False codling moth (FCM), Thaumatotibia leucotreta is an extremely important pest of citrus in South Africa and with the shift away from the use of chemicals, alternate control options are needed. One avenue of control which has only recently been investigated against the soil-borne life stages of FCM is the use of entomopathogenic fungi (EPF). In 2009, 12 entomopathogenic fungal isolates collected from South African citrus orchards showed good control potential during laboratory conducted bioassays. The aim of this study was to further analyse the potential of these isolates through concentration-dose and exposure-time response bioassays. After initial re-screening, concentration-dose response and exposure-time response sandconidial bioassays, three isolates were identified as exhibiting the greatest control potential against FCM in soil, Metarhizium anisopliae var. anisopliae (G 11 3 L6 and FCM Ar 23 B3) and Beauveria bassiana (G Ar 17 B3). Percentage mycosis was found to be directly related to fungal concentration as well as the amount of time FCM 5th instar larvae were exposed to the fungal conidia. LC50 values for the three isolates were not greater than 1.92 x 10⁶ conidia.ml⁻ₑ and at the LC₅₀, FCM 5th instar larvae would need to be exposed to the fungus for a maximum of 13 days to ensure a high mortality level. These isolates along with two commercially available EPF products were subjected to field persistence trials whereby net bags filled with a mixture of autoclaved sand and formulated fungal product were buried in an Eastern Cape citrus orchard. The viability of each isolate was measured on a monthly basis for a period of six months. All isolates were capable of persisting in the soil for six months with the collected isolates persisting far better than the commercially used isolates. Two of the isolates, G 11 3 L6 and G Ar 17 B3, were subjected to small scale laboratory application trials. Two formulations were investigated at two concentrations. For each isolate, each formulation and each concentration, FCM 5th instar larvae were applied and allowed to burrow into the soil to pupate before fungal application or after fungal application. Contact between fungi and FCM host is essential as, in contrast to pre-larval treatments, percentage mortality in post-larval treatments was low for both formulations and both isolates. For isolate G Ar 17 B3, a conidial suspension applied as a spray at a concentration of 1 x 10⁷ conidia.ml⁻ₑ obtained the highest percentage mortality (80 %). For isolate G 11 3 L6 however, both formulations performed equally well at a high, 1 x10⁷ conidia.ml⁻ₑ concentration (conidial suspension: 60 %; granular: 65 %) The results obtained thus far are promising for the control of FCM in citrus, but if these EPFs are to successfully integrate into current FCM control practices more research, some of which is discussed, is essential

The biology of key citrus pests Thaumatotibia leucotreta Meyrick (Lepidoptera: Tortricidae), Ceratitis capitata Wiedemann (Diptera: Tephritidae) and Ceratitis rosa Karsch (Diptera: Tephritidae) includes their dropping from host plants to pupate in the soil below citrus trees. Since most EP fungi are soil-borne microorganisms, the development and formulation of alternative control strategies using these fungi as subterranean control agents, targeted at larvae and pupae in the soil, can potentially benefit existing IPM management of citrus in South Africa. Thus, a survey of occurrence of entomopathogenic fungi was undertaken on soils from citrus orchards and natural vegetation (refugia) on conventionally and organically managed farms in the Eastern Cape Province in South Africa. A method for baiting soil samples with citrus pest T. leucotreta and C. capitata larvae, as well as with the standard bait insect, Galleria mellonella Linnaeus (Lepidoptera: Pyralidae), was implemented. Sixty-two potentially useful entomopathogenic fungal isolates belonging to four genera were collected from 288 soil samples, an occurrence frequency of 21.53%. The most frequently isolated entomopathogenic fungal species was Beauveria bassiana (Balsamo) Vuillemin (15.63%), followed by Metarhizium anisopliae var. anisopliae (Metschnikoff) Sorokin (3.82%). Galleria mellonella was the most effective insect used to isolate fungal species (χ2=40.13, df=2, P≤ 0.005), with a total of 45 isolates obtained, followed by C. capitata with 11 isolates, and T. leucotreta with six isolates recovered. There was a significantly (χ2=11.65, df=1, P≤ 0.005) higher occurrence of entomopathogenic fungi in soil samples taken from refugia compared to cultivated orchards of both organically and conventionally managed farms. No significant differences were observed in the recovery of fungal isolates when soil samples from both farming systems were compared. The physiological effects and host range of 21 indigenous fungal isolates obtained in the Eastern Cape were investigated in the laboratory to establish whether these isolates could be effectively used as biological control agents against the subterranean life stages of C. rosa, C. capitata and T. leucotreta. When these pests were treated with a fungal concentration of 1 x 10⁷ conidia ml⁻¹, the percentage of T. leucotreta adults which emerged in fungal treated sand ranged from 5 to 60% (F=33.295; df=21; P=0.0001) depending on fungal isolate and the percentage of pupae with visible signs of mycosis ranged from 21 to 93% (F= 96.436; df=21; P=0.0001). Based on fungal isolates, the percentage adult survival in C. rosa and C. capitata ranged from 30 to 90% and 55 to 86% respectively. The percentage of C. rosa and C. capitata puparia with visible signs of mycosis ranged from 1 to 14% and 1 to 11% respectively. Deferred mortality due to mycosis in C. rosa and C. capitata adult flies ranged from 1 to 58% and 1 to 33% respectively, depending on fungal isolate. Entomopathogenic fungal isolates had a significantly greater effect on the adults of C. rosa and C. capitata than they did on the puparia of these two fruit fly species. Further, C. rosa and C. capitata did not differ significantly in their response to entomopathogenic fungi when adult survival or adult and pupal mycosis were considered. The relative potency of the four most virulent Beauveria isolates as well as the commercially available Beauveria bassiana product, Bb Plus® (Biological Control Products, South Africa), were compared against one another as log-probit regressions of mortality against C. rosa, C. capitata and T. leucotreta which all exhibited a dose-dependent response. Against fruit flies the estimated LC50 values of all five Beauveria isolates ranged from 5.5 x 10¹¹ to 2.8 x 10¹² conidia/ml⁻¹. There were no significant differences between the relative potencies of these five fungal isolates. When T. leucotreta was considered, isolates: G Moss R10 and G 14 2 B5 and Bb Plus® were significantly more pathogenic than G B Ar 23 B3 and FCM 10 13 L1. The estimated LC₅₀ values of the three most pathogenic isolates ranged from 6.8 x 10⁵ to 2.1 x 10⁶ conidia/ml⁻¹, while those of the least pathogenic ranged from 1.6 x 10⁷ to 3.7 x 10⁷ conidia/ml⁻¹. Thaumatotibia leucotreta final instar larvae were exposed to two conidial concentrations, at four different exposure times (12, 48, 72 and 96 hrs) and showed an exposure time-dependant relationship (F=5.43; df=3; P=0.001). At 1 x 10⁷conidia/ml⁻¹ two Beauveria isolates: G Moss R10 and G 14 2 B5 were able to elicit a response in 50% of test insects at 72 hrs (3 days) exposure. Although a limited amount of mycosis was observed in the puparia of both fruit fly species, deferred adult mortality due to mycosis was high. The increased incidence of adult mortality suggests that post emergence mycosis in adult fruit flies may play a more significant role in field suppression than the control of fruit flies at the pupal stage. The increased incidence of pupal mortality, as well as the relatively low concentrations of conidia required to elicit meaningful responses in T. leucotreta pupae may suggest that pre-emergent control of false codling moth will play a more significant role in field suppression than the control of adult life stages using indigenous isolates of entomopathogenic fungi. Various entomopathogenic fungal application techniques targeted at key insect pests within integrated pest management (IPM) systems of citrus are discussed.

