bei Erwachsenen Anleitung zur medikamentösen Tumorschmerztherapie (Überarbeitet von Dr. Beekhchand Permar) Pflegeinterventionen in der Schmerztherapie Herausgeber: Arbeitsgruppe Schmerztherapie im Onkologischen Zentrum Westpfalz für die Arbeitsgruppe: Prof. Dr. Ch. Madler, Anästhesiologie Prof. Dr. H. Link, Medizinische Klinik
Dans la pharmacie en ligne Viagra-représenté Paris large éventail de la dysfonction érectile anti-plus consommée. Générique Levitra (vardenafil), Cialis (tadalafil) et achat viagra pour homme, dont le prix est acceptable pour tous les budgets.1
En internet farmacia empecé a pedir porque en la farmacia de al lado nunca había deseado surtido de medicamentos priligy precio Muy cómodo en el uso de la farmacia. Estuvimos en el restaurante a. aquí la tableta con la entrega en el lugar de.
This article appeared in a journal published by Elsevier. The attached
copy is furnished to the author for internal non-commercial research
and education use, including for instruction at the authors institution
and sharing with colleagues.
Other uses, including reproduction and distribution, or selling or
licensing copies, or posting to personal, institutional or third party
websites are prohibited.
In most cases authors are permitted to post their version of the
article (e.g. in Word or Tex form) to their personal website or
institutional repository. Authors requiring further information
regarding Elsevier's archiving and manuscript policies are
encouraged to visit:
Author's personal copy
Heterogeneity of selection and the evolution of resistance REX Consortium*,y The evolution of resistance to pesticides and drugs by herbicides ; antibiotics [2,3]; and HIV-1 protease inhibi- pests and pathogens is a textbook example of adapta- tion to environmental changes and a major issue in both The evolution of resistance to pesticides and drugs has public health and agronomy. Surprisingly, there is little offered several case studies of adaptive evolution and is a consensus on how to combine selection pressures (i.e., valuable example of other evolutionary changes that are molecules used in the treatment of pests or pathogens) more difficult to perceive and analyze. Hence, studies of over space and time to delay or prevent this evolutionary the evolution of resistance to various pesticides have process. By reviewing theoretical models and experi- improved understanding of the molecular mechanisms mental studies, we show that higher levels of heteroge- involved in adaptation  and dominance , the epistatic neity of selection are associated with longer-term relations between loci , and the fitness costs of adaptive sustainability of pest or pathogen control. The combina- mutations .
tion of molecules usually outcompetes other resistance The evolution of resistance to pesticides and drugs is not management strategies, such as Responsive alternation, only a textbook example of adaptation, but is also, and Periodic application, or Mosaic, because it ensures ‘mul- above all, a major issue for both public health and agrono- tiple intragenerational killing'. A strategic deployment my, because the number of drugs and pesticides with over space and/or time of several combinations can different mechanisms of toxicity and acting on indepen- ensure ‘multiple intergenerational killing', further delay- dent targets has proved to be limited (e.g., for antibiotics ing the evolution of resistance.
see  and for pesticides see ; Box 1). Only a few new active molecules have been discovered during the past 30 The worrying issue of the evolution of resistance years. A new wave of research and development (R&D) on Throughout history, humans have used a variety of strate- gies to control diseases and their vectors, as well as pests impacting crops and domestic animals. As far back as the 8th century BC, Homer referred to the use of sulfur to Cross-resistance: a resistance to a pesticide or drug that also confers resistance fumigate homes. Arsenic, an insecticide recommended by to another pesticide or drug.
the Roman naturalist Pliny the Elder during the 1st centu- Degree of treatment heterogeneity (DTH): the probability that a set of resistance genes is confronted by more than one pesticide or drug during a ry, was used during the 10th century by the Chinese to certain amount of time, be it within or between generations.
control garden pests. From the 1940s onwards, the discovery Insecticidal toxins: toxins produced by bacteria, mostly Bacillus thuringiensis of modern pesticides, such as DDT, and most of the major and Bacillus sphaericus, and used in sprays or in genetically engineered plants to control insects.
classes of antibiotic, appeared to offer the key to controlling Multiple intragenerational killing: a strategy that uses a variety of pesticides or pests and pathogens. Most of these measures were relatively drugs on each pest or pathogen individual, to maximize the probability that each individual is killed. An individual that is resistant to molecule A but cheap and ensured high levels of control. During the two susceptible to molecule B will be killed if treated simultaneously by molecules decades that followed, these pesticides were widely used in fields, farms, homes, and hospitals to treat crops, animals, Multiple intergenerational killing: a strategy that uses a variety of pesticides or drugs on successive generations of pests or pathogens, to maximize the and humans, saving yields and lives. Unfortunately, one of probability that the offspring of resistant individuals are killed. The offspring of the drawbacks of these treatments is that they exert selec- an individual that is resistant to molecule A but susceptible to molecule B will tion pressures on their target populations, leading to the be killed by molecule B.
Recessive resistance allele: an allele that confers resistance to diploid pests evolution of resistance mechanisms (see Glossary) that and pathogens only if present in a homozygous state. A dominant resistance reduce their efficacy [e.g., insecticides (Arthropod Pesticide allele confers resistance when it occurs in either a heterozygous or homozygous state.
Refuge: areas, fields, or group of pests or pathogens remaining untreated by pesticides or drugs.
Resistance: a heritable change in a population that is reflected in the ability of * Resistance to Xenobiotics (REX) Consortium members and affiliations: Denis individuals to survive and reproduce in the presence of environmental Bourguet (INRA, UMR Centre de Biologie pour la Gestion des Populations (CBGP), conditions that once killed most individuals of the same species.
F-34988 Montferrier/Lez, France), Franc¸ois Delmotte (INRA, ISVV, UMR1065 Sante´ Resistance cost: a negative pleiotropic effect of a resistant genotype that et Agroe´cologie du Vignoble, F-33140 Villenave d'Ornon, France), Pierre Franck results in a lower fitness of resistant individuals compared with susceptible (INRA, UR1115 Plantes et Syste mes de culture Horticoles, F-84914 Avignon cedex ones in the absence of a pesticide or drug.
9, France), Thomas Guillemaud (INRA-Universite´ de Nice-Sophia Antipolis-CNRS, Resistance gene: a gene at which one or more alleles confer resistance to UMR 1355, ISA, F-06903 Sophia Antipolis cedex, France), Xavier Reboud (INRA, pesticides or drugs.
