Methods of pest control

Abstract

Methods of pest control are provided. Accordingly, there is provided a method of protecting a plant at risk of pest infestation or treating a pest infested plant comprising contacting the plant with a pesticidal composition obtained or derived from an isolated Bacillus velezensis strain BX77 or a homolog thereof.

Description

[0001] METHODS OF PEST CONTROL

[0002] RELATED APPLICATION / S

[0003] This application claims the benefit of priority of U.S. Provisional Patent Application No. 63 / 692,753, filed on September 10, 2024, the contents of which are incorporated herein by reference in their entirety.

[0004] SEQUENCE LISTING STATEMENT

[0005] The XML file, entitled 104820.xml, created on September 1, 2025, comprising 4,152,034 bytes, submitted concurrently with the filing of this application is incorporated herein by reference.

[0006] FIELD AND BACKGROUND OF THE INVENTION

[0007] The present invention, in some embodiments thereof, relates to methods of pest control.

[0008] Most of the currently used chemical pesticides have a high potential for damaging the ecosystem, they are toxic to humans directly or through the food chain and their use is also impeded by development of genetic insect resistance.

[0009] In recent years, biological agents for plant protection have emerged as promising alternatives to chemical pesticides. These include beneficial organisms or natural enemies that reduce insect harm, pheromones that act as bait or disrupt reproduction, the release of sterile males to curb insect populations and more. In addition, microbial agents, such as bacteria, fungi, and viruses, and substances derived therefrom are also gaining traction for their ability to control pests without the adverse effects associated with chemical pesticides.

[0010] Additional background art includes:

[0011] Gollop R, et al. Front Plant Sci. 2024 15: 1287184, doi: 10.3389 / fpls(dot)2024.1287184, PMID: 38313804; and

[0012] International Patent Application Publication Nos. WO2022 / 195585; W02015184170; WO2014028521; WO2022245786; WO2020263812; and WO2024018434.

[0013] SUMMARY OF THE INVENTION

[0014] According to an aspect of some embodiments of the present invention there is provided a method of treating a pest infested plant, the method comprising contacting the plant with a pesticidal composition obtained or derived from an isolated Bacillus velezensis strain BX77 having a genome sequence as set forth in SEQ ID NO: 1 or a homolog thereof, thereby treating the pest infested plant.

[0015] According to an aspect of some embodiments of the present invention there is provided a method of protecting a plant at risk of pest infestation, the method comprising contacting the plant or with a pesticidal composition obtained or derived from an isolated Bacillus velezensis strain BX77 having a genome sequence as set forth in SEQ ID NO: 1 or a homolog thereof, thereby protecting the plant at risk of pest infestation.

[0016] According to some embodiments of the invention, the pesticidal composition comprises live cells of the Bacillus velezensis strain BX77 or a homolog thereof.

[0017] According to some embodiments of the invention, the pesticidal composition comprises a biofilm of the Bacillus velezensis strain BX77 or a homolog thereof.

[0018] According to some embodiments of the invention, the pesticidal composition comprises vegetative cells and / or dormant endospore of the Bacillus velezensis strain BX77 or a homolog thereof.

[0019] According to some embodiments of the invention, the pesticidal composition comprises a cells broth collected from a culture of the Bacillus velezensis strain BX77 or a homolog thereof.

[0020] According to some embodiments of the invention, the pesticidal composition comprises a cells free supernatant or filtrate obtained following culturing of the Bacillus velezensis strain BX77 or a homolog thereof.

[0021] According to some embodiments of the invention, the pesticidal composition comprises dead cells, an extract, cells fraction or a lysate of the Bacillus velezensis strain BX77 or a homolog thereof.

[0022] According to some embodiments of the invention, the pesticidal composition is obtained following culturing of the Bacillus velezensis strain BX77 or a homolog thereof under conditions allowing biofilm formation and / or sporulation.

[0023] According to some embodiments of the invention, the conditions comprise a medium selected from the group consisting of LB, NB and MRS.

[0024] According to some embodiments of the invention, the conditions comprise a LB medium.

[0025] According to some embodiments of the invention, the conditions comprise culturing for at least 36 hours.

[0026] According to some embodiments of the invention, the conditions comprise culturing for up to 72 hours.

[0027] According to some embodiments of the invention, the conditions comprise culturing while shaking at 100-200 rpm. According to some embodiments of the invention, the pesticidal composition is obtained following culturing of the Bacillus velezensis strain BX77 for 36-56 hours, at 28-32 °C, in a LB medium while shaking at 100-200 rpm.

[0028] According to some embodiments of the invention, the method comprising culturing the Bacillus velezensis strain BX77 under the conditions prior to the contacting.

[0029] According to some embodiments of the invention, the pesticidal composition is formulated as a liquid, solid, semi-solid, gel or powder.

[0030] According to some embodiments of the invention, the pesticidal composition is formulated as a liquid formulation.

[0031] According to some embodiments of the invention, the pesticidal composition is formulated as a dry formulation.

[0032] According to some embodiments of the invention, the contacting is effected once to twice a week.

[0033] According to some embodiments of the invention, upon the contacting the plant is at a post-flowering stage, a flowering stage, a pre-flowering stage, or any combination thereof.

[0034] According to some embodiments of the invention, the contacting is selected from the group consisting of spraying, immersing, drenching, coating, encapsulating, dusting, and fumigating.

[0035] According to some embodiments of the invention, the contacting is by spraying.

[0036] According to some embodiments of the invention, the contacting is onto or in the vicinity of a root, a stem, a trunk, a seed, a fruit, a flower, a leaf, or any combination thereof.

[0037] According to some embodiments of the invention, the contacting is by spraying a canopy of the plant and / or by drenching a root zone of the plant.

[0038] According to some embodiments of the invention, the pesticidal composition is provided in a concentration of at least 108CFU / ml.

[0039] According to some embodiments of the invention, the plant is a nightshade crop.

[0040] According to some embodiments of the invention, the plant is selected from the group consisting of tomato, cucumber, potato, eggplant, pepino, pepper, beet, citrul, physic nut, lucerne, spinach, bean, tobacco, maize and strawberries.

[0041] According to some embodiments of the invention, the plant is an ornamental plant.

[0042] According to some embodiments of the invention, the plant is a herbal plant.

[0043] According to some embodiments of the invention, the pest is an insect.

[0044] According to some embodiments of the invention, the insect is a moth.

[0045] According to some embodiments of the invention, the moth is Tula absoluta. According to some embodiments of the invention, the insect is Bemisia tabaci.

[0046] According to some embodiments of the invention, the insect is Aphis gossypii.

[0047] According to some embodiments of the invention, the pest is an arthropod.

[0048] According to some embodiments of the invention, the arthropod is a mite or a spidermite.

[0049] According to some embodiments of the invention, the spider- mite is Tetr any chus urticae.

[0050] According to some embodiments of the invention, the pest is a fungus.

[0051] According to some embodiments of the invention, the fungus is selected from the group consisting of Botrytis cinerea, Sclerotinia sclerotiorum and Oidium neolycopersici.

[0052] According to some embodiments of the invention, the pest is a virus.

[0053] According to some embodiments of the invention, the virus is a Tomato yellow leaf curl virus.

[0054] According to some embodiments of the invention, the plant is planted in an area known to be infested with the pest.

[0055] According to some embodiments of the invention, the area is a tropical region.

[0056] According to some embodiments of the invention, the area is a sub-tropical region.

[0057] According to some embodiments of the invention, the area is temperate region.

[0058] Unless otherwise defined, all technical and / or scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which the invention pertains. Although methods and materials similar or equivalent to those described herein can be used in the practice or testing of embodiments of the invention, exemplary methods and / or materials are described below. In case of conflict, the patent specification, including definitions, will control. In addition, the materials, methods, and examples are illustrative only and are not intended to be necessarily limiting.

[0059] BRIEF DESCRIPTION OF THE SEVERAL VIEWS OF THE DRAWINGS

[0060] Some embodiments of the invention are herein described, by way of example only, with reference to the accompanying drawings. With specific reference now to the drawings in detail, it is stressed that the particulars shown are by way of example and for purposes of illustrative discussion of embodiments of the invention. In this regard, the description taken with the drawings makes apparent to those skilled in the art how embodiments of the invention may be practiced.

[0061] In the drawings: FIGs. 1A-C demonstrate the pesticidal effect of Bacillus velezensis 'Ql l'l on the plant pathogen fungus Botrytis cinerea (gray mold) infestation of tomato plants. BX77 was grown for 24 hours at 37 °C in a laboratory medium [Nutrient broth (NB)] with 150 rpm shaking. The bacterium was applied by spray to the canopy of tomato plants or by drench to the root zone of the plants. Bacterium treatments were applied 4 and 0 days before the infection with B. cinerea conidia. Figure 1A demonstrates the effect on the severity of gray mold on tomato leaves of whole tomato plants (cv. Ikram). Results in the upper panel represent the average severity at each treatment applied by spray to the canopy of tomato plants or by drench; and in the lower panel the area under the disease progress curve (AUDPC). Figure IB demonstrates the effect on the severity (expressed as AUDPC) of gray mold (=Bolrylis cinerea-incitcd disease) on tomato (cv. Ikram) detached leaves in a humidity chamber that originated from bacterium treatments in a greenhouse at days 0, 7, and 14 before infection. Results represent the severity at each treatment applied by spray to the canopy of tomato plants calculated for the area under the disease progress curve (AUDPC). Figure 1C demonstrates the effect on the severity of gray mold on tomato (cv. Tilkram) detached leaves in a humidity chamber that originated from bacterium treatments in a greenhouse at days 0, 7, and 14 before infection. Results represent the severity at each treatment applied by spray to the canopy of tomato plants calculated for the area under the disease progress curve (AUDPC). In each figure, different letters attached to the columns represent statistical significance in Welch's ANOVA with a Dunnett post-hoc test, P<0.05 according to Dunnett Test. Bars = SE. Following the 24 hours of culturing the generated bacterial preparations were categorized as follows: In Figure 1A, bacteria were washed by centrifugation and resuspended in tap water at a concentration of ca. 108CFU / ml. In Figures 1B-C, Sporulation medium, washed - BX77 was grown in NB in a volume of 10 ml in a 50 ml tube and washed with double distilled water (DDW) (washing = centrifugation of the suspension, discarding of the supernatant and suspending the cells in DDW); Sporulation medium, unwashed - an unwashed BX77 grown in NB in a volume of 10 ml a 50 ml tube; Medium-density sporulation medium washed - BX77 grown in NB in a volume of 25 ml in a 50 ml tubes and washed in DDW; Aerated sporulation medium washed - BX77 grown in NB in a volume of 50 ml in 250 ml Erlenmeyer flask and washed in DDW.

[0062] FIG. ID demonstrates the pesticidal effect of Bacillus velezensis 'Ql l'l on the plant pathogen fungus Botrytis cinerea (gray mold) infestation of tomato plants. BX77 was grown for 48 hours at 30+1 °C in a lysogeny broth (LB) medium with 150 rpm shaking. Following, the generated bacterial preparations were categorized as follows: Alive - freshly grown BX77 cells suspended in the growth medium, without washing the cells; Alive washed - freshly grown cells washed from the medium and suspended in DDW; Heat killed / Washed heat killed - Bacteria killed in an autoclave using a standard program (121 °C for 15-20 minutes) unwashed or washed from the media; Supernatant - the media in which the bacteria were grown, collected and filtered through a 0.22-micron filter. The bacterial preparations were applied by spray to the canopy of tomato plants 4 days before the infection with B. cinerea conidia. Shown is reduction in disease severity following spraying with the indicated BX77-derived compositions.

[0063] FIG. 2 demonstrates the pesticidal effect of Bacillus velezensis 'Ql l'l on Sclerotinia sclerotiorum infestation of cucumber leaves of whole plants treated 3 days before inoculation with mycelium discs of the fungus. Results represent the average severity treatment applied by drench treatment. Different letters represent statistical significance in Welch's ANOVA with a Dunnett post-hoc test, P<0.05 according to Dunnett Test. Bars = SE.

