Promising Biological Control Research Articles

Wide interguild relationships among entomopathogenic and free-living nematodes in soil as measured by real time qPCR

Entomopathogenic nematodes (EPNs) are promising biological control agents of soil-dwelling insect pests of many crops. These nematodes are ubiquitous in both natural and agricultural areas. Their efficacy against arthropods is affected directly and indirectly by food webs and edaphic conditions. It has long been suggested that a greater understanding of EPN ecology is needed to achieve consistent biological control by these nematodes and the development of molecular tools is helping to overcome obstacles to the study of cryptic organisms and complex interactions. Here we extend the repertoire of molecular tools to characterize soil food webs by describing primers/probe set to quantify certain free-living, bactivorous nematodes (FLBNs) that interact with EPNs in soil. Three FLBN isolates were recovered from soil baited with insect larvae. Morphological and molecular characterization confirmed their identities as Acrobeloides maximum (RT-1-R15C and RT-2-R25A) and Rhabditis rainai (PT-R14B). Laboratory experiments demonstrated the ability of these FLBNs to interfere with the development of Steinernema diaprepesi, Steinernema riobrave and Heterorhabditis indica parasitizing the weevil Diaprepes abbreviatus (P<0.001), perhaps due to resource competition. A molecular probe was developed for the strongest competitor, A. maximum. We selected the highly conserved SSU rDNA sequence to design the primers/probe, because these sequences are more abundantly available for free-living nematodes than ITS sequences that can likely provide better taxonomic resolution. Our molecular probe can identify organisms that share ⩾98% similarity at this locus. The use of this molecular probe to characterize soil communities from samples of nematode DNA collected within a citrus orchard revealed positive correlations (P<0.01) between Acrobeloides-group nematodes and total numbers of EPNs (S. diaprepesi, H. indica and Heterorhabditis zealandica) as well as a complex of nematophagous fungi comprising Catenaria sp. and Monachrosporium gephyropagum that are natural enemies of EPNs. These relationships can be broadly interpreted as supporting Linford’s hypothesis, i.e., decomposition of organic matter (here, insect cadavers) greatly increases bactivorous nematodes and their natural enemies.

Effect of previous feeding by Falconia intermedia (Hemiptera: Miridae) on subsequent feeding activity on the invasive shrub Lantana camara (Verbenaceae)

After initially building up to high densities, populations of a promising biological control agent, Falconia intermedia (Distant) (Hemiptera: Miridae), imported to control the invasive plant Lantana camara L. (Verbenaceae) in South Africa, disappeared at almost all release sites in the Eastern Cape, Mpumalanga, KwaZulu-Natal and Limpopo provinces. In an attempt to explain this phenomenon, laboratory trials were conducted to determine the effect of previous feeding on the subsequent performance of F. intermedia on L. camara varieties from two of the Eastern Cape release sites; Lyndhurst Farm and Whitney Farm. F. intermedia feeding damage, number of adults, number of nymphs and oviposition on Whitney Farm, L. camara plants were 40.8% (p<0.001), 106% (p<0 .001), 81.5% (p <0.001) and 80% (p<0.001) higher, respectively, than on Lyndhurst Farm, indicating the superior suitability and quality of Whitney Farm plants for F. intermedia performance. A defence response was observed by plants from Lyndhurst Farm that were previously fed on (i.e. induced). The induced plants had significantly lower F. intermedia feeding damage (21.4% less, p=0.007), numbers of adults (187.5% less, p=0.034), numbers of nymphs (110% less, p=0.071) and oviposition (99.8% less, p=0.021) than plants that were not previously fed on. The defence responses were elicited and expressed throughout the plant in both damaged and undamaged leaves within five weeks after insect release (rapidly induced response). The significant decrease in herbivore performance suggests that some L. camara varieties possess factors that enable them to resist subsequent feeding activity, offering us some understanding of one of the factors that might have contributed to the failure of F. intermedia in South Africa.

