The generalist predatory mite Anystis baccarum (Acari: Anystidae) as a new biocontrol agent for western flower thrips, Frankliniella occidentalis (Thysanoptera: Thripidae).

A generalist predatory mite, Anystis baccarum (L.), was evaluated as a biological control agent against western flower thrips (WFT), Frankliniella occidentalis (Pergande). Laboratory assays showed A. baccarum was able to kill a mean of five WFT adult females or nine WFT larvae in 24 h, out-performing both Neoseiulus cucumeris (Oudemans) and Amblyseius swirskii Athias-Henriot. Next, a greenhouse assay was conducted to assess the performance of A. baccarum on potted chrysanthemums, comparing their efficacy to that provided by N. cucumeris slow-release sachets which represented the commercial standard in Canada. A combined treatment which incorporated both predatory mite species was also included to assess compatibility and potential additive effects of using both species together for WFT management. Introduction of two A. baccarum per pot was as efficacious as 125 N. cucumeris in terms of WFT control; however, despite the lack of significance between the level of WFT control obtained in the single predatory species treatments and the combined treatment, only the combination treatment suppressed WFT populations to levels that were almost unchanged over 8 weeks. There was no significant difference between the number of N. cucumeris recovered from plants in the single-species and the combination treatments, demonstrating the functional compatibility of the two predators. Additionally, WFT feeding damage was significantly lower on the A. baccarum-treated plants than on the untreated control and the N. cucumeris treatment. This study, together with our development of a prototype mass rearing method, shows that A. baccarum could be successfully used as a biocontrol agent for WFT.

Efficacy of Anystis baccarum against Foxglove Aphids, Aulacorthum solani, in Laboratory and Small-Scale Greenhouse Trials.

A generalist predatory mite, Anystis baccarum (L.), has been identified as a key predator of small, soft-bodied pest species in various agroecosystems around the world. The foxglove aphid Aulacorthum solani (Keltenbach) is a new problematic pest in Canadian greenhouses. Laboratory colonies of A. baccarum were established and its predatory efficacy against A. solani was assessed. In laboratory trials, A. baccarum ate approximately one adult aphid or seven first instar aphids in 24 h. In a greenhouse bench trial on sweet peppers with the free-flying aphid parasitoid, Aphidius ervi Haliday, the population dynamics of A. solani in the presence or absence of A. baccarum was evaluated. Although the parasitoid alone successfully eradicated A. solani, when A. baccarum were present on the plants, the aphid population was eradicated more rapidly. Fruit yield was also 15% higher from plants where A. baccarum was released than the control (without A. baccarum). Furthermore, plants were naturally infested by Frankliniella occidentalis (Pergande) during the trial, which caused visible feeding damage to the fruits. Anystis baccarum also predates on thrips and thrips' feeding damage to the fruits was reduced on plants where A. baccarum was released. Anystis baccarum was able to establish in sweet peppers and was determined to be complementary to the current practice of using A. ervi for the biological control of A. solani.

Risk of Spreading Soft Rot Through Cutting Dips Against Whiteflies in Greenhouse-Grown Poinsettia.

Dipping is a quick and cost-effective technique to reduce pest infestations on unrooted cuttings of greenhouse ornamental crops. Large numbers of cuttings are immersed in an insecticidal treatment, e.g., biopesticides and/or insecticidal soap, before they are stuck in the growing medium and rooted. This research investigated the risk of cross-contamination of poinsettia cuttings with Pectobacterium carotovorum subsp. carotovorum, a potentially devastating pathogen causing soft rot, through the dipping process. Sampling at four commercial greenhouses showed that P. carotovorum subsp. carotovorum was present in all dip suspensions during and after processing poinsettia cuttings; concentrations up to 1 × 105 CFU/ml were detected. A laboratory experiment determined that P. carotovorum subsp. carotovorum-infected cuttings could contaminate clean dip suspensions to similar levels. These results indicated that there is potential for disease transfer during dipping. The potential for cross-contamination of healthy cuttings was evaluated by immersing poinsettia cuttings in dip suspensions artificially inoculated with P. carotovorum subsp. carotovorum (from 1 × 103 to 1 × 107 CFU/ml). Disease incidence increased as P. carotovorum subsp. carotovorum concentrations in the dip suspension increased and the variety 'Prestige Red' was more susceptible than 'Freedom White.' However, even at the highest P. carotovorum subsp. carotovorum concentration of 1 × 107 CFU/ml, the proportion of diseased cuttings was low at 6% for var. 'Freedom White,' but higher at 21% for var. 'Prestige Red.' We conclude that P. carotovorum subsp. carotovorum transfer among unrooted poinsettia cuttings through the dipping process is relatively low although some varieties are sensitive to high levels of inoculum. Even so, strict sanitation practices are still important to prevent build-up of inoculum in the dip treatment.

