242 ENTOMOLOGIA SINICA Volume 1, Number 3, 1994, pp. 242-250

INSECTICIDE RESISTANCE OF COTTON APHID IN NORTH CHINA

Yunqin Sun, Guolei Feng, Jiagui Yuan and Kunyuan Gong Institute of Zoology. Academia Sinica. Beijing 100080, China (Accepted Apr. 26. 1993)

Abstract The cotton aphid (Aphis gossyp'i Glover) is one of the most important pests infesting cotton in the cotton areas of North China. Since 1953 organophosphorus insecticides such as parathion and systox have been used to control the aphids for keeping up good yield of cotton. After several years. the concentration and the amount of spray were increasing in the field. In the early 1980's highly effective pyrethroid insecticides such as decis and sumicidin were largely imported into China. When first used 2. 5% decis emulsion was diluted in the ratio from 1110000 to 1812000. However in 1985 the resistance of cotton aphid to pyrethroids increased by 171 times in general, 3230 times in some cotton fields. Thus it has prompted us to investigate the mechanism of resistance to insecticides and to search for the strategy to control the resistant aphids.

Experiments showed that the use of synergists including SV, (0, 0-diethyl, 0-phenyl p h w phorothionate) and PB has given evidence indicating mixed function oxidases (MFO) . a-NA esterases and a-NA carboxylesterase are involved in the formation of resistance. The results also showed that the sensitivity of AChE to paraoxon in resistant aphids was lower than that in susceptible aphids.

Experiments showed that SV1 was particularly synergistic to organophosphorus or pyrethroid insecti- cides and had played an excellent role in overcoming the resistance of cotton aphids to insecticides.

Key words Aphis @ssypiz' Glover, organophosphorus, pyrethroid, insecticide resistance

Cotton aphid (Aphis pssypii Glover) is one of the most serious insect pests in the cotton growing areas of North China. Evidently, control of cotton aphids during the cotton seeding stage is an important factor to ensure good crops of cotton. In the past, a vast amount of insecticides was used irrationally each year for protection of cotton plants, and the excessive use of insecticide consequently led to the Occurrence of aphid's resistance to in- secticides. There are roughly 4 stages of chemical control of cotton aphids in North China, and this paper summarizes the insecticides used in these stages and the transition of aphid's resistance to the insecticides applied.

The first stage (1953-1982) of chemical control was dominated by application of organophosphorus insecticides (OP) . At first, parathion and systox were used. In 1953, cotton aphids could be wiped out by parathion solution diluted in the ratio 1 : 2000 and sprayed twice: but by 1963. at least 5 times spraying ware needed by the solution diluted in the ratio 1 : 800. Therefore we were invited by the Chinese Ministry of Agriculture to conduct a survey to find out why parathion and systox became less effective to the cotton aphids. Table 1 shows the change of dosage of parathion to control cotton aphids during 1953-1963. The results of the survey showed that cotton aphids had developed their re- sistance to parathion and systox by 23 and 148 fold respectively in the cotton areas of North

Yunqin Sun et al. : Insecticide resistance of cotton aphid 2 4 3

China (Table 2). At the same time, we also synthesized many OP insecticides which were screened and tested both in the laboratory and in the field. Only methyl parathion, DDVP,

Table 1 Change of dosage of parathion to control cotton aphids during 1953-1963

concentration rate (g ai/mu) applications (g ai/mu) increase (%I

Year Effective Application Number of Total Ratio of Dilution

1953 0.0083 2. 5 1 2.50 1.0 5500

1956 0.0104 3.1 2 6.25 2.5 4400

1963 0.0420 J2.5 5 62.50 25.0 1100

Parathion 46%EC, 30kg/mu dilution

Table 2 Change of dosage of systox to control cotton aphids during 1953-1963 ~ ~~ ~~ ~

Year Effective Application Number of Total Ratio of Dilution concentration rate (g ai/mu) applications (g ai/mu) increase ( % I

