An ABC Transporter Mutation Is Correlated with Insect Resistance to Bacillus thuringiensis Cry1Ac
Assessment of Bt protein level in soil and effect of ... · Summary The effect of transgenic...
Transcript of Assessment of Bt protein level in soil and effect of ... · Summary The effect of transgenic...
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Assessment of Bt protein level in soil and effect of transgenic
Bt brinjal expressing cry1 Ac gene on soil microflora,
nematodes, collembola and earthworms
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Effect of transgenic brinjal expressing Bacillus thuringiensis cry1 Ac
gene on soil microflora, nematodes, collembola and earthworms.
Objective
The objectives of this study were:
1) to evaluate the effect of Bt brinjal expressing cry1Ac gene on soil microflora,
nematodes, collembola and earthworms, and
2) to determine the Cry1Ac protein levels in the Bt brinjal grown soil.
Study conducted by
Maharashtra Hybrid Seeds Co. Ltd., Jalna (Maharashtra).
Summary
The effect of transgenic brinjal expressing the cry1Ac gene from Bacillus
thuringiensis (Bt) on the soil microflora, nematodes, collembola, and earthworms has
been studied. Soil and insect samples were collected from the Bt brinjal experimental
trials conducted at Jalandhar, Mirzapur, Ahmad nagar, Bhopal, Dharmapuri,
Dharwad, and Karnool during 2004-05. Microflora populations were measured by
dilution plating method. There were no consistent significant differences between Bt
and non-Bt treatments in the numbers of total culturable bacteria and fungi.
Nematodes were extracted using Cobb’s decanting and sieving method. Collembola
populations were measured using pitfall traps There were no significant differences in
populations of nematodes and collembola between Bt and non-Bt treatments.
Earthworms were observed in all the seven locations studied. The Cry1Ac protein,
determined by insect bioassays with Helicoverpa armigera, was not detected in any
soil samples. These findings indicated that transgenic brinjal expressing the cry1Ac
gene does not have any adverse effect on the microflora, nematodes, collembola and
earthworms in soil.
Introduction
The incorporation into plants of genes from Bacillus thuringiensis (Bt) that code for
the production of insecticidal toxins reduces many problems associated with the use
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of chemical pesticides, as the toxins are produced continuously within these plants
and exhibit relatively high specificity. However, there is concern that genetically
engineered crops may pose ecological risks to natural and agricultural ecosystems
(Conway, 2000; Hails, 2000; Stotzky, 2000). While efforts have been made to
examine the environmental impacts of genetically modified plants “ above ground”,
comparatively little research has been directed at impacts of genetically modified
crops on soil organisms and processes, perhaps because of the greater difficulties
involved in the study of soil invertebrates and soil microorganisms. Concerns about
impacts of GM crops on soil biota have been raised, in part because of the chemical
and biological properties of soil (McGregor and Turner 2000). Soil materials have
large sorptive capacities for biological molecules, including insecticidal bacterial
proteins and DNA. Laboratory studies (Crecchio and Stotzky, 1998, 2001; Saxena and
Stotzky 2001; Saxena et al. 2002; Tapp et al. 1994; Tapp and Stotzky, 1995, 1998)
have shown that insecticidal Cry proteins from B. thuringiensis are readily adsorbed
at equilibrium and bound to clay minerals and humic acids, and persist in soil for up
to 350 days, the longest time studied. Zwahlen et al. (2003a) investigated the
degradation of Bt toxin in transgenic corn leaves under field conditions and reported
the persistence of the Cry1Ab protein up to 240 days. Consequently, the issue of the
impact of the Bt proteins released to soil from the roots and biomass of Bt crops on
soil organisms including soil microflora, collembola and earthworms is an important
one. In this study we evaluated the impact of this Bt brinjal on soil microflora,
nematodes, collembola, and earthworms. Also, we tried to quantify the Cry1Ac
protein levels in the Bt brinjal grown soils.
Materials and methods
Locations for sample collection
Soil and collembola samples were collected from the Bt brinjal experimental trials
conducted at Jalandhar, Mirzapur, Ahmad nagar, Bhopal, Dharmapuri, Dharwad and
Karnool during 2004-05. The field trial the following treatments:
a. Bt treatment : MHB 80 Bt/ MHB 4 Bt/ MHB 9 Bt or MHB 99 Bt
b. Near-isogenic non-Bt treatment: MHB 80 Bt/ MHB 4 Bt/ MHB 9 Bt or MHB 99,
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Plant
Zone for non-rhizosphere
soil sampling
Zone for rhizosphere
soil sampling
1
34
5
2
c. Non-Bt local check : Pusa purple round/ Gondegaon/ Arka Shirish/ Manjari kota
or Green +white local, and
d. Non-Bt commercial check: Navakiran/ Ajay/ Extra green long/ Manju or Harit.
Each treatment had four replications. Samplings were made in three replications
for each treatment.
Soil sampling
Pre-harvest soil samples were collected at 0, 30, 90, and 150 days after
transplantation. For sampling purpose, the area around the plants selected was divided
into root/rhizosphere and non-rhizosphere zones as shown in the following figure.
The root zone for rhizosphere sample collection extended to 20cm area around the
plant, and the non-rhizosphere zone represented the 20-40cm area around the plant.
To get one sample, five core samples (each from a 15-cm deep and 7.5-cm diameter
area) were taken around the plant and mixed thoroughly. From this mixture, 100 g of
soil was drawn as a representative sample. During the pre-harvest stage, a total of 3
rhizosphere and 3 non-rhizosphere samples per treatment were collected at every
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sampling time-point. Samples were transported to the Biotechnology R&D Center,
MAHYCO, Dawalvadi (Maharashtra) for laboratory analysis.
Determination of total bacterial and fungal population
To determine the numbers of total culturable bacteria and fungi, 10 g of rhizosphere
or non-rhizosphere soil was added to 90 ml of sterile distilled water and shaken for 20
min at 250 rpm on a Gyrorotary shaker. Soil suspensions were diluted further with
sterile distilled water and 100µl of 10-3
dilution was spread-plated onto selective
medium. Luria Bertani (LB) containing 100 mg/l cycloheximide (Sigma Chemical,
St.Louis, MO) and 1% soil extract, and Rose bengal streptomycin agar (Saxena and
Stotzky 2001) containing 50 mg/l streptomycin sulphate (Sigma Chemical, St.Louis,
MO) were used to enumerate bacteria and fungi, respectively. Total culturable
bacteria and fungi were counted following incubation at 28οC for 3 and 7 days,
respectively. Differences in numbers of culturable bacteria and fungi among
treatments were analyzed with Analysis of Variance (ANOVA) (SAS Institute, 1989).
Analysis of soil invertebrates
Populations of earthworms and collembola were enumerated in all the Bt and non-Bt
treatments. Earthworms were sampled at five time-points during the pre-harvest
season, i.e. 0, 30, 90, 120 and 150 days after transplantation. For measurement of
earthworm populations, holes (30-cm diameter and 90-cm deep) were dug in soil
along two rows of plants. The soil from each hole was spread on a plastic sheet, and
earthworm counts were made in the field.
Collembola populations inhabiting the soil surface were measured using pitfall traps.
The pitfall traps, placed along two rows of plants, consisted of a 300 ml plastic jar
with an inserted funnel made by cutting the bottom out of a 100 ml disposable plastic-
drinking cup. A solution containing 10% formalin and 1% copper sulphate was used
to trap the collembola. Each trap was inserted in the soil so that the lip of the cup was
level with the soil surface. Three traps were used per treatment. The traps were
removed 3 days after installation, capped, and brought to the laboratory where the
contents were poured into a Petri dish and collembola were counted.
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Nematodes were extracted using Cobb’s decanting and sieving method (Ayoub,
1980), from the soil samples collected at 0, 30, 60, 90, 120 and 150 days after
transplantation. Soil samples for nematode analysis was taken in the root zone. Each
sample was collected by initially taking five core samples (each from a 20 cm deep
and 10 cm diameter) around a plant and then mixing them thoroughly. All soil
samples were stored at 15ο
C to minimize the changes in nematode populations.
Nematodes were extracted within 7 days after sampling. Numbers of nematodes in
each treatment were counted in 250 g soil. An analysis of variance (ANOVA) was
carried out using the SAS statistical software (SAS Institute 1989).
Level of Bt protein in soil:
The level of Bt protein in soil samples was determined by insect bioassays with
Helicoverpa armigera.
Bioassays with soil samples: To assay for Cry1Ac protein, soil samples were
incorporated into the artificial diet and then presented to H. armigera neonates. Five
grams soil from each sample was thoroughly slurried with 20 ml of water in a 100 ml
centrifuge tube on a vortex mixer, of which 2.8 ml was mixed with artificial diet to
bring it up to a total volume of 14 ml. The soil-diet mixture of each sample was
added to 16 wells in the 128-well tray. After the soil-diet mixture was cooled and
solidified, one neonate of H. armigera was introduced into each well.
Standard mortality bioassays: To serve as a reference standard, standard mortality
bioassay was done, that involved exposure of neonate larvae to various concentrations
of diet incorporated Cry1Ac protein that caused 0-100% mortality and also to
calculate the percent surviving second instar larvae. The stock solution of Cry1Ac
was made in 0.2% agar solution and dilutions were made in deionized water. The
source of Cry1Ac protein used in the bioassays was the commercial formulation,
MVP II ® (Mycogen Corp., USA), which contained 19.7% (by weight) Cry1Ac.
Various dilutions of Cry1Ac were later mixed in the H. armigera diet for the
bioassays. There were 32 larvae per replication with a total of four replications for
each Cry1Ac concentration. The bioassays were done in 128-well trays with each
well consisting of 750 µL diet with Cry1Ac.
