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Pesticide Resistance: Strategies and Tactics for Management. 1986. National Academy Press, Washington, D.C. Preventing and Managing Fungicide Resistance JOHAN DEKKER Following a description of the term fungicide resistance, the fac- tors that govern the buildup of a resistant pathogen population in the field are covered. Short-term tactics and long-term strategies to counteract the development of resistance are discussed. INTRODUCTION When fungicide resistance became a problem shortly after the introduction of systemic fungicides in the 1960s, the reliability of chemical disease control was at stake, especially since we did not know how to cope with the problem. Farmers who saw that a fungicide was becoming less effective often increased the dose, thus increasing the selection pressure and aggravating the problem. The failure of some of the new, originally very effective fungicides to control disease was a surprise, and it created doubt about effective disease control using these fungicides. To better understand the resistance phenom- enon, biochemical and genetic studies were conducted. These studies revealed the mechanism of action of several new fungicides and the mechanism of resistance in fungi. Greenhouse and field experiments on the ecological and population aspects of resistance have yielded considerable information about fungicide resistance. This paper will discuss (1) the possibilities of using this information to develop tactics for preventing and managing fungicide resistance, (2) the research needed for further development and improvement of these strategies, and (3) what new approaches might offer prospects for dealing with the fungicide resistance problem. 347
348 TACTICS FOR PREVENTION AND MANAGEMENT FUNGICIDE RESISTANCE People define fungicide resistance in different ways, depending on their interests and concerns. Fungicide resistance occurs when a fungal cell or a fungal population that originally was sensitive to a fungicide becomes less sensitive by hereditable changes after a period of exposure to the fungicide. A panel of FAO (Food and Agriculture Organization of the United Nations) experts has recommended that the word "resistance" should apply only to a hereditable decrease in sensitivity. The word "tolerance" should not be used in this sense, since it is ambiguous (FAO, 19791. Use of the word "insensitivity" in place of resistance is also not recommended, because it suggests a complete loss of sensitivity, which occurs only rarely. The researcher first speaks of resistance after the emergence of less-sen- sitive cells has been observed in a petri dish and the hereditable nature of this phenomenon has been proved or seems likely. Resistance in the labo- ratory, however, does not mean that resistance will develop in the field. More important is a shift toward lower fungicide sensitivity in a field pathogen population, which may be called development of resistance, even if the fungicide still provides satisfactory disease control. The farmer speaks of fungicide resistance only when disease control fails. This definition is also often used by the agrochemical industry and the extension officer, who may be afraid that reports about laboratory resistance before problems in the field have occurred may confuse or even alarm the farmer. Because of the various meanings, one must define which type of resistance is being discussed: emergence of resistant cells in laboratory experiments; reduction of fungicide sensitivity in the field, but still with adequate control; or field resistance with loss of disease control. BUILDUP OF A RESISTANT POPULATION As Georgopoulos pointed out (this volume), the main mechanisms of resistance to fungicides are a change at the site of action in the fungal cell that decreases its affinity to the fungicide or a change in uptake of the chemical so that less of it reaches the site of action. Detoxification has rarely been reported as the cause of resistance in fungi, although it is the main mechanism of resistance in insects. Resistant populations develop and increase in different ways, often from forces we can control. The mechanism of resistance may influence the fitness of resistant cells, as compared with sensitive cells, which is important to the buildup of a resistant pathogen population. The fitness of resistant strains appears to vary considerably for different types of fungicides. For some, resistance appears to be linked to a decrease in fitness in the absence of the fungicide. Thus, fungicides may be classified as low-risk, moderate-risk, or
FUNGICIDE RESISTANCE 349 high-risk compounds (Dekker, 19841. With insecticides loss of fitness in resistant insects plays a lesser role. Here the strategy has been to change to an insecticide with a different mechanism of action. This strategy is possible only so long as new insecticides become available. The discovery of fun- gicides with a lower resistance risk offers possibilities for developing strat- egies to prolong their use. The life cycle of the pathogen and the nature of the disease may influence the speed of buildup of a resistant pathogen population. For example, resis- tance builds more rapidly in an abundantly sporulating pathogen on aerial plant parts than in a slowly expanding soil pathogen. Environmental con- ditions that increase the severity of the disease may also speed the devel- opment of resistance. Another important factor in this respect is the management of pesticide application: a continuously high selection pressure by one chem- ical or by more chemicals with the same mechanism of action favors the buildup of a resistant pathogen or insect population. SHORT-TERM TACTICS New Chemicals It would be very valuable to have information on the probability of resis- tance before a new chemical is used in the field. For example, experiments on an artificial medium, with or without mutagenic agents, may tell us whether emergence of resistant cells (by mutation or otherwise) is possible. If such experiments do not yield resistant cells, their emergence in the field should not be expected, but if they do, further testing should be done on the fitness of the resistant cells on the plant, compared with that of the wild- type