EPA's Records Disposition Schedule PEST 361 Scientific Data Reviews HED Records Center - File R139870 - Page 1 of 34

MEMORANDUM


### TO:

FROM:

PEERREVIEW:



### SUBJECT UNITED STATES ENVIRONMENTAL PROTECTION AGENC'I WASHING' ON DC 2,)460 

# AUG 2 "1" i".-li /' .' j

Susanne Cerrelli, Regulatory Action Leader Microbial Pesticides Branch Biopesticides and Pollution Prevention Division, 751 IC Robyn Rose, Entomologist Microbial Pesticides Branch Biopcsticides and Pollution Prevention Division, 751 IC Gail Tomimatsu, Ph.D., Plant Pathologist / ,,' 

### (:'j\, 

### , Microbial Pesticides Branch Biopesticides and Pollution Prevention Division, 7511C llFFICE ()\ pm 'JFt\ TION, PESTICIDES AND I '.<Ie SUBsTANCES

I "

Technical review summary ofnon-target, beneficial insect, avian and aquatic studies submitted by Kemira Agro Oy in support ofthe registration of Primastop® (EPA Reg. No. 64137-8) Gliocladium calenulatum Cell Mass (Technical Product) for outdooruses. Barcode Nos.: 0244243 & 0267225; Submission Nos.: S539189 & S580869; Chemical No.: 021009. 

### STUDY IDENTIFICATION I. MRiD #:445014-0I. A Dietary LC50 Toxicity and Pathogenicity Studywith the Northcl11 Boin'.hite, Colinus virginianus. 2. MR!D II 445014-()2. A Contact and Dietary LC,o Toxicity and Pathogenicity Study with the Rainh,m Trout, Oncorhynchus mykiss Under Static Renewal Conditions 3. MRID #445014-03. A 21-Day Life Cycle LC,o Toxicity Test with the Cladoceran Daphma mugllil under Static Renewal Conditions. 4. MR! [) #445014-04 .A Dietary LC,o ToxicityTest with the Green Lacewing, Chrysoperlu I',.'mea

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5 . .\1f{J[) #445014-05. A Dietary LCso Toxicity Test with the Parasitic Hymenopteran, iVasonhi vi.1 ri(JCIlnis 6. M RID 11445014-06. A Dietary LC,o Toxicity Test with the Ladybird Beetle, Hippodal1ll'f,',mvergens 7. \1RlD #.445014-07. A Dietary LC,o ToxicityTest with the Honey Bee Apis melli/"!',, 8. 

### MRID #44183-20
.Non-Target Plant Literature Search CLASSIFIC,\liON: Acceptable BACKGROliND: GliocladiulII (illenulalum cell mass (Registration #64137-1) is currentlyregistered for indoor greenhouse W;C. KemiraAgro Oy has submitted ecological effects studies required byEPAto register microbial pest control agents for outdooruse.. The end-use-product is called Primastop Biofungicidc Powder with the active ingredient listed as 37% CT/iocladium catenu/atum Strain J1446 (contains 1.0xlOX CFU/gram) 63%inert ingredients. The intended uses ofPrimastop are to control "damping-off, seedrot, root and stem rot, wilt and certain foliar dIseases caused by Rhizoctollia, P)'thium, Fusarium, Didymella,Bollyt;s, VerticilliulIJ. Aiternaria, Cladosporium, Helminthosporium, Penicillium, and Pi/curia on greenhouse or !Ield grown vegetables, herbs, ornamentals, turf, tree and forest seedlings." Primastop Illa\' be applied through incorporation inpottingmedia, treatment ofcuttings, bulb or tubers, as a "Oil drench or foliar spray. Three insect species including a neuropteran (Green lacewing, Chrysoperla carneal, parasitic hymenopteran I.tvasonia vitripenllis), a coleopteran (Convergent lady beetle, Hippodamia convergensl, and honey bees (Apis mellijera) were tested. In addition, potential effects of GliocladiulII c,lIcnulatum cell mass on a representative avian species ( northern bobwhite quail; Colinlls virgll1,al1us), freshwater fish species (rainbow trout; Oncoryhynchus mykiss) and aquatic invertebrate (L Jal'hnia magna) were evaluated. These are appropriate test speci<:s for the registration IlfIlllcrobial pest control products. CONCUJSIO'lS: Ellvi[QI]lIlelltal Fate An AGRlCUi \ search conducted byKemira Agro Oy using the key words Gliocladium catenlilatulII and Illsects resulted in no reports ofharmful effects ofCTliocladium catenulatum on insects (MRID if 441893-21). The only result ofthis search showed the potential forEPA's Records Disposition Schedule PEST 361 Scientific Data Reviews HED Records Center - File R139870 - Page 3 of 34

Gliocladiufll ,'(/fCllu/alum to degrade the insecticide aldicarb. A second AGRICOLA searchwas conducted IIsing the key words Gliocladium catenu/alum and avian, bird, shrimp, mollusk, or fish (MRID #441893-19). There were no items found bythis search for shrimp or fish. One item was ttJlmd tormollusks. howeverit addressed Gliocladium Vlrens and Gliocladium deliquescell,'; rather than Gliocladium catenulatum. A search using key words "avian" and "bird" resulted in 110 items relating to Gliocladiwn catenulalum. However, two items were found relating to imoi/avian and Gliocladium, These items did not relate to Gliocladium catenulatum. Studies idenlilied from the literature searches (MRID #441891-19 and 441893-21) do not suggest any potential for adverse effects presented to non-target species by Gliocladium catelllliatwJl applied in the field. Environmenlal fate studies are notrequired atthis time since no adverse risks weretriggered in Tier [Non-target Organisms Studies. Gliocladium catenulatum strainJl446 is a naturally occurring saprophytic soil fungus; as a species, it is found throughout the world in a variety of soil types, without pathogenic incidence to plant oranimal. The ubiquitous nature ofthe organism suggests that the envirorunental burden is not likely to increase substantial.iy to pose adverse risb 10 nontarget organisms as a result of the intended outdooruses. Members ol'th" genus have been reported as fungal biocontrol agents, e.g., Gliocladium virens; others may he involved in pathogenesis ofsomeplant species: ornamentals (G. vermoeseni : Gliociadiulll seem rot); citrus (Ci. roseum: pink mold) and carrot (G. roseum: hard rot). Estimated Environmental Concentration ofGliocladium catenulatllm Cell Mass: Based on the Illtended rates ofapplication, the estimated environmental concentrationof Gliocladium ('(Jtenulatllm Cell Mass is not anticipated to exceed

- 500 mg/L for soil incorporation applications; and**c~200** mg/l. for soil drench applications. Ecol.qgjcalEffects Results ofnon-target avian, freshwater fish, daphnia, honey bee and beneficial unseet studies submitted to tile Agency generally demonstrated a lack oftoxieityorpathogenicity to the indicator species. Results ofthese studies indicate that minimal toxic orpathogenic effects to terrestrial or n";:shwater wildlife are expected from currently intended applications of Glioe/adi1lm C,/fclluiat1lm cell mass. Summaries and conclusions ofthese studies arc presented below. OPPTS Guideline 885.4050: Terrestrial Avian (MRlD#445014--01): Under condit !OilS ofthe reported study, no adverse effects were exhibited byjuvenile northern bobwhlte ann .10 clays when administered (by oral gavage) daily doses of 1 x 1()4 Gliocladium catel1ulatulll-clllig body weight/day for a five day period. This dosage is approximately equivalent to a lotal dosage of7 x I ()O cfu/gbody weight tor 5 days. The no observed effect dosage of(jiwc/"dium catenll/atum cell mass was determined at approximately 1.4 x 10" cfu/g body weight 1;:1' five days. Therewere no evidences oftest substance replication, Icswns, 3EPA's Records Disposition Schedule PEST 361 Scientific Data Reviews HED Records Center - File R139870 - Page 4 of 34

plaquesor other indications ofinfectivity orpathogenicity for the groups treated with Gliocladiul!I ,«tenulatum cell mass in gross necropsies. This study showed a lack oftoxicityor pathogenic;IY0

## rCiliocladium ciltenulatum cell mass to northern bobwhite quail. This study is sCIentifically valid, satisfies current guideline requirements, and is acceptable for assessing shoJI-Ierm toxic andpathogenic effects ofGlioc/adium catenulatum Strain .11446 to terrestrial a\

### ;'111 species. OPPTS Guideline 885.4200: FreshwaterFish (MRID# 445014,,02): Under static-l
..,newal conditions, the no-observed effect concentration was 179 mg/L (1.3 x108 cfu/L) forjuvenile rainbow trout dual-exposed via aqueous tcst solutions and in a daily diet for 30 days. The no mortality concentration was 357 mg/L (2.5 X I0' dulL). The LC50 was calculated as ',04 mg/L (3.5x1 OS cfu/L). Therc were 110 signs ofinfection in fish tissues examined for necropsy, and all appeared internally and externallynormal. Ciliocladium catenulatum was isolated from the hemi, liver and brain samples

# or fish treated with 179 mg/L test substance; and from the kidney, heart, liver and brain samples offish treated with 357 mgiL test substance. Although the test substance was present, target organs did not appearto be affected, and there were no apparent effects uponthe health ofthe r,,;h. This study IS scientificallyvalid, satisfies current guideline requirements, and is acceptable for assessing Sh"'l'.-tcrm toxic and pathogenic effects ofGliocladium catenulatum Strain .11446 to freshwater Iisl, OPPTS Guideline 885,4240:Daphnia magna (MRID#**445014-0~:** Under static'l'cnewal conditions, the 21-day EC,o value for survival ofneonate Daphnia magna exposed aqucously to Gliocladium catenulatum was 7.8 mg/L (5.5 x 10° cfu/L). The LOEC and the NOEC Wei" 11 mg/L and 5.6 mg/L, respectively. Ofthe three p,;:'ameters assessed (survival, growth and reproduction), reproduction proved to be the most sellsiHc parameter. The EC,o value was 7.8 mglL**(~·5.5** x j06cfU/L); and the NOEC and the LOEC \\"re 2.8 (2.0 x 10" cfu/L) and 5.6 mg/L (3.9 x 10" cfu/L), respectively. This study is s,:ientifically valid, satisfies current guideline requirements, and is acceptable for assessing shorHenn toxic effects ofGiiocladium catenuiatum Strain .11446 to aquatic invertebrates The pathogenicity (orinfectivity) of Gliocladium catenulatum was not evaluated in this stud\' OPPTS Guideline 885.4300: Non Target Plants (MRID # 44183-20): Areview ofthe literature resulted in the utility ofCiliocladium catenulatum to control plant diseases caused by other fungi, Rhizoctonia, PYlhium, Botrytis, Fusarium, Didymella, Verti('illiulII, 'ii/crnaria, Cladosporium, Heiminthosporium, Penicillium andPi/caria. There 4EPA's Records Disposition Schedule PEST 361 Scientific Data Reviews HED Records Center - File R139870 - Page 5 of 34

were no rcpnn, ofpathogenicityto plants
.. Therefore, C. i'utenulatum is not expected to cause adverse effects to nontarget plants. The literature review satisfies current guidelines for non-target plant testing and may be used as the basis Jor W31\ ing lurthernon-target plant tests, pending appropriate product characterization and taxollOlnic ViC!']ficatioll. OPPTS Guideline 885-4380: Honey Bees (MRID#445014-07): This study 