UMR1347 Agroe´cologie, F-21000 Dijon, France), Corinne Vacher (INRA, UMR1202 Resistance management strategy: a strategy devoted to delay or prevent the BIOGECO, F-33612 Cestas, France; Univ. Bordeaux, BIOGECO, UMR 1202, F-33400 evolution of resistance in a population of pests or pathogens. Mosaic, Periodic Talence, France), and Anne-Sophie Walker (INRA, UR1290 BIOGER-CPP, F-78850 application, Combination, and Responsive alternation (Box 2) are simple resistance management strategies using more than one pesticide or drug.
0169-5347/$ – see front matter ! 2012 Elsevier Ltd. All rights reserved. http://dx.doi.org/10.1016/j.tree.2012.09.001 Trends in Ecology & Evolution, February 2013, Vol. 28, No. 2 Author's personal copy
Trends in Ecology & Evolution February 2013, Vol. 28, No. 2 Box 1. The evolution of resistance to pesticides and drugs Almost 8000 cases of resistance to 300 insecticide compounds have against the prevailing resistance mechanisms (see e.g., herbicides been reported in more than 500 species of arthropods (Arthropod , insecticides , antiviral drugs , and antibiotics ). The Pesticide Resistance Database; www.pesticideresistance.com). Simi- cost of developing new drugs and pesticides has been further larly, 300 cases of field resistance to 30 fungicides have been reported increased by the tightening of requirements by regulatory authorities, in 250 species of phytopathogenic fungi (Fungicide Resistance Action necessitating a larger number of toxicological, clinical, and environ- Committee database; http://www.frac.info). The International Survey of mental trials . Hence, according to Larson , it currently takes Herbicide-Resistant Weeds (http://www.weedscience.com) has sug- approximately 10 years and up to US$1 billion to develop a new gested that there are currently approximately 390 resistant biotypes in antibiotic. Similarly, 10–12 years are required to develop and launch a 210 weed species in 690 000 fields. The situation is most critical for new pesticide onto the market .
antibiotic resistance. Genes conferring resistance to antibiotics are At the turn of the 21st century, the combination of approaches, such ubiquitous in bacteria and are highly diverse. The Antibiotic Resistance as genomics , proteomics , and metabolomics , with Genes Database (http://ardb.cbcb.umd.edu/), developed by Liu and Pop target-based high-throughput screening strategies [70,74] appeared , lists more than 23 000 potential resistance genes of approximately promising for the discovery of new drugs and pesticides with little or 400 types, conferring resistance to 250 antibiotics in 1700 species of no impact on the environment and health. However, these new bacteria from 270 genera. Strains from highly pathogenic bacteria, such methods and strategies have proved relatively unsuccessful, for both as tuberculosis bacilli, that are resistant to all known classes of antibiotics  and pesticides . The situation is different for antibiotic have recently been described .
insecticidal toxins, mostly proteins from Bacillus thuringiensis, In addition, most of the major classes of antibiotic were first whether formulated for application in sprays or produced by isolated between 1940 and 1960 . The more recently commercia- transgenic plants. The number of toxins identified is increasing  lized drugs and pesticides are often variants of previously isolated or and the populations of most of the pests targeted remain resistance- synthesized compounds and, therefore, are not particularly effective free (, but see ).
drugs and pesticides, with the exception of that relating to In the literature, four principal basic strategies combin- insecticidal toxins, would be unlikely to yield substantial ing two (or more) molecules over time and/or space have public health and crop protection options within the next been considered, to delay the evolution of resistance to 10–15 years . In the meantime, there is a need to drugs and pesticides: ‘Responsive alternation', ‘Periodic protect the existing molecules. Fortunately, most classes application', ‘Mosaic', and ‘Combination' (Box 2). Is one of pesticide and antibiotic  include several molecules particular strategy intrinsically better than the others? that are still active and for which, at least in some cases, Conversely, does the ranking of strategies depend on the there is still no sign of resistance. This raises questions target organism or the pesticide or drug being considered? about how these molecules can be combined over time and Theoretical models predicting the outcome of selection space to preserve their efficacy for as long as possible.
pressures and experimental selection on pests and patho- Box 2. Strategies for combining molecules over time and space Four principal basic strategies combining two (or more) molecules over half-dose combinations have been proposed. In the full-dose time and/or space have been considered for drugs and pesticides. These Combination strategy, each pesticide or drug is applied at the dose strategies differ in the way that the pesticides or drugs are combined. In at which it would be used if applied alone, whereas in the half-dose the Periodic application and Responsive alternation strategies, molecule strategy, the dose of each pesticide or drug is half that used when the use is uniform over space but heterogeneous over time. Periodic compound is applied alone. Consequently, the final overall dose of application involves temporal cycles of pesticide or drug application, a the full-dose strategy is equivalent to twice that applied if each strategy first suggested by Coyne . By contrast, Responsive molecule were to be used alone, whereas the final overall dose of the alternation corresponds to successive applications of molecules, but half-dose Combination strategy corresponds to the dose at which without a cycle. In this approach, a molecule is used repeatedly until the each molecule would be applied if used alone.
emergence of resistance, after which the second molecule is introduced, Practical recommendations on the strategy to be used depend on and so on. Mosaic (a strategy first suggested by Muir ) concerns a the target organism. For instance, Combination is currently spatial pattern of application for at least two molecules. Molecule recommended in the treatment of HIV , tuberculosis , and application remains uniform over time and the spatial distributions of malaria . Pesticides are also increasingly used in combination the molecules used do not overlap. Finally, Combination is the rather than as individual compounds, as exemplified by the new concomitant use of two or more molecules over time and space.
generation of Bt crops, which produce several independent toxins Responsive alternation, Periodic application, Mosaic, and Combination against the target pests . However, Combination is not have been referred to by various names within and between the different the current default strategy in antibiotic treatment, particularly in classes of pesticides and drugs, as summarized in Table I.
the community, and is not recommended for several pesticides In practice, molecules in combinations are combined in variable (e.g., for the control of Anopheles, which is the malarial parasite ratios and at different doses. Strategies based on both full-dose and vector ).
