BIOPESTICIDES REGISTRATION ACTION DOCUMENT



## Pseudomonas fluorescens s train D7

Pesticide Chemical (PC) Code: 016418

U.S. Environmental Protection Agency Office of Pesticide Programs Biopesticides and P ollution Prevention Division

August 7 , 2014Pseudomonas fluorescens s train D7 Page 2 of 29

Biopesticides Registration Action Document

T ABLE OF CONTENTS

I. EXECUTIVE SUMMAR Y

................................ ................................ ................................ ................................ ..... 4

II. ACTIVE INGREDIE NT OVERVIEW

................................ ................................ ................................ ............... 6

III. REGULATORY BACK GROUND

................................ ................................ ................................ ...................... 6

A. Applications for Pesticide Product R egistration

................................ ................................ ................................ . 6

B. Food Tolerance Exemption

................................ ................................ ................................ ................................ ... 6

IV. RISK ASSESSMENT

................................ ................................ ................................ ................................ ........... 7

A. Product Analysis Assessment (40 CFR § 158.2120)

................................ ................................ ............................ 7

B. Human Health Assessment (40 CFR § 158.2140)

................................ ................................ ................................ 8

C. Environmental Assessm ent (40 CFR § 158.2150)

................................ ................................ .............................. 14

V. ENVIRONMENTAL JU STICE

................................ ................................ ................................ ......................... 17

VI. RISK MANAGEMENT DECISION

................................ ................................ ................................ ................ 17

A. Final Printed Labeling

................................ ................................ ................................ ................................ ......... 18

B. Terms of Registration

................................ ................................ ................................ ................................ .......... 18

C. Reporting of Adverse Effects and Hypersensiti vity Incidents

................................ ................................ ......... 18

VIII. GLOSSARY OF A CRONYMS AND ABBREVIA TIONS

................................ ................................ .......... 19

IX. BIBLIOGRAPHY

................................ ................................ ................................ ................................ .............. 20

A. Studies Submitted to Support the Active Ingredient, Pseudomonas fluorescens strain D7 and Pesticide Product Registrations for the Fluorescens Technical and the EP, D7

................................ ................................ .. 20

B. Environmental Protection Agency Risk Assessment Memoranda

................................ ................................ ... 22

C. Other References

................................ ................................ ................................ ................................ ................. 23

APPENDIX A. MICROBI AL PESTICIDES DATA REQUIREMENTS

................................ ........................... 24Pseudomonas fluorescens s train D7 Page 3 of 29

Biopesticides Registration Action Document

BIOPESTICIDES REGISTRATION ACTION DOCUMENT TEAM

Office of Pesticide Programs (OPP) Biopesticides and Pollution Prevention Division

Microbial Pesticides Branch

Science Reviews

Ibrahim S. Barso um, Ph.D. Product Analysis, Human Health

John L. Kough, Ph.D. Product Analysis, Human Health

In

- Soon You, Ph.D. Environmental Effects

Gail Tomimatsu , Ph.D. Environmental Effects Shannon Borges Environmental Effects

Regula t ory

Kimberly Nesci Chief, Microbial Pesticides Branch

Susanne Cerrelli Regulatory Action LeaderPseudomonas fluorescens s train D7 Page 4 of 29

Biopesticides Registration Action Document

I. EXECUTIVE SUMMARY

Background

On October 5, 2012, Ag

- Chem Consulting on behalf of Northwest Agricultural Products submitted application s for a new technical grade active ingredient (TGAI)

, Fluorescens

Technical (EPA File Symbol 71975

- G ), and a new end

- use product (EP)

, D7 (EPA File Symbol

71975

- U )

, to the United States Environmental Protection Agency (EPA) under section 3 of the

Federal Insectici de, Fungicide, and Rodenticide Act (FIFRA). These products both contain the new active ingredient, Pseudomonas fluorescens strain D7

.

Pseudomonas fluorescens strain D7 is a naturally occurring bacterium originally isolated from

winter wheat roots. On Fe bruary 14, 201 2

, an Experimental Use Permit no. 71975

- EUP

- 1 was issued to Northwest Agricultural products for experimental use on a total of 1,020 acres in Oregon and Washington states . This permit is effective from March 1 , 2012 to March 1, 20 15.

In S eptember 2009

, EPA completed a F inal R egistration R eview D ecision for Pseudomonas fluorescens , Registration Review Case 6006 ( http://www.regulations.gov/#!document Detail;D=EPA

- HQ

- OPP

- 2007

- 0567

- 0014 )

. In its decision, EPA concluded that the standards of R egistration R eview were met for the Pseudomonas fluorescens and identified no general risk concerns . This strain will now be considered as part of the Pseudomonas fluorescens Registration Review Case 6006

.

Th e EP containing Pseudomonas fluorescens strain D7 is proposed for use in field applications for suppression of d owny b rome (cheatgrass), m edusahead, Japanese b rome, and j ointed

g oatgrass on fields of turf and grasses grown for seed, alfalfa

, wheat, barley, triticale, oat

, and rangeland. The proposed EP label contains applications as a seed treatment and also by ground or aerial spray

. The label directs the user to apply the EP before weed seeds germinate in the spring or autumn when daytime temperatures are below 50 o F and just prior to expected rainfall . It can be applied up to 4 times per year for a maximum annual application of 8g/acre.

EPA s cientists reviewed product analysis, toxicology, and nontarget org anism data and information ( 40 CFR §§ 158.2120, 158.2140, and 158.2150, respectively) submitted to su pport the registration of the proposed pesticide product s

. They found that, o verall, such data and information are adequate for risk assessment purposes, f ulfill the current microbial pesticide data requirements, and support registration of the product under FIFRA section 3(c)(5)

.

Product Analysis

For the purposes of FIFRA section 3(c)(5) registration, the product analysis data requirements

for the TGAI (a lso the manufacturing use product) and EP were fulfilled by acceptable guideline

studies. These data requirements include product chemistry and composition, analysis of samples, physical and chemical characteristics

.

Toxicology

The applicant submitted a dequate mammalian toxicology data and information to support the pesticide products. A n acute injection toxicity/pathogenicity study submitted with the original application and conducted with high dosages of Pseudomonas fluorescens strain D7

, showedPseudomonas fluorescens s train D7 Page 5 of 29

Biopesticides Registration Action Document

adve rse effects including mortality, abnormal clinical signs

, transient weight loss with resultant decreased absolute body weight, abnormal gross necropsy findings, and altered organ weights

. This injection study was conducted at an inappropriate excessive dose

. T he applicant repeated

the acute injection study at an appropriate lower dose with additional co ntrols to account for

ob served results

. This repeated injection toxicity/pathogenicity stud y along with the a cute oral

and pulmonary toxicity/pathogenicity studies

, showed that Pseudomonas fluorescens strain D7 is

not toxic or p athogenic via thes e route s of exposure

. In l ight of the results of the acute toxicity/pathogenicity data , EPA did not require testing at higher tiers (i.e., Tiers II and III).

Tolerance Exemption

In its application to the EPA in 2012, Ag

- Chem Consulting on behalf of Nort hwest Agricultural

Products provided a petition, data and other information to support an exemption from the requirement of a tolerance for residues of Pseudomonas fluorescens strain D7 when used in or on growing crops and rangeland.

Occupational Expos ure

Despite the low toxicological profile of residues of the active ingredient, Pseudomonas fluorescens strain D7 , in or on all agricultural commodities

, the EPA is requiring baseline personal protective equipment (PPE) for handlers that, due to their occ upation

, may be subject to prolonged or repeated exposure to the active ingredient

. The label on D7

, the end

- use product,

directs h andlers working with Pseudomonas fluorescens strain D7 i n agricultural settings to wear a long

- sleeved shirt, long pants, soc ks, shoe s, waterproof gloves

, and 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

.

Nontarget Organisms

P. fluorescens occurs naturally in soil and water (P allerroni, 2005), and strain D7 is a root

associated bacterium that was isolated originally from the West Coast region of the United

States . S oil

- dwelling nontarget organisms in those areas are expected to currently have some exposure to this bacterium. T he nontarget organism data requirements have been adequately

addressed f

o r the purposes of FIFRA section 3(c)(5) registration, based on the submitted data, rationales, and limited anticipated exposure to nontarget organisms

. The specific findings for

each nontarget organism are addressed in Section IV below.

Public Participation

On October 1, 2009, the EPA announced a new policy to provide a more meaningful opportunity for the public to participate i n major registration decisions before they occur. Accord ing to this policy, the EPA intends to provide a public comment period prior to making a registration decision for, at minimum, the following types of appli cations: new active ingredients; first food uses; first outdoor uses; first residential use s; or any other registration actions for which the Agency believes there may be significant public interest

.

