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LEAF BLIGHT OF ROSE
Sensitivity of Botrytis cinerea to Carbendazim.
(IN VITRO STUDY)
Rosa floribunda plants exhibiting the leaf blight symptoms were collected from
different localities of Maharashtra from polyhouses and gardens. From these samples
ten isolates of Botrytis cinerea were obtained.
Sensitivity of Botrytis cinerea to carbendazim was determined by food
poisoning test ( Dakker and Gielink1979).The agar plates were prepared containing
different concentration of carbendazim. Disc (8mm) with fungal cultures were taken
from the margin of an actively growing colony and placed on agar surface, the plates
were then inoculated at 28+2 ºC in dark and linear growth was measured at different
time intervals .Plates without fungicides served as control.
(IN VIVO) STUDIES.
The sensitivity of ten isolates of Botrytis cinerea to carbendazim was
determined on rose plants. These plants were grown in earthen pots and kept in
departmental wire house. The different concentrations of fungicides were sprayed on
three month old rose plant. After 24 hrs. of fungicide treatment Botrytis cinerea
spore suspension was inoculated on the plants with the help of 0.5 mm Camlin hair
brush. The plants were covered with polyethen bags to maintain the r.h. The plants
sprayed with distilled water were treated as control .The plants without any treatment
were served as absolute control. After 10 days of inoculation the percentage of
infection of the disease was recorded, by following literature Kareem(2007)
It was observed that there was variation in MIC of the carbendazim against
Botrytis cinerea on Rosa floribunda (Table 48). The isolate BC-9 (MIC-60) from
156
Shindhudurg was most sensitive to carbendazim and isolate BC-8 (MIC-800) from
Pune was most resistant to carbendazim.
Dose response curve of Botrytis cinerea isolates to carbendazim are shown in
Figs. 1 to 5. In in vitro it appeared that with the increase concentration of carbendazim
in medium, there was decrease in the radial growth of Botrytis cinerea there are five
group of Botrytis cinerea isolates According to their sensitivity to carbendazim.
Group-A MIC from 100to200 µg/ml (BC-9,10), Group-B ranging MIC from 300 to
400 µg/ml- (BC-3,BC-6,BC-7) , Group-C- Ranging MIC from 500 to 600 µg/ml
(BC-4)., Group-D- Ranging MIC from700 to 800 µg/ml and (BC-2,BC-5) Group-E
Ranging MIC from 900 to 1000 µg/ml. (BC-1,BC-08 ) Isolate BC-8 from Pune was
highly resistant with MIC 1000 µg/ml. Somewhat similar results were obtained in in
vivo experiments.
157
Table 48: MIC of Carbendazim against Botrytis cinerea isolates causing leaf
blight of Rosa floribunda .
Sr.No. Locality Isolate In Vitro In Vivo
1 Kolhapur BC-1 900 µg/ml 700 µg/ml
2 Sangali BC-2 800 µg/ml 600µg/ml
3 Satara BC-3 300 µg/ml 200 µg/ml
4 Solapur BC-4 500 µg/ml 400 µg/ml
5 Nagpur BC-5 700 µg/ml 500 µg/ml
6 Aurangabad BC-6 400 µg/ml 300 µg/ml
7 Ratnagiri BC-7 300 µg/ml 250 µg/ml
8 Pune BC-8 1000 µg/ml 800 µg/ml
9 Shindhudurg BC-9 100 µg/ml 60 µg/ml
10 Osmanabad BC-10 200 µg/ml 150 µg/ml
158
0
10
20
30
40
50
60
70
80
90
contro
l 25 50 7510
0125 150 175 200 225
concentration of carbendazim in PPM
Line
argr
owth
inm
m
BC-9
BC-10
Fig : 56 Dose response curve of Botrytis cinerea isolates (BC-9 andBC-10) to various concentrations of carbendazim on agar plates.
0
10
20
30
40
50
60
70
80
90
contro
l 50 100 150
200 250 300
350 400 450 500
Concentration of carbendazim in PPM
Line
argr
owth
inm
m
BC-3
BC-6
BC-7
Fig : 57 Dose response curve of Botrytis cinerea (BC-3, BC-6 and BC-7)Isolates to various concentrations of carbendazim on agar plates.
159
0
10
20
30
40
50
60
70
80
90
contro
l100 200 300 40
0500 600 700
Concentration of carbendazim in PPM
Linea
rgro
wth
inm
m
BC-4
Fig : 58 Dose response curve of Botrytis cinerea (BC-4) isolatesto various concentrations of carbendazim on agar plates.
0
10
20
30
40
50
60
70
80
90
control
100200
300400
500 600700
800900
Concentration of carbendazim In PPM
Line
argr
owth
inm
m
BC_2
BC-5
Fig : 59 Dose response curve of Botrytis cinerea (BC-2and BC-5)isolate to various concentrations of carbendazim on agar plates.
160
0
10
20
30
40
50
60
70
80
90
contro
l10
0200 300 400 50
0600 700 800 90
010
0011
00
Concentration of carbendazim in PPM
Line
argr
owth
inm
m
BC-1
BC-8
Fig : 60 Dose response curves of Botrytis cinerea (BC-1 and, BC-8)isolates to various concentrations of carbendazim on agar plates.
0
20
40
60
80
100
120
Contro
l10
020
030
040
050
0600
concentration of carbendazim in ppm
Perc
enta
geof
infe
ctio
n
BC-4
Fig : 61 Dose response curve of Botrytis cinerea (BC-4) isolateto various concentrations of carbendazim (in vivo).
161
0
20
40
60
80
100
120
Control
100 200
300
400
500 600 700 800 900
Concentration of carbendazim in ppm
Perc
enta
geof
infe
ctio
n
BC-5
BC-2
BC-1
BC-8
Fig : 62 Dose response curves of Botrytis cinerea isolates (BC-5, BC-2,BC-1 and BC-8) to various concentrations of carbendazim (in vivo).
0
20
40
60
80
100
120
Contro
l 50 100
150
200 250 300 350
Concentration of carbendazim in ppm
Perc
enta
geof
infe
ctio
n BC-6
BC-7
Fig : 63 Dose response curves of Botrytis cinerea (BC-6and BC-7)isolates to various concentrations of carbendazim (in vivo).
162
0
20
40
60
80
100
120
Contr
ol 25 50 75100 12
515
017
520
022
5
Concentration of carbendazim in ppm
Perc
enta
geof
infe
ctio
n
BC-3
BC-10
Fig : 64 Dose response curve of Botrytis cinerea (BC-3 and BC-10)isolate to various concentrations of carbendazim (in vivo).
0
20
40
60
80
100
120
Control 10 20 30 40 50 60 70 80
concentration of carbendazim in ppm
Perc
enta
geof
infe
ctio
n
BC-9
Fig : 65 Dose response curve of Botrytis cinerea (BC-9) isolate to variousconcentrations of carbendazim (in vivo).
163
Effect of passage on development of carbendazim resistance in Botrytis cinerea
In vitro studies:-
Effect of continuous and alternate treatment of two fungicides with different
mode of action and a mixture of both on the development of carbendazim resistance
in Botrytis cinerea was studied In vitro. For this wild sensitive isolate (BC-9) was
cultured for eight successive passages on Czapek Dox Agar plates containing
Benomyl, Ridomil MZ ,Kocide, and Dhanuka alternating or mixed with carbendazim.
The concentration of fungicides was kept constant for all the passages.
Continuous and alternate treatment with carbendazim:-
Wild sensitive isolate BC-9 was cultured on plates containing carbendazim at
MIC level 100 μg/ml for eight successive passages. An 8 mm diameter agar disc from
the margin of actively growing colony taken from culture of the previous passage was
placed at the centre of each plate in triplicate. In each passage linear mycelium growth
was measured after 4 days.
The increase or decrease in the radial growth from passage to passage was
employed as a criterion for the development of carbendazim resistance. It was seen
that growing of Botrytis cinerea on the medium containing carbendazim for eight
successive passages continuously significantly increased the carbendazim resistance.
When the pathogen was cultured alternately with other fungicides there was
decrease in the development of carbendazim resistance. Carbendazim when used
alternately with Benomyl and Ridomil there was complete inhibitions of Botrytis
cinerea at 2nd passage only, Dhanuka inhibited the growth at fourth passage and
Kocide inhibited the growth of the pathogen at the sixth passage.
(Table 49. and Fig.66)
164
Table 49: Effect of exposure of Botrytis cinerea (In vitro) to Carbendazim
continuous and alternating with other fungicides on the development of resistance
(growth in mm) during eight successive passages.
Fungicide 1 2 3 4 5 6 7 8
Carbendazim
Individual
12.00 12.66 14.66 18.66 20.66 23.66 26.66 30.66
Cabendazim
Alt. Benomyl
12.00 00.00 00.00 00.00 00.00 00.00 00.00 00.00
Carbendazim
Alt. Ridomil,
12.00 00.00 00.00 00.00 00.00 00.00 00.00 00.00
Carbendazim
Alt.
kocide
12.00 13.33 11.33 10.33 09.00 00.00 00.00 00.00
Carbendazim
Alt.
Dhanuka
12.00 11.33 10.00 00.00 00.00 00.00 00.00 00.00
Mean ± SE ( standard error)Continuous carbendazim = 19.95 ± 2.39Carbendazim Alter Benomyl = 1.50 ± 1.5Carbendazim Alter Ridomil = 1.50 ± 1.50Carbendazim Alter Kocide = 6.99 ± 2.09Carbendazim Alter Dhanuka = 4.16 ± 2.04
0
5
10
15
20
25
30
35
1 2 3 4 5 6 7 8
Passage number
Line
argr
owth
inm
m
idividual carbendazim
Carbendazim alterRidomil
carbendazim alterBinomyl
Carbendazim alterKocide
carbendazim alterDhanuka
Fig: 66 Effect of exposure of Botrytis cinerea to carbendazimContinuous and alternating with other fungicides on the development ofResistance during eight successive passages (In Vitro)
165
Treatment of carbendazim in mixture:-
It was interesting to note that carbendazim when used along with Benomyl, and
Ridomil MZ there was complete inhibition of the pathogen at the first passage only
.Dhanuka inhibited the growth of the pathogen at the third passage. While Kocide
inhibited the growth of Botrytis cinerea at the fourth passage.
