Horizontal Transmission of the Entomopathogenic Fungus Beauveria bassiana among the Spruce Bark Beetle, Ips typographus (Col., Scolytidae) in the Laboratory and under Field Conditions

In laboratory and field cage experiments, the horizontal transmission of the entomopathogenic fungus Beauveria bassiana (Boverol®) between adults of the spruce bark beetle Ips typographus was examined. In a laboratory experiment, conidia transfer between treated and untreated beetles at different ratios resulted in 96% mortality (MST 4.3 days) at a ratio of 1:1 and 90% mortality (MST 4.5 days) at a ratio of 1:2. At ratios of 1:5, 1:10 and 1:20, the mortalities at 7 days were 83, 77 and 75%, respectively. In a second laboratory experiment, the efficacy of B. bassiana was investigated after transmission from contaminated to healthy beetles over a period of 5 days. Two weeks later, the mortality of treated and untreated beetles was 99% (control mortality was 44%), while the rate of mycosis was 79% in the treatment compared to 10% in the control beetles. A single contact between one treated male (2.0×10⁵ conidia/beetle) and an untreated female was found to be sufficient to transmit a lethal dose of B. bassiana of about 1.2×10⁴ conidia. Two experiments in field cages in a spruce stand demonstrated horizontal transmission of B. bassiana (Boverol®) between beetles. In the first experiment the transmission was investigated following the introduction of B. bassiana-inoculated beetles into a population of untreated beetles. Significant reductions were observed in the length of maternal galleries, and the number of larvae and pupae. In the second field cage experiment, natural populations of beetles were lured into a flight cage using a pheromone trap, and treated with B. bassiana or left untreated on alternate days. In this experiment, significant reductions were observed in the number of bore holes and the length of maternal galleries, while no larvae, pupae or juveniles were found under the bark of spruce trunks in the Boverol® transmission treatment. It is concluded that B. bassiana has potential for control of I. typographus, and further field experiments are warranted to investigate its efficacy under more practical conditions.     

Effect of Plant Type on the Persistence of Beauveria bassiana

Field evaluation of isolate MK 2001 of the entomopathogenic fungus Beauveria bassiana (Balsamo) Vuillemin, revealed conidia persistence and infectivity up to 26 days on foliage of lettuce and celery, two crops with substantial plant differences. Plants were treated at a rate of 100 ml/cage at a concentration of 1×10⁸ conidia/ml. The number of colony-forming units (CFU) recovered immediately after treatments from discs sampled on lettuce and celery leaves were not significantly different, due to similar foliar coverage of conidia by application. Significant differences were observed between both host plants during the 26 days of the field trial. The number of CFUs recovered on lettuce was significantly higher than that on celery leaves. However, for each host plant, there were no significant differences either between lettuce external leaves and internal leaves or between celery canopy and bottom leaves. An in vitro pathogenicity test carried out on Lygus lineolaris adults fed with leaf discs harvested from treated cages resulted in a high pathogenicity of B. bassiana isolate MK 2001. The mortality immediately after treatment did not differ from the death counts taken every other day for 26 days. However, efficacy was significantly different between both plants. Mortality was 91% on lettuce and 78% on celery, 7 days post-treatment. This study highlighted that plant type must be taken into account in foliar application of B. bassiana.     

Autodissemination of Beauveria bassiana by Sap Beetles (Coleoptera: Nitidulidae) to Overwintering Sites