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Bioassay trays were kept in dark at a temperature of 26 ± 1°C and 55-65% RH.
Mortality and instar stage of surviving larvae were recorded on seventh day. Larvae
that did not move when disturbed were considered to be dead.
Results
Microflora analysis
Total numbers of culturable bacterial and fungal colonies were counted, and analysis
of variance was done to determine the variations between Bt and non-Bt soils. No
consistent differences were observed in the culturable bacterial populations (Figures
1-7) between Bt, and non-Bt soil samples. The types of colonies, which grew from Bt
and non-Bt treatments, were similar.
The total fungal populations for the soil samples collected at pre- harvest stages are
presented in Figures 8-14. The differences in the total culturable fungal population
observed among the Bt and non-Bt treatments were not consistent. Species of
Aspergillus, Penicillium, and Rhizopus were the dominant fungi that were found from
these soil samples. Besides these, Trichoderma, Fusarium, Gliocladium,
Cephalosporium, Myrothecium, Cladosporium, Alternaria, Verticillium and
Rhizoctonia were some of the other fungi also seen in all six treatments.
Earthworms
Earthworm populations were measured at 0, 30, 60, 90, 120, and 150 days after
transplantation (DAT). The results are presented in Tables 1-3. At Jalandhar location
earthworms were observed only at 120 DAT. No significant (p>0.05) difference in the
earthworm populations was observed at other locations during the pre-harvest
sampling period. (Tables 1-3).
Enumeration of whole nematode communities, extracted from soil samples collected
from the Bt and non-Bt experimental plots, are presented in Figures 15-21. The
nematode population (Mean ± SE) estimated during the pre-sowing (0 DAT)
sampling period was 121 ± 8.9, 94.7 ± 3.03, 140 ± 6.2, 218 ± 6.7, 94.8 ± 12.3, 371
±13.3 and 243± 9.1 for Jalandhar, Mirzapur, Ahmad nagar, Bhopal, Dharmapuri,
Dharwad and Karnool, respectively. No significant difference in the total nematode
population was observed between Bt and non-Bt treatments irrespective of the
locations and sampling-timepoint.
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Collembola
Collembola were observed in all the seven locations studied. The mean (± SE)
number of collembola observed during the pre-transplantation time point at Jalandhar,
Mirzapur, Ahmad nagar, Bhopal, Dharmapuri, Dharward, and Karnool was
7.75±0.75, 4.75±0.43, 15.6±0.93, 19.6±0.74, 93.5±3.5, 26.4±1.48, and 5.42±0.47,
respectively. No significant (p > 0.05) differences in the number of collembola were
observed among Bt and non-Bt treatments irrespective of the locations during
different sampling time-points (Figures 22- 28).
Level of Bt protein in soil:
The level of Cry1Ac protein present in the soil samples from Bt brinjal field plots was
determined by insect bioassays with H. armigera. Results of the standard mortality
bioassay showed that 100% mortality occurs at 1.0 µg /ml of protein in the diet and
the lowest concentration (0.001 µg /ml) produced approximately 19% mortality
(Figure 29). Also, 46.50% of larvae were third instars at the lowest protein
concentration in the diet (Figure 30). Seventy nine percent of the larvae in the control
were third instars after six days of incubation and no mortality was observed in the
control treatment.
The results of insect bioassays with soil sample-diet mixture were presented in figures
31 to 37 for soil samples collected from different locations. Soil samples from
different locations when mixed individually with artificial diet produced no larval
mortality after six days incubation over all the sampling time points (Figures 31 – 37).
Also, the percent third instars were either equal to or higher in Bt soil samples than in
non-Bt, Local check and/or commercial check soil samples in all locations. The
average percent surviving third instar larvae were between 78.69-86.63% and 76.25-
88.0% when insect bioassays were done with pre-sown soil samples and control diet,
respectively.
The average percent surviving third instar larvae over all sampling time points was
around 87.5 % in Bt rhizosphere samples and 88.0% in Bt non-rhizosphere samples of
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North Zone (Jalandhar and Mirzapur), respectively (Figures 31 A, B and 32 A, B).
These values were comparable to 89.0% and 88.0 % in respective control samples
with no soil. The average percent surviving third instar larvae over different sampling
time points was around 81.5% in Bt rhizosphere and Bt non-rhizosphere soil samples
collected from Ahmadnagar, which was equal to the larvae obtained from control
(81.25%) (Figures 33 A and B). Similar trend was observed in the insect bioassays
containing Bt rhizosphere and Bt non-rhizosphere soil samples from Bhopal and
control sample, where there were 88.5% surviving third instars (Figures 34 A and B).
In the insect bioassays with soil samples collected from Dharmapuri, the average
percent third instar larvae was higher when Bt rhizosphere (86.0%) and Bt non-
rhizosphere (85.4%) soil samples were taken than when non-Bt soil samples and diet
control (84.0%) were taken (Figures 35 A and B). The percent third instars were 85-
87% in Bt rhizosphere and Bt non-rhizosphere samples from Dharwad and Kurnool as
compared to 87.0% when control (diet) was taken (Figures 36 A and B and 37 A and
B).
Discussion
An important aspect of the risk assessment of pesticidal transgenic plants encoding Bt
insecticidal genes is their impact on the soil ecosystem. We evaluated this by
measuring the populations of total culturable bacteria and fungi, numbers of
earthworms and collembola, and the level of Cry1Ac protein in soil on which Bt, and
non-Bt brinjal had been grown. We observed minimal differences, and none that was
persistent between the Bt and non-Bt brinjal treatments in the total population levels
of culturable soil bacteria and fungi. This is in agreement with our previous year study
where, we did not observe any significant effect of Bt brinjal on soil microflora (
“Effect of transgenic brinjal expressing Bacillus thuringiensis cry1 Ac gene on soil
microflora, collembola, and earthworms” - Report submitted to RCGM during July
2004). Even though no previous published reports on the effect of Bt brinjal on soil
bacteria and fungi are available, the effects of transgenic cotton and corn plants
expressing the Bt proteins have been studied. Donegan et al. (1995) evaluated the
effect of Bt cotton on the soil microflora by placing leaves of transgenic cotton plants
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in soil and monitoring numbers and species of indigenous soil bacteria and fungi.
They found that transgenic cotton lines expressing the cry1Ac gene caused a transient
increase in total bacteria and fungi population levels that were significantly higher on
several sample days than in the non-transgenic treatment. These authors suggested
that the genetic manipulation or the tissue-culturing of the plants might have resulted
in a change in plant characteristics that, other than production of Cry1Ac toxin, might
have impacted the soil microflora. Saxena and Stotzky (2001) showed that Bt corn
expressing the cry1Ab gene did not have any effect on culturable bacteria (including
actinomycetes), fungi, protozoa, nematodes, and earthworms. These findings
substantiate the findings of this study: i.e., the culturable microbial populations were
not adversely affected by Bt brinjal plants.
Collembola and earthworms aid in the decomposition process by fragmenting and
conditioning plant debris before further breakdown by microorganisms. The data of
our study indicated that soil collembola were not adversely affected by Bt brinjal
expressing cry1Ac gene. Our data are in relatively good agreement with the findings
of other researchers. Yu et al. (1997) evaluated the effect of transgenic potato
expressing Cry3A protein and transgenic cotton expressing cry1Ab or cry1Ac genes
on collembola and found that the time to attain reproductive maturity, egg production,
and final body length did not differ between collembola fed with transgenic or non-
transgenic plant tissue of these crops. Sims and Martin (1997) evaluated the dietary
toxicity of four Bt insecticidal proteins (Cry1Ab, Cry1Ac, Cry2A, and Cry3A) against
collembola and reported that purified Bt insecticidal proteins, equivalent to proteins
expressed in transgenic plants, posed no identifiable toxicological risk to soil-
inhabiting collembola. But, United States Environmental Protection Agency (EPA
2000) reported that the Cry1Ab protein found in one type of transgenic corn (event
176), when added to an artificial soil diet mix, caused significant mortality to
collembola and significantly reduced reproduction of the survivors. The LD50 was 240
mg Bt maize leaf protein/ kg of soil (95% CL 210-280). The lowest observed effect
level (LOEL) was 250 mg/ kg and the no observed effect level (NOEL) was 125 mg/
kg of soil. Even though EPA reported this non-target effect, they concluded that the
higher dose of Cry1Ab protein in corn (event 176) could have caused the adverse
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effect, and that there is a 200-fold safety factor in the levels of toxin that would occur
in the field.
Except in Jalandhar location, earthworms were observed at other locations during all
the pre-harvest sampling time-points of this study (Table 1-3). Published reports
indicate that Bt proteins do not have any adverse effect on earthworms (Saxena and
Stotzky 2001; Zwahlen et al. 2003b). Saxena and Stotzky (2001) did not find any
significant differences in mortality and weight of earthworms exposed to root
exudates or plant biomass of transgenic Bt corn or the near-isogenic control hybrid
after 40 or 45 days, respectively. Similarly, Zwahlen et al. (2003b) did not find any
significant differences in mortality between earthworms fed with biomass of Bt and
non-Bt corn. The nematode data of this study (Figures 15-21) indicated that Bt brinjal
appears not to be toxic to nematodes. This is in agreement with the published reports
(Saxena and Stotzky, 2001; Al-Deeb et al., 2003) indicating that Bt proteins do not
have any toxic effect to nematodes.
