2.1.1. Crop Rotation

Crop rotation is the successive cultivation of different crops in a specified order on the same fields, which prevents the cultivation of the same crop system in two consecutive years on the same field or even area [20]. It is often able to slow down CPB population buildups, but it has to be ensured that fields are properly isolated to avoid easy infestations [20,21]. The impacts of crop rotation and distances between fields on pest populations were first investigated in research from the 1990s [22]. Scientists observed that the distances between rotated fields and the closest potato fields of the previous year were highly related to pest outbreaks on current fields. The further the potato field of the current season was away from the previous season's field, the fewer pest problems the farmers had [9,21]. However, not many studies sugges<sup>t</sup> that crop rotation alone is a suitable method of controlling strong pests such as CPB, but that it could be integrated into a managemen<sup>t</sup> plan together with other alternative methods [20].

#### 2.1.2. Host Plant Resistance

Host plant resistance and tolerance can either occur naturally through evolution and selection or artificially through human, transgenic input [9]. Some are believed to be more successful against CPB infestations than others. However, there is no potato variety which is considered to be fully CPB resistant [23], although several potato plant varieties have shown e ffects in laboratory studies on CPB development time and mortality (e.g., Agria, Pasinler, Marfona, Granola, Caspar) often due to mortality of eggs and immature CPB stages and decrease of fecundity and reproductive rate [24]. Sablon et al. [10] summarized several studies indicating that genetic manipulation, the direct manipulation of an organism's genes using biotechnological methods, of potato plants can be used to intensify the expression of deterrent blends. The incorporation of genes that express leptin and other glycoalkaloids in conventional and wild potato varieties resulted in improved tuber yields and a protecting e ffect against CPB in field and laboratory trials [25].

The transgenic approach describes the strategy for the genetic modification of the pest's host plant. As a result, the plant produces certain substances that are avoided when parts of the plant are consumed or that cause severe damage to the pest [9,20,23]. As transgenic potato varieties are not approved for organic cultivation, basically all varieties that are currently open to the agricultural market are unusable. Furthermore, genetically modified plants that are resistant to insect pests are often not considered a suitable approach in IPM, at least in the case of potato farming. As IPM is mandatory in agricultural production, transgenic plants are not allowed to be used in EU countries, although some of the events (mainly maize varieties) have been registered in some countries. In these countries, farmers can use the seeds from GM plants but they are not eligible to receive state subsidies. Irrespective of their success, the use of genetically modified crops often represents a suitable control method, but also contemporarily inadequate to a majority of consumers, especially in Europe [9].

RNA interference (RNAi) is one additional biotechnology to preserve crops from being infested by pests which has gained a lot of attention within recent years. RNAi might successfully trigger the silencing of certain target genes causing mortality or at least reducing pest fertility and health [26,27]. Using plant protease proregions as regulators, directly induced through bacteria *Escherichia coli* of cysteine proteinases, is another possible alternative. In certain biotechnological systems, the ability to preserve the integrity of companion defense-related proteins from the action of insensitive proteases in target pests has been demonstrated [28]. Many more transgenic approaches to pest control by bacteria, fungi, and other microbes are now available on the market and could be used as an alternative to control pests such as CPB, especially outside of Europe in North and South American countries [9,29].

Overall, host plant resistance was proven to be an e ffective control mechanism against CPB through various studies tested in the field. The potato varieties showing full CPB resistance are those created by the transgenic approach. Even though these methods are not accepted by farmers and consumers who favor pure organic or ecological farming [20] and are not allowed in IPM, the adoption of GM plants resistant to pests and diseases can reduce pesticide use and ensure potato production. There is an open debate on the value of genetically modified food, its potential to solve many of the worlds' hunger and malnutrition problems, and its impact on the environment by increasing yield and reducing reliance upon chemical pesticides [10,23].