[0064] FIGs. 3A-D demonstrate the pesticidal effect of Bacillus velezensis 'Ql l'l on powdery mildew (Oidium neolycopersici) infestation of tomato grown in a greenhouse. Figure 3A demonstrates the effect of treatment with BX77 whole broth at different concentrations (106 / ml, 107 / ml, or 108 / ml) applied by spray to the canopy of tomato plants (cv. Tori). Plants were treated with the bacterial preparation twice, at days 0 and day 8 and disease severity was evaluated at days 6 and 13 after the first treatment. Results in the upper panel represent the severity at day 13 for each treatment; and in the lower panel calculated for the area under the disease progress curve (AUDPC) through 13 days. Figure 3B demonstrates the effect of treatment with BX77 whole broth (marked as “Bx77 EB 150”), washed bacterial cells or supernatant devoid of bacterial cells applied by spray to the canopy of tomato plants (cv. Tori). Plants were treated twice with the bacterial-derived preparation, at day 0 and day 6 and disease severity was evaluated at days 5 and 12 after the first treatment. Results in the upper panel represent the severity at day 12 for each treatment; and in the lower panel calculated for the area under the disease progress curve (AUDPC) through 12 days. Figure 3C demonstrates the effect of treatment with the indicated BX77-derived compositions applied by spray to the canopy on tomato leaves of whole tomato plants (cv. Ikram). Results in the upper panel represent the average severity at each treatment at day 19; and in the lower panel as an area under the disease progress curve (AUDPC) over 26 days. Figure 3D demonstrates the effect of treatment with the indicated BX77-derived compositions generated in sporulation supportive conditions and applied by spray to the canopy on tomato leaves of whole tomato plants (cv. Tilkram). Results represent the area under the disease progress curve (AUDPC) over 26 days (upper panel) and 32 days (lower panel). Different letters represent statistical significance in Welch's ANOVA with a Dunnett post-hoc test, P<0.05 according to Dunnett Test. Bars = SE. The generated bacterial preparations were categorized as follows: Bacillus 'Ql l'l was grown for 48 hours at 30+1 °C in a LB medium with 150 rpm shaking in Figures 3A-B and for 24 hours at 37+1 °C in a NB medium with 150 rpm shaking in Figures 3C-D. Following culture, in Figure 3A the whole broth was diluted in tap water to adjust to the required concentration. In Figure 3B, Washed Bx77- live BX77 cells washed from the medium and resuspended in tap water; BX77 LB 150 - live BX77 cells grown in LB medium at 150 rpm. Supernatant - the conditioning medium in which the bacteria were grown, collected and filtered through a 0.22-micron filter. In Figures 3C-D, Sporulation medium washed - BX77 grown in NB in a volume of 10 ml in a 50 ml tube and washed in DDW; Sporulation medium unwashed - unwashed BX77 grown in NB in a volume of 10 ml a 50 ml tube; Medium-density sporulation medium washed - BX77 grown in NB in a volume of 25 ml in a 50 ml tubes and washed in DDW; Aerated sporulation medium washed - BX77 grown in NB in a volume of 50 ml in 250 ml Erlenmeyer flask and washed in DDW.

[0065] FIGs. 4A-D demonstrate the pesticidal effect of Bacillus velezensis 'Ql l'l on Bemisia tabaci infestation of tomato plants. Treatment with the bacterial-based preparations was performed by drench or spray treatments on days 0, 7, 14, 21 and 28. Figure 4A demonstrates the effect of treatment with BX77 cultured under various conditions, which supports either biofilm formation or sporulation, and applied by drench or spray to the canopy on tomato leaves of whole tomato plants (cv. Ram 4107). Specifically, Bacillus 'Ql l'l was grown in LB, under conditions that stimulate biofilm formation in LB supplemented with glycerol and manganese (LB GM) medium (biofilm culture), or under conditions that stimulate sporulation NB medium (spore culture). The suspensions were washed with DDW and applied to plants by spray or drench at a concentration of ca. 108CFU / ml. Results in the upper panel represent the average incidence at each treatment presented as the incidence of individuals on day 20; and in the lower panel as the area under the Bemisia progress curve (AUDPC, lower). Figure 4B demonstrates the effect of treatment with the indicated BX77-derived compositions (obtained as described in Figures 1B-C and 3C-D), harboring sporulating cells were applied by spray to the canopy on tomato leaves of whole tomato plants (cv. Ikram). Results in the upper panel represent the average incidence at each treatment on day 33; and in the lower panel the area under B. tabaci progress curve (AUBPC) through 33 days. Figure 4C demonstrates the effect of treatment with the indicated BX77-derived compositions of sporulating BX77 cells applied by spray to the canopy on tomato leaves of whole tomato plants (cv. Tilkram). Results in the upper panel represent the average incidence at each treatment at day 26; and in the lower panel the area under B. tabaci progress curve (AUBPC) through 32 days. Different Letters represent statistical significance in Welch's ANOVA with a Dunnett post-hoc test, P<0.05 according to Dunnett Test. Bars = SE. Figure 4D demonstrates the effect of the bacterium that was sprayed at concentrations of 106to 109 / ml and the effect of the commercial product Serenade ASO Bacillus subtilis group BM02; BAYER, Crop Science, U.S.) on B. tabaci population on tomato leaves. Bacillus BX77 was grown for 48 hours at 30+1 °C in a LB medium with 150 rpm shaking and diluted in tap water to adjust the required concentrations. The suspensions were applied by spray on selected leaves in two weekly applications. Natural silverleaf whitefly population developed and counted 5 days after the second application. Silverleaf whitefly individuals were counted on leaf 6 that was treated by the bacterial suspensions and on leaf 8 that was not sprayed at all, in order to test systemic effect. The Bemisia population was suppressed by BX77 on the treated leaves and on the untreated leaves with better effect of concentrations 107to 109 / ml.

[0066] FIGs. 5A-D demonstrate the pesticidal effect of Bacillus velezensis 'Ql l'l on Tula absoluta infestation of tomato plants. Treatment with the bacterial-based preparations was performed by drench or spray treatments on a weekly basis and the incidence of galleries of 3 sizes formed by the larva of Tula absoluta on the leaves of whole tomato plants was determined. Additionally, the severity of damage inflicted by T. absoluta on the tomato leaves was quantified and calculated according to an index formula (described in the MATERIALS AND METHODS section of Example 5). Figure 5A demonstrates the effect of treatment with BX77 cultured under various conditions to promote either sporulation or vegetative growth (obtained as described in Figure 4A) and was applied by drench or spray to the canopy on tomato leaves of whole tomato plants (cv. Ram 4107). Results represent the average incidence at each treatment. The damage caused by the larva is presented for the incidence of small galleries (upper panel), the incidence of large galleries (second row panel), all galleries (third row panel), and indexed damage calculated for all galleries according to their size (bottom panel), at day 20 after the first set of treatments application. AUPPC = area under pest progress curve. Figure 5B demonstrates the effect of treatment with the indicated BX77-derived compositions (obtained as described in Figures 1B-C and 3C-D) of sporulating cells applied by spray to the canopy on tomato leaves of whole tomato plants (cv. Ikram). Results represent the average incidence at each treatment. The damage caused by the larva is presented for the incidence of small galleries (upper panel), the incidence of large galleries (second row panel), all galleries (third row panel), and indexed damage severity calculated for all galleries according to their size (lower panel), at day 19 after first set of treatments application. Figure 5C demonstrates the effect of treatment with the indicated BX77-derived compositions applied by spray to the canopy on tomato leaves of whole tomato plants (cv. Tilkram). Results represent the average incidence at each treatment. The damage caused by the larva is presented for the incidence of small galleries (upper panel), the incidence of large galleries (second row panel), all galleries (third row panel) and indexed damage severity calculated for all galleries according to their size (lower panel), at day 19 after the first set of treatments application. Figure 5D demonstrates the effect of treatment with the indicated BX77-derived composition. Bacillus BX77 was grown for 48 hours at 30+1 °C in an LB medium with 150 rpm shaking and diluted in tap water up to the required concentrations and applied by spray to the canopy on tomato leaves of whole tomato plants (cv. Tilkram) at final concentrations of 104 / ml, 105 / ml, 106 / ml, 107 / ml, and 108 / ml. Results represent the average incidence at each treatment. The damage caused by the larva is presented for the incidence of small galleries, medium galleries and large galleries (upper panel), the incidence of all galleries (second row panel) and indexed damage severity calculated for all galleries according to their size (lower panel), at day 12 after the first set of treatments application. Different letters represent statistical significance in Welch's ANOVA with a Dunnett post-hoc test, P<0.05 according to Dunnett Test. Bars = SE.

[0067] FIGs. 6A-B demonstrate the pesticidal effect of Bacillus velezensis 'Ql l'l on Tetranychus urticae infestation of tomato plants. Treatment with the bacterial-based preparations (obtained as described in Figures 1B-C and 3C-D) was performed by spray to the canopy treatments on a weekly basis. Figure 6A demonstrates the effect on tomato leaves of whole tomato plants (cv. Ikram). Results represent the average mites’ number at each treatment on day 33 (upper panel), the damage inflicted by the mites on day 26 (middle panel) and the area under the mites’ damage progress curve (AUMDPC) through 33 days (lower panel). Figure 6B demonstrates the effect on tomato leaves of whole tomato plants (cv. Tilkram). Results represent the average mites’ number at each treatment on day 32 (upper panel), the damage inflicted by the mites on day 32 (middle panel) and the area under the mite damage progress curve (AUMDC) through 32 days (lower panel). Different letters represent statistical significance in Welch's ANOVA with a Dunnett post-hoc test, P<0.05 according to Dunnett Test. Bars = SE.

[0068] FIG. 7 demonstrates the pesticidal effect of Bacillus velezensis 'Ql l'l on Tomato yellow leaf curl virus (TYLCV) infestation of tomato plants. Shown is the effect of treatment with the indicated BX77-derived compositions (obtained as described in Figures 1B-C and 3C-D) of sporulating cells were applied on a weekly basis (days 0, 7, 14, 21, and 28) by spray to the canopy on tomato leaves of whole tomato plants (cv. Ikram). Results in the upper panel represent the average symptoms severity in the various treatments; and in the lower panel the area under disease progress curve (AUDPC) through 33 days. Different letters represent statistical significance in Welch's ANOVA with a Dunnett post-hoc test, P<0.05 according to Dunnett Test. Bars = SE. FIGs. 8A-B demonstrate reduction in Botrytis (Figure 8A) or Tula absoluta (Figure 8B) disease severity on tomato leaves following spraying a BX77 suspension grown in LB or NB medium for 48 hours at 30+1 °C with 150 rpm shaking.

[0069] FIG. 9 demonstrates the pesticidal effect of Bacillus velezensis BX77 on Aphis gossypii infestation of whole tomato plants (cv. Tori). Treatment with the bacterial-based preparations (obtained as described in Figures 1B-C and 3C-D) was performed by spray to the canopy at 12 and 5 days before aphids’ evaluation. Results represent the average aphids’ number 5 days after the second application of the treatment. Different letters represent statistical significance in Welch's ANOVA with a Dunnett post-hoc test, P<0.05 according to Dunnett Test. Bars = SE.

[0070] DESCRIPTION OF SPECIFIC EMBODIMENTS OF THE INVENTION

[0071] The present invention, in some embodiments thereof, relates to methods of pest control.

[0072] Before explaining at least one embodiment of the invention in detail, it is to be understood that the invention is not necessarily limited in its application to the details set forth in the following description or exemplified by the Examples. The invention is capable of other embodiments or of being practiced or carried out in various ways.

[0073] Most of the currently used chemical pesticides have a high potential for damaging the ecosystem, they are toxic to humans directly or through the food chain and their use is also impeded by development of genetic insect resistance.

[0074] Whilst reducing specific embodiments of the present invention to practice, the present inventors have now uncovered that a bacterial isolate, BX77 has a pesticidal activity. As is illustrated hereinunder and in the examples section, which follows, the present inventors show that formulated suspensions of BX77 applied plants such as tomato and cucumber reduced gray mold caused by the fungal pest Botrytis cinereal, white mold disease caused by the fungal pest Sclerotinia sclerotiorum, and powdery mildew that is caused by Oidium neolycopersici. Application of the BX77 formulation also reduced the leaf damage caused by the tomato leaf miner Tuta absoluta, which is an insect moth, the red spider mite Tetranychus urticae and the populations of the silverleaf whitefly insect Bemisia tabaci on tomato leaves and of the insect pest Aphis gossypii on tomato plants. In addition, the formulation reduced the symptoms of Tomato yellow leaf curl virus infection, which is vectored by whiteflies. Importantly, both spray application to the leaves and root zone application had pesticidal activities.

[0075] Consequently, specific embodiments suggest that the Bacillus velezensis strain BX77 and pesticidal composition obtained or derived therefrom may be used in protecting and / or treating a plant from pest infestation. Thus, according to an aspect of the present invention, there is provided a method of treating a pest infested plant, the method comprising contacting the plant with a pesticidal composition obtained or derived from an isolated Bacillus velezensis strain BX77 having a genome sequence as set forth in SEQ ID NO: 1 or a homolog thereof, thereby treating the pest infested plant.

