Isaria fumosoroseus strain FE 9901 – Petition to Establish a Tolerance Exemption Page 1 of 11 

## Petition for an exemption from the requirement of a tolerance for residues of products containing the active ingredient

“ Isaria fumosoroseus strain FE 9901” in and on all food

commodities

Submitted for: Novozymes BioAg, Inc. (EPA Company No. 73314) 13100 W. Lisbon Road, Suite 600 Brookfield, Wisconsin 53005

Submitted by: Technology Sciences Group Inc.

1150 18 th 

### Street, NW, Suite 1000 Washington, DC 20036



## SECTION A Name, Chemical identity, and Composition of theIsaria fumosoroseus strain FE 9901 – Petition to Establish a Tolerance Exemption Page 2 of 11 

## Pesticide Chemical

Product Name: NoFLYTM Technical (EPA Reg. No. 73314-7) NoFLYTM WP (EPA Reg. No. 73314-6)

Active Ingredient: Isaria fumosoroseus strain FE 9901

Isaria fumosoroseus strain FE 9901 is a naturally occurring fungus found in most countries around the world and occurs in various soil types at very low densities. The fungus has frequently been isolated from Lepidoptera cadavers in Europe. Isolates occur in the Netherlands, Ireland, and Spain. The fungus has also been found in various non-European countries including China, Japan, Ghana, Pakistan, India, and the United States. Relevant information regarding the name, identity, and composition has been submitted to EPA and can be found in
MRID Nos. 47792001, 48051401, and 48051403.

Both the TGAI and end-use product formulation contain direct food additives as inert ingredients. The end-use product is currently used to control a variety of target insect pests (i.e. whiteflies, aphids, thrips, Psyllids, Mealybugs, and fungus gnats) on greenhouse nonfood plants but is proposed for use in/on growing agricultural crops, greenhouses, turf, nursery crops, commercial landscapes, and residential homes and gardens.

Mode of Action: As a representative and primary target pest species, Isaria

fumosoroseus strain FE 9901 infects adult whiteflies, all immature whitefly stages, and whitefly eggs. The ability to infect whitefly eggs is unique among entomopathogenic fungi. I. fumosoroseus strain FE 9901 is dispersed as conidia or blastospores, either by air or water movement or by other insects and/or mites. The mode of action of the various entomopathogenic fungi is functionally similar, and a typical infection cycle consists of: 1) conidial attachment; 2) germination; 3) penetration; 4) vegetative growth; and 5) conidiogenesis.

The infection cycle of I. fumosoroseus strain FE9901 is particularly rapid under optimal conditions. Initially, conidia are attached to cuticle or dorsum of immature or adult whiteflies. First symptoms of infection typically are apparent within one to two days after contact. Conidia and hyphae are present in the host hemocoel within one day. Fungal multiplication takes place through formation of hyphal bodies or blastospores. The mycelium is present on the dorsum of the insect body within two days. Sporulation occurs within three days and reaches a maximum within five to seven days. Relevant literature is summarized or

referenced in Attachment A of MRID No. 47792001.Isaria fumosoroseus strain FE 9901 – Petition to Establish a Tolerance Exemption Page 3 of 11 

## SECTION B Amount, Frequency, a nd Timing of Application of

the Pesticide Chemical

The intended use of Isaria fumosoroseus strain FE 9901 in the end-use product, NOFLYTM WP (EPA Reg. No. 73314-6) is as an insecticide for use on greenhouse and outdoor ornamentals, trees and shrubs, nursery crops, turf, production agriculture crops, commercial landscapes, and residential/home and garden ornamentals, plants, and lawns by foliar spray or soil drench application, by chemigation, and/or by seed treatment. Target pests include whiteflies, aphids, thrips, Psyllids, Mealybugs, and fungus gnats. NOFLYTM WP can be applied at 5-15 day intervals at rates of 1⁄2 - 2 pounds per acre or 2 – 8 ounces per 10,000 square feet on ornamentals, trees and shrubs, nonfood crops, and food crops (corn, potatoes, cucurbit vegetables, fruiting vegetables, grains, cereals, soybeans, cotton, oil crops, herbs and spices, leafy and cole vegetables, legume vegetables, root and tuber/bulb vegetables, berry crops, citrus, grape, pome fruit, stone fruit, nuts, tropical fruit, asparagus, banana, plantain, watercress, mushroom, olive, and hops) up to and including the day of harvest.