The absence of mortality in larvae released on soil-diet mixture of all locations clearly
suggests that soil samples do not contain any accumulated levels of Bt protein under
actual field conditions. Also, in insect bioassays done with soil samples from
different locations, the growth of the larvae as indicated by surviving percent third
instar larvae is equal to the growth observed when control (artificial diet alone) was
taken. This again suggests that there are no Bt accumulations that would cause
growth inhibition or mortality among the larvae. Other studies, performed under
actual conditions, have shown that Bt proteins produced in Bt transgenic crops are
rapidly degraded in soil. Palm et al. (1996) reported that in most of the experiments
with transgenic Bt cotton (Cry1Ab or Cry1Ac) and purified Bt toxin (Cry1Ac), an
initial rapid decline in extractable toxin concentration during the first 14 days,
followed by a slow decline until the end of the experiment. At the end of the
experiment, Bt toxin from transgenic cotton plants was
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and the half life of the protein in transgenic Bt cotton plant tissue was 41 days. Studies
of the environmental fate and degradation of other Bt proteins, such as Cry3A and
Cry1Ab, found that the half-lives of these proteins in soil was generally less than 20
days (Ream et al., 1994; Sims and Holden, 1996; Palm et al., 1996). Head et al.
(2002) reported that the quantity of Cry1Ac protein that accumulated as a result of
continuous culture of transgenic Bt cotton and subsequent incorporation of plant
residues into the soil by post-harvest tillage was low and did not result in any
detectable biological activity.
Conclusion
Our findings demonstrate that transgenic brinjal expressing Bt cry1 Ac gene does not
have any adverse effect on soil microflora (both fungi and bacteria), and soil
invertebrates such as earthworms, and collembola. This is not surprising since B.
thuringiensis and Bt proteins have shown to act specifically on the target insect pests
and do not have any deleterious effect on non-target organisms. Also, no Bt protein
was detected in any of the soil samples from Bt brinjal field plots.
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Figure 1. Effect of Bt brinjal on total rhizosphere and non-rhizosphere soil bacteria.
Colony forming units (± Standard error) x106
gram-1
soil. Location: Jalandhar (north
zone). Treatment details: T1 = MHB80 Bt, T2 = MHB80 non-Bt, T3 = non-Bt local
Check, T4 = Non-Bt commercial Check. P values for rhizosphere populations: 30
days after transplantation (DAP) = 0.0036, 60 DAP = 0.0407, 90 DAP = 0.1619, 120
DAP = 0.0301, and 150 DAP =0.0001. P values for non-rhizosphere samples: 30
DAP = 0.2469, 60 DAP = 0.0177, 90 DAP =0.7223, 120 DAP = 0.0011, and 150
DAP = 0.0001.
Jalandhar : rhizosphere bacteria
1.4
1.6
1.8
2
2.2
30 60 90 120 150
Sampling time-point: days after transplantation
Colo
ny f
orm
ing u
nit
s x
10
6 g
-1
soil
T1
T2
T3
T4
Jalandhar : non-rhizosphere bacteria
1
1.2
1.4
1.6
1.8
2
2.2
30 60 90 120 150
Sampling time-point : days after transplantation
Colo
ny f
orm
ing u
nit
s x 1
06 g
-1
soil
T1
T2
T3
T4
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Figure 2. Effect of Bt brinjal on total rhizosphere and non-rhizosphere soil bacteria.
Colony forming units (± Standard error) x106 gram
-1 soil. Location: Mirzapur (north
zone). Treatment details: T1 = MHB80 Bt, T2 = MHB80 non-Bt, T3 = non-Bt local
Check, T4 = Non-Bt commercial Check. P values for rhizosphere populations: 30
days after transplantation (DAP) = 0.0027, 60 DAP = 0.0001, and 90 DAP = 0.0008.
P values for non-rhizosphere samples: 30 DAP=0.0056, 60 DAP = 0.0001, and 90
DAP = 0.0022.
Mirzapur : rhizosphere bacteria
1
1.2
1.4
1.6
1.8
2
2.2
30 60 90
Sampling time-point: days after transplantation
Co
lon
y f
orm
ing
un
its
x 1
06 g
-1
soil
T1
T2
T3
T4
Mirzapur : non-rhizosphere bacteria
1
1.2
1.4
1.6
1.8
30 60 90
Sampling time-point : days after transplantation
Co
lon
y f
orm
ing
un
its
x 1
06 g
-1
soil
T1
T2
T3
T4
-
Page 17 of 53
Figure 3. Effect of Bt brinjal on total rhizosphere and non-rhizosphere soil bacteria.
Colony forming units (± Standard error) x106 gram
-1 soil. Location: Ahmad nagar
(central zone). Treatment details: T1 = MHB 4 Bt, T2 = MHB 4 non-Bt, T3 = non-Bt
local check- Gondegaon, T4 = non-Bt commercial check - Ajay. P values for
rhizosphere populations: 30 days after transplantation (DAP) = 0.0252, 60 DAP =
0.0004, 90 DAP = 0.0658, 120 DAP = 0.0001, and 150 DAP = 0.0802. P values for
non-rhizosphere samples: 30 DAP = 0.7792, 60 DAP = 0.0111, 90 DAP = 0.3459,
120 DAP = 0.1133, and 150 DAP = 0.0148.
Ahmad nagar : non-rhizosphere bacteria
1
1.2
1.4
1.6
1.8
30 60 90 120 150
Sampling time-point : days after transplantation
Co
lon
y f
orm
inh
un
its
x 1
06 g
-1 s
oil
T1
T2
T3
T4
Ahamad nagar: rizosphere bacteria
1
1.2
1.4
1.6
1.8
2
2.2
2.4
30 60 90 120 150
Sampling time-point : days after transplantation
Colo
ny f
orm
ing u
nit
s x 1
06 g
-1
soil
T1
T2
T3
T4
-
Page 18 of 53
Figure 4. Effect of Bt brinjal on total rhizosphere and non-rhizosphere soil bacteria.
Colony forming units (± Standard error) x106 gram
-1 soil. Location: Bhopal (central
zone). Treatment details: T1 = MHB80 Bt, T2 = MHB80 non-Bt, T3 = non-Bt local
check – Pusa purple round, T4 = non-Bt commercial check Navakiran. P values for
rhizosphere populations: 30 days after transplantation (DAP) = 0.0011, 60 DAP =
0.0208, 90 DAP = 0.0174, 120 DAP = 0.1157, and 150 DAP =0.0126. P values for
non-rhizosphere samples: 30 DAP = 0.0001, 60 DAP = 0.5142, 90 DAP = 0.0001,
120 DAP = 0.0078, and 150 DAP = 0.2863.
Bhopal : rhizosphere bacteria
1
1.2
1.4
1.6
1.8
2
60 90 120 150
Sampling time-point : days after transplantation
Co
lon
y f
orm
ing
un
its
x 1
06 g
-1
soil
T1
T2
T3
T4
Bhopal : non-rhizosphere bacteria
1
1.2
1.4
1.6
1.8
30 60 90 120 150
Sampling time-point : days afer transplantation
Co
lon
y f
orm
ing
un
its
x1
06 g
-1 s
oil
T1
T2
T3
T4
-
Page 19 of 53
Figure 5. Effect of Bt brinjal on total rhizosphere and non-rhizosphere soil bacteria.
Colony forming units (± Standard error) x106 gram
-1 soil. Location: Dharmapuri
(south zone). Treatment details: T1 = MHB 9 Bt, T2 = MHB 9 non-Bt, T3 = non-Bt
local check- Arka Shirish, T4 = non-Bt commercial check- Extra green long. P values
for rhizosphere populations: 30 days after transplantation (DAP) = 0.0135, 60 DAP =
0.0009, 90 DAP = 0.0963, 120 DAP = 0.0266, and 150 DAP = 0.0227. P values for
non-rhizosphere samples: 30 DAP = 01421, 60 DAP = 0.0039, 90 DAP = 0.6974, 120
DAP = 0.0041, and 150 DAP = 0.0001.
Dharmapuri : non-rhizosphere bacteria
1
1.2
1.4
1.6
1.8
30 60 90 120 150
Sampling time-point : days after transplantation
Colo
ny f
orm
ing u
nit
s x 1
06 g
-1
soil
T1
T2
T3
T4
Dharmapuri : rhizosphere bacteria
1
1.2
1.4
1.6
1.8
2
2.2
30 60 90 120 150
Sampling time-point : days after transplantation
Colo
ny f
orm
ing u
nit
s x 1
06 g
-1
soil
T1
T2
T3
T4
-
Page 20 of 53
Figure 6. Effect of Bt brinjal on total rhizosphere and non-rhizosphere soil bacteria.
Colony forming units (± Standard error) x106 gram
-1 soil. Location: Dharward (south
zone). Treatment details: T1 = MHB 10 Bt, T2 = MHB 10 non-Bt, T3 = non-Bt local
check- Manjari kota, T4 = non-Bt commercial check- Manju. P values for
rhizosphere populations: 30 days after transplantation (DAP) = 0.0012, 60 DAP =
0.0931, 90 DAP = 0.0002, 120 DAP = 0.0026, and 150 DAP = 0.0109. P values for
non-rhizosphere samples: 30 DAP=0.0036, 60 DAP = 0.386, 90 DAP = 0.0534, 120
DAP = 0.8172, and 150 DAP = 0.2765.
Dharward : rhizosphere bacteria
1
1.5
2
2.5
30 60 90 120 150
Sampling time-point : days after transplantation
Colo
ny f
orm
ing u
nit
s x10
6 g
-1 s
oil
T1
T2
T3
T4
Dharward : non-rhizosphere bacteria
1
1.2
1.4
1.6
1.8
2
2.2
30 60 90 120 150
Sampling time-point : days after transplantation
Colo
ny f
orm
ing u
nit
s x10
6 g
-1 s
oil
T1
T2
T3
T4
-
Page 21 of 53
Figure 7. Effect of Bt brinjal on total rhizosphere and non-rhizosphere soil bacteria.