[0076] According to an additional or an alternative aspect of the present invention, there is provided a method of protecting a plant at risk of pest infestation, the method comprising contacting the plant or with a pesticidal composition obtained or derived from an isolated Bacillus velezensis strain BX77 having a genome sequence as set forth in SEQ ID NO: 1 or a homolog thereof, thereby protecting the plant at risk of pest infestation.

[0077] As used herein, the term “pest" refers to an organism that negatively effects the health, growth and / or productivity of plants. The pest can directly harm the plant by feeding on it or competing with the plant on nutrients or indirectly by spreading a disease. Non-limiting examples of pests include insects (as defined herein), nematodes, fungi, bacteria and viruses.

[0078] According to specific embodiments, the pest is an insect.

[0079] The term "insect" is used herein includes species of the superphylum Panarthropoda (classification Systema Naturae, Brands, S.J. (comp.) 1989-2005. Systema Naturae 2000. Amsterdam, The Netherlands, [www(dot)sn2000(dot)taxonomy(dot)nl / ]), including the phyla Arthropoda, Tardigrada and Onychophora and includes all the different developmental phases of the life cycle, such as, but not limited to eggs, larvae, nymphs, pupae and adults.

[0080] According to specific embodiments, the insect belongs to the phylum Arthropoda (including, but not limited to the orders Archaeognatha, Thysanura, Paleoptera, Hemiptera and Neoptera, also ticks, mites and spiders); according to specific embodiments to the epiclass Hexapoda', according to specific embodiments, to the class Insecta. According to specific embodiments, the insect belongs to the order Lepidoptera. Non-limiting examples of insects include bedbugs, house flies, moths, beetles, mites, spiders, grasshoppers, caterpillars, aphids, mosquitos, fleas, horseflies, hornets, cockroaches and ants, such as, but not limited to:

[0081] • the order Lepidoptera, for example: Tuta absoluta, Acleris spp., Adoxophyes spp., Agrotis spp., Alabama argillacea, Amyelois spp., Anticarsia gemmatalis, Archips spp., Argyrotaenia spp., Autographa spp., Busseola fusca, Cadra cautella, Carposina nipponensis, Chilo suppressalis, Chilo spp., Choristoneura conflictana, Choristoneura fumiferana, Choristoneura occidentalis, Choristoneura rosaceana, Choristoneura spp., Clysia ambiguella, Cnaphalocrocis spp., Cnephasia spp., Cochylis spp., Coleophora spp., Crocidolomia binotalis, Cryptophlebia leucotreta, Cydalima perspectalis, Cydia inopinata, Cydia spp., Diatraea spp., Diparopsis castanea, Earias spp., Ephestia spp., Eucosma spp., Eupoecilia ambiguella, Euproctis spp., Euxoa spp., Grapholita prunivora, Grapholita spp., Hedya nubiferanal, Helicoverpa armigera, Helicoverpa zea, Helicoverpa spp., Heliothis spp., Hellula undalis, Hyphantria cunea, Keiferia lycopersicella, Eeucoptera scitella, Eithocolletis spp., Eobesia botrana, Lymantria spp., Eyonetia spp., Malacosoma spp., Mamestra brassicae, Manduca sexta, Numonia pyrivorella, Operophtera spp., Opogona sacchari, Ostrinia nubilalis, Pammene spp., Pandemis spp., Panolis flammea, Paysandisia archon, Pectinophora gossypiella, Phthorimaea operculella, Phyllonorycter spp., Pieris rapae, Pieris spp., Platynota rostrana, Plutella xylostella, Prays spp., Scirpophaga spp., Sesamia spp., Sesia spp., Sparganothis spp., Spodoptera dolichos, Spodoptera eridania, Spodoptera exigua, Spodoptera frugiperda, Spodoptera littoralis, Spodoptera litura, Spodoptera spp., Synanthedon spp., Tecia solanivora, Thaumatotibia leucotreta, Thaumetopoea processionea, Thaumetopoea spp., Tortrix spp., Trichoplusia ni, and Yponomeuta spp.;

[0082] • the order Coleoptera, for example, Agrilus anxius, Agrilus planipennis, Agriotes spp., Anomala orientalis, Anoplophora chinensis, Anoplophora glabripennis, Anoplophora spp., Anthonomus bisignifer, Anthonomus eugenii, Anthonomus grandis, Anthonomus quadrigibbus, Anthonomus signatus, Anthonomus spp., Apriona spp., Arrhenodes minutus, Atomaria linearis, Chaetocnema tibialis, Conotrachelus nenuphar, Cosmopolites spp., Curculio spp., Dendroctonus micans, Dendrolimus sibiricus, Dermestes spp., Diabrotica virgifera, Diabrotica virgifera zeae, Diabrotica virgifera, Diabrotica balteata, Diabrotica barberi, Diabrotica undecimpunctata howardi, Diabrotica undecimpunctata tenella, Diabrotica undecimpunctata, Diabrotica undecimpunctata, Diabrotica spp., Epilachna varivestis, Epilachna spp., Epitrix cucumeris, Eremnus cerealis, Eremnus spp., Gonipterus scutellatus, Ips amitinus, Ips cembrae, Ips duplicatus, Ips sexdentatus, Ips typographus, Ips spp., Eeptinotarsa decemlineata, Eeptinotarsa juncta, Eeptinotarsa texana, Eissorhoptrus spp., Eistronotus bonariensis, Melolontha spp., Monochamus spp., Naupactus leucoloma, Oryzaephilus spp., Otiorhynchus spp., Phlyctinus spp., Pissodes nemorensis, Pissodes strobi, Pissodes terminalis, Pissodes spp., Popilia japonica, Popilia spp., Premnotrypes spp., Pseudopityophthorus minutissimus, Pseudopity ophthorus pruinosus, Psylliodes spp., Rhizopertha spp., Rhynchophorus ferrugineus, Rhynchophorus palmarum, Scarabaeidae family spp., Scolytidae family spp., Sitophilus spp., Sitotroga spp., Sternochetus mangiferae, Tenebrio spp., Tribolium castaneum, Tribolium spp. and Trogoderma spp.;

[0083] • the order Orthoptera, for example, Gryllotalpa spp., Locusta spp., and Schistocerca spp.; • the order Blattaria, from example, Blatta spp., Blattella spp., Periplaneta spp., and Leucophaea maderae,

[0084] • the order Isoptera, for example, Coptotermes spp. and Reticulitermes spp.;

[0085] • the order Psocoptera, for example, Liposcelis spp.;

[0086] • the order Phthiraptera, suborder Anoplura, for example, Haematopinus spp., Einognathus spp., and Pediculus spp., and Trichodectes spp.;

[0087] • the order Phthiraptera, suborder Ischnocera, for example, Damalinia spp.;

[0088] • the order Thysanoptera, for example, Frankliniella occidentalis, Frankliniella platensis, Frankliniella spp., Hercinothrips spp., Taeniothrips spp., Thrips palmi, Thrips tabaci, Scirtothrips aurantii, Scirtothrips citri, Scirtothrips dorsalis, and Scirtothrips spp.;

[0089] • the order Hemiptera, suborder Heteroptera, for example, Cimex spp., Distantiella theobroma, Dysdercus spp., Euschistus spp., Eurygaster spp., Eeptocorisa spp., Nezara spp., Piesma spp., Rhodnius spp., Sahlbergella singularis, Scotinophara spp., Triatoma spp., Miridae family spp. such as Eygus hesperus and Eygus lineoloris, Eygaeidae family spp. such as Blissus leucopterus, and species of the Pentatomidae family;

[0090] • the order Hemiptera, suborder Sternorrhyncha, for example, Aleurocanthus spiniferus, Aleurocanthus woglumi, Aleurocanthus spp., Aleurothrixus floccosus, Aleyrodes brassicae, Aonidella citrina, Aonidiella spp., Aphididae family spp., Acyrthosiphon spp., Aphis fabae, Aphis glycines, Aphis gossypii, Aphis spp., Aspidiotus spp., Bemisia tabaci, Ceroplastes spp., Chrysomphalus aonidium, Chrysomphalus dictyospermi, Coccus hesperidum, Daktulosphaira vitifoliae, Diaphorina citri, Eriosoma larigerum, Gascardia spp., Lacanium corni, Eepidosaphes spp., Lopholeucaspis japonica, Macrosiphus spp., Margarodes prieskaensis, Margarodes vitis, Margarodes vredendalensis, Myzus persicae, Myzus spp., Parasaissetia nigra, Pemphigus spp., Phylloxera spp., Pianococcus spp., Pseudaulacaspis spp., Pseudococcus spp., Psylla spp., Pulvinaria aethiopica, Quadraspidiotus spp., Rhopalosiphum spp., Ripersiella hibisci, Saissetia spp., Schizaphis spp., Sitobion spp., Toxoptera citricida, Trialeurodes vaporariorum, Trioza erytreae, and Unaspis citri;

[0091] • the order Hemiptera, suborder Auchenorrhyncha, for example, Circulifer haematoceps, Circulifer tenellus, Draeculacephala minerva, Empoasca spp., Erythroneura spp., Graphocephala atropunctata, Hishimonus phycitis, Myndus crudus, Laodelphax spp., Nephotettix spp., Nilaparvata spp., Scaphoideus luteolus, Scaphoideus spp., and Xyphon fulgida;

[0092] • the order Hymenoptera, for example, Acromyrmex, Atta spp., Cephus spp., Diprionidae family spp. such as Diprion spp. and Gilpinia polytoma, Hoplocampa spp., Easius spp., Monomorium pharaonis, Neodiprion spp., Formicidae family spp. such as Solenopsis spp., and Vespa spp.;

[0093] • the order Diptera, for example, Aedes albopictus, Aedes cinereus, Aedes polynesiensis, Aedes spp., Amauromyza maculosa, Anastrepha fraterculus, Anastrepha ludens, Anastrepha obliqua, Anastrepha suspensa, Anastrepha spp., Anopheles gambiae, Anopheles spp., Aschistonyx eppoi, Atherigona soccata, Bactrocera spp., Bibio hortulanus, Calliphora erythrocephala, Cephalcia lariciphila, Ceratitis rosa, Ceratitis spp., Chrysomyia spp., Culex spp., Cuterebra spp., Dacus spp., Drosophila melanogaster, Dryocosmus kuriphilus, Euphranta canadensis, Euphranta japonica, Fannia spp., Gastrophilus spp., Gilpinia hercyniae, Glossina spp., Hypoderma spp., Hippobosca spp., Eiriomyza bryoniae, Eiriomyza huidobrensis, Eiriomyza sativae, Eiriomyza trifolii, Eiriomyza spp., Luc ilia spp., Melanagromyza spp., Musca spp., Oestrus spp., Orseolia spp., Oscinella frit, Pardalaspis cyanescens, Pardalaspis quinaria, Pegomyia hyoscyami, Phorbia spp., Rhagoletis pomonella, Rhagoletis spp., Sciara spp., Stomoxys spp., Tabanus spp., and Tipula spp.;

[0094] • the order Siphonaptera, for example, Ceratophyllus spp. and Xenopsylla cheopis; and the infraclass Thysanura, order Zygentoma, for example, Lepisma saccharina.

[0095] • the subclass Acari, for example, Tetranychus urticae, Tetranychus cinnabarinus, Panonychus citri, Aculops pelekassi, Tarsonemus spp.

[0096] According to specific embodiments, the insect is selected from the group consisting of Bemisia tabaci and Aphis gossypii.

[0097] According to specific embodiments, the pest is a mite or a spider mite.

[0098] Non-limiting examples of mite or a spider mite pests include, Tetranychus urticae, Tetranychus cinnabarinus, Panonychus citri, Oligonychus pratensis, Oligonychus ilicis, Aculops lycopersici, Polyphagotarsonemus latus, Phyllocoptruta oleivora, Tarsonemus confusus, and Tarsonemus spp.

[0099] According to specific embodiments, the pest is a Tetranychus urticae spider mite.

[0100] According to specific embodiments, the pest is a moth.

[0101] Non-limiting examples of moths include Tuta absoluta, Helicoverpa armigera, Cydia pomonella, Spodoptera frugiperda, Mythimna unipuncta, Helicoverpa virescens, Manduca quinquemaculata, Ostrinia nubilalis and Cydia pomonella.

[0102] According to specific embodiments, the pest is Tuta absoluta.

[0103] According to specific embodiments, the pest is a fungus.