Insecticidal Activity of the Granulosis Virus in Combination with Neem Products and Talc Powder Against the Potato Tuberworm Phthorimaea operculella (Zeller) (Lepidoptera: Gelechiidae)

The potato tuberworm Phthorimaea operculella (Zeller) is an important agricultural pest that causes significant economic losses to potato growers worldwide. The addition of an effective method of biological control for the potato tuberworm is greatly needed, and is currently unavailable in Brazil. The granulosis virus (Baculoviridae) is a promising biological control agent to protect post-harvest potatoes and in storage from the potato tuberworm. However, the control measure must be economically feasible. Liquid suspensions of a granulosis virus applied alone or in mixture with two commercial neem oil-based products (DalNeem™ and NeemAzal™), and a dry powder formulation of viral granules were evaluated for control of potato tuberworm larvae by treating potato tubers under laboratory conditions. High larval mortality (86.7%) was achieved when DalNeem and virus were applied together at 4 mg of azadirachtin/L and 10(4) occlusion bodies (OBs)/mL, respectively. This combination resulted in ≥50% efficacy in relation to their counterparts alone. Conversely, NeemAzal did not enhance virus effectiveness against larvae of the potato tuberworm. The talc-based virus formulation was used for dusting seed tubers at different concentrations and resulted in 100% larval mortality at 5 × 10(8) OBs/g. Formulated and unformulated virus provided 50% mortality at 166 OBs/g and at 5.0 × 10(5) OBs/mL, respectively. As a result, talc-based virus formulation had a better control efficiency on potato tuberworm than the aqueous virus suspension. The granulosis virus combined with DalNeem at low rates or formulated with talc powder is a viable option to control the potato tuberworm under storage conditions.

Pseudomonas putida NBRIC19 provides protection to neighboring plant diversity from invasive weed Parthenium hysterophorus L. by altering soil microbial community

Parthenium hysterophorus L. (Parthenium) is an invasive weed species which is spreading worldwide affecting natural ecological systems, biodiversity, crop production and human health. The present study was conducted to evaluate the potential of plant growth promoting Pseudomonas putida NBRIC19 in detoxifying the phytotoxic effect of Parthenium. Significant increase in C/N ratio, macronutrients, and micronutrients was observed in P. putida NBRIC19-treated soil. P. putida NBRIC19 treatment of the soil provided protection to plant communities in Parthenium invaded area, as the species diversity had increased in the treatment as compared to non-bacterized soil. P. putida NBRIC19 treatment besides Parthenium, also succeeded in controlling other weed species like Commelina benghalensis and Cynodon dactylon. In addition to this, the impact of Parthenium was also studied on functional microbial diversity based on carbon source utilization pattern. It was observed that P. putida NBRIC19 treatment of soil had shifted the microflora in such a manner that utilization of toxic allelochemicals increased to lessen their phytotoxic effect. Taken together, these results suggest that soil treatment with P. putida NBRIC19 may be used as a promising biological control measure for controlling the phytotoxic effect of Parthenium and in protecting ecosystem integrity of neighboring plants in Parthenium invaded areas.

Influence of five aphid species on development and reproduction of Propylaea japonica (Coleoptera: Coccinellidae)

Propylaea japonica is one of the most promising biological control agents in northern China. The objectives of the present study were to investigate the preimaginal development, immature survival, reproduction, longevity and fecundity of the ladybeetle, P. japonica, when reared on five different aphids species, Aphis gossypii Glover, Myzus persicae (Sulzer), Aphis craccivora Koch, Acyrthosiphon pisum (Harris) and Chaitophorus populeti (Panzer) and to quantify their relative suitability as prey. The results of the study indicate that the pre-adult development was shorter (9.9±0.1days) when fed on A. craccivora but was the longest (13.5±0.2days) on A. pisum. Immature survival, adult emergence, growth index, development rate, male and female longevity, oviposition period, and fecundity were maximal when P. japonica were fed on A. gossypii. In contrast these biological parameters were minimal on A. pisum. The ladybird adult’s fresh weight was maximal after feeding on A. gossypii and minimal on A. pisum. Further analysis revealed linear relationships between rate of development and weight of adult females; and longevity and fecundity of adult females. The results of the present study suggest the order of suitability of aphid species for P. japonica is A. gossypii>A. craccivora>M. persicae>C. populeti>A. pisum. The present study further propose that either the mass-rearing of P. japonica in the laboratory can be carried out by supplying the best food or the understanding its relative abundance in the field in relation to the availability of different prey species.