Table I. Names used to define strategies Antibiotics or antiviral drugs Insecticides or Bt toxins Sequence, sequential use, _ Sequence and threshold Cycling, antibiotic rotation, Periodic Rotation, alternation, Rotation, alternation, application, and sequential use and sequential use Mixing, 50-50 treatment, antibiotic diversity, Mosaic and multiple first-line therapy Combination Combination, antibiotic diversity, and Mixture and pyramiding Mixture and Combination Mixture, Combination, simultaneous strategy and double knockdown Author's personal copy
Trends in Ecology & Evolution February 2013, Vol. 28, No. 2 gens can be used to test such predictions. Here, we review contrast, Roush  and Lenormand and Raymond , the results obtained with theoretical models and in empir- who modeled the evolution of insecticide resistance, found ical studies for various pesticides and drugs (generally Periodic application > Mosaic.
considered separately, by ecologists and agronomists on the one hand and medical scientists on the other [12,13]).
Combination and multiple intragenerational killing at We show that some consensus can be reached on the the individual level deployment of selection pressures over time and space to Combination is effective due to multiple intragenerational delay or prevent the evolution of resistance in pest and killing , a key feature that can be explained as follows: pathogen populations if resistance alleles at each of two independent loci are present at low frequency in the pest or pathogen popula- Theoretical comparisons among strategies tion, then any given individual is extremely unlikely to We searched for articles that explicitly compared, in the carry resistance alleles at both loci. Moreover, if resistance same study, at least two of the four strategies [Responsive is recessive, then diploid pests and pathogens are only alternation, Periodic application, Mosaic, and Combination resistant if they are homozygous for the resistance allele at (whether half- or full-dose); Box 2] in terms of their efficacy both loci. When resistance alleles are at low frequencies, for delaying the evolution of resistance to more than one this probability is low. Thus, most individuals can be killed pesticide or drug. Therefore, we excluded all studies that by each one of the pesticides or drugs A and B. This is considered several molecules but modeled the evolution of described as multiple intragenerational killing, because resistance to only one molecule. A search of the Resistance most pest or pathogen individuals are susceptible to both to Xenobiotic (REX) bibliographic database [12,13] for molecules and, therefore, are ‘killed twice' (Figure 1).
articles relating to the modeling of resistance evolution The superiority of Combination over the other strategies identified 20 relevant articles. Further searches in the Web appears to be robust: in most models, this approach was of Science and Google Scholar, and screening of the articles effective for longer even if input and output parameters cited in the initial 20 articles, yielded an additional nine were varied. Its comparative advantage is particularly articles. Half of those articles were related to either insec- high when: (i) resistance to each pesticide or drug is ticide or antibiotic resistance.
initially rare [16–20]; (ii) resistance to each pesticide or Based on the 29 articles retained (Table S1 in the drug in the combination are controlled by independent loci supplementary material online), we identified a clear rank- (no cross-resistance) [16,21–23]; (iii) there is a high rate of ing of the strategies in terms of their efficacy for delaying recombination between the loci [16,19,22,23]; (iv) in resistance: Combination > Periodic application = Mosaic > diploids, homozygous susceptible individuals have a high Responsive alternation (Table 1). Combination was at least mortality [14,20]; (v) in diploids, resistance to each pesti- as good as, or outperformed Responsive alternation, Peri- cide is functionally recessive [16,22,24–26]; (vi) the pesti- odic application and Mosaic in more than 80% of the cides or drugs are of similar persistence [14,26]; and (vii) comparisons. Half-dose Combination was found to have some of the population remains untreated [19,21,23,24].
been little studied and comparisons of Combination with Even if these conditions are not completely met, Combina- other strategies were somewhat biased because a full-dose tion appears at least as good as the three other strategies.
Combination, by doubling the dose of pesticide or drug used, increases overall selection pressure. Responsive al- Degree of treatment heterogeneity and multiple killing ternation was less effective than Periodic application and The most recent approaches in medicine focus on antibiotic Mosaic in all comparisons. The ranking of Mosaic and heterogeneity [27,28], the idea being that higher degrees of Periodic application was, by contrast, not straightforward.
treatment heterogeneity (DTH) are associated with slower These two strategies were compared mostly to determine evolution of resistance. Mani  explored this idea for whether Periodic application (referred to as ‘cycling' in insecticide resistance more than 20 years ago. He showed clinical studies; Box 2) could delay resistance to antibiotics that, after Combination, the most promising strategy was more effectively than could Mosaic (referred to as ‘mixing' not to vary applications of a given molecule over time in clinical studies; Box 2) in hospitals or, more specifically, (Periodic application) or space (Mosaic), but to alternate in intensive care units. All the epidemiological models gave the insecticides used over both time and space, thereby the same answer: Mosaic > Periodic application. By maximizing the DTH.
Table 1. Side-by-side comparisons of the four strategies in terms of their relative efficacies for delaying or preventing resistance Theoretical studies Empirical studies Responsive alternation Periodic application Periodic application Responsive alternation Periodic application Responsive alternation an, number of comparisons in all theoretical and empirical studies.
bThe ranking of the strategies depends on the setting for one or several input or output parameters.
Author's personal copy
Trends in Ecology & Evolution February 2013, Vol. 28, No. 2 Dispersal < Mosaic scale Dispersal ≥ Mosaic scale Mul!ple intergenera!onal killing at the family or colony level Genera!on !me < Applica!on period Genera! on !me ≥ Applica!on period Mul!ple intergenera!onal killing at the family or colony level Mul!ple intragenera!onal killing TRENDS in Ecology & Evolution Figure 1. Schematic representation of the effect of the different strategies (Responsive alternation, Mosaic, Periodic application, and Combination) on the targeted pests or pathogens, here, a mosquito. These strategies can lead to multiple intragenerational killing at the individual level (for Combination) or multiple intergenerational killing at the family or colony level (for Periodic application and Mosaic). This depends on the balance between the spatial and temporal scales of the treatments and the dispersal capacities and generation time of the targeted pests or pathogens. At each generation (G), pests or pathogens are selected by molecule 1 (in blue patches), molecule 2 (in red patches), or a combination of these two molecules (in blue-and-red patches). Individuals S (black mosquitos), R mosquitos), and R susceptible, resistant to molecule 1, and resistant to molecule 2, respectively. Individuals R mosquitos), harboring genes conferring resistance to molecule 1 12 (blue-and-red as well as genes conferring resistance to molecule 2, can survive in a patch treated with a combination of these two molecules. The degree of treatment heterogeneity (DTH), defined here as the probability that a set of resistance genes is confronted by more than one pesticide or drug, varies among the strategies. Combination displays the largest DTH, followed by Periodic applications and Mosaic, depending on the generation time, dispersal distance, period, and spatial scales of application, and by To our knowledge, the relation between DTH, temporal Responsive alternation systematically results in the lowest or spatial selection heterogeneity, and the sustainability of DTH, because the offspring are treated with the same efficacy for a given molecule has never been clearly for- molecules as their parents until the population size (or malized. We suggest that DTH should be defined as the disease severity or yield loss) reaches unacceptable levels.