fungus. Such experiments may give an indication of the resistance risk of the fungicide and could be used in devising strategies to minimize the chance of resistance. Unfortunately, even when all possible laboratory and greenhouse experi- ments have been carried out, it is rarely possible to precisely predict what is going to happen in the field. Greenhouse conditions are never exactly the same as field conditions. Field experiments are indispensable, and they should be accompanied by careful monitoring (as outlined by Keiding, this volume). But even field experiments may not yield the results that can be obtained from large-scale application in practice, because the size of the area treated may play a role in the buildup of resistance. Thus, even if no resistance develops in laboratory, greenhouse, or field experiments, resistance could still appear in practice. For instance, with dodine and Kitazin-P, resistance problems arose only after many years. Nevertheless, experiments may in- dicate some of the risks involved, which is important for devising tactics to prevent or delay resistance.
350 TACTICS FOR PREVENTION AND MANAGEMENT High-Risk Chemicals High-risk compounds, such as the benzimidazole and acylalanine fungi- cides, should not be used to control risky diseases when other, less-risky chemicals provide satisfactory control. Diseases are considered risky when they allow a rapid buildup of a resistant pathogen population, for example, by a repeating infection cycle during the growing season with an abundant spore production. Other diseases may be less risky, for example, those caused by root and foot pathogens. If a high-risk fungicide is applied to control a risky disease, stringent tactics should be used to minimize the chance for development of resistance. To avoid having such fungicides and related chemicals exert a constant selection pressure, therefore, one must remember the following factors. · The amount of fungicide at risk applied to the crop should not exceed the minimum dose necessary for adequate disease control. · The mode of application should be considered, for example, soil drench may allow uninterrupted uptake of the chemical and a prolonged period of selection pressure by the fungicide. · Use of one particular fungicide or related fungicides for preharvest and postharvest application should be avoided, since the former may select for resistant cells, leading to problems in subsequent postharvest treatments. · Treatment of a large area (e.g., all fields with a particular crop in one region or country) with the same or related fungicides should be avoided. No sensitive forms will then be available to enter the crop from outside, which limits the competition between sensitive and resistant forms during intervals of low selection pressure. · A very thorough treatment of the crop, with little or no escape of sensitive cells, will eliminate competition between resistant and sensitive cells and thus favor the former. ~ The selection pressure exerted by a chemical at risk may be reduced by rotation or combined use of chemicals or by integrating chemical control with other control measures. Rotation or Combination Rotations or combinations of fungicides can reduce the risk of resistance, but only when certain guidelines are followed. To reduce selection pressure the fungicides should possess different mechanisms of action. Using two risky chemicals together is not recommended: the population of fungal cells is usually so high that the chance exists for simultaneous mutations in different genes toward resistance to both chemicals. One of the fungicides in the mixture should pose little or no risk, although the use of a mixture will not
FUNGICIDE RESISTANCE TABLE 1 Spraying Schemes and Risk for Development of Fungicide Resistance 351 Type Program Risk . . . a S - S - S -S Highest Risk b S - C - S -C Alternation c (S + C) - (S + C) - (S + C) - (S + C) Mixture d (S+C) - C - (S+C) - C Combination b and c e C - C - (S+C) - C Lowest Risk S = Fungicide at risk. C = Conventional fungicide. SOURCE: Dekker (unpublished). always stop the buildup of resistance to the compound at risk: resistance may occur in the pathogen population that is not eliminated by the nonrisky compound. Under certain conditions buildup of resistance in a mixture may occur at the same speed as when the compound is used alone (Kable and Jeffery, 1980~. Theoretically this happens when there is no escape, that is, when there are no fungal cells that are not hit by the fungicide mixture and when selection pressure is not reduced during spray intervals. Such situations are rare-e.g., with postharvest treatment of citrus fruit (Eckert, 1982) and usually do not occur in the field. In most cases the use of a mixture in the field will at least delay the buildup of resistance. If resistant strains have a reduced fitness, compared with sensitive strains, and if the interval between applications of the fungicide at risk is large enough to allow the proportion of the resistant pathogen population to drop to the preceding level, resistance problems might be avoided indefinitely. A resistant population may build up gradually in an alternation or rotation scheme. During the period that the nonrisky compound is used, the proportion of resistant strains will not increase, and may decrease if there is reduced fitness. Resistance could then be postponed indefinitely, depending on the degree of reduction in fitness and the length of the intervals between sprays of the compound at risk. The mixture may provide longer delay of resistance at higher escape; the alternations may provide more delay at lower escape (Kable and Jeffery, 19801. Both mixtures and alternations have some disadvantages. In mixtures the compound at risk is always present, which means constant selection pressure if the application is not interrupted. In an alternating scheme the use of the nonrisky compound is interrupted for no good reason. These disadvantages may be decreased by combining mixtures and rotations such that the nonrisky compound is constantly present and only the use of the risky compound is interrupted (Dekker, 19821. The chance for buildup of resistance may be delayed further by using the mixture only in critical situations (Table 11.
