The development of the truffle beetle Leiodes cinnamomeus at low temperature, a determining factor for the susceptibility of adults and larvae to entomopathogenic nematodes

Altres ajuts: acords transformatius de la UABThe European truffle beetle Leiodes cinnamomeus (Panzer) (Coleoptera: Leiodidae) is the most important pest in black truffle (Tuber melanosporum) plantations. Adults and larvae feed on truffles during the cold months of autumn and winter, while during spring and summer larvae are in diapause. This study aims to test the susceptibility of L. cinnamomeus adults and larvae to different entomopathogenic nematode (EPN) species at various temperatures under laboratory conditions. Different populations of Steinernema carpocapsae and Steinernema feltiae were applied against adults and mycophagous larvae at 20 °C, 10 °C and 10-15 °C (10 °C during 18 h and 15 °C during 6 h a day), while Heterorhabditis bacteriophora was only applied against diapause larvae at 25 °C. S. carpocapsae caused 100% mortality of adults three days after application at 20 °C. At 10 °C, adults were not susceptible to any EPN species seven days after treatment, while at 10-15 °C S. carpocapsae was the most virulent species (76.6-96.6% mortality). In the case of larvae, all EPN species were infective at 20 °C (43.3-83.3% mortality), despite differences among some populations. At 10 °C, only two populations of S. feltiae caused higher mortality of larvae (50-53.3%) than control seven days after treatment. H. bacteriophora caused 100% mortality against diapause larvae five days after application at 25 °C. Soil temperature was measured in a truffle plantation for each hour every day from September 2021 to April 2022 at 20 cm depth. From September to mid-October it was registered a temperature above 15 °C for more than 20 h a day. Temperatures were generally below 10 °C from November to March. An appropriate timing of field applications should be considered due to the short periods of time when temperature is optimal for each EPN species tested

Immune Response of Drosophila suzukii Larvae to Infection with the Nematobacterial Complex Steinernema carpocapsae-Xenorhabdus nematophila

Entomopathogenic nematodes have been proposed as biological agents for the control of Drosophila suzukii, an invasive pest of small-stone and soft-skinned fruits. Larvae of the fly are susceptible to Steinernema carpocapsae infection but the reaction of immune defenses of the host are unknown. To determine the immune response, larvae were infected with S. carpocapsae and Xenorhabdus nematophila to evaluate the effector mechanisms of both humoral and cellular processes. The symbiont bacteria presented an inhibitory effect on the phenoloxidase cascade with a low level of melanization. Besides, X. nematophila activated the synthesis of putative antimicrobial peptides on the hemolymph of infected larvae. However, those peptides presented a lower antimicrobial activity compared to hemolymph from larvae infected with non-symbiont bacteria. Xenorhabdus nematophila avoided also the phagocytosis response of hemocytes. During in vitro and in vivo assays, S. carpocapsae was not encapsulated by cells, unless the cuticle was damaged with a lipase-treatment. Hemocyte counts confirmed differentiation of lamellocytes in the early phase of infection despite the unrecognition of the nematodes. Both X. nematophila and S. carpocapsae avoided the cellular defenses of D. suzukii larvae and depressed the humoral response. These results confirmed the potential of entomopathogenic nematodes to control D. suzukii

Control biològic del Rhynchophorus ferrugineus a partir de diferents soques de nematodes entomopatògens i la seva problemàtica a Catalunya

