INSECTICIDE RESISTANCE AND RESISTANCE MANAGEMENT
Stage-Specific Expression of Resistance to Different Acaricides in Four Field Populations of Tetranychus urticae (Acari: Tetranychidae) XIAOFENG TANG, YOUJUN ZHANG, QINGJUN WU, WEN XIE,
AND
SHAOLI WANG1
Department of Plant Protection, Institute of Vegetables and Flowers, Chinese Academy of Agricultural Sciences, Beijing 100081, P. R. China
J. Econ. Entomol. 107(5): 1900Ð1907 (2014); DOI: http://dx.doi.org/10.1603/EC14064
ABSTRACT The two-spotted spider mite, Tetranychus urticae Koch, is a worldwide crop pest. The resistance to commonly applied acaricides (in this report, “acaricide” refers to both acaricides and insecticides that are toxic to mites) has seriously impaired T. urticae control in the Þeld. Here, the sensitivity of eggs, larvae, and adults of laboratory and Þeld populations of T. urticae to various acaricides was investigated. Based on data obtained with an acaricide-sensitive laboratory strain collected in 2009, abamectin was the most toxic of the tested acaricides. For each acaricide, susceptibility was greatest for larvae, least for adults, and intermediate for eggs. The egg was the most sensitive stage to abamectin, bifenazate, and hexythiazox; the larva was the most sensitive stage to abamectin, hexythiazox, bifenazate, propargite, and chlorfenapyr; and the adult was the most sensitive stage to abamectin, bifenazate, and chlorfenapyr. Based on the results obtained with the acaricide-sensitive laboratory strain, acaricides were selected to test against eggs, larvae, and adults of four Þeld populations of T. urticae from Beijing, China. Although the Þeld populations differed in their resistance to the acaricides in laboratory bioassays, the eggs, larvae, and adults of the four populations were sensitive to bifenazate and highly resistant to abamectin. Field trials for control of T. urticae in Beijing, China, should be conducted with bifenazate and other acaricides rather than with abamectin. KEY WORDS Tetranychus urticae, acaricide resistance, developmental stage, bifenazate, abamectin
The two-spotted spider mite, Tetranychus urticae Koch (Acari: Tetranychidae), is a cosmopolitan and destructive pest of agricultural crops in China and throughout the world. It can damage ⬎1,100 host plants, including ornamentals, vegetables, and fruit trees (Jeppson et al. 1975, Grbic´ et al. 2011, Ullah and Gotoh 2013, Vassiliou and Kitsis 2013). T. urticae damages plants by feeding on the leaves, resulting in premature cell death, leaf abscission, production loss, and even plant death (Gorman et al. 2002). Owing to its high reproductive rate, very short life cycle, parthenogenesis, and propensity to rapidly develop resistance to acaricides, management of this spider mite is often very difÞcult (Jeppson et al. 1975, Nauen et al. 2001, Van Pottelberge et al. 2009a, Van Leeuwen et al. 2010). Chemical control remains the most common strategy for managing spider mites on horticultural crops in China. However, the intensive use of acaricides or insecticides (in this paper, the term acaricide is hereafter used for acaricides and for insecticides with acaricidal activity) has seriously reduced the effectiveness of the chemicals (Zhu et al. 2013), mainly because of the development of resistance. Many papers have 1
Corresponding author, e-mail:
[email protected].
reported on the development of resistance by T. urticae to various acaricides, including organophosphates, pyrethroids, carbamates, and even some recently developed compounds (Herron et al. 2004, Suh et al. 2006, Van Leeuwen and Tirry 2007, Kwon et al. 2010a, Van Leeuwen et al. 2010, Vassiliou and Kitsis 2013). The resistance mechanisms of T. urticae to the acaricides are complex but mainly include enhancement of the cytochrome P450 mixed-function oxidase system and esterases and point mutations in sodium channels (Van Leeuwen et al. 2005, Tsagkarakou et al. 2009, Kwon et al. 2010a, Van Leeuwen et al. 2010, Tirello et al. 2012). When different acaricides with the same or similar mode of action have been used in the Þeld, cross-resistance has often enhanced the development of resistance (Uesugi et al. 2002). The T. urticae population from Shandong Province in China has developed high resistance to abamectin, pyridaben, and other commonly used acaricides (Zhao et al. 2001, Liu et al. 2012a). Even though T. urticae is highly resistant to many pesticides, acaricides are still important for its management in China. In some cases, a speciÞc pesticide is effective against only one or two developmental stages of a pest (Nauen et al. 2001, Sa´enz-de-Cabezo´ n et al. 2006). Thus, after T. urticae was subjected to
0022-0493/14/1900Ð1907$04.00/0 䉷 2014 Entomological Society of America
October 2014
TANG ET AL.: ACARICIDE RESISTANCE IN T. urticae
