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Bacillus thuringiensis var. kurstaki Cry2A - Pesticide Petition Filing 3/98

[Federal Register: April 8, 1998 (Volume 63, Number 67)]
[Notices]
[Page 17174-17176]
From the Federal Register Online via GPO Access [wais.access.gpo.gov]
[DOCID:fr08ap98-79]
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ENVIRONMENTAL PROTECTION AGENCY
[PF-800;FRL-5781-1]
Notice of Filing of Pesticide Petition
AGENCY: Environmental Protection Agency (EPA).
ACTION: Notice.
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SUMMARY: This notice announces the initial filing of petition (PP
7F4822), submitted by Monsanto Company, proposing the establishment of
a regulation for an exemption from the requirement of a tolerance for
residues of the plant pesticide, active ingredient, Bacillus
thuringiensis variety kurstaki (B.t.k.) insect control protein
(CryIIA), when used in or on all food and feed crops.
DATES: Comments, identified by the docket control number PF-800, must
be received on or before May 8, 1998.
ADDRESSEES: By mail submit written comments to: Public Information and
Records Integrity Branch (7502C), Information Resources and Services
Division, Office of Pesticides Programs, Environmental Protection
Agency, 401 M St., SW., Washington, DC 20460, In person bring comments
to: Rm. 119, CM #2, 1921 Jefferson Davis Highway, Arlington, VA, 22202.
    Comments and data may also be submitted electronically by following
the instructions under "SUPPLEMENTARY INFORMATION". No confidential
business information should be submitted through e-mail.
    Information submitted as a comment concerning this document may be
claimed confidential by marking any part of the information as
"Confidential Business Information" (CBI). CBI should not be
submitted through e-mail. Information marked as CBI will not be
disclosed except in accordance with procedures set forth in 40 CFR part
2. A copy of the comment that does not contain CBI must be submitted
for inclusion in the public record. Information not marked confidential
may be disclosed publicly by EPA without prior notice. All written
comments will be available for public inspection in Rm. 119 at the
address given above, from 8:30 a.m. to 4 p.m., Monday through Friday,
excluding legal holidays.

FOR FURTHER INFORMATION CONTACT: By mail: Willie H. Nelson,
Biopesticides and Pollution Prevention Division (7511W), Office of
Pesticides Programs, Environmental Protection Agency, 2800 Crystal
Drive, Arlington, VA 22202, (703) 308-8682; e-
mail:nelson.willie@epamail.epa.gov.

SUPPLEMENTARY INFORMATION: EPA has received petitions as follows
proposing the establishment and/or amendment of regulations for
residues of certain pesticide chemicals/microbials in or on various
food commodities under section 408 elements set forth in section
408(d)(2); however, EPA has not fully evaluated the sufficiency of the
submitted data at this time or whether the data supports granting of
the petition. Additional data may be needed before EPA rules on
petitions.
    The official record for this notice of filing, as well as the
public version, has been established for this notice of filing under
docket control number [PF-800] (including comments and data submitted
electronically as described below). A public version of this record,
including printed, paper versions of electronic comments, which does
not included any information claimed as CBI, is available for
inspection from 8:30 to 4 p.m., Monday through Friday, excluding legal
holidays. The official record is located at the address in
"ADDRESSES" at the beginning of this document.
    Electronic comments can be sent directly to EPA at:
opp-docket@epamail.epa.gov

[[Page 17175]]

    Electronic comments must be submitted as an ASCII file avoiding the
use of special characters and any form of encryption. Comments and data
will also be accepted on disks in Wordperfect 5.1/6.1 file format or
ASCII file format. All comments and data in electronic form must be
identified by the docket number [PF-800] and appropriate petition
number. Electronic comments on this notice may be filed online at many
Federal Depository libraries.

List of Subjects

    Environmental protection, Agricultural commodities, Food additives,
Feed additives, Pesticides and pests, Reporting and recordkeeping
requirements.

    Dated: March 20, 1998

Janet L. Andersen,
Director, Biopesticides and Pollution Prevention Division, Office of
Pesticides Programs.

Summary of Petition

    Below a summary of the pesticide petition is printed. The summary
of the petition was prepared by the petitioner. This petition summary
announces the availability of a description of the analytical methods
available to EPA for the detection and measurement of the pesticide
chemical residues or an explanation of why no such method is needed.