An autoinoculative device was used to test the ability of sap beetles (Coleoptera: Nitidulidae) to carry a specific strain of Beauveria bassiana (Balsamo) Vuillemin to overwintering sites in a multiyear field study. The device was baited with the pheromone and coattractants for the dusky sap beetle (Carpophilus lugubris Murray) and placed in the field in the fall of each year. The introduced strain occurred at high frequency among the B. bassiana isolated in the fall of all four years tested (100% of all isolates from 21 of 22 collection dates). The introduced strain of B. bassiana was isolated at high frequency from all the B. bassiana-contaminated sap beetles recovered from the overwintering traps (100% of all isolates from 13 of 23 trap/beetle species combinations) and was highest after the longest fall exposure. The introduced strain was primarily isolated from C. lugubris and C. antiquus, but species distribution was also dependent on the overwintering trap design used. Few non-sap beetles species of insects were recovered from the artificial overwintering sites. Although B. bassiana was isolated from free flying sap beetles caught in traps in the spring of each year, none were infected with the introduced strain. The autoinoculating device provides selective contamination of sap beetles in overwintering sites when used in the fall. It may be useful in providing some control of sap beetles or other insects where limited numbers of mass overwintering sites (such as tree holes) occur.     

Laboratory and glasshouse evaluation of entomopathogenic fungi against the two-spotted spider mite, Tetranychus urticae (Acari: Tetranychidae), on tomato, Lycopersicon esculentum

Forty isolates of entomopathogenic fungi from six genera were assessed against the two-spotted spider mite, Tetranychus urticae, in a single dose, direct application laboratory bioassay on tomato leaflets. Only three isolates caused greater mortality than the control: these were Metarhizium anisopliae 442.99, Hirsutella spp. 457.99, and Verticillium lecanii 450.99. These isolates were assessed in a multiple dose bioassay, together with three isolates cultured from commercial biopesticides as follows: Beauveria bassiana 432.99 (cultured from 'Naturalis-L', Troy Biosciences, Phoenix, TX, USA); Hirsutella thompsonii 463.99 (cultured from 'Mycar', Abbott Laboratories USA); and V. lecanii 19.79 (used in 'Mycotal' Koppert BV, The Netherlands). Beauveria bassiana 432.99, H. thompsonii 463.99, M. anisopliae 442.99, and V. lecanii 450.99 were all pathogenic to T. urticae in this bioassay. In addition, it was found that the mortality caused by B. bassiana 432.99 and Naturalis-L was increased when the mites were exposed to tomato leaflets sprayed previously with conidia suspensions, compared to spraying the mites directly. In a glasshouse experiment, sprays of B. bassiana 432.99, H. thompsonii 463.99, M. anisopliae 442.99, V. lecanii 450.99 and Naturalis-L reduced T. urticae populations in a tomato crop grown according to commercial practice. Naturalis-L reduced T. urticae numbers by up to 97%. In a second glasshouse experiment, single sprays of Naturalis-L and the acaricide fenbutatin oxide (Torq) were compared as supplementary treatments to release of the predatory mite, Phytoseiulus persimilis. Supplementary sprays of fenbutatin oxide reduced the numbers of T. urticae nymphs (80% reduction), while Naturalis-L reduced numbers of T. urticae adults, nymphs and eggs (98% reduction in all three cases). It is concluded that Naturalis-L has the potential to be used against T. urticae on glasshouse tomato crops.     

Honey bee dispersal of biocontrol agents: An evaluation of dispensing devices

Many reports have been published lately on the use of honey bees to disseminate biocontrol agents to flowers, using various kinds of inoculum dispensers. This technique is mainly used to deliver agents for controlling fire blight and gray mold. The present study evaluated the relative efficiency of a newly developed 'Triwaks' dispenser with three other dispenser types, 'Peng', 'Tub', and 'Harwood', with respect to effects of the dispensers on bee activity, and effectiveness for contaminating bees with high levels of inoculum for a long period of time after its application. We found differences in these parameters of performance between dispenser types, with the Triwaks dispenser having the overall best performance. This dispenser could prove to be an effective tool for biocontrol. The evaluation methods that we present could be used for evaluating other dispenser types.     

Do aphids infected with entomopathogenic fungi continue to produce and respond to alarm pheromone?