Table 50. Effect of exposure of Botrytis cinerea to the mixture of carbendazim with otherfungicides on the development of resistance during eight successive passagesFungicide 1 2 3 4 5 6 7 8
Carbendazim
Individual10.00 11.66 14.00 17.33 19.33 25.00 28.66 32.66
Carbendazim+ benomyl 00.00 00.00 00.00 00.00 00.00 00.00 00.00 00.00
Caebendazim+ ridomil, 00.00 00.00 00.00 00.00 00.00 00.00 00.00 00.00
Carbendazim+ Kocide 17.33 17.00 15.33 00.00 00.00 00.00 00.00 00.00
Carbendazim+Dhanuka 13.00 12.33. 00.00 00.00 00.00 00.00 00.00 00.00
Mean ± SE (standard error)Continuous carbendazim = 19.83 ± 2.90Carbendazim + Benomyl = 00.00 ± 00Carbendazim + Ridomil = 00.00 ± 00Carbendazim + Kocide = 6.20 ± 3.03Carbendazim +Dhanuka = 3.16 ± 2.07
0
5
10
15
20
25
30
35
1 2 3 4 5 6 7 8
Passage number
Line
argr
owth
inm
m
IndividualcarbendazimCarbendazim+BenomylCarbendazim +Ridomil
carbendazim + Kocide
carbendazim +Dhanuka
Fig : 67 Effect of exposure of Botrytis cinerea to the mixture of carbendazim withother fungicides on the development of resistance during eight successive passages(In Vitro)
166
In Vivo studies:
1. Continuous passage:
To study the effect of continuous passage, on the development of fungicide
resistance, in the pathogen, in vivo, 10 ml mycelial suspension of one culture tube of
wild sensitive isolate ( BC-9) was inoculated on the healthy rose plant, treated with 60
µg/ml Carbendazim, 24hrs before, continuously for eight successive passages. It was
observed that there was increased rate of infection to the rose plant.
2. Alternate passage:
To study the effect of alternate passage, on the development of fungicide
resistance, in the pathogen, in vivo, 10ml mycelial suspension of one culture tube of
wild sensitive isolate (BC-9) was inoculated on healthy plant of Rosa floribunda
treated with carbendazim altering with Benomyl, Ridomil MZ, Kocide, and Dhanuka
24hrs before, continuously for eight successive passages. Benomyl and Ridomil
prevented the infection of the pathogen at the second passage only. Kocide and,
Dhanuka prevented the infection of the pathogen at the third passage.
(Table 51 and Fig.68)
167
Table 51: Effect of exposure of Botrytis cinerea (In vivo) to Carbendazimcontinuous and alternating with other fungicides on the development ofresistance (growth in mm) during eight successive passages.
Fungicide 1 2 3 4 5 6 7 8
%ofinfection
10.00 15.00 20.00 25.00 30.00 40.00 50.00 60.00CarbendazimIndividual
grade 01 01 01 01 02 02 02 03
%ofinfection
10 00.00 00.00 00.00 00.00 00.00 00.00 00.00CabendazimAlt.Benomyl grade 01 00.00 00.00 00.00 00.00 00.00 00.00 00.00
%ofinfection
10 00.00 00.00 00.00 00.00 00.00 00.00 00.00CarbendazimAlt.Ridomil, grade 01 00.00 00.00 00.00 00.00 00.00 00.00 00.00
%ofinfection
10.00 05.00 00.00 00.00 00.00 00.00 00.00 00.00CarbendazimAlt.Kocide grade 01.00 01.00 00.00 00.00 00.00 00.00 00.00 00.00
%ofinfection
10.00 05.00 00.00 00.00 00.00 00.00 00.00 00.00CarbendazimAlt.Dhanuka grade 01.00 01.00 00.00 00.00 00.00 00.00 00.00 00.00
0
10
20
30
40
50
60
70
1 2 3 4 5 6 7 8
passage number
Per
cent
age
ofin
fect
ion
idividual carbendazim
Carbendazim alterRdomilCarbendazim AlterBinomylCarbendazim alterKocideCarbendazim alterDhanuka
Fig: 68 Effect of exposure of Botrytis cinerea to carbendazim continuous andalternating with other fungicides on the development of resistance during eightsuccessive passages (In Vivo).
168
3. Mixed Passage:
To study the effect of mixed passage, on the development of fungicide
resistance, in the pathogen, in vivo, 10 ml mycelial suspension of one culture tube of
wild sensitive isolate (BC-9) was inoculated on the healthy Plant, the plants were
treated with solution of carbendazim along with Benomyl, Ridomil MZ Kocide, and
Dhanuka in equal volume, 24hrs before, this was repeated for eight successive
passages.
In each type of passage mentioned above, the plants were covered with
polythene bags. Percentage of the infection on rose plant were recorded after each
passage and used as a criterion for the development of resistance in the pathogen.
Binomyl, Ridomil in mixture with carbendazim prevented the infection of the
pathogen to the rose plant at first passage .While kocide and Dhanuka prevented the
infection of the pathogen at the second passage. (Table 52 and Fig. 69)
169
Table 52.Effect of exposure of Botrytis cinerea (In Vivo) to the mixture of
Carbendazim with other fungicide on the development of resistance during eight
successive passages
Fungicide 1 2 3 4 5 6 7 8
%ofinfection
10.00 15.00 20.00 25.00 30.00 40.00 50.00 60.00CarbendazimIndividual
grade 01 01 01 01 02 02 02 03
%ofinfection
00.00 00.00 00.00 00.00 00.00 00.00 00.00 00.00Cabendazim+ Benomyl
grade 00.00 00.00 00.00 00.00 00.00 00.00 00.00 00.00
%ofinfection
00.00 00.00 00.00 00.00 00.00 00.00 00.00 00.00Carbendazim+ Ridomil,
grade 00.00 00.00 00.00 00.00 00.00 00.00 00.00 00.00
%ofinfection
15.00 00.00 00.00 00.00 00.00 00.00 00.00 00.00Carbendazim+Kocide
grade 01.00 00.00 00.00 00.00 00.00 00.00 00.00 00.00
%ofinfection
10.00 00.00 00.00 00.00 00.00 00.00 00.00 00.00Carbendazim+Dhanuka
grade 01.00 00.00 00.00 00.00 00.00 00.00 00.00 00.00
0
10
20
30
40
50
60
70
1 2 3 4 5 6 7 8
Passage number
Perc
enta
geof
infe
ctio
n
IndividualcarbendazimCarbendazim +BenomylCarbendazim +Ridomilcarbendazim +KocideCarbendazim +Dhanuka
Fig: 69 Effect of exposure of Botrytis cinerea to the mixture of carbendazim withother fungicides on the development of resistance during eight successive passages(In Vivo)
170
Physiology of sensitive and resistant isolates of Botrytis cinerea.
Effect of different sources like Carbon, Nitrogen, Phosphorus, Sulphate, Salts,
Micronutrients, Vitamins and Amino acids on the growth of Botrytis cinerea was
studied by amending them in the Czapek Dox Agar medium. Growth of sensitive and
resistant isolates of Botrytis cinerea was compared with control.
Carbohydrate nutrition:-
Various carbohydrates (3%) were incorporated in the CDA medium and linear
growth of sensitive BC-9 and resistant BC-8 isolate was recorded at various intervals.
It appeared that sugars are essential for growth of both the isolates of Botrytis cinerea
.There was highly significant increase in the growth of both the isolates when
compared with control. The growth rate of resistant BC-8was always higher than the
sensitive BC-9 isolate on the all sugars. Sucrose. Maltose, fructose, Dextrose,
stimulated the growth of sensitive and resistant isolates of Botrytis cinerea. While
Lactose inhibited the growth both the isolates of Botrytis cinerea.
(Table 53 and Fig.70 and 71)
Nitrogen nutrition:-
Peptone, ammonium nitrate, potassium nitrate and sodium nitrate were
incorporated in medium at 0.2%. There was slightly variation in the growth of both
the isolates, between nitrogen sources. Growth rate of resistant strain was found to be
always higher than the sensitive one. Growth rate of the resistant and sensitive
isolates were found to be higher on Peptone, sodium and ammonium nitrate. These
three nitrogen sources stimulate the growth of Botrytis cinerea. While potassium
nitrate inhibited the growth of both the isolates of Botrytis cinerea
(Table 54 and Fig.72 and 73)
171
Phosphate nutrition:-
Three phosphate sources at 0.1% concentration were used in this study. There
was slight variation in the growth of both the sensitive and resistant isolate at various
incubation periods. Ammonium dihydrogen orthophosphate and sodium dihydrogen
orthophosphate were inhibitory to the growth of sensitive and resistant isolates of
Botrytis cinerea. While potassium dihydrogen orthophosphate stimulated the growth
of sensitive and resistant isolates Botrytis cinerea. (Table 55 and Fig.74and 75 )
Sulphate nutrition:-
Magnesium sulphate and potassium sulphate were incorporated in the medium
at 0.05%. There was highly significant variation in the growth of both the sensitive
and resistant isolates between various sulphate sources and various incubation
periods. Among these sulphate sources it is found that Magnesium sulphate stimulated
the growth of the sensitive and resistant isolates. But potassium sulphate inhibited the
growth of the both isolates. (Table-56 and Fig. 76 and 77)
Effect of salts:-
Altogether 4 salts were used in this study. Magnesium chloride, Potassium
chloride, , sodium chloride, and calcium chloride were incorporated in medium at
0.05 % concentration and discs (8 mm) of sensitive and resistant isolates were
inoculated on the plates. There was variation in growth of both the isolates between
various salts sources and various incubation periods. Growth of resistant isolate was
found to be higher than the sensitive .All salts stimulated the growth Botrytis cinerea.
(Table-57 and Fig. 78 and 79)
172
Vitamin nutrition:-
Four vitamins were tested for the growth of the sensitive and resistant isolate at
the 0.01% concentration amended in CDA medium. There was variation in growth of
both the sensitive and resistant isolates between various incubation periods. Resistant
isolate showed higher growth rate than the sensitive isolate. All Vitamins stimulated
the growth of both the isolates of Botrytis cinerea
(Table-58 and Fig. 80 and 81).
Effect of Micronutrients:-
Mn, Cu, Co, Zn and Mo were tested for the growth of sensitive and resistant
isolate of Botrytis cinerea to carbendazim. Manganese stimulated the growth of
Pathogens while Copper, cobalt, zinc, and Molybdenum reduced the growth of both
the sensitive and resistant isolates of Botrytis cinerea
(Table-59and Fig. 82 and 83)
173
Amino acid nutrition:-
Altogether nine amino acids were used in this study. There was variation in the
growth of the sensitive and resistant isolates on different amino acids. T Cystine
Tryptophan and Tyrosine reduced the growth of the pathogen, While Glycine,
Phenylalanine, Proline, Serine, Alanine, and Histidine stimulated growth of both the
sensitive and resistant isolates of Botrytis cinerea.
