Revealing the compositions of the intestinal microbiota of three Anguillid eel species using 16S rDNA sequencing

Anguilla japonica, A. marmorata and A. bicolor pacifica are important eel species for aquaculture in Taiwan. In the past, seldom studies focused on the intestinal microbiome of them or the probiotics used in their cultivation. Probiotics are microbes that can improve the health of organisms, and the microbes isolated from the intestines of organisms might be the potential probiotics. Therefore, the aim of this study was to collect the intestinal mucus samples of these three eel species for investigating the autochthonous microbiome using 16S rDNA sequencing. The diversity, abundance of the intestinal microbiome and compositions of all the libraries were identified. Moreover, the composition of intestinal microbiome of eel was suggested that it would be affected by the waters and the characteristics of different eel species. Furthermore, most of the identified predominant genera of the intestinal microbiome of these three eel species, such as Cetobacterium, Clostridium, Shewanella, Acinetobacter and Bacteroides, seemed to have great potential as the probiotics. In the future, more studies shall be conducted to promote the practical application of these candidate probiotics for improving the techniques of eel farming.     

Comparison of glass eel stages of American eel and speckled worm eel in a northeast Florida estuary

Since 2001, a glass American eel, Anguilla rostrata (Lesueur), survey has been conducted at Guana River Dam, Florida, USA. Although present in earlier samples, glass speckled worm eel, Myrophis punctatus Lütken, were not identified until a large proportion appeared in samples in 2006. To determine whether the species could be separated in historical samples, weight–length relationships were compared between the two species. Then, with data from the full survey period, relationships between catch rate and annual percent contribution of glass speckled worm eel were explored in relation to temperature, water flow and salinity. Glass speckled worm eel were found to weigh significantly less and had distinguishing physical characteristics that could be used for identification. Glass speckled worm eel constituted between 0 and 34% of the catch, with no detectable pattern in timing or abundance among years. In 2006 and 2012, which had the two largest catches of glass speckled worm eel, relationship between nightly catch rate and measured environmental variables was inconsistent, even when measured variables had a similar range in values. It is recommended that south‐eastern, including Gulf of Mexico, USA, states restrict harvest of glass American eel to prevent the incidental capture and exportation of glass speckled worm eel. Efforts should be made to ensure proper species identification in glass eel surveys.     

Hen egg yolk and skinned krill as possible foods for rearing leptocephalus larvae of Anguilla japonica Temminck & Schlegel

Usual diets for rearing leptocephalus larvae of Japanese eel Anguilla japonica include eggs of the endangered spiny dogfish Squalus acanthias (SE). We investigated the effects of alternative food materials, hen egg yolk (HEY) and exoskeleton‐free (skinned) Antarctic krill (SAK), on the growth and survival of eel larvae. We found that feed comprising whole krill including exoskeleton (WAK) containing higher levels of fluoride (37.89 mg kg^?1) was acutely toxic to eel larvae exposed to this alone. In contrast, extract from SAK containing lower concentrations of fluoride (4.25 mg kg^?1) showed no apparent adverse effects. Growth of larvae fed a mixture of SE and SAK in a feed trial of 58 days [mean body weight (BW), 6.0 mg] was about twofold higher than that of larvae fed a mixture of SE and WAK (3.2 mg) (P < 0.01). A mixture of HEY and SAK also had some dietary benefits for eel larvae, enabling them to survive for up to 58 days and to grow significantly (mean BW, 2.4 mg), compared with their initial weight (mean BW, 0.2 mg) (P < 0.001). Although additional nutritional improvements are needed, the present results suggest that combination diet HEY and SAK may be a good alternative to SE as an effective diet for eel larvae.     

Ecology and conservation of the American eel in the Caribbean region

The majority of American eel, Anguilla rostrata LeSueur, knowledge is derived from temperate regions in the United States and Canada, with little known from its tropical Caribbean distribution. Findings of original research on American eel distribution, abundance, population biology, habitat ecology and threats from the Caribbean island of Puerto Rico were synthesised. American eel were captured from 48 of 116 sites (41.4%) in 26 of 49 river basins (53.1%) during 2005–2016, and it was extirpated upstream of dams and migration barriers >3.0 m high (38.9% of habitat). Mean density and biomass were 438.9 fish/ha and 23.44 kg/ha, respectively. Upstream habitats favoured larger individuals, and females were larger than males. The swim‐bladder parasite Anguillicoloides crassus Kuwahara, Niimi & Hagaki was not found in 120 eels examined. Realised threats include dams and other migratory barriers, habitat loss and alteration and pollution; exotic species and commercial fishing are impending threats; and the least understood is climate change.     

