Epidermis-Specific Metabolic Engineering of Sesquiterpene Formation in Tomato Affects the Performance of Potato Aphid Macrosiphum euphorbiae

Tomato produces a number of terpenes in their glandular trichomes that contribute to host plant resistance against pests. While glandular trichomes of cultivated tomato Solanum lycopersicum primarily accumulate a blend of monoterpenes, those of the wild tomato species Solanum habrochaites produce various sesquiterpenes. Recently, we have identified two groups of sesquiterpenes in S. habrochaites accessions that negatively affect the performance and choice behavior of the potato aphid (Macrosiphum euphorbiae). Aphids are piercing-sucking herbivores that use their mouthpart to penetrate and probe plant tissues in order to ultimately access vascular tissue and ingest phloem sap. Because secondary metabolites produced in glandular trichomes can affect the initial steps of the aphid feeding behavior, introducing the formation of defensive terpenes into additional plant tissues via metabolic engineering has the potential to reduce tissue penetration by aphids and in consequence virus transmission. Here, we have developed two multicistronic expression constructs based on the two sesquiterpene traits with activity toward M. euphorbiae previously identified in S. habrochaites. Both constructs are composed of sequences encoding a prenyl transferase and a respective S. habrochaites terpene synthase, as well as enhanced green fluorescent protein as a visible marker. All three coding sequences were linked by short nucleotide sequences encoding the foot-and-mouth disease virus 2A self-processing oligopeptide which allows their co-expression under the control of one promoter. Transient expression of both constructs under the epidermis-specific Arabidopsis CER5-promoter in tomato leaves demonstrated that formation of the two sets of defensive sesquiterpenes, β-caryophyllene/α-humulene and (−)-endo-α-bergamotene/(+)-α-santalene/(+)-endo-β-bergamotene, can be introduced into new tissues in tomato. The epidermis-specific transgene expression and terpene formation were verified by fluorescence microscopy and tissue fractionation with subsequent analysis of terpene profiles, respectively. In addition, the longevity and fecundity of M. euphorbiae feeding on these engineered tomato leaves were significantly reduced, demonstrating the efficacy of this novel aphid control strategy.

The results of catching winged aphids with water traps on potato plantings in the southern part of the Arkhangelsk region

Dynamics of the number and species composition of aphids were studied on seed potatoes using yellow water traps in the southern part of the Arkhangelsk region in 2018-2019. 12 species of winged aphids (165 insects) were identified in 2018, and 16 species (115 insects) - in 2019. The following types dominated: black bean aphid A. fabae Scop., bird cherry-oat aphid R. padi L., blackcurrant-sowthistle aphid H. lactucae L., glasshouse-potato aphid A. solani Kalt. The share of aphids as direct and indirect vectors of viral diseases was 91 % of the total number in 2018, and 79.1 % in 2019. The pest population of potato plants depended directly on weather conditions of the growing season. The sum of the average daily temperatures during the period of catching aphids was 1273.3 C in 2018, and 983.3 C - in 2019. During the experimental period, 131.6 mm of precipitation fell in 2018, and 280.4 mm - in 2019. Due to the constant presence of aphids on seed potato crops, annual monitoring of pest population and, in case of necessity, protective measures are required.

Biopesticide Evaluation from Lab to Greenhouse Scale of Essential Oils Used against Macrosiphum euphorbiae

Aphids are recognized as a major threat to economically important crops. Their control is predominantly based on synthetic insecticides that are detrimental to human health and the environment. Botanical pesticides based on essential oils (EOs) are a promising alternative. In this study, the entomotoxicity of green anise and fennel EO fumigation was tested on the potato aphid Macrosiphum euphorbiae. Three different settings of increasing scale were considered (leaflet, whole plant and greenhouse) to appraise the consistency of EO impact from controlled laboratory to greenhouse production conditions. LC50 values for green anise and fennel were 6.6 μl L−1air and 12.2 μl L−1air, respectively, based on dose-response curves in leaflet experiments but fennel EO induced phytotoxicity. EO efficiency was confirmed at the whole-plant scale. In the greenhouse experiment, fennel EO exhibited greater efficiency than at the laboratory scale equaling green anise EO efficiency but both EOs showed delayed phytotoxicity, illustrating the importance of long-term monitoring. The present study revealed the ability of both EOs to control M. euphorbiae populations under greenhouse conditions and hinted at the importance of assessing EO efficiency in realistic agronomic conditions (e.g., under the fluctuating environmental conditions usually occurring in greenhouses).

