Document ID: EPA-HQ-OPP-2018-0040-0004
Agency: epa
Document Type: Supporting & Related Material
Title: 
Posted Date: 2018-05-03T04:00Z

EPA BIOPESTICIDES AND POLLUTION PREVENTION DIVISION COMPANY NOTICE OF FILING FOR PESTICIDE PETITIONS PUBLISHED IN THE FEDERAL REGISTER  
(DRAFT January__, 2018)

EPA Biopesticides and Pollution Prevention Division contact: [Alan Reynolds 703-605-0515]

INSTRUCTIONS:  Please utilize this outline in preparing the pesticide petition.  In cases where the outline element does not apply, please insert "NA-Remove" and maintain the outline. Please do not change the margins, font, or format in your pesticide petition. Simply replace the instructions that appear in green, i.e., "[insert company name]," with the information specific to your action.

SUBMISSION: E-mail the completed template to: duggard.mari@epa.gov.

TEMPLATE:

Southern Gardens Citrus Nursery, LLC, 1820 County Road 833, Clewiston, FL 33440

PP XXXXX

	EPA has received a pesticide petition (XXXXX) from Southern Gardens Citrus Nursery, LLC, 1820 County Road 833, Clewiston, FL 33440 proposing, pursuant to section 408(d) of the Federal Food, Drug, and Cosmetic Act (FFDCA), 21 U.S.C. 346a(d), to amend 40 CFR part 180

   	1. by establishing a tolerance for residues of NA-Remove

	2. to establish an exemption from the requirement of a tolerance for the
	
	3. to establish an amendment/expansion of an existing tolerance exemption for the NA-Remove

   	1. microbial pesticide NA-Remove

	2. biochemical pesticide NA-Remove
	
	3. plant-pesticide Spinach Defensin Proteins in or on Citrus Fruits (Citrus Spp., Fortunella Spp., Crop Group 10-10; 40 CFR Section 180.41(c)(12)(13)).

	Pursuant to section 408(d)(2)(A)(i) of FFDCA, as amended, Southern Gardens Citrus Nursery, LLC has submitted the following summary of information, data, and arguments in support of their pesticide petition. This summary was prepared by Southern Gardens Citrus Nursery, LLC and EPA has not fully evaluated the merits of the pesticide petition. The summary may have been edited by EPA if the terminology used was unclear, the summary contained extraneous material, or the summary unintentionally made the reader conclude that the findings reflected EPA's position and not the position of the petitioner.

I. Southern Gardens Citrus Nursery, LLC Petition Summary
   
   PP XXXXX

A. Product Name and Proposed Use Practices

Modified spinach defensin genes 2 (SoD2), 2* (SoD2*), 7 (SoD7) and 8 (SoD8), respectively, have been incorporated into citrus, grapefruit, sweet orange, and lemon to confer tolerance/resistance to the plant disease Citrus Greening (CG), also known as Huanglongbing or HLB.

B. Product Identity/Chemistry

	1. Identity of the pesticide and corresponding residues. The pesticide and corresponding residues are identified as spinach defensin proteins. Defensin proteins are ubiquitous in plants and are known to exhibit antifungal and antibacterial activity.  Plant defensins are understood to act by binding to receptors on the membrane of plant pathogens and thereby causing the integrity of the host membrane to become compromised, leading to cell disruption and death.

2. Magnitude of residues at the time of harvest and method used to determine the residue. Spinach defensin proteins, expressed in disease resistant citrus, have been measured at very low levels (less than 25ppm in citrus leaf tissue (dry wt.)).  (Citrus fruits have yet to be produced on modified trees due to their juvenile nature. However, it is anticipated that levels of spinach defensin proteins expressed in fruits will be similarly low.) The petitioner believes no analytical method is needed because an exemption from the requirement is being sought. There is a long history of mammalian consumption of the entire spinach plant (both raw and cooked) as food, without causing any known deleterious human health effects or any evidence of toxicity.  Spinach plant leaves have long been part of the human diet and there have been no findings which indicate toxicity or allergenicity.  The portion of the citrus genome coding for spinach defensin proteins 2, 2*, 7 and 8, as well as for the marker genes neomycin phosphotransferase II (nptII) and ß-glucouronidase (gus) is well characterized.  An exemption from the requirement of a tolerance in all plants exists for both NPTII and GUS proteins at 40 CFR Section 174.521 and 40 CFR Section174.525, respectively. Non-occupational exposure such as drinking water exposure is minimal to non-existent since the genes are only expressed at very low levels within plant tissues. Disease resistant citrus expressing spinach defensins does not represent a source of novel potential allergenic or antinutrient proteins. 

C. Mammalian Toxicological Profile.  Spinach defensins are naturally occurring proteins with a long history of safe use in mammalian dietary consumption of food (spinach).  These proteins have no known deleterious human health effects or any evidence of toxicity to people, wildlife, or beneficial insects. 