probability that a set of resistance genes is confronted by Depending on the pattern of pest or pathogen dispersal, its more than one pesticide or drug within or between gen- generation time, and the temporal and spatial scales of erations. In case of Periodic application, offspring from treatment, higher DTH can be achieved with either Peri- individuals resistant to one molecule will be treated with odic application or Mosaic strategies, or through the use of another molecule depending on the generation time of the a combination of these two extreme strategies.
pathogen or pest and on the period of application of the drug or pesticide. These offspring would be expected to be Empirical comparisons between strategies susceptible to the second molecule, particularly in the In 1983, Georghiou  stated that: ‘Perusal of pertinent absence of cross-resistance and if resistance genes are literature reveals that there are more papers discussing independent of each other. In this case, DTH therefore the value of mixture [i.e., Combination] (as well as rotation ensures multiple intergenerational killing at the colony or [i.e., Periodic application]) than those that report actual family level (Figure 1), because the first molecule kills most research on the subject'. Unfortunately, this remains true individuals in the parental generation and the second in 2012. Using the Web of Science, Google Scholar, and the molecule then kills the offspring of the few survivors. As references cited in recent articles on this topic, we found explained above, Combination ensures multiple intragen- only 17 empirical studies (half of them being on insecticide erational killing and a maximal DTH because every indi- resistance) comparing at least two strategies under labo- vidual suffers both molecules simultaneously. In a Mosaic ratory, greenhouse, care units, or field conditions (Table S2 set up, the survivors to the first molecule can disperse and in the supplementary material online).
then be killed by the second molecule. If dispersal dis- In the 17 empirical studies identified, the ranking of tances are larger than the scale of Mosaic unit, then Mosaic efficacy was: Combination = Periodic application > Respon- can also lead to multiple intergenerational killing.
sive alternation (Table 1). Indeed, in five out of eight All things being equal, higher DTH should be associated comparisons, Combination was found to be as good as with longer-term sustainability of pesticides or drugs.
Periodic application, and Responsive alternation never Author's personal copy
Trends in Ecology & Evolution February 2013, Vol. 28, No. 2 outperformed either of these two strategies. It was not the success of Combination. However, nine of the 17 em- possible to rank Mosaic reliably, because too few compar- pirical studies were conducted without such refuges (Table isons included this strategy. Mosaic outperformed Periodic S2 in the supplementary material online). This is unfortu- application and Responsive alternation in two independent nate, because such refuges could easily be included in studies, but was found to be less effective than Combina- studies of the selection of resistance. Leaving a fraction tion, Periodic application, and Responsive alternation in of the population free of pesticide exposure would have the other four comparisons (Table 1).
better mimicked the conditions in fields, hospitals, and Although Combination appeared to be the best strategy care units. Indeed, a significant proportion of the pests or in theoretical models, it did not clearly outperform Periodic pathogens often remain untreated unintentionally. Dor- application in empirical studies. This discrepancy between mant weeds, resting spores of fungi, hidden mosquito theoretical and empirical results can simply reflect time breeding sites, soil seed banks or field borders, alternative constraints. Indeed, in most experimental studies, treat- hosts, or humans outside the medical system are common ments were applied during a fixed number of generations.
and constitute unplanned refuges of pests and pathogens.
In most cases, resistance emerged when molecules were used singly, but not when they were combined over space Can Combination be outcompeted? and/or time (Periodic application, Mosaic, or Combination).
One particular condition can render Combination inferior Thus, several studies reported an absence of resistance to other strategies. This condition is the occurrence of development for at least one molecule for both Combina- fitness costs, resulting in resistant individuals being less tion and Periodic application strategies (e.g., [31–35]), fit than susceptible individuals in the absence of the pesti- making it impossible to draw any firm conclusions con- cide or drug. Such costs might lead to the counterselection cerning possible differences in efficacy between these two of resistance alleles and, therefore, would delay, if not strategies. The conclusion that these two strategies are prevent, the development of resistance. The expression similar in efficacy is thus valid for the number of genera- of this cost would require spatial or temporal variation tions over which selection took place, but might not hold in pesticide or drug selection, with locations or periods of absolutely true per se.
time in which one of the pesticides or drugs is absent.
The discrepancy between theoretical and empirical Combination is the only strategy combining two molecules results can also result from the use of experimental set- that does not generate such variation and, therefore, it is tings that decrease the advantage of Combination over the only strategy that does not allow the expression of a other strategies. As mentioned above, empirical studies resistance cost. Consequently, fitness costs can facilitate are limited by the number of generations that can be run.
the mitigation of resistance in all strategies except Combi- They are also limited by the number of individuals per nation. This can explain why Dobson et al.  (theoreti- generation that can be manipulated. These experimental cally) and Immajaru et al.  (experimentally) found constraints have two important consequences. First, em- Combination to be less effective than Periodic application pirical studies focus on the evolution of resistance alleles and Mosaic (Tables S1 and S2 in the supplementary ma- already present in populations rather than on resistance terial online). Indeed, their theoretical and biological mod- alleles acquired de novo by mutation or horizontal transfer.
els were characterized by high fitness costs and an absence Hence, in all but one of the experimental studies (Table S2 in the supplementary material online), a deliberate deci- In practice, fitness costs might only rarely make Com- sion was taken to have a high frequency (i.e., >10!3) of bination worse than other strategies. First, the multiple resistance to at least one pesticide or drug at the start of selection, thereby decreasing the efficacy of Combination approaches might be sufficient to ensure the superiority by violating one of the favoring conditions [16–20]. Second, of this strategy in many cases, even in the presence of a sufficiently large number of individuals must survive fitness costs. Second, although mutations conferring resis- pesticide or drug treatments to establish the next genera- tance are often costly (see, e.g., herbicides , insecticidal tion. Consequently, the selection pressure applied in such proteins , antibiotics , and antivirus ), experiments generally varies between 0.5 and 0.8, corre- decreases in fitness can be attenuated or even completely sponding to low doses. In such cases, resistance can be abolished by compensatory mutations (see, e.g., herbicides functionally dominant, further decreasing the comparative , antimicrobial drugs , antibiotics , and antivi- advantage of Combination over the other strategies rus ) or through interactions with other resistance [16,22,24–26]. Experimental settings with high initial fre- mutations . Over time, costly resistance mutations quencies of resistance alleles and low selection pressures can also be replaced by resistance mutations associated can, in some cases, approach real conditions. Molecules with lower fitness costs . Finally, when part of the newly released onto the market are sometimes used in population remains untreated, fitness costs counteract combination with other molecules for which resistance has the selection of resistance alleles, even for Combination.
