Performing Department
(N/A)
Non Technical Summary
The proposed work directly responds to USDA SBIR Research Priority 8.7 Aquaculture, by developing methods to accelerate maturity in Seriola rivoliana (a high-value marine finfish). This will reduce generation time, allowing for accelerated selective breeding programs. The objectives of Phase II are to use hormonal and molecular tools to induce puberty and yield reproductively mature broodfish at the earliest time possible. This work targets broodstock, therefore no hormone-treated fish will be destined for human consumption.To effectively accelerate S. rivoliana puberty, we have chosen three hormonal treatments that we believe will condition the brain for earlier maturity, induce onset of ovarian development and vitellogenesis, and accelerate overall growth rate. A series of trials will be conducted over the two-year period. Initial work will refine the dose and administration of hormonal implants. Later, two six-month long trials will test and refine treatments. By analyzing levels of hormones in the brain and liver, and by comparing gonadal and somatic growth between treated fish and un-treated controls, we will identify if the methods are successful. The materials and procedures developed during this work can be used by the applicant (Ocean Era, Inc) and be licensed to other Seriola producers around the world.
Animal Health Component
(N/A)
Research Effort Categories
Basic
(N/A)
Applied
(N/A)
Developmental
100%
Goals / Objectives
This project will develop methods to accelerate maturity in Seriola rivoliana (a high-value marine finfish). This will reduce generation time, allowing for accelerated selective breeding programs. The objectives of Phase II are to use hormonal and molecular tools to induce puberty and yield reproductively mature broodfish at the earliest time possible.No hormone-treated fish will be destined for human consumption.Our working hypotheses are that:Stable and sustainable chronic low levels of E2 will increase the levels of ERa in the brain of juvenile female S. rivoliana;Addition of S. rivoliana IGF-1 to the E2 will mitigate any possible retardation of growth rate following E2 treatment; andElevated levels of Seriola FSH in the blood will further increase ERa levels and advance gonadal development into vitellogenesis.The ultimate objective of the proposed work is to identify an efficient method(s) of hastening puberty in juvenile S. rivoliana, thereby accelerating genetic improvement through compounding selective breeding benefits, and increasing production of high-quality seafood in the U.S. The Phase I work has demonstrated the fundamental feasibility of this approach, by showing the ability to manipulate hormonal levels in S. rivoliana that should lead to earlier maturation. The work will now seek to refine these manipulations, and reduce them to practice, by answering the following questions:1. Will prolonged, low levels of E2 at a dose of 5ng/Kg BW over 6-week intervals, administered by slow-release implant, induce upregulation of estrogen receptors in the brain of 12-month old, pre-pubescent S. rivoliana?2. What is the combined effect of IGF-1and FSH treatments (administered via plasmids expressing secretable hormones) in combination with E2 (administered by implant, 5ng/Kg BW) on growth rate, ERa and relevant miRNAs brain levels?3. What is the combined effect of IGF-1, FSH treatments and E2 on ovarian development and the onset of vitellogenesis?
Project Methods
To effectively accelerate S. rivoliana puberty, we have chosen three hormonal treatments that we believe will condition the brain for earlier maturity, induce onset of ovarian development and vitellogenesis, and accelerate overall growth rate. The treatments will consist of E2, IGF-1 and FSH. The responses to hormonal treatments in Phase I were smaller than anticipated, due to the (presumed) inconsistency in hormonal delivery from the implants. The Phase II research will therefore explore novel approaches to ensure that the hormone(s) are constantly present in the blood at low levels. Innovative Research America (http://www.innovrsrch.com/advantages.asp) offers commercially available subcutaneous E2 implants characterized by a steady release (zero order) kinetics, available in variable doses and sizes. For the proposed Phase II study, we will use these at a dose of 1µg E2 released over 90 days, predicted to release 11 ng/24 h or ~0.11 ng/g/day. Number of pellets, dose and duration will be adjusted to growth rate over the course of the six-month experiment. We will conduct a preliminary experiment to measure E2 blood levels, and adjust to maintain a range of 2-5 ng/ml circulating E2 levels.Since granting the Phase I award (but separate from the Phase I work plan), we have cloned S. rivoliana IGF-1. We will now therefore insert S. rivoliana IGF-1 cDNA into PE expression plasmids, and add a treatment of plasmid in PVP+ZP9 as one of the treatments. Through this approach, we will eliminate the presumed immune response, and ensure a prolonged exposure to IGF-1.We will administer plasmids via intramuscular injection with vectors expressing both Seriola FSH and IGF-1 (Sanchis-Benlloch, Nocillado, et al., 2017). Experiments comparing FSH circulating levels of FSH in seabass have shown that treatment with 1 µg/g BW plasmid exhibited higher and prolonged elevated levels of FSH, compared to treatment with the 0.5 µg recombinant FSH (Mazon, 2014).The Phase II experiments will focus on females, because onset of puberty in females occurs later than in males, and requires a larger size. We will perform two separate hormone treatments on cohorts of prepubescent S. rivoliana females, and compare metrics of maturation against a control group of female fish of the same age and size. For both treatments, these metrics include growth rate, onset of gonadal development, and upregulation of ERa in the brain. We will also determine how E2 and IGF-1 treatments affect FSH expression in the pituitary and FSH blood levels. Fish maintenance, treatment administration, and sample collection will take place at Ocean Era's facility at the Natural Energy Laboratory of Hawaii Authority (NELHA) in Kona, Hawaii. Tissue analysis will be led by our collaborators at the Institute of Marine and Environmental Technology (IMET) of University of Maryland Baltimore County (UMBC). The Zohar/Zmora lab at UMBC is specialized in translating the knowledge gained from genomic and cellular studies into manipulating reproductive outputs in striped bass and other cultured fish. Dr. Nilli Zmora, an Associate Professor, will lead the research, in close consultation with Dr. Yonathan Zohar, Chair of Department of Marine Biotechnology. This will include evaluation of E2 implants, preparation of the expression plasmids, and supply of the ingredients for the PVP preparations, qPCR analysis, and interpretation of the results. Dr. Zmora and a PhD student will travel to NELHA to conduct the preliminary experiment, and to train the Ocean Era staff in treatment preparations and sampling.