(Table-60 and Fig. 84 and 85)
174
Physiology of Sensitive and Resistant Isolates of Botrytis cinerea.
Table 53: Effect of different Carbon sources on the linear growth (mm) of
Botrytis cinerea, sensitive and resistant isolates to carbendazim on CDA medium.
Sensitive isolate Resistant isolateDays DaysSource
ofcarbon3%
1 2 3 4 1 2 3 4
Maltose 17.33 36.33 51.66 71.33 19.66 38.33 55.66 73.33
Fructose 15.66 33.33 48.66 64.66 17.66 36.33 53.66 67.33
Dextrose 16.33 33.66 49.33 67.33 17.33 35.33 53.66 69.66
Lactose 00.00 00.00 00.00 00.00 00.00 00.00 00.00 00.00
Sucrose 20.33 39.33 59.66 79.66 21.33 41.66 62.33 80.00
Control 12.00 21.00 33.66 45.33 13.33 23.66 35.00 48.66
Followed Tukey
Sensitive isolate (BC-9) Resistant isolate(BC-8)
Between sources F=14.87 Between sources F=16.06
C.D. at p value 0.05=28.05 C.D. at p value 0.05=28.56
175
0
10
20
30
40
50
60
70
80
90
1 2 3 4
Days
Line
argr
owth
inm
m
Control
Maltose
Fructose
Dextrose
Lactose
Sucrose
Fig : 70 Effect of different Carbon sources on the linear growth (mm) of Botrytis
cinerea sensitive isolate to carbendazim on Czapek Dox Agar Medium.
0
10
20
30
40
50
60
70
80
90
1 2 3 4
Days
linea
rgro
wth
inm
m
Control
Maltose
Fructose
Dextrose
Lactose
Sucrose
Fig : 71 Effect of different Carbon sources on the linear growth (mm) of Botrytiscinerea resistant isolate to carbendazim Czapek Dox Agar Medium.
176
Table 54: Effect of different nitrogen sources on the linear growth (mm) ofBotrytis cinerea, sensitive and resistant isolates to carbendazim on CDA medium.
Sensitive isolate Resistant isolateDays DaysNitrogen
sources0.2%
1 2 3 4 1 2 3 4
Peptone 17.66 36.33 54.33 76.33 18.66 37.66 56.66 78.66
AmmoniumNitrate
16.33 34.66 53.33 75.66 19.33 36.33 55.66 77.66
Potassiumnitrate
14.33 28.33 45.66 59.66 15.66 30.66 47.33 62.66
Sodiumnitrate
19.33 38.66 56.33 78.66 20.33 40.66 59.66 80.00
Control 15.33 32.66 49.66 72.66 17.33 35.33 51.66 74.33
Followed Tukey
Sensitive isolate (BC-9) Resistant isolate (BC-8)
Between sources F=11.5 between sources F=12.76
C.D. at p value 0.05=38.17 C.D. at p value 0.05=38.47
177
0
10
20
30
40
50
60
70
80
90
1 2 3 4
Days
Line
argr
owth
inm
m
control
Peptone
Ammonium nitrate
Potassium nitrate
sodium nitrate
Fig :72 Effect of different nitrogen sources on the linear growth (mm) of Botrytiscinerea sensitive isolate to carbendazim Czapek Dox Agar Medium.
0
10
20
30
40
50
60
70
80
90
1 2 3 4
Days
Linea
rgro
wth
inm
m
C0ntrol
Peptone
Amonium nitrate
Potassium nitrate
Sodium nitrate
Fig : 73 Effect of different nitrogen sources on the linear growth (mm) of Botrytiscinerea resistant isolate to carbendazim Czapek Dox Agar Medium.
178
Table 55: Effect of different phosphate sources on the linear growth (mm) ofBotrytis cinerea, sensitive and resistant isolates to carbendazim on CDA medium.
Sensitive isolate Resistant isolate
Days DaysPhosphatesources0.1%
1 2 3 4 1 2 3 4
Potassiumdihydrogenorthophosphate
20.33 38.33 59.66 78.33 21.33 40.33 62.66 79.33
Sodiumdihydrogenorthophosphate
16.33 31.33 46.66 63.66 18.66 33.66 47.66 66.33
Ammoniumdihydrogenorthophosphate
15.00 29.33 45.66 62.00 16.66 33.66 49.66 65.66
Control 19.33 36.33 55.33 71.66 21.66 38.33 57.66 72.66
Followed Tukey
Sensitive isolate (BC-9) Resistant isolate(BC-8)
Between sources F=18.20 Between sources F=13.70
C.D. at p value 0.05=35.14 C.D. at p value 0.05=35.38
179
0
10
20
30
40
50
60
70
80
90
1 2 3 4
Days
Line
argr
owth
nin
mm
Control
Potassiumdihydrogenorthophosphate
Sodium dihydrogenorthophosphate
Ammoniumdihydrogenorthophosphate
Fig : 74 Effect of different phosphate sources on the linear growth (mm) of Botrytiscinerea sensitive isolate to carbendazim Czapek Dox Agar Medium.
0
10
20
30
40
50
60
70
80
90
1 2 3 4
Days
Line
argr
owth
inm
m
Control
Potassium dihydrogenorthophosphate
Sodium dihydrogenorthophosphate
Ammoniumdihydrogenorthophosphate
Fig : 75 Effect of different phosphate sources on the linear growth (mm) ofBotrytis cinerea resistant isolate to carbendazim Czapek Dox Agar Medium.
180
Table 56: Effect of different sulphate sources on the linear growth (mm) ofBotrytis cinerea, sensitive and resistant isolates to carbendazim on CDA medium..Sensitive isolate Resistant isolate
Days DaysSulphatesources0.05%
1 2 3 4 1 2 3 4
MagnesiumSulphate
19.66 38.33 57.33 78.66 21.33 40.33 59.66 80.00
Potassiumsulphate
13.00 28.66 43.66 60.33 15.33 30.33 45.00 62.66
.control 18.66 33.66 51.66 71.66 19.66 36.66 55.66 73.66
Followed Tukey
Sensitive isolate (BC-9) Resistant isolate(BC-8)
Between sources F=22.10 Between sources F=22.67
C.D. at p value 0.05=36.29 C.D. at p value 0.05=36.48
181
0
10
20
30
40
50
60
70
80
90
1 2 3 4
Days
Line
argr
owth
inm
m control
Magnesium sulphate
Potassium sulphate
Fig : 76 Effect of different sulphate sources on the linear growth (mm) of Botrytiscinerea resistant isolate to carbendazim Czapek Dox Agar Medium.
0
10
20
30
40
50
60
70
80
90
1 2 3 4
Days
Linea
rgro
wth
inm
m
control
Magnesiumsulphate
Potassiumhydrogensulphate
Fig : 77 Effect of different sulphate sources on the linear growth (mm) ofBotrytis cinerea resistant isolate to carbendazim Czapek Dox Agar Medium.
182
Table 57: Effect of different salts sources on the linear growth (mm) of Botrytiscinerea, sensitive and resistant isolates to carbendazim on CDA medium.
Sensitive isolate Resistant isolateDays DaysSource of
salts 0.05%1 2 3 4 1 2 3 4
magnesiumChloride
19.00 35.33 52.33 72.66 21.00 36.66 54.66 73.66
Potassiumchloride
20.33 39.66 58.66 77.66 22.33 41.66 61.33 79.33
Sodiumchloride
19.33 38.33 55.66 75.33 20.00 38.33 57.33 77.33
CalciumChloride
18.66 35.33 51.66 69.33 20.33 36.33 52.00 71.33
Control 18.66 34.66 49.66 68.66 19.66 35.33 51.33 69.66
Followed Tukey
Sensitive isolate (BC-9) Resistant isolate(BC-8)
Between sources F=7.21 Between sources F=11.64
C.D. at p value 0.05=35.73 C.D. at p value 0.05=35.65
183
0
10
20
30
40
50
60
70
80
90
1 2 3 4
Days
Line
argr
owth
inm
m
Control
Mgcl2
Kcl2
Cacl2
Nacl2
Fig : 78 Effect of different salt sources on the linear growth (mm) of Botrytiscinerea sensitive isolate to carbendazim Czapek Dox Agar Medium.
0
10
20
30
40
50
60
70
80
90
1 2 3 4
Days
linea
rgro
wth
inm
m
control
Mgcl2
Kcl2
Cacl2
Nacl2
Fig : 79 Effect of different salt sources on the linear growth (mm) of Botrytiscinerea resistant isolate to carbendazim Czapek Dox Agar Medium.
184
Table 58: Effect of different vitamin sources on the linear growth (mm) ofBotrytis cinerea, sensitive and resistant isolates to carbendazim on CDA medium.
Sensitive isolate Resistant isolateDays DaysVitamins
0.01% 1 2 3 4 1 2 3 4Ascorbicacid
20.33 34.66 55.33 73.66 21.33 35.66 56.33 76.66
Thiamine 21.33 35.33 57.33 74.33 22.66 36.66 59.33 78.66
Riboflavin 19.33 32.33 53.66 71.66 20.66 34.33 54.66 74.33
Pyridoxine 22.33 36.33 59.33 76.66 24.33 37.66 61.33 80.00
Control 18.33 33.33 49.66 68.66 20.00 35.66 52.33 71.33
Followed Tukey
Sensitive isolate (BC-9) Resistant isolate(BC-8)
Between sources F=15.21 Between sources F=12.26
C.D. at p value 0.05=36.78 C.D. at p value 0.05=37.91
185
0
10
20
30
40
50
60
70
80
90
1 2 3 4
Days
Line
argr
owth
inm
m
Control
ascorbic acid
Thiamine
Riboflavin
Pyridoxime
Fig : 80 Effect of different Vitamin sources on the linear growth (mm) of Botrytiscinerea sensitive isolate to carbendazim Czapek Dox Agar Medium.
0
10
20
30
40
50
60
70
80
90
1 2 3 4
Days
Line
argr
owth
inm
m
Control
Ascorbic acid
Thiamine
Riboflavin
Pyridoxime
Fig : 81 Effect of different vitamin sources on the linear growth (mm) ofBotrytis cinerea resistant isolate to carbendazim Czapek Dox Agar Medium.
186
Table 59; Effect of different micronutrient sources on the linear growth (mm) ofBotrytis cinerea, sensitive and resistant isolates to carbendazim on CDA medium.
