Editing Bacillus thuringiensis (section)

==Use of Bt genes in genetic engineering of plants for pest control==
The Belgian company [[Plant Genetic Systems]] (now part of [[Bayer CropScience]]) was the first company (in 1985) to develop [[genetically modified crops]] ([[tobacco]]) with insect tolerance by expressing ''cry'' genes from ''B. thuringiensis''; the resulting crops contain [[delta endotoxin]].<ref name=Hofte_1986>{{cite journal | vauthors = Höfte H, de Greve H, Seurinck J, Jansens S, Mahillon J, Ampe C, Vandekerckhove J, Vanderbruggen H, van Montagu M, Zabeau M | title = Structural and functional analysis of a cloned delta endotoxin of Bacillus thuringiensis berliner 1715 | journal = European Journal of Biochemistry | volume = 161 | issue = 2 | pages = 273–80 | date = December 1986 | pmid = 3023091 | doi = 10.1111/j.1432-1033.1986.tb10443.x | display-authors = 6 | doi-access = free }}</ref><ref name=Vaeck>{{cite journal |doi=10.1038/328033a0 |title=Transgenic plants protected from insect attack |year=1987 | vauthors = Vaeck M, Reynaerts A, Höfte H, Jansens S, de Beuckeleer M, Dean C, Zabeau M, Van Montagu M, Leemans J | display-authors = 6 |journal=Nature |volume=328 |issue=6125 |pages=33–7 |bibcode=1987Natur.328...33V|s2cid=4310501 }}</ref> The Bt tobacco was never commercialized; tobacco plants are used to test genetic modifications since they are easy to manipulate genetically and are not part of the food supply.<ref>{{cite web | author = Staff | work = GMO Compass | date = 29 July 2010 | url = http://www.gmo-compass.org/eng/database/plants/304.tobacco.html | title = "Tobacco" entry in GMO Compass database | archive-url = https://web.archive.org/web/20131002090217/http://www.gmo-compass.org/eng/database/plants/304.tobacco.html | archive-date = 2 October 2013 }}</ref><ref>{{cite journal | vauthors = Key S, Ma JK, Drake PM | title = Genetically modified plants and human health | journal = Journal of the Royal Society of Medicine | volume = 101 | issue = 6 | pages = 290–8 | date = June 2008 | pmid = 18515776 | pmc = 2408621 | doi = 10.1258/jrsm.2008.070372 }}</ref>
[[Image:Bt plants.png|thumb|Bt toxins present in [[peanut]] leaves (bottom dish) protect it from extensive damage caused to unprotected peanut leaves by [[lesser cornstalk borer]] [[larva]]e (top dish).<ref>{{cite web | vauthors = Suszkiwn J |url= http://ars.usda.gov/is/ar/archive/nov99/pest1199.htm |title=Tifton, Georgia: A Peanut Pest Showdown |access-date=2008-11-23 |work= Agricultural Research magazine|date=November 1999| archive-url= https://web.archive.org/web/20081012182444/http://www.ars.usda.gov/is/AR/archive/nov99/pest1199.htm| archive-date= 12 October 2008 | url-status= live}}</ref>]]

===Usage===
In 1995, {{visible anchor|Bt potato|text=potato plants producing CRY 3A Bt toxin}} were approved safe by the [[Environmental Protection Agency]], making it the first human-modified pesticide-producing crop to be approved in the US,<ref>{{cite news|url = https://news.google.com/newspapers?id=A0YyAAAAIBAJ&pg=4631,1776980&dq=bacillus+thuringiensis+potato+1996+approved&hl= |title = Genetically Altered Potato Ok'd For Crops |work = Lawrence Journal-World |date = 6 May 1995|agency = AP|via = Google News}}</ref><ref>{{cite web |url=http://www.cera-gmc.org/files/cera/GmCropDatabase/docs/decdocs/02-269-004.pdf |title=Safety Assessment of NewLeaf ®Y Potatoes Protected Against Colorado Potato Beetle and Infection by Potato Virus Y Causing Rugose Mosaic |website=www.cera-gmc.org |access-date=31 August 2022 |archive-url=https://web.archive.org/web/20150927213821/http://www.cera-gmc.org/files/cera/GmCropDatabase/docs/decdocs/02-269-004.pdf |archive-date=27 September 2015 |url-status=dead}}</ref> though many plants produce pesticides naturally, including tobacco, [[coffee plant]]s, [[Theobroma cacao|cocoa]], [[cotton]] and [[black walnut]]. This was the 'New Leaf' potato, and it was removed from the market in 2001 due to lack of interest.<ref name=PotatoPro>{{cite news|title=The History and Future of GM Potatoes| vauthors = van Eijck P |url=http://www.potatopro.com/newsletters/20100310.htm|newspaper=PotatoPro Newsletter|date=March 10, 2010|access-date=October 5, 2013|archive-url=https://web.archive.org/web/20131012033805/http://www.potatopro.com/newsletters/20100310.htm|archive-date=October 12, 2013|url-status=dead}}</ref>