The diamondback moth, Plutella xylostella (Linnaeus 1758.) (Lepidoptera: Plutellidae), is a major insect pest of Brassica crops in many parts of the world leading to economic losses amounting to an estimated US$ 4-5 billion. Although parasitoids (biological control agents) play a major role in suppressing the pest populations during November – May in South Africa, the pest reaches outbreak status during September and October due to low impact of parasitoids, which has necessitated regular application of insecticides. However, insecticide applications have often resulted in the pest developing resistance, and when coupled with the negative effects of several insecticides on parasitoids, integration of the two pest control strategies for effective management of P. xylostella population density has been difficult to achieve. One approach that has received little attention is integration of host plant resistance (bottom-up effect) and biological control (top-down effect) for effective management of P. xylostella. However, the interaction between host plants, the insect pest, and parasitoids is not simple and straight forward, as strong negative impact of host plants on fitness of the insect pest can be cascaded up the food chain and have a negative impact on a given parasitoid, which in turn may reduce the desired complementary effect between the two pest control strategies. To identify optimal interactions between cabbage (Brassica oleracea L. var. capitata, Brassicaceae), P. xylostella and its larval parasitoid Cotesia vestalis (Haliday 1834) (Hymenoptera: Braconidae), this study investigated (i) the effects of seven cabbage cultivars (Empowa, Hollywood F1, Megaton, Leano, Menzania, Beverley Hills and Karabo) on fitness parameters (survival, developmental time, pupal weights, longevity without food and oviposition rates) of P. xylostella; (ii) the influence of the same host plant cultivars on fitness parameters (developmental time, pupal weights, longevity xi without food, fecundity, emergence rate and sex ratio) of C. vestalis. Furthermore, net reproductive rates and the intrinsic rates of natural increase were calculated for C. vestalis that emerged from hosts fed on each of the cultivars. All experiments were conducted in climate-controlled laboratory rooms maintained at 22 ± 1 ºC (mean ± S.D.), 60 ± 5 % RH and 16L: 8D photoperiod. Under the no choice test, overall survival of P. xylostella immature stages was highest on Karabo (67.26%) and lowest on Megaton (44.92%). The larval and pupal developmental period, and generation time was prolonged on Empowa (18.48 days), Karabo (14.64 days) and Beverly Hills (17.48 days), while developmental period on Hollywood F1 (13.79 days) was shortest. Male and female P. xylostella pupal weights were lighter from larvae that fed on Megaton (4.13 and 4.65 mg), Menzania (4.53 and 4.91 mg), and Hollywood F1 (4.11 and 5.08 mg), whereas pupal weights from Karabo (6.0 and 6.82 mg) were the heaviest. Unfed female moths originally reared on Beverley Hills had the highest longevity (5.05 days), whereas those reared on Leano (3.54 days) and Megaton (3.89 days) had the shortest life span. Under the choice-test, P. xylostella moth laid significantly more eggs on Empowa (48.8%) and Hollywood F1 (45.6%) and least on Menzania (11.8%) and Leano (10.6%). Megaton was more resistant to P. xylostella due to lower survival rates of immature stages, lower pupal weights and moth longevity. The generation time of C. vestalis was shortest on Karabo (10.10 days) and Leano (10.38 days), and longest on Megaton (12.57 days) and Empowa (12.80 days). The highest pupal weight of C. vestalis was obtained from parasitoids reared from P. xylostella fed Menzania (5.4 mg), Megaton (5.25 mg) and Beverly Hills (4.85 mg) and the lightest on Karabo (3.8 mg). Parasitoids reared on larvae that fed on Hollywood F1 lived the longest (2.28 days) followed by Menzania (1.94 days) and Beverly Hills (1.8 days), whereas those whose hosts fed on Leano had shortest life span (0.83 days). Despite the parasitoids from Megaton hosts being heavier, their fecundity and number of female progeny per female (16.87 and 3.60, respectively) were lowest. Cotesia vestalis fecundity and daughters produced per female were highest on hosts fed on Menzania (38.00 and 9.13, respectively) and Beverly Hills (32.87 and 9.07, respectively). As a consequence, the net reproductive rate (R0) and intrinsic rate of increase (r) were higher on Menzania (7.87 and 0.58, respectively) and Beverly Hills (8.29 and 0.62, respectively). As survival and overall fitness of P. xylostella was lower on Megaton, this cultivar can play a major role in restricting population growth of this pest and thus generational number of eggs deposited on it during September and October. However, this strong bottom-up effect of Megaton on P. xylostella was cascaded up the food chain, as overall fitness of C. vestalis was lower on hosts developing on it. In contrast, the overall fitness of C. vestalis was higher on hosts that developed on Menzania and Beverly Hills. As these cultivars showed potential to sustain population density of C. vestalis at higher levels, it is also assumed that the period required for the parasitoid to reach the critical density to suppress the host population at a lower average density will be reached quicker than on other cultivars. Thus, their cultivation may improve biological control of P. xylostella during November–May in South Africa / Agriculture, Animal Health and Human Ecology / M. Sc. (Agriculture)