NOFLYTM WP can be applied to golf course tees, greens, fairways, and roughs, commercial and residential lawns, athletic fields, parks, cemeteries, sod farms, turf landscapes, herbaceous and woody ornamentals, trees and plants against whitef lies, aphids, thrips, Psyllids, Mealybugs, and fungus gnats at rates of 1⁄2 - 2 pounds per acre or 2 – 8 ounces per 10,000 square feet at the same application interval (5-15 days) as crop use.

For residential/home and garden use, NOFLYTM WP can be applied as a foliar spray or soil drench to control whiteflies, aphids, thrips, Psyllids, Mealybugs, and fungus gnats on fruit and vegetable plants, annual and perennial bedding plants and flowers, roses, potted flowers, foliage plants, trees, shrubs, and lawns every 5-15 days at rates ranging from 2 – 8 tablespoons per gallon of water to 1⁄2 - 2 1⁄2 cups per 5 gallons of water.Isaria fumosoroseus strain FE 9901 – Petition to Establish a Tolerance Exemption Page 4 of 11 

## SECTION C Reports on Investigations Made with Respect to the Safety of the Pesticide Chemical

Toxicity of Isaria fumosoroseus strain FE 9901

Studies to evaluate the safety to mammals were conducted on the technical grade active ingredient Isaria fumosoroseus strain FE 9901 and are summarized herein (see Table 1). The studies performed include toxicity/pathogenicity studies via the oral, pulmonary, and intraperitoneal injection pathways as well as the conventional acute toxicity (“6-pack”) studies to assess acute oral, dermal, and inhalation toxicity, eye and skin irritation, and dermal sensitization. The oral, dermal, and inhalation routes of exposure are the routes relevant for

human health risk assessment. The results of thes e studies consistently demonstrated that

Isaria fumosoroseus strain FE 9901 was non-toxic by all routes of exposure and only minimally irritating. Additionally, there was no evidenc e of pathogenically observed; the product was not infective or pathogenic to test animals when administered via the oral, pulmonary, or intraperitoneal routes. Even though positive for skin s ensitization; no reports of hypersensitivity have been recorded from personnel working with this organism.



### Table 1. Summary of I. fumosoroseus strain FE 9901 Mammalian Toxicity Data Test Species Dose Tested Results
Acute oral toxicity

MRID No. 47791903 Rat Gavage dose of 5,000 mg/kg

2.0x10 9 CFU/g LD 50 > 5,000 mg/kg Category IV

Acute dermal toxicity

MRID No. 47791904 Rat Undiluted dose of 5,050 mg/kg

2.0x10 9 CFU/g LD 50 > 5,050 mg/kg Category III

Acute inhalation toxicity

MRID No. 47791905 Rat 2.18 mg/L for 4 hours

2.0x10 9 CFU/g LC 50 > 2.18 mg/L Category III

Primary eye irritation

MRID No. 47791906 Rabbit 0.1 gram undiluted product for 24 hours Slightly irritating Category IV

Dermal irritation

MRID No. 47791907 Rabbit 0.5 gram undiluted product for 4 hours Slightly irritating Category IV

Dermal Sensitization

MRID No. 47791908 Guinea Pig 1% intradermal injection + 25% topical application Localized area of erythema and swelling