Assessment of Yersinia entomophaga as a control agent of the diamondback moth Plutella xylostella.

The application of the biocontrol bacterium Yersinia entomophaga as a foliar spray was assessed for its efficacy against larvae of the diamondback moth, Plutella xylostella. The bacterium was applied as either a broth suspension, or as a biopolymer-based gel foliar spray and compared with commercial insecticides Dipel (Bacillus thuringiensis) and Spinosad. The performance of Y. entomophaga was comparable with that of Dipel. The gel-based formulation extended leaf persistence over that of the basic broth culture spray, while also providing higher initial foliar deposition rates. The bacterium was found to multiply within the P. xylostella larvae to 5.8 × 105 cells per larva, while the median lethal dose (LD50) was determined to be 2.69 × 103 cells per larva. Importantly, B. thuringiensis Cry1A-resistant, Cry1C-resistant, indoxacarb/pyrethroid-resistant, and Spinosad-resistant P. xylostella larvae were susceptible to Y. entomophaga.

Integration of microbial biopesticides in greenhouse floriculture: The Canadian experience.

Historically, greenhouse floriculture has relied on synthetic insecticides to meet its pest control needs. But, growers are increasingly faced with the loss or failure of synthetic chemical pesticides, declining access to new chemistries, stricter environmental/health and safety regulations, and the need to produce plants in a manner that meets the 'sustainability' demands of a consumer driven market. In Canada, reports of thrips resistance to spinosad (Success™) within 6-12 months of its registration prompted a radical change in pest management philosophy and approach. Faced with a lack of registered chemical alternatives, growers turned to biological control out of necessity. Biological control now forms the foundation for pest management programs in Canadian floriculture greenhouses. Success in a biocontrol program is rarely achieved through the use of a single agent, though. Rather, it is realized through the concurrent use of biological, cultural and other strategies within an integrated plant production system. Microbial insecticides can play a critical supporting role in biologically-based integrated pest management (IPM) programs. They have unique modes of action and are active against a range of challenging pests. As commercial microbial insecticides have come to market, research to generate efficacy data has assisted their registration in Canada, and the development and adaptation of integrated programs has promoted uptake by floriculture growers. This review documents some of the work done to integrate microbial insecticides into chrysanthemum and poinsettia production systems, outlines current use practices, and identifies opportunities to improve efficacy in Canadian floriculture crops.

Non-host larvae negatively impact persistence of the entomopathogen Beauveria bassiana in soil.

A better understanding of the ecology of the insect pathogenic fungus, Beauveria bassiana, in soil is needed to identify reasons behind the variable efficacy often seen after field application. A transformed strain of a candidate commercial strain of B. bassiana (F418 gfp tr3), expressing the green fluorescent protein and the hygromycin B resistance gene, was used to assess the effects of the larvae of a host insect, Tenebrio molitor L. (Coleoptera: Tenebrionidae), a non-host, Costelytra zealandica (Coleoptera: Scarabaeidae) and the absence of larvae on the persistence of F418 gfp tr3 in pasteurised and non-sterile soil over 4 months. In the presence of a T. molitor larvae, F418 gfp tr3 populations increased significantly in pasteurised and non-sterile soil; however, populations increased less in non-sterile soil than in pasteurised soil. Lower populations of F418 gfp tr3 were recovered in pasteurised soil in the presence of C. zealandica larvae than in pasteurised soil without larvae. No difference was observed between F418 gfp tr3 populations in non-sterile soil with a non-host larvae or without larvae. Accompanying studies showed that F418 gfp tr3 conidia germinated and produced appressoria on live and excised cuticle of non-host (C. zealandica) larvae but infection did not occur, leading to a net loss of viable conidia in the soil. Conidia administrated orally to C. zealandica larvae were viable on recovery from faecal samples, suggesting that ingestion of the fungus by the larvae had little impact on the viable fungal population. Soil bacterial and fungal community patterns were analysed using Single-Strand Conformation Polymorphism (SSCP) and showed a correlation between changes in F418 gfp tr3 persistence in pasteurised and non-sterile soil and changes in soil communities in the presence of a host insect, non-host insect or in the absence of insect. In pasteurised soil, non-specific germination of F418 gfp tr3 conidia on the non-host larval cuticle and the presence of antagonistic bacteria introduced with the field-collected larvae are most likely responsible for the differences observed. The more complex microbial community structures in non-sterile soil could lead to fungistasis, preventing potentially antagonistic bacteria degrading conidia or inhibiting attachment and germination on the non-host larval cuticle, resulting in the observed lack of difference between non-host and no larval treatments.