In 1996, {{visible anchor|Bt maize|Bt corn|text=[[genetically modified maize]] producing Bt Cry protein}} was approved, which killed the European corn borer and related species; subsequent Bt genes were introduced that killed corn rootworm larvae.<ref name=NatureEd>{{cite journal | vauthors = Hellmich RL, Hellmich KA | year = 2012 | title = Use and Impact of Bt Maize | url = http://www.nature.com/scitable/knowledge/library/use-and-impact-of-bt-maize-46975413 | journal = Nature Education Knowledge | volume = 3 | issue = 10| page = 4 }}</ref>

The Bt genes engineered into crops and approved for release include, singly and stacked: Cry1A.105, CryIAb, CryIF, Cry2Ab, [[Cry3Bb1]], Cry34Ab1, Cry35Ab1, mCry3A, and VIP, and the engineered crops include corn and cotton.<ref>{{cite web | vauthors = Bessin R | work = University of Kentucky College of Agriculture | orig-date = May 1996 | date = November 2010 | url = http://www2.ca.uky.edu/entomology/entfacts/ef118.asp | title = Bt-Corn for Corn Borer Control }}</ref><ref>{{cite book | vauthors = Castagnola AS, Jurat-Fuentes JL | chapter = Bt Crops: Past and Future. Chapter 15 | title = ''Bacillus thuringiensis'' Biotechnology | veditors = Sansinenea E | publisher = Springer | date = March 2012 | isbn = 978-94-007-3020-5 }}</ref>{{rp|285ff}}

Corn genetically modified to produce VIP was first approved in the US in 2010.<ref>{{cite web | vauthors = Hodgson E, Gassmann A | work = Iowa State Extension, Department of Entomology. | date = May 2010 | url = http://www.extension.iastate.edu/CropNews/2010/0510hodgsongassman.htm | title = New Corn Trait Deregulated in U.S. }}</ref>

In India, by 2014, more than seven million cotton farmers, occupying twenty-six million acres, had adopted {{visible anchor|Bt cotton}}.<ref>{{cite magazine | url = http://www.newyorker.com/magazine/2014/08/25/seeds-of-doubt | title = Seeds of Doubt: An activist's controversial crusade against genetically modified crops. | vauthors = Specter M | magazine = [[The New Yorker]] | date = 25 August 2014 }}</ref>

[[Monsanto]] developed a {{visible anchor|Bt soybean|text=soybean expressing Cry1Ac}} and the [[glyphosate]]-resistance gene for the Brazilian market, which completed the Brazilian regulatory process in 2010.<ref>{{cite web | author = Staff | work = Monsanto | date = August 2009 | url =  http://www.gmo-compass.org/pdf/regulation/soybean/MON87701xMON89788_soybean_application_food_feed.pdf | title = Application for authorization to place on the market MON 87701 × MON 89788 soybean in the European Union, according to Regulation (EC) No 1829/2003 on genetically modified food and feed | archive-url = https://web.archive.org/web/20120905233938/http://www.gmo-compass.org/pdf/regulation/soybean/MON87701xMON89788_soybean_application_food_feed.pdf | archive-date=2012-09-05 }} Linked from the {{cite web | work = GMO Compass | url = http://www.gmo-compass.org/eng/gmo/db/147.docu.html | title = MON87701 x MON89788 | archive-url = https://web.archive.org/web/20131109152621/http://www.gmo-compass.org/eng/gmo/db/147.docu.html | archive-date=2013-11-09 }}</ref><ref>{{Cite web|url=http://www.isaaa.org/kc/cropbiotechupdate/article/default.asp?ID=6565|title=Monsanto's Bt Roundup Ready 2 Yield Soybeans Approved for Planting in Brazil| work = Crop Biotech Update | date = 27 August 2010 | publisher =  International Service for the Acquisition of Agri-biotech Applications (ISAAA) }}</ref>

{{visible anchor|Bt aspen|Bt-transformed [[aspen]]s}} - specifically ''[[Populus]]'' hybrids - have been developed. They do suffer lesser leaf damage from insect [[herbivory]]. The results have not been entirely positive however: The intended result - better [[timber]] yield - was not achieved, with no growth advantage despite that reduction in herbivore damage; one of their major pests still preys upon the transgenic trees; and besides that, their [[leaf litter]] decomposes differently due to the transgenic toxins, resulting in alterations to the [[aquatic insect]] populations nearby.<ref name="Stange-et-al-2021">{{cite journal | vauthors = Stange M, Barrett RD, Hendry AP | title = The importance of genomic variation for biodiversity, ecosystems and people | journal = Nature Reviews. Genetics | volume = 22 | issue = 2 | pages = 89–105 | date = February 2021 | pmid = 33067582 | doi = 10.1038/s41576-020-00288-7 | publisher = [[Nature Research]] | s2cid = 223559538 }}</ref>

[[Image:Btcornafrica.jpg|thumb|Agriculture enthusiasts examining insect-resistant [[transgenic]] Bt corn]]

