ISIS Report 11/07/12
Bt Toxicity Confirmed: Flawed Studies Exposed
Researchers confirm Bt toxicity to non-target beneficial insects
and show how experiments claiming to refute their results were designed not to
find the effect Dr Eva Sirinathsinghji
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A new
study confirms that the Cry1Ab Bacillus
thuringiensis (Bt) toxin present in genetically modified (GM) crops kills
the larvae of the two-spotted ladybird (Adalia bipunctata L.), a
species that GM supporters claim to be unaffected by the toxin [1].
The
study raises questions regarding the integrity of previous work published by GM
proponents, whose experimental protocols were re-tested and shown to lack the
scientific rigour required to pick up signs of toxicity even in target insects
that the pesticide is designed to kill.
Bt
toxins are present in many GM crops including cotton and maize. Monsanto’s Mon 810
Bt maize is currently approved for cultivation in Europe, although it has been
banned by individual nations including Hungary, France, Austria, Germany,
Greece and Luxembourg due to health and environmental concerns. Many previous
studies have found effects on health and the
environment (see [2] Bt Crops Failures
and Hazards,
SiS 53, [3] More Illnesses
Linked to Bt Crops, SiS 30).
Previous
Bt toxicity studies slandered by GM proponents
GM
proponents claim that certain Bt toxins are effective against limited orders of
insects, with Cry1Ab killing only Lepidoptera (butterflies and moths) such as
the common maize pest the European cornborer. However, a peer-reviewed study
published by Angela Hilbeck and colleagues at the Swiss Federal Institute of
Technology in 2009 showed increased mortality in ladybirds exposed to the
‘activated’ toxin that had been coated on their food – meal moth eggs [4]; the team had found similar effects in green
lacewings previously [5-7]. The activated version refers to the cleavage of the
pro-toxin to produce the actual toxin.
In
response to their original publication, a coordinated effort aimed at
discrediting their findings appeared in the journal Transgenic Research,
which included two highly charged critiques [8, 9] and a study led by Jörg
Romeis from Agroscope, Switzerland, which failed to detect any toxicity [10].
They concluded that the results of Hilbeck’s team were ‘false-positives’ and
artefacts of a poor study design. One critique went as far as suggesting the
work was ‘pseudo-science’. Agroscope, a Swiss federal governmental research
organisation, is linked to the agrotech giant Syngenta, which along with
Monsanto, produces Cry1Ab GM crops. These hostile
attacks were triggered by the ban of Mon 810 maize in Germany based on results
obtained by Hilbeck’s team among 30 other scientific publications showing harmful
effects from the pesticide.
Addressing
discrepancies between previous studies
The
new work from Hilbeck’s team aimed to address the discrepancies between their
own findings and those of their critics. First they conducted a
‘proof-of-concept’ experiment where they tested both their original protocol and
Agroscope’s protocol on the target species, the European cornborer.
In the Hilbeck team’s original study, the ladybird larvae were
exposed continuously for 10 days to a microbially-produced purified version of
Cry1Ab or a microbially
produced ‘empty’ version lacking the toxin. They were exposed through coating
their food - meal moth eggs - with the toxin. The
Agroscope protocol on the other hand, exposed the larvae for only 24 hours at a
time through a sugar/water droplet with or without the toxin. As the larvae are
carnivorous and cannot survive on a sugar diet alone, they were transferred to
petri dishes with untreated moth eggs, thus giving them a period to recover
from the exposure. This exposure/recovery was apparently repeated 4 times in
total.
So, the aim of the new study was to understand if the differences
in these protocols may have accounted for the opposing results obtained by
Hilbeck’s team and by Agroscope. By testing target species that the toxin is
designed to kill, any weakness in the protocol would become apparent.
Hilbeck’s team repeated the basic protocols by exposing 4 day old
larvae to Bt maize as well as near isogenic non-GM maize sprayed with Bt toxins
either continuously for 7 days, or for 24 hours followed by untreated
non-transgenic maize for 6 days. They found high levels of mortality following
continuous exposure as expected (just below 100 % with both types of exposure).
The mortality rates dropped by half when animals were exposed to Bt sprayed
plants for 24 hours only. Exposure to Bt maize for only 24 hours did not even
cause mortality rates to rise above unexposed control groups.
An experimental protocol that cannot detect toxicity of a pesticide
on a target species is clearly not fit for testing potential harm to non-target species.
