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Abstract Objective The ayu or sweetfish Plecoglossus altivelis is ray-finned fish that is widely distributed in East Asia. The genome size of ayu was estimated at approximately 420 Mb. Previously, we reported on ayu draft genome assembly by whole-genome shotgun using Illumina short reads and PacBio long reads; however, the assembly was not to chromosome level. Therefore, to improve the draft genome sequence of ayu to chromosome level, we performed in situ Hi-C sequencing as a source of linkage information. Results The ayu genome assembly yielded 28 large scaffolds that corresponded to the karyotype of ayu (n = 28). The resulting ayu genome assembly has a N50 scaffold length of 17.0 Mb, improved from 4.3 Mb. The high-quality reference genome will be helpful for phylogenetic research on bony fishes and for breeding programs in ayu aquaculture.

Objective: The ayu or sweetfish Plecoglossus altivelis is ray-finned fish that is widely distributed in East Asia. The genome size of ayu was estimated at approximately 420 Mb. Previously, we reported on ayu draft genome assembly by whole-genome shotgun using Illumina short reads and PacBio long reads; however, the assembly was not to chromosome level. Therefore, to improve the draft genome sequence of ayu to chromosome level, we performed in situ Hi-C sequencing as a source of linkage information. Results: The ayu genome assembly yielded 28 large scaffolds that corresponded to the karyotype of ayu (n = 28). The resulting ayu genome assembly has a N50 scaffold length of 17.0 Mb, improved from 4.3 Mb. The high-quality reference genome will be helpful for phylogenetic research on bony fishes and for breeding programs in ayu aquaculture. [ABSTRACT FROM AUTHOR]

Abstract Ayu (Plecoglossus altivelis altivelis) is an important freshwater fisheries resource and popular recreational fishing species in Japan that lives for only 1 year, with a single breeding season. To supplement increased recreational fishing demand for this species, more than 13 million wild‐born landlocked juveniles are translocated every year from Lake Biwa into Japanese rivers, and more than 52 million hatchery‐reared juveniles born from captive‐reared parents have been extensively released. The translocation of landlocked juveniles and hatchery release has continued for more than 100 and 30 generations, respectively. Previous studies have reported that landlocked and amphidromous forms of Ayu easily hybridise, leading to concerns that the fitness of hybrid progeny would be reduced in the wild. However, limited information exists regarding the reproductive success of landlocked Ayu in translocated rivers, and to date no studies have evaluated the effects of translocation on population structure. Demonstrating that hybridisation occurs between the two forms is central to future management and conservation of this species. To address this, a literature search to summarise biological knowledge of Ayu and population genetic data screening was undertaken, and published genetic data sets covering a distribution range in Japan were reanalysed. Analyses provide strong evidence for very high gene flow between sampling locations. Genetic diversity is homogeneous in amphidromous samples. Bayesian admixture analysis infers widespread hybridisation in Japanese rivers (24 ± 8%). Maximum likelihood admixture graphs detect two migration events from Lake Biwa to rivers in the northern Sea of Japan and Hokkaido. Analyses consistently indicate that hybridisation between translocated landlocked juveniles and native amphidromous Ayu occurs throughout Japanese rivers. These results are discussed in relation to the management of this species.

For the first time, natural entry of ayu or sweetfish into the river of Peter the Great Bay basin (Japan Sea) and generally into the waters of Russian Federation is registered. This finding extends the list of fish species and families dwelling in the Russian waters.

With the rapid development of intensive farming, the aquaculture industry uses a great many antibiotics for the prevention and treatment of bacterial diseases. Despite their therapeutic functions, the overuse and accumulation of antibiotics also pose a threat to aquaculture organisms. In the present study, ayu (Plecoglossus altivelis) was used as a fish model to study the impacts of ciprofloxacin (CIP) overuse on intestinal homeostasis and immune response during subsequent Pseudomonas plecoglossicida infection. Based on 16S rRNA gene amplification and Illumina sequencing, we found that CIP pre-exposure caused significant variation in intestinal microbiota, including increased species richness, altered microbiota composition and interaction networks, and increased metabolic dysfunction. Furthermore, immunohistochemical analysis indicated that CIP pre-exposure resulted in severe mucosal layer damage, goblet cell reduction, and epithelial cell necrosis of the intestinal barrier in infected ayu. Quantitative real-time polymerase chain reaction (qRT-PCR) showed that disruption of intestinal homeostasis impaired systemic antiinfection immune responses in the intestine, gill, spleen, and head kidney, while inhibiting IL-1β, TNFα, and IL-10 expression and promoting TGF-β expression. Our findings indicated that CIP administration can directly affect intestinal microbiota composition and intestinal integrity in ayu fish. This perturbation of intestinal homeostasis is likely responsible for the lower survival rate of hosts following subsequent infection as the capacity to mount an effective immune response is compromised. This study also provides preliminary clues for understanding the effects of antibiotic overuse on higher vertebrates through trophic transfer. [ABSTRACT FROM AUTHOR]

The complete genome sequences of seven Edwardsiella ictaluri strains, isolated from the kidneys of dead ayu ( Plecoglossus altivelis altivelis ) in Kochi's Kagami and Shimanto Rivers, Japan, were determined. Multilocus sequencing typing revealed that their genotypes were sequence-type ST26., Competing Interests: The authors declare no conflict of interest.