## "'b conducted in compliance with Good Laboratory Practice Standards as published by the U.S. I!nvironmental Protection Agency, Office ofPesticide Programs in 40 CFR Part 160. Reported be:c mortalities were not dose-responsive nor statistically significant. Thereforethe dietary Leu v,due for honey bees exposed to G. catenulatum cell mass for 10 days is estimated to be greater than 5200 ppm (3.6xI0" CFlJ/mL), the highest concentration tested, and the no observed eficcconcentration is estimated to be 5200 ppm (3.6x10" CFU/mL). An 

### LD5(J value >3.6)( I0(' 

# en 'tnL and an NOEL of3.6xI0' CFU/mL were reported. This study showed a lack oftoxicity of I ;flOcladium catenuiatum cell mass to honeybees. Mortalities were therefore presumed ttl Ix associated with otherenvironmental variables associated with the test arena. There were 110 evaluations ofinfectivity or pathogenicity. This study IS scientificallyvalid, satisfies current guideline requirements, and is acceptable for assessing s]l\)Jt-1erm toxic effects ofGliocladium catenulatum Strain Jl446 to honeybees.

OPPTS Guideline 885.4340: Nontarget Insects

- Green Lacewing (MRID#445014-04): The rates oflllllrtality and pupation were similarin the treatment groups as in the untreated control group. Reported mortalities were nottreatmentrelated. Therefore, the dietary LC50 value for green lacewing larvae exposed to G. catenulatum cell mass for 12 days is presumed greater than 5200 ppm (J.6xI0" CFUlg) and the no observed effect concentration is**~5200** ppm (3.6xI06 CFU/g). This study showed a lack oftoxicity ofGliocladium catenulatum cell mass to green lacewings. There wcre no evaluations ofinfectivity or pathogenicity. This study i" scientifically valid, satisfies current guideline requirements, and is acceptable for assessing shm,-term toxic effects ofGliocladiul1l catellulatum Strain .11446 to green lacewings. OPPTS Guideline 885-4340: Nontarget Insects

- Parasitic Hymenoptera (MRID# 445014-**~:** Mortalities '"ere related to the test substance in one group--treated with 5,200 ppm G. catenlllatlllJl c\.:lllllass. Therefore the dietary LC50 value for parasitic Hymenoptera exposed to G. catcnulu/l(lu cel1mass for eight days was calculated approximately 1992 ppm (lAxI0" CFU/lllL) and the no observed effect concentration dctemlined to be 520 ppm (3.6xlOj CFU/mL).TIllS study showed a lack oftoxicity ofGliocladium calenulatum cell mass to 5EPA's Records Disposition Schedule PEST 361 Scientific Data Reviews HED Records Center - File R139870 - Page 6 of 34

parasitic HYlrenoptera. There were no evaluations ofinfectivity orpathogenicity. This study

# j, ·;cicntifically valid, satisfies current guideline requirements, and is acceptable for assessing shor:-term toxic effects ofGliocladium catenu/atum Strain Jl446 to parasitic wasps. OPPTS Guideline 885.4340:**Nontan~et** Insects

- Lady Beetles 

### (MRID#445014-06): Mortalities were not dose-responsive, nor associated with Gliocladium catenu/atum cell mass. Therefore the dietary LC50 value for ladybird beetles exposed to 

### G. catenu/atum cell mass for 16 days is estimaled to be greater than 5200 ppm (3.6xI0" CFU/mL), the highest concentration tested, and 

### 1.11(' no observed effect concentration to be 520 ppm (3.6xl05 CFUlmL). An 

### LDso >3.6< 10" Cf-{iimL and an NOEL of3.6xIOS CFUlmLwas reported. This study showed a lack of toxicity oflihue/adium catenutatum cell mass to lady beetles. There wereno evaluations of infectivity (II' pathogenicity. This study i," ,'cientifically valid, satisfies current guideline requirements, and is acceptable for assessing short-(crm toxic effects ofG/iocladium catenu/atum Strain ]1446 to ladybird beetles. Imp/!.f:fl· toEndangeredSpecies Impacts to endangered species [rom intended exposures ofPrimastopTM are anticipatedto be negligible or nonexistent. This assessment is premisedupon a combination ofseveral factors: (I) low BEC values; (2) data and evidence from the public literature as summarized in the Environmental Fateportion ofthis assessment; and (3) the practically nontoxic (andnoninfective) status oftheMPCA in surrogate toxicity/pathogenicity studies summarized in the Ecological Effects portion.EPA's Records Disposition Schedule PEST 361 Scientific Data Reviews HED Records Center - File R139870 - Page 7 of 34

DATA EVALUATION 

### REPORT Reviewed bv: Robyn Rose, Entomologist, BPPD

## ~t ':?

# I}\ i:)\**"\~'** I Secondary Reviewer: Gail Tomimatsu, Plant Pathologist, BPPD"n I 'I; I ' \c' I
STUDY TY\)I:: MRlDNO. CHEMICAL 

### ',CL TEST MA**T!~RIAL:** STUDYNO,: SPONSOR TESTING F\CIUTY: TITLE OF \,FPORT:
AUTHORS STUDY COMPLETED: CLASSIFlCAnON: Study Summary; Green Lacewing (Chrvsoperla carnea) Larvae Dietary Pathogenicity and Toxicity Study (l:54A-23; OPPTS 885.4340) 445014-()4 021009 Gliocladium catellulatum strain 

### J1446 (Technical Powder) 2')3-113A Kcmira Agro Oy, P.O. Box 330, Helsinki, Finland Wildlife International Ltd., 8598 Commerce Dr.. Easton, MD 21601 Gliocladium catcnulatum Cell Mass (Tec1mical Product): A Dietary Pathogenicity and Toxicity Study With Green Lacewing Larvae (Chrvsoperla carnea) Kimberly A. Hoxter, Susan J. Palmer, and Henry O. Krueger, PhD. 2/5/98 Acceptable

Objectivc: "The objective ofthe study was to evaluate the pathogenicity and toxicity of Gliociadilllll cutellulalum cell mass administered to green lacewing larvae (Chrvsoperla carneal in the diet." Test**Materi~\1:** G/ioc/adiufI/ ,urenulalum Cell Mass (Technical Product) Batch No. 44/95 111 ; a light brown powderwith a reported activity of7x10' colony forming units per gram (cfu/g) was received from Kemira Agro Oy (3/26/97; Wildlife International ID#3996). Test substance viability was determined 11011] 0, Ig tcst substance 

### + ImL 0.3% TRITON-X sol. + 99mL sterile**H20'~JO-J** dilution. TIll' Illixture was homogenized for**~** 

### 1 0 sec. beforepreparing final solutions (10-5, 

### 10-", \0.7 ) 10 bepial,xi on potato dextrose agar. The activity ofthe G. calenulalum was verified at Wildlife lntclllatiollal using the equation: Viability (in tenns ofCFU/g) = L:C"IV" x N

# xL (Dn) [cfu/gJ: C:""h ",'colonieson the agar plate, Vn=volume plated(mL), N=# ofreplicate plates, D,,=dilution, 

## 0: the sample(g/mL). Tcst Organism. Green LacewtJ':" (Chrysoperlu c({rnea) larvae from Rincon-Vitova Insectaries, Inc., P.O. Box 1555, Ventura. CA 93002. 7EPA's Records Disposition Schedule PEST 361 Scientific Data Reviews HED Records Center - File R139870 - Page 8 of 34

TestMethqd,: The pathogenicity and toxicity ofGliocladium calenulatum cell mass fed to green lacewing (GL) larvae was evaluated. Threenominal concentrations (52, 520, and 5200ppm) ofG. catenulatum and a negative control (moth eggs without test substance) were fed to GL larvae viamoth (Sitotroga sp. From Rincon-VitovaInsectaries, Inc., Ventura, CAl eggs mixed with thetest substance; the moths were given new diet material weeklyadlihitum. The concentrations were equivalent (0 :u,xI04, 3.6x 10', and 3.6xI0" CFU/g respectively based on a verified activity (verifIed 5 tnncs on two dates: May 10 and May 25) of7x 10' colony forming units pergram (CFU/g). The test diet wasprepared bycalculating: Desired Concentration (ppm or fig/g) = Test Substance (gj Final Volume ofStock Suspension (mL) x Volume ofStock Suspension Mixed With Moth lOggs (mL) -;- Final Weight ofDiet (g) x 10" fig/g. Sampleswere taken from thetest diets and the control on Day 0 and Day 7 to determine stability andhomogeneity ofthe stock suspensiolls hv serial dilution plating onto potato dextrose agar. Each concentration and the control received thirty test chamberswithone GL in each chamber. GLs were reared and fed the test material in the lab. Test chambers consisted ofone ounce clear semitransparent plastic Solo@ (No. 601P) cups with clear semitransparent plastic lids. GLs were kept in incubators at 20.8 ± 0.2°C (SD) WIth a rangc of20.8 to 21 "C, average relative humidity 87 ± 6% (Sf)i (range 66

- 90%), and a photoperiod of 12: 12 with overhead fluorescent lighting. Observations ofmortality andpupation weremade within four hours (-3/4 ofan hour) aftertest initiation and Ihen daily for 12 days. Observationswere discontinued after the 12th day when pupation exceeded 50% and mortality exceeded 20% inthe negative control. The LC,O values were determined by the cumulative mortalitypercentages observed in thetreatment groups. The concentration resulting in no effects was determined through mortality, pupation, and observation,
_ Reported**Res"l1~:** Cumulative percent mortality ofgreen lacewing (GL, Chrysoper/a carnea) larvac exposedto Giiocladiulli ,-"renulalum cell mass and percent pupation ofgreen lacewing larvae exposed to G. catenu/atuII! cell mass were' Control 52 ppm 520 ppm 5200ppm



