BIOPESTICIDES REGISTRATION ACTION DOCUMENT
Candida oleophila Strain O
PC Code: 021010
U.S. Environmental Protection Agency
Office of Pesticide Programs
Biopesticides and Pollution Prevention Division
Last Updated - July 15, 2009
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Candida oleophila Strain O
Biopesticides Registration Action Document (BRAD) July 2009
Table of Contents
I. EXECUTIVE SUMMARY 4
II. ACTIVE INGREDIENT OVERVIEW 6
III. REGULATORY BACKGROUND 6
IV. RISK ASSESSMENT 7
A. PRODUCT ANALYSIS ASSESSMENT 8
1. Product Chemistry and Composition 8
2. Analysis and Certified Limits 9
3. Physical and Chemical Characteristics 9
B. HUMAN HEALTH ASSESSMENT 9
1. Toxicology 9
2. Dietary Exposure and Risk Characterization 13
3. Drinking Water Exposure and Risk Characterization 13
4. Acute and Chronic Dietary Exposure and Risks for Sensitive
Subpopulations, Particularly Infants and Children 14
5. Occupational, Residential, School, and Day care Exposure
and Risk Characterization 14
6. Aggregate Exposure from Multiple Routes Including
Dermal, Oral, and Inhalation 15
7. Cumulative Effects 16
8. Risk Characterization 16
C. ENVIRONMENTAL ASSESSMENT 16
1. Ecological Hazards 16
2. Environmental Fate and Ground Water Data 17
3. Ecological Exposure and Risk Characterization 17
4. Endangered Species Assessment 17
V. ENVIRONMENTAL JUSTICE 17
VI. RISK MANAGEMENT AND REGISTRATION DECISIONS 18
A. DETERMINATION OF ELIGIBILITY 18
B. REGISTRATION REVIEW 19
C. REGULATORY DECISION 19
D. LABELING 20
VII. ACTIONS REQUIRED BY THE REGISTRANT 20
A. REPORTING OF ADVERSE EFFECTS 20
B. REPORTING OF HYPERSENSITIVITY INCIDENTS 20
VIII. GLOSSARY OF ACRONYMS AND ABBREVIATIONS 21
IX. BIBLIOGRAPHY 22
A. STUDIES SUBMITTED IN SUPPORT OF THIS REGISTRATION 22
B. EPA RISK ASSESSMENT MEMORANDUMS 23
C. OTHER REFERENCES 24
APPENDIX A 25
APPENDIX B 31
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Candida oleophila Strain O
Biopesticides Registration Action Document (BRAD) July 2009
BIOPESTICIDES REGISTRATION ACTION DOCUMENT TEAM
Office of Pesticide Programs
Biopesticides and Pollution Prevention Division
Microbial Pesticides Branch
Science Reviews
Joel Gagliardi, Ph.D. Product Analysis, Human Health
John Kough, Ph.D. Product Analysis, Human Health
Gail Tomimatsu, Ph.D. Environmental Effects
Regulations
Sheryl Reilly, Ph.D. Chief, Microbial Pesticides Branch
Jeannine Kausch, M.S.E.L. Regulatory Action Leader
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Candida oleophila Strain O
Biopesticides Registration Action Document (BRAD) July 2009
I. EXECUTIVE SUMMARY
Candida oleophila Strain O is a single-celled yeast found naturally on plant tissues
(fruits, flowers, and wood) and in water. It was originally isolated from golden delicious
apples and is intended for use as an antagonist to control the fungal pathogens, gray mold
(Botrytis cinered) and blue mold (Penicillium expansum), which cause post-harvest decay
on apples and pears. The mode of action for Candida oleophila Strain O is primarily
through competition for nutrients and pre-colonization of plant wound sites (Jijakli etal.
1993), although information provided to the Agency suggests that production of beta-1,3-
glucanases (i.e., hydrolytic enzymes that can degrade fungal phytopathogen cell walls)
may also contribute to its antagonistic activity.
To decide whether to grant a unconditional registration under Section 3(c)(5) of the
Federal Insecticide, Fungicide, and Rodenticide Act (FIFRA) to NEXY, a proposed end-
use product containing Candida oleophila Strain O as an active ingredient at 57.0% by
weight, the Biopesticides and Pollution Prevention Division (BPPD) reviewed microbial
pesticide product chemistry and composition, analysis and certified limits, physical and
chemical characteristics, mammalian and non-target organism toxicology, and
environmental fate data [40 Code of Federal Regulations (CFR) §§ 158.2120, 158.2140,
and 158.2150]. It was determined that the data/information submitted adequately satisfy
current data requirements.
Adequate mammalian toxicology data/information were submitted to support the
registration of NEXY. Acceptable acute toxicity guideline studies were submitted, and
data waivers were granted by the Agency to fulfill the remaining Tier I acute toxicity data
requirements based on the lack of toxicity, infectivity, and/or pathogenicity of Candida
oleophila Strain O, as well as the nature of the inert ingredients in the NEXY
formulation. Additionally, two mutagenicity studies not required for the registration of
this active ingredient were voluntary submitted by the registrant and indicated that
Candida oleophila Strain O did not have mutagenic potential.
Although dietary exposure to Candida oleophila Strain O may occur through
consumption of treated apples and pears, the following factors were considered by the
Agency in making its decision to register this microbial pesticide: (1) lack of acute oral
toxicity/pathogenicity, (2) inability of Candida oleophila Strain O to grow at mammalian
body temperatures, (3) absence of clinical reports of infections despite the common
occurrence of Candida oleophila on various food commodities consumed by the United
States population, (4) standard practices for processing fruit (i.e., washing, peeling, and
cooking), which would reduce residues of Candida oleophila Strain O several orders of
magnitude below the dose tested in the acute oral toxicity/pathogenicity study, and (5)
prior establishment of an exemption from the requirement of a tolerance for residues of a
similar strain of yeast, Candida oleophila isolate 1-182, when used as a post-harvest
biological fungicide in or on all raw agricultural commodities (40 CFR § 180.1144).
Furthermore, drinking water and non-dietary, non-occupational exposures to Candida
oleophila Strain O are unlikely to occur because of the proposed use pattern, use sites,
and application methods specified on NEXY's product label. Cumulative effects are not
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Candida oleophila Strain O
Biopesticides Registration Action Document (BRAD) July 2009
anticipated because Candida oleophila Strain O has no demonstrated toxicity. The
Environmental Protection Agency (EPA) concluded that the data submitted for the
NEXY registration meet the safety requirements of the Food Quality Protection Act
(FQPA) of 1996, and support an exemption from the requirement of a tolerance for the
residues of Candida oleophila Strain O on apples and pears. If the microbial active
ingredient is used in accordance with the product label, there is a reasonable certainty that
no harm will result from aggregate exposure to the United States population, including
infants and children, to the residues of Candida oleophila Strain O. As a result of this
determination, EPA established a permanent exemption from the requirement of a
tolerance for residues of the microbial pesticide, Candida oleophila Strain O, on apples
and pears when applied/used as a post-harvest biofungicide (40 CFR § 180.1289).
Personal protective equipment (PPE) is required for mixers, loaders, and applicators that
may be exposed for prolonged periods to Candida oleophila Strain O due to their
occupation. Mixers and loaders must wear a dust/mist filtering respirator meeting
National Institute for Occupational Safety and Health (NIOSH) standards of at least N-
95, R-95, or P-95 because of an incident in which three workers reported clinical
symptoms of a respiratory reaction after being exposed to large amounts of Candida
oleophila Strain O and wearing no PPE. No adverse dermal effects have been reported.
Nevertheless, the Agency believes that the potential for dermal hypersensitivity incidents
is best mitigated by label requirements that a long-sleeved shirt, long pants, socks, shoes,
and waterproof gloves be worn when mixing, loading, or applying the product, or when
handling crates containing treated apples and pears. Protective eyewear is also required
when mixing, loading, or applying the product or when handling crates containing treated
apples and pears, based upon the results of the submitted acute eye irritation study.
Adequate non-target toxicology data/information are available to support registration of
NEXY, which contains Candida oleophila Strain O, for intended indoor uses on pome
fruits (pears, apples). Because of these intended uses, direct exposures to most terrestrial
and all aquatic environments are not expected, and therefore, all non-target toxicology
data requirements for Candida oleophila Strain O were satisfied with appropriate data
waiver requests and rationales. No risks to non-target organisms (to include federally
listed endangered and threatened species) or to the environment are anticipated as a result
of the intended indoor uses of Candida oleophila Strain O. If future uses include outdoor
uses where exposures of non-target organisms would be increased, additional data and/or
information must be submitted to and reviewed by the EPA.
