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Surge in insect resistance to transgenic crops and prospects for sustainability

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Transgenic crops have revolutionized insect pest control, but their effectiveness has been reduced by evolution of resistance in pests. We analyzed global monitoring data reported during the first two decades of transgenic crops, with each case representing the responses of one pest species in one country to one insecticidal protein from Bacillus thuringiensis (Bt). The cases of pest resistance to Bt crystalline (Cry) proteins produced by transgenic crops increased from 3 in 2005 to 16 in 2016. By contrast, in 17 other cases there was no decrease in pest susceptibility to Bt crops, including the recently introduced transgenic corn that produces a Bt vegetative insecticidal protein (Vip). Recessive inheritance of pest resistance has favored sustained susceptibility, but even when inheritance is not recessive, abundant refuges of non-Bt host plants have substantially delayed resistance. These insights may inform resistance management strategies to increase the durability of current and future transgenic crops.
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926 VOLUME 35 NUMBER 10 OCTOBER 2017 NATURE BIOTECHNOLOGY
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Genetically engineered crops have the potential to help meet the chal-
lenge of sustainably providing food and fiber for the world’s growing
population1. In particular, transgenic crops producing insecticidal
proteins from Bt have revolutionized pest control2–5. Bt proteins kill
some voracious insect pests, but cause little or no harm to most other
organisms, including humans, wildlife, and most beneficial insects6–8.
The hectares (ha) planted with Bt crops worldwide increased from
1.1 million in 1996 to 98.5 million in 2016, with a cumulative total
of more than 830 million4 (Fig. 1a). Bt corn, cotton, and soybean
have accounted for >99% of this total. In addition to the crystalline
(Cry) proteins from Bt produced by transgenic crops for the past
two decades, some recently introduced types of Bt corn and cotton
produce a vegetative insecticidal protein (Vip) from Bt9–13. When
produced naturally by the bacteria, Cry proteins are produced dur-
ing sporulation and retained within the cell wall, whereas Vips are
produced during the vegetative phase and secreted11. Benefits of Bt
crops include pest suppression, decreased use of conventional insec-
ticides, conservation of beneficial natural enemies, increased yields,
and higher farmer profits14–20.
The benefits of Bt crops, however, are threatened by the evolu-
tion of pest resistance3,12,21–24. The scientific literature on this topic
has exploded; a Web of Science search for “Bacillus thuringiensis and
resistance” identified >1,100 papers published from 2013 to April
2017. Here, we analyze the relevant literature from the past two dec-
ades to elucidate the current status of pest resistance to transgenic
crops, to better understand how we got where we are, and to deter-
mine how we can move forward effectively. We simplify the criteria for
classifying resistance to Bt crops, update the global status of resistance
to Bt crops, briefly summarize theory and tactics for delaying resist-
ance to Bt crops (Box 1), and test the theory with data from the field.
We end by assessing the future prospects for managing resistance to
recently introduced Bt crops that produce Vip3Aa and transgenic
crops in the pipeline that combine RNA interference (RNAi) with Bt
proteins for pest control.
Compared with previous reviews on this topic24–26, the field-moni-
toring data analyzed here represent a more diverse set of Bt toxins
(one Vip and nine Cry toxins), crops (corn, cotton, and soy), pests (15
species from two insect orders), and countries (ten countries on six
continents). Strikingly, the number of cases of resistance to Bt crops
with practical consequences for pest control has more than tripled
(Fig. 1b) since completion of our previous review based on monitor-
ing data published as of 2012 (B.E.T., Y.C. et al.24).
Field-evolved resistance
Previous publications provide detailed discussion of various defini-
tions and criteria for resistance to Bt crops24,27,28. In this Review, we
define field-evolved resistance as a genetically based decrease in sus-
ceptibility of an insect population to a Bt toxin caused by selection in
the field. This is similar to, but broader than, our previous definitions
(B.E.T., Y.C. et al.)24,27 because it includes the possibility of selection
in the field by one toxin that causes cross-resistance to another toxin.
As in previous work, each case reviewed here represents responses of
one pest species in one country to one Bt toxin24.
Although we have previously used up to six categories of suscepti-
bility and resistance to Bt crops24, here we classify each case into one of
three categories: category 1, practical resistance; category 2, no decrease
in susceptibility; or category 3, early warning of resistance. Practical
resistance to a Bt crop is field-evolved resistance that reduces the effi-
cacy of the Bt crop and has practical consequences for pest control27.
The criteria for practical resistance are that >50% of individuals in a
population are resistant and the efficacy of the Bt crop is reduced in
the field27. The percentage of resistant individuals can be estimated
from survival of insects exposed to a concentration of Bt toxin that
kills all, or nearly all, susceptible individuals24,27 (Supplementary
Methods). Exposure can be mediated by allowing insects to eat Bt
Surge in insect resistance to transgenic
crops and prospects for sustainability
Bruce E Tabashnik & Yves Carrière
Transgenic crops have revolutionized insect pest control, but their effectiveness has been reduced by evolution of resistance in
pests. We analyzed global monitoring data reported during the first two decades of transgenic crops, with each case representing
the responses of one pest species in one country to one insecticidal protein from Bacillus thuringiensis (Bt). The cases of pest
resistance to Bt crystalline (Cry) proteins produced by transgenic crops increased from 3 in 2005 to 16 in 2016. By contrast, in
17 other cases there was no decrease in pest susceptibility to Bt crops, including the recently introduced transgenic corn that
produces a Bt vegetative insecticidal protein (Vip). Recessive inheritance of pest resistance has favored sustained susceptibility,
but even when inheritance is not recessive, abundant refuges of non-Bt host plants have substantially delayed resistance. These
insights may inform resistance management strategies to increase the durability of current and future transgenic crops.
Department of Entomology, University of Arizona, Tucson, Arizona, USA.
Correspondence should be addressed to B.E.T. (brucet@cals.arizona.edu).
Received 4 May; accepted 25 August; published online 11 October 2017;
doi:10.1038/nbt.3974
© 2017 Nature America, Inc., part of Springer Nature. All rights reserved.
NATURE BIOTECHNOLOGY VOLUME 35 NUMBER 10 OCTOBER 2017 927
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plants, Bt plant tissues, or diet containing a ‘diagnostic concentration’
of Bt toxin24,27. Large increases in the concentration of toxin killing
50% of the insects tested (LC50) also indicate >50% of individuals in
a population are resistant27. Cases fit category 2 when the monitoring
data show no statistically significant decrease in susceptibility after
field populations have been exposed to a Bt crop24. Whereas defini-
tions for the first two categories are the same as before, for simplicity
here, we broaden the intermediate category of ‘early warning of resist-
ance’ to include all cases of field-evolved resistance where monitor-
ing data show a statistically significant decrease in susceptibility, yet
reduced efficacy of the Bt crop has not been reported.
Global status of insect resistance to Bt crops
The 36 cases reviewed here consist of 16 cases of practical resistance,
17 cases of no decrease in susceptibility, and 3 cases of early warning of
resistance (Fig. 1, Tab l es 1 and 2, and Supplementary Notes 1 and 2).
Of the 15 pest species monitored, 14 are lepidopterans and one is a
coleopteran (Diabrotica virgifera virgifera, western corn rootworm).
Increasingly rapid evolution of practical resistance. The cumula-
tive number of cases of practical resistance to the Bt toxins in trans-
genic crops surged from 3 in 2005 to 16 in 2016 (Fig. 1b and Tab l e 1).
These 16 cases represent resistance of some populations of seven
major pests in five countries to each of the nine Cry toxins produced
by widely grown Bt crops: Cry1Ab, Cry1Ac, Cry1A.105, Cry1Fa,
Cry2Ab, Cry3Bb, mCry3A, eCry3.1Ab, and Cry34/35Ab (Tab le 1).
For these 16 cases of practical resistance, the average time from the
first commercial planting of a Bt crop in a region to the first sampling
of field populations in the region that provided evidence of resistance
was 5.2 years (s.e.m. = 0.7, range: 0 to 10 years) (Tab l e 1). Practical
resistance to Bt corn has been documented for some populations of
five pest species (Busseola fusca, Diatraea saccharalis, D. v. virgifera,
Spodoptera frugiperda, and Striacosta albicosta), to Bt cotton for one spe-
cies (Pectinophora gossypiella), and to both Bt corn and cotton for the
remaining species (Helicoverpa zea). The rise in total reported cases of
practical resistance from 5 in 2012 (ref. 24) to 16 in 2016 reflects 2 cases
for species that had no practical resistance before (D. saccharalis and
S. albicosta) and 9 additional cases for four species that had practical
resistance to only one Bt toxin previously and now resist up to four toxins
(Tab l e 1). Practical resistance has reduced the number of Bt toxins in
transgenic crops that are effective against some populations of major
pests to two, one, or none (Table 3).
Evolution of practical resistance has accelerated over the past two
decades, as shown by the significant negative association between
the time for practical resistance to occur and the year when a Bt crop
was first grown commercially (Fig. 2). Cross-resistance to one Bt
toxin caused by selection with another Bt toxin is an important factor
accelerating the evolution of practical resistance (B.E.T.)29 (Fig. 2).
For D. v. virgifera, cross-resistance caused by resistance to Cr y3Bb
is implicated in resistance to the closely related toxins mCry3A and
eCry3.1Ab30,31. Resistance to eCry3.1Ab was detected in the field
before plants producing this toxin were commercialized, providing
a
c
0
10
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50
60
70
80
90
100
Bt crops (million ha)
b
0
2
4
6
8
10
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18
1996
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1996
1998
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2002
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Total cases of practical
resistance to Bt crops
Year Year
Practical resistance
Early warning
Susceptible
Figure 1 Global status of pest resistance to Bt crops. (a) Hectares planted to Bt crops each year. (b) Cumulative cases of field-evolved practical
resistance to Bt crops. (c) Each symbol represents 1 of 36 cases indicating responses of one pest species in one country to one toxin in Bt corn, cotton,
or soy (Tables 1 and 2).