Consistent with the policy of making registration actions more transparent, the pesticide product containing Pseudomonas fluorescens strain D7 is subject to a 15

- day comment period prior to registration of the manufacturing and end

- use products . In addition to containing a new activePseudomonas fluorescens s train D7 Page 6 of 29

Biopesticides Registration Action Document

ingredient, the registration of the pesticide product would result in the first outdoor and food uses

for Pseudomonas fluore scens strain D7 . The docket identification number associated with these registration actions

, and accessed through either http://www.regulations.gov or http://www.epa.gov/pesticides/regulating/registration

- status.html , is EPA

- HQ

- OPP

- 201 3

- 0 570

. The following documents are being made available for comment in EPA

- HQ

- OPP

- 201 3

- 0 570 : (1) draft Pseudomonas fluorescens strain D7 Registration Action Document (BRAD); (2) draft

label for the manufacturing

- use product

, Fluorescens Technical (File Symbol 71975

- G ), and ( 3 ) draft label for the end

- use pesticide product, D7 (EPA File Symbol 71975

- U ).

Registration Decision

W hile a final decision on registration is contingent upon review and consideration of public comments, EPA presently believes that the risk assessment and information submitted support registration of the Fluorescens Technical (TGAI) and the D7 EP. The basis for this preliminary

d ecision can be found in the risk assessment for Pseudomonas fluorescens strain D7 which is characterized throughout this BRAD

.

II. ACTIVE INGREDIENT OVERVIEW

Biological Name: Pseudomonas fluorescens strain D7

Culture Deposit: Agricul tural Research Service Culture Collection NRRL B

- 18293

OPP Chemical Code: 107200

Type of Pesticide: Mi crobial Pesticide

– Herbicide

See Appendix B for specific information (e.g., use sites, application rate s , methods of

app lication, formulation type s , and target pests) regarding the pesticide products containing this active ingredient.

III. REGULATORY BACKGROUND

A. Applications for Pesticide Product Registration

On October 5, 2012, Ag

- Chem Consulting (address: 12208 Quin que Lane, Clifton, VA 20124) on behalf of Northwest Agricultural Products (address: 821 South Chestnut Avenue, Pasco, WA

99301 ) submitted application s for a new TGAI, Fluorescens Technical (EPA File Symbol 71975

- G), and a new EP

, D7 (EPA File Symbol 71975

- U )

, to the EPA under section 3 of FIFRA.

On October 30 , 201 3 ( 7 8 FR 64937 ) , EPA announced receipt of the application to register a pesticide product containing a new active ingredient for use in agriculture (outdoors)

. EPA opened a 3 0

- day public comment period pursuant to the provisions of FIFRA section 3(c)(4). No

comme nts were received following this publication.

B. Food Tolerance Exemption

Concurrent with its registration applications and under Federal Food, Drug, and Cosmetic Act (FFDCA) section 408(d), Northwest Agricultural Products submitted a petition to establish an exemption from the requirement of a tolerance for residues of the pesticide active ingredient,Pseudomonas fluorescens s train D7 Page 7 of 29

Biopesticides Registration Action Document

Pseudomonas fluorescens strain D7 [Pesticide Petition (PP) 2F8103 ] in or on all foo d

commodities

. EPA published a Notice that Northwest Agricultural Products filed a petition to establish an exemption fr om the re qui re m e nt of a tol e r a n c e f or r e sidu es of Pseudomonas fluorescens strain D7 in or on growing crops and rangeland in the Federal Register of September
12, 2013 (78 FR 56185) and opened a 30

- day comment period. One comment was received in

the docket for the petition for Pseudomonas fluorescens strain D7 tolerance exemption that expressed general opposition to the sale of pesticides, and to their negative effects on bees and human beings. The comment was not specific to Pseudomonas fluorescens strain D7.

IV. RISK ASSESSMENT

In the Federal Register of October 26, 2007 ( 72 FR 61002 )

, the EPA issued a Final Rule on the data requirements to support registration of microbial pesticides and updated the definition for microbial pesticides

. The rule became effective on December 26, 2007. The data and information

ev aluated for this BRAD were considered in light of these requirements.

The EPA classifies each data submission with an indication of the usefulness of the information contained in the documents for risk assessment. A rating of “ acceptable ” indicates the st udy 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 : upgradeable ”). If a study

is rated as “supplemental: upgradeable,” EPA 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 wi ll often state that t he study is not required by 40 CFR P art 158. Both

“ acceptable” and “supplemental ” studies may be used in the risk assessment process as appropriate. An “unacceptable ” rating indicates that new data must be submitted.

For the acu te tox icity data requirements, Toxicity C ategories are assigned based on the hazard(s) identified from studies and/or other infor mation submitted to the EPA in support of a pesticide registration. The EPA classifies t he active ingredient or particular product i nto Toxicity Category I, II, III, or IV, where Toxicity Category I indicates the highest toxicity and Toxicity Category IV indicates the lowest toxicity (see 40 CFR § 156.62 )

.

A. Product Analysis Assessment ( 40 CFR § 158.2120 )

For purposes of registration under FIFRA section 3(c)(5), the EPA determined that all product analysis data requirements for Pseudomonas fluorescens strain D7 have been fulfilled. Refer to

T ables 1, and 2, in Appendix A for a summary of the data requirements, including both generic and product

- specific information.Pseudomonas fluorescens s train D7 Page 8 of 29

Biopesticides Registration Action Document

B. Human Health Assessment ( 40 CFR § 158.2140 )

1. Toxicity

All a ppli ca ble m a mm a li a n to x i c ology d a ta re qui r e m e nts suppo r ting the r e q u e st f or a n e x e mption

fr o m the re qui re m e nt of a tol er a n c e f or r e sidu e s of Pseudomonas fluorescens strain D7 i n or on

a ll f ood c ommoditi e s h a ve b e e n f ul f ill e d with d a ta submitt e d b y the applicant or scientific

rationale. Furthermore, Tier II and Tier III studies were not required for Pseudomonas fluorescens strai n D7 because of the lack of acute toxicity/pathogenicity in the Tier I studies.

The to x i c i t y and pathogenicity t e sts ( d er m a l, to x i c i t y ) a nd i r r it a tion t e sts (ac ute e y e a nd

p r im a r y d er m a l i rr it a tion) th a t a d d re ss pot e nti a l r out e s of e x posu r e to the ac tive i n g r e di e nt a r e

a ll c l a ssi f i e d in To x i c i t y C a t e g o r y I V ( s e e below or U.S. EPA 2014a and b) a nd re v ea l e d little

to no to x i c i t y a tt r ib u t e d to Pseudomonas fluorescens strain D7

. F in a l l y
, the applicant re po r t e d

th a t no h y p e r s e nsitivi t y in c id e n t s o cc u rr e d du r i n g the r e s earc h, d e v e lopm e nt, a nd t e sting of this active ingredient.

The ov e r a ll c on c lusions f r om a ll to x i c olo g i ca l in f

o r m a tion submitt e d b y the applicant ar e b r i ef l y d e s cr ib e d b e low, in sections IV(B)(1)(a) and IV(B)(1)(b) (see pages 8

– 1 1 ), and summarized in

Table 3 in Appendix A

.

a. Acute Toxicity/Pathogenicity

– Tier I

Acute Injection Toxicity/Pathogenicity

– Rat ( OCSPP Guideline Number 885.3200 ) (MRID 49349701 ) A study was repeated to re

- evaluate the acute intravenou s toxicity infectivity and pathogenicity, of a microbial pest control agent (MPCA), Pseudomonas fluorescens D7, at a single exposure of

10 6 and 10 7 CFU/rat doses. The MPCA test substance, inactivated MPCA, or sterile filtrate was injected into the tail vei n of one of four groups of rats. A control group (untreated) was conducted concurrently. The animals were observed frequently on day of dosing for mortality and signs of pharmacological and/or toxicological effects and once daily thereafter for 21 days.

Ti ssue and blood samples taken at interim sacrifices from the group receiving the active test substance were cultured to provide quantitative measurements of the test microbe's clearance pattern. There was no mortality in any group during the study. During o bservations for clinical signs, all animals appeared normal for the duration of the study. The gross necropsy conducted at termination of the study revealed no observable abnormalities. The test organism had cleared completely from Groups IV and V (MPCA) b lood, kidneys, mesenteric lymph nodes, lungs, and brain, liver, spleen and cecum contents by Day 7. After two consecutive interim sacrifices showed no growth of test organism in the organs plated, no further tissue samples were taken.

The test substance Ps eudomonas fluorescens D7 was determined to be non

- toxic and not pathogenic or infective to rats when injected intravenously at a dose of 3.4 x 10 7 CFU/rat or at
3.7 x 10 6 CFU/rat. EPA determined this study to be acceptable

.

This study (MRID 49349701) add resses EPA recommendations to repeat an acute injection toxicity study using a lower dose of the microorganisms that includes additional controls such as filtered culture medium, and autoclaved medium to account for the observed injection results.