1956 0.0073 2.2 2 4.4 1 6800 1959 0.015 4.4 3 13.2 3 3400 1961 0.029 8.8 4 35.2 8 1700 1963 0.037 11.0 5 66.0 15 1400

Systox 46% EC, 30 kg/mu dilution

monocrotophos and omethoate could still exert good control. In fact, since the beginning of the seventies, cotton aphids have developed strong resistance to almost all OP insecticides (Table 3) . Table 3 The changing concentrations of organophosphorus insecticides used

Inkticide Years of Effective conc. (ai%) Times of applications Beginning Final increasing

DDVP 1963-1979 0.01 0.025-0.05 2.5-5 Phosphamidon 1965- 1979 0.01 0.025-0.05 2.5-5 Methyl parathion 1965-1979 0.00625 0.025-0.05 4.0-8.0 Omethoate 1972-1979 0.01 0.020-0.025 2.0-2.5 Monocrotophos 1973-1979 0.01 0.016-0.025 1.6-2.5

The second stage was from 1982 to 1986. In the early 1980's. highly effective pyrethroid pesticieds such as deltamethrin. fenvalerate and cypermethrin were imported in large quantities. The pyrethroids proved around 30 times more effective than the

244 ENTOMOLOGIA SINICA Volume 1, Number 3, 1994

organophosphorous insecticides. Their usage spread widely and rapidly in North China, and the OP pesticides were almost driven out of the market. In 1982, 2.' 5% deltamethrin emulsion was diluted in the ratio about 18 16000 for usage. It was diluted in the ratio 1 : 8000 in 1983. By 1985, cotton aphid's resistance (LDS0 ratio) to deltamethrin had increased by 359 times in general. 3230 times in some cotton fields even higher in some other fields. The resistance to pyrethroids in cotton aphids developed rapidly in a short span of 3 or 4 years (Table 4).

Table 4 The resistance of cotton aphid to deltamethrin durinR 1980-1985

Deltamethrin Fenvalerate Year LD50 Times of LD50 Times of

(pglaphid) resistance (pg/aphid) resistance 1980 1. lox 10-5 1 1983 3.95 X 359 8.18X10-' 1 1984 9 . 2 0 ~ 10-3 836 4.58 X lo-' 56 1985 3.55x 10-2 3200 2.88 X lo- ' 350

The third stage was from 1987 to 1989. In order to understand further the cause of aphid's resistance to pesticides and look for more effective strategies to delay the Occurrence of resistance, it is necessary to investigate the biochemical mechanism of pesticide resistance of the aphids in detail. The results revealed that the development of resistance of cotton aphids to insecticide was caused by several factors to which we have made the following studies.

The penetration rates (Sun 1987) of 3 5 S - ~ y ~ t ~ ~ topically applied in acetone to the cotton aphid are lower in the resistant population than those in the susceptible one (Table 5).

Table 5 Relationship between mortalities and penetration rates of 35 S-systox in acetone topically applied to the cotton aphid

Time after Resistant population Susceptible population topical application Mortality (%) Penetration (%I Mortality (%) Penetration (%I

(min)

5 0 2.0 13.0 4.9 10 0 3.2 49.0 7.4 20 1.0 3.9 81.1 8.8 40 11.0 5.9 98.0 9.2 80 73.0 8.7

Table 6 shows that polybutadiene (PB) had significant synergism to malaoxon and in-

Yunqin Sun et al. : Insecticide resistance of cotton aphid 245

dicates Mixedfunction oxidases (MFO) is involved in cotton aphid resistance. In vitro condition (Wilkinson 1969, Brooks 19701, aldrin oxidation to dieldrin by MFO not only can be detected in aphid homogenate, but is less in the mixed homogenate of aphids and houseflies, than in housefly homogenate alone. This shows that MFO plays a role in the aphid resistance, and an endogenous inhibitor may be present in the aphids which affects housefly MFO (Table 7 ) .

