Two epizootic Perkinsus spp. events in commercial oyster farms at Santa Catarina,Brazil

Perkinsus spp. have been detected in various bivalve species from north‐east Brazil. Santa Catarina is a South Brasil state with the highest national oyster production. Considering the pathogenicity of some Perkinsus spp., a study was carried out to survey perkinsosis in two oyster species cultured in this State, the mangrove oyster Crassostrea gasar and the Pacific oyster Crassostrea gigas. Sampling involved eight sites along the state coast, and oyster sampling was collected during the period between January 2013 and December 2014. For the detection of Perkinsus, Ray's fluid thioglycollate medium (RFTM) and histology were used, and for the identification of the species, PCR and DNA sequencing were used. Perkinsus spp. was found by RFTM in C. gigas and C. gasar from São Francisco do Sul. This pathology was also detected in C. gasar from Balneário Barra do Sul both, by RFTM and histology. Perkinsus marinus was identified in C. gigas and C. gasar from São Francisco do Sul and Perkinsus beihaiensis in C. gasar from Balneário Barra do Sul. This is the first report of P. marinus in C. gigas from South America. Results of this preliminary study suggest that both oyster species tolerate the species of Perkinsus identified, without suffering heavy lesions.     

Pathogenic threats and probiotic use in larviculture of the scallop,Pecten maximus

During a series of experiments, bacteriological elements in scallop larval rearing were investigated: larvae susceptibility to pathogens as a function of their age, and the use of probiotic bacteria during larviculture. Younger larvae (d5 PF) were highly more susceptible to pathogenic‐challenge than their older siblings, which were challenged at an older age (d15 PF). A challenge with 10⁴ CFU mL^?1 of V. pectenicida killed 100% of d5 PF larvae 7 days following challenge, yet killed only 9% of d15 PF larvae 9 days following challenge. Use of the probiotics Phaeobacter gallaeciensis, Alteromonas macleodii 0444 and Neptunomonas sp. 0536, provided for larger larvae, a high yield of competent larvae and, perhaps more importantly, protection against pathogen‐challenge similar to levels achieved from antibiotic use. When challenged with V. pectenicida, d29 survivals were 20.3%, 85.1% and 75.0% respectively for control (no probiotic), antibiotic treated, and ‘probiotic mix’ administered larvae. Use of potential probiotic Pseudoalteromonas sp. D41 appeared to hinder scallop larvae. Future use of probiotics in scallop larval rearing would benefit from combined use of P. gallaeciensis, A. macleodii 0444 and Neptunomonas sp. 0536.     

Recent advances of genome mapping and marker‐assisted selection in aquaculture

Aquaculture is the fastest growing sector in agriculture. Substantial genetic gains have been achieved in a few cultured species using conventional selective breeding approaches. However, the majority of fish and shellfish species remain in their wild state. Due to the recognition of the enormous potential of marker‐assisted selection (MAS) to speed up genetic gain through early selection, aquaculture scientists have constructed linkage maps in over 40 species and mapped quantitative trait loci (QTL) for important traits in over 20 species since the 1990s. Although MAS and genomic selection (GS) have not been widely used in aquaculture, their application in breeding programmes is expected to be a fertile area of research. In this paper, I summarized the recent advances of linkage and QTL mapping, as well as MAS in aquaculture species. I also discussed the potentials of genome‐wide association studies (GWAS) and GS in aquaculture species.     

Declines in the energy content of yearling non‐native alewife associated with lower food web changes in Lake Michigan

Juveniles of non‐native alewife, Alosa pseudoharengus (Wilson), were collected in Lake Michigan in 1998, 1999, 2010, 2011 and 2013 to evaluate changes in energy content during a period of major ecosystem changes. Consistent with historical data, energy content of yearling alewife declined from late winter into late spring and was at its lowest point in June. Energy density and length‐adjusted, entire‐body energy were lower in 2010, 2011 and 2013 than in 1998 and 1999. Energy losses over the first winter in the lake were more severe for the 2010 year class (56% decrease) than for the 1998 year class (28% decrease). Alewife diets in late spring of 2010–2013 reflected the loss of major prey such as Diporeia spp. and a shift towards lower energy prey. The recent decline in energy content of yearling alewife can be linked to recent changes in productivity and abundance of key components of the lower food web of Lake Michigan following the dreissenid invasion.     