En el present treball s'ha avaluat el potencial dels nemàtodes entomopatògens per a controlar la plaga de R. ferrugineus. Per fer-ho, s'ha determinat la susceptibilitat d'aquesta a 4 espècies diferents de nemàtodes: Steinernema carpocasae (soca B14, IDEBIO, BIOVERD), Steinernema feltiae (soca D114), Steinernema sp. (D122) i Heterorhabditis bacteriophora (soca DG46). D'altra banda, s'ha determinat la predació de Steinernema carpocapsae per part de l'àcar Centroupeda almerodai (Acari: Acaridae) per comprovar si aquest pot influir negativament en l'efectivitat de S. carpocapsae com agent de control biològic. S'ha vist que el morrut de les palmeres és molt susceptible als nemàtodes entomopatògens en especial una soca comercial (S. carpocapsae), la qual produeix mortalitats del 91,67%. Hi ha evidències de que l'àcar C. almerodai depreda les formes infectives de S. carpocapsae encara que no és suficient important com perquè es vegi compromès l'efectivitat com a bioinsecticida. L'ús de nemàtodes entomopatògens com a control biològic és una alternativa viable als mètodes químics de eficàcia similar però menys respectuosos amb el medi ambient.In this work the potential of the entomopathogenic nemàtodes to control the pest of R. ferrugineus was evaluated. Four species of nemàtodes were used: Steinernema carpocasae (B14, IDEBIO, BIOVERD strain), Steinernema feltiae (D114 strain), Steinernema gliseri group (D122 strain) and Heterorhabditis bacteriophora (DG46 strain). On the other hand, the predation of Steinernema carpocapsae by the acar Centroupeda almerodai (Acari: Acaridae) was determined, to evaluate if it could negatively influence in the effectiveness of S. Carpocapsae as a biological control agent. We have seen that the red plam weevil is very susceptible to the entomopathogens nemàtodes, specialy the comercial stain IDEBIO (S. carpocapsae), that produce a mortality of 91,67%. There is an evidence that the acar C. almerodai depredate the infective forms of S. carpocapsae even if it's not enought to compromise the bioinsecticide effectiveness. The use of entomopathogens nemàtodes as a biological control is a viable alternative to the chemical methods that have a similar efficiency, but not so respectuous with the environment

Los Nematodos entomopatógenos (Rhabditida: Steinernematidae y Heterorhabditidae) para el control del gusano cabezudo, Capnodis tenebrionis (Coleoptera: Buprestidae)