© 2017 Nature America, Inc., part of Springer Nature. All rights reserved.
928 VOLUME 35 NUMBER 10 OCTOBER 2017 NATURE BIOTECHNOLOGY
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Box 1 Resistance management from theory to practice
The most widely used strategy for delaying evolution of pest resistance to Bt crops is to grow ‘refuges’ of host plants that do not make Bt
toxins and thereby boost survival of susceptible pests16,21,24,66,132,133. The hope is that rare resistant pests that survive on Bt plants will
mate with the relatively abundant susceptible pests that thrive in refuges. If inheritance of resistance is recessive, the resulting hetero-
zygous offspring will die on Bt crops, greatly delaying the evolution of resistance. This is sometimes called the ‘high-dose refuge strategy
because it works best if the dose of toxin ingested is high enough to kill all, or almost all, of the heterozygous insects that feed on Bt plants.
In principle, the high-dose standard can be assessed by measuring the survival of resistant insects, susceptible insects, and their F1
progeny on Bt plants24. This allows calculation of the dominance parameter h, which varies from 0 for completely recessive to 1 for com-
pletely dominant134. Values of h less than 0.05 satisfy the high-dose criterion24. Alternatively, several indirect tests measure survival of
susceptible insects on Bt plants135. The US Environmental Protection Agency (EPA) indicates that Bt plants meet the high-dose standard
if they kill at least 99.99% of susceptible insects135. This criterion reflects the concept that if Bt plants do not kill all or nearly all suscep-
tible insects, they probably will not kill nearly all insects that are heterozygous for resistance. If survival of susceptible insects is >0.01%,
then survival is likely to be higher for the heterozygotes than for the homozygous susceptible insects, which yields nonrecessive inheritance
of resistance that accelerates adaptation24. If a high dose is not achieved, resistance can be delayed by increasing refuge abundance,
which compensates for survival of heterozygous progeny on Bt plants by reducing the proportion of the population selected for resistance24.
Extending the efficacy of Bt crops by increasing refuge abundance can impose a short-term cost of greater pest damage to the non-Bt
crop refuge. However, in the United States, planting non-Bt corn refuges yielded growers $4.3 billion in short-term benefits because of
the lower cost of non-Bt seed and regional suppression of the primary pest O. nubilalis by Bt corn16. Also, in China, millions of farmers
voluntarily increased planting of non-Bt cotton refuges, apparently to achieve short-term economic gains124 (see below).
Overall, five factors favor success of the refuge strategy for delaying resistance: recessive inheritance of resistance (i.e., plants meet
the high-dose standard), low resistance allele frequency, abundant refuges of non-Bt host plants near Bt plants, fitness costs, and
incomplete resistance24,136. When fitness costs occur, fitness on non-Bt host plants is higher for susceptible insects than resistant
insects, so refuges select against resistance. Incomplete resistance occurs when homozygous resistant insects can survive on Bt plants,
but they suffer a disadvantage relative to resistant insects on non-Bt plants. When a potentially heterogeneous mixture of resistant and
susceptible individuals is tested, less than 100% survival on Bt plants is expected, so such results alone are not sufficient to infer
incomplete resistance (Supplementary Methods).
Whereas each of the first-generation Bt plants makes a single Bt toxin, second-generation Bt plants each produce two or more Bt toxins
to address one or more of the following goals: to delay or counter resistance, improve efficacy, and broaden the spectrum of pests killed64.
Bt crop ‘pyramids’ are designed to delay the evolution of resistance by producing two or more distinct toxins or other traits that kill the
same pest34,64. Based on modeling and experimental evidence, pyramids are considered more effective for delaying resistance than tempo-
ral alternations or spatial mosaics of crops with different Bt toxins137. First commercialized in 2003, Bt crop pyramids have become preva-
lent globally, with the notable exception of transgenic cotton producing a single Bt toxin still grown throughout China24. Although some
cotton producing a cowpea trypsin inhibitor plus Bt toxin Cry1Ac has been planted in China, a three-year field study found that relative to
cotton producing Cry1Ac alone, addition of the trypsin inhibitor did not significantly decrease the population density of H. armigera138.
The five factors listed above favor durability of pyramids as well as single-toxin transgenic crops. In addition, the following three fac-
tors are especially important for delaying pest resistance to pyramids: the concentration of each toxin in the pyramid is high enough to
kill all or nearly all susceptible insects, no cross-resistance occurs between toxins in the pyramid, and pyramids are not grown simultane-
ously with single-toxin plants that produce one of the toxins in the pyramid34,64,139. In some cases, the efficacy and durability of Bt crop
pyramids has been reduced by resistance to single-toxin crops producing the same toxins used in pyramids, as well as cross-resistance
and antagonism between Bt toxins34,64.
Although refuges are essential for durability of both pyramids and single-toxin transgenic crops, the optimal spatial configuration of
refuges remains unresolved. Blocks of non-Bt plants called ‘structured refuges’ have been cultivated in separate fields or within fields of
Bt crops to delay pest resistance since 1996 (refs. 34,45,64). In 2007, to delay pest resistance to Bt cotton pyramids, the US EPA ap-
proved ‘natural refuges’ consisting of non-Bt host plants other than cotton46. This approach has been effective for prolonging the efficacy
of Bt cotton against H. virescens, but not against H. zea (Tab les 1 and 2). Although both of these pests use many non-Bt host plants
other than cotton, the Cry1 and Cry2 toxins in Bt cotton meet the high-dose standard for H. virescens, but not for H. zea (Tab les 1 and 2).
Natural refuges have helped to slow, but not stop, adaptation to Bt cotton by H. armigera in northern China74 (Table 2). The effective
refuge percentage including non-cotton host plants was estimated as 56% for H. armigera in northern China74 versus 39% for H. zea in
Arkansas and Mississippi of the southeastern United States140. Cultivation of non-Bt host plants and Bt cotton in small fields that are
close to each other may also boost the success of the natural refuge strategy in northern China74.
Since 2010, seed mixtures (also called ‘refuge-in-a-bag’ or RIB) yielding a random mixture of Bt plants and non-Bt plants side-by-
side within fields have been planted to delay pest resistance to Bt corn pyramids64. Seed mixtures solve the problem of farmers not com-
plying with block refuge requirements. However, results from modeling and small-scale experiments indicate that if larvae move between
Bt and non-Bt plants, seed mixtures may accelerate evolution of resistance by reducing the survival of susceptible insects and the effec-
tive refuge size, or by increasing the survival of heterozygotes relative to susceptible homozygotes, thereby increasing the dominance of
resistance in seed mixtures relative to blocks of Bt crops (Y.C., B.E.T. et al.)64.
In the Yangtze River Valley of China, millions of growers serendipitously implemented a novel seed mixture strategy by planting
second-generation seeds from crosses between Bt and non-Bt cotton, which yields a refuge of 25% non-Bt plants randomly interspersed
within fields of Bt cotton124. Analysis of 11 years of field monitoring data from six provinces implies that this approach delayed or even
reversed P. gossypiella resistance to single-toxin Bt cotton while sustaining pest suppression124.
© 2017 Nature America, Inc., part of Springer Nature. All rights reserved.
NATURE BIOTECHNOLOGY VOLUME 35 NUMBER 10 OCTOBER 2017 929
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direct evidence of cross-resistance31. Several lines of evidence suggest
that cross-resistance to Cry2Ab caused by resistance to Cry1Ac in
Bt cotton hastened the markedly decreased susceptibility of H. zea
field populations to Cry2Ab in the southern United States in 2005,
only two years after commercial planting of Bt cotton producing
Cry2Ab32,33. Although strong cross-resistance generally does not
occur between Cry1 and Cr y2 toxins, statistically significant but
weak cross-resistance is typical34–37 . Resistance to Cry1Ab in H. zea
probably caused some cross-resistance to Cry1A.10536,95. In Brazil,
cross-resistance between Cry1Ab and Cry1Fa cannot be excluded as
a factor accelerating evolution of practical resistance to these toxins
in S. frugiperda, which occurred two years after Bt corn producing
each toxin was grown commercially38–40.
In addition to cross-resistance, factors favoring faster evolution of
resistance to more recently commercialized Bt crops are increased
adoption rates and concomittantly reduced percentages of host plants
that are refuges of non-Bt crops. The increase in cases of practical
resistance in the second decade of Bt crops is also associated with
an increase in the total area planted with Bt crops (Fig. 1), expo-
sure of pests to Bt crops in more countries, and longer cumulative
exposure of pests to Bt crops. The number of new cases of practical
resistance reported relative to the yearly mean area planted with Bt
crops globally is similar for the first and second decades of Bt crops:
2.3 and 2.0 new cases per 10 million ha, respectively (3 cases/12.9
million ha for 1996–2005 and 13 cases/63.8 million ha for 2006–
2016). Nonetheless, the 16 cases of practical resistance analyzed here
underestimate the situation in the field, in part because of the delay
between field sampling that yields evidence of resistance and publica-
tion of the data (mean = 3.4 years, s.e.m. = 0.7, n = 16 cases).
We tested the hypothesis that resistance evolved faster for pests that
fed on Bt crops for more generations per year (GPY). The negative
association between time to practical resistance and GPY was not
significant when all 16 cases of practical resistance were considered
(n = 16, P = 0.19), but it was significant when the five cases of cross-
resistance were excluded (n = 11, P = 0.022) (Supplementary Table 1
and Supplementary Fig. 1). Multiple regression for all 16 cases
also shows that the negative relationship between time to practical
resistance and GPY was significant (P = 0.049) after accounting for the
significant effect of cross-resistance (P = 0 . 00 5) ( Supplementary Table 2).
