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Wireworms are the subterranean larval stage of click beetles (Coleoptera: Elateridae), generalist herbivores that feed on a wide range of crops and cause serious damage. Wireworm management is challenging due to their long life cycle, subterranean habitat, wide range of host plants, and lack of effective chemical approach in most crops. Thus, developing an effective alternative control approach is an urgent need. The overall aim of this research study was to evaluate alternative [to insecticides] tactics that could be used as components of effective IPM strategies against wireworms. In Chapter 1 we provide an overview of wireworm's biology and management practices. In Chapter 2 we focused on biological control to evaluate the efficacy of entomopathogenic nematodes and fungi as single and mixed application against Limonius californicus and L. infuscatus in organic vegetable farms. The results showed the higher efficacy of commercial EPN, Steinernema feltiae, against Limonius species than entomopathogenic fungi. In Chapter 3, we focused on cultural control as a component of integrated pest management for wireworm control. We evaluated the efficacy of brown and yellow mustard and their byproducts, including defatted seed meal and seed meal concentrated extract against Limonius spp. Our finding indicated the effectiveness of brown mustard concentrated extract on reducing the wireworm population. However, it has a negative impact on the field-residing entomopathogenic nematodes. In Chapter 4, in the component of cultural control, we evaluated the wireworm, L. californicus, preference to three different crops, pea, bean, and wheat, for identifying an effective trap crop. We also quantified CO2 and organic volatile compounds released from each plant's root and evaluated the wireworm response toward the synthetic volatiles. Detailed results are provided in chapter 4. Finally, we looked at the novel new technology of RNA interference to find the effective target genes in wireworms for application as bioinsecticides and/or transgenic plants to control wireworms. In the concluding chapter, Chapter 6, we summarized our findings on the efficacy of different management tactics, biological, cultural, and molecular, to control Limonius species in cereals and organic production systems. / Doctor of Philosophy / Wireworms, the larval stage of click beetles (Coleoptera: Elateridae), are belowground pests that feed on a wide range of crops such as potatoes, cereals, vegetables, and sugar beet and can cause serious economic damage. Wireworm management is challenging because they live for long periods in the soil and feed on almost all crops. Moreover, most insecticides available for wireworm control, such as neonicotinoids, are not effective in reducing wireworm populations and just serve as feeding deterrents. Thus, developing an effective alternative control approach is an urgent need. The overall aim of this research study was to evaluate alternative [to insecticides] tactics that could be used as components of effective IPM strategies against wireworms. In Chapter 1 we provide an overview of wireworm's biology and management practices. In Chapter 2 we focused on biological control to evaluate the efficacy of entomopathogenic nematodes and fungi when applied individually and in combination with each other against sugar beet wireworm in organic vegetable farms. The results showed the commercial nematode, Steinernema feltiae was more effective in reducing sugar beet wireworm than fungi. In Chapter 3, we focused on cultural control as a component of integrated pest management for wireworm control. We evaluated the efficacy of brown and yellow mustard and their byproducts including defatted seed meal and seed meal concentrated extract against sugar beet wireworms in spring wheat. Our finding indicated the effectiveness of brown mustard concentrated extract in reducing the wireworm population. In Chapter 4, in the component of cultural control, we evaluated the sugar beet wireworm preference for three different crops, pea, bean, and wheat, for identifying an effective trap crop. We also quantified CO2 and organic volaille compounds released from each plant's root and evaluated the wireworm response toward the synthetic volatiles. Detailed results are provided in chapter 4. Finally, we looked at the novel new technology of RNA interference to find the effective target genes in wireworms for application as bioinsecticides and/or transgenic plants to control wireworms. In the concluding chapter, Chapter 6, we summarized our findings on the efficacy of different management tactics, biological, cultural, and molecular, to control wireworm species in cereals and organic production systems.

Thesis (PhD)--Stellenbosch University, 2011. / ENGLISH ABSTRACT: Codling moth, Cydia pomonella (L.) (Lepidoptera: Tortricidae) is a devastating pest of pome fruit in temperate regions of the world. Control of this pest, previously involved the extensive use of broad-spectrum insecticides. However, concerns over human safety, environmental impact, widespread dispersal of resistant populations of codling moth and the sustainability of synthetic pesticides in agroecosystems, has encouraged the development and use of alternative environmentally-friendly pest management technologies including the use of entomopathogenic nematodes. These nematodes are lethal pathogens of insects and belong to the families Steinernematidae and Heterorhabditidae, and are ideal candidates for incorporation into the integrated pest management programme currently being developed for residue-free pome fruit production in South Africa. The biological and ecological factors pertaining to the successful use of these nematodes for the control of codling moth were investigated in this study. Their use for bin-disinfestations was evaluated, focusing on the optimum handling conditions to ensure the survival and subsequent efficacy of the nematodes. The study proved that the local isolate SF41 of Heterorhabditis zealandica Poinar 1990 could be used for successful bin-disinfestation. The use of the same nematode isolate was also investigated for the disinfestation of mulch layers of diapausing codling moth larvae. An insect containment device which allowed for direct trial efficacy evaluation was identified and ecological factors pertaining to the successful use of nematodes for mulch disinfestation were investigated. The biological control potential of local nematode isolates, which had previously never been tested against codling moth larvae, was investigated in the laboratory under conditions as can be expected during orchard applications. The efficacy of the selected isolates was confirmed in field experiments. Innovative insect containment methods for above-ground trial efficacy evaluation in the field were investigated. Desiccation proved to be the most limiting factor to the survival and subsequent efficacy of the nematodes during field applications in temperate regions. The effect of low moisture levels on H. zealandica’s efficacy to control diapausing codling moth larvae was subsequently investigated and a starch-based formulation was further tested to overcome the issue of desiccation. Conclusive results indicated that there were several biological and ecological factors