MEMORANDUM

SUBJECT: Review of Toxicology Data for Metarhizium anisopliae F52 in Support of a 

Section Three Registration for Metarhizium anisopliae F52 from Taensa, Inc., 

Fairfield, CT.

FROM: Chris A. Wozniak, Ph.D., Primary Reviewer

Microbial Pesticides Branch

Biopesticides and Pollution Prevention Division

THROUGH: John L. Kough, Ph.D., Senior Scientist

Microbial Pesticides Branch

Biopesticides and Pollution Prevention Division

TO: Leonard Cole, Regulatory Action Leader

Biopesticides and Pollution Prevention Division

ACTION REQUESTED: 

Taensa Inc. has applied for a Section 3 registration for indoor and greenhouse use of Metarhizium 

anisopliae strain F52. This fungus in an entomopathogen and will be used to control household 

and greenhouse insect pests. This is considered a non-food use and therefore no associated 

tolerance is requested at this time. The attached data evaluation reports were originally reviewed 

by Agency contract employees and were subsequently reviewed by BPPD scientists.

BACKGROUND:

The active ingredient, M. anisopliae Strain F52, will be produced using two different 

manufacturing processes in place at four different locations (manufacturers). M. anisopliae F52 is 

proposed for use in greenhouses and indoors on non-food applications. This deuteromycetous 

fungus has a host range primarily affecting coleopterans of the families Elateridae and 

Curculionidae, although other insect groups are known to be within the host range of this 

pathogen. This species is known worldwide from insects and non-insect sites, such as soil, river 

sediments, associated with nematodes, and as a saprophyte on organic detritus.
2

DATA REVIEW RECORD

Active Ingredients: Metarhizium anisopliae strain F52

Product Name: TAE-001 Technical Bioinsecticide

ID No.: 072098-T TAE-001 Technical Bioinsecticide

Submission No.: S569284

Chemical No.: 029056 Metarhizium anisopliae strain F52

DP Barcode: D271484

Sponsor: Taensa, Inc., 26 Sherman Court, P.O. Box 764, Fairfield, CT 06430

MRID/Study Titles: 448447-09 - Acute Oral Toxicity / Pathogenicity - Rat (885.3050)

448447-10 - Acute Dermal Toxicity / Pathology Study of Metarhizium 

anisopliae Strain 52 in Rabbits

448447-11 - Toxicity / Pathogenicity Testing of Metarhizium anisopliae

Strain 52 Following Acute Intratracheal Challenge in Rats

448447-12 - Toxicity / Pathogenicity Testing of Metarhizium anisopliae

Strain 52 Following Acute Intraperitoneal Challenge in Rats

448447-13 - Primary Eye Irritation Study of Metarhizium anisopliae, Strain 

F52, in Rabbits

448447-15 - Dermal Sensitization Study of Metarhizium anisopliae, Strain 

F52, in Guinea Pigs Using the Buehler Method

SUMMARY OF REVIEWS:

448447-09 - Acute Oral Toxicity / Pathogenicity - Rat (885.3050) - Fifty-six males and 56 

female CD Rats, approximately 58 days old, were divided into treatment group (TG), killed 

treatment group (KTG) and naive control (NC) groups. A shelf control (SC) group was also not 

treated and housed with the TG group. TG group rats were dosed orally with approximately 1.04 x 

108cfu of Metarhizium anisopliae per animal in a 1 mL volume. KTG rats were treated with heatkilled test substance and naive control group rats were not dosed. Body weights were recorded at 

the time of dosing and on days 3 and 7. The test animals were observed for clinical signs twice 

daily. The animals were fasted overnight prior to sacrifice for test material enumeration from 

tissue. Three rats/sex/group from the TG, KTG, and NC groups were scheduled to be killed and 

necropsied on days 0, 3, 7, 14, and 21. Due to the clearance of the test material by day 3, the 

animals scheduled to be sacrificed on days 14 and 21 were killed on day 7 and not necropsied. 

The SC group rats were necropsied on day 7. The blood, brain, lungs, spleen, liver, kidneys, 

mesenteric lymph nodes, stomach and small intestines, cecum, and feces were removed for 

microbial enumeration and weight determination. Samples from these tissues and organs 

recovered the test substance on day 0, but by day 3 the test substance was below the detection 

limit. Metarhizium anisopliae Strain F52 does not appear to be toxic and/or pathogenic in rats, 
3

when dosed at 1.04 x 108cfu/animal. 

