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Biological control using predatory mites is an important component of pest management in a wide range of crops, including grapes. However, the trophic relationships established in these environments are still not fully understood. The use of molecular tools can contribute to identify the food sources of predatory mites and disentangle the complex relationships occurring in the field. We hypothesized that using qPCR and TaqMan probes we could quantify the proportion of plant based food sources and animal diet of various predatory mites throughout the season. Hence, our specific objectives were: 1) to design speciesspecific primers and TaqMan probes to identify the main mite groups in vineyards from La Rioja DOC Region (northern Spain), 2) to detect prey and plant products in predatory mites via PCR and qPCR, and 3) to validate these methods using predatory mites collected from the field. First, we designed specific primersprobe sets to identify the main predatory mite species in the region (Typhlodromus pyri and Kampimodromus aberrans) as well as to detect their potential prey (e.g. tetranychiids, tydeiids and eriophyid mites) and plant food sources. Then, we developed a predator-prey system (T. pyri and Tetranychus urticae) to quantify DNA by qPCR and establish post-consumption detection times of plant and animal food sources. Third, we collected predatory mites from the vineyard to unravel their prey items and establish their trophic interactions. Ultimately, detecting food sources within predatory mites could provide a useful tool to better understand the effectiveness of biological control using predatory mites in vineyards

[ES] Los nematodos entomopatógenos (NEPs) son agentes de control biológico comúnmente presentes en el suelo de los cultivos. Las prácticas agrícolas de laboreo convencional y aplicación de agroquímicos predisponen a la pérdida de biodiversidad del suelo, comprometiendo su salud y alterando el equilibrio natural de factores abióticos y bióticos que modulan la abundancia y actividad de los NEPs. El viñedo, soporte de un sector socioeconómico de gran relevancia mundial, es uno de los sistemas de cultivo más más se apoya en las prácticas del laboreo intensivo y la aplicación de productos agroquímicos. De este modo, con el fin de avanzar en su sostenibilidad, son necesarios nuevos enfoques en el manejo del viñedo. Nuestra hipótesis es que las estrategias alternativas al laboreo y a la liberación excesiva de agroquímicos en los manejos del suelo y de plagas, enfermedades y malas hierbas en vid, como son la implementación de cubiertas vegetales o acolchados y la agricultura ecológica, favorecerán la comunidad nativa de NEPs en el suelo del viñedo. De este modo, nuestro objetivo fue evaluar el impacto de diferentes prácticas vitivinícolas sobre las poblaciones nativas de NEPs y otros organismos específicos asociados a su red trófica del suelo y cómo sus interacciones podrían ser indicativo de la salud del suelo en los viñedos. Para ello, combinamos metodologías tradicionales e innovadoras con el fin de aislar e identificar organismos de la mesofauna del suelo. En primer lugar, estimamos diferentes actividades del suelo, incluidas aquellas asociadas a los NEPs, empleando larvas de Galleria mellonella como cebos en las muestras de suelo usadas como trampas de insectos. Además, utilizamos cebadores y sondas específicos para identificar y cuantificar, mediante qPCR, la presencia y abundancia de 10 especies de NEPs y otros 12 organismos asociados a su red trófica: cuatro nematodos de vida libre (NVLs), seis hongos nematófagos (HNs) y dos bacterias ectoparásitas (BcPs). Por últim, [EN] Entomopathogenic nematodes (EPNs) are biological control agents that often occur naturally in crop soils. The conventional agricultural practices of regular tillage and agrochemical applications predispose to soil biodiversity losses, compromising soil health and disrupting the natural balance of abiotic and biotic factors that might modulate EPN abundance and activity. The vineyard, which supports a relevant socio-economic sector worldwide, is one of the most intensively managed cropping systems. Therefore, approaches rather than reliance on mechanization and agrochemicals are needed to achieve more sustainable viticulture. We hypothesized that alternative strategies to tillage for soil management and the release of agrochemicals for pests, diseases, and weed management, such as cover cropping, mulching, and organic farming, would favor the native EPN community in vineyard soils. Therefore, our objective was to evaluate the impact of differentiated viticulture practices on native EPNs and other targeted organisms associated with their soil food web and how their assemblage might signal soil health in vineyards. We implemented traditional and innovative methodologies to isolate and identify mesofauna to achieve this aim. Firstly, we estimated different soil activities, including those associated with EPNs, by baiting the soil samples with Galleria mellonella larvae. Besides, we used speciesspecific primers/probe qPCR sets to screen and quantify the occurrence and abundance of 10 EPN species and 12 organisms linked to their soil food web: four free-living nematodes (FLNs), six nematophagous fungi (NF), and two ectoparasitic bacteria (EcPB). Lastly, a third soil subsample set was employed to estimate the soil properties. Following this procedure, we performed three independent studies to evaluate the impact of different management practices on the EPN community and associated soils organisms in The Appellation of Origin (DOCa) Rioja vineyards (Northern Spain): (i

