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OCR for page 121
6
Answers to Charge Questions
On the basis of the reviews presented in Chapters 2-5, the committee con-
cluded that the available studies of Fusarium oxysporum f.sp. cannabis, F. ox-
ysporum f.sp. erythroxyli, and Crivellia papaveracea or Brachycladium pa-
paveris are preliminary and exploratory. Thus, in answering the specific charge
questions, the committee attempted to capture what conclusions could be drawn
from the limited dataset and what might be expected on the basis of what is
known about fungal pathogens and mycoherbicides in general.
EFFECTIVENESS
Are the drug crops (cannabis, coca, and opium poppy) known to be
susceptible to the proposed mycoherbicides?
o The committee found documented evidence from greenhouse and
small field studies that cannabis (Cannabis sativa), coca (Erythroxy-
lum coca and E. novogranatense), and opium poppy (Papaver som-
niferum) are susceptible to F. oxysporum f.sp. cannabis, F. ox-
ysporum f.sp. erythroxyli, and C. papaveracea/B. papa-veris, re-
specttively. However, the few studies available involved controlled
applications to the target plants and often used artificial environ-
mental conditions that favored infection. Thus, the committee
judged that the information is insufficient to conclude that the pro-
posed mycoherbicides would be efficacious for the control of illicit-
drug crops under field conditions.
Have the mechanisms of action of the proposed mycoherbicides’
toxicity to illicit-drug crops been established?
o The types of diseases produced by the proposed mycoherbicide
strains are wilt diseases in cannabis and coca and a blight of the ae-
rial parts of opium poppy. Some members of the fungal species to
which the proposed mycoherbicide strains belong can produce tox-
121
OCR for page 122
122 Feasibility of Using Mycoherbicides for Controlling Illicit Drug Crops
ins involved in disease development under laboratory conditions, but
no reliable data are available on the proposed mycoherbicide strains.
The molecular mechanisms underlying the host-pathogen interac-
tions are unknown, and the nature of disease progression and fungal
spread after disease onset, which are critical determinants of myco-
herbicide efficacy, are not documented.
Are the proposed mycoherbicides host-plant-specific?
o The proposed mycoherbicide strains can cause disease in their target
plants, including those grown legally and those which occur natu-
rally near the illicit crops. However, the few host-range studies con-
ducted with nonrelated species were of little value because they only
report that the mycoherbicide strains did not cause disease in some
native plants and crop species and do not provide experimental de-
tails (or in some cases even the names of the plants). Furthermore,
none of the available studies used a standard, systematic process to
select the most relevant plants to test in host-range studies. For ex-
ample, of about 200 species of Erythroxylum native to South Amer-
ica, only two were tested for susceptibility to the coca mycoherbi-
cide. Thus, the data are insufficient to conclude that the proposed
mycoherbicides would not pose a risk to other plants or crops.
What quantities of mycoherbicides would be needed to eradicate il-
licit drug crops?
o Workable mycoherbicide formulations, delivery methods, and appli-
cation strategies need to be developed in partnership with an indus-
trial producer before this question can be answered. The committee
made estimates for a single application per hectare solely on the ba-
sis of published laboratory production methods, which may or may
not be realistic. Its estimates indicate that tens to hundreds of kilo-
grams of dry formulations per hectare or hundreds to thousands of
liters of liquid-spray formulation (containing billions to trillions of
spores) per hectare would be required for a single application of the
mycoherbicides.
How would the method of delivery affect the effectiveness of the
mycoherbicides in eradicating the drug crops?
o For all three mycoherbicides, on-ground application would be the
most precise and uniform. However, it is not feasible, because the
growers of illicit drug crops would be uncooperative and possibly
hostile. Aerial application is probably the most practical approach,
but it would need to be from altitudes greater than those normally
used for agricultural applications.