### - Percent M()] 1 ality 27 20 13 13 Percent Pupat1011 57 67 70 70 The results showed the dietary LC,o value for green lacewing larvae exposedto G. carenulatum cell mass for 12 days to be greaterthan 5200 ppm (3.6xI0" CFUlg) and the no observed effect concentration to be 5200 ppm (3.6x10' CFU/g). There were no signs ofovert toxicity noted in the surVIving larvae in the untreated control ortreatmcnt groups. The rates ofmortality and pupation werc similar in the treatment groups as to the untreated control group. Percent mortality and percent pupation were respectively slightly higher and slightly lower than groups exposed to'; c"tenu/alum. 8EPA's Records Disposition Schedule PEST 361 Scientific Data Reviews HED Records Center - File R139870 - Page 9 of 34

For nominal stc)Ck solutionconcentrations of312, 3,120 and 31,200 mg/L; mean viable counts of 

### G. calenu/aillfl' were reported: from Day 0 verification samples: 7,5 x 104, 1.8 x 10" and >2.0 x 10" CFU/mL. respectively; and from Day 7 stability samples: 5.4 x 104,8.3 x 10' ami 1.5 x 10 7 CFU/mL. Ua:- () verification and Day 7 stability samples each demonstrated mean value of0.0 CFU/mL. Reviewer C()Jll)11ents: A. Methods This study \\a,; conducted in compliance with Good Laboratory Pf31ctice Standards as published by the U.S. hlvironmental ProtectionAgency, Office ofPesticidePrograms in 40 CFR Part 160. I. Thi; study did not include two controls: an infectivity control group (inactivated Gliocladium e,denu/alum) and a treatment in which lacewings are dosed with sterile filtrate from produclion cultures. However, absence ofthese treatments did not influence the overall conclusions 01 the study. B. ConcluslOllS The results ofIhis study showed the dietary 

### Le,o value for grecn lacewing larvae exposed to G. catenulatulI! ("II mass for 12 days is greater than 5200 ppm (3.6x106 CFU/g) and the no observed eflccl concentration is 5200ppm (3.6xI0" CFU/g). This study showed a lack of toxicity of(,'lll)ciadium catenulatum cell mass to green lacewings. Therates ofmortality and pupation were similarin the treatment groups as (0 the untreated control group. Percent mortality and percentpupation were respectively slightly higher and slightly lowerthan groups exposed to (, ,·alenu/atum.

9EPA's Records Disposition Schedule PEST 361 Scientific Data Reviews HED Records Center - File R139870 - Page 10 of 34

DATA EVALUATION REPORT Reviewed by: Robyn Rose, Entomologist, BPPD

### l~ i:"J(JI

### (co 

### I Secondary RC\lCwer: Gail Tomimatsu, Plant Pathologist,

## BPPD:;~:~ ,;/? i 

## i t'l
STUDY TYPI.: MRID NO. CHEMICAL 1\0.: TEST MATFR1AL: STUDY NO
.. SPONSOR TESTING F,\,'IUTY: TITLE OF REPORT:
AUTHORS STUDY COMPLETED: CLASSIFICATJON: Study Summan: Parasitic Hymenoptera (Nasonia vi/rtpenl1is) Dietary Pathogenicity and Toxicity Study (154A-23; OPPTS 885.4340) 445014-05 021009 Gliocladiurn catenulaturn strain .11446 (Technical Powder) 293-112 Kcmira Agro Oy, P.O. Box 330, Helsinki, Finland Wildlife International Ltd., 8598 Commerce Dr., Easton, MD 21601 Gliocladiurn catenulaturn Cell Mass (TechnicalProduct): A Dietary Pathogenicity and Toxicity Study With Parasitic Hymenoptera (Nasonia vi/ripennis) Kimberly A. Hoxter, Susan 1. Palmer, and Joann B. Beavers. 2/5/98 Acceptable

Objective: "The objective ofthe study was to evaluate the pathogenicity and toxicity of Gliocilldiulli (illel/ulaturn cell mass administered to parasitic Hymenoptera (Nasonia l'itripel1l1is) in the JieL" Test Matena!: GliocladiullI uucilulatum Cell Mass (Technical Product) Batch No. 44/95 JJl; a light brown powderwith ;" reported activity of7x 108 colony fomling units pergram (cfu/g) was received from Kemira .\gro Oy (3/26/97; Wildlife International ID#3996). Test substance viability was determined Ir,Jln O.lgtest substance 

### -+ ImL 0.3°1t. TRITON-X soL 

### -+ 99mL sterile H20=]()·3 dilution. The mixture was homogenized for -10 sec. before preparing final solutions (10-', ]()-", J0-7). The activity ofthe G. catenulatum was verified at Wildlife International using the equation: Vlabiilly (in tenus ofCFU/g)**.~** [C./V" x N x 

## L (D,,) [cfu/g]; C,,=# ofcolonies onthe agar plate, \,.\·olumeplated(mL), N=# ofreplicate plates, D,,=dilutions ofthe sample(g/mL). Test OrgamslI\: Parasitic**HYI11~noptera** (Nasonia vitripcnnis) obtained from CarolinaBiological Supply Company, :C/[)O York Rd., Burlington, NC 27215-3398 as larvae inblowfly (Sarcophaga sp.) pupae Adulh were obtained from pupae after 3 days in an incubator set to maintain a 10EPA's Records Disposition Schedule PEST 361 Scientific Data Reviews HED Records Center - File R139870 - Page 11 of 34

temperature t)f

. HIto 

### 28°C, with relative humidity above**~40%.** Test**Method~:** ThepathogenIcity and toxicity ofGliocladiwn catenulatum cell mass fed to Nasonia vitripennis was evaluated Three nominal concentrations (52, 520, and 5200 ppm) ofGliocladium and a negative control were presented to N vifripennis twice each weekvia a honey and deionized waterdiet cOal'cd on a cotton swab. Samples weretaken from the tcst diets and the control on Day 0 and Dll', 7 to determine stability and homogeneity ofthe stock suspensions by serial dilution platll1g onpotato dextrose agar. The nominal concentrations were equivalentto 3.6x

### 10\ 3.6xI0" and} (" I0" respectivelybased on an activity of7x 10' colony forming units per gram (CFUig). The test substance was combined with 10mL deionized water and honey was added until IOOml. ",as reached; diets were prepared immediately before testinitiation and on Day 7. Twenty five wasps ornearly identical size were immobilized with nitrogen gas and impartially distributed am,lIlg the test chambers. Wasps were allowed adlihitum access to diet and waterin test chambers. Test chambel" consisted ofone-pint rolled papercontainers (9cm diameter and 9 cm high) covered with" disposableplastic petri dish**(~** 10cm diameter) with an inverted 20mL glass vial containing dci''Ilized water inserted through it. Test dosages and the negative control were replicated in lilplicate; 75 adult wasps were tested pertreatment group. N. vitripennis werekept in incubators ,,'. 27.2 ± 1.3°C (SD) with a range of24.9 to 28.9°C, average relative humiditywas 82 ± 8'% (range: 7()

- 93%), and a photoperiod of 12: 12 with overhead fluorescent lighting. Observations 

### ,)1'mortality and clinical signs oftoxicology were made one and 1.5 hours afterthe initiation ofthl: test and then daily for eight days. Observations were discontinued afterthe eighth day \'. h,:n mortality exceeded 20% in the negative control. The C.E. SlCP'1a1l computerprogram was used to analyze mortality results at test termination. The 

## to,,, vallie and the 95% confidence interval (by probit analysis), and the moving average method or\)]nomial probability with nonlinear interpolation were calculatedviathe C..E. Stephan computerprnl:ram. The concentration resulting in no effects was determined through visual inspection 0['mortality and clinical observation. Reported ResuIt,;: Cumulative perccllt mortality ofparasitic Hymenoptera (N. vitripennis) exposed to Gliocladium catenulatulJI cell mass were:



### - Control 52 ppm 520 ppm 5200 ppm Percent IVImtality 

### 23 35 25 68



### - # thal did 17175 26/75 119175 51/75



### - Mortality Il·om the 52 ppm and 520 ppm treatment groups were comparable to the negative control group i'ercent mortality at test termination in the 5200 ppm treatment group was 68% (51 of75) \. sl11all number ofwasps exhibited a loss ofequilibrium orwere immobile during 11EPA's Records Disposition Schedule PEST 361 Scientific Data Reviews HED Records Center - File R139870 - Page 12 of 34

the test. All olher surviving wasps in this treatment group were normal in appearance and behaviorthroughout the test period. Mortality in the 5200ppm treatment group began to increase on da\" 2 and continueduntil test termination; and in this treatment mortalities appeared to be related I" treatment with the test substance. A small numb"r ofwasps were immobile or lethargic in the 52 ppm treatment group during the test. All other surviving wasps in the 52 and 520ppmtreatment groups were normal in appearance an,1 behaviorthroughout the test. Mortality at test termination in the 52 and 520 ppm treatment grolips were comparableto the negative control group. Therefore, the mortalities and clinical signs observed were not considered to be related to treatment with the test substance. The results showed the dietary LC50 value for parasitic Hymenoptera exposed to G. calenu/aluill cdlmass for eight days to be approximately 1992 ppm (IAxIO" CFUlmL) and the no ohserved ·;::Te,:t concentration to be 520 ppm (3.6>< 10' CFU/mL). Viahle populations of

### G. calenu/alllm from verification and stability samples ofnominal concentration" of52, 520 and 5200 mg/L were as follows: Day 0 verification samples demonstrated mean values of2.3 x 104,2.7 X 10' and 1.3 x 10" CFU/mL respectively. Day 

### 7 stability samples demonstrated meanvalues of2.2 

### x 103,4.0

### x 10', and 1.5 

### x 105 CFU/mL respectively. Day 0 verification and Day 7 stability samples from Ih<: negative control diet showed 0.0 ('U/mL. Thenumber ofplates used to determine mean values was not reported. Reviewer C'2!nments: A. Method, This study \va" conducted in compliance with Good Laboratory Practice Standards as published by the U.S. L:llvironmental Protection Agency, Office ofPesticide Programs in 40 CFR Part 160. 1. '11m study did not include two controls: an infectivity control group (inactivated Gliocllldiulil ,,,{clIu/alum) and atreatment in whichparasitic wasps are dosed with sterile filtrate from productun cultures. However, absence ofthese treatments did not influence the overall conclusions o! tile study. B. Conclusio"s Mortality ohs,:rvcd at the 52 and 520 ppm were comparable to the control and, therefore, not considered to be treatment related. Mortality in the 5200 ppm treatment group beganto increase on day 2 and ,:ontmued until test termination; and in this treatment mortalities appeared to be related to treatll1ent withthe test substance. Therefore the dietary LC50 value for parasitic Hymenoptera 'exposed to G. ealenu/alum cell mass tilr eight days was calculated as .- 1992 ppm (lAx I0" CHIllL) and the no observed elfect concentrationwas calculated as 520 ppm (3.6x I0' CFU/mL).1 his study showed a lack oftoxicity ofGlioc/adiun1 c"tenu/atum cell mass to parasitic H\'!1",noptera.