The entomopathogenic nematodes (EPNs) are biological control agents that are widespread in crop soils. However, traditional agricultural management practices such as cultivation and agrochemical usage can alter the soil balance that enables their occurrence and activity. Alternative strategies like mulching are commonly employed to prevent weed growth, enhance below-ground biodiversity by improving soil, organic matter content, fertility, and moisture. We hypothesized that organic mulches would favor biotic conditions for nematofauna development in crop soil, including EPNs, compared to herbicide application or tillage. Traditional (insect baits) and molecular (qPCR analysis) tools were used in this study to assess the abundance and activity of native EPNs, and the abundance of potential natural enemies, such as free-living nematode (FLN) competitors, nematophagous fungi (NF), and ectoparasitic bacteria, in soils managed with different organic mulches or traditional practices. As a model agroecosystem, we selected the vineyard, one of the most intensively managed crop systems. We compared mulches of grape pruning debris (GPD-M), straw (Str-M), and spent mushroom compost (SMC-M) in two commercial vineyards, which employed either integrated or organic pest and disease management. Following a completely randomized design, we retrieved two composite samples per plot (n = 3 per treatment in each vineyard) in April, June, and October 2020. Numbers of EPNs and selected members of their soil food web were higher in the organic than the integrated managed vineyard. Supporting our hypothesis, organic mulching overall favored nematode occurrence in both vineyards. We found higher NF abundance for Str-M, and GPD-M in the organic vineyard, which plausibly explained the lower EPN activity and occurrence compared to SMC-M in both vineyards. We conclude that the organic mulches can provide appropriate conditions for increasing nematofauna numbers but, depending on the mulch type, m

Vineyards and their associated socio-economic activities are relevant sectors worldwide. Still, this agroecosystem is one of the most intensely managed crops and erosion-prone land areas. The conventional viticulture practices to control pests, diseases, and weeds, like tillage and agrochemical applications, accelerate the loss of soil biodiversity and compromise the presence of beneficial soil organisms such as the entomopathogenic nematodes (EPNs). Such human disturbances in the agroecosystems can strongly affect abiotic (e.g., soil texture and properties) and biotic factors (natural enemies and potential competitors) that modulate the EPN activity as biological control agents. For the first time in viticulture, this study aimed to investigate the impact of differentiating management on the EPN community and associated soil organisms and if their assemblage will provide indicators of better practices for sustainable farming. We hypothesized that organic pest management and alternative strategies to tillage might enhance the abundance and activity of the native EPN community in vineyard soils. In autumn 2019, we collected two composite soil samples from 80 vineyards distributed across the Guaranteed Designation of Origin (denominated DOCa) Rioja region. The sites belonged to one category of each of the two factors: pest management (integrated vs. organic, 40 plots each) and soil managing (tillage vs. cover cropping, 48 and 32 vineyards, respectively). Isolated through sucrose-gradient centrifugation and employing species-specific primers/probe qPCR sets, we investigated the presence of ten EPN species and associated soil organisms: four free-living nematodes (FLNs), six nematophagous fungi (NF), and two ectoparasitic bacteria (EcPB). Besides, we estimated the EPN activity using the traditional insect-bait method. We included in the analysis twenty soil variables to characterize the evaluated treatments and assess their impact on soil organism distribution. Our resu