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Candida oleophila Strain O
Biopesticides Registration Action Document (BRAD) July 2009
II. ACTIVE INGREDIENT OVERVIEW
Biological Name: Candida oleophila Strain O
Culture Deposit: Belgian Coordinated Collections of Microorganisms
(BCCM)/Mycotheque de 1'Universite Catholique de
Louvain (MUCL) in Louvain-la-Neuve, Belgium
under Accession Number MUCL 40564
Office of Pesticide
Programs (OPP)
Chemical Code: 021010
Type of Pesticide: Microbial Pesticide - Post-Harvest Biofungicide
See Appendix B for specific information (i.e.,
use sites, application rate, method of application,
formulation type, and target pests) regarding
the end-use product, NEXY, containing this active
ingredient.
III. REGULATORY BACKGROUND
On December 28, 2007, SynTech Global, LLC (address: P.O. Box 640, Hockessin,
Delaware 19707), acting as the United States authorized agent for BioNext sprl (address:
Passage des deportes, 2, B-5030 Gembloux, Belgium), submitted an application to
register NEXY (EPA File Symbol 84863-R) under Section 3 of the Federal Insecticide,
Fungicide, and Rodenticide Act. On March 28, 2008, the EPA announced receipt of this
application to register a pesticide product containing a new active ingredient [73 Federal
Register (FR) 16676] and opened a 60-day public comment period.
One comment was received in response to the notice of receipt. A private citizen
expressed opposition to Candida oleophila Strain O's introduction into the United States
in light of the "thousands of chemicals already out there" and implied a concern about the
effects of "toxic chemicals" on human health and the environment. Pursuant to its
authority under FIFRA, the Agency conducted a rigorous assessment of Candida
oleophila Strain (as described throughout this document) and concluded that it is not
expected to cause any unreasonable adverse effects to human health or the environment.
Further, Candida oleophila Strain O is not considered a conventional pesticide.
Conventional pesticides generally consist of synthetic materials, may affect abroad
spectrum of non-target organisms, and may be inherently more toxic. Candida oleophila
Strain O, however, is a naturally occurring yeast isolated from golden delicious apples
and found on various food commodities, and has been classified as a microbial pesticide.
Candida oleophila Strain O is intended for use as an antagonist to specifically control the
fungal pathogens, gray mold (Botrytis cinered) and blue mold (Penicillium expansum),
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Candida oleophila Strain O
Biopesticides Registration Action Document (BRAD) July 2009
which cause post-harvest decay on apples and pears. The mode of action for Candida
oleophila Strain O is primarily through competition for nutrients and pre-colonization of
plant wound sites, and use of Candida oleophila Strain O may result in decreased
conventional pesticide applications to apples and pears after harvest.
Concurrent with their registration application, and under the Federal Food, Drug, and
Cosmetic Act (FFDCA), as amended by the Food Quality Protection Act of 1996,
SynTech Global, LLC submitted a petition to establish an exemption from the
requirement of a tolerance for Candida oleophila Strain O [Pesticide Petition (PP)
7F7310]. In the Federal Register dated March 28, 2008 (73 FR 166731 the EPA
announced that SynTech Global, LLC proposed to establish an exemption from the
requirement of a tolerance for residues of the microbial pesticide, Candida oleophila
Strain O, on apples and pears when used as a post-harvest biofungicide and opened a 30-
day public comment period. No comments were received following this publication.
On May 13, 2009, the Agency established a permanent exemption from the requirement
of a tolerance for residues of the microbial pesticide, Candida oleophila Strain O, on
apples and pears when applied/used as a post-harvest biofungicide (40 CFR § 180.1289;
74 FR 22460]. Shortly thereafter and pursuant to FIFRA section 3(c)(5), an unconditional
registration was issued for NEXY on June 1, 2009 (EPA Registration Number 84863-1).
IV. RISK ASSESSMENT
On October 26, 2007, the Agency issued a Final Rule in the Federal Register on the data
requirements to support registration of biochemical and microbial pesticides, and updated
the definitions for biochemical and microbial pesticides (72 FR 61002). The rule became
effective on December 26, 2007. The data and information evaluated for this
Biopesticides Registration Action Document (BRAD) were considered in light of these
requirements.
The classifications that are found for each data submission are assigned by EPA science
reviewers and are an indication of the usefulness of the information contained in the
documents for risk assessment. A rating of "ACCEPTABLE" indicates the study is
scientifically sound and is useful for risk assessment. A "SUPPLEMENTAL" rating
indicates the data provide some information that can be useful for risk assessment. The
studies may have certain aspects determined not to be scientifically acceptable
("SUPPLEMENTAL: UPGRADABLE"). If a study is rated as "SUPPLEMENTAL:
UPGRADABLE," the Environmental Protection Agency always provides an indication
of what is lacking or what can be provided to change the rating to "ACCEPTABLE." If
there is simply a "SUPPLEMENTAL" rating, the reviewer will often state that the study
is not required by the current 40 CFR Part 158. Both "ACCEPTABLE" and
"SUPPLEMENTAL" studies may be used in the risk assessment process as appropriate.
An "UNACCEPTABLE" rating indicates that new data need to be submitted.
For product-specific acute toxicity data requirements, toxicity categories are assigned
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Candida oleophila Strain O
Biopesticides Registration Action Document (BRAD) July 2009
based on the hazard(s) identified from studies and/or information submitted to the
Agency. The product is classified into Toxicity Category I, II, III, or IV, where Toxicity
Category I indicates the highest toxicity and Toxicity Category IV indicates the lowest
toxicity.
A. Product Analysis Assessment (40 CFR § 158.2120)
All product analysis data requirements for Section 3(c)(5) registration of NEXY,
containing Candida oleophila Strain O as an active ingredient, have been satisfied by
either acceptable guideline studies or waiver rationales. For a comprehensive guideline-
by-guideline summary of the product analysis data requirements described in sections
IV(A)(1), IV(A)(2), and IV(A)(3), refer to Table 1 in Appendix A.
1. Product Chemistry and Composition [Office of Prevention,
Pesticides, and Toxic Substances (OPPTS) Guidelines 885.1100, 885.1200,
and 885.1300]
Candida oleophila Strain O is a single-celled yeast [approximately 1 micrometer (um) in
diameter] in the phylum Ascomycota, the class Ascomycetes, and the family
Saccharomycetaceae. Found naturally on plant tissues (fruits, flowers, and wood) and in
water, it was originally isolated from golden delicious apples. Candida oleophila Strain O
reproduces asexually by budding. After 3 days at 25°C, a few cells are spheroidal but
most are ovoidal or long-ovoidal to elongate. The cells occur singly, in pairs, and in
short-branched and unbranched chains. No production of antibiotics or toxic volatile
compounds has been observed. According to reported testing, Candida oleophila Strain
O does not grow above 33 °C, is sensitive to ultraviolet light, and is dependent on a
carbon source for growth (Jijakli et al. 1993). Some species of Candida, namely Candida
albicans and Candida glabrata, have been found as opportunistic pathogens to humans
(mainly the immunosuppressed) and/or animals (Yemma and Berk 1994). Candida
oleophila, however, is an environmental microbe unable to grow at mammalian body
temperatures and is easily distinguished from the Candida species reported as clinical
isolates. More importantly, no pathogenic effects or infections from Candida oleophila
Strain O were seen in the submitted infectivity studies as described in section
IV(B)(l)(a). Candida oleophila has not been reported to produce toxins against plants or
animals, and no production of toxins has been observed under laboratory conditions.
Candida oleophila Strain O is intended for use as an antagonist to control the fungal
pathogens, gray mold (Botrytis cinered) and blue mold (Penicillium expansuni), that
cause post-harvest decay on apples and pears. The mode of action for Candida oleophila
Strain O is primarily through competition for nutrients and pre-colonization of plant
wound sites (Jijakli etal. 1993). Submitted data also suggest that production of beta-1,3-
glucanases (i.e., hydrolytic enzymes that can degrade fungal phytopathogen cell walls)
may contribute to its antagonistic activity.