already been selected in the targeted pest or pathogen Untreated individuals can be actively preserved by the use populations, for economic purposes. Selection at low doses or maintenance of refuges for pests and pathogens. The use can also occur in field conditions because of the dilution of of refuges is not possible in hospitals, because it would be the molecules and their degradation over time.
unethical not to treat infected humans with antibiotics or Nevertheless, one specific feature of empirical studies other drugs. However, the community outside hospitals clearly differs from practice. As pointed out above, the constitutes a refuge for most pathogen populations and presence of untreated individuals from refuges increases individuals carrying pathogenic bacteria or viruses but Author's personal copy
Trends in Ecology & Evolution February 2013, Vol. 28, No. 2 displaying no symptoms, or only minor symptoms, are left second antibiotic in a Mosaic implemented at the scale of untreated. Finally, pathogens or pests generally escape the bed or at the scale of the care unit than in a Periodic treatments even within the host or the field, because application based on the cycling of antibiotics over several treatment coverage is rarely complete.
months. Another hypothesis has been put forward by Boni et al.  to explain the higher performance of Mosaic over Increasing the DTH of Combination Periodic application: Periodic application degrades the The number of molecules that can actually be used in a mean fitness of the parasite population more quickly than Combination is limited by resistances that have already does Mosaic, making it easier for new resistant types to developed (Box 1) and by several challenges outlined in Box invade and spread in the population.
3. Generally, the concomitant use of a large number of Although difficult to implement, we suggest that Peri- molecules entails higher costs, which can outweigh the odic application at the level of the patient, rather than the benefits of delaying or preventing resistance in the eyes of hospital (or care unit), might result in greater DTH than the stakeholders. Thus, combinations containing all the using a Mosaic approach. Alternating antibiotics to treat available molecules are unlikely to be used. However, it patients would increase the likelihood of multiple inter- might be possible to use several different combinations to generational killing (i.e., the probability of colonies resis- treat a given pest or pathogen. These combinations would tant to a given antibiotic being treated and, therefore, ideally be used to ensure the highest DTH, yielding multi- killed by another antibiotic in the next generation).
ple intragenerational killing (at the individual level) and multiple intergenerational killing (at the colony or family Beyond Combination and DTH: protecting populations level). Depending on the distances over which dispersal against the emergence of resistance alleles occurs and on generation time, the highest DTH can be The question of how to combine pesticides and drugs over provided by a complex temporal and spatial arrangement time and space is only one part of the overall debate on of the various combinations.
resistance management. The dose of the molecules used This might have practical consequences. For example, must also be considered. Resistance management strate- in antibiotic resistance management, treatment heteroge- gies sometimes include the use of high doses of pesticides neity is currently defined at the level of the hospital rather and drugs. For bacterial and HIV infections, this has been than the pathogen. The theoretical and empirical studies referred to as the ‘Hit hard and early' approach .
reviewed here show that diversity in antibiotic use be- Interestingly, different rationales are applied to pesticides tween care units or beds at a given time (i.e., a Mosaic and drugs. For drugs, the reason for treating ‘hard' is to strategy) is more sustainable than is cycling different decrease the size of the pathogen population as much as antibiotic regimens over time (i.e., a Periodic application possible, to prevent the appearance of resistance alleles.
strategy). This is because, at the scale relevant to bacterial For pesticides, high-dose strategies are designed to avoid populations, Mosaic imposes greater DTH than does Peri- not only the emergence of new resistance alleles [48,49], odic application [28,29,48,49]. This is particularly true but also the building of polygenic resistance , and to when the cycle of each antibiotic regimen is long, extending make resistance of diploid pests functionally recessive .
over several months. Indeed, due to its short generation The use of a high dose can also enlarge the spectrum of time, a bacterial colony is more likely to encounter the pests targeted. This is particularly true for herbicides, Box 3. Challenges with combinations Imagine that two molecules are available and that all conditions are limiting the adverse effects of treatment. Adverse effects occur when satisfied for their combination to outperform all other strategies for single molecules are used, but they are probably worsened by the use delaying the evolution of resistance. Would Combination become the of combinations, because synergy between molecules  can optimal strategy for use with any given set of pesticides and drugs? increase the threat to the environment  and human health .
This is unlikely because there are several obstacles to the universal Stakeholders (i.e., companies, users, prescribers, and public recommendation and implementation of this strategy.
authorities) diverge on their respective interests, goals, and their The possibility of antagonistic effects between molecules (which sensitivity to the costs of the strategy, depending on the policy may seriously reduce pest or pathogen control) constitutes a first implemented. For example, refuges increase the risk of pest and/or obstacle to the use of the Combination strategy . Synergistic pathogen damage and, in the short term, this cost is met directly by molecule combinations can be advantageous in controlling pests and users. Similarly, Combination, by multiple intragenerational killing, pathogens. However, resistance to such combinations can evolve can be more efficient for controlling pests and/or pathogens, but faster than can resistance to antagonistic molecule combinations because of the higher dose applied, implies financial costs to farmers and, in some cases, to individual molecules themselves .
or patients (or public authorities, if there is social health coverage), as A second obstacle for using Combination is that the molecules well as an increased magnitude of adverse effects on health and the prescribed by physicians and used by farmers not only control pests environment and, thus, the costs to be covered by public authorities.
and pathogens, but may also injury crops and have adverse effects on The willingness of the various stakeholders to share the costs non-target organisms and human health. The WHO has reported that depends directly on the extent to which they are likely to be affected there are approximately three million human cases of pesticide by or considered responsible for the emergence of resistance. Hence, poisoning annually, resulting in 220 000 deaths worldwide , as users are confronted with the so-called ‘Tragedy of the Commons' well as hepatoxicity, neurotoxicity, and lipodystrophy [89–91].
when exploiting a common property resource , even if they are Chemical pesticides have a significant impact on non-target plants, likely to be strongly affected by the evolution of resistance. In most fungi, and arthropods . Pesticide use can disrupt biological cases, by not playing their part in the management of resistance, each control through direct toxicity , indirectly changing the commu- user maximizes their own short-term benefit but favors the selection nity structure  and natural predators or parasitoids . A trade- of resistant pests and/or pathogens, thus having potentially long- off thus exists between controlling the pest with the right dose and term negative effects for the community.