Selection of promising fungal biological control agent of the western flower thrips Frankliniella occidentalis (Pergande)

Larval stages of Frankliniella occidentalis are known to be refractory to fungal infection compared with the adult stage. The objective of this study was to identify promising fungal isolate(s) for the control of larval stages of F. occidentalis. Ten isolates of Metarhizium anisopliae and eight of Beauveria bassiana were screened for virulence against second-instar larvae of F. occidentalis. Conidial production and genetic polymorphism were also investigated. Metarhizium anisopliae isolates ICIPE 7, ICIPE 20, ICIPE 69 and ICIPE 665 had the shortest LT(50) values of 8.0-8.9 days. ICIPE 69, ICIPE 7 and ICIPE 20 had the lowest LC(50) values of 1.1 × 10(7), 2.0 × 10(7) and 3.0 × 10(7) conidia ml(-1), respectively. Metarhizium anisopliae isolate ICIPE 69 produced significantly more conidia than M. anisopliae isolates ICIPE 7 and ICIPE 20. Internally transcribed spacers sequences alignment showed differences in nucleotides composition, which can partly explain differences in virulence. These results coupled with the previous ones on virulence and field efficacy against other species of thrips make M. anisopliae isolate ICIPE 69 a good candidate. Metarhizium anisopliae isolate ICIPE 69 can be suggested for development as fungus-based biopesticide for thrips management.

Survival patterns under Costa Rican field conditions of the gregarious caterpillar Euselasia chrysippe (Lepidoptera: Riodinidae), a potential biological control agent of Miconia calvescens (Melastomataceae) in Hawaii

Survival of Euselasia chrysippe, a natural herbivore of Miconia calvescens, was investigated under field and laboratory conditions in Costa Rica as part of a biological control program in Hawaii. Following its introduction to Pacific islands, M. calvescens has become a dominant invasive species and the gregarious caterpillar of E. chrysippe has emerged as a promising biological control candidate. Under laboratory conditions, survivorship from egg to adult was 63%. E. chrysippe produced viable clutches in an experimental field plot. Similar clutch sizes occurred in both natural habitats and the experimental plot. Stage-specific life tables encompassing two years in the experimental plot indicate that larval survivorship from egg to the end of the sixth instar was about 10%. Egg parasitism was low in natural habitats and nil under experimental conditions. Larval group sizes were similar in natural and experimental field conditions, suggesting that survivorship is similar in both environments. During the coldest dry period no larvae survived past the fifth instar. Rainfall was a limiting factor for the survivorship of E. chrysippe in the experimental plot, but temperature appears to be the factor that would limit the effect of E. chrysippe on M. calvescens in Hawaii. Efforts should be invested in natural quarantine facilities to provide a testing ground for this species in target environments.

An intensive search for promising fungal biological control agents of ticks, particularly Rhipicephalus microplus

Entomopathogenic fungi have been investigated worldwide as promising biological control agents of the cattle tick Rhipicephalus microplus. The current study evaluates the virulence of several fungal isolates to R. microplus larva in the laboratory as part of an effort to identify isolates with promise for effective biocontrol of R. microplus in the field. Sixty fungal isolates, encompassing 5 Beauveria spp. and 1 Engyodontium albus (=Beauveria alba), were included in this study. In addition to bioassays, the isolates were characterized morphologically and investigated as to their potential for conidial mass production. These findings were correlated with previous reports on the same fungal isolates of their natural UV-B tolerance (Fernandes et al., 2007), thermotolerance and cold activity (Fernandes et al., 2008), and genotypes (Fernandes et al., 2009). R. microplus larvae obtained from artificially infested calves were less susceptible to Beauveria bassiana infection than ticks acquired from naturally infested cattle from a different location. Isolates CG 464, CG 500 and CG 206 were among the most virulent Beauveria isolates tested in this study. All fungal isolates presented morphological features consistent with their species descriptions. Of the 53 B. bassiana isolates, five (CG 481, CG 484, CG 206, CG 235 and CG 487) had characteristics that qualified them as promising candidates for biological control agents of R. microplus, viz., mean LC50 between 107 and 108conidiaml−1; produced 5000 conidia or more on 60mm2 surface area of PDAY medium; and, in comparison to untreated (control) conidia, had the best conidial tolerances to UV-B (7.04kJm−2) and heat (45°C, 2h) of 50% or higher, and conidial cold (5°C, 15d) activity (mycelial growth) higher than 60%. The current study of 60 Beauveria spp. isolates, therefore, singles out a few (five) with high potential for controlling ticks under field conditions.