Consultable des del TDXTítol obtingut de la portada digitalitzadaEl gusano cabezudo, Capnodis tenebrionis (L.) (Coleoptera: Buprestidae), es una importante plaga de cultivos de frutales de hueso y de pepita en zonas mediterráneas. Este trabajo engloba los estudios llevados a cabo para probar la eficacia de los nematodos entomopatógenos en el control de Capnodis tenebrionis. Se realizó el aislamiento de nematodos entomopatógenos de cultivos de frutales de hueso en dos regiones mediterráneas de la Península Ibérica. Se recogieron un total de 630 muestras de suelo (de 210 campos) en Cataluña y 90 muestras de suelo (de 30 campos) en la región de Murcia, resultando un 5,2% y un 20% de suelos positivos para la presencia de nematodos entomopatógenos. Se recogieron diez aislados de steinernemátidos y tres heterorhabdítidos utilizando el método de Galleria mellonella. Las especies aisladas fueron identificadas como Steinernema feltiae y Heterorhabditis bacteriophora. Se evaluó la eficacia de los nematodos entomopatógenos contra la larva neonata de Capnodis tenebrionis. Las cepas de nematodos utilizadas en este ensayo fueron dos Steinernema carpocapsae, una S. feltiae, una S. arenarium y una Heterorhabditis bacteriophora. Las larvas neonatas fueron expuestas a 3 y 48 IJs/cm2 en tubos de ensayo con arena estéril, resultando una mortalidad de 60-91% y 96-100%, respectivamente. A la concentración de 150 IJs/larva, todas las cepas de nematodos fueron muy virulentas. Sin embargo, en el ensayo a baja concentración (10 IJs/larva), S. arenarium resulto ser la cepa más virulenta. Se evaluó también el rango de penetración como un indicador de la capacidad de infección de los nematodos. El porcentaje más elevado fue obtenido por S. arenarium (36%), seguido por H. bacteriophora (30.6%), S. feltiae (23.1%), y S. carpocapsae (20.7%). Se evaluó también la virulencia de cepas contra larvas, pupas y adultos de C. tenebrionis. La mortalidad larvaria varió de 50 a 100%, la de pupas de 0 a 70% y la de adultos de 66.7% a 100%. Los resultados de este estudio sugieren que algunos de los aislados de los nematodos testados pueden ser utilizados para cotrolar C. tenebrionis. Se evaluó la tolerancia al calor, desecación e hipoxia, el efecto de la temperatura en la capacidad de infección y reproducción y la capacidad de migración vertical en columnas de arena de diferentes cepas de nematodos entomopatógenos. Los resultados mostraron diferencias entre especies y dentro de cada una de las especies. Se analizaron todos los factores importantes para la presencia de C. tenebrionis, destacando tres cepas de S. feltiae. Se estudió la susceptibilidad de las larvas de C. tenebrionis en el interior de plántulas. Los nematodos fueron capaces de localizar y matar las larvas de C. tenebrionis en el interior de las raíces. S. feltiae obtuvieron una eficacia de entre 79.68% y 88.24%, las cepas de H. bacteriophora strains entre 71.66 y 76.47%. S. carpocapsae (B14) y S. affine (Gspe3) causaron el menor control de C. tenebrionis larvae (62.03% y 34.76%, respectivamente). Se realizó un ensayo de campo con una cepa de S. feltiae aislada de una larva muerta de C. tenebrionis. Se aplicaron los nematodos por riego superficial e inyección. En ambos tratamientos se utilizó una dosis de 1 millón de IJ por arbol durante 4 u 8 semanas. Se evaluó número, estadio y localización de cada insecto en el tronco del árbol. Steinernema feltiae Bpa redujo la población de C. tenebrionis en ambos tratamientos, con un cotrol entre 88,3% y 97%. No se observaron diferencias significativas entre ambos tratamientos. La persistencia de los nematodos se observó hasta 6 semanas después de haber sido aplicados. Los resultados indican que S. feltiae Bpa es adecuada para el control de C. tenebrionis en cultivos de cerezos.The Mediterranean flat-headed rootborer, Capnodis tenebrionis (L.) (Coleoptera: Buprestidae), is an economically important pest of stone fruit and seed fruit in Mediterranean areas. This report summarizes the studies carried out to prove the efficacy of entomopathogenic nematodes to control C. tenebrionis: Entomopathogenic were isolated from stone-fruit orchards in two Mediterranean regions of Spain. A total of 630 soil samples from Catalonia and 90 soil samples from Murcia were evaluated resulting in 5.2% and 20% of the soils testing positive for nematodes, respectively. Ten steinernematid isolates and three heterorhabditid isolates were recovered using the Galleria mellonella baiting method. Based on morphometric data, molecular data and cross-breeding experiments the nematode species were identified as Steinernema feltiae and Heterorhabditis bacteriophora. The efficacy of five entomopathogenic nematode strains was tested against the neonate larvae of Capnodis tenebrionis. The nematode strains screened included two Steinernema carpocapsae, one S. feltiae, one S. arenarium and one Heterorhabditis bacteriophora. Exposure of neonate larvae to 3 and 48 IJs/cm2 resulted in mortality between 60-91% and 96-100%, respectively. At a concentration of 150 IJs/larva, all of the nematode strains were highly virulent. S. carpocapsae caused infection and mortality to larvae more quickly than the other strains. The penetration rate as an indicator of entomopathogenic nematode infection was also evaluated. The highest value was recorded for S. arenarium (36%), followed by H. bacteriophora (30.6%), S. feltiae (23.1%), and S. carpocapsae (20.7%). The virulence of 14 Steinernema feltiae isolates, one S. carpocapsae and three Heterorhabditis bacteriophora isolates was compared in the laboratory against larvae, pupae and adults of Capnodis tenebrionis. Larval mortality ranged from 50 to 100% and pupae mortality from 0 to 70% and adults from 66.7% to 100%. Significant differences in mortality between C. tenebrionis males and females were observed. The results of this study suggest that some of the isolate strains have potential for improved biological control of C. tenebrionis. Environmental tolerance to heat, desiccation and hypoxia, the effect of temperature on infectivity and reproduction and nematode migration in sand columns were compared among isolates and one S. carpocapsae strain. Results showed differences among species and a great variability within species. Beneficial traits for each strain were added up to identify a superior candidate to control Mediterranean flat-headed rootborer, Capnodis tenebrionis. When all analyzed factors were considered, three S. feltiae isolates (Bpa, Sor and M116) obtained the best scores, and when hypoxia was removed, two of the strains (Bpa and Sor) continued ranking superior to other strains. The susceptibility of larvae of Capnodis tenebrionis to entomopathogenic nematodes was examined using GF-677@ potted trees (peach x almond hybrid) as the host plant. The nematode strains tested included nine Steinernema feltiae, one S. affine, one S. carpocapsae and two Heterorhabditis bacteriophora. Nematodes showed the ability to locate and kill larvae of C. tenebrionis just after they enter into the roots of the tree. S. feltiae strains provided an efficacy ranging from 79.68% to 88.24%. H. bacteriophora strains resulted in control of 71.66-76.47%. S. carpocapsae (B14) and S. affine (Gspe3) caused lower control of C. tenebrionis larvae (62.03% and 34.76%, respectively). The influence of foraging strategy and the use of native nematodes to control C. tenebrionis larvae inside the roots is discussed. The potential control of the entomopathogenic nematode Steinernema feltiae (Filipjev) (strain Bpa), isolated from a dead C. tenebrionis larva, was tested in a cherry tree orchard in Ullastrell, Barcelona (Spain). Nematode infective juveniles (IJs) were applied by drench and injection. In both the treatments, a rate of 1 million IJs was applied per tree every week during 4 or 8 weeks, with a total dose of 4 x 106 IJs/tree and 8 x 106 IJs/tree. Number, stage and localization of insects in each tree trunk were recorded. In both the experiments, S. feltiae significantly reduced the population of C. tenebrionis providing control ranging from 88.3% to 97%. No significant differences were recorded between the different treatments. Persistence of nematodes was recorded until 6 weeks after application. Results indicate that the application of S. feltiae (Bpa) provides adequate control of C. tenebrionis in cherry trees

