Document ID: EPA-HQ-OPP-2010-0944-0002
Agency: epa
Document Type: Supporting & Related Material
Title: 
Posted Date: 2011-02-02T05:00Z

EPA BIOPESTICIDES AND POLLUTION PREVENTION DIVISION

NOTICE OF FILING TEMPLATE FOR PESTICIDE PETITIONS PUBLISHED IN THE

FEDERAL REGISTER 

EPA Biopesticides and Pollution Prevention Division contact: Susanne
Cerrelli, 703-308-8077 

EPA has received a pesticide petition [insert petition number] from
Certis USA LLC, 9145 Guilford Road, Suite 175, Columbia, MD 21046,
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 to
establish an exemption from the requirement of a tolerance for residues
of the microbial pesticide Bacillus subtilis var. amyloliquefaciens
strain D747 in or on all agricultural commodities. 

Pursuant to section 408(d)(2)(A)(i) of the FFDCA, as amended, Certis USA
LLC has submitted the following summary of information, data, and
arguments in support of their pesticide petition. This summary was
prepared by Certis USA 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. Certis USA LLC Petition Summary 

0F7760

A. Product name and Proposed Use Practices 

Bacillus subtilis var. amyloliquefaciens strain D747, microbial pest
control product, is a broad-spectrum preventative biofungicide for
control or suppression of fungal and bacterial plant diseases.  The
product contains a naturally occurring strain of the beneficial
rhizobacterium Bacillus subtilis var. amyloliquefaciens, which colonizes
leaves, roots, and other plant surfaces, killing pathogenic organisms
which disrupt pathogen cell wall production.  Bacillus subtilis var.
amyloliquefaciens provides efficacy by demonstrating antagonistic
action, competing for nutrient content of the plant body and habitat;
thus, .preventing establishment of disease-causing fungi and bacteria.

The proposed uses for Bacillus subtilis var. amyloliquefaciens strain
D747 include application alone or in combination and/or rotation with
chemical fungicides as a tool for integrated disease management in
agricultural crops, ornamental and nursery plants, and turfgrass. 
Bacillus subtilis var. amyloliquefaciens strain D747 offers a valuable
tool for management of resistance to chemical fungicides through its
multiple and unique modes of action.

B. Product Identity/Chemistry 

Identity of the pesticide and corresponding residues. 

Bacillus subtilis var. amyloliquefaciens strain D747 is a rod shaped,  
HYPERLINK "http://en.wikipedia.org/wiki/Gram-positive" \o
"Gram-positive"  Gram-positive ,   HYPERLINK
"http://en.wikipedia.org/wiki/Catalase" \o "Catalase"  catalase
-positive, mobile   HYPERLINK "http://en.wikipedia.org/wiki/Bacterium"
\o "Bacterium"  bacterium  commonly found in soil.  Bacillus subtilis
var. amyloliquefaciens strain D747 has the ability to form a tough,
protective   HYPERLINK "http://en.wikipedia.org/wiki/Endospore" \o
"Endospore"  endospore , allowing the organism to tolerate extreme
environmental conditions. Bacillus subtilis var. amyloliquefaciens
strain D747 is not known as an aquatic bacterium, and is not expected to
proliferate in aquatic environments.

The culture is deposited in the International Patent Organism
Depository, National Institute of Advanced Industrial Science and
Technology, as Bacillus sp. D747.  The deposit number is FERM BP-8234.

Bacillus subtilis var.amyloliquefaciensis strain D747 is not considered
to be a human pathogen.  Bacillus subtilis species, including Bacillus
subtilis var. amyloliquefaciens D747, produce proteolytic enzymes and
are widely used as additives in   HYPERLINK
"http://en.wikipedia.org/wiki/Laundry_detergent" \o "Laundry detergent" 
laundry detergents .  Bacillus subtilis var.amyloliquefaciens D747 is
used as a   HYPERLINK "http://en.wikipedia.org/wiki/Soil_inoculant" \o
"Soil inoculant"  soil inoculant  in   HYPERLINK
"http://en.wikipedia.org/wiki/Horticulture" \o "Horticulture" 
horticulture  and   HYPERLINK "http://en.wikipedia.org/wiki/Agriculture"
\o "Agriculture"  agriculture , and can be used as a plant growth
promoter.  

Magnitude of residue at the time of harvest and method used to determine
the residue. 

Bacillus subtilis var. amyloliquefaciens strain D747 is ubiquitous in
the environment, especially in soils and agricultural environments. all
over the world. As a result, dietary exposure to background levels of
the naturally occurring microbe likely is already occurring and likely
will continue to occur. Because of its ubiquitous presence in the
environment, no increase in exposure to Bacillus subtilis var.
amyloliquefaciens strain D747 resulting from the existing and proposed
pesticidal uses when compared to existing exposure to background levels
of Bacillus subtilis var.amyloliquefaciens strain D747 is expected to
occur. 

Any potentially occurring residual deposits on crops will not harm
humans because the strain shows no toxicity, infectivity, or
pathogenicity in appropriate tests.  

3. A statement of why an analytical method for detecting and measuring
the levels of the pesticide residue are not needed. 