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123
Answers to Charge Questions
o F. oxysporum f.sp. cannabis and F. oxysporum f.sp. erythroxyli are
soilborne, root-infecting pathogens; for greatest efficacy, they
should be delivered to the soil. Aerial application of dry formula-
tions, such as prills and pellets, could reduce the efficacy of such
soilborne pathogens because the formulations would be subject to
scattering by wind, which would lead to nonuniform, discontinuous
placement of the inoculum over the target area and reduce the size of
the plant-pathogen interface.
o C. papaveracea/B. papaveris is a pathogen that attacks primarily the
aerial parts of opium poppy; for greatest efficacy, it should be
sprayed on the foliage of the plants. All tests of this pathogen have
used a wet application. Aerial application of liquid formulations
would be subject to wind-driven drift, which would lead to irregular
deposition on the target area. The large amount of water needed for
aerial delivery of the mycoherbicide could be an even more impor-
tant limiting factor. Low-volume and ultra-low-volume applications
have not been tried with the proposed mycoherbicides.
FEASIBILITY OF LARGE-SCALE MANUFACTURE AND DELIVERY
What sort of facility would be required for the large-scale Indus-
trial manufacture of the proposed mycoherbicides?
o An entity with industrial-scale liquid or solid-state fermentation and
formulation capabilities and capacity to produce sufficient tonnage
of the finished products per year would be needed for large-scale
manufacture of mycoherbicides. The experience gained in industrial
production of registered mycoherbicides could be adapted to pro-
duce the materials needed.
What sort of equipment and technology would be required for the
delivery of the proposed mycoherbicides on a large scale?
o Delivery of the mycoherbicides is expected to pose a challenge. The
most likely scenario is aerial application of the proposed mycoherbi-
cides from an aircraft capable of delivering a dry or liquid formula-
tion.
o On the basis of published data, the Fusarium mycoherbicides would
be best delivered as dry formulations and the Crivellia/Brachy-
cladium mycoherbicide as a spray. However, the latter appears im-
practical because of the amount of water required for spraying and
the possible inability to apply from low altitudes.
What is the overall technical feasibility of the large-scale industrial
manufacture of the proposed mycoherbicides?
o Technology for large-scale industrial production of commercial my-
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124 Feasibility of Using Mycoherbicides for Controlling Illicit Drug Crops
coherbicides against weeds exists. It probably could be adapted for
production of the proposed mycoherbicides. However, there are no
data on whether the proposed strains could be mass-produced on a
large scale.
What consideration would need to be made for large-scale delivery
of the proposed mycoherbicides?
o In addition to the equipment and technology required for large-scale
manufacture and application, consideration should be given to the
potential impediments to implementing the use of the mycoherbi-
cides. The impediments include the need for international approval
and cooperation with the countries in which the mycoherbicides are
to be used and the uncooperative and possibly hostile growers who
might try to prevent or counteract the effects of mycoherbicide ap-
plication.
What types of field trials of the proposed agents are needed? For
example, is testing in tropical or arid environments required?
o More epidemiological data on the proposed mycoherbicides from
experiments with consistent protocols are needed before extensive
field tests can be conducted. In general, the field-test conditions
should mimic the climatic and environmental conditions under
which the illicit-drug crops are grown, and the mycoherbicides
should be applied in a manner that simulates the intended delivery
method.
o The published data shed some light on the environmental conditions
that favor the mycoherbicides’ performance. This information is
useful for guiding further research. There would have to be several
on-site trials over a few years and in different areas where the drug
crops are grown to identify field conditions that are conducive to the
efficacious performance of the mycoherbicide and to establish the
best timing for application. This phase of research should be done in
cooperation with industry.
o As part of any future research program, ecoclimatic data on major
drug-producing regions should be gathered to select test sites and
regions where the mycoherbicides could be tested and used success-
fully. Climate-matching computer programs could help identify
geographic regions where the climatic conditions would be condu-
cive for the mycoherbicides to cause substantial damage on their
target crops.
PERSISTENCE IN THE ENVIRONMENT
How long are the mycoherbicides likely to persist in the environ-
ment after application?
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125
Answers to Charge Questions
o Persistence of the mycoherbicide fungi would depend on environ-
mental conditions at the application site and whether the type of
fungal inoculum (asexual or sexual spores, microsclerotia, pseudo-
thecia, or mycelia) applied would be able to survive environmental
stress. Simply determining the abiotic conditions that favor or re-
duce persistence under controlled environments is insufficient be-
cause persistence would also depend on interactions with other liv-
ing organisms and on environmental conditions that might not be
replicable in controlled settings.