The mean GPY did not differ significantly between the 16 cases of
practical resistance (4.2, s.e.m. = 0.9) and the 17 cases of sustained
susceptibility (4.1, s.e.m. = 0.5) (t-test, t = 0.04, df = 31, P = 0.97)
(Supplementary Tables 1 and 3).
Sustained susceptibility. In contrast with the 16 cases of practical
resistance described above, the global monitoring data reveal 17 cases
where no significant decrease in susceptibility occurred after 1 to
19 years of exposure to Bt crops (mean = 10.6 years, s.e.m. = 1.4;
Tab l e 2). These 17 cases include data from six countries indicating
susceptibility to five toxins in Bt crops for populations of nine species
of lepidopteran pests: Chrysodeixis includens, Diatraea grandiosella,
Helicover pa armigera, Helicoverpa punctigera, Heliothis virescens,
Ostrinia nubilalis, P. gossypiella, S. frugiperda, and Sesamia nona-
groides. In 11 of these 17 cases, no decrease in susceptibility has been
demonstrated for at least 10 years (Tab l e 2). Moreover, in all of these
cases, the currently available monitoring data underestimate the ulti-
mate duration of sustained susceptibility, because this can be known
only after resistance occurs.
Early warning of resistance. The three cases of early warning of
resistance involve responses of Diatraea saccharalis in the United
States and Ostrinia furnacalis in the Philippines to Cry1Ab, and
H. armigera in China to Cry1Ac (Table 2 and Supplementary Note 2).
Testing theory with data
Consistent with previous results from smaller data sets24,25, the data
from the 36 cases reviewed here support the main predictions from
the evolutionary theory underlying the refuge strategy (Box 1). When
the high-dose standard is met, which indicates recessive inheritance
of resistance, resistance is less likely to evolve rapidly. In the 30 cases
where the available data enable evaluation of this factor, the high-dose
standard was met for 69% (9 of 13) of the cases with no decrease in
susceptibility and with none of the 17 cases showing either practical
resistance or early warning of resistance (Tables 1 and 2). This pattern
demonstrates a significant association between meeting the high-dose
standard and a lower risk of rapid evolution of resistance (Fishers
exact test, n = 30, P < 0.0001).
In two of the four exceptional cases where the high-dose stand-
ard was not met and susceptibility did not decrease (Tab l e 2), pest
exposure to the relevant Bt toxins was limited: O. nubilalis exposure
to Cry1F corn in the United States41 and C. includens exposure
to Cry1Ac soy in Brazil42 . In the third exceptional case, O. nubi-
Table 1 Practical resistance to Bt crops
Insect Crop Toxin Country Year marketedaYearsbHigh dosecReferencesd
B. fusca Corn Cry1Ab S. Africa 1998 8 No 87,88
D. saccharalis Corn Cry1A.105 Argentina 2010 4 ? 51,89–91
D. v. virgifera Corn Cry3Bb USA 2003 6 No 28,86
D. v. virgifera Corn Cry34/35Ab USA 2006 7 No 28,92,93
D. v. virgifera Corn mCry3A USA 2007 4eNo 28,94
D. v. virgifera Corn eCry3.1Ab USA 2014 0eNo 28,30,31
H. zea Corn Cry1Ab USA 1996 8 No 95,96
H. zea Corn Cry1A.105 USA 2010 6eNo 95
H. zea Cotton Cry1Ac USA 1996 6 No 24,97,98
H. zea Cotton Cry2Ab USA 2003 2eNo 24,32,99
P. gossypiella Cotton Cry1Ac India 2002 6 No 50,100–102
P. gossypiella Cotton Cry2Ab India 2006 8 ? 75
S. albicosta Corn Cry1Fa USA 2003 10 No 103–106
S. frugiperda Corn Cry1Ab Brazil 2008 2eNo 40
S. frugiperda Corn Cry1F Brazil 2009 2 No 38,107
S. frugiperda Corn Cry1F USA 2003 4 No 60,108
aFirst year of commercial planting of a Bt crop in the region monitored. bYears from the first commercial planting of a Bt crop in the region to the first sampling of field populations in
the region yielding evidence of resistance. cTest for the high-dose standard based on direct or indirect evidence (Box 1). If both types were available, the table reflects the direct evidence.
“?” indicates data not available. dEach reference provides evidence of practical resistance, data for evaluating the high-dose criterion, or both. eCross-resistance suspected or known as a factor
contributing to resistance.
© 2017 Nature America, Inc., part of Springer Nature. All rights reserved.
930 VOLUME 35 NUMBER 10 OCTOBER 2017 NATURE BIOTECHNOLOGY
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lalis and Cry1Ab corn, lar vae from the resistant strain tested did
not sur vive on young vegetative-stage plants, but larvae from the
resistant strain and the progeny from a cross between the resistant
strain and a susceptible strain survived on older reproductive-stage
plants, yielding partially recessive resistance that does not meet
the high-dose standard (h = 0.31) (ref. 43). Larvae surviving on
reproductive-stage corn were found feeding on reproductive tis-
sues that have lower toxin concentration than the leaves eaten by
larvae on vegetative-stage corn43. However, this evaluation of the
high-dose criterion is based on survival after only 15 days, which
could overestimate dominance43.
The fourth exceptional case, the sustained efficacy of Cry1Ac
against H. armigera in Australia for two decades (Tab le 2), is par-
ticularly instructive. Recognizing that the high-dose standard was
not fully satisfied, the Australian resistance management plan for
cotton producing Cry1Ac proactively required a non-Bt cotton
refuge of at least 70% from 1996 to 2003 (ref. 44). Susceptibility was
maintained to Cry1Ac cotton until it was replaced in 2004 by two-
toxin cotton producing Cry1Ac and Cry2Ab. When the two-toxin cot-
ton was introduced, the refuge requirement dropped to 10% unsprayed
non-Bt cotton or its equivalent44. This approach has yielded no net
decrease in susceptibility of H. armigera to either of the two toxins.
By contrast, practical resistance to both toxins has evolved in the
closely related species H. zea in the United States (Ta bl e 1), where
refuge requirements have been less stringent. In the United States,
the minimum refuge required was 4% unsprayed non-Bt cotton for
Cry1Ac cotton when it was first grown commercially in 1996 (ref. 45);
and in most regions, no refuge has been required for two-toxin Bt
cotton since 2007 (refs. 24,46).
The outcomes summarized above imply that when the high-
dose standard is not met, proactive deployment of abundant ref-
uges can substantially delay resistance. Conversely, along with failure
to meet the high-dose standard, the scarcity of refuges seems
to be a key factor contributing to the cases of practical resist-
ance in Argentina, Brazil, India, South Africa, and the United
States24,28,38,39,47–51 (Ta bl e 1).
Managing resistance of lepidopteran pests to Vip3Aa
Compared with the extensive exposure of pests to Cry proteins dur-
ing the past two decades, exposure to vegetative insecticidal proteins
(Vips) from Bt has been limited and no field-evolved resistance has
been reported. Bt produces both Cry and Vip toxins11, which could
be a ‘natural pyramid strategy’ (Box 1). Although four Vip families
with a total of >100 toxins are known11, Vip3Aa is the only Vip in
commercialized transgenic crops. Because Vip3Aa19 in Bt corn
and Vip3 Aa20 in Bt cotton are 99.9% identical in their amino acid
sequence (Y.C., B.E.T. et al.)34, we refer to both as Vip3Aa. Vip3Aa
kills some lepidopteran pests and is produced in combination with
the lepidopteran-active proteins Cry1Ab, Cry1F, or both in Bt corn
and with Cry1 and Cry2A toxins in Bt cotton34,52.
Table 2 No decrease in susceptibility and early warning of resistance to Bt crops
Insect Crop Toxin Country Year marketedaYearsbHigh dosecReference
No decrease in susceptibility
C. includens SoydCry1Ac Brazil 2013d1dNo 42,109
D. grandiosella Corn Cry1Ab USA 1999 6 ? 110
H. armigera Cotton Cry1Ac Australia 1996 19 No 111–113
H. armigera Cotton Cry2Ab Australia 2004 11 Yes 12,114
H. punctigera Cotton Cry1Ac Australia 1996 19 ? 111
H. punctigera Cotton Cry2Ab Australia 2004 11 Yes 12,114
H. virescens Cotton Cry1Ac Mexico 1996 11 ? 115
H. virescens Cotton Cry1Ac USA 1996 11 Yes 98,115,116
H. virescens Cotton Cry2Ab USA 2003 2 Yes 32,117–119
O. nubilalis Corn Cry1Ab Spain 1998 15 ? 120,121
O. nubilalis Corn Cry1Ab USA 1996 15 No 43,122
O. nubilalis Corn Cry1Fa USA 2003 8 No 41,123
P. gossypiella Cotton Cry1Ac China 2000 15 Yes 124
P. gossypiella Cotton Cry1Ac USA 1996 12 Yes 17
P. gossypiella Cotton Cry2Ab USA 2003 5 Yes 17,125,126
S. frugiperda Corn Vip3Aa Brazil 2010 5 Yes 10
S. nonagroides Corn Cry1Ab Spain 1998 15 Yes 127
Early warning of resistancee
D. saccharalis Corn Cry1Ab USA 1999 10 No 24,128,129
H. armigera Cotton Cry1Ac China 1997 16 No 74
O. furnacalis Corn Cry1Ab Philippines 2003 6 No 130,131
aFirst year of commercial planting of a Bt crop in the region monitored. bFor cases with no decrease in susceptibility, this column shows years of documented susceptibility, calculated as the year
of the most recent monitoring data cited minus the first year of commercialization in the region. For early warning of resistance, this column shows the years from the first year of commercial
planting in the region to the most recent year of monitoring data reviewed here. cTest for the high-dose standard based on direct or indirect evidence (Box 1). If both types were available, the
table reflects the direct evidence. ‘?’ indicates data not available. dThe first season of commercial planting in Brazil for transgenic plants producing Cry1Ac was 2013-14 for soy and 2006–2007
for cotton, which is an occasional host of C. includens42. Based on monitoring data from the 2014–2015 season, documented susceptibility to Cry1Ac is 1 year since introduction of Bt soy
and 8 years since introduction of Bt cotton. eThe highest percentage of resistant individuals reported for any field population screened (based on survival at a diagnostic concentration in diet
bioassays) was 2.4% for D. saccharalis, 11.3% for H. armigera, and 5.5% for O. furnacalis.