This st udy supersedes the p reviously submitted acute injection toxicity/pathogenicity in rat study (MRID 48966402) that tested Pseudomonas fluorescens strain D7 and showed adverse effects on

rats following a single intravenous injection administration of 1.3 x 10 8 CFU/rat , includingPseudomonas fluorescens s train D7 Page 9 of 29

Biopesticides Registration Action Document

mortality, abnormal clinical signs; transient weight loss, abnormal gross necropsy findings, and altered organ weights.

Acute Oral Toxicity/Pathogenicity

– Rat ( OCSPP Guideline Number 885.3050 ) (MRID 48966403) Sprague

- Dawley rats , 8 week old, (12/sex) were given a single oral dose of MPCA

, Pseudomonas fluorescens strain D7

, of 1.3 X 10 9 CFU/animal. The animals were observed three times on day 0 after dosing and daily through day 21 with interim scheduled sacrifices on Days
3, 7, a nd 14. Five males and five females were treated with autoclaved Pseudomonas fluorescens strain D7 as inactive MPCA controls, two untreated animals per sex were used as “shelf controls,” and two animals per sex were designated as untreated controls.

No ob servable abnormalities were found during observations for clinical signs or at necropsy, and there were no toxicologically relevant differences between the body weights of the treated animals and those of controls. Relative to their respective controls, M PCA

- treated females had increased relative liver weights (+16.0%; p<0.05), and MPCA

- treated males had increased relative spleen weights (+16.7%; p<0.05), and these differences correlated with increased mean absolute weights of these same organs (+11.7% and +27.1% for MPCA

- treated female absolute

liver and MPCA

- treated male absolute spleen weights, respectively). No CFUs (or in one instance a single individual count of <100 CFU/g) were recovered from the brain, lungs, liver, kidneys, or lymph nodes of any a ctive

- treated animal. Pseudomonas fluorescens strain D7 was found in feces, urine, and cecum contents of MCPA

- treated animals and appeared to be completely cleared by day 14 following oral administration to rats. Based on the results of this study

, Pseud

omonas fluorescens strain D7 does not appear to be toxic, infective, and/or pathogenic in rats when dosed at 1.3 X 10 9 CFU/animal. The EPA rated this study as acceptable.

Acute Pulmonary Toxicity/Pathogenicity

– Rat ( OCSPP Guideline Number 885.3150 ) (MRI D 489664

- 04) Groups of 12 week old Sprague Dawley rats (3/sex/Group) were exposed by the intratracheal

route to Pseudomonas fluorescens strain D7 in sterile PBS at a constant dose volume of 0.1 mL/

animal and a dose of 4.6 x 10 8 cfu/animal. Eight males an d eight females were treated with

autoclaved Pseudomonas fluorescens strain D7 as autoclaved controls; five males and five females were not treated and used as untreated control; and four males and four females were not

treated and used as shelf controls. The animals were then observed for up to 21 days, with interim sacrifices on day 0 (all groups) and days 3, 7, and 14 (active

- MCPA group).

There were no test substance related clinical signs, gross necropsy findings, or differences in organ weights. Two males in the active test material treated group sacrificed on day 3 lost weight and one female in autoclaved test material treated group sacrificed on day 21 did not gain weight during the first week but gained weight by day 14. All other animals gain ed weight prior to scheduled sacrifice. The test organisms were not seen or were present at < 100 cfu/g in lungs from the animals sacrificed after dosing on day 0. The test organisms were not seen or were present at < 100 cfu/g in blood, brain, lungs, sp leen, liver, kidneys, mesenteric lymph nodes, and cecum content removed from animals sacrificed on days 3 and 7. Based on these results, Pseudomonas fluorescens strain D7 does not appear to be toxic, infective, and/or pathogenic in

rat when dosed at 4.6 x 10 8 cfu /animal . The EPA rated this study as acceptable.Pseudomonas fluorescens s train D7 Page 10 of 29

Biopesticides Registration Action Document

Acute Dermal Toxicity/Pathology

– Rabbit ( OCSPP Guideline Number 870.1200 ) ( MRID 489664

- 05) Five male and five female New Zealand White rabbits, 16

- 18 weeks old, were dermally exposed

to a 2000 mg/ kg bw dose of Pseudomonas fluorescens strain D7 moistened with 2.0 mL of deionized water/g test substance for 24 hours to an area of approximately 10% of body surface area. Following exposure, the animals were observed for a period of 14 days. All animals survived and had no abnormal systemic clinical signs during the study. Very slight to well

defined Erythema was noted on all dose sites on day 1 and resolved by day 4. With the exception of one female that lost weight during the second week, all animals g ained weight throughout the study. No observable abnormalities were found at necropsy. The dermal LD 50 for males was greater than 2000 mg/kg bw; for females was greater than 2000 mg/kg bw; and for both sexes combined was greater than 2000 mg/kg bw. Base d on the results of this study, Pseudomonas fluorescens strain D7 is of low toxicity

. EPA classified the acute dermal toxicity

of Pseudomonas fluorescens strain D7 as Toxicity Category IV and rated this study as acceptable.

Primary Eye Irritation

– Rabbi t ( OCSPP Guideline Number 870.2400 ) (MRID 49100801) 100 mg of undiluted Pseudomonas fluorescens strain D7 (Batch No. 201109210301 ; purity 95%; pH not reported) was instilled as supplied into the conjunctival sac of the right eye of three male

and three female New Zealand White rabbits. Untreated left eyes served as controls. Animals were observed at 1, 24, 48, and 72 hours after test material instillation. Irritation was scored by the method of Draize and classified by the system of Kay and Calandra.

No corneal opacity, iritis, or conjunctival irritation was noted on any rabbit throughout the study. Pseudomonas fluorescens strain D7 was not irritating to the eye and EPA classified the acute eye irritation as Toxicity Category IV for primary eye irrita tion. The EPA rated this study as acceptable.

Primary Dermal Irritation

– Rabbit ( OCSPP Guideline Number 870.2500 ) (MRID 49100802 ) Three male and three female New Zealand White rabbits were dermally exposed to 500 mg of Pseudomonas fluorescens strain D7 moistened with 1.0 mL of DI water for 4 hours on an approximately 2.5 x 2.5 cm area of the body surface. The animals were observed at 1, 24, 48, and 72 hours after patch removal. Irritation was scored by the method of Draize. No dermal

irritation was no ted on any animal during the study. The primary irritation index was 0.0. Pseudomonas fluorescens strain D7 was not irritating and EPA classified the dermal irritation toxicity as Toxicity Category IV. The EPA rated this study as acceptable.

H y p e rs e nsit i v ity in c id

e nts ( OCSPP Guideline Number 885.3400 )

The applicant re po r t e d th a t no h y p er s e n sitivi t y in c id e nts, in c lud i ng imm e di a t e

- t y p e or d e l a y e d

t y pe re a c tions of h u m a ns a nd dom e stic a nim a ls, o c c u rre d d u r ing re s e a r c h, d e v e lopm e nt, or

t e sti n g of Pseudomonas fluorescens strain D7

b. Acute Toxicology and Subchronic Toxicity/Pathogenicity

– Tier II; Reproductive

, Fertility Effects, Carcinogenicity, Immunotoxicity, and Infectivity/Pathogenicity Analysis

– Tier III

Tier II and Tier III studies were not requir ed for Pseudomonas fluorescens s train D7 based on the lack of acute toxicity/pathogenicity in the Tier I data requirements.Pseudomonas fluorescens s train D7 Page 11 of 29

Biopesticides Registration Action Document

c. Endocrine Disruptors

As required by the Administrator under the Federal Food, Dru g, and Cosmetic Act ( FFDCA )

, the EPA has developed the Endocrine Disruptor Screening Program (EDSP) and has begun to implement the screening program that is to be used to test all pesticides to determine whether certain substances (including pesticide acti ve and other ingredients) may have an effect in humans or wildlife similar to an effect produced by a “naturally occurring estrogen, or other such endocrine effects as the Administrator may designate.” FFDCA section 408(p)(4) authorizes the Administrator, by order, to exempt from the requirements of the Endocrine Disruptor Screening Program a biologic substance or other substance if a determination is made that the substance is anticipated to not produce any effect in humans similar to an effect produced by a naturally occurring estrogenic substance. Between October 2009 and February 2010, EPA issued test

orders/data call

- ins for the first group of 67 chemicals, which contains 58 pesticide active ingredients and 9 inert ingredients. Pseudomonas fluorescens s train D7 is not among the group of 58 pesticide active ingredients on the initial list to be screened under the EDSP.

The EPA believes that for Pseudomonas fluorescens s train D7 is a substance that would not produce any effect in humans similar to an ef fect produced by a naturally occurring estrogenic substance. As such, and pursuant to Section 408(p)(4), the EPA will determine in the future whether it can exempt for Pseudomonas fluorescens s train D7 from the requirements of the Section 408(p)(4) EDSP. I n the event the EPA does determine to exempt this substance from the EDSP, an order will be issued. For further information on the status of the EDSP, the policies and procedures, the list of 67 chemicals, the test guidelines and the Tier 1 screening batte ry, please visit our website: http://www.epa.gov/endo/

.