influencing the survival of nematodes and illustrated how these factors could be manipulated to overcome these issues and thereby ensure the efficacy of treatments. This is the first report of its kind to comprehensively investigate the use of South African entomopathogenic nematodes for the control of diapausing codling moth larvae and all results emanating from the study can be integrated into a framework for the commercial use of these nematodes in this regard in future. / AFRIKAANSE OPSOMMING: Kodlingmot, Cydia pomonella (L.) is ‘n ernstige sleutelplaag in appel- en peerboorde in gematigde klimaats gebiede wêreldwyd. In die verlede is hoofsaaklik breëspektrum insektedoders gebruik vir die beheer van hierdie plaaginsek. Maar, kommer oor veiligheid vir die mens, impak op die omgewing, verspreiding van weerstandbiedende populasies van kodlingmot en beperkte volhoubaarheid van sintetiese plaagdoders het die ontwikkeling en gebruik van alternatiewe plaagbeheer tegnologieë, insluitend die gebruik van entomopatogeniese nematodes, genoodsaak. Entomopatogeniese nematodese horende tot die families Steinernematidae en Heterorhabditidae, is ideale kandidate vir insluiting in die geïntegreerde plaagbestuur programme wat huidiglik ontwikkel word vir gebruik in plaaslike boorde met die uiteindelike doel om residu-vrye vrugte te produseer. In hierdie studie word die biologiese en ekologiese faktore bestudeer wat die sukses van ‘n nematode-toediening gemik op kodlingmot beïnvloed. Hierdie studie het bewys dat die lokale SF41 isolaat van Heterorhabditis zealandica Poinar 1990 gebruik kan word om vrugtekratte te disinfesteer van kodlingmot. Die gebruik van dieselfde isolaat vir die disinfestasie van deklae is ook ondersoek. ‘n Metode van insek-inkamping is ook ontwikkel wat die evaluering van toedienings vergemaklik en meer effektief maak. Die omgewings-toestande wat ook bydrae tot die oorlewing en gevolglike sukses van ‘n toediening is ook ondersoek. Die biologiese beheer potensiaal van ‘n paar lokale isolate wat nog nooit voorheen teen kodlingmot getoets is nie, is ook bestudeer. Die isolate se effektiwiteit is ook bevestig in veldproewe en insek-bekampings metodes wat meer van toepassing is vir bogrondse plaaginsekte is ook geïdentifiseer. Resultate dui daarop dat vogverlies en gevolglike uitdroging van nematodes die grootste beperkende faktor is vir hierdie tipe toedienings in gematigde gebiede en ‘n stysel-gebaseerde formulasie is dus ondersoek om hierdie probleem te oorkom. Die uiteindelike gevolgtrekking van die studie was, dat alhoewel daar verskeie biologiese en ekologiese faktore is wat die oorlewing van nematodes beperk, daar tog verskeie maniere is om hierdie faktore te manipuleer en sodoende te oorkom, wat bydrae tot die uiteindelike sukses van ‘n toediening. Hierdie is die eerste studie wat werklik die praktiese gebruik van lokale entomopatogeniese nematodes vir die beheer van kodlingmot ondersoek en alle bevindinge kan geïntegreer word in toekomende riglyne vir die kommeriële gebruik van nematodes vir die beheer van kodlingmot.

Thesis (MSc)--Stellenbosch University, 2013. / ENGLISH ABSTRACT: Planococcus ficus (Signoret) (Hemiptera: Pseudococcidae), the vine mealybug, is of economic importance to the wine and table grape (viticulture) industries, as it characteristically causes more damage than other mealybug species. Mealybug infestations contaminate grapes with their waxy secretions, egg-sacs and honeydew production, on which sooty mould grows, resulting in the fruit being unmarketable. Many export grapes are rejected, prior to shipment, as a result of infestations and phytosanitary concerns with regard to mealybug infestations. They are also vectors for various plant viruses. Up to date, the most common method of mealybug control in South Africa has been the use of chemical insecticides. Unfortunately, mealybugs are difficult to control chemically, due to their secretive/hidden lifestyle, where chemicals do not reach them. Of great concern is the ability of mealybugs to rapidly build up resistance to insecticides as well as the negative environmental effects associated with chemical pesticide use. Alternatively, entomopathogenic nematodes (EPNs), belonging to the families Heterorhabditidae and Steinernematidae, have been identified as lethal insect pathogens and their insecticidal action, towards a variety of insect pests, has proven them to be valuable and effective biocontrol agents. Laboratory bio-assays, to determine the ability of eight different EPN isolates to infect and kill P. ficus, were conducted. Six of the isolates were indigenous species and the other two, Heterorhabditis bacteriophora and Steinernema feltiae, were produced in Germany and are commercially available in South Africa. Planococcus ficus was highly susceptible to two indigenous species, Heterorhabditis zealandica and Steinernema yirgalemense; responsible for 96% ± 2% and 65% ± 10% mealybug mortalities, respectively. Biological studies illustrated that both H. zealandica and S. yirgalemense are able to complete their life cycles within adult female P. ficus. There was no significant difference in the pathogenicity of commercially produced H. bacteriophora, recycled through an insect host, and those from the formulated commercial product. However, commercially produced S. feltiae individuals, that were recycled through an insect host, were statistically more effective than those that were not. The LC50 and LC90 values for H. zealandica, in the current study, were 19 and 82 infective juveniles (IJs) respectively, which were similar to the LC50 and LC90 values for S. yirgalemense at 13 and 80, respectively. The LC50 and LC90, for commercially available H. bacteriophora, were greater than they were for both H. zealandica and S. yirgalemense, with values of 36 and 555, respectively. Such results indicate that there is a definite positive relationship which exists between the concentration of IJs of all three nematode species, used for inoculation, and the percentage mortality of P. ficus. Sand column tests resulted in S. yirgalemense outperforming H. zealandica significantly, with average mortalities of 95% ± 1.4% and 82% ± 4.1%, respectively. As a result S. yirgalemense was chosen for further studies in the field. IJs of commercially produced H. bacteriophora and S. feltiae were exposed to imidacloprid in laboratory bioassays to determine the effect on survival and infectivity. This study established the fact that these two EPN species can be applied, in combination with imidacloprid, in an integrated pest management scheme. Soil application field trials at Welgevallen and Nietvoorbij, using S. yirgalemense and mealybugs in Eppendorf tubes, buried 15 cm in the soil, resulted in 50% ± 10% and 52% ± 12% mealybug mortalities, respectively, when applying IJs at a concentration of 80 IJs/cm2. No significant difference was found between mealybug mortalities as a result of the three IJ concentrations applied (20, 40 and 80 IJs/cm2) for both vineyards. Persistence trials indicated that after four months post application, Cydia pomonella larval mortalities showed no significant reduction in infectivity on the Welgevallen vineyard, while on the Nietvoorbij vineyard there were no larval mortalities. Tests to establish whether or not S. yirgalemense and H. zealandica produced ant deterrent