[0104] As used herein, the term "fungus" or "fungi" includes a nucleated spore bearing organisms without chlorophyll. Examples of fungi include yeast, molds, rusts, and mushrooms. Non- limiting examples of fungal pests include fungi of the phyla Plasmodiophoromycota, Chytridiomycota, Oomycota, Zygomycota, Ascomycota and Basidiomycota. Examples of fungal pest include, but not limited to Botrytis cinerea, Sclerotinia sclerotiorum, Oidium neolycopersici, Phytophthora infestans, Puccinia graminis, Fusarium oxysporum, Verticillium dahliae, Colletotrichum spp., Rhizoctonia solani. and Pythium spp.

[0105] According to specific embodiments, the fungus is selected from the group consisting of Botrytis cinerea, Sclerotinia sclerotiorum and Oidium neolycopersici

[0106] According to specific embodiments, the pest is a virus.

[0107] Non-limiting examples of viruses include Tomato yellow leaf curl virus (TYLCV), Cucumber mosaic virus (CMV), Tomato spotted wilt virus (TSWV), Potato virus Y (PVY), Pepper mild mottle virus (PMMoV), Tobacco mosaic virus (TMV), Zucchini yellow mosaic virus (ZYMV), Papaya ringspot virus (PRSV), Watermelon mosaic virus (WMV), Beet curly top virus (BCTV), Maize streak virus (MSV), and Banana bunchy top virus (BBTV), some of which are transmitted by insect vectors such as whiteflies, aphids, thrips, or leafhoppers.

[0108] According to specific embodiments, the virus is a Tomato yellow leaf curl virus.

[0109] The term ’’plant” as used herein encompasses whole plants, a grafted plant, ancestors and progeny of the plants and plant parts, including seeds, shoots, stems, roots (including tubers), rootstock, scion, and plant cells, tissues and organs. The plant may be in any form including suspension cultures, embryos, meristematic regions, callus tissue, leaves, gametophytes, sporophytes, pollen, and microspores. Plants that are particularly useful in the methods of the invention include all plants which belong to the superfamily Viridiplantee, in particular monocotyledonous and dicotyledonous plants including a fodder or forage legume, ornamental plant, food crop, tree, or shrub selected from the list comprising Acacia spp., Acer spp., Actinidia spp., Aesculus spp., Agathis australis, Albizia amara, Alsophila tricolor, Andropogon spp., Arachis spp, Areca catechu, Astelia fragrans, Astragalus cicer, Baikiaea plurijuga, Betula spp., Brassica spp., Bruguiera gymnorrhiza, Burkea africana, Butea frondosa, Cadaba farinosa, Calliandra spp., Camellia sinensis, Cannabaceae, Cannabis indica, Cannabis, Cannabis sativa, Hemp, industrial Hemp, Capsicum spp., Cassia spp., Centroema pubescens, Chacoomeles spp., Cinnamomum cassia, Coffea arabica, Colophospermum mopane, Coronillia varia, Cotoneaster serotina, Crataegus spp., Cucumis spp., Cupressus spp., Cyathea dealbata, Cydonia oblonga, Cryptomeria japonica, Cymbopogon spp., Cynthea dealbata, Cydonia oblonga, Dalbergia monetaria, Davallia divaricata, Desmodium spp., Dicks onia squarosa, Dibeteropogon amplectens, Dioclea spp., Dolichos spp., Dorycnium rectum, Echinochloa pyramidalis, Ehraffia spp., Eleusine coracana, Eragrestis spp., Erythrina spp., Eucalypfus spp., Euclea schimperi, Eulalia vi / losa, Pagopyrum spp., Feijoa sellowlana, Fragaria spp., Flemingia spp, Freycinetia banksli, Geranium thunbergii, GinAgo biloba, Glycine javanica, Gliricidia spp, Gossypium hirsutum, Grevillea spp., Guibourtia coleosperma, Hedysarum spp., Hemaffhia altissima, Heteropogon contoffus, Hordeum vulgare, Hyparrhenia rufa, Hypericum erectum, Hypeffhelia dissolute, Indigo incamata, Iris spp., Eeptarrhena pyrolifolia, Eespediza spp., Eettuca spp., Eeucaena leucocephala, Eoudetia simplex, Eotonus bainesli, Eotus spp., Macrotyloma axillare, Malus spp., Manihot esculenta, Medicago saliva, Metasequoia glyptostroboides, Musa sapientum, Nicotianum spp., Onobrychis spp., Ornithopus spp., Oryza spp., Peltophorum africanum, Pennisetum spp., Persea gratissima, Petunia spp., Phaseolus spp., Phoenix canariensis, Phormium cookianum, Photinia spp., Picea glauca, Pinus spp., Pisum sativam, Podocarpus totara, Pogonarthria fleckii, Pogonaffhria squarrosa, Populus spp., Prosopis cineraria, Pseudotsuga menziesii, Pterolobium stellatum, Pyrus communis, Quercus spp., Rhaphiolepsis umbellata, Rhopalostylis sapida, Rhus natalensis, Ribes grossularia, Ribes spp., Robinia pseudoacacia, Rosa spp., Rubus spp., Salix spp., Schyzachyrium sanguineum, Sciadopitys vefficillata, Sequoia sempervirens, Sequoiadendron giganteum, Sorghum bicolor, Spinacia spp., Sporobolus fimbriatus, Stiburus alopecuroides, Stylosanthos humilis, Tadehagi spp., Taxodium distichum, Themeda triandra, Trifolium spp., Triticum spp., Tsuga heterophylla, Vaccinium spp., Vicia spp., Vitis vinifera, Watsonia pyramidata, Zantedeschia aethiopica, Zea mays, amaranth, artichoke, asparagus, broccoli, Brussels sprouts, cabbage, canola, carrot, cauliflower, celery, collard greens, flax, kale, lentil, oilseed rape, okra, onion, potato, rice, soybean, straw, sugar beet, sugar cane, sunflower, tomato, squash tea, trees. Alternatively algae and other non-Viridiplantae can be used for the methods of some embodiments of the invention.

[0110] According to specific embodiments, the plant is a crop.

[0111] “Crop” as used herein means a plant species or variety that is grown to be harvested as food, livestock fodder, fuel raw material, or for any other economic purpose. As non-limiting examples, the crop can be maize, cereals, such as wheat, rye, barley and oats, sorghum, rice, sugar beet and fodder beet, fruit, such as pome fruit (e.g., apples and pears), citrus fruit (e.g., oranges, lemons, limes, grapefruit, or mandarins), stone fruit (e.g., peaches, nectarines or plums), nuts (e.g., almonds or walnuts), soft fruit (e.g., cherries, strawberries, blackberries or raspberries), the plantain family or grapevines, leguminous crops, such as beans, lentils, peas and soya, oil crops, such as sunflower, safflower, rapeseed, canola, castor or olives, cucurbits, such as cucumbers, melons or pumpkins, fiber plants, such as cotton, flax or hemp, fuel crops, such as sugarcane, miscanthus or switchgrass, vegetables, such as potatoes, tomatoes, pepino, peppers, lettuce, spinach, onions, carrots, eggplants, asparagus or cabbage, ornamentals, such as flowers (e.g., petunias, pelargoniums, roses, tulips, lilies, or chrysanthemums), shrubs, broad-leaved trees (e.g., poplars or willows) and evergreens (e.g., conifers), grasses, such as lawn, turf or forage grass or other useful plants, such as coffee, tea, tobacco, hops, pepper, rubber or latex plants.

[0112] According to specific embodiments, the plant is a nightshade crop.

[0113] “Nightshade crop” as used herein refers to a plant species or variety that belongs to the botanical family Solanaceae, commonly known as the nightshade family. This family includes a wide range of agriculturally important food and industrial crops that are grown for their edible fruits, tubers, or leaves, or for other economic purposes. Non-limiting examples of nightshade crops include tomato (Solatium lycopersicum), potato (Solatium tuberosum), eggplant (Solatium melongena), pepper (Capsicum annuum, Capsicum frutescens), tobacco (Nicotiana tabacum), pepino (Solatium muricatum), and Physalis species (e.g., groundcherry, cape gooseberry). In some embodiments, the nightshade crop is a cultivated variety of one or more of the aforementioned species, grown for human consumption, industrial use, or horticultural purposes.

[0114] According to specific embodiments, the plant is selected from the group consisting of tomato, cucumber, potato, eggplant, pepino, pepper, beet, citrul, physic nut, lucerne, spinach, bean, tobacco, maize and strawberries.

[0115] According to specific embodiments, the plant is selected from the group consisting of tomato, cucumber, grape, lettuce, potato, eggplant, pepper, tobacco and potato.

[0116] According to specific embodiments, the plant is a tomato and / or cucumber plant.

[0117] According to specific embodiments, the plant is an ornamental plant.

[0118] Non-limiting examples of ornamental plants include flowering plants such as petunia, rose, chrysanthemum, pelargonium, tulip, and lily, as well as ornamental shrubs such as boxwood and azalea, and foliage ornamentals such as coleus and caladium.

[0119] According to specific embodiments, the plant is a herbal plant.

[0120] Non-limiting examples of herbal plants include basil (Ocimum basilicum), mint (Mentha spp.), chamomile, sage, thyme, rosemary, and lavender.

[0121] According to specific embodiments, the plant is infested with a pest.

[0122] As used herein, the phrase “pest infested plant” or “plant infested with a pest”, refers to a plant that is infested with a pest even if symptoms are not detectable. Methods of determining plant infestation are well known in the art and depend on the type of pest and plant. These include, but not limited to visual inspection (signs may include damaged leaves or fruits, holes, wilting, discoloration, excessive leaf drop, deformities, presence of mold or mildew, reduced growth, visible pest on the plant surface etc.), traps and monitoring devices, sampling of plant part followed by analysis to detect pests or their damage, soil / root inspection, remote sensing and imaging, and molecule techniques (e.gh. PCR, ELISA).

[0123] According to specific embodiments, the plant is at risk of pest infestation.

[0124] As used herein, the phrase “at risk of pest infestation” refers to a plant pre-disposed or susceptible to pest infestation. This predisposition or susceptibility may arise from genetic factors or other factors such as environmental conditions (e.g., high humidity, temperature extremes), stress, poor nutrition etc. The plant’s vulnerability may pertain to specific plant-pest combinations or may encompass such combinations. According to specific embodiments, the plant is located in an area known to be at risk of the pest infestation or confirmed to be infested with the pest. For example, this could apply to areas where the pest has been identified on neighboring plants, or where environmental conditions (e.g., high humidity) favor reproduction of the pest.

[0125] According to specific embodiments, the plant is growing under conditions of high humidity, such as relative humidity levels exceeding 70%, 75%, 80%, 85%, 90%, or 95%.

[0126] According to specific embodiments, the plant is growing under drought stress.

[0127] According to specific embodiments, the plant is growing under conditions of temperature extremes, including unusually high or low temperatures, such as temperatures exceeding 35 °C, 36 °C, 37 °C, 38 °C, 39 °C, 40°C, or 45°C, or falling below 5 °C, 0 °C, -5 °C or -10 °C.

[0128] According to specific embodiments, the plant is experiencing nutritional deficiency, such as low levels of nitrogen, phosphorus, potassium and / or elements such se, Fe, Mg. Ca, Zn, Mn, Mo, Si, B, Cu, Ni (boron, chlorine, copper, iron, manganese, molybdenum, nickel, and zinc).

[0129] According to specific embodiments, the plant is cultivated in poor soil quality or degraded soil.

[0130] According to specific embodiments, the plant is subjected to mechanical or chemical stress, including pruning, wind damage, salinity stress, herbicide exposure, or physical damage.

[0131] According to specific embodiments, the plant is part of a monoculture or dense planting, which may facilitate pest propagation.

[0132] According to specific embodiments, the plant is planted in an area known to be infested with the pest.

[0133] According to specific embodiments, the plant is located near other host plants known to harbor the pest.

[0134] According to specific embodiments, the plant is cultivated in an area with a known history of recurring pest outbreaks.

[0135] According to specific embodiments, the area is a tropical region. According to specific embodiments, the area is a sub-tropical region.

[0136] According to specific embodiments, the area is temperate region.

[0137] The methods disclosed herein comprise contacting the plant with a pesticidal composition.

[0138] As used herein the term “pesticidal composition” refers to a composition capable of preventing, inhibiting or reducing the ability of a pest to feed, grow, move, spread, develop, survive and / or reproduce; and / or to mitigate pest-related damage. Such ability may be manifested by e.g., killing of the pest; decreasing pest survival or longevity; suppressing pest's fecundity and / or fertility; inhibiting or arresting pest's feeding, growth, development and / or mobility; alleviating plant symptoms caused by the pest; and / or preventing infestation by pest.

