TEST PROTOCOL FOR EFFICACY EVALUATION OF INSECTICIDES
Introduction
The experiment is conducted in well-drained fertile and good quality soil of the research farm of the institution,
with all the normal recommended package of practice to raise the relevant crop for undertaking bioefficacy
evaluation of insecticides. The bioefficacy trial may be monitored by high-power group of scientist, nominated by
the Director of Research of the institution Heads of the relevant plant protection departments ( Entomology, Plant
Pathology / Nematology, senior experienced superannuated eminent scientists ( if needed ), along with
representation of the client industry. A checklist of protocols may be drawn up during each evaluation after
discussing with the relevant pesticide manufacturer ( client ) and the scientist ( evaluator ). The check-list may
draw up the High-power scientific group.
A project document (PD) is drawn up, with due identification number and authentication of the institution,
containing the under-mentioned details, before the commencement of the experiment and approved by the
competent authority of the institution. The data generated may be recorded in permanent work dairies and need
to be preserved in the institution for reasonable duration till the pesticide registration process is completed.
I) Bioefficacy-assessment evaluation of insecticides for tissue-chewing, cutting, boring insects
This group of insects from the insect Orders Orthoptera, Isoptera, Lepidoptera, Coleoptera, Hymenoptera, and
Diptera have to be separately considered while the PD of the evaluation protocol is designed for each agro-climate.
The natural enemies of each of these may be different from those of other sector of pests and careful consideration
of the type of species involved ( based on Indian published literature ) for these may be drawn up for agro-climate
where the bioefficacy evaluations are undertaken.
1.0
EXPERIMENTAL CONDITION
1.1
Crop, Variety and Target insect (Scientific names).
The name of the crop along with that of variety and target test organism (s), i.e. the relevant target tissuechewing, cutting, boring insects may be mentioned in the PD.
1.2
Trial conditions
Field trial should be set up where target crop of moderate susceptibility to the target insect
species. Standard crop husbandry practices like application of FYM, fertilizers including
micronutrients and other maintenance treatments including pest management practices for pests
other than the target pest should be specified. Separate trials needs to be undertaken for
evaluation of the given formulation against defined pest species in the same crop, if these pests
are invading the crop together. Also ensure that that the targeted species is not influenced by
previous pesticide application. Trial should be carried out in different agroclimatic regions.
1.3
Design and layout of the trial
1.3.1
Treatments
Test product(s), reference product(s) and untreated control, arranged in a randomized block
design or any other statistically suitable design.
1.3.2
Plot size and replication
Net plot size at least 15 – 20 sq.m. per replication per treatment but according to the formulation
or application equipment, it may be necessary to use a larger plot size
Replicates – at least three, provided the error degrees of freedom are at least 12.
1.2.0
APPLICATION OF TREATMENT
1.2.1
Test product (s)
The formulated product under investigation is applied according to the PD.
1.2.2
Reference product
Registered product, if any / recommended product for the control of target insect pest.
1.2.3
Mode of application
Application should comply with good agricultural practices. Details on the spray equipment, type
of nozzle and water volume used at different stages of crop growth should be elucidated. Addition
of sticker, if any (for monsoon sprays) must be specified. Sticker should be tried in the check plot
with water spray.
1.2.3.1 Type of application
The type of application (e.g. spray, soil incorporation, broad casting) as proposed by the sponsor.
1.2.3.2 Time and frequency of application
The time and frequency of application will normally be specified by the manufacturer/ sponsor.
The time and frequency will normally depend on the pest species, & local environmental
conditions & pest ETL’S. The number of applications and the date of each application should be
recorded.
1.2.3.3 Doses and Volumes used
The product should be tested at the proposed dose and may also be used other doses. The dosage
will normally be expressed in g a.i./ha and formulation ml or gm per ha.
1.3.0 MODE OF ASSESSMENT, RECORDING & MEASUREMENTS
1.3.1
Weather data
1.3.1.1 Weather data during experimental duration for each location should be provided and co-related
with the field data that is generated on the disease incidence and plant mortality of each treatment
across replications. The inference shall be directed towards the influence of test substance in
reducing the target insect / mite damage.
1.3.1.2
Soil
Soil type should be mentioned where the crop is cultivated for such evaluations.
1.3.2
Observations / Assessments
1.3.2.1 Insect pest Assessments
The PD shall contain the details of the observations / assessments of treatment and the
frequency as well as the various scientific basis ( published literature reference ) for those.
1.3.2.1.1
Helicoverpa armigera
Larval count before and after treatment be made and evaluated statistically for significance of
treatment effects. Run ‘t’-test between pre-count and post-count to see if the build up has been
significant in check plots at each date of observation.
The no. of damaged pods/fruits/balls/squares/loculi is recorded on suitable no. of randomly
selected plants per replication.
The observations are taken either at suitable interval or each picking or at maturity depending on
the crop.
Work out the % pod/fruit/ball/square/loculi damage reduction over control using either Abbott’s
formula or Henderson & Tilton formula :
Abbott’s formula :
Corrected %
Pod/fruit/ball/square/loculi =
Damage reduction
n in T after treatment
1- ------------------------------
x 100
n in Co after treatment
Where, n = no. of damaged pods/fruits/balls/squares/loculi; T = Treated; Co = Control
Reference: Abbott, W.S. (1925). A method of computing the effectiveness of an insecticide. J.
Econ. Entomol.; 18:265-267.
Henderson & Tilton formula :
Corrected %
n in Co before treatment x n in T after treatment
Pod/fruit/ball/square/loculi = 1- ---------------------------------------------------------------- x 100
Damage reduction
n in Co after treatment x n in T before treatment
Where, n = no. of damaged pods/fruits/balls/squares/loculi; T = Treated; Co = Control
Reference: Henderson, C.F. and E. W. Tilton, 1955. Tests with acaricides against the brow
wheat mite, J. Econ. Entomol. 48:157-161.
OR
1.3.2.1.2 Spodoptera litura
The larval population of insect is recorded on suitable no. of randomly selected plants per
replication.
Pre-treatment larval population of insect is recorded one day before the spray of insecticide.
The post-treatment larval population of insect is recorded at suitable interval after each spray.
Run ‘t’-test between pre-count and post-count to see if the build up has been significant in check
plots at each date of observation.
Work out the % reduction in larval population over control using Henderson & Tilton formula :
Corrected %
reduction in
=
larval population
n in Co before treatment x n in T after treatment
1- ----------------------------------------------------------------n in Co after treatment x n in T before treatment
x 100
Where, n = larval population; T = Treated; Co = Control
Reference: Henderson, C.F. and E. W. Tilton, 1955. Tests with acaricides against the brow
wheat mite, J. Econ. Entomol. 48:157-161.
OR
1.3.2.1.3 Leaf Folder (Rice - Cnaphalocrosis medinalis)
Observations are recorded before & after each application of insecticides.
Record the number of freshly damaged or folded leaves/hill
Observe ten hills per replication
Work out per cent damage using IRRI scale.
Scale
Score
Damage
0
No Damage
1
1 – 10%
3
11-20%
5
21 – 35%
7
36 – 50%
9
51% & above
Source : Standard Evaluation System for Rice, IRRI, 1996.
OR
1.3.2.1.3 Leaf Folder (Rice - Cnaphalocrosis medinalis)
The no. of freshly damaged leaves/hill are recorded on suitable no. of randomly selected hills per
replication.
Pre-treatment freshly damaged leaves are recorded one day before the spray of insecticide.
The post-treatment freshly damaged leaves are recorded at suitable interval after each spray.
Run ‘t’-test between pre-count and post-count to see if the build up has been significant in check
plots at each date of observation.
Work out the % reduction in freshly damaged leaves over control using Henderson & Tilton
formula :
Corrected %
Reduction in
n in Co before treatment x n in T after treatment
Freshly damaged = 1- ----------------------------------------------------------------Leaves
n in Co after treatment x n in T before treatment
x 100
Where, n = no. of freshly damaged leaves; T = Treated; Co = Control
Reference: Henderson, C.F. and E. W. Tilton, 1955. Tests with acaricides against the brow
wheat mite, J. Econ. Entomol. 48:157-161.
1.3.2.1.4 Stem Borer (Paddy Stem Borer - Scirpophaga incertulas)
Dead Hearts : Observe 10 hills/ plot at maximum tillering to stem elongation stage and work out
percent damage (based on tiller count) using IRRI scale.
White Earheads : Observe 10 hills per plot at dough stage & work out percent damage (based on
productive tillers count) using IRRI scale.
Scale
Score
Damage
0
No Damage
1
1 – 10%
3
11-20%
5
21 – 30%
7
31 – 60%
9
61% & above
Source : Standard Evaluation System for Rice, IRRI, 1996.
OR
1.3.2.1.4 Stem Borer (Paddy Stem Borer - Scirpophaga incertulas)
The no. of dead-hearts/white-earheads per hill are recorded on suitable no. of randomly selected
hills per replication.
The post-treatment dead-hearts are recorded at suitable interval after each spray.
The post-treatment white-earheads are recorded at dough stage.
Run ‘t’-test between pre-count and post-count to see if the build up has been significant in check
plots at each date of observation.
Work out the % dead-heart/white-earhead reduction over control using Abbott’s formula formula
Corrected %
Dead-heart/White-earhead
=
Damage reduction
n in T after treatment
1- ------------------------------
x 100
n in Co after treatment
Where, n = no. of Dead-heart/White-earhead; T = Treated; Co = Control
Reference: Abbott, W.S. (1925). A method of computing the effectiveness of an insecticide. J.
Econ. Entomol.; 18:265-267.
1.3.2.1.5 Fruit & Shoot Borer (Okra - Earias vitella; Brinjal – Leucinodes arbonalis)
The no. of damaged fruits/shoots are recorded on suitable no. of randomly selected plants per
replication.
Observations on damaged shoots are taken at suitable interval after each spray.
Observations on damaged fruits are taken at each picking.
Work out the % shoot/fruit damage reduction over control using Abbott’s formula :
Abbott’s formula :
Corrected %
Shoot/fruit
Damage reduction
=
n in T after treatment
1- ------------------------------
x 100
n in Co after treatment
Where, n = no. of damaged fruits/shoots; T = Treated; Co = Control
Reference: Abbott, W.S. (1925). A method of computing the effectiveness of an
insecticide. J. Econ. Entomol.; 18:265-267.
1.3.2.1.6 Diamond Back Moth, DBM (Plutella xylostella)
The larval population of insect is recorded on suitable no. of randomly selected plants per
replication.
Pre-treatment larval population of insect is recorded one day before the spray of insecticide.
The post-treatment larval population of insect is recorded at suitable interval after each spray.
Run ‘t’-test between pre-count and post-count to see if the build up has been significant in check
plots at each date of observation.
Work out the % reduction in DBM larval population over control using Henderson & Tilton
formula :
Corrected %
n in Co before treatment x n in T after treatment
Reduction in = 1- -----------------------------------------------------------------
DBM popul.
x 100
n in Co after treatment x n in T before treatment
Where, n = no. of DBM larvae; T = Treated; Co = Control
Reference: Henderson, C.F. and E. W. Tilton, 1955. Tests with acaricides against the brow
wheat mite, J. Econ. Entomol. 48:157-161.
1.4
Phyto-toxicity
Phyto-toxicity at ‘X’ and ‘2X’ dose shall be recorded.
Phytotoxicity Rating Scale (PRS)
Crop response/ Crop injury
Rating
0-00
0
1-10%
1
11-20%
2
21-30%
3
31-40%
4
41-50%
5
51-60%
6
61-70%
7
71-80%
8
81-90%
9
91-100%
10
Rating shall be recorded individually for yellowing, stunting, necrosis, epinasty
hyponasty etc.,
T
Treatment
No
Dose / ha
Days after 1st spray
g. a. i.
1
Formulation
3
5
7
10
1
2
3
4
1.5
Effect on Natural Enemies
The natural enemy spectrum of the target pest, as catalogued in the literature and perceived in the
region on the crop species in which this evaluation is done, may be recorded in the field collected
sample test insects and the impact of the targeted pesticide treatments in terms of mortality,
moribund condition, lack of natural enemy invasion unlike in untreated check plots may be
recorded.
1.5.1
Other new pests ( insects / mites / nematodes / diseases ) in the experiment
The investigator (evaluator) may look for the occurrence of any new pests ( insects / mites /
nematodes / diseases ) in each of the evaluation plots. The help of relevant specialists for each of
these may be taken and need to be recorded throughout the experiment period.
1.6.
Yield
The crop yield data may be recorded for each treatment and may be statistically analysed. The
conclusion on the treatment effect alone on enhancement of crop yield has to be brought out
using appropriate statistical and other tools so that there is no confused statement in the inference
that the pesticide dosage enhanced yield, without quantifying the avoidable crop loss due to given
pesticide dose application against the target pests in the evaluation.
1.7
Statistical Analysis
The data thus obtained shall be subjected to appropriate transformations and analyzed using
standard experimental designs.
1.8
RESULT & CONCLUSION
1.8.1
Result
Results to be submitted in table format mentioning the statistical parameters of assessment
such CD.CV%
1.8.2
Inferences and Conclusion
Brief not on the trial observations and performance of the test substance based on the results
may be provided with critical assessment of tested pesticide efficacy on the target pest(s)
along with opinion and conclusion on its impact on the target species on natural enemies of
the given agro-climate in which the bioefficacy evaluation was undertaken. Any
observations on the upsurge of any other pests ( insects / mites / nematodes / disease causing
organisms ) may be specifically mentioned. The possible role of good agricultural practice
by the new pesticide formulation may be inferred. The conclusion shall mention the most
effective concentration / dose of the pesticide to suppress the target pests. The results may be
given only quantifying the avoidable crop loss due to given pesticide dose application
against the target pests in the evaluation.
II)
Bioefficacy-assessment evaluation of insecticides for sap-sucking insects ( Jassids / Aphids /
Scale insects / Mealybugs / Whiteflies / Thrips )
Saps sucking insects pests of crops are generally early-season pests. These may be jassids, aphids,
whitefly, or tissue-lacerating pests such as thrips. Each of these insect groups has to be handled
differently in bioefficacy evaluation as their biology, metamorphic cycles and biology on the
given crop species varies. Often they may have the role of disease vectors too. Accordingly, the
design of evaluation in the PD may be developed. The natural enemies of these may be different
from those of other sector pests and careful consideration of the type of species involved ( based
on Indian published literature ) for these may be drawn up for agro-climate where the bioefficacy
evaluations are undertaken.
2.0
EXPERIMENTAL CONDITION
2.1
Crop, Variety and Target insect (Scientific names).
The name of the crop along with that of variety and target test organism (s), i.e. sap-sucking insects ( Jassids /
Aphids / Scale insects / Mealybugs / Whiteflies / Thrips ) may be mentioned in the PD.
2.2
Trial conditions
Field trial should be set up where target crop of moderate susceptibility to the target insect
species. Standard crop husbandry practices like application of FYM, fertilizers including
micronutrients and other maintenance treatments including pest management practices for pests
other than the target pest should be specified. Separate trials needs to be undertaken for
evaluation of the given formulation against defined pest species in the same crop, if these pests
are invading the crop together. Also ensure that that the targeted species is not influenced by
previous pesticide application. Trial should be carried out in different agro-climatic regions.
2.3
Design and layout of the trial
2.3.1
Treatments
Test product(s), reference product(s) and untreated control, arranged in a randomized block
design or any other statistically suitable design.
2.3.2
Plot size and replication
Net plot size at least 15 – 20 sq.m. per replication per treatment but according to the formulation
or application equipment, it may be necessary to use a larger plot size
Replicates – at least three, provided the error degrees of freedom are at least 12.
2.4
APPLICATION OF TREATMENT
2.4.1
Test product (s)
The formulated product under investigation is applied according to the PD. The appliance shall be
to facilitate uniform high volume application on the crop at its given age and canopy size.
2.4.2
Reference product
Registered product, if any / recommended product for the control of target insect pest.
2.4.3
Mode of application
Application should comply with good agricultural practices. Details on the spray equipment, type
of nozzle and water volume used at different stages of crop growth should be elucidated. Addition
of sticker, if any (for monsoon sprays) must be specified. Sticker should be tried in the check plot
with water spray.
2.4.3.1 Type of application
The type of application (e.g. foliar spray, soil incorporation, broad casting) as proposed by the
sponsor.
2.4.3.2 Time and frequency of application
The time and frequency of application will normally be specified by the manufacturer/ sponsor.
The time and frequency will normally depend on the pest species, & local environmental
conditions & pest ETL’S. The number of applications and the date of each application should be
recorded.
2.4.3.3. Doses and Volumes used
The product should be tested at the proposed dose and may also be used other doses. The dosage
will normally be expressed in g a.i./ha and formulation ml or gm per ha.
2.5 MODE OF ASSESSMENT, RECORDING & MEASUREMENTS
2.5.1
Weather data
2.5.1.1 Weather data during experimental duration for each location should be provided and co-related
with the field data that is generated on the disease incidence and plant mortality of each treatment
across replications. The inference shall be directed towards the influence of test substance in
reducing the target insect / mite damage.
2.5.1.2
Soil
Soil type should be mentioned where the crop is cultivated for such evaluations.
2.5.2
Observations / Assessments
2.5.2.1 Insect pest Assessments
The PD shall contain the details of the observations / assessments of treatment and the
frequency as well as the various scientific basis ( published literature reference ) for those.
The population of insect ( Jassids, Aphids, Mealybugs-nymph & adult; Whiteflies-Nymphs )
is recorded on a suitable no. of randomly selected plants/trees per replication.
Suitable no. of randomly selected leaves/panicles/twigs/inflorescence ( or target plant part )
per plant / tree are selected for observations.
Suitable method is used to count the insect population
Pre-treatment population of insect is recorded one day before the spray insecticide.
The post-treatment population of insect, is recorded at suitable intervals after each spray.
Run ‘t’-test between pre-count and post-count to see if the build up has been significant in check
plots at each date of observation.
Work out the % reduction in target insect population over control using Henderson & Tilton
formula :
Henderson & Tilton formula :
Corrected %
n in Co before treatment x n in T after treatment
Pod/fruit/ball/square/loculi = 1- ---------------------------------------------------------------- x 100
Damage reduction
n in Co after treatment x n in T before treatment
Where, n = no. of damaged pods/fruits/balls/squares/loculi; T = Treated; Co = Control
Reference: Henderson, C.F. and E. W. Tilton, 1955. Tests with acaricides against the brow
wheat mite, J. Econ. Entomol. 48:157-161.