In addition to insufficient exposure time, other flaws in the
Agroscope experiments were noted by Hilbeck. Sugar/water droplets to which Bt
toxin was added were found to dry up overnight, leaving the levels of exposure
undetermined. Only one dose was tested, as opposed to three tested in the
original study by Hilbeck’s team [4]. There was no clear description of
the number of animals used or the number of times the experiments were
replicated, whereas performing 3 replicates is standard in laboratory studies.
Re-testing
effects of Bt toxin on ladybird larvae with a new combined protocol
To counter the criticisms aimed at their previous study, Hilbeck’s
team adopted a
combined protocol consisting of 7 days continuous
exposure to a sugar/water solution with or without the Bt toxin placed on
cotton balls to prevent them drying up. After 24 hours, instead of allowing a
recovery period, the cotton balls were replaced with fresh cotton balls
with or without Bt toxin solutions. Additional meal moth eggs coated in the
toxin were given to provide an adequate diet and
ensure continuous exposure to the toxin.
After only 6 days of exposure, mean mortality rate was 40 % compared to
around 25 % in unexposed larvae. The greatest difference in mortality between
treated and untreated animals peaked at 4 days where there was around a 20 % increase in mortality over untreated
animals, after which it began to level off.
The new work not only corroborates the team’s previous findings [4],
but also Agroscope’s failure to detect toxicity on non-target insects [10].
It is important to distinguish the difference between the natural
bacterial toxin and the modified version inserted into GM plants. Neither of
the original studies by Hilbeck or Agroscope used the versions expressed
in GM crops, which are significantly modified. Modifications are made to
increase the ‘performance’ of the toxin, including changing the promoter and
enhancer elements to increase production of the protein; changes in sequence to
increase solubility of the toxin, as well as altering the final portions of the
gene to ensure the termination of gene expression.
In reality, it is difficult for researchers to obtain the
transgenes made by industry, as there is strict patent laws and resistance to
giving permission to conduct independent research on their products. Previous
studies have shown that the modified toxin is more toxic than their naturally
produced counterparts, with green lacewings suffering from delayed development
and reduced survival (see [11] GM Food & Feed Not Fit for
"Man or Beast", ISIS
Report). The new study from Hilbeck’s team was carried out with Cry1Ab toxin
from another independent lab, which is not the same as that produced in Bt
maize or those in earlier work that highlights these differences in toxicity.
The story remains unclear as to which versions are more toxic, or if there is
any non-toxic version. (The variable expression of transgenes found in
different Bt crops under different environmental conditions also complicate matters
(see [12] Scientists Confirm
Failures of Bt-Crops, SiS 28)). Currently, no regulatory body requires testing
of the modified transgene, which means that their effects have not been properly
assessed in any version.
Attacks
on researchers with ‘inconvenient’ results
The
attack on scientists who publish data that happen to
go against the safety of biotech products are under immense pressure from
GM proponents, industry and even regulatory bodies. The work is highly
scrutinised in a manner rarely seen in other non-profit-driven subject areas. As Hilbeck said in a comment piece, deliberate
counter-studies and confrontational attacks have also been witnessed with other
commercial products such as bisphenol A, asbestos and tobacco [13]. The team were
never given the opportunity to respond to their critiques.
In the case of Bt toxicity, this is not the first time that the
researchers have faced such scrutiny; the publications on lacewing lethality [5-7]
drew a similar response from some of the same authors that targeted the
ladybird study.
To
conclude
Studies into the toxic effects of the Bt toxins are beginning to
shed light on the wider effects of Bt toxins to non-target insects. This
knowledge is critical to agricultural success with insects like the ladybirds
playing an important biological function due to their predation on crop pests
such as aphids and white flies. As it stands, the full off-target effects of
the Bt
toxins are not understood, with a reported 91 % of Bt toxins tested on 10 or
less species, most of which are presumed target species [14]. Independent studies
have however, also linked Bt exposure to abnormal growth in snails [15] and
caddisflies (see [16] Bt Crops Threaten
Aquatic Ecosystems,
SiS 36) and reduced fitness of water fleas [17]. Off-target
effects need to be investigated thoroughly prior to the release of such
products. With Bt crops already widely commercialised, we are left with the
option of withdrawing them from the market until irrefutable evidence of their
safety becomes available
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