Monsanto Company

PP 7F4822

    1. Plant-pesticide uses. Cotton, Gossypium hirsutum, has been
genetically engineered to be resistant to selected insect pests of the
taxonomic order Lepidoptera. Insect protection was accomplished by the
insertion of the cryIIA gene from Bacillus thuringiensis subsp.
kurstaki (B.t.k.) which encodes for the production of a protein
specifically insecticidal to Lepidopteran larvae in cotton but safe to
nontarget organisms such as mammals, birds, fish and beneficial
insects. Larvae of Lepidopteran pests are the most important insect
pests impacting successful cotton production and numerous chemical
insecticide treatments are typically applied for their control. The
production of cotton varieties containing the CryIIA gene from B.t.k.
is expected to significantly reduce chemical insecticide use in cotton
and; therefore, provide a major benefit to cotton growers and the
environment.
    2. Safety. The CryIIA protein produced in BollgardTM
Cotton is >99.9% identical to the protein produced by the B.t.k. HD-1
bacterial strain found in nature and in commercial B.t.k. formulations
registered with the EPA. These microbial B.t.k. formulations have been
commercially available for the last 30 years. This strain controls
insect pests by the production of crystalline insecticidal proteins
known as delta-endotoxins. To be active against the target insect, the
protein must be ingested. In the insect gut, the protein binds to
specific receptors on the insect mid-gut, inserts into the membrane and
forms ion-specific pores. These events disrupt the digestive processes
and cause the death of the insect.
    There are no receptors for the protein delta-endotoxins of B.
thuringiensis subspecies on the surface of mammalian intestinal cells;
therefore, humans are not susceptible to these proteins. This has been
confirmed in numerous safety studies carried out in laboratory animals
which are traditionally experimental surrogates for humans. The results
of some of these studies have been published in scientific reviews
(Ignoffo,1973; Shadduck et al., 1983; Siegel and Shadduck, 1990).
Results of unpublished safety studies generated by registrants of B.
thuringiensis commercial preparations have also been summarized in a
recently issued EPA Registration Standard for Bt Formulations (EPA,
1988). In published reviews and the EPA document, studies are
referenced in which large doses (5,000 mg/kg) of B. thuringiensis
formulations were administered as single or multiple oral doses (up to
2 years) to different laboratory animals, with no adverse effects.
    Avian and aquatic organisms have also been fed B. thuringiensis
formulations, with no adverse effects. A typical formulation is
composed of Bt spores and Bt protein endotoxin, the latter compromising
up to one-third of the weight of the spores. While target insects are
susceptible to oral doses of B.t.k. proteins, there was no evidence of
any toxic effects observed in non-target laboratory mammals, fish or
birds given the equivalent of up to 106 g of protein per
gram of body weight. No deleterious effects were observed on non-target
insects at doses over 100 fold higher than needed to control target
insects (EPA 1988).
    In addition to the lack of receptors for the B.t.k. proteins, the
absence of adverse effects in non-target animals is further supported
by the poor solubility and stability of the B.t.k. proteins in the acid
milieu of the stomach. The acid conditions in the stomach and the
presence of bile acids denature the B.t.k. proteins facilitating their
rapid degradation by pepsin. In vitro enzymatically activated delta-
endotoxins are also non-toxic when administered orally to laboratory
animals (Nishitsutsuji-Uwo et al. 1980). Even if activated B.t.k.
protein toxins could enter the mammalian gastrointestinal tract, there
are no receptors on the surface of gastrointestinal tissues to permit
binding of the protein toxin to the cell surface. These scientific
considerations are experientially support by the history of completely
safe use of B. thuringiensis preparations. Based on the available
scientific data, EPA and other regulatory scientists worldwide have
determined that use of registered B. thuringiensis products pose no
risks to human health or non-target organisms.
    Monsanto Company has also submitted several toxicology studies in
support of the CryIIA protein as a plant pesticide. According to
Monsanto Company, there is no acute toxicity of the CryIIA protein. In
addition, the CryIIA protein is also produced at low levels by Bollgard
cotton plants and is contained within the cells of the cotton plant.
Consequently, there would be negligible exposure to the protein from
handling cottonseed, leaf tissue or lint at planting, during growth, or
at harvest. In addition, there would be no potential hazard during
storage, transportation, or disposal of Bollgard cottonseed as the
protein cannot drift or volatilize from the plant and its bioactivity
is rapidly lost upon decomposition of the plant tissue.
    The following mammalian toxicity studies have been conducted to
support this exemption from the requirement of a tolerance:
    i. A mouse acute oral gavage study in which the No-Observed-Effect-
Level (NOEL) for toxicity of the CryIIA protein administered as a
single dose was considered to be 4,000 mg/kg (the highest tested dose).
    ii. In vitro digestive fate of the CryIIA protein in simulated
gastric and intestinal fluids. The results of this study established
that the CryIIA protein and its associated functional activity will be
efficiently degraded upon exposure to gastric and intestinal fluids in
the mammalian digestive tract. A lack of stability to digestion is a
characteristic of proteins which are non-allergens.
    iii. Amino acid sequence homology assessment of the CryIIA protein
to known allergens and toxins. The results of this analysis establish
that the CryIIA protein expressed in Bollgard cotton shares no
significant sequence similarity with known toxins, allergens or gliadin
proteins. In addition, the CryIIA protein