Gossypium hirsutum, le coton Upland, représente plus de 95% de la fibre de coton produite annuellement dans le monde et est cultivé dans environ 40 pays. La protection des champs de coton contre l’attaque des arthropodes herbivores nécessite des quantités importantes d’insecticides de synthèse, environ 18% de la consommation mondiale en 2000, bien que cela ait beaucoup diminué grâce au coton Bt et aux programmes d’éradication de certains ravageurs. Les composés organiques volatils (COV) naturellement émis par les plantes cultivées peuvent réduire les attaques d'insectes, les COV ayant une influence sur le comportement des arthropodes herbivores et des arthropodes auxiliaires. La recherche scientifique sur les COV des plantes a beaucoup augmenté depuis deux ou trois décennies. La stimulation des émissions de COV dans les champs de coton est désormais recommandée par les entomologistes du Cirad en tant que composante de la stratégie de protection. L’écimage (topping en anglais), c'est-à-dire couper la tête des cotonniers au cours de la floraison, stimule l'émission de COV de défense, une protection écologique qui limite les risques pour la santé dus aux insecticides synthétiques, pour les agriculteurs et les personnes des environs. L'objectif de la thèse était d'améliorer nos connaissances sur les bases génétiques des émissions de COV après l’écimage. La combinaison de plusieurs disciplines telles que la bio-informatique, l'écologie chimique et la génétique moléculaire nous a permis de: 1) analyser les séquences génomiques des gènes des voies de biosynthèse des COVs terpènes et terpénoïdes et des facteurs de transcription (TF) liés à la réponse au stress, à l'aide des bases de données publiées sur les génomes de trois espèces de coton, G. raimondii, G. arboreum (cotons diploïdes) et G. hirsutum (coton tétraploïde), 2) étudier les émissions de COV par les feuilles de coton des plants de G. hirsutum en réponse à l’écimage, en capturant ces molécules en serre et en caractérisant leurs profils cinétiques par chromatographie en phase gazeuse-spectrométrie de masse (GC-MS), incluant des mesures quantitatives, et 3) étudier les modifications de l'expression ARN des plants de coton G. hirsutum après l’écimage, pour 44 gènes impliqués dans la biosynthèse des COV, et également par une comparaison du transcriptome complet au moyen d'une analyse RNA-seq. Les résultats des trois domaines scientifiques, bio-informatique, analyse chimique et expression des gènes, ont pu être liés dans notre thèse de recherche: par ex., deux des gènes initialement identifiés par la bio-informatique, correspondant à deux enzymes, TPS50 (EC: 4.2.3.106 - (E ) bêta-ocimène synthase) et TPS16 (EC: 4.2.3.111 - alpha-terpinéol synthase), ont montré une augmentation de leur expression après l’écimage, et l'analyse GC-MS montre une modification correspondante des profils d'émission de COV. Ces mêmes composés avaient déjà été caractérisés dans d'autres plantes en réponse aux dégâts d’arthropodes. Ce travail de thèse constitue une première exploration des bases génétiques des émissions défensives de COV par les cotonniers cultivés G. hirsutum. La variabilité des comportements d’expression génique observés entre les trois génotypes étudiés de coton Upland africain G. hirsutum, permet de supposer qu’une diversité génétique est présente pour les émissions défensives de COV, ce qui pourrait permettre d’améliorer et d’adapter ces mécanismes de défense naturels et leur réponse à l’écimage, dans la perspective d'une protection naturelle plus efficace des champs de coton. / Gossypium hirsutum, the Upland cotton, represents more than 95% of the cotton fiber annually produced worldwide and is grown in about 40 countries. The protection of cotton fields against the attack of herbivorous arthropods needs important quantities of synthetic insecticides, around 18% of the world consumption for the year 2000, although this decreased very much thanks to Bt cotton and eradication programs for some pests. Volatile organic compounds (VOCs) naturally emitted by crop plants can reduce insect attacks through the influence of VOCs on the behaviors of herbivorous arthropods and auxiliary arthropods. Scientific research