What is Bacillus thuringiensis (Bt)?
Bacillus thuringiensis (Bt) is a Gram-positive, rod-shaped, and spore-forming bacterium that produces a parasporal body, known as crystal, that consists of one or more intracellular proteinaceous crystallin inclusions with pesticidal properties.
Bt is a facultative pathogenic bacterium but it is also able to live in the environment, outside its host. However, it is thought that insects are the optimal site to multiplicate and exchange genetic material.
The Bt life cycle has two distinct growth phases (Figure 1).The first phase, named the vegetative growth phase, occurs when the nutritional and aeration conditions are beneficial. The spores germinate and cells grow and reproduce by binary fission as long as nutrients are available. When encountering unfavorable conditions, Bt enters the stationary phase, also known as the sporulation growth phase, where the cells form a resistant spore and one or more parasporal crystal inclusions of proteinaceous nature.
Figure 1. Schematic representation of the life cycle of Bacillus thuringiensis. During the vegetative phase, the bacterium grows exponentially and each cell is divided into two new cells. When faced with a lack of nutrients, it enters the stationary phase in which it sporulates and the proteins aggregate to form the parasporal crystals. Finally, cells undergo autolysis and release the crystal and the spore, which will germinate again when the conditions turn favorable.
What do Bt crystal proteins do?
Bt may become infective once the spores and crystals are ingested by the host and reach the midgut, where the crystal bodies are solubilized in its alkaline pH into protoxins and, subsequently, proteolytically activated by gut enzymes into toxins. Once this occurs, the toxins bind to specific receptors in the apical face of the epithelial cells from the mesenteron and form a pore in the cell membrane. This produces an osmotic imbalance in the cells and, eventually, their lysis. Host death results from the disruption of the epithelial cells, which ultimately, leads to its paralysis and inefficient feeding (Figure 2).
Figure 2. Schematic overview of the mode of action of B. thuringiensis crystal proteins. Once the spores and crystals reach the host midgut (1), crystal proteins are solubilized (2) and processed by midgut proteases (3). Activated toxins are able to cross the peritrophic matrix (4) and bind to specific membrane receptors (5) located in the midgut epithelial cells. This binding allows toxins to form pores (6), which leads to osmotic cell lysis. Massive cell death disrupts the integrity of the midgut epithelial layer (7), which is enough to produce host death.
What proteins does Bt produce and what are they for?
The most studied Bt crystal proteins are members of the Cry (crystal) protein class, which have a 3-domain structure (such as Cry1, Cry2, Cry3, Cry4, Cry5, Cry9, etc.). Other crystal proteins are the Cyt (cytolytic) toxins, whose active form contains a single domain (Cyt2Aa).
Bt crystal toxins have target specificity, meaning that a given protein will have a controlling effect on a specific pest but may not work on a different one.
The literature often describes many proteins as being active against a specific insect order such as lepidopteran, dipteran, and coleopteran, or against different nematode species. In principle, the range of proteins that function effectively against a given number of targets is rather narrow.
This is evidenced by the Bt strains developed as commercial products, which usually harbor similar gene contents for the control of species belonging to the same insect order.
Why are Bioinsectis Bt strains different?
The current Bt solutions in the crop protection market are mainly aimed at controlling lepidopteran pests. Most of the available commercial products rely either on strains of the serovar kurstaki, which mainly express combinations of Cry1A and Cry2A protein variants, and serovar aizawai that, in addition, include Cry1C and/or Cry1D. The result, is a highly redundant offer of active ingredients that translates into a lower differentiation between products.
In order to provide the market with new and differentiated Bt strains, the BIOINSECTIS R&D department has dedicated itself to building a new collection of thousands Bt isolates of which, up to date, over 600 unique strains have been sequenced. Today, the team continues to characterize those with new combinations of insecticidal genes at the biological level, providing companies of the sector with unprecedented and competitive active ingredients that outperform the currently available Bt-based products (Table 1).
Table1. Insecticidal gene content of Bioinsectis Bt strains (BST-Bt) in comparison to serovars kurstaki and aizawai commercial strains