Submitted data also adequately describe the production process, potential microbial
contaminants, and potential carryover impurities from the inert ingredients; therefore, the
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Biopesticides Registration Action Document (BRAD) July 2009
requirements for manufacturing process and discussion of formation of unintentional
ingredients have been satisfied.
2. Analysis and Certified Limits (OPPTS Guidelines 885.1400 and 885.1500)
Results of a five-batch preliminary analysis were provided and the requirement for
analysis of samples has been satisfied.
The certified limits for the active and inert ingredients exceed the OPPTS Guideline
830.1750 specified ranges, but an acceptable explanation was provided; therefore, the
requirement for certified limits has been satisfied.
3. Physical and Chemical Characteristics (OPPTS Guidelines 830.6302,
830.6303, 830.6304, 830.6313, 830.6317, 830.6319, 830.6320, 830.7000,
830.7100, and 830.7300)
Submitted data adequately describe the physical and chemical characteristics and waiver
rationales provided are acceptable; therefore, the requirements for color, physical state,
odor, stability to normal and elevated temperatures, metals, and metal ions, storage
stability, miscibility, corrosion characteristics, pH, viscosity, and density/relative
density/bulk density (specific gravity) have been satisfied.
B. Human Health Assessment
1. Toxicology
Adequate mammalian toxicology data/information are available to support the
registration of NEXY, which contains Candida oleophila Strain O. All Tier I toxicology
data requirements for Candida oleophila Strain O and its associated end-use product have
been satisfied by guideline studies or waiver rationales. Tier II and Tier III studies were
not required for Candida oleophila Strain O based on the lack of acute
toxicity/pathogenicity in the Tier I studies. Two mutagenicity studies were submitted and
reviewed, although they were not required for Candida oleophila Strain O.
For a comprehensive guideline-by-guideline summary of the toxicology data
requirements and additional studies described in sections IV(B)(l)(a), IV(B)(l)(b), and
IV(B)(l)(c), refer to Table 2 and Table 3 in Appendix A.
a. Acute Toxicity/Pathogenicity - Tier I (40 CFR § 158.2140)
Acute Oral Toxicity andPathogenicity - Rat [OPPTS Guideline 885.3050; Master
Record Identification (MRID) Number (No.) 473138-071: In an acute oral toxicity and
pathogenicity study, groups of rats were given a single oral dose of Candida oleophila
Strain O at a dose of 2.3-3.8 x 108 colony-forming units (cfu)/animal. The animals were
observed for a period of up to 22 days with interim scheduled sacrifices on days 4, 8, and
15. There were no treatment-related clinical signs, necropsy findings, or changes in body
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Biopesticides Registration Action Document (BRAD) July 2009
weight. No test organisms were recovered from the gastrointestinal contents, any organs,
or blood of any animal or feces from treated animals sacrificed on day 22. Based on the
results of this study, Candida oleophila Strain O does not appear to be toxic, infective,
and/or pathogenic in rats. This study was rated "ACCEPTABLE" for risk assessment
purposes.
Acute Pulmonary Toxicity andPathogenicity - Rat (OPPTS Guideline 885.3150; MRID
No. 473138-09): In an acute pulmonary toxicity and pathogenicity study, groups of rats
were exposed by the intratracheal route to Candida oleophila Strain O at a dose of 1.2-
5.2 x 108 cfu/animal. The animals were observed for up to 22 days. There were no test
substance-related clinical signs, necropsy findings, or changes in body weight. Test
organisms were recovered in the lungs from the treated males and females sacrificed one
hour post dosing with clearances by day 4. No test organisms were recovered from the
gastrointestinal contents, organs, or blood of any animal or feces from treated animals
sacrificed on day 22. Based on these results, Candida oleophila Strain O does not appear
to be toxic, infective, and/or pathogenic to rats at 1.2-5.2 x 108 cfu/animal. This study
was rated "ACCEPTABLE" for risk assessment purposes.
Acute Subcutaneous Injection Toxicity andPathogenicity - Rat (OPPTS Guideline
885.3200; MRID No. 473138-08): In an acute subcutaneous injection toxicity and
pathogenicity study, groups of rats were injected subcutaneously with Candida oleophila
Strain O with a dose of 1.1-2.0 x 107 cfu/animal. The animals were observed for up to 22
days. There were no treatment-related clinical signs, necropsy findings, or changes in
body weight. No test organisms were recovered from the gastrointestinal contents,
organs, blood, or the injection site of any animal. Based on the results of this study,
Candida oleophila Strain O does not appear to be toxic, infective, and/or pathogenic in
rats when dosed at 1.1-2.0 x 107 cfu/animal. This study was rated "ACCEPTABLE" for
risk assessment purposes.
Hvversensitivity Incidents (OPPTS Guideline 885.3400; MRID No. 473138-12): During a
pilot-plant production trial of NEXY using fermentation vessels and involving large
amounts of Candida oleophila Strain O, three of six workers (all on the same work team)
not wearing personal protective equipment reported clinical symptoms of a respiratory
reaction. As a result, the product label requires that mixers and loaders wear a dust/mist
filtering respirator meeting NIOSH standards of at least N-95, R-95, or P-95, which will
also be worn by workers in the production facility. Due to the application methods
(dipping and drenching), respiratory exposure to operators during pesticide application is
not anticipated. No adverse dermal effects have been reported by workers, and dermal
sensitization to operators or consumers is not anticipated. Nevertheless, potential for
dermal hypersensitivity incidents has been mitigated by label requirements that a long-
sleeved shirt, long pants, socks, shoes, and waterproof gloves be worn when mixing,
loading, or applying the product or when handling crates containing treated apples and
pears. Any future hypersensitivity incidents must be reported per OPPTS Guideline
885.3400.
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Cell Culture (OPPTS Guideline 885.3500): This study is not required because Candida
oleophila Strain O is not a virus (refer to test note #4 of 40 CFR § 158.2140(d)).
Acute Oral Toxicity (OPPTS Guideline 870.1100; MRID No. 475610-01): Waived based
on the results of MRID No. 473138-07, which showed that Candida oleophila Strain O
was not toxic, infective, and/or pathogenic via oral route of exposure, and the nature of
the inert ingredients in NEXY's formulation. This waiver request was rated
"ACCEPTABLE" and NEXY was classified as TOXICITY CATEGORY IV.
Acute Dermal Toxicity (OPPTS Guideline 870.1200; MRID No. 473227-01): Waived
based on the results of MRID No. 473138-08 (which showed that Candida oleophila
Strain O was not toxic, infective, and/or pathogenic via subcutaneous route of exposure),
MRID No. 473138-11 (which showed that NEXY was not irritating to the skin of rabbits
at a dose of 0.5 gram/animal), and the nature of the inert ingredients in NEXY's
formulation. This waiver request was rated as "ACCEPTABLE" and NEXY was
classified as TOXICITY CATEGORY III.
Acute Inhalation Toxicity (OPPTS Guideline 870.1300; MRID No. 475610-01): Waived
based on the results of MRID No. 473138-09, which showed that Candida oleophila
Strain O was not toxic, infective, and/or pathogenic via pulmonary route of exposure, and
the nature of the inert ingredients in NEXY's formulation. This waiver request was rated
"ACCEPTABLE" and NEXY was classified as TOXICITY CATEGORY III.
Acute Eve Irritation - Rabbit (OPPTS Guideline 870.2400; MRID No. 473138-10): In an
acute eye irritation study, 100 milligrams (mg) of NEXY was instilled into the
conjunctival sac of the right eye of four rabbits. Thirty seconds after treatment, the eye of
the first rabbit was rinsed with saline. Animals were observed for 15 days. Irritation was
scored and classified by the system of Kay and Calandra. No corneal opacity or iritis was
noted on the unwashed eyes of three rabbits. Positive conjunctival irritation was noted on
1 of 3 rabbits 24 hours after test material instillation, with reduction to score 1 by 48
through 72 hours, and with clearance by day 8. In this study, NEXY was minimally
irritating to the eye based on the highest maximum mean total score of 6.7, recorded 1
hour after test material instillation. This study was rated "ACCEPTABLE" for risk
assessment purposes and NEXY was classified as TOXICITY CATEGORY III for acute
eye irritation by dose of 100 mg per animal.