Positive for skin sensitization

Acute oral toxicity/pathogenicity

MRID No. 47792002 Rat 100mg of 10 8

CFU/rat Non-toxic, non-infective, nonpathogenic

Acute pulmonary toxicity/pathogenicity

MRID Nos. 47792004

49044301 Ferret 1.0x10 8 (in 0.5mL)

CFU/ferret Non-toxic, non-infective, nonpathogenic

Acute intraperitoneal Toxicity/pathogenicity

MRID Nos. 47792005

49044301 Rat 10 7 CFU/rat Non-toxic, non-infective, nonpathogenicIsaria fumosoroseus strain FE 9901 – Petition to Establish a Tolerance Exemption Page 5 of 11 The results of toxicity testing on the organism show there is no risk to human health from the active ingredient. Isaria fumosoroseus strain FE 9901 is not toxic, not pathogenic, not infective, and only minimally irritating to mammals. Further, it is important to note that Isaria

fumosoroseus strain FE 9901 does not grow or develop at temperatures in excess of 35°C (95°F) (MRID No. 47792001) and therefore is essentially inactivated in mammalian systems.

Literature searches have been conducted against the Agency’s list of open literature database searches (July 26, 2010 edition). Those searches have demonstrated that there are no reports of ecological or human health hazards caused by Isaria fumosoroseus strain FE 9901. It does not produce recognized toxins, enzymes, or virulence factors normally associated with mammalian invasiveness or toxicity.

A special analysis was conducted across five production batches of NOFLYTM Technical to determine the prevalence of beauverolide metabo lites, beauvericin, and 2,6-pyridinedicarboxylic acid (PDCA) following the fermentation process. The results of this analysis (MRID No. 49118301) concluded that there was no quantitation of beauvericin and 2,6-PDCA was only quantified at levels of 1-10 μ g/g. Across the fifteen beauverolide peptide species analyzed, Beauverolide I was determined to be the most prevalent (18% of the overall peptides combined), and 100% of the peptides combined were well below 0.1% of NOFLYTM Technical; therefore, no metabolite formation rises to a level of toxicological concern. This is further confirmed due to the low toxicity evidenced across the suite of mammalian studies on Isaria

fumosoroseus strain FE 9901.

As has been demonstrated, dietary exposure to Isaria fumosoroseus strain FE 9901 meets the FQPA 1996 safety standard. Studies submitted with this petition showed no evidence of toxicity, pathogenicity, or infectivity when this strain was tested in laboratory animals.



### Aggregate Exposure

1) Dietary Exposure:

Dietary exposure from use of Isaria fumosoroseus strain FE 9901, as proposed, is

minimal. The intended use of Isaria fumosoroseus strain FE 9901 is to agricultural crops, greenhouse and outdoor food and nonfood crops, nursery crops, trees and shrubs, commercial turf and landscapes, and residential lawns and gardens for the purposes of insect control. Isaria fumosoroseus strain FE 9901 does not grow or develop at temperatures in excess of 35°C (95°F), and residues are not expected due to its degradation in sunlight and in high temperatures (MRID No. 47792001). Any remaining residues would be removed during common post-harvest treatments including washing, packing, peeling, etc.

The results of toxicity testing indicate there is no risk to human health or the environment

from Isaria fumosoroseus strain FE 9901. There are no reports of ecological or human health hazards caused by the microorganism Isaria fumosoroseus . It does not produce recognized toxins, enzymes, or virulence factors normally associated with mammalian invasiveness or toxicity. The absence of acute toxicity or pathogenicity in laboratory animals demonstrates the benign nature of this strain. Dietary exposures would not be expected to pose any quantifiable risk, due to a lack of residues of toxicological concern.Isaria fumosoroseus strain FE 9901 – Petition to Establish a Tolerance Exemption Page 6 of 11 2) Drinking Water Exposure:

Similarly, exposure to humans from residues of Isaria fumosoroseus strain FE 9901 in consumed drinking water would be unlikely. Potential exposure to surface water would be negligible and exposure to drinking water (well or ground water) would be impossible

to measure. The intended use of Isaria fumosoroseus strain FE 9901 is to agricultural crops, greenhouse and outdoor food and nonfood crops, nursery crops, trees and shrubs, commercial turf and landscapes, and residential lawns and gardens for the purposes of insect control. The risk of the microorganism passing through the soil to ground water is minimal to unlikely. Additionally the fungi would not tolerate the conditions water is subjected to in a drinking-water treatment facility (including: chlorination, pH adjustments, high temperatures and/ or anaerobic conditions) (MRID No. 47792001).