Table 6 Synergism of PB and SV, to malaoxon and sumicidin in cotton aphid

Insecticide Y =a+bX LD50 (pg/aphid) SR

Sumicidin Y =7.33+0.301X 4.31 x 10-4

Sumicidin t PB Y =7.49+0.305X 2 . 8 4 ~ 10-4 1.52

Sumicidin + SV, Y =8.02+0.270X 1.38 X 31.2

Malaoxon Y =9.97+1.178X 1 .47x

Malaoxon t PB Y =11.52+1.086X 2 . 4 7 ~ 10-3 5. 96

Malaoxon t SV, Y =lo. 62+0.911X 2.09 X 7.05

SR: synergism ratio=LDso of unsynergixd/LD5,, of synergized treatment

Table 7 MFO activity of cotton aphids and houseflies and their mixture

Sample E” Ratio2’ 1n3)

100 Aphids

100 Aphids+2.5 Houseflies *)

2.5 Houseflies ‘)

0.00

0.175 28.5

0.164 100

71. 5

0

1) MFO activity, dieldrin pruduced (pg/30min), 2) MFO activity ratio (%), 3) MFO activity inhibition (%) of aphids to houseflies, 4) houseflies abdomen.

The enzyme assays for esterases (Asperen 1962) or acetylcholinesterases (AChE) are conducted at optical conditions that provide linear rates of product formation. Activity of AChE (Ellman 1961) (with or without eserine inhibition) and their sensitivity to paraoxon and SV2 are found significantly different between the resistant and susceptible populations (Table 8). Eserine-insensitive esterase, i. e. carboxylesterases, occupies 9. 94% of ester- ases in the susceptible aphids, but 45.8% in the resistance ones. These indicated that high- er esterase activity especially from a-NA carboxylesterases is one of the main factors in cotton aphid resistance (Table 9). High carboxylesterases activity in the resistant aphid hy- drolyzed a large amount of insecticides, and thus caused high resistance. Isoelectric focusing of esterases was studied and its scanning enzymograms from resistant and susceptible cotton aphids were shown in Fig. 1.

246 ENTOMOLOGIA SINICA Volume 1, Number 3, 1994

1. 0

F‘,

A

0. 5 F’ 1

9 i k 27 36 i s 54 83 i z i i

Distance (mm)

Fig. 1 Scanning zymograms of esterases from Gaomi and Beijing aphids by isoelectric focusing. A. Gaomi: B. Beijing.

Yunqin Sun et d. : Insecticide resistance of cotton aphid 247

Table 8 Inhibition of paraoxon and SV,to AChE activity in susceptible and resistant cotton aphids

Inhibition

Control” 1 . 3 8 X lo-‘ 2.14X10-‘ 1.55

Paraoxon I,, (M) 4.47 x 10-9 6.34 X lo-’ 14.20

sv, 150 (M) 1.65 x 10-5 4.31 X lo-‘ 2 . 6 1

1) AChE activity: pM/aphid*min

Table 9 Activities of a-NA esterase and a-NA carboxylesterase from aphids in different regions and their sensitivity to paraoxon

Enzyme Ekijing 6) Gaomi (R) Ratio (R/S)

Esterase activity” 1.61 24.56 15.25

Carboxylesterase activity” 0.16 11.25 70.31

150 (E600) (M) 1.39 X lo-’ 2.95 X lo-’ 213.6

150 6v2) (M) 2.97 X lo-’ 1 . 3 7 ~ 10-5 462.1

1) Esterase and carboxylesterase activities: pM/aphid emin

Fig. 2 shows the relationship between the frequency of individual esterases and resistant level. There is 71.1 % of optical density less than 0.05 and 28.9% of 0.05-1.0 OD for sus- ceptible aphids, 95.2% of 2.0-3.5 OD for resistant ones. The results suggested that each population with or without a period of insecticide selection was homogeneous and this might be used to predict resistance development.