Primary Dermal Irritation - Rabbit (OPPTS Guideline 870.2500; MRID No. 473138-11):
In a primary dermal irritation study, three rabbits were dermally exposed to 0.5 grams (g)
of NEXY for 4 hours to a 25 millimeter (mm) x 25 mm area of body surface. The animals
were observed for 72 hours. Irritation was scored by the method of Draize. No dermal
irritation was noted on any rabbit during the study. The Primary Irritation Index (PII) was
0.0, the study was rated "ACCEPTABLE" for risk assessment purposes, and NEXY was
classified as TOXICITY CATEGORY IV for primary dermal irritation by dose of 0.5 g
per animal.
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Biopesticides Registration Action Document (BRAD) July 2009
b. Acute Toxicology and Subchronic Toxicity/Pathogenicity - Tier II;
Reproductive Fertility Effects, Carcinogenicity, Immunotoxicity, and
Infectivity/Pathogenicity Analysis - Tier III (40 CFR $ 158.2140)
Tier II and Tier III studies were not required for Candida oleophila Strain O based on the
lack of acute toxicity/pathogenicity in the Tier I studies.
c. Additional Studies
Additional studies, which are not included in the current microbial pesticide toxicology
data requirements and are not required for Candida oleophila Strain O, were submitted to
the Agency. These studies are as follows:
Bacterial Reverse Mutation Test (OPPTS Guideline 870.5100; MRID No. 473138-13)
and In vitro Mammalian Cell Gene Mutation Test (OPPTS Guideline 870.5300; MRID
No. 473138-14): Two mutagenicity studies were reviewed and indicated that Candida
oleophila Strain O did not have mutagenic potential. These studies were rated
"SUPPLEMENTAL" for the purposes of risk assessment and are not required for
Candida oleophila Strain O.
d. Effects on the Endocrine System
Section 408(p) of the Federal Food, Drug, and Cosmetic Act (FFDCA) requires EPA to
develop a screening program to determine whether certain substances (including all
pesticide active and other ingredients) "may have an effect in humans that is similar to an
effect produced by a naturally occurring estrogen, or such other endocrine effect as the
Administrator may designate." Following the recommendations of its Endocrine
Disrupter Screening and Testing Advisory Committee (EDSTAC), EPA determined that
there were scientific bases for including, as part of its program, androgen and thyroid
hormone systems, in addition to the estrogen hormone system. The Environmental
Protection Agency also adopted EDSTAC's recommendation that the Program include
evaluations of potential effects on wildlife.
The Agency has no knowledge of Candida oleophila Strain O being an endocrine
disrupter, nor is this microbe related to any class of known endocrine disrupters.
Following several routes of exposure in rodents, the Tier I toxicology data (see section
IV(B)(l)(a)) indicated that the immune system was still intact and able to process and
clear Candida oleophila Strain O from a variety of organs and tissues. Additional data,
specifically on the endocrine effects of this microbial pesticide, are not required at this
time. When the appropriate screening and/or testing protocols being considered under the
Agency's Endocrine Disrupter Screening Program (EDSP) have been developed and
vetted, Candida oleophila Strain O may be subjected to additional screening and/or
testing to better characterize effects related to endocrine disruption.
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2. Dietary Exposure and Risk Characterization
Dietary exposure to Candida oleophila Strain O is likely to occur, mainly through food.
Candida oleophila Strain O is naturally present on apples as it was originally isolated in
1991 from the surface of golden delicious apples. Based on information submitted to the
Agency, population densities of white yeasts are estimated to reach 1.5 x 103 cfu/square
centimeter (cm2) on harvested apples, which includes the natural population of Candida
oleophila. Background levels of Candida oleophila Strain O on apples are expected to be
below 1.5 x 103 cfu/cm2. Post-harvest treatment with Candida oleophila Strain O will
probably lead to a temporary increased level of this yeast on apples. The mode of action
of Candida oleophila Strain O is primarily based on competition for nutrients; therefore,
sufficient colonization of apple surfaces has to be reached to ensure efficacy of the active
ingredient. The recommended application rate of Candida oleophila Strain O leads to an
expected residual Candida oleophila Strain O population of approximately 4 x 104
cfu/cm2 (105 cfu/apple). Standard practices of washing, peeling, cooking, or processing
fruits further reduces residues of Candida oleophila Strain O and minimizes dietary
exposure. Actual dietary exposure is expected to be several orders of magnitude lower
than the dose used in the acute oral toxicity/pathogenicity test described in section
IV(B)(l)(a), during which no toxic or pathogenic effects were observed in rats.
Additionally, Candida oleophila Strain O has exhibited no ability to grow at mammalian
body temperatures (see section IV(A)(1)), there have been no clinical reports of
infections despite the ubiquitous nature of Candida oleophila on various food
commodities (e.g., olives, apples, strawberries, fermenting grapes, and tomatoes)
consumed by the United States population, and an exemption from the requirement of a
tolerance for residues of Candida oleophila isolate 1-182, which is a similar active
ingredient when compared to Candida oleophila Strain O, was previously established by
the Agency for post-harvest biofungicide use in or on all raw agricultural commodities
(40 CFRS 180.1144).
Based on all of the considerations as outlined above, the Agency concludes that there is a
reasonable certainty that no harm will result from the aggregate exposure to the residues
of Candida oleophila Strain O in food.
3. Drinking Water Exposure and Risk Characterization
Exposure of humans to residues of Candida oleophila Strain O in drinking water is
unlikely. The use pattern, use sites, and application methods for Candida oleophila Strain
O (i.e., indoor dip or drench application to apples and pears after harvest and prior to
storage) does not include direct application to aquatic environments. In the unlikely event
that Candida oleophila Strain O was transferred to surface or ground water intended for
eventual human consumption, the microbe would not survive the conditions water is
subjected to in a drinking water treatment facility, including flocculation, chlorination,
pH adjustments, and/or filtration. Even if oral exposure should occur through drinking
water, the Agency concludes that there is a reasonable certainty that no harm will result
from the exposure to the residues of Candida oleophila Strain O in all the anticipated
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drinking water exposures because of the lack of acute oral toxicity/pathogenicity to
mammals (see section IV(B)(l)(aT) and the ubiquitous nature of the microbe.
4. Acute and Chronic Dietary Exposure and Risks for Sensitive
Subpopulations, Particularly Infants and Children
Section 408(b)(2)(C) of the FFDCA provides that EPA shall assess the available
information about consumption patterns among infants and children, special
susceptibility of infants and children to pesticide chemical residues, and the cumulative
effects on infants and children of the residues and other substances with a common
mechanism of toxicity. In addition, FFDCA section 408(b)(2)(C) also provides that EPA
shall apply an additional tenfold margin of safety for infants and children in the case of
threshold effects to account for prenatal and postnatal toxicity and the completeness of
the database unless EPA determines that a different margin of safety will be safe for
infants and children. Margins of exposure (safety), which are often referred to as
uncertainty factors, are incorporated into EPA risk assessment either directly or through
the use of a margin of exposure analysis, or by using uncertainty (safety) factors in
calculating a dose level that poses no appreciable risk.
Based on the acute toxicity and pathogenicity data discussed in section IV(B)(l)(a), the
Environmental Protection Agency concludes that there is a reasonable certainty that no
harm to sensitive Subpopulations, including infants, children, and adults, will result from
the use of Candida oleophila Strain O in the end-use product, NEXY. This includes all
anticipated dietary exposures and all other exposures for which there is reliable
information. The Agency has arrived at this conclusion because the data available on
Candida oleophila Strain O do not demonstrate toxic, pathogenic, or infective potential to
mammals. Thus, there are no threshold effects of concern and, as a result, the provision
requiring an additional margin of safety does not apply. Further, the considerations of
consumption patterns, special susceptibility, and cumulative effects do not apply to
pesticides without a demonstrated significant adverse effect.