How to Start with a Clean Crop: Biopesticide Dips Reduce Populations of Bemisia tabaci (Hemiptera: Aleyrodidae) on Greenhouse Poinsettia Propagative Cuttings.

(1) Global movement of propagative plant material is a major pathway for introduction of Bemisia tabaci (Hemiptera: Aleyrodidae) into poinsettia greenhouses. Starting a poinsettia crop with high pest numbers disrupts otherwise successful biological control programs and widespread resistance of B. tabaci against pesticides is limiting growers' options to control this pest; (2) This study investigated the use of several biopesticides (mineral oil, insecticidal soap, Beauveria bassiana, Isaria fumosorosea, Steinernema feltiae) and combinations of these products as immersion treatments (cutting dips) to control B. tabaci on poinsettia cuttings. In addition, phytotoxicity risks of these treatments on poinsettia cuttings, and effects of treatment residues on mortality of commercial whitefly parasitoids (Eretmocerus eremicus and Encarsia formosa) were determined; (3) Mineral oil (0.1% v/v) and insecticidal soap (0.5%) + B. bassiana (1.25 g/L) were the most effective treatments; only 31% and 29%, respectively, of the treated B. tabaci survived on infested poinsettia cuttings and B. tabaci populations were lowest in these treatments after eight weeks. Phytotoxicity risks of these treatments were acceptable, and dip residues had little effect on survival of either parasitoid, and are considered highly compatible; (4) Use of poinsettia cutting dips will allow growers to knock-down B. tabaci populations to a point where they can be managed successfully thereafter with existing biocontrol strategies.

Effect of Temperature and Host Life Stage on Efficacy of Soil Entomopathogens Against the Swede Midge (Diptera: Cecidomyiidae).

The Swede midge, Contarinia nasturtii Kieffer, is an economically significant pest of cruciferous crops in Canada and the northeastern United States. The effect of temperature on the virulence of three entomopathogenic nematode species, Heterorhabditis bacteriophora, Steinernema carpocapsae, and Steinernema feltiae, the entomopathogenic fungus Metarhizium brunneum, and a H. bacteriophora+M. brunneum combination treatment to C. nasturtii larvae, pupae, and cocoons was investigated. In the laboratory, all three nematode species successfully reproduced inside C. nasturtii larvae: H. bacteriophora produced the highest number of infective juveniles per larva, followed by S. carpocapsae and S. feltiae. H. bacteriophora and the H. bacteriophora+M. brunneum combination treatment generally caused the highest mortality levels to all C. nasturtii life stages at 20°C and 25°C, whereas S. feltiae caused the highest mortality to larvae and pupae at 16°C. No nematode species caused significant mortality when applied in foliar treatments to the infested host plant meristem and, in spite of high mortality, an antagonistic interaction was observed in the H. bacteriophora+M. brunneum combination treatment when compared with expected mortality. In trials conducted in broccoli fields in Elora, Ontario, M. brunneum suppressed adult emergence of C. nasturtii from infested soil in 2012 and all nematode treatments successfully suppressed adult emergence in 2013; however, no significant effects were observed in field trials conducted in Baden, Ontario.

Association of entomopathogenic fungi with exotic bark beetles in New Zealand pine plantations.

Hylastes ater and Hylurgus ligniperda are introduced pests of re-established Pinus radiata in New Zealand. Both species breed under the bark of stumps in recently harvested areas. Adult maturation feeding on pine seedlings planted in adjacent areas can significantly impact seedling growth, and in severe cases seedlings will die. Entomopathogenic fungi are important natural mortality factors in bark beetle populations, and Beauveria spp. are predominant. Here, we report on the isolation of other fungal species from H. ater in New Zealand. Based on morphological characteristics and sequencing data, two species, Metarhizium flavoviride var. pemphigi and Hirsutella guignardii, were recovered from H. ater. Both are new records for New Zealand and appear to be the first records of these species from bark beetles worldwide.