12EPA's Records Disposition Schedule PEST 361 Scientific Data Reviews HED Records Center - File R139870 - Page 13 of 34

DATA EVALUATION REPORT Reviewed by: Robyn Rose, Entomologist, BPPD 

# {L

# Ii 8/:21**(0\,,~,** 

# I Secondary Re"iewer: Gail Tomimatsu, Plant Pathologist, BPPDk)' I, I -:
STUDY TYPf: MRID NO
.. CHEMICAL ,',0: TEST'vIATERIAL: STUDY NO : SPONSOR TESTING FACILITY: TITLE OF RFPORT:
AUTHORS. STUDY COMPLETED: CLASSIFIC'l.T10N: Study**Summar~':** 

### " Ladybird Beetle (Hippodamia convergens) Dietary Pathogenicity and Toxicity Study (154A-23; OPPTS 885.4340) 445014-06 021009 Gliocladiul11 catenu/atum strain J1446 (Technical Powder) 293-11 I Kemira Agro Oy, P.O, Box 330, Helsinki, Finland Wildlife International Ltd., 8598 Commerce Dr., Easton, MD 21601 Gliocladiwn catenu/atul11 Cell Mass (Technical Product): A Dietary Pathogemcity and Toxicity Study With The Ladybird Beetle (Hipl'odamia convergel1s) Kimberly A. Hoxter, Susan 1. Palmer, and Henry O. Krueger, PhD, 2/5/98 Acceptable

Objective: "fhe: objective ofthe study was to evaluatethepathogenicity and toxicity of G/ioc/adilllll UI{Cl1u/atum cell mass administered to the ladybird beetle (Hippodmnia convergens) in the diet." Test Material: G/iocladiul1l (Olcflu/alum Cell Mass (Technical Product) Batch No. 44/95 111 ; a light brown powderwith a reported activity of7xI08 colony forming units per gram (cfu/g) was received from Kemira·\gro Oy (3/26/97; Wildlife International ID #3996), Test substance viability was deternlined limn O.lg test substance 

### + ImL 0.3% TR1TON-X sol. 

### + 99mL sterile H,O=IO'] dilution. The :nixture was homogenized for -10 sec. beforepreparing final solutions (10-5
, 10-6, 10,7). The actlvity ofthe 

### G. catenu/alum was verified at Wildlife International using the equation: y",hdity (interms ofCFU/g) =**~C,/V,** x N x**~** (0,) [cfu/g]; C,=# ofcolonies onthe agarplate, V"l'olumeplated(mL), N=# ofreplicate plates, D,=dilutions ofthe sample(g/rnL). Test Orgamsill. Adult Ladybird Beetles (Hipl'odamia convergens) obtained from A-I Unique Insect Control, 5504 Sperrv L}.'i\'e, Citrus Heights, CA 95IJ2L

13EPA's Records Disposition Schedule PEST 361 Scientific Data Reviews HED Records Center - File R139870 - Page 14 of 34

Test Methods Thepathogenicity and toxicity ofGliocladium catenulatum cell mass fed to adult ladybird beetles (Hippodamia convergen.\') was evaluated. Three nominal concentrations (52, 520, and 5200 ppm) 0 I' Gliocladium and a negative control were fed to N. vitripennistwice each week via a honey and d"ionized water diet coated on a cotton swab
.. Thetest substance was combined with IOmL deionized water and honey was added until IOOmL was reached. The concentrations were equivaiclilio 3.6xI04, 3.6X 105, and 3,6xI0" respectivelybased on an activity of7x 1 08 colony fomling units pergram (CFU/g). All treatments and the negative control were replicated three times (7' ladybird beetles pertreatment). Samples werc taken from thetcst diets and the control on Da\' () and Day 7 to determine stability and homogeneity ofthe stock suspensions Test chambers consisted ofone-pintrolled papercontainers (<Jcm diameter and 9 cm high) covered willi :.1 disposableplastic petri dish**(~I**Ocm diameter) with an inverted 20mL glass vial containing deionized water inserted through it. Adult ladybird beetles werekept in an incubator for four days pl'ior to test initiation; temperature was sct at 26 to 28°C and the relative humidity was above approximately40%. Duringthe test, the temperature in chamberswas 27.2 ± 1.3°C (SD) \\'ith a r,mgc of24.9 to 28.9°C, relative humidity was 82 ± 8% (SD) with a range 000to 93%, and the photoperiod was 12: 12 with overhead floureseent lighting. Each test chamber consisted of2'i IJ. convergen.\'. Observations ol'mortalitywere made approximately one hour and 1.5 hours ;Inertest initiation and then daily for 16 days (except Day 12); negative control mortality reach 20% on Day 16. Samples were taken from the test diets and the control on Day 0 and Day 7 to**t!\~lcrmine** stability and homogeneity ofthe stock suspensions. An estimation '.l

## rmortality patterns were determined by visual inspection ofdata. The concentration ,csL!lting in no effects was determined through mortality and clinical observation data. The LC 5, was determined bynominal concentrations. Reported Results: At telmination orthe test, the percentmortalities were:



### - Control 52 ppm 520 ppm 5200ppm



### - Percent M

### onality 

### 23 17 19 17



### - /I that died 17175 13175 14/75 13175

.. Mortalities were not the result ofthe treatment substance sll1ce the mortahtles did not occur 111 a dose-responSive pattern and the percent mortality in all treatment groups was less than that ofthe control on the last 2 days ofthe test. A small number ofimmobilebeetles were observed during the tests. All other surviving beetles in the 52, 520, and 5200 ppm treatment groups were normal in app<:arance and behaviorthroughout the study. Result:; ()

## rplate counts for nominal test concentrations used in the study were 3.6 x 104
, 3.6 x 105, alld .l,b x 10" CFU/mL, respectively. Samples collected on the day aftest initiation (Day 0) demonstrated mean values of2.3 x 104.2.7 X lOS, and 1.3 x 10" CFU/mL, respectively. 14EPA's Records Disposition Schedule PEST 361 Scientific Data Reviews HED Records Center - File R139870 - Page 15 of 34

Stability samples from Day 7 demonstrated mean values of2.2 x 103,4.0 X 103 , and 1.5 x 105 CFUimL. Dav 0 verification and Day 7 stability samples from the negative control diet showed 0.0 CHJ/mL Reviewer C:OI)ll11Cnts: A. Methods This study "as conductedin compliance with Good Laboratory Practice Standards as published by the U.S. i-:t'vironmental Protection Agency, Office ofPesticide Programs in 40 CFR Part 160. I. Till> study did not include two controls: an infectivity control group (inactivated GliocladiulIl cUlcnulatum) and 

### atreatment in which ladybird beetles are dosed with sterile filtrate from producti,)]) cultures. However, absence ofthese treatments did not influence the overall conclusions oC the study. B. Results and Discussion The pattem 01 mortalities exhibited did not establish that expected ofatypical dosage response. Ladybird bectle mortalities were noted in all treatments, including the negative control; cumulative murtalities were slightly greater in the negative control group than 

### in any ofthe groups treated with logarithmic increments ofGliocladium catcnulatum cell mass. The dietary LC50 value I,)r ladybird beetles exposed to G. catenulatum cell mass for 16 days was estimated to bc greater tnan 5200 ppm (3.6xI0" CFt;/mL), the highest concentration tested, and the no observed elTcd concentration estimatedto be 520 ppm (3.6x 105 CFU/mL). An LD,o >3.6xI0" CFU!mL and an NOEL of3.6x 105 CFU/mL was reported When comparedto the negative controls, resul1s showed a lack oftoxicity ofGliocladium catcnulatum cell mass to lady beetles.

15EPA's Records Disposition Schedule PEST 361 Scientific Data Reviews HED Records Center - File R139870 - Page 16 of 34

DATA EVALUATION REPORT Reviewed by: Robyn Rose, Entomologist, BPPD

### (.~ 

# t?IJ:IA Secondary Reviewer: Gail Tomimatsu, Plant Pathologist,**BPPD~~,\"_'''':+v'** i, (', STUDY TYPL: MRlDNO,. CHEMICAL NO.: TEST MATERIAL: STUDY NO
. SPONSOR TESTING FACIUTY: TITLE OF REPORT:
AUTHORS STUDY COMPLETED: CLASSIFIC.A.T10N: Study Summary: Honey bee (Apis mellifera) Dietary Pathogenicity and Toxicity Study (154A-23; OPPTS 885.4380) 445(J14-07 021009 Gliocladium catenu/alum strain J1446 (Technical Powder) 293-110 Kemira Agro Oy, P.O. Box 330, Helsinki, Finland Wildlife International Ltd., 8598 Commerce Dr., Easton, MD 21601 Gliocladium catenuialum Cell Mass (Technical Product): A Dietary Pathogenicity and Toxicity Study With The HOlley Bee (Api.\' melli{era) Kimberly A. Hoxter, Susan

## .r. Palmer, and Henry O. Krueger, PhD, 2/5/08 Acceptable.
Objective: "The objective ofthe study was to evaluate the pathogenicity and toxicity of Glioe/adium (cilcl1uiatum cell mass administered to the honey bee (Apis mellifera) in the diet." Test Material: GliocladiulII (il/cl1uiatum Cell Mass (Technical Product) Batch No. 44/95,,1; a light brown powder with a reported activity of7x 108 colony fonning units pergram (cfu/g) was received from Kemira ·\gro Oy (3/26/97; Wildlife InternationallD#3996). Test substance viability was determined I'rom 0.1 g test substance 

### + I mL 0.3% TRITON-X sol. 

### +99mL sterile H"O=I 0-3 dilution. The mixture was homogenized for -10 sec. before preparing final solutions (105, 1O-", 10-7 serial dilutions). The activity ofthe G. catcnuiatum was verified at Wildlife International using the equation: Viability (in terms ofCFU/g) =

# LC,IV, x N x

# L(0,) [cfu/g];**c,~#** of colonies 011 the agar plate, V",ovolume plated(mL), N=# ofreplicate plates, D"o'dilutions ofthe samplc(g/mL, Test Organisn! Honey bee (Al'is melli/era), an importantplant pollinator, obtained as capped brood from hives maintained b\ Wildlife Intemational Ltd. and maintained according to practices recommended by the State ,)!'Maryland.
16EPA's Records Disposition Schedule PEST 361 Scientific Data Reviews HED Records Center - File R139870 - Page 17 of 34