352 TACTICS FOR PREVENTION AND MANAGEMENT Low- or Moderate-Risk Chemicals A fungicide may be at low risk when resistant strains show a strongly reduced fitness in greenhouse experiments or when it has been used for a few years without resistance problems. Because resistance could develop in later years, fields should be monitored. Monitoring will show whether a shift toward reduced sensitivity of the pathogen population occurs during the growing season or during consecutive seasons. Depending on the degree of this shift, measures (as discussed above for the risky compounds) can be taken to prevent or delay the buildup. LONG-TERM STRATEGIES To prevent or delay the buildup of a resistant pathogen population, different chemicals that are effective against a particular disease must be available. One way of increasing the number of available chemicals is to search for new site-specific inhibitors. Before the introduction of fungicides with site- specific action, little was known about the metabolic differences between the cells of a pathogen and a host, with the exception of the wall-constituent chitin, present in most fungi but not in plants. This difference is exploited by the polyoxin antibiotics, which interfere with chitin synthesis in the fungal cell wall. Examples of sites found since then include differences in tubulins constituting the spindle in fungi and plants; differences in sterols, namely ergosterol in fungi versus lanosterol in plants; and differences in the protein synthesizing apparatus, in the respiratory chain, or in enzymes involved in RNA synthesis between plants and certain fungi. More such sites will prob- ably be discovered. Special attention should be given to site-specific inhib- itors that show a low risk to development of resistance. Further, the search should be intensified for disease-control agents that are not fungicidal in vitro, but increase the resistance of the host plant or decrease the pathogenicity of the parasite. Some of these chemicals might not or might less-readily encounter resistance. Another concept is that of developing compounds with negatively corre- lated cross-resistance: a mutational change in a pathogen that confers resis- tance to fungicide A and, at the same time, increased sensitivity to fungicide B. and vice versa. Thus, when a combination of A and B is used, B will eliminate strains resistant to A, and A will eliminate strains resistant to B. Several combinations of such compounds have been described in the liter- ature, but in most cases the phenomenon did not occur with all resistant strains. Moreover, occurrence of other resistance mechanisms, which do not result in negative cross-resistance to A and B. cannot be excluded. Never- theless the phenomenon deserves further exploration. Another phenomenon is synergism between two fungicides, A and B.