5. Occupational, Residential, School, and Daycare Exposure and Risk
Characterization
Significant additional human exposure to Candida oleophila Strain O from its pesticidal
use is not expected in occupational, residential, school, or daycare areas.
a. Occupational Exposure and Risk Characterization
Even though a report was submitted to the Agency indicating that three workers exhibited
clinical symptoms of a respiratory reaction, potential worker and handler exposure to
Candida oleophila Strain O is not expected to pose any undue risk if the end-use product
is used as labeled. As explained in section IV(B)(l)(a), the three workers were exposed to
large amounts of Candida oleophila Strain O during a pilot-plant production trial of
NEXY using fermentation vessels and were not wearing any personal protective
equipment. To mitigate the possibility for occupational exposure to Candida oleophila
Strain O and potential for respiratory allergic sensitization, mixers and loaders working
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with NEXY must wear a dust/mist filtering respirator meeting NIOSH standards of at
least N-95, R-95, or P-95. Furthermore, although no adverse dermal effects have been
reported by workers and dermal sensitization to operators is not anticipated, potential for
dermal hypersensitivity incidents is mitigated by label requirements that a long-sleeved
shirt, long pants, socks, shoes, and waterproof gloves be worn when mixing, loading, or
applying the product or when handling crates containing treated apples and pears. Lastly,
protective eyewear is also required when mixing, loading, or applying the product or
when handling crates containing treated apples and pears because of exposure potential
and the results of the acute eye irritation study.
b. Residential, School, andDaycare Exposure and Risk Characterization
According to the label, NEXY is to be applied to pears and apples after harvest and
before storage, which is a distinct agricultural use. No indoor residential, school, or
daycare uses currently appear on the label; thus, human exposure to Candida oleophila
Strain O should not occur in these areas.
6. Aggregate Exposure from Multiple Routes Including Dermal, Oral, and
Inhalation
In examining aggregate exposure, Section 408 of the FFDCA directs EPA to consider
available information concerning exposures from the pesticide residue in food and all
other non-occupational exposures, including drinking water from ground water or surface
water and exposure through pesticide use in gardens, lawns, or buildings (residential and
other indoor uses).
Potential non-occupational dermal or inhalation exposure is considered unlikely for this
distinctly agricultural use (i.e., post-harvest treatment of the harvested portions—pears
and apples—of agricultural plants). Oral exposure, via food and drinking water, was
described in section IV(B)(2) and section IV(B)(3), respectively. While a general
residential, school, and daycare exposure and risk characterization is provided in section
IV(B)(5)(b), additional discussion of non-occupational dermal and inhalation exposures
are found below.
a. Dermal Exposure
Non-occupational dermal exposure to Candida oleophila Strain O, when used as labeled,
is expected to be negligible because it is limited to post-harvest agricultural treatment of
apples and pears. However, should non-occupational dermal exposure occur through
treated food commodities, the risk posed by this low toxicity microbe is likely to be
minimal based on the toxicity and pathogenicity tests described in section IV(B)(l)(a).
Furthermore, exposure would not be expected to exceed background as similar yeasts and
those in the genus and species, Candida oleophila., are commonly associated with
particular food commodities.
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b. Inhalation exposure
Non-occupational inhalation exposure to Candida oleophila Strain O, when used as
labeled, is expected to be negligible because of the method of application (i.e., dipping
and drenching of apples and pears), which then allows sufficient time for drying prior to
distribution to consumers. Furthermore, most of the residual yeast on apples and pears is
trapped in the cuticular wax and it is unlikely to be inhaled by consumers.
7. Cumulative Effects
Section 408(b)(2)(D)(v) of the FFDCA requires the Agency to consider the cumulative
effects of exposure to Candida oleophila Strain O and to other substances that have a
common mechanism of toxicity. These considerations include the possible cumulative
effects of such residues on infants and children. Candida oleophila Strain O is not toxic
or pathogenic to mammals via several routes of exposure. Additionally, there are no other
Candida oleophila strains currently registered as pesticides with the Agency.
Consequently, since this microbial pesticide has no demonstrated toxicity, there is no
reason to anticipate cumulative effects from the residues of this product with other related
microbial pesticides.
8. Risk Characterization
The Agency considered human exposure to Candida oleophila Strain O in light of the
standard for registration and safety factors in FIFRA and FFDCA, as amended by the
FQPA. A determination has been made that no unreasonable adverse effects to the United
States population in general, and to infants and children in particular, will result from the
use of Candida oleophila Strain O when used in accordance with EPA-approved labeling.
C. Environmental Assessment
1. Ecological Hazards (40 CFR § 158.2150)
Adequate non-target toxicology data/information are available to support registration of
NEXY, which contains Candida oleophila Strain O, for intended indoor uses on pome
fruits (pears, apples). Because of these intended uses, direct exposures to most terrestrial
and all aquatic environments are not expected, and therefore, all non-target toxicology
data requirements for Candida oleophila Strain O were satisfied with appropriate data
waiver requests and rationales.
Based on the summary data and information submitted in MRID No. 473138-15,
Candida oleophila Strain O is not toxic to adult rainbow trout (Oncorhynchus mykiss) nor
to freshwater cladocerans (Daphnia magnd). As agreed by participants during a pre-
registration conference held on November 21, 2006, only summary information of testing
were submitted for the federal registration for intended indoor uses of Candida oleophila
Strain O. If future uses include outdoor uses where exposures of non-target organisms
would be increased, complete studies must be submitted and reviewed by appropriate
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Agency personnel. Other data requirements may be triggered dependent on the intended
uses and exposures to non-target organisms and the environment.
For a comprehensive guideline-by-guideline summary of the non-target toxicity data
requirements, refer to Table 4 in Appendix A.
2. Environmental Fate and Ground Water Data
Tier II environmental fate and groundwater data requirements were not triggered because
the product is labeled for indoor use on pome fruit (apples and pears) only. Furthermore,
as agreed by participants during a pre-regi strati on conference held on November 21,
2006, summary information of two studies were submitted for the federal registration:
(1) soil survival "kinetics" and (2) survival in sterile distilled water by Candida oleophila
Strain O.
(1) Survival of Candida oleophila Strain O in soil: The viability and survival of
Candida oleophila Strain O was demonstrated in an in vitro study using two
soil microcosms (acidic and limy-clay). After 115 days, it was reported that
"this strain of yeast can survive in heat-treated soil at high density ( >105 cfu/g
soil)" and populations of this yeast did not increase when introduced into
soils.
(2) Survival of Candida oleophila Strain O in sterile distilled water: The viability
and survival of Candida oleophila Strain O decreased and disappeared after
21 days; in aqueous suspensions of 107cells/milliliter (mL). It was reported
that Candida oleophila Strain O is capable of surviving in freshwater lakes
and sediments.
3. Ecological Exposure and Risk Characterization
No risks to non-target organisms or to the environment are anticipated as a result of the
intended indoor uses of Candida oleophila Strain O.
4. Endangered Species Assessment
No risks to endangered species are anticipated as a result of the intended indoor uses of
Candida oleophila Strain O.
V. ENVIRONMENTAL JUSTICE
The Environmental Protection Agency seeks to achieve environmental justice—the fair
treatment and meaningful involvement of all people regardless of race, color, national
origin, or income—with respect to the development, implementation, and enforcement of
environmental laws, regulations, and policies. Fair treatment means that no group of
people, including racial, ethnic, or socioeconomic groups, should bear a disproportionate
share of the negative environmental consequences resulting from industrial, municipal,
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and commercial operations or the execution of federal, state, local, and tribal
environmental programs and policies. Meaningful involvement means that (1) potentially
affected community residents have an appropriate opportunity to participate in decisions
about a proposed activity that will affect their environment and/or health; (2) the public's
contribution can influence the regulatory agency's decision; (3) the concerns of all
participants involved will be considered in the decision-making process; and (4) the
decision-makers seek out and facilitate the involvement of those potentially affected.
EPA has this goal for all communities and persons across the United States.
At this time, the Environmental Protection Agency does not believe the use of NEXY,
which contains Candida oleophila Strain O as an active ingredient, will cause harm or a
disproportionate impact on at-risk communities.
For additional information regarding environmental justice issues, please visit EPA's web
site at http://www.epa.sov/compliance/environmentaliustice/mdex.html.
VI. RISK MANAGEMENT AND REGISTRATION DECISIONS
A. Determination of Eligibility
Section 3(c)(5) of FIFRA permits for the registration of a pesticide provided that all the
following determinations are made:
(1) Its composition is such as to warrant the proposed claims for it;
(2) Its labeling and other material required to be submitted comply with the
requirements of FIFRA;
(3) It will perform its intended function without unreasonable adverse effects on
the environment; AND
(4) When used in accordance with widespread and commonly recognized
practice, it will not generally cause unreasonable adverse effects on the
environment.