Classification: Acceptable.

448447-10 - Acute Dermal Toxicity / Pathology Study of Metarhizium anisopliae Strain 52 in 

Rabbits - Five male and five female New Zealand white rabbits were received from Kuiper 

Rabbit Ranch, Gary, IN. The male and female rabbits weighed 2.3-2.6 kg and 2.3-2.4 kg, 

respectively, on the day of treatment. The test material (2000 mg/kg body weight; 3.63-4.42 x 

1010 cfu/animal) was applied evenly to the clipped dorsal trunk in an area of approximately 10% 

(about 240 cm2). The application site was covered with a surgical dressing and a plastic film, then 

secured with a lint-free cloth and an elastic adhesive bandage. The skin was checked for irritation 

within one hour after patch removal and daily thereafter for 14 days. The rabbits were weighed 

prior to treatment and on days 7 and 14. The rabbits were euthanized on day 14 without gross 

necropsy. No sign of overt toxicity was noted from any rabbit. Very slight to severe erythema was 

noted on all rabbits after patch removal with clearance by the second week. Very slight to 

moderate edema was noted on one male and five females after patch removal with clearance by

day 6 of the study. Eschar, scabs, cracking skin, and/or scaly skin were noted on a few animals 

during the first week of the study. All animals recovered from these irritation symptoms before 

the end of the study. No deaths and no clinical toxicity were observed. The acute lethal dose 

(LD50) is greater than 2000 mg/kg. 

Classification: Acceptable - Toxicity category III.

448447-11 - Toxicity / Pathogenicity Testing of Metarhizium anisopliae Strain 52 Following 

Acute Intratracheal Challenge in Rats - Eighty male and 80 female CD®rats, approximately 70 

days old, were assigned to groups: treatment group (TG), killed treatment group (KTG), naive 

control (NC), and shelf control (SC). The TG group rats were dosed with approximately 1.17 

108cfu/0.1 mL/animal. Body weights were recorded weekly (0, 7, 14, 21, 28, and 35). The test 

animals were observed for clinical signs twice daily. Five rats/sex/group from the TG, KTG, and 

NC groups were killed and necropsied on days 0, 7, 21, and 35. The SC group rats were 

necropsied on day 35. The blood, brain, lungs and associated lymph nodes, spleen, liver, kidneys, 

and cecum were removed for microbial enumeration and weight determination. Mean body weight 

of males and females and mean body weight gains of males indicated no statistically significant 

effects in any test groups during the study. The mean body weight gains of the females in the TG 

group were statistically lower than the control groups between days 14 and 21. One TG male (No. 

515) and one NC female (No. 621) lost weight during the first week. One KTG female (No. 605) 

and one NC female (No. 626) lost weight during the second week. One TG female (No. 586) lost 

weight during the third week. One TG female (No. 590) lost weight during the fourth week. No 

adverse signs were observed from any group. Three females (Nos. 585, 586, and 589) in the TG 

group that were sacrificed on day 21 showed brown and/or mottled lungs. All other rats had no 

gross lesions. Decreased relative spleen, kidney, brain, and liver weights appeared to be a 

transitory effect. No statistically significant effects in relative organ weights in other tissues were 

observed. Since these weight differences were transitory, the observation is considered as not 

biologically meaningful and likely reflects the normal variation present in organ weights of 
4

animals. Metarhizium anisopliae was only detected in the rats treated with the test material (TG 

group). Metarhizium anisopliae was detected in the lungs and associated lymph nodes of males on 

days 0 and 7 and were below the detection limit by day 21. The test organism was detected in the 

lungs and associated lymph nodes of females on days 0, 7, and 21 and were below the detection 

limit by day 35. The test organism was detected in the cecum of males and was below the 

detection limit by day 7. The test organism was detected in the spleen of females and was below 

the detection limit by day 21. Metarhizium anisopliae, Strain F52 does not appear to be toxic 

and/or pathogenic in rats, when dosed intratracheally at approximately 1.17  108cfu/animal.

Classification: Acceptable.