FUNGlClDE RESISTANCE 353 especially if A is more active on strains resistant to B than on wild-type strains. The effect of respiration-inhibiting fungicides on fenarimol-resistant strains of Aspergillus nidulans and Penicillium italicum illustrates such syn- ergism (de Waard and Dekker, 19831. Fungicide-resistant strains do not accumulate fenarimol, due to the presence of a constitutive energy-dependent efflux. Adding a respiration-inhibiting compound results in fenarimol ac- cumulation and fungicide sensitivity. Researchers should further explore the existence of additional combinations of compounds that might counteract resistance in a similar way. The search for integrated disease-control measures should be intensified. The use of cultivars with a certain degree of natural resistance, cultural practices, and biological control measures might be integrated with chemical control. For example, microorganisms used for biological control could be made resistant to a fungicide so that both can be used at the same time (Papavizas et al., 19821. In addition to strengthening our research efforts, we must ensure that the number of conventional, nonsystemic fungicides is not needlessly decreased by regulatory agencies. Although these chemicals cannot perform all the tasks of systemic fungicides, they remain a reliable and invaluable tool for controlling disease when resistance to a new specific compound occurs. They can also be used as companion compounds of systemic fungicides, in mix- tures, or in rotation. The need for a varied arsenal of fungicides to cope with fungicide resistance requires that barriers for introducing new chemicals are not made higher than necessary. The risks of a new fungicide to nontarget organisms and the environment should be carefully weighed against the benefits. Not using chemicals is risky, not only for the economy and world food production but also for toxicological reasons: some fungi occurring in the crop or in the harvested product may produce mycotoxins, of which some may be carcin- ogenic. A senate committee appointed by President Kennedy, reporting on pesticides and public policy after the appearance of Rachel Carson's book Silent Spring, stressed the importance of a balanced benefit-risk equation and noted that the public lacked information concerning stringent precautions taken by the government to limit possible risks of the application of pesticides (U. S. Senate, 19661. Finally, any long-term strategy must create possibilities for implementing tactics to prevent and manage fungicide resistance. We must develop an efficient system to communicate information among growers, extension of- ficers, teachers, research workers, manufacturers, salesmen, the press, reg- ulatory agencies, and the government. One example of an information distribution effort was the International Course for Southeast Asia on Fun- gicide Resistance in Crop Protection, held in Malaysia from October 17 to 24, 1984. The course was organized by the crop protection departments of
354 TACTICS FOR PREVENTION AND MANAGEMENT the agricultural universities at Wageningen, the Netherlands, and at Serdang, Malaysia, in collaboration with the Food and Agricultural Organization, the Chemical Control Committee of the International Society of Plant Pathology, and the Fungicide Resistance Action Committee of the International Group of National Associations of Agrochemical Manufacturers. CONCLUSION Although it is not yet possible to predict the development of resistance to a new fungicide with certainty, as much information as possible should be obtained about the potential of plant pathogens to become resistant to such a fungicide. This can be done in appropriate laboratory, greenhouse, and field experiments. If fungicides are applied that have been proved to be risky with respect to development of resistance, it is of prime importance to avoid a continuous and high selection pressure by such fungicides by using different fungicides in a mixture or in alternation. For flexible management it is also important to have a range of chemicals available. This can be achieved by the devel- opment of more fungicides with different mechanisms of action and by a reticent policy with respect to Reregistration of old, conventional fungicides. In order to alleviate the resistance problem in the future, attention should also be given to disease-control agents that increase the resistance of the host plant and to the phenomena of synergism and negatively correlated cross- resistance. REFERENCES Dekker, J. 1982. Counter measures for avoiding fungieide-resistanee. Pp. 177-186 in Fungieide- Resistanee in Crop Proteetion, J. Dekker and S. G. Georgopoulos, eds. Wageningen, Netherlands: Centre for Agrieultural Publishing and Documentation. Dekker, J. 1984. The development of resistance to fungicides. Prog. Pest. Bioehem. Toxieol. 4: 165- 218. de Waard, M. A., and J. Dekker. 1983. Resistance to pyrimidine fungicides which inhibit ergosterol biosynthesis. Pp. 43-49 in Human Welfare and the Environment, Vol. 3, Y. Miyamoto and P. C. Kearney, eds. Oxford: Pergamon. Eekert, J. W. 1982. Penieillium decay of citrus fruits. Pp. 231-250 in Fungieide-Resistanee in Crop Proteetion, J. Dekker and S. G. Georgopoulos, eds. Wageningen, Netherlands: Centre for Ag- rieultural Publishing and Documentation. Food and Agriculture Organization. 1979. Pest resistance to pesticides and crop loss assessment. FAG Plant Prod. and Prot. Paper, No. 6/2. Kable, P. F., and H. Jeffery. 1980. Selection for tolerance in organisms exposed to sprays of biocide mixtures: A theoretical model. Phytopathology 70:8-12. Papavizas, G. C., J. A. Lewis, and T. H. Abd-el Moity. 1982. New biotypes of Trichoderma harzianum with tolerance to benomyl and enhanced biocontrol activities. Phytopathology 72: 126- 132. U.S. Senate, Committee on Government Operations. 1966. Report on Pesticides and Public Policy. Washington, D.C.: U.S. Government Printing Office.