[[Page 17176]]

appears to contain no sequences relevant to allergy or coeliac disease.
    3. Threshold effects-- i. Acute toxicity. Based on the available
acute toxicity data for the CryIIA protein and on the safe use of
microbial Bacillus thuringiensis foliar formulations containing the
same protein and registered with the EPA and used commercially for 30
years, no acute dietary risks are posed.
    ii. Chronic effects. The CryIIA protein is degraded upon exposure
to gastric and intestinal fluids in the mammalian digestive tract.
Consequently, no chronic effects are expected. In addition, in
published reviews and the EPA Registration Standard for Bt Formulations
(EPA, 1988) studies are referenced where large doses (5,000 mg/kg) of
B. thuringiensis formulations were administered as single or multiple
oral doses (up to 2 years) to different laboratory animals, with no
adverse effects.
    4. Non-threshold effects. Carcinogenicity: Proteins are not
considered to be carcinogenic (Pareza and Foster, 1983) and
consequently, there is no carcinogenic risk associated with the CryIIA
protein.
    5. Aggregate exposure. Cottonseed meal is not currently used for
human consumption in the United States (Morgan, 1990; Cottonseed Oil,
1990). The presence of gossypol and cyclopropenoid fatty acids in
cottonseed also limits its use as a protein supplement in animal feed
except for cattle, which are unaffected by these components.
Inactivation or removal of these components during processing, which
entails heating and chemical treatment, enables the use of some
cottonseed meal for catfish, poultry and swine. However, as the CryIIA
protein is heat labile, the biological activity of the protein is
expected to be lost upon processing as demonstrated by Sims and
Berberich with other B.t. proteins (1996).
    Refined cottonseed oil and cottonseed linters (the fiber remaining
after ginning seed cotton) are also highly processed and are the only
cotton products consumed as food by humans. Cottonseed oil is typically
removed from the meal by direct solvent extraction with hexane and is
further processed and refined by exposure to extreme heat and alkaline
pH. Processed cottonseed oil contains no detectable protein (Fuchs,
1994; Fuchs et. al., 1993). Cotton linters are essentially comprised
only of cellulose (>99.9%) and Sims et. al. (1996) have demonstrated
that processed linters, which also undergo exposure to temperatures
exceeding 100 deg.C and alkaline treatment do not contain detectable
levels of transgenic proteins such as CryIIA.
    Based on these results, aggregate exposure to the CryIIA protein
through ingestion of cottonseed oil and linters derived from bollgard
cotton would be negligible.
    6. Determination of safety for U.S. population. The toxicity data
support an exemption from the requirement of a tolerance for the CryIIA
protein expressed in Bollgard cotton indicate that there would be no
risk from exposure to the CryIIA protein by the overall U.S.
population. In addition, the CryIIA protein expressed in Bollgard is
more than 99.9% identical to the natural protein, which is component of
microbial Bacillus thuringiensis subsp. kurstaki formulations that have
been registered with the EPA and available commercially for the last 30
years. The EPA and other regulatory scientists worldwide have
determined that use of registered B. thuringiensis products pose no
significant risks to human health or non-target organisms (EPA, 1988).
    7. Determination of safety for infants and children. Monsanto
considers the acute toxicity data, the rapid degradation of the CryIIA
protein in the mammalian digestive system, the lack of homology to
known proteinaceous allergens or toxins and a 30 year history of safe
use of microbial B. thuringiensis  containing the near identical CryIIA
protein as ample evidence to support the safety of this protein to
neonatal infants, infants and children.
    8. Estrogenic effects Not applicable. Proteins are not capable of
direct estrogenic activity as they are incapable of binding to an
estrogen receptor.
    9. Chemical residue. Not applicable. In the United States, only
refined cottonseed oil and cottonseed linters (the fiber remaining
after ginning seed cotton), which are highly processed, are the only
cotton products consumed as food by humans. Cottonseed oil is typically
removed from the meal by direct solvent extraction with hexane and is
further processed and refined by exposure to extreme heat and alkaline
pH. Processed cottonseed oil contains no detectable protein (Fuchs,
1994; Fuchs et. al., 1993). Cotton linters are essentially comprised
only of cellulose (>99.9%) and Sims et. al. (1996) have demonstrated
that processed linters, which also undergo exposure to temperatures
exceeding 100 deg.C and alkaline treatment do not contain detectable
levels of transgenic proteins such as CryIIA.
    10. Environmental fate. The CryIIA protein expressed in Bollgard
cotton plant tissue was evaluated over 120 d in both a laboratory
microcosm and under field conditions. DT50 values were 15.5
d and 31.7 d for the laboratory and field respectively. These results
demonstrate that CryIIA protein, as a component of post-harvest
Bollgard cotton plants, will dissipate when cultivated into soil.