Sensitive isolate Resistant isolate
Days DaysSource of
Micronutrients.0.01% 1 2 3 4 1 2 3 4
Manganese 20.33 39.33 58.66 77.33 21.66 40.66 61.33 79.66
Copper 14.33 30.33 49.33 65.66 16.33 32.66 51.66 68.66
Cobalt 15.33 31.33 52.66 67.33 18.66 33.33 54.33 69.33
Zinc 13.33 28.33 47.66 63.33 15.66 30.66 49.33 65.33
Molybdenum 12.00 25.33 38.66 55.66 14.00 28.66 42.33 57.66
Control 19.33 34.33 56.66 75.66 20.00 36.66 59.33 76.33
Followed Tukey
Sensitive isolate (BC-9) Resistant isolate(BC-8)
Between sources F=22.10 Between sources F=22.67
C.D. at p value 0.05= C.D. at p value 0.05=
187
0
10
20
30
40
50
60
70
80
1 2 3 4
Days
Linea
rgro
wth
inm
m
Control
manganese
Copper
Cobalt
Zinc
Molybdenum
Fig : 82 Effect of different micronutrients sources on the linear growth (mm)of Botrytis cinerea sensitive isolate to carbendazim Czapek Dox Agar Medium.
0
10
20
30
40
50
60
70
80
90
1 2 3 4
Days
Line
argr
owth
inm
m
Control
Manganese
Copper
Cobalt
Zinc
Molybdenum
Fig : 83 Effect of different micronutrients sources on the linear growth (mm)of Botrytis cinerea resistant isolate to carbendazim Czapek Dox Agar Medium.
188
Table 60;Effect of different amino acids sources on the linear growth (mm) of
Botrytis cinerea, sensitive and resistant isolates to carbendazim on CDA medium.
.Sensitive Resistant
Days DaysSourceAmino acid(0.2%)
1 2 3 4 1 2 3 4
Glycine 17.33 33.39 56.33 71.33 18.66 41.66 59.33 72.33
Phenylalanine 20.33 39.33 57.33 74.66 21.33 40.33 58.66 76.66
Tyrosine 14.66 28.33 45.66 64.33 15.33 30.66 47.66 66.33
Proline 21.33 40.66 56.33 75.33 22.33 42.33 58.66 77.33
Tryptophan 14.00 35.33 45.66 55.33 16.33 36.66 47.66 57.33
Cystine 13.66 28.33 42.66 53.66 15.00 30.66 44.33 54.66
Serine 20.00 40.33 58.66 75.33 22.66 42.33 69.33 77.66
Alanine 22.33 41.33 57.66 76.66 22.66 42.66 59.33 78.33
Histidine 19.33 35.66 55.33 72.33 20.33 37.33 56.66 73.66
Control 16.66 31.66 52.33 69.33 17.66 34.33 55.66 71.00
Followed Tukey
Sensitive isolate (BC-9) Resistant isolate(BC-8)
Between sources F=20.83 Between sources F=19.71
C.D. at p value 0.05=34.98 C.D. at p value 0.05=34.42
189
0
10
20
30
40
50
60
70
80
90
1 2 3 4
Days
Line
argr
owth
inm
m
control
Glycine
Phenyl alanine
Tyrosine
Proline
Cystine
serine
Alanine
Histidine
Fig: 84 Effect of different amino acid sources on the linear growth (mm) of Botrytiscinerea sensitive isolate to carbendazim Czapek Dox Agar Medium.
0
10
20
30
40
50
60
70
80
90
1 2 3 4
Days
Line
argr
owth
inm
m
control
Glycine
Phenyl alanine
Tyrosine
Proline
Cystine
serine
Alanine
Histidine
Fig: 85 Effect of different amino acid sources on the linear growth (mm) of Botrytiscinerea resistant isolate to carbendazim Czapek Dox Agar Medium.
190
BIOCHEMICAL CHAACTERISICS OF SENSITIVE AND RESITANT
ISOLATE OF BOTRYTIS CINEREA.
This was studied by inoculating the Rosa floribunda plants with spore
suspension of resistant isolate and sensitive isolate of Botrytis cinerea. After 10 days
of inoculation, the infected plants were collected and dried at 40 C̊ in hot air oven
and powder was obtained in grinder. The oven dried leaf powder used for analysis of
total soluble sugar, reducing sugar,starch, polyphenols ,DNA,RNA, and total ash.
Results in (Table 61) indicated that when compared with healthy plant, total sugar,
reducing sugar, DNA, RNA content of host plant were reduced due to infection of
sensitive and resistant isolates of Botrytis cinerea. While starch ,total ash and phenol
content were increased in the host plant which is infected by resistant isolate of
Botrytis cinerea.
Table 61: BIOCHEMICAL CHARACTERISTICS OF LEAF SPOT OF ROSE
INFECTED WITH SENSITIVE AND RESISTANT ISOLATE OF BOTRYTIS
CINEREA.
Sr.no Estimation Healthy Sensitive Resistant
1 Total sugar(g/100g) 1.90 1.29 1.20
3 Reducing sugar(g/100g) 0.47 0.32 0.29
4 Starch(g/100g) 0.110 0.15 0.23
5 Polyphenols(g/100g) 1.56 2.75 2.90
6 DNA (mg/ml) 0.40 0.029 0.022
7 RNA(mg/ml) 1.32 1.19 1.10
8 Total ash (%) 7.00 8.00 11.00
191
Synergistic effects of agrochemicals on the carbendazim resistance in Botrytis
cinerea.
In vitro studies:-
Agrochemicals such as Fungicides, Insecticides, Herbicides, Antibiotics, Salts,
Fertilizers and Micronutrients were mixed with carbendazim in the CDA medium.
Carbendazim resistant BC-8 isolate was inoculated on to the plates and linear growth
was measured after four days. Increased growth over carbendazim alone ‘control’ was
considered as increase in the resistance or vice-versa.
Fungicides:-
Benomyl, Ridomil MZ, and Kocide-101 were used for this study. These
fungicides were mixed with carbendazim (1000 µg/ml) at various concentrations (25,
50, 75 and 100 µg /ml). Benomyl and Ridomil MZ, fungicides inhibited the growth of
the pathogen at 25 µg /ml. While Kocide reduced the growth of the pathogen with the
increase in its concentration in the plates, as compared to carbendazim alone
‘control’. (Table 62 and Fig.86)
Insecticides:-
Dunet, Krinet and Monosaan were mixed with carbendazim (1000 µg/ml). It
was observed that Dunet and Krinet insecticides inhibited the growth of resistant
isolate of Botrytis cinerea at 25µg/ml of insecticides. While Monosaan inhibited the
growth of the pathogen at 75 µg/ml. (Table 63 and Fig.87)
192
Herbicides:-
Sencor, Kleen-80, Krizin and Atrazin were mixed with carbendazim to study
the effect of herbicides on the carbendazim resistant isolate (BC-8). Kleen-80 and
Krizin inhibited the growth of the pathogen at 50µg/ml, while Atrazin inhibited the
growth of the pathogen at 75µg/ml. And Sencor reduced growth of pathogen with
increased its concentration in the medium. (Table 64 and Fig.88)
Antibiotics:-
Streptomycin, Oflaxacin-400, Griseofulvin and Cefixime were mixed with
carbendazim (1000 µg/ml) at 0.1 to o.4 µg/ml. All these antibiotics reduced growth of
the pathogen with increase in their concentration in the medium.
(Table 65 and Fig.89)
Salts:-
Altogether four salts were incorporated in the medium along with carbendazim
(1000 µg/ml) All these salts reduced the growth of pathogen with increase in their
concentration in plates as compared with carbendazim alone ‘control’. (Table 66and
Fig.90)
193
Fertilizers:-
Urea, Muriate of potash, Samarth (10:26:26) and DAP (18:46:0) were mixed
with carbendazim (1000 µg/ml) at 0.1 to 0.4 % concentration. From these Samarth
and DAP inhibited the growth of the pathogen at 0.1%, and muriate of potash
inhibited the growth of the pathogen at 0.2%. While Urea reduced the growth of
pathogen with increased in its concentration in the plates. (Table 67 and Fig.91)
Micronutrients:-
Fe, Cu, Mn, Mo, Zn and Co was mixed with carbendazim (1000 µg/ml) at
various concentrations ranging from 0.1 to 0.4µg/ml in the medium. It was observed
Zinc and cobalt inhibited the growth of the pathogen completely at 0.1µg/ml. While
remaining micronutrients inhibited the growth of the pathogen as their concentration
increased in the medium (Table 68 and Fig.92)
.
194
Table 62: Synergistic effect of fungicides on the development of carbendazim
resistance in Botrytis cinerea in vitro
Fungicides with Carbendazim
1000 μg/ml
Concentration
Latent
Period Days
Linear
Growth in
mm.
25 - 00.00
50 - 00.00
75 - 00.001 Benomyl (µg/ml)
100 - 00.00
25 - 00.00
50 - 00.00
75 - 00.002 Ridomil (µg/ml)
100 - 00.00
25 2 14.33
50 2 13.33
75 2 11.663 Kocide-10 (µg/ml)
100 2 10.33
4Control (Carbendazim
1000 μg/ml)2 12.00
Mean ± SE (Standard Error)6.85 ± 2.19
CD at P value 0.05 = 4.6
195
0
2
4
6
8
10
12
14
16
Control 25 50 75 100
conc.of fungicides in ppm
Line
argr
owrt
inm
m
Binomyl
Ridomil
Kocide
Fig .86 Synergistic effects of fungicides on the development of
Carbendazim resistance in Botrytis cinerea in vitro
196
Table 63: Synergistic effect of Insecticides on the development of carbendazim
resistance in Botrytis cinerea (In Vitro)
Insecticides withCarbendazim 1000 μg/ml Concentration
LatentPeriod Days
LinearGrowth inmm.
25 - 00.00
50 - 00.00
75 - 00.001
Dunet (µg/ml)
100 - 00.00
25 - 00.00
50 - 00.00
75 - 00.002 Krinet(µg/ml)
100 - 00.00
25 2 12.33
50 2 10.66
75 - 00.003 Monosaan (µg/ml)
100 - 00.00
4Control
Carbendazim alone1000 μg/ml2 12.00
Mean ± SE (Standard Error)2.69 ± 1.42CD at P value 0.05 = 2.01
197
0
2
4
6
8
10
12
14
Control 25 50 75 100
Concentration of insecticides in ppm
Radi
algr
owth
inm
m
Dunet
Krinet
Monasaan
Fig. 87 Synergistic effect of Insecticides on the development
of carbendazim resistance in Botrytis cinerea (In Vitro)
198
Table 64: Synergistic effect of Herbicides on the development of carbendazimresistance in Botrytis cinerea.(In Vitro)
Herbicides with Carbendazim
1000 μg/mlConcentration
Latent
Period Days
Linear
Growth in
mm.