The response of pea aphids, Acyrthosiphon pisum, to aphid alarm pheromone was not modified by infection with Beauveria bassiana. Approximately 50% of uninfected and infected aphids responded to synthetic alarm pheromone. The simulated attack of aphids infected with B. bassiana did not elicit a response in uninfected aphids. Preliminary air entrainment experiments of both uninfected aphids and aphids at different stages of B. bassiana (generalist pathogen) or P. neoaphidis (obligate pathogen of aphids) demonstrated that B. bassiana infected aphids produced less alarm pheromone than uninfected aphids and, conversely, P. neoaphidis infected aphids produced more alarm pheromone than uninfected aphids. These results are discussed with particular emphasis on the different life history strategies of these two pathogens. We hypothesise that the obligate, specialist pathogen, P. neoaphidis, is under greater selection pressure to increase pathogen transmission and survival resulting in modified host behaviour, than the generalist pathogen, B. bassiana.     

Microencapsulation: a Strategy for Formulation of Inoculum

A non-toxic phase separation method was developed for microencapsulation of inoculum used in biological control. Aqueous sodium alginate or gelatin and agar was mixed with inocula of various biopesticides and emulsified in a mixture of corn oil, n-hexadecane, and lecithin. Gelatin and agar globules gelled in the emulsion; alginate globules gelled after settling into a lower phase of aqueous CaCl₂. A layer of gelatinous material thus surrounded the inoculum as 'capsules'. Mixing with n-hexadecane reduced the specific gravity and surface tension of the oil, allowing aqueous extraction of the capsules. Successful extraction of alginate capsules depended upon lecithin (>0.17%), n-hexadecane (>30%), and CaCl₂ (>0.01 M) concentrations. Alginate-encapsulated macroconidia of Fusarium avenaceum caused 23±3% leaf area damage to seedlings of marsh reed grass, versus 4±3% for unformulated controls. In green foxtail seedlings, gelatin and agar-encapsulated conidia of Bipolaris sorokiniana caused 21.3 vs. 7.9 lesions per plant for encapsulated versus unformulated conidia. Mortality of Douglas-fir tussock moth larvae caused by a nuclear polyhedrosis virus was delayed when 23 polyhedral inclusion bodies (PIB) were incorporated into alginate capsules, but it proceeded normally for 2.3 PIB/capsule, where efficacy was also higher versus positive controls. Microencapsulation enhances the activity of biological control agents and protects them from adverse conditions.     

小菜蛾应对球孢白僵菌感染的免疫应答基因分析

通过室内生物测定,筛选对小菜蛾Plutella xylostella具有高毒力的球孢白僵菌Beauveria bassiana菌株,分析小菜蛾应对球孢白僵菌浸染的免疫应答基因。采用新一代Illumina高通量测序技术对接种/感染48h与未接种的小菜蛾样品分别进行转录组测序,筛选差异表达基因;结合生物信息学分析差异表达基因的功能、分类及涉及的信号通路等。结果表明接种小菜蛾48h后诱导2 434个基因发生差异表达,包括1 080个上调基因和1 354个下调基因。GO分类结果表明,这些差异表达基因（DEGs）被注释到45个GO term中,包括23个生物学过程,12个细胞组分和10个分子功能。KEGG分析表明,1 308个DEGs被富集到25个功能途径,这些DEGs包括497个上调基因和811个下调基因。分析发现,DEGs编码蛋白包含肽聚糖识别蛋白、丝氨酸蛋白酶55、丝氨酸蛋白酶抑制剂以及细胞色素P450 6K1类的免疫相关蛋白。另外,组蛋白、磷酸泛酰半胱氨酸脱羧酶、双链RNA结合蛋白、黑芥子酶等基因在球孢白僵菌侵染过程中也高表达,推测这些基因可能参与小菜蛾应对球孢白僵菌侵染的免疫反应。研究结果为进一步研究小菜蛾免疫相关基因的功能奠定了基础。     

Interaction Between the Entomopathogens Beauveria bassiana, Metarhizium anisopliae and Paecilomyces fumosoroseus and the Mycoparasites Clonostachys spp., Trichoderma harzianum and Lecanicillium lecanii