2.6
Phyto-toxicity
Phyto-toxicity at ‘X’ and ‘2X’ dose shall be recorded.
Phytotoxicity Rating Scale (PRS)
Crop response/ Crop injury
Rating
0-00
0
1-10%
1
11-20%
2
21-30%
3
31-40%
4
41-50%
5
51-60%
6
61-70%
7
71-80%
8
81-90%
9
91-100%
10
Rating shall be recorded individually for yellowing, stunting, necrosis, epinasty
hyponasty etc.,
T
No
Treatment
Dose / ha
Days after 1st spray
g. a. i.
1
Formulation
3
5
7
10
1
2
3
4
2.7
Effect on Natural Enemies
The natural enemy spectrum of the target pest, as catalogued in the literature and perceived in the
region on the crop species in which this evaluation is done, may be recorded in the field collected
sample test insects and the impact of the targeted pesticide treatments in terms of mortality,
moribund condition, lack of natural enemy invasion unlike in untreated check plots may be
recorded.
2.7.1
Other new pests ( insects / mites / nematodes / diseases ) in the experiment
The investigator (evaluator) may look for the occurrence of any new pests ( insects / mites /
nematodes / diseases ) in each of the evaluation plots. The help of relevant specialists for each of
these may be taken and need to be recorded throughout the experiment period.
2.8
Crop Yield
The crop yield data may be recorded for each treatment and may be statistically analysed. The
conclusion on the treatment effect alone on enhancement of crop yield has to be brought out
using appropriate statistical and other tools so that there is no confused statement in the inference
that the pesticide dosage enhanced yield, without quantifying the avoidable crop loss due to given
pesticide dose application against the target pests in the evaluation.
2.9
Statistical Analysis
The data thus obtained shall be subjected to appropriate transformations and analyzed using
standard experimental designs.
2.10
RESULT & CONCLUSION
2.10.1 Result
Results to be submitted in table format mentioning the statistical parameters of assessment
such CD.CV%, SD etc.
2.10.2 Inferences and Conclusion
Brief not on the trial observations and performance of the test substance based on the results
may be provided with critical assessment of tested pesticide efficacy on the target pest(s)
along with opinion and conclusion on its impact on the target species on natural enemies of
the given agro-climate in which the bioefficacy evaluation was undertaken. Any
observations on the upsurge of any other pests ( insects / mites / nematodes / disease causing
organisms ) may be specifically mentioned. The possible role of good agricultural practice
by the new pesticide formulation may be inferred. The conclusion shall mention the most
effective concentration / dose of the pesticide to suppress the target pests. The results may be
given only quantifying the avoidable crop loss due to given pesticide dose application
against the target pests in the evaluation.
III)
Bioefficacy-assessment evaluation of insecticides for phytophagous mites
Various species of phytophagous mites that infest crops are targeted for evaluation of acaricides /
insecticides. Each of these phytophagous mite groups has to be handled differently in bioefficacy
evaluation as their biology, metamorphic cycles and biology on the given crop species species
varies. Accordingly, the design of evaluation in the PD may be developed. The natural enemies of
these may be different from those of other sector pests and careful consideration of the type of
species involved ( based on Indian published literature ) for these may be drawn up for agroclimate where the bioefficacy evaluations are undertaken.
3.0
EXPERIMENTAL CONDITION
3.1
Crop, Variety and Target insect (Scientific names).
The name of the crop along with that of variety and target test organism (s), i.e. the target phytophagous mite pest
species may be mentioned in the PD.
3.2
Trial conditions
Field trial should be set up where target crop of moderate susceptibility to the target insect
species. Standard crop husbandry practices like application of FYM, fertilizers including
micronutrients and other maintenance treatments including pest management practices for pests
other than the target pest should be specified. Separate trials needs to be undertaken for
evaluation of the given formulation against defined pest species in the same crop, if these pests
are invading the crop together. Also ensure that that the targeted species is not influenced by
previous pesticide application. Trial should be carried out in different agro-climatic regions.
3.3
Design and layout of the trial
3.3.1
Treatments
Test product(s), reference product(s) and untreated control, arranged in a randomized block
design or any other statistically suitable design.
3.3.2
Plot size and replication
Net plot size at least 15 – 20 sq.m. per replication per treatment but according to the formulation
or application equipment, it may be necessary to use a larger plot size
Replicates – at least three, provided the error degrees of freedom are at least 12.
3.4
APPLICATION OF TREATMENT
3.4.1
Test product (s)
The formulated product under investigation is applied according to the PD. The appliance shall be
to facilitate uniform high volume application on the crop at its given age and canopy size.
3.4.2
Reference product
Registered product, if any / recommended product for the control of target insect pest.
3.4.3
Mode of application
Application should comply with good agricultural practices. Details on the spray equipment, type
of nozzle and water volume used at different stages of crop growth should be elucidated. Addition
of sticker, if any (for monsoon sprays) must be specified. Sticker should be tried in the check plot
with water spray.
3.4.3.1 Type of application
The type of application (e.g. foliar spray, soil incorporation, broad casting) as proposed by the
sponsor.
3.4.3.2 Time and frequency of application
The time and frequency of application will normally be specified by the manufacturer/ sponsor.
The time and frequency will normally depend on the pest species, & local environmental
conditions & pest ETL’S. The number of applications and the date of each application should be
recorded.
3.4.3.3 Doses and Volumes used
The product should be tested at the proposed dose and may also be used other doses. The dosage
will normally be expressed in g a.i./ha and formulation ml or gm per ha.
3.5 MODE OF ASSESSMENT, RECORDING & MEASUREMENTS
3.5.1
Weather data
3.5.1.1 Weather data during experimental duration for each location should be provided and co-related
with the field data that is generated on the disease incidence and plant mortality of each treatment
across replications. The inference shall be directed towards the influence of test substance in
reducing the target insect / mite damage.
3.5.1.2
Soil
Soil type should be mentioned where the crop is cultivated for such evaluations.
3.5.2
Observations / Assessments
3.5.2.1 Insect pest Assessments
The PD shall contain the details of the observations / assessments of treatment and the
frequency as well as the various scientific basis ( published literature reference ) for those.
The population of insect the target phytophagous mite pest species ( Jassids, Aphids,
Mealybugs-nymph & adult; Whiteflies-Nymphs ) is recorded on a suitable no. of randomly
selected plants/trees per replication.
Suitable no. of randomly selected leaves/panicles/twigs/inflorescence ( or target plant part )
per plant / tree are selected for observations.
Suitable method is used to count the insect population
Pre-treatment population of insect is recorded one day before the spray insecticide.
The post-treatment population of insect, is recorded at suitable intervals after each spray.
Run ‘t’-test between pre-count and post-count to see if the build up has been significant in check
plots at each date of observation.
Work out the % reduction in target insect population over control using Henderson & Tilton
formula :
Henderson & Tilton formula :
Corrected %
n in Co before treatment x n in T after treatment
Pod/fruit/ball/square/loculi = 1- ---------------------------------------------------------------- x 100
Damage reduction
n in Co after treatment x n in T before treatment
Where, n = no. of damaged pods/fruits/balls/squares/loculi; T = Treated; Co = Control
Reference: Henderson, C.F. and E. W. Tilton, 1955. Tests with acaricides against the brow
wheat mite, J. Econ. Entomol. 48:157-161.
Saps sucking insect pests of crops are generally early-season pests. These may be jassids, aphids, whitefly, or
tissue-lacerating pests as thrips. Each of these insect groups has to be handled differently in bioefficacy evaluation
as their biology, metamorphic cycles and biology on the given crop species varies. Often they may have the role
of disease vectors too. Accordingly, the design of evaluation in the PD may be developed. The natural enemies of
these may be different from those of other sector of pests and careful consideration of the type of species involved
( based on Indian published literature ) for these may be drawn up for agro-climate where the bioefficacy
evaluations are undertaken.
The mite population (motile stage) is recorded on randomly selected plants/trees/bushes per replication as per the
protocol fixed for the given species and crop.
Suitable no. of randomly selected leaves (or target plant part) per plant / tree are selected for observations.
Suitable approved method is used to count the mite population.
Pre-treatment population of mite is recorded one day before the spray of acaricide.
The post-treatment population of mite is recorded at suitable intervals after each spray.
Run ‘t’-test between pre-count and post-count to see if the build up has been significant in check plots.
Work out the % reduction in mite population over control using Henderson & Tilton formula :
Corrected %
n in Co before treatment x n in T after treatment
reduction in = 1- ----------------------------------------------------------------mite population
n in Co after treatment x n in T before treatment
x 100
Where, n = mite population; T = Treated; Co = Control
Reference: Henderson, C.F. and E. W. Tilton, 1955. Tests with acaricides against the brow
wheat mite, J. Econ. Entomol. 48:157-161.
3.6
Phyto-toxicity
Phyto-toxicity at ‘X’ and ‘2X’ dose shall be recorded.
Phytotoxicity Rating Scale (PRS)
Crop response/ Crop injury
Rating
0-00
0
1-10%
1
11-20%
2
21-30%
3
31-40%
4
41-50%
5
51-60%
6
61-70%
7
71-80%
8
81-90%
9
91-100%
10
Rating shall be recorded individually for yellowing, stunting, necrosis, epinasty
hyponasty etc.,
T
No
Treatment
Dose / ha
Days after 1st spray
g. a. i.
1
Formulation
3
5
7
10
1
2
3
4
3.7
Effect on Natural Enemies
The natural enemy spectrum of the target pest, as catalogued in the literature and perceived in the
region on the crop species in which this evaluation is done, may be recorded in the field collected
sample test insects and the impact of the targeted pesticide treatments in terms of mortality,
moribund condition, lack of natural enemy invasion unlike in untreated check plots may be
recorded.
3.7.1
Other new pests ( insects / mites / nematodes / diseases ) in the experiment
The investigator (evaluator) may look for the occurrence of any new pests ( insects / mites /
nematodes / diseases ) in each of the evaluation plots. The help of relevant specialists for each of
these may be taken and need to be recorded throughout the experiment period.
3.8
Crop Yield
The crop yield data may be recorded for each treatment and may be statistically analysed. The
conclusion on the treatment effect alone on enhancement of crop yield has to be brought out
using appropriate statistical and other tools so that there is no confused statement in the inference
that the pesticide dosage enhanced yield, without quantifying the avoidable crop loss due to given
pesticide dose application against the target pests in the evaluation.
3.9
Statistical Analysis
The data thus obtained shall be subjected to appropriate transformations and analyzed using
standard experimental designs.
3.10
RESULT & CONCLUSION
3.10.1 Result
Results to be submitted in table format
3.10.2 Inferences and Conclusion
Brief not on the trial observations and performance of the test substance based on the results
may be provided with critical assessment of tested pesticide efficacy on the target pest(s)
along with opinion and conclusion on its impact on the target species on natural enemies of
the given agro-climate in which the bioefficacy evaluation was undertaken. Any
observations on the upsurge of any other pests ( insects / mites / nematodes / disease causing
organisms ) may be specifically mentioned. The possible role of good agricultural practice
by the new pesticide formulation may be inferred. The conclusion shall mention the most
effective concentration / dose of the pesticide to suppress the target pests. The results may be
given only quantifying the avoidable crop loss due to given pesticide dose application
against the target pests in the evaluation.
IV)
Bioefficacy-assessment evaluation of insecticides for phytophagous mites
Introduction
The experiment is conducted in well-drained fertile and good quality soil of the research farm of the institution,
with all the normal recommended package of practice to raise the relevant crop for undertaking bioefficacy
evaluation of insecticides. The bioefficacy trial may be monitored by high-power group of scientist, nominated by
the Director of Research of the institution Heads of the relevant plant protection departments ( Entomology, Plant
Pathology / Nematology, senior experienced superannuated eminent scientists ( if needed ), along with
representation of the client industry. A checklist of protocols may be drawn up during each evaluation after
discussing with the relevant pesticide manufacturer ( client ) and the scientist ( evaluator ). The check-list may
draw up the High-power scientific group.
A project document (PD) is drawn up, with due identification number and authentication of the institution, containing the
under-mentioned details, before the commencement of the experiment and approved by the competent authority of the
institution. The data generated may be recorded in permanent work dairies and need to be preserved in the institution for
reasonable duration till the pesticide registration process is completed.
Various vertebrate pests such as rodents ( rats, bandicoots, squirrels etc. ) , blue bulls, wild boars, porcupines, monkeys
depredatory birds and others are well known to damage standing crops as well as commodity in storage in farms, markets
and the like. Effective pesticides to suppress their build up or to repel and cause abnoxious reactions would save the
agricultural farms from their damage to crops in the field as well as storage. Many rodenticides and other chemicals are
used for such purposes. Standard protocols are essential to evaluate the newly developed chemistries and their formulations
for suppression of vertebrate pests.
V. *General guidelines for the evaluation of bio-efficacy of pesticide substances
Presently, the pesticide industry utilizes the laboratory ( under the given state agricultural university (SAU) or crop institute
under the Indian Council of Agricutlural Research ( ICAR or any other similar institution ) of any agro-ecological region of
the country ) may take up the evaluation through the approved and well-planned research project, with due identification
number and authentication of the institution.
Central Insecticide Board may encourage the organization(s) with such desirable field and laboratory facility to accredit
that with the globally-recognized standards organizations for the purpose for which the service is offered.
The organization where the bio-efficacy test / evaluation facility is available may intimate the Registration Committee of
the Central Insecticide Board ( RC of CIB ) about is readiness and willingness to conduct, record and share the data, so
evaluated with the R&D system of the pesticide industry as well as, if need arises, with the RC of CIB. They may mention
the details of the various laboratory and field facility including manpower that it has to take up, undertake, analyse, infer
and report the results and available data management system of evaluation with regard to the given pesticide substance
against the target pest (s) in crops.
Such information from various institutions shall be kept in the web-site of Central Insecticide Board to facilitate the
prospective companies to undertake the evaluation of the pesticide substances for the desired bio-efficacy against the target
pest(s) in the designated crops.
Periodic assessment and update of this information may be provided by the respective organization(s), which is keen to
take up this service for the pesticide industry. The pesticide industry through its association(s0 may facilitate this so that a
minimum number of designated laboratories are available for this prupose.
The organization(s) may spell out the capacity / expertise in undertaking such evaluations in order to facilitate the pesticide
industry to choose from the available list of specific purpose for which it may desire to charter the service in order to
evaluate its target pesticide substance.
Such R&D organizations may also undertake original research to publish in peer-reviewed globally recognized research
scientific journals methods of bio-efficacy evaluations of pesticide substance(s) against new and emerging pest(s) as well
the refined and upgrade methods for the existing pest(s) of crops. The organization may also publish its research findings
on these evaluations as and when they deem fit.
Ann: A (ii)
ANNEXURE- II
Insect pest assessment ( Time of application : Based on ETL assessment )
Name of the insect
Formula
Helicoverpa armigera
As per Henderson and tilton formula
Tobacoo caterpillar
As per Henderson and tilton formula
Pink boll worm
As per Henderson and tilton formula
Stem borer
As per IRRI
As per IRRI
Leaf folder, caseworm, gall midge
As per IRRI
Whorl maggot
Fruit and shoot borer
As per Abbots formula
DBM
As given
BPH / WBPH
Tap 4 rice hills vigorously count the number of plant
hoppers floating and jumping on the water surface
between the four hills. Ten such places are to be
observed at random within each plot.
GLH
Tap 4 rice hills vigorously count the number of plant
hoppers floating and jumping on the water surface
between the four hills. Ten such places are to be
observed at random within each plot.
0=0, 1=1-5, 3=5-10, 5=11-15, 7=51 to 100, 9=more than
100 ( assessment should be made between 1-3 days and
1 week after treatment.
DBM
Count the number of caterpillars of different ages on all
plants in each plot.
Assessment :1-3 days after treatment
2nd assessment:7-14 days after treatment
Mealy bugs, aphids, scale insects
As given
Whiteflies
Observation to be made on 4 top leaves from 10
randomly selected plants in each treatment. Preassessment and Post-assessment observations to be
taken.
Thrips
Observation to be made on 15 leaves ( Top 5, Middle 5
and bottom 5 ) 10 randomly selected plants in each
treatment. Pre-assessment and Post-assessment
observations at an interval of one week may be taken.
Other insect pest ( mites / nematodes )
As given
Other insect pest of major crops
Not given
Household insect pest
Not given
Stored grain insect pest
Not given
STANDARD EVALUATION SYSTEM
BIOEFFICACY – ASSESSMENT EVALUATION OF RODENTICIDE /
PESTICIDE AGAINST VERTEBRATE PEST IN CROPS
TEST PROTOCOL FOR BIO-EFFICACY EVALUATION OF RODENTICIDES
Introduction
The experiment is conducted in well-drained fertile and good quality soil of the research farm of the institution,
with all the normal recommended package of practice to raise the relevant crop for undertaking bio-efficacy
evaluation of insecticides. The bio-efficacy trial may be monitored by high-power group of scientist, nominated by
the Director of Research of the institution Heads of the relevant plant protection departments ( Entomology, Plant
Pathology / Nematology, senior experienced superannuated eminent scientists ( if needed ), along with
representation of the client industry. A checklist of protocols may be drawn up during each evaluation after
discussing with the relevant pesticide manufacturer ( client ) and the scientist ( evaluator ). The check-list may
draw up the High-power scientific group.
A project document (PD) is drawn up, with due identification number and authentication of the institution,
containing the under-mentioned details, before the commencement of the experiment and approved by the
competent authority of the institution. The data generated may be recorded in permanent work dairies and need
to be preserved in the institution for reasonable duration till the pesticide registration process is completed.
PROTOCOLS FOR EVALUATION OF RODENTICIDES IN LABORATORY AND FIELD LABORATORY
AND FIELD METHODOLOGY FOR EVALUATION BIOEFFICACY OF RODENTICIDES.