many generations of selection by spirodiclofen, the resistance level was much higher for larvae than for eggs (Van Pottelberge et al. 2009b). Sensitivity to a particular acaricide can even differ among the different periods in the same developmental stage of the pest (Marcic 2003, Kramer and Nauen 2011). Pest managers can delay the development of resistance by spraying mixtures of acaricides that do not exhibit cross-resistance and that do not interfere with each otherÕs efÞcacy (Herron et al. 2003). In addition to delaying the development of resistance, the application of pesticide mixtures can also provide better pest control than the application of single pesticides (Ahmad et al. 2009, Khan et al. 2013). Development of resistance can also be reduced by alternating or rotating pesticides (Vassiliou and Kitsis 2013). For the effective use of pesticides in mixtures or rotation, information is required about which stages of the pest are sensitive to which pesticides (Cloyd 2009). Beginning in 2012, growers in some areas in Beijing, China, found that abamectin and other commonly used acaricides failed to control T. urticae. This failure has been attributed to acaricide resistance and the improper use of acaricides (S. W., unpublished data). In this study, we Þrst investigated whether the sensitivity to 11 acaricides differed among eggs, larvae, and adults of an acaricide-sensitive laboratory strain of T. urticae. Based on the results obtained with the laboratory strain, the most effective acaricides were selected to test against eggs, larvae, and adults of four Þeld populations of T. urticae from Beijing, China. The results obtained from this laboratory study will be useful for improving the control and resistance management of T. urticae in the Þeld. Materials and Methods Chemicals. The following 11 pesticides, which are commercially available, were used in the bioassays: abamectin (18 EC; Hebei Veyong biochemical Co., Ltd., Hebei, China); propargite (730 EC; Chemtura Corporation, Shanghai, China); hexythiazox (50 EC; Jiangsu Rotam Chemistry Co., Ltd, Jiangsu, China); pyridaben (150 EC; Jiangsu Kesheng Group Co., Ltd, Jiangsu, China); chlorfenapyr (100 SC; BASF Chemical Co., Ltd, Germany); bifenthrin (100 EC; Bayer Cropscience China Co., Ltd, China); bifenazate (430 SC; Chemtura Corporation); profenofos (400 EC; Nanjing Red Sun Group Corporation, Nanjing, China); azocyclotin (250 WP; Syngenta Crop Protection Company, Shandong, China); clofentezine (200 SC; Shandong Heze beilian pesticide manufacture Co., Ltd, Shangdong, China); and spirotetramat (240 SE; Bayer CropSciences, Hangzhou, China). Among the 11 pesticides, abamectin, bifenthrin, and chlorfenapyr are registered as both insecticide and acaricides, and propargite, hexythiazox, pyridaben, bifenazate, azocyclotin, and clofentezine are registered as acaricides. The remaining two pesticides (profenofos and spirotetramat) are insecticides that have some activity against spider mites and are occasionally used as acaricides in China.
1901
Table 1. Background information on the T. urticae field populations used in this study Population HD CP HR MY
Collection location in Beijing Haidian Changping Huairou Miyun
Collection date
Host plant
May 2013 Sept. 2013 June 2013 Oct. 2013
Eggplant Eggplant Strawberry Cucumber
Mite Strains. Laboratory Strain of T. urticae. In 2009, specimens of an acaricide-susceptible strain of T. urticae were collected from an apple tree in TaiÕan, Shandong, China. The main chemicals used to control the mites in the apple orchard had been bifenthrin and profenofos. Before it was used in bioassays, the laboratory mite strain was maintained for over 50 generations on bean leaf discs (ÔBifengÕ) without exposure to any pesticides in an incubator at 26 ⫾ 1⬚C, 60 Ð 80% relative humidity (RH), and a photoperiod of 16:8 (L:D) h each day. Field Populations of T. urticae. T. urticae specimens were collected in 2013 from three host plants (eggplant, cucumber, and strawberry) in Haidian (HD), Changping (CP), Huairou (HR), and Miyun (MY) in Beijing, China (Table 1). Among the crops grown in these four locations, strawberry and eggplant experience the most severe T. urticae infestations and have been commonly treated with abamectin. As noted earlier, growers in these locations reported that abamectin had failed to control T. urticae from 2012 to the present (2014). Three subsamples of T. urticae were collected from each location by the Þve-spot-sampling method. Subsampling areas were separated by at least 1,000 m, and 500 Ð1000 spider mites were collected in each subsample. The spider mites in the three subsamples were combined to obtain one sample per location. The specimens from each of the four locations were considered to represent one Þeld population. T. urticae adults were transported to the laboratory with the original leaves and then transferred to the bean leaf discs in the incubator as described earlier. Before the bioassays, the four Þeld populations were maintained on bean leaf discs for three generations; this was done to reduce differences in their physiology and to generate the numbers required (Vassiliou and Kitsis 2013). Egg and Larva Bioassays. The ovicidal and larvicidal toxicity of the acaricides to T. urticae eggs and larvae were assessed with the leaf dip method according to Tirello et al. (2012). For eggs, 10 adult females (3Ð5 d old) were placed on a 20-mm-diameter bean leaf disc in a petri dish, where they deposited eggs. After 12 h, the females were removed, and the leaf discs (each with ⬇35 eggs) were dipped into one of the acaricide solutions (Þve to seven concentrations were used for each acaricide) for 5 s according to the method of He et al. (2011) and Roh et al. (2011). After the discs were removed from the acaricide solution, the excess solution on each leaf disc was removed by contact with a small piece of Þlter paper, and the discs