To satisfy criterion 1, Candida oleophila Strain O has well-known properties. The
Agency has no knowledge that would contradict the claims made on the NEXY label and
Candida oleophila Strain O is not expected to cause unreasonable adverse effects when
used according to the label instructions. Criterion 2 is satisfied by the current label and
the data presented in this document. It is believed that Candida oleophila Strain O will
not cause any unreasonable adverse effects, and it is likely to provide protection against
blue and gray mold as claimed, satisfying criterion 3. Criterion 4 is satisfied in that
Candida oleophila Strain O is not expected to cause unreasonable adverse effects when
used according to label instructions. Therefore, NEXY, containing Candida oleophila
Strain O as a new active ingredient, is eligible for registration under FIFRA section
3(c)(5) for the labeled uses. If the registrant proposes more extensive uses (e.g., outdoor
uses), a registration amendment with additional data will be required.
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B. Registration Review
The Federal Insecticide, Fungicide, and Rodenticide Act, as amended by the FQPA of
1996, mandated the continuous review of existing pesticides. All pesticides distributed
and sold in the United States must generally be registered by the EPA, based on scientific
data showing that they will not cause unreasonable risks to human health, workers, or the
environment when used as directed in product labeling. The registration review program
is intended to make sure that, as the ability to assess risk evolves and as policies and
practices change, all registered pesticides continue to meet the statutory standard of no
unreasonable adverse effects to human health or the environment. Changes in science,
public policy, and pesticide use practices will occur over time. Through the registration
review program, the Agency periodically reevaluates pesticides to make sure that as
change occurs, products in the marketplace can be used safely. Information on this
program is provided at http://www.epa.gov/oppsrrdl/registration_review/.
The Agency has implemented the registration review program pursuant to FIFRA section
3(g) and will review each registered pesticide every 15 years to determine whether it
continues to meet the FIFRA standard for registration. A registration review decision is
the Agency's determination whether a pesticide meets, or does not meet, the standard for
registration in the Federal Insecticide, Fungicide, and Rodenticide Act. Candida
oleophila Strain O will be included in the schedule for registration review at the end of
the Fiscal Year when schedules are updated.
C. Regulatory Decision
As of June 2009, the data requirements have been fulfilled for NEXY's labeled sites
(Appendix B). Consequently, under FIFRA section 3(c)(5), the Biopesticides and
Pollution Prevention Division recommends unconditional registration of the end-use
product, NEXY, containing Candida oleophila Strain O as an active ingredient.
Additionally, the toxicological database submitted in support of NEXY and Candida
oleophila Strain O meets the safety requirements of the Food Quality Protection Act of
1996, provided that the end-use product is used as labeled; therefore, a permanent
exemption from the requirement of a tolerance has been established for the residues of
the microbial pesticide, Candida oleophila Strain O, on apples and pears when applied as
a post-harvest biofungicide (40 CFR § 180.1289).
To mitigate the possibility for occupational exposure to Candida oleophila Strain O and
potential for respiratory allergic sensitization, mixers and loaders working with NEXY
must wear a dust/mist filtering respirator meeting NIOSH standards of at least N-95, R-
95, or P-95. Furthermore, although no adverse dermal effects have been reported by
workers and dermal sensitization to operators is not anticipated, potential for dermal
hypersensitivity incidents is mitigated by label requirements that a long-sleeved shirt,
long pants, socks, shoes, and waterproof gloves be worn when mixing, loading, or
applying the product or when handling crates containing treated apples and pears. Lastly,
protective eyewear is also required when mixing, loading, or applying the product or
when handling crates containing treated apples and pears because of exposure potential
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and the results of the acute eye irritation study.
D. Labeling
The label for the registered end-use product containing the active ingredient, Candida
oleophila Strain O, is available at http://oaspub.epa.gov/pestlabl/ppls.home.
VII. ACTIONS REQUIRED BY THE REGISTRANT
Before releasing pesticide products containing Candida oleophila Strain O for shipment,
the registrant is required to provide appropriate final printed labeling to the Agency.
The Agency evaluated the data submitted in connection with the initial registration of an
end-use product, NEXY, containing Candida oleophila Strain O as a new active
ingredient. It was determined that these data fulfill current registration guideline
requirements. No additional data are required to be submitted to the Agency at this time.
For new uses and/or changes to existing uses, additional data may be required.
Notwithstanding the information stated in the previous paragraph, it should be clearly
understood that certain, specific data are required to be reported to the Agency as a
requirement for maintaining the federal registration for a pesticide product. A brief
summary of these types of data are listed below.
A. Reporting of Adverse Effects
Reports of all incidents of adverse effects to the environment must be submitted to the
Agency under the provisions stated in FIFRA section 6(a)(2).
B. Reporting of Hypersensitivity Incidents
Additionally, all incidents of hypersensitivity (including both suspected and confirmed
incidents) must be reported to the Agency under the provisions of 40 CFR § 158.2140(d).
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VIII. GLOSSARY OF ACRONYMS AND ABBREVIATIONS
July 2009
BCCM Belgian Coordinated Collections of Microorganisms
BPPD Biopesticides and Pollution Prevention Division
BRAD Biopesticides Registration Action Document
CFR Code of Federal Regulations
cfu colony-forming unit
cm2 square centimeter
°C degrees Celsius
EDSP Endocrine Disrupter Screening Program
EDSTAC Endocrine Disrupter Screening and Testing Advisory Committee
EP end-use product
EPA Environmental Protection Agency (the "Agency")
FFDCA Federal Food, Drug, and Cosmetic Act
FIFRA Federal Insecticide, Fungicide, and Rodenticide Act
FQPA Food Quality Protection Act
FR Federal Register
g gram
kg kilogram
L Liter
LCso median lethal concentration. A statistically derived concentration of a
substance that can be expected to cause death in 50% of test animals. It is
usually expressed as the weight of substance per weight or volume of water,
air, or feed (e.g., mg/L, mg/kg, or ppm).
LLC limited liability company
LOEC lowest observable effect concentration
MRID No. Master Record Identification Number
um micrometer
mg milligram
mL milliliter
mm millimeter
MUCL Mycotheque de I'Universite Catholique de Louvain
NIOSH National Institute for Occupational Safety and Health
NOEC no observable effect concentration
OPP Office of Pesticide Programs
OPPTS Office of Prevention, Pesticides, and Toxic Substances
PII Primary Irritation Index
P.O. Box Post Office Box
PP Pesticide Petition
PPE personal protective equipment
TGAI technical grade of the active ingredient
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IX. BIBLIOGRAPHY
A. Studies Submitted in Support of This Registration
473138-01 Servajean E. 2006. GLP Analysis of a WG Bio-Formulation. Part 1:
Physico-Chemical Properties Upon Receipt (Final Report). Project
Number: 05-75-022 ES. Unpublished report prepared by PHYTOSAFE
s.a.r.l., 31 pages.
473138-02 Servajean E. 2006. GLP Analysis of a WG Bio-Formulation. Part 2:
Stability Upon 6-Month Storage at Room Temperature (Final Report).
Project Number: 05-75-022-ES. Unpublished report prepared by
PHYTOSAFE s.a.r.l., 40 pages.
473138-03 Servajean E. 2006. GLP Analysis of a WG Bio-Formulation. Part 3:
Stability Upon 18-Month Storage at 4 ± 2°C (Final Report). Project
Number: 05-75-022-ES. Unpublished report prepared by PHYTOSAFE
s.a.r.l., 31 pages.
473138-04 Wagner JM. 2007. NEXY Candida oleophila Strain O: Physical and
Chemical Properties Rationale and Request for Waivers for Certain Data.
Unpublished report prepared by JM Wagner, 5 pages.
473138-05 BioNext sprl. 2007. Candida oleophila Strain O: Product Identity,
Manufacturing Process, Analysis of Samples, and Discussion of
Formation of Unintentional Ingredients. Unpublished report prepared by
BioNext sprl, 91 pages.
473138-06 Jij akli MH. 2005. Cell Counting Method for Viable Yeasts in Dry
Products Containing Candida oleophila Strain O. Unpublished report
Prepared by Laboratoire de Pathologie Vegetale, 5 pages.
473138-07 Arrowsmith W. 2005. Candida oleophila (Montrocher, Strain O): Acute
Oral Toxicity and Pathogenicity to the Rat. Project Number:
BTF 001/052303. Unpublished report prepared by Huntingdon Life
Sciences Limited, 58 pages.