about plant VOCs has been increasing much since two or three decades. The stimulation of VOCs emissions by cultivated cotton plants is now recommended by entomologists of CIRAD as a component of the cotton fields protection strategy. “Topping", that is, cutting the head of the cotton plants during the useful floriferous period, stimulates the emission of defense VOCs. It is an environmentally friendly method and it limits health hazards due to the use of synthetic insecticides for the farmers and the surrounding human populations. The objective of the thesis was to improve our knowledge about the genetic bases of VOCs emissions after topping. The combination of several disciplines such as bioinformatics, chemical ecology and molecular genetics allowed us to: 1) analyze the genomic sequences of VOCs genes of the terpene and terpenoid biosynthesis pathways and transcription factors (TF) related to stress response, using the published genome databases of three cotton species, G. raimondii, G. arboreum (both diploid cottons) and G. hirsutum (tetraploid cotton), 2) study the VOCs emissions by cotton leaves of G. hirsutum plants in response to topping, by capturing these molecules in greenhouse and then characterizing their kinetic profiles by means of gas-chromatography mass-spectrometry (GC-MS), with quantitative measurements, and, 3) study the modifications of the RNA expression of G. hirsutum cotton plants after topping, for genes involved in VOCs biosynthesis through quantitative PCR measurements on 44 targeted genes and also by means of a whole-transcriptome comparison through an RNA-seq analysis. The results from the three different fields, bioinformatics, chemical analysis and gene expression, could be interrelated in our research thesis: e.g., two of the genes initially identified by bioinformatics, corresponding to two enzymes, TPS50 (EC: 4.2.3.106 - (E) -beta-ocimene synthase) and TPS16 (EC: 4.2.3.111 - alpha-terpineol synthase), were shown to increase their expression after topping, while the GC-MS analysis showed an modification of the corresponding VOCs in emission profiles. These compounds have been already characterized in other organisms in response to wounds produced by herbivorous insects. This thesis work is a first exploration of the genetic bases of defensive VOCs emission by G. hirsutum cultivated cottons. The variability of genic expression behaviors observed amongst the three genotypes of African Upland cotton G. hirsutum that were studied permits to hypothesize that a genetic diversity is present for defensive VOCs emissions, that could permit to improve and adapt by breeding these natural defense mechanisms and the response to topping, in perspective of a more efficient natural protection of cotton fields.

Thesis (Ph. D. Agriculture (Plant Protection)) -- University of Limpopo, 2013 / Global withdrawal of synthetic fumigant and non-fumigant nematicides due to their ecounfriendly impacts and high toxicity to non-target organisms, respectively, increased the research and development of alternatives for managing population densities of plantparasitic nematodes, particularly the root-knot (Meloidogyne species) nematodes. Although Meloidogyne species had been managed using genotypes that are resistant to plant-parasitic nematodes in various crops, various challenges negate the available or introgressed nematode resistance. In tomato (Solanum lycopersicum) production, nematode races and instability of nematode resistant genotypes under certain conditions necessitated the continued research and development of alternatives since most of the existing commercial tomato cultivars are highly susceptible to various biological races of Meloidogyne species. The aim of the study was to research and develop appropriate dosages of two phyto- nematicides which could be applied through drip irrigation system in open field tomato production systems, while the specific objectives were to: (1) determine whether a computer-based model could provide nonphytotoxic concentrations to tomato plants using fresh fruits of wild watermelon (Cucumis africanus) and wild cucumber (C. myriocarpus) under greenhouse conditions, (2) determine whether computer-based concentrations from the two plant species when using dried fruits would be less phytotoxic and more suppressive to nematodes, (3) investigate application time intervals for the two products, (4) determine responses of plant growth in tomato and nematode suppression in respect to the derived dosages, and and (5) validate dosages of fermented crude extracts from the two plant species with respect to plant growth of tomato and suppression of nematode numbers. xxxiii Greenhouse, microplot and field studies were set to test the hypotheses intended to achieve the stated objectives, with reliability of measured variables being ensured by using statistical levels of