Recipient Organization
MICROBIO ENGINEERING, INC
3988 SHORT ST STE 100
SAN LUIS OBISPO,CA 934017574
Performing Department
(N/A)
Non Technical Summary
The major innovation proposed herein is to develop more productive and easily cultivated, high value, freshwater strains of microalgae for use in hatchery-nursery operations. Avoiding the need for seawater recycle and re-use will be a major advance in this business, addressing a major opportunity. A joint venture between MicroBio Engineering Inc. (MBE), a microalgae technology developer, and Reed Mariculture Inc. (RMI) the largest global supplier of algal feeds to the aquaculture industry, this project focuses on the near-term commercial applications of microalgae to supply hatchery/ nursery operations, produced at relatively small scale, one and two acres, for applications and marketing by RMI. There are considerable opportunities for process improvements and cost reductions, as well as improved quality, specifically a higher content of omega-3 fatty acids. Although essentially all work in this field has used marine diatoms, the use of freshwater species has significant advantages in their production, and potentially even in their utilization. The need for water recirculation in algae production is a challenge, in particular where seawater supply and/or discharge is a limiting factor in production and economics. For RMI, the production of freshwater strains of Thalassiosira or, possibly, the closely related Cyclotella would solve one of the major current costs and bottlenecks in their operations. In sum, this project addresses a major opportunity for improving the competitiveness of the current US microalgae aquaculture feed industry in the global marketplace and provides a pathway to larger-scale production of microalgae feeds for aquaculture.Phase I will demonstrate the cultivation and utilization of freshwater microalgae, specifically diatoms, for aquaculture feeds in hatchery and nursery operations. Freshwater diatom species, obtained from culture collections and other sources, will be tested for growth rates and productivity in laboratory batch and continuous cultures and selected strains will be cultivated in 3.5 m2 raceway ponds in a greenhouse. The objective is to obtain biomass productivity data over six months, allowing extrapolation to year-round production. Algae will be harvested by centrifugation and supplied to RMI as refrigerated concentrates, and used to feed rotifers, to demonstrate their suitability for use in aquaculture feed applications. Successful demonstration of the efficacy of using freshwater diatoms as feeds will allow a major reduction in costs by eliminating the expense of seawater supply and recirculation. Phase II will demonstrate increased production rates to further reduce costs and increase outputs. Freshwater microalgae feed production will allow RMI to retain its position as the leading supplier of microalgae feeds to the hatcheries and nurseries worldwide, protecting and expanding US jobs and exports. This project will also provide a stepping-stone towards the longer-term development of bulk microalgae feeds for aquaculture, to replace fish oil and other high-value ingredients.In brief, the specific, near-term benefits of this project are in the development of new lower cost sources of diatom aquaculture feeds for hatchery/nursery operations. RMI faces significant competitive pressures from abroad in the marketing of its microalgae products - achieving the goals of the Phase I and II projects will allow it to remain competitive, and even expand operations, with both preservation and expansion of well-paying jobs and US exports. The broader national benefits are in providing a novel approach for lower cost and greater resource potential (by not requiring seawater availability) for future, larger-scale production of bulk algae biomass for animal feed ingredients.
Animal Health Component
100%
Research Effort Categories
Basic
(N/A)
Applied
100%
Developmental
(N/A)
Goals / Objectives
The goal of Phase I is to demonstrate the cultivation and utilization of freshwater microalgae, specifically diatoms, for aquaculture feeds in hatchery and nursery operations. The successful demonstration of the efficacy of using freshwater diatoms as feeds will allow a major reduction in costs by eliminating the expense of seawater supply and recirculation.To achieve these goals the following objectives will be addressed:The growth rate and productivity of freshwater diatom strains, particularly species of Thalassiosira, will be measured in laboratory reactors in batch and continuous cultures.Following laboratory screening, chosen strains will be cultivated in 3.5m2 raceway systems in a greenhouse with the objective of obtaining biomass productivity data over six months, which would allow extrapolation to year-round production.Biomass will be harvested and supplied to Reed Mariculture Inc. and used to feed rotifers to demonstrate suitability for use in aquaculture feed applications. Biomass compositional analysis of the algae and rotifers will be performed.Objective of Phase I and II of this project is to reduce diatom costs to below $200/kg (dry basis).
