I asked a more competent colleague and here is his quick response: "The study concerns the safety of Bacillus thuringiensis strains as used in organic agriculture - there is no direct link to genetically modified Bt-crops. Because the proteins are different (Protoxins) the data are not directly transferable to Bt-crops. The authors do not seem to be very convinced that their data are conclusive as they use the very common backdoor opener "suggesting further studies are required to clarify ..." . Hope this helps...

yes they say that a couple of times. Wouldn't hurt to do the research. I don't think its ever been done?

B. thuringiensis preparations containing CRY δ-endotoxins are considered as the most effective, specific and environmentally-friendly bioinsecticides; they have been used as biological pesticides in agriculture, forestry and in human health for the elimination of vectors of diseases for more than 60 years and their implementation far exceeds other microbial agents such as fungi, protozoa or viruses (Sanchis 2010). Both the long history of safe use of B.thuringiensis and the acute oral toxicity data allow for a conclusion that these and other δ-endotoxins pose negligible toxicity risk to humans (Glare and O.Callaghan 2000, OECD 2007).

There is actually some scientific debate how exactly these δ-endotoxins mechanistically work: In their critical review, Vachon et al. (2012) concluded that the presently available information still supports the notion that Bt Cry proteins act by forming pores in insect guts, but most physiological events leading to their formation, following binding of the activated toxins to their receptors, remain relatively poorly understood. Nevertheless, each Cry protein is highly specific to species groups like lepidoptera and coleoptera and are targeted at pests of these groups (O’Callaghan et al., 2005).

A paper by the 'hematology' authors in Food and Chemical Toxicology, published in Elsevier, claiming that Bacillus thuringiensis showed toxic effects in the blood of Swiss albino mice has been withdrawn.

Citation: Mezzomo BP, Miranda-Vilela AL, de Souza Freire I, Barbosa LC, Portilho FA, Lacava ZG, Grisolia CK, 'WITHDRAWN: Effects of oral administration of Bacillus thuringiensis as spore-crystal strains Cry1Aa, Cry1Ab, Cry1Ac or Cry2Aa on hematologic and genotoxic endpoints of Swiss albino mice', Food Chem Toxicol. 2012 Nov 9. pii: S0278-6915(12)00777-6. doi:10.1016/j.fct.2012.10.032

Now the same authors published the story we discuss here as

Citation: Mezzomo BP, Miranda-Vilela AL, Freire IdS, Barbosa LCP, Portilho FA, et al. (2013) Hematotoxicity of Bacillus thuringiensis as Spore-crystal Strains Cry1Aa, Cry1Ab, Cry1Ac or Cry2Aa in Swiss Albino Mice. J Hematol Thromb Dis 1:104. doi: 10.4172/jhtd.1000104

The new journal is a brand new open access journal (Volume 1 • Issue 1) with 21 days rapid review process plus quality and quick editorial, review and publication processing.

After a second careful reading, I have several critical remarks now to the authors:

The material and methods (MM) part is unclear in respect whether pure toxin crystals or a spore/crystal mixture was used (compare Mezzomo’s MM with those MM in citation 26 (Santos et al. 2009 of their literature list). They write ‘spore-crystals’ but I think they mean ‘spore/crystal mixtures’. There is also no information on the details genetic modification of the Bt strains used. It is well known that – contrary to CRY – the CYT protein is a mammalian toxic component of Bt crystals (Glare & OCallaghan 2000, chapter 7.3: Mammals p.63-70).

Citation: “These strains were genetically modified to express individually Cry1Aa, Cry1Ab, Cry1Ac or Cry2A, and bioassays using the purified proteins from these genetically modified (GM) Bt spore-crystals have been published [26].” There is no information whether the authors checked for presence of CYT proteins so that they can exclude that any effect is cause by other components of the spore/crystal mix. The authors indirectly confirmed that they have not used the purified toxin forms. There is also no information whether they have tested for the concentration of active CRY proteins in their assay.