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were then air-dried in a fume hood. As controls, discs with eggs were dipped into double-distilled water. Each concentration of each acaricide was represented by four replicate discs. After the discs had dried, the eggs were counted, and the petri dishes with the treated leaf discs were placed in an incubator as described earlier. Mortality was assessed daily starting with the eclosion of the Þrst protonymph (⬇6 d after treatment) and continuing for Þve successive days; by 6 d after treatment, all the eggs in the controls had eclosed except those that had died because of physiological causes. Egg mortality was determined by subtracting the number of protonymphs from the total number of eggs. For bioassays with larvae, adult females of T. urticae were placed on the leaf discs and allowed to lay eggs for 12 h as described in the previous paragraph. Once larvae emerged from the ⬇35 eggs per disc, the discs were dipped into the acaricide solutions (Þve to seven concentrations were used for each acaricide) for 5 s. The leaf discs with larvae were dried and stored in an incubator as described in the previous paragraph. After 4 Ð5 d, all larvae in the control treatment had developed into adults, and the larval mortality rate for each treatment was calculated based on the number of larvae and the number of adults that developed per treatment. Adult Female Bioassay. Bioassays with adult females of T. urticae were conducted using the slide-dip method recommended by FAO (Dittrich et al. 1980). Brießy, double-sided sticky tape was cut into 2- to 3-cm-long pieces, which were attached to one end of a glass slide. Healthy adult females of the same size were gently stuck to the tape by their backs (not by their feet or mouthparts) using a small brush. Three rows with 15 individuals per row were stuck to each slide. Slides with mites were placed in an incubator for 4 h. The mites were then examined with a dissecting microscope, and dead and inactive individuals were removed. Then the ends of the slides with mites were immersed and gently shaken for 5 s in acaricide solutions at different concentrations, and excess liquid was promptly removed with absorbent paper. Slides dipped in water served as the controls. The slides were subsequently placed in a porcelain container, which had saturated gauze on the bottom and top and which was placed in the incubator. Each concentration of each acaricide was represented by four replicate slides. After 24 h, the dead mites were counted with a dissecting microscope. A mite was regarded as dead if it did not respond when gently touched with a small brush. If the mortality rate in the control treatment exceeded 10%, the assay was rejected, and a new assay was performed. Data Analysis. Mortality data for each stage were corrected using the AbbottÕs formula (Abbott 1925), and the LC50 values of the tested acaricides and their 95% Þducial limits and slope ⫾ SE were calculated from probit analysis using the Polo Plus Version 1.0 software. The relative toxicity (RT) of each acaricide against the three stages (egg, larva, and adult) was calculated by dividing the LC50 value of a stage by the
Vol. 107, no. 5
LC50 value of the most susceptible stage. Resistance ratios (RRs) were calculated by dividing the LC50 value of the Þeld population by the LC50 value of the laboratory strain. Results Baseline Sensitivity of Eggs, Larvae, and Adults to 11 Acaricides. Bioassay results with the acaricidesensitive T. urticae laboratory strain showed that acaricide toxicity depended on both the acaricide and the T. urticae developmental stage (Table 2). In the case of adult females, abamectin with an LC50 of 0.091 mg/liter was more toxic than the other acaricides, and bifenazate and chlorfenapyr were the next most toxic. The eggs of T. urticae were more sensitive to abamectin, hexythiazox, and bifenazate than to the other chemicals; abamectin, hexythiazox, and bifenazate had LC50 values ⬍0.60 mg/liter. For larvae, the most toxic materials were abamectin, hexythiazox, bifenazate, chlorfenapyr, and propargite (Table 2). When all acaricides tested in this study were considered, susceptibility was greatest for larvae, intermediate for eggs, and least for adults. RT values for eggs ranged from 1.18 to 207.22. RT values for adults ranged from 4.24 to 187,500 (Table 2). Resistance Monitoring With Egg Bioassay. Of the three acaricides that were tested against eggs of the four Þeld populations, bifenazate and hexythiazox were much more toxic than abamectin (Table 3). CP and MY eggs were not resistant to hexythiazox but HR and HD eggs were, with RR values of 636 and 3,121, respectively. CP, MY, and HR eggs had low resistance to bifenazate, with RR values ⬍7. The resistance of eggs against bifenazate and hexythiazox was highest in the HD population. The resistance of eggs to abamectin was extremely high in all four Þeld populations, with RR values ⬎20,000 (Table 3). Resistance Monitoring With Larval Bioassay. Of the Þve acaricides tested against larvae of the four Þeld populations, bifenazate