Salinity extrusion and resilience of coastal aquaculture to the climatic changes in the southwest region of Bangladesh.

The southwest coastal belt of Bangladesh is characterized by a fresh and saline water interaction which gives rise to a discrete inter-saline freshwater convergence zone (ICZ). Hydrology and farming along this transition zone is influenced by upstream and downstream abiotic factors including salinity intrusion and water flow. To better understand the changing geography of the transitional ICZ line and the relative influence of these hydrological events on farming therein, the recent study compared relative changes from 2010 to 2014 based on qualitative and quantitative survey work with 80 households of 4 villages (Shobna, Faltita, Badukhali and Rudaghora) from Khulna and Bagerhat district. Contrary to the conventional hotcake climate change induced salinity intrusion the study found a significant decrease in saltwater influx and increased freshwater volume in the ICZ villages, reflecting a seaward movement trend. Farmer perception shifted to low saline and freshwater in many areas where it was high saline and medium saline in 2010. The factual and perceived salinity were varied from 1 ± 0.44 to 2 ± 0.77 ppt in the studied villages. To confront the condition farmer diversified their farming pattern from single crop like either only shrimp or prawn culture to concurrent culture of shrimp-prawn, shrimp, prawn and rice with an increased production of (68-204 kg/ha), finfish (217-553 kg/ha) and dyke crop (92-800 kg/ha). Thus, affecting the socioeconomic condition of the farmer with an increase in average monthly income, reported for the better-off classes in 2014, ranged from 14,300 to 51,667 BDT and for the worse-off ranged from 5000 to 9900 BDT. In contrast, this average monthly income was 9500- 27,000 for better-off and 3875 - 8600 for worse-off classes, reported in 2010. Besides, farming areas (average 17% for better-off and -0.5% for worse-off) and land leasing (average increment rate per ha 50%) also increased among the surveyed farmers, reported in 2014 compared to 2010. In addition, several adaptation strategies like unrefined salt use, change of water use, diversification through prawn, finfish and dyke crops along with traditional shrimp and overall land use change have a positive impact on farmer's economic and nutritional security as well as farming intensity. The study showed a unique attributes of salinity extrusion in micro-level of ICZ line where farmers intensified farming system with indigenous knowledge to secured their livelihoods.

Gene Editing of the Catfish Gonadotropin-Releasing Hormone Gene and Hormone Therapy to Control the Reproduction in Channel Catfish, Ictalurus punctatus.

Transcription activator-like effector nuclease (TALEN) plasmids targeting the channel catfish gonadotropin-releasing hormone (cfGnRH) gene were delivered into fertilized eggs with double electroporation to sterilize channel catfish (Ictalurus punctatus). Targeted cfGnRH fish were sequenced and base deletion, substitution, and insertion were detected. The gene mutagenesis was achieved in 52.9% of P1 fish. P1 mutants (individuals with human-induced sequence changes at the cfGnRH locus) had lower spawning rates (20.0−50.0%) when there was no hormone therapy compared to the control pairs (66.7%) as well as having lower average egg hatch rates (2.0% versus 32.3−74.3%) except for one cfGnRH mutated female that had a 66.0% hatch rate. After low fertility was observed in 2016, application of luteinizing hormone-releasing hormone analog (LHRHa) hormone therapy resulted in good spawning and hatch rates for mutants in 2017, which were not significantly different from the controls (p > 0.05). No exogenous DNA fragments were detected in the genome of mutant P1 fish, indicating no integration of the plasmids. No obvious effects on other economically important traits were observed after the knockout of the reproductive gene in the P1 fish. Growth rates, survival, and appearance between mutant and control individuals were not different. While complete knock-out of reproductive output was not achieved, as these were mosaic P1 brood stock, gene editing of channel catfish for the reproductive confinement of gene-engineered, domestic, and invasive fish to prevent gene flow into the natural environment appears promising.

Effects of CRISPR/Cas9 dosage on TICAM1 and RBL gene mutation rate, embryonic development, hatchability and fry survival in channel catfish.