Ever since the technology is developed for rodent management in the developed countries, several guidelines were
issued by Environmental Protection Agency ( 1982 ), European Plant Protection Organization ( 1975 ), Food and
Agricultural Organization of United Nations ( 1992 ) for the evaluation of the effecti of different rodenticides
currently considered by different countries. The test method has been set up in such a way that they form a series
of stages in development of rodenticides from their preliminary screening as a potentially useful chemical to the
final evaluation in the field in some suitable formulation. Thus, a scientist may select the tests appropriate to their
needs. Keeping this in view, procedure for evaluating the biological efficacy of various rodenticides have been
prepared by the Scientific Advisory Committee ( SAC ) on Rodenticides constituted by Central Insecticides Board,
Government of India. While preparing this, the guidelines issued by the above International Agencies have been
taken into consideration. The chairman of the committee, being ICAR Project Coordinator AICRP on Rodent
Control has obtained the opinion of Pricipal Investigators ( PIs ) of ten ICAR Project Centres before circulating
the above Scientific Advisory Committee members. The suggestion of the PIs as well as SAC members has been
incorporated. Prof. Iswar Prakash an eminent and leading Rodent Specialist of the Country also offered his
considered opinion, which has also been incorporated. Broadly, the procedure involves10 Laboratory trials
initially followed by ii) field bioefficacy trials as given below :
A. Procedure for screening rodenticides in laboratory conditions
In all the laboratory evaluations, the following parameters may be followed. The field rodents should be preferably
trapped from infestation without recent history of rodenticide treatment and should be acclimatized the the
laboratory cages for 10 days before the experimentation. Constant daily food intake may be taken as completion
of acclimatization. The dimensions of the cages may be 45x30x30 cm. Constant laboratory conditions are to be
maintained. The room temperature should be between 19-23oC , relative humidity of 40-70% with a light
phtoperiod of 12h. light / 12-h. dark. During the acclimatization, plain baits, which are normally staple food of the
area and water, should be provided ad libitum. In each experimental design a minimum of six experimental adult
animals with more or less same body weight are to be used. Whenever the outcome of any particular test is
recorded as critical, at least 20 animals may be employed using equal number of each sex.
The animals are to be retained for at least two weeks after poison treatment. Dead animals should be autopsied to
know the reason for the death. The bait consumption and the mortality should be recorded every day. During the
eriod of experimentation, disturbance in the laboratory is to be avoided.
1. Test for acute toxicity
It is an important principle that a minimum quantity of toxicant is required to mix in the bait material to avoid
danger to the non-target species and also to get adequate bait palatability by the test species. The lethal dose
otherwise called as toxicity is expressed as a percentage of the mortality of a test species to a given mg/kg body
weight. Usually, the toxicity of a chemical is expressed in LD50 since greater confidence may be attached to the
estimates in the middle range rather than at the extreme. To arrive at the LD50, six animals are to be individually
caged and permitted for acclimatization on the laboratory conditions. The dosage are generally selected in
arithmetic progression 50,100,150,200 mg a.i./kg etc. Solvent may be used as per the label specification of the
toxicant. If the mid-point can be estimated, the progression will be centered there and work both ways. An
example zinc phosphide concentration for Mus musculus at 250 mg/kg is 150, 200, 300, 350 with a mid point of
250 mg/kg. The dosage for each individual animal is computed below :
Mg/kg= Weight of the toxicant for individual animal
Average weight of the animal (g)
For example for house mouse weighing 15g with a concentration of 250 mg/kg:
250/1000=x/15 or 1000x =250x15 or x = 3.750
X=3.75 mg is the quantity of the toxicant needed
The toxicant should be finally ground in a mortar and dissolved in water or in any suitable solvent. At different
concentrations, each solution thus prepared be administered to six caged animals as per the dosage calculated
depending upon their body weight. Water and laboratory food are to be provided at ad libitum and the animals be
inspected for every hour thereafter. Symptoms and time on onset of illness and time of death should be recorded.
Percentage mortality may be plotted against dosage of log probability paper and the LD50 be estimated using
probit analysis.
2. No choice feeding test
Having obtained an approximate LD50 for the new toxicant, it is appropriate to observe results of the tests in
which the poison is offered to the rodent without alternative food and to compare the mortality achieved with other
conventional rodenticides. It is suggested that 8 times the LD50 value can be used as the starting point. Taking with
an average sized rodents, each consume about 10 percent of its weight per day, for zinc phosphide at 250 mg/kg
LD50, the consumption will be as follows :
250 mg/kg LD50, the comutation will be as follows :
8x=
0.25 g/kg
= 1.3% zinc phosphide in bait
1.5g ( 10% of 15g ) body weight
In all the tests a minimum of 5 adult animals belonging to each sex at nearly equal body weight are to be used.
Exposure period will depend as per label specification. The dailly consumption of the poison bait be measured
during the experimental period for each dosage. The mortality is to be noted along with the time of onset of the
illness.
3. Choice feeding tests
The next stage in evaluation of the toxicant is to test the palatability of the toxicant in bait offered with a choice.
Plain bait is to be offered as a coice food to the test rodents and daily consumption is to be measured. The principal
criterion for determining the palatability will be total intake of poisoned bait in comparison with plain baits, when
other test conditions are maintainded same as for the earlier tests.
Thus above tests are designed to screen the experimental toxicant in laboratory conditions followed later by field
screening trails.
B. Procedure for screening rodenticides in field conditions
1.0
EXPERIMENTAL CONDITION
1.1
Selection of test site : the field may be selected covering major cropping systems of the area /
region. The cropping system may include rice, wheat, or sugarcane based with different crops in the selected area.
1.2
Trial conditions : The field should be infested as uniformly as possible by the rodent pests. This
can be determined with pre baiting census ( with bait stations for about 4-5 days pre baiting ) or trapping ( for
about 3-4 days ) or active burrow or damage incidence. Cultural conditions should be uniform for all plots of
trials and should conform to local agricultural conditions.
Trial should be carried out in different regions with distinct environmental conditions and preferably in two
different seasons.
The evaluation may be conducted during lien period, tillering or pre flowering stages ensuring a minimum rodent
infestation of 100 rodent burrows in the selected area.
1.3
Design and layout of the trial
1.3.1
Treatments
The application of the treatments should be done in burrows, on bunds as well as in crop fields.
When burrows are not visible or inaccessible and during rainy season, bait stations at a distance
of 10m apart should be placed on the periphery of the selected area.
1.3.2
Plot size and replication
Net plot size may be 1hectares of compact field in an area of reasonably synchronized planting.
In hilly area, 0.5 hectare may be taken. The total infestation should be at least 25 burrows/ha in
the selected area. At least three replication ( which may be three different locations ) may be
used.
2.0
APPLICATION OF TREATMENT
2.1
Test product (s)
The test product is the formulated product under investigation.
2.2
Reference product
Registered product known to be satisfactory for the control of rodent pests and may be close to
the test product depending on the aim of the trial.
1.2.3
Mode of application
Application should comply with good agricultural practices. Details on the spray equipment, type
of nozzle and water volume used at different stages of crop growth should be elucidated. Addition
of sticker, if any (for monsoon sprays) must be specified. Sticker should be tried in the check plot
with water spray.
2.3
Type of application Application should comply with Good Agricultural Practices
2.3.1.
Type of application : The bait may be applied in paper packets inside the burrows. The quantity
may be as per label specifications. Where the burrows cannot be located, bait stations like pieces
of bamboo, or pvc pipes or coconut shells or disposed scooter tyres may be used.
Type of equipment used : Any equipment used for placement should be the ones in current use.
Plastics pipes for fumigants and tracking box for tracking dusts may be used. Bait stations as
above may be also be used.
Time and frequency of application : The number of application(s), date of each application,
stage of the crop at each application should be recorded. If baits are used, baiting frequency as
recommended on the (proposed) label be indicated.
Doses and Volumes used : The product should be treated at recommended dosage as well as at
other doses with different concentrations, frequency, and / or changing number of bait stations.
2.3.2.
2.3.3.
2.3.4.
3.0 MODE OF ASSESSMENT, RECORDING & MEASUREMENTS
3.1 Meteorological and edaphic data
3.1.1
Meteorological data : On the date of application the meteorological data should be recorded,
which often affect the quality and persistence of the treatment. Any significant change in weather should
be noted. All data should be preferably recorded on the trial site, but may be obtained from a nearby
meteorological station.
3.1.2
Edaphic data : Type of soil and other edaphic parameters should be recorded while evaluating
fumigants.
3.2 Type, Time and frequency of assessment
3.2.1 Type : Assessment should be made in any three of the following methods :
Nut damage incidence
Rate of rodent infestation
3.3
Time and frequency : Assessment should be done prior to trial and one week after the (last)
application of the product for acute poison and two weeks after application for anticoagulants.
Effect on non-target organisms : Any effect on non-target prganism should be recorded. Any
observed effects on human safety should be recorded.
3.4
4. Results : Results should be reported in a systematic form and report should include an analysis and
evaluation. Original ( raw ) data should be available. Statistical analysis should be done using standard
methods which should be indicated.
C. Procedure for screening rodenticides in field conditions ( Coconut )
1.0
EXPERIMENTAL CONDITION
1.1
Selection of test site : the field may be selected covering major cropping systems of the area /
region. The cropping system may include rice, wheat, or sugarcane based with different crops in the selected area.
1.2
Trial conditions : The field should be infested as uniformly as possible by the rodent pests. This
can be determined with pre baiting census ( with bait stations for about 4-5 days pre baiting ) or trapping ( for
about 3-4 days ) or active burrow or damage incidence. Cultural conditions should be uniform for all plots of
trials and should conform to local agricultural conditions.
Trial should be carried out in different regions with distinct environmental conditions and preferably in two
different seasons.
The evaluation may be conducted during lien period, tillering or pre flowering stages ensuring a minimum rodent
infestation of 100 rodent burrows in the selected area.
1.3
Design and layout of the trial
1.3.1
Treatments
The application of the treatments should be done in burrows, on bunds as well as in crop fields.
When burrows are not visible or inaccessible and during rainy season, bait stations at a distance
of 10m apart should be placed on the periphery of the selected area.
1.3.2
Plot size and replication
Net plot size may be 1hectares of compact field in an area of reasonably synchronized planting.
In hilly area, 0.5 hectare may be taken. The total infestation should be at least 25 burrows/ha in
the selected area. At least three replication ( which may be three different locations ) may be
used.
2.0
APPLICATION OF TREATMENT
2.1
Test product (s)
The test product is the formulated product under investigation.
2.2
Reference product
Registered product known to be satisfactory for the control of rodent pests and may be close to
the test product depending on the aim of the trial.
1.2.3
Mode of application
Application should comply with good agricultural practices. Details on the spray equipment, type
of nozzle and water volume used at different stages of crop growth should be elucidated. Addition
of sticker, if any (for monsoon sprays) must be specified. Sticker should be tried in the check plot
with water spray.
2.3
Type of application Application should comply with Good Agricultural Practices
2.3.5.
Type of application : The bait may be applied in paper packets inside the burrows. The quantity
may be as per label specifications. Where the burrows cannot be located, bait stations like pieces
of bamboo, or pvc pipes or coconut shells or disposed scooter tyres may be used.
Type of equipment used : Any equipment used for placement should be the ones in current use.
Plastics pipes for fumigants and tracking box for tracking dusts may be used. Bait stations as
above may be also be used.
Time and frequency of application : The number of application(s), date of each application,
stage of the crop at each application should be recorded. If baits are used, baiting frequency as
recommended on the (proposed) label be indicated.
Doses and Volumes used : The product should be treated at recommended dosage as well as at
other doses with different concentrations, frequency, and / or changing number of bait stations.
2.3.6.
2.3.7.
2.3.8.
3.0 MODE OF ASSESSMENT, RECORDING & MEASUREMENTS
3.1 Meteorological and edaphic data
3.1.1
Meteorological data : On the date of application the meteorological data should be recorded,
which often affect the quality and persistence of the treatment. Any significant change in weather should
be noted. All data should be preferably recorded on the trial site, but may be obtained from a nearby
meteorological station.
3.1.3
Edaphic data : Type of soil and other edaphic parameters should be recorded while evaluating
fumigants.
3.2 Type, Time and frequency of assessment
3.4.1 Type : Assessment should be made in any three of the following methods :
Bait acceptance and census baiting
Damage incidence
Number of active burrows
Tacking with tracking tile or dust
Trapping index using trap line at a distance of 10m apart
3.5
Time and frequency : Assessment should be done prior to trial and one week after the (last)
application of the product for acute poison and two weeks after application for anticoagulants.
Effect on non-target organisms : Any effect on non-target prganism should be recorded. Any
observed effects on human safety should be recorded.
3.6
4. Results : Results should be reported in a systematic form and report should include an analysis and
evaluation. Original ( raw ) data should be available. Statistical analysis should be done using standard
methods which should be indicated.
D. Procedure for screening rodenticides in storage / residential conditions
1.0
EXPERIMENTAL CONDITION
1.1
Selection of test site : The site may be selected covering general storage godowns or residential
premises with commodity storage . Since rodents normally inhabit premises with commodity storage, storage
godowns are ideal for evaluation.
1.2
Trial conditions : The storage units should be infested as uniformly as possible by the rodent
pests. This can be determined with pre baiting census ( with bait stations for about 4-5 days pre baiting ) or
trapping ( for about 3-4 days ) or active burrow or damage incidence. Cultural conditions should be uniform for
all plots of trials and should conform to local agricultural conditions.
Trial should be carried out in different regions with distinct environmental conditions and preferably in two
different seasons.
The evaluation may be conducted during maximum storage period.
1.3
Design and layout of the trial
1.3.1
Treatments
The application of the treatments should be done in bait stations inside the premises, burrows
present outside the premises.
1.3.2
Plot size and replication
Net plot size may be 1 stoarge unit in an area with visible rodent movement and with a total rate
of 100 percent tracking and /or significant bait consumption in census bait method. At least three
replications may be used in similar rural / urban situations.
2.0
APPLICATION OF TREATMENT
2.1
Test product (s)
The test product is the formulated product under investigation.
2.2
Mode of application : Application should comply with Good Agricultural Practices
2.2.1 Type of application : The bait may be applied in paper packets inside the burrows. The quantity
may be as per label specifications. Where the burrows cannot be located, bait stations like pieces of
bamboo, or pvc pipes or coconut shells or disposed scooter tyres may be used.
2.2.2
2.2.3
2.2.4
Type of equipment used : Any equipment used for placement should be the ones in current use.
Plastics pipes for fumigants and tracking box for tracking dusts may be used. Bait stations as
above may be also be used.
Time and frequency of application : The number of application(s), date of each application,
stage of the crop at each application should be recorded. If baits are used, baiting frequency as
recommended on the (proposed) label be indicated.
Doses and Volumes used : The product should be treated at recommended dosage as well as at
other doses with different concentrations, frequency, and / or changing number of bait stations.
3.0 MODE OF ASSESSMENT, RECORDING & MEASUREMENTS
3.1 Meteorological and edaphic data
3.1.1
Meteorological data : On the date of application the meteorological data should be recorded,
which often affect the quality and persistence of the treatment. Any significant change in weather should
be noted. All data should be preferably recorded on the trial site, but may be obtained from a nearby
meteorological station.
3.1.4
Edaphic data : Type of soil and other edaphic parameters should be recorded while evaluating
fumigants.
3.2 Type, Time and frequency of assessment
3.2.1 Type : Assessment should be made in any three of the following methods :
Bait acceptance and census baiting
Tacking with tracking tile or dust
Trapping index
3.2.2
3.2.3
Time and frequency : Assessment should be done prior to trial and one week after the (last)
application of the product for acute poison and two weeks after application for anticoagulants. If
pulsed baiting is used, assessment should be made 15 days be made 15 days after the first pulsing and
15 days after second pulsing.
Effect on non-target organisms : Any effect on non-target prganism should be recorded. Any
observed effects on human safety should be recorded.
4. Results : Results should be reported in a systematic form and report should include an analysis and
evaluation. Original ( raw ) data should be available. Statistical analysis should be done using standard
methods which should be indicated.
*General guidelines for the evaluation of bio-efficacy of pesticide substances
Presently, the pesticide industry utilizes the laboratory ( under the given state agricultural university (SAU) or crop institute
under the Indian Council of Agricutlural Research ( ICAR or any other similar institution ) of any agro-ecological region of
the country ) may take up the evaluation through the approved and well-planned research project, with due identification
number and authentication of the institution.
Central Insecticide Board may encourage the organization(s) with such desirable field and laboratory facility to accredit
that with the globally-recognized standards organizations for the purpose for which the service is offered.
The organization where the bio-efficacy test / evaluation facility is available may intimate the Registration Committee of
the Central Insecticide Board ( RC of CIB ) about is readiness and willingness to conduct, record and share the data, so
evaluated with the R&D system of the pesticide industry as well as, if need arises, with the RC of CIB. They may mention
the details of the various laboratory and field facility including manpower that it has to take up, undertake, analyse, infer
and report the results and available data management system of evaluation with regard to the given pesticide substance
against the target pest (s) in crops.
Such information from various institutions shall be kept in the web-site of Central Insecticide Board to facilitate the
prospective companies to undertake the evaluation of the pesticide substances for the desired bio-efficacy against the target
pest(s) in the designated crops.
Periodic assessment and update of this information may be provided by the respective organization(s), which is keen to
take up this service for the pesticide industry. The pesticide industry through its association(s0 may facilitate this so that a
minimum number of designated laboratories are available for this prupose.
The organization(s) may spell out the capacity / expertise in undertaking such evaluations in order to facilitate the pesticide
industry to choose from the available list of specific purpose for which it may desire to charter the service in order to
evaluate its target pesticide substance.
Such R&D organizations may also undertake original research to publish in peer-reviewed globally recognized research
scientific journals methods of bio-efficacy evaluations of pesticide substance(s) against new and emerging pest(s) as well
the refined and upgrade methods for the existing pest(s) of crops. The organization may also publish its research findings
on these evaluations as and when they deem fit.
TEST PROTOCOL FOR EFFICACY EVALUATION OF BIO-INSECTICIDES
1.0
EXPERIMENTAL CONDITION
1.1
Crop, Variety and Target insect (Scientific names).
1.2
Trial conditions
Field trial should be set up where target crop of moderate susceptibility to the target insect
species. Standard crop husbandry practices like application of FYM, fertilizers including
micronutrients and other maintenance treatments including pest management practices for pests
other than the target pest should be specified. Separate trials needs to be undertaken for
evaluation of the given formulation against defined pest species in the same crop, if these pests
are invading the crop together. Also ensure that that the targeted species is not influenced by
previous pesticide application.Trial should be carried out in different agroclimatic regions.
1.3
Design and layout of the trial
1.3.1
Treatments
Test product(s), reference product(s) and untreated control, arranged in a randomized block
design or any other statistically suitable design.
1.3.2
Plot size and replication
Net plot size at least 15 – 20 sq.m. per replication per treatment but according to the formulation
or application equipment, it may be necessary to use a larger plot size
Replicates – at least three, provided the error degrees of freedom are at least 12.
2.0
APPLICATION OF TREATMENT
2.1
Test product (s)
The formulated product under investigation.
2.2
Reference product
Registered product, if any / recommended product for the control of target insect pest.