Table 3 Limited availability of transgenic crops to control pests
that have practical resistance to two or more Bt toxins
Insect Crop Country
Practical resistance
reporteda
Practical resistance
not reported and
toxin effective
D. v. virgifera Corn USA Cry3Bb, eCry3.1Ab,
mCry3A, Cry34/35Ab
Noneb
P. gossypiella Cotton India Cry1Ac, Cry2Ab Noneb
H. zea Corn and
cotton
USA Cry1Ab, Cry1Ac,
Cry1A.105, Cry2Ab
Vip3Aa
S. frugiperda Corn Brazil Cry1Ab, Cry1F Cry2Ab, Vip3Aac
aSee Table 1. bIn the country listed, no Bt toxins in currently commercialized transgenic crops
remain effective against all populations of the pest, but some Bt toxins in currently com-
mercialized transgenic crops do remain effective against some populations of the pest. Note:
Vip3Aa is not effective against D. v. virgifera or P. gossypiella. cThe combination of Cry2Ab
and Vip3Aa is not produced by any widely adopted transgenic corn hybrids, so each toxin acts
alone against pest populations resistant to Cry1 toxins.
© 2017 Nature America, Inc., part of Springer Nature. All rights reserved.
NATURE BIOTECHNOLOGY VOLUME 35 NUMBER 10 OCTOBER 2017 931
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In the United States, corn and cotton producing Vip3Aa were first
registered in 2008 (ref. 33) and first grown commercially in 2011 and
2014, respectively (Supplementary Table 4). In the United States,
Vip3Aa corn accounted for roughly 1.1% and 3.5% of all corn planted in
2011 and 2013, respectively, while Vip3Aa cotton was less than 1% of all
cotton planted annually from 2014 to 2016 (Supplementary Table 4).
The only type of Vip3Aa cotton grown commercially in the
United States from 2014 to 2016 also produces Cry1F and Cry1Ac
(Widestrike 3) (Supplementary Table 4); introduction is expected in
2017 for cotton producing Cry1Ac + Cry2Ab + Vip3Aa (Bollgard 3)
and Cry1Ab + Cry2Ae + Vip3Aa (Twinlink Plus)53.
In Brazil, Vip3Aa corn was approved in 2009 and has accounted
for <5% of all corn10, while Vip3Aa cotton has not been grown com-
mercially. In Australia, Bt corn is not grown and the percentage of
hectares planted with cotton producing Cry1Ac + Cry2Ab + Vip3Aa
increased from about 7.6% in the 2015–2016 season to >90% in the
2016–2017 season (Supplementary Table 4). Aside from Australia,
the low adoption of Vip3Aa crops to date suggests their value is pri-
marily for managing resistance, rather than providing immediate
economic benefits.
The genetic potential for field-evolved resistance to Vip3Aa is dem-
onstrated by laboratory selection with Vip3Aa that yielded 285- to
>3,000-fold resistance to this toxin in five major lepidopteran pests
(H. armigera, H. punctigera, H. virescens, S. frugiperda, and Spodoptera
litura)9,54–57. Based on F2 screens conducted from 2009 to 2013,
before field populations in Australia were exposed to crops produc-
ing Vip3Aa, the estimated frequency of alleles conferring resistance
to Vip3Aa was higher than expected: 0.034 for H. armigera11 and
0.010 for H. punctigera12. A similar study in Brazil10 estimated the
frequency of Vip3Aa resistance alleles in S. frugiperda as 0.0009 in
2013 to 2014.
Although Cry and Vip toxins have the same general mode of action,
they have no structural homology and bind to different sites in the
insect midgut, so cross-resistance between them is predicted to be
low or nil11,34,57. The experimental evidence supports this hypothesis.
In 11 evaluations of cross-resistance based on comparisons between
related strains of five pest species, 57- to 20,000-fold resistance to
Cry1Ac or Cry1F caused at most 3.2-fold cross-resistance to Vip3A
(Supplementary Table 5). In four comparisons between related
strains of H. virescens, 2,000-fold resistance to Vip3A caused up to
sevenfold cross-resistance to Cry1Ab or Cry1Ac56. Analysis of all
15 experiments mentioned above reveals weak, but statistically sig-
nificant cross-resistance between Vip3Aa and Cry1Ab, Cry1Ac, or
Cry1F (mean = 1.8-fold cross-resistance, n = 15, one-sample t-test of
log-transformed resistance ratios, P = 0.01).
Comparisons between unrelated insect strains or populations,
which can be influenced by differences in genetic background and are
less rigorous than the aforementioned comparisons between related
strains, also show no strong cross-resistance between Vip3Aa and
Cry1Ac9,13,58,59. For 142 families of S. frugiperda produced by sin-
gle pairs of field-collected adults, survival on corn producing Vip3Aa
was not correlated with survival on corn producing Cry1F60. For
H. armigera, a Cry2Ab-resistant strain generated by CRISPR–Cas9
knockout of the ABC transporter gene HaABCA2 wa s not cro ss-r esis t-
ant to Vip3Aa13. Likewise, strains of H. armigera and H. punctigera
selected for >200-fold resistance to Vip3Aa had either increased sus-
ceptibility to Cry2Ab or at most 1.7-fold cross-resistance to Cry2Ab
relative to unrelated susceptible strains9.
Pyramiding Vip3Aa and Cry toxins diminishes the risk of resist-
ance in pest populations that are susceptible to both types of toxin34
(Box 1). For example, in Australia, cotton producing Cry1Ac, Cry2Ab,
and Vip3Aa was introduced when populations of H. armigera and
H. punctigera were susceptible to all three toxins, which greatly lowers
the risk of resistance12.
Conversely, in the United States, some populations of H. zea are
already resistant to the Cry1 and Cry2A toxins used in combination
with Vip3Aa in Bt corn and cotton (Ta bl e 1 and Supplementary
Tab l e 4 ). Moreover, while plants producing Vip3Aa are gradually being
more widely adopted in the United States, resistance of H. zea to Cry1
and Cry2A toxins is likely to become more widespread. This increases
the chances that in the near future, Vip3Aa will be the only toxin in
commercialized Bt plants that is effective against some populations
of H. zea (Ta b le 3), which markedly raises the risk of resistance.
In addition, exposure of H. zea to Vip3Aa in both corn and cotton
increases selection for resistance to this toxin. According to the US
Environmental Protection Agency, cotton producing Vip3Aa alone met
the high-dose standard against H. zea in one test, but not in an othe r61.
Corn producing Vip3Aa alone does not meet the high-dose criterion
against H. zea62,63. Therefore, if corn producing Vip3A is widely
deployed, large refuges will be needed to delay resistance in H. zea.
For S. frugiperda, resistance to Cry1 toxins (Tab le 1) increases
the risk of resistance to corn and cotton producing Cry1 toxins in
combination with Vip3Aa53. Yet, corn producing Vip3Aa meets the
high-dose standard against S. frugiperda52,55,62,63, which lowers the
risk of resistance. Also, field-evolved resistance to Cry2A toxins has
not been reported for S. frugiperda, and Cry2A toxins are effective
against strains of S. frugiperda resistant to Cry1F or Vip3Aa52,53,60.
Therefore, the cotton pyramids producing Vip3Aa together with
Cry2Ab or Cry2Ae could help to delay resistance. In corn, however,
some current hybrids produce either Vip3Aa or Cry2Ab64,65, but
recently developed hybrids producing both of these toxins have not
been widely adopted. Thus, in effect, Cry1-resistant populations on
corn may be selected by a mosaic of single-toxin plants exposing some
larvae to Vip3Aa and others to Cry2Ab (Tab l e 3), which is considered
the least durable way to deploy two toxins66.
0
2
4
6
8
10
12
Years to resistance
1996
1998
2000
2002
2004
2006
2008
2010
2012
2014
Year first commercialized
Figure 2 Increasingly rapid evolution of pest resistance to Bt crops.
For the 16 cases of practical resistance to Bt crops (Table 1), the time
from the first commercial planting of a Bt crop to the first evidence of
resistance (years to resistance) decreased over the past two decades
(linear regression: y = 0.32x + 643, R2 = 0.35, df = 14, P = 0.016). The
squares indicate five cases where cross-resistance is suspected or known
to have shortened the time to resistance, including western corn rootworm
resistance to eCry3.1Ab detected in the field in 2014 before plants
producing this toxin were grown commercially.
© 2017 Nature America, Inc., part of Springer Nature. All rights reserved.
932 VOLUME 35 NUMBER 10 OCTOBER 2017 NATURE BIOTECHNOLOGY
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Next-generation insect control
RNAi offers great promise as an alternative, or complement, to Bt tox-
ins in transgenic crops for managing insect pests67–69. In RNAi, small
double-stranded RNA (dsRNA) causes sequence-specific suppression
of target gene expression. To achieve safe and effective pest control
with RNAi, the goal is to reduce expression of genes encoding proteins
that are essential to pests, but not to other organisms. Because the
mode of action differs markedly between Bt toxins and RNAi, strong
cross-resistance between them is not expected.