Prospective Agents for the Biological Control ofTithonia rotundifolia(Mill.) S.F.Blake andTithonia diversifolia(Hemsl.) A.Gray (Asteraceae) in South Africa

Starting in 2007, two weedy sunflower species, Tithonia rotundifolia (Mill.) S.F.Blake and Tithonia diversifolia (Hemsl.) A.Gray (Asteraceae: Heliantheae), were targeted for biological control in South Africa. Surveys conducted in their native range (Mexico) revealed that there were five potential biological control agents for T. rotundifolia, and three of these are currently undergoing host-specificity and performance evaluations in South Africa. Two leaf-feeding beetles, Zygogramma signatipennis (Stål) and Zygogramma piceicollis (Stål) (Coleoptera: Chrysomelidae), are the most promising biological control agents for T. rotundifolia: preliminary host-specificity trials suggest that they are adequately host-specific. The stem-boring beetle, Lixus fimbriolatus Boheman (Coleoptera: Curculionidae), is also highly damaging to T. rotundifolia, but its host range is yet to be determined. Two other stem-boring beetles, Canidia mexicana Thomson (Coleoptera: Cerambycidae) and Rhodobaenus auctus Chevrolat (Coleoptera: Curculionidae), have also been recorded on T. rotundifolia, and these will be considered for further testing if L. fimbriolatus is found to be unsuitable for release in South Africa. Only two insect species were imported as candidate agents on T. diversifolia, the leaf-feeding butterfly Chlosyne sp. (Lepidoptera: Nymphalidae), and an unidentified stem-boring moth (Lepidoptera: Tortricidae): the latter was tested in quarantine but rejected because it attacked several sunflower cultivars. Only one pathogen, Puccinia enceliae Dietel & Holw. (Uredinales: Pucciniaceae), was found that could potentially have been used as a biological control agent against the Tithonia species, but attempts to culture this rust were unsuccessful.

Parasitoids of Dipteran leafminers, Diglyphus spp. (Insecta: Hymenoptera: Eulophidae)

This 3-page illustrated fact sheet describes these promising biological control agents for agromyzid leafminers (Diptera: Agromyzidae) — distribution, description, hosts, and economic importance. Includes references. Written by Jian Li and Dakshina R. Seal, Published by the UF Department of Entomology and Nematology as part of the Featured Creatures Collection, December 2010. EENY 484/IN877: Parasitoids of Dipteran leafminers, Diglyphus spp. (Insecta: Hymenoptera: Eulophidae) (ufl.edu)

Performance of potential European biological control agents of Vincetoxicum spp. with notes on their distribution

Exploratory surveys were conducted in Europe to locate potential biological control agents of Vincetoxicum nigrum and Vincetoxicum rossicum and to collect known specialist herbivores of Vincetoxicum hirundinaria (Abrostola asclepiadis, Chrysolina a. asclepiadis, Eumolpus asclepiadeus and Euphranta connexa). Development of the most abundant herbivores found attacking leaves, roots, and developing seeds was evaluated in the laboratory on three Vincetoxicum spp. Field surveys revealed that the highest diversity of herbivores was associated with the abundant and widespread V. hirundinaria. No new herbivores were reported from V. hirundinaria or V. nigrum, but the leaf-feeding noctuid Hypena opulenta was recorded for the first time attacking V. rossicum and V. scandens in Ukraine. Based upon larval survival and development and adult fecundity, the leaf feeders A. asclepiadis, C. a. asclepiadis, and H. opulenta, perform better on V. nigrum and V. rossicum than on V. hirundinaria. Larval performance of the root feeder E. asclepiadeus followed a similar pattern, but adult fecundity of this insect did not vary among host plant species. Immature development time of the pre-dispersal seed feeder E. connexa is similar among hosts, but larvae grow larger on V. nigrum and V. hirundinaria than on V. rossicum. All herbivores are promising biological control agents of Vincetoxicum in North America and we have prioritized host range testing with H. opulenta and E. asclepiadeus.