[0139] The pesticidal composition described herein is obtained or derived from an isolated Bacillus velezensis strain BX77 or a homolog thereof.

[0140] As used herein, the phrase “derived from an isolated Bacillus velezensis strain BX77 or a homolog thereof’ refers to a composition that is produced by, extracted from, isolated from, secreted by, cultured from, or otherwise obtained using a Bacillus velezensis strain or a homolog thereof, and encompasses cellular forms of the organism (e.g., viable cells, dormant endospores, biofilms, dead cells etc.) and cells-derived preparations (e.g., cell-free supernatants, lysates, extracts, cell fractions etc.), as further described herein.

[0141] According to specific embodiments, the pesticidal composition is obtained from bacterial cells of Bacillus velezensis strain BX77 or a homolog thereof.

[0142] According to other specific embodiments, the pesticidal composition comprises purified components obtained from cultures of Bacillus velezensis strain BX77 or a homolog thereof.

[0143] As used herein, the Bacillus velezensis strain BX77 is defined by a genome sequence set forth in SEQ ID NO: 1 (deposited as Accession No. CP166773 at GenBank) or a homolog of SEQ ID NO: 1 (referred to herein as “BX77 homolog”), such as disclosed in Gollop R, et al. Front Plant Sci. 2024 15: 1287184, doi: 10.3389 / fpls.2024.1287184, PMID: 38313804, and International Patent Application Publication No. WO2022 / 195585.

[0144] As used herein, the term “BX77 homolog” refers to a bacterial strain having a genomic sequence which is at least 99.1 % identical to the genome sequence set forth in SEQ ID NO: 1, and maintains the pesticidal activity of Bacillus velezensis strain BX77.

[0145] According to specific embodiments, the homolog exhibits at least 99.2%, 99.3%, 99.4%, 99.5%, 99.6%, 99.7%, 99.8%, or 99.9% sequence identity to SEQ ID NO: 1. According to specific embodiments, the sequence identity refers to global identity, an identity over the entire nucleic acid sequences disclosed herein and not over portions thereof (e.g., whole-genome sequence).

[0146] Sequence identity or homology can be determined using any nucleic acid sequence alignment algorithm such as BlastN, ClustalW, or MUSCLE or by genome-wide similarity measures such as OrthoANI.

[0147] According to specific embodiments, sequence identity or homology is determined by genome-wide similarity measures such as OrthoANI.

[0148] According to specific embodiments, the pesticidal composition is derived from BX77 strain defined by a genome sequence as set forth in SEQ ID NO: 1.

[0149] According to specific embodiments, the pesticidal composition comprises live cells of Bacillus velezensis strain BX77 or a homolog thereof. Such cells may be present in various forms, including isolated planktonic cells, a biofilm, vegetative cells, and / or dormant endospores, or as part of a cell broth comprising both the cells and the liquid medium in which they were cultured.

[0150] According to specific embodiments, the pesticidal composition comprises a biofilm of a Bacillus velezensis strain BX77 or a homolog thereof.

[0151] As used herein, the term "biofilm” refers to an aggregate of living Bacillus velezensis strain BX77 or a homolog thereof cells which adhere to each other and / or immobilized onto a surface as colonies. The bacterial cells in a biofilm are typically embedded within a self-secreted matrix of extracellular polymeric substance (EPS), which is a polymeric sticky mixture of nucleic acids, proteins and polysaccharides. The cells of Bacillus velezensis strain BX77 or a homolog thereof growing in a biofilm are physiologically distinct from cells in the "planktonic form" of the same organism, which by contrast, are single-cells that may float or swim in a liquid medium.

[0152] Methods of detecting and analyzing a biofilm are well known in the art and include, but are not limited to, microscopy [e.g., Atomic Force Microscopy, Transmitting Electron Microscopy, Scanning Transmitting Electron Microscopy, light microscopy, epifluorescence microscopy, scanning electron microscopy, confocal microscopy), histology, histochemistry, immunohistochemistry, micro-CT, X-ray diffraction (XRD) and FTIR].

[0153] According to specific embodiments, the pesticidal composition comprises a vegetative cell and / or dormant endospore of the Bacillus velezensis strain BX77 or a homolog thereof. As used herein, the term “vegetative cell” refers to a Bacillus velezensis strain BX77 or a homolog thereof cell that is metabolically active. These cells are capable of growth, reproduction, consuming nutrients, and performing all necessary functions for life.

[0154] As used herein, the term “dormant endospore” refers to a metabolically inactive non- reproductive structure formed by Bacillus velezensis strain BX77 or a homolog thereof as a survival mechanism, which is highly resistant compared to a vegetative cell. Once favorable conditions return, dormant endospores can germinate back into vegetative cells.

[0155] According to specific embodiments, the pesticidal composition comprises a cell broth collected from a culture of the Bacillus velezensis strain BX77 or a homolog thereof.

[0156] As defined herein, “cell broth”, refers to a liquid culture containing both cells of Bacillus velezensis strain BX77 or a homolog thereof and a liquid medium. It should be noted that a cell broth can be obtained by growing the bacteria in a liquid medium, or by suspending (also referred to as harvesting) Bacillus velezensis strain BX77 or a homolog thereof grown on an agar plate in a liquid medium.

[0157] According to specific embodiments, the pesticidal composition comprises cells of Bacillus velezensis strain BX77 or a homolog thereof and the liquid medium they were cultured in (i.e., unwashed cells).

[0158] According to specific embodiments, the pesticidal composition comprises dead cells, an extract, a cell fraction or lysate of the Bacillus velezensis strain BX77 or a homolog thereof.

[0159] Such materials may be obtained by disrupting the bacterial cells using physical, chemical, or enzymatic methods well known in the art, including but not limited to sonication, mechanical shear, freeze-thaw cycling, enzymatic digestion, or chemical lysis. The resulting preparations may contain proteins, nucleic acids, lipids, polysaccharides, or other metabolites, and may be used either as crude mixtures or following enrichment or separation of particular components, such as a cell wall fraction, membrane fraction, cytoplasmic fraction, or spore coat. The extract, cell fraction or lysate may further undergo a process of sterilization and / or filtration.

[0160] According to specific embodiments, the pesticidal composition comprises a cell free supernatant or filtrate obtained following culturing of the Bacillus velezensis strain BX77 or a homolog thereof.

[0161] Such a preparation may be obtained by removing the bacterial cells from a culture using methods well known in the art including, but not limited to, centrifugation, sedimentation, decanting, or filtration. The resulting liquid fraction may contain secreted proteins, peptides, nucleic acids, secondary metabolites, lipids, polysaccharides, or other bioactive substances produced during bacterial growth, which exhibit pesticidal activity. The cell-free supernatant or filtrate may further undergo a process of sterilization and / or filtration to ensure removal of residual cells and to improve stability. According to specific embodiments, the pesticidal composition is obtained following culturing of the Bacillus velezensis strain BX77 or a homolog thereof.

[0162] Thus, according to specific embodiments, the methods disclosed herein comprise culturing the Bacillus velezensis strain BX77 or a homolog thereof under prior to the contacting.

[0163] Culturing may be carried out under conditions suitable for maintaining the strain in culture and for supporting or enhancing the generation of pesticidal activity. Such conditions generally include selection of an appropriate medium, incubation temperature, pH, aeration, agitation, and duration of growth, as further described herein and exemplified in the Examples section.

[0164] According to specific embodiments, the medium is a solid medium. Non-limiting examples include LB agar, MRS agar, nutrient agar, Czapek agar, CASO agar, mannitol agar, marine agar, oatmeal agar, saccharose agar, and tomato juice agar.

[0165] According to other specific embodiments, the medium is a liquid medium. Non-limiting examples include lysogeny broth (LB), nutrient broth (NB), de Man-Rogosa-Sharpe medium (MRS), LBGS, TSB (tryptic soy broth), Terrific Broth, M9 minimal medium, M17 medium, Czapek medium, CASO medium, glucose-yeast extract medium, mannitol broth, marine broth, oatmeal medium, saccharose medium, SOB medium, SOC medium, tomato juice medium, whey medium, YPD (yeast extract-peptone-dextrose), YPG (yeast extract-peptone-galactose), YPM (yeast extract-peptone-maltose), and YMG (yeast-malt extract with glucose).

[0166] According to specific embodiments, the medium is selected from the group consisting of LB, NB and MRS.

[0167] According to specific embodiments, medium is a LB medium.

[0168] According to specific embodiments, culturing is carried out for at least 6, 12, 24, 36, 48, or 56 hours.

[0169] According to specific embodiments, culturing is carried out for at least 36 hours.

[0170] According to specific embodiments, culturing is carried out for up to 1 week, up to 144 hours, up to 120 hours, up to 96 hours, or up to 72 hours or up to 56 hours.

[0171] According to specific embodiments, culturing is carried out for up to 72 hours.

[0172] According to specific embodiments, culturing is carried out for 24-96 hours, 24-72 hours, 36-72 hours, or 36-56 hours.

[0173] According to specific embodiments, the culturing temperature is within a range of 20-40 °C, such as 25-35 °C, 28-32 °C, 28-35 °C, 30-35 °C, or 28-37 °C.

[0174] According to specific embodiments, the culturing temperature is in the range of 28-32 °C According to specific embodiments, the culturing temperature is about 30 °C.

[0175] According to specific embodiments, the culturing conditions comprise shaking or agitation, which terms are used interchangeably herein and refer to the mechanical movement of the culture vessel to maintain homogeneity of the culture, improve aeration, and promote nutrient availability. Methods of providing agitation are well known in the art and include, without limitation, orbital shaking of flasks, mechanical stirring in bioreactors, and impeller- driven mixing in fermenters.

[0176] According to specific embodiments, the shaking speed may be at least 50, 80, 100, 120, 150, 180, 200, 250, 300, 350, or 400 rpm, or within ranges such as 50-200, 80-200 rpm, 100- 200 rpm, 150-250 rpm, 200-300 rpm, or 250-400 rpm.

[0177] According to specific embodiments, the shaking is in a range of 100-200 rpm.

[0178] According to specific embodiments, the culturing is for 36-56 hours, at 28-32 °C, in a LB medium while shaking at 100-200 rpm.

[0179] According to other specific embodiments, static incubation is used. Such conditions may promote pellicle biofilm formation at the air-liquid interface.

[0180] According to specific embodiments, the pesticidal composition is obtained following culturing of the Bacillus velezensis strain BX77 or a homolog thereof under conditions allowing biofilm formation and / or sporulation. Such conditions are described herein and also exemplified in the Examples section which follows (see e.g., Example 8).

[0181] According to specific embodiments, culturing is continued to the late logarithmic to stationary growth phase and the culture is harvested at that stage.

[0182] According to specific embodiments, the late logarithmic to stationary growth phase is optimal for pesticidal activity.

[0183] Other culturing parameters, such as pH, aeration, inoculum density, the type of culture vessel and the like, may be adjusted as appropriate, and such adjustments are within the routine abilities of a person skilled in the art.

[0184] According to specific embodiments, the culturing is carried out in a bioreactor.

[0185] In such embodiments, a standard equipment may be used to maintain controlled conditions of temperature, pH, aeration, and mixing, and the system may be operated in batch, fed-batch, or continuous modes. Selection of reactor type (e.g., stirred-tank, airlift, packed-bed, or flow-cell) and operating parameters (including agitation speed, gas flow, feed strategy, and working volume) is within the routine abilities of a person skilled in the art. According to specific embodiments, internal supports or carriers are included to permit biofilm formation. Parameters such as shear and residence time may be adjusted to influence growth characteristics and the yield of pesticidal factors.

[0186] Once the pesticidal composition is obtained, it is to be contacted with the plant.

[0187] The pesticidal composition may be formulated, by methods well known in the art, as a solid, liquid, semi- solid, gel, emulsion, or powder, including seed-coating formulations and formulations suitable for application to plants or to a plant growth medium. Formulation aspect are well known in the art. Non-limiting examples that can be used with specific embodiments of the invention are provided infra.

[0188] According to specific embodiments, the pesticidal composition is formulated as a liquid, solid, semi- solid, gel or powder.

[0189] According to specific embodiments, the pesticidal composition is formulated as a liquid formulation.