===Safety studies===
The use of Bt [[toxin]]s as [[Insecticide#Plant-incorporated protectants|plant-incorporated protectants]] prompted the need for extensive evaluation of their safety for use in foods and potential unintended impacts on the environment.<ref>{{Cite web|url=https://gmoscience.org/2015/08/10/is-bt-toxin-safe/|title=Are all forms of Bt toxin safe?|website=Gmoscience.org|access-date=9 April 2022}}</ref>

====Dietary risk assessment====
Concerns over the safety of consumption of genetically modified plant materials that contain [[Delta endotoxin|Cry proteins]] have been addressed in extensive dietary risk assessment studies. As a toxic mechanism, ''cry'' proteins bind to specific receptors on the membranes of mid-gut ([[epithelial]]) cells of the targeted pests, resulting in their rupture. While the target pests are exposed to the toxins primarily through leaf and stalk material, Cry proteins are also expressed in other parts of the plant, including trace amounts in maize kernels which are ultimately consumed by both humans and animals.<ref>{{cite journal | vauthors = Fearing PL, Brown D, Vlachos D, Meghji M, Privalle L | title = Quantitative analysis of CryIA (b) expression in Bt maize plants, tissues, and silage and stability of expression over successive generations. | journal = Molecular Breeding | date = June 1997 | volume = 3 | issue = 3 | pages = 169–176 | doi = 10.1023/A:1009611613475 | s2cid = 34209572 }}</ref> However, other organisms (including humans, other animals and non-targeted insects) that lack the appropriate receptors in their gut cannot be affected by the ''cry'' protein, and therefore are not affected by Bt.<ref name="auto"/><ref name="auto1"/>

=====Toxicology studies=====
Animal models have been used to assess human health risk from consumption of products containing Cry proteins. The United States Environmental Protection Agency recognizes mouse acute oral feeding studies where doses as high as 5,000&nbsp;mg/kg body weight resulted in [[No-observed-adverse-effect level|no observed adverse effects]].<ref>{{cite web|publisher=US EPA|date=2001|url=http://www3.epa.gov/pesticides/chem_search/reg_actions/pip/bt_brad2/2-id_health.pdf |archive-url=https://web.archive.org/web/20151211051629/http://www3.epa.gov/pesticides/chem_search/reg_actions/pip/bt_brad2/2-id_health.pdf |archive-date=2015-12-11 |url-status=live|title=Bt Plant-Incorporated Protectants October 15, 2001 Biopesticides Registration Action Document|access-date=2022-04-09}}</ref> Research on other known toxic proteins suggests that {{clarify|text=toxicity occurs at much lower doses|reason=This statement is likely the opposite of what was intended.|date=November 2020}}, further suggesting that Bt toxins are not toxic to mammals.<ref>{{cite journal | vauthors = Sjoblad RD, McClintock JT, Engler R | title = Toxicological considerations for protein components of biological pesticide products | journal = Regulatory Toxicology and Pharmacology | volume = 15 | issue = 1 | pages = 3–9 | date = February 1992 | pmid = 1553409 | doi = 10.1016/0273-2300(92)90078-n | url = https://zenodo.org/record/1258447 }}</ref> The results of toxicology studies are further strengthened by the lack of observed toxicity from decades of use of ''B. thuringiensis'' and its crystalline proteins as an insecticidal spray.<ref>{{cite journal | vauthors = Koch MS, Ward JM, Levine SL, Baum JA, Vicini JL, Hammond BG | title = The food and environmental safety of Bt crops | journal = Frontiers in Plant Science | volume = 6 | pages = 283 | date = April 2015 | pmid = 25972882 | pmc = 4413729 | doi = 10.3389/fpls.2015.00283 | doi-access = free }}</ref>

=====Allergenicity studies=====
Introduction of a new protein raised concerns regarding the potential for allergic responses in sensitive individuals. [[Bioinformatics|Bioinformatic]] analysis of known [[allergen]]s has indicated there is no concern of [[Allergy|allergic reactions]] as a result of consumption of Bt toxins.<ref>{{cite journal | vauthors = Randhawa GJ, Singh M, Grover M | title = Bioinformatic analysis for allergenicity assessment of Bacillus thuringiensis Cry proteins expressed in insect-resistant food crops | journal = Food and Chemical Toxicology | volume = 49 | issue = 2 | pages = 356–62 | date = February 2011 | pmid = 21078358 | doi = 10.1016/j.fct.2010.11.008 }}</ref> Additionally, [[Allergy#Skin prick testing|skin prick testing]] using purified Bt protein resulted in no detectable production of toxin-specific [[Immunoglobulin E|IgE]] antibodies, even in [[Atopy|atopic]] patients.<ref>{{cite journal | vauthors = Batista R, Nunes B, Carmo M, Cardoso C, José HS, de Almeida AB, Manique A, Bento L, Ricardo CP, Oliveira MM | title = Lack of detectable allergenicity of transgenic maize and soya samples | journal = The Journal of Allergy and Clinical Immunology | volume = 116 | issue = 2 | pages = 403–10 | date = August 2005 | pmid = 16083797 | doi = 10.1016/j.jaci.2005.04.014 | url = http://repositorio.insa.pt/bitstream/10400.18/114/1/Lack%20of%20detectable%20allergenicity.pdf | hdl = 10400.18/114 }}</ref>

