Bt

There’s a lot of interesting and sometimes conflicting things about Bt out there. Most people know that the Bt gene originally came from Bacillus thuringiensis bacteria, which are common in soil, but there’s a lot more to know! Here, I’ll discuss the differences and similarities between Bt from bacteria and Bt which is found in genetically engineered crops, as well as the way these similarities and differences are considered for purposes of toxicology testing.
A field is a complex place, even without consideration of the different types of Bt. Before we get too far into the “weeds”, let’s look at Life in a standard and in Bt maize field, a video produced for the European MOBITAG project. Discussion of Bt begins at 16:40.
[kad_youtube url=”https://youtu.be/oU3X3OLOlBw?t=16m40s” ]

Many forms of Bt

Bt as found in B. thuringiensis bacteria is not quite the same protein as that which is used in transgenic crops. There are 2 key differences:
1) The DNA sequence has been converted from the language (aka codon specificity or codon preference) recognized by bacterial DNA transcription/translation machinery to the language recognized by plant machinery. While the gene sequences are different, this does not change the actual sequence of the resulting protein.
2) The DNA sequence from the bacteria was shortened before being used to transform plants. The native protein crystallizes while the altered one does not. Both proteins must be activated by specific enzymes in the basic pH of an insect gut, and this active form of the protein then binds to specific receptors in the membrane of cells in the insect’s gut, creating holes in the gut, which kills the insect. Since the protein only works when the receptor is present, only certain types of insects are susceptible to the Bt toxin.
The part of the Bt protein that binds to receptors in the insect’s gut can be altered to become more or less specific. If the receptor is the key, and the Bt is the lock – we can make the lock fit a set of similar keys or change it to fit a different key. The DNA sequence of the original Bt has been changed by humans to create a series of Bt proteins that affect different insects. It’s not just people that have altered Bt. Good old mother nature has been modifying the Bt protein as well – there have been close to 200 different types of Bt identified in different strains of B. thuringiensis.

The different forms of the protein are given different names. While the common name of the group of proteins is Bt (and that name works well enough for most purposes), when we want to be specific, the proteins are called Cry (for crystalline protein) with a combination of letters and numbers, such as Cry1A. Another way to know what type of Bt is used is to look at the crop variety. For example, MON810 expresses Cry1A(b). Note that various forms of Bt are not just used in engineered crops – the proteins are also expressed in bacteria and used as whole bacteria or Bt extracts in sprays in organic and conventional farming, including many of the same Bt proteins that are used in biotech crops.

What about safety?

Because Bt is a pesticide (insecticide, to be exact), in the United States, the EPA (Environmental Protection Agency) is responsible for determining its safety before use is approved. The EPA has approved 12 different Bt proteins for use in corn and/or cotton since 1995, some of which are no longer in use. The links here are from the Biosafety Clearing-House, set up by the Cartagena Protocol on Biosafety.

• Cry1Ab in corn from B. thuringiensis subsp. kumamotoensis
• Cry1Ac in cotton from B. thuringiensis subsp. kumamotoensis (truncated Cry1Ac aka X gene not registered in US, from B. thuringiensis subsp Dakota)
• Cry1F in corn, cotton from B. thuringiensis var. aizawai
• Cry2Ab2 in cotton (Cry2Ab not registered in US) from B. thuringiensis subsp. kumamotoensis
• Cry34Ab in corn (Cry34Ab1 not registered in US) from B. thuringiensis strain PS149B1
• Cry35Ab in corn
• Cry3A in potato, corn from B. thuringiensis subsp. kumamotoensis
• Cry3Bb in corn
• Cry3Bb1 in corn from B. thuringiensis subsp. kumamotoensis
• Cry9C in corn from B. thuringiensis subsp. kumamotoensis
• CryAb2 in corn, cotton
• Cry1A.105 in corn from B. thuringiensis subsp. kumamotoensis, chimeric gene including Cry1Ab, Cry1F, and Cry1Ac proteins

EDIT: For specific information on Bt traits currently on the market in the US, check out the Handy Bt Trait Table from University of Wisconsin Extension.