Biocontrol agents of numerous insect pests and fungal pathogens exist but virtually nothing is known about their interaction if used simultaneously. Our objective was to investigate the compatibility of the entomopathogens Beauveria bassiana, Metarhizium anisopliae and Paecilomyces fumosoroseus, and the broad host-range mycoparasites Clonostachys spp., Trichoderma harzianum and Lecanicillium lecanii. In vitro host-range tests revealed that M. anisopliae was highly susceptible to all mycoparasites tested. B. bassiana was attacked by Clonostachys rosea, and P. fumosoroseus. was resistant to mycoparasites. M. anisopliae but not P. fumosoroseus killed nymphs of Bemisia tabaci in bioassays. B. bassiana and M. anisopliae proved lethal to Cosmopolites sordidus, Diatraea saccharalis and Sitophilus oryzae. Coapplication of mycoparasites with entomopathogens did not affect their biocontrol efficacy in vivo, although the reisolation success of entomopathogens could be significantly reduced, especially from smaller insect species. Trichoderma spp. were reisolated from mycoparasite-treated insects more frequently than C. rosea. The coapplication of the highly susceptible M. anisopliae generally enhanced mycoparasite recovery. Mycofungicide preparations caused some insect mortality but less than a copper hydroxide fungicide which is still permissible in organic agriculture. We concluded that the tested entomopathogens and mycoparasites are compatible elements of integrated pest management.     

Beauveria bassiana for the control of Sunn Pest (Eurygaster integriceps) (Hemiptera: Scutelleridae) and aspects of the insect's daily activity relevant to a mycoinsecticide

A series of investigations was carried out at ICARDA during April-June 2004 and May-June 2005 to investigate the use of the entomopathogenic fungus Beauveria bassiana to control Eurygaster integriceps and to determine aspects of the insect's biology that could be relevant to control with the fungus. Application in 2004 of an oil-based formulation of B. bassiana showed distribution of the spray decreasing from the top of the wheat to the bottom, although the proportion of tillers with coverage at the base was still 80%. Seventy percent of Sunn Pest had received direct hits, with the highest amount on the dorsal surface, followed by the ventral surface, then the legs and antennae. Mortality of field-treated, then laboratory-maintained insects, ranged from 47.6% for insects exposed only to the B. bassiana spray, 74.2% for unsprayed insects exposed to treated wheat and 86.4% for insects receiving direct spray and being exposed to treated wheat. Some field results demonstrated mortality. Mortality assessments following the 2005 treatments were conducted solely in the field, there was no apparent effect upon Sunn Pest numbers. The majority of studies on Sunn Pest biology were carried out on untreated insects outside the trial plots. Different assessment methods resulted in varying estimates of adults and juveniles in the field. The numbers of Sunn Pest found and the proportion of these at the top of the wheat crop, declined after early morning and increased again in the early evening, suggesting sheltering deeper in the crop or in the soil during the middle part of the day. Mobility and location of marked insects was followed in micro-plots and by mark and recapture studies, demonstrating a range of activities, from movement around the crop for a number of metres, including contact with numerous tillers to almost total immobility; Sunn Pest generally avoided movement on the soil. The potential for Sunn Pest control with a mycoinsecticide and how Sunn Pest behaviour is relevant to a mycoinsecticide application are discussed.     

Laboratory investigations on the potential of entomopathogenic fungi for biocontrol of Helicoverpa armigera (Lepidoptera: Noctuidae) larvae and pupae

The susceptibility of third instar Helicoverpa armigera to seven strains of three entomopathogenic fungal species, i.e. Metarhizium anisopliae, Beauveria bassiana and Paecilomyces fumosoroseus, was tested under laboratory conditions using the larval immersion method. High efficacies ranging from 68 to 100% corrected mortality were recorded with more profound effects in treatments with B. bassiana and P. fumosoroseus strains. The median lethal concentration (LC₅₀) for L3 was 6.0×10⁵ in M. anisopliae 79, 1.5×10⁵ in B. bassiana 124 and 4.2×10⁴ in P. fumosoroseus 14. These three strains were further used to characterize the age-dependent mortality of different larval stages (L2-L5) and the effect against pupae of H. armigera. Larval stages did not differ in their mortality but differed i in median lethal time, with shorter values recorded in the second instar. Tested fungi also caused a high reduction between 74.4 and 100% in the emergence of pupae using the soil inoculation method and the pupal immersion technique. All three fungal species, especially P. fumosoroseus, have a high potential for biocontrol of H. armigera larvae and also as a soil treatment targeting the pupae.     