significance (P ≤ 0.05) and coefficients of determination (R2), while validity was ensured by conducting experiments at the same location over two seasons and/or by setting up factorial treatments. Firstly, fermented plant extracts of fresh fruits from C. africanus and C. myriocarpus consistently reduced population densities of Meloidogyne species by 80-92% and 50-90%, respectively. Tomato plants were highly sensitive to the two products as shown by the total degree of sensitivities (Σk) and biological index of 0 and 3, respectively. Also, the mean concentration stimulation range (MCSR) of 11% and 7% concentrations, respectively, attested to this phytotoxicity. Secondly, fermented crude extracts of dried fruits from C. africanus and C. myriocarpus also reduced population densities of Meloidogyne species by 78-97% and 87-97%, respectively. Tomato plants were highly tolerant to the two products in dried form as shown by the total degree of sensitivities (Σk) and biological index of 4 and 3, respectively. The MCSR values for C. africanus and C. myriocarpus dried fruits on tomato were 2.64% and 2.99%, respectively, which for the purpose of this study were individually adjusted to 3%, which translated to 36 L undiluted material/ha of 4 000 tomato plants. In subsequent studies, 3% concentration was used as the standard, along with double strength concentration, namely, 6% concentration. Thirdly, the MCSR values derived in Objective 4, namely 3% and 6% concentration for both Cucumis species using the CARD model were used in the optimisation of application time interval using the innovative concept of weeks (0, 1, 2, 3 and 4) in a 30-day month period. Application time interval for 3% and 6% concentrations of C. africanus fruits was xxxiv optimised at 2.40 and 2.61 weeks in a 30-day month period, respectively, which translated to 18 days [(2.4 weeks/4 weeks) × 30 days] and 20 days [(2.6 weeks/4 weeks) × 30 days], respectively. In contrast, for both concentrations from fermented crude extracts of C. myriocarpus fruits, application time interval was optimised at 16 days for 2.2 and 2.1 weeks, respectively. During optimisation of application frequencies, fermented crude extracts from C. africanus and C. myriocarpus reduced final population densities of M. incognita race 2 by 70-97% and 76-96%, respectively. Fourthly, optimum application intervals (time), allowed computation of dosage, which is a product of concentration and application frequency (dosage = concentration × application frequency). Fifthly, validation of the dosages under open field conditions suggested that 6% × 16-day dosage under crude extracts from C. myriocarpus fruit significantly (P ≤ 0.05) improved growth of tomato plants when compared with those of either 0% (untreated control) or 3% at 16 days. In contrast, dosages of C. africanus fruit at two application frequency had no effect on growth of tomato plants – suggesting that either of the dosages was suitable for use in tomato production since both reduced nematode numbers. During validation, the materials reduced nematode numbers by margins similar to those observed previously under other environments. In conclusion, crude extracts of the two Cucumis species have stimulatory concentrations which have potential similar reductive effects on population densities of Meloidogyne species and could serve as botanical nematicides. However, since plant responses to the two products differed in terms of their respective dosages and active ingredients, it implied that for further improvement of the two, the overriding focus should be on their interaction with the protected plants and nematode numbers. Ideally, future research xxxv should include environmental impact studies, especially on the influence of the products fruit quality of tomato, earthworms, fish and bees.

Thesis (Ph. D. Agriculture (Plant Protection)) -- University of Limpopo, 2013 / Global withdrawal of synthetic fumigant and non-fumigant nematicides due to their ecounfriendly impacts and high toxicity to non-target organisms, respectively, increased the research and development of alternatives for managing population densities of plantparasitic nematodes, particularly the root-knot (Meloidogyne species) nematodes. Although Meloidogyne species had been managed using genotypes that are resistant to plant-parasitic nematodes in various crops, various challenges negate the available or introgressed nematode resistance. In tomato (Solanum lycopersicum) production, nematode races and instability of nematode resistant genotypes under certain conditions necessitated the continued research and development of alternatives since most of the existing commercial tomato cultivars are highly susceptible to various biological races of Meloidogyne species. The aim of the study was to research and develop appropriate dosages of two phyto- nematicides which