Transgenic crops under development that kill pests with a combi-
nation of Bt toxins and RNAi include corn targeting D. v. virgifera68
and cotton targeting H. armigera69. The corn produces two Bt toxins
(Cry3Bb and Cry34/35Ab) active against Diabrotica species and a
dsRNA transcript. The dsRNA transcript contains a 240-base pair
fragment of the D. v. virgifera gene encoding a protein (DvSnf7) vital
for intracellular protein sorting68,70. In diet bioassays, the DvSnf7
dsRNA killed larvae of D. v. virgifera a nd Diabrotica undecimpunctata
howardi, but not larvae from five other genera of beetles or insects
from seven species representing three other orders71.
The LC50 of DvSn7 dsRNA was 2.7-fold higher for a Cry3Bb-resist-
ant strain (Gass-R) than a related Cry3Bb-susceptible strain (Gass-S)
of D. v. virgifera, indicating that resistance to Cry3Bb caused statisti-
cally significant, but weak cross-resistance to this form of RNAi72.
Although no significant correlation occurred between susceptibil-
ity to Cry3Bb and DvSn7 dsRNA across eight unrelated strains of
D. v. virgifera72, this type of analysis is less rigorous than the compari-
son between related strains noted above, because differences among
strains in genetic background could mask weak cross-resistance. In
greenhouse bioassays, beetle emergence was higher for Gass-R than
Gass-S on corn producing DvSn7 dsRNA either alone, with Cry3Bb,
or with Cry3Bb + Cry34/35Ab; but this difference was statistically
significant only for DvSn7 dsRNA with Cry3Bb72. In field tests where
resistance to Cry3Bb was likely, DvSn7 dsRNA reduced emergence
of D. v. virgifera adults by about 80–95% (ref. 68). Corn producing
DvSn7 dsRNA, Cry3Bb, and Cry34/35Ab for rootworm control, along
with Cry1A.105, Cry1Fa, and Cry2Ab for control of caterpillar pests
is under consideration for registration by the US Environmental
Protection Agency68.
In related work, Ni et al.69 developed two kinds of transgenic
cotton plants (JHA and JHB) producing dsRNA that kills larvae of
H. armigera by interfering with their juvenile hormone (JH). JH is
critical for insect development, yet absent from most other organ-
isms73. JHA cotton suppresses JH acid methyltransferase, which is
crucial for JH synthesis, while JHB cotton suppresses JH binding
protein, which transports JH to organs69. In bioassays with cotton
leaves, mortality of H. armigera larvae was 57–64% for JHA cotton
and 66–70% for JHB cotton69. Mortality caused by JHA and JHB cot-
ton did not differ significantly between a Cry1Ac-resistant strain and
a related susceptible strain of H. armigera, indicating no cross-resist-
ance. For cotton plants protected by both RNAi (either JHA or JHB)
and a Bt toxin similar to Cry1Ac, these two traits acted independently
and caused 92–93% mortality of a susceptible strain.
Results from modeling suggest that combining RNAi-mediated pro-
tection with one or more Bt toxins can delay the evolution of resist-
ance, but the gain in durability depends on the refuge percentage69. For
example, under realistic assumptions, the predicted delay for resist-
ance to evolve to a cotton combining both Bt and RNAi (Bt+RNAi)
relative to Bt cotton is 4 years with a 5% refuge versus 14 years with a
50% refuge69. For Bt+RNAi corn, a 5% refuge was simulated, and the
increase in durability for Bt+RNAi corn relative to Bt corn was 1 to 5
years under some realistic scenarios (fig. 5b,d in ref. 68). In northern
China where Bt cotton is grown extensively, abundant non-Bt host
plants provide an effective refuge estimated as 56% for H. armigera74.
By contrast, for many populations of D. v. virgifera in the midwestern
United States, the refuge percentage may be close to 5%, which is the
minimum under current regulations48,65.
Because resistance to RNAi has not been reported yet in the lab-
oratory or field, the assumptions in models about this adaptation
remain to be tested. Nonetheless, the qualitative effects of refuge
percentage were similar across a broad range of assumptions, which
suggests that these trends are robust and larger refuges can greatly
extend efficacy69.
Outlook for managing resistance
When the first Bt crops were commercialized more than 20 years
ago, strategies for delaying pest resistance relied entirely on theo-
retical projections from modeling. Since then, global monitoring has
documented both remarkable successes and disappointing failures
in terms of managing pest resistance to Bt crops (Ta bl e s 1 and 2). In
the best cases, despite high adoption of Bt crops, pest resistance has
been delayed for close to two decades, with excellent prospects for
continued pest suppression. These successes include sustained sus-
ceptibility of H. armigera and H. punctigera in Au stra lia; H. virescens,
O. nubilalis, and P. g o s s y p i e l l a in the United States; and P. g o s s y p i e l l a
in China (Tab l e 2).
Conversely, for the 16 cases of practical resistance, the average time
for evolution of resistance was only 5.2 years (Ta b le 1). In four situ-
ations, practical resistance has reduced the number of Bt toxins that
are available in commercialized transgenic crops and still effective
against some pest populations to two, one, or none (Tab l e 3). In India,
no transgenic cotton is available now or expected to be available in
the next several years to control P. g o s s y p i e l l a populations resistant
to Cry1 and Cry2 toxins75 (Ta b le 1). Cotton producing Vip3Aa alone
had minimal efficacy against this pest61, and cotton has not been
engineered to produce the genetically modified Cry toxins that kill
P. g o s s y p i e l l a r esistant t o Cry1 and Cry2 toxi ns (B. E.T. et al.)76. In the
United States, practical resistance to each of the four coleopteran-
active Bt toxins produced by corn has been documented for some
populations of D. v. virgifera (Ta bl e 1 and Supplementary Note 1).
Transgenic corn protected from this pest by RNAi might be available
commercially in a few years68. However, this new trait does not meet
the high-dose standard68, and its efficacy may be short-lived unless
refuge requirements are markedly increased, and the transgenic corn
is used with other control measures such as crop rotation48,77. We
also caution that, despite the encouraging results with Bt+RNAi cot-
ton against H. armigera6 9, it has generally been difficult to control
lepidopteran pests with RNAi78.
Modeling results and empirical evidence show that refuge require-
ments must be tailored to each pest–transgenic-crop combination. For
plants producing a single Bt toxin, when the high-dose standard is met
and resistance is rare, refuges accounting for as little as 20% of a pest’s
host plants may be sufficient to delay resistance for a decade or more.
Conversely, when the high-dose standard is not met and resistance is
not rare, larger refuges (e.g., 50%) are needed to substantially delay pest
adaptation21,24,25,48. Similar principles apply to pyramids. Modeling
and empirical evidence suggest that refuges of 10% can be effective
for delaying resistance of pest populations that are highly suscepti-
ble to each of two or more independently acting toxins or traits in a
pyramid12,44,79. However, smaller refuges are risky even under optimal
conditions79,80, and much larger refuges are needed to substantially
delay resistance if each of the toxins or traits in a pyramid is not highly
effective, either inherently or because of field-evolved resistance34,64.
© 2017 Nature America, Inc., part of Springer Nature. All rights reserved.
NATURE BIOTECHNOLOGY VOLUME 35 NUMBER 10 OCTOBER 2017 933
REVIEW
Insects are remarkably adaptable and are expected to evolve
resistance to any control method, including transgenic plants with
combinations of protective traits as different as Bt toxins and RNAi.
Innovations such as genetically modified Bt toxins that kill pests
resistant to native Bt toxins81,82 and discovery of insecticidal proteins
from bacteria other than Bt83,84 will continue to provide new tools for
pest control. In turn, pests will adapt. The analyses of global patterns
of field-evolved resistance to transgenic crops presented here provide
empirical support for a framework to effectively manage pest resist-
ance to current and future transgenic crops.
The primary lesson from the past two decades is that abundant ref-
uges can delay pest resistance to transgenic crops. In practical terms,
transgenic crops are most durable when used in combination with other
control tactics in integrated pest management17,64,77,85,86. The sustain-
ability of transgenic crops for pest control depends largely on the will
to implement this knowledge.
Note: Any Supplementary Information and Source Data files are available in the
online version of the paper.
ACKNOWLEDGMENTS
This work was supported by USDA Biotechnology Risk Assessment Grant 2014-
33522-22214.
COMPETING FINANCIAL INTERESTS
The authors declare competing financial interests: details are available in the online
version of the p aper.
Reprints and permissions information is available online at http://www.nature.com/
reprints/index.html. Publisher’s note: Springer Nature remains neutral with regard to
jurisdictional claims in published maps and institutional affiliations.
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Supplementary Information
Surge in Insect Resistance to Transgenic Crops and Prospects for Sustainability
Bruce E Tabashnik & Yves Carrière
Department of Entomology, University of Arizona, Tucson, Arizona, USA.
Address correspondence to Bruce Tabashnik: brucet@cals.arizona.edu.
This Supplementary Information contains:
Supplementary Methods
Supplementary Notes 1-2
Supplementary Tables 1-5
Supplementary Figure 1
!
2
Supplementary Methods
Distinguishing between incomplete resistance and heterogeneous populations:
Calculating the minimum percentage of resistant individuals in a potentially
heterogeneous population when some susceptible individuals survive in a bioassay
Ideally, a diagnostic test with either a plant or diet bioassay kills 100% of susceptible individuals, but
0% or close to 0% of resistant individuals. If so, the percentage of resistant individuals is readily
calculated as the survival in the diagnostic test adjusted for control mortality on either non-Bt plants
or untreated diet.
Here we provide a formula to calculate the minimum percentage of resistant individuals in a
potentially heterogeneous population when the survival of susceptible individuals tested separately
using the same bioassay is substantially greater than zero. We also apply this formula to the data
from Minnesota of Ludwick et al.93 for western corn rootworm seedling and greenhouse bioassays
with Cry34/35Ab corn, and use the conceptual framework provided by the formula to clarify the
evidence for resistance to Cry34/35Ab in both Iowa92 and Minnesota93.
