On August 1, 2013, EPA received TSCA Experimental Release Applications (TERAs) R-13-0003, -0004, -0005, -0006 and -0007 from Sapphire Energy, Inc. to test five different intergeneric strains of the photosynthetic green algae Scenedesmus dimorphus in open ponds. The purpose of the field test is to (1) evaluate the translatability of the genetically modified strains from the laboratory to an outdoor setting, and (2) to characterize the potential ecological impact (dispersion and invasion) of the genetically-modified microalgae. The introduced intergeneric DNA sequences include certain metabolism genes and an intergeneric marker gene that enables detection of the microorganism from environmental samples. Also, different regulatory elements controlling expression of the genes were used, resulting in the five intergeneric strains. The field trials are to be conducted at the University of California San Diego Biology Field Station (BFS) in La Jolla, CA.

Proposed Use and Field Study

The submission algal strains were created within the lab space in an enclosed building that is subject to an NIH R&D exemption (40 CFR parts 725.234 and 725.235). The strains will be transported on solid agar media to the BFS (3.3 miles away). The recipient and subject microorganisms will be grown up in culture flasks and then scaled up in hanging bag photobioreactors contained within one of the four greenhouses at the BFS. When the appropriate density is reached in the photobioreactor bags, they will be used to inoculate the outdoor miniponds.

The five algal strains are to be grown semi-continuously in up to eight 1,200 L capacity miniponds and the biomass will be harvested from the ponds. Ponds will operate with volumes of 600-800 L. The production volume (PV) for each of the five submission strains is estimated by EPA to be 9.8 x 10¹³ cfu for the 2-month long field test for a total algal cell production volume of 4.9 x 10¹⁴ cfu, based on the data provided by the submitter.

Algal samples will be sent for analysis/testing at the Sapphire Energy facility in San Diego, CA. As part of its program to characterize the potential ecological impact, the submitter will test the ability of the subject microorganisms to disperse aerially into trap ponds positioned at various distances at the field site, In addition they will perform a series of incubation experiments in the laboratory inoculating the subject microorganism into intact wild communities in water obtained from local lakes.

Samples will be taken periodically during the 2-month field test. Once the field experiment has been terminated, all biomass will be inactivated by bleaching or autoclaving. The submitter performed inactivation tests by bleaching and/or autoclaving, and EPA determined that all treatments are sufficient to inactivate the algal strains. Any pond spill will be contained within the secondary containment and treated with bleach, and liquid disposed of in the sewage system.

Conclusions of EPA's Review

EPA granted these TERA submissions on September 25, 2013. EPA determined that the proposed field tests will not present unreasonable risks to human health or the environment. This small-scale field test is necessary to test the outdoor growth of the algal strains, and to generate data regarding dispersal and competition. These data will be required before future larger scale field tests are approved.

For a copy of the original nonconfidential TERAs and the nonconfidential approval letter, please contact the TSCA Non-Confidential Information Center (NCIC) by phone at (202) 566-0280, or by fax at (202) 566-9744.