Get Complete Information on Bacillus Thuringiensis

Microbial preparations that kill the pests and pathogens in their natural habitat are called microbial pesticides or biopesticides. In this unit preparations of Bacillus thuringiensis (BtJ and Trichoderma spp. have been discussed in detail.

Bacillus Thuringiensis

The cells of Bacillus are rod shaped. There are several species of Bacillus, many of them are dangerous pathogens such as Bacillus anthracis causing anthrax disease in animals. The others are: B. megaterium, B. polymyxa, B. thuringiensis, etc. Moreover, B. thuringiensis is commonly found in soil, insect-rich environments, mills and warehouses. Their active ingredients are known to be insecticidal crystalline proteins produced during spore formation.

It produces many toxins e.g. a-, (3-, and 5-endotoxins. Different toxins have different spectra of activity. The genes involved to produce Bt toxins are called Bt genes or cry genes. Many Bt genes (cry IA) have been isolated. Different strains and serotypes have been developed by different companies. In addition to production of endotoxins, many strains of Bt are potent insect pathogens also.

The crystalline protein (8-endotoxin) is believed to be a protoxin because it is only highly toxic when ingested in susceptible insects.

Strains used in crop protection are selected from those habitats, isolated from nature on the basis of their potency on selected insects, spectrum of host insects and the ease with which they can be grown in fermenters.

1. Insect Pests:

The insects of Lepidopteran, particularly the American bollworm (Helicoverpa armigera), pink bollworm (Pectinophera species), spotted bollworm (Erias insulana) diamond back moth (.Plutela xylostella) and other vegetable pests such as Colorado potato beetle (Leptinotarsa decemlineota) and forest insects seriously damage the plants.

They cause a serious agricultural problem leading to yield losses and reduced product quality. Insects can damage both in the field and during storage. Each year, insects destroy about 25% of food crops worldwide.

The larvae of Ostrinia nubilalis, the European corn borer, can destroy up to 20% of a maize crop. It is a major pest in southern and central Europe. Western corn rootworm beetles feeding on a maize cob is shown. Certain cultivars of Bt maize are resistant to this serious pest.

GM rootworm resistant crops are not approved for cultivation in the European Union but are now being grown in the U.S. The insects cause damage to several plants such as vegetables, fruit, maize, small grain cereals and forests, orchards.

2. Mode of Action:

B. thuringiensis produces para-sporal, proteinaceous crystal inclusion bodies during sporulation. Upon ingestion, these become insecticidal to larvae of the order Lepidoptera and to both larvae and adults of a few Coleoptera. Once the crystal proteins are solubilized in the insect, the gut proteases converts the original pro-toxin into a combination of up to four smaller toxins.

These hydrolyzed toxins bind to the insects mid gut cells at high affinity at specific receptor binding sites where they interfere with the potassium-ion dependent active amino acid symport mechanism. This disruption causes the formation of the large pores that increase the water permeability of the cell membrane.

A large uptake of water causes cell swelling and eventual rupture, disintegrating the mid gut lining. Different toxins bind to different receptors in different insect species and with varying intensities. This explains species specificities.

Many Bt genes (cry IA) have been isolated and used to transform crop plants. Such crops are known as Genetically Modified (GM) Crops or Transgenic Crop such as Cotton ECH-162, MECH- 184, MECH-12. This makes them resistant to insect pests.

3. Bulk Inoculum Production:

Bulk inoculum of B. thuringiensis is produced in controlled fermentor in deep tanks of sterilized liquid nutrient medium. The endotoxins and living spores are harvested as water dispersible liquid concentrates for subsequent formulation.

The potency of Bt formulation indicates the dose-dependent, lethal activity of the product as compared to that of an accepted standard Bt. The first protocols were based on spore counts. But the number of spores did not reflect the number of crystals present in bacteria. This method was unreliable and resulted in some major product failures in the field.

With the development of commercial Bt products, a titration method based on comparison of the LC₅₀s of a product, standard method was developed. This method led to rapid improvements not only in product reliability, but also in increased biological activity of the products. The new technique became an instrument for industrial fermentation and formulation development.

It shows scanning electron micrograph (SEM) of spores and insecticidal crystal proteins (ICPs) of B. thuringiensis on a culture medium. The white spots are the spores and black dots are the ICPs.

The Bt insecticides are sold as a combination of S-endotoxin crystals and living bacterial spores. The products are formulated as a suspension concentrate (SC), a granular bait (GB), a ready to use bait (RB), a suspo-emulsion (SE), a granule (GR), an oil miscible flowable concentrate (oil miscible suspension) (OS), a dispersible powder (DP) and a wettable powder (WP).

Trade Names of Commercial Products Many countries are producing the Bt products. Trade names of some of the products are: Dipel BL, Delfin WG Bactin, Bio-Tek, Bio Bit WP, WOCK Biological (Halt-Bt/’

5. Specification of Products:

The products must bear the following properties:

(i) All formulations should be standardized. The toxin content should be expressed in terms of international units active against a target pest per mg of product.

(ii) The products should not be exposed to direct sunlight, and kept in cool conditions. Light instability can cause problems if exposed to high light intensities. If stored under cool dark conditions, the products remain viable for two years or more.

(iii) The products should not be used in combination with broad spectrum biocides such as Chlorothalonil. However, it is compatible with a wide range of acaricides, insecticides, fungicides, stickers, spreaders and wetter’s. Do not use water with a pH above 8.0. In alkaline condition Bt crystals are denatured.

6. Environmental Impact and Non-target Toxicity:

Bt has a short persistence in the open environment due to its sensitivity to U.V. light. No adverse effects have been recorded in approved field and none are anticipated. Bt should not be used near water-courses. However, no adverse effect has been observed on birds, fishes and honeybees.

7. Application:

The Bt insecticides should be used at rates of 100-300 g active ingredient (a.i.) per hectare. It should be ensured that the crop is well covered with the spray suspension. It should be applied while larvae are small and repeated every five to seven days if infestations are high. Br-based sprays can be applied up to the day of harvest.