[0190] When the pesticidal composition is prepared as a liquid formulation for application to plants, it can be prepared in a concentrated formulation (e.g., emulsifiable concentrate, capsule suspension, soluble liquid, flowable concentrate) or a ready-to-use (working) formulation. Nonlimiting examples of liquid formulations include aqueous solutions, aqueous suspension concentrates (SC), soluble liquids (SL), flowable concentrates, capsule suspensions (CS), oil-in- water emulsions (EW), water-in-oil emulsions (WO), microemulsions (ME), suspoemulsions (SE), ultra-low-volume (ULV) oil formulations, and chemigation concentrates. Non-limiting examples of liquid carriers include water, buffered water (e.g., phosphate- or citrate-buffered), saline, culture medium or conditioned medium, and oils such as vegetable oils (e.g., soybean, canola, com, sunflower) or mineral / paraffinic oils, as well as mixtures thereof to form EW or WO systems. In embodiments comprising live cells or spores, the liquid carrier may be physiologically compatible and may include stabilizers (e.g., glycerol, trehalose) to preserve viability during storage and use.

[0191] According to specific embodiments, the pesticidal composition is formulated as a dry formulation.

[0192] When the pesticidal composition is prepared as a solid formulation for application to plants, it can be prepared as a granular formulation or a powder agent (e.g., wettable powder, water-dispersible granules, granules, dry flowables, dusts, pellets, tablets, briquettes, baits). Non-limiting examples of solid carriers and excipients include clays (e.g., kaolin, bentonite), talc, diatomaceous earth, silica, calcium carbonate, starches, maltodextrin, microcrystalline cellulose, lignosulfonates, and polymeric binders, optionally with wetting agents and dispersants to facilitate re-suspension. Dry formulations may be produced by spray-drying, fluid-bed granulation, extrusion, lyophilization / freeze-drying, or drum / vacuum drying. Where reconstitution is intended, the product may be designed for dispersion in water prior to application. In embodiments comprising live cells or spores, the dry formulation may include protectants or stabilizers (e.g., trehalose, skim milk powder, sucrose, or maltodextrin) and be packaged to limit moisture uptake.

[0193] According to specific embodiments, the pesticidal composition is provided in a form suitable for agricultural application, including dips, sprays, seed coatings, concentrates, dusts, drenches, chemigation concentrates, fogs, or mists. Formulations may be prepared by conventional means (e.g., lyophilization, freeze-drying, desiccation, or dispersion in aqueous carriers, buffers, or oils).

[0194] According to specific embodiments, the formulated compositions may be in the form of a dust or granular material, powder, gel, cream (where appropriate for nursery or protected-culture uses), paste, pellet, tablet or a suspension in oil (vegetable or mineral), or water or oil / water emulsions, capsule suspension, emulsifiable concentrate, or as a wettable powder, wettable granules, water dispersible granules, aerosols, foam, slurries or flowable concentrates

[0195] According to specific embodiments, concentrates (e.g., liquid concentrate, slurry, or “wet cake”) are supplied for end-user dilution, dispersion, suspension, emulsification, or reconstitution prior to use.

[0196] According to specific embodiments, the pesticidal composition is formulated for delivery by spraying, irrigation and / or fumigation , and in certain embodiments by chemigation, seed treatment, in-furrow application, root dip, soil drench, banded application, or foliar misting.

[0197] According to specific embodiments, the compositions disclosed herein are stable, both during storage and during utilization, meaning that the integrity of the composition is maintained under storage and / or utilization conditions of the composition, which may include elevated temperatures, freeze-thaw cycles, dilution, reconstitution, changes in pH or in ionic strength, UV-irradiation, presence of harmful chemicals and the like.

[0198] According to specific embodiments, the integrity and activity of the composition is maintained under open field or controlled conditions, e.g., greenhouse.

[0199] It should be noted that the compositions disclosed herein can be formulated with various carriers designed to increase e.g. delivery, stability, permeability and the like.

[0200] A "carrier", as used herein, means any solid, semi-solid or liquid carrier in or on(to) which a pesticidal composition can be suitably incorporated, included, immobilized, adsorbed, absorbed, bound, encapsulated, embedded, attached, or comprised. Non-limiting examples of such carriers include nanocapsules, microcapsules, nanospheres, microspheres, nanoparticles, microparticles, liposomes, vesicles, beads, a gel, weak ionic resin particles, cochleate delivery vehicles, small granules, granulates, nano-tubes, bucky-balls, water droplets that are part of a water-in-oil emulsion, oil droplets that are part of an oil-in-water emulsion, organic materials such as cork, wood or other plant-derived materials (e.g. in the form of seed shells, wood chips, pulp, spheres, beads, sheets or any other suitable form), paper or cardboard, inorganic materials such as talc, clay, microcrystalline cellulose, silica, alumina, silicates and zeolites, or even microbial cells (such as yeast cells) or suitable fractions or fragments thereof.

[0201] According to specific embodiments, the carriers are such that they have immediate or gradual or slow-release characteristics, for example over several minutes, several hours, several days or several weeks. Also, the carriers may be made of materials (e.g. polymers) that rupture or slowly degrade (for example, due to prolonged exposure to high or low temperature, sunlight, high or low humidity or other environmental factors or conditions) over time (e.g. over minutes, hours, days or weeks) and so release the pesticidal composition from the carrier. According to specific embodiments, the carrier is coupled, bound, linked or otherwise attached to or associated with the pesticidal composition. According to some embodiments, coupling is covalent; according to others, association is non-covalent (e.g., adsorption or entrapment).

[0202] The compositions disclosed herein may be provided per-se, or may be formulated as an agrochemical composition that is mixed with suitable agriculturally acceptable carriers or excipients.

[0203] Herein the term "active ingredient" refers to the pesticidal composition obtained or derived from Bacillus velezensis strain BX77 that is responsible for the pesticidal effect.

[0204] An "agrochemical formulation" as used herein means a composition for agricultural use, comprising one or more of the active ingredients described with other chemical components such as agriculturally acceptable carriers and excipients.

[0205] As used herein the term "agriculturally acceptable carrier" refers to a material that facilitates application of the pesticidal composition disclosed herein to the plant. According to specific embodiments, the term covers all adjuvants, e.g., inert components, dispersants, surfactants, tackifiers, binders, etc. that are ordinarily used in insecticide formulation technology and are well known to the skilled artisan. Carriers used in compositions for application to plants and plant material are preferably non- phytotoxic or only mildly phytotoxic. A suitable carrier may be a solid, liquid or gas depending on the desired formulation. Suitable carriers include mineral or vegetable oils, water, clays, silica, calcium carbonate, starches, polymeric binders, and conventional formulation aids such as surfactants, dispersants, wetting agents, thickeners, anti-caking agents, stabilizers, preservatives, colorants, and coatings; selection is within routine skill and may consider strain, crop, soil, climate, and intended use pattern.

[0206] Herein the term "excipient" refers to an inert substance added to a composition to further facilitate administration of an active ingredient. Examples, without limitation, of excipients include calcium carbonate, calcium phosphate, various sugars and types of starch, cellulose derivatives, surfactant, gelatin, vegetable oils, polyethylene glycols, wetting agents, spreading agents, buffering agents, acidifiers and the like.

[0207] Additional excipients and formulation aids may include oils (vegetable, mineral, or heavy petroleum distillates), gums and resins, clays and silicates (e.g., kaolin, bentonite, diatomaceous earth), polyoxyethylene and polyethylene glycol derivatives, terpenes, fatty acid esters, sulfated alcohols, sulfonates (alkyl or petroleum), alkyl phosphates, aryl / alkyl polyoxyethylene glycols, alkyl amine acetates, polyhydric alcohols, and anti-caking salts (e.g., sodium sulfite or sulfate, naphthalene sulfonate salts, calcium carbonate), used singly or in combination.

[0208] According to specific embodiments, the pesticidal compositions disclosed herein may be the only active substance in the composition.

[0209] According to other specific embodiments, the composition comprises one or more additional active agents in addition to the pesticidal composition described herein. Non-limiting examples of such agents include herbicides, other pesticides, plant growth regulators, safeners, fertilizers, micronutrients, biostimulants, attractants / repellents and the like.

[0210] According to specific embodiments, the pesticidal composition and the additional active agent are in a co-formulation.

[0211] According to specific embodiments, the pesticidal composition and the additional active agent are in separate containers.

[0212] As used herein the term "contacting" refers to the positioning or applying the pesticidal composition(s) disclosed herein such that they are in direct or indirect contact with the plant and, in some embodiments, with the target pest.

[0213] According to specific embodiments, contacting is effected via direct application to the plant (e.g., seed treatment, canopy spray, foliar misting etc.).

[0214] According to specific embodiments, contacting is effected by indirect application to plant-associated substrates (e.g., soil or growth medium, irrigation water, hydroponic systems etc.).

[0215] Application may occur prophylactically or therapeutically, i.e., before or after pest presence is detected.

[0216] According to specific embodiments, the contacting is with a pest infested plant. According to other specific embodiments, the contacting is with a plant at risk of pest infestation prior to detection of infestation.

[0217] Contacting can be effected using any suitable method known in the art, including without limitation spraying, spreading, wetting, immersing, dipping, coating / painting, dusting, soaking, drenching, foaming, fogging, misting, dressing, encrusting, banded application, soil drench, infurrow application, or irrigation / chemigation.

[0218] According to specific embodiments, contacting is performed by spraying, immersing, drenching, coating, encapsulating, dusting, and / or fumigating.

[0219] According to specific embodiments, contacting is by spraying.

[0220] According to specific embodiments, contacting is effected by spraying, irrigating and / or fumigation.

[0221] The contacting may be effected at any stage of the plant life cycle. According to specific embodiments, upon the contacting the plant is at a post-flowering stage, a flowering stage, a preflowering stage, or any combination thereof.

[0222] According to specific embodiments, contacting is onto or in the vicinity of a root, a stem, a trunk, a seed, a fruit, a flower, a leaf, or any combination thereof.

[0223] According to specific embodiments, contacting is by spraying a canopy of the plant and / or by drenching a root zone of said plant.

[0224] According to an embodiment, application is carried out in an open field.

[0225] According to an embodiment, application is carried out in a greenhouse.

[0226] In some embodiments, application is carried out in protected cultivation or soilless / hydroponic systems.

[0227] Selection of dose, timing, and frequency is within the routine abilities of a skilled person and may be adjusted based on crop, growth stage, pest species and pressure, environmental conditions, and application method.

[0228] According to specific embodiments, the dose regimen is performed such as to control the spread of a pest and / or eliminate a pest and / or eliminate / reduce / minimize any damage that can be caused by the pest.

[0229] According to an embodiment, the pesticidal composition is provided in a concentration of at least 106, at least 107, at least 108or at least 109CFU / ml.

[0230] According to an embodiment, the pesticidal composition is provided in a concentration of at least 107CFU / ml.

[0231] According to an embodiment, the pesticidal composition is provided in a concentration of at least 108CFU / ml. According to specific embodiments, the pesticidal composition is provided in a concentration of up to IO10, 109or 108CFU / ml

[0232] According to an embodiment, the pesticidal composition is provided in a concentration of 106- IO10, 107- 109or 107- 108CFU / ml.

[0233] According to specific embodiments, the application is carried out at least once.

[0234] According to specific embodiments, the applying comprises repeated application (2 or more applications e.g., every day, every other day, twice a week, every week, every two weeks, twice a month, every month, seasonal, etc.). Repeated applications are especially envisaged for field / greenhouse treatments.

[0235] According to an embodiment, the composition is applied daily, bi-weekly, weekly, bimonthly or monthly.

[0236] According to specific embodiments, contacting is effected once to twice a week.

[0237] According to specific embodiments, contacting is performed at intervals of about 3-21 days (e.g., 7-14 days), with a total of 1-6 applications per season, and may be timed relative on a calendar basis or to agronomic events (e.g., seeding, planting, transplanting, pre-flowering, flowering, or fruit set) and / or pest monitoring thresholds.

[0238] According to specific embodiments, re-application may be performed following significant rainfall or irrigation events, or as dictated by pest pressure and label constraints.

[0239] As used herein the term “about” refers to ± 10 %.

[0240] The terms "comprises", "comprising", "includes", "including", “having” and their conjugates mean "including but not limited to".

[0241] The term “consisting of’ means “including and limited to”.

[0242] The term "consisting essentially of" means that the composition, method or structure may include additional ingredients, steps and / or parts, but only if the additional ingredients, steps and / or parts do not materially alter the basic and novel characteristics of the claimed composition, method or structure.

[0243] As used herein, the singular form "a", "an" and "the" include plural references unless the context clearly dictates otherwise. For example, the term "a compound" or "at least one compound" may include a plurality of compounds, including mixtures thereof.

