Crossley MS, Schoville SD, Haagenson DM, Jansky SH. J Econ Entomol. 2018;111(4):1875-1884. doi: 10.1093/jee/toy120.

Colorado potato beetle, Leptinotarsa decemlineata Say (Coleoptera: Chrysomelidae), is a serious global pest of potato, Solanum tuberosum L. Management of L. decemlineata has relied heavily on insecticides, but repeated evolution of insecticide resistance has motivated the exploration and development of alternative strategies, such as plant resistance. The recent development of two diploid potato families derived from crosses between cultivated and wild potato species (S. chacoense and S. berthaultii) has provided a unique opportunity to reexamine plant traits for resistance breeding. In this 2-yr study, we surveyed select F2 clones for the induction of L. decemlineata mortality and a reduction in defoliation in no-choice feeding assays when challenged with adults and larvae from three sites in Wisconsin. We tested for an association with glandular trichome density and foliar levels of the glycoalkaloids chaconine and solanine. Several potato clones demonstrated resistance in specific feeding assays, but none excelled consistently across experiments. Mortality and defoliation generally differed significantly among L. decemlineata populations, which could be indicative of heritable variation in beetle responses to plant defenses or variation in the physiological status of the beetle populations tested. Contrary to expectations, higher trichome density increased mortality or decreased defoliation in only a few cases, and levels of mortality and defoliation were unrelated to foliar glycoalkaloid content, warranting further investigation of the defense mechanisms of resistant clones. In addition to identifying several potential L. decemlineata resistance sources, this study underscores the need to include multiple insect populations in surveys of plant resistance to this diverse pest.

Cingel A, Savić J, Lazarević J, et al. Insect Science. 2017;24(5):768-780. doi: 10.1111/1744-7917.12364.

Colorado potato beetle (CPB; Leptinotarsa decemlineata Say, Coleoptera: Chrysomelidae) has shown a remarkable adaptability to a variety of control measures. Although oryzacystatin I and II (OCI and OCII) have potential in controlling pests that use cysteine proteinases for food digestion, expression of a single OC gene in potato exhibited a minimal or no effect on CPB fitness traits. The aim of this study was to examine the effect of coexpressed OCI and OCII in potato (Solanum tuberosum L.) cultivars Desiree, Dragačevka and Jelica on CPB larvae. Growth parameters, consumption rates and food utilization, as well as activity of proteases of CPB larvae were assayed. Second and third instar larvae fed on transformed leaves molted earlier and had higher relative growth and consumption rates than larvae fed on nontransformed leaves, while efficiency of food utilization was unaffected. In contrast, fourth instar maximum weight gain and amount of leaves consumed were about 20% lower for the larvae fed on transgenic potato. Analysis of total protease activity of third instar larvae revealed reduction in overall proteolytic activity measured by azocasein hydrolysis, accompanied with inhibition of cysteine proteinase activity 24 h after ingestion of potato leaves expressing OCI and OCII. However, after long-term feeding on transformed leaves proteolytic activities of larvae became similar to the controls. Although feeding on OCI/OCII leaves did not affect larval survival, coexpression of OC genes reduced the development time and thus significantly decreased plant damage caused by CPB larvae.

Guo, W., et al. 2018 J Agric Food Chem. 66(45):11990-11999. doi: 10.1021/acs.jafc.8b03914.

RNA interference (RNAi) has been developed for plant pest control. In this study, hairpin-type double-stranded RNA (dsRNA) targeting the juvenile hormone (JH) acid methyltransferase (JHAMT) gene (dsJHAMT) was introduced in potato plants via Agrobacterium-mediated transformation. The results indicated that the transcriptional RNA of dsJHAMT accumulated in the transgenic plants. The transcripts and proteins of the L. decemlineata JHAMT gene were significantly reduced in larvae feeding on dsJHAMT transgenic foliage. The dsJHAMT had a significant negative effect on the growth and development of L. decemlineata, especially resulting in less oviposition. Importantly, in the field trials, transgenic plants are high-efficiently protected from insect damage mainly because surviving insects laid fewer or no eggs. Even full protection from beetle damage can be acquired by continuously lowering insect population size at large scale in the field over the years. Therefore, the transgenic plants expressing dsJHAMT successfully provided an additional option for plant pest control.