The Ayu (Plecoglossus altivelis altivelis) population on Yaku-shima Island, Kagoshima Prefecture, Japan represents the southernmost population of the subspecies and is considered to be facing extinction. We investigated the genetic characteristics of Ayu in the Miyanoura River (MYU) in the northeast of the island and the Kurio River (KRO) in the southwest of the island, using partial sequences of mitochondrial (mt) control region and polymorphic microsatellite (simple sequence repeat: SSR) loci. The mtDNA analysis revealed that the Yaku-shima Island population was significantly differentiated from other Ayu populations; additionally, the MYU and KRO populations were significantly different from each other in terms of mtDNA (pairwise ΦST = 0.5826, P < 0.01) and SSR (pairwise ΦST = 0.1598, P < 0.01) analyses, indicating very little or no gene flow between them. The mtDNA haplotype diversity values were minimal for KRO and somewhat lower for MYU (h = 0.8176) than for each population from the mainland of Japan (Honshu–Kyushu) and the Korean Peninsula (h = 0.9905–1.0000). The mean values of expected heterozygosity (He) of SSRs were also lower in KRO (mean He = 0.555) than in MYU (mean He = 0.649). A considerably small effective population size (Ne = 100.1 for MYU, 151.2 for KRO) and a bottleneck effect for Yaku-shima Island population were suggested by SSR analysis. These findings underscore the importance of monitoring the genetic diversity of Ayu on Yaku-shima Island and the necessity of designing conservation policies for each river's population. [ABSTRACT FROM AUTHOR]

The Ryukyu ayu Plecoglossus altivelis ryukyuensis is an endangered amphidromous fish that inhabits rivers in the Ryukyu Archipelago (Japan). Populations of the species have declined dramatically. Consequently, the Ryukyu ayu has been registered as a natural monument in Japan and monitoring surveys with direct catching are restricted legally. This restriction, unfortunately, makes monitoring of population abundances difficult and creates a barrier to both advancing understanding of the species' status and the development of appropriate conservation plans. We developed a non-invasive monitoring methodology using eDNA analyses. We designed a specific quantitative PCR assay for the Ryukyu ayu using the mitochondrial ND4 region. Using this primer/probe set, we conducted eDNA analyses in three rivers on Amami-Ohshima Island. The DNA fragments were amplified from the eDNA extracted from natural water in each river. The numbers of DNA fragments detected were positively correlated with individual counts of fish obtained by visual snorkelling surveys. Our method does not contravene restrictions and facilitates abundance monitoring of this endangered fish species. [ABSTRACT FROM AUTHOR]

Classical Fc receptors (FcRs) mediate the binding to and recognition of the Fc portion of antibodies and play an important role during immune responses in mammals. Although proteins similar to soluble FcRs have been identified in fish, little is known about the role of such proteins in fish immunity. Here, we cloned a cDNA sequence encoding a soluble Fc receptor for an immunoglobulin G (FcγR) homolog from ayu (Plecoglossus altivelis) (PaFcγRl). The predicted protein was composed of two immunoglobulin C2-like domains but lacked a transmembrane segment and a cytoplasmic tail. The PaFcγRl transcripts were distributed at low levels in all tested tissues, but significantly increased after Vibrio anguillarum infection. The PaFcγRl protein was expressed in the head kidney, trunk kidney, and neutrophils. Recombinant PaFcγRl (rPaFcγRl) was secreted when transfected into mammalian cells and the native protein was also detected in serum upon infection. rPaFcγRl was also demonstrated to bind to ayu IgM, as assessed by cell transfection. Suppressive activity of the recombinant mature protein of PaFcγRl (rPaFcγRlm) on in vitro antisheep red blood cell (SRBC) responses was detected by a modified hemolytic plaque forming cell assay. In conclusion, our study revealed that PaFcγRl is closely involved in the negative regulation of IgM production in the ayu spleen. [ABSTRACT FROM AUTHOR]

A new bio-economic model for managing population of non-renewable inland fishery resource in uncertain environment is presented. Population dynamics of the resource is described with stochastic differential equations (SDEs) having ambiguous growth and mortality rates. The performance index to be maximized by the manager of the resource while minimized by nature is presented in the context of differential game theory. The dynamic programming principle leads to a Hamilton-Jacobi-Bellman-Isaacs (HJBI) equation that governs the optimal resource management strategy under the ambiguity. The main contribution of this paper is a series of theoretical analysis on the reduced HJBI equation for non-renewable fishery resources in a broad sense, indicating that the ambiguity critically affects the resulting optimal controls. Practical implications of the theoretical analysis results are also presented focusing on artificially hatched Plecoglossus altivelis (Ayu), an important inland fishery resource in Japan. [ABSTRACT FROM AUTHOR]

Tissue factor (TF) plays an important role in the host's immune system as the principal initiator of coagulation. However, the precise function of TF in teleosts remains unclear. We determined the cDNA sequence of TF from ayu Plecoglossus altivelis (PaTF). The PaTF transcript was expressed in all tested tissues, and changes in expression were observed in tissues and monocytes/macrophages (MO/MФ) upon infection with Vibrio anguillarum . PaTF was prokaryotically expressed and purified to prepare anti-PaTF antibodies. Western blot analysis revealed that native PaTF was glycosylated in thrombocytes, but not in ayu MO/MФ. Microparticles could transfer PaTF to thrombocytes. PaTF neutralization or knockdown led to anti-inflammatory status in ayu MO/MФ upon V. anguillarum infection. PaTF neutralization reduced the apoptosis of ayu MO/MФ and improve survival rate in V. anguillarum -infected ayu. Our results indicate that PaTF plays a role in ayu immune response against bacterial infection as a pro-inflammatory mediator. [ABSTRACT FROM AUTHOR]