2. Federal Food, Drug, and Cosmetic Act (FFDCA) Considerations

Section 408(c)(2)(A)(i) of FFDCA allows the EPA to establish an exemption from the requireme nt for a tolerance (the legal limit for a pesticide chemical residue in or on a food) only

if the EPA determines that the exemption is “safe.” Section 408(c)(2)(A)(ii) of FFDCA defines “safe” to mean that “there is a reasonable certainty that no harm will result from aggregate exposure to the pesticide chemical residue, including all anticipated dietary exposures and all other exposures for which there is reliable information.” This includes exposure through drinking water and in residential settings

, but i t does not include occupational exposure. Pursuant to section 408(c)(2)(B) of FFDCA, in establishing or maintaining in effect an exemption from the requirement of a tolerance, the EPA must take into account the factors set forth in section 408(b)(2)(C) of FFDCA, which require the EPA to give special consideration to exposure of infants and children to the pesticide chemical residue in establishing a tolerance exemption

, and to “ensure that there is a reasonable certainty that no harm will result to infants and children from aggregate exposure to the pesticide chemical residue....” Additionally, section 408(b)(2)(D) of FFDCA requires that the EPA consider “available information concerning the cumulative effects of [a particular pesticide's] . . . residues and other substances that have a common mechanism of toxicity.”

The EPA performs a number of analyses to determine the risks from aggregate exposure to pesticide residues. First, the Agency determines the toxicity of a pesticide . Second, the EPA examines exp osure to the pesticide through food, drinking water, and through other exposures that occur as a result of pesticide use in residential settings. Consistent with section 408(b)(2)(D)Pseudomonas fluorescens s train D7 Page 12 of 29

Biopesticides Registration Action Document

of FFDCA, the EPA has reviewed the available scientific data and other r elevant information, and considered its validity, completeness, and reliability and the relationship of this information

to human risk. The Agency also considered available information concerning the variability of the sensitivities of major identifiable s ubgroups of consumers, including infants and children. Based on the acute toxicity/pathogenicity data and information discussed previously and presented in Table 3 in Appendix A , t he data required for a FFDCA risk assessment fo r Pseudomonas fluorescens s train D7 have been fulfilled.

a. Aggregate Exposure

In examining aggregate exposure, section 408 of FFDCA directs EPA to consider available information concerning exposures from the pesticide residue in food and all other nonoccupational 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).

Food Exposure and Risk Characterization : The EPA found that incr eased d ietary exposure to Pseudomonas fluorescens s train D7

, a naturally occurring bacterium

, is anticipated to be negligible

. For the proposed use of Pseudomonas fluorescens strain D7 as an herbicide, the applications are made before crop plants emerge, o r as seed treatment

, and consequently oral

exposure to residues from such use is expected to be minimal. Pseudomonas fluorescens s train

D7 and other closely related Pseudomonas fluorescens bacteria already exist in the soil

environment

. The EPA concluded that t he risk posed to adults, infants, and children is likely to be minimal because of the low acute oral toxicity/ p athogenicity potential of Pseudomonas fluorescens strain D7

.

Drinking Water Exposure and Risk Characterization : Exposure to residues of Pseudomonas fluorescens strain D7 in consumed drinking water is not likely to increase because there are no use sites for the pesticide with direct applications to water. There is a possibility of spray drift from aerial applications or runoff of prepared fields and rangelands into surface waters. G round water is not expected to have significant exposure to Pseudomonas fluorescens strain D7 since,

like other microorganisms, this microbial pesticide would likely be filtered out by the particulate

nature of many soil types. If it were to be transferred to surface or ground waters that are intended for eventual human consumption (e.g., through spray drift or runoff) and directed to wastewater treatment systems or drinking water facilities, it likely would not survive the conditions water is subjected to in such systems or facilities, including chlor ination, pH adjustments, filtration , and/or occasionally high temperatures (Centers for Disease Control and Prevention 2009; U.S. EPA 2004 , and DeFelice et.al., 1993 ). In the remote likelihood that Pseudomonas fluorescens strain D7 is present in drinking water (e.g., water not subject to treatment systems or facilities), its target pest specificity and available data indicate no toxicity and/or pathogenicity is likely to occur with any drinking water exposure to Pseudomonas fluorescens strain D7 that results from pesticide application s made in accordance with good agricultural practice s (see section IV(B)(1)(a) on pages 8

– 10 and Table 3 in A ppendix A ).

Non

- occupational, Residential Risk Characterization : Given that Pseudomonas fluorescens

strain D7 use sites do n ot include residential settings and because the bacterium is naturally

occurring

, EPA determined that non

- occupational exposure to the bacterium is unlikely

. Repeated exposure s to the Pseudomonas fluorescens strain D7 microorganism from pesticidal applications do not exceed EPA’s level of concern, particularly in light of available data thatPseudomonas fluorescens s train D7 Page 13 of 29

Biopesticides Registration Action Document

demonstra te Pseudomonas fluorescens strain D7 is not toxic (acute dermal toxicity and acute pulmonary toxicity/ pathogenicity), is non

- irritating (primary dermal irritation), and is not pathogenic when used as labeled in accordance with good agricultural practices (acute pulmonary toxicity/pathoge nicity , and acute injection toxicity/pathogenicity ) (see section IV(B)(1)(a) on pages 8

– 1 1 and Table 3 in Appendix A ).

b. Cumulative Effects from Substances with a Common Mechanism of Toxicity

Section 408(b)(2)(D)(v) of FFDCA requires that, when considering whether to establish, modify, or revoke a tolerance exemption, the EPA consider “available information concerning the cumulative effects of [a particular pesticide’s] . . . residues and other substances that have a

com mon mechanism of toxicity.” The EPA has not found Pseudomonas fluorescens strain D7 to share a common mechanism of toxicity with other substances. Pseudomonas fluorescens strain

D7 does not appear to be toxic to humans via dietary, dermal and pulmonary ex posure. For the purposes of the tolerance action, therefore, the EPA has assumed that Pseudomonas fluorescens

strain D7 does not have a common mechanism of toxicity with other substances. Thus, section 408(b)(2)(D)(v) of the FFDCA does not apply. For infor mation regarding the EPA’s efforts to determine which chemicals have a common mechanism of toxicity and to evaluate the cumulative effects of such chemicals, see the EPA’s website at http://www.epa.go v/pesticides/cumulative

.

c. Determination of Safety for the United States Population, Infants and Children

In considering the establishment of a tolerance or tolerance exemption for a pesticide chemical residue, FFDCA section 408 (b) (2) (C) provides that the 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) provides that the EPA shall apply an additional tenfold (10X) margin of exposure ( safety ) for infants and children in the case of threshold effects to account for prenatal and postnatal toxicity and the completeness of the database on toxicity and exposure

unless the EPA determines that a different margin of exposure ( safety ) will be safe for infants and children. This additional margin of exposure ( safety ) is commonly referred to as the Food Quality Protection Act Safety Factor. In applying this provision, the EPA either retains the default value of 10X or uses a different additional safety factor when reliable data available to

the EPA support the choice of a different factor.

Based on the acute toxicity an d pathogenicity data/information discussed in section IV(B)(1)(a) (see pages 8

– 1 1 ) and Table 3 in Appendix A

, the EPA concludes that there are no threshold

effects of concern to infants, children, or adults when Pseudomonas fluorescens strain D7 is used as labeled in accordance with good agricultural practices. As a result, the EPA concludes that no additional margin of exposure (safety) is necessary to protect infants and childre n and that not adding any additional margin of exposure (safety) will be safe for infants and children.

Moreover, based on the same data /information and EPA analysis as presented directly above, the Agency is able to conclude that there is a reasonable ce rtainty that no harm will result to the

United States population, including infants and children, from aggregate exposure to the residues

of Pseudomonas fluorescens strain D7 when it is used

— as labeled and in accordance with good agricultural practices

— as a n herbicide . Such exposure includes all anticipated dietary exposuresPseudomonas fluorescens s train D7 Page 14 of 29

Biopesticides Registration Action Document

and all other exposures for which there is reliable information. The EPA has arrived at this conclusion because, considered collectively, the data and information available on Pseudomon as fluorescens strain D7 do not demons trate toxic or pathogenic potential to mammals, including

infants and children.

3. Occupational Exposure and Risk Characterization

The EPA does not expect h andler exposure to Pseudomonas fluorescens strain D7 to po se any undue risk. Regardless, appropriate personal protective equipment (PPE) and precautionary statements are required on pesticide product labels to mitigate any potential risks to pesticide handlers due to prolonged or numerous exposures. Handlers appl ying D7 in agricultural settings

must wear a dust/mist filtering respirator , waterproof gloves, long

- sleeved shirt, long pants,

socks and shoes.