=====Digestibility studies=====
Studies have been conducted to evaluate the fate of Bt toxins that are ingested in foods. Bt toxin proteins have been shown to digest within minutes of exposure to simulated [[Gastric acid|gastric fluids]].<ref>{{cite journal | vauthors = Betz FS, Hammond BG, Fuchs RL | title = Safety and advantages of Bacillus thuringiensis-protected plants to control insect pests | journal = Regulatory Toxicology and Pharmacology | volume = 32 | issue = 2 | pages = 156–73 | date = October 2000 | pmid = 11067772 | doi = 10.1006/rtph.2000.1426 }}</ref> The instability of the proteins in digestive fluids is an additional indication that Cry proteins are unlikely to be allergenic, since most known food allergens resist degradation and are ultimately [[Small intestine#Absorption|absorbed]] in the small intestine.<ref>{{cite journal | vauthors = Astwood JD, Leach JN, Fuchs RL | title = Stability of food allergens to digestion in vitro | journal = Nature Biotechnology | volume = 14 | issue = 10 | pages = 1269–73 | date = October 1996 | pmid = 9631091 | doi = 10.1038/nbt1096-1269 | s2cid = 22780150 }}</ref>

=====Persistence in environment=====
Concerns over possible environmental impact from accumulation of Bt toxins from plant tissues, pollen dispersal, and direct secretion from roots have been investigated. Bt toxins may persist in soil for over 200 days, with [[half-life|half-lives]] between 1.6 and 22 days. Much of the toxin is initially degraded rapidly by microorganisms in the environment, while some is [[adsorbed]] by organic matter and persists longer.<ref name="Helassa">{{cite book | vauthors = Helassa N, Quiquampoix H, Staunton S | veditors = Xu J, Sparks D | title=Molecular Environmental Soil Science|date=2013|publisher=Springer Netherlands|isbn=978-94-007-4177-5|pages=49–77|chapter=Structure, Biological Activity and Environmental Fate of Insecticidal Bt (Bacillus thuringiensis) Cry Proteins of Bacterial and Genetically Modified Plant Origin| doi = 10.1007/978-94-007-4177-5_3 }}</ref> Some studies, in contrast, claim that the toxins do not persist in the soil.<ref name="Helassa" /><ref>{{cite journal | vauthors = Dubelman S, Ayden BR, Bader BM, Brown CR, Jiang, Vlachos D | year = 2005 | title = Cry1Ab Protein Does Not Persist in Soil After 3 Years of Sustained Bt Corn Use | journal = Environ. Entomol. | volume = 34 | issue = 4| pages = 915–921 | doi=10.1603/0046-225x-34.4.915| doi-access = free }}</ref><ref>{{cite journal | vauthors = Head G, Surber JB, Watson JA, Martin JW, Duan JJ | year = 2002 | title = No Detection of Cry1Ac Protein in Soil After Multiple Years of Transgenic Bt Cotton (Bollgard) Use | journal = Environ. Entomol. | volume = 31 | issue = 1| pages = 30–36 | doi=10.1603/0046-225x-31.1.30| doi-access = free }}</ref> Bt toxins are less likely to accumulate in bodies of water, but pollen shed or [[soil runoff]] may deposit them in an aquatic ecosystem. Fish species are not susceptible to Bt toxins if exposed.<ref>{{cite journal | vauthors = Clark BW, Phillips TA, Coats JR | title = Environmental fate and effects of Bacillus thuringiensis (Bt) proteins from transgenic crops: a review | journal = Journal of Agricultural and Food Chemistry | volume = 53 | issue = 12 | pages = 4643–53 | date = June 2005 | pmid = 15941295 | doi = 10.1021/jf040442k | hdl = 10161/6458 | url = http://dukespace.lib.duke.edu/dspace/bitstream/handle/10161/6458/Clark%20et%20al.%202005%20Bt%20Review.pdf?sequence=1 }}</ref>

=====Impact on non-target organisms=====
The toxic nature of Bt proteins has an adverse impact on many major crop pests, but some ecological risk assessments has been conducted to ensure safety of beneficial non-target organisms that may come into contact with the toxins. Toxicity for the monarch butterfly, has been shown to not reach dangerous levels.<ref>{{cite journal | vauthors = Sears MK, Hellmich RL, Stanley-Horn DE, Oberhauser KS, Pleasants JM, Mattila HR, Siegfried BD, Dively GP | title = Impact of Bt corn pollen on monarch butterfly populations: a risk assessment | journal = Proceedings of the National Academy of Sciences of the United States of America | volume = 98 | issue = 21 | pages = 11937–42 | date = October 2001 | pmid = 11559842 | pmc = 59819 | doi = 10.1073/pnas.211329998 | bibcode = 2001PNAS...9811937S | doi-access = free }}</ref> Most soil-dwelling organisms, potentially exposed to Bt toxins through root exudates, are probably not impacted by the growth of Bt crops.<ref>{{cite journal | vauthors = Saxena D, Stotzky G | year = 2000 | title = ''Bacillus thuringiensis'' (Bt) toxin released from root exudates and biomass of Bt corn has no apparent effect on earthworms, nematodes, protozoa, bacteria, and fungi in soil | url = http://www.ask-force.org/web/Bt/Saxena-Stotzky-2001.pdf | journal = Soil Biology & Biochemistry | volume = 33 | issue = 9| pages = 1225–1230 | doi=10.1016/s0038-0717(01)00027-x}}</ref>