The word Ayurveda comes from the Sanskrit root Ayu which means span of life. The purpose of examination of Ayu is to obtain knowledge regarding longevity, residual span of life in diseased person and rate of the mortality. The purpose of Dashavidha Pariksha is to obtain knowledge regarding Dosha Bala (Vyadhi Bala), Bala (Rogibala) and Ayu of the patient as described in Charak Samhita. Commonly Ayu and Vaya are regarded as synonyms but they are not similar terms. Hence, description of Vaya and Ayu is separately mentioned in Charaka as well as Sushruta Samhita. Description exclusively on Ayu Pareeksha described in Charaka Viman Sthana, Charaka Chikitsasthana, Charaka Indriya Sthana and Charaka Shareer Sthana. In Indriya Sthana, Ayu Pariksha indicates the residual life-span of the diseased person and where as in Jaatisutriya Adhyaya in Shareer Sthana mentioned the characteristic properties of the child who is going to survive for longer period of time. In Sushruta Samhita, Ayu is classified into three broad headings namely Dhirghayu, Madhyamayu and Alpayu and explained the characteristic properties of all the three.

Excessive predation pressure from the waterfowl Phalacrocorax carbo (Great Cormorant) on Plecoglossus altivelis (Ayu) has recently been a severe problem of river environment in Japan. Local fishery cooperatives are currently suffering from economic difficulties due to decrease of the fish catch of P. altivelis. Local fishery cooperatives and municipalities have been enthusiastically trying to develop countermeasures that can effectively reduce the predation pressure; however, their effectiveness and efficiency have not been systematically quantified well. This aim can be achieved with the help of an appropriate mathematical model. In this paper, based on a pure death process, a practical stochastic control model for population dynamics of released P. altivelis in river environment under predation pressure from P. carbo, harvesting by human, and environmental fluctuations is proposed. Finding an optimal management strategy ultimately reduces to solving a 2D Hamilton-Jacobi-Bellman equation, which is performed with a finite element scheme. Its application to a Japanese river environment successfully computes the optimal management strategy that is consistent with the reality. Numerical sensitivity analysis of the presented mathematical model is also performed for comprehension of dependence of the optimal strategy on the model parameters. [ABSTRACT FROM AUTHOR]

Liver X receptors (LXR) are modulators of metabolic processes and inflammation in mammals as nuclear receptors. However, the precise function of LXR in teleosts remains unclear. Here, we characterized a LXR gene (PaLXR) from ayu, Plecoglossus altivelis . The PaLXR transcript was expressed widely in all tissues studied, and changes in expression were observed in tissues and monocytes/macrophages (MO/MΦ) upon infection with the bacterium Edwardsiella ictaluri . PaLXR activation decreased the mRNA expression of interleukin-1β (IL-1β), tumor necrosis factor-α (TNF-α), and IL-10 upon E. ictaluri infection, while their expression was increased following the knockdown of PaLXR by siRNA. Moreover, E. ictaluri infection induced the apoptosis of ayu neutrophils and PaLXR activation enhanced the internalization of E. ictaluri -infected apoptotic neutrophils by MO/MΦ (efferocytosis), while PaLXR knockdown led to decreased efferocytosis. Furthermore, PaLXR activation inhibited intracellular bacterial survival during efferocytosis, while PaLXR knockdown enhanced survival. In conclusion, our results indicate that PaLXR plays a role in the modulation of innate immune responses in ayu MO/MФ. [ABSTRACT FROM AUTHOR]

Prostaglandin E2 (PGE2) plays an important role in a broad spectrum of physiological and pathological processes by interacting with E-type prostanoid receptors (EPs). EP4 is one of four EP subtypes known to mediate the immune response in mammalian monocytes/macrophages. However, the precise function and characteristics of EP4 in fish remain unclear. In this study, we characterized a novel EP4-like (PaEP4L) gene from ayu, Plecoglossus altivelis. The cDNA sequence of PaEP4L is 2781 nucleotides (nts) in length, encoding a polypeptide of 459 amino acid residues with a calculated molecular weight of 51.17 kDa. Sequence comparison and phylogenetic tree analysis showed that PaEP4L shared 76% amino acid identity with that of the Atlantic salmon (Salmo salar). PaEP4L mRNA was detected by real-time quantitative PCR (QPCR) in all tested tissues and head kidney-derived monocytes/macrophages (MO/MФ). It varied greatly in liver, spleen and MO/MФ upon Vibrio anguillarum infection. Western blot analysis revealed a significant increase of PaEP4L in cell homogenates from ayu MO/MФ upon V. anguillarum infection. Moreover, anti-PaEP4L IgG reversed the down-regulation of interleukin 1β (IL-1β) and tumor necrosis factor α (TNF-α) mRNA expression as well as phagocytosis in ayu MO/MФ caused by PGE2. There were no significant differences in the respiratory burst response between PGE2 treated and untreated cells. We further found that cAMP mediated PGE2/PaEP4L signal in ayu MO/MФ. In conclusion, our results indicate that PaEP4L mediates PGE2 effects on ayu MO/MФ function, revealing that EP4 also plays a role in the modulation of cells of the fish’s innate immune system. [ABSTRACT FROM AUTHOR]