Author's personal copy
Trends in Ecology & Evolution February 2013, Vol. 28, No. 2 because fields generally contain more than one weed spe- 4 Doyon, L. et al. (2009) Resistance to HIV-1 protease inhibitors. In cies that must be controlled.
Antimicrobial Drug Resistance (Mayers, D., ed.), pp. 477–492, Springer 5 Ranson, H. et al. (2002) Evolution of supergene families associated with The drawback of hitting hard is that it increases the insecticide resistance. Science 298, 179–181 costs associated with resistance management (Box 3). This 6 Bourguet, D. (1999) The evolution of dominance. Heredity 83, 1–4 strategy can be counterproductive if resistant pathogens 7 Raymond, M. et al. (1989) Amplification of various esterase B's are already established [49,54]. However, in the case of responsible for organo-phosphate resistance in Culex mosquitoes.
new molecules for which no resistance has been detected, Biochem. Genet. 27, 417–423 8 Coustau, C. et al. (2000) Resistance to xenobiotics and parasites: can we this approach can be the most appropriate, provided that count the cost? Trends Ecol. Evol. 15, 378–383 the costs are sustainable. Unfortunately, most mathemat- 9 Davies, J. (2007) Microbes have the last word. A drastic re-evaluation of ical models of the evolution of pesticide resistance assume antimicrobial treatment is needed to overcome the threat of antibiotic- that population size is infinite, but see, for example, resistant bacteria. EMBO Rep. 8, 616–621 [52,55]. Consequently, resistance alleles are generally as- 10 Woods, D.J. and Williams, T.M. (2007) The challenges of developing novel antiparasitic drugs. Invert. Neurosci. 7, 245–250 sumed to be initially present at all resistance loci in the 11 Theuretzbacher, U. (2009) Future antibiotics scenarios: is the tide population. In fact, natural populations are limited in size starting to turn? Int. J. Antimicrob. Agents 34, 15–20 and might contain no resistance alleles. In such cases, the 12 REX Consortium (2007) Structure of the scientific community appearance and early increase in the frequency of resis- modelling the evolution of resistance. PLoS ONE 2, e1275 tance alleles is a stochastic process that is dependent on 13 REX Consortium (2010) The skill and style to model the evolution of resistance to pesticides and drugs. Evol. Appl. 3, 375–390 the balance between mutation rates and population size.
14 Roush, R.T. (1989) Designing resistance management programs: how This stochasticity is also largely ignored in empirical stud- can you choose? Pestic. Sci. 26, 423–441 ies. As indicated above, empirical studies are always per- 15 Lenormand, T. and Raymond, M. (1998) Resistance management: the formed at locations or using strains in which resistance to stable zone strategy. Proc. R. Soc. Lond. B: Biol. Sci. 265, 1985–1990 16 Comins, H.N. (1986) Tactics for resistance management using multiple at least one molecule occurs at a relatively high frequency, pesticides. Agric. Ecosyst. Environ. 16, 129–148 which constitute an unfavorable situation for high-dose 17 Barnes, E. et al. (1995) Worm control and anthelmintic resistance: adventures with a model. Parasitol. Today 11, 56–63 Therefore, there is a need for both theoretical and 18 Bonhoeffer, S. et al. (1997) Evaluating treatment protocols to prevent empirical studies to further investigate the evolution of antibiotic resistance. Proc. Natl. Acad. Sci. U.S.A 94, 12106–12111 19 Curtis, C.F. and Otoo, L.N. (1986) A simple model of the build-up of resistance in conditions allowing stochastic events . In resistance to mixtures of anti-malarial drugs. Trans. R. Soc. Trop. Med.
such situations, ‘hitting hard' probably results in a greater Hyg. 80, 889–892 efficacy of Combination than of other strategies and full- 20 Roush, R.T. (1998) Two-toxin strategies for management of insecticidal dose Combination certainly provides populations with the transgenic crops: can pyramiding succeed where pesticide mixtures highest level of protection against the emergence of resis- have not? Philos. Trans. R. Soci. Lond. B: Biol. Sci. 353, 1777–1786 21 Caprio, M.A. (1998) Evaluating resistance management strategies for tance alleles. The use of high-dose Bt crops could be seen as multiple toxins in the presence of external refuges. J. Econ. Entomol.
a life-size experiment testing this hypothesis. Interesting- ly, in the USA, populations of the pink bollworm, Pectino- 22 Diggle, A.J. et al. (2003) Herbicides used in combination can reduce the phora gossypiella, targeted by Bt cotton, and of the probability of herbicide resistance in finite weed populations. Weed Res.
European corn borer, Ostrinia nubilalis, targeted by Bt 23 Mani, G.S. (1985) Evolution of resistance in the presence of 2 maize, have been declining from year to year [57,58]. For insecticides. Genetics 109, 761–783 the pink bollworm, sterile moth releases have been suc- 24 Argentine, J. et al. (1994) Computer simulation of insecticide cessful in suppressing the emergence of resistance alleles resistance management strategies for control of Colorado potato to Bt cotton , a cornerstone for the multi-tactic eradi- beetle (Coleoptera: Chrysomelidae). J. Agric. Entomol. 11, 137–155 25 Curtis, C.F. (1985) Theoretical models of the use of insecticide mixtures cation program of this pest . This provides some hope for the management of resistance. Bull. Entomol. Res. 75, 259–265 that pest populations could be eliminated over a wide area 26 Curtis, C.F. et al. (1993) Are there effective resistance management before resistance alleles emerge and spread.
strategies for vectors of human disease? Biol. J. Linn. Soc. 48, 3–18 27 Bal, A. et al. (2010) Antibiotic heterogeneity: from concept to practice.
