United States Prevention, Pesticides EPA712-C-96-338
Environmental Protection and Toxic Substances February 1996
Agency (7101)
&EPA Microbial Pesticide
Test Guidelines
OPPTS 885.5200
Expression in a
Terrestrial Environment
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INTRODUCTION
This guideline is one of a series of test guidelines that have been
developed by the Office of Prevention, Pesticides and Toxic Substances,
United States Environmental Protection Agency for use in the testing of
pesticides and toxic substances, and the development of test data that must
be submitted to the Agency for review under Federal regulations.
The Office of Prevention, Pesticides and Toxic Substances (OPPTS)
has developed this guideline through a process of harmonization that
blended the testing guidance and requirements that existed in the Office
of Pollution Prevention and Toxics (OPPT) and appeared in Title 40,
Chapter I, Subchapter R of the Code of Federal Regulations (CFR), the
Office of Pesticide Programs (OPP) which appeared in publications of the
National Technical Information Service (NTIS) and the guidelines pub-
lished by the Organization for Economic Cooperation and Development
(OECD).
The purpose of harmonizing these guidelines into a single set of
OPPTS guidelines is to minimize variations among the testing procedures
that must be performed to meet the data requirements of the U. S. Environ-
mental Protection Agency under the Toxic Substances Control Act (15
U.S.C. 2601) and the Federal Insecticide, Fungicide and Rodenticide Act
(7U.S.C. I36,etseq.).
Final Guideline Release: This guideline is available from the U.S.
Government Printing Office, Washington, DC 20402 on The Federal Bul-
letin Board. By modem dial 202-512-1387, telnet and ftp:
fedbbs.access.gpo.gov (IP 162.140.64.19), internet: http://
fedbbs.access.gpo.gov, or call 202-512-0132 for disks or paper copies.
This guideline is also available electronically in ASCII and PDF (portable
document format) from the EPA Public Access Gopher (gopher.epa.gov)
under the heading "Environmental Test Methods and Guidelines."
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OPPTS 885.5200 Expression in a terrestrial environment.
(a) Scope—(1) Applicability. This guideline is intended to meet test-
ing requirements of the Federal Insecticide, Fungicide, and Rodenticide
Act (FIFRA) (7 U.S.C. 136, et seq.}.
(2) Background. The source material used in developing this har-
monized OPPTS test guideline is OPP guideline 155A-10.
(b) Test standards. This guideline is to be used with OPPTS
885.5000. (1) Tests shall be conducted in a greenhouse environment to
determine whether the microbial pest control abent (MPCA) is able to sur-
vive, persist, and replicate in a terrestrial environment consisting of soil
and vegetation representative of the proposed use site. The following pa-
rameters should be varied to determine their effect on the survival and
growth of the MPCA population:
(i) Temperature.
(ii) Humidity.
(iii) Precipitation (amount, frequency, pH).
(iv) Sunlight.
(v) pH (soil and foliar surfaces).
(vi) Nutrients (soil, vegetation).
(2) The values selected for each parameter should be selected to ap-
proximate the conditions expected at the intended use site.
(3) Laboratory studies designed to determine the microbial agent's
growth requirements (e.g., temperature, humidity, pH, sunlight and nutri-
ents) may supplement the greenhouse study described in paragraph
(d)(l)(i) of OPPTS 885.5000. Laboratory studies may demonstrate that the
MPCA will be unable to survive and the Agency will consider studies
on a case-by-case basis to meet the intent of testing in lieu of the green-
house study.
(c) Test substance. A typical end-use product or the technical grade
of the active ingredient shall be tested.
(d) Test duration. Data to establish a population decline curve shall
be collected at intervals until two half-life determinations have been made
or until data establish that the microbial agent population is able to main-
tain itself in the terrestrial environment at or above the level present imme-
diately after test initiation.
(e) Reporting and evaluation of data. The reporting and evaluation
provisions are the same as those set forth in OPPTS 885.5000.
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(f) Tier progression. If results of this study indicate that the MPCA
is able to persist in the terrestrial environment such that susceptible nontar-
get organisms tested in Tier I are likely to be exposed, then the appropriate
testing in Tier III (OPPTS 885.4550, 885.4600, or 885.4850) is required
as specified in 40 CFR 158.740.
(g) References. The following references contain information for de-
veloping acceptable protocols:
(1) Anthony, D.W.et al. Field test with Nosema algerae Vavra and
Undeen (Microsporida, Nosematidae) against Anopheles albimanus Wiede-
mann in Panama. Miscellaneous Publications of the Entomological Society
of America 11:17-28 (1978).
(2) Armstrong, J.L et al. A microcosm method to assess survival of
recombinant bacteria associated with plants and herbivorus insects. Cur-
rent Microbiology: in press (1987).
(3) Cunningham, J.C. Persistence of the nuclear polyhedrosis virus
of the eastern hemlock looper on balsam foliage. Insect Pathology Re-
search Institute, Sault Ste. Marie, Ontario, Canada. Bimonthly Research
Notes 26:24-25 (1970).