===Insect resistance===
Multiple insects have developed a resistance to ''B. thuringiensis''. In November 2009, [[Monsanto]] scientists found the [[pink bollworm]] had become [[Pesticide resistance|resistant]] to the first-generation [[Bt cotton]] in parts of [[Gujarat]], India - that generation expresses one Bt gene, ''Cry1Ac''. This was the first instance of Bt resistance confirmed by Monsanto anywhere in the world.<ref name=MonsantoWebCotton>{{cite web|url=http://www.monsanto.com/newsviews/Pages/india-pink-bollworm.aspx |title=Cotton in India |publisher=Monsanto.com |date=2008-11-03 |access-date=2013-07-09}}</ref><ref>{{cite journal | vauthors = Bagla P | title = India. Hardy cotton-munching pests are latest blow to GM crops | journal = Science | volume = 327 | issue = 5972 | pages = 1439 | date = March 2010 | pmid = 20299559 | doi = 10.1126/science.327.5972.1439 | bibcode = 2010Sci...327.1439B | doi-access = free }}</ref> Monsanto responded by introducing a second-generation cotton with multiple Bt proteins, which was rapidly adopted.<ref name=MonsantoWebCotton /> Bollworm resistance to first-generation Bt cotton was also identified in Australia, China, Spain, and the United States.<ref>{{cite journal | vauthors = Tabashnik BE, Gassmann AJ, Crowder DW, Carriére Y | title = Insect resistance to Bt crops: evidence versus theory | journal = Nature Biotechnology | volume = 26 | issue = 2 | pages = 199–202 | date = February 2008 | pmid = 18259177 | doi = 10.1038/nbt1382 | s2cid = 205273664 }}</ref> Additionally, resistance to Bt was documented in field population of [[diamondback moth]] in Hawaii, the continental US, and Asia.<ref>{{cite journal | vauthors = Tabshnik BE |title=Evolution of Resistance to Bacillus Thuringiensis |journal=Annual Review of Entomology |date=January 1994 |volume= 39 |pages=47–79 |doi= 10.1146/annurev.en.39.010194.000403}}</ref>  Studies in the [[cabbage looper]] have suggested that a mutation in the membrane transporter ABCC2 can confer resistance to Bt ''Cry1Ac''.<ref>{{cite journal | vauthors = Baxter SW, Badenes-Pérez FR, Morrison A, Vogel H, Crickmore N, Kain W, Wang P, Heckel DG, Jiggins CD | title = Parallel evolution of Bacillus thuringiensis toxin resistance in lepidoptera | journal = Genetics | volume = 189 | issue = 2 | pages = 675–9 | date = October 2011 | pmid = 21840855 | pmc = 3189815 | doi = 10.1534/genetics.111.130971 }}</ref>