Based on the research on resistance mechanism, strategies were sought to prevent and delay the development of resistance in cotton aphids. SV1 (0, O-diethyl, O-phenyl phos- phorothionate) is a inhibitor of MFO and carboxylesterase (Oppenoorth 1974). A series of experiments to determine the synergistic effects of SV1 and PB and to determine if SVland insecticides exhibited co-toxicity yield results a convincing manner (Tables 6 and 10). PB has significant synergism to malaoxon; SV, is good to both of them (Table 11). This suggests that resistance mechanism of cotton aphids to organophosphate relates to mixed function oxidases and esterases. For this reason, a synergist as carboxylesterases inhibitor, especially SV1 mixed with OP or pyrethroids for cotton aphid control, is one of the most ef- fective tactics of insecticide resistance management. We had synthesized SV1 and introduced it to a pesticide factory for production and it has been registered in China. From 1986 to 1990, the factory had produced 1781 tons of SV1 emulsion and 3574 tons of preparation mixtures of pyrethroid and SV,. At present, there are 5 factories producing SV, in China.

2 4 8 ENTOMOLOGIA SINICA Volume 1, Number 3. 1994

30

20

10

h

- P

- Ir

- I L -

t 40 c Beijing 0 Gaomi

~ -- 0.005 0. 0110. 051 0. 11 0. 51 1. 01 1. 51 2. 01 2. 51 3. 01 3. 51

I I l l @ I I I I I I 0.010 0.0500. 10 0.50 1.00 1. 50 2.00 2.50 3.00 3.50 4.00

O D aphid-1

Fig. 2 Relationship between the frequency of individual activity of esterases and resistance level.

Table 10 The co-toxicity coefficients of some organophosphoruses and pyrethroids to cotton aphids when mixture with SVl

LD50 SV, +insecticide Co-toxicity @/aphid) LD5, $/aphid) Coefficient

Insecticide

SVl 3.063

Malaoxon 0.0134 0.0073 365.5

Phosphamidon 0.0257 0.0384 132.74

Omethoate 0.041 0.046 175.91

Sumicidin 4.3180X 4.0699X 900.8837

De-cis 1. 6479X10-4 2.6158X 6329.4688

Specifically, SVI played a noticeable role in controlling insecticide resistance of cotton aphid. Regretfully until 1989 even the synergistic phyrethroids were unable to control the resistant cotton aphids in the key cotton areas of North China.

The fourth stage is from 1990 up to now. Cotton aphids have developed very high re- sistance to pyrethroid insecticides. so the insecticides are no longer used to control the cotton aphids in the key cotton areas in North China. In 1989 the resistance to pyrethroids in cotton bollworm ( Heliothis armigem 1 incregsed 20-30 times as much as that in susceptible

Yunqin Sun et al. : Insecticide resistance of cotton aphid 2 4 9

Table 11 Field block tests for synergistic effects of SV, to cotton aphids when SV, mixed with insecticides

Insecticide Times of No. of (% of control after days) dilution aphids 1. 3 5 7

40% omethoate Ec 1000 964 98. 2 99. 8 SV, fomethoate 1000 633 96.2 98.4 95.2 50% malaoxon EC 2000 675 94. 5

SV, +malaoxon 2000 768 98.4 80% malaoxon 2000 698 83.5 98.2 SV, +malaoxon 2000 960 95.6 98.7 50% zazodrin EC 3000 614 88.9 96.6 78.0 SV, +azodrin 3000 1000 88.0 92.8 79.4 20% sumicidin EC 8000 872 98.9 99.7 99.4 99.2 SV, +sumicidin 8000 765 95.6 94.0 94.0 92.3 2.5% decis EC 10000 1109 95.6 92.0 94.7 81.0 SV, +deck 10000 1015 92.5 97.3 94.5 88.4