2.3
Mode of application
Application should comply with good agricultural practices. Details on the spray equipment, type
of nozzle and water volume used at different stages of crop growth should be elucidated. Addition
of sticker, if any (for monsoon sprays) must be specified. Sticker should be tried in the check plot
with water spray.
2.3.1
Type of application
The type of application (e.g. spray, soil incorporation, broad casting) as proposed by the sponsor.
2.3.2
Time and frequency of application
The time and frequency of application will normally be specified by the manufacturer/ sponsor.
The time and frequency will normally depend on the pest species, & local environmental
conditions & pest ETL’S. The number of applications and the date of each application should be
recorded.
2.3.3
Doses and Volumes used
The product should be tested at the proposed dose and may also be used other doses. The dosage
will normally be expressed in g a.i./ha and formulation ml or gm per ha.
3.0 MODE OF ASSESSMENT, RECORDING & MEASUREMENTS
3.1
Weather data
3.1.1 Weather data during experimental duration for each location should be provided
3.1.2
Soil
Soil type should be mentioned
3.2
Observations / Assessments
3.2.1
Insect pest Assessments
3.2.1.1 Helicoverpa armigera
Larval count before and after treatment be made and evaluated statistically for significance of
treatment effects. The no. of damaged pods/fruits/balls/squares/loculi is recorded on suitable no.
of randomly selected plants per replication.
The observations are taken either at suitable interval or each picking or at maturity depending on
the crop.
Work out the % pod/fruit/ball/square/loculi damage reduction over control using either Abbott’s
formula or Henderson & Tilton formula :
Abbott’s formula :
Corrected %
Pod/fruit/ball/square/loculi =
Damage reduction
n in T after treatment
1- ------------------------------
n in Co after treatment
x 100
Where, n = no. of damaged pods/fruits/balls/squares/loculi; T = Treated; Co = Control
Reference: Abbott, W.S. (1925). A method of computing the effectiveness of an insecticide. J.
Econ. Entomol.; 18:265-267.
Henderson & Tilton formula :
Corrected %
n in Co before treatment x n in T after treatment
Pod/fruit/ball/square/loculi = 1- ---------------------------------------------------------------- x 100
Damage reduction
n in Co after treatment x n in T before treatment
Where, n = no. of damaged pods/fruits/balls/squares/loculi; T = Treated; Co = Control
Reference: Henderson, C.F. and E. W. Tilton, 1955. Tests with acaricides against the brow
wheat mite, J. Econ. Entomol. 48:157-161.
3.2.1.2 Spodoptera litura
The larval population of insect is recorded on suitable no. of randomly selected plants per
replication.
Pre-treatment larval population of insect is recorded one day before the spray of insecticide.
The post-treatment larval population of insect is recorded at suitable interval after each spray.
Work out the % reduction in larval population over control using Henderson & Tilton formula :
Corrected %
reduction in
=
larval population
n in Co before treatment x n in T after treatment
1- ----------------------------------------------------------------n in Co after treatment x n in T before treatment
x 100
Where, n = larval population; T = Treated; Co = Control
Reference: Henderson, C.F. and E. W. Tilton, 1955. Tests with acaricides against the brow
wheat mite, J. Econ. Entomol. 48:157-161.
3.2.1.3 Leaf Folder (Rice - Cnaphalocrosis medinalis)
Observations are recorded before & after each application of insecticides.
Record the number of freshly damaged or folded leaves/hill
Observe ten hills per replication
Work out per cent damage using IRRI scale.
Scale
Score
Damage
0
No Damage
1
1 – 10%
3
11-20%
5
21 – 35%
7
36 – 50%
9
51% & above
Source : Standard Evaluation System for Rice, IRRI, 1996.
OR
3.2.1.3 Leaf Folder (Rice - Cnaphalocrosis medinalis)
The no. of freshly damaged leaves/hill are recorded on suitable no. of randomly selected hills per
replication.
Pre-treatment freshly damaged leaves are recorded one day before the spray of insecticide.
The post-treatment freshly damaged leaves are recorded at suitable interval after each spray.
Work out the % reduction in freshly damaged leaves over control using Henderson & Tilton
formula :
Corrected %
Reduction in
n in Co before treatment x n in T after treatment
Freshly damaged = 1- ----------------------------------------------------------------Leaves
n in Co after treatment x n in T before treatment
x 100
Where, n = no. of freshly damaged leaves; T = Treated; Co = Control
Reference: Henderson, C.F. and E. W. Tilton, 1955. Tests with acaricides against the brow
wheat mite, J. Econ. Entomol. 48:157-161.
3.2.1.4 Stem Borer (Paddy Stem Borer - Scirpophaga incertulas)
Dead Hearts : Observe 10 hills/ plot at maximum tillering to stem elongation stage and work out
percent damage (based on tiller count) using IRRI scale.
White Earheads : Observe 10 hills per plot at dough stage & work out percent damage (based on
productive tillers count) using IRRI scale.
Scale
Score
Damage
0
No Damage
1
1 – 10%
3
11-20%
5
21 – 30%
7
31 – 60%
9
61% & above
Source : Standard Evaluation System for Rice, IRRI, 1996.
OR
3.2.1.4 Stem Borer (Paddy Stem Borer - Scirpophaga incertulas)
The no. of dead-hearts/white-earheads per hill are recorded on suitable no. of randomly selected
hills per replication.
The post-treatment dead-hearts are recorded at suitable interval after each spray.
The post-treatment white-earheads are recorded at dough stage.
Work out the % dead-heart/white-earhead reduction over control using Abbott’s formula formula
Corrected %
Dead-heart/White-earhead
Damage reduction
=
n in T after treatment
1- ------------------------------
x 100
n in Co after treatment
Where, n = no. of Dead-heart/White-earhead; T = Treated; Co = Control
Reference: Abbott, W.S. (1925). A method of computing the effectiveness of an insecticide. J.
Econ. Entomol.; 18:265-267.
3.2.1.5 Fruit & Shoot Borer (Okra - Earias vitella; Brinjal – Leucinodes arbonalis)
The no. of damaged fruits/shoots are recorded on suitable no. of randomly selected plants per
replication.
Observations on damaged shoots are taken at suitable interval after each spray.
Observations on damaged fruits are taken at each picking.
Work out the % shoot/fruit damage reduction over control using Abbott’s formula :
Abbott’s formula :
Corrected %
Shoot/fruit
Damage reduction
=
n in T after treatment
1- ------------------------------
x 100
n in Co after treatment
Where, n = no. of damaged fruits/shoots; T = Treated; Co = Control
Reference: Abbott, W.S. (1925). A method of computing the effectiveness of an
insecticide. J. Econ. Entomol.; 18:265-267.
3.2.1.6 Diamond Back Moth, DBM (Plutella xylostella)
The larval population of insect is recorded on suitable no. of randomly selected plants per
replication.
Pre-treatment larval population of insect is recorded one day before the spray of insecticide.
The post-treatment larval population of insect is recorded at suitable interval after each spray.
Work out the % reduction in DBM larval population over control using Henderson & Tilton
formula :
Corrected %
n in Co before treatment x n in T after treatment
Reduction in = 1- -----------------------------------------------------------------
DBM popul.
x 100
n in Co after treatment x n in T before treatment
Where, n = no. of DBM larvae; T = Treated; Co = Control
Reference: Henderson, C.F. and E. W. Tilton, 1955. Tests with acaricides against the brow
wheat mite, J. Econ. Entomol. 48:157-161.
3.2.1.7 Sucking Pest (Jassids / Aphids / Thrips / Mealybugs / Whiteflies)
The population of insect (Jassids, Aphids, Thrips, Mealybugs - nymph & adult; Whiteflies –
Nymphs) is recorded on suitable no. of randomly selected plants/ trees per replication.
Suitable no. of randomly selected leaves/panicles/twigs/inflorescence (or target plant part) per
plant / tree are selected for observations.
Suitable method is used to count the insect population.
Pre-treatment population of insect is recorded one day before the spray of insecticide.
The post-treatment population of insect, is recorded at suitable intervals after each spray.
Work out the % reduction in target insect population over control using Henderson & Tilton
formula :
Corrected %
n in Co before treatment x n in T after treatment
Insect Population = 1- ----------------------------------------------------------------reduction
n in Co after treatment x n in T before treatment
x 100
Where, n = insect population; T = Treated; Co = Control
Reference: Henderson, C.F. and E. W. Tilton, 1955. Tests with acaricides against the brow
wheat mite, J. Econ. Entomol. 48:157-161.
3.2.1.8 Mites
The mite population (motile stage) is recorded on randomly selected plants/trees/bushes per
replication as per the protocol fixed for the given species and crop.
Suitable no. of randomly selected leaves (or target plant part) per plant / tree are selected for
observations.
Suitable approved method is used to count the mite population.
Pre-treatment population of mite is recorded one day before the spray of acaricide.
The post-treatment population of mite is recorded at suitable intervals after each spray.
Run ‘t’-test between pre-count and post-count to see if the build up has been significant in check
plots.
Work out the % reduction in mite population over control using Henderson & Tilton formula :
Corrected %
n in Co before treatment x n in T after treatment
reduction in = 1- ----------------------------------------------------------------mite population
n in Co after treatment x n in T before treatment
x 100
Where, n = mite population; T = Treated; Co = Control
Reference: Henderson, C.F. and E. W. Tilton, 1955. Tests with acaricides against the brow
wheat mite, J. Econ. Entomol. 48:157-161.
3.2.2
Phyto-toxicity
Phyto-toxicity at ‘X’ and ‘2X’ dose shall be recorded.
Phytotoxicity Rating Scale (PRS)
Crop response/ Crop injury
Rating
0-00
0
1-10%
1
11-20%
2
21-30%
3
31-40%
4
41-50%
5
51-60%
6
61-70%
7
71-80%
8
81-90%
9
91-100%
10
Rating shall be recorded individually for yellowing, stunting, necrosis, epinasty
hyponasty etc.,
T
Treatment
No
Dose / ha
Days after 1st spray
g. a. i.
1
Formulation
3
5
7
10
1
2
3
4
3.2.3
Effect on Natural Enemies
3.2.4
Yield
Yield data shall be given.
3.2.5
Statistical Analysis
The data thus obtained shall be subjected to appropriate transformations and analyzed using
standard experimental designs.
4.0 RESULT & CONCLUSION
4.1
Result
Results to be submitted in table format
4.2
Conclusion
Brief note on the trial observations and performance of the test substance.
STANDARD EVALUATION SYSTEM
BIOEFFICACY – ASSESSMENT EVALUATION OF BIOEFFICACY OF
PLANT PATHOGENS IN CROPS
TEST PROTOCOL FOR EFFICACY EVALUATION OF FUNGICIDES AGAINST
VARIOUS FUNGAL PATHOGENS
Bioefficacy evaluation of pesticide substances as formulations against plant pathogens ( fungi, bacteria, virus and
other relevant ones ) in the designated crop would enable the identification of apt dose of the formulation for
effective suppression the plant disease infection organ(s). The academic evaluation in designated research
institutions under both laboratory and field situations would follow the under-mentioned protocol and produce
results that is useful for considering for pesticide registration.
Before taking up field evaluation, it is desirable to undertake laboratory evaluation of the candidate pesticide
substance following standard internationally accepted protocol. The dosage arrived at from from such laboratory
evaluation, as machted up with the dosage proposed by the sponsor of the evaluation, shall be considered for field
evaluation of candidate disease causing organism(s).
The field experiment is conducted in well-drained fertile and good quality soil of the research farm of the
institution, with all the normal recommended package of practice to raise the relevant crop for undertaking
bioefficacy evaluation of insecticides. The bioefficacy trial may be monitored by high-power group of scientist,
nominated by the Director of Research of the institution Heads of the relevant plant protection departments (
Entomology, Plant Pathology / Nematology, senior experienced superannuated eminent scientists ( if needed ),
along with representation of the client industry. A checklist of protocols may be drawn up during each evaluation
after discussing with the relevant pesticide manufacturer ( client ) and the scientist ( evaluator ). The check-list may
draw up the High-power scientific group.
A project document (PD) is drawn up, with due identification number and authentication of the institution, containing the
under-mentioned details, before the commencement of the experiment and approved by the competent authority of the
institution. The data generated may be recorded in permanent work dairies and need to be preserved in the institution for
reasonable duration till the pesticide registration process is completed. The impact of the test-pesticide substance on the
crop in terms of its susceptibility to other pests / vectors ( insects, mites, other non-target pathogens ( virus, mycoplasma,
bacteria, and other fungi ) shall be carefully observed and recorded. The data may be recorded on the impact on
communities of natural enemies ( based on Indian Published literature ) of pests ( insects, mites, pathogens ) for thegiven
agro-climate where the bioefficacy evaluations are undertaken. The data on the impact of the test-substance on natural
enemies shall also be taken up under the study of eco-toxicology of the pesticide substance.
1) Protocol for efficacy evaluation of fungicides against various fungal pathogens
The experiment is conducted in well-drained fertile and good quality soil of the research farm of the institution, with all the
normal recommended package of practice to raise the relevant crop for undertaking bioefficacy evaluation of insecticides.
The bioefficacy trial may be monitored by high-power group of scientist, nominated by the Director of Research of the
institution Heads of the relevant plant protection departments ( Entomology, Plant Pathology / Nematology, senior
experienced superannuated eminent scientists ( if needed ), along with representation of the client industry. A checklist of
protocols may be drawn up during each evaluation after discussing with the relevant pesticide manufacturer ( client ) and
the scientist ( evaluator ). The check-list may draw up the High-power scientific group.
A project document (PD) is drawn up, with due identification number and authentication of the institution, containing the
under-mentioned details, before the commencement of the experiment and approved by the competent authority of the
institution. The data generated may be recorded in permanent work dairies and need to be preserved in the institution for
reasonable duration till the pesticide registration process is completed.
I)
Bioefficacy-assessment evaluation of fungicides against fungal pathogen
The target fungal disease(s) and the crop commodity may be spelt out while the PD of the evaluation protocol is
designed for each agro-climate. The fungal biology has to be kept in view for deciding on the mode and time of
application of the relevant test dose that is finalized based on the laboratory evaluation data.
1.0
EXPERIMENTAL CONDITION
1.1
Crop, Variety and Target disease(s) (Scientific names)
The name of the crop along with that of variety and target test organism (s), i.e. the relevant target tissuechewing, cutting, boring insects may be mentioned in the PD.
1.2
Trial conditions
Field trial should be set up where the target crop of moderate susceptibility to the target disease is likely
to be infected by the pathogen. Standard crop husbandry practices like application of FYM, fertilizers
including micronutrients and other maintenance treatments including pest management practices for pests
other than the target pest should be specified. Trial should be carried out in different agro-climatic regions
1.3
Design and layout of the trial
1.3.1
Treatments
Test product(s), reference product(s) and untreated control, arranged in a randomized block design or any
other statistically suitable design.
1.3.2
Plot size and replication
Net plot size at least 15-20 sq.m per treatment but according to the formulation or application
equipment, it may be necessary to use a larger plot size. In case of tree crops, 1-2 trees per replication per
treatment. In vine crops like grapes, 1 m row length can be considered.
Replicates – at least three, provided the error degrees of freedom are at least 12.
1.2.0
APPLICATION OF TREATMENT
1. 2.1
Test product (s)
The formulated product under investigation is applied according to the PD. The appliance shall be
chosen to facilitate uniform high volume application on the crop at its given age and canopy size.
1.2.2
Reference product
Registered product if any / Recommended product for the control of target insect pest.
1.2.3
Mode of application
Application should comply with good agricultural practices. Details on the spray equipment, type
of nozzle and water volume used at different stages of crop growth should be elucidated. Addition
of sticker, if any (for monsoon sprays) must be specified. Sticker should be tried in the check plot
with water spray.
1.2.3.1. Type of application
The type of application (e.g. Foliar spray, soil drenching, stem painting etc) as proposed by the
sponsor
1.2.3.2 Time and frequency of application
The time and frequency of application will normally be specified by the manufacturer/sponsor.
The time and frequency will normally depend on the pest species, & local environment conditions
& pest ETL’S. The number of applications and the date of each application should be recorded.
1.2.3.3 Doses and Spray volumes used
The product should be tested at the proposed dose and may also be used with other doses, usually
one dose higher and lower than the proposed dose. The doses will normally be expressed in g
a.i/ha and formulation in terms of ml/ha, g/ha.
1.3
MODE OF ASSESSMENT, RECORDING & MEASUREMENTS
1.3.1
Weather Data
1.3.1.1 Weather data during experimental duration for each location should be provided and co-related
with the field data that is generated on the disease incidence and plant mortality of each treatment
across replications. The inference shall be directed towards the influence of test substance in
reducing the target insect / mite damage.
1.3.1.2
Soil
Soil type should be mentioned where the crop is cultivated for such evaluations
1.3.2
Observations / Assessments
The PD shall contain the details of the observations / assessments of treatment and the
frequency as well as the various scientific basis ( published literature reference ) for those.
Disease Assessments
Disease assessments can be carried out either as disease severity (e.g. % infected surface area) or
incidence (proportion of the sample with any infection e.g. leaves, panicles & fruit etc).
Severity: Assess and record disease severity on a % -scale for each disease (0% = disease free,
100% = complete disease cover). The % infected surface area can be used for whole plots, whole
plants or individual plant parts.
Incidence: For incidence, the number of samples with infection and the total number of samples
per plot should be assessed and recorded in the rating description e.g. number of diseased leaves
per 30 leaves assessed or total number of infected ears/area.
Calculate the percent disease Index using the following formula
PDI =
1.7
Sum of all disease Ratings
____________ x 100
Total no of leaves/bunches assessed x Maximum Disease grade
Effect on Natural Enemies
The natural enemy spectrum of the target pest, as catalogued in the literature and perceived in the
region on the crop species in which this evaluation is done, may be recorded in the field collected
sample test insects and the impact of the targeted pesticide treatments in terms of mortality,
moribund condition, lack of natural enemy invasion unlike in untreated check plots may be
recorded.
1.7.1
Other new pests ( insects / mites / nematodes / diseases ) in the experiment
The investigator (evaluator) may look for the occurrence of any new pests ( insects / mites /
nematodes / diseases ) in each of the evaluation plots. The help of relevant specialists for each of
these may be taken and need to be recorded throughout the experiment period.
Phytotoxicity: Phytotoxicity at ‘X’ and ‘2X’ dose as per the rating scale given below.
Phyto-toxicity at ‘X’ and ‘2X’ dose shall be recorded at 1, 3, 5, 7 and 10 days after application.