Characterization of Growth and Conidia Production of Exserohilum monoceras on Different Substrates

On agar media the maximum conidia production of Exserohilum monoceras occurred on V-8 juice agar (VA) or centrifuged V-8 juice agar, whereas the optimal radial mycelial growth occurred on Czapek-Dox agar. The optimal temperatures for radial mycelial growth and conidia production were 28 and 27°C respectively. Light prohibited E. monoceras conidia production. The best sporulation occurred under continuous dark conditions. Echinochloa leaf decoction significantly increased conidia production on potato dextrose agar (PDA) and VA, and significantly increased germ tube length on PDA, lima bean agar and VA, but did not affect conidia germination. No conidia were produced in liquid media. Of 22 agricultural-based products evaluated as solid substrates, the most abundant sporulation (1.8 × 10⁶ conidia g^-1 of dry weight) occurred on corn leaves. The conidia production of E. monoceras on corn leaves was affected by incubation period, moisture content and substrate quantity. There were no differences in germination rate, germ tube length and virulence of conidia produced on agar media or corn leaves.     

A Stilbene Optical Brightener can Enhance Bacterial Pathogenicity to Gypsy Moth (Lepidoptera: Lymantriidae) and Colorado Potato Beetle (Coleoptera: Chrysomelidae)

Stilbene optical brighteners were first investigated to protect biological control agents such as viruses, fungi, and nematodes against ultraviolet light. Some are known to enhance the activity of insect viruses in Lepidoptera. In this work, one stilbene brightener, Tinopal LPW, also increased mortality of gypsy moth and Colorado potato beetle larvae when treated with bacteria/optical brightener combinations. This increase in mortality, however, did not occur for every bacteria/insect combination. In gypsy moth, a significant increase in larval mortality was observed only with Bacillus thuringiensis combined with Tinopal LPW. In Colorado potato beetle, however, the addition of Tinopal LPW increased larval mortality with all bacteria tested (B. thuringiensis, Serratia marcescens, Photorhabdus luminescens, and Chromobacterium sp.). The brightener also decreased the time to kill for these pathogens. This decrease in LT₅₀ was observed not only for bacteria+Tinopal LPW combinations, but also for combinations of Chromobacterium sp. toxin+Tinopal LPW. The mechanism for increase in bacterial toxicity by optical brighteners is compatible with mechanisms proposed for enhancement based on viral/lepidopteran/optical brightener systems that are not dependent on replication.     

Effect of combination treatment with entomopathogenic fungi Beauveria bassiana and Nomuraea rileyi (Hypocreales) on Spodoptera litura (Lepidoptera: Noctuidaeae)

In biocontrol of insect pests, efficacy of treatment with multiple pathogens has not been frequently investigated but may have potential for effective management. The possible advantage of a combination treatment with two entomopathogenic fungi - Beauveria bassiana and Nomuraea rileyi - was assessed in laboratory bioassays on second instar Spodoptera litura. From among the fungal isolates of an epizootic population, two isolates of each fungus differing in virulence to S. litura were chosen, one highly virulent and the other with low virulence. The bioassays were carried out at either a continuous temperature of 25±1°C or at a temperature cycle of 32±2°C 8 h/21±2°C 16 h to mimic the field temperatures during the epizootic. Treatments with the two fungi were done both simultaneously and sequentially. In combination treatments at 25±1°C, in all isolate combinations, a majority of the larvae showed N. rileyi induced mycosis; the percentage mortality and speed of kill of insects in these treatments was similar to the N. rileyi isolate used in the combination treatments. At the temperature cycle of 32±2°C 8 h/21±2°C 16 h, in all combination treatments, all the dead insects exhibited B. bassiana mycosis; the mortality pattern was similar to the B. bassiana isolate used in the combination treatments. The adverse effect of high temperature on virulence of N. rileyi was however, not evident in in vitro growth assays. Combination treatment with both fungi did not have a synergistic effect on insect mortality.     