Natural dietary additive yellow loess as potential antibiotic replacer in Japanese eel,Anguilla japonica: Effects on growth,immune responses,serological characteristics and disease resistance against Edwardsiella tarda

The present experiment was conducted to evaluate the efficacy of an additive derived from the nature as an alternative of dietary antibiotic in Japanese eel, Anguilla japonica. Six experimental diets were formulated to contain no antibiotics or additive (yellow loess/YL) (control/CON), three graded levels of yellow loess at 5 (YL₅), 10 (YL₁₀) and 20 g/kg (YL₂₀), oxytetracycline at 5 (OTC) and amoxicillin at 10 g/kg amoxicillin (AMX) of diet. Weight gain (WG) and specific growth rate (SGR) from fish fed CON or YL₅ diets were significantly lower than those of fish fed YL₂₀ or OTC diets. Among non‐specific enzyme, lysozyme activity of fish fed YL₂₀, OTC or AMX was detected to be significantly higher than that from fish fed CON or YL₅ diets, whereas superoxide dismutase (SOD) activity of the fish fed CON was significantly lower than that for fish fed other experimental diets. Challenge test with bacteria, Edwardsiella tarda, showed improved disease resistance among the fish fed different levels of natural additive without any statistical difference from those fed antibiotics (OTC and/or AMX) supplemented diets. Therefore, these results demonstrated the potential of natural feed additive, yellow loess to replace oxytetracycline and/or amoxicillin in Japanese eel, A. japonica.     

Evaluation and characterization of a novel probiotic Lactobacillus pentosus PL11 isolated from Japanese eel (Anguilla japonica) for its use in aquaculture

In the present study, a potential Lactobacilli probiotics were isolated from Japanese eels (Anguilla japonica) and characterized and evaluated for their possible use in eel farming. Sixteen Lactobacilli were isolated from intestines of Japanese eels, using selective media. The lactobacilli strains (represented as PL1 to PL16) were screened by their ability to produce digestive enzyme. Among these, three strains (PL11, PL13 and PL16) producing four digestive enzymes (amylase, cellulase, protease and phytase) simultaneously were characterized further using API ZYM kit. From these, PL11 (Lactobacillu (L.) pentosus) was identified as potential probiotics candidate producing 15 enzymes among 20 tested. Further examination of biological activities of PL11 revealed tolerance against pH, artificial bile juice and antibacterial activity against several fish pathogenic bacteria. The in vitro competitive exclusion assay also revealed 88.4% reduction in adhesion of fish pathogen (Edwardsiella tarda) by PL11 to host intestinal mucus. In vitro incubation of Japanese eel foregut with Baclight‐labelled PL11 showed colonization of the enterocyte surface by confocal and scanning electron microscopy. In summary, PL11 isolated from eels could serve as a potential probiotics with acid and bile tolerance, production of digestive enzymes, antibacterial activity and inhibition of fish pathogen adhesion to intestinal mucus.     

Development and validation of a real‐time PCR assay for the detection of anguillid herpesvirus 1

Anguillid herpesvirus 1 (AngHV1) causes a haemorrhagic disease with increased mortality in wild and farmed European eel, Anguilla anguilla (L.) and Japanese eel Anguilla japonica, Temminck & Schlegel). Detection of AngHV1 is currently based on virus isolation in cell culture, antibody‐based typing assays or conventional PCR. We developed, optimized and concisely validated a diagnostic TaqMan probe based real‐time PCR assay for the detection of AngHV1. The primers and probe target AngHV1 open reading frame 57, encoding the capsid protease and scaffold protein. Compared to conventional PCR, the developed real‐time PCR is faster, less labour‐intensive and has a reduced risk of cross‐contamination. The real‐time PCR assay was shown to be analytically sensitive and specific and has a high repeatability, efficiency and r²‐value. The diagnostic performance of the assay was determined by testing 10% w/v organ suspensions and virus cultures from wild and farmed European eels from the Netherlands by conventional and real‐time PCR. The developed real‐time PCR assay is a useful tool for the rapid and sensitive detection of AngHV1 in 10% w/v organ suspensions from wild and farmed European eels.     