4. Human Health Risk Characterization

The EPA considered human exposure to Pseudomonas fluorescens strain D 7 in light of the standard for registration in FIFRA and the relevant safety factors in FFDCA. The Agency

determin ed that no unrea sonable adverse effects to the United States population in general, and to infants and children in particular, will result whe n products containing Pseudomonas fluorescens strain D7 are used in accordance with EPA

- approved labeling.

C. Environmental Assessment ( 40 CFR § 158.2150 )

Four original guideline studies with the TGAI and three sets of data waiver rationale were submitted to meet data requirements for nontarget organisms per 40 CFR Part 158.2150 for Pseudomonas flu orescens strain D7 , which are sufficient to fulfill the relevant microbial pesticide data requirements and for risk assessment purposes

. Further testing of nontarget organisms at higher tier levels (i.e., Tiers I I, III, and IV) is not required. Adverse eff ects to nontarget organisms, including federally listed threatened and endangered species, are not expected to result from the proposed registration of P. fluorescens strain D7 when applied in

accordance with the directions o n the proposed label.

For a summary of the generic data requirements described in sections IV(C)(1) (see pages 14

– 1 5 ) , refer to Table 4 in Appendix A

.

1. Ecological Exposure and Risk Characterization

According to the proposed label for the EP, P. flu orescens strain D7 is intended for use in suppression of downy brome, medusahead, Japanese brome, and jointed goatgrass on wheat, barley, triticale, oats, and rangeland. The EP containing 95% P. fluorescens strain D7 is a freeze

dried powder that is to be mixed in water and applied as a direct spray to the soil surface at a rate of 2 g /acre (4 x 10 11 cells/acre). The label directs the user to apply the EP before weed seeds germinate in the spring or autumn when daytime temperatures are below 50 o F. The EP i s to be applied just prior to expected rainfall, and can be applied up to 4 times per year for a maximum annual application of 8g/acre. The EP may also be used as a seed treatment to control the listed weeds at planting.Pseudomonas fluorescens s train D7 Page 15 of 29

Biopesticides Registration Action Document

The proposed EP label indicates th at the product is applied by spray (except for seed treatments), and may be applied using aerial equipment. Aerial sprays are generally expected to have greater potential for nontarget exposure compared to other application methods (e.g., ground sprays,

sp rays using hand

- held equipment). However, because the EP is to be applied directly to the soil surface, in autumn or spring when temperatures are cooler, and at preemergence, the potential exposure of nontarget organisms to the EP is expected to be more li mited. Spray drift from aerial applications may result in some exposure to nearby areas, including aquatic environments. However, based on the Tier I aerial agricultural application scenario in the AgDRIFT spray drift model, assuming fine to medium droplet size, a maximum of 12.5% of the amount applied is expected to reach aquatic areas at the water’s edge. P. fluorescens also occurs naturally in soil and water (Pallerroni, 2005 ). As discussed above, strain D7 is a root

- associated bacterium which

was isolat ed originally from the West Coast region of the United States, so soil

- dwelling nontarget organisms in those areas currently have some exposure to this bacterium

.

Birds and Mammals

The guideline study submitted for the avian oral toxicity/pathogenicity testing requirement

showed no adverse effects in the bird species tested (Northern bobwhite, Colinus virginianus ). P. fluorescens strain D7 also is not likely to grow at a bird’s high body temperatures (see McNab, 1966). Based on expected relatively low e xposure and lack of adverse effects observed in birds exposed to P. fluorescens strain D7, risk to nontarget birds resulting from the proposed registration is minimal

.

An acute oral toxicity/pathogenicity study with laboratory rats indicated no adverse ef fects of P

. fluorescens strain D7 when administered at a dose of 1.3 X 10 9 CFU/animal (MRID No. 48966403)

. As discussed above, other studies with laboratory mammals also showed no adverse effects upon exposure to P

. fluorescens strain D7 (USEPA, 2014a; 20 14b). Some wild mammals may be exposed to P. fluorescens strain D7 in treated areas, but the exposure of most wild

mammals is expected to be limited because of the root

- borne nature of P. fluorescens strain D7 (i.e., it is a rhizobacterium), application ti ming (e.g., pre

- emergence) and type of use (e.g., applied to soil surface). Since adverse effects were not observed in the mammalian studies, risk to wild mammals is expected to be low from the proposed registration of the new a.i.

Nontarget Insects and Honey Bees

The guideline study submitted to meet the requirement for nontarget insect testing did not show toxicity or pathogenicity to ladybird beetles ( Hippodamia convergens ) upon a 7

- day dietary exposure to corn earworm ( Helicoverpa zea ) sprayed with P. fluorescens strain D7 at 1 x 10 6

cfu/ml. The applicant also provided a study with honey bees to support a previous Experimental

Use Permit for P. fluorescens D7 (MRID No. 48668301) which showed no adverse effects at a

dose of 1 x 10 7 cfu/ml over seven days. This study was classified as supplemental due to the short duration (USEPA, 2012), so the applicant submitted rationale (MRID No. 49177701) to address the honey bee data requirement for the proposed section 3 registration of P. fluorescens

strain D7. The rationale referenced the former study and also cited a literature study (Johnson et al., 1993) that involved honey bees as a vector for the dispersal of another strain of this microorganism, Pseudomonas fluorescens strain A506, to apple and pear blos soms without observable impact on honey bees.Pseudomonas fluorescens s train D7 Page 1 6 of 29

Biopesticides Registration Action Document

Exposure to nontarget insects and honey bees may occur in the treated areas. However, under the recommended application conditions (cool, below 50oF, and wet), insects including honey bees are not likely ac tive, thus no significant exposure of these insects to P. fluorescens strain D7 is expected.

Based on the above information (no hazard and low exposure), adverse effects to nontarget honey bees and insects are not likely from the proposed registration of the new a.i.

Nontarget Plants

P. fluorescens occurs naturally in soil and water. P. fluorescens strain D7, the a.i., was isolated initially from the West Coast region of the United States. No adverse effects of the a.i. on tested

dicots and most monoc ots were shown in greenhouse, growth chamber or field studies. Exposure to nontarget plants is expected with the proposed registration of P. fluorescens strain D7. Scientific rationale submitted for a waiver of the requirement for Nontarget Plant Testing is based on its selective mode of action on monocotyledonous weeds (particularly to Bromus spp.) and limited number of seasonal applications to soil surfaces, and the microorganism’s reported ecology. The rationale provided was sufficient to conclude that the proposed uses of P. fluorescens strain D7 to suppr ess certain weeds (downy brome, medusa head, Japanese brome, and join ted goat grass) is not expected to result in increased exposure to, or adverse effects in nontarget plants of economic importance

.

Freshwater Fish and Invertebrates

The studies submitted for the nontarget freshwater fish and invertebrate testing requirements showed no toxicity/pathogenicity of P. fluorescens strain D7 to Oncorhynchus mykiss (rainbow trout) and Daphnia mag n a when t hese aquatic animals were exposed to P. fluorescens strain D7. Some exposure to freshwater fish and invertebrates in nearby aquatic environments may occur with proposed applications of P. fluorescens D7, especially if the proposed treatments are applied in aerial sprays. However, as described above, the exposure of freshwater fish and invertebrates would be somewhat limited because of the type of use (e.g., applied directly to soil surfaces and before the rain) of P. fluorescens D7 and limited number of ap plications in autumn and spring, respectively, when it is cool (below 50 o C) and wet. Based on the studies submitted and anticipated low exposure to strain D7 sprayed onto soil surfaces, adverse effects on nontarget freshwater organisms are not likely.

M arine/Estuarine Fish and Invertebrates

The applicant did not submit a guideline study to meet the requirement for nontarget marine/estuarine fish and invertebrate testing, but submitted scientific rationale in lieu of testing. This testing is conditional ly required if significant exposure of nontarget marine/estuarine fish

and invertebrates to P. fluorescens strain D7 is expected. Similar to the nontarget organisms in fresh water environments, P. fluorescens strain D7 is not expected to reach marine/estua rine environments in significant quantities, thus significant exposure of the estuarine/marine fish and invertebrates to stain D7 is not likely as a result of the proposed registration of the a.i. Therefore,

adverse effects to marine/estuarine fish and inv ertebrates are not anticipated.Pseudomonas fluorescens s train D7 Page 17 of 29

Biopesticides Registration Action Document

2. Environmental Fate Data

As t he data and information provided are sufficient to fulfill the Tier I nontarget organism data requirements and allow for nontarget organism risk asse ssment for Pseudomonas fluorescens

s tra in D7

, f

urther testing at higher levels (Tiers II, III, and IV) is not required.

3. Threatened and Endangered Species Assessment

Since EPA has determined that no effects are anticipated for any nontarget species exposed to Pseudomonas fluorescens stra in D7 as a result of the proposed labeled applications, effects to federally

- listed threatened and endangered species and their designated critical habitats are also not expected. Therefore, a “No Effect” determination is made for direct and indirect effe cts to listed species and their designated critical habitats resulting from the proposed uses of Pseudomonas fluorescens strain D7 , as labeled.