#### *2.2. Direct Methods for CPB Control*

#### 2.2.1. Behavioral Interference Methods for CPB

Chemical signals that regulate the behavior of insects usually consist of a mixture of odor substances emitted by plants, insects, and other animals. One type of signal is the aggregation pheromones [30]. For instance, the male-insect-produced unique pheromone (S)-3,7-dimethyl-2-oxo-oct-6-ene-1,3-diol was identified for CPB, and both male and female were found to be attracted to the pheromone in laboratory bioassays [31]. The potential for the use of aggregation pheromone in CPB managemen<sup>t</sup> was observed in the early 2000s in field trials where the pitfall traps baited with the aggregation pheromone were used to capture the colonizing CPB adults coming to the newly planted field. Even though over five times as many CPBs were captured in pheromone traps in comparison to controls, the efficacy decreased after only five days. Nevertheless, the potential for the use of the aggregation pheromone in CPB managemen<sup>t</sup> was observed [32]. This method could only be effective at the beginning of the beetle emergence and colonization of newly planted potato fields, when mean daytime temperatures are below 20 ◦C and adults are not able to fly.

In field experiments, synthetic blends of (R)-and (S)-enantiomers of the same pheromone were tested. Mixtures with as much as 87% or higher optical purity of the second enantiomer attracted CPBs most effectively [33]. The compound was also tested as a combination with various other potato volatiles (e.g., 2-phenylethanol and nonanal) in field and laboratory tests. A mixture with a three-component plant attractant was detected to be most successful. In another experiment, scientists treated potato plants with (Z)-3-hexenyl acetate (þ/–)-linalool and methyl salicylate, a synthetic host volatile mixture, to test the attraction to few days old adult CPBs. For the plant treatment, they used four different doses and compared them to potato plants treated with azadirachtin-based antifeedant as well as untreated plants as control. All the experiments were conducted in greenhouses. The researchers figured out that the beetles favored plants nursed with the attractants over the ones with antifeedant, while only the highest antifeedant dose showed better results than the control. This shows the potential of synthetic attractants as components of a "stimulo-deterrent strategy", rather than antifeedants (at low doses) alone [34].

Behavioral responses of CPB were investigated through bioassays in the laboratory. The scientists tested a variety of 13 different compounds all emitted naturally from potato plants. In addition, they used compounds from tomatoes and soybeans [35]. Beetles were attracted by potato volatiles of damaged foliage, but not by tomato plants. Among the 16 odor components, six blends were attractive, two repellents, and eight without preferences. Even at low concentrations, (Z)-3-hexenyl acetate (+/−)-linalool and methyl salicylate were most attractive, while blends with rather high quantities of volatiles from leaves indicated opposite effects. In general, it was revealed that there are certain blends, even within the compound portfolio of potato (and tomato) host plants, that should be further investigated and considered for CPB managemen<sup>t</sup> [35].

The efficacy of limestone dust as a deterrent at two different concentrations was tested as well. It was successful in decreasing the number of CPBs (eggs and larvae) during the individual stages of development, such as against eggs and from first to fourth instar larval stages [36].

Sablon et al. [10] also summarized several chemical compounds categorizing them into masking odors, trap crops including attractants and aggregation pheromones, and antifeedants [10]. In addition to some of the above-described extracts, they listed a multitude of antifeedants including hydroxides, alcohol extracts of the leaves and bark of *Quercus alba* L., limonin, α-mangostin, sesquiterpenes, terpenoids, lactones, and extracts from various plants including wild species of potatoes. Most of these compounds led to a successful decrease in beetle feeding behavior, but also could potentially prevent female oviposition. This was detected for citrus limonoids, but also some other blends [10]. Therefore, behavioral interference methods can be an efficient and environmentally friendly way of CPB management. They represent a strongly increasing application in potato managemen<sup>t</sup> through laboratory and field studies, while the most widely used methods such as chemical treatments decrease [10].