A size‐age model based on bootstrapping and Bayesian approaches to assess population dynamics of Anguilla anguilla L. in semi‐closed lagoons

A size‐age modelling technique is presented for assessing the vital rates, stock and recruitment of eel populations in semi‐closed lagoons with fully monitored migration of silver eels. Data for yellow and silver European eels (Anguilla anguilla L.) were obtained in 2011 from the Comacchio lagoon (Italy). The analysis was performed in three steps: (i) correction of yellow eel data, which are affected by the fyke nets selectivity during samplings, (ii) estimation of survival curve, stock, recruitment and metamorphosis rates of the population (calibration using data from 2011) and (iii) validation of the model using the observed amount of silver eel migrating population of the next year. A bootstrap procedure was used to assess the level of uncertainty for each parameter using the 95% intervals of the highest posterior density distribution HPDD (Bayesian approach). The measured abundance of silver eels was 0.56 ind·ha^?1, while the yellow eel abundance and recruitment were estimated by the model for 2011 at 8.77 ind·ha^?1 and 5.99 ind. ha^?1 respectively. The model performance during validation was satisfactory as the observed total mass of migrating population of 2012 (3777 kg) was inside the 95% HPDD intervals (3197–3839 kg) of model's predictions. The estimated stocks and recruitment were at least ten times lower from the respective estimations of previous studies of 1989 highlighting the crucial conditions of the population. The proposed modelling approach can provide significant information about eel population conditions, facilitating the evaluation of a range of management options in the context of eel conservation plans.     

The effect of foraging and ontogeny on the prevalence and intensity of the invasive parasite Anguillicola crassus in the European eel Anguilla anguilla

Infection patterns of the invasive Anguillicola crassus nematode were investigated in a population of the European eel Anguilla anguilla where parasite invasion is very recent, Loch Lomond, Scotland. Intensity levels of the parasite were associated with differences in fish ontogeny and trophic ecology. Although eels foraged on both fish and invertebrates, individuals which were smaller and fed on invertebrates (>70% contribution to diet) were found to contain a greater number of swim bladder parasites compared to larger eel with a predominance of fish (>60% contribution) in their diet. Within affected fish, a significant negative relationship was found between fish length and parasite intensity, with smaller individuals having higher parasite intensity than larger individuals. This study indicates that food intake and infection risk are linked in this recently infected host–parasite system. From a management perspective increasing our understanding of how infection intensity and repeated exposure is linked to resource use in an ecosystem is important for the future management of this endangered species in Europe.     

Aeromonas veronii caused disease and pathological changes in Asian swamp eel Monopterus albus

More and more diseases were occurred in the aquaculture of Asian swamp eel. Bacterial isolates were collected from the pathogenic organs of diseased Asian swamp eels. A dominant strain, Aeromonas veronii JL‐01, was isolated and identified from the liver and intestine tissue of diseased Asian swamp eels. The symptoms of fish infected artificially with A. veronii were the same as those observed under natural infection. The mortality rate was 40%–80% and infected Asian swamp eels died 2–3 days after infection. Drug sensitivity tests showed that the isolated bacterium was highly sensitive to seven kinds of antibiotics, including cephalosporins, aminoglycoside, quinolones, tetracyclines (except Tetracycline), pheniclos, nitrofurans and sulfonamides. The bacterium was resistant to penicillins (except Piperacillin), macrolides, polypeptides and lincomycin. The histological changes in Asian Swamp Eel induced by A. veronii infection mainly occurred in the liver, intestines and kidney. A. veronii showed strong pathogenicity towards the Asian swamp eel, causing tissue damage and death. Gentamycin, doxycycline and sulfamethoxazole could be used for preventing and treat this disease.     

Mortality of European eel after downstream migration through two types of pumping stations

Although numerous pumping stations (PS) have been used by water managers for numerous applications on rivers, canals and other water bodies, their impact on fish populations is poorly understood. This study investigates European eel, Anguilla anguilla (L.), mortality after natural downstream passage through a propeller pump and two Archimedes screw pumps at two PSs on two lowland canals in Belgium. Fyke nets were mounted permanently on the outflow of the pumps during the silver eel migration periods. Based on the condition and injuries, maximum eel mortality rates were assessed. Mortality rates ranged from 97 ± 5% for the propeller pump to 17 ± 7% for the large Archimedes screw pump and 19 ± 11% for the small Archimedes screw pump. Most injuries were caused by striking or grinding. The results demonstrate that PSs may significantly threaten escapement targets set in eel management plans.     