was the most toxic, with LC50 values ⬍1 mg/liter; moderate resistance to bifenazate, however, was evident with populations HR, CP, and HD (Table 4). The second most toxic acaricide against larvae of the four Þeld populations was propargite, with LC50 values ⬍7 mg/liter. Larval resistance to the acaricides differed among the four populations. The LC50 values for hexythiazox and bifenazate were signiÞcantly higher for HD larvae than for the larvae of the other three populations. The HR and HD larvae were more resistant than the CP or MY larvae to hexythiazox and propargite. Larvae of all four populations were highly resistant to abamectin, with RR values ⬎10,000 (Table 4). Resistance Monitoring With Adult Bioassay. Adult females from the four Þeld populations did not show resistance to bifenazate, and the RR values were close to 1 (Table 5). The adult females were sensitive or somewhat resistant to chlorfenapyr depending on the Þeld population. In the case of abamectin, however, adult females from all four Þeld populations were highly resistant in that LC50 values exceeded 150 mg/
October 2014 Table 2. Acaricide Abamectin Bifenazate Chlorfenapyr Profenofos Propargite Bifenthrin Pyridaben Azocyclotin Hexythiazox Clofentezine Spirotetramat
TANG ET AL.: ACARICIDE RESISTANCE IN T. urticae
1903
Baseline sensitivity of eggs, larvae, and adult females of the laboratory strain of T. urticae to 11 acaricides Stage
na
LC50 (95% FL; mg/liter)b
RTc
Egg Larval Adult Egg Larval Adult Egg Larval Adult Egg Larval Adult Egg Larval Adult Egg Larval Adult Egg Larval Adult Egg Larval Adult Egg Larval Adult Egg Larval Adult Egg Larval Adult
1,016 954 473 1,056 830 506 1,010 890 560 1,252 691 585 968 1,093 508 1,028 804 625 882 723 597 835 881 511 885 977 569 980 740 532 1,117 774 574
0.0019 (0.0017Ð0.0022) 0.0011 (0.00079Ð0.0015) 0.091 (0.065Ð0.15) 0.57 (0.47Ð0.66) 0.027 (0.020Ð0.035) 23.92 (16.56Ð33.42) 3.85 (2.94Ð5.22) 0.55 (0.39Ð0.75) 107.33 (90.13Ð125.74) 239.29 (183.43Ð319.63) 66.65 (37.36Ð151.82) 282.90 (245.06Ð325.09) 39.37 (32.28Ð47.18) 0.19 (0.16Ð0.23) 364.82 (259.57Ð502.13) 18.61 (16.53Ð20.84) 6.30 (4.41Ð8.77) 366.75 (314.67Ð426.95) 28.67 (18.70Ð41.34) 7.74 (6.36Ð9.45) 714.60 (553.98Ð987.05) 3.24 (2.24Ð4.36) 1.24 (0.90Ð1.59) 1, 010.71 (761.22Ð1, 453.53) 0.032 (0.026Ð0.040) 0.016 (0.014Ð0.017) ⬎3,000 24.11 (17.47Ð31.82) 20.40 (17.18Ð23.79) ⬎5,000 21.57 (16.73Ð27.83) 6.05 (4.91Ð7.26) ⬎5,000
1.73 1 82.73 21.11 1 885.93 7 1 195.15 3.59 1 4.24 207.22 1 1,920.11 2.95 1 58.21 3.70 1 92.33 2.61 1 815.09 2 1 ⬎1,87500 1.18 1 ⬎245 3.57 1 ⬎826
Fit of probit analysis Slope ⫾ SE
2
df
1.80 ⫾ 0.13 2.45 ⫾ 0.17 1.19 ⫾ 0.20 3.19 ⫾ 0.23 2.20 ⫾ 0.16 1.75 ⫾ 0.18 2.03 ⫾ 0.16 1.46 ⫾ 0.12 2.02 ⫾ 0.19 1.53 ⫾ 0.10 1.18 ⫾ 0.13 2.45 ⫾ 0.21 3.41 ⫾ 0.24 2.78 ⫾ 0.17 1.78 ⫾ 0.17 2.41 ⫾ 0.16 1.53 ⫾ 0.11 1.89 ⫾ 0.16 1.69 ⫾ 0.12 1.42 ⫾ 0.16 1.26 ⫾ 0.15 2.42 ⫾ 0.21 2.50 ⫾ 0.18 2.26 ⫾ 0.25 2.76 ⫾ 0.18 3.48 ⫾ 0.25 N/A 2.31 ⫾ 0.16 2.17 ⫾ 0.16 N/A 1.78 ⫾ 0.12 1.66 ⫾ 0.14 N/A
1.49 7.89 0.23 3.59 5.06 3.56 5.55 3.50 2.75 7.50 5.67 0.22 4.66 6.30 3.69 2.53 3.76 0.23 5.77 2.93 1.46 5.22 5.13 3.67 4.00 1.25
3 3 3 3 3 3 3 3 3 4 3 3 3 4 3 4 3 3 3 3 4 3 3 3 3 3
6.87 1.50
3 3
6.87 1.35
4 3
For simplicity, the term “acaricide” is used for both acaricides and insecticides in all tables. a Number of specimens used in the bioassay, including controls. b Lethal concentration (95% Þducial limit). c RT, LC50 of a stage/LC50 of the most susceptible stage for the same acaricide.
liter, and RR values ranged from 1,926 Ð 4,989 (Table 5). Discussion T. urticae is a serious threat to horticultural crops, especially when it has developed resistance to many insecticides. Chemical control is an important comTable 3. Acaricide Abamectin
Hexythiazox
Bifenazate
a b c
ponent of integrated pest management (IPM) for this pest mite in China, and monitoring of local populations for resistance to acaricides is essential for T. urticae control and resistance management. The current study shows that resistance to various acaricides differs among T. urticae developmental stages and Þeld populations from Beijing, China.
Resistance of eggs from four field populations of T. urticae to selected acaricides Population
na
LC50 (95% FL; mg/liter)b
RRc
RR (95% FL)
HR CP MY HD HR CP MY HD HR CP MY HD
990 787 940 905 1,017 833 981 920 1,004 1,007 759 799
76.04 (58.74Ð104.64)ab 109.28 (87.90Ð136.45)b 52.01 (36.33Ð71.44)a 64.53 (51.81Ð84.32)ab 20.65 (17.90Ð23.52)b 0.049 (0.033Ð0.071)a 0.055 (0.045Ð0.065)a 101.31 (76.66Ð138.32)c 3.44 (2.69Ð4.65)b 1.69 (1.38Ð1.99)a 2.45 (2.18Ð2.73)b 6.40 (5.33Ð7.65)c
39,504 56,771 27,019 30,970 636 1.51 1.68 3,121 6.09 2.99 4.33 11.32
28,729Ð54,319 43,721Ð73,716 21,918Ð33,306 25,378Ð44,283 532Ð761 1.23Ð1.86 1.44Ð1.96 2,545Ð3,827 5.06Ð7.33 2.61Ð3.43 3.73Ð5.03 9.83Ð13.04
Fit of probit analysis Slope ⫾ SE
2
df
1.01 ⫾ 0.11 1.34 ⫾ 0.13 1.72 ⫾ 0.13 1.11 ⫾ 0.13 2.30 ⫾ 0.15 1.65 ⫾ 0.11 2.70 ⫾ 0.18 1.54 ⫾ 0.15 1.91 ⫾ 0.17 3.85 ⫾ 0.36 2.99 ⫾ 0.23 2.74 ⫾ 0.25
3.46 1.44 5.04 2.46 1.98 5.38 3.57 3.25 5.62 3.81 2.89 3.33
4 3 3 3 3 3 3 3 4 3 3 3
Number of eggs used in the bioassay, including controls. Different letters indicate nonoverlap of Þducial limits (P ⬍ 0.05). RR, the LC50 of a Þeld population/the LC50 of the laboratory strain (for the same developmental stage).