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 1.3 × 10⁹–10 × 10¹¹ 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.     

Natural occurrence of Bacillus thuringiensis and Bacillus cereus in eukaryotic organisms: a case for symbiosis

Bacillus thuringiensis and Bacillus cereus, two members of the Bacillus cereus sensu lato group, are most noted for their virulence in, respectively, arthropods and mammals including humans. Because of their pathogenicity to insects in particular, and their narrow host range, strains of B. thuringiensis have been utilised successfully as biocontrol agents of insect pests. Whereas B. cereus is not an established entomopathogen, certain strains of this species are well known to be etiological agents of gastrointestinal and emetic syndromes in humans. While much is known about the taxonomic properties and molecular basis for virulence of B. thuringiensis and B. cereus, comparatively less is known about their ecology in natural environments. Thus, there are limited data regarding their resilience, i.e. recycling of vegetative and sporulated phases of growth in soil, ecolgical niches including symbiotic interactions with other organisms, and the impact on ecosystems in which they proliferate. Nevertheless, based on recent data, a picture is beginning to emerge that B. thuringiensis and B. cereus are capable of establishing mutual and commensal relationships with both animals and plants. In this regard, these bacilli can proliferate in the digestive tracts of animals, where upon defecation they form dormant spores in the soil, and to a lesser extent on the phylloplane and rhizospheres of plants. Altogether, current evidence strongly suggests that B. thuringiensis and B. cereus are capable of completing their life cycles and recycling through various reservoirs, including animals, plants, and soil. This review focuses on the current knowledge pertaining to the ecologic interactions between B. thuringiensis and B. cereus and eukaryotic hosts, with special emphasis on symbiosis.     

Microbial metabolism of 1-aminoanthracene by Beauveria bassiana

The carcinogen and mutagen, 1-aminoanthracene, was efficiently metabolized by the fungal strain Beauveria bassiana ATCC 7159 to yield three new metabolites identified as 1-acetamido-5-[(4′-O-methyl-β-d-glucopyranosyl)oxy]anthracene, 1-acetamido-8-[(4′-O-methyl-β-d-glucopyranosyl)oxy]anthraquinone, and 1-acetamido-6-[(4′-O-methyl-β-d-glucopyranosyl)oxy]anthraquinone, together with 1-acetamidoanthracene and 1-acetamidoanthraquinone. Formation of these metabolites suggests that the metabolic pathways of 1-aminoanthracene in B. bassiana ATCC 7159 involve acetylation, oxidation, hydroxylation, and O-methylglucosylation.     

Hirsutella thompsonii and Metarhizium anisopliae as potential microbial control agents of Varroa destructor,a honey bee parasite