The results of toxicity testing indicate there is no risk to human health or the environment

from Isaria fumosoroseus strain FE 9901. There are no reports of ecological or human health hazards caused by the microorganism Isaria fumosoroseus . It does not produce recognized toxins, enzymes, or virulence factors normally associated with mammalian invasiveness or toxicity. The absence of acute toxicity or pathogenicity in laboratory animals demonstrates the benign nature of this strain. Drinking water exposures would not be expected to pose any quantifiable risk, due to a lack of residues of toxicological

concern.

3) Non-Dietary Exposure:

The potential for non-dietary exposure to the general population, including infants and children, is unlikely. The intended use of Isaria fumosoroseus strain FE 9901 is to agricultural crops, greenhouse and outdoor food and nonfood crops, nursery crops, trees and shrubs, commercial turf and landscapes, and residential lawns and gardens for the purposes of insect control. The non-toxic and non-pathogenic nature of this product demonstrates that other incidental non-dietary exposures are not of concern.

The results of toxicity testing indicate there is no risk to human health or the environment

from Isaria fumosoroseus strain FE 9901. There are no reports of ecological or human health hazards caused by the microorganism Isaria fumosoroseus . It does not produce recognized toxins, enzymes, or virulence factors normally associated with mammalian invasiveness or toxicity. The absence of acute toxicity or pathogenicity in laboratory animals demonstrates the benign nature of this strain. Non-dietary exposures would not be expected to pose any quantifiable risk due to a lack of residues of toxicological

concern.



### Cumulative Effects

It is not expected that, when used as proposed, Isaria fumosoroseus strain FE 9901 would result in residues that are of toxicological concern. The intended use of Isaria fumosoroseus strain FE 9901 is to agricultural crops, greenhouse and outdoor food and nonfood crops,Isaria fumosoroseus strain FE 9901 – Petition to Establish a Tolerance Exemption Page 7 of 11 nursery crops, trees and shrubs, commercial turf and landscapes, and residential lawns and gardens for the purposes of insect control. The results of toxicity testing indicate there is no risk

to human health or the environment from Isaria fumosoroseus strain FE 9901. There are no reports of ecological or human health hazards caused by Isaria fumosoroseus strain FE 9901. It does not produce recognized toxins, enzymes, or virulence factors normally associated with mammalian invasiveness or toxicity. The potential for toxic metabolite formation is diminutive in

nature as demonstrated in the de minimus findings of beauverolide formation. The absence of acute toxicity or pathogenicity in laboratory animals demonstrates the benign nature of this strain. Any exposures would not be expected to pose any quantifiable risk due to a lack of residues of toxicological concern.



### Safety Determination

1) General US Population:

Acute toxicity studies have demonstrated that Isaria fumosoroseus strain FE 9901 is not toxic, pathogenic, or infective to mammals. Other ingredients associated with the TGAI and end-use product are also direct food additives. The intended use of Isaria

fumosoroseus strain FE 9901 is to agricultural crops, greenhouse and outdoor food and nonfood crops, nursery crops, trees and shrubs, commercial turf and landscapes, and residential lawns and gardens for the purposes of insect control. The results of toxicity testing indicate there is no risk to human health or the environment from Isaria

fumosoroseus strain FE 9901, and there is a lack of residues on treated commodities. There are no reports of ecological or human hazards caused by the microorganism

Isaria fumosoroseus . It does not produce recognized toxins, enzymes, or virulence factors normally associated with mammalian invasiveness or toxicity. The absence of acute toxicity or pathogenicity in laboratory animals demonstrates the benign nature of this strain. There is a reasonable certainty of no harm to the general US population from exposure to this active ingredient.