Colony forming units (± Standard error) x106 gram
-1 soil. Location: Karnool (south
zone). Treatment details: T1 = MHB 99 Bt, T2 = MHB 99 non-Bt, T3 = non-Bt local
check - green + white local, T4 = non-Bt commercial check- Harit. P values for
rhizosphere populations: 30 days after transplantation (DAP) = 0.0001, 60 DAP =
0.0001, 90 DAP = 0.0066, 120 DAP = 0.0036, and 150 DAP = 0.0122. P values for
non-rhizosphere samples: 30 DAP = 0.0020, 60 DAP = 0.6019, 90 DAP = 0.6934,
120 DAP = 0.1486 and 150 DAP = 0.3102.
Karnool : non-rhizosphere bacteria
1
1.2
1.4
1.6
1.8
2
30 60 90 120 150
Sampling time-point : days after transplantation
Colo
ny f
orm
ing u
nit
s x 1
06 g
-1 s
oil
T1
T2
T3
T4
Karnoo l : rh izo sphere bact eria
1
1 .2
1 .4
1 .6
1 .8
2
2 .2
2 .4
30 60 90 12 0 1 50
Sam plin g t im e-p oin t : day s aft er t ransp lan t at io n
Colo
ny f
orm
ing u
nit
s 10
6 g
-1 s
oil
T 1
T 2
T 3
T 4
-
Page 22 of 53
Figure 8. Effect of Bt brinjal on total rhizosphere and non-rhizosphere fungal
population. Colony forming units (±Standard error) x104
gram-1
soil. Location:
Jalandhar (north zone). Treatment details: T1 = MHB80 Bt, T2 = MHB80 non-Bt, T3
= non-Bt local check – Pusa purple round, T4 = non-Bt commercial check- Navakiran.
P values for rhizosphere populations: 30 days after transplantation (DAP) =0.0248,
60 DAP = 0.2375, 90 DAP = 0.3870, 120 DAP = 0.5408, and 150 DAP =0.0065. P
values for non-rhizosphere samples: 30 DAP = 0.7072, 60 DAP = 0.3283, 90 DAP =
0.4548, 120 DAP = 0.0417, and 150 DAP =0.0123.
Jalandhar : rhizosphere fungi
15
20
25
30
30 60 90 120 150
Sampling time-point: days after transplantation
Colo
ny f
orm
ing u
nit
s x
10
4 g
-1
soil
T1
T2
T3
T4
Jalandhar : non-rhizosphere fungi
15
20
25
30
30 60 90 120 150
Sampling time-point : days after transplantation
Colo
ny f
orm
ing u
nit
s x 1
04 g
-1
soil
T1
T2
T3
T4
-
Page 23 of 53
Figure 9. Effect of Bt brinjal on total rhizosphere and non-rhizosphere fungal
population. Colony forming units (±Standard error) x104 gram
-1 soil. Location:
Mirzapur (north zone). Treatment details: T1 = MHB 80 Bt, T2 = MHB 80 non-Bt,
T3 = non-Bt local check – Pusa purple round, T4 = non-Bt commercial check-
Navakiran. P values for rhizosphere populations: 30 days after transplantation (DAP)
= 0.0689, 60 DAP = 0.4574, and 90 DAP = 0.0055. P values for non-rhizosphere
samples: 30 DAP = 0.798, 60 DAP = 0.6673, and 90 DAP = 0.0417.
Mirzapur : rhizosphere fungi
15
20
25
30
30 60 90
Sampling time-point: days after transplantation
Co
lon
y f
orm
ing
un
its
x 1
04 g
-1
soil
T1
T2
T3
T4
Mirzapur : non-rhizosphere fungi
12
16
20
24
28
30 60 90
Sampling time-point : days after transplantation
Colo
ny f
orm
ing u
nit
s x 1
04 g
-1
soil
T1
T2
T3
T4
-
Page 24 of 53
Figure 10. Effect of Bt brinjal on total rhizosphere and non-rhizosphere fungal
population. Colony forming units (±Standard error) x104 gram
-1 soil. Location:
Ahmad nagar (central zone). Treatment details: T1 = MHB 4 Bt, T2 = MHB 4 non-Bt,
T3 = non-Bt local check- Gondegaon, T4 = non-Bt commercial check - Ajay. P values
for rhizosphere populations: 30 days after transplantation (DAP) = 0.5043, 60 DAP =
0.6021, 90 DAP = 0.0178, 120 DAP = 0.0358, and 150 DAP = 0.0001. P values for
non-rhizosphere samples: 30 DAP = 0.0903, 60 DAP = 0.0231, 90 DAP = 0.0001,
120 DAP = 0.8311, and 150 DAP = 0.0006.
Ahmad nagar: rhizosphere fungi
10
14
18
22
26
30 60 90 120 150
Sampling time-point : days after transplantation
Colo
ny f
orm
ing u
nit
s x 1
04 g
-1
soil
T1
T2
T3
T4
Ahmad nagar : non-rhizosphere fungi
10
12
14
16
18
20
22
24
30 60 90 120 150
Sampling time-point : days after transplantation
Colo
ny f
orm
ing u
nit
s x 1
04 g
-1
soil
T1
T2
T3
T4
-
Page 25 of 53
Figure 11. Effect of Bt brinjal on total rhizosphere and non-rhizosphere fungal
population. Colony forming units (±Standard error) x 104 gram
-1 soil. Location:
Bhopal (central zone). Treatment details: T1 = MHB 80 Bt, T2 = MHB 80 non-Bt, T3
= non-Bt local check- Pusa purple round, T4 = non-Bt commercial check - Navakiran.
P values for rhizosphere populations: 30 days after transplantation (DAP) = 0.2749,
60 DAP = 0.0015, 90 DAP = 0.4667, 120 DAP = 0.0405 and 150 DAP = 0.1848. P
values for non-rhizosphere samples: 30 DAP = 0.0611, 60 DAP =0.3230, 90 DAP =
0.0382, 120 DAP = 0.0002, and 150 DAP = 0.2670.
Bhopal : rhizosphere fungi
10
15
20
25
30
30 60 90 120 150
Sampling time-point : days after transplantation
Co
lon
y f
orm
ing
un
its
x 1
04 g
-1
soil
T1
T2
T3
T4
Bhopal : non-rhizosphere fungi
10
15
20
25
30
30 60 90 120 150
Sampling time-point : days afer transplantation
Colo
ny f
orm
ing u
nit
s x10
4 g
-1 s
oil
T1
T2
T3
T4
-
Page 26 of 53
Figure 12. Effect of Bt brinjal on total rhizosphere and non-rhizosphere fungal
population. Colony forming units (±Standard error) x104
gram-1
soil. Location:
Dharmapuri (south zone). Treatment details: T1 = MHB 9 Bt, T2 = MHB 9 non-Bt,
T3 = non-Bt local check- Arka Shirish, T4 = non-Bt commercial check – Extra green
long. P values for rhizosphere populations: 30 days after transplantation (DAP) =
0.0094, 60 DAP = 0.0180, 90 DAP = 0.9485, 120 DAP = 0.3392, and 150 DAP =
0.4556. P values for non-rhizosphere samples: 30 DAP=0.0896, 60 DAP = 0.0478, 90
DAP = 0.0001, 120 DAP = 0.5526, and 150 DAP = 0.7728.
Dharmapuri : rhizosphere fungi
10
15
20
25
30
35
30 60 90 120 150
Sampling time-point : days after transplantation
Colo
ny f
orm
ing u
nit
s x 1
04 g
-1
soil
T1
T2
T3
T4
Dharmapuri : non-rhizosphere fungi
10
12
14
16
18
20
22
24
26
30 60 90 120 150
Sampling time-point : days after transplantation
Colo
ny f
orm
ing u
nit
s x 1
04 g
-1
soil
T1
T2
T3
T4
-
Page 27 of 53
Figure 13. Effect of Bt brinjal on total rhizosphere and non-rhizosphere fungal
population. Colony forming units (± Standard error) x104 gram
-1 soil. Location:
Dharward (south zone). Treatment details: T1 = MHB 10 Bt, T2 = MHB 10 non-Bt,
T3 = non-Bt local Manjari Kota, T4 = non-Bt commercial check - Manju. P values
for rhizosphere populations: 30 days after transplantation (DAP) = 0.1762, 60 DAP =
0.7600, 90 DAP = 0.0901, 120 DAP = 0.9435, and 150 DAP = 0.1310. P values for
non-rhizosphere samples: 30 DAP = 0.0435, 60 DAP = 0.0161, 90 DAP = 0.5399,
120 DAP = 0.9811, and 150 DAP = 0.2882.
Dharward : rhizosphere fungi
15
20
25
30 60 90 120 150
Sampling time-point : days after transplantation
Colo
ny f
orm
ing u
nit
s x10
4 g
-1 s
oil
T1
T2
T3
T4
Dharward : non-rhizosphere fungi
14
18
22
26
30 60 90 120 150
Sampling time-point : days after transplantation
Colo
ny f
orm
ing u
nit
s x10
4 g
-1 s
oil
T1
T2
T3
T4
-
Page 28 of 53
Figure 14. Effect of Bt brinjal on total rhizosphere and non-rhizosphere fungal
population. Colony forming units (± Standard error) x 104 gram
-1 soil. Location:
Karnool (south zone). Treatment details: T1 = MHB 99 Bt, T2 = MHB 99 non-Bt, T3
= non-Bt local check - Green+white local, T4 = non-Bt commercial check - Harit. P
values for rhizosphere populations: 30 days after transplantation (DAP) = 0.1240, 60
DAP = 0.1255, 90 DAP = 0.0605, 120 DAP = 0.0021, and 150 DAP = 0.141. P
values for non-rhizosphere samples: 30 DAP = 0.4103, 60 DAP = 0.1124, 90 DAP =
0.5024, 120 DAP = 0.0001, and 150 DAP = 0.0327.