factors, showed no significant differences between the number of intact cadavers for both nematode species and for cadavers that were either four or six days old. There is, however, indication that deterrent factors may be in action in cadavers that were used six days after inoculation with 60% and 49% remaining intact for H. zealandica and S. yirgalemense infected cadavers respectively. All freeze killed cadavers were consumed by Linepithema humile (Mayr) (Argentine ant). The effects of low temperatures on EPN movement and infectivity were tested for H. zealandica and S. yirgalemense in the laboratory. The mortality of P. ficus at 14˚C, as opposed to 25˚C, for S. yirgalemense and H. zealandica were found to be 9.1% ± 2.6% and 2.5% ± 1.2% respectively. Vertical sand column tests were also conducted at 14˚C for S. yirgalemense and H. zealandica, which produced low mealybug mortalities of 3.5% ± 2.4% and 8.5% ± 1.4% respectively. This illustrates the low infectivity of the two local species at low temperatures. Laboratory persistence trials, conducted over a period of four months with S. yirgalemense, showed steady persistence of 100%, while H. zealandica had a statistically significant decrease of codling moth mortalities to 44% ± 5%. A three armed olfactometer was designed to establish if S. yirgalemense responds and moves towards chemical cues in the soil. A significant greater average number of IJs moved towards the grape vine roots (246 ± 0.124 IJs), than to the mealybugs (133 ± 0.168 IJs) and to the control (4 ± 1.02 IJs). This demonstrates that S. yirgalemense does actively seek out its hosts and that volatile cues produced by damaged grape vine roots, are more attractive to the EPN than cues produced by P. ficus. This study illustrates that S. yirgalemense has great potential as a biopesticide for controlling P. ficus in the soil of South African grape vineyards. Emphasis was placed on soil application of S. yirgalemense in the field, which produced good results, while laboratory tests indicate the potential for further aerial field application trials on grape vines. As the EPNs are not negatively affected by the agrochemical imidacloprid, the simultaneous use and combined action of both agents will potentially provide the farmer with excellent control against P. ficus. Further field- and aerial application studies will complement the current study and hopefully provide positive results for the efficient control of P. ficus found on grape vines. / AFRIKAANSE OPSOMMING: Planococcus ficus (Signoret) (Hemiptera: Pseudococcidae), die wingerd witluis, is van groot ekonomiese belang vir die wyn en tafeldruif industrieë, aangesien dit kenmerkend meer skade veroorsaak as enige ander witluis spesies. Witluis infestasies besmet druiwe met hulle wasagtige afskeidings, eierssakke en heuningdou produksie, waarop swamme groei, wat tot gevolg het dat die druiwe onbemarkbaar is. Baie besendings druiwe, bestem vir uitvoer, word afgekeur weens witluis besmettings en ook as gevolg van fitosanitêre oorwegings. Hulle tree ook op as vektore van verskeie plantvirusse. Die mees algemene manier waarmee witluis in Suid-Afrika beheer word, is chemiese behandeling. Ongelukkig is witluis baie moeilik om chemies te beheer vanweë hulle verskuilde lewenswyse wat dit moeilik maak vir chemikalieë om hulle te bereik. Die vermoë van witluis om vinnig weerstand op te bou teen insekdoders, asook die negatiewe effek van chemiese middels op die omgewing, is kommerwekkend. Alternatiewelik, kan entomopatogeniese nematodes (EPNs) van die families Heterorhabditidae en Steinernematidae gebruik word vir die beheer van witluis. Hierdie nematodes is geïdentifiseer is as dodelike insek patogene, vir ʼn groot verskeidenheid van pes insekte en daar is bewys dat hulle as waardevolle en effektiewe biologiese beheer agente kan optree. Laboratorium biotoetse is gedoen om die vermoë van agt EPN isolate te evalueer om P. ficus te beheer. Ses van die EPN isolate is inheems, terwyl die ander twee, Heterorhabditis bacteriophora en Steinernema feltiae, in Duitsland produseer is en kommersieel beskikbaar is in Suid-Afrika. Planococcus ficus is hoogs vatbaar vir die twee inheemse EPN spesies, naamlik Heterorhabditis zealandica en Steinernema yirgalemense en hulle is verantwoordelik vir 96% ± 2% en 65% ± 10% van witluis mortaliteit. Biologiese studies het aangetoon dat beide H. zealandica en S. yirgalemense in staat is om hul lewensiklus te voltooi in volwasse wyfies van P. ficus. Daar is geen beduidende verskil gevind in die patogenisiteit van die geformuleerde kommersiële produk H. bacteriophora en dié wat in vivo geproduseer is nie. Daar is egter in die geval van S. feltiae, gevind dat die nematodes, wat in insekte produseer is, statisties beduidend meer effektief was, as dié wat kommersieel beskikbaar was. Die LC50 en die LC90 waardes van H. zealandica, in die huidige studie, was 19 en 82 infektiewe larwes (IJs), wat baie naby die LC50 en LC90 waarders van S. yirgalemense van 13 en 80 was. Die LC50 en LC90 vir die kommersieel beskikbare H. bacteriophora was groter as vir beide H. zealandica en S. yirgalemense, met waardes van 36 en 555 onderskeidelik. Hierdie resultate dui daarop dat daar ʼn positiewe verwantskap bestaan tussen die konsentrasie van die IJs van drie EPN spesies en die persentasie mortaliteit van P. ficus. Sand kolomtoetse dui daarop dat S. yirgalemense baie beter vaar as H. zealandica met gemiddelde mortaliteite van 95% ± 1.4% en 82% ± 4.1% onderskeidelik. Op grond van hierdie resultate is S. yirgalemense gebruik vir verdere veld studies. IJs van kommersieel geproduseerde H. bacteriophora en S. feltiae is in laboratorium biotoetse blootgestel aan imidacloprid om die effek op die oorlewing en infektiewe vermoë vas te stel. Hierdie studie het aangetoon dat die twee EPN spesies aangewend kan word saam met imidacloprid in ʼn geïntegreerde plaagbestuur opset. Grond aanwendings is in veld proewe by Welgevallen en Nietvoorbij gedoen deur gebruik te maak van S. yirgalemense en P. ficus volwasse wyfies in Eppendorf buisies, 15 cm in die grond begrawe, het albei 50% ± 10% en 52% ± 10% witluis mortaliteit, respektiewelik, tot gevolg gehad, met die toediening van nematodes teen ʼn konsentrasie van 80 IJs/cm2. Geen beduidende verskille is gevind tussen die witluismortaliteit en die resultate van die verskillende EPN konsentrasies (20, 40 en 80 IJs/cm2) wat op beide wingerde toegedien is nie. Oorlewings toetse het aangedui dat, drie maande na toediening, met Cydia pomonella as indikator, geen beduidende verskille in die infeksie potensiaal van die Welgevallen wingerd to gevolg gehad het nie, terwyl daar op die Nietvoorbij wingerd geen verdere larvale mortaliteit gevind is was nie. Toetse om vas te stel of S. yirgalemense en H. zealandica afkrikmiddels vir miere in besmette insek kadawers produseer het aangetoon dat daar geen beduidende verskil is tussen die getal kadawers wat intakt is vir beide EPN spesies en kadawers wat vier of ses dae oud is nie. Daar is egter aangetoon dat die afskrikmiddels wel ses dae na infeksie deur insek kadawers afgeskei word; aangesien 60% en 49% van die oorblywende kadawers nog volledig was toe dit geïnfekteer was met H. zealandica en S. yirgalemense, onderskeidelik. Al die insek kadawers, wat deur bevriesing doodgemaak is, was deur Linepithema humile (Mayr) (Argentynse mier) verorber.