1904 Table 4. Acaricide Abamectin
Hexythiazox
Bifenazate
Propargite
Chlorfenapyr
a b c
JOURNAL OF ECONOMIC ENTOMOLOGY Resistance of larvae from four field populations of T. urticae to selected acaricides Population
na
LC50 (95% FL; mg/liter)b
RRc
RR (95% FL)
HR CP MY HD HR CP MY HD HR CP MY HD HR CP MY HD HR CP MY HD
654 1,107 754 676 812 783 1,019 658 665 711 809 617 713 613 608 763 730 762 674 775
38.13 (28.77Ð53.04)b 47.64 (34.17Ð66.97)b 44.37 (33.70Ð63.30)b 16.31 (12.25Ð22.58)a 46.05 (31.33Ð70.90)b 0.13 (0.11Ð0.16)a 0.20 (0.15Ð0.25)a 89.00 (51.49Ð142.73)b 0.47 (0.36Ð0.59)b 0.33 (0.29Ð0.39)b 0.12 (0.084Ð0.16)a 0.82 (0.61Ð1.22)c 6.79 (5.70Ð7.90)b 1.13 (0.83Ð1.53)a 1.94 (0.98Ð4.15)a 4.94 (3.05Ð10.20)ab 113.99 (79.66Ð161.58)a 335.98 (215.80Ð928.17)b 182.29 (129.22Ð323.83)ab 490.36 (374.82Ð728.74)b
33,923 42,378 39,475 14,506 2,955 8.47 12.56 5,710 17.45 12.42 4.37 30.52 35.77 6.00 10.20 25.98 207.52 611.65 331.86 892.70
24,535Ð46,902 33,864Ð53,032 28,381Ð54,905 10,468Ð20,103 2423Ð3603 7.05Ð10.17 10.44Ð15.11 4,543Ð7,176 14.66Ð20.76 10.18Ð15.15 3.58Ð5.34 21.45Ð43.42 29.43Ð43.48 4.89Ð7.25 7.52Ð13.85 18.45Ð36.58 155.87Ð276.29 409.66Ð913.23 207.72Ð530.17 617.14Ð1291.30
Table 5. Acaricide Abamectin
Bifenazate
Chlorfenapyr
b c
Fit of probit analysis Slope ⫾ SE
2
df
0.99 ⫾ 0.12 1.25 ⫾ 0.087 0.99 ⫾ 0.13 1.00 ⫾ 0.10 1.44 ⫾ 0.13 2.12 ⫾ 0.16 2.06 ⫾ 0.14 1.52 ⫾ 0.14 3.01 ⫾ 0.26 2.19 ⫾ 0.20 2.06 ⫾ 0.14 1.14 ⫾ 0.16 2.15 ⫾ 0.18 1.99 ⫾ 0.18 1.07 ⫾ 0.10 1.04 ⫾ 0.11 1.46 ⫾ 0.15 1.26 ⫾ 0.20 1.19 ⫾ 0.19 1.31 ⫾ 0.18
1.11 6.02 1.67 2.88 9.28 3.78 4.59 5.71 4.04 2.72 4.72 1.49 0.98 3.67 6.13 6.01 4.78 3.08 2.78 0.89
3 4 3 3 4 4 4 3 3 3 3 3 3 3 3 4 4 3 3 3
Number of larvae used in the bioassay, including controls. Different letters indicate nonoverlap of Þducial limits (P ⬍ 0.05). RR, the LC50 of a Þeld population/the LC50 of the laboratory strain (for the same developmental stage).
For each insecticide or acaricide, the target pest or developmental stage differs depending on the chemicalÕs mode of action. In the current study, toxicity bioassays were conducted with a laboratory strain of T. urticae to identify the most effective, stage-speciÞc acaricide; as noted earlier, the term “acaricide” in this paper is used for both acaricides and insecticides to simplify the presentation. Across all of the tested acaricides, susceptibility was greatest for larvae, intermediate for eggs, and least for adults. Relative toxicity (RT) values for adults ranged from 4.24 to 187,500 (Table 2). This is in accord with the other reports, which showed that larvae of the German susceptible strain of T. urticae (GSS) were more sensitive than adults to many acaricides including abamectin, spirodiclofen, and chlorpyrifos (Nauen et al. 2001, Rauch and Nauen 2002, Stumpf and Nauen 2002, Khajehali et al. 2010, Ay and Kara 2011). It follows that chemical
a
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control of T. urticae in the Þeld should focus on the adult stage followed by the egg stage. In the bioassays with the T. urticae laboratory strain, hexythiazox was highly toxic to eggs and larvae but not to adult females, and hexythiazox was therefore chosen to test against the eggs and larvae of the four Þeld populations. In the tests with Þeld populations (HR, HD, CP, and MY), HR and HD eggs and larvae were highly resistant to hexythiazox but CP and MY eggs and larvae were not (Tables 3 and 4). Thus, when eggs or larvae are the dominant stage(s), hexythiazox could be used to control T. urticae in the Changping and Miyun Districts but not in the Huairou and Haidian Districts of Beijing. More specially, CP and MY populations have developed low resistance (8 ⬍ RR ⬍ 13) to hexythiazox in the larval stage but remain susceptible in the egg stage (RR ⬍ 2). In addition, the four Þeld populations shared low to high resistance to bife-
Resistance of adult females from four field populations of T. urticae to selected acaricides Population
na
LC50 (95% FL; mg/liter)b
RRc
RR (95% FL)
HR CP MY HD HR CP MY HD HR CP MY HD
442 450 556 411 501 485 612 634 430 434 576 450
268.27 (131.66Ð475.50)ab 452.46 (363.23Ð562.41)b 174.64 (115.69Ð263.78)a 204.55 (115.45Ð308.58)a 26.86 (17.26Ð42.04)a 19.63 (14.55Ð25.89)a 25.24 (18.27Ð35.05)a 30.04 (21.03Ð44.72)a 600.36 (441.98Ð841.32)ab 255.02 (179.11Ð446.01)a 597.24 (455.26Ð749.78)b 120.44 (56.21Ð208.46)a
2,958 4,989 1,926 2,255 1.12 0.82 1.06 1.26 5.59 2.38 5.57 1.12
1,826Ð4,791 3,156Ð7,886 1,248Ð2,972 1,409Ð3,611 0.84Ð1.50 0.58Ð1.16 0.83Ð1.35 0.96Ð1.64 3.93Ð7.97 1.50Ð3.75 4.56Ð6.80 0.83Ð1.51
Fit of probit analysis Slope ⫾ SE
2
df
1.33 ⫾ 0.14 1.97 ⫾ 0.21 2.00 ⫾ 0.21 1.67 ⫾ 0.22 1.50 ⫾ 0.17 1.30 ⫾ 0.16 2.08 ⫾ 0.17 1.71 ⫾ 0.17 1.23 ⫾ 0.20 1.17 ⫾ 0.21 3.13 ⫾ 0.35 1.67 ⫾ 0.26
5.46 1.44 6.03 3.65 3.94 0.27 8.54 7.41 1.68 1.82 4.86 4.91
3 3 3 3 3 3 4 4 3 3 3 3
Number of adult females used in the bioassay, including controls. Different letters indicate nonoverlap of Þducial limits (P ⬍ 0.05). RR, the LC50 of a Þeld population/the LC50 of the laboratory strain (for the same developmental stage).