(4) Elgee, E. Persistence of a virus of the white-marked tussock moth
on balsam fir foliage. Maritime Forest Research Centre, Fredericton, New
Brunswick, Canada. Bimonthly Research Notes 31:33-34 (1975).
(5) Grison, P. et al. Microbial pesticides and environment. Annals
of Zoological Ecology of Animals 8:133-160 (1976).
(6) Harcourt, D.J. Persistence of a granulosis virus of Pieris rapae
in soil. Journal of Invertebrate Pathology 11:142-143 (1968).
(7) Hukuhara, T. and H. Namura. Distribution of a nuclear
polyhedrosis virus of the fall webworm Hyphantria cunea in soil. Journal
of Invertebrate Pathology 19:308-316 (1972).
(8) Ignoffo, C.M. et al. Stability of conidia of an entomopathogenic
fungus Nomuraea rileyi in and on soil. Environmental Entomology 7:724-
727 (1978).
(9) Ignoffo, C.M. et al. Vertical movement of conidia of Nomuraea
rileyi through sand and loam soils. Environmental Entomology 7:270-272
(1977).
(10) Jaques, R.P. The persistence of a nuclear polyhedrosis virus in
the habitat of the host insect Trichoplusia ni 11. Polyhedra in soil. Cana-
dian Entomology 99:820-829 (1967).
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(11) Jaques, R.P. Leaching of the nuclear polyhedrosis virus of
Trichoplusia ni from soil. Journal of Invertebrate Pathology 13:256-263
(1969).
(12) Jaques, R.P. Occurrence and accumulation of viruses of
Trichoplusia ni in treated field plots. Journal of Invertebrate Pathology
23:140-152(1974).
(13) Jaques, R.P. Occurrence and accumulation of the granulosis virus
of Pieris rapae in treated field plots. Journal of Invertebrate Pathology
23:351-359(1974).
(14) Kerr, A. Soil microbiological studies on Agrobacterium
radiobacter and biological control of crown gall. Soil Science 118:158-
172 (1974).
(15) Ladd, Jr. T.L. and P.J. McCabe. Persistence of spores of Bacillus
popilliae, the causal organism of Type A milky disease of Japanese beetle
larvae in New Jersey soils. Journal of Economic Entomology 60:493-495
(1967).
(16) Liang, L.N. et al. Fate in model ecosystems of microbial species
of potential use in genetic engineering. Applied Environmental Microbi-
ology 44:708-714 (1982).
(17) Lingg, A.J. and J.J. McMahon. Survival of lyophilized Bacillus
popilliae in soil. Applied Microbiology 17:718-720 (1969).
(18) Milner, R.J. and Lutton, G.G. Metarrhizium anisoplia: Survival
of Conidia in the soil. Proceedings of the First International Colloquium
on Invertebrate Pathology. Queen's University at Kingston, Canada, pp.
428-429 (1976).
(19) Morris, O.N. A method of visualizing and assessing deposits of
aerially sprayed insect microbes. Journal of Invertebrate Pathology
22:115-121(1973).
(20) Narayanan, K. et al. Preliminary observations on the persistence
of nuclear polyhedrosis virus of Spodoptera litura F. Madras Agricultural
Journal 64:487-488 (1977).
(21) Pinnock, D.E. et al. Effect of tree species on the coverage and
field persistence of Bacillus thuringiensis spores. Insect biological control.
Journal of Invertebrate Pathology 25:209-214 (1975).
(22) Roone, R.E. and R.A. Dacust. Survival of the nuclear
polyhedrosis virus of Heliothis armigera on crops and in soil in Botswana.
Journal of Invertebrate Pathology 27:7-12 (1976).
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(23) Swift, M.J. Microbial succession during the decay of organic
matter. In: Burns, R.G.; Slater, J.H. (eds.) Experimental microbial ecology.
Oxford: Blackwell, 164-177 (1982).
(24) Thomas, E.D. et al. The effect of soil pH of cabbage looper
nuclear polyhedrosis virus in soil. Journal of Invertebrate Pathology
21:21-25 (1973).
(25) Vankova, J. and M. Svestka, M. The field persistence and effi-
cacy of Bacillus thuringiensis formulations. Biological control of forest
pests. Anz Schadlingskd Pflanzenschutz 49:33-38. (English Summary)
(1976).
(26) Walter, M.V. et al. A method to evaluate survival of genetically
engineered microorganisms in soil extracts. Current Microbiology: in press
(1987).
(27) Walter, M.V. et al. Measuring genetic stability in bacteria of
potential use in genetic engineering. Applied Environmental Microbiology
53:105-109(1987).
(28) Wojciechlowska, M. et al. Duration of activity of
entomopathogenic microorganisms introduced into the soil. Polish Eco-
logical Studies 3:141-148 (1977).
(29) Young, S.Y. Pre- and posttreatment assessment of virus levels.
Pages 139-142 In selected papers from EPA-USDA Working Symposium.
M. Summers, R. Engler, L. Falcon, and P. Vail (eds.) American Society
of Microbiology, Washington, DC (1975).
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