[0244] Throughout this application, various embodiments of this invention may be presented in a range format. It should be understood that the description in range format is merely for convenience and brevity and should not be construed as an inflexible limitation on the scope of the invention. Accordingly, the description of a range should be considered to have specifically disclosed all the possible subranges as well as individual numerical values within that range. For example, description of a range such as from 1 to 6 should be considered to have specifically disclosed subranges such as from 1 to 3, from 1 to 4, from 1 to 5, from 2 to 4, from 2 to 6, from 3 to 6 etc., as well as individual numbers within that range, for example, 1, 2, 3, 4, 5, and 6. This applies regardless of the breadth of the range.

[0245] Whenever a numerical range is indicated herein, it is meant to include any cited numeral (fractional or integral) within the indicated range. The phrases “ranging / ranges between” a first indicate number and a second indicate number and “ranging / ranges from” a first indicate number “to” a second indicate number are used herein interchangeably and are meant to include the first and second indicated numbers and all the fractional and integral numerals therebetween.

[0246] As used herein the term "method" refers to manners, means, techniques and procedures for accomplishing a given task including, but not limited to, those manners, means, techniques and procedures either known to, or readily developed from known manners, means, techniques and procedures by practitioners of the chemical, pharmacological, biological, biochemical and medical arts.

[0247] As used herein, the term “treating” includes abrogating, substantially inhibiting, slowing or reversing the progression of a condition, substantially ameliorating clinical or aesthetical symptoms of a condition or substantially preventing the appearance of clinical or aesthetical symptoms of a condition.

[0248] When reference is made to particular sequence listings, such reference is to be understood to also encompass sequences that substantially correspond to its complementary sequence as including minor sequence variations, resulting from, e.g., sequencing errors, cloning errors, or other alterations resulting in base substitution, base deletion or base addition, provided that the frequency of such variations is less than 1 in 50 nucleotides, alternatively, less than 1 in 100 nucleotides, alternatively, less than 1 in 200 nucleotides, alternatively, less than 1 in 500 nucleotides, alternatively, less than 1 in 1000 nucleotides, alternatively, less than 1 in 5,000 nucleotides, alternatively, less than 1 in 10,000 nucleotides.

[0249] It is appreciated that certain features of the invention, which are, for clarity, described in the context of separate embodiments, may also be provided in combination in a single embodiment. Conversely, various features of the invention, which are, for brevity, described in the context of a single embodiment, may also be provided separately or in any suitable subcombination or as suitable in any other described embodiment of the invention. Certain features described in the context of various embodiments are not to be considered essential features of those embodiments, unless the embodiment is inoperative without those elements. Various embodiments and aspects of the present invention as delineated hereinabove and as claimed in the claims section below find experimental support in the following examples.

[0250] EXAMPLES

[0251] Reference is now made to the following examples, which together with the above descriptions illustrate some embodiments of the invention in a non-limiting fashion.

[0252] Generally, the nomenclature used herein and the laboratory procedures utilized in the present invention include molecular, biochemical, microbiological and recombinant DNA techniques.

[0253] EXAMPLE 1

[0254] CONTROL OF BOTRYTIS CINEREA

[0255] MATERIALS AND METHODS

[0256] Preparation of bacterial compositions - A bacterial isolate, BX77, was isolated from food products, characterized by whole genome sequencing, and identified as Bacillus velezensis. BX77 genome sequence is provided in SEQ ID NO: 1 (deposited as Accession No. CP166773 at GenBank). Following, several BX77 preparations were prepared, as detailed in the description of Figures 1A-D.

[0257] Evaluating the pesticidal effect on tomato plants - Tomato plants were grown from seedlings at the age of 4-5 weeks after seeding. The plants grown from seeds were transplanted to 750 ml pots and maintained in a greenhouse for 3-4 weeks until exposure to the bacterium treatments. Suspension of the pest Botrytis cinerea conidia served as inoculum for infection of tomato leaves on whole plants or on detached leaves. Specifically, drops of B. cinerea conidia suspension (5xl05 / ml) were placed on leaves attached to the plants. Inoculated whole plants were incubated in a humidity chamber at a temperature of 22 ± 1 °C in an illuminated walk-in growth chamber. Detached leaves were used in some assays. For the detached leaves assay, following treatments with the bacterium as mentioned above, leaves were detached from the treated plants, placed in a humidity chamber, inoculated as mentioned above and incubated as mentioned above.

[0258] The bacterium was applied 4 days and 4 hours (= day 0) before infection. Drench treatments were applied by pouring 30 ml into the root zone of each of the treated plants. Spray treatment to the canopy of the leaves of the plants were applied using 10 ml to each plant. Five to six replicate plants were used for each bacterium treatment and 5 to 6 plants served as untreated control. Control plants were treated with water. The rot development under each drop was evaluated on a 0-100% scale where 0=symptomless plant tissue and 100=full rot development. Area Under the Disease progress Curve (AUDPC) was calculated for each plant and averaged for each treatment.

[0259] Additional description on Botrytis evaluation is provided in Swartzberg, D., Kirshner, B., Elad, Y. and Granot, D. (2008) European Journal of Plant Pathology 120:289-297. DOI: 10.1007 / S10658-007-9217-6.

[0260] RESULTS

[0261] Botrytis cinerea is a fungal pest of plants that infects more than 1400 species. It is a necrotroph that thrives at high humidity and infects plants at optimal temperatures of up to 25 °C. The typical symptoms are rot and gray mold. Important crops that are affected are grape, tomato, cucumber and many more species.

[0262] Drench and spray treatments reduced gray mold on whole tomato plants (Figure 1A) and on detached leaves (Figure 1B-C).

[0263] In the next step, the effect of various BX77-derived compositions (e.g., alive cells, alive and washed, heat killed, heat killed and washed, supernatant). As shown in Figure ID, all bacterial-derived preparations had a pesticidal activity.

[0264] EXAMPLE 2

[0265] CONTROL OF SCLEROTINIA SCLEROTIORUM

[0266] MATERIALS AND METHODS

[0267] Preparation of bacterial compositions - As described in Example 1, with the following modifications: isolated BX77 was grown for 48 hours at 30+1 °C in a Lysogeny broth (LB) medium with 150 rpm shaking.

[0268] Evaluating the pesticidal effect on cucumber plants - Cucumber (cv. Mani) were grown from seeds in 800 ml pots containing growth medium. Sclerotinia sclerotiorum was grown in PDA Petri dishes and discs taken from the edges of 4 days old culture of the fungus were placed on cucumber leaves. Inoculated plants were incubated in a humidity chamber at a temperature of 22 + 1 °C in an illuminated walk-in growth chamber.

[0269] The bacterium was drench-applied 3 days before infection. Five to six replicate plants were used for each bacterium treatment and 5 to 6 plants served as untreated control, water treated.

[0270] Disease was evaluated by measuring the diameter of the developed rot. RESULTS

[0271] Sclerotinia sclerotiorum is a fungal pathogen of hundreds of plant species. It causes white mold on crops such as cucumber, tomato, lettuce, rape seed. The pest sclerotia survives for many years and forms appotecia fruiting bodies that release spores. Infection occurs at high humidity and optimal temperatures of 15-25 °C.

[0272] Drench treatment reduced white mold on whole cucumber plants (Figure 2).

[0273] EXAMPLE 3

[0274] CONTROL OF OIDIUM NEOLYCOPERSICI

[0275] MATERIALS AND METHODS

[0276] Preparation of bacterial-derived compositions - Several BX77 preparations were prepared, as detailed in the description of Figures 3A-D.

[0277] Evaluating the fungicidal effect on tomato plants - Tomato plants were kept in a greenhouse. The fungal pest Oidium neolycopersici infested the plants naturally. Disease symptoms were evaluated on a 0-100% severity index where 0=no symptoms and 100=leaves fully covered by symptoms of the disease. The above described bacterial-derived compositions were applied weekly by spray to the canopy of tomato plants. Five to six replicate plants were used for each bacterium treatment and 5 to 6 plants served as untreated control (solely water). RESULTS

[0278] Powdery mildew of tomato is caused by the obligate fungal pathogen Oidium neolycopersici. Natural infection by conidia occurred in the greenhouse and the typical whitish symptoms were observed on the leaves. Following treatments, the severity of coverage of the symptoms on leaves of whole plants was evaluated.

[0279] Spray treatment with BX77 bacterial preparation reduced the disease at all concentrations tested (ranging from 106to 108 / ml, Figure 3A).

[0280] In the next step, the effect of whole cell bacterial culture on powdery mildew was compared with the effect of other BX77-derived compositions (e.g., cells of the bacterium washed of the supernatant and with the supernatant containing no bacteria). As shown in Figures 3B-D, all bacterial-derived preparations had a pesticidal activity in three different tomato plants. EXAMPLE 4

[0281] CONTROL OF BEMISIA T ABACI

[0282] MATERIALS AND METHODS

[0283] Preparation of bacterial-derived compositions - Several BX77 preparations were prepared, as detailed in the description of Figures 4A-D.

[0284] Evaluating the insecticidal effect on tomato plants - Tomato plants were kept in a glass- covered greenhouse. The insect pest Bemisia tabaci appeared naturally in the greenhouse where tomato plants were grown. The number of individual adults of the B. tabaci was counted on the lower side of the 7thleaf. Three weekly applications of the above described bacterial-derived compositions were applied by drench or spray to the canopy of the plant (Bacterial cone. 108) and adult B. tabaci density was evaluated by counting one week following each treatment, and prior to the next treatment. Five to six replicate plants were used for each bacterium treatment and 5 to 6 plants served as untreated control (water treated). Additionally, treatment of a single leaf was performed by spraying the bacteria at concentrations of 106to 109 / ml on selected leaves in two weekly applications. Natural silverleaf whitefly population developed and counted 5 days after the second application. Silverleaf whitefly individuals were counted on leaf 6 that was treated by the bacterial suspensions and on leaf 8 that was not sprayed at all, in order to test systemic effect. For comparative analysis, the commercial product Serenade ASO (Bacillus subtilis group BM02; BAYER, Crop Science, U.S.) was applied as well by drench or spray to the canopy of the plants at a concentration of 1.0% preparation.

[0285] RESULTS

[0286] Silverleaf whitefly Bemisia tabaci Gennadius), also called sweet potato whitefly, occurred naturally on treated tomato plants. The insect causes damage to many plant crops by feeding on them and by transmission of virus plant pathogens. It sucks phloem liquid from the leaves, causes whitish dots, and secretes honeydew, promoting the development of sooty molds on plant canopies. Many crop plants may be affected.

[0287] Treatment by drench or spray to the canopy of tomato plants with various BX77-derived compositions suppressed Bemisia tabaci population on the tomato leaves of three different tomato plants (Figures 4A-C). Treating a single leaf (leaf 6) instead of treating the whole plants and counting the number of Bemisia individuals on each treated leaf and on untreated leaves showed a reduction of Bemisia on the untreated leaves (Figure 4D) indicating a systemic effect with a plant signal that moved from the treated leaves to the untreated leaves. Serenade did not affect the Bemisia incidence on both tested leaves. EXAMPLE 5

[0288] CONTROL OF TUTA ABSOLUTA

[0289] MATERIALS AND METHODS

[0290] Preparation of bacterial-derived compositions - Several BX77 preparations were prepared, as detailed in the description of Figures 5A-D.

[0291] Evaluating the pesticidal effect on tomato plants - Tomato plants were kept in a greenhouse at 18-28 °C. The moth Tula absoluta infested the plants naturally. The insect larva feed on tomato plants, especially on leaves, producing large galleries in the leaflets and feeding on young fruits and apical buds. Mining symptoms incidence and severity were evaluated on all leaves of each plant; small galleries had size of 1-3 mm, medium-sized galleries measured 4-6 mm and large galleries measured 7 mm dimeter and larger. Each size group of galleries was counted separately and the summation of all galleries sized according to the three sizes was calculated and described as number of all galleries. The severity of T. absoluta damage was calculated as follows: {T. absoluta Index = (number of small galleries) + (number of medium galleries)*3 + (number of medium galleries)*6}. Three weekly applications were of the above described bacterial-derived compositions were applied by drench or spray to the canopy of the plants. The number of small, medium and large galleries per plant were evaluated on a weekly basis. Five to six replicate plants were used for each bacterium treatment and 5 to 6 plants served as untreated control (water treated).

[0292] RESULTS

[0293] The moth Tuta absoluta Meyrick lay eggs on the leaves of tomato plants. Those eggs hatch to larva, which eat the leaves in a process known as leaf mining. Tomato is the main host plant, but T. absoluta also attacks other crop plants including potato, eggplant, pepper, tobacco, and other Solanaceous plants. The insect can cause total yield loss.