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TANG ET AL.: ACARICIDE RESISTANCE IN T. urticae
nazate and chlorfenapyr as larvae or eggs but were susceptible as adults (RR ⬍ 1.3). Interestingly, similar Þndings were found for another T. urticae strain and for Panonychus ulmi and Bemisia tabaci (Nauen et al. 2008, Van Pottelberge et al. 2009b, Kramer and Nauen 2011). Stage-speciÞc resistance to insecticides or acaricides is likely related to differences in metabolic detoxiÞcation (resulting from differences in the expression of detoxifying enzymes) among life stages and differences in the expression of resistance genes such as glutathione S-transferase (GST) and the cytochrome P450 (Yang et al. 2013). In the current study, the broad-spectrum insecticide and acaricide abamectin was highly toxic to eggs, larvae, and adult females of the T. urticae laboratory strain. This was not the case, however, for the four Þeld populations of T. urticae from the Beijing area. For the Þeld populations, RR values exceeded 1,000 for adult females and exceeded 10,000 for eggs and larvae. These high RR values are consistent with recent statements from growers in Beijing that this mite cannot be controlled by abamectin. High abamectin RR values were also reported for the T. urticae populations collected in Changle and Shouguang, Shandong Province, China (Liu et al. 2012b), and also for adult females collected from Korea and Cyprus (Kwon et al. 2010b, Vassiliou and Kitsis 2013). T. urticae resistance to abamectin may result from enhanced mixed-function oxidase and esterase activities (Kwon et al. 2010b) and also due to a point mutation (G323D) in the glutamate-gated chloride channel (Kwon et al. 2010c). A PCR assay was developed to monitor the frequency of this G323D point mutation in the same four populations from Beijing, China, investigated in the current study; the PCR assay detected a high frequency (85Ð100%) of the resistant genotype in all four populations (Tang et al. 2014). Therefore, the use of abamectin for controlling T. urticae populations in the four tested areas in Beijing should be immediately terminated. Bifenazate was discovered in 1990, Þrst commercialized in 1999, and approved for Þeld application in China in 2008 (Van Leeuwen et al. 2006a, Cao et al. 2011). The toxicity and selectivity of bifenazate against pest mites have been previously reported (Ochiai et al. 2007, Ullah and Gotoh 2013). The current study conÞrmed that bifenazate is highly toxic to eggs and larvae of the T. urticae laboratory strain (with LC50 values of 0.027 and 0.57 mg/liter, respectively) but less toxic to adult females (LC50 ⫽ 23.92 mg/liter; Table 2). For the three developmental stages of the four T. urticae Þeld populations, the LC50 values were obviously lower for bifenazate than for the other acaricides. Another advantage of bifenazate is that it does not reduce the survival or fecundity of the predatory mites Phytoseiulus persimilis and Neoseiulus californicus (Ochiai et al. 2007) and is therefore compatible with biological control. It follows that bifenazate should be considered a promising acaricide against T. urticae. The four Þeld populations, however, did exhibit tolerance or a low to moderate level of resistance to
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bifenazate. The bifenazate RR values were ⱕ1.26 for adults but ranged from 4.37 to 30.52 for larvae and from 2.99 to 11.32 for eggs. Similar resistance levels were found for T. urticae in South Korea, Jordan, and Cyprus (Lee et al. 2003, Al-Antary et al. 2012, Vassiliou and Kitsis 2013). Therefore, pest managers must be careful to avoid selecting for bifenazate resistance in pest mite populations. Although cross-resistance between abamectin and bifenazate seems unlikely because the four T. urticae Þeld populations were susceptible to bifenazate but highly resistant to abamectin, abamectin and bifenazate should not be used together because these chemicals have similar activities against T. urticae life stages. Also, bifenazate should not be mixed with carbamates or organophosphates in the Þeld, because organophosphates and carbamates interfere with bifenazate efÞcacy (Van Leeuwen et al. 2007). Because the resistance of T. urticae to various acaricides differs among locations, the management of T. urticae insecticide resistance must be based on local monitoring. The data presented in the current study indicate that resistance to more than one acaricide, including the new acaricide bifenazate, is common among the Þeld populations of T. urticae in Beijing. Therefore, substantial attention should be paid to resistance development. Avoiding the further development of resistance will require that acaricides with different modes of action be mixed or alternated. Mixing acaricides that are active against different life stages should reduce the development of resistance and increase control efÞcacy. Although the RRs of T. urticae adult females from Beijing to chlorfenapyr were ⬍6, chlorfenapyr toxicity was nevertheless low because its LC50 was ⬎100 mg/liter. In addition, chlorfenapyr and bifenazate exhibit positive cross-resistance (Van Leeuwen et al. 2006b) and should not be used simultaneously or more than twice in the same growing season. Based on the current results for T. urticae populations in Beijing, mixing and alternating bifenazate and hexythiazox or propargite should provide a high level of control while delaying the development of acaricide resistance. Spirodiclofen and fenpyroximate are the most toxic insecticides against T. urticae eggs in Shandong and Henan Provinces, China (Liu et al. 2012b). Because of the rapid development of acaricide resistance in some T. urticae populations, the use of selective acaricides or growth regulators, e.g., bifenazate or spiromesifen, in association with predatory mites should be considered for the management of T. urticae (Rhodes and Liburd 2006, Sato et al. 2011). Given the current status of acaricide resistance in T. urticae populations, biological control would be a good choice for the sustainable management of T. urticae. Acknowledgments This research was supported by the 863 Program (2012AA101502), the Special Fund for Agro-scientiÞc Research in the Public Interest (201103020), the China Agriculture Research System (CARS-26-10), and the Beijing Key Laboratory for Pest Control and Sustainable Control. The
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JOURNAL OF ECONOMIC ENTOMOLOGY
granting agencies had no role in study design, data collection and analysis, decision to publish, or manuscript preparation.