Viticulture is a valuable sector worldwide with an extraordinary socio-economic impact in Spain. Numerous pests and diseases threaten vineyards, and their management primarily relies on the use of conventional agrochemicals. The current paradigm of sustainability pursues the implementation of ecologically sound strategies in vineyard ecosystems. The use of cover crops is arising as an alternative with numerous benefits, including favoring above-belowground biodiversity and the presence of beneficial soil organisms such as the entomopathogenic nematodes (EPNs). We hypothesized that the use of specific cover crops in vineyards might enhance the natural occurrence and activity of EPNs by modulating the assemblage with associated organisms. We performed the experiments in an ongoing experimental vineyard (Vitis vinifera var Tempranillo, clon RJ-26, rootstock ‘110-Richter’) located in Logroño (Spain), drove with different soil management systems (three replicates each): conventional tillage practice and the cover crops (i) seeded with Bromus catharticus (Poaceae), (ii) flower-driven, and (iii) spontaneous. We took four soil composite samples per plot (n = 48 per sampling time) late spring and early autumn in two consecutive years (2017 and 2018). By using species-specific primers/probes qPCR sets, we screened for the presence and abundance of eight EPNs species and 12 related soil organisms: six nematophagous fungi, four free-living nematodes, and two ectoparasitic bacteria. Additionally, we assessed the EPN activity by the traditional insect-bait method. Overall, we recorded higher EPN numbers or activity rates on cover crops than on bare soils. However, some of the results were divergent among no-till treatments. We observed not only higher EPN abundance and activity on spontaneous covers but lower numbers of antagonistic organisms, particularly endoparasitic nematophagous fungi. Thus, according to our results, the use of spontaneous covers could be the most promising

Most predatory mites belong to the family Phytoseiidae (Acari). Throughout the world, phytoseiids are involved in the biological control of phytophagous mites in vineyards. Conservative strategies, including cover-vegetation management, are essential to achieve environmentally friendly viticulture. The abundance and diversity of phytoseiid mites in the grapevine canopy and the vegetal ground cover of a Mediterranean vineyard were surveyed by weekly samplings, from early May until the end of September for two years (2016 and 2017). Three types of soil management without herbicide application were analysed and referred to as >Tillage>, >Spontaneous Cover>, and >Flower-driven Cover> treatments. Six phytoseiid species were collected on the grapevine canopy, with Typhlodromus pyri being the dominant species (99.5 %). Five phytoseiid species were recorded in the ground cover, with Typhlodromus and Neoseiulus as the major genera. The Flower-driven Cover treatment showed the highest abundance of phytoseiids in the grapevine canopy. However, both species richness and abundance of phytoseiid mites on the ground-cover vegetation were highest in the Spontaneous Cover treatment. These observations suggest that improving vegetation cover would promote both the abundance and diversity of phytoseiid mites in vineyards because the greater supply of pollen would enhance their survival. Therefore, the use of cover crops in vineyards represents a means of improving vineyard ecosystems by conservative biological control.

Entomopathogenic nematodes (EPNs) are well-studied biocontrol agents of soil-dwelling arthropod pests. The insecticidal efficiency of EPNs is modulated by food web dynamics. EPNs can reproduce in freeze-killed insect larvae, even in competition with free-living bacterivorous nematodes (FLBNs) in the genus Oscheius. The objective of this study was to assess the efficiency of EPNs as scavengers when competing with free-living saprophagous nematodes and fungi, and to determine the possible impact on subsequent EPN offspring fitness. Live and freeze-killed larvae of Galleria mellonella were used to evaluate the reproduction rate and progeny fitness of two EPN species, Heterorhabditis bacteriophora and Steinernema feltiae, applied individually or combined with the FLBN species Oscheius onirici or Pristionchus maupasi, or Aspergillus flavus, an opportunistic saprophytic fungus. We hypothesized that (1) EPN scavenging behaviors previously observed (for H. megidis and S. kraussei) apply to other EPN species, (2) infective juveniles (IJs) emerging from freeze-killed larvae will display reduced pathogenicity and reproduction, and (3) fitness reduction will be amplified by exposure to other organisms competing for the resources. The reproduction rate of S. feltiae was lower in freeze-killed larvae than in larvae infected and killed by the nematode, whereas H. bacteriophora failed to reproduce as a scavenger. The S. feltiae F1 IJs that emerged from freeze-killed larvae exhibited lower pathogenicity rates than IJs resulting from entomopathogenic activity, and also lower reproductive rates if they experienced high FLBN competitive pressure during development. This study illustrates that scavenging is a suboptimal alternative pathway for EPNs, especially in the face of scavenger competition, even though it provides a means for some EPN species to complete their life-cycle.