Les Nématodes Entomopathogènes (NEP) de la famille des Steinernematidae et Heterorhabditidae sont des parasites pour les insectes. Ils hébergent dans leur intestin un symbiote bactérien (genres Xenorhabdus et Photorhabdus) essentiel au succès parasitaire. Les NEP sont présents dans les sols des cinq continents de la planète. Leur diversité génétique et leurs propriétés biologiques constituent une intéressante ressource biologique. Grâce à leur entomotoxicité, les NEP sont de bons outils de lutte biologique en agriculture et en culture ornementale un peu partout dans le monde. La diversité et la biogéographie des NEP dans les cinq continents de la Terre (à l'exception de l'Antarctique) ont été étudiées partout dans le monde mais le Liban est parmi les rares pays du moyen orient où aucune prospection de ces nématodes n'a été réalisée alors que des NEP en Turquie, Syrie, Jordanie, Palestine et Egypte ont déjà été trouvés et caractérisés. L'objet de la thèse est de procéder à une étude biogéographique dans le but de connaître la diversité des NEP au Liban. L'enjeu scientifique est donc de combler un « vide » dans la connaissance de la répartition et de la biodiversité mondiale des NEP. Pour cela, un échantillonnage à l'échelle des étages de végétation est mené au Liban. Des échantillons de sol sont ainsi prélevés, mis en contact avec des larves de Galleria mellonella pour isoler les nématodes entomopathogènes et leurs symbiotes. Les nématodes et leurs symbiotes sont par la suite identifiés morphologiquement et moléculairement. Par la suite, une approche à l'échelle de l'habitat fait l'objet de cette thèse également pour étudier les interactions biotiques et abiotiques influençant la présence des nématodes entomopathogènes dans le sol. Les enjeux technologiques, exposés au second volet de la thèse, sont liés aux propriétés biologiques des nématodes et de leurs symbiotes afin de valoriser leur entomotoxicité en lutte biologique. Dans ce cadre, la sensibilité des Cephalcia tannourinensis, ravageur des cédraies au Liban, par rapport aux nématodes entomopathogènes est exploitée in vitro ; différentes espèces de nématodes entomopathogènes sont testées pour suivre leur cycle à l'intérieur des Cephalcia. / Entomopathogenic nematodes (EPNs) are parasites of soil-dwelling insects that occur in natural and agricultural soils around the world. Thanks to their entomotoxicity, EPNs are good tools for biological control in agriculture almost everywhere in the world. They are ubiquitous, having been isolated from every inhabited continent (except Antartica) from a wide range of ecologically diverse soil habitats including cultivated fields, forests, grasslands, deserts, and even ocean beaches. Biogeographic assessments of EPNs in the Eastern Mediterranean basin have been conducted in several countries such as Turkey, Syria, Jordan, Israel, Palestine and Egypt. Lebanon is among the few countries of the Middle East for which no survey of EPNs has been done. The scientific stake is thus to fill a gap in our knowledge of EPNs distribution in the Mediterranean basin. Survey of EPNs was conducted in this framework to cover the different vegetation levels defined in Lebanon. Soil samples were removed placed in contact with Galleria mellonella to isolate entomopathogenic nematode and their symbiotic bacteria. EPNs and their bacteria were then identified morphologically and molecularly. On the other hand, despite the different national surveys conducted on EPNs distribution around the world, habitat preferences remain inadequately known for entomopathogenic nematodes. Therefore, a comprehensive understanding of their distribution and the various biotic and abiotic factors influencing their presence is also a second object of our work. Beside a technological approach related to the biological properties of the nematodes and their symbiotics: valorisation of the entomotoxicity in biological control will be part of the third shutter of the thesis. In this framework, the sensibility of cedar pests, Cephalcia tannourinensis against entomopathogenic nematodes is exploited in vitro; different EPNs species were tested to study their life cycle inside Cephalcia larvae.

Bacteria of the genus Xenorhabdus live as the natural symbionts of the entomopathogenic nematodes of the family Steinernematidae. They produce a wide range of toxic secondary metabolites of different chemical structure and effect. The synthesis of particular products is specific for each strain of Xenorhabdus. The thesis is focused on evaluating the impact of bacterial metabolites on facultatively entomoparasitic nematodes Oscheius myriophila, the free-living nematode Caenorhabditis elegans and the phytophagous potato cyst nematode Globodera rostochiensis. Target species of nematodes were exposed to 37 strains of Xenorhabdus bacteria, isolated from various species of Steinernematidae. Testing the impact of bacterial metabolites on O. myriophila and C. elegans nematodes was performed both by direct cultivation of target species on solid medium with live bacterial cultures as well as in sterilized liquid bacteria cultures. The effect of toxic substances on G. rostochiensis was evaluated only in selected sterilized bacteria cultures.

The potential synergism between chosen species of entomopathogenic nematodes (Steinernema arenarium, Steinernema feltiae) and entomopathogenic fungi (Beauveria bassiana, Isaria fumosorosea, Lecanicillium lecanii) is investigated in this study. It is theoretically possible to expect increasing of their efficiency at the collective introduction into environment. Creating of uniform laboratory method which will be possible to use at the research of this interaction system is a part of this study.