Test Methods: The pathogenicity and toxicity ofGliocladium catenulatum cell mass fed to honey bees was evaluated. Three nominal concentrations (52,520, and 5200 ppm) ofGliocladium and a negative control (honey and water) were fed to honeybees adlibitum via a honey and deionized water (20 mL) diet coated on a cOllon swab. The concentrations were equivalentto 3.6xI04, 3.6xl05, and 3.6x 10" respectively based on an activity of7x 10' colony forming units pergram (CFU/g). Viability ol'thetest substance was determined priorto test initiationin CFUlmL. The test substance Vias combined with 20mL deionized water and honey was added unlil200mL was reached; diets were prepared fresh and presented to the bees immediatelybefore test initiation and on Da}' 7 Twenly five apparently healthy bees (approximately 1to 3 days old) were aclded to each test chamber. Thebees were obtainedby removing frames containingpupae from hives on site. During a 3-d"y acclimationperiod before test initiation, the [rames were housed in clear acrylic boxes in an environmental chamber at an**averag,~** temperature of-32°C.; emerging bees were allowed to feed adlibitum on honey and pollen remaining in the cells ofthe frames. Three test chambers pertreatment were evaluated. Test chambers consisted ofone-pint rolled papel" containers (9cm diameter and 9 cm high) covered with a disposable plastic petri dish ("IOem diameter) with an inverted 20 mL glass vial containing the appropriate diet inserted through it, and another glass vial containing deionized water was also inserted through the lid. Honey bees were kept in tcst chambers at 26°C to 28°C**(x'~** 27.2 ± 0.2°C), rellativc humidity ranging from b2 to 96% (x = 77 ± 85), and total darkness except for when diet was replaced and observations were: made. Diets were available to the bees to feed adlibitum throughout the test perioa. Ohservations ofmortality and clinical signs oftoxicology were made 

### y, and I 3/4 hours after the initiation ofthe test and then daily for ten days. Observations were di.seontinued after the tenth day IYhen mortality exceeded 

### 20'Yo in the negative control. Samples ofthe diets for all treatment grollpS and the negative control were collected after preparation 011 Day 0 and after diets had been stored under test conditions onDay 7. These samples were assayed to determine homogeneity, test concentration measurements and establish Day 0 and Dav 7 values for evalluating stability ofthe test substancein the diets, and under test conditions. The 

### C"E'. Stephan computerprogram was used to analyze mortalityresults at test termination. The LOS II value and the 95% confidence interval (hy probit analysis), and the moving average method orbinomial probability with nonlinear interpolation were calculated via the C.E. Slephan computerprogram. The concentration resulting in no effects was determined through v'lsual inspection ofmortality and clinical observation.

17EPA's Records Disposition Schedule PEST 361 Scientific Data Reviews HED Records Center - File R139870 - Page 18 of 34

Rep0l1ed Results: At termination (lfthe test (after 10 days), the percent cumulative mortalities were: Control 52 ppm 520ppm 5200ppm Percent lV!ortalit)I 25 37 37 36 Ii that died 19175 28/75 

### 28175 27175 Bee mortalttlcs were noted first in the highest nominal concentrationtested (5200 mg/L) on Day 1ofthe teS1. Before the end ofDay 3, bee mortalities were noted in all treatments, mcluding the negative control. Percent mortalities increased erratically in all groups through Day 10. Mortalities were not the result ofthe treatment substance since the mortalities did not occurin a dose-reSpOnS1YC pattern and the percent mortality in all treatment groups was comparable to that ofthe control. A small numberofbees (l pertreatment) exhibited lethargy or loss ofequilibrium (unbalanced) III all groups on the last day ofthe test. Lethargic or unbalanced bees were noted also in the group treated with 520 mg/L on Day 5 and in the group treated with 5200 mg/L on Day 7.**Durill.~** the test, a small number ofthe bees in all treatments and one bee in the control group appeared lethargic or exhibited a loss ofequilibrium. The results ofthis study showed the dietalY LCso yalue for honeybees exposed to G. ca/enu/alum celi mass for 10 days to be greater than 5200 ppm (3.6x1 0" CFU/mL), the highest concentration tested and the no observed effect concentration to be 5200 ppm (3.6x10" CFU/mL). ;\11 

### LDso >3.6x 10" CFUlmL and an NOEL of3.6x 10' CFU/mL was reported. These values are not statistically significant. Viable plate counts for homogeneity/stability samples on Day 0 were 2.1 x 102
, 4.8 x 103, and (,.4 x 104 CFU/mL for nominal concentrations 52 (3.6 x 10" CFU/mL diet) , 520 (3.6 x \0'

# cn'irnL diet) and 5200 mg/L (3.6 x 10" CFU/mL diet) respectively. The Day 0 verification samples and tbe Day 7 stability samples for the negative control on Day 0 were 0.0 CFUhnL. Reviewer C\'lll!!l<:!ltS: A. Methods This study was conducted in compliance with Good Laboratory Practice Standards as published by the

## u.s. Environmental Protection Agency, Office ofPesticide Programs in 40 CFRPart 160. 1
.. Jhi., studydid not include two controls: an infectivity control group (inactivated Glivcladium (-ulel/ulalum) and a treatment in which honeybees are dosed with sterile: nltrate from producll()n cultures. However, absence ofthese treatments did not influence the overall conclLlsions Of1ilC study

J8EPA's Records Disposition Schedule PEST 361 Scientific Data Reviews HED Records Center - File R139870 - Page 19 of 34

2. ViabilIty determination, and stability/homogeneity oftest substance: The viability counts appear remarkably low for results ofthe nominal test concentrations. B. Conclusiolls The reported percent mOlialities did not reveal a pattern in response to logarithmic increments 01' test substances dosages, and although percent mortalities in the treated groups were slightly higher than in the negative control on the last 3 days ofthe test, they were not statistically Significant. Mortalities were therefore presumed to be associated with other environmental variables associated with the test arena. The dietary LCIO value for honey bees exposedto C. catenuiatlll1J cell mass for 10 days is greater than 520() ppm (3.6x10" CFU/mL), the highest concentrationtested and the no observed etTect concentration to be 5200 ppm (3.6XI06 CFUlmL). An LOso value >3.6x ilO" CFU/mL and an NOEL 01' 3.6'; 10' CFUlrnL were reported. These values are not statistically significant. There wcre no otherobservations ofinfectivity orpathogenicity.

19EPA's Records Disposition Schedule PEST 361 Scientific Data Reviews HED Records Center - File R139870 - Page 20 of 34

DATA E:VALUATION REPORT Reviewedby: Robyn Rose, Entomologist, BPPD 

# (L**d~** 

# 61Jlivl Secondary Reviewer: Gail Tomimatsu, Plant Pathologist, BPPo;**~:t,.·,\,-·c!"** 

## ,I .,,, STUDY TYPI-. MRID NO. CHEMICAL NO.: TEST**MATI~RIAL:** STUDY NO.: SPONSOR TESTING FACILITY:**TITLE~** OF RHORT:
AUTHORS: STUDY COMPLETED: CLASSIFICA liON: Study**Summllr~'::** Avian Oral Pathogenicity and Toxicity Study (154A-23; OPPTS 885.4050) 445014-0I 021009 Gliac/adium ('{{Ienu/alum strain Jl446 (Technical Powder) 293-109 Kemira Agro Oy, P.O. Box 330, Helsinki, Finland Wildlife International Ltd., 8598 Commerce Dr., Easton
.. MD 21601 Gilacladiwn calenu/alum Cell Mass (Technical Product): An Avian Oral Pathogenicity and Toxicity Study With The Northern Bobwhite Quail Kimberly A. Hoxter, Susan 

### .T. Palmer, and Henry O. Krueger, PhD. 5/29/97 Acceptable

Objective: "The objective ofthis study was to evaluate the acute toxicity and pathogenicity of GliacladiufII ""lcl/u/alum cell mass when administered orallyto northern bobwhite (Colinus virginianusi. Test Materia!: Gliacladiufll c:llellulalum Cell Mass (Technical Product) Batch No. 44/95; a lightbrown powder with a reported activity of7xl0scolony forming units pergram (cfu/g) was received from Kemira Agro Uy Ireported 3/26/97; Wildlife International TD #3723), and stored under refrigeration c,mditions. The viability ofthe test substance was verified byWildlife International Ltd. and was 6 % )< 10'cfu/g. Test Organism. Seeminglyhealthy northern bobwhite quail (Calinus virginianus) from a f10ck maintained by Wildlife International Ltd. that hatched September 9, 1996. Chicks were fed a vitamin supplement daily until test initiation. Waterfrom the local town (Easton) supply and feed were provided ad Iifoillll1l hefore and dunng the test. Prior to testing, birds were acclimated (the precise duration ofthe acclimationperiod was not reported) and observed daily for behavior or physical injuries. Only apparently healthy .2 I-day old birds were included in the test. 20EPA's Records Disposition Schedule PEST 361 Scientific Data Reviews HED Records Center - File R139870 - Page 21 of 34

Test l\1etho"lf,: The pathogenicity and toxicity ofGliocladium catenulatum cell mass fed tojuvenilebobwhite quail was evaluated basedupon procedures specified in OPPTS 885.4050 (previously the Pesticide Assessment Guidelines, FIFRA Subdivision 1\1, Section 154A-16). Birds were dosed daily for 5 days with a mixture ofthe test substance and reverse-osmosis (RO) water; ncgati\'c controlbirds were given onlyRO (1% v/w) water. The dosing suspensions for the treatment group were prepared daily by measuring a calculatedvolume ofthetest substance and mixing thoroughly with RO water. Dosing stocks were preparedbyweighing I() g oftest substance in 'I 100 mL beaker and sufficient amount ofRO water was added to bring the final volumc to 50 lilt and blended with a glass stir rod. During dosing, the stock was stin'ed constantly by placing on a magnetic stirplate. Eight pens With five birds each were evaluated. Two pens were used as negative controls (ten birds total) and six pens were used as treatments. A total of30 birds were individually treated with l.4x 1 ()" cfulg bodyweight over a five day dosing period; the resultant total dosage was approximated at 7 x 10" cfu/g ofbody weight over the five day period. All birds were weighed individually and administered the appropriate dosing suspension directly into the crop or proventriculus using alectubcrculin syringe. Treatment and control groups were given a total of1() mLlkg hody weight solution. No antibiotic medications were administered to the chicks during the 5tlIdv. Birds ofunknown sex were randomly assigned to pens that measured approximately 72x90x23 cm high. AII sides consisted ofgalvanized wire mesh and the walls and ceiling also had galvanized siIeeting. Floors ofthe pens consisted ofgalvanized wire mesh. Conditions during testing were 21.5o, 1.8°C (SO), with an average relative humidity of62%H2°"'i. Aphotoperiod of 1() hours 0 