Functional identification of ToLAAO genes and polymorphism association analysis of Cryptocaryon irritans resistance in Trachinotus ovatus

‘Marine white spot disease’ is caused by Cryptocaryon irritans infection and can lead to high mortality in Trachinotus ovatus. L-Amino acid oxides (LAAOs) play a key role in antibacterial activity and parasitic activity. To investigate the function of the LAAO (ToLAAO) and LAAO-like (ToLAAO-like) genes of T. ovatus, this study explored the sequence characteristics and relationship between polymorphisms and traits of anti-C. irritans. The ToLAAO and ToLAAO-like ORF sequences obtained from the whole genome of T. ovatus were 1563 and 1584 bp, which encoded 520 and 527 amino acids respectively. Both sequences contained a highly conserved flavin adenine dinucleotide-binding domain and a similar amino oxidase domain. Sequence multiple alignment analysis showed that ToLAAO and ToLAAO-like had the highest homology to the LAAO sequence of Larimichthys crocea. Quantitative real-time polymerase chain reaction (qRT-PCR) results showed that ToLAAO and ToLAAO-like mRNA were generally expressed in 10 tissues. ToLAAO mRNA was highly expressed in the testis, while ToLAAO-like mRNA was highly expressed in muscle tissue. After C. irritans infection, ToLAAO and ToLAAO-like mRNA were significantly upregulated in the skin and spleen, while only ToLAAO mRNA was significantly upregulated in the liver and head kidney, and only ToLAAO-like mRNA was significantly upregulated in the gills. Five SNP sites were identified from the ToLAAO and ToLAAO-like genomic sequence fragments, and two sites (6200C/T and 6237G/A) of LAAO were significantly associated with resistance to C. irritans. These results suggest that ToLAAO and ToLAAO-like genes play crucial roles in defending against the immune response to C. irritans.     

Development of an in vitro model to assess protein bioavailability in diets for common octopus (Octopus vulgaris)

An in vitro model designed to assess protein bioavailability in diets for growing Octopus was developed. The model required a previous assessment of some functional features of protein digestion in this species like the main producing organs, optimum pH for activity and total production per g tissue. The main producing organs identified were the salivary glands and the hepatopancreas (HP), being optimum pH for protease activity quite different in both organs (mostly alkaline in the posterior salivary glands and acid in the HP). In spite of the high‐specific protease activity measured in the salivary glands, a major role of the HP in protein hydrolysis is suggested due to the much bigger size of this viscera. All this information was used as a basis to develop an in vitro two‐step hydrolysis process, which simulated protein hydrolysis performed by these two organs using the Octopus enzymes. The assay was used to evaluate differences in amino acid bioavailability from several protein sources (casein, gelatin, fish meal, squid meal and krill meal) that could be used as feed ingredients for this species. As significant differences were detected both in total amount and in rate of release of the amino acids from such proteins, the model is proposed as a complementary tool in the selection and nutritional evaluation of protein ingredients to be used in diets for Octopus.     

Multiscalar land suitability assessment for aquaculture production in Uruguay

This article presents a highly detailed territorial assessment for aquaculture production in Uruguay and a methodology to identify the most suitable areas in order to develop the activity. Furthermore, it proposes the basis to generate a dynamic modelling tool of high spatial resolution to support the decision‐making process regarding national aquaculture activities. The modelling structure of aquaculture suitability was developed by the construction of a hierarchical model, combining a multicriteria assessment and a geographic information system. Forty‐one attributes were integrated in six aptitude models: one model for each one of the five common production systems intended to be the most viable in Uruguay and a sixth aptitude indicator that summarizes those five systems, representing the maximum suitability for aquaculture activities. In addition, the hierarchical model allows measurement and identification of relative weights of each model, according to the planning scale, ranging from basin to cell.     

Use of planted biofilters in integrated recirculating aquaculture‐hydroponics systems in the Mekong Delta,Vietnam

The feasibility of using planted biofilters for purification of recirculated aquaculture water in the Mekong Delta of Vietnam was assessed. The plant trenches were able to clean tilapia aquaculture water and to maintain good water quality in the fish tanks without renewal of the water. NH₄‐N was removed efficiently in the plant trenches, particularly in the trenches with Canna glauca L., probably because of plant uptake and nitrification–denitrification. Plant uptake constituted 6% of N and 7% of P in the input feed. Approximately 1.0 m³ of water was needed per kg of fish produced, and 370, 97 and 2842 g fresh aboveground biomass of Ipomoea aquatica Forssk., Lactuca sativa L. and C. glauca, respectively, were produced. The leafy vegetables provide some extra income besides fish products, whereas C. glauca provides nice flowers and contributes to a significant nutrient removal with annual uptake rates of 725 kg N and 234 kg P ha^?1 year^?1. This research demonstrates that integrated recirculating aquaculture‐hydroponics (aquaponics) systems provide significant water savings and nutrient recycling as compared with traditional fish ponds.     