[0294] Treatment by drench or spray to the canopy of tomato plants with various BX77-derived compositions (bacterial concentration 108 / ml) reduced Tuta absoluta damage on the tomato leaves of three different tomato plants (Figures 5A-C). In addition, spray treatment of tomato plants at varied bacterial concentrations resulted in a significant reduction in the incidence of T. absoluta galleries by bacterium concentrations of 105-108 / ml; and a significant reduction in the severity of damage intensity by concentrations of 106-108 / ml (Figure 5D). EXAMPLE 6

[0295] CONTROL OF TETRANYCHUS URTICAE

[0296] MATERIALS AND METHODS

[0297] Preparation of bacterial-derived compositions - Several BX77 preparations were prepared, as detailed in the description of Figures 6A-B.

[0298] Evaluating the pesticidal effect on tomato plants - Tomato plants were kept in a greenhouse at 18-28 °C. The spider mite Tetranychus urticae infested the plants naturally. Five weekly applications of the above described bacterial-derived compositions (bacterial concentration 108 / ml) were applied by spray to the canopy of the tomato plants (days 0, 7, 14, 21, and 28). Typical scratching symptoms severity were evaluated on leaves at each plant; 0 = no damage (all leaves are symptomless) and 100 = leaves were fully affected by scratching symptoms. The number of mite individuals was assessed on the under-side of a leaf on each plant. Five to six replicate plants were used for each bacterium treatment and 5 to 6 plants served as untreated control (Water sprayed).

[0299] RESULTS

[0300] The red spider mite, Tetranychus urticae Koch, also known as the two-spotted spider mite, occurred naturally on the treated tomato plants. Tetranychus urticae Koch is a plant-feeding mite of the family Tetranychidae, which can feed on tomato, pepper, and potato. It sucks the cell contents in leaves causing whitish scratching symptoms on the leaf surface. Further, it reduces the photosynthetic ability of the plants, eventually causing leaf mortality and major yield losses.

[0301] Treatment by spray to the canopy of tomato plants with various BX77-derived compositions reduced the number of Tetranychus urticae mites and their induced damage on the tomato leaves of two different tomato plants (Figures 6A-B).

[0302] EXAMPLE 7

[0303] CONTROL OF TOMATO YELLOW LEAF CURL VIRUS

[0304] MATERIALS AND METHODS

[0305] Preparation of bacterial-derived compositions - Several BX77 preparations were prepared, as detailed in the description of Figure 7.

[0306] Evaluating the suppression effect on tomato plants - Tomato plants were kept in a greenhouse at 18-28 °C. Spread of the disease onto treated tomato plants was achieved by the migration of the insect vector, Bemisia tabaci, from the infected plants. Disease severity was evaluated on each plant according to a % coverage key, whereby 0% is defined as all leaves with no disease symptoms, and 100% is defined as all leaves fully covered by disease symptoms. Five weekly applications of the above described bacterial-derived compositions (bacterial concentration of 108 / ml) were applied by spray to the canopy of the tomato plants (days 0, 7, 14, 21 and 28). Five to six replicate plants were used for each bacterium treatment and 5 to 6 plants served as untreated control (Water sprayed).

[0307] RESULTS

[0308] Tomato yellow leaf curl virus (TYLCV) is a DNA virus from the genus Begomovirus, transmitted by an insect vector - Silverleaf whitefly (Bemisia tabaci). The infected plants include tomato, eggplant, potato, tobacco, and pepper. Virus infection causes stunting, reduction of leaf size, upward cupping / curling of leaves, chlorosis on leaves and flowers, and reduction of fruit production. It was found that spraying the bacterium.

[0309] Treatment by spray to the canopy of tomato plants with various BX77-derived compositions reduced the symptoms of the disease (Figure 7).

[0310] EXAMPLE 8

[0311] THE EFFECT OF DIFFERENT CULTURE CONDITIONS

[0312] The effects of different culturing conditions on the composition and the pesticidal activity of Bacillus velezensis 'Ql l'l cells were determined. The results are summarized in Figure 8A-B and Table 1 hereinbelow. It was found that growing the cells in Lysogeny broth (LB) at 30 °C for 48 hours triggers biofilm formation but not sporulation. Further, diluting the LB medium (1:3) induced efficient sporulation. Another tested growth medium - nutrient broth (NB) did not induce biofilm formation and induced some sporulation. BX77 cells grown under all conditions had pesticidal activity, however, the conditions affected its extent.

[0313] Table 1: The effect of different growth conditions on the level of BX77 sporulation, biofilm formation, and antagonistic activity against the gray mold disease and the pest Tula absoluta.

[0314] * - (no aggregates), + (aggregates < 50), ++( aggregates > 50), +++( aggregates > 50+ biofilm matrix).

[0315] EXAMPLE 9

[0316] CONTROL OF APHIS GOSSYPII

[0317] MATERIALS AND METHODS

[0318] Preparation of bacterial-derived compositions - Several BX77 preparations were prepared, as detailed in the description of Figure 9.

[0319] Evaluating the insecticidal effect on tomato plants - Tomato plants were kept in a greenhouse at 18-28 °C. The melon and cotton aphid Aphis gossypii infested the plants naturally. Two weekly applications of the above described bacterial-derived compositions (bacterial concentration 108 / ml) were applied by spray to the canopy of the tomato plants (days 20 and 27 after planting). The number of aphid individuals was assessed 5 days after the 2ndtreatment on the whole plant while observing all plants. Six replicates of 4 plants each were used for the bacterium treatment the same number of replicates and plants served as untreated control (Water sprayed).

[0320] RESULTS

[0321] The melon and cotton aphid, Aphis gossypii Koch, also known as the two- spotted spider mite, occurred naturally on the treated tomato plants. Aphis gossypii Glover is a plant-feeding aphid that, of the family Aphididae the which can feed on a variety of plant species in the families

[0322] Rosaceae, Chenopodiaceae, Malvaceae, Cruciferae, Cucurbitaceae, Solanaceae, Compositae and others. It sucks sap from plants, causes leaf curling and distortion and finally the foliage may become chlorotic and die prematurely, and major yield losses.

[0323] Treatment by spray to the canopy of tomato plants with the BX77-derived composition reduced the number of A. gossypii aphids on the tomato canopy (Figures 9).

[0324] Although the invention has been described in conjunction with specific embodiments thereof, it is evident that many alternatives, modifications and variations will be apparent to those skilled in the art. Accordingly, it is intended to embrace all such alternatives, modifications and variations that fall within the spirit and broad scope of the appended claims. It is the intent of the applicant(s) that all publications, patents and patent applications referred to in this specification are to be incorporated in their entirety by reference into the specification, as if each individual publication, patent or patent application was specifically and individually noted when referenced that it is to be incorporated herein by reference. In addition, citation or identification of any reference in this application shall not be construed as an admission that such reference is available as prior art to the present invention. To the extent that section headings are used, they should not be construed as necessarily limiting. In addition, any priority document(s) of this application is / are hereby incorporated herein by reference in its / their entirety.

Claims

WHAT IS CLAIMED IS:

1. A method of treating a pest infested plant, the method comprising contacting the plant with a pesticidal composition obtained or derived from an isolated Bacillus velezensis strain BX77 having a genome sequence as set forth in SEQ ID NO: 1 or a homolog thereof, thereby treating the pest infested plant.

2. A method of protecting a plant at risk of pest infestation, the method comprising contacting the plant or with a pesticidal composition obtained or derived from an isolated Bacillus velezensis strain BX77 having a genome sequence as set forth in SEQ ID NO: 1 or a homolog thereof, thereby protecting the plant at risk of pest infestation.

3. The method of any one of claims 1-2, wherein said pesticidal composition comprises live cells of said Bacillus velezensis strain BX77 or a homolog thereof.

4. The method of any one of claims 1-3, wherein said pesticidal composition comprises a biofilm of said Bacillus velezensis strain BX77 or a homolog thereof.

5. The method of any one of claims 1-2, wherein said pesticidal composition comprises a vegetative cell and / or dormant endospore of said Bacillus velezensis strain BX77 or a homolog thereof.

6. The method of any one of claims 1-3, wherein said pesticidal composition comprises a cell broth collected from a culture of said Bacillus velezensis strain BX77 or a homolog thereof.

7. The method of any one of claims 1-2, wherein said pesticidal composition comprises a cell free supernatant or filtrate obtained following culturing of said Bacillus velezensis strain BX77 of a homolog thereof.

8. The method of any one of claims 1-2, wherein said pesticidal composition comprises dead cells, an extract, a cell fraction or a lysate of said Bacillus velezensis strain BX77 or a homolog thereof.

9. The method of any one of claims 1-8, wherein said pesticidal composition is obtained following culturing of said Bacillus velezensis strain BX77 or a homolog thereof under conditions allowing biofilm formation and / or sporulation.

10. The method of claim 9, wherein said conditions comprise a medium selected from the group consisting of LB, NB and MRS.

11. The method of claim 9, wherein said conditions comprise a LB medium.

12. The method of any one of claims 9-11, wherein said conditions comprise culturing for at least 36 hours.

13. The method of any one of claims 9-12, wherein said conditions comprise culturing for up to 72 hours.

14. The method of any one of claims 9-13, wherein said conditions comprise culturing while shaking at 100-200 rpm.

15. The method of any one of claims 1-8, wherein said pesticidal composition is obtained following culturing of said Bacillus velezensis strain BX77 for 36-56 hours, at 28-32 °C, in a LB medium while shaking at 100-200 rpm.

16. The method of any one of claims 9-15, comprising culturing said Bacillus velezensis strain BX77 under said conditions prior to said contacting.

17. The method of any one of claims 1-16, wherein said pesticidal composition is formulated as a liquid, solid, semi-solid, gel or powder.

18. The method of any one of claims 1-16, wherein said pesticidal composition is formulated as a liquid formulation.

19. The method of any one of claims 1-16, wherein said pesticidal composition is formulated as a dry formulation.

20. The method of any one of claims 1-19, wherein said contacting is effected once to twice a week.

21. The method of any one of claims 1-20, wherein upon said contacting the plant is at a post-flowering stage, a flowering stage, a pre-flowering stage, or any combination thereof.

22. The method of any one of claims 1-21, wherein said contacting is selected from the group consisting of spraying, immersing, drenching, coating, encapsulating, dusting, and fumigating.

23. The method of any one of claims 1-21, wherein said contacting is by spraying.

24. The method of any one of claims 1-23, wherein said contacting is onto or in the vicinity of a root, a stem, a trunk, a seed, a fruit, a flower, a leaf, or any combination thereof.

25. The method of any one of claims 1-21, wherein said contacting is by spraying a canopy of said plant and / or by drenching a root zone of said plant.

26. The method of any one of claims 1-6 and 9-25, wherein said pesticidal composition is provided in a concentration of at least 108CFU / ml.

27. The method of any one of claims 1-26, wherein said plant is a nightshade crop.

28. The method of any one of claims 1-26, wherein said plant is selected from the group consisting of tomato, cucumber, potato, eggplant, pepino, pepper, beet, citrul, physic nut, lucerne, spinach, bean, tobacco, maize and strawberries.

29. The method of any one of claims 1-26, wherein said plant is an ornamental plant.

30. The method of any one of claims 1-26, wherein said plant is a herbal plant.

31. The method of any one of claims 1-30, wherein said pest is an insect.

32. The method of claim 31, wherein said insect is a moth.

33. The method of claim 32, wherein said moth is Tula absoluta.

34. The method of claim 31, wherein said insect is selected from the group consisting of Bemisia tabaci and Aphis gossypii.

35. The method of any one of claims 1-30, wherein said pest is an arthropod.

36. The method of claim 35, wherein said arthropod is a mite or a spider mite.

37. The method of claim 36, wherein said spider mite is Tetranychus urticae.

38. The method of any one of claims 1-27, wherein said pest is a fungus.

39. The method of claim 38, wherein said fungus is selected from the group consisting of Botrytis cinerea, Sclerotinia sclerotiorum and Oidium neolycopersici.

40. The method of any one of claims 1-27, wherein said pest is a virus.

41. The method of claim 40, wherein said virus is a Tomato yellow leaf curl virus.

42. The method of any one of claims 1-41, wherein said plant is planted in an area known to be infested with said pest.

43. The method of claim 42, wherein said area is a tropical region.

44. The method of claim 42, wherein said area is a sub-tropical region.

45. The method of claim 42, wherein said area is temperate region.

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