Statistics: My own experience with the Man-Whitney U test is that one gets more false positive effects the more individual 2-2- tests are performed.

There is also no clear dose-response relationship, with no explanation why the different CRY toxin treatments show ‘shaping’ in various but no common direction.

Further, I don’t see any physiological explanation or a clear discussion on the biological relevance of the author’s findings, specifically in the light that CRY endotoxins so far showed no in vitro or in vivo toxicity to mammalian cultured cell lines (overview in Glare and O’Callaghan 2000).

In this case an experiment carried out with a lot of open questions is not sufficient to change the long history of safe use of Bacillus thuringiensis spore/crystal mixtures.

Scientific research is a form of organized scepticism, and therefore necessary to constantly challenge our current knowledge. In this case the challenge is not evidenced enough. More research does not hurt, and in this case is probably necessary in order to confirm the preliminary results.

More literature:

Glare, T.R., O’Callaghan, M. (2000): Bacillus thuringiensis: Biology, Ecology and Safety. John Wiley and Sons, Ltd., West Sussex, UK. 350 pages.

O'Callaghan, M., Glare, T.R., Burgess, E.P.J., Malone, L.A. (2005): Effects of plants genetically modified for insect resistance on nontarget organisms. Annual Review of Entomology 50: 271-292. DOI:10.1146/annurev.ento.50.071803.130352.

OECD (2007): Consensus document on safety information on transgenic plants expressing Bacillus thuringiensis - derived insect control proteins. Series on Harmonisation of Regulatory Oversight in Biotechnology

Sanchis, V. (2010): From microbial sprays to insect-resistant transgenic plants: history of the biospesticide Bacillus thuringiensis. A review. Agronomy for Sustainable Development. DOI:10.1051/agro/2010027.

Vachon, V., Laprade, R., Schwartz, J-L., (2012): Current models of the mode of action of Bacillus thuringiensis insecticidal crystal proteins: a critical review, Journal of Invertebrate Pathology (2012), doi: http://dx.doi.org/10.1016/j.jip.2012.05.001

I found it fascinating that the article was withdrawn from the initial journal and then published in an OMICS journal, which is basically a well-known pay-to-publish predatory publisher (on J. Beall's list). At the same time, it has been publicized recently that Food Chem Toxicol has appointed a Monsanto-tied "editor" that has been suspected of censoring articles unfavorable for GMO.

(link: http://independentsciencenews.org/science-media/the-goodman-affair-monsanto-targets-the-heart-of-science/ )

I wish an indication from the authors were available with regard to why their paper was withdrawn from a legitimate journal. It creates an aura of a scandal around it.

BTW I am not involved in agricultural research at all, just got interested in the story because of these publishing intricacies.

It becomes not clear in material and method part what was used in this study. We only know that it is a lyophilised powder of viable spores and crystal toxins from recombinant Bt strains. It is not clear how the lyophilised spore-crystal powder was prepared. Can this powder contain chemical residues from sample preparation? In this case a pure water control wasn’t sufficient for this study.

Spore-crystal preparations with cry1 and cry2 proteins have been tested in oral applications several times in much higher dosages without toxic effects. This fact is mentioned but not discussed. However, the authors refer to a study with Bt israelensis (page 7, citation no. 33), showing that alkaline solubilised Bt toxins have haemolytic effects. It is demonstrated already that this effect was due to a Bt israelensis specific toxin called cyt which is not related to cry toxins and for sure the used Bt strains don’t produce the cyt toxin. Furthermore, cyt is not toxic when applied orally. Therefore, this has nothing to do with the observations in this paper.

The presented study is not appropriate to indicate toxic effects of the cry toxins. Effects could be due to the recombinant Bt spores or impurities in the used Bt spore-toxin samples. The obvious conflicts with previous toxicological studies with Bt strains producing the same toxins are not discussed in a sufficient manner.