V. ENVIRONMENTAL JUSTICE

The EPA seeks to achieve environmental justice

— the fair treatment and meaningful inv olvement 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, and commercial operations or the execution of federal, state, local, and tribal environmental programs an d 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. The EPA has this goal for all communities and persons across the United States.

To help address potential environmental justice issues, during the 15

- day public participation comment period, the EPA sought information on any groups or segments of the population who

, as a result their location, cultural practices, or other factors, may have atypical, unusually high

exposure to Pseudomonas fluorescens strain D7

, compared to the general population. The EPA received no public comments on this particular matter

.

For add itional information regarding environmental justice issues, please visit the EPA’s website

at http://www.epa.gov/compliance/environmentaljustice/index.html

.

V I

. RISK MANAGEMENT DECISION

Section 3(c)(5) of FIFRA permits for the registration of a pestici de provided that all the following determinations are made:

(1) Its composition is such as to warrant the claims for it; (2) Its labeling and other material required to be submitted comply with the requirements of FIFRA;Pseudomonas fluorescens s train D7 Page 18 of 29

Biopesticides Registration Action Document

(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, the Pseudomonas fluorescens s train D7 product s, the TGAI and EP, ha ve well

- known properties. The EPA has no knowledge that would contradict the claims made on the

label s
, and th e p esticide product s are not expected to cause unreason able adverse effects on the

environment when used according to label instructions. Criterion 2 is t he Agency believe s that

the Pseudomonas fluorescens s train D7 p esticide product s will not cause any unreasonable

adverse effects on the environment and based on cited field testing publications these products

offer biocontrol of certain weed species satisfying criterion 3. Criterion 4 is satisfied in that the Pseudomonas fluorescens s train D7 pesticide product s are not expected to cause unreasonable

adverse ef

fects when used according to label instructions. Therefore, both the MP and the EP,

D7

, which contain s Pseudomonas fluorescens s train D7 as a new active ingredient, are eligible for registration under FIFRA section 3(c)(5) for the labeled uses.

V II

. ACT IONS REQUIRED OF THE REGISTRANT

A. Final Printed Labeling

Before the manufacturing

- use product, Pseudomonas fluorescens s train D7 TGAI and the end

use product, D7 , may be released for shipment, the applicant is required to provide appropriate final prin ted labeling to the EPA

.

B. Terms of Registration

The EPA is not requiring additional information on the registration of these two products

.

C. Reporting of Adverse Effects and Hypersensitivity Incidents

Notwithstanding the information stated in th e previous sections, it should be clearly understood

that certain specific data must be reported to the EPA as a requirement for maintaining the federal registration for a pesticide product. Reports of all incidents of adverse effects to the

environment mu st be submitted to the EPA under the provisions stated in FIFRA section 6(a)(2). Additionally, all incidents of hypersensitivity (including both suspected and confirmed incidents) must be reported to the EPA under the provisions of 40 CFR § 158.2140(d).Pseudomonas fluorescens s train D7 Page 19 of 29

Biopesticides Registration Action Document

V III. GLOSSARY OF ACRONYMS AND ABBREVIATIONS

BRAD Biopesticides Registration Action Document

CFR Code of Federal Regulations

CFU colony forming units

cSt centistokes

EDSP Endocrine Disruptor Screening Program

EP end

- use pesticide product

EPA United State s Environmental Protection Agency (the “Agency”)

FFDCA Federal Food, Drug, and Cosmetic Act

FIFRA Federal Insecticide, Fungicide, and Rodenticide Act

FR Federal Register g/mL grams per milliliter lb/gal pounds per gallon

LD 50 median lethal dose. A statisti cally derived single dose that can be expected

to cause death in 50% of the test animals when administered by the route indicated (oral, dermal, or inhalation). It is expressed as a weight of substance per unit weight of animal (e.g., mg/kg). mg/kg m illigrams per kilogram

MP manufacturing

- use pesticide product

MPCA microbial pest control agent

MRID No. Master Record Identification Number

NIOSH National Institute for Occupational Safety and Health

OPP Office of Pesticide Programs

PC Code Pesticide Ch emical Code

PP Pesticide Petition

PPE personal protective equipment

TGAI technical grade of the active ingredient

U.S. United StatesPseudomonas fluorescens s train D7 Page 20 of 29

Biopesticides Registration Action Document

I X. BIBLIOGRAPHY

A . Studies Submitted to Support the Active Ingredient, Pseudomonas fluoresce ns strain D7

and Pesticide Product Registration s for the Fluorescens Technical and the EP

, D7

MRID No. Study Information

48497800 Northwest Agricultural Products (2011) Submission of Product Chemistry Data in

Support of the Experimental Use of Bacterium Pseudomonas fluorescens Strain D7 on Aggressive Weeds. Transmittal of 1 Study.

48497801 Brooks, M. (2011) D7, Product Chemistry, Group: Product Identity, Composition

and Analytical Test Guidelines and Group B: Physical and Chemical Properties Test Guidelines. Project Number: NAP01. Unpublished study prepared by NorthWest Agricultural Products. 21 p.

48524500 Northwest Agricultural Products (2011) Submission of Product Chemistry Data in

Support of the Experimen tal Use of D7. Transmittal of 1 Study.

48524501 Brooks, M. (2011) D7, Product Chemistry, Deposition of a Sample in a Nationally

Recognized Cullture Collection. Discussion of Formation of Unintentional Impurities. Analysis of Samples. Project Number: NAP02 . Unpublished study prepared by Ag

- Chem Consulting. 7 p.

48548901 Kennedy, A. (2011) Effect of Pseudomonas Strain D7 on Nontarget Vegetation.

Project Number: NAP003. Unpublished study prepared by Washington State University. 13p.

48668300 Northwest Agric ultural Products (2011) Submission of Toxicity Data in Support of

the Experimental Use of Pseudomonas fluorescens Strain D7. Transmittal of 1 Study.

48668301 Younger, C. (2011) Pseudomonas fluorescens D7: Honey Bee, Apis mellifera, Non

target Insect Micro bial Testing: Final Report. Project Number: 14619/10. Unpublished study prepared by Stillmeadow, Inc. 12p.

48724301 Brooks, M. (2012) Metabolite Analysis of Pseudomonas Strain D7. Project

Number: NAP04. Unpublished study prepared by Ag

- Chem Consulting, LL C. 33p.

48966400 Northwest Agricultural Products (2012) Submission of Product Chemistry and

Toxicity Data in Support of the Application for Registration of D7 and Fluorescens

Technical. Transmittal of 10 Studies.

48966401 Wend, C. (2012) Identification o f Pseudomonas fluorescens Strain D7. Project

Number: NAP20121004. Unpublished study prepared by Northwest Ag. Products. 27p.

48966402 Monds, K. (2011) Pseudomonas fluorescens D7: Intravenous Toxicity /

Pathogenicity Study in Rats: Final Report. Project Nu mber: 14620/10. Unpublished study prepared by Stillmeadow, Inc. 34p.Pseudomonas fluorescens s train D7 Page 21 of 29

Biopesticides Registration Action Document

48966403 Doig, A. (2012) Pseudomonas fluorescens Strain D7: Acute Oral Toxicity /

Pathogenicity Study in Rats: Final Report. Project Number: 15730/11. Unpublished study prepared by Still meadow, Inc. 29p.

48966404 Monds, K. (2012) Pseudomonas fluorescens Strain D7: Acute Pulmonary Toxicity /

Pathogenicity Study in Rats: Final Report. Project Number: 15728/11. Unpublished study prepared by Stillmeadow, Inc. 30p.

48966405 Monds, K. (2012) Pseudomonas fluorescens Strain D7: Acute Dermal Toxicity /

Pathogenicity Study in Rabbits: Final Report. Project Number: 15729/11. Unpublished study prepared by Stillmeadow, Inc. 12p.

48966406 Brooks, M. (2012) D7, Product Chemistry, Stability to Normal a nd Elevated

Temperatures, Metals and Metal Ions, Analysis of Samples, Storage Stability and Corrosion Characteristics. Project Number: NAP20121002. Unpublished study prepared by Northwest AG Product. 17p.

48966407 Younger, C. (2012) Pseudomonas fluorescen s Strain D7: Ladybird Beetle Non

target Insect Microbial Testing. Project Number: 16338/12. Unpublished study prepared by Stillmeadow, Inc. 11p.

48966408 Hartwell, T. (2011) Pseudomonas fluorescens Strain D7: Microbial Pest Control

Agent (MPCA) Freshwater Fish Test with Oncorhynchus mykiss (Rainbow Trout): Final Report. Project Number: 14622/10. Unpublished study prepared by Stillmeadow, Inc. 17p.

48966409 Younger, C. (2011) Pseudomonas fluorescens D7: Acute Oral Toxicity Study in

Bobwhite Quail: Final Re port. Project Number: 14716/10. Unpublished study prepared by Stillmeadow, Inc. 17p.