Natural occurrence of entomopathogenic nematodes (Steinernema and Heterorhabditis) and Pristionchus nematodes in black truffle soils from Spain

Altres ajuts: acords transformatius de la UABThe European truffle beetle Leiodes cinnamomeus is the most important pest in black truffle (Tuber melanosporum) plantations. Current control methods against it are inefficient, so entomopathogenic nematodes (EPNs) could play an important role in their population regulation due to their efficacy against many soil-dwelling insect pests. A survey of EPNs and Pristionchus nematodes was conducted in truffle soils of Spain, considering environmental and physical-chemical soil factors. A total of 164 soil samples were collected from forests, productive plantations and null-low productive plantations, representing three distinct black truffle-growing habitat types. EPNs were isolated from seven soil samples (4.3%); four nematodes were identified as Steinernema feltiae and three as Heterorhabditis bacteriophora. Both species were sampled in three types of soil texture (loam, sandy loam or sandy clay loam), characterized by alkaline pH (7.5 to 8.5) and high organic matter (2.1-11.04%). The presence of these EPNs was influenced by habitat type and organic matter content. Pristionchus nematodes were isolated from truffle soil, around truffle fruit bodies and under the elytra of L. cinnamomeus, with Pristionchus maupasi being the most commonly identified species. No significant associations were found between environmental and soil factors and the occurrence of Pristionchus nematodes. These nematodes were found in alkaline soils (pH 7.75 to 8.7), across all seven sampled soil textures, with variable organic matter content (0.73%-5.92%). The ecological trends and the presence of Pristionchus may affect the occurrence of EPNs and their prospective use as biological control agents against L. cinnamomeus in black truffle plantations

Field efficacy against the hazelnut weevil, Curculio nucum and short-term persistence of entomopathogenic nematodes