Karnool : rhizosphere fungi
10
15
20
25
30
30 60 90 120 150
Sampling time-point: days after transplantation
Colo
ny f
orm
ing u
nit
s 10
4 g
-1 s
oil
T1
T2
T3
T4
Karnool : non-rhizosphere fungi
10
14
18
22
26
30 60 90 120
Sampling time-point : days after transplantation
Colo
ny f
orm
ing u
nit
s x 1
04 g
-1
soil
T1
T2
T3
T4
-
Page 29 of 53
Figure 15. Effect of Bt brinjal on soil nematode population. Total number of
nematodes (±Standard error) 250 gram-1
soil. Each value is a mean of three
replications. Location: Jalandhar (north zone). Treatment details: T1 = MHB 80 Bt,
T2 = MHB 80 non-Bt, T3 = non-Bt local check – Pusa purple round, T4 = non-Bt
commercial check - Navakiran. P values: 30 days after transplantation (DAP) =
0.4541, 60 DAP = 0.9879, 90 DAP = 0.8970, 120 DAP = 0.7529, and 150 DAP =
0.1656.
Jalandhar
0
50
100
150
200
250
300
350
30 60 90 120 150
Sampling time-point : days after transplantation
Nu
mb
er o
f n
emat
od
es 2
50
g -1
so
ilT1
T2
T3
T4
-
Page 30 of 53
Figure 16. Effect of Bt brinjal on soil nematode population. Total number of
nematodes (±Standard error) 250 gram-1
soil. Each value is a mean of three
replications. Location: Mirzapur (north zone). Treatment details: T1 = MHB 80 Bt,
T2 = MHB 80 non-Bt, T3 = non-Bt local check – Pusa purple round, T4 = non-Bt
commercial check – Navakiran. P values: 30 days after transplantation (DAP) =
0.9692, 60 DAP = 0.8043, and 90 DAP = 0.0002.
Mirzapur
0
50
100
150
200
30 60 90
Sampling time-point: days after transplantation
Nu
mb
er o
f n
emat
od
es 2
50
g-1
soil
T1
T2
T3
T4
-
Page 31 of 53
Figure 17. Effect of Bt brinjal on soil nematode population. Total number of
nematodes (±Standard error) 250 gram-1
soil. Each value is a mean of three
replications. Location: Ahmad nagar (central zone). Treatment details: T1 = MHB 4
Bt, T2 = MHB 4 non-Bt, T3 = non-Bt local check Gondegaon, T4 = non-Bt
commercial check- Ajay. P values: 30 days after transplantation (DAP) = 0.7676, 60
DAP = 0.7554, 90 DAP = 0.7928, 120 DAP = 0.9147, and 150 DAP = 0.8990.
Ahmad nagar
0
50
100
150
200
250
30 60 90 120 150
Sampling time-point : days after transplantation
Nu
mb
er o
f n
emat
od
es 2
50
g-1
soil
T1
T2
T3
T4
-
Page 32 of 53
Figure 18. Effect of Bt brinjal on soil nematode population. Total number of
nematodes (±Standard error) 250 gram-1
soil. Each value is a mean of three
replications. Location: Bhopal (central zone). Treatment details: T1 = MHB 80 Bt,
T2 = MHB 80 non-Bt, T3 = non-Bt local check – Pusa purple long, T4 = non-Bt
commercial check- Navakiran. P values: 30 days after transplantation (DAP) =
0.6761, 60 DAP = 0.7015, 90 DAP = 0.5386, 120 DAP = 0.0838 and 150 DAP =
0.0326.
Bhopal
0
100
200
300
400
500
600
30 60 90 120 150
Sampling time-point : days after transplantation
Nu
mb
er o
f n
emat
od
es 2
50
g-1
soil
T1
T2
T3
T4
-
Page 33 of 53
Figure 19. Effect of Bt brinjal on soil nematode population. Total number of
nematodes (±Standard error) 250 gram-1
soil. Each value is a mean of three
replications. Location: Dharmapuri (south zone). Treatment details: T1 = MHB 9 Bt,
T2 = MHB 9 non-Bt, T3 = non-Bt local check – Arka Shirish, T4 = non-Bt
commercial check – Extra green long. P values: 30 days after transplantation (DAP)
= 0.3694, 60 DAP = 0.5745, 90 DAP = 0.0583, 120 DAP = 0.8492, and 150 DAP =
0.2438.
Dharmapuri
0
50
100
150
200
250
300
350
400
450
30 60 90 120 150
Sampling time-point: days after transplanation
Nu
mb
er o
f n
emat
od
es 2
50
g-1
so
ilT1
T2
T3
T4
-
Page 34 of 53
Figure 20. Effect of Bt brinjal on soil nematode population. Total number of
nematodes (±Standard error) 250 gram-1
soil. Each value is a mean of three
replications. Location: Dharward (south zone). Treatment details: T1 = MHB 10 Bt,
T2 = MHB 10 non-Bt, T3 = non-Bt local check- Manjari kota, T4 = non-Bt
commercial check- Manju. P values: 30 days after transplantation (DAP) = 0.3092,
60 DAP = 0.6835, 90 DAP = 0.6861, 120 DAP = 0.1006, and 150 DAP = 0.3296.
Dharward
0
100
200
300
400
500
600
30 60 90 120 150
Sampling time-point : days after transplantation
Nu
mb
er o
f n
emat
od
es 2
50
g-1
soil
T1
T2
T3
T4
-
Page 35 of 53
Figure 21. Effect of Bt brinjal on soil nematode population. Total number of
nematodes (±Standard error) 250 gram-1
soil. Each value is a mean of three
replications. Location: Karnool (south zone). Treatment details: T1 = MHB 99 Bt,
T2 = MHB 99 non-Bt, T3 = non-Bt local check –Green + white local, T4 = non-Bt
commercial check- Harit. P values for rhizosphere populations: 30 days after
transplantation (DAP) = 0.4283, 60 DAP = 0.5717, 90 DAP = 0.1112, 120 DAP =
0.8720, and 150 DAP = 0.032.
Karnool
0
100
200
300
400
500
600
30 60 90 120 150
Sampling time-point: days after transplantation
Nu
mb
er o
f n
emat
od
es 2
50
g-1
soil
T1
T2
T3
T4
-
Page 36 of 53
Figure 22. Effect of Bt brinjal on soil Collembola. Each value is a mean of three
replications. Location: Jalandhar (north zone). Treatment details: T1 = MHB80 Bt, T2
= MHB 80 non-Bt, T3 = non-Bt local check - Pusa purple round, T4 = non-Bt
commercial check- Navakiran. P values: 30 days after transplantation (DAP) = 0.359,
60 DAP = 0.376, 90 DAP = 0.859, 120 DAP = 0.095, and 150 DAP = 0.0645.
Jalandhar
0
5
10
15
20
25
30 60 90 120 150
Sampling time-point: days after transplantation
Co
llem
bo
la p
op
ula
tio
n :
Mea
n ±
SE
T1
T2
T3
T4
-
Page 37 of 53
Figure 23. Effect of Bt brinjal on soil Collembola. Each value is a mean of three
replications. Location: Mirzapur (north zone). Treatment details: T1 = MHB80 Bt, T2
= MHB 80 non-Bt, T3 = non-Bt local check - Pusa purple round, T4 = non-Bt
commercial check- Navakiran. P values: 30 days after transplantation (DAP) = 0.481,
60 DAP = 0.981, and 90 DAP = 0.0383.
Mirzapur
0
4
8
12
16
20
30 60 90
Sampling time-point; days after transplantation
Co
llem
bo
la p
op
ula
tio
n :
Mea
n ±
SE
T1
T2
T3
T4
-
Page 38 of 53
Figure 24. Effect of Bt brinjal on soil Collembola. Each value is a mean of three
replications. Location: Ahmad Nagar (central zone). Treatment details: T1 = MHB 4
Bt, T2 = MHB 4 non-Bt, T3 = non-Bt local check- Gondegaon, T4 = non-Bt
commercial check - Ajay. P values: 30 days after transplantation (DAP) = 0.433, 60
DAP = 0.847, 90 DAP = 0.121, 120 DAP = 0.734, and 150DAP = 0.1296.
Ahmad nagar
0
10
20
30
40
50
60
30 60 90 120 150
Sampling time-point: days after transplantation
Co
llem
bo
al p
op
ula
tio
n:
Mea
n ±
SE
T1
T2
T3
T4
-
Page 39 of 53
Figure 25. Effect of Bt brinjal on soil Collembola. Each value is a mean of three
replications. Location: Bhopal (central zone). Treatment details: T1 = MHB80 Bt, T2
= MHB 80 non-Bt, T3 = non-Bt local check - Pusa purple round, T4 = non-Bt
commercial check- Navakiran. P values: 30 days after transplantation (DAP) = 0.777,
60 DAP = 0.793, 90 DAP = 0.394, 120 DAP = 0.223, and 150 DAP = 0.3376.
Bhopal
0
1
2
3
4
5
30 60 90 120 150
Sampling time-point : days after transplantation
Co
llem
bo
la p
op
ula
tio
n :
Mea
n ±
SE
T1
T2
T3
T4
-
Page 40 of 53
Figure 26. Effect of Bt brinjal on soil Collembola population. Each value is a mean of
three replications. Location: Dharmapuri (south zone). Treatment details: T1 = MHB
9 Bt, T2 = MHB 9 non-Bt, T3 = non-Bt local check- Arka Shirish, T4 = non-Bt
commercial check- extra green long. P values: 30 days after transplantation (DAP) =
0.385, 60 DAP = 0.683, 90 DAP = 0.708, 120 DAP = 0.564, and 150 DAP = 0.2561.