Water sounds are believed to be important in triggering the upriver ascent behavior of migratory fish species. Modeling the effect of underwater sound on fish preference represents a promising technique to determine migratory paths for the comprehensive analysis of river habitat suitability. In this study, we aimed to determine the preference and weight of underwater sounds by laboratory experiments using adult and juvenile ayu (Plecoglossus altivelis). A watercourse with an underwater speaker installed at one end was used for the experiments. Sound was emitted from the speaker at different sound pressure levels. The sound sources included pure tones (100 Hz, 200 Hz, 400 Hz, and 800 Hz), white noise, recorded sound from a weir on the Fushino River, and recorded sound at a fish ladder on the Misumi River (Japan). The results showed that juvenile and adult ayu avoided the pure sound of 100 Hz and recorded sound of the Fushino River weir. In contrast, both adults and juveniles preferred the pure sound of 200 Hz and recorded sound of the fish ladder. Moreover, by comparing the obtained quantitative preference parameters for sounds, we found that adult ayu expressed higher preference for sound than juveniles. We designed a procedure to calculate sound preference and built it into our fish behavior simulation model on geographic information system (GIS) software. We believe that the model could successfully reproduce observed fish migration behavior in rivers. This technique could be used for designing an underwater soundscape to attract fish to the entrances of fish ladders or to keep fish away from hydropower plants. [ABSTRACT FROM AUTHOR]

Ayu Plecoglossus altivelis , a herbivorous fish, is an important fishery resource and key component of the foodweb in many Japanese streams. Radionuclide contamination of this species is likely transferred to higher trophic levels, include humans, in the food chain. After the Fukushima accident in March 2011, ayu were exposed to highly contaminated silt while feeding on algae attached to the riverbed stones. To understand the route by which herbivorous fish are exposed to radionuclides, the activity concentrations of sum of 134 Cs and 137 Cs (radiocesium) were analyzed in riverbed samples (algae and silt) and in the internal organs and the muscle of ayu in five river systems in the Fukushima Prefecture between summer 2011 and autumn 2013. Although there was a positive correlation between the radiocesium activity concentrations in the muscle and the internal organs of ayu, the median activity concentration in the muscle was much lower than those in the internal organs. The activity concentrations of radiocesium in the riverbed samples and the internal organs and the muscle of ayu were correlated with contamination levels in soil samples taken from the watershed upstream of the sample sites. The results of the generalized linear mixed models suggest that the activity concentrations in both the internal organs and the muscle of ayu declined over time. Additionally, the activity concentrations in the internal organs were correlated with those in the riverbed samples that were collected around the same time as the ayu. The activity concentrations in the muscle were correlated with ayu body size. Our results suggest that ayu ingest 134 Cs and 137 Cs while grazing silt and algae from the riverbed, and a part of the 134 Cs and 137 Cs is assimilated into the muscle of the fish. [ABSTRACT FROM AUTHOR]

Pseudomonas plecoglossicida is a lethal pathogen of ayu (Plecoglossus altivelis) in Japan and is responsible for substantial economic costs to ayu culture. Previously, we demonstrated the efficacy of phage therapy against P. plecoglossicida infection using two lytic phages (PPpW-3 and PPpW-4) (S. C. Park, I. Shimamura, M. Fukunaga, K. Mori, and T. Nakai, Appl Environ Microbiol 66:1416 -1422, 2000, http://dx.doi.org/10.1128/AEM.66.4.1416-1422.2000; S. C. Park and T. Nakai, Dis Aquat Org 53:33-39, 2003, http://dx.doi.org/10.3354/dao053033). In the present study, the complete genome sequences of these therapeutic P. plecoglossicida phages were determined and analyzed for deleterious factors as therapeutic agents. The genome of PPpW-3 (myovirus) consisted of 43,564 bp with a GC content of 61.1% and 66 predicted open reading frames (ORFs). Approximately half of the genes were similar to the genes of the Escherichia coli phage vB-EcoM-ECO1230-10 (myovirus). The genome of PPpW-4 (podovirus) consisted of 41,386 bp with a GC content of 56.8% and 50 predicted ORFs. More than 70% of the genes were similar to the genes of Pseudomonas fluorescens phage ϕIBB-PF7A and Pseudomonas putida phage ϕ15 (podoviruses). The whole-genome analysis revealed that no known virulence genes were present in PPpW-3 and PPpW-4. An integrase gene was found in PPpW-3, but other factors used for lysogeny were not confirmed. The PCR detection of phage genes in phage-resistant variants provided no evidence of lysogenic activity in PPpW-3 and PPpW-4. We conclude that these two lytic phages qualify as therapeutic agents. [ABSTRACT FROM AUTHOR]

Competing Interests: The author declares that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.