473138-08 Arrowsmith W. 2005. Candida oleophila (Montrocher, Strain O): Acute
Subcutaneous Toxicity and Pathogenicity to the Rat. Project Number:
BTF 004/052304. Unpublished report prepared by Huntingdon Life
Sciences Limited, 41 pages.
473138-09 Arrowsmith W. 2005. Candida oleophila (Montrocher, Strain O): Acute
Pulmonary Toxicity and Pathogenicity to the Rat. Project Number:
BTF 002/052490. Unpublished report prepared by Huntingdon Life
Sciences Limited, 57 pages.
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July 2009
473138-10 Rees PB. 2005. Candida oleophila (Montrocher, Strain O): Eye Irritation
to the Rabbit. Project Number: BTF 006/053099. Unpublished report
prepared by Huntingdon Life Sciences Limited, 23 pages.
473138-11 Rees PB. 2005. Candida oleophila (Montrocher, Strain O): Skin Irritation
to the Rabbit. Project Number: BTF 005/052854. Unpublished report
prepared by Huntingdon Life Sciences Limited, 18 pages.
473138-12 BioNext sprl. 2007. Exposure to NEXY Biofungicide: Sensitization Risk
Assessment to Workers, Operators, and Consumers. Unpublished report
prepared by BioNext sprl, 7 pages.
473138-13 Nesslany F. 2005. Mutagenicity Test on Salmonella typhimurium HIS
Using B.N. Ames's Technique with Candida oleophila Montrocher, Strain
O. Project Number: FSR-IPL 041202/Candida oleophila Montrocher,
Strain O/BIONEXT. Unpublished report prepared by Institut Pasteur de
Lille, 39 pages.
473138-14 Nesslany F. 2005. Mutation Assay at the TK Locus in L5178Y Mouse
Lymphoma Cells Using a Microtiter Cloning Technique
(Trifluorothymidine Resistance) Carried Out with Candida oleophila
Montrocher, Strain O. Project Number: FSR-IPL 041203/Candida
Oleophila Montrocher, Strain O/BIONEXT. Unpublished report prepared
by Institut Pasteur de Lille, 61 pages.
473138-15 Wagner JM. 2007. NEXY (Containing Candida oleophila Strain O):
Microbial Nontarget Organisms and Environmental Fate Data
Requirements Rationale and Request for Waivers for Certain Data.
Unpublished report prepared by JM Wagner, 9 pages.
473227-01 Wagner JM. 2008. NEXY Candida oleophila Strain O: Rationale and
Request for Waiver. Unpublished report prepared by JM Wagner,
5 pages.
475610-01 Wagner JM. 2008. NEXY Candida oleophila Strain O: Rationale and
Request for Data Waivers. Unpublished report prepared by JM Wagner,
6 pages.
B. Environmental Protection Agency Risk Assessment Memorandums
Gagliardi JV, Kough JL. 2008. NEXY Containing 57% Candida oleophila Montrocher,
Strain O. U.S. EPA, Washington, D.C. Memorandum dated November 13, 2008.
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Tomimatsu G. 2009. Cover Memo for the Ecological Risk Assessment and
Environmental Effects Chapter for the Biopesticides Registration Action
Document of Candida oleophila Strain O. U.S. EPA, Washington, D.C.
Memorandum dated May 12, 2009.
C. Other References
Jijakli MH, Lepoivre P, Tossut P, Thornard P. 1993. Biological control ofBotrytis
cinerea and Penicillium sp. on post-harvest apples by two antagonistic yeasts.
Med. Fac. Landbouww. Univ. Gent 53(3b): 1349-1358.
Yemma JJ, Berk MP. 1994. Chemical and physiological effects of Candida albicans
toxin on tissues. Cytobios 77(310): 147-158.
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APPENDIX A - MICROBIAL PESTICIDE DATA REQUIREMENTS
TABLE 1. Product Analysis Data Requirements for the Technical Grade of the Active Ingredient (TGAI),
Candida oleophila Strain O, and Its Associated End-Use Product (EP), NEXY (40 CFR § 158.2120)
Data Requirement
(OPPTS Guideline)
Results
TGAI
EP
MRID
Number
Product Chemistry and Composition
Product Identity
(885.1100)
Manufacturing Process
(885.1200)
Deposition of a Sample in a
Nationally Recognized
Culture Collection
(Not applicable)
Discussion of Formation of
Unintentional Ingredients
(885.1300)
Not
applicable
Candida oleophila Strain O is a single-celled yeast in
the phylum Ascomycota, the class Ascomycetes, and
the family Saccharomycetaceae. Found naturally on
plant tissues (fruits,
flowers, and wood) and in water,
it was originally isolated from golden delicious
apples.
NEXY, containing Candida oleophila Strain O, is
intended for use as an antagonist to control the
fungal pathogens, gray mold (Botrytis cinerea) and
blue mold (Penicillium expansum), that cause post-
harvest decay on apples and pears. The mode of
action for Candida oleophila Strain O is primarily
through competition for nutrients and pre-
colonization of plant wound sites (Jijakli et al. 1993).
Submitted data also
1,3-glucanases (i.e.,
suggest that production of beta-
hydrolytic enzymes that can
degrade fungal phytopathogen cell walls) may
contribute to its antagonistic activity.
Classification: Acceptable
Submitted data satisfy the requirements of manufacturing process
for both the TGAI and EP.
Classification: Acceptable
Candida oleophila Strain O is
on deposit with the Belgian
Coordinated Collections of
Microorganisms
(BCCM)Mycotheque de
I'Universite Catholique de
Louvain (MUCL) in Louvain-
la-Neuve, Belgium under
Accession Number MUCL
40564.
Not applicable
Submitted data satisfy the requirements of discussion of formation
of unintentional ingredients for both the TGAI and EP.
Classification: Acceptable
473138-05
473138-05
473138-05
473138-05
Analysis and Certified Limits
Analysis of Samples
(885.1400)
Submitted data satisfy the requirements of analysis of samples for
both the TGAI and EP.
Classification: Acceptable
473138-05
473138-06
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Data Requirement
(OPPTS Guideline)
Certification of Limits
(885.1500)
Results
TGAI
Not applicable
EP
The certified limits for the
active and inert ingredients
exceed the OPPTS Guideline
830. 1750 specified ranges, but
an acceptable explanation was
provided.
Classification: Acceptable
MRID
Number
Not applicable
Physical and Chemical Characteristics
Color
(830.6302)
Physical State
(830.6303)
Odor
(830.6304)
Stability to Normal and
Elevated Temperatures,
Metals, and Metal Ions
(830.6313)
Storage Stability
(830.6317)
Miscibility
(830.6319)
Corrosion Characteristics
(830.6320)
pH
(830.7000)
Viscosity
(830.7100)
Density /Relative Density /Bulk
Density
(830.7300)
Yellow
Solid (micro-granules)
Yeast
Waived due to the dry basis of
the product, the stability at
normal temperatures, and
packaging in non-metallic
containers.
Classification: Acceptable
Not applicable
Not applicable
Not applicable
Not applicable
6-month study at 20±2°C, the active ingredient was higher then
the acceptance value (8. 10 x 109 cfu/g), though a significant
decrease was seen after 5 and 6 months of storage.
18-month study at 4±2°C, the active ingredient was higher then
the acceptance value (6. 10 x 109 cfu/g), though a significant
decrease was seen at 18 months of storage.
Not applicable
Not applicable
6.24 (1% weight/volume in
water)
Not applicable
Bulk density = 0.597 kilogram
(kg)/Liter (L)
Tap density = 0.667 kg/L
Not required because NEXY is
not an emulsifiable form of
microbial pesticide (refer to test
note#2of40CFR§
158.2120(d)).
Waived because NEXY is a dry
product on an inert carrier
packaged in polyethylene
materials highly resistant to
corrosion.
Classification: Acceptable
Not applicable
Not required because NEXY is
not a liquid form of microbial
pesticide (refer to test note #4
of40CFR§ 158.2120(d)).