Do the mycoherbicides have geographic or climatic boundaries?
o F. oxysporum f.sp. cannabis, F. oxysporum f.sp. erythroxyli, and C.
papaveracea/B. papaveris occur naturally in the native geographic
ranges of their target crops. Particular strains probably have geo-
graphic and climatic boundaries that would limit their survival and
infectivity, but the strains that were tested were evaluated under par-
ticular environmental conditions, so it is not possible to define those
boundaries.
What combination of environmental conditions (such as tempera-
ture, depth in soil, and pH) would favor persistence after application?
o Moisture (from dew, high relative humidity, or rainfall) for several
hours over several days with favorable temperatures would probably
be required for the pathogens to become established on the target
crops.
o Information on moisture, temperature, and other requirements for
pathogen survival are based on data on a few strains collected from
very small areas relative to the entire geographic and climatic ranges
of the target drug crops. The available data are not sufficient to con-
clude that the strains studied would be effective in all areas where
the illicit crops are grown.
o Studies have not examined the survival of various types of fungal
propagules (spores, mycelia, or other structures) either in general or
derived from the particular strains studied for development as my-
coherbicides.
What conditions would shorten the persistence of the proposed
mycoherbicides in the environment?
o In general, a variety of factors could shorten the persistence of fungi,
such as moisture conditions (too much or too little moisture), ex-
treme temperatures (too high or too low), ultraviolet radiation, soil
depth and compaction, and the presence of antagonistic organisms,
including other microorganisms, plants, and microfauna.
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126 Feasibility of Using Mycoherbicides for Controlling Illicit Drug Crops
Could persistence of the mycoherbicides in the environment be
controlled?
o Shortening persistence: The persistence of mycoherbicides in the
environment could be shortened by the application of chemicals,
such as fungicides or soil fumigants. Such a control strategy would
be effective for reducing mycoherbicide agents only in small areas
and would be impractical or impossible for large areas. Controlling
the mycoherbicides in this manner carries substantial risk of harm-
ing the environment or other organisms in the treated area.
o Prolonging persistence: The available data support the hypothesis
that increased inoculum levels could be maintained for a few months
after application but do not support the hypothesis that the mycoher-
bicide strains could persist indefinitely at higher population densities
than those of the indigenous strains of the same fungi.
TOXICITY TO NONTARGET ORGANISMS
Would the proposed mycoherbicides harm licit crops or kill other
soil fungi?
o Licit crops of cannabis, coca, and poppies could be adversely af-
fected if they were exposed to the proposed mycoherbicides. Such
exposures could occur through accidental drift of aerially applied
mycoherbicides; through transport by human, insect, or rodent carri-
ers; through seed transmission; or through long-distance aerial
transport of spores. The available data are insufficient to determine
whether any of the proposed mycoherbicides would harm other
types of licit crops, native plants, or soil microbiota.
Would the proposed mycoherbicides threaten biodiversity or pose
other risks to the environment?
o There are few data on risks posed by the application of the proposed
mycoherbicides. To the extent that a large volume of fungal inocu-
lum would be introduced by the application of the mycoherbicides,
the biodiversity and ecology of the target regions could be affected.
The risk of disease in related, nontarget species of Cannabis,
Erythroxylum, and Papaver and to native plants caused by the my-
coherbicide strains and the consequent indirect effects on microor-
ganisms and fauna are unknown.
What is the nature of the health risks to animals and humans posed
by the use of the proposed mycoherbicides?
o There are no reliable data on the health risks posed by F. oxysporum
f.sp. cannabis, F. oxysporum f.sp. erythroxyli, and C. papav-
eracea/B. papaveris to animals or humans. Some strains of F. ox-
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127
Answers to Charge Questions
ysporum can infect humans or produce low levels of mycotoxins
under laboratory conditions, but the variation within this species
complex and the relative rarity of human infections and known my-
cotoxin-producing strains means that other strains in the species
complex cannot serve as surrogates for the proposed mycoherbicide
strains.
o There are no data on the types or quantities of toxins or allergens
produced, if any, by the proposed mycoherbicide fungi. Without
such data, risks to human and animal health cannot be evaluated.
What would be the range of transmission of the proposed myco-
herbicides and what factors would influence their spread?
o There is potential for the spread of the mycoherbicides beyond the
site of application by a variety of means, including dispersal by
wind, rain, insects, animals, and humans and spread with seed or
soil. Therefore, neither the range nor the speed of transmission of
the proposed mycoherbicides can be predicted.