Mammalian safety studies have been conducted with microbial-produced SoD2 and SoD7 proteins that have been demonstrated as equivalent to SoD2 and SoD7 expressed in disease resistant citrus. Results of these safety studies confirm that spinach defensin proteins are not toxic and lack potential to become allergens.  In an acute oral toxicity study (up and down method) in the mouse (female), the estimated acute lethal dose, LD50, was determined to be over 5,000 mg/kg for SoD2. In an in vitro study, microbial-produced SoD2 and SoD7 proteins were rapidly and extensively degraded in simulated gastric and intestinal conditions in the presence of pepsin (at pH 1.2) and pancreatin, respectively. Both microbial-produced SoD2 and SoD7 proteins demonstrated half-lives of approximately five minutes when subjected to pepsin digest, and both proteins were completely proteolyzed to amino acids and small peptide fragments in less than 1 minute in the presence of 0.15 mg/mL pancreatin. A modelling study indicated that SoD8 would be similarly rapidly degraded in simulated mammalian digestive fluids. These results indicate that spinach defensin proteins are highly susceptible to digestion in the human digestive tract and that the potential for adverse health effects from chronic exposure is virtually nonexistent. Moreover, proteins in general are not known to be carcinogenic. A search of relevant literature and databases indicated that the amino acid sequences of the disease resistant citrus-expressed SoD2, SoD2*, SoD7 and SoD8 proteins exhibit no significant homology to the sequences of known allergens or protein toxins. Thus, spinach defensin proteins are highly unlikely to trigger an allergic response, do not share sequence similarity to known toxins and therefore do not warrant additional toxicity testing.

The genetic material necessary for the production of the spinach defensin proteins are nucleic acids (DNA) which are common to all forms of plant and animal life. There are no known instances in which nucleic acids have caused toxic effects as a result of dietary exposure.

Collectively, the available data on spinach defensin proteins along with the history of safe use of consumption of spinach, establish the safety of the plant pesticides spinach defensin proteins and the genetic material necessary for their production in all citrus fruit commodities.

D. Aggregate Exposure

	1. Dietary exposure. 

	             i.     Food

Significant dietary exposure to spinach defensin proteins is unlikely to occur. Dietary exposures, to the degree they may occur via ingestion of fresh and processed citrus commodities, will be at levels far below existing dietary exposure to these proteins from regular consumption of spinach.  Moreover, such exposures are unlikely to be problematic because of the low toxicity and the high degree of digestibility of the proteins. 
         
            ii. Drinking water

The spinach defensin proteins are not likely to be present in drinking water because the proteins are only present in minute quantities within the citrus plant.  Such small quantities are further confined to the upper part of the plant, and there is no evidence that the proteins may enter the soil. 
         
         2.  Non-dietary exposure
         
Spinach defensin proteins are tightly bound membrane proteins which are expressed at extremely low levels in citrus (less than 25ug/g (ppm) leaf tissue dry wt.).  These proteins bind tightly in plant tissue within the plant matrix, which greatly reduces availability for any dermal or respiratory exposure. 
         
E. Cumulative Effects

Common modes of toxicity are not relevant to consideration of the cumulative exposure to spinach defensin proteins. The product has demonstrated low mammalian toxicity and there are currently no EPA-registered pesticide products containing defensin proteins.  Furthermore, there exists a long and documented history of safe use and consumption of these naturally occurring proteins in spinach such that biological effects do not appear to be cumulative with any other known compounds.

F. Safety Determination

	1. U.S. population. There is a long history of human consumption of spinach plants (both raw and cooked), without causing any known deleterious human health effects or any evidence of toxicity.  On the contrary, spinach is commonly regarded as a "super food" that serves as an excellent source of vitamins, minerals, and antioxidants.   ["Food $ense," Utah State Univ. Cooperative Extension (Apr. 2011) (available online at: http://extension.usu.edu/files/publications/publication/FN_Food$ense_2100-08pr.pdf).]  Recent U.S. consumption statistics indicate that, on average, 2 lbs. of spinach are consumed per person per year in the United States.  ["Spinach Profile," Agricultural Marketing Resource Center (June 2013) (http://www.agmrc.org/commodities__products/vegetables/spinach-profile/).]  

Similarly, citrus whole fruits and juices have been an important part of the American and international diets for centuries.  ["History of Citrus," All Foods Natural (2013) (available online at: http://www.allfoodsnatural.com/article/history-of-citrus.html).]  Non-occupational exposure such as drinking water exposure is minimal-to-non-existent since the gene is only expressed within plant tissues. Disease resistant citrus, expressing SoD proteins, does not represent a source of novel potential allergenic or antinutrient proteins. Finally, the proteins will be expressed at very low levels.

	2. Infants and children. Based on the lack of demonstrated toxicity, absence of allergenic potential, the high degree of digestibility of spinach defensin proteins and their documented history of safe use, dietary exposure is anticipated to be at very low levels, indicating with reasonable certainty that no harm to infants, children or adults will result from aggregate exposure to these proteins.  Exempting spinach defensin proteins from the requirement of a tolerance should pose no significant risk to humans or the environment.

G. Effects on the Immune and Endocrine Systems

Given the rapid digestibility of spinach defensin proteins, no chronic effects are expected. Spinach defensin proteins (or degradate products of these proteins) are not known, nor expected, to have any effect on the immune or endocrine systems. Proteins in general are not carcinogenic, therefore, no carcinogenic risk is associated with spinach defensin proteins.

H. Existing Tolerances

	NA-Remove

I. International Tolerances

	NA-Remove