References Cited Abbott, W. S. 1925. A method of computing the effectiveness of an insecticide. J. Econ. Entomol. 18: 265Ð267. Ahmad, M., M. A. Saleem, and A. H. Sayyed. 2009. EfÞcacy of insecticide mixtures against pyrethroid- and organophosphate-resistant populations of Spodoptera litura (Lepidoptera: Noctuidae). Pest Manage. Sci. 65: 266 Ð274. Al-Antary, T. M., M.R.L. Al Lala, and M. I. Abdel-Wali. 2012. Response of seven populations of the two-spotted spider mite (tetranychus urticae koch) for bifenazate acaricide on cucumber (cucumis sativus l.) under plastic houses in Jordan. Adv. Environ. Biol. 6: 2203Ð2207. Ay, R., and F. E. Kara. 2011. Toxicity, inheritance and biochemistry of clofentezine resistance in Tetranychus urticae. Insect Sci. 18: 503Ð511. Cao, W., G. B. Chen, Y. F. Guan, and Y. J. Li. 2011. Determination of Bifenazate by HPLC. Agrochemicals 50: 37Ð 38. Cloyd, R. A. 2009. Getting mixed-up: are greenhouse producers adopting appropriate pesticide mixtures to manage arthropod pests? HortTechnology 19: 638 Ð 646. Dittrich, V., J. E. Cranham, L. R. Jepson and W. Helle. 1980. Revised method for spider mites and their eggs (e.g. Tetranychus spp. and Panonychus ulmi Koch), FAO method No. l0a. FAO Plant Production and Protection Paper 21, pp. 49 Ð53. FAO, Rome, Italy. Gorman, K., F. Hewitt, G. Devine, and I. Denholm. 2002. New developments in insecticide resistance in the glasshouse whiteßy (Trialeurodes vaporariorum) and the two spotted spider mite (Tetranychus urticae) in the UK. Pest Manage. Sci. 58: 123Ð130. Grbic´ , M., T. Van Leeuwen, R. M. Clark, S. Rombauts, P. Rouze´, V. Grbic´ , E. J. Osborne, W. Dermauw, P. C. Ngoc, F. Ortego, et al. 2011. The genome of Tetranychus urticae reveals herbivorous pest adaptations. Nature 479: 487Ð 492. He, H. G., H. B. Jiang, Z. M. Zhao, and J. J. Wang. 2011. Effects of a sublethal concentration of avermectin on the development and reproduction of citrus red mite, Panonychus citri (McGregor) (Acari: Tetranychidae). Internat. J. Acarol. 37: 1Ð9. Herron, G. A., J. Rophail, J. Holloway, and I. Barchia. 2003. Potentiation of a propargite and fenpyroximate mixture against two-spotted spider mite, Tetranychus urticae (Acari: Tetranychidae). Exp. Appl. Acarol. 29: 115Ð119. Herron, G. A., J. Rophail, and L. J. Wilson. 2004. Chlorfenapyr resistance in two-spotted spider mite (Acari: Tetranychidae) from Australian cotton. Exp. Appl. Acarol. 34: 315Ð321. Jeppson, L. R., H. Keifer, and E. W. Baker. 1975. Mites injurious to economic plants. University of California Press, Berkley, CA. Khajehali, J., T. Van Leeuwen, M. Grispou, E. Morou, H. Alout, M. Weill, L. Tirry, J. Vontas, and A. Tsagkarakou. 2010. Acetylcholinesterase point mutations in European strains of Tetranychus urticae (Acari: Tetranychidae) resistant to organophosphates. Pest Manage. Sci. 66: 220 Ð 228. Khan, H.A.A., W. Akram, S. A. Shad, and J. J. Lee. 2013. Insecticide mixtures could enhance the toxicity of insecticides in a resistant dairy population of Musca domestica L. PloS ONE 8: e60929. Kramer, T., and R. Nauen. 2011. Monitoring of spirodiclofen susceptibility in Þeld populations of European red
Vol. 107, no. 5
mites, Panonychus ulmi (Koch) (Acari: Tetranychidae), and the cross-resistance pattern of a laboratory-selected strain. Pest Manage. Sci. 67: 1285Ð1293. Kwon, D. H., D. Y. Song, S. Kang, J. J. Ahn, J. H. Lee, B. R. Choi, S. W. Lee, J. H. Kim, and S. H. Lee. 2010a. Residual contact vial bioassay for the on-site detection of acaricide resistance in the two-spotted spider mite. J. Asia-PaciÞc Entomol. 13: 333Ð337. Kwon, D. H., G. M. Seong, T. J. Kang, and S. H. Lee. 2010b. Multiple resistance mechanisms to abamectin in the twospotted spider mite. J. Asia-PaciÞc Entomol. 13: 229 Ð232. Kwon, D. H., K. S. Yoon, J. M. Clark, and S. H. Lee. 2010c. A point mutation in a glutamate-gated chloride channel confers abamectin resistance in the two-spotted spider mite, Tetranychus urticae Koch. Insect Mol. Biol. 19: 583Ð 591. Lee, Y. S., M. H. Song, K. S. Ahn, K. Y. Lee, J. W. Kim, and G. H. Kim. 2003. Monitoring of acaricide resistance in two-spotted spider mite (Tetranychus urticae) populations from rose greenhouses in Korea. J. Asia-PaciÞc Entomol. 