25 2 13.66
50 2 12.33
75 2 10.331 Sencor (µg/ml)
100 2 09.33
252 12.66
50 - 00.00
75 - 00.00
2 Kleen-80(µg/ml)
100 - 00.00
252 10.00
50- 00.00
75 - 00.00
3 Krizin (µg/ml)
100 - 00.00
25 2 13.33
50 2 11.33
75 - 00.004 Atrazin (µg/ml)
100 - 00.00
5 Control
Carbendazim alone1000 μg/ml2 12.00
Mean ± SE (Standard Error)6.15 ± 1.47CD at P value 0. 05 = 3.12
199
0
2
4
6
8
10
12
14
Control 25 50 75 100
Concentration of herbicides in ppm
Line
argr
owth
inm
m
sencor
Kleen
Krizin
Atrazin
Fig. 88 Synergistic effect of herbicides on the development of
carbendazim resistance in Botrytis cinerea (In Vitro)
200
Table 65; Synergistic effect of Antibiotics on the development of carbendazim
resistance in Botrytis cinerea (In Vitro)
Antibiotics withCarbendazim 1000 μg/ml Concentration
LatentPeriod Days
LinearGrowth inmm.
0.1 2 12.33
0.2 2 11.33
0.3 2 10.331 Streptomycin (µg/ml)
0.4 2 09.00
0.1 2 12.66
0.2 2 11.33
0.3 2 10.002 Oflaxacin 400 (µg/ml)
0.4 2 09.00
0.1 2 13.00
0.2 2 12.00
0.3 2 11.663 Griseofulvin (µg/ml)
0.4 2 10.00
0.1 2 13.33
0.2 2 12.00
0.3 2 11.334 Cefixime (µg/ml)
0.4 2 10.00
5Control Carbendazim
alone1000 μg/ml2 12.00
Mean ± SE (Standard Error)11.52 ± 0.32CD at P value 0.05 = 0.68
201
0
2
4
6
8
10
12
14
Control 0.1 0.2 0.3 0.4
Concentration of antibiotics in ppm
Line
argr
owth
inm
m
Streptomycin
Griseofulvin
Cefixime
Oflaxacine
Fig. 89 Synergistic effect of Antibiotics on the development
of carbendazim resistance in Botrytis cinerea (In Vitro)
202
Table 66: Synergistic effect of Salts on the development of carbendazim
resistance in Botrytis cinerea.(In Vitro)
Salts withCarbendazim 1000 μg/ml Concentration Latent
Period Days
LinearGrowth inmm.
0.1 2 12.33
0.2 2 11.00
0.3 2 10.001 Sodium chloride (µg/ml)
0.4 2 09.00
0.1 2 12.00
0.2 2 11.33
0.3 2 10.002 Potassium chloride (µg/ml)
0.4 2 09.00
0.1 2 12.66
0.2 2 11.33
0.3 2 10.333
Magnesium chloride (µg/ml)
0.4 2 09.00
0.1 2 12.66
0.2 2 11.33
0.3 2 10.004 Calcium chloride (µg/ml)
0.4 2 09.00
5
Control
Carbendazim alone1000 μg/ml2 12.00
Mean ± SE (Standard Error)
10.76 ± 0.32
CD at P value 0.05 = 0.68
203
0
2
4
6
8
10
12
14
Control 0.1 0.2 0.3 0.4
Concentration of salts in ppm
Line
argr
owth
inm
m
Nacl2
Kcl2
Mgcl2
Cacl2
Fig. 90 Synergistic effect of salts on the development
of carbendazim resistance in Botrytis cinerea (In Vitro)
204
Table 67: Synergistic effect of Fertilizers on the development of carbendazimresistance in Botrytis cinerea (In Vitro)
Fertilizers withCarbendazim 1000 μg/ml
ConcentrationLatentPeriod Days
LinearGrowth inmm.
0.1 2 15.66
0.2 2 13.33
0.3 2 12.001 Urea (%))
0.4 2 10.00
0.1 2 13.00
0.2 - 00.00
0.3 - 00.002 Muriate of potash (%)
0.4 - 00.00
0.1- 00.00
0.2- 00.00
0.3 - 00.00
3 Samarth (10:26:26) (%)
0.4 - 00.00
0.1 - 00.00
0.2 - 00.00
0.3 - 00.004 DAP (%)
0.4 - 00.00
Control
Carbendazim alone1000 μg/ml2 12.00
Mean ± SE (Standard Error)
7.41 ± 1.58
CD at P value 0.05 = 3.3
205
0
2
4
6
8
10
12
14
16
18
Control 0.1 0.2 0.3 0.4
Concentration of fertilizers in ppm
Line
argr
owth
inm
m
Urea
Murate ofpotashSamarath
DAp
Fig. 91 Synergistic effect of Fertilizers on the development
of carbendazim resistance in Botrytis cinerea (In Vitro)
206
Table 68 : Synergistic effect of Micronutrients on the development ofcarbendazim resistance in Botrytis cinerea (In Vitro)
Micronutrients withCarbendazim 1000 μg/ml
Concentration LatentPeriod Days
LinearGrowth inmm.
0.1 2 13.33
0.2 2 12.33
0.3 2 11.331 Iron (µg/ml)
0.4 2 09.00
0.1 2 12.00
0.2 2 11.00
0.3 2 10.002 Copper (µg/ml)
0.4 - 00.00
0.1 2 13.00
0.2 2 12.00
0.3 2 11.003 Mangenese (µg/ml)
0.4 2 00.00
0.1 - 00.00
0.2 - 00.00
0.3 - 00.004 Zinc (µg/ml)
0.4 - 00.00
0.1 - 00.00
0.2 - 00.00
0.3 - 00.005 Cobalt (µg/ml)
0.4 - 00.00
6Control Carbendazim
alone1000 μg/ml2 12.00
Mean ± SE (Standard Error)
6.04 ± 1.30
CD at P value 0.05 = 2.72
207
0
2
4
6
8
10
12
14
Control 0.1 0.2 0.3 0.4
Concentration of micronutrients in ppm
Linea
rgro
wth
inm
m
Iron
Copper
Manganese
Zinc
Cobalt
Fig. 92 Synergistic effect of micronutrients on the development
of carbendazim resistance in Botrytis cinerea (In Vitro)
208
In vivo studies:
To study synergistic effect of agrochemicals on the development of resistance in
Botrytis cinerea against carbendazim, 10 ml mycelial suspension of one culture tube
of resistant isolate (BC-8) was inoculated on healthy rose plant treated with resistant
dose of carbendazim (800 µg/ml) and other agrochemicals at different concentrations,
24 hrs before. Each rose plant was covered with polythene bag. and Percentage of
infection on rose plant was determined.
Fungicides:-
Benomyl, Ridomil MZ, µg/ml Kocide-101 were mixed with carbendazim (800
µg/ ml) at various concentrations (25, 50, 75 and 100 µg/ml). Benomyl, Ridomil MZ
completely prevented the infection of pathogen to rose plant at 25µg/ml with
carbendazim, while Kocide prevented the infection of pathogen at 75µg/ml with
carbendazim, (Table 69 and Fig. 93)
Insecticides:-
Dunet, Krinet and Monosaan were mixed with carbendazim (800 µg/ ml) at
various concentrations (25, 50, 75 and 100 µg/ml).plants of Rosa floribunda were
treated with these mixtures and inoculated with resistant isolate. All above said
insecticides completely prevented the infection of pathogen to rose plant at 25µg/ml
with carbendazim, , (Table 70 and Fig. 94)
Herbicides:-
Sencor, Kleen-80 and Krizin and Atrazine were mixed with carbendazim (800
µg/ml) at various concentrations (25, 50, 75 and 100 µg/ml). Kleen-80 and Krizin
completely prevented the infection of pathogen to rose plant at 50µg/ml with
carbendazim, and Atrazine prevented the infection of pathogen to rose plant at
209
75µg/ml with carbendazim .While Sencor reduced the infection to the plant as there
was increase in its concentration . , (Table 71 and Fig. 95)
Antibiotics;-
Streptomycin, Oflaxacin-400, Griseofulvin and Cefixime were used for this
study. Among these antibiotics Streptomycin and Oflaxacin-400 prevented the
infection of pathogen to rose plant at 0.1µg/ml with carbendazim. While Griseofulvin
and Cefixime prevented the infection to rose plants by Botrytis cinerea at 0.4µg/ml
(Table 72 and Fig. 96)
Salts:-
Altogether four salts were mixed with carbendazim at (0.1to0.4µg/ml) .
Magnesium chloride and calcium chloride prevented the infection of pathogen to rose
plant at 0.1µg/ml with carbendazim .while potassium chloride and sodium chloride
prevented the infection of pathogen to rose plant at 0.3µg/ml and 0.4µg/ml
respectively with carbendazim, (Table 73 and Fig. 97)
Fertilizers:-
Urea, Muriate of potash, Samarth, and DAP were mixed with carbendazim (800
µg/ml) at (0.1 to 0.4 %) concentrations. DAP and muriate of potash were found very
effective and protected the rose plants from the infection of pathogen at 0.2 and 0.3%
respectively with carbendazim. While Urea and Samarth prevented the infection 0f
Botrytis cinerea to rose plant at 0.4% with carbendazim, (Table 74and Fig. 98).