could be applied through drip irrigation system in open field tomato production systems, while the specific objectives were to: (1) determine whether a computer-based model could provide nonphytotoxic concentrations to tomato plants using fresh fruits of wild watermelon (Cucumis africanus) and wild cucumber (C. myriocarpus) under greenhouse conditions, (2) determine whether computer-based concentrations from the two plant species when using dried fruits would be less phytotoxic and more suppressive to nematodes, (3) investigate application time intervals for the two products, (4) determine responses of plant growth in tomato and nematode suppression in respect to the derived dosages, and and (5) validate dosages of fermented crude extracts from the two plant species with respect to plant growth of tomato and suppression of nematode numbers. xxxiii Greenhouse, microplot and field studies were set to test the hypotheses intended to achieve the stated objectives, with reliability of measured variables being ensured by using statistical levels of significance (P ≤ 0.05) and coefficients of determination (R2), while validity was ensured by conducting experiments at the same location over two seasons and/or by setting up factorial treatments. Firstly, fermented plant extracts of fresh fruits from C. africanus and C. myriocarpus consistently reduced population densities of Meloidogyne species by 80-92% and 50-90%, respectively. Tomato plants were highly sensitive to the two products as shown by the total degree of sensitivities (Σk) and biological index of 0 and 3, respectively. Also, the mean concentration stimulation range (MCSR) of 11% and 7% concentrations, respectively, attested to this phytotoxicity. Secondly, fermented crude extracts of dried fruits from C. africanus and C. myriocarpus also reduced population densities of Meloidogyne species by 78-97% and 87-97%, respectively. Tomato plants were highly tolerant to the two products in dried form as shown by the total degree of sensitivities (Σk) and biological index of 4 and 3, respectively. The MCSR values for C. africanus and C. myriocarpus dried fruits on tomato were 2.64% and 2.99%, respectively, which for the purpose of this study were individually adjusted to 3%, which translated to 36 L undiluted material/ha of 4 000 tomato plants. In subsequent studies, 3% concentration was used as the standard, along with double strength concentration, namely, 6% concentration. Thirdly, the MCSR values derived in Objective 4, namely 3% and 6% concentration for both Cucumis species using the CARD model were used in the optimisation of application time interval using the innovative concept of weeks (0, 1, 2, 3 and 4) in a 30-day month period. Application time interval for 3% and 6% concentrations of C. africanus fruits was xxxiv optimised at 2.40 and 2.61 weeks in a 30-day month period, respectively, which translated to 18 days [(2.4 weeks/4 weeks) × 30 days] and 20 days [(2.6 weeks/4 weeks) × 30 days], respectively. In contrast, for both concentrations from fermented crude extracts of C. myriocarpus fruits, application time interval was optimised at 16 days for 2.2 and 2.1 weeks, respectively. During optimisation of application frequencies, fermented crude extracts from C. africanus and C. myriocarpus reduced final population densities of M. incognita race 2 by 70-97% and 76-96%, respectively. Fourthly, optimum application intervals (time), allowed computation of dosage, which is a product of concentration and application frequency (dosage = concentration × application frequency). Fifthly, validation of the dosages under open field conditions suggested that 6% × 16-day dosage under crude extracts from C. myriocarpus fruit significantly (P ≤ 0.05) improved growth of tomato plants when compared with those of either 0% (untreated control) or 3% at 16 days. In contrast, dosages of C. africanus fruit at two application frequency had no effect on growth of tomato plants – suggesting that either of the dosages was suitable for use in tomato production since both reduced nematode numbers. During validation, the materials reduced nematode numbers by margins similar to those observed previously under other environments. In conclusion, crude extracts of the two Cucumis species have stimulatory concentrations which have potential similar reductive effects on population densities of Meloidogyne species and could serve as botanical nematicides. However, since plant responses to the two products differed in terms of their respective dosages and active ingredients, it implied that for further improvement of the two, the overriding focus should be on their interaction with the protected plants and nematode numbers. Ideally, future research xxxv should include environmental impact studies, especially on the influence of the products fruit quality of tomato, earthworms, fish and bees.