Sustainable agriculture requires the support of new tools for successful and low-impact management of pests and diseases at global and local scales. The implementation of cover crops is considered a promising alternative ecological structure for sustainable agriculture, because they provide refuge to natural enemies of pests. Specifically, we hypothesised that implementing cover crop vegetation in a vineyard can enhance populations of natural enemies as a measure to support conservation biological control. We analysed insect abundance and diversity on Vitis vinifera var. Tempranillo (clon RJ-26, rootstock “110-Richter”) in a Mediterranean vineyard located in La Rioja, Spain from spring to autumn 2016–2017. Two types of sampling systems (pitfall traps and vacuum sampling) were used in each of the three evaluated ground cover management approaches (n = 3): (i) tillage, (ii) spontaneous cover, and (iii) flower-driven cover. A total of 58 families that belonged to four functional subgroups of insects (predators, parasitoids, phytophagous insects, and pollinators) were found. Overall, differences among the three treatments were only detected for total abundance of insects collected in the ground traps, and the “spontaneous cover” treatment showed significantly higher predator richness and abundance. However, the higher Shannon–Wiener value was recorded for “flower-driven cover” at both the ground and canopy levels. In addition, the abundance of predators, parasitoids and phytophagous insects in the ground traps for both cover crop treatments was more than two times higher than that observed in “tillage” treatment. Several predator taxa, such as ants, ground beetles, earwigs, and vespoid wasps had significantly higher representation under spontaneous cover treatment. Moreover, for parasitoids, the “flower-driven cover” treatment revealed higher Hill numbers values (0D, 1D, and 2D) at the canopy level, although no significant differences were observed. These results indica

Viticulture is a worldwide valuable sector, with special relevance in Spain for their socio-economic impact. Vineyards are threatened by numerous pests and diseases, and their management relies primarily on use of conventional agrochemicals. Current paradigm of sustainability pursues the implementation of ecologically sound strategies. The use of cover crops is arising as an alternative with numerous benefits, including favouring above-belowground biodiversity. We hypothesize that vineyard with cover crops might increase the prevalence of entomopathogenic nematodes (EPNs), biocontrol agents of soil pests, modulating the assemblage of other associated organisms. The presence and activity of native EPNs were evaluated in an ongoing experiment in La Grajera (Logroño, Spain) on Vitis vinifera var Tempranillo (clon RJ-26, rootstock “110-Richter”) investigating tillage, and three types of covers: spontaneous, flower-driven, and seeded with Bromus perenne (Poaceae) (n = 3 per treatment). During late-spring and early-autumn 2017, 48 composite soil samples (12 cores, 2.5 cm diam X 20 cm depth) were taken to ensure balanced spatial distribution (two pair’s rows/inter-rows per plot). Each sample was assessed for the presence and abundance of 9 EPNs and other related organisms: 6 nematophagous fungi, 5 free-living nematodes (FLNs), and 2 ectoparasitic bacteria, by qPCR approaches. Moreover, traditional insect-bait provided the measurement of EPN activity. In spring 2017, nematode activity was slightly higher (P = 0.068) in inter-rows than in rows. However, this trend was only observed for the tillage and seeded treatments in later autumn (P < 0.05). Two EPN species were isolated: Steinernema feltiae and, for the first time in La Rioja, Heterorhabditis bacteriophora. The insect-baits detected low EPN-FLN incidence in the isolates, indicating that EPNs could suffer low competition with other nematodes for the cadaver under this scenario. The ongoing surveys in 2018 will supply ad

Conservative techniques, such as ground cover management, could help promote viticulture sustainability, which is a goal of conservation biological control, by providing shelter and food sources for predatory insects. A field experiment was conducted in a Mediterranean vineyard to evaluate ground cover management impacts on predatory insect and potential grapevine pest abundance and diversity, both on the ground and in the grapevine canopy. Three different ground cover management techniques (tillage, spontaneous cover and flower-driven cover) were tested for two years (2016 and 2017). Overall, the ground cover management significantly affected the abundance of important epigeal predators, of which carabids, forficulids and staphylinids were the most captured. The carabid abundances under both the cover crop treatments were found to be approximately three times higher compared with that under the tillage treatment. In contrast, the canopy insect abundance in the vineyard was similar among the treatments for both the predators and the potential grapevine pest species. These results indicate that cover crop vegetation can be used in vineyards to enhance predatory insect abundance and may improve agroecosystem resilience