The Value of 
IN VITRO FUNGICIDE TESTS 
Because chemists now seem able to synthesize an 
unlimited number of compounds, we can expect that 
many fungicides developed in the future will prove 
to be better than those we are using at the present 
time. Appropriate methods must be employed to select 
the better fungicides from among the mass of candi- 
date materials. 
Separating the successful and unsuccessful fungi- 
cides is an expensive procedure. Wellman (1967) 
estimated industry’s cost of research on one new fungi- 
cide to be $3 million. Since it becomes increasingly 
difficult to find a fungicide that is more successful 
than one just introduced, these costs can be expected 
to increase. With this vast expense involved, research 
agencies must develop and use techniques and methods 
that are both efficient and reliable. 
The three traditional steps in the search for an 
effective new fungicide are laboratory evaluation, 
greenhouse evaluation, and field evaluation. With the 
proper tests and the proper interpretation of the test 
results, this still seems to be the only practical system. 
Opinions vary among investigators as to the importance 
of each of the steps. This paper will review the test 
methods used to seek and evaluate fungicides in the 
laboratory, and specifically the in vitro tests. It will be 
further limited to those methods used to develop pro- 
tective foliar fungicides. The specialized methods 
used to develop eradicative, systemic, and soil fungi- 
cides are not included. 
The value of in vitro tests to appraise fungicides 
in the laboratory is determined by the ease, speed, 
and simplicity of test manipulations and the reliability 
and usability of test results. Alternative in vivo 
methods are time consuming, which prohibits exten- 
sive use of greenhouse and field techniques in the 
initial evaluation of chemicals. In vivo tests are essen- 
tial, however, in determining how effectively chemi- 
cals control plant diseases. 
The literature concerning in vitro testing of fungi- 
cides has been reviewed by several investigators 
(Horsfall 1945a, 1945b, 1956; McCallan 1947, 1959), 
most recently by Torgeson (1967). In this paper I 
wish to emphasize the variety of in vitro techniques 
that can be used as the basis for selecting fungicides 
This paper is published by authority of the State of Illinois, IRS 
Ch. 127, Par. 58.21, and is a contribution from the Section of Botany 
and Plant Pathology of the Natural History Survey. It was earlier 
presented in England, in a symposium on methods of testing fungicides, 
at the First International Congress of Plant Pathology, London, July 
14-26, 1968. Dr. Dan Neely is a plant pathologist at the Survey. 
2 
Dan Neely 
for greenhouse and field studies. Many of these in 
vitro techniques have additional value, for in recent 
years they have been extensively used in studies seek- 
ing the toxic principles of fungicide groups and the 
mode of action of chemical structures on plant patho- 
gens. 
The significance of fungicide tests in an artificial 
rather than a natural situation is occasionally ques- 
tioned. Marsh (1936) and Howard (1939) compared 
spore germination on leaves and on glass slides and 
found comparable results. Miller (1943) compared 
the retention of copper on leaves and on pyralin plates 
and found laboratory methods to be fairly accurate. 
FUNGICIDE-FUNGUS INTERACTIONS 
In vitro tests are used to measure and rank the 
fungitoxicity of fungicides, to measure fungal spe- 
cificity, and to measure fungicide deposition, redistri- 
bution, tenacity, persistence, stability, and volatility. 
Chemicals toxic to fungi have been observed to 
affect the fungi in various ways. The fungus mycelium 
may cease growing, change metabolic processes, or be 
killed. The fungus spores may fail to swell before 
germinating, fail to germinate, plasmolyze, or be 
killed. Each of these reactions is measurable. Each 
has been used in the past to determine fungicide toxi- 
city in vitro. Standard methods of determining toxicity 
have not been universally accepted. 
Fungitoxicity 
While we accept the general definition of a fungi- 
cide as “any substance that destroys fungi or inhibits 
the growth of the spores or hyphae,” we should restrict 
the use of the adjective fungicidal to the conditions 
that kill fungi and the term fungistatic to those condi- 
tions that prevent additional growth or sporulation of 
fungi without killing them (McCallan & Wellman 
1942). The term fungitoxic has come into use in those 
situations where detrimental morphological or physio- 
logical fungal changes occur but no effort is made to 
determine whether or not the fungi are killed by the 
treatment. 
Many techniques have been developed during the 
last 40 years that attempt to measure the fungitoxicity 
of chemicals or compare the fungitoxic values of 
chemicals. These techniques are usually bioassays in- 
volving a chemical and a test fungus. The use of a 
fungus introduces biological variability and reduces 