We begin with an equation for total survival (VT) (adjusted for control mortality) observed in a
bioassay of a potentially heterogeneous population:
VT = (PS X VS) + (PR X VR)
where PS and P
R are the proportion of phenotypically susceptible and resistant individuals in the
tested population, respectively, and VS and VR are the survival (adjusted for control mortality) of the
susceptible and resistant individuals, respectively. (Note: Values of VR < 1 indicate incomplete
resistance, whereas PR < 1 indicates the population includes some susceptible individuals.)
To estimate the minimum percentage of resistant individuals, we assume the survival of resistant
individuals (VR) = 1. This assumpiton maximizes the contribution of each resistant individual to total
survival (VT), which yields the minimum proportion of resistant individuals needed to achieve any
given value of VT.
Because PS + PR = 1, we can substitute 1- PR for PS. By substitution and rearrangement we get
PR = (VT - VS)/(1- VS)
which enables calculation of PR from any values of VT and VS.
From the greenhouse bioassays of Ludwick et al.93, VT = 0.63 and VS = 0.25, which yields PR = 0.51.
From their seedling bioassays, VT = 0.68 and V
T = 0.27, which yields P
R = 0.56. Thus, in the
potentially heterogeneous population tested, the results of the greenhouse and seedling bioassays
show that at least 51% and 56% of the individuals were resistant, respectively.
In the greenhouse bioassays, larval weight was about 65% lower on Cry34/35Ab corn than non-Bt
corn for both the resistant Minnesota population and a control population93. Given that >50% of the
population was resistant based on the survival data noted above, we infer these survivors had
incomplete resistance because larval weight on Cry34/35Ab was not significantly higher for this
population than for the control population.
In laboratory plant bioassays, survival of larvae from the Iowa populations with resistance to
Cry34/35Ab was 72 to 73% lower on Cry34/35Ab corn relative to non-Bt corn92. The root injury data
from the populations in Iowa show no difference between Cry34/35Ab corn and non-Bt corn92,
implying all or nearly all individuals were resistant. This high proportion of resistant individuals
coupled with the significantly lower survival on Cry34/35Ab corn than non-Bt corn also indicates
incomplete resistance.
Nature Biotechnology: doi:10.1038/nbt.3974
!
3
Supplementary Note 1
Practical resistance to Bt corn in D. v. virgifera and S. albicosta
Practical resistance of D. v. virgifera has been documented to all four coleopteran-active Bt toxins
produced by corn: Cry3Bb, mCry3Aa, eCry3.1Ab, and Cry34/35Ab (Table 1). Resistance to
Cry3Bb, which was first detected in 2009, causes strong cross-resistance to mCry3Aa and
eCry3.1Ab, but not to Cry34/35Ab28-31,85,94. Two papers report evidence of practical resistance to
Cry34/35Ab in five field populations sampled in 2013, four from central and eastern Iowa92 and
one from Minnesota93. At all five sites, growers or crop consultants complained of greater than
expected damage to Bt corn producing Cry34/35Ab alone (two sites) or pyramids of Cry34/35Ab
plus either Cry3Bb or mCry3Aa (three sites). Larval survival on Cry34/35Ab corn relative to non-
Bt corn was significantly higher for the progeny of adults collected from the five problem field sites
than for 11 unselected populations tested as controls92,93. These results show that the reduced
efficacy observed in the field was associated with genetically based, field-evolved resistance.
The field-selected resistant population from Minnesota had 63 to 68% survival on
Cry34/35Ab corn relative to non-Bt corn93, which indicates >50% of the individuals in this
population were resistant to Cry34/35Ab (Supplementary Methods). For three of the four sites in
Iowa, root injury data were obtained for non-Bt corn (control) and Bt corn producing Cry34/35Ab
either alone or in pyramids with Cry3Bb and mCry3Aa. At these three sites, the mean root injury
was not lower for the Bt corn (1.59) than the non-Bt corn (1.57), indicating that Cry34/35Ab (as
well as Cry3Bb and mCry3Aa) provided no protection against rootworm damage92. These results
suggest that all or nearly all of the individuals in these three populations were resistant to
Cry34/35Ab.
Relative to their performance on non-Bt corn, the populations with resistance to
Cry34/35Ab suffered disadvantages on Cry34/35Ab corn, including lower survival92,93. The
observed disadvantages on Cry34/35Ab corn relative to non-Bt corn imply that these populations
included some susceptible individuals, resistance in homozygous resistant individuals was
incomplete, or both (Box 1 and Supplementary Methods). The evidence points to incomplete
resistance in both the Minnesota and Iowa populations with practical resistance to Cry34/35Ab
(Supplementary Methods).
For S. albicosta (western bean cutworm), the evidence of practical resistance to Cry1F
produced by Bt corn includes laboratory bioassay data and widespread reports of reduced
efficacy in the field104-107. The LC50 of Cry1F for a population sampled from Texas in 2013 was
2000 times higher than the LC50 values for both the most susceptible field population tested in
2013 and a laboratory strain104. Overall, a statistically significant, 5.2-fold increase in the LC50 of
Cry1F occurred in data pooled from 13 bioassays of populations sampled in 2013 and 2014 from
Nebraska, New Mexico and Texas relative to 19 bioassays of populations sampled from Iowa and
Nebraska in 2003 and 2004 (ref. 104).
Based on extensive field trials in Iowa, Nebraska, Colorado, Texas and New Mexico from
2002 to 2006, the mean percentage of ears damaged by S. albicosta was reduced by about 85%
in corn producing Cry1F relative to non-Bt corn (P < 0.001) (ref. 107). In July 2016, five extension
entomologists from Nebraska concluded, “When first introduced to the market, Cry1F provided
approximately 80% control of western bean cutworm. However, recent research has shown that
its effectiveness has decreased in some areas, such as parts of southwest and central Nebraska,
in the last 10 years”106. Moreover, extension entomologists from six other states wrote an open
letter to the seed industry in October 2016 declaring, “Wherever Cry1F is challenged by WBC
[western bean cutworm], it fails to provide observable benefit to producers.”105.
Nature Biotechnology: doi:10.1038/nbt.3974
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4
Supplementary Note 2
Early warning of resistance
Zhang et al.142 used the term ‘early warning’ of resistance to describe the statistically significant
increase in the percentage of individuals with resistance to Cry1Ac in H. armigera from northern
China. Their 2010 survey showed that mean survival at a diagnostic concentration of Cry1Ac was
significantly higher for 13 field populations from northern China (1.3%), where exposure to Bt
cotton producing Cry1Ac was extensive, relative to two field populations from northwestern China
(0%) where exposure to Bt cotton was limited. Subsequent monitoring in northern China showed
further increases in the mean survival at the diagnostic concentration, which by 2013 had
increased to 5.5% (range: 0.3 to 11.3%)73. As in the other cases of early warning of resistance
described below, reduced efficacy of Bt plants in the field has not been reported.
For D. saccharalis in Louisiana, the frequency of alleles conferring resistance to Cry1Ab
in Bt corn increased 9-fold in 2009 relative to 2004 to 2008 (ref. 129). Based on the 2009
monitoring data and related work showing non-recessive inheritance of D. saccharalis resistance
to Cry1Ab129,130, the maximum percentage of resistant individuals was estimated as 2.4% (ref.
24). During 2009 field trials in Louisiana, Huang et al.143 observed a substantial number of live
larvae and injury to multi-toxin Bt corn. However, during 2010 and 2011, abundance of this pest in
the region was so low that not enough individuals were sampled for screening129.
For O. furnacalis in the Philippines, where Bt corn producing Cry1Ab was commercialized
in 2003, the maximum survival at a diagnostic concentration of Cry1Ab increased 14-fold from
0.38% in 2007-2008 to 5.5% in 2009 (ref. 131). Also, field populations with >1% survival at the
diagnostic concentration increased from 0% (0 of 11) in 2007-2008 to 62% (5 of 8) in 2009
(Fisher’s exact test, P = 0.0048) (ref. 131). During 2010, efficacy in 198 fields based on mean
percentage of damaged plants for Bt corn relative to non-Bt corn was 49% (including corn with
and without herbicide tolerance)144. Meanwhile, cultivation of Bt corn in the Philippines increased
from 400,000 ha in 2009 to 760,000 ha in 2014, with over 80% adoption in some areas132,145.
References
142. Zhang, H. et al. Early warning of cotton bollworm resistance associated with intensive
planting of Bt cotton in China. PLoS ONE 6, e22874. (2011).
143. Huang, F., Ghimire, M.N., Leonard, B.R., Zhu, Y.-C. & Head, G.P. Susceptibility of field
populations of sugarcane borer from non-Bt and Bt maize plants to five individual Cry
toxins. Insect Sci. 19, 570-578 (2012).
144. Afidchao, M. M., Musters, C.J.M., Wossink, A. & Balderama, O.F. Analysing the farm level
economic impact of GM corn in the Philippines. NJAS Wageningen J. Life Sci. 70-71,
113-121 (2014).
145. Adlemita, R.R., Villena, M.M.C.A. & James, C. Biotech Corn in the Philippines: A Country
Profile. Los Baños, Laguna: ISAAA and Southeast Asian Regional Center for Graduate
Study and Research in Agriculture - Biotechnology Information Center (2015).
Nature Biotechnology: doi:10.1038/nbt.3974
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Supplementary Table 1. The number of generations per year pests fed on Bt crops for the
16 cases of practical resistance.