Project Methods
In Phase I, MBE will demonstrate the ability to cultivate two strains of freshwater diatoms, within the order Thalassiosirales, in small-scale (3.4 m2) raceway ponds in a greenhouse, and harvesting by (centrifugation). The concentrated algal biomass will then be used in feeding Brachionus plicatilis rotifers by RMI. Biomass composition analysis of the algae and rotifers for C20 and C22 omega-3 fatty acid content will further support the utility and value of the algal and rotifer biomass. The anticipated results for the rotifer feeding by RMI are that both the feed utilization (in terms of rotifer growth rates, yields and composition) and quality would be similar to the marine T. weissflogii and T. pseudonanna feeds currently marketed by the company.Task 1. Initial Culture Selection. Initially, freshwater diatom strains will be screened for growth potential in Bubble Columns Photobioreactors (BC-PBR). These reactors allow comparison of cultures in parallel, under set light intensity, pH, temperature and media composition. Suitable strains are those that exhibit fast growth rates (in dilute cultures), high productivities (in light-limited cultures), and a high omega-3 fatty acid content. Initial strain selection will be based on two runs for each strain, perfromed under low and high light intensities (150 and 1500 uE/m2-s), with otherwise similar conditions (pH 7.8-8.0, with 2-4 meq alkalinity and temperature at 25 oC). Strains will be selected based on maximum growth rates (log phase of growth) and maximum productivity (during the linear, light limited, growth phase). This task will require one month, which includes project initiation and initial growth experiments. Using standard methods routinely performed at MBE, biomass determinations (ash-free dry weights, OD) and biochemical content (proteins, lipids, and carbohydrates) analysis will be performed on diatom cultures.Task 2. Greenhouse Pond Operations. The second and major technical project objective of this Phase I, is the outdoor mass cultivation of selected strains in raceway ponds inside a greenhouse at the MBE field site in Paso Robles, California. To start, two strains selected with the BC-PBR (Task 1) will be tested in duplicate ponds, and replaced by other strains if they do not grow well. During Phase I, four 3.4 m2 ponds will be operated in duplicate over six months, from August 1st to January 31st, thus permitting summer, fall and winter seasonal productivities to be monitored. The process begins with production of inoculum at the MBE laboratory in 800 mL bubble columns. At density, the culture is then transferred to 15 L panels which are used to inoculate 100 liter 0.5 m2 ponds (operated at 20 cm). These ponds are then used to inoculate the 1,000 liter 3.4 m2 ponds. It takes approximately one to two weeks to build up such a culture. The cultures will be grown with one to three harvest/dilutions per week, adjusted by season and weather to maximize productivity. The concentrated algae will be refrigerated and kept at near 0 oC until used. A two-month shelf-life has been determined by RMI to be safe for use as rotifer feeds. Algae will be supplied to RMI starting within two months of project initiation and continuing through the end of the project. The main data collected by MBE will be on productivity, determined by ash free dry weight measurements and hydraulic dilutions. Samples will also be analyzed for proteins-carbohydrates and lipids.Task 3. Rotifer Feeding Trials. The final technical objective is to test the algal biomass for suitability in microalgae feeds. This work will be carried out by rotifer feeding studies under a subcontract by Reed Mariculture Inc. (RMI) using the compact Feeding System for the rotifer Brachionus plicatilis.Rotifer feed tests will be carried out using the Compact Culture System™ (CCS) production platform, developed by RMI, and using procedures routinely employed in RMI's rotifer production systems. The CCS is essentially a scaled-down version of the 1,000 L continuous-production culture systems used by RMI to produce billions of rotifers per day. The MBE freshwater algae concentrate feed trials will employ six CCS units. One control unit will be fed RMI's standard rotifer feed, and the other test units will be fed test algae concentrates. Feed will be dosed into the CCS units hourly using multi-channel timer-controlled peristaltic pumps. Rotifers will be cultured at their optimal salinity and temperature of 15 psu and 26 °C. To prevent toxicity resulting from the accumulation of ammonia excreted by the rotifers, ClorAm-X ammonia neutralizer and pH buffer will be pre-mixed with the algae concentrate in amounts based on the nitrogen content of the feeds. Ammonia tests (API Ammonia Test Kit) will be performed daily to confirm that ammonia remains below 0.5 mg/L NH3 (the toxic form of ammonia).Daily rotifer counts will be made using Sedgewick-Rafter counting slides, and rotifer densities and egg ratios (which indicate rotifer reproductive status and overall culture condition) will be recorded. Before the daily harvest, the waste trap is removed from the CCS and the accumulated waste is flushed away with a jet of clean water, and the trap is sanitized by heating to 60 °C for 1 h before re-installing in the CCS. Daily harvests will be made by removing the appropriate volume of the culture and pouring through a 40 µm mesh screen (the waste trap of the CCS ensures that negligible amounts of particulate matter contaminate rotifer samples). Rotifer samples will then be rinsed with isotonic ammonium bicarbonate solution to remove salts and stored frozen at -20 °C for subsequent analysis (biomass dry weight yield, proximate nutritional content, and fatty acid profile). The data will be analyzed for any statistical differences in feed quality, and correlated with the algae composition.