Intron-Containing Hairpin RNA Interference Vector for OBP8 Show Promising Mortality in Peach Potato Aphid

Myzus persicae is a devastating pest affecting potato production. RNA Interference technology is used against essential odorant binding protein 8 (OBP8) to enhance protection against Myzus persicae in potato. Gene was isolated, sequenced and GenBank IDs were allotted and ERNAi was used to design siRNA targets from OBP8 with no off-targets. Multiple Sequence Alignment show M. persicae OBP8 resemblance with Acyrthosiphon pisum, Rhopalosiphum maidis, Aphis fabae, and Sitobion avenae. DsRNA (7 µg/µl) oral acquisition had resulted in 69% mortality and 58% reduction in OBP8 expression 8D post dsRNA feeding in comparison to control. Golden Gate (GG) cloning based RNAigg is used for RNA interference taking advantage of type IIs restriction enzyme Eco31I. Agro infiltration assay was used for introduction of intron-containing hairpin RNA (ihpRNA) in Solanum tuberosum. Aphids feeding on transgenic S. tuberosum show 57.6% mortality and 49% reduction in OBP8 expression 8d post-feeding in comparison to control. This work proves OBP8 as promising ihpRNA targets in potato and related crops for whom Myzus is a devastating target.

Induced Local and Systemic Defense Responses in Tomato Underlying Interactions Between the Root-Knot Nematode Meloidogyne incognita and the Potato Aphid Macrosiphum euphorbiae

Plants mediate interactions between different herbivores that attack simultaneously or sequentially aboveground (AG) and belowground (BG) organs. The local and systemic activation of hormonal signaling pathways and the concomitant accumulation of defense metabolites underlie such AG-BG interactions. The main plant-mediated mechanisms regulating these reciprocal interactions via local and systemic induced responses remain poorly understood. We investigated the impact of root infection by the root-knot nematode (RKN) Meloidogyne incognita at different stages of its infection cycle, on tomato leaf defense responses triggered by the potato aphid Macrosiphum euphorbiae. In addition, we analyzed the reverse impact of aphid leaf feeding on the root responses triggered by the RKN. We focused specifically on the signaling pathways regulated by the phytohormones jasmonic acid (JA), salicylic acid (SA), abscisic acid (ABA), and indole-3-acetic acid (IAA) as well as steroidal glycoalkaloids as induced defense compounds. We found that aphid feeding did not induce AG hormonal signaling, but it repressed steroidal glycoalkaloids related responses in leaves, specifically when feeding on plants in the vegetative stage. Root infection by the RKN impeded the aphid-triggered repression of the steroidal glycoalkaloids-related response AG. In roots, the RKN triggered the SA pathway during the entire infection cycle and the ABA pathway specifically during its reproduction stage. RKN infection also elicited the steroidal glycoalkaloids related gene expression, specifically when it was in the galling stage. Aphid feeding did not systemically alter the RKN-induced defense responses in roots. Our results point to an asymmetrical interaction between M. incognita and Ma. euphorbiae when co-occurring in tomato plants. Moreover, the RKN seems to determine the root defense response regardless of a later occurring attack by the potato aphid AG.

Macrosiphum euphorbiae (potato aphid).

Host-alternating populations of M. euphorbiae disperse through migratory flights in the spring and autumn. The spring migration takes aphids from the over-wintering host-plant (Rosa spp.) to a wide range of secondary host-plants, including potato, tomato, lettuce and other cultivated plants. Non-host alternating populations can survive year-round on secondary hosts, especially in greenhouse and other indoor environments. Aphids can be carried on foliage in trade. There is little evidence to suggest that the geographical range of this aphid is currently expanding.

Repellent and Antifeedant Activities of Citral-Derived Lactones against the Peach Potato Aphid

Citral is well known for its antimicrobial, antifungal, and insecticidal activities. Natural sesquiterpene α-methylenelactones also exhibit a broad spectrum of biological activities. The aim of the study was to explore the effect of structural changes to citral molecules on citral behavior-modifying activity towards Myzus persicae. Specifically, the effects of the introduction of a γ-lactone moiety and methylene groups in α and γ positions of the lactone ring were investigated. The lactones were obtained in five-step (saturated lactone and γ-methylenelactone) or six-step (α-methylenelactone and α,γ-dimethylenelactone) syntheses from citral. The synthetic procedures and physical and spectral data of the lactones are presented. The settling behavior of freely moving aphids in choice and no-choice situations was monitored. The probing behavior of tethered M. persicae using the Electrical Penetration Graph (EPG) technique was also analyzed. Citral appeared a strong repellent and pre-ingestive and ingestive probing deterrent to M. persicae. The incorporation of a lactone moiety caused the loss of the repellent activity. α-Methylenelactone inhibited aphid settling and probing activities at pre-ingestive and ingestive phases. The saturated γ-lactone and α,γ-dimethylenelactone were the settling post-ingestive deterrents to M. persicae, which did not affect aphid probing activity. γ-Methylenelactone did not affect aphid behavior.