Phytotoxicity Rating Scale (PRS)
Crop response/ Crop injury
Rating
0-00
0
1-10%
1
11-20%
2
21-30%
3
31-40%
4
41-50%
5
51-60%
6
61-70%
7
71-80%
8
81-90%
9
91-100%
10
Rating shall be recorded individually for yellowing, stunting, necrosis, epinasty
hyponasty etc.,
T
Treatment
No
Dose / ha
Days after 1st spray
g. a. i.
1
Formulation
3
5
7
10
1
2
3
4
1.8.
Crop Yield
The crop yield data may be recorded for each treatment and may be statistically analysed. The
conclusion on the treatment effect alone on enhancement of crop yield has to be brought out
using appropriate statistical and other tools so that there is no confused statement in the inference
that the pesticide dosage enhanced yield, without quantifying the avoidable crop loss due to given
pesticide dose application against the target pests in the evaluation.
1.8.1
Statistical Analysis
The data thus obtained shall be subjected to appropriate transformations and analyzed using
standard experimental designs.
1.8.2
RESULT & CONCLUSION
1.8.2.1 Result
Results to be submitted in table format mentioning the statistical parameters of assessment
such CD.CV%
1.8.2
Inferences and Conclusion
Brief note on the trial observations and performance of the test substance based on the results
may be provided with critical assessment of tested pesticide efficacy on the target pest(s)
along with opinion and conclusion on its impact on the target species on natural enemies of
the given agro-climate in which the bioefficacy evaluation was undertaken. Any
observations on the upsurge of any other pests ( insects / mites / nematodes / disease causing
organisms ) may be specifically mentioned. The possible role of good agricultural practice
by the new pesticide formulation may be inferred. The conclusion shall mention the most
effective concentration / dose of the pesticide to suppress the target pests. The results may be
given only quantifying the avoidable crop loss due to given pesticide dose application
against the target pests in the evaluation.
Certain model examples for fungal diseases
1. Example for some prominent fungal disease are as below :
Assessment units and disease rating scales for various fungal diseases
Downy mildew of grape (Plasmopara viticola)
Examine 50 leaves, inflorescence or bunches per plot from disease infection stage to berry development
and grade the disease incidence as per the scale below:
Score
1
2
3
4
5
6
Symptoms
No infection
<5% of the area infected
5-10% of the area infected
11-25% of the area infected
26-50% of the area infected
>50% of the area infected
Powdery mildew of grape (Uncinula necator)
Examine 50 leaves or bunches per plot from disease infection stage to berry development and grade the
disease incidence as per the scale below:
Score
1
2
3
4
5
6
Symptoms
No infection
<5% of the area infected
5-10% of the area infected
11-25% of the area infected
26-50% of the area infected
>50% of the area infected
Die-back & Fruit rot of chilli (Colletotrichum capsici)
Examine 5 plants within the plot during development stages and grade the disease incidence as per the
scale below.
Score
Symptoms
0
1
2
3
4
5
No infection
First symptom, 19% infection
20-39% infection
40-59% infection
60-79% infection
80-100% infection
Powdery mildew of chilli (Leveillula taurica)
Examine 5 plants within the plot during development stages and grade the disease incidence as per the
scale below.
0
1
3
5
7
9
No symptom on any plant
Small powdery specks on the leaves covering 1% or less area
Powdery lesions small, scattered covering 1 – 10% of leaf area
Powdery patches big, scattered covering 11 – 25% of the leaf area
Powdery patches big, coalescing covering 26 – 50% of leaf area
Powdery growth covering 51% or more of leaf area. Leaf turn yellow and dry up
Late Blight of potato / Tomato (Phytophthora infestans)
Score
Symptoms
Score
0
Not seen on field
1
Only a few plants affected here and there, upto 1 or 2 spots in 12 yards radius. Upto 10
spots per plant, or general light spotting
3
About 50 spots per plant, or upto 1 leaf let in 10 attacked
5
Nearly every leaflet with lesions, plants still retaining normal form; field may smell of
blight but looks green, although every plant is affected
7
Every plant affected and about ½ of leaf area destroyed by blight; field looks green
flecked with brown
9
About ¾ of leaf area destroyed by blight; field looks either predominantly brown or
green. In some varieties the youngest leaves escape infection. Only few leaves left
green but stem remain green. All leaves dead, stems dead or drying.
Black Scurf of potato (Rhizoctonia solani)
Observe tubers from 5 plants per plot and score on a 0 – 9 scale as follows:Score
Symptoms
0
No symptoms on potato tuber
1
1% or less area affected
2
1-10% area affected
5
11-20% area affected
9
51% or more area affected
Early Blight of tomato (Alternaria solani)
Score
Symptoms
0
No symptom on the leaf
1
Small irregular brown spots covering 1% or less of the leaf area
3
Small, irregular, brown spots with concentric rings covering 1 – 10% of the leaf
area
5
Lesions enlarging, irregular, brown with concentric rings, cover 11 – 25% of leaf
area
7
Lesions coalesce to form irregular, dark brown patches with concentric rings
covering 26 – 50% of leaf area. Lesions on stems and petioles
9
Lesions coalesce to form irregular, dark brown patches with concentric rings
covering more than 51% of leaf area. Lesions on stems and petioles
Leaf Blast (Pyricularia oryzae)
Observe 5 hills per replication at maximum tillering stage and grade leaves using IRRI scale.
Score
Symptoms
0
No visible lesions
1
Small brown specs of pin point size or larger brown specks without sporulating
center
3
Small, roundish to slightly elongated necrotic sporulating spots, about 1-2 mm in
diameter with a distinct brown margin or yellow hao
5
Narrow or slightly elliptical lesions, 1-2 mm in breadth, more than 3 mm long with
a brown margin
7
Broad spindle shaped lesion with yellow, brown or purple margin
9
Rapidly coalescing small, whitish, grayish or bluish lesions without distinct
margins
Panicle Blast (Pyricularia oryzae)
At 20-25 days after heading at Dough to Mature stage; observe 25 panicles per replication and grade
using following scale and calculate disease severity using the formula given below:
Score
Symptoms
0
No visible lesions or observed lesions on only few pedicels
1
Lesions on several pedicels or secondary branches
3
Lesions on a few primary branches or middle part of panicle axis
5
Lesions partially around the base (node) or the uppermost internode or
the lower part of the panicle axis near the base
7
Lesions completely around panicle base or uppermost internode or
panicle axis near base with more than 30% filled grains
9
Lesions completely around panicle base or uppermost internodes or the
panicle axis near the base with less than 30% filled grains
Sheath Blight (Rhizoctonia solani)
Observe 3 hills/ plot and observe tillers score on scale as follows.
Grade
Description
0
No Incidence
1
Vertical spread of the lesions up to 20% of plant height
3
Vertical spread of the lesions up to 21-30% of plant height
5
Vertical spread of the lesions up to 31-45% of plant height
7
Vertical spread of the lesions up to 46-65% of plant height
9
Vertical spread of the lesions up to 66-100% of plant height
Blister Blight (Exobasidium vexans):
Observations to be recorded on pluckable shoots (2 leaves & a bud) and individually examined for blister
blight and scored on a 0-4 scale (UPASI)
0=
1=
2=
3=
4=
No infection;
One or two lesions;
Three to Five lesions;
More than 5 lesions;
Stalk Infection).
Coffee leaf rust (Hemelia vastotrix):
Observations to be recorded on pluckable shoots (2 leaves & a bud) and individually examined for blister
blight and scored on a 0-4 scale (UPASI)
0=
1=
2=
3=
4=
5=
No infection;
1-20% leaf area infected;
21-40% leaf area infected;
41-60% leaf area infected;
61-80% leaf area infected;
81 %and above leaf area infected
2) Bioefficacy-assessment evaluation of fungicides against bacterial pathogen
Introduction
The bacterial pathogens are significant in infecting certail crops. It would be very important to screen and assess
effective pesticide formulations to contain the bacterial disease in crops. The bacterial diseases need to be separately
evaluated with candidate pesticide formulation. The bacterial diseases in conjunction with fungal and other pest
association need to be separately assessed for the bioefficacy of pesticides that are proposed to be registered for
managing bacterial crop diseases.
Bioefficacy-assessment evaluation of fungicides against fungal pathogen
The target fungal disease(s) and the crop commodity may be spelt out while the PD of the evaluation protocol is
designed for each agro-climate. The fungal biology has to be kept in view for deciding on the mode and time of
application of the relevant test dose that is finalized based on the laboratory evaluation data.
2.0
EXPERIMENTAL CONDITION
2.1
Crop, Variety and Target disease(s) (Scientific names)
The name of the crop along with that of variety and target test organism (s), i.e. the relevant target tissuechewing, cutting, boring insects may be mentioned in the PD.
2.2
Trial conditions
Field trial should be set up where the target crop of moderate susceptibility to the target disease is likely
to be infected by the pathogen. Standard crop husbandry practices like application of FYM, fertilizers
including micronutrients and other maintenance treatments including pest management practices for pests
other than the target pest should be specified. Trial should be carried out in different agro-climatic regions
2.3
Design and layout of the trial
2.3.1
Treatments
Test product(s), reference product(s) and untreated control, arranged in a randomized block design or any
other statistically suitable design.
2.3.2
Plot size and replication
Net plot size at least 15-20 sq.m per treatment but according to the formulation or application
equipment, it may be necessary to use a larger plot size. In case of tree crops, 1-2 trees per replication per
treatment. In vine crops like grapes, 1 m row length can be considered.
Replicates – at least three, provided the error degrees of freedom are at least 12.
2.2.0
APPLICATION OF TREATMENT
2.2.1
Test product (s)
The formulated product under investigation is applied according to the PD. The appliance shall be
chosen to facilitate uniform high volume application on the crop at its given age and canopy size.
2.2.2
Reference product
Registered product if any / Recommended product for the control of target insect pest.
2.2.3
Mode of application
Application should comply with good agricultural practices. Details on the spray equipment, type
of nozzle and water volume used at different stages of crop growth should be elucidated. Addition
of sticker, if any (for monsoon sprays) must be specified. Sticker should be tried in the check plot
with water spray.
2.2.3.1. Type of application
The type of application (e.g. Foliar spray, soil drenching, stem painting etc) as proposed by the
sponsor
2.2.3.2 Time and frequency of application
The time and frequency of application will normally be specified by the manufacturer/sponsor.
The time and frequency will normally depend on the pest species, & local environment conditions
& pest ETL’S. The number of applications and the date of each application should be recorded.
2.2.3.3 Doses and volumes used
The product should be tested at the proposed dose and may also be used with other doses, usually
one dose higher and lower than the proposed dose. The doses will normally be expressed in g
a.i/ha and formulation in terms of ml/ha, g/ha.
2.3
MODE OF ASSESSMENT, RECORDING & MEASUREMENTS
2.3.1
Weather Data
2.3.1.1 Weather data during experimental duration for each location should be provided and co-related
with the field data that is generated on the disease incidence and plant mortality of each treatment
across replications. The inference shall be directed towards the influence of test substance in
reducing the target insect / mite damage.
2.3.1.2
Soil
Soil type should be mentioned where the crop is cultivated for such evaluations
2.3.2
Observations / Assessments
The PD shall contain the details of the observations / assessments of treatment and the
frequency as well as the various scientific basis ( published literature reference ) for those.
Disease Assessments
Disease assessments can be carried out either as disease severity (e.g. % infected surface area) or
incidence (proportion of the sample with any infection e.g. leaves, panicles & fruit etc).
Severity: Assess and record disease severity on a % -scale for each disease (0% = disease free,
100% = complete disease cover). The % infected surface area can be used for whole plots, whole
plants or individual plant parts.
Incidence: For incidence, the number of samples with infection and the total number of samples
per plot should be assessed and recorded in the rating description e.g. number of diseased leaves
per 30 leaves assessed or total number of infected ears/area.
Calculate the percent disease Index using the following formula
PDI =
2.4
Sum of all disease Ratings
____________ x 100
Total no of leaves/bunches assessed x Maximum Disease grade
Effect on Natural Enemies
The natural enemy spectrum of the target pest, as catalogued in the literature and perceived in the
region on the crop species in which this evaluation is done, may be recorded in the field collected
sample test insects and the impact of the targeted pesticide treatments in terms of mortality,
moribund condition, lack of natural enemy invasion unlike in untreated check plots may be
recorded.
2.4.1
Other new pests ( insects / mites / nematodes / diseases ) in the experiment
The investigator (evaluator) may look for the occurrence of any new pests ( insects / mites /
nematodes / diseases ) in each of the evaluation plots. The help of relevant specialists for each of
these may be taken and need to be recorded throughout the experiment period.
2.5
Phytotoxicity: Phytotoxicity at ‘X’ and ‘2X’ dose as per the rating scale given below.
Phyto-toxicity at ‘X’ and ‘2X’ dose shall be recorded at 1, 3, 5, 7 and 10 days after application.
Phytotoxicity Rating Scale (PRS)
Crop response/ Crop injury
Rating
0-00
0
1-10%
1
11-20%
2
21-30%
3
31-40%
4
41-50%
5
51-60%
6
61-70%
7
71-80%
8
81-90%
9
91-100%
10
Rating shall be recorded individually for yellowing, stunting, necrosis, epinasty
hyponasty etc.,
T
No
1
Treatment
Dose / ha
Days after 1st spray
g. a. i.
1
Formulation
3
5
7
10
2
3
4
2.5
Crop Yield
The crop yield data may be recorded for each treatment and may be statistically analysed. The
conclusion on the treatment effect alone on enhancement of crop yield has to be brought out
using appropriate statistical and other tools so that there is no confused statement in the inference
that the pesticide dosage enhanced yield, without quantifying the avoidable crop loss due to given
pesticide dose application against the target pests in the evaluation.
2.5.1
Statistical Analysis
The data thus obtained shall be subjected to appropriate transformations and analyzed using
standard experimental designs.
2.5.2
RESULT & CONCLUSION
2.5.2.1 Result
Results to be submitted in table format mentioning the statistical parameters of assessment
such CD.CV%
2.5.2.2 Inferences and Conclusion
Brief note on the trial observations and performance of the test substance based on the results
may be provided with critical assessment of tested pesticide efficacy on the target pest(s)
along with opinion and conclusion on its impact on the target species on natural enemies of
the given agro-climate in which the bioefficacy evaluation was undertaken. Any
observations on the upsurge of any other pests ( insects / mites / nematodes / disease causing
organisms ) may be specifically mentioned. The possible role of good agricultural practice
by the new pesticide formulation may be inferred. The conclusion shall mention the most
effective concentration / dose of the pesticide to suppress the target pests. The results may be
given only quantifying the avoidable crop loss due to given pesticide dose application
against the target pests in the evaluation.
3) Bioefficacy-assessment evaluation of fungicides against Virus disease pathogen
Introduction
Crop damage due to viral diseases is quite common. Pesticide substances that are targeted towards control of viral
disease might be evaluated in times to come. The nioefficacy protocol need to look into the reduction in infection
process as well as their carry over through tissues including through planting material and seeds.
Bioefficacy-assessment evaluation of fungicides against fungal pathogen
The target fungal disease(s) and the crop commodity may be spelt out while the PD of the evaluation protocol is
designed for each agro-climate. The fungal biology has to be kept in view for deciding on the mode and time of
application of the relevant test dose that is finalized based on the laboratory evaluation data.
2.0
EXPERIMENTAL CONDITION
2.1
Crop, Variety and Target disease(s) (Scientific names)
The name of the crop along with that of variety and target test organism (s), i.e. the relevant target tissuechewing, cutting, boring insects may be mentioned in the PD.
2.2
Trial conditions
Field trial should be set up where the target crop of moderate susceptibility to the target disease is likely
to be infected by the pathogen. Standard crop husbandry practices like application of FYM, fertilizers
including micronutrients and other maintenance treatments including pest management practices for pests
other than the target pest should be specified. Trial should be carried out in different agro-climatic regions
2.3
Design and layout of the trial
2.3.1
Treatments
Test product(s), reference product(s) and untreated control, arranged in a randomized block design or any
other statistically suitable design.
2.3.2
Plot size and replication
Net plot size at least 15-20 sq.m per treatment but according to the formulation or application
equipment, it may be necessary to use a larger plot size. In case of tree crops, 1-2 trees per replication per
treatment. In vine crops like grapes, 1 m row length can be considered.
Replicates – at least three, provided the error degrees of freedom are at least 12.
The experiment is conducted in well-drained fertile and good quality soil of the research farm of the institution, with all the
normal recommended package of practice to raise the relevant crop for undertaking bioefficacy evaluation of insecticides.
The bioefficacy trial may be monitored by high-power group of scientist, nominated by the Director of Research of the
institution Heads of the relevant plant protection departments ( Entomology, Plant Pathology / Nematology, senior
experienced superannuated eminent scientists ( if needed ), along with representation of the client industry. A checklist of
protocols may be drawn up during each evaluation after discussing with the relevant pesticide manufacturer ( client ) and
the scientist ( evaluator ). The check-list may draw up the High-power scientific group.
A project document (PD) is drawn up, with due identification number and authentication of the institution, containing the
under-mentioned details, before the commencement of the experiment and approved by the competent authority of the
institution. The data generated may be recorded in permanent work dairies and need to be preserved in the institution for
reasonable duration till the pesticide registration process is completed.
The target fungal disease(s) and the crop commodity may be spelt out while the PD of the evaluation protocol is
designed for each agro-climate. The fungal biology has to be kept in view for deciding on the mode and time of
application of the relevant test dose that is finalized based on laboratory evaluation data.
3.0
EXPERIMENTAL CONDITION
3.1
Crop, Variety and Target disease(s) (Scientific names)
The name of the crop along with that of variety and target test organism (s), i.e. the relevant target tissuechewing, cutting, boring insects may be mentioned in the PD.
3.2
Trial conditions
Field trial should be set up where the target crop of moderate susceptibility to the target disease is likely
to be infected by the pathogen. Standard crop husbandry practices like application of FYM, fertilizers
including micronutrients and other maintenance treatments including pest management practices for pests
other than the target pest should be specified. Trial should be carried out in different agro-climatic regions
3.3
Design and layout of the trial
3.3.1
Treatments
Test product(s), reference product(s) and untreated control, arranged in a randomized block design or any
other statistically suitable design.
3.3.2
Plot size and replication
Net plot size at least 15-20 sq.m per treatment but according to the formulation or application
equipment, it may be necessary to use a larger plot size. In case of tree crops, 1-2 trees per replication per
treatment. In vine crops like grapes, 1 m row length can be considered.
Replicates – at least three, provided the error degrees of freedom are at least 12.