Field trials to evaluate effects of Bt-transgenic silage corn expressing the Cry1Ab insecticidal toxin on non-target soil arthropods in northern New England, USA.

Traditionally, control of European corn borer (Ostrinia nubilalis) Hübner has been achieved through the use of chemical insecticides. With increasing emphasis on reducing pesticide inputs in agricultural production, alternative management technologies are now being used including transgenic silage corn modified to express Cry1Ab protein toxins derived from Bacillus thuringiensis (Bt) Berliner. The Cry1Ab toxin is expressed by all plant cells and throughout the growing season. Furthermore, the toxins are exuded from corn plant roots into the rhizosphere, raising concerns over possible side-effects on non-target beneficial organisms in the same habitat. In addition, detrivores are exposed to crop residues containing the toxin when incorporated into the soil. The current 2-year study (2003, 2004) evaluated effects of two silage-corn varieties: Pioneer var. 38A25 (Bt-corn expressing the Cry1Ab toxin) and Pioneer var. 38A24 (parent isoline) on species diversity and evenness of carabid beetles and Collembola. Pitfall traps were used to collect surface-dwelling species on a bimonthly schedule from April to October. Soil cores were taken once a month from April to October to sample subterranean species, which were extracted using Berlese funnels. All individuals were recorded and identified where possible to species level for analysis in the Simpson's D and Shannon-Wiener H' diversity indices. Evenness was measured using Simpson's E', after which dominant species were analyzed in a multivariate ordination analysis. Results showed Bt-corn had no negative effects on any of the organisms analyzed. There was a significant year effect on the abundance of surface-dwelling Collembola and on species diversity of soil-dwelling Collembola. Our findings suggest that crop management practices and/or environmental conditions (e.g., heavy rainfall during the 2004 growing season) had the greatest impact on species diversity and evenness, rather than the crop itself (Bt or isoline).

Whey for mass production of Beauveria bassiana and Metarhizium anisopliae.

Spore production of Beauveria bassiana and Metarhizium anisopliae was studied in a novel whey-based culture media. Spore yield and viability were determined for two B. bassiana (GHA-726 and CA-603) and two M. anisopliae (CA-1 and IMI 330189) isolates following production in three whey-based systems: solid, liquid, and a diphasic production system. Our study indicated that whey permeate can be used effectively for production of spores of entomopathogenic fungi. However, spore yield and viability were significantly influenced by fungal isolate, whey concentration, and the type of production process used. Under the conditions defined in the present study, spore yields ranging from 1.3x10(9)-10x10(11) spores l(-1) of whey medium could be obtained depending on the strain and production process used. Our study revealed that spores produced by all strains in whey-based solid and liquid media showed between 73-99% viability; germination rates were comparable with those obtained using the standard SDA medium. In the two-stage production process, the viabilities of conidia produced by GHA-726, CA-603, and CA-1 were 35-86, 32-98, and 6-29%, respectively; viability was correlated with whey concentration and isolates. Whey permeate can be used as a growth substrate for mass production of biocontrol fungi. We hypothesize that spore yield and viability could be improved by careful selection of whey content in the medium, incorporation of critical additives and optimization of culture conditions.

Effects of tobacco genetically modified to express protease inhibitor bovine spleen trypsin inhibitor on non-target soil organisms.

Effects of tobacco genetically modified to express the protease inhibitor bovine spleen trypsin inhibitor (BSTI) were examined in laboratory assays against three earthworm and one collembolan species. BSTI is a serine protease inhibitor that can bind to the digestive trypsins of insects feeding on modified plants, resulting in reduced growth and survival. Protease inhibitors are active against a broad range of insects, so may have a large impact on non-target organisms. Survival and fecundity of the collembolan Folsomia candida were unaffected by consumption of artificial diet containing BSTI-expressing tobacco leaf or powdered freeze-dried BSTI-expressing tobacco leaf that was added to soil. Similarly, mortality and growth of earthworms Aporrectodea caliginosa and Lumbricus rubellus did not differ significantly between soil augmented with BSTI-expressing tobacco leaves or unmodified control leaves. The redworm Eisenia fetida gained less weight when provided with BSTI-expressing leaves in one assay, but when the experiment was repeated, there was no significant difference between treatments. BSTI-expressing tobacco and unmodified control leaves decomposed at the same rate, indicating that the inhibitor had no effect on the overall function of the decomposer community of micro-flora and fauna in soil.