#### 2.2.2. Physical and Mechanical Control

One approach that aroused bigger attention in the 1980s and 1990s was the bug vacuum [37]. With this method, insects were sucked from the plants into a large machine combined with a tractor pulling it and killed. However, the machine was never a grea<sup>t</sup> success as it came with too many agricultural and environmental disadvantages. It also killed useful and beneficial insects and other animals, but also caused heavy soil compaction due to its weight. The biggest problem was the low success against pests which were found deeper within the crop canopy that could not be reached, so a wide application of this method was never an option [37].

Assays in laboratories identified the usage of wood ash as a possible compound for CPB managemen<sup>t</sup> due to its toxicity against adults and larval stages [38]. When exposing beetles permanently to wood ash for up to 10 days, all beetles of all stages were killed. The decreasing e fficacy after repeated usage as well as the decreasing activity in the field within moist environments were the main detected problems. Nevertheless, the author of the study suggested that thick layers of ash applied as strips around the base of potato plants could act as a physical barrier like a fence, limiting big colonization of beetles as CPBs avoid crossing it [38].

Another alternative possibility to decrease CPB populations is physical control. Pneumatic and thermal pest controls can be used to control various stages within the development of the plant and beetle [39]. Here, scientists use the fact that in the early season, when potato plants start to grow out of the soil, plants are supposed to be less vulnerable to heat than adult beetles and eggs. Propane burners are directed towards the crop rows and eliminate most beetles while plants remain rather healthy [40]. Therefore, to control overwintering CPBs, the e fficacy of flame technology was demonstrated for plant sizes of around eight inches in height [41]. The highest control e fficacy was reached during sunny, warm days as CPBs are more active and often feed on the top of potato plants. Compared to most common insecticides which achieve a control rate of normally 25–50% of overwintering adults, flaming can be very e fficient. In field tests, burning of beetles obtained up to a 90% fatality rate as well as a 30% reduction in the number of eggs to hatch [41]. Under laboratory and field conditions, a single-row insect scorcher for CPB control was tested accordingly. By controlling the temperature of the gasses and contact time in adult beetles, 60% of individuals were injured while the potato plants were not damaged [42].

Laboratory and field investigations illustrated that a combination of steam and air left more than 50% of adult CPBs incapacitated while barely damaged the potato plants. In detail, steam of low pressure was injected into a plant-covering hood [39,43]. It is also known that the CPB answers to disturbances by undergoing a defense strategy defined as thanatosis. By that, the beetles release hold from their host plants and just fall to the ground. Thanatosis can be initiated by using hot air and blowing it on CPBs feeding on the plant [40]. In some studies, researchers found out that the main causes of falling were related to certain exposure time, temperatures, and air velocity [39,40]. Afterward, the apparatus collected the air blown insects with its equipped collection device. Around 65% of fallen beetles could be collected this way [44]. For removed beetles that fall to the ground between crop rows, shielded propane burners were applied to kill them. Here, the e ffectiveness was at least as e fficient as chemical insecticides [39,40].

Another way to hinder pest insects to enter crop fields is the usage of plastic-lined trenches and row covers which can function as physical barriers [45]. For that, synthetic fabric is used to avoid CPBs entering the potato fields. The material can be improved by fine soil particles and arranged in an angle wider than 46◦ to make it impossible for most beetles to have a firm hold on the surface. Even though small numbers of beetles may be able to escape during rain showers when washed away, the material regains its protection as soon as it is dried afterward [45]. A portable variant was

developed by Canadian scientists. With this version placed at field-edges, CPBs are able to walk up the sides of the trap, but from there ge<sup>t</sup> captured when falling into the inside [24].

The CPB is mainly diurnal, but can also be active at night. In one experiment, it was tested how strong the positive phototactic behavior of the beetle was in darkness, when stimulated with different wavelengths of light [46], respectively if low-intensity yellow light was favored over pheromones [47]. In both experiments, continuous yellow light (and in the first experiment also green light) was the most successful wavelength source to affect, capture, and control CPB individuals [46,47]. Physical and mechanical methods are promising alternatives to not only control pest populations efficiently, but also contribute to clean air and water by eliminating insecticide spraying completely.