First report of Shewanella sp. and Listonella sp. infection in freshwater cultured loach,Misgurnus anguillicaudatus

A novel disease outbreak occurred among cultured loach, Misgurnus anguillicaudatus in China. The diseased fish displayed lethargy, loss of appetite, black discolouration, anasarca and ventral skin hyperaemia. Clinical signs were observed in most of the fish examined, which consisted of discoloured liver, swollen spleen and petechial haemorrhage in the intestine and peritoneal membranes. The causative agent was found to be two species of bacteria by experimental infection, identified as Shewanella sp. and Listonella sp. The result showed that these were likely to be Shewanella putrefaciens and Listonella anguillarum respectively. Bacterial identification consisted of physiological and biochemical tests as well as 16S rRNA sequence analysis. In addition, antibiotic susceptibility testing was performed, revealing that Shewanella putrefaciens and Listonella anguillarum were susceptible to enrofloxacin, ciprofloxacin, levofloxacin, pipemidicacid and norfloxacin. This is the first report of S. putrefaciens and L. anguillarum from cultured loach.     

Bromodeoxyuridine DNA labelling reveals host and parasite proliferation in a fish‐myxozoan model

Enteromyxum leei is a myxozoan parasite responsible for enteritis in gilthead sea bream (Sparus aurata). The parasite proliferates in the paracellular space of the intestinal epithelium and induces an inflammatory reaction. To assess intestinal cell turnover and parasite proliferation, fish were infected with the parasite by anal intubation; after 17 and 64 days, the cell proliferative marker bromodeoxyuridine (BrdU) was administered; and after 24 hr, tissue samples were taken for immunohistochemical detection. Parasite exposure induced increased epithelial and immune cell proliferation in all intestinal segments at all time points, even before parasite establishment. This increased turnover was triggered early after intubation and mainly at a local level, as shown by an increased proliferating cell nuclear antigen (pcna) gene expression only at the posterior intestine after 17 days (not found in lymphohaematopoietic organs). Incorporation of BrdU in parasite secondary and tertiary daughter cells indicated that parasite endogeny is not by schizogonial division, which uses de novo synthesis pathway of pyrimidines. Altogether, BrdU immunolabelling and pcna gene expression showed the rapid proliferative response of the fish intestines upon a myxozoan infection and how this response is effectively triggered even before the parasite reaches or establishes in the site.     

Recent update on the prevalence of Vibrio species among cultured grouper in Peninsular Malaysia

Vibrio infections are common among marine fish and lead to serious problems in the aquaculture sector. This study reports a recent occurrence of Vibrio species (spp.) isolated from cultured groupers in Peninsular Malaysia using the gyrB and pyrH genes. A total of 147 Vibrio strains were successfully isolated from 77 (64%) groupers using culture method and subjected to gyrB and pyrH sequencing for species identification and confirmation. Results showed that 89% of Vibrio strains were identified and clustered to six groups of Vibrio spp., while 11% were not clustered to any Vibrio spp. using the gyrB sequences. Meanwhile, by analysis of the pyrH sequences all the 147 Vibrio strains (100%) were successfully identified and clustered into 11 groups of Vibrio spp., including the gyrB non‐identified strains. The pyrH gene provides a better resolution for identification of Vibrio spp. compared with the gyrB gene. Thus, the pyrH gene was more suitable for a rapid determination of Vibrio spp. distribution in Peninsular Malaysia. Using the pyrH gene, our study found higher prevalence of Vibrio vulnificus (33%), V. alginolyticus (24%) and V. parahaemolyticus (22%), followed by V. rotiferianus (5%), V. harveyi (3%), V. tubiashii (2%), V. campbellii (2%), V. ponticus (1%), V. diabolicus (1%), V. owensii (1%) and others Vibrio sp. (7%). Thus, the results of this study revealed that the occurrence of pathogenic vibrios among grouper fish is still high in Malaysian aquaculture. In addition, the pyrH gene was proved as a suitable marker for rapid identification of Vibrio species compared with the gyrB gene.     

The efficacy of MS‐222 as anaesthetic agent in four freshwater aquarium fish species

The efficacy of anaesthetic tricaine methanesulfonate (MS‐222) was evaluated in four freshwater aquarium fish species, Zebrafish (Danio rerio), Guppy (Poecilia reticulata), Discu (Symphysodon discus) and Green swordtail (Xiphophorus helleri). The correct dose of anaesthetic should induce the plane 4 of anaesthesia in less than 180 s, recovery in less than 300 s and must survive when exposed during 30 min to anaesthetic. Fishes were exposed to six concentrations of anaesthetic (75, 100, 125, 150, 200 and 250 mg L^?1) and the time of fish reaching plane 4 of anaesthesia, post exposure recovery, and the percentage of survival when fish were subject to 30 min in the anaesthetic were recorded. The optimal doses varied according to the species: D. rerio – 75, 100 and 125 mg L^?1, P. reticulata – 125, 150 and 200 mg L^?1, S. discus – 75 and 100 mg L^?1 and X. helleri – 125 and 150 mg L^?1. The induction time generally decreased significantly with increasing concentration of MS‐222 for all of the species evaluated. The recovery time had a tendency to increase with the increase of the MS‐222 concentration for D. rerio, P. reticulata and S. discus. On the other hand, X. helleri recovery time decreased with the increase of MS‐222 concentration. MS‐222 proved to be effective in anaesthesia for all the freshwater ornamental species studied. The main results clearly show that the optimal dose to anesthetize is fish species dependent and it is completely wrong to extrapolate optimal anaesthetic concentrations between different species.     