Once released, would the pathogens uncontrollable?
o Control or containment of the mycoherbicide strains after they are
released would be all but impossible. The fungal strains are living
organisms that interact with and adapt to their environment. Their
ability to survive, propagate, and disperse beyond the target area
would depend on environmental factors that can be neither predicted
nor controlled. The persistence of indigenous strains of the fungi
across the native range of their hosts is consistent with the conclu-
sion that introduced mycoherbicide strains are unlikely to be con-
tained or eradicated once they are released.
POTENTIAL FOR MUTATION AND RESULTING TOXICITY
TO TARGET AND NONTARGET ORGANISMS
What would be the potential of the pathogens to mutate?
o The potential of mycoherbicide fungi to mutate would be similar to
that of fungi in general. The genomes of fungi could change by nu-
cleotide substitution, the gain of genetic material either from closely
related species (by introgression) or from distantly related fungi (by
horizontal gene transfer), the duplication of genetic material, and the
loss of genetic material. New genetic variation could become estab-
lished in fungal populations by natural selection or by chance. Natu-
ral selection results in adaptation to changing environments, includ-
ing adaptation to new cultivars of the target crops or to new host
species (nontarget crops). Adaptation might occur in fungi that re-
produce sexually or asexually. Sexual reproduction allows new
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128 Feasibility of Using Mycoherbicides for Controlling Illicit Drug Crops
genotypes to be produced more quickly through recombination than
does mutation alone, as would occur in asexually reproducing or-
ganisms.
o One place where the mycoherbicides would be expected to adapt
would be during the fermentation necessary to produce large
amounts of inoculum. In such a case, adaptation to the fermentation
conditions might alter the virulence of the mycoherbicides. It seems
likely that the alteration would be to lower virulence rather than to
raise it. Thus, as in any industrial fermentation, quality control of
both the product at the end of fermentation and the inoculum that is
used to start the fermentation process would be essential.
o F. oxysporum reproduces clonally; no sexual stage is reported.
However, the distribution of mating type alleles and genotypic di-
versity in soil populations of F. oxysporum are consistent with sex-
ual recombination in field populations of this fungus. C. papav-
eracea reproduces clonally and by sexual recombination. B.
papaveris reproduces clonally and, although it also reproduces sexu-
ally, is self-fertile; the relative proportions of inbreeding and out-
breeding are unknown.
Are there environmental or other conditions that would drive mu-
tations in the mycoherbicides?
o Potential mutagenic changes in the proposed mycoherbicide strains
should be the same as in any fungus. There is no reason to expect
the proposed mycoherbicides to be more or less susceptible to muta-
tion due to environmental or other conditions than indigenous
strains.
Does the potential for mutation pose additional risks to nontarget
organisms (including other plants, fungi, animals, and humans) or the envi-
ronment?
o It is not possible to predict what types of mutations that might ad-
versely affect nontarget organisms would occur.
How might mutations affect the susceptibility of the target crops to
the mycoherbicides?
o Planting resistant crop plants selects for new fungal genotypes that
can overcome the host plant’s resistance. Selection for crop varieties
that are resistant to existing fungal genotypes follows, and the cycle
repeats. Selection for resistant drug crop varieties would reduce the
effectiveness of the mycoherbicides and would require continued
development of new mycoherbicide strains that could attack the
newly developed target crop genotypes.
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Answers to Charge Questions
o Mutation of the proposed mycoherbicide strains could reduce, in-
crease, or have no effect on the efficacy of the mycoherbicides on a
target crop. It is not possible to predict what type of mutations might
occur, how the pathogen or host plant might be affected, or whether
the mutations would be favored by natural selection.
How might mutations affect the toxicity of mycoherbicides generally?
o Given that F. oxysporum and C. papaveracea/B. papaveris are al-
ready present where the illicit-drug crops are grown, there is no rea-
son to expect that toxins, or mutations that affect toxins, found in the
mycoherbicides would differ from those already present in indige-
nous strains.