The potential of Hirsutella thompsonii Fisher and Metarhizium anisopliae (Metschinkoff) as biological control agents of the parasitic mite, Varroa destructor Anderson and Trueman was evaluated in the laboratory and in observation hives. In the laboratory, time required for 90% cumulative mortality of mites (LT(90)) was 4.16 (3.98-4.42) days for H. thompsonii and 5.85 (5.48-7.43) days for M. anisopliae at 1.1 x 10(3) conidia mm(-2). At a temperature (34+/-1 degrees C) similar to that of the broodnest in a honey bee colony, Apis mellifera L., H. thompsonii [LC(90)=9.90 x 10(1) (5.86-19.35) conidia mm(-2) at Day 7] and M. anisopliae [LC(90)=7.13 x 10(3) (2.80-23.45) conidia mm(-2) at Day 7] both showed significant virulence against V. destructor. The applications of H. thompsonii to observation hives resulted in significant mortality of mites, and reduction of the number of mites per bee 21 and 42 days post-treatments. The treatments did not significantly affect the mite population in sealed brood. However, the fungus must have persisted because infected mites were still observed [82.97+/-(0.6)%] 42 days post-treatment. In addition, the fungus was found to sporulate on the host. A small percentage [2.86+/-(0.2)%] of dead mites found in the control hives also showed fungal infection, suggesting that adult bees drifted between hives and disseminated the fungus. H. thompsonii was harmless to the honey bees at the concentrations applied and did not have any deleterious effects on the fecundity of the queens. Microbial control with fungal pathogens provides promising new avenues for control of V. destructor and could be a useful component of an integrated pest management program for the honey bee industry.     

Production, Survival and Evaluation of Liquid Culture-produced Inocula of Coniothyrium minitans Against Sclerotinia sclerotiorum

Growth of Coniothyrium minitans on potato dextrose broth was compared with that on an inexpensive molasses-yeast liquid medium at 18-22°C in static culture. Biomass and conidial production were, in general, similar, although the rate of biomass production was quicker and conidial production was slightly greater per unit volume of medium in the molasses-yeast medium. Air-dried biomass from molasses-yeast liquid culture containing mycelia, pycnidia and conidia of C. minitans was mixed (12%, w/w) with kaolin to give a kaolin-biomass dust. The ability of C. minitans to survive and subsequently infect and reduce the viability of sclerotia of Sclerotinia sclerotiorum from this kaolin-biomass dust was found to be little affected by storage for 48 weeks between 4 and 15°C but was decreased by higher storage temperatures. The kaolin-biomass dust preparation did not differ from a standard maizemeal-perlite inoculum of C. minitans in its ability to infect sclerotia of S. sclerotiorum or reduce their viability or carpogenic germination in glasshouse and field pot bioassays. Further, when either inoculum was applied once to glasshouse soil naturally infested with S. sclerotiorum prior to planting three successive crops of lettuce, the pattern of disease control, reduction of sclerotial numbers/ plot, infection of sclerotia, reduction of sclerotial viability and survival in soil were similar for both inocula. The potential for the commercial development of liquid-culture-produced inocula of C. minitans is discussed.     

Nutritional effects of supplementary diets on brood development,biological activities and honey production of Apis mellifera L

The present research work was conducted to assess the impact of nutrient-enriched diet on the physiological activities and subsequently honey yield. Eighteen colonies of Apis mellifera L. were selected from Dera Ismail Khan region, KPK, Pakistan, during the winter and summer seasons, 2019–2020. Five pollen supplement diets were prepared and provided to screen out the palatable one to be fed as pollen alternative nutrition to bee bread. Results of diet consumption regarding mean data for consumption rate displayed that soybean flour enriched artificial diet was maximally consumed (74.34 g) by honey bees per week. Minimum consumption was observed for grinded groundnut enriched diet (64.62 g) which was relatively lesser than the other tested artificial diets. Results of area of worker brood disclosed that soybean flour fortified diet (1489.27 cm²/colony) statistically noteworthy than the other artificial diets whereas control (463.51 cm²/colony) was least effective. Highest bee strength (10.00 bee frames/colony) was noted in the bee colonies fed with soybean flour fortified diet, date paste (8.0 bee frames/colony) was the next effective one, among the tested pollen replacement diets whereas relatively least (5 bee frames/colony) was noticed in case of grinded groundnut. Highest body weight (12.41 g) of neonate bees was noted in case of soybean enriched diet while lowermost (5.31 g) was noted in control bees. Results of wax cell built up and foraging efficiency were also superior in artificial diets than respective control bees. Hence, artificial diets especially soybean-enriched pollen alternative diet can boost up the physiology of honey bee leading to increased honey yield and profit.