The EPA has a very nuanced stance on testing of Bt for human health, which they discuss in the Biopesticides Registration Action Document for Bt.
Some testing has been done with protein either extracted from various strains of B. thuringiensis or extracted from E. coli engineered to express Bt (or E. coli engineered to express the plant language version of Bt). These early tests were useful, and can to some degree be extrapolated to the more recent forms of Bt that have been engineered to be more specific to different insects. The proteins are very similar to one another, break down in similar ways, etc. However, we can’t assume that they’re exactly the same – they are at least slightly different protein sequences after all! Each new version of Bt has undergone new testing for allergenicity and toxicity both by the company wanting to sell crops expressing the new protein and by independent agencies all around the world.

The basic premise relied on for the toxicology assessment is the fact that all the Bt plant-incorporated protectants are proteins. Proteins are commonly found in the diet and, except for a few well described phenomena, present little risk as a mammalian hazard. In addition, for the majority of Bt proteins currently registered, the source bacterium has been a registered microbial pesticide which has been approved for use on food crops without specific restrictions. Because of their use as microbial pesticides, a long history of safe use is associated with many Bt products.

Does that mean that the EPA considers Bt to be GRAS (generally recognized as safe)? Definitely not. The EPA collects 3 types of data for each new type of Bt “to provide a reasonable certainty that no harm will result from the aggregate exposure to these proteins”. The tests are intended to show that the Bt protein:

• Behaves as would be expected of a dietary protein, as determined with an in vitro digestion assay.
• Is not structurally related to any known food allergen or protein toxin, as determined with amino acid sequence homology comparisons, and
• The Bt protein does not display any oral toxicity when administered at maximum hazard dose using purified protein of the plant incorporated protectant as a test substance. Due to limitations of obtaining sufficient quantities of pure protein test substance from the plant itself, an alternative production source of the protein is often used such as the Bacillus thuringiensis source organism or an industrial fermentation microbe.

Note that they aren’t testing oral toxicity of any old Bt protein, they are looking at the specific Bt protein in question, albeit expressed in a bacteria or yeast instead of in the crop itself – because the crop doesn’t produce enough of the protein to result in a toxic response.

EPA believes that protein instability in digestive fluids and the lack of adverse effects using the maximum hazard dose approach in general eliminate the need for longer-term testing of Bt protein plant-incorporated protectants. Dosing of these animals with the maximum hazard dose, along with the product characterization data should identify potential toxins and allergens, and provide an effective means to determine the safety of these protein.

Despite the fact that the EPA and various scientific bodies find it unnecessary to preform additional toxicity testing of Bt proteins, scientists do it anyway. Numerous tests have been conducted feeding Bt crops (mostly corn, but also cotton and potato) to livestock, other animals such as quail, and yes, even monarch butterfly. Few have found significant differences between Bt and non-Bt feed, and those that have found differences often have methodological or statistical errors that have been well covered elsewhere.
Ironically, crops expressing Bt have reduced toxin exposure all over the world. Corn expressing Bt has dramatically reduced incidence of fungal infection and of potentially deadly mycotoxins (toxins produced by fungus). Why? The Bt corn has fewer insect bite marks, which is how the fungus enters the kernels to colonize the ear. There are other ways to prevent fungus growth, but none are better than Bt, especially in tropical and sub-tropical areas such as southern Africa and Central America where maize aka corn is a staple. In addition, use of Bt has allowed farmers to use fewer broad spectrum insecticides, letting more non-pest insects live and reducing exposure to farmers and neighbors during pesticide application. Finally, because Bt protects corn from insect damage, Bt corn has higher yields so less land is needed to grow the same amount of food.

I love corn, and knowing that Bt is a safe protein for animal (and human!) consumption, and knowing the risks of mycotoxins (as well as how gross earworm can be!) I would be happy to eat Bt corn, whether in field corn ground for tortillas, masa, or other foods, or in delicious sweet corn*.

Further reading

• There’s no Bt in your blood – Biofortified Blog
• Plant Incorporated Protectants – EPA
• EPA’s Regulation of Bacillus thuringiensis (Bt) Crops – EPA
• Bacillus thuringiensis: Profile of a bacterium – GMO Safety
• The cry proteins – Protein Spotlight
• Bt search results – Biosafety Clearing-House
• Bacillus Thuringiensis – Extension Toxicology Network
• Feeding Transgenic Crops to Livestock – Federation of Animal Science Societies
• Mycotoxins in Crops: A Threat to Human and Domestic Animal Health – APS Net
• Genetically Engineered Crops—What, How and Why – Scientific American

* To date, while Bt sweet corn varieties do exist, they aren’t widely used.