Wetzel WC, Aflitto NC, Thaler JS. 2018. Ecology. 99(10):2338-2347. doi: 10.1002/ecy.2472.

A growing number of studies have manipulated intraspecific plant diversity and found dramatic changes in the densities of associated insect herbivores and their predators. While these studies have been essential for quantifying the net ecological consequences of intraspecific plant diversity, they have been less effective at uncovering the ways in which plant diversity alters trophic interactions within arthropod communities. We manipulated intraspecific plant diversity and predation risk in the field in a factorial design to reveal how a mixture of plant genotypes changes the response of an herbivorous beetle (Leptinotarsa decemlineata) to a common stink bug predator (Podisus maculiventris). We repeated the manipulations twice across the ontogeny of the beetle to examine how the effects of diversity on the predator-prey interaction differ between larval and adult stages. We found that intraspecific plant diversity, mixtures of susceptible and resistant varieties of potato (Solanum tuberosum), reduced larval survival by 20% and adult oviposition by 34%, which surprisingly put survival and oviposition lower in the mixed-genotype plots than in the resistant monocultures. Moreover, we found that predation risk reduced larval survival 25% and 11% in resistant and susceptible monocultures, respectively, but had no effect in the mixture. This result indicated that our genotypic mixing treatment interacted nonadditively with predation risk such that plant diversity altered the predator-prey interaction by changing the responses of the beetles to their stink bug predators. In addition, even though predation risk reduced larval survival, it increased adult overwintering survival by 9%, independently of plant treatment, suggesting that these interactions change through ontogeny. A key implication of our study is that plant diversity influences arthropod communities not only by changing resource quality, as past studies have suggested, but also by changing interactions between species within the arthropod community.

Gabaston J, El-Khawand T, Waffo-Teguo P, et al. Journal of Pest Science. 2018;91(2):897-906. doi: 10.1007/s10340-018-0956-2.

Stilbenes are phenolic compounds which are produced in large amounts in vine and are involved in plant defence as phytoalexins. Oligomeric forms have recently proven to be the most active compounds against a wide range of parasites such as fungi, bacteria or algae. The aim of this study was to investigate the activity of a grapevine root extract which is a stilbene oligomer pool against Leptinotarsa decemlineata, a major pest of Solanaceae crops. Analysis by UHPLC-DAD-MS of the stilbene-enriched extract obtained from grapevine root (Riparia Gloire de Montpellier rootstock) highlighted twelve stilbenes at 25% (w/w). The major stilbenes found in root extract were isolated such as the tetramers vitisin B, vitisin A and hopeaphenol; the dimers ampelopsin A and E-ε-viniferin and the monomer E-resveratrol. The insecticidal effects of this extract as well as the main compounds were investigated against L. decemlineata larvae. The extract caused chronic toxicity, inhibited larval development and, to a lesser extent, inhibited food intake. The high concentrations of vitisin A and vitisin B in grapevine root contributed to this effect as they are the most toxic compounds. Outdoor pot experiments revealed the efficacy of stilbene-enriched extract with high mortality of L. decemlineata and protection of potato plants. The extract also revealed an absence of toxicity against non-targeted organisms such as earthworms (Eisenia fetida). Thus, these results strongly suggest that grapevine roots are a promising source of bioactive stilbenes for the development of natural insecticides.

Ahmed HAA, Onarıcı S, Bakhsh A, et al. Plant Biotechnology Reports. 2017;11(5):315-329. doi: 10.1007/s11816-017-0453-8.