Bait worms: a valuable and important fishery with implications for fisheries and conservation management

Bait is an integral part of coastal life, but is perceived as a low‐value resource as fisheries are data‐limited, locally focussed and largely unregulated even though the ecological impacts of collection are considerable. An empirical assessment of three UK‐based ragworm fisheries combined with an analysis of published literature has produced the first global assessment of polychaete bait fisheries. The five most expensive (retail price per kg) marine species sold on the global fisheries market are polychaetes (Glycera dibranchiata, Diopatra aciculata, Nereis (Alitta) virens, Arenicola defodiens and Marphysa sanguinea). We estimate that 1600 t of N. virens per annum (worth £52 million) are landed in the UK with approximately 121 000 tonnes of polychaetes collected globally valued at £5.9 billion. Using remote closed circuit television (CCTV) cameras to monitor collectors, activity at local sites is considerable with a mean of 3.14 collectors per tide (day and night) at one site and individuals digging for up to 3 h per tide, although intensity differed seasonally and between sites. Collectors removed on average 1.4 kg of N. virens per person per hour, walking a considerable distance across the intertidal sediment to reach areas that were usually already dug. The implications of these human activity and biomass removal levels are explored in the context of fisheries and conservation management. At local, regional and national scales, polychaete bait fisheries are highly valuable, extract significant biomass and have considerable impacts; therefore, they urgently require governance equivalent to other fisheries.     

Effects of temperature on amoebic gill disease development: Does it play a role?

A relationship between increasing water temperature and amoebic gill disease (AGD) prevalence in Atlantic salmon (Salmo salar) has been noted at fish farms in numerous countries. In Scotland (UK), temperatures above 12°C are considered to be an important risk factor for AGD outbreaks. Thus, the purpose of this study was to test for the presence of an association between temperature and variation in the severity of AGD in Atlantic salmon at 10 and 15°C. The results showed an association between temperature and variation in AGD severity in salmon from analysis of histopathology and Paramoeba perurans load, reflecting an earlier and stronger infection post‐amoebae exposure at the higher temperature. While no significant difference between the two temperature treatment groups was found in plasma cortisol levels, both glucose and lactate levels increased when gill pathology was evident at both temperatures. Expression analysis of immune‐ and stress‐related genes showed more modulation in gills than in head kidney, revealing an organ‐specific response and an interplay between temperature and infection. In conclusion, temperature may not only affect the host response, but perhaps also favour higher attachment/growth capacity of the amoebae as seen with the earlier and stronger P. perurans infection at 15°C.     

Virus infections of the European Eel in North Rhine Westphalian rivers

Viral infections have been suggested to play a role in the decline of the panmictic population of the European eel (Anguilla anguilla). However, despite the importance of knowledge about pathogenic eel viruses, little is known about their spread in the wild European eel population and only a few eel pathogenic viruses have been described so far. In this study, we aimed to investigate the health status of the A. anguilla stock in North Rhine Westphalia (NRW) State of Germany. For this purpose, we examined tissue samples of 16 elvers, 100 yellow eels and 6 silver eels, sampled from the rivers Rhine, Lippe and Ems. Virus detection was performed via a combination of cell culture and PCR. Next to the detection of frequently encountered pathogenic eel viruses (anguillid herpesvirus 1 and eel virus European X (EVEX)), we isolated the eel picornavirus 1 (EPV-1) from tissue of yellow eels and elvers and demonstrate the distribution of EPV-1 in wild eel population in NRW.     

Antibiotic resistance and repetitive‐element PCR fingerprinting in Aeromonas veronii isolates

This study evaluated antibiotic resistance and the related genes in total 47 Aeromonas veronii isolates from pet fish, eel (Anguilla japonica) and koi (Cyprinus carpio) in Korea. In comparison with the antibiotic susceptibilities of isolates from eel and koi, those of pet fish were more resistant to ceftiofur, aminoglycosides, tetracycline and nitrofurantoin. And isolates from pet fish showed high prevalences of class 1 integron, quinolones and tetracycline resistance determinants than those from eel and koi. Repetitive‐element palindromic PCR (rep‐PCR) showed larger diversities among A. veronii isolates. Collectively, pet fish may be a reservoir for multiple clones of A. veronii involved in antibiotic resistance. In this aspect, imported fish in the aquaculture trade should be steadily and continually screened for bacterial antibiotic resistance and related genes.     