The influence of aquatic predators on mosquito abundance in animal drinking troughs in New Zealand

The occurrence and abundance of mosquito populations may be associated with the abundance of predators. We examined the relationship between aquatic predators and populations of mosquitoes in animal water troughs in Waikanae, New Zealand. We also investigated the effects of water volume and environmental factors (temperature, rainfall, wind speed, humidity, and pressure) in order to further understand factors influencing mosquito and predator populations. Logistic regression indicated that the presence or absence of mosquitoes was primarily affected by three factors: predator abundance, week of observation, and water volume. Pearson's correlation indicated that the presence of predators had a positive correlation with water volume (r² = 0.176, p< 0.05). Otherwise, the presence of mosquito larvae in water troughs was negatively correlated with water volume (r² =-0.159, p=0.022) and wind speed (r² =0.142, p=0.041). We established a translocation experiment in which predators or mosquitoes were moved between troughs in order to examine the prey survival rate after exposure to Anisops wakefieldi predators. The survival rate of mosquitoes was not significantly different, between 0-0.1%, irrespective of the number of predators translocated (1-9) or the initial mosquito density (20-70 larvae). Our results suggested that A. wakefieldi predators may have the potential to be a promising biological control tool for the control of mosquito populations by altering mosquito population dynamics.

Statistical approach to determine the effect of combined environmental parameters on conidial development of Trichoderma atroviride (T‐15603.1)

Trichoderma atroviride (T-15603.1) is a promising fungal agent for biological control of wood decay fungi in urban tree wounds. The aim of this work was to determine the combined effects of water activity (a(w), 0.998-0.892), temperature (10-30 °C) and pH (3-7) on the development of conidia on low-nutrient agar (LNA). Lag phase prior to germination (h), germination rates (µ(m)) and germ-tube elongation were obtained at each set of conditions. The experimental data were used to fit a response surface model for predicting the germination rates of T-15603.1 and to analyze the effect of the abiotic parameters tested. The polynomial response surface model was mathematically evaluated using graphical plots and several statistical indices (RMSE, %SEP, A(f), B(f), pRE). Data analyses showed a highly significant effect on conidial development of a(w) and temperature (P < 0.001), whereas no significant effect of pH (P ≥ 0.05) was observed. The germination rate dropped and the lag prior to germination increased as the temperature and a(w) decreased, but T-15603.1 appeared to be more sensitive to a(w) reduction than to temperature. The minimum a(w) level for germination was 0.910 at 15-25 °C, and maximum germination rates were obtained at a(w) = 0.998, 25-30 °C and pH 5. The response surface model was useful and widely accurate (R² = 0.983) for predicting the germination rate of T-15603.1 and complemented the experimental results. These findings contribute to better understanding how combined environmental factors affect the environmental parameter tolerance levels of T-15603.1 and to the development of an adequate delivery system for optimized application of T-15603.

Postharvest biological control of melon pathogens using Bacillus subtilis EXWB1.

Melon (Cucumis melo L.) is a widely used fruit which suffers from postharvest decay resulting in large economic losses. In this work we investigated the biological control of melon postharvest diseases using Bacillus subtilis EXWB1, which we had previously isolated and identified from melon in lab experiments. We found that this bacterium excretes a bio-surfactant in Luria- Bertani broth (pH 5-8) that helps it to stick to the hydrophobic melon skin, and spreads at a speed of 125 µm per h over the surface at room temperature and high humidity. During storage time we found that the peak of respiration of melon fruits increased on day 6, and was reduced by 26.1% and 71.9%, respectively in the presence of B. subtilis EXWB1 in inoculated and uninoculated treatments. The ethylene production peak was delayed by about 2 days and reduced by 72.3% and 61.6% on day 4. Growth of one of the main fungal pathogens of melon, Alternaria alternata, was limited around inoculated sites on the fruit surface, and necrotic lesions induced by this pathogen were significantly reduced by 77.2% after coinoculation with B. subtilis EXWB1. Treated fruits retained high levels of sugar and vitamin C, and low levels of organic acids, and maintained water content and turgidity at room temperature. Our findings provide an alternative and promising biological control agent, B. subtilis EXWB1, which can suppress postharvest diseases caused by A. alternata, and may effectively control postharvest physiological changes caused by fungal pathogens.