The hazelnut weevil, Curculio nucum L. (Coleoptera: Curculionidae) is a pest affecting hazelnut orchards in Europe, with an important economical repercussion. Its potential control, short-term field persistence and the vertical distribution of native entomopathogenic nematode strains were tested in Muntanyes de Prades, Tarragona (NE Iberian Peninsula) over two consecutive years. Steinernema feltiae strain D114, Steinernema sp. strain D122 and Heterorhabditis bacteriophora strain DG46 were used in summer and spring applications at a dosage of 5 · 105 IJs m�2. The three nematode species reduced the hazelnut weevil population, ranging from 32% to 88% efficacy, without significant differences in efficacy or between the two applications. Persistence evaluation was carried out during 9 weeks for S. feltiae (D114), Steinernema sp. (D122) and H. bacteriophora (DG46) and showed all species capable of lasting for this period. Nematodes and larval vertical distribution was assessed. Most of the hazelnut weevil stayed within the first 25 cm although some were found as deep as 40 cm. Entomopathogenic nematodes were found along all 40 cm depth. This study proves the suitability of entomopathogenic nematodes to control the hazelnut weevil

Field efficacy against the hazelnut weevil, Curculio nucum and short-term persistence of entomopathogenic nematodes

The hazelnut weevil, Curculio nucum L. (Coleoptera: Curculionidae) is a pest affecting hazelnut orchards in Europe, with an important economical repercussion. Its potential control, short-term field persistence and the vertical distribution of native entomopathogenic nematode strains were tested in Muntanyes de Prades, Tarragona (NE Iberian Peninsula) over two consecutive years. Steinernema feltiae strain D114, Steinernema sp. strain D122 and Heterorhabditis bacteriophora strain DG46 were used in summer and spring applications at a dosage of 5 · 10 IJs m. The three nematode species reduced the hazelnut weevil population, ranging from 32% to 88% efficacy, without significant differences in efficacy or between the two applications. Persistence evaluation was carried out during 9 weeks for S. feltiae (D114), Steinernema sp. (D122) and H. bacteriophora (DG46) and showed all species capable of lasting for this period. Nematodes and larval vertical distribution was assessed. Most of the hazelnut weevil stayed within the first 25 cm although some were found as deep as 40 cm. Entomopathogenic nematodes were found along all 40 cm depth. This study proves the suitability of entomopathogenic nematodes to control the hazelnut weevil

Attraction of entomopathogenic nematodes to black truffle and its volatile organic compounds : A new approach for truffle beetle biocontrol

Altres ajuts: acords transformatius de la UABThe European truffle beetle, Leiodes cinnamomeus, is the most important pest in black truffle (Tuber melanosporum) plantations. Entomopathogenic nematodes (EPNs) are a promising biological control agents against L. cinnamomeus. EPNs may employ multiple sensory cues while seeking for hosts, such as volatile organic compounds (VOCs) and CO gradients. We report for the first time the attraction of EPNs to truffle fruitbodies, and identified some VOCs potentially playing a key role in this interaction. We conducted olfactometer assays to investigate the attraction behavior of Steinernema feltiae and Steinernema carpocapsae towards both T. melanosporum fruitbodies and larvae of L. cinnamomeus. Subsequently, a chemotaxis assay using agar plates was performed to determine which of the 14 of the main VOCs emitted by the fruitbodies attracted S. feltiae at low (0.1 %) and high (mg/100 g truffle) concentrations. Both EPN species were attracted to mature fruitbodies of T. melanosporum, which may enhance the likelihood of encountering L. cinnamomeus during field applications. L. cinnamomeus larvae in the presence of truffles did not significantly affect the behavior of EPNs 24 h after application, underscoring the importance of the chemical compounds emitted by truffles themselves. Chemotaxis assays showed that four long-chain alcohol compounds emitted by T. melanosporum fruitbodies attracted S. feltiae, especially at low concentration, providing a first hint in the chemical ecology of a little-studied ecological system of great economical value. Further studies should be conducted to gain a finer understanding of the tritrophic interactions between T. melanosporum, EPNs, and L. cinnamomeus, as this knowledge may have practical implications for the efficacy of EPNs in the biological control of this pest

Control biològic del Rhynchophorus ferrugineus a partir de diferents soques de nematodes entomopatògens i la seva problemàtica a Catalunya