Gens. per yeara
Insect
Crop
Toxin
Country
Reference
B. fusca
Corn
Cry1Ab
S. Africa
2
146
D. saccharalis
Corn
Cry1A.105
Argentina
3.5
147
D. v. virgifera
Corn
Cry3Bb
USA
1
28
D. v. virgifera
Corn
Cry34/35Ab
USA
1
28
D. v. virgifera
Corn
mCry3A
USA
1b
28
D. v. virgifera
Corn
eCry3.1Ab
USA
1b
28
H. zea
Corn
Cry1Ab
USA
2
148
H. zea
Corn
Cry1A.105
USA
2b
148
H. zea
Cotton
Cry1Ac
USA
3
141
H. zea
Cotton
Cry2Ab
USA
3b
141
P. gossypiella
Cotton
Cry1Ac
India
7
149
P. gossypiella
Cotton
Cry2Ab
India
7
149
S. albicosta
Corn
Cry1Fa
USA
1
150
S. frugiperda
Corn
Cry1Ab
Brazil
11b
38
S. frugiperda
Corn
Cry1F
Brazil
11
38
S. frugiperda
Corn
Cry1F
USA
10
47
aGenerations per year that the pest fed on the Bt crop in the region monitored. When a range
was reported, we used the midpoint of the range (i.e., 3.5 represents the range of 3-4 generations
reported).
b!Cross-resistance suspected or known as a factor contributing to resistance.
References
146. Van Rensburg, J.B.J. et al. Geographical variation in the seasonal moth flight activity of
the maize stalk borer, Busseola fusca (Fuller) in South Africa. S. African J. Plant Soil 2,
123-126 (1985).
147. Moré, M. et al. Influence of corn, Zea mays, phenological stages in Diatraea saccharalis
F. (Lep. Crambidae) oviposition. J. Appl. Entomol. 127, 512-515 (2003).
148. Storer, N.P. et al. Spatial processes in the evolution of resistance in Helicoverpa zea
(Lepidoptera: Noctuidae) to Bt transgenic corn and cotton in a mixed agroecosystem: a
biology-rich stochastic simulation model. J. Econ. Entomol. 96,156-172 (2003).
149. Mohan, K.S. An area-wide approach to pink bollworm management on Bt cotton in India
a dire necessity with community participation. Curr. Sci. 112, 1988-1989 (2017).
150. Hanson et al. Degree-day prediction models for the flight phenology of western bean
cutworm (Lepidoptera: Noctuidae) assessed with the concordance correlation coefficient.
J. Econ. Entomol. 108,1728-1738 (2015).
Nature Biotechnology: doi:10.1038/nbt.3974
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6
Supplementary Table 2. Multiple regression testing the effects on years to evolve
practical resistance of two factors: cross-resistance and the number of
generations per year that pests fed on Bt crops.
_______________________________________
Source Estimate (SE) P value
_______________________________________
Intercept 5.76 (0.79) <0.0001
Cross-resistance -1.86 (0.55) 0.005
Generations per year -0.31 (0.14) 0.049
_______________________________________
The fit of the model including both main effects (cross-resistance and generations per
year) is significant (P = 0.007, R2 = 0.53). None of the interactions between factors is
significant. The multiple regression analysis was performed in JMP 12, SAS Institute Inc.
2015. Cary, NC.
Nature Biotechnology: doi:10.1038/nbt.3974
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Supplementary Table 3. The number of generations per year pests fed on Bt crops
for the 17 cases of sustained susceptibility.
Gens.
Insect
Crop
Toxin
Country
per yr. a
Referencesd
C. includens
Soy
Cry1Ac
Brazil
3
151
D. grandiosella
Corn
Cry1Ab
USA
3.5
152
H. armigera
Cotton
Cry1Ac
Australia
4
25
H. armigera
Cotton
Cry2Ab
Australia
4
25
H. punctigera
Cotton
Cry1Ac
Australia
3
153
H. punctigera
Cotton
Cry2Ab
Australia
3
153
H. virescens
Cotton
Cry1Ac
Mexico
6
154
H. virescens
Cotton
Cry1Ac
USA
4
25
H. virescens
Cotton
Cry2Ab
USA
4
25
O. nubilalis
Corn
Cry1Ab
Spain
2.5
155
O. nubilalis
Corn
Cry1Ab
USA
3
25
O. nubilalis
Corn
Cry1Fa
USA
3
25
P. gossypiella
Cotton
Cry1Ac
China
3
125
P. gossypiella
Cotton
Cry1Ac
USA
5
25
P. gossypiella
Cotton
Cry2Ab
USA
5
25
S. frugiperda
Corn
Vip3Aa
Brazil
11
38
S. nonagroides
Corn
Cry1Ab
Spain
3
25
______________________________________________________________________________________________________________________
aGenerations per year that the pest fed on the Bt crop in the region monitored. When a range
was reported, we used the midpoint of the range (i.e., 3.5 represents the range of 3-4
generations reported).
References
151. Palma, J. et al. Molecular variability and genetic structure of Chrysodeixis includens
(Lepidoptera: Noctuidae), an important soybean defoliator in Brazil. an J. Plant SoilPLoS
ONE 10(3): e0121260 (2015).
152. Baldwin, J.L. et al. Corn borer pests in Louisiana corn. LSUAg. Pub. 2947.
http://www.lsuagcenter.com/NR/rdonlyres/A626E99C-CF02-480C-97D1-
1F922408A2CC/23629/pub2947cornborerLOWRES.pdf
153. Baker, G.H. & Tann, C.R. Long-term changes in the numbers of Helicoverpa punctigera
(Lepidoptera: Noctuidae) in a cotton production landscape in northern New South Wales,
Australia. Bull. Entomol. Res. 107, 174-187 (2016).
154. Molina-Ochoa, J. et al. Current status of Helicoverpa zea and Heliothis virescens within a
changing landscape in the southern United States and Mexico. Southwestern Entomol.
35, 347-354 (2010).
155. Farinós, G.P. et al. Resistance monitoring of field populations of the corn borers Sesamia
nonagrioides and Ostrinia nubilalis after 5 years of Bt maize cultivation in Spain. Entomol.
Exp. Appl. 110, 23-30 (2004).
Nature Biotechnology: doi:10.1038/nbt.3974
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Supplementary Table 4. Planting of transgenic corn and cotton producing Vip3Aa.
Country
Crop
Other Bt toxinsa
Year
%b
References
Australiac
Cotton
Cry1Ac, Cry2Ab
2015
7.6
156-158
Australiac
Cotton
Cry1Ac, Cry2Ab
2016
92
156-158
Brazild
Corn
Cry1Ab
2015
<5
10
USA
Corn
Cry1Ab, Cry1F
2011
1.1
159
USA
Corn
Cry1Ab, Cry1F
2013
3.5
160
USA
Cotton
Cry1Ac, Cry1Fa
2014
<0.01
161
USA
Cotton
Cry1Ac, Cry1Fa
2015
0.48
162
USA
Cotton
Cry1Ac, Cry1Fa
2016
0.65
163
aFor the years indicated above, the lepidopteran-active Bt toxins listed were produced in
combination with Vip3Aa in all of the Vip3Aa cotton and in some types of Vip3Aa corn. Although
some corn hybrids produce coleopteran-active Bt toxins in combination with Vip3Aa, only the
lepidopteran-active Bt toxins are listed.
bPercentage of the total area planted with the crop listed that produced Vip3Aa. For example, in
Australia in 2015, cotton producing Vip3Aa accounted for 7.6% of all cotton planted.
cSown in the year listed (2015 or 2016), full seasons are 2015-16 and 2016-17, respectively.
dBernardi et al.10 indicates that Vip3Aa corn was approved for commercial planting in 2009 and
"was planted in less than 5% of the total corn-growing area in Brazil (Syngenta information)."
References
156. Queensland Country Life. Aust cotton growers rush to use Bollgard 3.
http://www.queenslandcountrylife.com.au/story/4245922/bollgard-3-dominates-aust-
cotton-crop/ (2016).
157. Cotton Australia. Statistics. http://cottonaustralia.com.au/cotton-library/statistics (2017)
158. Monsanto Australia. Anthony May email April 9, 2017.
159. Christensen, P. Chinese approval of Syngenta Agrisure Viptera. Seed in Context Blog.
http://www.intlcorn.com/seedsiteblog/?p=268 (2012).
160. Christensen, P. Viptera could have been approved for importation into China, but was
not. Seed in Context Blog. http://www.intlcorn.com/seedsiteblog/?p=1891 (2014).
161. U.S. Department of Agriculture, Agricultural Marketing Service. Cotton varieties planted
2014. crop.https://search.ams.usda.gov/mndms/2014/09/CN20140912AVAR.PDF
162. U.S. Department of Agriculture, Agricultural Marketing Service. Cotton varieties planted
2015. crophttps://search.ams.usda.gov/mndms/2015/09/CN20150915AVAR.PDF
163. U.S. Department of Agriculture, Agricultural Marketing Service. Cotton varieties planted
2016. crop. https://www.ams.usda.gov/mnreports/cnavar.pdf
Nature Biotechnology: doi:10.1038/nbt.3974
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9
Supplementary Table 5. Cross-resistance between Vip3A and Cry1 toxins in related strains of five species of
noctuid lepidopteran pests.
Related
Tested for
Unselected
selected
Selected
cross-
Insect
strain
strain
with
resistance to
Metrica
RRb
CRRc
log(CRR)
Ref.
