Friday, 16 January 2015

Bt maize and mouse immune responses - what's wrong with this picture?

I have to admit, I haven't been paying close attention to anti-GMO material lately. I've been very busy writing my PhD thesis* and so have largely left GM related myth detection and busting in other very capable hands, such as the guys at the Genetic Literacy Project and the Plant Science Panel at Sense about Science. I also don't get a whole lot of exposure to unscientific anti-GM arguments during my average work day, what with working in a plant science lab.

However, by night (and morning, and weekend), I'm a fancy rat owner and breeder, which exposes me to all sorts of different people and views, especially where rat nutrition is concerned. I've become quite used to discussing (dare I say debunking) concerns relating to the safety of Roundup Ready (RR) maize in rat feed that followed the widely refuted, retracted and then re-plublished Seralini study on long term exposure to glyphosate and RR maize. Less expected, however, were concerns regarding the below poster shared from the GMO Free USA facebook account, referencing a recently published study on mouse immune responses to the Bacillus thuringiensis (Bt) protein 'Cry1Ab' - a naturally occurring insecticidal protein which has been modified and inserted into maize and other crop species. This makes the crops more resistant to some insect pests, thus making growers less reliant on pesticide sprays.

Dare I say it? I smelled a rat. So I decided to review the evidence behind the poster. (I will confess, for the sake of my sanity I stuck to the referenced publication on inhalation of Bt proteins. I believe studies on gut responses have been reviewed elsewhere but if I fail to find the links I may do it myself.)

It has to be said, the creators of the poster have done a good job of evoking fear without saying anything too outlandish - note the use of the word 'may'. However, the implication was that the referenced study proved that inhalation of GM Bt maize induced an allergic reaction in mice. The creators also cherry-picked the data which could potentially support their cause, without acknowledging the data that didn't...

What the paper says

The researchers exposed the mice to a number of treatments by anaesthetising them and then dripping liquid solutions directly into their nostrils. The treatments were purified Cry1Ab from B. thuringiensis, the transgenic Cry1Ab found in maize, pollen and leaf preparations of GM Bt maize, and pollen and leaf preparations from non-Bt maize as controls. This was accompanied by a group which were just treated with liquid buffer. From what I can tell, no other purified protein was used as a control. They then measured the immune response of the mice (looking at variables such as IgE antbibody production, which is associated with allergic responses, and proliferation of different sorts of white blood cells and proteins important for immune responses).

The main piece of 'positive' data presented in this study, which the poster takes inspiration from, is the finding that the pure Bt protein treatments resulted in Cryp1Ab-specific antibody production. However, these antibodies were not detected from the mice treated with the Bt maize extracts, or the non-GM maize extracts. None of the other immune response variables (with the exception of one type of cytokine in one experiment) differed significantly between mice treated with the Bt protein containing extracts and those treated with extracts from non-Bt plants. However, this is somewhat obfuscated in the paper:

The authors repeatedly, and rather confusingly, refer to the mice treated with liquid buffer as the controls, rather than those treated with the non-GM maize extracts. This allows the authors to highlight the significant differences between the GM plant extract treatments and their 'control group'. However, as they quite rightly point out, there is no significant difference between the GM and non-GM plant treatments, indicating that the observed responses are not due to the Bt protein. Indeed, one type of white blood cell (eosinophils) were elevated in response to plant extracts, but not in response to the purified Bt protein, which rather suggests that this response was caused by another plant component. Indeed, the authors conceded in their discussion that many of the observed responses were likely a generalised response to plant extracts rather than to Bt protein.

So to summarise - purified Bt protein (Cry1Ab) treatment resulted in antibody production, but neither this nor the other investigated variables differed between mice treated with Bt maize extracts compared to non-Bt maize extracts.