## rflourescent light 1.0 approximate noon day sunlight was maintained by a time clock during acclimation and throughout the study. The birds were exposedto approximately 177 ± 55 lux ofillurt1lf1ation. In general, the National Institutes ofHealth recommended housing and husbandry practices were followed. Birds were evaluated for illness,. abnomlal behavior, and mortality at test initiation and twice daily until tcnnlllation onday 30. Bodyweights were determined after birds were dosed on days (), 1, 2, 3, 4
.. I ;. 18,25, and 30. Feed consumption was determined on days 4, 11, 18, 25, and 30. Attes! termination, birds were euthanized using carbon dioxide gas and subjected to gross necropsy.·j IS';UCS were examined for abnormalities, lesions, and any sign ofpathogenicity. Reported**Rcsult~:** There were 110 Illortalities nor abnormalities observed in the negative control group. Therewere no mortalil:!cs. obvious signs oftoxicity, pathogenicity, abnormal appearance orbehavior demonstrated hy Gliocladium-treated birds. Ofthe treated birds, one bird fractured awing on day 25 and ()i' bird demonstrated subcutaneous emphysema ( a pocket ofair under the skin) from 21EPA's Records Disposition Schedule PEST 361 Scientific Data Reviews HED Records Center - File R139870 - Page 22 of 34

days 15-30. l\"itherofthese occurrences were treatmentrelated. There was also no effect of GliociadiulII clltenulatum cell mass treatment on body weight orfeed consumption of1:he birds. Gross necropsy ill1dings included a bird inthe negative control group with a fractured right humenls; and a bird in the Gliocladium-treated group had subcutaneous emphysema in the abdominal and thoracic regions and pale breast muscle with petechial hemorrhaging. These results were considered unrelated to treatment. There were no evidences oftest substance replication, lesions, plaques orother indications ofpathogenicity for the groups treated with GliocladiulII ('iI{('lmlatum cell mass . Reviewer COml1lS'Ilts: A. Method" 1. This study was conducted in compliance with Good Laboratory Practice Standards as published by the U.S. Environmc:ntal Protection Agency, Office ofPesticide Programs in40 CFR Part lbO, except: Verification ofthe test concentrations, stability, and homogeneity ofthe tcst substance i11 deionized waterwere not determined. 2. This study did not includetwo controls: an ini,;ctivity control group (inactivated Gliocladium catenuiatul1I) ancl a treatment in which birds arc dosed with sterile filtrate from production cultures. Hnwcvcr, absence ofthese treatments did not influence the overall conclusions ofthe study B. Results and Conclusions Under conditions ofthe reported study, no adverse effects were exhibited byjuvenile northern bobwhite aficr .10 clays when administered daily doses (by oral gavage) of 1 x 104 (;{iocladium catenuiatul1l··cftug body weight/day for a five day period. This dosage is approximately equivalent to a total dosage of7 :< 

### 10" cfu/gbody weight for 5 days. The no observed effect dosage ofGliucl"dium catellulatum cell mass was determined at approximately 1.4 x 10" cfulg body \Veight t,)r tive days. There were no evidences oftest substance replication, lesions, plaques or otherindlcallons ofinfectivity or pathogenicity for the groups treated with Gliocladium cl1tenuil1tul1l cell mass in gross necropsies. This study showed an lack oftoxicity or pathogenicity ofGllOc!adillll, (utenullltum cell mass to northern bobwhite quail.

22EPA's Records Disposition Schedule PEST 361 Scientific Data Reviews HED Records Center - File R139870 - Page 23 of 34

DATA EVALUATION REPORT Reviewed by: Robyn Rose, Entomologist, BPPD 

## fI,IL 'J,

## :;;1.\1**,j~** " 

## Ie' Secondary Reviewer: Gall T0I11Imatsu, Plant PathologIst,**BPPD:'l~"h** \ ,I
STUDY TY!'!·: MRIDNO.: CHEMICAL "-0.: TEST MATERIAL: STUDY NO.: SPONSOR TESTING i',\CILlTY: TITLE OF REPORT:
AUTHORS STUDY COMPLETED: CLASSIFICA nON:
STUDY SUMMARY: Rainbow Trout Toxicity and Pathogenicity Test (154A-19; OPPTS 885.4200) 445014-02 021009 (;lioc/adium catenulatum strain Jl446 (Technical Powder) 293A-I04 Kcmira Agro Oy, P.O. Box 330, Helsinki, Finland Wildlife International Ltd., 8598 Commerce Dr., Easton, MD 21601 Gliocladium catenalatum Cell Mass (Technical Product): A FiveConcentrationToxicity and PathogenicityTestWith the Rainbow Trout (Oncoryhynchus mykiss)-Static Renewal SusanJ. Palmer. and Henry O. Krueger, PhD. 211 0/98 Acceptable

Objective: "The objective ofthis study was to evaluate the toxic and pathogenic ct1ixts of Gliocladiw." c"tenulatum cell mass (technical product) to the rainbow trout (OnchoIJ}/lchus mykiss) dun1lf! a 30-day aqueous and dietary exposureperiod under static-renewal tcst conditions
..' TestMaterial: Glioc/adnan Ccilcl1ulatum Cell Mass (Technical Product) Batch No. 44/95:; a light brown powder witb a reporrcd activity of7xIOX colony fonning units per gram (cfu/g) was received from Kemira Agro Oy (3/26/97; Wildlife International ID #3996). Jest substance viability was determined 11'0111 0.1g test substance 

### + ImL 0.3°;', TRITON-X sol. 

### + 99mL sterile H,o=10-3 dilution. The mixture was homogenized for**~**I0 sec. before preparing final solutions (10-', 10-", 10'). The activity ofthe G. eatenu/alum was verified at Wildlife International using the equation: Viahllity (in tenns ofCFU/g) = IC/V, x N x I (D,,) [cfll/g]; 

### C,,=# ofcolonies on the agar plate, 

### 'v.- volume plated(mL), N=# ofreplicate plates, D,,=dilutions ofthe sample(g/mL). Test Organism: Juvenile rainhow trout (Onehorvnchus mykiss) obtained as green eggs and sperm from Mt. Lassen Trout I ann, Red Bluff; California on Aprill )(), 1997 were used in this tcst. '\iegativeEPA's Records Disposition Schedule PEST 361 Scientific Data Reviews HED Records Center - File R139870 - Page 24 of 34

control fish measured at the end ofthe test were 36 to 40 mm (x = 39 mm) in length and weighed 0.70 to 1.3 g 

### !< .0 I.I g). Fish wereheld for 40 dlays afterhatching and allowed to acclimateto test conditions li)r 52 hours priorto testing. For 14 days priorto testing, fish wereheld at 11.8°C

- 12.5"C,**8.~** ... S.4 pH, and 9.0

- 9.9 mg/L dissolved oxygen. Priorto a 7(, hour starvation period before test lIlitiation, rainbow trout were fed salmon mash and/or salmon starter supplied by Zeigler Bros. Inc., Gardners, PA. Apparently disease and stress lree fish that were from the same source. vearclass, and approximate standard length were used for testing. Test Methods: This test was conducted according to guidelines outlined in OPPTS 885.4200 (previously OPP Section ]S4A·](), SubdivisionM ofthe Pesticide Testing Guidelines) and ASTM Standard E729-88a, Standard Guide for Conducting Acute Toxicity Tests with Fishes, Macroinvertebrates and Amphlbi'lI1s. Test chambers consisted of38-L glass aquariacontaining 15 L oftest solution measuring 12 em deep. Chambers were kept in a controlled room at 12±1°C, 16 hours offlourescent light similar to natural sunlight and eighthours ofdark regulated by a timer, and gently aerated throughout the test. This 1.es! was conducted for 30 days. A YeUow Springs Instrument Model 51 B dissolved oxygen meter and a FisherAccumet Model 915 pH meterwere used to measure dissolved oxygen and pH respectively in each test chamber at thebeginning and end ofobservations. A Yellow Springs Instrument Model 33 Salinity-Conductivity- Temperature meterwas llsed to detemline specllic: conductance and hardness and alkalinity measurements, and were based on procedures in Standard Methods for the Examination ofWater and Wastewater. The routes or({'st substance administration were aqueous and dietary. Forthe aqueous exposures, the lestmaterial was pre-welted and mixed thoroughly in dilution wateron a weight to volume

# b,,,,s hefore adding to the test chambers. Aftermixing, it was reportedthat test Solutl()nS appeared tan in colorand were cloudy with particulate matterthroughout the water column; and lhat the cloudiness increased [romthe lowest to higherconcentrations. Test solutions were prepared andrenewed every Monday, Wednesday, and Friday in separate aquaria that trout were transferred into. Waterwas obtained from 

### a 40m deep well located at Wildlife International Ltd and charactenzed as moderately-hard. Pmticlcs largerthan 2511m were removed by"tiing through a sand filter. Diet was prcpared by adding 6.6 mg ofGlioc/adium catenu/atum to 100g oftrout chow. The diet was mixed for 1.5 hours on ajarmill rotating at 50 rpm. Trout chow and 4.62x 107 cfu/kg was the resullant mixture. The diet was provided in each tcst chambertwice daily. Control fish wereprovided with untreated trout chow. One test chambercontaining ten trout were evaluated for each concentration and control. Nominal concentrations ofthe technical productwere 45,89, 179, :l57, anO 714 mg/L. Based on an activity of7:< 108cfu/g (verified by laboratory), these concentratlon-. clrc equivalent to 3 15 x1 07, b.23>: 107
, 1.25xI0', 2.50X 10', and 5.0x 11)" cfu/L, rcspectivelv.
24EPA's Records Disposition Schedule PEST 361 Scientific Data Reviews HED Records Center - File R139870 - Page 25 of 34

Water samples ofnew and old test solutions werc collected from each test chamber during the first week ofthe study; and then at weekly intervals for the remainder ofthe study. Samples were placed In sterile glass scintillation vials and analyzed immediately or stored refi-igerated (Oto SOC) up to 3 weeks until they couldbe processed and analyzed bydilution plating for viabilily. Serial dilutions ofthe Ilest solution wereprepared before spreading 0.1 mL onto plates containingpotato dcxtrose agar. After 3 days ofincubation at 28°C, all plateswere examined for growth and enumerated for viable Gliocladium cateuu/atum growth (CFU/mLl. Observations ofmortality and clinical signs oftoxicology were made daily throughout the test by sampling for dead or abnonmtl fish by running a dip net through the tankno more than three times. Watel· Illr all treatments was somewhat cloudy. Obscrvations onday I for the 179 mg/L concentration and on all days the 357 and 714 mg/L treatment groups were difficult becausethe water was panicularly cloudy. Therefore, a dip net was used to observe fish in these treatments. The actual number ofmortalities was determined at test tennination. A sterilenecropsywas perfornled on tllree fish from each treatment and control group to evaluate external or internal signs oflesions inflammation, areas ofdiscoloration orother abnonnalities. Fins, eyes, nares, oral cavity, anus, and gills were externally examined. Liver, kidney, heart, andbrain were evaluated fOI·necropsy. Samples from these organtissues were streakedonto plates ofpotato dextrose agar lor presence ofGliocladiwn catenulatum. The CE. Stephan computer programwas used to analyze mortalityresults at test tern1ination. The LD", vallie and the 95% con:fidence interval (by probit analysis), and the moving average method orbinomial probability with nonlinear interpolation wcre calculated via the C. E. Stephan computer program. The concentration resulting in no effects wasdetenninedthrough visual inspection ,",1"mortality and clinical observation.