The expression of insecticidal genes must be induced at appropriate time and in sufficient amount to confer protection against targeted pests. However, the increased scientific reports of resistance development in insect pest against insecticidal delta-endotoxins, produced by Bacillus thuringiensis, provide impetus for the development of alternative insect management strategies. The present study was conducted to investigate the importance of targeted expression of a hybrid insecticidal gene (SN19) in potatoes. For this purpose, two plant expression vectors were constructed by cloning hybrid SN19 gene (cry1Ba-domain I-III and cry1Ia-domain II) under the control of a wound-inducible promoter isolated from Asparagus officinalis (AoPR1) and CaMV 35S promoter, and were transferred to Agrobacterium tumefaciens strain EHA 105. Four potato genotypes (Marabel, Innovator, Tokat 10/1 and Tokat 6/24) were transformed with EHA 105 strain harboring pTF101.1 35S-SN19 and pTF101.1 AoPR1-SN19 constructs. Phosphinothricin (PPT) was used at concentration of 1 mg/l for selection of primary transformants. PCR results showed the presence of both introduced SN19 and bar genes in 43 plants out of total 154 putative transgenics. Expression of SN19 protein in primary transformants was confirmed by Western blot assays. The mechanical wounding of transgenic plants exhibited more accumulated levels of SN19 proteins during post wounding period. Leaf biotoxicity assays with Colorado potato beetle (Coleoptera) and tomato leafminer (Lepidoptera) exhibited 100% mortality of the pests in primary transformants. Based on our mortality results with both constructs, we concluded that the potato transgenic lines exhibited targeted expression of insecticidal gene under the control of AoPR1 promoter upon insect wounding with eliminated toxicity of Cry protein and hence can be further used effectively in potato breeding programme.

Hussain T, Aksoy E, Çalișkan ME, Bakhsh A. Transgenic Res. 2019;28(1):151-164. doi: 10.1007/s11248-018-0109-7.

Most of the commercialized insect resistant transgenic crops express cry gene(s) isolated from Bacillus thuringiensis; however, intensive cultivation of Bt crops over almost two decades has been questioned regarding its sustainability and durability in pest management. The present study focused on silencing of highly specific molting-associated Ecdysone receptor (EcR) gene of Colorado potato beetle (CPB) using RNA interference (RNAi) approach. The partial cDNA of EcR gene of CPB was amplified using specific primers in sense and anti-sense orientations, and cloned in pRNAi-GG vector flanked by an intronic sequence (pdk). Leaf and internodal explants of Agria and Lady Olympia potato cultivars were infected with Agrobacterium strain LBA4404 harboring constructs under the control of CaMV 35S promoter. Standard molecular analysis of primary transformants showed proper integration of T-DNA in plant genome. The transgenic plants of both cultivars were evaluated for their efficacy against first, second and third instar CPB larvae. The leaf biotoxicity assays revealed 15-80% of CPB mortality. A significantly lower fold-change (0.87-4.14×) in larval weight was observed in insects fed on transgenic plants compared to the ones fed on control plants (1.87-6.53×). Furthermore, CPB larvae fed on transgenic plants exhibited reduced EcR transcripts, indicating the functionality of dsRNA EcR in silencing EcR gene expression. This study is an excellent example of the integration of an alternative, effective and reliable method to cope with potato insect pests that incur significant losses to potato production in the world.

Molnár I, Besenyei E, Thieme R, et al. Pest Manag Sci. 2017;73(7):1428-1437.