Limitations of the study

Aside from the aforementioned lack of a non-allergenic protein control, this study has a few limitations that I've picked up on, many of which the authors themselves highlight in their discussion:

The premise for this research was to analyse the risks of allergic reactions to inhaled Bt protein, either from insecticide sprays (more on that later) on inhaling GM Bt pollen. However, it's not clear how well their experimental design of dripping liquid into the nostrils of unconscious mice placed on their backs reflects natural nasal exposure of humans to the investigated substances. Indeed, the authors state that "the model is optimised to induce allergy", indicating that it may not reflect normal exposure in humans. This contradicts the GMO Free USA statement accompanying the poster that "the protein may actually be MORE [sic] allergenic and immunotoxic than this study suggests"

Furthermore, again acknowledged by the study authors, "the clinical relevance of the detected specific IgE was not evaluated" meaning that no assessment was made of whether the IgE production led to a symptomatic allergic reaction in mice. Further work is therefore required to demonstrate whether, as the poster puts it, "mice that inhale Bt toxin insecticidal proteins from GM corn may develop immune and allergic reactions".

Bt isn't just a 'GM' thing

One major caveat of this study's use by anti-GMO campaigners is that the use of Bt proteins isn't unique to GM crop production. On the contrary, B. thuringiensis is a widely used 'biopesticide', and has been used in organic crop production for over 50 years. Opponents of GM have previously argued that the 'natural' Bt protein and the transgene product are not substantially equivalent, since the naturally occurring protein is activated in insect guts, while the transgenic protein is 'pre-activated', which could cause health issues in humans and animals. However, this particular study does not support that argument, since both forms of the protein resulted in antibody production when applied in purified form. The fact that this was not the case for Bt protein containing maize extracts again rather turns this argument on its head. The authors state in their conclusions that:

"Given the importance of Bt-transgenic maize as food and feed across the world, a considerable number of individuals may be exposed to Cry1Ab by inhalation, in the field as well as along the food/feed chain."

However, since antibody production was induced by purified Bt protein but not by plant extracts, the relevance of transgenic Bt-crops as opposed to Bt treatment of conventional and organic crops in terms of human exposure is arguably overstated.


The referenced study does provide evidence that purified Bt protein Cry1Ab can induce an immunogenic response in mice, when applied in liquid form directly to their nasal passages. However, this does not translate into evidence that Bt maize causes allergic reactions (in mice or humans), especially since no clear immunological response to Bt-maize extracts was observed compared to non-GM plants.

I for one will be leaving my face mask in the growth room, where it protects me from mycotoxin producing Fusarium species. Incidentally, there is evidence that cultivation of Bt-maize might help to lower contamination of maize cobs with fungal mycotoxins, since the fungi normally infect the plants through holes made by certain insects (which are killed by Bt protein). I personally would rather risk consuming Bt protein than deoxinivalenol (a.k.a vomitoxin), but perhaps that's just me.

Don't get me wrong, I'm not saying Cry1Ab isn't a potential human allergen. It might well be, but the findings from this study are a long way from demonstrating it. In addition, this study suggests that all forms of Bt protein might be equally allergenic, rather than just those produced by GM crops.

Finally, and only because I'm being very pedantic, that's a baby rat wearing a mask, not a mouse. Here's a picture of a baby rat without a mask, which is much cuter.

*As a side note, this blog post has taken me over a day to research and write. This included reading the paper a couple of times, brushing up on my knowledge of antigens, allergens and the human immune system (quite different to plants) and reading other relevant documents, and being accused of working for Monsanto when I tried to engage with other commenters on the original post. All this when I arguably should have been writing up papers from my PhD thesis in order to increase the likelihood of securing funding to carry on doing research into how to protect plants from diseases. This was without addressing other questions such as the credibility of the journal, the interests and funding source of the authors, and the validity of the statistical tests used. Small wonder, then, that many anti-GMO statements like this one go unrefuted by scientists. It's not that they're irrefutable, rather that doing all the research required to present a solid, evidence-based critique is likely far more laborious than generating the statements in the first place.

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