A. Test Subsl,lllCC Viability and Verification ofTest Concentrations The aVGrage concentration from each group measured during the test were 8.3x 105
, 9.76X J()5, 2.32·!O", 5.58xlO", and2.08x 107 CFUiL (fornominal test concentrations 45, 89,179, 357 and 714 mgiL). "[TJhere was no measured activity inthe negative controls indicating that the negative l·omrol solutions were free ofcontamination." B. Water Chell1lstry Temperatures (measured continuously) ranged from approximately 11.5 to l3.0 DC; dissolved oxygcn levels ranged from 10.2 to 1.9 mgdissolved oxygen/L**(~94%** saturation in freshwater at l2°C), and pH ranged Irom 8.1 to 8.6 during the 30 day study. Specific conductance ofthe negative comrol ranged from 290 to 320 1l11lhos/cl1l (x**~** 303); alkalinity ofthe negative control solution ranged from 178 to 182 mg/L as CaCO) (x '0 180); and values for hardness ranged fonn 132 to do ( 134).
25EPA's Records Disposition Schedule PEST 361 Scientific Data Reviews HED Records Center - File R139870 - Page 26 of 34

C. Mortality and Clinical and Necropsy Observations There was 110 mOliality and fish appeared and behavednormal in the negative control and in the groups receiving 4:5,89, and 179 mgtest substanc:e/L treatments gletll's. There was no mortality in thc 357 mgl l treatment group but observations ofsublethal signs such as lethargy and lying on the bottom ofthe test chamberwith only gill movement were made the second week after test initiation. The 714 mg/L treatmentresulted in 100% mortality eight days after test initiation. Of the surviving lIsh examined for necropsy (3 from each ofthe treatment groups and negative control), all appeared internally and externally normal and there were no signs ofinfection. The 30 day LC,,, was 504mg/L (3.5xlO8 cfu/L). The lower 95% confidence limit was 357 mg/l (2.5xIOS cfull.) clnd upper confidence limit was 714 mg/L (5.0xI08cfu/L). Tissue samples collected from [ish in the negative control, 45 andl 89 mg/L treatment groups demonstrated no Gliocladium ('(/tenulatum cell mass (technical product) colony growth. The 179 mg/l treatment group showed () to 16 colonies forming (from heart, liver and brain tissues sampled) and 1to 24 colonies (from samples ofkidney, heart, liver and brain tissues) werc observed from lish tissues sampled from the 357 mg/Ltreatment group. Although the test substance was present, target organs did not appear to be affected, and there were no apparent effects upon the health ofthe fish. Reviewer Comments: A. Methods This study was conducted in compliance with Good Laboratory Practicc Standards as published by the U.S. EllVlronmental ProtectionAgency, Office ofPesticide Programs in40 C'FR Part 160. This study was conducted according to EPA guidelines outlines in OPPTS 885.4200 titled Freshwater Fish Testing, Tier I. Inclusion ofa control group offish exposed (aqueous and dietary) to sterile filtrates from production cultures ofGliac/adium catenulatum might supplementthe negativepathologies noted inthe studv; however absence ofthis control does not significantly alter the conclusions. B. Conclusion" The 1(1 day LC", was calculated as 504 mg/L (3.5xI()< cfu/L). The no-observed effect concentration W,IS 179 mg/l (1.3x10'cfu/L). Th" 30 day no mortality concentration was 357 mg/L (2.5xII)' cfu/l). Ther,: were no signs ofinfection in fish tissues examined for necropsy, and all appeared internally and**~xtemally** normal. Gliocladium cotcnulatum could not be recovered from tissue samples collected Irom fish in the negative control, 45 and 89 mg/L treatment groups; however several colony forming units ofG. catenulatum were recovered from the heart, liver and brain samples offish treated with 179 mg/L test substance, and from the kidney., heart, liver and brain samples of Cish treated with 357 mg/L test substance. Although the test substance was present, target organs did not appear to be affected, and there were no apparent effects upon the health of the fish. 2()EPA's Records Disposition Schedule PEST 361 Scientific Data Reviews HED Records Center - File R139870 - Page 27 of 34

DATA EVALUATION REPORT Reviewed by: Robyn Rose, Entomologist, BPPD

### I~J,- 

## ':";(:,J.,l

# lD\, Secondary Reviewer: Gail Tomimatsu, Plant Pathologist, BPPD

## ",-,~, 1-, III:>( ! STUDYTYPE MRIDNO: CHEMICAL NO.: TEST MATERIAL: STUDY NO.' SPONSOR. TEST1NG FACILITY: TITLE OF REPORT:
AUTHORS STUDY COMPLETED: CLASSIFICATION:
STUDY StlMMARY: Daphnia 21-Day Life-Cycle Toxicity and Pathogenicitv Test (154A-20; OPPTS 885.4240) 445014-03 021009 Gliociadiul1I catenulatum strain J1446 (Technical Powder) 293A-I03 Kemira Agro Oy, P.O. Box 330, Helsinki, Finland Wildlife International Ltd., 8598 Commerce Dr., Easton, MD 21601 Gliocladium catenulalum Cell Mass (Technical Product): A 21- Day Life-Cycle Toxicity and Pathogenicity Tesl! With The Cladoceran (Daphnia magna) SusanJ. Palmer, andHenry 0. Krueger, PhD. 211 0/98 Acceptable

Objective: "[he objective ofthis study was to evaluatethe toxic and pathogenic etTects of Giiociadiwil (([Ienulalum cell mass (technical product) onthe Cladoceran, Daphnia magna, during a 21 ··day exposure period under static-renewal test conditions." Test Materi.;ll: Gliodadiwl1 I <ltenulatum Cell Mass (Technical Product) Batch No. 44/95; a light brownpowder with all activity nf7x108 colony forming units per gram (cfu/g) was received from Kemira Agro Oy (3/26/97: Wildlife InternationallD#3996). Test substance viability was determined from O.lg (cst substance 

### + ImL 0.3% TRITON-X sol. 

### + 99mL sterile**H20=IO~'** dilution. The mixture was homogelllw:i for -10 sec. beforepreparing final solutions (10.5, I**O~",** 10"). The activity of the U catenllilllllnt was verified at Wildlife Intemational using the equation: Viability (in terms ofCFU/g)
. ::.:C/V" x N x 

## L (D,,) [cfu/g]; C"=#ofcolonies on the agar plate, V"=volume platedlmL). N'iIofreplicate plates, D,,=dilutions ofthe sample(g/mL). The test concentration was verified weekly by taking samples from alternating replicates from each treatment level. Test Orgal}ism: Neonate 1),("/1111([ magna «24 hOllrs old) obtained from cultures maintained by Wildlife Internatiol1.dl. [.ld
.. Easton, MD were tested as a representative aquatic invertebrate. Neonate 27EPA's Records Disposition Schedule PEST 361 Scientific Data Reviews HED Records Center - File R139870 - Page 28 of 34

juveniles were rcared and collected from 15 to 21 day old adult daphnids and used in this test. The adult daplmids werereared in water supplemented with selenium that was 20.6 to 2l.5 DC, 8.2 to 8.6 pH, and 8.4 to 9.0 mg dissolved oxygenJL, and showed no signs ofdisease or stress during the 14-day periodprecedingthe test. Test Methods: At test initiation, neonate daphnids were col1ected from cultures and transferred to IO-mL glass beakers contall1l1lg a small amount ofdilution water. Progeny from six adults were used inthe study. Ten daphnids were addeclto 12 separate 10 mL glass beakers containing a small amount ofdilution water. These daphnids were then transferred into a test chamber. Test chambers were 21 L glass aquaria containing 5 L oftest solution. Each test chamber consisted 01 Iwo compartments constructed from 500 mL glass beakers (,·8 em in diameter and 13 em in height) containing five daphnids for the first seven days ofthe study. A nylon mesh screen attached to the side oftest compartments allowed water was to flow through. On the seventh day, dapl111ids were individuallyplaced in a compartment in the test chamber. These compartments were -60 mm in diameter and 15 mm high culture dishes with 350 11m nylon screen attached to the sides (,,1() cm in height). Test solution was added to chambers to a height of-5.7 cm. Daphnids were fed three times daily throughout the test and once on the last day. A mixture ofyeast, Cerophyll®, and trout chow were feti to daphnids during culture and testing in addition to 

### Selenastrum caprlcornutum, freshwater algae. During the test, the target temperature was 20,,,1 DC (measuredby a hand·held thernlOmetcr) and the photoperiod was 16 hours oflight and 8 hours ofdark, controlled by an automatic tltner. The temperature was measured continuously in one negative control replicate using a Fulsco]Je ERJC Recorder. To aVOid sudden changes in lighting, a 30·mintransition period oflow light intensity was provided: light intensity during the test was approximately 350 lux at the wakr surface and provided by fluorescent tubes (Colortone®). Dissolved oxygen and pH were measured. in each lest chamber al the beginning and end ofthe test, daily for the first 7 days, and at the beginning and end ofone renewal period each week. Measurements ofpH were made using a Fisher Accumet Model 915 pH meter, and dissolved oxygen was measured using a Yellow Sprin,>;s Instrument Model 51 Bdissolved oxygen meter. Hardness and alkalinity and specitic conductance were measured in the dilution water at the beginning and end ofthe test, and at appruxllnately weekly intervals during the test. Test solutions wererenewed OIl a Monday/WcdJlC'sday/Friday schedule. Nominal concentrations ofthe test substance tested were 2.0x1O(', 3,).jO". 7.7xIO", 1.6xIO', and 3.Z<I07 CFU/L(2.8, 5J), 11,23, and 45 mg/L). The**MlUrc'~** ofwater used for testing was freshwater obtained from an approximately 40 m deep well located on Wildlifc International Ltd. site; and characterized as moderately·hard. The W.lter WCI' passed through a sand tilter to remove particles> 25 11m, pumped in 1.0 a 37,800 L storage tank ,lI1d aerated with spmy nozzles. Befbre testing, the water was filtered again and passed througl': all ultraviolet sterilizer and aerated overnight.


