Testes em condições para o controle de Dysmicoccus texensis (Tinsley) (Hemiptera, Pseudococcidae) em cafeeiro com nematóides entomopatogênicos do gênero Heterorhabditis(Rhabditida, Heterorhabditidae)

Os nematóides entomopatogênicos (NEPs) apresentam potencial para o controle biológico de pragas e têm sido usados com sucesso, em vários países, no controle de pragas de solo e de ambientes crípticos, como a cochonilha-da-raiz-do-cafeeiro Dysmicoccus texensis (Tinsley). Testes de laboratório demonstram que estes agentes apresentam alta virulência sobre este inseto, no entanto, são necessários testes que avaliem a eficiência dos NEPs em condições de casa-de-vegetação e campo, sendo este o objetivo do presente trabalho. O experimento em condição de casa-de-vegetação para o controle da cochonilha foi realizado em vasos infestados, usando dois isolados do nematóide e dois métodos de aplicação (cadáver infectado e suspensão aquosa), conduzido em delineamento inteiramente casualisado com cinco repetições. O experimento em condições de campo foi conduzido em blocos casualisados (6 blocos), para avaliar a eficiência de dois isolados heterorhabditídeos no controle da cochonilha-da-raiz-do-cafeeiro. Os resultados mostraram que, em casa-de-vegetação, o método de suspensão aquosa apresentou melhores resultados para os dois isolados, sendo que JPM3 aplicado em suspensão aquosa foi o melhor tratamento, apresentando eficiência de controle de 70%. No experimento de campo, apenas o tratamento com o inseticida Actara 250 WG (thiamethoxam), usado como padrão de comparação, e JPM3, aplicado em suspensão aquosa, diferiram da testemunha, apresentando 81 e 65% de eficiência de controle, respectivamente.

Application technology for the entomopathogenic nematodes Heterorhabditis indica and Steinernema sp. (Rhabditida: Heterorhabditidae and Steinernematidae) to control Spodoptera frugiperda (Smith) (Lepidoptera: Noctuidae) in corn

O efeito de diversas tecnologias de aplicação foi avaliado sobre a concentração, viabilidade e eficácia dos juvenis infectantes dos nematóides Heterorhabditis indica Poinar, Karunakar & David (IBCB-n5) e Steinernema sp. (IBCB-n6) no controle da lagarta-do-cartucho Spodoptera frugiperda Smith na cultura do milho. Para o controle da lagarta-do-cartucho no terceiro estádio em placas de Petri foram necessários 280 juvenis infectantes de Steinernema sp., enquanto que 400 juvenis infectantes de H. indica controlaram apenas 75% das lagartas. Podem-se pulverizar os entomopatógenos, sem que haja perda significativa na sua concentração e viabilidade, com equipamentos que forneçam carga elétrica à calda, ponta centrífuga e pontas hidráulicas. Entretanto, o emprego de pulverizadores com pontas que requerem elementos filtrantes com malha igual a 100 resultou em decréscimo na concentração de juvenis infectantes de H. indica e Steinernema sp., de 28% e 53%, respectivamente. Os tensoativos organosiliconado e etoxilados não afetaram a viabilidade dos juvenis infectantes de Steinernema sp. Nos experimentos de pulverização em plantas de milho (V6) com Steinernema sp., doses equivalentes a até 288 milhões de juvenis infectantes por hectare, diluídos em volume de calda de até 800 L ha-1 com 0,01 % do tensoativo etoxilado, ou nesse volume seguido de exposição a chuva artificial (lâmina de água de 6 mm), não foram suficientes para o controle de S. frugiperda em casa-de-vegetação.

Movement of Heterorhabditis amazonensis and Steinernema arenarium in search of corn fall armyworm larvae in artificial conditions

Spodoptera frugiperda (Smith, 1797) (Lepidoptera: Noctuidae) is considered to be the main pest of maize crops in Brazil. Entomopathogenic nematodes (EPN) may be used to control this pest and exhibit different, unique abilities to search for their hosts. The movement of EPN in relation to S. frugiperda was evaluated. To test for horizontal movement, a styrofoam enclosure filled with sand was divided into segments, nematodes were placed at the entrance to the enclosure and a larva was placed at the end of each division. The same approach was used to evaluate vertical movement; however, PVC pipes were used in this case. In general, the mortality was inversely proportional to the initial distance between host and nematodes. In the vertical displacement test, both nematodes were able to kill the larvae up to a distance of 25 cm. Therefore, the infective juveniles of H. amazonensis and S. arenarium can search out, infect and kill larvae of S. frugiperda at distances of up to 60 cm and 25 cm of horizontal and vertical displacement, respectively.