Ann. N. Y. Acad. Sci. 1213, 81–91 We would like to thank the De´partement Sante´ des Plantes et 28 Masterton, R.G. (2010) Antibiotic heterogeneity. Int. J. Antimicrob.
Environnement (SPE) of INRA for financial support. This paper is Agents 36, S15–S18 dedicated to the memory of Jean-Batiste Berge´, who had a leading 29 Mani, G.S. (1989) Evolution of resistance with sequential application of influence on the study of xenobiotic resistance in our institute.
insecticides in time and space. Proc. R. Soc. Lond. B: Biol. Sci. 238, 30 Georghiou, G. (1983) Management of resistance in arthropods. In Pest Appendix A. Supplementary data Resistance to Pesticides (Georghiou, G. and Saito, T., eds), pp. 769–792, Supplementary data associated with this article can be found, in the online version, at http://dx.doi.org/10.1016/ 31 Burden, G.S. et al. (1960) Development of chlordane and malathion resistance in the German cockroach. J. Econ. Entomol. 53, 1138–1139 32 Castle, S.J. et al. (2002) Field evaluation of different insecticide use strategies as resistance management and control tactics for Bemisia tabaci (Hemiptera: Aleyrodidae). Bull. Entomol. Res. 92, 449–460 1 Powles, S.B. and Yu, Q. (2010) Evolution in action: plants resistant to 33 McKenzie, C.L. and Byford, R.L. (1993) Continuous, alternating, and herbicides. Annu. Rev. Plant Biol. 61, 317–347 mixed insecticides affect development of resistance in the horn fly 2 Bush, K. et al. (2011) Tackling antibiotic resistance. Nat. Rev.
(Diptera, Muscidae). J. Econ. Entomol. 86, 1040–1048 Microbiol. 9, 894–896 34 Pimentel, D. and Burgess, M. (1985) Effects of single versus 3 Davies, J. and Davies, D. (2010) Origins and evolution of antibiotic combinations of insecticides on the development of resistance.
resistance. Microbiol. Mol. Biol. Rev. 74, 417–433 Environ. Entomol. 14, 582–589 Author's personal copy
Trends in Ecology & Evolution February 2013, Vol. 28, No. 2 35 Prabhaker, N. et al. (1998) Evaluation of insecticide rotations and 63 Walsh, C. and Wright, G. (2005) Introduction: antibiotic resistance.
mixtures as resistance management strategies for Bemisia argentifolii Chem. Rev. 105, 391–393 (Homoptera: Aleyrodidae). J. Econ. Entomol. 91, 820–826 64 Erickson, J.M. et al. (1985) Herbicide resistance and cross-resistance: 36 Dobson, R.J. et al. (1987) A genetic model describing the evolution of changes at three distinct sites in the herbicide-binding protein. Science levamisole resistance in Trichostrongylus colubriformis, a nematode parasite of sheep. IMA J. Math. Appl. Med. Biol. 4, 279–293 65 Tabashnik, B.E. et al. (2009) Field-evolved insect resistance to Bt crops: 37 Immaraju, J.A. et al. (1990) Field-evaluation of insecticide rotation and definition, theory, and data. J. Econ. Entomol. 102, 2011–2025 mixtures as strategies for citrus thrips (Thysanoptera, Thripidae) 66 Hertogs, K. et al. (2000) Phenotypic and genotypic analysis of clinical resistance management in California. J. Econ. Entomol. 83, 306–314 HIV-1 isolates reveals extensive protease inhibitor cross-resistance: a 38 Vila-Aiub, M.M. et al. (2009) Fitness costs associated with evolved survey of over 6000 samples. AIDS 14, 1203–1210 herbicide resistance alleles in plants. New Phytol. 184, 751–767 67 Sanders, C.C. et al. (1984) Selection of multiple antibiotic-resistance by 39 Gassmann, A.J. et al. (2008) Fitness costs of insect resistance to quinolones, beta-lactams, and aminoglycosides with special reference Bacillus thuringiensis. Annu. Rev. Entomol. 54, 147–163 to cross-resistance between unrelated drug classes. Antimicrob. Agents 40 Anderson, R.M. and May, R.M. (1991) Infectious Diseases of Humans: Chemother. 26, 797–801 Dynamics and Control, Oxford University Press 68 Li, J.W.H. and Vederas, J.C. (2009) Drug discovery and natural 41 Nijhuis, M. et al. (2001) Implications of antiretroviral resistance on products: end of an era or an endless frontier? Science 325, 161–165 viral fitness. Curr. Opin. Infect. Dis. 14, 23–28 69 Larson, E. (2007) Community factors in the development of antibiotic 42 Paris, M. et al. (2008) The effects of the genetic background on herbicide resistance. Annu. Rev. Public Health 435–447 resistance fitness cost and its associated dominance in Arabidopsis 70 Lein, W. et al. (2004) Target-based discovery of novel herbicides. Curr.
thaliana. Heredity 101, 499–506 Opin. Plant Biol. 7, 219–225 43 Maisnier-Patin, S. and Andersson, D.I. (2004) Adaptation to the 71 Dougherty, T.J. et al. (2002) Microbial genomics and novel antibiotic deleterious effects of antimicrobial drug resistance mutations by discovery: new technology to search for new drugs. Curr. Pharm. Des. 8, compensatory evolution. Res. Microbiol. 155, 360–369 44 Andersson, D. and Hughes, D. (2010) Antibiotic resistance and its cost: 72 Freiberg, C. et al. (2004) The impact of transcriptome and proteome is it possible to reverse resistance? Nat. Rev. Microbiol. 8, 260–271 analyses on antibiotic drug discovery. Curr. Opin. Microbiol. 7, 451– 45 Menendez-Arias, L. et al. (2003) Fitness variations and their impact on the evolution of antiretroviral drug resistance. Infect. Dis. 3, 73 Aliferis, K. and Chrysayi-Tokousbalides, M. (2011) Metabolomics in pesticide research and development: review and future perspectives.
46 Cong, M. et al. (2007) The fitness cost of mutations associated with Metabolomics 7, 35–53 human immunodeficiency virus type 1 drug resistance is modulated by 74 Geary, T.G. et al. (2004) The changing landscape of antiparasitic drug mutational interactions. J. Virol. 81, 3037–3041 discovery for veterinary medicine. Trends Parasitol. 20, 449–455 47 Guillemaud, T. et al. (1998) Evolution of resistance in Culex pipiens: 75 Bro¨tz-Oesterhelt, H. and Sass, P. (2010) Post-genomic strategies in allele replacement and changing environment. Evolution 52, 443–453 antibacterial drug discovery. Future Microbiol. 5, 1553–1579 48 Gressel, J. (2011) Global advances in weed management. J. Agric. Sci.