population. The Chinese government has been deeply concerned with this serious situation . In 1991 National and International symposia on strategies against cotton pest resistance to pesticides were held in kijing. In 1977, Georghiou mentioned that resistance develops more rapidly with dominance, higher gene frequencies, and a greater fitness advantage to re- sistant genotypes. We have made great efforts to create necessary conditions for the devel- opment of the cultural control against the cotton aphid in cotton growing areas of North China. Meanwhile. other preventive measures in field management are taken as follows:

may suppress the population growth of cotton aphids in cotton seedling stage. Because the cotton aphids multiply relatively late in the temporal scale, its natural enemies can multiply on wheat before hand and move from wheat to cotton. In this way the cotton aphid damage can be put under control without using any pesticide. Generally speaking, if the proportion between the beneficial insects such as lady beetles or Aphidius and the aphid is at 1 : 120 or mummified aphid is over 30%. no pesticide is needed.

To dress the seeds with carbofuran and the effective period may last 50 days, so the time of cotton aphid damage can be postponed.

The resistance to insecticides may be delayed by increasing economic thresholds and re- ducing the use of pesticides. If the drawn aphids multiply seriously, monocrotophos or methomyl will be used for a single treatment.

To utilize alternatively different pesticides is another practice. Insecticide rotation and insecticide mixtures are the common tactics. The common practice with mixtures can reduce

Intercropping of wheat and cotton

250 ENTOMOLOGIA SINICA Volume 1. Number 3. 1994

the usage rates of each insecticide, which sometimes may not be sufficient to delay re- sistance. As the resistance to pyrethroids of cotton aphid and cotton bollworm has become serious, we have to limit the use of pyrethroids. The pyrethroids can be used once to e- liminate the second generation of cotton bollworm based on the amount of 100-300 eggs on a hundred cotton plants. Bacillus thuringigimis was used to eliminate the third generation of cotton bollworm only in the moist weather. In dry weather monocrotophos or methomyl can be used.

Rational pest-control strategies must be designed to manage insecticide resistance of cotton aphids, both to prolong the effectiveness of pesticides and to reduce the environmental pollution by excessive use of chemicals. These strategies should be based on integrated pest management (IPM) techniques. At present. an insecticide resistance man- agement CIRM) strategy for cotton pest insects has been used in the world. The IPM has improved insect control on crops and has been implemented to suppress pyrethroid re- sistance, to prevent reselection of some historical insecticide resistance and to solve future problems with other insecticides.

References

Asperen, K. Van, J. Ins. Physiol 8: 401.

Brooks, G. T., A. Harrison and S. E. Lewis 1970 Cyclodiene epoxide ring hydration by microsomes from mammalian livers and houseflies. B i o c h . Pharmacol. 19: 255.

Ellman, G. L. and K. D. Courtney 1961 A new and rapid colorimetric determination of acetyl- cholinesterase activity. Biochem. Ibid. harmacol. 7: 88.

Georghiou. G. P. and E. T. Charles 1977 Genetic and biological influences in the evolution of insecti- cide resistance. J. Econ. Entomol. 70: 319-323.

Oppenoorth, F. J. 1974 Development of resistance to insecticides. InThe future for insecticides: needs and prospects. R. L. Metealf and J. Mckelvey (eds. 1, Whiley, New Yrok, 6: 41-59.

Sun, Yunqin. Guolei Feng, Jiagui Yuan, Ping Zhu and Kunyuan Gong 1987 Biochemical mechanism of resistance of cotton aphids to organophosphorus insecticides. 30 (1) : 13-20.

Wilkinson. C. F. and L. J. Hicks 1969 Micrwmal metabolism of the 1, 3-benzodioxole ring and its possible significance in synergistic action.

1962 A study of housefly esterases by means of a sensitive colorimetric method.

Acta Ent. Sinica

J. Agr. Food C h . 17: 829.