Dharmapuri
0
5
10
15
20
25
30 60 90 120 150
Sampling time-point : days after transplantation
Co
llem
bo
la p
op
ula
tio
n :
Mea
n ±
SE
T1
T2
T3
T4
-
Page 41 of 53
Figure 27. Effect of Bt brinjal on soil Collembola population. Each value is a mean of
three replications. Location: Dharward (south zone). Treatment details: T1 = MHB
10 Bt, T2 = MHB 10 non-Bt, T3 = non-Bt local check- Manjari kota, T4 = non-Bt
commercial check- Manju. P values: 30 days after transplantation (DAP) = 0.71, 60
DAP = 0.68, 90 DAP = 0.793, 120 DAP = 0.802, and 150 DAP = 0.0553.
Dharward
0
1
2
3
4
30 60 90 120 150
Sampling time-point : days after transplantation
Co
llem
bo
la p
op
ula
tio
n :
Mea
n±
SE
T1
T2
T3
T4
-
Page 42 of 53
Figure 28. Effect of Bt brinjal on soil Collembola. Each value is a mean of three
replications. Location: Karnool (south zone). Treatment details: T1 = MHB 99 Bt,
T2 = MHB 99 non-Bt, T3 = Non-Bt local check - Green + white local, T4 = non-Bt
commercial check- Harit. P values: 30 days after transplantation (DAP) = 0.86, 60
DAP = 0.841, 90 DAP = 0.802, 120 DAP = 0.455, and 150 DAP = 0.057.
Karnool
0
1
2
3
4
5
6
7
30 60 90 120 150
Sampling time-point: days after transplantation
Co
llem
bo
la p
op
ula
tio
n :
Mea
n ±
SE
T1
T2
T3
T4
-
Page 43 of 53
Figure 29. Dose-response curve for mortality of H. armigera neonate larvae exposed
to various concentrations of diet-incorporated Cry1Ac protein (MVP II) (n = 896)
Figure 30. Percent second instar larvae of H. armigera surviving post-exposure to
various concentrations of diet-incorporated Cry1Ac protein (MVP II). This data was
obtained from the standard mortality bioassay of which the mortality results are
presented in Figure 29.
0
10
20
30
40
50
60
70
80
90
100
0.001 0.01 0.1 1
Cry1Ac (ug/mL diet)
Pe
rce
nt
Mo
rta
lity
0
10
20
30
40
50
60
70
80
90
100
0 0.001 0.004 0.012 0.037 0.111 0.333 1
Cry1Ac (ug/ml diet)
Perc
en
t T
hir
d I
nsta
r L
arv
ae
-
Page 44 of 53
Figure 31. Insect bioassays with Helicovepa armigera to detect the levels of Cry1Ac protein
present in the A) rhizosphere and B) non-rhizosphere soil samples collected from transgenic
brinjal field plots at Jalandhar (Kharif 2004) at different crop growth stages. T1, T2, T3, T4
and C represent MHB-80 (Bt), MHB-80 (Non-Bt), Local Check (Pusa Purple Round),
Commercial Check (Navakiran - Sungro seeds) and control, respectively. Neonates of H.
armigera were fed with artificial diet +/- soil samples to detect Cry1Ac protein. Growth stages
of larvae were recorded on 7th day of release.
0
10
20
30
40
50
60
70
80
90
100
T1 T2 T3 T4 C T1 T2 T3 T4 C T1 T2 T3 T4 C T1 T2 T3 T4 C
Treatments (Rhizosphere)
Pe
rce
nt
thir
d i
ns
tar
larv
ae
30 DAT 60 DAT 90 DAT 120 DAT
Jalandhar (Kharif '04) A
0
10
20
30
40
50
60
70
80
90
100
T1 T2 T3 T4 C T1 T2 T3 T4 C T1 T2 T3 T4 C T1 T2 T3 T4 C
Treatments (Non-Rhizosphere)
Pe
rce
nt
thir
d i
ns
tar
larv
ae
30 DAT 60 DAT 90 DAT 120 DAT
Jalandhar (Kharif '04) B
-
Page 45 of 53
Figure 32. Insect bioassays with Helicovepa armigera to detect the levels of Cry1Ac protein
present in the A) rhizosphere and B) non-rhizosphere soil samples collected from transgenic
brinjal field plots at Mirzapur (Kharif 2004) at different crop growth stages. T1, T2, T3, T4
and C represent MHB-80 (Bt), MHB-80 (Non-Bt), Local Check (Pusa Purple Round),
Commercial Check (Navakiran - Sungro seeds) and control, respectively. Neonates of H.
armigera were fed with artificial diet +/- soil samples to detect Cry1Ac protein. Growth stages
of larvae were recorded on 7th day of release.
0
10
20
30
40
50
60
70
80
90
100
T1 T2 T3 T4 C T1 T2 T3 T4 C T1 T2 T3 T4 C
Treatments (Rhizosphere)
Pe
rce
nt
thir
d i
ns
tar
larv
ae
30 DAT 60 DAT 90 DAT
Mirzapur (Kharif '04) A
0
10
20
30
40
50
60
70
80
90
100
T1 T2 T3 T4 C T1 T2 T3 T4 C T1 T2 T3 T4 C
Treatments (Non-Rhizosphere)
Pe
rce
nt
thir
d i
ns
tar
larv
ae
30 DAT 60 DAT 90 DAT
Mirzapur (Kharif '04) B
-
Page 46 of 53
Figure 33. Insect bioassays with Helicovepa armigera to detect the levels of Cry1Ac protein present in
the A) rhizosphere and B) non-rhizosphere soil samples collected from transgenic brinjal field plots at
Ahmadnagar (Kharif 2004) at different crop growth stages. T1, T2, T3, T4 and C represent MHB-4
(Bt), MHB-4 (Non-Bt), Local Check (Gondegaon Local), Commercial Check (Ajay Hybrid - Ankur
seeds) and control, respectively. Neonates of H. armigera were fed with artificial diet +/- soil samples
to detect Cry1Ac protein. Growth stages of larvae were recorded on 7th
day of release.
0
10
20
30
40
50
60
70
80
90
100
T1 T2 T3 T4 C T1 T2 T3 T4 C T1 T2 T3 T4 C T1 T2 T3 T4 C
Treatments (Rhizosphere)
Pe
rce
nt
thir
d in
sta
r la
rva
e
30 DAT 60 DAT 90 DAT 120 DAT
Ahmadnagar (Kharif '04) A
0
10
20
30
40
50
60
70
80
90
100
T1 T2 T3 T4 C T1 T2 T3 T4 C T1 T2 T3 T4 C T1 T2 T3 T4 C
Treatments (Non-Rhizosphere)
Pe
rce
nt
thir
d in
sta
r la
rva
e
30 DAT 60 DAT 90 DAT 120 DAT
Ahmadnagar (Kharif '04) B
-
Page 47 of 53
Figure 34. Insect bioassays with Helicovepa armigera to detect the levels of Cry1Ac protein present in
the A) rhizosphere and B) non-rhizosphere soil samples collected from transgenic brinjal field plots at
Bhopal (Kharif 2004) at different crop growth stages. T1, T2, T3, T4 and C represent MHB-80 (Bt),
MHB-80 (Non-Bt), Local Check (Pusa Purple Round), Commercial Check (Navakiran - Sungro seeds)
and control, respectively. Neonates of H. armigera were fed with artificial diet +/- soil samples to detect
Cry1Ac protein. Growth stages of larvae were recorded on 7th
day of release.
0
10
20
30
40
50
60
70
80
90
100
T1 T2 T3 T4 C T1 T2 T3 T4 C T1 T2 T3 T4 C T1 T2 T3 T4 C
Treatments (Rhizosphere)
Pe
rce
nt
thir
d in
sta
r la
rva
e
30 DAT 60 DAT 90 DAT 120 DAT
Bhopal (Kharif '04) A
0
10
20
30
40
50
60
70
80
90
100
T1 T2 T3 T4 C T1 T2 T3 T4 C T1 T2 T3 T4 C T1 T2 T3 T4 C
Treatments (Non-Rhizosphere)
Pe
rce
nt
thir
d in
sta
r la
rva
e
30 DAT 60 DAT 90 DAT 120 DAT
Bhopal (Kharif '04) B
-
Page 48 of 53
Figure 35. Insect bioassays with Helicovepa armigera to detect the levels of Cry1Ac protein
present in the A) rhizosphere and B) non-rhizosphere soil samples collected from transgenic
brinjal field plots at Dharmapuri (Kharif 2004) at different crop growth stages. T1, T2, T3,
T4 and C represent MHB-9 (Bt), MHB-9 (Non-Bt), Local Check (Arka Shirish), Commercial
Check (Extra green long - Sungro seeds) and control, respectively. Neonates of H. armigera
were fed with artificial diet +/- soil samples to detect Cry1Ac protein. Growth stages of larvae
were recorded on 7th day of release.