76 Woods, D.J. and Knauer, C.S. (2010) Discovery of veterinary antiparasitic agents in the 21st Century: a view from industry. Int.
49 van den Bosch, F. et al. (2011) The dose rate debate: does the risk of J. Parasitol. 40, 1177–1181 fungicide resistance increase or decrease with dose? Plant Pathol. 60, 77 Bravo, A. et al. (2011) Bacillus thuringiensis: a story of a successful bioinsecticide. Insect Biochem. Mol. Biol. 41, 423–431 50 Boni, M. et al. (2008) Benefits of using multiple first-line therapies 78 Gassmann, A.J. et al. (2011) Field-evolved resistance to Bt maize by against malaria. Proc. Natl. Acad. Sci. U. S. A. 105, 14216–14221 western corn rootworm. PLoS ONE 6, e22629 51 Ho, D.D. (1995) Time to hit HIV, early and hard. N. Engl. J. Med. 333, 79 Coyne, F.P. (1951) Proper use of insecticides. Br. Med. J. 2, 911–912 80 Muir, D.A. (1977) Genetic Aspects of Developing Resistance of Malaria 52 Renton, M. et al. (2011) Does cutting herbicide rates threaten the Vectors. 2. Gene Flow and Control Pattern, WHO sustainability of weed management in cropping systems? J. Theor.
81 WHO (2010) Antiretroviral Therapy for HIV Infection in Adults and Biol. 283, 14–27 Adolescents: Recommendations for a Public Health Approach – 2010 53 Alstad, D.N. and Andow, D.A. (1995) Managing the evolution of insect resistance to transgenic plants. Science 268, 1894–1896 82 WHO (2009) Treatment of Tuberculosis: Guidelines, WHO 54 Read, A.F. et al. (2011) The evolution of drug resistance and the curious 83 WHO (2006) Guidelines for the Treatment of Malaria, WHO orthodoxy of aggressive chemotherapy. Proc. Natl. Acad. Sci. U. S. A.
84 Cui, J. et al. (2011) Effect of pyramiding Bt and CpTI genes on 108, 10871–10877 55 Sisterson, M.S. et al. (2004) Effects of insect population size on Noctuidae) under laboratory and field conditions. J. Econ. Entomol.
evolution of resistance to transgenic crops. J. Econ. Entomol. 97, 85 Enayati, A. and Hemingway, J. (2010) Malaria management: past, 56 Renton, M. (2012) Shifting focus from the population to the individual present, and future. Annu. Rev. Entomol. 55, 569–591 as a way forward in understanding, predicting and managing the 86 Green, J.M. (1989) Herbicide antagonism at the whole plant level.
complexities of evolution of resistance to pesticides. Pest Manag.
Weed Technol. 3, 217–226 87 Hegreness, M. et al. (2008) Accelerated evolution of resistance in 57 Carrie re, Y. et al. (2003) Long-term regional suppression of pink multidrug environments. Proc. Natl. Acad. Sci. U. S. A. 105, 13977– bollworm by Bacillus thuringiensis cotton. Proc. Natl. Acad. Sci.
U. S. A. 100, 1519–1523 88 WHO (1992) Our Planet, Our Health, WHO 58 Hutchison, W.D. et al. (2010) Areawide suppression of European corn 89 Caron-Debarle, M. et al. (2010) HIV-associated lipodystrophy: from fat borer with Bt maize reaps savings to non-Bt maize growers. Science injury to premature aging. Trends Mol. Med. 16, 218–229 90 Polson, J.E. (2007) Hepatotoxicity due to antibiotics. Clin. Liver Dis.
59 Tabashnik, B.E. et al. (2010) Suppressing resistance to Bt cotton with sterile insect releases. Nat. Biotechnol. 28, 1304–1307 91 Toovey, S. (2009) Mefloquine neurotoxicity: a literature review. Travel 60 Grefenstette, B. et al. (2009) Pink Bollworm Eradication Plan in the Med. Infect. Dis. 7, 2–6 92 McLaughlin, A. and Mineau, P. (1995) The impact of agricultural 61 Liu, B. and Pop, M. (2009) ARDB: Antibiotic Resistance Genes practices on biodiversity. Agric. Ecosyst. Environ. 55, 201–212 Database. Nucleic Acids Res. 37, D443–D447 93 Hossain, M.B. and Poehling, H.M. (2006) Non-target effects of three 62 Velayati, A.A. et al. (2009) Emergence of new forms of totally drug- biorationale insecticides on two endolarval parasitoids of Liriomyza resistant tuberculosis bacilli, super extensively drug-resistant sativae (Dipt., Agromyzidae). J. Appl. Entomol. 130, 360–367 tuberculosis or totally drug-resistant strains in Iran. Chest 136, 94 Hanazato, T. (1998) Response of a zooplankton community to insecticide application in experimental ponds: a review and the Author's personal copy
Trends in Ecology & Evolution February 2013, Vol. 28, No. 2 implications of the effects of chemicals on the structure and functioning 97 Peng, Y. et al. (2010) Dimethoate, fenvalerate and their mixture affects of freshwater communities. Environ. Pollut. 101, 361–373 Hylyphantes graminicola (Araneae: Linyphiidae) adults and their 95 Hardin, M.R. et al. (1995) Arthropod pest resurgence: an overview of unexposed offspring. Agric. For. Entomol. 12, 343–351 potential mechanisms. Crop Prot. 14, 3–18 98 Uchino, S. et al. (2005) Acute renal failure in critically ill patients: 96 Cedergreen, N. et al. (2008) A review of independent action compared to a multinational, multicenter study. J. Am. Med. Assoc. 294, concentration addition as reference models for mixtures of compounds with different molecular target sites. Environ. Toxicol. Chem. 27, 99 Hardin, G. (1968) The tragedy of the commons. Science 162, 1243–1248
HARNESSING INNATEAND ADAPTIVE IMMUNITY IN In 1975 Kohler and Milstein solved a technical problem that scientists had been battling for years – how to produce large amounts of antibodies in the laboratory in such a way that all antibody-molecules were identical. This invention earned them the Nobel price in 1984. In the wake of their discovery, hopes were high that