3.4
APPLICATION OF TREATMENT
3.4.1
Test product (s)
STANDARD EVALUATION SYSTEM
BIOEFFICACY – ASSESSMENT EVALUATION OF BIOEFFICACY OF
PESTICIDE
FOR SEED TREATMENT
EVALUATION TEST PROTOCOL FOR BIOEFFICACY – ASSESSMENT
Introduction
The field experiment is conducted in well-drained fertile and good quality soil of the research farm of the institution, with all the
normal recommended package of practice to raise the relevant crop for undertaking bioefficacy evaluation of insecticides. The
bioefficacy trial may be monitored by high-power group of scientist, nominated by the Director of Research of the institution
Heads of the relevant plant protection departments ( Entomology, Plant Pathology / Nematology, senior experienced
superannuated eminent scientists ( if needed ), along with representation of the client industry. A checklist of protocols may be
drawn up during each evaluation after discussing with the relevant pesticide manufacturer ( client ) and the scientist
(
evaluator ). The check-list may draw up the High-power scientific group.
A project document (PD) is drawn up, with due identification number and authentication of the institution, containing the undermentioned details, before the commencement of the experiment and approved by the competent authority of the institution. The
data generated may be recorded in permanent work dairies and need to be preserved in the institution for reasonable duration till
the pesticide registration process is completed.
I) EVALUATION OF FUNGICIDES FOR SEED TREATMENT
Seed treatment with fungicides is intended for the control of internally and externally seed-borne plant
pathogens and also of soil-born plant pathogens. The manufacture and / or the sponsor of the bioefficacy evaluation research study of test pesticide substance may approach the relevant plant protection
department of state agriculture university (SAU) or crop institute under the Indian Council of
Agricultural Research ( ICAR or any other similar institution* ) of any agro-ecological region of the
country ). The evaluation of the test pesticide substance is intended to assess the efficiency of disease
reduction of soil-born and seed-borne pathogens during the early crop growth of the desired crop
commodity as also to measure all relevant effects on phyto-toxicity, non-target soil organisms as well as
on plant growth characteristics.
1.0
EXPERIMENTAL CONDITION
The laboratory may take up the evaluation through the approved and well-planned research project, with
due identification number and authentication of the institution.
1.1
Selection of crop, cultivar and test organisms
This test protocol is concerned with the efficacy evaluation of seed treatment fungicides for the control of
seed-borne diseases (surface-borne or internally seed-borne) like Loose smut and Bunt in wheat and Soilborne diseases like Downy mildew in maize, sorghum, bajra and sunflower, wilt pathogens in soil that
infect may field and horticulture crops.
1.2
Trial conditions
Field trial / evaluation of the candidate pesticide ( fungicide ) substance should be set up using a stock of
naturally infected seeds (10-30%), available in the test-laboratory that has proficiency for this purpose.
The evaluation should be carried out in specified regions during normal crop season(s). As far as possible,
try to ensure uniform disease infection through out the trial site.
Before commencing the field trial / evaluation, the investigator / scientist of the designated laboratory
may take up laboratory evaluation of the pesticide substance that is subjected to assess field efficacy
against the target soil borne seed-borne pathogen, following the standard globally recognized
methodology. The finalization of the evaluation protocol such as design and layout of ( laboratory & field
) shall be through formal consultation with the industry R&D representative and the designated scientist
and Head of the department with CIB designated organization.
1.3
Design and layout of the trial
1.3.1
Treatments
Test product(s), reference product(s) and untreated control, arranged in a randomized block design or any
other appropriate statistical design (Consult statistician in this regard).
1.3.2
Plot size and replication
Net plot size of at least 25 m2 or as required by target crop is recommended. But the plot size or
replications should be increased as the level of infection drops or inoculated seed may be used, depending
upon the disease to be tested. Replicates – at least three, provided the error degrees of freedom are at least
12.
2.0
APPLICATION OF TREATMENT
2.1
Test product (s)
The formulated pesticide substance with the proposed dosage of the sponsor and / or as arising from the
laboratory evaluation results may be considered.
2.2
Reference product
Pesticide substances of equal potency and efficacy that are already recommended for the control of the
disease through seed treatment should be included. If available, products with specific seed treatment
formulations such as WS, FS, and DS should be included.
2.3
Mode of application
Method of application of seed dresser should be specified and undertaken with details of the
process that is expressed explicitly, viz., dry coating, slurry coating, pelleting etc, as applicable.
Application should comply with good agricultural practices ( may spell it out before the
evaluation is taken up ). Mention the use of any polymers/stickers /other additives for better
adhesion of the test substance. These materials extrinsic to the test substance, should be
separately assessed for their influence / efficacy on target disease causing organism other than
with the pesticide substance in the evaluation process.
2.3.1
Type of sowing
The type of sowing (e.g. broadcasting, drilling and dribbling) as per recommended practice or as
suggested by the manufacturer.
2.3.2
Time and frequency of application
Usually, a single application as seed treatment, unless otherwise specified by the manufacturer/
sponsor. The date of application should be recorded. The date shall be fixed, as emerging out of
the discussion with the pesticide industry sponsor of the research evaluation or as independently
worked for the specific crop seed by the R&D organization that is involved in the bio-efficacy
evaluation.
2.3.4
Doses and Volumes used
The product should be tested at the proposed dose and may also be used other doses. The dosage
will normally be expressed in g a.i/kg seed or grams or milliliter of formulated product per kg of
seed. For slurry preparation, the water volume needs to be specified according to quantity of seed
to be treated. The hundred / thousand seed weight shall be the criteria for fixing the dosage as
well as for recommending the appliance that can be used to ensure that each grain carries the
lethal dosage prescribed to ensure optimum seed health and full plant stand per unit area.
2.0 MODE OF ASSESSMENT, RECORDING & MEASUREMENTS
2.1
Meteorological data
Weather data during experimental duration for each location should be provided and co-related
with the field data that is generated on the disease incidence and plant mortality of each treatment across
replications. The inference shall be directed towards the influence of test substance in reducing the pathogenesis.
2.1.2 Soil
2.1.3
Soil physical and chemical parameters should be mentioned in the test report as also for deciding
the treatments for bioefficacy evaluation of the given test substance.
Crop
The peculiarities of response in the crop, right from emergence to over one month may be
recorded in all treatments.
3.0
Observations
3.1
Disease Assessments
Disease assessments can be carried out either as disease severity (e.g. % infected surface area) or
incidence (proportion of the sample with any infection e.g. leaves, panicles & fruit etc).
3.1.2
Severity: Assess and record disease severity on a % -scale for each disease (0% = disease free,
100% = complete disease cover). The % infected surface area can be used for whole plots, whole
plants or individual plant parts.
3.1.3
Incidence: For incidence, the number of samples with infection and the total number of samples
per plot should be assessed and recorded in the rating description e.g. number of diseased leaves
per 30 leaves assessed or total number of infected ears/area.
3.1.4
Calculate the percent disease Index using the following formula
PDI =
Sum of all disease Ratings
____________ x 100
Total no of leaves/bunches assessed x Maximum Disease grade
3.1.5
% germination to be assessed
3.2
Yield
At harvest the grain yield shall be determined plotwise and expressed using appropriate measure
viz kgs or quintals or tones per hectare. Test weight (weight of 100 seeds) be recorded and
expressed in grams. This has to be coorelated with total plant stand of each treatment to relate the
influence of disease incidence in the crop and crop yield, keeping all other agronomic practices (
spacing, fertilizer application, irrigation ( if relevant ) other pesticide application, growth
hormone etc. ) are constant. The research may bring out any observed effect on quality / quantity
of the crop commodity and may be reported with suitable scientific reason.
3.3
Phytotoxicity
Assess the detailed phytotoxicity / growth observations in the field at ‘X’ and ‘2X’ dose as per
the standard rating scale.The laboratory or field germination tests should be carried out
immediately after seed treatment.
4.0
Statistical Analysis
The data shall be subjected to appropriate transformations where necessary and analyzed using
standard experimental designs and computer software package.
Assessment units and disease rating scales for various fungal diseases
The published literature supported assessment and interpretation of the results may be made in respect of
the disease(s) under the present bioefficacy evaluation.
As examples, the following may be considered. However, the research study could modify and develop
its own protocol, as supported by scientific publications / reasoning.
Loose smut of wheat (Ustilago tritici) and Bunts (Telletia spp.)
Examine plants within 15 feet radius from heading to harvest and grade the disease incidence as per the scale
below both in terms of plants and tillers.
Score
Symptoms
0
No plants containing loose smut
1
1% or less plants with smutted heads
3
1-10% or less plants with smutted heads
5
11-20% or less plants with smutted heads
7
21-50% or less plants with smutted heads
9
51% ad above plants with smutted heads
______________________________________________________________________________
Source: Phytopathometry, Mayee & Datar, 1986
Downy mildew of sorghum (Peronosclerospora sorghi)
Examine plants within the entire plot from seedling, booting, panicle initiation and seed setting and grade the
disease incidence as per the scale below.
Score
Symptoms
0
1
Plants free from infection
Upto 1% of the plants showing infection. No shredding
3
Top leaves and bases of lower ones turn whitish or yellow. Partial shredding of
leaves. 1-10% plants affected.
5
Leaves turn whitish or yellow with downy growth on lower side. Complete
shredding of leaves. Ears partially filled. 11-20% plants affected.
7
Leaves narrow, pale yellow with downy growth on both surfaces. Complete
shredding of leaves. Ears not formed. 121-50% plants affected.
9
All leaves at seedling stage showing pale yellow patches with downy growth.
Premature death of seedling. Above 50% plants affected.
Source: Phytopathometry, Mayee & Datar, 1986
Brown stripe downy mildew of maize (Scleropthora rayssiae)
Examine 10 plants within the plot from two leaves to maturity and grade the disease incidence as per the
scale below.
Score
Symptoms
0
No symptoms on leaf
1
Upto 1% of the leaf area covered with necrotic lesions
3
1-10% of the leaf area covered with necrotic lesions
5
11-25% of the leaf area covered with necrotic lesions
7
26-50% of the leaf area covered with necrotic lesions
9
More than 50% of the leaf area covered with necrotic lesions
Source: Phytopathometry, Mayee & Datar, 1986
STANDARD EVALUATION SYSTEM
BIOEFFICACY – ASSESSMENT EVALUATION OF BIOEFFICACY OF
HERBICIDE AGAINST WEED
TEST PROTOCOL FOR EFFICACY EVALUATION OF HERBICIDES AGAINST
WEEDS
1.0
EXPERIMENTAL CONDITION
1.1
Crop, Variety and Target weeds (Scientific names)
1.1.2
Variety : Popular variety or common grown variety grown in the area
1.1.3
Target Weeds : with other predominant weeds of the crop
1.1.4
Type of Land : Low land or up land
1.2
Trial conditions
Field trial should be set up where target crop are likely to be infested by target weed species and where
target crop are likely to be infecsted by target weed species and where the weeds become major
problem. Standard crop husbandry practices like application of FYM, fertilizers including micronutrients
and other maintenance treatments including pest management practices for pests other than the target pest
should be specified. Trial should be carried out in different agro-climatic regions as specified by ICAR
or Planning Commission
1.3
Design and layout of the trial
1.3.1
Treatments
Test product(s), reference product(s) and untreated control, arranged in a randomized block design or any
other suitable statistical design which represent the whole field.
1.3.2
Plot size and replication
Net plot size at least 15 – 20 sq.m. per replication per treatment but according to the formulation or
application equipment, it may be necessary to use a larger plot size ( at least 25 sq.m). The plot size
must be the same at all location.
Replicates – at least three, provided the error degrees of freedom are at least 12. Minimum 7 treatments in
which one standard check, one Weed Free and one Weekly Check must be included with minimum 4
treatments of test product.
2.0
APPLICATION OF TREATMENT
2.1
Test product (s)
The formulated product under evaluation. Three treatments.
2.2
Reference product
Registered product if any / Recommended product for concerned weed management. As decided
by the respective AICRP / ICAR Institute / SAU in consultation with sponsor.
2.3
Mode of application
Application should comply with good agricultural practices. Details on the spray equipment, type
of nozzle (Flat fan or flood jet) and water volume used at different stages of crop growth should
be given. Addition of sticker, to improve the uptake of active ingredient, if any must be specified.
Sticker should be tried in the check plot with water spray. The above requirement should be
followed uniformly in all location in both the season.
2.3.1
Type of application
The type of application (e.g. foliar spray, soil incorporation, broad casting) as proposed by the
sponsor
2.3.2
Time and frequency of application
The time and frequency of application will normally be specified by the manufacturer/ sponsor.
The time of application should be conveniently mentioned based on the time of weed emergence
(PPI, Pre-emergence, early post emergence or late post emergence) or based on the stage of the
weed (1-2 leaf stage, 3-4 leaf stage etc) or DAS ( days after sowing ).
2.3.5
Doses and Volumes used
The product should be tested at the proposed dose(s), as desired. The doses will normally be
expressed in g a.i/ha and formulation ml/ha.
3.0 MODE OF ASSESSMENT, RECORDING & MEASUREMENTS
3.1
Meteorological Data
Weather data during experimental duration for each location should be provided
3.1.2
Soil
Soil type should be mentioned
4.0
Observations
4.1
Weed intensity
4.1.1
Weed count: The number of species wise weeds to be determined at pre-spray, 30DAS,
60DAS and at harvest at about 5 location per plot using a square quadrant, 1
sq.m.using a square quadrant, 1sq.m at about five locations per plot.
4.1.2
Weed Dry Matter : Dry or / fresh weight of weeds species wise to be determined at prespray, 30DAS, 60DAS and at Harvest and statistically analysed after transformation.
4.1.3
Percent weed control efficiency (WCE): Weed control efficiency should be mentioned
species wise as per label claim at 30DAS, 60DAS and at harvest.
WCE =
No of weeds in UTC – No of weeds in treated
------------------------------------------------------- x 100
No of weeds in UTC
WCE =
WDM in UTC – WDM in treated
------------------------------------------------------- x 100
WDM in UTC
And / or
WDM = Weed Dry Matter
UTC = Untreated control
4.1.4
Dry or/ Fresh Weight: Dry or fresh weight of weeds at last observation.
4.2
Yield: Crop yield data should be given along with Yield attributing character and statistically
analysed. Enhancement of the crop has to be brought out using appropriate statistical ando
other tools so that there is no confusion statement in drawing the inference.
4.3
Phytotoxicity: Phytotoxicity at ‘X’ and ‘2X’ dose as per the rating scale given below.
Phyto-toxicity at ‘X’ and ‘2X’ dose shall be recorded at 1, 3, 5, 7 and 10 days after application.
.
4.3.1 Photographs with label at each observation
Recovery Photographs if initially there is phytotoxicity
Phytotoxicity Rating Scale (PRS)
Crop response/ Crop injury
Rating
0-00
0
1-10%
1
11-20%
2
21-30%
3
31-40%
4
41-50%
5
51-60%
6
61-70%
7
71-80%
8
81-90%
9
91-100%
10
Rating shall be recorded individually for yellowing, stunting, necrosis, epinasty
hyponasty etc.
T
Treatment
No
Dose / ha
Days after 1st spray
g. a. i.
1
Formulation
3
5
7
10
1
2
3
4
5.0
Results: Results to be submitted in table format after appropriate statistical analysis along with
inferences.
6.0
Conclusion:
Brief note on the trial observations and performance of the test substance.
TEST PROTOCOL FOR EFFICACY EVALUATION OF PGR
1.0
EXPERIMENTAL CONDITION
1.1
Crop, Variety and Target weeds (Scientific names)
1.2
Trial conditions
Field trial should be set up where target crop are likely to be infested by target weed species. Standard
crop husbandry practices like application of FYM, fertilizers including micronutrients and other
maintenance treatments including pest management practices for pests other than the target pest should be
specified. Trial should be carried out in different agro-climatic regions
1.3
Design and layout of the trial
1.3.1
Treatments
Test product(s), reference product(s) and untreated control, arranged in a randomized block design or any
other suitable statistical design.
1.3.2
Plot size and replication
Net plot size at least 15 – 20 sq.m. per replication per treatment but according to the formulation or
application equipment, it may be necessary to use a larger plot size
Replicates – at least three, provided the error degrees of freedom are at least 12.
2.0
APPLICATION OF TREATMENT
2.1
Test product (s)
The formulated product under investigation.
2.2
Reference product
Registered product if any / Recommended product for the control of the target insect pest.
2.3
Mode of application
Application should comply with good agricultural practices. Details on the spray equipment, type
of nozzle (Flat fan or flood jet) and water volume used at different stages of crop growth should
be given. Addition of sticker, to improve the uptake of active ingredient, if any must be specified.
Sticker should be tried in the check plot with water spray.
2.3.1
Type of application
The type of application (e.g. foliar spray, soil incorporation, broad casting) as proposed by the
sponsor
2.3.2
Time and frequency of application
The time and frequency of application will normally be specified by the manufacturer/ sponsor.
The time and frequency will normally depend on critical growth stage of the crop & local
environment conditions. The number of applications and the date of each should be recorded.
2.3.6
Doses and Volumes used
The product should be tested at the proposed dose(s), and may also be used other doses. The
dosage will normally be expressed in g a.i/ha and formulation ml/ha.
4.0 MODE OF ASSESSMENT, RECORDING & MEASUREMENTS
3.1
Meteorological Data
Weather data during experimental duration for each location should be provided
3.1.2
Soil
Soil type should be mentioned
3.1.3
Age of the plant
3.2
Observations / Assessments
3.2.1
Growth parameters
Plant height
No. of primary branches / plant at 15 or 30 days interval after application
Yield and Yield attributes parameters ( no. of fruits, fruit length, fruit weight, yield
etc. )
3.2.2
.
Phytotoxicity: Phytotoxicity at ‘X’ and ‘2X’ dose shall be recorded.
Phytotoxicity Rating Scale (PRS)
Crop response/ Crop injury
Rating
0-00
0
1-10%
1
11-20%
2
21-30%
3
31-40%
4
41-50%
5
51-60%
6
61-70%
7
71-80%
8
81-90%
9
91-100%
10
Rating shall be recorded individually for yellowing, stunting, necrosis, epinasty
hyponasty etc.
T
Treatment
No
Dose / ha
Days after 1st spray
g. a. i.
1
Formulation
3
5
7
10
1
2
3
4
3.2.4 Yield
Yield data shall be given
3.2.5 Statistical Analysis
The data thus obtained shall be subjected to appropriate transformations and analyzed using standard
experimental designs. Enhancement of the crop has to be brought out using appropriate statistical and
other tools so that there is no confusion statement in drawing the inference.