Thesis (PhD)--Stellenbosch University, 2013. / ENGLISH ABSTRACT: Entomopathogenic nematodes have the potential to be outstanding biocontrol agents against agricultural pest insects. Combined with their bacterial symbionts, these biocontrol agents have proven to be very effective against numerous pests. The nematodes belong to the families Steinernematidae and Heterorhabditidae, and are ideal to be used in, and integrated with, pest management systems. There is a dire need for new and innovative methods to control agricultural pests, as numerous pest insects have developed resistance against broad-spectrum insecticides. Together with the environmental impact of these insecticides and the safety aspect regarding humans and animals, the need to develop new technologies, including entomopathogenic nematodes for pest management, is high. In this study, the associated symbiotic bacteria of three entomopathogenic nematodes species were isolated, and the potential of two nematode species to be successfully mass cultured in liquid medium was evaluated. Regarding the symbiotic bacteria, results from the study showed that bacteria species from all three nematode species, Heterorhabditis noenieputensis, Steinernema khoisanae and Heterorhabditis zealandica, were novel. Heterorhabditis noenieputensis was isolated in the Mpumalanga province during a previous survey conducted in citrus orchards. The bacterium isolated from this nematode belongs to the genus Photorhabdus, and bear closest similarity (98.6%) to the type strain of P. luminescens subsp laumondii (TT01T). Photorhabdus luminescens subsp. noenieputensis subsp. nov., derives its name from the area where the nematode was sourced, namely the farm Springbokvlei, near the settlement Noenieput close to the Namibian border. Thus far, 85 Steinernema spp. have been described worldwide, including S. khoisanae which was isolated in the Western Cape province of South Africa. Four S. khoisanae strains, namely SF87, SF80, SF362 and 106-C, were used for characterisating the new bacteria from different localities in South Africa. Using the neighbor-joining method, all the strains were aligned with 97% homology to the 16S rRNA sequences of several Xenorhabdus- type strains, indicating that they belonged to the same genus. The multigene approach was used to distinguish between the Xenorhabdus spp. and partial recA, dnaN, gltX, gyrB and infB gene sequences of the various strains were analysed. The bacterium species was named Xenorhabdus khoisanae sp. nov. after the nematode from which it was isolated. The results showed that the third bacterium species, which was isolated from H. zealandica, was new. The sequence of the bacteria strain clustered with the type strains of P. temperata and P. asymbiotica, indicate that it belonged to the genus Photorhabdus. This is the first study to show that H. zealandica associates with a luminescent Photorhabdus species, rather than with the known non-luminescent P. temperata. The potential of H. zealandica and Steinernema yirgalemense mass culture in liquid was investigated. Results illustrated that H. zealandica and its P. luminescens symbiont can be successfully cultured in liquid. However, two generations occurred during the process time, instead of the desirable one-generation. The growth curve of the symbiotic bacteria during the process time was measured, in order to determine when the stationary phase was reached, with the results showing this to occur after 36 h. Therefore, the optimum amount of time required for inoculating the IJs and for aiding in maximum infective juvenile (IJ) recovery is 36 h for adding the nematodes post pre-culturing of the bacteria. Future research goals should be to increase the percentage recovery in liquid culture, which would increase the number of nematodes produced per ml, which would, therefore, reduce the processing time significantly. The results from mass culturing the second nematode species, S. yirgalemense, indicated an asynchronous nematode development in the first generation. Growth curves were performed with the symbiotic bacteria that showed the exponential phase of Xenorhabdus started after 15 h, and that, after 42 h, the stationary phase was reached, with an average of 51 × 107 cfu·ml-1. Bioassays were performed to compare the virulence between in vitro- and in vivo-produced nematodes, with the results showing that the in vitro-produced nematodes were significantly less virulent than were the nematodes produced in vivo. The success obtained with the production of S. yirgalemense in liquid culture can serve as the first step in the optimising and upscaling of the commercial production of nematodes in industrial fermenters. The last aim of the current study was to determine when Xenorhabdus reached the stationary phase, when it is grown in a 20-L fermenter, as this would be the optimum time at which to add the IJs of S. yirgalemense. Such characteristics as the effect of stationary phase conditions on the bacterial cell density and on the DO2 rate in the fermenter were investigated. The results showed that the stationary phase of Xenorhabdus was reached after 36 h at 30˚C, which took 6 h less than did the same procedures followed with the Xenorhabdus sp. cultured in Erlenmeyer flasks on orbital shakers. This is the first step toward the liquid mass culturing of S. yirgalemense in industrial-size fermenters. Data from this study indicated the optimum amount of time that is required for adding nematodes to the bacterial culture in the fermenter, and for ensuring the optimum recovery of IJs, as well as a subsequent high yield of nematodes within a minimum processing time. This is the first report of its kind to investigate comprehensively the successful liquid culture of two South African entomopathogenic nematode species for the sole purpose of evaluating potential commercialisation. Results emanating from this study could be used as groundwork in future, in combination with similar research such as culturing nematodes intensively in large fermenters. / AFRIKAANSE OPSOMMING: Entomopatogeniese nematodes het die potensiaal om as doeltreffende biologiese beheeragente teen sleutelplaaginsekte gebruik te word. Elke nematood werk interaktief met ‘n spesifieke bakterium. Entomopatogeniese nematodes, behorende tot die families Steinernematidae en Heterorhabditidae, is ideale kandidate vir gebruik in ‘n geïntegreerde plaagbestuurprogram. Tans is daar ʼn behoefte vir nuwe metodes vir die beheer van plaaginsekte, omdat meeste insekte reeds weerstand opgebou het teen bestaande plaagdoders. As gevolg van die negatiewe impak van plaagdoders op die omgewing, asook kommer oor veiligheid vir die mens en diere, is die ontwikkeling en gebruik van alternatiewe plaagbeheermiddels noodsaaklik. In die eerste deel van die studie word drie nuwe bakterie spesies geïsoleer en beskryf. Resultate van hierdie studie het aangetoon dat die bakterië spesies vanuit die nematode spesies, Heterorhabditis noenieputensis, Steinernema khoisanae, en Heterorhabditis zealandica, tot dusver onbeskryf was. Eersgenoemde, H. noenieputensis, is afkomstig