LAMP) assay was developed to amplify the fragments of the O9 Salmonella-specific insertion element and evaluated in the laboratory for its potential use in a field situation, such as poultry farms. Among the bacteria tested, a positive reaction was observed only for 128 strains of 6 serovars of the O9 group Salmonella, such as Enteritidis (SE) and Pullorum. The detection limit of the LAMP assay was 10(3)CFU/ml, which was more sensitive than that of the polymerase chain reaction (PCR) assay with the same target gene (10(6)CFU/ml). The final results were obtained within 30 min for the LAMP assay, while the PCR assay needed a total of 120 min. When the LAMP assay was applied to the enrichment broth mixed with cecal dropping samples either spiked with SE in vitro or excreted by SE-inoculated hens, the results were comparable to those of the conventional plating method including 2 separate enrichments. In conclusion, the LAMP assay developed in the present study is an effective method for the specific detection of the O9 group Salmonella serovars, including SE.     

Isolation and culture of rabbit primordial germ cells

Primordial germ cells (PGCs) are embryonic precursors of the gametes of adult animals and are considered stem cells of the germline. Since their proliferation in vitro correlates well with the schedule of developmental changes in vivo, they might be interesting research tools for genomic imprinting, germ-cell tumors and fertility. Furthermore, once primordial germ cells are separated and placed on a feeder layer with cytokines, they become cultured pluripotent cell lines called embryonic germ (EG) cells. EG cells share several important characteristics with embryonic stem (ES) cells as they can also contribute to the germ line of chimeras. To investigate the characteristics of PGCs and establish rabbit EG (rEG) cells, we cultured rabbit PGCs (rPGCs) in vitro with various combinations of leukemia inhibitory factor (LIF), basic fibroblast growth factor (bFGF) and forskolin on inactivated mouse embryonic fibroblast (MEF) feeder layers. The present study found PGC proliferation in early cultures and induction of rEG-like colonies. These cells expressed pluripotent markers, such as alkaline phosphatase activity, OCT-4, Sox-2 and SSEA-1, in the undifferentiated state; however, the cells did not develop into a teratoma when injected into the kidney capsules of SCID mice, although the restricted differentiation potentials to neural cells were determined via embryoid body formation. From these characteristics and further characterization of the germ stem cell markers Vasa, SCP-1 and SCP-3, we suggested that these were hybrid cells with characteristics somewhere between PGC and EG cells.     

Construction of an expression vector for improved secretion of canine IL-18

To construct a vector for caspase-1 independent expression of canine IL-18, the signal sequence of canine IL-12p40 was fused to the sequence of mature IL-18 on the NdeI restriction site which is located at the 3' end of the signal sequence. The resulting vector expressed coding protein from transfected mammalian cells. The expressed protein was shown to have IL-18 bioactivity in a INF-gamma-inducing assay. These results suggest that the expression vector is the desired tool for advancement of dendritic cell (DC)-based cancer therapy, provided that the vector can successfully be transfected into dendritic cells. We propose a simple and widely applicable method for providing the signal sequence.     

Effect of niacin supplementation in long‐distance transported steer calves

Long‐distance transportation has negative impacts on production and health in cattle. Feed and water are routinely deprived from cattle during transportation. We investigated whether niacin supplementation could improve niacin nutrition and mitigate the adverse effect of transportation with feed and water deprivation in steer calves. We also studied the adverse effect of feed and water deprivation in nontransported steer calves. Twelve calves were assigned to feed and water deprivation for 2 days, or full access to feed and water in experiment 1. Ten calves were assigned to 2‐day transportation with feed and water deprivation, or the transportation with feed and water deprivation, but with supplementation of rumen‐protected niacin at 100 g/day per head in experiment 2. Bodyweight was measured and blood was collected for 32 days in each experiment. Feed and water deprivation temporarily decreased serum glucose concentrations and bodyweight gain. Transportation with deprivation of feed and water caused a temporal decrease in bodyweight gain and serum albumin concentration, and a continuous decrease in serum glucose and total cholesterol concentrations, which was suppressed by niacin supplementation. Niacin supplementation increased blood niacin concentration. These results suggest that niacin supplementation mitigates adverse effects of transportation with feed and water deprivation in steer calves.     

Identification of a novel filovirus in a common lancehead (Bothrops atrox (Linnaeus, 1758))

I performed metaviromic analysis of publicly available RNA-seq data from reptiles to understand the diversity of filoviruses (family Filoviridae). I identified a coding-complete sequence of a filovirus from the common lancehead (Bothrops atrox (Linnaeus, 1758)), tentatively named Tapajós virus (TAPV). Although the genome organization of TAPV is similar to mammalian filoviruses, our phylogenetic analysis showed that TAPV forms a cluster with a fish filovirus. However, TAPV is still distantly related to all the known filoviruses, suggesting that TAPV can be assigned as a species of a novel genus in Filoviridae. To our knowledge, this is the first report identifying a filovirus in reptiles, and thus contributes to a deeper understanding of the diversity and evolution of filoviruses.     