5.0 RESULT & CONCLUSION :
5.1 Result
Results to be submitted in table format with writeups.
5.2 Conclusion
Brief note on the trial observations and performance of the test substance.
COMMENTS OF BIO EFFICACY EXPERTS
(CIB & RC)
Protocol for bio-efficacy evaluation for registration of Insecticides
A: General Insecticides:
Operation:
Trial protocols should be elaborated, before starting. The organization should elaborate
standard operating procedures (SOPs) or standard modes of operation for testing,
measurements, data collection and reporting procedures or other tasks that are carried
repetitively, to ensure consistency and repeatability of the activities.
Verification:
1. Persons who are scientifically responsible for the trial within the organization should be
able to check and validate the trial throughout its course and ascertain that the protocol
and SOPs are being followed.
2. Organizations should accept at any time inspections that may be organized by the
registration authority or another competent national authority to verify the compliance of
trial/protocols. Organization must intimate to the Secretary CIB&RC about the beginning &
closure of the trials.
3. All trials need proper display. Experimental sites and display boards needs to be
photographed. All spray operations, observations in the experimental fields, from time to
time and crop growth stage from time to time needs to be well documented/ photographed
for submission to registration authority.
Experimental conditions:
1. Bio efficacy trials should strictly be conducted/data generated at SAU/ICAR institutes and
deemed to be Universities which have been listed on the ICAR website.
2. Consistency in methodology of trials/experiments should be followed at all the trial
locations in respect of frequency of recording bio efficacy data, frequency of treatment
applications, Number of trials , number of application and unit of evaluation of bio efficacy
as per standard scale for specific pest.
3. Trials should be conducted in locations which represent the range of agronomic, plant
health, environmental and climatic conditions likely to be encountered in practice in the
area of proposed use.
4. Selection of location for trial should be done keeping in view the prevalence of target pest
on crop species (Endemic areas) in that particular area (identified/ declared by
ICAR/SAUs/Indian published literature).
5. The crops in the experimental locations may be specifically grown for the trials or be part of
crop grown for commercial purposes, but in all cases should be grown according to good
agronomic practice (GAP).
6. Any cultural operations in the fields apart from the one being tested should be according to
Good Agronomic Practice (GAP).
7. If there is a possibility that soil types may affect the efficacy of the insecticide, the various
trial sites should be chosen to be representative of the range of soil types that can be
encountered in the proposed field use of the product.
8. Within each trial site, environmental and agronomic conditions should be as uniform as
possible.
9. In case Infestation levels are low or incidence of target insect pest does not occurs at the
location of trial in the season then the trial should be conducted in the next season at the
same location.
Bio-Efficacy evaluation:
1. Effects on non-target organisms (e.g. impact on pollinators and pollination, effects on
parasitoids/parasites and predators specific to the target pest and generally
found/reported in that crop eco-system.
2. Brief details of all cultural/agronomical practices, methods of fertilizer doses and its
application etc should be submitted to the registration authority in respect of trial
plots/standard check plots and control plots.
3. All cultural/agronomical practices, fertilizer dosages, irrigation and any other treatment
except the application of test chemical should be at par in trial plots, standard check plots
and control plots, so as level field comparative study of bio efficacy of the test product.
4. Insecticides which were approved/recommended by CIB&RC with doses against specific
crop pest should be taken as standard check during B/E trials.
5. The bio efficacy data of those cases which have been rejected by the Registration
Committee in the past due to non conformity of active ingredient contents during the lab
test of the samples, these bio efficacy data will not be considered /accepted for fresh
application of the same product by the same applicant with a new sample test report.
6. Details of Cost: Benefit ratio in terms of input cost, cost of product (Yield/Ha.) should be
submitted to the Registration Authority, in case of combination products. (Input cost should
be inclusive of the cost of cultural, agronomical, mechanical practices as per prevailing local
rates at the site of trial).
7. In case the insecticide is applied as a mixture with fungicide / PGR/fertilizer or other
chemical, the compatibility as claimed by the applicant should be supported with the
bioefficacy data in respect of other constituents of such mixture should be observed/
recorded to evaluate the efficacy of the constituent to prove the compatibility claim.
8. The test product should be safe to the environment, non target organisms and having
better efficacy and also cost effective in comparison to the standard checks.
9. i) In case of formulations- for new molecule additional data may be collected/
supported on effects on succeeding crops in the trial field.
ii) Effect on the test product on other non-target organism and eco-friendly insects in
adjacent field crops should also be recorded and submitted to registration authority.
10. All the activities should be well documented and supported with appropriate photographs
authenticated by concerned scientist and the director (Research) may be submitted to the
registration authority.
Experimental design:
i) Efficacy trials should be designed in such way so that appropriate standard statistical
analysis of the data (with minimum degree of freedom) can be performed to ensure that
any effects attributable to the insecticide can be distinguished from other forms of
experimental variability.
ii) A fully randomized design can be applied only if the trial environment/agro ecosystem is
completely homogeneous.
iii) Trial plot should be at least 25m2 in size with four replications each, however depending
upon the other factors viz. mobility of the target pest/organism, crop species/
canopy/spacing like sugarcane, sorghum, maize, bajra, pigeon pea and cotton etc., the plot
size should be 50m2 or more may be taken for trials.
Untreated control:
i) An untreated control should be included in the trial.
ii) Should confirm the presence of an adequate pest population/infestation and all activities
under GAP should be at par with the treated plots except the application of test chemicals
during the course of the trial.
iii) In case there is reduction in population of pest/NE, reason may be recorded thereof in the
trial data.
Application of Insecticide:
1. Treatment application should be made after observing ETL of the target pests in the
proposed crop.(ETL value for major pest in crops is given in Annexure-I)
2. The tentative schedule of treatment application should be planned/done keeping in view
the occurrence of pest at a particular crop stage and should be spread over the entire
susceptible/vulnerable stage of the crop at all the trial locations.
3. The equipment and method of application of the insecticide may be same in all trials handheld equipment may be used.
4. Application rates for sprays, the volume of application and dilution ratios (as appropriate)
may be as per recommendation for use.
5. Treatments doses, time/stage of application (tentative schedule), number of applications,
application intervals, method of application and spray volume may be recommended by the
concerned scientist based on the B/E trials results.
6. Application equipment should be properly calibrated before the treatment application for
obtaining intended application rate and droplet spectrum.
7. Details of the sprayer, its operating pressure, nozzle type and position of nozzle(s) relative
to crop, forward speed and volume of application rate are need to be specified by the
concerned scientist in the recommendation.
Weather:
1. Weather conditions viz. Ambient temperature, relative humidity, precipitation (if any), wind
speed at the time of application may be recorded on the day of treatment.
2. Temperature, relative humidity, precipitation and extreme weather conditions may also
need to be recorded on a regular basis throughout the trial period. The exact parameters
and frequency of recording will depend on the type of crop/pest/product under study.
Biological efficacy:
1. The variables that need to be assessed to determine the efficacy of the plant protection
product will strongly depend on the crop and pest being studied. They may include density
or incidence of the pest, infestation levels, percentage mortality or control, severity of
symptoms or damage to the crop, quantity of yield, quality of the product, etc. should be
recorded in the trial reports.
2. Pre-treatment assessments of target pests and non target pests (natural enemies) including
honey bees* & other pollinators should be carried out and recorded invariably in the B/E
trails reports/observation tables.
3. Parameters such as population reduction over control expressed as percent mortality,
4. The parasitoids, parasites/predators, honey bees/other pollinators* and pathogens of
target pest should be counted/observed before & after each application as in case of target
pest covered under specific safe insect order/crop ecosystem and it should be recorded in
trails reports/observation tables.
5. A separate bio efficacy study on the effect of neonicotinoid on honey bees & other
pollinators may be conducted and reported to the registration committee. For bio efficacy
(Residue) the following data may also be generated and submitted.
i) Estimation of level of residue in pollen and nectar resulting from spray treatment of
crops.
ii) Estimation of level of residue in pollen and nectar resulting from soil/seed treatment.
iii) Estimation of level of residue in pollen and nectar by calculating based on the spray
application rate.
*In case of neonicotinoid
Phytotoxcity: As given
Mode of application: As given
Time and frequency of application: As given
Dose and Volume used: As given
Results: As given
Statistical analysis: As given
Inferences and conclusion: As given
Study on persistence in soil, water and plant may be carried out as per existing data
requirement in the guidelines.
Study on residue in soil and plant may be carried out as per existing data requirement in the
guidelines.
B. House-hold Insecticide:
1. Trial should be conducted as per the WHO protocol/guidelines.
(Source: www.who/htm/ntd/whopes/2009.3)
2. More government (NABL/GLP) accreditated laboratories may be added in the existing list of
national laboratories for Bio efficacy trials for household insecticide category.
3. The proposed addition of government (NABL/GLP) accreditated laboratories will be added
after due verification of available trial facilities for conducting B/E trials.
C. Public Health Pesticides (PHP):
Bio efficacy study should be conducted as per NVBDCP approved SOP.
(Source: www.nvbdcp.nic.in)
D. Seed Treatment (Insecticide):
i)
The general requirements for registration of insecticide will be the same except the
method of application of insecticide.
ii)
Phytotoxicity study/trial must include comparative study of % seed germination in
control, standard check vis-à-vis the treatment.
iii)
Study on persistence in soil, water and plant may be carried out as per existing data
requirement in the guidelines.
iv)
Study on residue in soil and plant may be carried out as per existing data requirement
in the guidelines.
Test Protocol for bio-efficacy evaluation of Bio-insecticides
(Bio pesticides Unit)
The following parameters may be included in the test protocol of Bio-insecticides:Operation:
Trial protocols should be elaborated, before starting. The organization should elaborate
standard operating procedures (SOPs) or standard modes of operation for testing,
measurements, data collection and reporting procedures or other tasks that are carried
repetitively, to ensure consistency and repeatability of the activities.
Verification:
4. Persons who are scientifically responsible for the trial within the organization should be
able to check and validate the trial throughout its course and ascertain that the protocol
and SOPs are being followed.
5. Organizations should accept at any time inspections that may be organized by the
registration authority or another competent national authority to verify the compliance of
trial/protocols. Organization must intimate to the Secretary CIB&RC about the beginning &
closure of the trials.
6. All trials need proper display. Experimental sites and display boards needs to be
photographed. All spray operations, observations in the experimental fields, from time to
time and crop growth stage from time to time needs to be well documented/ photographed
for submission to registration authority.
Experimental conditions:
1. Bio efficacy trials should strictly be conducted/data generated at SAU/ICAR institutes and
Institutes recognized by ICAR under central universities engaged in field oriented research
trials. A list of all such SAU/ICAR institutes and other institutes attached at Annexure-I.
2. Consistency in methodology of trials/experiments should be followed at all the trial
locations in respect of frequency of recording bioefficacy data, frequency of treatment
applications, Number of trials , number of application and unit of evaluation of bioefficacy
as per standard scale for specific pest.
3. Trials should be conducted in locations which represent the range of agronomic, plant
health, environmental and climatic conditions likely to be encountered in practice in the
area of proposed use.
4. Selection of location for trial should be done keeping in view the prevalence of target pest
on crop species (Endemic areas) in that particular area (identified/ declared by
ICAR/SAUs/Indian published literature).
5. The crops in the experimental locations may be specifically grown for the trials or be part of
crop grown for commercial purposes, but in all cases should be grown according to good
agronomic practice (GAP).
6. Any cultural operations in the fields apart from the one being tested should be according to
Good Agronomic Practice (GAP).
7. If there is a possibility that soil types may affect the efficacy of the bio-insecticide, the
various trial sites should be chosen to be representative of the range of soil types that can
be encountered in the proposed field use of the product.
8. Within each trial site, environmental and agronomic conditions should be as uniform as
possible.
9. In case Infestation levels are low or incidence of target insect pest does not occurs at the
location of trial in the season then the trial should be conducted in the next season at the
same location.
Bio-Efficacy evaluation:
1 Effects on non-target organisms (e.g. impact on pollinators and pollination, effects on
parasitoids/parasites and predators specific to the target pest and generally
found/reported in that crop eco-system.
2.
Brief details of all cultural/agronomical practices, methods of fertilizer doses and its
application etc should be submitted to the registration authority in respect of trial
plots/standard check plots and control plots.
3. All cultural/agronomical practices, fertilizer dosages, irrigation and any other treatment
except the application of test biopesticides/chemicals should be at par in trial plots,
standard check plots and control plots, so as level field comparative study of bio efficacy of
the test product.
4. Insecticides which were approved/recommended by CIB&RC with doses against specific
crop pest should be taken as standard check during B/E trials. In case of unavailability of
such insecticide which have been approved /recommended 9under AICRP)by
ICAR/SAUs/Agriculture Institutes under Central Universities may be taken as standard
check( Authenticated published copy of such recommendation should be submitted with
reports/data).
5. Trial plot should be at least 20m2 in size with four replications each, however depending
upon the other factors viz. mobility of the target pest/organism, crop species/ canopy
/spacing like sugarcane ,sorghum, maize ,bajara, pigeon pea and cotton etc. ,the plot size
should be 50m2 or more may be taken for trials.
6. The bio efficacy data of those cases which have been rejected by the Registration
Committee in the past due to non conformity of active ingredient contents during the lab
test of the samples, these bioefficacy data will not be considered /accepted for fresh
application of the same product by the same applicant with a new sample test report.
7. Details of Cost: Benefit ratio in terms of input cost, cost of product (Yield/Ha.) should be
submitted to the Registration Authority, in case of combination bio-products. (Input cost
should be inclusive of the cost of cultural, agronomical, mechanical practices as per
prevailing local rates at the site of trial).
8. The test product should be safe to the environment, non target organisms and having
better efficacy and also cost effective in comparison to the standard checks.
9. i) In case of formulations- for new bio-pesticides product/ strain additional data may be
collected/ supported on effects on succeeding crops in the trial field.
iii) Effect on the test product on other non-target organism and eco-friendly insects in
adjacent field crops should also be recorded and submitted to registration authority.
10. All the activities should be well documented and supported with appropriate photographs
authenticated by concerned scientist and the director (Research) may be submitted to the
registration authority.
Experimental design:
I.
Efficacy trials should be designed so that appropriate statistical analysis of the data can
be performed to ensure that any effects attributable to the insecticide can be
distinguished from other forms of experimental variability.
II. A fully randomized design can be applied only if the trial environment/agro ecosystem is
completely homogeneous.
III. Plot size and shape -the most suitable plot size (at least 25m2) depends upon many
factors such as the mobility of the target organism (larger plots are generally needed
for more mobile organisms), Replicates Four.
Untreated control:
i.
An untreated control should be included in the trial.
ii.
Should confirm the presence of an adequate pest infestation and to confirm that there
were no natural reductions in their numbers or levels during the course of the trial.
Application of Insecticide:
1.Treatment application should be made after observing ETL of the target pests in the
proposed crop.
2.
The timing of treatment application should be planned/done keeping in view the
occurrence of pest at a particular crop stage and should be spread over the entire
susceptible/vulnerable stage of the crop.
3.
The equipment and method of application of the insecticide should be the same
In small-scale trials hand-held equipment may be used,
4. Application rates for sprays, the volume application rate and dilution ratios (as appropriate)
should be the same as recommended for use.
5. Treatments doses, time/stage of application (tentative schedule), number of applications,
application intervals, method of application and spray volume should be recommended by
the concerned scientist based on the B/E trials results.
6. Application of equipment should be properly calibrated to give the intended application
rate and droplet spectrum.
7. Details of the sprayer, its operating pressure, nozzle type and position of nozzle(s) relative
to crop, forward speed and volume of application rate need to be specified as well as dose,
spray concentration, and formulation used.
Weather:
1.Weather conditions should be measured at the trial site on the day of treatment. Ambient
temperature, relative humidity, precipitation, wind speed and direction should be recorded,
where relevant.
2.Temperature, relative humidity, precipitation and extreme weather conditions may also
need to be recorded on a regular basis throughout the trial period. The exact parameters and
frequency of recording will depend on the type of crop/pest/product under study.
Biological efficacy:
1.The variables that need to be assessed to determine the efficacy of the plant protection product
will strongly depend on the crop and pest being studied. They may include density or incidence of
the pest, infestation levels, percentage mortality or control, severity of symptoms or damage to the
crop, quantity of yield, quality of the product, etc. should be recorded in the trial reports.
2.Pre-treatment assessments of target pests and non target pests (natural enemies) should be
carried out and recorded invariably in the B/E trails reports/observation tables.
3.Parameters such as population reduction over control expressed as percent mortality,
4. The parasitoids, parasites/predators and pathogens of target pest should be
counted/observed after each application as in case of target pest covered under specific
safe insect order/crop ecosystem and it should be recorded in trails reports/observation
tables.
Insect pest assessment (Time of application: Based on ETL assessment)
Name of the insect
Helicoverpa armigera
Tobacco caterpillar
Pink boll worm
Stem borer
Leaf
folder,caseworm,gall
midge
Whorl maggot
Fruit and shoot borer
DBM
BPH/ WBPH
Formula
As per Henderson and Tilton formula
As per Henderson and Tilton formula
As per Henderson and Tilton formula
As per IRRI
As per IRRI
As per IRRI
As per Abbotts formula
As given
Tap 4 rice hills vigorously count the number of plant
hoppers floating and jumping on the water surface
between the four hills. Ten such places are to be
observed at random within each plot.
0=0, 1=1-10, 3=10-25, 5= 25 to 50, 7=51 to 100
9=more than 100(assessment should be made
between 1-3 days and I week after treatment.
GLH
DBM
Mealy bugs,aphids,scale
insects
Whiteflies
Thrips
Other
insect
pest
(mites/nematodes)
Other insect pest on major
crops
House hold insect pest
Stored grain insect pest
Tap 4 rice hills vigorously count the number of plant
hoppers floating and jumping on the water surface
between the four hills. Ten such places are to be
observed at random within each plot.
0=0, 1=1-5, 3=5-10, 5=11-25, 7=26-50
9=more than 50(assessment should be made between
1-3 days and I week after treatment.
Count the number of caterpillars of different ages on
all plants in each plot.
Assessment:1-3 days after treatment
2nd assessment: 7-14 days after treatment.
As given
Observation to be made on 4 top leaves from 10
randomly selected plants in each treatment. Pre
assessment and Post assessment observations to be
taken.
Observation to be made on15 leaves (Top 5, Middle
5 and Bottom 5) 10 randomly selected plant in each
treatment. Pre Assessment and Post assessment at an
interval of one week may be taken
As given
Not given
Not given
Not given
Phytotoxcity: As given in the protocol.