CXCL8, a CXC-type chemokine, plays a crucial role in acute inflammation by recruiting and mediating neutrophils and other cells. In this study, the cDNA and genomic DNA sequence of a CXCL8-like protein (PaCXCL8l) from ayu (Plecoglossus altivelis) was determined. Sequence analysis showed that PaCXCL8l represented the typical structure of animal CXCL8s. Phylogenetic tree analysis indicated that PaCXCL8l was closest to CXCL8 of Atlantic cod (Gadus morhua). Constitutive expression of PaCXCL8l was detected in all tested tissues and monocytes/macrophages, and its expression dramatically increased upon Listonella anguillarum infection. In vitro, recombinant PaCXCL8l exhibited a significant chemotactic effect on neutrophils at 0.1 μg/ml and on monocytes/macrophages at 1.0 μg/ml. In vivo, the numbers of peritoneal neutrophils and monocytes/macrophages were both up-regulated following intraperitoneal administration of recombinant PaCXCL8l. These results suggest that PaCXCL8l is crucially involved in the immune response of ayu by mediating chemotaxis of neutrophils and monocytes/macrophages. [ABSTRACT FROM AUTHOR]

We evaluated the effects of turbidity on school formation in ayu (Plecoglossus altivelis) [24.5 ± 2.2 mm standard length (Ls)], Japanese anchovy (Engraulis japonicus) (29.1 ± 3.1 mm Ls) larvae, which often live in turbid coastal waters, and yellowtail (Seriola quinqueradiata) juveniles (37.1 ± 2.5 mm Ls), which live in clear offshore waters. Fish were introduced into experimental tanks at one of five turbidity levels obtained by dissolving 0, 5, 20, 50, or 300 mg l−1 of kaolin in seawater. Their behaviour was video recorded, and the nearest neighbour distance (DNN) and separation angle (AS) were compared among turbidity levels. Mean DNN of ayu was significantly smaller at 20 and 50 mg l−1 than any other level of turbidity, as was AS at 20 mg l−1 compared with 0 mg l−1. Mean AS of anchovy was smaller at 50 mg l−1 of turbidity than any others. In contrast, mean DNN of yellowtail was larger at 300 mg l−1 than any others. These results suggest that moderate turbidities enhance schooling behaviour in ayu and Japanese anchovy larvae, whereas turbidity has an inhibitive effect on schooling of yellowtail juveniles, corresponding well to the habitat characteristics of each species. [ABSTRACT FROM PUBLISHER]

It is widely believed that attached diatoms are the best food source for freshwater ayu Plecoglossus altivelis (Temminck et Schlegel) in Japan. However, sensory testing by a panel consisting of 40 untrained tasters and analyses of gut content showed that there was no significant correlation between preference scores for fish taste and the dominance of diatoms in the algal assemblages fed upon by ayu in the Yahagi River, Aichi Prefecture, Central Japan. Ayu that fed on the filamentous blue-green algae Homeothrix varians was evaluated higher for aroma and fat content than those that fed on diatoms. Ayu feeding on diatoms were collected in the lower reaches of the Yahagi River which receive domestic sewage. In the absence of more detailed analyses, exposure to sewage is considered to confer a characteristic aroma on Ayu. [ABSTRACT FROM AUTHOR]

P2X purinoceptor 7 (P2X7R), an ATP-gated ion channel, plays an important role during the innate immune response in mammals. However, relatively little is known about the role of P2X7R in the fish immune system. Here, we cloned a cDNA sequence encoding ayu (Plecoglossus altivelis) P2X7R (aP2X7R). The predicted protein was composed of 574 amino acid residues with a P2X family signature, two transmembrane domains, and a long C-terminal. aP2X7R transcripts were mainly distributed in ayu immune tissues and significantly increased in all tested tissues and in macrophages after Listonella anguillarum infection. The aP2X7R protein was upregulated significantly in macrophages upon bacterial challenge. An antibody against the ectodomain of aP2X7R (aEPAb) and an antagonist (oATP) were used to block aP2X7R. aP2X7R siRNA was also used to knockdown the receptor expression in ayu macrophages. Cell death induced by ATP was significantly inhibited in ayu macrophages after aEPAb, oATP, or siRNA treatment. Moreover, aP2X7R ablation also resulted in suppression of phagocytic activity and ATP-induced bacterial killing in ayu macrophages. Our results indicated that aP2X7R was upregulated after infection and mediated cell death, phagocytosis, and bacterial killing of ayu macrophages. [ABSTRACT FROM AUTHOR]

P2X purinoceptor 7 (P2X7R), an ATP-gated ion channel, plays an important role during the innate immune response in mammals. However, relatively little is known about the role of P2X7R in the fish immune system. Here, we cloned a cDNA sequence encoding ayu (Plecoglossus altivelis) P2X7R (aP2X7R). The predicted protein was composed of 574 amino acid residues with a P2X family signature, two transmembrane domains, and a long C-terminal. aP2X7R transcripts were mainly distributed in ayu immune tissues and significantly increased in all tested tissues and in macrophages after Listonella anguillarum infection. The aP2X7R protein was upregulated significantly in macrophages upon bacterial challenge. An antibody against the ectodomain of aP2X7R (aEPAb) and an antagonist (oATP) were used to block aP2X7R. aP2X7R siRNA was also used to knockdown the receptor expression in ayu macrophages. Cell death induced by ATP was significantly inhibited in ayu macrophages after aEPAb, oATP, or siRNA treatment. Moreover, aP2X7R ablation also resulted in suppression of phagocytic activity and ATP-induced bacterial killing in ayu macrophages. Our results indicated that aP2X7R was upregulated after infection and mediated cell death, phagocytosis, and bacterial killing of ayu macrophages. [ABSTRACT FROM AUTHOR]