### 28EPA's Records Disposition Schedule PEST 361 Scientific Data Reviews HED Records Center - File R139870 - Page 29 of 34

Water samples ofnew and old test solutions were collected from each test chamber during the lI
..s! week ofthe study; and then also at weekly intervals from alternating replicates of new test solutions for the remainder ofthe study, Samples were placed in sterile**glas~;** scintillation vials and analyzed immediately or stored refrigerated (0 to 8°C) up to 3 weeks until they could be processed and analyzed by serial dilution plating for viability onloplates containing potato dextrose agar. After 3 days ofincubation at 28°C, all plates were examined for growth and enumerated for viable Gliocladiul11 catCl1u/atlll11 growth (CFU/mLl, Obs,ervations ofmortality, immobility, or clinical signs oftoxicity for first generation daphnids were recorded every Monday, Wednesday, and Friday during the test Daphnids were recorded as dead when a heartbeat was absent, coloration was white opaque, there was a lack of movement and response to gentle prodding. Signs ortoxicity included inabilityto maintain position in the water column, uncoordinated swimming, lethargy, opaque color and ceased feeding. Nl:Ollate daphnids were discarded when observations were made and at test termination. Length (from the apex ofthe head to the base onhe spine) and dry weight ofthe daphnids were measured at the end ofthe 21-day exposure period. Statistical analyseswereperformt'd on the survival ofthe first-generation daphnids on the 7"'
.. 14'h. and 21" days as well as onthe length and dry weight, and the number ofliveyoung produced by each surviving first-generation daphnids. A 2 X 2 contingency table was used to analyze survival data and significant differences between the treatment and control groups were detennined bv a chi-square test. The CB. Stephan program was used to calculate a 21-Day EC,O value. Reproduction and growth were evaluated for normality via the Shapiro Wilds test and homogeneity orvariance was evaluated using Bartlett's test. Bonferroni tcst determined statistical di ITcrences. Reported Results:: A. Test Suhslance Viability ActivJ[Y ofthe test substance, Gliocladiunl catenu/alum cell mass was measuredpriorto the test; and all nominal concentrations (see Methodology) were based 011 a verified and Sponsor-approved average of 4 x lOx 

### CFU/g test substance. The average Gliocladillin catenulalum concentration from each group measured during the test (average ofnew solutions for Days 2. 7 and 14) were 3.0>< 104, Ll XI0', 7,9xI05, 9.4x 10', and 3.3x1 0" CFUlL (0.04, 0,07, 0.16. i,l, I,. and4.7mg/L). B. Welter ('Ill'l11istry Measurements Measurements oftemperature were well within the range 0[20 ± 1°C (range: 20.1 to 20.6°('); dissllived oxygen levels were within the range of8.5 to 9.0 mg/L (? 93 % saturation at 20 0 e); and water pH ranged from 8.1 to 8 6.

29EPA's Records Disposition Schedule PEST 361 Scientific Data Reviews HED Records Center - File R139870 - Page 30 of 34

C. Mortality alld Survival, and Other Clinical Observations There were no significant differences betweenthe percent mortality in any ofthe treatment groups and negative control on Day 7 Crable I). On Day 14 there were no significant differences belween the 2.8 and 5.6 mg/Ltreatment groups and the control; howeverthere was significantly more mortality in the 11, 23, and 45 mg/Ltreatment groups than the negative control group (Table 

### I). Thetest was terminated after 21 days because mortality in the negative control group exceeded 20%. There were no signs ofimmobility orunusual behavior in surviving organisms through test termination. Therefore, Wildlife International concluded that the 2I-day IT," value for Daphnia magna exposed to Gliocladium catenuiatum cell mass (technicalproduct} was 7.8 mg/L. The 95% confidence limits were 5.2 and II mg/L. The LOEC was] I mg/L and the NOEC was 5.6 mg/L. Table 1. Summary ofCumulative Percent Mortality

Nominal Tesi Cur'cent IlL ) raliOn mg/L i ( 'F I :'\.egati\(' (,,1ntroI 2.8 (2.0:< II)" I 'FU/L) ,'i.e) (3.9>< 1 n{' I TI IIL)

- ; I (7.7x \.)1' { II! L)**2~(1'()xl'J** ( r't ilI.) 4\ (3.2"' ).' ( lUI.) -. Day 7 (%**mortalit~,)** 0 0 0 15 0 5 Day 14**(~':)** mortality) Day 21 (%, mortality)
5 ,W 5 :20 j J5 4() * 70 * ()O * 75 * 85 * 95 * *Percent mortalities significantly different (p s OJJ5) fro111 the negative controL Reproduction The ncgative control group produced an average of59 live young. The 2.8 mg/L Glioe/adiulII nllelluiatum cell mass treatment group produced 57 live young which was not statistically dtiTcrent from the control group. With 51 young adults produced, the 5.6 mg/L GliocladiulII ,·,tfenuiatum cell mass treatment group was statistically less (p :> 0.05) than the control group. Significant mortalities were noted in groups treated withi 1,23, and 45 mg test substancc/L, therefore, reproductionresults were not analyzed for these groups. The 21·day 

## Ee

## ,O value was 7.8 mg/L (-5.5 x IOI'du/L) w 95% confidence intervals of5.2 and II mgiL (3.6 x 10" and 7.7 x 10" dulL). The NOEC and the LOEC were 2.8 (2.0 x 10" cfu/L) and 5.6 mg/L (3.9 x 10" cfu/L), m;pectively. GrowthDapillll(is in the groups treated with 2.8 and 5.6 mg test substanceiL grew to a similar size as the dapbnids in the untreated group. The mean length and mean dry weight for adults inthe control group were 4.0 mm and 0.649 mg. Significant effects on survival were noted in groups 30EPA's Records Disposition Schedule PEST 361 Scientific Data Reviews HED Records Center - File R139870 - Page 31 of 34

recclving 11,23., and 45 mg test substancc/L treatments, therefore, growthresults were not analyzed statlsl1cally. The LOEC for growth was cstimated to be> 5.6 mg/L and the NOEC was 5.6 Illg/L. 

### Reviewer Comments: A. Methods This studywas conducted in compliance with Good LaboratoryPractice Standards as published by the U.S. Environmental Protection Agency, Office ofPesticide Programs in40 CFR Part I(,0 This study meets the guidelines outlined in OPPTS 885.4340 (updated from FIFRA Subdivision M, Series 154A-23). There were no verifications or observatiOons of infectivity. The inclusion ofa control group using sterile filtrate from production cultures mighthave aided in assessing pathogenesis. The absence ()fthis treatment docs not significantly alterthe overall conclusions, however, for hazard testing ofGliocladium catenulatum for the current intended uses. B. Conclm.ions Under conditions ofthe reported study, the 21-day ECsll value for survival fOor Daphnia magna exposed 10 Gliocladium catenulatum cell mass (technical product) was 7.8 mglL (5.5 x 10' dulL). The LOEC was 11 mg/L (3.6 x 10" cfu/L) and the NOEC was 5.6 mg/L. Ofthe three parameters assessed (survival, growth and reproduction), reproduction proved to he the most sensitive parameterin this study. The 21-day EC,o vaille was 7.8 mg/L**(~5.5** li. IO"clu/L); and theNOEC and the LOEC were 2.8 (2.0 x 10" cfu/L) and 5.6 rng/L (3.9 x 10" cfu/L), Ic,;pcctively.

31EPA's Records Disposition Schedule PEST 361 Scientific Data Reviews HED Records Center - File R139870 - Page 32 of 34

DATA EVALUATION REPORT Reviewed by Robyn Rose, Entomologist, BPPD 

## f2,t ':)(;! (

## I,) I Secondary Reviewer: Gail Tomimatsu, Plant Pathologist, BPPD'lf 

# &/z-'/of
STUDYTYPE: MRIDNO.: CHEMICAL NO.: TEST MATFRIAL: STUDYNO
.. SPOJ\SOR TESTING 1·'i\ClLITY: TITLE OF REPORT: AUTHORS. STUDY COMPLETED: CLASSIFI

### c;\nON: Study Summary:: Non-Target Plant Literature Search (OPPTS Guideline 885.4300) 44183-20 021009 Gliocladium calenu/alum strain 

### J1446 (Technical Powder) NIA Kemira Agro Oy, P.O. Box 330, Helsinki, Finland 'vVildlife International Ltd
.. 8598 Commerce Dr., Easton, MD 21:601 Gliocladium catenu/atum

- Literature Search for NontargetPlants Oili Mohammadi 11122/96 Acceptable

ObjC.f.tivc: Summary ofthe literature search on Gliocladiwn catenu/atum and plants Results of**;~carcll** A search on AGRICOLA, CA SEARCH, CAB Abstracts, BIOSIS PREVIEW. EMBASE, Enviroline, and Life Sciences Collection withthe key words Gliocladium catenu/atum and plants resulted in ,21 items found. Discussion: Ofthe 21 abstracts found, 14 related to the biological control ofGliocladium catenu/atunl against plant diseases, one related to nematode control, and two articles addressed pine seed stimulation. Fourabstracts reported Gliocladium catcnulatum isolated from diseased or dead planh. Two abstracts reported damping,·offofcoffee in Cuba, on reported a memberof the pea rool rot pathogen complex, and an isolation from dead birchtrees was reported. However, these articles did not show that Gliocladium catenulatum was the causal agent ofthe disease.**Prc,~nce** ofGlioc/ariium calenu{alum is usually a result ratherthan a cause ofthe infection. Reviewer (.·omments: Giiociadiwil calcllulatum is a fungal pathogen that controlsplant diseases such as damping-off, seed rot, root and stem rot, wilt and certain foliar diseases caused by Rhizoctonia, Pythium, Fusarium, /){{(nnella, Botrytis, Vcrticilliumc Alternaria, Cladosporium, Helminthosporium, Penicillium. and Pilcaria. Therefore, Gliocladium catenulatum is not expected to adversely 32EPA's Records Disposition Schedule PEST 361 Scientific Data Reviews HED Records Center - File R139870 - Page 33 of 34

effect non-target plant species. In addition a literature search conducted byKemira Agro Oy did not show any potential adverse effects ofCiliocladium catenulatum onnon-target plants. This literature review ofstudies relating to potential adverse effects ofGliocladium catenulatum on no-target planl species is sufficient to satisfy current guidelines for non-targetplants a\ this time.

33EPA's Records Disposition Schedule PEST 361 Scientific Data Reviews HED Records Center - File R139870 - Page 34 of 34

# ~ 13544



# • R139870

Chemical: Gliocladium ('atenulatuUl strain J1446

HED File Code: l\1emo nate: File ID: Accession #: PC Code: 021009 41300 BPPIl Eco Elfects 812112001 DPD244243 DPD267225 000-00-900I
HED Records Reference Center 3/2212007 

# •

•