van ʼn sitrusboord in die Mpumalanga Provinsie. Die bakterie hieruit geïsoleer behoort tot die genus Photorhabdus en is biologies verwant (98.6%) aan P. luminescens subsp laumondii (TT01T). Die bakterie is benaam as Photorhabdus luminescens subsp. noenieputensis nov. en is na die nematood waaruit dit geïsoleer is vernoem. Tot dusver is wêreldwyd 82 spesies van Steinernema spp. beskryf, insluitende S. khoisanae van die Weskaap provinsie. Vier bakterie isolate is van S. khoisanae, SF87, SF80, SF362 en 106-C geïsoleer. Die buur-koppeling metode was gebruik om te bepaal dat hierdie bakterie isolate tot 97% ooreenstem met verskeie isolate van Xenorhabdus se 16S rRNA DNS volgordebepalings. Om tussen Xenorhabdus spp. te onderskei is ʼn multi-geen benadering gebruik deur gedeeltelike recA, dnaN, gltX, gyrB en infB DNS basispaar volgordebepalings van die verskeie isolate te bepaal. Hierdie bakterie isolaat is soortgelyk ook vernoem as, Xenorhabdus khoisanae sp. nov., na die nematood waaruit dit geïsoleer is. Die derde onbekende bakteriële spesie is uit H. zealandica geïsoleer. Die DNS basispaar volgordebepaling van die 16S geen van SF41 toon aan dat dit in dieselfde groep as P. temperata en P. asymbiotica val en sodoende aan die genus Photorhabdus behoort. Hierdie is die eerste studie met die bevinding dat H. zealandica ook met ʼn ander bakterie spesie geassosieer kan word buiten die normale P. temperata spesie. Die tweede deel van die studie gaan oor die teling van twee nematood spesies, H. zealandica en Steinernema yirgalemense, en hulle is geëvalueer vir hulle potensiaal om geteel te word in ʼn vloeibare medium. Die resultate het gewys dat H. zealandica met sy P. luminescens simbiont suksesvol in vloeistof aangeteel kan word, ten spyte van die feit dat daar twee generasies ontwikkel het, in plaas van die meer ideale enkel generasie. Die groeikurwe van die simbiotiese bakterie was gemonitor om te bepaal wanneer die stasionêre fase bereik word. Die resultate toon dat hierdie fase na 36 uur bereik was. Dus was die infektiewe nematode larwes eers na 36 uur tot die vloeibare medium waarin die bakterie geteel was bygevoeg. Navorsing in die toekoms moet dus gefokus wees om die persentasie herwinning van die infektiewe larwes te verhoog. Dit sal daartoe lei dat meer nematodes per ml geproduseer kan word en ook die prosesseringstyd van die nematodes verminder. ʼn Tweede nematode spesie, S. yirgalemense, was ook in vloeistof geteel. Hier het ʼn asinkroniese ontwikkeling in die eerste generasie plaasgevind wat problematies is. Groeikurwes is bepaal van die bakteriële simbiont en die resultate het gewys dat die groeifase van Xenorhabdus na 15 uur in aanvang geneem het en dat die stasionêre fase bereik was na 42 uur met ʼn gemiddelde van 51 × 107 selle·ml-1. Die virulensie van nematodes wat in vitro geteel is, is vergelyk met die virulensie van nematodes wat in vivo geteel is en die resultate het getoon dat die in vitro geteelde nematodes minder virulent was. Die teling van S. yirgalemense in vloeistof was oor die algemeen meer suksesvol as die teling van H. zealandica in dieselfde medium. Die doelwit van die laaste gedeelte van hierdie studie was om te bepaal wanneer Xenorhabdus die stasionêre fase bereik wanneer dit in ʼn 20-L fermenter gekweek word. Dit bepaal sodoende die optimale tyd wanneer die infektiewe larwes van S. yirgalemense bygevoeg behoort te word. Die uitwerking van die stasionêre fase op die bakteriële selle, asook die DO2-konsentrasie in die fermenter, was geëvalueer. Resultate het gewys dat die stasionêre fase van Xenorhabdus na 36 uur bereik was, wat 6 uur korter is as toe dit gekweek is in Erlenmeyer flesse. Hierdie studie is die eerste stap om die massa teling van S. yirgalemense in industriële fermenters suksesvol te bemeester. Die data wat verkry was, het aangedui wat die ideale tydsduur sal wees om die bakteriegetalle te vermeerder voordat die nematode bygevoeg word. Hierdie is die eerste studie wat die teling van twee Suid-Afrikaanse nematode spesies omvattend in vloeistof evalueer het. Die hoof doelwit is om die potensiaal van hierdie nematode spesies, met die oog op kommersiële gebruik, te meet. Die resultate van hierdie studie kan gekombineer word met toekomstige studies in hierdie spesifieke navorsingsveld.

Thesis (MScAgric)--Stellenbosch University, 2012. / ENGLISH ABSTRACT: Planococcus citri (Risso) (Hemiptera: Pseudococcidae), the citrus mealybug, is a highly destructive pest of citrus in South Africa. The ability of mealybugs to develop resistance to chemicals, as well as their cryptic nature and protective wax coverings, all individually and combined, impair the ability of insecticides to control them. Furthermore, chemicals deplete natural enemy populations that usually establish control over mealybug populations well before harvest in March to May. The inability of chemicals to control P. citri successfully and the growing public awareness of the detrimental environmental effects and health risks that are associated with pesticides are pressuring citrus growers to find alternative methods to chemical control. Entomopathogenic nematodes of the Rhabditida order, belonging either to the families Heterorhabditidae or Steinernematidae, have proven to be valuable biocontrol agents of a variety of insect pest species. These nematodes are, however, not yet commercially available in South Africa. Various laboratory bioassays were conducted to determine the potential of entomopathogenic nematodes to control P. citri. Adult female P. citri were screened for susceptibility to six indigenous nematode species. Planococcus citri was found to be most susceptible to Steinernema yirgalemense and Heterorhabditis zealandica, causing 97% and 91% mortality, respectively. Both H. zealandica and S. yirgalemense completed their life cycles inside adult female P. citri. Under optimal conditions, H. zealandica and S. yirgalemense, at an application rate of 170 infective juveniles (IJs) / insect (LD90), were able to control P. citri efficiently with a discriminating dosage of 11(LD50). The water activity (aw) bioassay indicated that S. yirgalemense was two times more tolerant to lower levels of free water than H. zealandica, with aw50 = 0.96 and aw90 = 0.99, compared to H. zealandica with aw50 = 0.98 and aw90 = 1.01. Furthermore, S. yirgalemense proved able to locate and infect P. citri at a faster rate than H. zealandica. Nematode activity was not significantly affected when exposed to 15°C, 20°C and 25°C. Results of the exposure trial also showed that the first 2−4 h post-application was the most decisive time for establishing successful infection of mealybugs. Further experiments were conducted to determine the ability of two polymer products, Zeba® and Xanthan gum, and a surfactant, Nu-Film-P®, to improve nematode application suspensions. Despite not being as effective as Xanthan gum, Zeba® still retarded sedimentation significantly. The addition of 0.02% Xanthan gum did not improve the ability of nematodes to control P. citri at 60% and 80% relative humidity containing H. zealandica and 0.03% Zeba® increased mortality