With the rapid development of intensive farming, the aquaculture industry uses a great many antibiotics for the prevention and treatment of bacterial diseases. Despite their therapeutic functions, the overuse and accumulation of antibiotics also pose a threat to aquaculture organisms. In the present study, ayu ( Plecoglossus altivelis ) was used as a fish model to study the impacts of ciprofloxacin (CIP) overuse on intestinal homeostasis and immune response during subsequent Pseudomonas plecoglossicida infection. Based on 16S rRNA gene amplification and Illumina sequencing, we found that CIP pre-exposure caused significant variation in intestinal microbiota, including increased species richness, altered microbiota composition and interaction networks, and increased metabolic dysfunction. Furthermore, immunohistochemical analysis indicated that CIP pre-exposure resulted in severe mucosal layer damage, goblet cell reduction, and epithelial cell necrosis of the intestinal barrier in infected ayu. Quantitative real-time polymerase chain reaction (qRT-PCR) showed that disruption of intestinal homeostasis impaired systemic anti-infection immune responses in the intestine, gill, spleen, and head kidney, while inhibiting IL-1β , TNF-α , and IL-10 expression and promoting TGF-β expression. Our findings indicated that CIP administration can directly affect intestinal microbiota composition and intestinal integrity in ayu fish. This perturbation of intestinal homeostasis is likely responsible for the lower survival rate of hosts following subsequent infection as the capacity to mount an effective immune response is compromised. This study also provides preliminary clues for understanding the effects of antibiotic overuse on higher vertebrates through trophic transfer.

Abstract: Error-prone polymerase chain reactions (epPCRs) are often used to introduce mutations in random mutagenesis, which has been used as a tool in protein engineering. Here, we developed a new method of epPCR using heavy water as a solvent instead of normal water (H2O). Rhodopsin cDNA of the Ayu fish (Plecoglossus altivelis) was used as a template and was amplified using five different conditions: (A) 100% H2O with no Mn2+, (B) 100% H2O/0.6mM Mn2+, (C) 99% D2O with no Mn2+, (D) 99% D2O/0.6mM Mn2+ and (E) 99% H2 18O with no Mn2+. The 13,960 (for each of the conditions A to D) and 33,504 (for condition E) base pairs were sequenced. A maximum error rate of 1.8×10−3 errors/bp was detected in condition D, without any particular hot-spot mutations. A high preference for AT→GC transitions was observed in condition D, whereas a high preference for transitions over transversions was observed in condition C. All of the mutations observed in condition E were transversions. When conditions A and C were applied to another template, the honeybee actin gene, the results were comparable to those for Ayu rhodopsin. Based on these results, the use of heavy water, instead of H2O, as a solvent for epPCR can introduce random mutations without positional bias, template dependency or decreased yield. Our new epPCR method, and possibly combining the use of D2O and H2 18O, may be a powerful random mutagenesis technique. [Copyright &y& Elsevier]

The present study was performed to determine the current infecion status of Metagonimus yokogawai metacercariae in sweetfish, P/ecog/ossus attire/is, collected from several streams in eastern and southern coastal areas of Korea. The sweetfish collected were artificially digested with pepsin-HCI solution and examined under a stereomicroscope in August and September, 2007. Out of 145 sweetfish collected from 10 streams in eastern coasts (Gangwon-do and Gyeongsangbuk-do), 88 (60.7%) were infected with Metagonimus metacercariae. The average metacemarial density was 61 per infected fish. Among 141 sweetfish collected from 10 streams in southern coasts (Gyeongsangnam-do, Ulsan Metropolitan city, and Jecllanam-do), 140 (99.3%) were infected with Metagonimus metacercariae, and their average density was 949 per infected fish. The present study confirmed that M. yokogawai metacercariae are still prevalent in the sweetfish from several eastern and southern coastal localities. The prevalence and metacemarial density were much higher in the sweetfish from streams of southern coasts than in those of eastern coasts. Therefore, attention should be paid to this small fluke infection, and consumption of raw sweetfish naturally produced in these areas should be prohibited. [ABSTRACT FROM AUTHOR]

We examined the influence of several hydrological and meteorological parameters on the migratory movements of ayu Plecoglossus altivelis altivelis in central Japan. When comprehensively evaluating rivers and ayu behaviour on a catchment scale, the subjects of analysis typically include human activities and hydrological and meteorological phenomena. However, limiting analyses to such factors may be too restrictive when human activities are being conducted. Accordingly, we incorporated a biological viewpoint into the evaluation method, analysing hydrological data (river discharge, river water temperature, sea water temperature) to determine watershed characteristics and examining the relationship between these characteristics and the habitat conditions of ayu. Then we constructed a numerical model for ayu migratory runs that incorporated ayu ecology and watershed characteristics. Analyses of ayu movements from a lower estuarine dam demonstrated that downstream displacements were associated with high water flows of more than 200m3 s−1 at the beginning of summer. We conclude that it is important to consider the effects of environmental parameters on the movements of different fish species to understand the causes of spatial variation in fish distribution in lowland rivers. [Copyright &y& Elsevier]

The article presents a study which examined the effect of oral administration of humus extract on the protection against Flavobacterium psychrophilum infection in ayu fish (Plecoglossus altivelis) in Japan. It notes that farmed Ayu fish and other freshwater fishes are affected by the cold water disease caused by gram-negative bacterium, Flavobacterium psychrophilum. Results of the study revealed that the administration of humus extract orally induces effective protection against cold water disease caused by F. psychrophilum infections. It is noted that fish mortality and development of skin lesions, such as hemorrhages on the skin, gill cover on mouth, were significantly inhibited in fish treated with different levels of humus extract adsorbed on dry pellets.