Performance of three endophytic actinomycetes in relation to plant growth promotion and biological control of Pythium aphanidermatum, a pathogen of cucumber under commercial field production conditions in the United Arab Emirates

In the current study, the performance of three endophytic actinomycetes identified as Actinoplanes campanulatus, Micromonospora chalcea and Streptomyces spiralis previously shown to reduce seedling damping-off, and root and crown rots of mature cucumber (Cucumis sativus) caused by Pythium aphanidermatum in pots under greenhouse conditions were further evaluated to determine their potential as biological control agents and as plant growth promoters in the field under the conditions of commercial production of cucumbers in the United Arab Emirates (UAE). When applied individually or in combination to cucumber seedlings, the three isolates significantly promoted plant growth and yield and reduced seedling damping-off and root and crown rots of mature cucumber plants. Individually the performance level of S. spiralis was relatively the best followed by A. campanulatus and then by M. chalcea. The three isolates (which were not inhibitory to each other) performed better, both as biological control agents as well as plant growth promoters, when applied together than when they were inoculated individually. The ability of these three isolates to colonize the internal tissues of roots, stems and leaves under field conditions, and to persist up to 8 weeks after seedling inoculation, showed that they can easily adapt to an endophytic habit systemically within healthy cucumber plants. As the three endophytic actinomycete isolates also colonized the rhizosphere and showed outstanding rhizosphere competency it is clear that they are facultative and not obligate endophytes. The success with the three inoculants indicated that they could well be used in place of the fungicide metalaxyl which is currently recommended for the management of Pythium diseases in the UAE. This is the first successful field use of endophytic actinomycetes as promising plant growth promoters and biological control agents against Pythium diseases of cucumber.

Effects of leaf and root herbivory by potential insect biological control agents on the performance of invasive Vincetoxicum spp.

The European leaf-feeding moth Abrostola asclepiadis and root-feeding beetle Eumolpus asclepiadeus are promising biological control agents for two European swallow-worts ( Vincetoxicum rossicum and Vincetoxicum nigrum) in North America, however, their impact on plant performance is uncertain. Densities of each herbivore were manipulated in a common garden to determine whether leaf and root herbivory affect the performance of these plants. During the second year of the experiment, V. rossicum and V. nigrum unexpectedly became infected with the fungal pathogens Ascochyta sp. and Cercospora sp. (Ascomycota), respectively. Although pathogen infection mainly reduced shoot height and delayed reproduction, herbivore effects on plant growth were still evident. Leaf herbivory by A. asclepiadis had no effect on plant growth 1 year after defoliation. Root herbivory by E. asclepiadeus reduced shoot height and plant biomass and decreased the ability of plants to compensate for pathogen attack. Pathogen infection prevented detection of herbivore effect on reproduction. Due to its substantial impact on plant biomass, E. asclepiadeus should be further evaluated as a biological control agent against Vincetoxicum spp. populations invading open habitats in North America. Further research is needed to evaluate the impact of A. asclepiadis in combination with E. asclepiadeus and plant competition under high and low light conditions.

Induction of Tolerance to Fusarium Wilt and Defense-Related Mechanisms in the Plantlets of Susceptible Berangan Banana Pre-Inoculated with Pseudomonas sp. (UPMP3) and Burkholderia sp. (UPMB3)