by 14% at 60% RH and by 22% at 80% RH. The same polymer formulation was tested with S. yirgalemense, with mortality being found to have increased by 21% at 60% RH and by 27% at 80% RH. The combined addition of Nu-Film-P® and Zeba® was able to retard sedimentation significantly, increasing the average number of nematodes deposited on 2-cm2 leaf discs. To illustrate those factors that should be taken into consideration before applying nematodes in an integrated pest management (IPM) programme for citrus, the compatibility of H. zealandica and S. yirgalemense with biological control agents and agrochemicals to which they are most likely to be exposed was determined. Bioassays showed the coccinellid beetle, Cryptolaemus montrouzieri, to be susceptible to both nematode species. Beetle larvae proved to be highly susceptible, with H. zealandica obtaining 80%, and S. yirgalemense obtaining 92% control. Adult beetles were found to be twice as susceptible to S. yirgalemense, with 64% mortality recorded, as to H. zealandica, with 30% mortality recorded. Tolerance of H. zealandica and S. yirgalemense infective juveniles to the affect of aqueous solutions of an insecticide (Cyperphos 500 E.C.®), two biopesticides (CryptogranTM and HelicovirTM), and two adjuvants (Nu-Film-P® and Zeba®) on the infectivity and survival of H. zealandica and S. yirgalemense was evaluated. Heterorhabditis zealandica proved to be highly compatible with all products tested. Significant increase in mortality of S. yirgalemense was recorded after 12 h exposure to CryptogranTM, HelicovirTM and Cyperphos 500 E.C.®, and after 6 h exposure to Nu-Film-P®. However, there was no decrease in the ability of S. yirgalemense to infect the insect hosts. Towards selecting the appropriate nematode species for field studies, the ability of S. yirgalemense and H. zealandica to control P. citri was first evaluated under less harsh simulated glasshouse conditions. The ability of both the above-mentioned nematode species to control P. citri, with and without the addition of 0.03% Zeba® and 0.06% Nu-Film-P®, was evaluated in a growth chamber at 75 ± 8% relative humidity and 22°C. All treatments resulted in significantly higher mortality of adult female P. citri than did the control. The addition of 0.03% Zeba® and 0.06% Nu-Film-P® to an H. zealandica and an S. yirgalemense suspension increased resultant mortality from 26% to 30%, and from 34% to 45%, respectively. This increase in mortality was, however, not significant. The ability of this formulation to prolong the ability of S. yirgalemense to infect P. citri and to prolong nematode survival was also evaluated under the same conditions and showed the formulation to improve both infectivity and survival for up to 2–3 h post-application. In the semi-field trial, S. yirgalemense was unable to obtain significant control of P. citri without the addition of 0.03% Zeba® to nematode application suspensions. However, with the addition of 0.03% Zeba® S. yirgalemnse was able to obtain up to 53% control. The study established that the polymer product Zeba® improves the ability of S. yirgalemense to infect P. citri by retarding desiccation and by buffering nematodes from suboptimal environmental conditions. / AFRIKAANSE OPSOMMING: Planococcus citri (Risso) (Hemiptera: Pseudococcidae), die sitrus witluis, is ʼn baie skadelike pes van sitrus in Suid Afrika. Die vermoë van witluise om weerstand teen chemiese middels te ontwikkel, hul kriptiese lewenswyse en die beskermende waslaag wat hul liggame omhul inhibeer gesamentlik en individueel die vermoë van insektisiede om witluis bevolkings te beheer. Chemiese middels verminder ook die natuurlike vyande wat gewoonlik witluis bevolkings beheer voor die sitrus oestydperk in Maart tot Mei. Die onvermoë van chemiese middels om P. citri suksesvol te beheer en verhoogde bewustheid van die publiek rond om die vernietigende omgewings impak en gesondheidsrisiko’s verbonde aan chemiese insek beheer, noodsaak sitrus produsente om alternatiewe beheermetodes te ontwikkel. Entomopatogeniese nematodes, van die orde Rhabditida wat aan die families Heterorhabditidae of Steinernematidae behoort, is bekend as effektiewe biologiese beheeragente van ʼn verskeidenheid insek pes spesies. Hierdie nematodes is egter tans nie kommersieel beskikbaar in Suid- Afrika nie. Om ten einde die vermoë van plaaslike nematode spesies te bepaal om P. citri te beheer, is verskeie biotoetse in die laboratorium uitgevoer. ʼn Vinnige siftings proses is uitgevoer om vas te stel watter plaaslike nematode spesies die hoogste persentasie mortaliteit van P. citri wyfies veroorsaak. Daar is bevind dat P. citri die mees vatbaarste is vir Steinernema yirgalemense en Heterorhabditis zealandica wat 97% en 91% mortaliteit respektiewelik veroorsaak het. Die ontwikkeling van beide H. zealandica en S. yirgalemense na die infektering van volwasse P. citri wyfies is gevolg en daar is bevind dat beide nematode spesies hul lewensiklusse kon voltooi in insek kadawers. Onder optimale toestande was H. zealandica en S. yirgalemense in staat om P. citri effektief te beheer, as hul teen ʼn konsentrasie van 170 infektiewe larwes (JIs) per insek (LD90) toegedien word, met ʼn diskriminerende toedienings dosis van 11 (LD50). ʼn Water aktiwiteit biotoets het gewys dat S. yirgalemense twee keer so verdraagsaam is teenoor laer vlakke van vrye water as wat H. zealandica is met aw50 = 0.96 en aw90 = 0.99, in vergelyking met H. zealandica met aw50 = 0.98 en aw90 = 1.01. Resultate van ʼn blootstellings toets het ook gewys dat S. yirgalemense die vermoë het om P. citri vinniger op te spoor en te infekteer as H. zealandica. Die blootstellings toets was uitgevoer teen 15°C, 20°C en 25°C en daar is bevind dat die aktiwiteit van H. zealandica nie beduidend verhoog het met ʼn verhoging in temperatuur nie. Resultate vir die blootstellings toets het ook gewys dat die eerste 2 tot 4 uur na toediening van nematodes die mees kritieke tyd is om suksesvolle infektering van witluise te bevestig. Verdere eksperimente is uitgevoer om te bepaal of die byvoeging van twee polimeer produkte nl. Zeba® en Xanthan gum en ʼn benatter Nu-Film-P®, nematode toedienings suspensies kan verbeter. Deur 0.02% Xanthan gum by nematode toedienings suspensies te voeg is die vermoë van nematodes om P. citri by 60% en 80% relatiewe humiditeit (RH) te beheer nie beduidend verhoog nie, terwyl die byvoeging van 0.03% Zeba® wel ʼn beduidende verhoging in mortaliteit veroorsaak het. ʼn Suspensie van H. zealandica en 0.03% Zeba® het beheer met 14% by 60% RH en met 22% by 80% RH verhoog. Die invloed van dieselfde polimeer formulasie was ook getoets op S. yirgalemense en mortaliteit het verhoog met 21% by 60% RH en met 27% by 80% RH. Ten spyte daarvan dat “Xanthan gum” nie so effektief was om afsakking van nematodes te verhoed nie, het die byvoeginging van 0.03% Zeba® by toedienings suspensies steeds ʼn beduidende invloed gehad. Die gekombineerde byvoeging Nu-Film-P® en Zeba® was in staat om die gemiddelde aantal nematodes gedeponeer op 2-cm2 blaar skyfies te verhoog. Om te wys watter faktore in ag geneem moet word voor EPNs as deel van ʼn geïntegreerde pes beheer program toegedien word, is die verenigbaarheid van H. zealandica en S. yirgalemense vir biologiese beheer agente en landbouchemikalieë, waaraan hulle heel waarskynlik blootgestel gaan word, bepaal. Biotoetse het gewys dat die liewenheersbesie, Cryptolaemus montrouzieri, vatbaar is vir beide nematode spesies. Die larwale fase was hoogs vatbaar met mortaliteit van 80% en 92% verkry deur die toedeining van H. zealandica en S. yirgalemense respektiewelik. Daar is ook bepaal dat volwasse besies twee maal so vatbaar is vir S. yirgalemense wat 64% mortaliteit veroorsaak het, as vir H. zealandica met 30% mortaliteit aangeteken. Verdraagsaamheid van IJs van H. zealandica en S. yirgalemense met oplossings van ʼn insektisied (Cyperphos 500 E.C.