===Secondary pests===
Several studies have documented surges in "sucking pests" (which are not affected by Bt toxins) within a few years of adoption of Bt cotton. In China, the main problem has been with [[mirids]],<ref>{{cite journal | vauthors = Lu Y, Wu K, Jiang Y, Xia B, Li P, Feng H, Wyckhuys KA, Guo Y | title = Mirid bug outbreaks in multiple crops correlated with wide-scale adoption of Bt cotton in China | journal = Science | volume = 328 | issue = 5982 | pages = 1151–4 | date = May 2010 | pmid = 20466880 | doi = 10.1126/science.1187881 | bibcode = 2010Sci...328.1151L | s2cid = 2093962 | doi-access = free }}</ref><ref>{{cite conference | vauthors = Just DR, Wang S, Pinstrup-Andersen P |year=2006 |title=Tarnishing Silver Bullets: Bt Technology Adoption, Bounded Rationality and the Outbreak of Secondary Pest Infestations in China |conference=American Agricultural Economics Association Annual Meeting |location=Long Beach, CA |url=http://purl.umn.edu/21230}}
* {{cite news | vauthors = Lang S |date=July 25, 2006 |title=Seven-year glitch: Cornell warns that Chinese GM cotton farmers are losing money due to 'secondary' pests |newspaper=Cornell Chronicle |url=http://www.news.cornell.edu/stories/July06/Bt.cotton.China.ssl.html |archive-url=https://web.archive.org/web/20060811215559/http://www.news.cornell.edu/stories/July06/Bt.cotton.China.ssl.html |archive-date=2006-08-11}}</ref> which have in some cases "completely eroded all benefits from Bt cotton cultivation".<ref>{{cite journal |doi=10.1504/IJBT.2008.018348 |title=Bt-cotton and secondary pests |year=2008 | vauthors = Wang S, Just DR, Pinstrup-Andersen P |journal=International Journal of Biotechnology |volume=10 |issue=2/3 |pages=113–21}}</ref> The increase in sucking pests depended on local temperature and rainfall conditions and increased in half the villages studied. The increase in insecticide use for the control of these secondary insects was far smaller than the reduction in total insecticide use due to Bt cotton adoption.<ref>{{cite journal | vauthors = Wang Z, Lin H, Huang J, Hu R, Rozelle S, Pray C |doi=10.1016/S1671-2927(09)60012-2 |title=Bt Cotton in China: Are Secondary Insect Infestations Offsetting the Benefits in Farmer Fields? |year=2009 |journal=Agricultural Sciences in China |volume=8 |pages=83–90}}</ref> Another study in five provinces in China found the reduction in pesticide use in Bt cotton cultivars is significantly lower than that reported in research elsewhere, consistent with the hypothesis suggested by recent studies that more pesticide sprayings are needed over time to control emerging secondary pests, such as aphids, spider mites, and lygus bugs.<ref>{{cite journal | vauthors = Zhao JH, Ho P, Azadi H | title = Benefits of Bt cotton counterbalanced by secondary pests? Perceptions of ecological change in China | journal = Environmental Monitoring and Assessment | volume = 173 | issue = 1–4 | pages = 985–994 | date = February 2011 | pmid = 20437270 | doi = 10.1007/s10661-010-1439-y | s2cid = 1583208 }}; Erratum published 2012 Aug 5: {{cite journal | vauthors = Zhao JH, Ho P, Azadi H |doi=10.1007/s10661-012-2699-5 |title=Erratum to: Benefits of Bt cotton counterbalanced by secondary pests? Perceptions of ecological change in China |year=2012 |journal=Environmental Monitoring and Assessment |volume=184 |issue=11 |page=7079 |doi-access=free }}</ref>

Similar problems have been reported in India, with both [[mealy bugs]]<ref>{{cite web | vauthors = Goswami B | work = InfoChange | url = http://infochangeindia.org/200709026463/Other/Features/Making-a-meal-of-Bt-cotton.html | archive-url = https://web.archive.org/web/20080616053151/http://infochangeindia.org/200709026463/Other/Features/Making-a-meal-of-Bt-cotton.html | archive-date = 16 June 2008 | title = Making a meal of Bt cotton | access-date = 6 April 2009 }}</ref><ref>{{cite news|url=http://www.gmwatch.org/en/news/archive/2007/7640-bug-makes-meal-of-punjab-cotton-whither-bt-magic-492007 |access-date=14 March 2018 |title=Bug makes meal of Punjab cotton, whither Bt magic? |date=4 September 2007 |newspaper=The Economic Times}}</ref> and aphids<ref>{{cite journal |doi=10.1016/j.worlddev.2010.09.008 |title=Field versus Farm in Warangal: Bt Cotton, Higher Yields, and Larger Questions |year=2011 | vauthors = Stone GD |journal=World Development |volume=39 |issue=3 |pages=387–98}}</ref> although a survey of small Indian farms between 2002 and 2008 concluded Bt cotton adoption has led to higher yields and lower pesticide use, decreasing over time.<ref>{{cite journal |doi=10.1016/j.agsy.2011.11.005 |title=Bt cotton and sustainability of pesticide reductions in India |year=2012 | vauthors = Krishna VV, Qaim M |journal=Agricultural Systems |volume=107 |pages=47–55}}</ref>

===Controversies===
The controversies surrounding Bt use are among the many [[genetically modified food controversies]] more widely.<ref>{{Cite web|url=https://www.pbs.org/wgbh/harvest/viewpoints/|title= Harvest of fear: viewpoints | work = Frontline/NOVA | publisher = Public Broadcasting Service | date = 2001 |access-date=9 April 2022}}</ref>