This study is aimed at assessing the ability of two endophytic bacteria originally isolated from healthy oil palm roots, Pseudomonas sp. (UPMP3) and Burkholderia sp. (UPMB3) to induce resistance in susceptible Berangan banana against Fusarium oxysporum race 4 (FocR4). Increased accumulation of resistance-related enzymes such as peroxidase (PO), phenylalanine ammonia lyase (PAL), lignithioglycolic acid (LTGA), and pathogenesis-related (PR) proteins (chitinase and β-1,3-glucanase) has been observed in plantlets treated with endophytic bacteria UPMP3 and UPMB3 singly or as mixture under glasshouse conditions. Pre-inoculation of banana plantlets with UPMP3 showed a significant reduction in Fusarium wilt incidence 72 d after challenged inoculation with FocR4. UPMB3 was less effective in suppressing Fusarium wilt compared to UPMP3, whereas, the mixture of both endophytes showed an intermediate effect. Based on these results, it is concluded that UPMP3 could be a promising biological control agent that can trigger resistance against Fusarium wilt in susceptible Berangan banana.

Differentiation of entomopathogenic fungus Beauveria bassiana (Ascomycetes: Hypocreales) isolates by PCR-RFLP

The entomopathogenic fungus Beauveria bassiana is a promising biological control agent of several insect pests in agriculture. Molecular approaches (PCR, DNA sequence analysis and PCR-RFLP) were used in our research as tools for the identification of different B. bassiana isolates. Our work consisted in identifying the 18S, ITS1, 5.8S, ITS2 and 28S regions of B. bassiana ribosomal DNA. The DNA sequences of the amplified regions showed that the 18S rDNA is the most conserved unit, with a high homology (99.5%) between the isolates studied, while the 3’ end of the 28S rDNA has a great variability, which makes it possible to differentiate the isolates. The PCR-RFLP method was used to monitor isolates of B. bassiana and distinguish them in a target pest, Lygus lineolaris. This method involved two main steps. First, PCR was used to amplify a region of the 28S gene of B. bassiana. Second, this PCR product was digested using restriction endonucleases, and the fragments produced were compared using gel electrophoresis. Because of the high specificity and sensitivity of PCR-RFLP, it was possible to discriminate between B. bassiana isolates using spores scraped from the surface of an infected insect as samples.

Natural enemies of balloon vine Cardiospermum grandiflorum (Sapindaceae) in Argentina and their potential use as biological control agents in South Africa

Exploratory field surveys of the natural enemies associated with balloon vine Cardiospermum grandiflorum Swartz, an environmental weed in South Africa, Australia and other countries, were conducted in northern Argentina from 2005 to 2009. The surveys included other plant species in the genus Cardiospermum and other native Sapindaceae, permitting an assessment of the distribution and host range of the natural enemies. Seventeen phytophagous insects in five orders and ten families, and two fungal pathogens were found. The nature of the potential agents’ damage, their field distribution and abundance, and the results of preliminary host-specificity testing indicated that the seedfeeding weevil Cissoanthonomus tuberculipennis (Coleoptera: Curculionidae) and the fruit-galling midge Contarinia sp. (Diptera: Cecidomyiidae) were the most promising biological control agents for C. grandiflorum outside of its native range.

Pathogenicity of Pochonia species on eggs of Meloidogyne javanica

Among fungi, species of the genus Pochonia Batista & O.M. Fonseca are considered as promising biological control agents with high potential to reduce root-knot nematode (RKN) and nematode populations. In this research we investigated Fars province of Iran for the presence of Pochonia spp., compared pathogenicity of different Pochonia species on eggs of RKN in vitro, and selected the best isolates for further studies. During 2004–2006, 128 soil samples of fields infested with cyst nematodes and 18 soil samples infested with RKN were collected from Fars province of Iran. In vitro pathogenicity tests were carried out on 36 isolates of Pochonia spp. obtained from CBS and IRAN culture collections. The seven best isolates of this experiment were selected for greenhouse test and their ability in controlling RKN was examined in natural soil. In greenhouse test fresh weight of plant’s tops and roots, gall index, nematode multiplication, second-stage juveniles’ population in soil, reproduction rate ( P f/ P i), proportion of infected eggs, control efficacy, root colonization and soil colony forming units were determined. In vitro pathogenicity of Pochonia on RKN eggs varied between 39% and 95% eggs infected. In greenhouse experiment, three isolates are promising for control of RKN and selected isolates are subjected to more extensive testing to determine their effectiveness in a range of conditions before being developed as commercial biological control agents.