Micronutrients:-
Fe, Cu, Mn, Mo, Zn and Co were used for this study. From these Cu, Mn, Mo,
Zn and Co were found to very effective in controlling the disease of rose plants at
0.1µg/ ml with carbendazim. While iron protected the rose plants from disease at 0.3
µg/ ml along with the carbendazim. (Table 75and Fig. 99)
210
Table 69: Synergistic effect of fungicides on the development of carbendazimresistance in Botrytis cinerea (In vivo)
Fungicides with Carbendazim
800 µg/ml
ConcentrationPercentageof infection
Grade
25 00.00 00.00
50 00.00 00.00
75 00.00 00.001 Benomyl (µg/ml)
100 00.00 00.00
25 00.00 00.00
50 00.00 00.00
75 00.00 00.002 Ridomil (µg/ml
100 00.00 00.00
25 15.00 01.00
50 10.00 01.00
75 05.00 00.003 Kocide-10 (µg/ml)
100 00.00 00.00
4Control 800 µg/ml
Carbendazim alone25.00 01.00
Mean ± SE (Standard Error)
4.23 ± 2.18
CD at P value 0.05 = 4.7
211
0
5
10
15
20
25
30
Control 25 50 75 100
Concentration of fungicides in ppm
perc
enta
geof
infe
ctio
n
Binomyl
Ridomil
Kocide
Fig.93 Synergistic effect of fungicides on the development ofcarbendazim resistance in Botrytis cinerea (In vivo)
212
Table 70: Synergistic effect of Insecticides on the development of carbendazim
resistance in Botrytis cinerea ( In vivo)
Insecticides with
Carbendazim 800 µg/mlConcentration
Percentage
of infection
Grade
25 00.00 00.00
50 00.00 00.00
75 00.00 00.001 Dunet (µg/ml)
100 00.00 00.00
25 00.00 00.00
50 00.00 00.00
75 00.00 00.002 Krinet(µg/ml)
100 00.00 00.00
25 00.00 00.00
50 00.00 00.00
75 00.00 00.003
Monosaan (µg/ml)
100 00.00 00.00
4
Control 800 µg/ml
Carbendazim alone25.00 01.00
Mean ± SE (Standard Error)
1.92 ± 1.92CD at P value 0.05 = 4.19
213
0
5
10
15
20
25
30
Control 25 50 75 100
Concentration of insecticides in ppm
Perc
enta
geof
infe
ctio
n
Dunet
Krinet
Monasaan
Fig.94 Synergistic effect of Insecticides on the development
of carbendazim resistance in Botrytis cinerea ( In vivo)
214
Table 71: Synergistic effect of Herbicides on the development of carbendazim
resistance in Botrytis cinerea (In vivo)
Herbicides with
Carbendazim 800 µg/mlConcentration
Percentage
of infectionGrade
25 20.00 01.00
50 15.00 01.00
75 10.00 01.001 Sencor (µg/ml)
100 05.00 01.00
25 15.00 01.00
50 00.00 00.00
75 00.00 00.002 Kleen-80(µg/ml
100 00.00 00.00
25 10.00 01.00
50 00.00 00.00
75 00.00 00.003 Krizin (µg/ml)
100 00.00 00.00
25 15.00 01.00
50 10.00 01.00
75 00.00 00.004 Atrazin (µg/ml)
100 00.00 00.00
5
Control 800 µg/ml
Carbendazim alone25.00 01.00
Mean ± SE (Standard Error)
7.35 ± 2.01
CD at P value 0.05 = 4.27
215
0
5
10
15
20
25
30
Control 25 50 75 100
Concentration of herbicides in ppm
Perc
enta
geof
infe
ctio
n
sencor
Kleen
Krizin
Atrazin
Fig. 95 Synergistic effect of Herbicides on the development
of carbendazim resistance in Botrytis cinerea (In vivo)
216
Table 72; Synergistic effect of Antibiotics on the development of carbendazimresistance in Botrytis cinerea (In vivo)
Antibiotics with
Carbendazim 800 µg/mlConcentration
Percentage
of infectionGrade
0.1 15.00 01.00
0.2 10.00 01.00
0.3 00.00 00.001 Streptomycin (µg/ml)
0.4 00.00 00.00
0.1 20.00 01.00
0.2 15.00 01.00
0.3 00.00 00.002
Oflaxacin 400 (µg/ml)
0.4 00.00 00.00
0.1 15.00 01.00
0.2 10.00 01.00
0.3 05.00 01.003 Griseofulvin (µg/ml)
0.4 00.00 00.00
0.1 15.00 01.00
0.2 10.00 01.00
0.3 05.00 01.004 Cefixime (µg/ml)
0.4 00.00 00.00
5Control 800 µg/ml
Carbendazim alone25.00 01.00
Mean ± SE (Standard Error)
8.52 ± 1.95CD at P value 0.05 = 4.14
217
0
5
10
15
20
25
30
Control
0.1 0.2 0.3 0.4
Concentration of antibiotics in ppm
Perc
enta
geof
infe
ctio
n
Streptomycin
Griseofulvin
Cefixime
Oflaxacime
Fig.96 Synergistic effect of Antibiotics on the development of carbendazimresistance in Botrytis cinerea (In vivo)
218
Table 73; Synergistic effect of Salts on the development of carbendazim
resistance in Botrytis cinerea (In vivo)
Salts with Carbendazim800 µg/ml Concentration
Percentageofinfection
Grade
0.1 15.00 01.00
0.2 10.00 01.00
0.3 05.00 01.001 Sodium chloride (µg/ml)
0.4 00.00 00.00
0.1 15.00 01.00
0.2 10.00 01.00
0.3 00.00 00.002 Potassium chloride (µg/ml)
0.4 00.00 00.00
0.1 00.00 00.00
0.2 00.00 00.00
0.3 00.00 00.003 Magnesium chloride (µg/ml)
0.4 00.00 00.00
0.1 00.00 00.00
0.2 00.00 00.00
0.3 00.00 00.004 Calcium chloride (µg/ml)
0.4 00.00 00.00
5 Control 800 µg/mlCarbendazim alone
25.00 01.00
Mean ± SE (Standard Error)
5.88 ± 1.91
CD at P value 0.05 = 4.05
219
0
5
10
15
20
25
30
Control 0.1 0.2 0.3 0.4
Concentration of salts in ppm
Perc
enta
geof
infe
ctio
n
Nacl2
Kcl2
Mgcl2
Cacl2
Fig. 97 Synergistic effect of Salts on the development
of carbendazim resistance in Botrytis cinerea ( In vivo)
220
Table 74: Synergistic effect of Fertilizers on the development of carbendazim
resistance in Botrytis cinerea (In vivo)
Fertilizers withCarbendazim 800 µg/ml
ConcentrationPercent ofinfection
Grade.
0.1 20.00 01.00
0.2 10.00 01.00
0.3 05.00 01.001 Urea (%))
0.4 00.00 00.00
0.1 10.00 01.00
0.2 05.00 01.00
0.3 00.00 00.002
Murate of potash (%)
0.4 00.00 00.00
0.1 15.00 01.00
0.2 10.00 01.00
0.3 05.00 01.003 Samarth (10:26:26) (%)
0.4 00.00 00.00
0.1 10.00 01.00
0.2 00.00 00.00
0.3 00.00 00.004 DAP %
0.4 00.00 00.00
5Control 800 µg/ml
Carbendazim alone25.00 01.00
Mean ± SE (Standard Error)
6.76 ± 1.86
CD at P value 0.05 = 2.08
221
0
5
10
15
20
25
30
Control 0.1 0.2 0.3 0.4
Concentration of fertilizers in percentage
Perc
enta
geof
infe
ctio
n
Urea
Murate of potash
Samarath
DAP
Fig.Synergistic effect of Fertilizers on the development of carbendazim resistance in
Botrytis cinerea (In vivo)
222
Table 75: Synergistic effect of Micronutrients on the development of
carbendazim resistance in Botrytis cinerea . (In Vivo)
Micronutrients withCarbendazim 800 µg/ml
Concentration Percent ofinfection
Grade
0.1 10.00 01.00
0.2 05.00. 01.00
0.3 00.00 00.001 Iron (µg/ml)
0.4 00.00 00.00
0.1 00.00 00.00
0.2 00.00 00.00
0.3 00.00 00.002 Copper (µg/ml)
0.4 00.00 00.00
0.1 00.00 00.00
0.2 00.00 00.00
0.3 00.00 00.003 Mangenese (µg/ml)
0.4 00.00 00.00
0.1 00.00 00.00
0.2 00.00 00.00
0.3 00.00 00.004 Zinc (µg/ml)
0.4 00.00 00.00
0.1 00.00 00.00
0.2 00.00 00.00
0.3 00.00 00.005 Cobalt (µg/ml)
0.4 00.00 00.00
6 Control 800 µg/mlCarbendazim alone
25.00 01.00
Mean ± SE (Standard Error)
1.90 ± 1.26
CD at P value 0.05 = 2.64
223
0
5
10
15
20
25
30
Control 0.1 0.2 0.3 0.4
Concentration of micronutrients in ppm
Perc
enta
geof
infe
ctio
n
Iron
Copper
Manganese
Zinc
Cobalt
Fig. 99 Synergistic effect of Micronutrients on the development
of carbendazim resistance in Botrytis cinerea . (In Vivo)
224
Percentage control efficacy (PCE) of agrochemicals:-
The percentage control efficacy (PCE) of each agrochemical was calculated by using
the formula given by (Kamble 1991).
PCE=100(1-x/y) where,
X = diameter of colony on agar medium amended with fungicide.
Y= diameter of colony on agar medium without fungicide.
In Vitro study
100 percent control of leaf blight of rose was due to the treatment of
carbendazimwith other agrochemicals.Moreover, this control was seen to be at
thelower concentration 100% PCE was found in the mixture of carbendazim with
Benomyl, Ridomil, Dunet, krinet at 25 µg/ ml.In kleen and Krizin 100% PCE was at
50 µg/ ml and in Monosaan and atrazin 100% PCE found at 75µg/ ml. with
carbendazim. Fertilizers like Muriate of potash, Samarth and DAP shown 100% PCE
at 0.1%. In copper. 100% PCE was found at 100µg/ ml with carbendazim.
In vivo study
Benomyl, Ridomil, gave 100 % PCE at 25 µg/ml with carbendazim. While Kocide
shown 100 % PCE at 100 µg/ml with carbendazim. Dunet, Krinet, Monosan, gave
100 % PCE at 25 µg/ml with carbendazim. Herbicides like Atrazin gave 100% PCE at
25 µg/ml with carbendazim. Kleen and Krizin gave 100% PCE at 50 µg/ml with
carbendazim. Antibiotics like Streptomycin and Oflaxacine-400 gave 100% PCE at
0.3 µg/ml with carbendazim. While Griseofulvin and Cefixime gave 100% PCE at 0.4
µg/ml with carbendazim .Salts like calcium chloride, magnesium chloride gave 100 %
PCE at 0.1µg/ml with carbendazim while potassium chloride and Sodium chloride
gave 100 % PCE at 0.3µg/ml and 0.4 µg/ml respectively with carbendazim. Fertilizers
225
like DAP gave 100% PCE at 0.2 % and Muriate of potash gave 100% PCE at 0.3
While Urea and Samarth gave 100 % PCE at 0.4% with carbendazim .Micronutrients
like Copper, Manganese, Zinc, and Cobalt gave 100 % PCE at 0.1µg/ml with
carbendazim, While Iron gave 100 % PCE at 0.3µg/ml with carbendazim.