Heterorhabditis amazonensis RSC5 (Rhabditida: Heterorhabditidae) movement and host recognition

Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)

Movement of Heterorhabditis amazonensis and Steinernema arenarium in search of corn fall armyworm larvae in artificial conditions

Spodoptera frugiperda (Smith, 1797) (Lepidoptera: Noctuidae) is considered to be the main pest of maize crops in Brazil. Entomopathogenic nematodes (EPN) may be used to control this pest and exhibit different, unique abilities to search for their hosts. The movement of EPN in relation to S. frugiperda was evaluated. To test for horizontal movement, a styrofoam enclosure filled with sand was divided into segments, nematodes were placed at the entrance to the enclosure and a larva was placed at the end of each division. The same approach was used to evaluate vertical movement; however, PVC pipes were used in this case. In general, the mortality was inversely proportional to the initial distance between host and nematodes. In the vertical displacement test, both nematodes were able to kill the larvae up to a distance of 25 cm. Therefore, the infective juveniles of H. amazonensis and S. arenarium can search out, infect and kill larvae of S. frugiperda at distances of up to 60 cm and 25 cm of horizontal and vertical displacement, respectively.

Effects of Galleria mellonella cadavers infected with Heterorhabditis bacteriophora and Steinernema rarum, their macerates and dead infective juveniles on Meloidogyne javanica suppression

Nematodes of the Meloidogyne genus affect to most of crops of an economic importance in Argentina. Researches related to new control strategies are needed to reduce the damage produced by these organisms. The objective of this work was to compare the effects of Galleria mellonella cadavers infected with the Argentine isolates Heterorhabditis bacteriophora Rama Caída and Steinernema rarum NOE, cadaver macerates and dead infective juveniles (IJs) on M. javanica suppression. Experiments were performed using 24-well plates and pepper plants grown in a growth chamber. The entomopathogenic nematodes-infected G. mellonella cadavers, their cadaver macerates and dead IJs were effective in suppressing M. javanica second-stage juveniles under laboratory conditions. The use of H. bacteriophora-infected cadavers caused a significant decrease in the number of galls and egg masses on pepper plants parasitized by M. javanica, in a growth-chamber.

Opções de manejo de Neoleucinodes elegantalis (Guenée), (Lepidoptera: Crambidae) com bases biológicas e controle mecânico, biológicos e extratos de plantas

A broca-pequena-do-fruto, Neoleucinodes elegantalis (Guenée) (Lepidoptera: Crambidae) é uma praga de grande importância na cultura do tomateiro. Desta forma, métodos de manejo que auxiliem no controle dessa praga e que reduzam a aplicação de agrotóxicos devem ser estudados. O objetivo do presente trabalho foi avaliar métodos alternativos de manejo da broca-pequena-do-fruto, como: o controle biológico, através da utilização de parasitoides do gênero Trichogramma e nematoides entompatogênicos (Heterorhabditis indica e Sterneneima carpocapsae); o controle físico, por meio do ensacamento de cachos de tomate com o tecido-não-tecido (TNT); e o controle químico através do estudo da atividade inseticida de plantas como a pimenta, o fumo, o alho e a mamona nas fases embrionárias, larval, pupal e adulta. Desta forma, por meio das análises foi possível verificar que a espécie e/ou linhagem de Trichogramma que mais se destacou foi T. galloi (Tg1) com características biológicas favoráveis ao manejo de N. elegantalis. Para os nematoides entomopatogênicos, S. carpocapsae, foi o mais efetivo, causando uma mortalidade de 82,93% a uma concentração de 65 juvenis infectivos por pré-pupa da broca-pequena-do-fruto. Através do ensacamento dos frutos, foi possível verificar a redução na oviposição da praga em frutos do tomateiro para quase zero com sacolas de TNT com fundo fechado, não ocorrendo alteração no peso, pH e graus brix do fruto, enquanto que no controle químico através de inseticidas botânicos foi possível verificar o destaque do extrato aquoso de fumo, reduzindo a oviposição, entrada e saída das lagartas nos frutos; causando mortalidade na fase embrionária, lagarta, pré-pupa e pupa da broca-pequena-do-fruto.

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.