Literature Cited

    1. Cottonseed Oil. 1990. eds. L.A. Jones and C.C. King. National
Cottonseed Products Association, Inc. and The Cotton Foundation,
Memphis.
    2. EPA, 1988. Guidance for the Reregistration of Pesticide Products
Containing Bacillus thuringiensis as the Active Ingredient. NTIS PB 89-
164198.
    3. Fuchs, R.L., Berberich, S.A. and Serdy, F.S. 1993. Safety
evaluation of genetically engineered plants and plant products: insect-
resistant cotton. In "Biotechnology and Safety Assessment." J.A.
Thomas and L.A. Myers, eds. Raven Press Ltd., New York, pp. 199-212.
    4. Fuchs, R.L., 1994. Gene Expression and Compositional Analysis
from Field-Grown Insect Resistant Cotton Tissues, Study Number 92-01-
36-07, an unpublished study conducted by Monsanto Company. EPA MRID#
43168701
    5. Ignoffo, C.M. 1973. Effects of Entomopathogens on Vertebrates.
Ann. N.Y. Acad. Sci. 217:144-172.
    6. Morgan, S.E. 1990. Gossypol Residues in Organ Meats vs
Thresholds of Toxicity. Vet. Hum. Toxicol. 32S:76.
    7. Nishitsutsuji-Uwo and Yasuhisa Endo et. al. 1980. Mode of Action
of Bacillus thuringiensis -Endotoxin: Effect on TN-368 Cells. Appl.
Ent. Zool. 15:133-139.
    8. Pareza, M.W. and Foster, E.M. 1983. Determining the stability of
enzymes used in food processing. J. Food. Prot. 46:453-468.
    9. Siegel and Shadduck, 1989, Safety of Microbial Insecticides to
Vertebrates and Humans, In Safety of Microbial Insecticides. CRC Press,
Inc., FL. pp 101-113.
    10. Sims, S.R. and Berberich, S.A. 1996. Bacillus thuringiensis
CryIA protein levels in raw and processed cottonseed of transgenic
cotton: determination using insect bioassay and ELISA. J. Econ.
Entomol. 89:247-251.
    11. Sims, S.R., Berberich, S.A., Nida, D.L., Segalini, L.L., Leach,
J.N., Ebert, C.C. and Fuchs, R.L. 1996. Analysis of expressed proteins
in fiber fractions from insect-protected and glyphosate-tolerant cotton
varieties. Crop Physiol. Metabol.36:1212-1216.

[FR Doc. 98-8659 FIled 4-7-98; 8:45 a.m.]
BILLING CODE 6560-50-F