(Tables 76 to 82)
Table 76: Percentage control efficacy (PCE) of fungicides in combination withthe carbendazim controlling the resistant isolate of Botrytis cinerea. In vitro andin vivo.
Fungicides withCarbendazim Concentration
PCEIn vitro
PCEIn Vivo
25 100 100.00
50 100 100.0075 100 100.00
1
Benomyl (µg/ml)
100 100 100.00
25 100 100.00
50 100 100.0075 100 100.00
2
Ridomil (µg/ml)
100 100 100.00
25 82.08 85..00
50 83.33 90.00
75 86.25 95.00
3
Kocide-10 (µg/ml)
100 87.08 100.00
226
Table 77: Percentage control efficacy (PCE) of insecticides in combination withthe carbendazim controlling the resistant isolate of Botrytis cinerea In vitro andin vivo.
Insecticides withCarbendazim
ConcentrationPCEIn vitro
PCEIn Vivo
25 100.00 100.00
50 100.00 100.0075 100.00 100.00
1Dunet (µg/ml)
100 100.00 100.00
25 100.00 100.00
50 100.00 100.0075 100.00 100.00
2Krinet(µg/ml)
100 100.00 100.00
25 84.88 100.00
50 86.87 100.00
75 100.00 100.00
3 Monosaan (µg/ml)
100 100.00 100.00
227
Table 78: Percentage control efficacy (PCE) of herbicides in combination withthe carbendazim controlling the resistant isolate of Botrytis cinerea In vitro andin vivo.
Herbicides with Carbendazim Concentration PCEIn vitro
PCEIn Vivo
25 82.92 80.00
50 84.58 85.0075 87.00 90.00
1 Sencor (µg/ml)
100 88.33 95.00
25 84.17 85.00
50 100.00 100.0075 100.00 100.00
2 Kleen-80(µg/ml)
100 100.00 100.00
25 87.00 90.00
50 100.00 100.00
75 100.00 100.00
3 Krizin (µg/ml)
100 100.00 100.0025 83.33 100.0050 85.33 100.0075 100.00 100.00
4 Atrazin
100 100.00 100.00
228
Table 79: Percentage control efficacy (PCE) of antibiotics in combination withthe carbendazim controlling the resistant isolate of Botrytis cinerea In vitro andin vivo.
Antibiotics withCarbendazim
ConcentrationPCEIn vitro
PCEIn Vivo
0.1 84.58 80.00
0.2 85.33 85.000.3 87.00 90.00
1 Streptomycin(µg/ml)
0.4 88.75 95.00
0.1 84.17 85.00
0.2 85.33 100.000.3 87.50 100.00
2 Oflaxacin 400(µg/ml)
0.4 88.75 100.000.1 83.75 90.00
0.2 85.00 100.00
0.3 86.25 100.00
3 Griseofulvin (µg/ml)
0.4 87.00 100.000.1 83.33 100.000.2 85.00 100.000.3 85.33 100.00
4 Cefixime (µg/ml)
0.4 87.50 100.00
229
Table 80: Percentage control efficacy (PCE) of salts in combination with thecarbendazim controlling the resistant isolate of Botrytis cinerea. In vitro and invivo.
Salts withCarbendazim
ConcentrationPCEIn vitro
PCEIn Vivo
0.1 84.58 85.00
0.2 86.25 90.000.3 87.50 95.00
1 Sodium chloride(µg/ml)
0.4 88.00 100.000.1 85.00 85.00
0.2 85.33 90.000.3 87.50 100.00
2 Potassium chloride(µg/ml)
0.4 88.50 100.000.1 84.17 100.00
0.2 85.33 100.00
0.3 87.08 100.00
3 Magnesium chloride(µg/ml)
0.4 88.75 100.000.1 84.17 100.000.2 85.33 100.000.3 87.50 100.00
4 Calcium chloride(µg/ml) 0.4 88.75 100.00
230
Table 81: Percentage control efficacy (PCE) of fertilizers in combination with thecarbendazim controlling the resistant isolate of Botrytis cinerea In vitro and invivo..
Fertilizers withCarbendazim
ConcentrationPCEIn vitro
PCEIn Vivo
0.1 80.42 80.00
0.2 83.33 90.000.3 85.00 95.00
1 Urea (%))
0.4 87.00 100.000.1 100 90.00
0.2 100 95.000.3 100 100.00
2 Muriate of potash(%)
0.4 100 100.000.1 100 85.00
0.2 100 90.00
0.3 100 95.00
3Samarth (10:26:26)(%)
0.4 100 100.000.1 100 90.00
0.2 100 100.00
0.3 100 100.004 DAP (18:46:0) %
0.4 100 100.00
231
Table 82: Percentage control efficacy (PCE) of micronutrients in combinationwith the carbendazim controlling the resistant isolate of Botrytis cinerea. In vitroand in vivo.
Micronutrients withCarbendazim
ConcentrationPCEIn vitro
PCEIn Vivo
0.1 83.00 90.00
0.2 84.00 95.000.3 85.00 100.00
1 Iron (µg/ml)
0.4 86.00 100.000.1 85.00 100.00
0.2 86.00 100.000.3 87.00 100.00
2 Copper (µg/ml)
0.4 100.00 100.000.1 84.00 100.00
0.2 85.00 100.00
0.3 86.00 100.00
3 Mangenese (µg/ml)
0.4 87.00 100.000.1 86.00 100.000.2 87.00 100.000.3 87.00 100.00
4 Zinc (µg/ml)
0.4 88.00 100.000.1 8400 100.000.2 85.00 100.000.3 87.00 100.005 Cobalt (µg/ml)
0.4 88.00 100.00
232
Survival ability of carbendazim resistant Botrytis cinerea in mixed population
The survival ability of resistant BC-8 isolates in the mixed population was
studied by inoculating Rosa floribunda plant leaves with mixture of carbendazim
sensitive BC-9 and resistant BC-8 isolates of Botrytis cinerea. The mycelial
suspensions from the sensitive and resistant isolates of Botrytis cinerea were mixed in
the following proportions.
Sensitive : Resistant
90 : 10
75 : 25
50 : 50
Rosa floribunda plant were subsequently inoculated according to the method
described earlier. Ten days after inoculations infected leaves of Rosa floribunda were
used for preparations of mycelial suspension. The suspensions of each variant was
used to inoculate second batch of rose plant. For next passage, prior to inoculation
5ml of sample was taken from each mycelial suspension and adjusted to the described
concentration of 100 mycelial fragments per ml approximately. One ml of each
suspension was then pipetted and poured onto the surface of 15 ml water agar plates.
These plates were incubated for 4 days till the colonies of Botrytis cinerea become
visible. At least 100 colonies from each variant were transferred to the plates of
Czapek Dox Agar medium containing carbendazim at 500 μg/ml concentration lethal
to sensitive isolate of Botrytis cinerea. The plates were again incubated for 4 days
.The surviving colonies were counted and their composition of each mixture was
examined in this way for four successive passages on Rose plant leaves treated and
233
untreated with carbendazim. In these variants without carbendazim treatment it was
seen that resistant population in the mixture reduces from passage to passage when
the resistant population is increased in the mixture still there is reduction in the
population. But in case of carbendazim treated passage resistant population increased
from passage to passage. This increased is also seen when its proportion is increased
in the mixture with the increased of passage number. Thus carbendazim treatment to
rose plant imposes a selection pressure on the population that only resistant
propagules are able to survive at fourth passage.
It is clear that in population from untreated plant, the reproduction ratio was always
smaller than on the carbendazim treated rose plant. This shows that resistant strain
may slowly disappear from the mixed population in due course if the application of
fungicide is discontinued on the other hand if the rose plant were treated with
carbendazim reproduction ration of the resistant strain was greater than that of the
sensitive strain. In this situation resistant strain finally dominated the sensitive
population in the presence of carbendazim. (Table 83)
Table 83: Survival of Carbendazim resistant Botrytis cinerea isolates in mixed
population during four successive passage on Rosa floribunda. Leaves were
treated with 400 µg/ml Carbendazim or remain untraded
Original
Ratio
Untreated Passage Treated with carbendazim
passage
1 2 3 4 1 2 3 4
S: R S: R S: R S: R S: R S: R S: R S: R S: R
90:10 91:09 94:06 95:05 96:04 80:20 55:45 36:64 16:84
75:25 65:35 69:31 70:30 76:24 70:30 25:75 23:77 12:88
50:50 52:48 56:44 56:44 58:42 25:75 10:90 05:95 03:97
234
Survival of Carbendazim resistant Botrytis cinerea isolates in mixed population
during four successive passage on Rosa floribunda. Leaves were treated with 400
µg/ml Carbendazim or remain untraded
235
Use of Bio pesticides in the management of leaf Blight Rose
Fresh leaves of Melia azedarach L., Clerodendrum inerme (L.) Gaertn, Hyptis
suaveolen (L.)Poit, Swietenia macrophylla King and, Tagetes erecta L, were
collected, washed, dried and pulverised to obtain dry powders. 100 gm of powder was
taken. Extract of each plant was prepared with 95% alcohol by (1:5 w/v) and
condensed to serve as stock extract. The toxicity of stock extract was determined
against sensitive and resistant is isolate of Botrytis cinerea to carbendazim by food
poisoning technique (Mishra and Tiwari, 1992) at four different concentrations. Plates
containing CDA supplemented with different plant extracts at four concentrations
with three replicates were inoculated with fresh 6 days old culture of Botrytis cinerea
8 mm agar disc of fungal culture was prepared with the help of sterile cork borer and
kept upside down on agar plates, these plates were incubated at 28 ± 2ºC. Plates
without plant extract served as control. Linear growth of Botrytis cinerea was
measured at different intervals.
The above procedure was repeated for the water extracts of the same plants
instead of alcoholic extracts. The proportion of water and alcohol was 95:5
respectively
The 25% alcoholic leaf extract of all above plants were very effective and
completely inhibited growth of sensitive and resistant isolates of Botrytis cinerea
.Water extract of above plants showed little inhibition of both the sensitive and
resistant isolate of Botrytis cinerea .But water leaf extracts of Hyptis and Tagetes
completelely inhibit the growth of the pathogen at 50%.