Molecular and morphological characterization of heterorhabditid entomopathogenic nematodes from the tropical rainforest in Brazil

Despite massive losses of primary forest, the Amazonian rainforest remains an extremely rich source of biodiversity. In recent years, entomopathogenic nematodes (EPNs) have been isolated from soil in various parts of the world and used successfully as biological control agents against numerous insect pests. Therefore, a sampling in the rainforest of Monte Negro, Rondônia, Brazil was conducted with the aim of discovering new strains and/or species of EPNs for future development as biological control agents. From 156 soil samples taken at nine collecting sites, 19 isolates were obtained, all of them belonging to the genus Heterorhabditis. Four strains were subjected to detailed morphological and molecular evaluation. Based on morphometrics and internal transcribed spacer (ITS) sequence data, the strains LPP1, LPP2 and LPP4 were identified as Heterorhabditis indica, whereas LPP7 was considered Heterorhabditis baujardi. Comparative analysis of the ITS1 sequence of H. indica and H. baujardi isolates showed a polymorphic site for the restriction enzyme Tth 111 that could be used to distinguish the two species. Consequently, strains LPP1, LPP2, LPP3, LPP4, and LPP9 were identified as H. indica, whereas LPP5, LPP7, LPP8 and LPP10 were identified as H. baujardi.

Direct and indirect root defences of milkweed (Asclepias syriaca): trophic cascades, trade-offs and novel methods for studying subterranean herbivory

P>1. Entomopathogenic nematodes can function as indirect defence for plants that are attacked by root herbivores. By releasing volatile organic compounds (VOCs), plants signal the presence of host insects and thereby attract nematodes.2. Nonetheless, how roots deploy indirect defences, how indirect defences relate to direct defences, and the ecological consequences of root defence allocation for herbivores and plant biomass are essentially unknown.3. We investigate a natural below-ground tritrophic system, involving common milkweed, a specialist root-boring beetle and entomopathogenic nematodes, and asked whether there is a negative genetic correlation between direct defences (root cardenolides) and indirect defences (emission of volatiles in the roots and nematode attraction), and between constitutive and inducible defences.4. Volatiles of roots were analysed using two distinct sampling methods. First, we collected emissions from living Asclepias syriaca roots by dynamic headspace sampling. This method showed that attacked A. syriaca plants emit five times higher levels of volatiles than control plants. Secondly, we used a solid phase micro-extraction (SPME) method to sample the full pool of volatiles in roots for genetic correlations of volatile biosynthesis.5. Field experiments showed that entomopathogenic nematodes prevent the loss of biomass to root herbivory. Additionally, suppression of root herbivores was mediated directly by cardenolides and indirectly by the attraction of nematodes. Genetic families of plants with high cardenolides benefited less from nematodes compared to low-cardenolide families, suggesting that direct and indirect defences may be redundant. Although constitutive and induced root defences traded off within each strategy (for both direct and indirect defence, cardenolides and VOCs, respectively), we found no trade-off between the two strategies.6. Synthesis. Constitutive expression and inducibility of defences may trade off because of resource limitation or because they are redundant. Direct and indirect defences do not trade off, likely because they may not share a limiting resource and because independently they may promote defence across the patchiness of herbivore attack and nematode presence in the field. Indeed, some redundancy in strategies may be necessary to increase effective defence, but for each strategy, an economy of deployment reduces overall costs.

Influence of temperature on the production and infectivity of entomopathogenic nematodes against larvae and pupae of vine weevil, Otiorhynchus sulcatus, (Coleoptera: Curculionidae)

Susceptibility of late instar vine weevil Otiorhynchus sulcatus larvae and pupae to four species entomopathogenic nematodes were tested. Bioassays on production and infectivity to larvae and pupae were compared for two steinernematids and two heterorhabditis such as Steinernema carpocapsae, S. feltiae, Heterorhabditis indica and H. bacteriophora. Nematodes production of all species was determined by the number infective juveniles (IJs) established in vine weevil larvae and pupae O. sulcatus using sand and filter paper bioassay. S. feltiae produced the maximum number in larvae and pupae at 20°C as compared to other nematodes but production of H. indica, was better at 25°C in larvae and pupae followed by H. bacteriophora, S. carpocapsae and Infectivity test of larvae and pupae was also done in sand media. Infective juveniles recovered from larvae and pupae when infected with S. feltiae produced maximum infective juveniles at 20°C temperatures than all other isolates. H. bacteriophora produced higher number of IJs in larvae and pupae than all other nematode isolates at 25°C. This paper indicates the application of nematodes with the knowledge of insect pest biology represents a possible new strategy for O. sulcatus larvae and pupae.