2) Infants and Children:

As mentioned above, it is not expected that, when used as proposed, Isaria

fumosoroseus strain FE 9901 would result in residues that are of toxicological concern. There is a reasonable certainty of no harm for infants and children from exposure to

Isaria fumosoroseus strain FE 9901 from the proposed uses.



### Information on Endocrine Effects

To date there is no evidence to suggest that Isaria fumosoroseus strain FE 9901 functions in a manner similar to any known hormone, or that it acts as an endocrine disrupter.Isaria fumosoroseus strain FE 9901 – Petition to Establish a Tolerance Exemption Page 8 of 11 

## Section D Tests on the Amount of Residue Remaining

Not applicable, Novozymes BioAg, Inc. is petitioning for an exemption from the requirement of a

tolerance for residues of Isaria fumosoroseus strain FE 9901.

Analytical Method: An analytical method for residues is not applicable. It is expected
that, when used as directed, Isaria fumosoroseus strain FE 9901 will not result in residues that are of toxicological concern.Isaria fumosoroseus strain FE 9901 – Petition to Establish a Tolerance Exemption Page 9 of 11 

## SECTION E - Practical Methods for Removing Residues

Not applicable, Novozymes BioAg, Inc. is petitioning for an exemption from the requirement of a

tolerance for residues of Isaria fumosoroseus strain FE 9901.Isaria fumosoroseus strain FE 9901 – Petition to Establish a Tolerance Exemption Page 10 of 11 

## SECTION F – Proposed Tolerance Exemption

Novozymes BioAg, Inc. is proposing that the following Exemption from the need of a tolerance be established for the pesticide, Isaria fumosoroseus strain FE 9901, similar to the current tolerance exemption in 40 CFR §180.1306 for a similar fungal strain, Isaria fumosorosea Apopka strain 97:

“An exemption from the requirement of a tolerance is established for residues of Isaria

fumosoroseus strain FE 9901 in or on all food commodities when applied as an insecticide or miticide and used in accordance with good agricultural practices.”Isaria fumosoroseus strain FE 9901 – Petition to Establish a Tolerance Exemption Page 11 of 11 

## SECTION G Reasonable Grounds in Support of an Exemption

from the Need of a Tolerance

EXISTING TOLERANCES

US EPA Tolerance -

While there is not a specific tolerance for Isaria fumosoroseus strain FE 9901, there is an established and existing tolerance exemption for a similar fungal strain ( Isaria fumosorosea Apopka strain 97), codified under 40 CFR §180.1306, which states:

“An exemption from the requirement of a tolerance is established for residues of Isaria

fumosorosea Apopka strain 97 in or on all food co mmodities when applied as an insecticide or miticide and used in accordance with good agricultural practices.”

International -

A Codex Alimentarium Commission Maximum Residue Level (MRL) is neither established nor required for Isaria fumosoroseus strain FE 9901.

It is not expected that, when used as proposed, Isaria fumosoroseus strain FE 9901 would result in residues that are of toxicological concern. The intended use of Isaria fumosoroseus strain FE 9901 is to agricultural crops, greenhouse and outdoor food and nonfood crops, nursery crops, trees and shrubs, commercial turf and landscapes, and residential lawns and gardens for the purposes of insect control. The results of toxicity testing indicate there is no risk

to human health or the environment from Isaria fumosoroseus strain FE 9901. There are no reports of ecological or human health hazards caused by Isaria fumosoroseus strain FE 9901. It does not produce recognized toxins, enzymes, or virulence factors normally associated with mammalian invasiveness or toxicity. The absence of acute toxicity or pathogenicity in laboratory animals demonstrates the benign nature of this strain, and in conjunction with the established tolerance for similar fungal strains ( Isaria fumosorosea Apopka strain 97), serves to demonstrate reasonable grounds in support of the pet ition for exemption from the requirement

of a tolerance.