®), twee bio-plaagdoders (CryptogranTM en HelicovirTM), en twee byvoegmiddels (Nu-Film-P® en Zeba®), vir infektiwiteit en oorlewing is bepaal. Resultate het gewys dat H. zealandica hoogs verenigbaar is met alle produkte wat getoets is. ʼn Beduidende verhoging in mortaliteit van S. yirgalemense is aangeteken na 12 ure se blootstelling aan CryptogranTM, HelicovirTM en Cyperphos 500 E.C.®, en na 6 ure se blootstelling aan Nu-Film-P®. Resultate het egter ook gewys dat die vermoë van S. yirgalemense om om gashere te infekteer nie deur beinvloed word nie. Om ten einde te bepaal watter een van H. zealandica of S. yirgalemense die beter isolaat sou wees om te gebruik in veldproewe, is hul vermoë om P. citri te beheer eers evalueer onder minder ongunstige gesimuleerde glashuis toestande. Die vermoë van beide nematode spesies om P. citri te beheer is bepaal in ʼn groeikamer by 75 ± 8% (RH) met en sonder die byvoeging van 0.03% Zeba® en 0.06% Nu-Film-P®. Mortaliteit verkry deur alle behandelings was beduidend hoër as die kontrole. Deur 0.03% Zeba® en 0.06% Nu-Film-P® by toedienings suspensies van H. zealandica en S. yirgalemense te voeg is die gevolglike mortaliteit van P. citri verhoog van 26% na 30% en van 34% na 45%, respektiewelik. Hierdie verhoging in mortaliteit was egter nie beduidend nie. Die vermoë van die bo-genoemde formulasie om oorlewing van S. yirgalemense en infeksie tydperk van P. citri te verleng was bepaal onder dieselfde toestande en daar is gevind dat die formulasie beide die infektiwiteit en oorlewing van nematodes met 2 tot 3 uur na toediening verleng het. Tydens die semi-veldproef was S. yirgalemense nie in staat om beduidend hoër mortaliteit van P. citri te verkry, in vergelyking met die kontrole, sonder die byvoeging van 0.03% Zeba® by toedienings suspensies nie. Met die byvoeging van Zeba® is daar tot 53% beheer verkry. Die studie het onweerlegbaar gewys dat die polimeer produk Zeba® die vermoë van S. yirgalemense om P. citri te infekteer verbeter deur die uitdroging van nematodes te vertraag en deur hul te beskerm teen ongunstige omgewings toestande.

Submitted by Renata Lopes (renatasil82@gmail.com) on 2016-10-20T13:47:58Z No. of bitstreams: 1 juliocesarrochacosta.pdf: 1044594 bytes, checksum: 36dcb55eeb5afb30dd503104cc262f69 (MD5) / Approved for entry into archive by Adriana Oliveira (adriana.oliveira@ufjf.edu.br) on 2016-10-25T12:27:57Z (GMT) No. of bitstreams: 1 juliocesarrochacosta.pdf: 1044594 bytes, checksum: 36dcb55eeb5afb30dd503104cc262f69 (MD5) / Made available in DSpace on 2016-10-25T12:27:57Z (GMT). No. of bitstreams: 1 juliocesarrochacosta.pdf: 1044594 bytes, checksum: 36dcb55eeb5afb30dd503104cc262f69 (MD5) Previous issue date: 2006-02-22 / CAPES - Coordenação de Aperfeiçoamento de Pessoal de Nível Superior / Aphitobius diaperinus (Panzer), conhecido como o cascudinho ou "lesser mealworm" coloniza o substrato encontrado em granjas, criando-se em altas populações, causando problemas sanitários e econômicos. Eles afetam a saúde e a desempenho das aves, sendo também transmissores de agentes de doenças. Seu controle baseia-se em aplicações de inseticidas, que aumentam os custos de produção e são prejudiciais aos animais. Assim sendo, as infectividades de Heterohabditis riobravus e de Steinernema carpocapsae diante dos estádios de larva, pupa e 20 adulto deste inseto para a primeira espécie de nematóide citada, e de larva e adulto para a segunda espécie, foram avaliadas em laboratório, utilizando os seguintes tipos de substratos: cepilho + ração, papel de filtro e terra. O objetivo foi determinar o grau de virulência dos juvenis infectivos (J Is) destes nematóides em cada tratamento. Os resultados indicaram que tanto H. riobravus, quanto S. carpocapsae apresentam potencial para infectar todos os estádios de desenvolvimento da espécie A. diaperinus analisados; e que os J Is de S. carpocapsae mostraramse mais eficientes contra larvas e adultos de A. diaperinus, do que os J Is de H. riobravus, em todos os tipos de substratos testados. Como já observado em outro estudo, os estágios imaturos se mostraram mais susceptíveis aos nematóides do que os adultos. Quando comparou - se os tratamentos com as duas espécies de nematóides sobre indivíduos adultos no substrato cepilho + ração, ambiente natural da espécie a ser controlada, verificou - se que as DL50 e DL90 de J Is de S. carpocapsae, corresponderam respectivamente a apenas 54,18% da DL50 e a 40,34% da DL90, encontradas para o tratamento envolvendo o nematóide da espécie H. riobravus. / Aphitobius diaperinus (Panzer), known as cascudinho or "lesser mealworm" colonizes the substrate found at farms, developing itselves in high populations, causing sanitary and economical problems. They affect the health and performance of the birds, being also transmitters of diseases. Their control is based on the application of insecticides, which increase the production costs and are harmful to the animals. Therefore, the infectivity of Heterohabditis riobravus and of Steinernema carpocapsae in front the life stages of larvae, pupae and adult of 21 this insect - for the first mentioned species of nematode - and of larva and adult for the second species, were evaluated in laboratory, using the following types of substrate: small plane + ration, filter paper and earth. The objective was to determine the degree of virulence of the J Is of these nematodes in each treatment. The results indicated that so much H. riobravus, as well as S. carpocapsae presented potential to infect all of the life stages of development of the species A. diaperinus analyzed; and that the infective juvenile of S. carpocapsae were shown more efficient against larvae and adults of A. diaperinus, that the infective juvenile of H. riobravus, in all of the types of tested substrate. As already observed in another study, the immature life stages were shown to be more susceptible to the nematodes than the adult life stage. When we compared the treatments with the two species of adult individuals constituted by the nematodes in the substrate of small plane + ration, natural environment for the species to be controlled, we verified that DL50 and DL90 of J Is of S. carpocapsae, corresponded respectively only 54.18% of DL50 and to 40.34% of DL90, found for the treatment involving the nematode of the species H. riobravus.