A comparative study on the effectiveness of laboratory bioassays of entomopathogenic nematodes against desert locust nymphs Schistocerca gregaria (Acrididae:Orthoptera)

Entomopathogenic nematodes, Steinernema carpocapsae, S. feltiae (Steinernematids) Heterorhabditis indica and H. bacteriophora (Heterorhabditids) were studied to control nymphs of desert locust Schistocerca gregaria. Results of all experiments showed a significant difference in mortality percentage among all isolates. All nematodes were found more effective when exposure time was increased up to 10 days. On the other hand, both Heterorhabditids caused maximum mortality as compared to Steinernematids at 30 degree C. When different moisture levels were tested in the sand arena, a medium level of moisture (1%) caused maximum insect mortality in all isolates. However, highest concentration of each isolate (200 IJs per ml) proved to be most appropriate for maximum insect death. Similarly, both Heterorhabditis nematodes when orally applied to insects killed maximum nymphs as compared to other two Steinernematids. A similar response was observed in infectivity test when maximum percentage of IJs of both isolates of Heterorhabditis successfully penetrated into the body of locust nymphs. This research suggests some useful basic findings in developing biocides with suitable virulent of entomopathogenic nematode for controlling nymphs of desert locust.

Facultative scavenging as a survival strategy of entomopathogenic nematodes

Entomopathogenic nematodes cannot be considered only as parasitic organisms. With dead Galleria mellonella larvae, we demonstrated that these nematodes use scavenging as an alternative survival strategy. We consider scavenging as the ability of entomopathogenic nematodes to penetrate, develop and produce offspring in insects which have been killed by causes other than the nematode-bacteria complex. Six Steinernema and two Heterorhabditis species scavenged but there were differences among them in terms of frequency of colonisation and in the time after death of G. mellonella larvae that cadavers were penetrated. The extremes of this behaviour were represented by Steinernema glaseri which was able to colonise cadavers which had been freeze-killed 240 h earlier and Heterorhabditis indica which only colonised cadavers which had been killed up to 72 h earlier. Also, using an olfactometer, we demonstrated that entomopathogenic nematodes were attracted to G. mellonella cadavers. (c) 2007 Australian Society for Parasitology Inc. Published by Elsevier Ltd. All rights reserved.

Scavenging or infection? Possible host choosing by entomopathogenic nematodes

Entomopathogenic nematodes are able to survive by scavenging. We tested Steinernema feltiae, S. affine and Heterorhabditis megidis alone or in different combinations to evaluate the responses of these nematodes when dead or live Galleria mellonella larvae were offered. Steinernema feltiae and S. affine scavenged upon dead G. mellonella larvae and about 30% more dead larvae were penetrated than live ones. By contrast, H. megidis penetrated more live larvae than dead ones. When the nematode species were combined, the results varied among the combinations, but the dead larvae were always used as a host. The behaviour of natural field populations of S. feltiae and S. affine was also compared. Steinernema feltiae showed no difference between scavenging and performing 'normal infections', whereas S. affine scavenged to a reduced amount (around 60% less); this difference could be related to the particular foraging strategy of these nematodes.

The lethal and sub-lethal consequences of entomopathogenic nematode infestation and exposure for adult pine weevils, Hylobius abietis (Coleoptera: Curculionidae)

Entomopathogenic nematodes (EPN) frequently kill their host within 1–2 days, and interest in EPN focuses mainly on their lethality. However, insects may take longer to die, or may fail to die despite being infected, but little is known about the effects of EPN infection on insects, other than death. Here we investigate both lethal and sub-lethal effects of infection by two EPN species, Steinernema carpocapsae and Heterorhabditis downesi, on adults of the large pine weevil, Hylobius abietis. Following 12 h nematode–weevil contact in peat, S. carpocapsae killed a significantly higher proportion of weevils (87–93%) than H. downesi (43–57%) at all concentrations tested. Less than 10% of weevils were dead within 2 days, and weevils continued to die for up to 10 days after exposure (LT50 of 3 days or more). In a separate experiment, live weevils dissected 6 days after a 24 h exposure to nematodes on filter paper harbored encapsulated and dead nematodes, showing that weevils could defend themselves against infection. Some live weevils also harbored live nematodes 6 days after they had been removed from the nematode infested medium. Feeding by weevils was not affected by infection with, or exposure to, either species of EPN. We discuss these results in relation to the use of EPN in biological control against H. abietis.