En el present treball s'ha avaluat el potencial dels nemàtodes entomopatògens per a controlar la plaga de R. ferrugineus. Per fer-ho, s'ha determinat la susceptibilitat d'aquesta a 4 espècies diferents de nemàtodes: Steinernema carpocasae (soca B14, IDEBIO, BIOVERD), Steinernema feltiae (soca D114), Steinernema sp. (D122) i Heterorhabditis bacteriophora (soca DG46). D'altra banda, s'ha determinat la predació de Steinernema carpocapsae per part de l'àcar Centroupeda almerodai (Acari: Acaridae) per comprovar si aquest pot influir negativament en l'efectivitat de S. carpocapsae com agent de control biològic. S'ha vist que el morrut de les palmeres és molt susceptible als nemàtodes entomopatògens en especial una soca comercial (S. carpocapsae), la qual produeix mortalitats del 91,67%. Hi ha evidències de que l'àcar C. almerodai depreda les formes infectives de S. carpocapsae encara que no és suficient important com perquè es vegi compromès l'efectivitat com a bioinsecticida. L'ús de nemàtodes entomopatògens com a control biològic és una alternativa viable als mètodes químics de eficàcia similar però menys respectuosos amb el medi ambient.In this work the potential of the entomopathogenic nemàtodes to control the pest of R. ferrugineus was evaluated. Four species of nemàtodes were used: Steinernema carpocasae (B14, IDEBIO, BIOVERD strain), Steinernema feltiae (D114 strain), Steinernema gliseri group (D122 strain) and Heterorhabditis bacteriophora (DG46 strain). On the other hand, the predation of Steinernema carpocapsae by the acar Centroupeda almerodai (Acari: Acaridae) was determined, to evaluate if it could negatively influence in the effectiveness of S. Carpocapsae as a biological control agent. We have seen that the red plam weevil is very susceptible to the entomopathogens nemàtodes, specialy the comercial stain IDEBIO (S. carpocapsae), that produce a mortality of 91,67%. There is an evidence that the acar C. almerodai depredate the infective forms of S. carpocapsae even if it's not enought to compromise the bioinsecticide effectiveness. The use of entomopathogens nemàtodes as a biological control is a viable alternative to the chemical methods that have a similar efficiency, but not so respectuous with the environment

Compatibility of entomopathogenic nematodes and essential oils : A new step for integrated pest management of the truffle beetle

Altres ajuts: acords transformatius de la UABThe European truffle beetle, Leiodes cinnamomeus, is the most important pest in black truffle (Tuber melanosporum) plantations. Entomopathogenic nematodes (EPNs) are promising biological control agents against L. cinnamomeus. Essential oils (EOs) are also recently being investigated for the control of the adults of this pest. Therefore, both control methods could be combined in Integrated Pest Management (IPM) programs to enhance their efficacy. However, limited information exists regarding the effects of the EOs on EPNs and so their compatibility. The aims of our work were to study the effects of three previously described insecticidal and nematicidal essential oils, Allium sativum, Mentha suaveolens, and Satureja montana, on the survival, infectivity, reproduction, and attraction behaviour of three EPN species: Steinernema feltiae, Steinernema carpocapsae and Heterorhabditis bacteriophora. Therefore, we conducted three experiments under laboratory conditions to observe the lethal and sublethal effects by direct contact, the lethal effect by fumigation, and the chemotaxis response. Allium sativum caused the highest mortality rates in all three EPN species at 24 and 72 h post application whether by direct contact (97-99%) or fumigation (40-42%), and it also reduced their infective capacity on Galleria mellonella. Satureja montana EO caused low mortality rates compared to A. sativum, but it was significantly more lethal (6-8%) than the control (0-3%) at 72 h in the direct contact assay. It also displayed repellent properties against S. feltiae and H. bacteriohora in the chemotaxis assay. In contrast, M. suaveolens EO exhibited minimal impact on the survival, infectivity and reproduction of all three EPN species. Therefore, our results suggest M. suaveolens oil may be the most compatible EO for use integrated with EPNs. Further validation under field conditions and in the presence of L. cinnamomeus is necessary to confirm the practical applicability of these findings