Not applicable
473138-01
473138-01
473138-01
473138-04
473138-02
473138-03
473138-04
473138-04
473138-01
473138-04
473138-01
26
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Candida oleophila Strain O
Biopesticides Registration Action Document (BRAD)
July 2009
TABLE 2. Toxicology Data Requirements for the Technical Grade of the Active Ingredient (TGAI), Candida
oleophila Strain O, and Its Associated End-Use Product (EP), NEXY (40 CFR § 158.2140)
Data Requirement
(OPPTS Guideline)
Results
TGAI
I
EP
MRID
Number
Tier I
Acute Oral
Toxicity/Pathogenicity
(885.3050)
Not toxic, infective, and/or
pathogenic to rats by oral dose
of 2.3-3.8 x 108cfu/animal.
Classification: Acceptable
Not applicable
473138-07
Acute Pulmonary
Toxicity/Pathogenicity
(885.3150)
Not toxic, infective, and/or
pathogenic to rats by
pulmonary dose of 1.2-5.2 x
108cfu/animal.
Classification: Acceptable
Not applicable
473138-09
Acute Injection
Toxicity/Pathogenicity
(885.3200)
Not toxic, infective, and/or
pathogenic to rats by
subcutaneous dose of 1.1-2.0
x 107 cfu/animal.
Classification: Acceptable
Not applicable
473138-08
Hypersensitivity Incidents
(885.3400)
During a pilot-plant production trial of NEXY using fermentation
vessels and involving large amounts of Candida oleophila Strain
O, three of six workers not wearing personal protective equipment
reported clinical symptoms of a respiratory reaction. No adverse
dermal effects have been reported by workers. Potential for
hypersensitivity incidents has been mitigated by label
requirements that a long-sleeved shirt, long pants, socks, shoes,
and waterproof gloves be worn when mixing, loading, or applying
the product or when handling crates containing treated apples and
pears. Additionally, mixers and loaders must wear a dust/mist
filtering respirator meeting NIOSH standards of at least N-95, R-
95, orP-95. Any future hypersensitivity incidents must be reported
per OPPTS Guideline 885.3400.
473138-12
Cell Culture
(885.3500)
Not required because Candida
oleophila Strain O is not a
virus (refer to test note #4 of
40 CFR § 158.2140(d)).
Not applicable
Not applicable
Acute Oral Toxicity
(870.1100)
Not applicable
Waived based on the results of
MRID Number 473138-07 and
the nature of the inert ingredients
in NEXY's formulation.
Classification: Acceptable
TOXICITY CATEGORY IV
475610-01
Acute Dermal Toxicity
(870.1200)
Not applicable
Waived based on the results of
MRID Numbers 473138-08 and
473138-11 and the nature of the
inert ingredients in NEXY's
formulation.
Classification: Acceptable
TOXICITY CATEGORY III
473227-01
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Candida oleophila Strain O
Biopesticides Registration Action Document (BRAD)
July 2009
Data Requirement
(OPPTS Guideline)
Acute Inhalation Toxicity
(870.1300)
Acute Eye Irritation
(870.2400)
Primary Dermal Irritation
(870.2500)
Results
TGAI
Not applicable
Not applicable
Not applicable
EP
Waived based on the results of
MRID Number 473 138-09 and
the nature of the inert ingredients
in NEXY's formulation.
Classification: Acceptable
TOXICITY CATEGORY III
NEXY was minimally irritating
to rabbits at a dose of 100
mg/animal.
Classification: Acceptable
TOXICITY CATEGORY III
NEXY was not irritating to
rabbits at a dose of 0.5 g/animal.
Classification: Acceptable
TOXICITY CATEGORY IV
MRID
Number
475610-01
473138-10
473138-11
Tiers II and III
Not required for Candida oleophila Strain O based on the lack of acute toxicity/pathogenicity in the Tier I studies.
TABLE 3. Additional Studies Submitted to Support the Registration of the End-Use Product (EP),
NEXY, Containing Candida oleophila Strain O as an Active Ingredient
Data Requirement
(OPPTS Guideline)
Results
MRID
Number
Mutagenicity Testing
Bacterial Reverse Mutation
Assay
(870.5100)
In vitro Mammalian Cell
Assay
(870.5300)
Indicates Candida oleophila Strain O does not have mutagenic
potential.
Classification: Supplemental (Not a required study for this
active ingredient)
Indicates Candida oleophila Strain O does not have mutagenic
potential.
Classification: Supplemental (Not a required study for this
active ingredient)
473138-13
473138-14
28
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Candida oleophila Strain O
Biopesticides Registration Action Document (BRAD)
July 2009
TABLE 4. Non-Target Organisms and Environmental Fate Data Requirements for the Technical Grade of the
Active Ingredient (TGAI), Candida oleophila Strain O (40 CFR § 158.2150)
Data Requirement
(OPPTS Guideline)
Results
Toxicity
Category/Description
MRID
Number
Tier I
Avian Oral Toxicity
(885.4050)
Not required for the intended indoor
use pattern (refer to test note #1 of 40
CFR § 158.2150(e)).
Submitted waiver rationale was
Acceptable.
Not applicable
473138-15
Avian Inhalation
Toxicity/Pathogenicity
(885.4100)
Not required for the intended indoor
use pattern (refer to test note #1 of 40
CFR § 158.2150(e)).
Submitted waiver rationale was
Acceptable.
Not applicable
473138-15
Wild Mammal
Toxicity/Pathogenicity
(885.4150)
Not required for the intended indoor
use pattern.
Submitted waiver rationale was
Acceptable.
Not applicable
473138-15
Freshwater Fish
Toxicity/Pathogenicity
(885.4200)
Not required for the intended indoor
use pattern (refer to test note #1 of 40
CFR § 158.2150(e)).
Submitted waiver rationale was
Acceptable. Submitted summary
information indicated no mortalities on
adult rainbow trout under semi-static,
daily water renewal conditions. The
reported median lethal concentration
(LC50) was greater than 333 mg
TGAI/L after 96 hours.
Not acutely toxic to
adult rainbow trout1
473138-15
Freshwater Invertebrate
Toxicity/Pathogenicity
(885.4240)
Not required for the intended indoor
use pattern (refer to test note #1 of 40
CFR § 158.2150(e)).
Submitted waiver rationale was
Acceptable. Submitted summary
information indicated no mortalities on
adult daphnids under semi-static,
frequent renewal conditions. The
reported no observable effect
concentration (NOEC) (measured) was
equal to 15.7 mg TGAI/L; reported
lowest observable effect concentration
(LOEC) (measured) was greater than
15.7 mg TGAI/L after 21 days.
Not toxic to adult
daphnids1
473138-15
1 Only summary information of testing were submitted for the federal registration for intended indoor uses
of Candida oleophila Strain O, as agreed by participants during a pre-registration conference held on
November 21, 2006.
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Candida oleophila Strain O
Biopesticides Registration Action Document (BRAD)
July 2009
Data Requirement
(OPPTS Guideline)
Estuarine/Marine Fish Testing
Estuarine and Marine
Invertebrate Testing
(885.4280)
Non-Target Plant Testing
(885.4300)
Non-Target Insect Testing
(885.4340)
Honey Bee Testing
(885.4380)
Results
Not required for the intended indoor
use pattern.
Not required for the intended indoor
use pattern (refer to test note #1 of 40
CFR § 158.2150(e)).
Submitted waiver rationale was
Acceptable.
Not required for the intended indoor
use pattern.
Submitted waiver rationale was
Acceptable.
Not required for the intended indoor
use pattern.
Submitted waiver rationale was
Acceptable.
Toxicity
Category/Description
Not applicable
Not applicable
Not applicable
Not applicable
MRID
Number
Not applicable
473138-15
473138-15
473138-15
Tiers II, III, and IV
Not required for Candida oleophila Strain O because its associated end-use product, NEXY, is labeled for indoor use
on pome fruit (apples and pears) only.
30
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Candida oleophila Strain O
Biopesticides Registration Action Document (BRAD)
July 2009
APPENDIX B - CANDIDA OLEOPHILA STRAIN O END-USE PRODUCTS
EPA
Registration
Number
84863-1
Registration
Name
NEXY
Percentage
Active
Ingredient
57.0%
Formulation
Type
Dilutable
Powder
Use Site
Indoor
(Apples
and
Pears)
Method of
Application
Dip or Drench
Application
Rate
0.9 ounces of
NEXY and
5.4 ounces of
NEXY
ADDITIVE
per 20 gallons
of water
2.4 to 7. 3
gallons of
mixture per
ton of apples
or pears
Target Pests
Gray Mold
(Botrytis
cinerea) and
Blue Mold
(Penicillium
expansum)
31
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