Abstract: Hydraulic structures such as a dam, weir and water gate affect on the flow depth, velocity, bed topology, water quality and so on, so that the hydraulic structures may affect on the water environment for plants and aquatic lives in and around rivers. Therefore, before construction of such hydraulic structures, it is necessary to conduct the environmental assessment. However, the accuracy of the environmental assessment is not so high at present, because the preference curves of the flow depth, velocity and so on are not clear. Recently, Nagaya et al. [Nagaya, T., Onitsuka, K., Higashino, M., Takami, T., Otsuka, N., Akiyama, J., Matsumoto, K., 2004. Evaluation of weight of parameter for spawning of ayu, Plecoglossus altivelis, and prediction of spawn density per unit area. In: Lee, J.H.W., Lam, K.M. (Eds.), Environmental Hydraulics and Sustainable Water Management, vol. 2. AA. Balkema Publishers, UK, pp. 947–954] pointed out that the velocity, flow depth and substrate affect on the suitability of spawning for ayu, Plecoglosses altivelis altivelis on the basis of statistical investigation. Preference curves of velocity and flow depth were made clear by Onitsuka et al. [Onitsuka, K., Nagaya, T., Higashino, M., Takami, T., Otsuka, N., Akiyama, J., Ozeki, H., Matsumoto, K., Shiraishi, Y., 2005. Suitable flow depth and velocity for spawning of ayu, Plecoglossus altivelis. In: Proceedings of the 31th Congress of IAHR, Korea, pp. 1850–1858]. On the other hand, the preference curve of substrate has not been investigated. In this study, the preference curve of substrate was investigated on the basis of field survey. Further, a numerical simulation by making use of a horizontal 2D numerical model was carried out with changing the discharge at the Gokasegawa River due to a lot of ayu inhabits in the Gokasegawa River. The numerical model is based on the model which was suggested by Nagata et al. [Nagata, N., Hosoda, T., Muramoto, Y., 2000. Numerical analysis of river channel processes with bank erosion. Journal of Hydraulic Engineering, ASCE 126 (4), 243–252]. The suitability of spawning for ayu is predicted with the preference curves of the flow depth, velocity and substrate. [Copyright &y& Elsevier]

Abstract: A rhamnose-binding lectin, named SFL, was isolated from the eggs of ayu (sweet fish, Plecoglossus altivelis) by affinity and ion-exchange chromatographies. SFL revealed 287 amino acid residues with 3 tandemly repeated domains, and contained 8 half-Cys residues in each domain. The lectin was shown to have a highly specific binding affinity to globotriaosylceramide (Gb3) by frontal affinity chromatography using 100 oligosaccharides. SFL was localized in several tissues and serum of both male and female ayu, such as gill, liver, ovary, testis, intestine, stomach, brain, kidney and serum. The lectin agglutinated the spores of the microsporidian Glugea plecoglossi, which is a pathogen of ayu. Although SFL bound to glycoproteins and glycolipids of G. plecoglossi spores, Gb3 could not be detected in either of them. The results suggest that SFL could interact with various glycoconjugates of pathogens to play a role in the adhesion of microorganisms invading in the body. [Copyright &y& Elsevier]

To clarify the feeding habits of tidal mudflat fishes, the gut contents of 29 fish species, collected from unvegetated tidal mudflats in Tokyo Bay, central Honshu, Japan, were examined. Onto- genetic changes in food preference were recognized in 21 species, including several of commercial importance (e.g. Acanthogobius flavimanus, Konosirus punctatus, Mugil cephalus cephalus, Plecoglossus altivelis altivelis, and Sardinella zunasi). In general, larvae and/or juveniles of these species fed mainly on small zooplankton or benthic harpacticoid copepods, later switching to other prey items with growth (e.g. gammaridean amphipods, mysids, polychaetes, detritus, bivalves, and juvenile fishes). A cluster analysis based on dietary overlaps showed that the tidal mudflat fish assemblage comprised six feeding guilds (small benthic and epiphytic crustacean, zooplankton, detritus, mollusc, polychaete, and fish feeders). Of these, small benthic and epiphytic crustacean feeders were the most abundantly represented in the number of species. [ABSTRACT FROM AUTHOR]

Metallothionein (MT) has been used widely as a potential molecular marker to detect the deleterious effects of heavy metals in aquatic ecosystem. Here we exposed ayu, Plecoglossus altivelis, to zinc (Zn) and tested the distribution as well as the induction of MT in various tissues such as liver, kidney, intestine and stomach. MT induction was significant in liver tissue, followed by kidney and intestine, whereas no induction was detected in stomach. The gene encoding ayu MT was successfully cloned and characterized. Complete nucleotide sequencing and analysis of the 4.5 kb DNA fragment containing the ayu MT gene revealed that the gene has three exons interrupted by two introns, a 5′-flanking region of about 2.5 kb and about 1.6 kb of 3′-flanking region. In grouper heart and kidney cells, the 2.5 kb promoter containing eight metal responsive elements (MREs), two hepatic nuclear factor 5 responsive elements (HNF5REs) and one cAMP responsive element (CRE) had the highest reporter activity. [Copyright &y& Elsevier]

In order to detect sex-linked DNA markers, linkage analysis was performed in each of three ayu half-sib families using 32 amplified fragment length polymorphism primer combinations, four microsatellite DNA primer sets and phenotypic sex (male). In each family the sex-determining locus appeared in a distal position on one of the linkage groups in which the D7-141* marker was strongly associated with male gender. The D7-141* marker was amplified in all of the male ayu (100%), and 8.7% of the female ayu. The D7-141* marker was detected in all male ayu at a level similar to that in the 40 wild individuals. These results suggest that the D7-141* marker is a male-specific marker linked with the sex-determining locus of the male in amphidromous ayu. [ABSTRACT FROM AUTHOR]