236
Table 84 ; Effect of Melia azedarach L. leaf extracts (alcoholic and aqueous) on
linear growth (mm) of sensitive isolate Botrytis cinerea causing leaf blight of
rose
DaysConcentration
in %
Leaf extract
1 2 3 4
Alcoholic 00.00 00.00 00.00 00.0025
Aqueous 16.33 33.66 41.66 53.66
Alcoholic 00.00 00.00 00.00 00.0050
Aqueous 14.33 32.33 39.66 50.66
Alcoholic 00.00 00.00 00.00 00.0075
Aqueous 13.33 30.33 35.33 47.66
Alcoholic 00.00 00.00 00.00 00.00100
Aqueous 12.33 28.66 34.33 45.66
Control. 20.33 41.33 61.66 80.00
Followed TukeySensitive isolate (BC-8)Between concentration F= 10.15Between Days F=16.05C.D. at p value 0.05=67.68
237
Fig. 101 Effect of Melia azedarach L. leaf extracts (alcoholic and aqueous) on
linear growth (mm) of sensitive isolate Botrytis cinerea causing leaf blight of
rose
238
Table 85 : Effect of Melia azedarach L. leaf extracts (alcoholic and aqueous) on
linear growth (mm) of resistant isolate Botrytis cinerea causing leaf blight of
rose
DaysConcentration
In percentage.Leaf extract
1 2 3 4
Alcoholic 00.00 00.00 00.00 00.0025
Aqueous 17.66 35.33 43.66 56.33
Alcoholic 00.00 00.00 00.00 00.0050
Aqueous 15.66 33.33 41.66 53.33
Alcoholic 00.00 00.00 00.00 00.0075
Aqueous 14.33 32.66 40.33 51.33
Alcoholic 00.00 00.00 00.00 00.00100
Aqueous 13.00 30.33 37.66 49.66
Control 21.66 42.33 63.66 80.00
Followed TukeySensitive Resistant isolate (BC-9)Between concentration F= 10.74Between Days F=17.94C.D. at p value 0.05=72.33
239
Fig. 102 Effect of Melia azedarach L. leaf extracs (alcoholic and aqueous) t on
linear growth (mm) of resistant isolate Botrytis cinerea causing leaf blight of
rose
240
Table 86: Effect of Clerodendrum inerme L. leaf extracts (alcoholic and aqueous)
on linear growth (mm) of sensitive isolate botrytis cinerea causing leaf blight of
rose
Daysconcentration
In percentageLeaf extract
1 2 3 4
Alcoholic 00.00 00.00 00.00 00.0025
Aqueous 17.33 35.66 45.33 56.33
Alcoholic 00.00 00.00 00.00 00.00
Aqueous 16.33 33.66 43.33 53.6650
Alcoholic 00.00 00.00 00.00 00.00
Aqueous 15.66 31.66 41.33 50.6675
Alcoholic 00.00 00.00 00.00 00.00
100 Aqueous 14.33 28.66 39.66 48.33
Control. 20.33 41.33 61.66 80.00
Followed TukeySensitive isolate (BC-8)Between concentration F= 8.59Between Days F=65.54C.D. at p value 0.05= 72.15
241
Fig. 103 Effect of Clerodendrum inerme L. leaf extracts (alcoholic and aqueous)
on linear growth (mm) of sensitive isolate botrytis cinerea causing leaf blight of
rose
242
Table 87: Effect of Clerodendrum inerme L. leaf extracts (alcoholic and aqueous)
on linear growth (mm) of resistant isolate of botrytis cinerea causing leaf blight
of rose
Daysconcentration In
percentage
Leaf extract
1 2 3 4
Alcoholic 00.00 00.00 00.00 00.0025
Aqueous 18.33 36.33 47.66 59.33
Alcoholic 00.00 00.00 00.00 00.0050
Aqueous 17.33 34.66 45.33 56.66s
Alcoholic 00.00 00.00 00.00 00.0075
Aqueous 16.66 33.33 42.33 54.66
Alcoholic 00.00 00.00 00.00 00.00100
Aqueous 15.33 31.33 41.66 51.66
Control. 21.66 42.33 63.66 80.00
Followed TukeyResistant isolate(BC-9)Between concentration F= 10.13Between Days F=93.91C.D. at p value 0.05= 76.35
243
Fig. 104Effect of Clerodendrum inerme L. leaf extracts (alcoholic and aqueous)
on linear growth (mm) of resistant isolate of botrytis cinerea causing leaf blight
of rose
244
Table : 88 Effect of Hyptis suaveolens L.Poit leaf extracts (alcoholic and
aqueous) on linear growth (mm) of sensitive isolate Botrytis cinerea causing leaf
blight of rose.
Daysconcentration
In percentageLeaf extract
1 2 3 4
Alcoholic 00.00 00.00 00.00 00.0025
Aqueous 14.66 21.33 30.66 42.66
Alcoholic 00.00 00.00 00.00 00.0050
Aqueous 00.00 00.00 00.00 00.00Alcoholic 00.00 00.00 00.00 00.00
75Aqueous 00.00 00.00 00.00 00.00
Alcoholic 00.00 00.00 00.00 00.00
Aqueous 00.00 00.00 00.00 00.00100
20.33 41.33 61.66 80.00
Followed TukeySensitive isolate (BC-8)Between concentration F= 16.07Between Days F=2.17C.D. at p value 0.05= 14.90
245
Fig. 105 Effect of Hyptis suaveolens L.Poit leaf extracts (alcoholic and aqueous)
on linear growth (mm) of sensitive isolate Botrytis cinerea causing leaf blight of
rose.
246
Table 89: Effect of Hyptis suaveolens L.Poit leaf extracts (alcoholic and aqueous)
on linear growth (mm) of resistant isolate Botrytis cinerea causing leaf spot of
rose.
Daysconcentration
In percentageLeaf extract
1 2 3 4
Alcoholic 00.00 00.00 00.00 00.0025
Aqueous 15.66 23.33 32.66 44.33
Alcoholic 00.00 00.00 00.00 00.0050
Aqueous 00.00 00.00 00.00 00.00
Alcoholic 00.00 00.00 00.00 00.0075
Aqueous 00.00 00.00 00.00 00.00
Alcoholic 00.00 00.00 00.00 00.00100
Aqueous 00.00 00.00 00.00 00.00
Control 21.66 42.33 63.66 80.00
Followed TukeyResistant isolate (BC-9)Between concentration F= 17.33Between Days F=2.20C.D. at p value 0.05=15.50
247
Fig. 106 Effect of Hyptis suaveolens L.Poit leaf extracts (alcoholic and aqueous)
on linear growth (mm) of resistant isolate Botrytis cinerea causing leaf spot of
rose.
248
Table 90: Effect of Swietenia macrophylla. King. Leaf extracts (alcoholic and
aqueous) on linear growth (mm) of sensitive isolate Botrytis cinerea causing
leaf blight of rose
Daysconcentration
In percentageLeaf extract
1 2 3 4
Alcoholic 00.00 00.00 00.00 00.0025
Aqueous 15.33 28.66 37.00 42.00
Alcoholic 00.00 00.00 00.00 00.0050
Aqueous 14.00 22.33 30.00 38.00
Alcoholic 00.00 00.00 00.00 00.0075
Aqueous 12.33 20.66 25.33 34.00
Alcoholic 00.00 00.00 00.00 00.00100
Aqueous 10.00 17.00 20.00 30.00
Control 20.33 41.33 61.66 80.00
Followed TukeySensitive isolate (BC-8)Between concentration F= 11.17Between Days F=15.22C.D. at p value 0.05= 48.12
249
Fig.107 Effect of Swietenia macrophylla. King. Leaf extracts(alcoholic and
aqueous) on linear growth (mm) of sensitive isolate Botrytis cinerea causing
leaf blight of rose
250
Table 91: Effect of Swietenia macrophylla. King. Leaf extracts (alcoholic and
aqueous) on linear growth (mm) of resistant isolate Botrytis cinerea causing leaf
blight of rose
Daysconcentration
In percentageLeaf extract
1 2 3 4
Alcoholic 00.00 00.00 00.00 00.0025
Aqueous 17.66 30.00 39.00 45.00
Alcoholic 00.00 00.00 00.00 00.0050
Aqueous 15.00 24.00 35.00 42.00
Alcoholic 00.00 00.00 00.00 00.0075
Aqueous 12.00 22.00 30.00 38.00
Alcoholic 00.00 00.00 00.00 00.00100
Aqueous 15.33 18.33 25.33 32.00
Control 21.66 42.33 63.66 80
Followed TukeyResistant isolate (BC-9)Between concentration F= 10.93Between Days F=18.70C.D. at p value 0.05=53.00
251
Fig . 108 Effect of Swietenia macrophylla. King. Leaf extracts (alcoholic and
aqueous) on linear growth (mm) of resistant isolate Botrytis cinerea causing leaf
blight of rose
252
Table 92: Effect of Tagetes erecta. L. Leaf extracts (alcoholic and aqueous) on
linear growth (mm) of sensitive isolate Botrytis cinerea causing leaf blight of rose
Daysconcentration
In percentageLeaf extract
1 2 3 4
Alcoholic 00.00 00.00 00.00 00.0025
Aqueous 14.33 27.66 37.33 42.33
Alcoholic 00.00 00.00 00.00 00.0050
Aqueous 00.00 00.00 00.00 00.00
Alcoholic 00.00 00.00 00.00 00.0075
Aqueous 00.00 00.00 00.00 00.00
Alcoholic 00.00 00.00 00.00 00.00100
Aqueous 00.00 00.00 00.00 00.00
Control 20.33 41.33 61.66 80.00
Followed TukeySensitive isolate (BC-8)Between concentration F=16.62Between Days F=2.18C.D. at p value 0.05= 15.92
253
Fig. 109 Effect of Tagetes erecta. L. Leaf extracts (alcoholic and aqueous) on
linear growth (mm) of sensitive isolate Botrytis cinerea causing leaf blight of rose
254
Table 93: Effect of Tagetes erecta. L Leaf extracts (alcoholic and aqueous) onlinear growth (mm) of resistant isolate Botrytis cinerea causing leaf blight ofrose
DaysconcentrationIn percentage Leaf extract
1 2 3 4Alcoholic 00.00 00.00 00.00 00.0025 Aqueous 15.66 28.66 39.33 43.33Alcoholic 00.00 00.00 00.00 00.0050Aqueous 00.00 00.00 00.00 00.00Alcoholic 00.00 00.00 00.00 00.0075 Aqueous 00.00 00.00 00.00 00.0000.00 00.00 00.00 00.00 00.0010000.00 00.00 00.00 00.00 00.00
Control. 21.66 42.33 63.66 80.00
Followed TukeyResistant isolate(BC-9)Between concentration F= 17.91Between Days F=2.20C.D. at p value 0.05=16.35
Fig. 110 Effect of Tagetes erecta. L Leaf extracts (alcoholic and aqueous)
on linear growth (mm) of resistant isolate Botrytis cinerea causing leaf blight of
rose