0
10
20
30
40
50
60
70
80
90
100
T1 T2 T3 T4 C T1 T2 T3 T4 C T1 T2 T3 T4 C T1 T2 T3 T4 C
Treatments (Rhizosphere)
Pe
rce
nt
thir
d i
ns
tar
larv
ae
30 DAT 60 DAT 90 DAT 120 DAT
Dharmapuri (Kharif '04) A
0
10
20
30
40
50
60
70
80
90
100
T1 T2 T3 T4 C T1 T2 T3 T4 C T1 T2 T3 T4 C T1 T2 T3 T4 C
Treatments (Non-Rhizosphere)
Pe
rce
nt
thir
d i
ns
tar
larv
ae
30 DAT 60 DAT 90 DAT 120 DAT
Dharmapuri (Kharif '04) B
-
Page 49 of 53
Figure 36. Insect bioassays with Helicovepa armigera to detect the levels of Cry1Ac protein
present in the A) rhizosphere and B) non-rhizosphere soil samples collected from transgenic
brinjal field plots at Dharwad (Kharif 2004) at different crop growth stages. T1, T2, T3, T4
and C represent MHB-10 (Bt), MHB-10 (Non-Bt), Local Check (Manjari gotta), Commercial
Check (Manju - Syngenta seeds) and control, respectively. Neonates of H. armigera were fed
with artificial diet +/- soil samples to detect Cry1Ac protein. Growth stages of larvae were
recorded on 7th day of release.
0
10
20
30
40
50
60
70
80
90
100
T1 T2 T3 T4 C T1 T2 T3 T4 C T1 T2 T3 T4 C T1 T2 T3 T4 C
Treatments (Rhizosphere)
Pe
rce
nt
thir
d i
ns
tar
larv
ae
30 DAT 60 DAT 90 DAT 120 DAT
Dharwad (Kharif '04) A
0
10
20
30
40
50
60
70
80
90
100
T1 T2 T3 T4 C T1 T2 T3 T4 C T1 T2 T3 T4 C T1 T2 T3 T4 C
Treatments (Non-Rhizosphere)
Pe
rce
nt
thir
d i
ns
tar
larv
ae
30 DAT 60 DAT 90 DAT 120 DAT
Dharwad (Kharif '04) B
-
Page 50 of 53
Figure 37. Insect bioassays with Helicovepa armigera to detect the levels of Cry1Ac
protein present in the A) rhizosphere and B) non-rhizosphere soil samples collected
from transgenic brinjal field plots at Kurnool (Kharif 2004) at different crop growth
stages. T1, T2, T3, T4 and C represent MHB-99(Bt), MHB-99 (Non-Bt), Local Check
(Green +White Local), Commercial Check (Harit - Syngenta seeds) and control,
respectively. Neonates of H. armigera were fed with artificial diet +/- soil samples to
detect Cry1Ac protein. Growth stages of larvae were recorded on 7th
day of release.
0
10
20
30
40
50
60
70
80
90
100
T1 T2 T3 T4 C T1 T2 T3 T4 C T1 T2 T3 T4 C T1 T2 T3 T4 C
Treatments (Rhizosphere)
Pe
rce
nt
thir
d i
ns
tar
larv
ae
30 DAT 60 DAT 90 DAT 120 DAT
Kurnool (Kharif '04) A
0
10
20
30
40
50
60
70
80
90
100
T1 T2 T3 T4 C T1 T2 T3 T4 C T1 T2 T3 T4 C T1 T2 T3 T4 C
Treatments (Non-Rhizosphere)
Pe
rce
nt
thir
d i
ns
tar
larv
ae
30 DAT 60 DAT 90 DAT 120 DAT
Kurnool (Kharif '04) B
-
Page 51 of 53
Table 1. Earthworm populations measured in the Bt brinjal experimental plots at
North zone locations.
Mean number of earthworms a
Sampling time-point : days after transplantation
Location Treatmentsc
0 b
30 60 90 120 150
T1 0.00 0.00 0.00 0.00 1.70 0.00
T2 0.00 0.00 0.00 0.00 0.00 0.00
T3 0.00 0.00 0.00 0.00 0.33 0.00
T4 0.00 0.00 0.00 0.00 0.00 0.00
Jala
ndhar
P value 0.00 0.00 0.00 0.00 0.22 0.00
T1 1.33 1.33 1.33 0.33 YDd
YD
T2 1.33 1.00 1.00 0.00 YD YD
T3 2.00 1.33 1.30 0.33 YD YD
T4 0.67 1.33 0.67 0.33 YD YDMir
zapur
P value 0.7583 0.9849 0.7091 0.8022 YD YD
a. Each value is a mean of three replicationsb. Earthworm populations measured in the respective plots before transplantation.c. Treatment details :
Jalandhar: T1 = MHB 80 Bt, T2 = MHB 80 non-Bt, T3 = non-Bt
local check – Pusa purple round , and T4 = non-Bt
commercial check – Navakiran.
Mirzapur: T1 = MHB 80 Bt, T2 = MHB 80 non-Bt, T3 = non-Bt
local check – Pusa purple round, and T4 = non-Bt
commercial check – Navakiran.
d. YD = yet to be done.
-
Page 52 of 53
Table 2. Earthworm populations measured in the Bt brinjal experimental plots at
Central zone locations.
Mean number of earthworms a
Sampling time-point : days after transplantation
Location Treatmentsc
0 b
30 60 90 120 150
T1 0.33 3.33 2.67 3.00 5.00 1.00
T2 0.33 1.67 2.33 3.67 1.67 0.33
T3 0.67 1.33 1.00 3.67 1.33 0.67
T4 0.00 2.00 2.33 4.00 1.67 1.00
Ahm
ad
Nag
ar
P value 0.4547 0.7487 0.4547 0.8652 0.1050 0.7718
T1 0.33 6.33 2.33 2.00 1.33 0.33
T2 2.00 1.67 1.00 0.33 0.33 0.33
T3 0.67 1.67 0.33 0.33 0.33 0.00
T4 1.67 4.00 1.33 0.67 0.00 0.33Bhopal
P value 0.7537 0.4333 0.3533 0.1984 0.112 0.4757
a. Each value is a mean of three replicationsb. Earthworm populations measured in the respective plots before transplantation.c. Treatment details :
Ahmad nagar: T1 = MHB 4 Bt, T2 = MHB 4 non-Bt, T3 = non-Bt
local check – Gondegaon , and T4 = non-Bt
commercial
check – Ajay.
Bhopal: T1 = MHB 80 Bt, T2 = MHB 80 non-Bt, T3 = non-Bt
local check – Pusa purple round, and T4 = non-Bt
commercial check – Navakiran.
-
Page 53 of 53
Table 3. Earthworm populations measured in the Bt brinjal experimental plots at
South zone locations.
Mean number of earthworms a
Sampling time-point : days after transplantation
Location Treatmentsc
0 b
30 60 90 120 150
T1 0.67 0.00 1.33 1.67 2.33 2.33
T2 0.67 1.33 3.33 0.00 4.33 1.33
T3 0.67 2.00 1.67 0.67 4.33 1.00
T4 0.67 3.67 2.33 0.00 2.67 2.33
Dhar
map
uri
P value 0.8927 0.6030 0.8363 0.8363 0.6414 0.3556
T1 7.67 2.33 2.67 0.00 1.67 2.33
T2 17.00 2.67 3.33 0.00 0.00 1.00
T3 12.67 6.33 5.00 0.33 0.33 0.00
T4 10.00 5.00 6.00 0.00 0.00 1.00
Dhar
war
d
P value 0.6536 0.4925 0.0295 0.45 0.2241 0.1889
T1 0.00 0.00 1.33 2.33 1.33 1.67
T2 0.00 0.33 4.00 3.00 3.00 2.67
T3 0.00 0.00 2.67 3.00 2.67 3.00
T4 0.00 0.00 3.33 1.67 2.33 2.67
Kar
nool
P value 0.00 0.4547 0.0590 0.2287 0.1692 0.5376
a. Each value is a mean of three replicationsb. Earthworm populations measured in the respective plots before transplantation.c. Treatment details:
Dharmpuri: T1 = MHB 9 Bt, T2 = MHB 9 non-Bt, T3 = non-Bt
local
check – Arka Shirish, and T4 = non-Bt commercial
check – Extra green long.
Dharward: T1 = MHB 10 Bt, T2 = MHB 10 non-Bt, T3 = non-Bt
local
check – Manjari kota, and T4 = non-Bt commercial
check – Manju.
Karnool: T1 = MHB 99 Bt, T2 = MHB 99 non-Bt, T3 = non-Bt
local
check – Green + white local, and T4 = non-Bt
commercial
check – Harit.
-
Bt brinjal- pollen flow study
Page 1 of 17
Assessment of Pollen Flow from Bt brinjal
-
Bt brinjal- pollen flow study
Page 2 of 17
CONTENTS
S. No Particulars Page no
1 Introduction 3
2 Materials and Methods 5
2.1 Sampling procedures for pollen flow studies 5
3 Results and Discussion 7
3.1 Estimation of Pollen Dispersal 7
4 Conclusions 8
5 Summary 8
6 References 9
7 Tables 1 – 8 12
-
Bt brinjal- pollen flow study
Page 3 of 17
1. Introduction
In India among the Solanaceous vegetables, brinjal Solanum melongena Linn. is the
most important vegetable crop. It occupies 0.39 m.ha.(Sardana, 2002) with an
average production of 76 tonnes per ha. In recent years, however, the production of
brinjal not only in the Indian sub-continent but also in East and South Africa, Congo,
Malaysia, Thailand, Germany, Sri Lanka and Burma has been seriously affected due
to steady increase in insect pest infestation, especially the shoot and fruit borer,
Leucinodes orbonalis (Guen.). The young larvae of the pest bore in to petioles and
midribs of large leaves and tender shoots causing shoot tips to wilt and later they
bore in to flower buds and fruits. The affected fruits loose their market value besides
considerable reduction in yield as well as vitamin 'C ' content. The pest poses a
serious problem because of its high reproductive potential, rapid turnover of
generations and intensive cultivation of brinjal both in wet and dry seasons of the
year. In India it has been estimated that shoot and fruit borer causes damage to fruits
ranging from 25.8 - 92.5 % and yield reduction from 20.7 – 60% (Mall et.al., 1992).