48966410 Hartwell, T. (2011) Pseudomonas fluorescens D7: Microbial Pest Control Agent

(MPCA) Freshwater Aquatic Invertebrate Test with Daphnia magna: Final Report. Projec t Number: 14623/10. Unpublished study prepared by Stillmeadow, Inc. 27p.

49021100 Northwest Agricultural Products (2012) Submission of Fate Data in Support of the

Application for Registration of Fluorescens Technical and D7. Transmittal of 1 Study.

49021 101 Brooks, M. (2012) Effect of Pseudomonas Strain D7 on Nontarget Vegetation

-

Supplemental Report. Project Number: NAP003A. Unpublished study prepared by U.S. Dept. of Agriculture/Pacific West Area. 165p.

49100800 Northwest Agricultural Products (2013) Submission of Toxicity Data in Support of

the Applications for Registration of D7 and Fluorescens Technical. Transmittal of 2

Studies.

49100801 Kuhn, J. (2013) Pseudomonas fluorescens Strain D7 Acute Eye Irritation in

Rabbits. Project Number: 17085/12. Un published study prepared by Stillmeadow, Inc. 16p.

49100802 Kuhn, J. (2013) Pseudomonas fluorecens Strain D7: Acute Dermal Irritation in

Rabbits. Project Number: 17085/12. Unpublished study prepared by Stillmeadow, Inc. 10p.Pseudomonas fluorescens s train D7 Page 22 of 29

Biopesticides Registration Action Document

49157600 Northwest Agricultur al Products (2013) Submission of Toxicity Data in Support of

the Applications for Registration of Fluorescens Technical and D7. Transmittal of 1 Study.

49157601 Younger, C. (2013) Pseudomonas fluorescens Strain D7: Ladybird Beetle Non

Target Insect Microb ial Testing. Project Number: 16338/12. Unpublished study prepared by Stillmeadow, Inc. 12p.

49177700 Northwest Agricultural Products (2013) Submission of Product Chemistry,

Toxic i ty, and Fate Data in Support of the Applications for Registration of

Fluores cens Technical and D7. Transmittal of 3 Studies.

49177701 Brooks, M. (2013) Pseudomonas Strain D7 Nontarget Insect Testing/ Supplemental

Report. Project Number: NAP7162012A1. Unpublished study prepared by Ag

Chem Consulting. 71p.

49177702 Brooks, M. (20 13) Effect of Pseudomonas Strain D7 on Nontarget Vegetation

Supplemental Report 2. Project Number: NAP003B. Unpublished study prepared by Washington State University and U.S. Dept. of Agriculture. 36p.

49177703 Brooks, M. (2013) Pseudomonas Strain D7 Dis cussion of Formation of

Unintentional Ingredients: Supplemental Report. Project Number: NAP71612013. Unpublished study prepared by Northwest Agricultural Products. 10p.

49349700 Northwest Agricultural Products (2014) Submission of Toxicity Data in Support of

the Applications for Registration of Fluorescens Technical and D7. Transmittal of 1 Study.

49349701 Doig, A. (2014) Pseudomonas fluorescens D7: Intravenous Toxicity/Pathogenicity

Study in Rats. Project Number: 17873/13. Unpublished study prepared by

S tillmeadow, Inc. 38p.

B. Environmental Protection Agency Risk Assessment Memoranda

US Environmental Protection Agency (USEPA). 2012. Memorandum from S. Borges (OPP/BPPD) to S. Cerrelli, Re: Environmental risk assessment for Pseudomonas fluorescens

s train D7 in response to an application for a FIFRA Section 5 Experimental Use Permit. Dated February 6, 2012.

USEPA. 2014a. Memorandum from I. Barsoum (OPP/BPPD) to S. Cerrelli, Re: Review of Acute Toxicity/Pathogenicity study for section 3 registrati on of the TGAI, Fluorescens Technical, and the EP, D7, containing the active ingredient Pseudomonas fluorescens strain D7. Dated April 15, 2014.

USEPA. 2014b. Memorandum from I. Barsoum (OPP/BPPD) to S. Cerrelli, Re: Review of Acute Toxicity/Pathogenic ity study for section 3 registration of the TGAI, Fluorescens Technical, and the EP, D7, containing the active ingredient Pseudomonas fluorescens strain D7. Dated April 15, 2014.

USEPA. 2014 c. Memorandum from I. You, Ph.D. through S

. Borges (OPP/BPPD) to S. Cerrelli Environmental risk assessment for the FIFRA Section 3 registration of the TGAI, FluorescensPseudomonas fluorescens s train D7 Page 23 of 29

Biopesticides Registration Action Document

Technical (EPA Reg. No. 71975

- G), and the EP, D7 (EPA Reg. No. 71975

- U), containing the active ingredient Pseudomonas fluorescens Strain D7 . Dated July 18, 2014.

C. Other References

Centers for Disease Control and Prevention. 2009. Drinking Water

- Water Treatment. Available

from http://www.cdc.gov/healthywater/drink ing/public/water_treatment.html

.

DeFelice K, Wollenhaupt N, Buchholz D. 1993. Aquifers and Soil Filter Effect. Available from http://extension.missouri.edu/p/WQ24

.

U.S. EPA. 2004. Primer for Municipal Wastewater Treatment Systems. EPA 832

- R

- 04

- 001.

Available from http://www.epa.gov/npdes/pubs/primer.pdf

Johnson, K. B., V. O. Stockwell, D. M. Burgett, D. Sugar, and J. E. Loper. 1993. Dispersa l of Erwinia amylovora and Pseudomonas fluorescens by Honey Bees from Hives to Apple and Pear Blossoms. Phytopathology 83:478

- 484.

Kremer, R. J. and A. C. Kennedy. 1996. Rhizobacteria as Biocontrol Agents for Weeds. Weed Technology 10:601

- 609.

McNab , B. K. 1966. An Analysis of the Body Temperatures of Birds. The Condor 68:47

- 55.

Palleroni, N. J. 2005. Pseudomonas. Pp. 323

– 379 . In: Bergey’s Manual of Systematic Bacteriology, Vol. 2. D. J. Brenner, N. R. Krieg, and J. T. Staley (Eds.) Springer, New York,

New York

Skipper, H. D. A. G. Ogg, and A. C. Kennedy. 1996. Root Biology of Grasses and Ecology of Rhizobacteria for Biological Control. Weed Technology 10: 610

- 620.Pseudomonas fluorescens s train D7 Page 24 of 29

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APPENDIX A. MICROBIAL PESTICIDES DATA REQUIREMENTS

(40 CFR PART 158

– SUBP ART V)

TABLE 1. Product Analysis Data Requirements for the Active Ingredient (TGAI), Pseudomonas fluorescens strain D7 and the

End

- Use Pesticide Product (EP), D7 (40 CFR § 158.2120)

Harmonized

G uideline

N umber Data Requirement Results MRID No.

885 .1100 Product Identity Submitt ed data fulfill the requirement for product identity and composition. 48966401

885.1200 Manufacturing

Process Submitted data fulfill the requiremen t for manufacturing process

. 48497801

885.1250 Deposition of a

Sample in a Nationally Recognized Culture

Collection Pseudomonas fluorescens strain D7 is on deposit at Agricultural Research

Service Culture Collection as NRRL B

- 18293

885.1300 Discussion of

Formation of

Unintentional Ingredients Submitted data fulfill the requ irement for preventing and detecting any

microbial contaminants. 4917703

885.1400 Analysis of S amples Submitted data fulfill the requirement for analysis of samples. 48966406

8 85 .1 50 0 Certification of

L imits Submitted data fulfill the requirement for m anufacturing process. CSFs dated

9/25/12Pseudomonas fluorescens s train D7 Page 25 of 29

Biopesticides Registration Action Document

TABLE 2 . Physical and Chemical Characteristics for the Technical Grade of the Active Ingredient (TGAI)

, Pseudomonas

fluorescens strain D7 and the End

- Use Pesticide Product (EP),D7 (40 CFR § 158.2120)

Harmoniz ed

G uideline N umber Data Requirement Results MRID No.

830.6302 Color Brown

48497801 830.6303 Physical State Granular

830.630 4 Odor Sweet sugar smell

830.6313 Stability to Normal and

E levated Temperatures, Metals,

and Metal I ons Stability to Met als: N/A

Stability to Elevated Temperatures: This product is not stable at elevated temperatures and must be stored refrigerated or frozen. This product will be stored in paper and used

immediately when tank mixed. N/A

830.6317 Storage Stability Submitte d data fulfill the requirement for storage stability and demonstrate stabilty for s test period of 361 days. 48966406

830.6319 Miscibility Not applicable (chemical is solid).

830.6320 Corrosion

Characteristics Submitted data fulfill the requirement for corrosion

characteristics 48966406

830.7000 pH The pH of a 1% solution is 5.1 48497801

830.7100 Viscosity Not applicable to a solid

830.7300 Density/Relative

Density/Bulk Density (Specific Gravity) 12.48 lbs/ ft 3 48497801Pseudomonas fluorescens s train D7 Page 26 of 29

Biopesticides Registration Action Document

TABLE 3

. Toxicology D ata Requirements for the Active Ingredient Pseudomonas fluorescens strain D7

and the End Use Pesticide Product D7 (40 CFR § 158.2140)

Harmonized

Guideline

Number Data Requirement Results MRID No.