Mode of application: As given in the protocol.
Time and frequency of application: As given in the protocol.
Dose and Volume used: As given in the protocol.
Results: As given in the protocol.
Statistical analysis: As given
Inferences and conclusion: As given
ADDITIONAL REQUIREMENTS
INSECTICIDES PRODUCTS:
FOR
EVALUATION
OF
EFFICACY
OF
BIO
Antagonistic /Entomo-pathogenic fungi:
1. Information on the particular strain and location from where the strain was isolated (Source of
strain) should be mentioned in the bio efficacy trial reports.
2. Information on soil pH from where the particular strain has been isolated should be given in B/E
trails reports in case of efficacy of nematode species.
3. Information about the isolation of NPV (specific pest species) should be given in case of NPV.
4. Evaluation of bio-efficacy trial of bio-agents on nematodes should be carried –out in vitro.
5. Bio-pesticides product strain should be deposited in National Repository and submit Accession
Number to Sectt. of CIB& RC
6. Bio-pesticides / bio-agent (talc based formulation @ 0.4 -1.0 kg should be enriched /added
with 1 ton of Farm Yard Manure (FYM) or compost and the same should be left under shade for
a period of 15 days, maintaining optimum moisture and in between it should be mixed
thorough at an interval of 5 days for proper enrichment of FYM with this bio-agent.
7. In vivo parasitisation of eggs/ egg masses, hatching suppression of nematodes & computing the
nematodes etc. internationally accepted Methodology should be followed for bio-efficacy
studies should be in the field conditions.
8. Lc50 value for each insect species should be generated in minimum two laboratories of
ICAR/SAU’s/CSIR/ICMR and the same be submitted to the Registration Authority.
9. Data on bio effectiveness and phyto-toxicity generated at ICAR, SAUs, CSIR / ICMR institutes. The data
should be certified either by the Director or Head of the Institute.
10. Data on non-target organisms: One season / one year data on the effect of the
product on natural predators / parasites. In case of nematodes natural soil predatory
nematodes & Natural Rhizospheric soil micro-organism may also be considered.
Test Protocol for bio-efficacy evaluation of fungicide
(Fungicide unit)
The following parameters may be included in the test protocol of Bio-fungicides.
Operation:
Trial protocols should be elaborated, before starting. The organization should elaborate
standard operating procedures (SOPs) or standard modes of operation for testing,
measurements, data collection and reporting procedures or other tasks that are carried
repetitively, to ensure consistency and repeatability of the activities.
Verification:
1. Persons who are scientifically responsible for the trial within the organization should be
able to check and validate the trial throughout its course and ascertain that the protocol
and SOPs are being followed.
2. Organizations should accept at any time inspections that may be organized by the
registration authority or another competent national authority to verify the compliance of
trial/protocols. Organization must intimate to the Secretary CIB&RC about the beginning &
closure of the trials.
3. All trials need proper display. Experimental sites and display boards needs to be
photographed. All spray operations, observations in the experimental fields, from time to
time and crop growth stage from time to time needs to be well documented/ photographed
for submission to registration authority.
Experimental conditions:
1. Bio efficacy trials should strictly be conducted/data generated at SAU/ICAR institutes and
Institutes recognized by ICAR under central universities engaged in field oriented research
trials. A list of all such SAU/ICAR institutes and other institutes attached at Annexure-I.
attached with the insecticide unit protocol .
2. Consistency in methodology of trials/experiments should be followed at all the trial
locations in respect of frequency of recording bio efficacy data, frequency of treatment
applications, Number of trials , number of application and unit of evaluation of bio efficacy
as per standard scale for specific pest.
3. Trials should be conducted in locations which represent the range of agronomic, plant
health, environmental and climatic conditions likely to be encountered in practice in the
area of proposed use.
4. Selection of location for trial should be done keeping in view the prevalence of target
disease on crop species (Endemic areas) in that particular area (identified/ declared by
ICAR/SAUs/Indian published literature).
5. The crops in the experimental locations may be specifically grown for the trials or be part of
crop grown for commercial purposes, but in all cases should be grown according to good
agronomic practice (GAP).
6. Any cultural operations in the fields apart from the one being tested should be according to
Good Agronomic Practice (GAP).
7. If there is a possibility that soil types may affect the efficacy of the fungicide, the various
trial sites should be chosen to be representative of the range of soil types that can be
encountered in the proposed field use of the product.
8. Within each trial site, environmental and agronomic conditions should be as uniform as
possible.
9. In case Infection levels are low or incidence of target insect pest does not occurs at the
location of trial in the season then the trial should be conducted in the next season at the
same location.
Bio-Efficacy evaluation:
1. Brief details of all cultural/agronomical practices, methods of fertilizer doses and its
application etc should be submitted to the registration authority in respect of trial
plots/standard check plots and control plots.
2. All cultural/agronomical practices, fertilizer dosages, irrigation and any other treatment
except the application of test chemical should be at par in trial plots, standard check plots
and control plots, so as level field comparative study of bio efficacy of the test product.
3. Fungicides which were approved/recommended by CIB&RC with doses against specific crop
disease should be taken as standard check during B/E trials. In case of unavailability of such
insecticide, fungicide which have been approved /recommended under AICRP) by
ICAR/SAUs/Agriculture Institutes under Central Universities may be taken as standard
check( Authenticated published copy of such recommendation should be submitted with
reports/data).
4. The bio efficacy data of those cases which have been rejected by the Registration
Committee in the past due to non conformity of active ingredient contents during the lab
test of the samples, these bio efficacy data will not be considered /accepted for fresh
application of the same product by the same applicant with a new sample test report.
5. Details of Cost: Benefit ratio in terms of input cost, cost of product (Yield/Ha.) should be
submitted to the Registration Authority, in case of combination products. (Input cost should
be inclusive of the cost of cultural, agronomical, mechanical practices as per prevailing local
rates at the site of trial).
6. In case the fungicide is applied as a mixture with insecticide / PGR/fertilizer or other
chemical, the compatibility as claimed by the applicant should be supported with the bioefficacy data in respect of other constituents of such mixture should be observed/ recorded
to evaluate the efficacy of the constituent to prove the compatibility claim.
7. The test product should be safe to the environment, non target organisms and having
better efficacy and also cost effective in comparison to the standard checks.
8. All the activities should be well documented and supported with appropriate photographs
authenticated by concerned scientist and the director (Research) may be submitted to the
registration authority.
Experimental design:
i) Efficacy trials should be designed in such way so that appropriate standard statistical
analysis of the data (with minimum degree of freedom) can be performed to ensure that
any effects attributable to the fungicide can be distinguished from other forms of
experimental variability.
ii) A fully randomized design can be applied only if the trial environment/agro ecosystem is
completely homogeneous.
iii) Trial plot should be at least 25m2 in size with four replications each, however depending
upon the other factors viz. mobility of the target disease /organism, crop species/
canopy/spacing like sugarcane, sorghum, maize, bajra, pigeon pea and cotton etc., the plot
size should be 50m2 or more may be taken for trials.
Untreated control:
i) An untreated control should be included in the trial.
Application of fungicide:
1. The tentative schedule of treatment application should be planned/done keeping in view
the occurrence of disease at a particular crop stage and should be spread over the entire
susceptible/vulnerable stage of the crop at all the trial locations.
2. The equipment and method of application of the fungicide may be same in all trials handheld equipment may be used.
3. Application rates for sprays, the volume of application and dilution ratios (as appropriate)
may be as per recommendation for use.
4. Treatments doses, time/stage of application (tentative schedule), number of applications,
application intervals, method of application and spray volume may be recommended by the
concerned scientist based on the B/E trials results.
5. Application equipment should be properly calibrated before the treatment application for
obtaining intended application rate and droplet spectrum.
6. Details of the sprayer, its operating pressure, nozzle type and position of nozzle(s) relative
to crop, forward speed and volume of application rate are need to be specified by the
concerned scientist in the recommendation.
Weather:
1. Weather conditions viz. Ambient temperature, relative humidity, precipitation (if any), wind
speed at the time of application may be recorded on the day of treatment.
2. Temperature, relative humidity, precipitation and extreme weather conditions may also
need to be recorded on a regular basis throughout the trial period. The exact parameters
and frequency of recording will depend on the type of crop/pest/product under study.
Biological efficacy:
1. The variables that need to be assessed to determine the efficacy of the plant protection
product will strongly depend on the crop and pest being studied. They may include density
disease infestation levels, percentage mortality or control, severity of symptoms or damage
to the crop, quantity of yield, quality of the product, etc. should be recorded in the trial
reports.
2. Pre-treatment assessments of target disease and non target disease & pollinators should be
carried out and recorded invariably in the B/E trails reports/observation tables.
3. The pathogens of target pest should be counted/observed before & after each application
as in case of target pest covered crop ecosystem and it should be recorded in trails
reports/observation tables.
Test Protocol for bio-efficacy evaluation of
Herbicides/PGR
The following parameters may be included:
Operation:
Trial protocols should be elaborated, before starting. The organization should elaborate
standard operating procedures (SOPs) or standard modes of operation for testing,
measurements, data collection and reporting procedures or other tasks that are carried
repetitively, to ensure consistency and repeatability of the activities.
Verification:
1. Persons who are scientifically responsible for the trial within the organization should be
able to check and validate the trial throughout its course and ascertain that the protocol
and SOPs are being followed.
2. Organizations should accept at any time inspections that may be organized by the
registration authority or another competent national authority to verify the compliance of
trial/protocols. Organization must intimate to the Secretary CIB&RC about the beginning &
closure of the trials.
3. All trials need proper display. Experimental sites and display boards needs to be
photographed. All spray operations, observations in the experimental fields, from time to
time and crop growth stage from time to time needs to be well documented/ photographed
for submission to registration authority.
Experimental conditions:
1. Bio efficacy trials should strictly be conducted/data generated at SAU/ICAR institutes and
Institutes recognized by ICAR under central universities engaged in field oriented research
trials.
2. Consistency in methodology of trials/experiments should be followed at all the trial locations in
respect of frequency of recording bio efficacy data, frequency of treatment applications,
Number of trials , number of application and unit of evaluation of bio efficacy as per standard
scale for specific pest.
3. Trials should be conducted in locations which represent the range of agronomic, plant health,
environmental and climatic conditions likely to be encountered in practice in the area of
proposed use.
4. Selection of location for trial should be done keeping in view the prevalence of target pest
on crop species in that particular area (identified/ declared by ICAR/SAUs/Indian published
literature).
5. The crops in the experimental locations may be specifically grown for the trials or be part of
crop grown for commercial purposes, but in all cases should be grown according to good
agronomic practice (GAP).
6. Any cultural operations in the fields apart from the one being tested should be according to
Good Agronomic Practice (GAP).
7. If there is a possibility that soil types may affect the efficacy of the insecticide, the various
trial sites should be chosen to be representative of the range of soil types that can be
encountered in the proposed field use of the product.
8. Within each trial site, environmental and agronomic conditions should be as uniform as
possible.
Bio-Efficacy evaluation:
1. Effects on non-target weeds.
3. Brief details of all cultural/agronomical practices, methods of fertilizer doses and its
application etc should be submitted to the registration authority in respect of trial
plots/standard check plots and control plots.
4. All cultural/agronomical practices, fertilizer dosages, irrigation and any other treatment
except the application of test chemical should be at par in trial plots, standard check plots
and control plots, so as level field comparative study of bio efficacy of the test product.
5. Insecticides which were approved/recommended by CIB&RC with doses against specific
crop pest should be taken as standard check during B/E trials. In case of unavailability of
such insecticide, insecticide which have been approved /recommended under AICRP) by
ICAR/SAUs/Agriculture Institutes under Central Universities may be taken as standard check
( Authenticated published copy of such recommendation should be submitted with
reports/data).
6. The bio efficacy data of those cases which have been rejected by the Registration
Committee in the past due to non conformity of active ingredient contents during the lab
test of the samples, these bio efficacy data will not be considered /accepted for fresh
application of the same product by the same applicant with a new sample test report.
7. Details of Cost: Benefit ratio in terms of input cost, cost of product (Yield/Ha.) should be
submitted to the Registration Authority, in case of combination products. (Input cost should
be inclusive of the cost of cultural, agronomical, mechanical practices as per prevailing local
rates at the site of trial).
8. In case the herbicide is applied as a mixture with other chemical, the compatibility as
claimed by the applicant should be supported with the bio-efficacy data in respect of other
constituents of such mixture should be observed/ recorded to evaluate the efficacy of the
constituent to prove the compatibility claim.
9. The test product should be safe to the environment, non target organisms and having
better efficacy and also cost effective in comparison to the standard checks.
10. i) In case of formulations- for new molecule additional data may be collected/
supported on effects on succeeding crops in the trial field.
iii. Effect on the test product on other non-target organism and eco-friendly weeds in
adjacent field crops should also be recorded and submitted to registration authority.
11. All the activities should be well documented and supported with appropriate photographs
authenticated by concerned scientist and the director (Research) may be submitted to the
registration authority.
Experimental design:
iv.
v.
vi.
Efficacy trials should be designed in such way so that appropriate standard statistical
analysis of the data (with minimum degree of freedom) can be performed to ensure that
any effects attributable to the insecticide can be distinguished from other forms of
experimental variability.
A fully randomized design can be applied only if the trial environment/agro ecosystem is
completely homogeneous.
Trial plot should be at least 25m2 in size with Three replications each, however
depending upon the other factors viz., crop species/ canopy/spacing like sugarcane,
sorghum, maize, bajra, pigeon pea and cotton etc., the plot size should be 50m2 or more
may be taken for trials.
Untreated control:
vii.
An untreated control should be included in the trial.
viii.
Should confirm the presence of an adequate pest population/infestation and all
activities under GAP should be at par with the treated plots except the application of
test chemicals during the course of the trial.
ix.
In case there is reduction in population of weeds, reason may be recorded thereof in the
trial data.
Application of Herbicide:
1.Treatment application should be made as recommendation made in the L/L.
2. The tentative schedule of treatment application should be planned/done keeping in view
stage of crop and weeds at a particular crop stage and should be spread over the entire
susceptible/vulnerable stage of the crop at all the trial locations.
3. The equipment and method of application of the insecticide may be same in all trials handheld equipment may be used.
4. Application rates for sprays, the volume of application and dilution ratios (as appropriate)
may be as per recommendation for use.
5. Treatments doses, time/stage of application (tentative schedule), number of applications,
application intervals, method of application and spray volume may be recommended by the
concerned scientist based on the B/E trials results.
6. Application equipment should be properly calibrated before the treatment application for
obtaining intended application rate and droplet spectrum.
7. Details of the sprayer, its operating pressure, nozzle type and position of nozzle(s) relative
to crop, forward speed and volume of application rate are need to be specified by the
concerned scientist in the recommendation.
Weather:
1. Weather conditions viz. Ambient temperature, relative humidity, precipitation (if any),
wind speed at the time of application may be recorded on the day of treatment.
2. Temperature, relative humidity, precipitation and extreme weather conditions may also
need to be recorded on a regular basis throughout the trial period. The exact parameters
and frequency of recording will depend on the type of crop/pest/product under study.
Bio-efficacy:
1. The variables that need to be assessed to determine the efficacy of the plant protection
product will strongly depend on the crop and pest being studied. They may include density,
infestation levels, percentage control, damage to the crop, quantity of yield, quality of the
product, etc. should be recorded in the trial reports.
2 .Pre-treatment assessments of target and non target weeds.
3. Parameters such as population reduction over control expressed
Phytotoxcity: As given
Mode of application: As given
Time and frequency of application: As given
Dose and Volume used: As given
Results: As given
Statistical analysis: As given
Experimental Condition:
1.1 - Target Weeds with others Predominant Weeds in the Experimental crop.
1.2 – Type of Land- low Land , Upland or Dry Land.
1.2- Trial Condition:
Field trial should be set up major crop growing area and where the weeds become problem along
with Target Weeds and other condition mentioned in Note. Trial should be carried out in
different agro-climatic regions as specified by ICAR or Planning commission.
Design and Lay Out :
Test product(s), reference product(s) and untreated control, arranged in a randomized block design or any
other suitable statistical design which represent the whole field.
1.3.2
Plot size and replication
Net plot size at least 20 sq.m. per replication per treatment but according to the formulation or
application equipment, it may be necessary to use a larger plot size (at least 25sq m.).The plot size must
be the same at all location.
Replicates – at least three, provided the error degrees of freedom are at least 12. Minimum 7
treatments in which one standard check, one Weed Free and one Weedy Check must be included with
minimum 4 treatment of test product.
2.1
Test product (s)
The formulated product under evaluation minimum Three treatments.
2.3
Method of application
Application should comply with good agricultural practices. Details on the spray equipment, type
of nozzle (Flat fan or flood jet) and water volume used at different stages of crop growth should
be given. Addition of sticker, to improve the uptake of active ingredient, if any must be specified.
Sticker should be tried in the check plot with water spray. The above requirement should be
followed uniformly in all location in both the season.
2.3.2
Time and frequency of application
The time and frequency of application will normally be specified by the manufacturer/
sponsor. The time of application should be conveniently mentioned based on the time of
weed emergence (PPI, Pre-emergence, early post emergence or late post emergence) or
based on the stage of the weed (1-2 leaf stage, 3-4 leaf stage etc) or DAS (days after
sowing).
3.3 – Irrigation --- The crop are irrigated, Rain fed or Dry land forming.
4 --- Observation :
4.1.--- Weed Intensity :
4.1.1 --- Weed Count ; The number of species wise Weeds to be determine at pre-spray , 30 DAS , 60
DAS and at Harvest at about 5 location per plot using a squire quadrate , 1 sq. m. and statistically
analysed after transformation.
4.1.2 --- Weed Dry matter – Dry or/ fresh weight of weeds species wise to be determine at pre-spray,
30 DAS, 60 DAS and at Harvest and statistically analysed after transformation.
4.1.3 ----- Percent Weed Control efficiency (WCE) : Weed control efficiency should be mentioned
species wise as per label claim at 30 DAS , 60 DAS and at Harvest.
WDM in UTC --- WDM in treated
WCE =
--------------------------------------------------------- x 100
WDM in UTC
WDM == Weed Dry Matter.
UTC = Untreated control.
4.3 -- Yield and Yield attributing: character with statistical analysed. Enhancement of the crop has to
be brought out using appropriate statistical and other tools so that there is no confusion statement in
drawing the inference.
Phytotoxicity ;
Photographs at each observation with label.
Recovery Photographs of initial phytotoxicity and their recovery.