The acute toxicity and studies discussed in Section C of this document
are sufficient to show that there are no foreseeable human or domestic
health hazards likely to arise from the use of the product as proposed. 
Certis USA is requesting an exemption from the requirement for a
tolerance for any residues remaining in/on all agricultural commodities.
 Since enforcement of residue levels would not be needed, an enforcement
analytical method is not required.

C. Mammalian Toxicological Profile 

Toxicological data on the active ingredient are submitted to support the
request for exemption from the requirement of a tolerance for residues
of Bacillus subtilis var. amyloliquefaciens strain D747 when used as a
foliar spray in/on all raw agricultural commodities. The submitted
studies on the active ingredient include the following: 

An acute oral toxicity/ pathogenicity study performed in rats
demonstrated the lack of mammalian toxicity at a high level of exposure
to Bacillus subtilis.var. amyloliquefaciens strain D747. In this study,
Bacillus subtilis var.amylokiquefaciens D747 was not toxic, infective
nor pathogenic to rats given an oral dose of 1 x 108 colony forming
units (CFU) per animal.  As a result no mortality or abnormalities were
demonstrated in any clinical signs, body weight changes, or at necropsy.
Test microbes were not detected in kidney, brain, liver, lung, spleen,
small intestine, large intestine, lymph or blood.  Excreted test
material was not detected in the rat feces at 14 days after
administration. 

An acute pulmonary toxicity/pathogenicity study in rats demonstrated
that Bacillus subtilus var. amyloliquefaciens strain D747 was neither
toxic, pathogenic nor infective (residual microbial viability) to rats
dosed intra-tracheally with 1.0 x 108 CFU of the test material. 
Bacillus subtilis var. amyloliquefaciens strain D747 demonstrated a
gradual decrease in microbes in the nasal cavity, trachea, and lungs
over time.  The test microbes were not detected in other organs,
including the kidney, brain, liver, spleen and blood, during the test
period.

An acute (intravenous) injection toxicity/pathogenicity study in rats
demonstrated no mortality or abnormalities in the clinical observations,
body weight, and at necropsy.  Animals were dosed with 1x108 CFU/animal.
Although the microbe was detected primarily in kidney, liver, spleen,
and blood initially after administration, the test microbes could not be
detected in the blood from 14 days after administration,  The test
microbe did not completely disappear from the liver and spleen during
the observation period of 60 days, but test microbes decreased
significantly within a short period after administration; therefore,
proliferation of the test microbes were not observed in any organ during
the test period.  As a result Bacillus subtilis var. amyloliquefaciens
strain D747 was concluded to have no infectiousness, pathogenicity,
toxicity, or residual microbial viability in the body.

An acute oral toxicity study demonstrated no acute toxicity potential of
Bacillus subtilis var. amyloliquefaciens strain D747 enduse   product,
(CX-9030), when administered to female albino rats.  The acute oral LD50
is greater than 5000 mg/kg in females.

An acute dermal toxicity study in rabbits demonstrated that Bacillus
subtilis var. amyloliquefaciens strain D747 enduse product, (CX-9030),
was not toxic to rabbits when a single 5 x 1010 dose was administered
dermally. The acute dermal toxicity LD50 is greater than 5050 mg/kg in
males and females.

A primary eye irritation study in rabbits demonstrated that Bacillus
subtilis var. amyloliquefaciens strain D747 enduse product (CX-9030),
produced no positive effect, (cornea opacity, Iritis, conjunctival
redness or chemosis), in any eyes at 48 hours after treatment, when a
single 0.1 gram ocular dose was administered.  The test substance was
rated mildly irritating and assigned Toxicity Category III.

An acute inhalation toxicity study in rats demonstrated that Bacillus
subtilis var. amyloliquefaciens strain D747 enduse product (CX-9030),
produced no mortality during the study. The acute inhalation LC50 is
greater than 2.18 mg/L in males and females.

An acute dermal irritation study in rabbits demonstrated no signs of
irritation at any time throughout the duration of the study.  Bacillus
subtilis var. amyloliquefaciens D747 enduse product (CX-9030) was
determined to be non-irritating and assigned Toxicity Category IV.

D. Aggregate Exposure 

1. Dietary exposure.   

food

Bacillus subtilis var. amyloliquefaciens strain D747 is ubiquitous in
the environment, especially in soils and agricultural environments. As a
result, dietary exposure to background levels of the naturally occurring
microbe likely is already occurring and likely will continue to occur.
Because of its ubiquitous presence in the environment, no increase in
exposure to Bacillus subtilis var. amyloliquefaciens strain D747,
resulting from the existing and proposed pesticidal uses is expected
when compared to existing exposure to background levels of Bacillus
subtilis var. amyloliquefaciens strain D747.