Graduation date: 1995

House flies (Musca domestica L.) are common pests affecting horses and their owners. Control of house flies in stable yards is currently based on the use of pesticides. However, the development of resistance by these flies to most pesticide groups has motivated horse owners to seek alternative methods of fly control. An entomopathogenic fungus, Beauveria bassiana (Bb) and an entomopathogenic bacterium, Bacillus thuringiensis var. israelensis (Bti) are two biological agents known to have activity against house flies. The broad objective of this study was to evaluate the effect of these two biological control agents on house flies in an equine environment. Using a structured questionnaire, presented in Chapter 2, thirty horse owners in KwaZulu-Natal were asked about the nuisance value of house flies, their current control measures, the potential market for biocontrol agents against house flies, and each owner’s perception of biocontrol methods. The horse owners were using three methods of house fly control namely, physical, chemical and biological. Most horse owners (97%) wanted access to effective biocontrol agents for control of house flies. Most horse owners (80%) stabled their horses at night, some or all of the time. The resultant manure piles in the stable yard were considered to be the primary cause of house fly problems. About 64% of the horse owners were dissatisfied with the currently available methods of controlling house flies in this situation. Chapter 3 covers two observational trials in which varying doses of Bacillus thuringiensis var. israelensis (Bti) were fed to horses, in order to identify a baseline dosage to give to horses in order to adequately control house fly populations growing in horse manure. The bacterium Bti, grown on wheat bran, was fed to six miniature horses at doses of 0, 0.125, 0.25, 0.5, 0.75, and 1.0 g per meal in Trial 3a, and at 0, 0.5, 1, 2, 4 and 8 g per meal in Trial 3b. Faeces were collected three times a week for 11 weeks and placed in incubation trays to allow the number of emerging adult house flies and closed pupae to be counted. In Trial 3a, there was a significant reduction in the number of closed pupae with an increase in Bti in the feed. The regression equation suggests that there will be 3.1 times as many closed pupae in the faeces when horses are fed 1 g of Bti in their feed, than when horses are fed no Bti. This dosage is the minimum baseline dosage for future trials. v Using manure from horses dosed in Trials 3a and 3b, the survival of the bacterium through the gut of horses was evaluated using a standard isolation technique. The growth of Bt colonies on the manure after the Bt isolation technique showed that some of the bacterial cells survived transition through the digestive tract of the horse. This study was qualitative in nature and did not attempt to quantify the level of Bti spore survival. These two observations suggest that Bacillus thuringiensis var. israelensis has the potential to be used as a biocontrol agent, applied via horse feed, for the control of house flies in stable yards. Future clinical trials, with appropriate replication, should be conducted using 1 g Bti/meal as the lowest test dosage. The objective of Chapter 4 was to determine whether spraying Bti or Bb on to horse manure is effective in the control of house flies. Over a six week period, two spraying trials were conducted in which increasing doses of Bb and Bti were sprayed on to 500 g samples of horse manure. Counts of house fly pupae and adults were taken. The doses of Bb and Bti tested were 0, 1, 2, 4 g in Trial 4a, and 0, 4, 8 and 12 g in Trial 4b. The research reported in Chapter 4 was characterized by the unexpectedly high levels of biological variation in egg, larvae and pupae numbers that were found in samples of horse manure, taken from the same skip two days apart. The statistical design of the two trials conducted was inadequate to cope with the high level of variation about treatment means for fly and larval counts. However, despite the lack of significant differences between treatment means, there is observational evidence that suggests that both Bb and Bti do have an effect on house fly survival. A simplified statistical model, which compared the number of hatched house flies on untreated manure, with the number on manure treated with any level of Bb (1 to 4 g /250 ml water), found a significant reduction in the number of hatched flies on treated manure. There was no significant corresponding reduction in the number of closed pupae, which suggests that Bb acts primarily before the larva pupates. The optimal dose of Bb and Bti to be sprayed on to manure could not be determined because of the high variation about treatment means. It is suggested that, in future trials similar dosages for Bb could be tested, but that higher dosages of Bti (starting at 2 g/250 ml water) should be used. Trial periods should be extended and replication increased dramatically to reduce variation about treatment means. Transformation of data before analysis may also be necessary to equalize variation about treatment means. / M.Sc.Agric. University of KwaZulu-Natal, Pietermaritzburg 2013.

Thesis (Ph.D. (Horticulture)) -- University of Western Sydney, 2002. / "A thesis submitted to the University of Western Sydney for the fulfillment of study for a degree of Doctor of Philosophy in Horticulture" "Principal supervisor: Robert Spooner-Hart, co-supervisor: Andrew Beattie, co-supervisor: Alfie Meats" Bibliography : leaves 231-265.

Thesis (M.Sc. (Biochemistry, Microbiology & Biotechnology)) - Rhodes University, 2008.