The Ryukyu-ayu, Plecoglossus altivelis ryukyuensis, is an amphidromous fish that is endemic to Amami-oshima Island in southernmost Japan. Its abundance, however, has been appreciably reduced during the last two decades such that the subspecies is now considered to be endangered. The variation of the mitochondrial DNA control region was investigated among specimens of the extant populations in the eastern and western parts of Amami-oshima Island (Sumiyo Bay and Yakeuchi Bay areas, respectively), using restriction fragment length polymorphism analysis in order to estimate the extent of gene flow between the two areas. Of a total of 165 fish including temporally different samples, four haplotypes were detected and each area possessed the two haplotypes. However, the common haplotypes shared between these areas were not observed, which indicates that recent gene flow has not occurred between these populations. The nucleotide divergence between populations was much higher than the nucleotide diversity within each population, and the neighbor-joining phylogram among haplotypes showed that the haplotypes are associated with their geographic area. These results suggest that the two populations of Ryukyu-ayu on Amami-oshima Island have been historically formed. [ABSTRACT FROM AUTHOR]

Effects of macromolecule synthesis inhibitors on the light-induced phase shift of the circadian clock in the photoreceptive pineal organ of a teleost, ayu (Plecoglosus altivelis) were investigated using metatonin release as an indicator. A single light pulse during the early- and late-subjective night delayed and advanced the phase of the circadian rhythm in melatonin release, respectively. During the late subjective-night, protein synthesis inhibitor cycloheximide (CHX) delayed the rhythm while RNA synthesis inhibitor 5,6-dichlorobenzimidazole riboside (DRB) had little effect. Light-induced phase advance was diminished by the treatment of CHX but not by DRB. During the early subjective-night. DRB, CHX. light and combination of these (DRB+light, CHX+light) all phase-delayed the rhythm. There were no additive effects of light and DRB or CHX. These results indicate that macromolecule synthesis is somehow involved in generation of circadian oscillation, and that de novo protein synthesis is required for light-induced phase shift of the circadian clock in the ayu pineal organ. [ABSTRACT FROM AUTHOR]

The landlocked dwarf ayu (koayu) Plecoglossus altivelis altivelis in Lake Biwa is one of the most important predators of planktonic crustacea. As part of predation ecology of koayu, the process of gastric evacuation was examined by two kinds of aquarium experiments. In the first experiment, koayu was fed for 1 h and the stomach contents after feeding was enumerated. Stepwise multiple regression analysis of the number of prey in the stomach on eight independent variables (numbers of six kinds of behavior during feeding, food concentration, and predator mass) selected only the number of foraging attacks as the predictor variable. In the second experiment, koayu was starved for 2 h after 1 h feeding. Prey number in the stomach before starvation was calculated from the number of foraging attacks and was compared with the number actually enumerated after starvation. Almost all stomach contents were digested and evacuated during the 2-h starvation. The stomach contents of koayu were, hence, ingested less than 2 h before the moment of collection. The present results enabled the estimation of predation rate in the field from the mass of stomach contents. [ABSTRACT FROM AUTHOR]

Ayu (Plecoglossus altivelis) is a teleost whose gonadal development is stimulated by shortened daylength and is a useful model to study the mechanism of photoperiodism. However, localization and characteristics of the photoreceptor that mediates photoperiodism in gonadal development remain to be determined. To identify the photoreceptive molecule that regulates photoperiodic responses, in the present study, we have cloned and characterized the cDNA encoding an opsin gene expressed in the ayu brain, a putative site of the photoreceptor for photoperiodism. The identified opsin was rhodopsin that is identical to the rhodopsin expressed in the retina. Phylogenetic analysis demonstrated that this rhodopsin belongs to the retina-type but not to the pineal-specific rhodopsin group. Genomic polymerase chain reaction (PCR) demonstrated that the ayu rhodopsin gene is intron-less. Southern and Northern blots and reverse-transcription PCR analyses indicate that the same rhodopsin gene is expressed in the retina and the brain but not in the pineal organ of ayu. These results indicate that the rhodospin gene is expressed in the retina and brain and mediates not only visual but also nonvisual functions such as photoperiodism and entrainment of the circadian clock. [ABSTRACT FROM AUTHOR]

Larval and juvenile ayu from seven year-groups (1986, 1987, 1992, 1995, 1996, 1999 and 2000) were collected in the Shimanto Estuary and adjacent coastal waters. The present study focuses on the variation among year-groups in the hatching period estimated by age determination using the otolith. The hatching period of the dominant cohorts in the 1986, 1987 and 1992 year-groups occurred from late October to mid-November. A delay of the hatching period of the dominant cohort was observed from the 1995 year-group, which occurred in late November to early December, and the dominant periods in the 1996, 1999 and 2000 year-groups were observed from early to late December. It appears that the delay of the hatching period of dominant cohorts was not due to a delay of spawning, but a high mortality of early hatched larvae. The water temperature in autumn in the coastal waters adjacent to the Shimanto River has tended to rise over the past 20 years and this trend was especially notable in the late 1990s. The delay in the hatching period of the dominant cohort observed from the 1995 year-group was likely to be related to the rise in seawater temperatures in autumn. [ABSTRACT FROM AUTHOR]