Tier I

885.3050 Acute Oral

Toxicity/Pathogenicity Ps eudomonas fluorescens s train D7 does not appear to be toxic, infective, and/or pathogenic in rats, when dosed at 1.3 X 10 9

CFU/animal. Classification: Acceptable 489664 03

885.3150 Acute Pulmonary

Toxicity/Pathogenicity Pseudomonas fluorescens s train D7 do es not appear to be toxic, infective, and/or pathogenic in rat, when dosed at 4.6 x 10 8 cfu /

animal. Classification: Acceptable 48966404

885.3200 Acute Injection

Toxicity/ Pathogenicity Average CFU counts of the cecum contents were 8.6 x 10 4 , 5.5 x 10 5

,

2.9 x 10 5 , and 2.1 x 10 2 CFU/g on days 3, 7, 14, and 21, respectively. Pseudomonas fluorescens strain D7 was TOXIC and/or

PATHOGENIC as tested in this study.
Classification: Acceptable , however, injection with the active MPCA

at a dose of 1.3 x 10 8 CFU/ra t resulted in mortality, abnormal clinical signs; transient weight loss with resultant decreased absolute body weight, abnormal gross necropsy findings, and altered organ weights. 48966402

This study supersedes MRID 48966402 and was conducted to evaluat e the acute intravenous toxicity and infectivity of the microbial pest control agent (MPCA), Pseudomonas fluorescens D7, at two concentration exposures and at adequate post

- exposure observation period. No abnormalities were seen during daily observations o r at necropsy. There was no mortality in any group during the study. The test organism had cleared completely from the active 10 6 CFU/rat and

active 10 7 CFU/rat groups blood, kidneys, mesenteric lymph nodes, lungs, brain, liver, spleen and cecum contents b y Day 7. The observed differences in spleen weights for groups treated wi th live Pseudomonas fluorescens D7 are considered a normal immune response during the clearance phenomenon. The test substance Pseudomonas fluorescens

D7 was determined to be non

- toxi c to rats when injected intravenously

at a dose of 3.4 x 10 7 CFU/rat or at 3 .7 x 10 6 CFU/rat

. Classification: Acceptable 49349701Pseudomonas fluorescens s train D7 Page 27 of 29

Biopesticides Registration Action Document

TABLE 3

. Toxicology D ata Requirements for the Active Ingredient Pseudomonas fluorescens strain D7

and the End Use Pesticide Product D7 (40 CFR § 158.2140)

Harmonized

Guideline

Number Data Requirement Results MRID No.

885.3400 Hypersensitivity

Incidents The applicant reported that n o hypersensitivity incidents, including

immediate

- type or delayed

- type reactions of humans and domestic animals, occurred durin g research, development, or testing of the

TGAI. Any future hypersensitivity incidents must be reported to EPA (refer to test note #3 of 40 CFR § 158.2140(d))

. 885.3500 Cell Culture N/ A

870.1100 Acute Oral Toxicity This requirement addressed by Acute Oral Toxicity /pathogenicity study described above. Classification: Acceptable 48966403

870.1200 Acute Dermal Toxicity Dermal LD 50 Males >2000 mg/kg bw; Females > 2000 mg/kg bw;

Combined > 2000 mg/kg bw. Based on the results of this study, Pseudomonas fluorescens strain D7 is of low toxicity. Classification: Acceptable

TOXICITY CATEGORY IV 48966405

870.1300 Acute Inhalation

Toxicity This requirement addressed by Acute pulmonary

Toxicity/ pathogenicity study described above Classification: Acceptable 48966404

870.2400 Acute Eye Irritation No corneal opacity, iritis, or conjunctival irritation was noted on any rabbit throughout the study. Pseudomonas fluorescens s train D7 was

not irritatin g to the eye

. Classification: Acceptable

TOXICITY CATEGORY IV 49100801

870.2500 Primary Dermal

Irritation No dermal irritation was noted on any animal during the study. The

primary irritation index was 0.0. Classification: Acceptable

TOXICITY CATEGORY I V 49100802

Tiers II and III

Not required for Pseudomonas fluorescens s train D7 based on the lack of acute toxicity/pathogenicity in the Tier I studies.Pseudomonas fluorescens s train D7 Page 28 of 29

Biopesticides Registration Action Document

TABLE 4

. Nontarget Organism Toxicity and Environmental Fate Data Requirements for the Technical Grade of the Active Ingredient TGAI

, Pseudomonas fluorescens strain D7. ( 40 CFR § 158.2150)

Harmonized

Guideline

Number Data Requirement Results MRID No.

Tier I

885.4050 Avian Oral Toxicity An avian oral test indicated that P. fluorescens strain D7 is not toxic or pathogenic to Northern bobwhite ( Colinus virginianus ) dosed at 2.1 x

10 9 cfu/kg body wt/day. Classification: Acceptable 48966409

885.4100 Avian Inhalation

Toxicity/Pathogenicity Not required as the nature of the microbial pesticide does no t indicate potential pathogenicity to birds or relatedness to any known bird pathogens (refer to test note #3 of 40 CFR § 158.2150(e)). N/A

885.4150 Wild Mammal

Toxicity/Pathogenicity Wild mammal testing was not conducted nor was a waiver requested.

Studi es conducted with P. fluorescens strain D7 on laboratory animals are appropriate for use in risk assessment. Acceptable studies with laboratory rats indicated no concerns for toxicity/pathogenicity in

mammals

. Classification: Acceptable 48966403,

48966402

885.4200 Freshwater Fish

Toxicity/Pathogenicity A freshwater fish test with P. fluorescens strain D7 showed no toxic or pathogenic effect on Rainbow trout ( Oncorhynchus mykiss ) dosed at 1 x

10 6 cfu/ml. Classification: Acceptable 48966408

885.4240 Fre shwater Invertebrate

Toxicity/Pathogenicity A freshwater invertebrate study with P. fluorescens strain D7 showed no adverse effects on mobility or reproduction of Daphnia magna at exposure to 1 x 10 6 cfu/ml of test medium. Classification: Acceptable 489 66410

885.4280 Estuarine/Marine Fish

and Invertebrate Testing Scientific rationale was insufficient to conclude that no adverse effects

are expected on nontarget fish and invertebrates for all applications. P. fluorescens strain D7 is not to be applied d irectly to water and thus is not expected to reach estuarine or marine environments in significant amounts. However, aerial applications over large areas may lead to some exposure of P. fluorescens D7 in nearby estuarine/marine

environments.

Classificati on: Supplemental No MRID, but a letter (dated June 3, 2013) requesting a

waiver for

Estuarine/

Marine Fish and

Invertebrate Testing was

submitted.

885.4300 Nontarget Plant Testing Additional data and published literature submitted were sufficient to

supp ort the nontarget plant testing requirement for P. fluorescens strain

D7. Classification: Acceptable 49021101,

48966401

885.4340 Nontarget Insect

Testing A laboratory bioassay with adult ladybird beetles ( Hippodamia

convergens ) at 1X the field applicatio n rate showed no adverse effects

of P. fluorescens strain D7 to the test organism ( Helicoverpa zea ) within 7 days of testing. The study is not of sufficient duration and was not

carried out at the maximum hazard dose of 10X. Classification: Supplemental 49157601

49177701

885.4380 Honey Bee Testing Scientific rationale was sufficient to conclude that no adverse effects of P. fluorescens strain D7 on honey bees are likely to occur. Because of recommended preemergence uses in cool and wet (< 50 o F) condition s, the exposure of nontarget honey bees to D7, even when it occurs, is expected to be low. Classification: Acceptable 49177701

Tiers II, III, and IV Not required for based on the acceptability of the data and other information provided for Tier I

.Pseudomonas fluorescens s train D7 Page 29 of 29

Biopesticides Registration Action Document

A PPENDIX B. PESTICIDE PRODUCT S

EPA

File Symbol Registration

Name Percentage

Active Ingredient Formulation

Type Use Site(s) Method(s) of Application Application Rate (s) Target Pest

71975

- G Fluorescens

Technical 95% Technical Manufacturing use only N/A N/ A

D owny Brome (cheatgrass), Medusahead, Japanese Brome,

and Jointed Goatgrass 71975

- U D7 95% End

- use product Agricultural

turf and grasses grown for seed, alfalfa; wheat, barley, triticale, oat, and rangeland Ground

application,

Aerial 2g/acre (see lab el for

dilution instructions)

Seed

treatments 2

- 4 g/100 lbs. of seed