Growth differences of Japanese eels Anguilla japonica between fresh and brackish water habitats in relation to annual food consumption in the Kojima Bay‐Asahi River system,Japan

To learn about the relationships between feeding and growth of temperate eels in freshwater and brackish water habitats, we analysed 533 yellow‐phase Japanese eels Anguilla japonica collected in both types of habitats in southeastern Japan. Because male eels were very rare in each habitat (FW, N = 1; BW, N = 20), characteristics of female eels were compared between the different habitats. Annual food consumption was evaluated with the consideration of instantaneous food consumption and annual activity period. Stomach fullness index (stomach content weight/body weight) was used as an indicator of instantaneous food consumption. The ratios of number of months with eel catch to those when eel sampling was conducted were used as an indicator of activity period. Female yellow eels tended to be older and slower growing in fresh water (N = 78; age, mean ± SD = 7.9 ± 2.4 years; growth rate, 59.8 ± 14.0 mm year^?1) than in brackish water (N = 229; age, 5.5 ± 1.8 years; growth rate, 90.1 ± 24.4 mm year^?1). Irrespective of sex, yellow eels in brackish water had a higher stomach fullness index and a greater ratio of months with eel catches, indicating greater annual food consumption by brackish water eels. These results indicate that greater annual food consumption contributes to the greater growth rates of Japanese eels in brackish water habitats.     

Ex‐post facto analysis of diseases of the Gulf of Mexico′s white shrimp Litopenaeus setiferus

This study presents an Ex‐Post Facto analysis of diseases of wild juvenile and adults of Litopenaeus setiferus collected from a field survey at the Natural Protected Area of Terminos lagoon, southern Mexico. The objective of the present approach was to determine if sampling site and/or shrimp age were contributing risk factors for disease between juvenile and adult shrimp; if there was a determined period of time in a year cycle when diseases were more critical, and if the analysis would help to decision‐ making considering what population would pose less risk of disease‐carrying when withdrawn for experimental purposes; all under an after‐the‐fact (ex‐post facto) approach. We identified that juvenile shrimp were at more risk of contracting some diseases in the estuarine environment and June, July and August months, were found to be a critical period when colonizing and parasitic diseases maintained a significant high prevalence in the shrimp population. These assumptions may help for decision‐making when wild shrimp have to be withdrawn from their natural environment for research purposes.     

clpV is a key virulence gene during in vivo Pseudomonas plecoglossicida infection

Interaction between bacterial pathogen and aquatic animal host is exceedingly complex, which involves large dynamic changes in gene expression during different stages of the disease. However, research on identifying key virulence genes based on the dynamics of gene expression changes of a one‐sided bacterial pathogen in tissue has not been reported so far across different stages of infectious disease. The clpV for the T6SS of Pseudomonas plecoglossicida was identified for a candidate for key virulence gene based on dynamic changes of gene expression. For the Epinephelus coioides infected using clpV‐RNAi strain, no deaths were observed up to 20 dpi. The spleens, kidneys and livers of all the E. coioides that received clpV‐RNAi strain failed to develop visible nodules at 5–8 dpi, with the swelling gradually disappearing. The burdens of clpV‐RNAi strain in the spleen and blood were greatly reduced at most of the time points after injection, and the burdens of clpV‐RNAi strain in the head kidneys and trunk kidneys also had a sharp reduction from 72 to 120 hpi. This paper provides a new insight into the discovery of key virulence genes of pathogens in infected tissue systems.     

Study of the distribution of active caspase‐3‐positive cells in turbot,Scophthalmus maximus (L.), enteromyxosis

Enteromyxosis caused by Enteromyxum scophthalmi is one of the parasitizations with a higher economic impact on turbot, Scophthalmus maximus (L.), aquaculture. This myxosporean produces severe catarrhal enteritis with abundant inflammatory infiltrates in the lamina propria‐submucosa (LP), epithelial detachment and leucocyte depletion of the lymphohaematopoietic organs. Some advances made on the pathogenesis pointed to a role of apoptosis in the enteromyxosis. Therefore, the main aim of this work was to employ the TUNEL assay and the anti‐(active caspase‐3) immunohistochemical assay to detect apoptotic cells in both healthy and E. scophthalmi‐infected turbot in order to establish the presence and distribution of apoptotic cells during development of the disease. More apoptotic cells located within the gastrointestinal epithelium were observed in the initial stages of the infection in E. scophthalmi‐infected turbot compared with non‐infected turbot. As the infection progressed, a higher degree of apoptosis occurred in the epithelium of folds heavily parasitized. In the severely infected turbot, apoptosis was also found among the leucocytes of the intestinal inflammatory infiltrates. Moreover, the number of active caspase‐3‐positive cells in the lymphohaematopoietic organs tended to increase with disease severity. In view of the results, increased apoptosis in the epithelium may favour the scaling that occurs during enteromyxosis and cell death of leucocytes in the intestinal LP, contributing to leucocyte depletion in severe cases.