As discussed above, dietary exposure to the naturally occurring microbe
likely is already occurring and likely will continue to occur. Notably,
similar Bacillus subtilis strains are used internationally in the
production of food grade products and in fermented foods in Japan and
Thailand. Reports in the literature implicating Bacillus subtilis sp. in
food-borne illness do not describe any pathogen or toxin production, but
rather simple spoilage from Bacillus subtilis growth in dough. Such
low-quality dough would not be suitable for bread production by
commercial bakeries and so the Agency considers this particular food
exposure scenario to be unlikely and the risk to be negligible. The risk
posed to adults, infants and children from food-related exposures to
Bacillus subtilis var. amyloliquefaciens strain D747 is minimal due to
the demonstrated lack of acute oral toxicity/pathogenicity associated
with the microbial pesticide. Based on the submitted data, it is
concluded that there are no dietary risks that exceed the Agency’s
level of concern. 

  

 Drinking water. 

Bacillus subtilis var. amyloliquefaciens strain D747 is ubiquitous in
the environment, and exposure to the microbe through drinking water may
already be occurring and likely will continue to occur. While the
proposed and existing use sites do not include direct application to
aquatic environments, the intended use of Bacillus subtilis treatment of
growing crops or seed for the control of plant disease might result in
the potential for pesticide spray drift or runoff that might reach
surface or ground waters. However, the potential for human exposure to
Bacillus subtilis var. amyloliquefaciens strain D747 residues in
drinking water is very low.  Bacillus subtilis species are not
recognized aquatic bacteria; thus, are not expected to grow in aquatic
environments. Also, Municipal drinking water treatment processes and
deep water wells would both further reduce any such residues. More
importantly, even if oral exposure to this ubiquitous microbe should
occur through drinking water, due to its demonstrated lack of acute oral
toxicity/ pathogenicity, it is concluded that there is a reasonable
certainty that no harm will result from such exposure. 

2. Non-dietary exposure.

The pesticide uses of Bacillus subtilis var. amyloliquefaciens strain
D747, both those currently allowed and the additional proposed uses  are
limited to commercial agriculture naturally occurring and ubiquitous in
the environment, the potential for non-dietary, non-occupational
exposure to its residues for the general population, including infants
and children, is likely since populations have probably been previously
exposed (and likely will continue to be exposed) to background levels of
the microbe. However, neither such common human exposures to Bacillus
subtilis var. amyloliquefaciens strain D747 naturally present in soils,
waters and plants, nor exposures associated with similar Bacillus
subtilis strains used internationally in producing food-grade products
and fermented foods, have resulted in reports of infections in humans
(which consistently are reported only in otherwise-compromised
individuals), those reports are most notable for their rare and
exceptional nature. 

EPA’s previous evaluation of the high-dose Tier I acute toxicity and
pathogenicity tests resulted in the assignment of Toxicity Category IV
(least toxic), and determinations of not infective and not pathogenic,
for all exposure routes. No toxicological end points of concern were
identified. There are no dietary endpoints that exceed EPA’s Level of
Concern (LOC). Therefore, it is concluded  that any additional exposure
to the microbe resulting from residues attributable to Bacillus subtilis
var. amyloliquefaciens strain pesticide use will not result in
additional aggregate non-occupational risk from dermal and inhalation
exposures. This conclusion, based solely on non-occupational exposures,
is consistent with previous EPA determinations for Bacillus subtilis
strains that no occupational risks exceed the Agency’s LOC, meaning
that even regular occupational exposures associated with this active
ingredient pose negligible risk.

E. Cumulative Exposure 

No mechanism of toxicity in mammals has been identified for Bacillus
subtilis.var. amyloliquefaciens strain D747.  Therefore, no cumulative
effect with other related organisms is anticipated. Because the
available data demonstrate a lack of toxicity/pathogenicity potential
for the active ingredient, adverse dietary effects are unlikely.

F. Determination of Safety for U.S. population, Infants and Children 

Based on the acute toxicity information discussed in 3.C., it is
concluded that there is a reasonable certainty that no harm will result
to the United States population, including infants and children, from
aggregate exposure to residues of Bacillus subtilis var.
amyloliquefaciens strain D747.  This includes all anticipated dietary
exposures and all other exposures for which there is reliable
information. This conclusion is based on the data available on Bacillus
subtilis species, including Bacillus subtilis var. amyloliquefaciens
strain D747, which demonstrate a lack of toxicity/ pathogenicity
potential. Thus, there are no threshold effects of concern and, as a
result, it is concluded that the additional tenfold margin of safety for
infants and children is unnecessary in this instance. Further, the need
to consider consumption patterns, special susceptibility, and cumulative
effects does not arise when dealing with pesticides with no demonstrated
significant adverse effects. 

G. Effects on the Immune and Endocrine Systems 

Bacillus subtilis var. amyloliquefaciens strain D747 is a ubiquitous
organism in the environment that is non-toxic to mammals. To date, there
is no evidence to suggest that Bacillus subtilis var. amyloliquefaciens
strain D747 affects the immune system, functions in a manner similar to
any known hormone, or that it acts as an endocrine disruptor. Indeed,
the submitted toxicity/ pathogenicity studies in rodents indicate that,
following several routes of exposure, the immune system is intact and
able to process and clear the active ingredient. Therefore, it is
unlikely that this organism will have estrogenic or endocrine effects. 

H. Existing Tolerances 

There are no existing tolerance exemptions or tolerances established for
Bacillus subtilis var. amyloliquefaciens strain D747

I. International Tolerances