Brinjal crop is also known to be attacked by a range of sucking pests, which include
leafhoppers, aphids, white flies, thrips and mites. Losses due to these sucking pests
range from 25 – 40% (Natarajan et.al., 1986, Anonymous, 1999).
Farmers all over the world use large quantities of chemical insecticides singly or in
combination to get blemish free fruits, which fetch premium price in the market. This
practice of indiscriminate use of insecticides has resulted in the build up of pesticide
residues in the produce, destruction of natural enemies, pest resurgence and
environmental pollution. As such published reports are not available on the
quantification of the chemicals used in brinjal for shoot and fruit borer control.
However, based on a sample estimate that the brinjal crop receives 5-10 sprays in a
season against the pest and each spray costing Rs.875 per ha, for the area of 0.39
m ha the total cost of insecticide sprays will be in the range of Rs. 1706-3412 million.
Considering 50% of the total insecticidal sprays in brinjal crops are targeted against
Fruit & Shoot borer, for a minimum of 2 - 5 insecticide sprays the cost would come in
the range of Rs. 1750 to 4375 per ha. If the Bt brinjal is adopted over 40% of 0.39m
ha brinjal area in India, it can
be projected that, there can be a national level saving of Rs 273 - 683 million
annually.
Crop protection to control insect pests in several crops through transgenic approach
is effectively used around the world. The development of transgenic cotton that
-
Bt brinjal- pollen flow study
Page 4 of 17
expresses insecticidal crystal protein genes from Bacillus thuringiensis (Bt) var.
kurstaki had resulted in lines with improved resistance to lepidopteran insect pests
(Perlak et al., 1990). Fischhoff et al., (1987), developed insect tolerant transgenic
tomato plants showing resistance to tobacco hornworm ( Manduca sexta L.) and
tobacco budworm (Heliothis virescens F.). Perlak et al., (1993) developed genetically
improved potatoes producing Cry IIIA proteins having protection from the damage by
Colorado potato beetles. Several successful attempts at introducing insect resistant
genes into other crops such as corn (Koziel et al., 1993), tobacco (Warren et al.,
1992) and others have been reported. Rao et al., (1999) studied the relative efficacy
of seven lepidopteran specific Bt ä-endotoxin genes on Leucinodes orbanalis and
concluded that cry1A(c) gene was effective.
Plant transformation protocols have been developed for many crops using both
particle bombardment method and Agrobacterium mediated transformation methods.
We have been able to optimize a protocol for Brinjal transformation using
Agrobacterium system with relatively high frequency. Using this system, many
transgenic plants have been obtained and characterized. To reduce pests-linked
damage in brinjal crop as well as to protect the environment from adverse affects of
pesticides, our scientists at Maharashtra Hybrid Seed Company have developed
transgenic brinjal lines expressing the lepidopteran specific cry1A(c) gene under the
control of enhanced CaMV 35 S promoter for high level expression in brinjal to
provide an effective built-in control for shoot and fruit borer. This would result in
bringing down the cultivation costs of brinjal, as contribution of chemical pesticides to
brinjal cultivation is sizable. In addition, as the protein acts only on the target pests
(lepidopteran pests), it is not reported to reduce the population of beneficial insects,
which also helps to keep the destructive pest population under control.
The study briefed here aims to assess Pollen flow from transgenic Brinjal and the
performance of Bt brinjal transgenic lines under two different agro-climatic regions
-
Bt brinjal- pollen flow study
Page 5 of 17
2. Materials and Methods
A field trial consisting of four brinjal lines was laid out during the K-02 season of
2002-03 in a randomized block design with four replications at Mahyco farms at
Mandwa, Jalna District, Maharashtra. Two transgenic brinjal lines, TB2 and TB3
which had a single insert of cry1Ac gene along with their non-Bt counter part (brinjal
variety Mahyco line 60208) with check (Manjri gota) were evaluated under two agro-
climatic regions which were approved by RCGM on 9.6.03, Ref No: BT/BS/17/02/94-
PID(Vol.III). Brinjal seedlings were transplanted in a plot of 29.4 X 18 m and spaced
at 90 X 60 cm. Each treatment plot was laid out with 5 rows of 10 plants each on a
plot of 4.5 X 6 m. The transgenic block was surrounded by rows of non-transgenic
trap rows as shown in Fig 1. Recommended agronomic practices, which include
basal dose of NPK (80:40:40 kg/ha) and subsequent top dressings, were applied.
Sprayings were initiated on attaining ETL of the sucking pests (2 leafhoppers or 5
white flies per leaf). A total of two pesticide sprays were imposed, with one
application of each selected spray for providing protection against sucking pest
complex. (Confidor-200SL @ 0.5 ml/liter for white flies and Dicofol @ 2.7 ml/liter for
control of mites).
2.1 Sampling procedures for pollen flow studies
Thirteen squares were planted with a non-spiny brinjal variety Pusa Kranti
surrounding the Bt experimental plots at varying distances (Fig 1). Fruit samples
were drawn from each row from plants at a distance of five meters on all side of the
square thrice at an interval of one month. The seeds were extracted from the fruits.
The seeds from plants were planted on the nursery beds for the squares from each
of the pickings. Data was recorded on the progeny rows grown from these plants for
all the three pickings. The
numbers of spiny seedlings were counted and their percentage was worked out to
know the out crossing.
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Fig 1. Schematic field layout (not to scale) for the estimation of pollen dispersal
by honeybees from transgenic Bt brinjal.
Central 29.4m x 18.0m Replicated Trial Block Containing Bt Brinjal Having Spiny Character
Row 6 Rows 1 to 5
Row 7
Row 8
Row 9
Row 10
Row 11
Row 12
Row 13
BeeHives
Rows 1 to 13 are planted with non-Bt, non-spiny brinjal plants forming concentric pollen-trap rows. Nos. 1 to 5 is 2 meters apart, at 2 to 10m distances and Nos. 6 to 13 is 5 m apart, at 15 to 50m distances from the transgenic plot.
Note: indicates honey bee hives at the four corners of the transgenic trial block.
Bee hives
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3. Results and Discussion
3.1 Estimation of Pollen Dispersal
3.1.1 At Mandwa
First Harvest
Out crossing was observed to an extend of 6.25 % on the western side in the eighth
Square and no out crossing was observed on other three sides in this square.
No out crossing was observed in squares 9 to 13. (Table 1).
Second Harvest
No out crossing was recorded in any of the square in the fruits harvested from
second Harvest study. (Table 2).
Third Harvest
No out crossing was recorded in any of the squares in the fruits harvested from third
harvest study. (Table 3).
A total of 1351 plant progenies (450 progenies from I, 451 progenies from II and 450
progenies from III picking) were observed for outcrossing. One progeny in the eighth
square has shown 0.6% outcrossing. Out of the 1351 progenies studied over squares
8 to 13, only one progeny has shown out crossing thus the outcrossing comes to
0.07%. Only one outcrossing event has been recorded in the eighth square, which is
situated at a distance of 25 meters from the source (Spiny Plants). No outcrossing
were recorded in squares 9,10,11,12 and 13 situated at a distance of 30, 35, 40, 45
and 50 meters from the source. (Table 4).
3.1.2 At Ranebennur
First Harvest
Out crossing was observed to an extend of 9.1 % on the western side in the eighth
square.
No out crossing was observed on other three sides to this square. No out crossing
was observed in squares 9 to 13. (Table 5).
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Second Harvest
No out crossing was recorded in any of the squares in the fruits harvested from
second harvest study. (Table 6).
Third Harvest
No out crossing was recorded in any of the squares in the fruits harvested from third
harvest study. (Table 7).
A total of 1183 plant progenies (395 progenies from I, 394 progenies from II and 395
progenies from III picking) were observed for out crossing. One progeny in the eighth
square has shown 0.8% out crossing. Out of the 1183 progenies studied over
squares 8 to 13, only one progeny has shown out crossing thus the outcrossing %
comes to 0.08%. Only one out crossing event has been recorded in the 8 th square
which is situated at a distance of 25 meters from the source (Spiny Plants). No
outcrossing were recorded in squares 9,10,11,12 and 13 situated at a distance of 30,
35, 40, 45 and 50 meters from the source. (Table 8).
Conclusions
4.1 Studies on Pollen Dispersal
Pollen dispersal studies indicate that maximum pollen transfer takes place up to 2
meters only, followed by 4 meters and 6 meters. The least dispersal was observed
at 20 meters at Mandwa location and at 15 meters at Ranebennur location from the
transgenic Bt brinjal lines (spiny plants).
Summary
Crop protection to control insect pests through transgenic approach is very effective
and forms an integral component of the integrated pest management (IPM)
strategies. The maximum amount of pollen flow from transgenic brinjal was found to
occur upto six meters and the extent of dispersal was upto a distance of 20 meters
only. It is in the best interests of the farming community the benefits of such
technology be conserved and extended for the longest possible time.
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