Development of a novel bioassay for estimation of median lethal concentrations (LC50) and doses (LD50) of the entomopathogenic fungus Beauveria bassiana, against western flower thrips, Frankliniella occidentalis.

To conduct laboratory experiments aimed at quantifying secondary acquisition of fungal conidia by western flower thrips (Frankliniella occidentalis), an efficient assay technique using Beauveria bassiana as the model fungus was developed. Various application protocols were tested and it was determined that the percent mortality did not vary among protocols. Peak mortality of second-instar nymphs, under constant exposure to conidia, occurred 5 days post-inoculation. Second-instar thrips that were exposed to conidia within 24 h of the molt to second instar were more susceptible to Beauveria bassiana than thrips exposed after times greater than 24 h post-molt. Conidia efficacy, which was monitored at 24 h intervals, did not differ significantly within 72 h. A test of the final bioassay system was conducted in a series of assays aimed at determining the LD50 of B. bassiana technical powder against second-instar western flower thrips. It was determined that B. bassiana (strain GHA) is highly effective at very low doses (LD50 of 33-66 conidia/insect).

Characterization of Beauveria bassiana and Metarhizium anisopliae isolates for management of tarnished plant bug, Lygus lineolaris (Hemiptera: Miridae).

Selected morphological and physiological characteristics of four Beauveria bassiana (Balsamo) Vuillemin isolates and one Metarhizium anisopliae (Metschnikoff) Sorokin isolate, which are highly pathogenic to Lygus lineolaris (Palisot de Beauvois) (Hemiptera: Miridae), were determined. There were significant differences in conidial size, viability, spore production, speed of germination, relative hyphal growth, and temperature sensitivity. Spore viability after incubation for 24h at 20 degrees C ranged from 91.4 to 98.6% for the five isolates tested. Spore production on quarter-strength Sabouraud dextrose agar plus 0.25% (w/v) yeast extract after 10 days incubation at 20 degrees C ranged from 1.6x10(6) to 15.5x10(6)conidia/cm(2). One B. bassiana isolate (ARSEF 1394) produced significantly more conidia than the others. Spore germination was temperature-dependant for both B. bassiana and M. anisopliae. The time required for 50% germination (TG(50)) ranged from 25.0 to 30.9, 14.0 to 16.6, and 14.8 to 18.0h at 15, 22, and 28 degrees C, respectively. Only the M. anisopliae isolate (ARSEF 3540) had significant spore germination at 35 degrees C with a TG(50) of 11.8h. A destructive sampling method was used to measure the relative hyphal growth rate among isolates. Exposure to high temperature (40-50 degrees C) for 10min had a negative effect on conidial viability. The importance of these characteristics in selecting fungal isolates for management of L. lineolaris is discussed.

Pathogenicity of Beauveria bassiana, Metarhizium anisopliae (Deuteromycotina: Hyphomycetes), and other entomopathogenic fungi against Lygus lineolaris (Hemiptera: Miridae).

The pathogenicity of 32 fungal isolates from the genera of Beauveria, Verticillium, Paecilomyces, Metarhizium, Mariannaea, and Hirsutella to second-instar tarnished plant bug, Lygus lineolaris (Palisot de Beauvois), was tested under laboratory conditions. These isolates originated from various insect hosts and substrates from France, Denmark, Austria, Italy, Turkey, Syria, and the United States. A single exposure concentration (1 x 10(7) conidia/ ml) assay for each isolate was first conducted by immersing the insects in 10 ml of a fungal suspension for 5s. These were followed by concentration-mortality assays on five of the most pathogenic isolates using four test concentrations ranging from 2 x 10(4) to 2 x 10(7) conidia/ml. B. bassiana 726 (Bemisia-passaged GHA strain) was used as a standard for comparison in all of the assays. Among the test isolates, three produced mortality not significantly different from the water control. Mortality ranged from 35 to 98% among the other 29 isolates. The LC50 values of the five most pathogenic isolates ranged from 0.8 to 5.0 x 10(5) conidia/ml. The LT50 values for these isolates ranged from 6.0 to 6.9, 3.1 to 5.1, and 2.5 to 4.0 d for concentrations of 2 x 10(5), 2 x 10(6), and 2 x 10(7) conidia/ml, respectively. Two strains of B. bassiana (ARSEF 1394,5665) and one M anisopliae (ARSEF 3540) were more pathogenic to the nymphs than the standard, having significantly lower LC50 and LT50, values. Our results demonstrated that several genera of entomopathogenic fungi have promise as microbial control agents against L. lineolaris.