H. armigera
96S
Cry1Ac-R
Cry1Ac
Vip3Aa
LC50
2970
1.7
0.23
164
H. armigera
SCD-r1
SCD
Cry1Ac
Vip3Aa
LC50
440
1.0
-0.01
13
H. virescens
YDK
YHD2
Cry1Ac
Vip3Aa
LC50
20,000
1.2
0.068
57
H. virescens
Vip-Unsel
Vip-Sel G15
Vip3Aa
Cry1Ab
LC50
2000
3.2
0.51
55
H. virescens
Vip-Unsel
Vip-Sel G18
Vip3Aa
Cry1Ab
LC50
2000
6.7
0.83
55
H. virescens
Vip-Unsel
Vip-Sel G15
Vip3Aa
Cry1Ac
LC50
2000
7.1
0.85
55
H. virescens
Vip-Unsel
Vip-Sel G18
Vip3Aa
Cry1Ac
LC50
2000
1.0
0.00
55
H. zea
SC
AR
Cry1Ac
Vip3Aa
LC50
100
0.94
-0.027
165
H. zea
GA
GA-R
Cry1Ac
Vip3Aa
EC50
57
1.6
0.20
36
S. frugiperda
SS
RR
Cry1F
Vip3Aa
LC50
>62
1.5
0.18
52
S. frugiperda
SS
RR
Cry1F
Vip3Aa
IC50
930
0.6
-0.26
52
T. ni d
SS
RR
Cry1Ac
Vip3Aa
IC50
2054
2.1
0.33
166
T. ni
SS
RR
Cry1Ac
Vip3AaAc
IC50
2054
1.8
0.26
166
T. ni
SS
RR
Cry1Ac
Vip3Ac
IC50
2054
1.02
0.0078
166
T. ni
SS
RR
Cry1Ac
Vip3AcAa
IC50
2054
3.2
0.51
166
aConcentrations causing 50% response in tested insects based on mortality (LC50), efficacy (EC50), or growth inhibition
(IC50).
bResistance ratio (RR) is the LC50 (or EC50 or IC50) of a toxin for the resistant strain that was selected with that toxin
divided by the LC50 (or EC50 or IC50) of the same toxin for a related, unselected strain.
c Cross-resistance ratio (CRR) is the LC50 (or EC50 or IC50) of a toxin not used for selection (e.g., Vip3Aa) for a strain
selected with another toxin (e.g., Cry1Ac), divided by the LC50 (or EC50 or IC50) of the toxin not used for selection for a
related, unselected control strain. The expected value of CRR is 1 if cross-resistance is absent and >1 if cross-resistance
is present.
dTrichoplusia ni
References
164. Zhang, Q., Chen, L.-Z., Lu, Q., Zhang, Y., Liang, G.-M. Toxicity and binding analyses of Bacillus thuringiensis toxin
Vip3A in Cry1Ac-resistant and -susceptible strains of Helicoverpa armigera (Hü bner). J. Integr. Agric. 14, 347354
(2015).
165. Anilkumar, K. J. et al. Production and characterization of Bacillus thuringiensis Cry1Ac-resistant cotton bollworm
Helicoverpa zea (Boddie). Appl. Environ. Microbiol. 74, 462-469 (2008).
166. Fang, J. et al. Characterization of chimeric Bacillus thuringiensis Vip3 toxins. Appl. Env. Microbiol. 73, 956-961
(2007).
Nature Biotechnology: doi:10.1038/nbt.3974
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10
a
b
Supplementary Figure 1. Negative association between time to practical resistance and the
number of generations per year (GPY) that pests fed on Bt crops. (a) With all 16 cases, this
association is not significant (linear regression: R2 = 0.12, df = 14, P = 0.19). Squares represent
the five cases where cross-resistance is suspected or known to have shortened the time to
resistance. (b) The negative association is significant when considering only the 11 cases where
cross-resistance is not involved (y = -0.42x + 8.1, R2 = 0.46, df = 9, P = 0.022). Multiple
regression for all 16 cases shows a significant negative relationship between time to practical
resistance and GPY (P = 0.049) after accounting for the significant effects of cross-resistance (P
= 0.005) (Supplementary Table 2).
0
2
4
6
8
10
12
0 2 4 6 8 10 12
Years to resistance
Generations per year
0
2
4
6
8
10
12
0 2 4 6 8 10 12
Years to resistance
Generations per year
Nature Biotechnology: doi:10.1038/nbt.3974
... 5 However, when pests evolve resistance to Bt toxins, the advantages of these Bt crops are reduced. [6][7][8][9][10] Practical resistance is field-evolved resistance that decreases the efficacy of a Bt crop and has practical consequences for pest control. 8 Scientists have documented practical resistance to Bt crystalline (Cry) toxins in some populations of at least nine species of major pests targeted by Bt crops. ...
... [8][9][10] Thus, farmers have begun to increasingly adopt multi-toxin transgenic crops that produce the Bt vegetative insecticidal protein Vip3Aa together with Cry proteins. 7,11 Although we are not aware of documented cases of practical resistance to Vip3Aa, an early warning of field-evolved resistance to this toxin has been reported for Helicoverpa zea in the United States. 11 Vip3 and Cry proteins share little amino acid sequence similarity. ...
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Background: Transgenic crops that make insecticidal proteins from Bacillus thuringiensis (Bt) have revolutionized management of some pests. However, evolution of resistance to Bt toxins by pests diminishes the efficacy of Bt crops. Resistance to crystal (Cry) Bt toxins has spurred adoption of crops genetically engineered to produce the Bt vegetative insecticidal protein Vip3Aa. Here we used lab diet bioassays to evaluate responses to Vip3Aa by pink bollworm (Pectinophora gossypiella), one of the world's most damaging pests of cotton. Results: Against pink bollworm larvae susceptible to Cry toxins, Vip3Aa was less potent than Cry1Ac or Cry2Ab. Conversely, Vip3Aa was more potent than Cry1Ac or Cry2Ab against lab strains highly resistant to those Cry toxins. Five Cry-susceptible field populations were less susceptible to Vip3Aa than a Cry-susceptible lab strain (APHIS-S). Relative to APHIS-S, significant resistance to Vip3Aa did not occur in strains selected in the lab for >700-fold resistance to Cry1Ac or both Cry1Ac and Cry2Ab. Conclusions: Resistance to Cry1Ac and Cry2Ab did not cause strong cross-resistance to Vip3Aa in pink bollworm, which is consistent with predictions based on the lack of shared midgut receptors between these toxins and previous results from other lepidopterans. Comparison of the Bt toxin concentration in plants relative to the median lethal concentration (LC50 ) from bioassays may be useful for estimating efficacy. The moderate potency of Vip3Aa against Cry1Ac- and Cry2Ab-resistant and susceptible pink bollworm larvae suggests that Bt cotton producing this toxin together with novel Cry toxins might be useful as one component of integrated pest management. This article is protected by copyright. All rights reserved.
... Only a handful of Bt crops (expressing the Bacillus thuringiensis Cry toxins that are insecticidal per se) such as Bt cotton and Bt corn got regulatory approval for field trial in USA, China, India, Brazil and EU. Further, a number of insect pests are developing Bt resistance worldwide (Tabashnik and Carrière, 2017). A number of RNAi GMO crops have been marketed harboring the novel traits such as nicotinefree tobacco, decaffeinated coffee, nutrient fortified and hypoallergenic crops, Banana Bract Mosaic Virus (BBrMV) resistant crops etc. RNAi generated healthier oil production by suppressing the enzyme that converts oleic acid into a different fatty acid (Plenish ® high oleic acid soybean from DuPont Pioneer). ...
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... In the existing environmental monitoring process, some agricultural concerns associated with commercial cultivation of GM crops have been reported [11]. In Bt crops for example, the target insect pests may develop Bt toxin resistance over time, making them more difficult to be controled in the future [12,13]. The fear is that the transgene flow from Bt crops to surrounding plant diversity, and the potential development of 'super weed' is one of the examples [14]. ...
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In recent years, transgenic technology has developed rapidly, but the risk of the environmental release of transgenic organisms is still a key issue. Research on the impact on biodiversity is an effective way to objectively evaluate the risk. By taking transgenic maize HGK60 with insect-resistant gene Cry1Ah and common maize Zheng 58 as control, a 2-year experiment of arthropod community biodiversity in fields of them were studied using three methods.in 2019 and 2020. The results showed that a total of 124 species and 38537 individuals were observed from the experiment, belonging to 11 orders and 40 families. There was no significant difference in the individual number and species number of herbivorous, predatory and parasitic groups in the two kinds of maize in two years. Only the individual number of HGK60 was significantly higher than that of common maize Zheng 58 at heading stage in 2019. And the percentages of individual number and species number in different groups were basically the same in the two kinds of maize at each stage in two years. Analyses of Richness index, Shannon-Wiener diversity index, Dominance index and Evenness index showed no significant difference between the two kinds of maize in two years. The similarity coefficient of the arthropod community suggested that the arthropod community composition of HGK60 was similar to that of common maize Zheng 58. Furthermore, HGK60 had no significant effect on the relative stability of the arthropod community. These results indicated that despite the presence of a relatively minor difference in arthropod community between the two kinds of maize, the planting of HGK60 had little effect on arthropod community biodiversity. The results provided some data and support for the further studies of environmental risk of transgenic crops.
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Enhancement of plant defense by exogenous elicitors is a promising tool for integrated pest management strategy. In the present study, cotton plants were treated with different concentrations (0, 0.01, 0.1, and 1.0 mM) of the natural plant defense elicitor, jasmonic acid (JA), and defense-related indicators in the plants were then determined. The cotton bollworm larvae were fed with JA-treated cotton leaves and larvae performances were discussed in terms of larvae relative growth rate (RGR), larval duration, pupal mass, humoral immunity, and activities of a target enzyme, three detoxification enzymes and two metabolic enzymes. Research results showed that JA treatment increased the contents of gossypol and H2O2, and decreased that of the total soluble carbohydrates, and 0.1 mM JA was more powerful in the induction of defense-related parameters. As a consequence, cotton bollworm larvae reared on JA-treated cotton leaves showed slower RGR, prolonged larvae duration, and decreased pupal mass. In addition, when larvae were fed with JA-treated cotton leaves, activities of phenoloxidae (an indicator of humoral immunity) and acetylcholinesterase (AchE, a target enzyme), alkaline phosphatases (ALP), acidic phosphatase (ACP), and three detoxification enzymes, carboxylesterase (CarE), glutathione S-transferase (GST), and cytochrome P450 (P450), were all reduced compared to the control. Taken together, the results suggest that JA can be an alternative agent for pest management by delaying insect growth and inhibiting immune defense and detoxification capacity of the cotton bollworm, which may reduce the use of synthetic pesticides.