Quantitative correlation between molecular similarity and receptor-binding activity of neonicotinoid insecticides

Quantitative correlation between molecular similarity and receptor-binding activity of neonicotinoid insecticides such as imidacloprid and acetamiprid was studied by using a method of similarity index and semi-empirical molecular orbital calculations. A series of compounds having an aromatic ring and a cyclic or acyclic amine moiety with an electron-withdrawing group were subjected to the similarity-activity analysis. Energy-minimum structures and electrostatic properties of the molecules were obtained by MNDO-PM3. The electrostatic similarity of each molecule compared with the most active compounds was found to correlate significantly with the binding activity to nicotinic acetylcholine receptor (nAChR) in honey bee when the two molecules were superimposed to maximize the molecular shape similarity by simplex procedure. This indicates that molecular similarity in terms of electrostatic properties is important for activity, as well as superimposability in terms of molecular shape. A schematic model of interaction between neonicotinoids and nAChR is proposed according to the results of similarity-activity analyses. © 1998 SCI.     

Correlation of changes in seasonal distribution and catch of red sea bream Pagrus major with winter temperature in the eastern Seto Inland Sea,Japan (1972–2010)

Red sea bream Pagrus major is a commercially important fish in Japan. In eastern Seto Inland Sea (SIS), the catch has increased from 297 tons in 1972 to 2,039 tons in 2010. We examined the relationship, 1972–2010, between increase in catch and winter temperature, based on the catch in February and March and the lowest water temperature at 10 m depth. In 1972–1986, the lowest water temperatures in the inner SIS areas (Osaka Bay, Harima‐nada, and Bisan‐seto) were <8°C, which is physiologically unfavorable for red sea bream. However, in 1987–2010 temperatures were generally ≥8°C. In the inner areas, the catch during winter had been minimal until the early 1980s, presumably because most red sea breams moved to the Kuroshio‐influenced (warmer) Kii Channel area. However, the winter catch in the inner areas of SIS increased from the late 1980s with warm winters. In addition, the catch between April and June, the spawning season, increased in the inner areas from the 1990s, and the catch rate of the inner areas was more than twice higher in the 2000s than in the 1980s. The results suggest that expansion of the distribution area during winter due to warm winter and increase in egg production in the inner areas greatly contributes to the increasing in catch in the eastern SIS.     

Low-fidelity homing behaviour of Biwa salmon <Emphasis Type="Italic">Oncorhynchus</Emphasis> sp. landlocked in Lake Biwa as inferred from otolith elemental and Sr isotopic compositions

Biwa salmon Oncorhynchus sp. is endemic to Lake Biwa, Japan, where it is an important commercial and recreational fisheries species. However, no information is currently available on its population structure and migration ecology. Therefore, here we evaluated whether otolith Sr/Ca, Ba/Ca and ⁸⁷Sr/⁸⁶Sr ratios can be used as natural signatures in Biwa salmon and then used these to determine the natal origins of lake-migration-phase individuals and spawning adults, and the homing ability of spawning adults in the Lake Biwa water system. Quadratic discriminant function analysis demonstrated that the lake-migration school comprised individuals with multiple origins, including rivers to the east, west and north of Lake Biwa, and that the homing rate of spawning adults was low (18 out of 80 individuals), with ca. 78% of fish straying into non-natal rivers. However, this straying behaviour was not spatially random, with fish tending to migrate upstream in rivers neighbouring their natal rivers. The high rate of straying in spawning adults is considered important for establishing and maintaining this species, which is highly adapted to life in the Lake Biwa water system where environmental disturbances often occur.     

Interspecific differences in the post-settlement survival of <Emphasis Type="Italic">Acropora</Emphasis> corals under a common garden experiment

Extremely low post-settlement survival is one of the largest barriers for artificial rehabilitation of Acropora corals. However, little data have been found for interspecific difference of the post-settlement survival probably because the observation of coral juvenile is difficult in the field. Here, we analyzed the survival of three dominant species of Acropora corals (A. digitifera, A. tenuis, A. yongei), with different colony morphologies and habitat preferences, for 2 years after settlement under the same environmental conditions. The post-settlement survival was significantly higher for A. tenuis than for A. digitifera 3 months after settlement. Two years later, the survival rate of A. tenuis was approximately 15 times higher than A. digitifera. In a separate analysis of three bottle-brush species (A. awi, A. echinata, A. subglabra) and A. tenuis, post-settlement survival was always higher for A. awi than for other two bottle-brush species, suggesting that the initial survival was different among morphologically sister species. Low survival was possibly associated with slow growth rates during the first 7 months. Thus, species selection is important for successful artificial coral rehabilitation, with A. tenuis being the most viable option. Alternatively, new techniques are required to improve post-settlement survival of slow growing coral species.