448447-12 - Toxicity / Pathogenicity Testing of Metarhizium anisopliae Strain 52 Following 

Acute Intraperitoneal Challenge in Rats - Thirty-nine male and 39 female CD® rats, 

approximately 51 days old, were assigned to groups: treatment group (TG), killed treatment group 

(KTG), naive control (NC), and shelf control (SC). The TG group rats were dosed with approximately 1  107

cfu / mL/animal. Three rats/sex/group from the TG, KTG, and NC groups were 

killed and necropsied on days 0, 3, 7, and 14. The SC group rats were necropsied on day 14. The 

blood, brain, lungs and associated lymph nodes, spleen, liver, kidneys, and cecum were removed

for microbial enumeration and weight determination. No adverse signs were observed from any

group. No gross lesions were noted from any rat. Decreased relative spleen, brain, and liver 

weights all appeared to be transitory in effect. Based upon the overall weight gain and lack of 

pathogenic effects noted upon necropsy, the differences in organ weights are not considered as 

resulting from primary treatment with M. anisopliae F52. No statistically significant effects on 

relative organ weights for other tissues were observed. Metarhizium anisopliae, Strain F52 does 

not appear to be toxic and/or pathogenic in rats, when dosed intraperitoneally at approximately 1 

107cfu/animal.

Classification: Acceptable.

448447-13 - Primary Eye Irritation Study of Metarhizium anisopliae, Strain F52, in Rabbits -

Three male and three female New Zealand white rabbits were treated with test material (0.1 g with 

6.3 x 108cfu/eye/animal), applied in the everted lower lid of the right eye, and the eye held closed 

for approximately 2 seconds. The eyes were rinsed with lukewarm water 24 hours after test 

material instillation. The contralateral eye served as control. The eyes were examined and scored 

1, 24, 48 and 72 hours and 4, 7, 10, 14, 17, and 21 days after test material instillation. No deaths 

were observed in any of the dosed test animals. Excluding one rabbit because of an unrelated 

infection, three rabbits had corneal opacity 24 hours after test material instillation. Four rabbits 

had iritis and five rabbits had positive conjunctival irritation one hour after test material 

instillation. By day 4, no irritation was noted on any rabbits with the exception of the male 

tagged as No. 497. Since rabbit No. 497 was checked for infection by swabbing and culture onto 

TSA and Sabouraud media and no M. anisopliae was recovered, the infection in this animal is 

considered to be unrelated to treatment with the test substance. Eliminating rabbit No. 497 from 

the calculations, the highest primary eye irritation score was 23.2 at 24 hours after test material 

instillation. The lack of test substance in the rabbit No. 497 indicates the secondary infection 

responsible for the substance was not primarily caused or associated with treatment with M. 

anisopliae F52. Metarhizium anisopliae Strain F52 at 0.1 g/animal (6.3 x 108cfu/animal) was 
5

moderately irritating causing corneal opacity, iritis, and conjunctival irritation with resolution by

day 4. 

Classification: Acceptable. Toxicity category III for eye irritation.

448447-15 - Dermal Sensitization Study of Metarhizium anisopliae, Strain F52, in Guinea 

Pigs Using the Buehler Method - The backs of 40 male Hartley guinea pigs were clipped one 

day prior to each treatment. For the induction phase, 0.3 g of the test material (2.37 x 109cfu 

Metarhizium anisopliae Strain F52) was applied with an occlusive 25 mm Hilltop Chamber for 

six hours once each week for three weeks to the pre-moistened clipped skin of 20 test animals. 

Two weeks after the third induction, the animals were challenged with 0.3 g of test material under 

occlusion to naive sites for 6 hours. A naive (negative) control group (10 animals) was treated 

with 0.3 g of test material at challenge (i.e., two weeks after application of the third induction 

dose) only at the same site as the treated guinea pigs (i.e., lower left quadrant). The ten positive 

control animals were induced with 0.3 mL of alpha-hexylcinnamaldehyde (HCA) and challenged 

with 0.3 mL of 50 % w/v HCA in acetone. All wrapping materials were removed 6 hours after the 

challenge dose and the test site rinsed with water and wiped with gauze. Reactions were scored 24 

and 48 hours after the first induction and challenge. No deaths were observed in any of the guinea 

pigs. The animals had normal body weight gains. After three consecutive inductions, 1/20 treated 

test animals had a positive reaction 48 hours after challenge. No reactions were noted on any

naive control animal. As for the positive control animals, 6/10 and 7/10 animals had positive 

reactions at 24 and 48 hours, respectively, after challenge. Based upon the submitted data, 

Metarhizium anisopliae Strain F52 does not appear to be a dermal sensitizer when induced and 

challenged at 2.37 x 109cfu.

Classification: Acceptable. MaF52 is not considered a dermal sensitizer.