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.     

Winter kill in intensively stocked channel catfish (Ictalurus punctatus): Coinfection with Aeromonas veronii,Streptococcus parauberis and Shewanella putrefaciens

Unusual persistent natural mortality occurred in a floating in‐pond raceway system intensively stocked with channel and hybrid catfish beginning in early November 2016 up until March 2017. The temperature during the period of outbreak ranged from 7.2 to 23.7°C. Gross examination of freshly dead and moribund fish revealed pale gills, slight abdominal distension and swollen inflamed vents. Comprehensive necropsy of 20 fish demonstrated vast amounts of bloody ascitic fluid in the coelomic cavity, visceral congestion, splenomegaly and pale friable livers but macroscopically normal kidneys, suggesting systemic bacterial infection. Bacterial cultures were initiated from skin, gills and major internal organs. Following incubation, a mixture of three bacterial colony phenotypes was observed on agar plates. Presumptive biochemical characterization of the isolates followed by 16S‐rRNA sequence analysis resulted in the identification of Aeromonas veronii, Streptococcus parauberis and Shewanella putrefaciens. Channel catfish juveniles were experimentally infected with the recovered isolates to fulfil Koch's postulates. Moreover, an antibiogram was used to evaluate the susceptibility of the isolates to antimicrobial drugs approved for use in aquaculture. Aquaflor was used successfully for treatment. Here, we report bacterial coinfection lead by A. veronii and the first identification of S. parauberis and S. putrefaciens from cultured catfish in North America.     

Current knowledge of nocardiosis in teleost fish

Nocardia sp. is the causative agent of nocardiosis, a lethal granulomatous disease of the skin, muscle, and various inner tissues affecting various teleost and shellfish. Four species of Nocardia have been isolated from diseased fish and shellfish, namely Nocardia asteroides, Nocardia seriolae, Nocardia salmonicida and Nocardia crassostreae. Therefore, in fish aquaculture, nocardiosis has caused severe economic losses, especially in the Asian region. Considerable research has been performed, since the first report of identified Nocardia sp. in fish, to characterize Nocardia sp. and identify rapid detection techniques, immune response against infection and prophylactic approaches. In this review, the current state of knowledge about nocardiosis in fish has been presented, including the pathogenesis, diagnosis, host immune response and vaccine development.     

Identification and pathogenicity study of emerging fish pathogens Acinetobacter junii and Acinetobacter pittii recovered from a disease outbreak in Labeo catla (Hamilton, 1822) and Hypophthalmichthys molitrix (Valenciennes, 1844) of freshwater wetland in West Bengal,India

The disease outbreaks in aquaculture system of wetlands are the major cause of fish mortality. Among various bacterial septicaemic diseases, fish mortality caused by Acinetobacter spp. is recently reported in different fish species. Fish disease outbreak was investigated in a wetland of West Bengal, India to identify the aetiological factors involved. The moribund fish were examined and subjected to bacterial isolation. Two bacterial causative agents were identified as Acinetobacter junii and Acinetobacter pittii by biochemical characterization and 16S rRNA gene amplification. Both the isolates were oxidase‐negative, nitrate‐negative, catalase‐positive and indole‐negative. The molecular identification using 16S rRNA gene sequencing and phylogenetic tree analysis further confirmed the two Acinetobacter spp. with 97%–99% similarity. The antibiotic resistance patterns of these two bacteria revealed that both of them were resistant to β‐lactam, cefalexin, cephalothin, amoxyclav, cefuroxime, cefadroxil, clindamycin, vancomycin and penicillin. In addition, A. pittii was also resistant to other antibiotics of cephams group such as ceftazidime and cefotaxime. In the challenge experiment, both A. junii and A. pittii were found to be pathogenic with LD₅₀ of 1.24 × 10⁵ and 1.88 × 10⁷ cfu/fish respectively. Histopathological examination of gill, liver and kidney revealed prominent changes supporting bacterial septicaemia. The investigation reports for the first time on concurrent infection by A. junii and multidrug‐resistant (MDR)‐A. pittii as emerging fish pathogens to cause severe mortality in Labeo catla and Hypophthalmichthys molitrix in a freshwater wetland.