====Lepidopteran toxicity====
The most publicised problem associated with Bt crops is the claim that pollen from Bt maize could kill the [[monarch butterfly]].<ref>{{cite journal | vauthors = Losey JE, Rayor LS, Carter ME | title = Transgenic pollen harms monarch larvae | journal = Nature | volume = 399 | issue = 6733 | pages = 214 | date = May 1999 | pmid = 10353241 | doi = 10.1038/20338 | bibcode = 1999Natur.399..214L | s2cid = 4424836 | doi-access = free }}</ref> The paper produced a public uproar and demonstrations against Bt maize; however by 2001 several follow-up studies coordinated by the USDA had asserted that "the most common types of Bt maize pollen are not toxic to monarch larvae in concentrations the insects would encounter in the fields."<ref name=Waltz>{{cite journal | vauthors = Waltz E | journal = Nature News | date = 2 September 2009 | doi = 10.1038/461027a | title = GM crops: Battlefield | volume = 461 | issue = 7260 | pages = 27–32 | pmid = 19727179 | s2cid = 205048726 }}</ref><ref>{{cite journal | vauthors = Mendelsohn M, Kough J, Vaituzis Z, Matthews K | title = Are Bt crops safe? | journal = Nature Biotechnology | volume = 21 | issue = 9 | pages = 1003–9 | date = September 2003 | pmid = 12949561 | doi = 10.1038/nbt0903-1003 | s2cid = 16392889 | url = https://zenodo.org/record/1233343 }}</ref><ref>{{cite journal | vauthors = Hellmich RL, Siegfried BD, Sears MK, Stanley-Horn DE, Daniels MJ, Mattila HR, Spencer T, Bidne KG, Lewis LC | title = Monarch larvae sensitivity to Bacillus thuringiensis- purified proteins and pollen | journal = Proceedings of the National Academy of Sciences of the United States of America | volume = 98 | issue = 21 | pages = 11925–30 | date = October 2001 | pmid = 11559841 | pmc = 59744 | doi = 10.1073/pnas.211297698 | bibcode = 2001PNAS...9811925H | display-authors = 6 | jstor = 3056825 | doi-access = free }}</ref><ref>{{cite web |url=http://www.ars.usda.gov/is/br/btcorn/ |title=Bt Corn and Monarch Butterflies |date=2004-03-29 |work=USDA Agricultural Research Service |access-date=2008-11-23| archive-url= https://web.archive.org/web/20081106062846/http://www.ars.usda.gov/is/br/btcorn/| archive-date= 6 November 2008 | url-status= live}}</ref> Similarly, ''B. thuringiensis'' has been widely used for controlling ''[[Spodoptera littoralis]]'' larvae growth due to their detrimental pest activities in Africa and Southern Europe. However, ''[[Spodoptera littoralis|S. littoralis]]'' showed resistance to many strains of ''B. thuriginesis'' and were only effectively controlled by a few strains.<ref>{{cite journal|vauthors=Salama HS, Foda MS, Sharaby A|title=A proposed new biological standard for bioassay of bacterial insecticides vs. Spodoptera spp.|journal=Tropical Pest Management|date=1989|volume=35|issue=3|pages=326–330|url=https://www.cabi.org/isc/abstract/19901181560|doi=10.1080/09670878909371391|access-date=2017-11-12|archive-date=2018-09-29|archive-url=https://web.archive.org/web/20180929194738/https://www.cabi.org/isc/abstract/19901181560|url-status=dead}}</ref>

====Wild maize genetic mixing====
A study published in ''Nature'' in 2001 reported Bt-containing maize genes were found in maize in its center of origin, [[Oaxaca]], Mexico.<ref>{{cite journal | vauthors = Quist D, Chapela IH | title = Transgenic DNA introgressed into traditional maize landraces in Oaxaca, Mexico | journal = Nature | volume = 414 | issue = 6863 | pages = 541–3 | date = November 2001 | pmid = 11734853 | doi = 10.1038/35107068 | bibcode = 2001Natur.414..541Q | s2cid = 4403182 }}</ref> Another ''Nature'' paper published in 2002 claimed that the previous paper's conclusion was the result of an [[Artifact (error)|artifact]] caused by an [[inverse polymerase chain reaction]] and that "the evidence available is not sufficient to justify the publication of the original paper."<ref>{{cite journal | vauthors = Kaplinsky N, Braun D, Lisch D, Hay A, Hake S, Freeling M | title = Biodiversity (Communications arising): maize transgene results in Mexico are artefacts | journal = Nature | volume = 416 | issue = 6881 | pages = 601–2; discussion 600, 602 | date = April 2002 | pmid = 11935145 | doi = 10.1038/nature739 | bibcode = 2002Natur.416..601K | s2cid = 195690886 }}</ref> A significant controversy happened over the paper and ''Nature''{{'}}s unprecedented notice.<ref name="pbs.org">{{Cite web | url=https://www.pbs.org/now/science/genenature.html | archive-url = https://web.archive.org/web/20030220201414/https://www.pbs.org/now/science/genenature.html | archive-date = 20 February 2003 | title=Seeds of Conflict: NATURE Article Debate | work = NOW with Bill Moyers. Science & Health. | publisher = [[PBS]] }}</ref>