BACKGROUND: Colorado potato beetle (CPB) has become the biggest enemy of cultivated potato worldwide. One of the most effective sources of resistance to CPB is Solanum chacoense, an accession with a high leptine glycoalkaloid content. The aim of our study was to assay the repellence and toxicity of S. chacoense, its somatic hybrids (SHs) and their backcross progenies (BC1) with potato for CPB adults and larvae. Transgenic S. chacoense, deficient in DNA mismatch repair (MMR), was also used to produce SHs, in order to increase homeologous recombination and hence introgression of wild-species DNA into the potato gene pool. RESULTS: Wild-type SH was highly resistant to CPB. Resistance to CPB of BC1 progenies showed a 1:3 inheritance pattern. MMR-deficient SHs performed better in the resistance analysis. Most MMR-deficient SHs had a similar toxicity as S. chacoense and an intensely repellent effect on CPB adults. Resistance of SHs and BC1 clones may be attributed to leptine biosynthesis, which was confirmed using a RAPD marker. CONCLUSION: This is the first report of SHs and their progenies exhibiting both antibiosis and antixenosis against CPB. Resistant SHs are an important step forward in combating this voracious pest of potato.

Malik RJ, Ali JG, Bever JD.  Pedobiologia. 2018;66:29-35. doi: 10.1016/j.pedobi.2017.12.004.

While arbuscular mycorrhizal (AM) fungi may have a prominent role in trophic ecology, mycorrhizal improvement or reduction on herbivore growth and survival may also be dependent on herbivore's stage of development. Solanum lycopersicon (tomato) was grown on sterile background soil treated with either mycorrhizal inoculant (AM+) or non-mycorrhizal control (AM-). Mycorrhizal treatments included four single species of AM-fungi (Entrophospora infrequens, Funneliformis mosseae, Claroideoglomus claroideum, and Racocetra fulgida) and a mixture of all four species (fungal community). To determine if mycorrhizal treatment indirectly alters the ability of beetle larvae (Leptinotarsa decemlineata) to access plant resources, plant damage and trichome density were quantified as plants were infested with a single neonate (early-stage) for 96 hours (h). In a second experiment, beetle growth rate was assessed as plants were infested with a single third-instar (late-stage). After 72 h of late-stage beetle infestation, beetle mass was measured. It was found that early-stage beetles inflicted more damage on AM+ tomatoes. Interestingly, this corresponds with fewer trichomes on AM+ tomatoes, as well as higher early-stage beetle survivorship. Specifically, AM taxon, C. claroideum increases herbivory and thereby reduces beetle mortality. Among late-stage beetles, C. claroideum does not improve beetle growth nor rate of survival. This suggests that AM taxa that are beneficial to early-stage beetles may not necessarily provide an advantage to late-stage beetles. Taken together, these findings highlight potential dependencies of AM-fungal effects on herbivory and herbivore life history, including growth and life-stage specific survival.

Andrés MF, Rossa GE, Cassel E, et al. Food and Chemical Toxicology. 2017;109:1086-1092.

In this study we evaluated the effect of a pressure gradient (1-2 atm) in the extraction and composition of the essential oil (EO) of Piper hispidinervum by steam distillation. We also evaluated the insect antifeedant effects (Spodoptera littoralis, Leptinotarsa decemlineata, Myzus persicae and Rhopalosiphum padi) and nematicidal activity (Meloidogyne javanica) of the oils, their major components and their synergistic interactions. Safrole was the major component (78-81%) followed by terpinolene (5-9%). The EOs tested were effective insect antifeedants. Safrole, explained most of the insect antifeedant action of P. hispidinervum EOs. When safrole and terpinolene were tested in binary combinations, low ratios of safrole improved the antifeedant effects of terpinolene. P. hispidinervum EOs caused higher mortality of M. javanica juveniles than their major components. In binary combinations, low ratios of terpinolene increased the nematicidal effects of safrole. The EO treatment strongly suppressed nematode egg hatching and juvenile infectivity. P. hispidinervum EOs affected the germination of S. lycopersicum and L. sativa mostly at 24 h of treatment, being L. sativa the most sensitive. Safrole moderately affected germination and root growth of L. sativa, S. lycopersicum and L. perenne. Terpinolene only affected S. lycopersicum root growth.