RESEARCH AND DEVELOPMENT NEEDS
Could the effectiveness of the mycoherbicides in eradicating illicit-
drug crops be improved through research and development?
o Only a few strains of the proposed mycoherbicides considered in
this report have been investigated. Before improvements can be con-
sidered, more research is needed to obtain and identify the most ef-
ficacious strains available (preferably from plants common to the
target regions), the most susceptible stage in the life cycle of the tar-
get crop, and the climatic and environmental conditions required for
disease development.
o Once those conditions are met, there are ways to improve the effec-
tiveness of the mycoherbicides, such as the use of adjuvants and the
development of formulation and delivery methods to facilitate and
maximize infection.
What types of expertise would be most relevant for improving the
effectiveness and safety of the proposed mycoherbicides?
o At a minimum, a team with expertise in the growing practices and
conditions under which the drug crops are grown; in plant ecology;
in plant disease epidemiology; in plant pathology; in fungal genet-
ics; in fermentation, formulation, and application technology; and in
nontarget risk assessment is needed to develop the proposed strains
as mycoherbicides, to assess their effectiveness and safety, and to
gather data for Environmental Protection Agency (EPA) registra-
tion.
o Partnerships between industry and science are needed to guide tech-
nical development and to assess the feasibility of producing the pro-
posed mycoherbicides.
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130 Feasibility of Using Mycoherbicides for Controlling Illicit Drug Crops
What types of research and technology would improve the produc-
tion and delivery of the mycoherbicides?
o Research is needed to determine whether the proposed mycoherbi-
cide strains could be produced on a large scale. Research is also
needed to guide product formulation, storability, and delivery. The
technology to support production and delivery could be identified or
developed as needed by an industrial producer.
What type of testing would be needed before mycoherbicides could
be safely and effectively used to eradicate illicit-drug crops (for example,
mode-of-action studies)?
o A full complement of research documenting the efficacy, specificity,
and mode of action of the proposed mycoherbicide products is
needed.
o For safety assessments, at a minimum, EPA’s toxicology, nontarget-
organism, and environmental-fate data requirements for microbial
pesticides should be met.
o Testing in countries where the mycoherbicides would be used
should be done, particularly to address local biodiversity and non-
target-risk considerations.
What would be required under U.S. federal and state laws to test
and approve a mycoherbicide of this type, and what guidelines of the Inter-
national Organisation for Biological and Integrated Control of Noxious
Animals and Plants1 would apply?
o The committee did not feel comfortable in developing a checklist of
testing requirements that must be met, especially with regard to state
and international rules, which differ by state and country. The ex-
perience from testing registered mycoherbicides suggests that the
requirements become part of an evolving process as dictated by the
experimental findings with regard to individual organism, host, and
conditions. The committee therefore took a more general approach
of reviewing and identifying the types of data required for registra-
tion in the United States and the special considerations for domestic
and international uses of the proposed mycoherbicides.
1
The International Organisation for Biological and Integrated Control of Noxious
Animals and Plants is a professional society that promotes the use of biological control.
While the organization and some of its members have been involved in the development
of certain international standards for testing of pesticides and guidelines for transport and
release of biological control agents, the IOBC does not have its own set of requirements.
The committee interpreted this question to be a more general one about what interna-
tional guidelines are relevant to mycoherbicides, and reviewed pertinent international
treaties and guidance from the United Nations.
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131
Answers to Charge Questions
o For a microbial pesticide to be registered by EPA for use in the
United States, a variety of tests and reports are required, for exam-
ple, on product analysis, pesticide residues, toxicology, effects on
nontarget organisms, and environmental fate of the pesticide. Such
testing is required for each strain of a mycoherbicide to be regis-
tered. An environmental assessment or an environmental impact
statement would probably be required for each mycoherbicide and
perhaps for each strain.
o At the international level, applicable guidelines and potential limita-
tions for testing, approval, and application of mycoherbicides
against illicit crops fall under the International Plant Protection
Convention, the International Standards for Phytosanitary Measures,
the Biological Weapons Convention of 1972, and legal requirements
in the country where the mycoherbicides would be used. Collec-
tively, the guidelines are intended to prevent the spread and intro-
duction of plant pests by requiring their safe export, import, ship-
ment, and release. Some of the requirements involve documentation
for the transport of agents, conducting risk assessments, and imple-
menting risk-management strategies.