A subsequent large-scale study in 2005 failed to find any evidence of genetic mixing in Oaxaca.<ref>{{cite journal | vauthors = Ortiz-García S, Ezcurra E, Schoel B, Acevedo F, Soberón J, Snow AA | title = Absence of detectable transgenes in local landraces of maize in Oaxaca, Mexico (2003-2004) | journal = Proceedings of the National Academy of Sciences of the United States of America | volume = 102 | issue = 35 | pages = 12338–43 | date = August 2005 | pmid = 16093316 | pmc = 1184035 | doi = 10.1073/pnas.0503356102 | bibcode = 2005PNAS..10212338O | jstor = 3376579 | doi-access = free }}</ref> A 2007 study found the "transgenic proteins expressed in maize were found in two (0.96%) of 208 samples from farmers' fields, located in two (8%) of 25 sampled communities." Mexico imports a substantial amount of maize from the U.S., and due to formal and informal seed networks among rural farmers, many potential routes are available for transgenic maize to enter into food and feed webs.<ref>{{cite journal |doi=10.1890/1540-9295(2007)5[247:TPIMIT]2.0.CO;2 |year=2007 |volume=5 |pages=247–52 |title=Transgenic proteins in maize in the Soil Conservation area of Federal District, Mexico | vauthors = Serratos-Hernández J, Gómez-Olivares J, Salinas-Arreortua N, Buendía-Rodríguez E, Islas-Gutiérrez F, De-Ita A |journal=Frontiers in Ecology and the Environment |issue=5 |issn=1540-9295}}</ref> One study found small-scale (about 1%) introduction of transgenic sequences in sampled fields in Mexico; it did not find evidence for or against this introduced genetic material being inherited by the next generation of plants.<ref>{{cite journal | vauthors = Piñeyro-Nelson A, Van Heerwaarden J, Perales HR, Serratos-Hernández JA, Rangel A, Hufford MB, Gepts P, Garay-Arroyo A, Rivera-Bustamante R, Alvarez-Buylla ER | title = Transgenes in Mexican maize: molecular evidence and methodological considerations for GMO detection in landrace populations | journal = Molecular Ecology | volume = 18 | issue = 4 | pages = 750–61 | date = February 2009 | pmid = 19143938 | pmc = 3001031 | doi = 10.1111/j.1365-294X.2008.03993.x | display-authors = 6 }}</ref><ref>{{cite journal | vauthors = Dalton R | title = Modified genes spread to local maize | journal = Nature | volume = 456 | issue = 7219 | pages = 149 | date = November 2008 | pmid = 19005518 | doi = 10.1038/456149a | doi-access = free }}</ref> That study was immediately criticized, with the reviewer writing, "Genetically, any given plant should be either non-transgenic or transgenic, therefore for leaf tissue of a single transgenic plant, a GMO level close to 100% is expected. In their study, the authors chose to classify leaf samples as transgenic despite GMO levels of about 0.1%. We contend that results such as these are incorrectly interpreted as positive and are more likely to be indicative of contamination in the laboratory."<ref>{{cite journal | vauthors = Schoel B, Fagan J | title = Insufficient evidence for the discovery of transgenes in Mexican landraces | journal = Molecular Ecology | volume = 18 | issue = 20 | pages = 4143–4; discussion 4145–50 | date = October 2009 | pmid = 19793201 | doi = 10.1111/j.1365-294X.2009.04368.x | s2cid = 205362226 | doi-access = free }}</ref>

====Colony collapse disorder====
As of 2007, a new phenomenon called [[colony collapse disorder]] (CCD) began affecting [[Honey bee|bee]] hives all over North America. Initial speculation on possible causes included new parasites, pesticide use,<ref>{{cite web |url=http://www.ars.usda.gov/News/docs.htm?docid=15572 |title=ARS: Questions and Answers: Colony Collapse Disorder |work=ARS News | publisher = Agricultural Research Service, United States Department of Agriculture |date=2008-05-29 | access-date=2008-11-23| archive-url = https://web.archive.org/web/20081105121119/http://www.ars.usda.gov/News/docs.htm?docid=15572 | archive-date = 5 November 2008 | url-status= dead }}</ref> and the use of Bt transgenic crops.<ref>{{cite news | vauthors = Latsch G | title = Are GM Crops Killing Bees? |work=[[Spiegel Online]] |date=March 22, 2007 |url=http://www.spiegel.de/international/world/collapsing-colonies-are-gm-crops-killing-bees-a-473166.html}}</ref> The [[Mid-Atlantic Apiculture Research and Extension Consortium]] found no evidence that pollen from Bt crops is adversely affecting bees.<ref name=Waltz/><ref>{{cite journal |doi=10.1051/apido:2007022 |title=Effects of Bt corn pollen on honey bees: Emphasis on protocol development |year=2007 | vauthors = Rose R, Dively GP, Pettis J |journal=Apidologie |volume=38 |issue=4 |pages=368–77|s2cid=18256663 |url=https://hal.archives-ouvertes.fr/hal-00892271/document }}</ref> According to the USDA, "Genetically modified (GM) crops, most commonly Bt corn, have been offered up as the cause of CCD. But there is no correlation between where GM crops are planted and the pattern of CCD incidents. Also, GM crops have been widely planted since the late 1990s, but CCD did not appear until 2006. In addition, CCD has been reported in countries that do not allow GM crops to be planted, such as Switzerland. German researchers have noted in one study a possible correlation between exposure to Bt pollen and compromised immunity to ''[[Nosema (microsporidian)|Nosema]]''."<ref>{{cite web | publisher = United States Department of Agriculture | url = http://www.ars.usda.gov/is/AR/archive/jul12/colony0712.htm | title = Colony Collapse Disorder: An Incomplete Puzzle | work = Agricultural Research Magazine | date = July 2012 }}</ref> The actual cause of CCD was unknown in 2007, and scientists believe it may have multiple exacerbating causes.<ref>{{cite news |url=http://news.bbc.co.uk/1/hi/sci/tech/7925397.stm |title='No proof' of bee killer theory | vauthors = McGrath M |date=5 March 2009 |work=BBC News}}</ref>