The current study was conducted to assess the effects of microinjection of different dosages of guide RNA (gRNA)/Cas9 protein on the mutation rate, embryo survival, embryonic development, hatchability and early fry survival in channel catfish, Ictalurus punctatus. Guide RNAs targeting two of the channel catfish immune-related genes, toll/interleukin 1 receptor domain-containing adapter molecule (TICAM 1) and rhamnose binding lectin (RBL) genes, were designed and prepared. Three dosages of gRNA/Cas9 protein (low, 2.5 ng gRNA/7.5 ng Cas9, medium, 5 ng gRNA/15 ng Cas9 and high, 7.5 ng gRNA/22.5 ng Cas9) were microinjected into the yolk of one-cell embryos. Mutation rate increased with higher dosages (p < 0.05). Higher dosages increased the mutation frequency in individual embryos where biallelic mutations were detected. For both genes, microinjection procedures increased the embryo mortality (p < 0.05). Increasing the dosage of gRNA/Cas9 protein increased the embryo mortality and reduced the hatching percent (p < 0.05). Embryonic development was delayed when gRNAs targeting RBL gene were injected. Means of fry survival time were similar for different dosages (p > 0.05). The current results lay the foundations for designing gene editing experiments in channel catfish and can be used as a guide for other fish species.

Generation of Myostatin Gene-Edited Channel Catfish (Ictalurus punctatus) via Zygote Injection of CRISPR/Cas9 System.

The myostatin (MSTN) gene is important because of its role in regulation of skeletal muscle growth in all vertebrates. In this study, CRISPR/Cas9 was utilized to successfully target the channel catfish, Ictalurus punctatus, muscle suppressor gene MSTN. CRISPR/Cas9 induced high rates (88-100%) of mutagenesis in the target protein-encoding sites of MSTN. MSTN-edited fry had more muscle cells (p < 0.001) than controls, and the mean body weight of gene-edited fry increased by 29.7%. The nucleic acid alignment of the mutated sequences against the wild-type sequence revealed multiple insertions and deletions. These results demonstrate that CRISPR/Cas9 is a highly efficient tool for editing the channel catfish genome, and opens ways for facilitating channel catfish genetic enhancement and functional genomics. This approach may produce growth-enhanced channel catfish and increase productivity.

Aquaculture genomics, genetics and breeding in the United States: current status, challenges, and priorities for future research.

Advancing the production efficiency and profitability of aquaculture is dependent upon the ability to utilize a diverse array of genetic resources. The ultimate goals of aquaculture genomics, genetics and breeding research are to enhance aquaculture production efficiency, sustainability, product quality, and profitability in support of the commercial sector and for the benefit of consumers. In order to achieve these goals, it is important to understand the genomic structure and organization of aquaculture species, and their genomic and phenomic variations, as well as the genetic basis of traits and their interrelationships. In addition, it is also important to understand the mechanisms of regulation and evolutionary conservation at the levels of genome, transcriptome, proteome, epigenome, and systems biology. With genomic information and information between the genomes and phenomes, technologies for marker/causal mutation-assisted selection, genome selection, and genome editing can be developed for applications in aquaculture. A set of genomic tools and resources must be made available including reference genome sequences and their annotations (including coding and non-coding regulatory elements), genome-wide polymorphic markers, efficient genotyping platforms, high-density and high-resolution linkage maps, and transcriptome resources including non-coding transcripts. Genomic and genetic control of important performance and production traits, such as disease resistance, feed conversion efficiency, growth rate, processing yield, behaviour, reproductive characteristics, and tolerance to environmental stressors like low dissolved oxygen, high or low water temperature and salinity, must be understood. QTL need to be identified, validated across strains, lines and populations, and their mechanisms of control understood. Causal gene(s) need to be identified. Genetic and epigenetic regulation of important aquaculture traits need to be determined, and technologies for marker-assisted selection, causal gene/mutation-assisted selection, genome selection, and genome editing using CRISPR and other technologies must be developed, demonstrated with applicability, and application to aquaculture industries.Major progress has been made in aquaculture genomics for dozens of fish and shellfish species including the development of genetic linkage maps, physical maps, microarrays, single nucleotide polymorphism (SNP) arrays, transcriptome databases and various stages of genome reference sequences. This paper provides a general review of the current status, challenges and future research needs of aquaculture genomics, genetics, and breeding, with a focus on major aquaculture species in the United States: catfish, rainbow trout, Atlantic salmon, tilapia, striped bass, oysters, and shrimp. While the overall research priorities and the practical goals are similar across various aquaculture species, the current status in each species should dictate the next priority areas within the species. This paper is an output of the USDA Workshop for Aquaculture Genomics, Genetics, and Breeding held in late March 2016 in Auburn, Alabama, with participants from all parts of the United States.