6: 91Ð96. Liu, Q. J., Y. J. Liu, Y. Yu, X. H. Zhou, and H. Ma. 2012a. The study on resistance and its mechanism of Tetranychus urticae to several common insecticides. Chinese J. Appl. Entomol. 49: 376 Ð381. Liu, Q. J., Y. J. Liu, X. H. Zhou, A. S. Zhang, L. L. Li, X. Y. Men, S. C. Zhang, and Y. Yu. 2012b. Differences in the sensitivities of the eggs of four Tetranychus urticae geographical strains to ten acaricides. Plant Prot. 38: 178 Ð180. Marcic, D. 2003. The effects of clofentezine on life-table parameters in two-spotted spider mite Tetranychus urticae. Exp. Appl. Acarol. 30: 249 Ð263. Nauen, R., N. Stumpf, A. Elbert, C. P. Zebitz, and W. Kraus. 2001. Acaricide toxicity and resistance in larvae of different strains of Tetranychus urticae and Panonychus ulmi (Acari: Tetranychidae). Pest Manage. Sci. 57: 253Ð261. Nauen, R., P. Bielza, I. Denholm, and K. Gorman. 2008. Age-speciÞc expression of resistance to a neonicotinoid insecticide in the whiteßy Bemisia tabaci. Pest Manage. Sci. 64: 1106 Ð1110. Ochiai, N., M. Mizuno, N. T. Miyake, M. Dekeyser, L. J. Canlas, and M. Takeda. 2007. Toxicity of bifenazate and its principal active metabolite, diazene, to Tetranychus urticae and Panonychus citri and their relative toxicity to the predaceous mites, Phytoseiulus persimilis and Neoseiulus californicus. Exp. Appl. Acarol. 43: 181Ð197. Rauch, N., and R. Nauen. 2002. Spirodiclofen resistance risk assessment in Tetranychus urticae (Acari: Tetranychidae): a biochemical approach. Pestic. Biochem. Physiol. 74: 91Ð101. Rhodes, E. M., and O. E. Liburd. 2006. Evaluation of predatory mites and Acramite for control of twospotted spider mites in strawberries in north central Florida. J. Econ. Entomol. 99: 1291Ð1298. Roh, H. S., E. G. Lim, and J. Kim. 2011. Acaricidal and oviposition deterring effects of santalol identiÞed in sandalwood oil against two-spotted spider mite, Tetranychus urticae Koch (Acari: Tetranychidae). J. Pest. Sci. 84: 495Ð 501. Sa´ enz-de-Cabezo´ n, F. J., I. Pe´rez-Moreno, F. G. Zalom, and V. Marco. 2006. Effects of lufenuron on Lobesia botrana (Lepidoptera: Tortricidae) egg, larval, and adult stages. J. Econ. Entomol. 99: 427Ð 431. Sato, M. E., M. Z. da Silva, A. Raga, K. G. Cangani, B. Veronez, and R. L. Nicastro. 2011. Spiromesifen toxicity to the spider mite Tetranychus urticae and selectivity to the predator Neoseiulus californicus. Phytoparasitica 39: 437Ð 445.
October 2014
TANG ET AL.: ACARICIDE RESISTANCE IN T. urticae
Stumpf, N., and R. Nauen. 2002. Biochemical Markers Linked to Abamectin Resistance in Tetranychus urticae (Acari: Tetranychidae). Pestic. Biochem. Physiol. 72: 111Ð121. Suh, E., S. H. Koh, J. H. Lee, K. I. Shin, and K. Cho. 2006. Evaluation of resistance pattern to fenpyroximate and pyridaben in Tetranychus urticae collected from greenhouses and apple orchards using lethal concentrationslope relationship. Exp. Appl. Acarol. 38: 151Ð165. Tang, X. F., S. L. Wang, Y. J. Zhang, Q. J. Wu, and W. Xie. 2014. Abamectin resistance of Tetranychus urticae and detection for resistance-related gene by PASA technique. Acta Phytophylacica Sin. 41: 67Ð73. Tirello, P., A. Pozzebon, S. Cassanelli, T. Van Leeuwen, and C. Duso. 2012. Resistance to acaricides in Italian strains of Tetranychus urticae: toxicological and enzymatic assays. Exp. Appl. Acarol. 57: 53Ð 64. Tsagkarakou, A., T. Van Leeuwen, J. Khajehali, A. Ilias, M. Grispou, M. S. Williamson, L. Tirry, and J. Vontas. 2009. IdentiÞcation of pyrethroid resistance associated mutations in the para sodium channel of the two-spotted spider mite Tetranychus urticae (Acari: Tetranychidae). Insect Mol. Biol. 18: 583Ð593. Uesugi, R., K. Goka, and M. H. Osakabe. 2002. Genetic basis of resistances to chlorfenapyr and etoxazole in the twospotted spider mite (Acari: Tetranychidae). J. Econ. Entomol. 95: 1267Ð1274. Ullah, M. S., and T. Gotoh. 2013. Laboratory-based toxicity of some acaricides to Tetranychus macfarlanei and Tetranychus truncatus (Acari: Tetranychidae). Int. J. Acarol. 39: 244 Ð251. Van Leeuwen, T., and L. Tirry. 2007. Esterase-mediated bifenthrin resistance in a multiresistant strain of the twospotted spider mite, Tetranychus urticae. Pest Manage. Sci. 63: 150 Ð156. Van Leeuwen, T., S. Van Pottelberge, and L. Tirry. 2005. Comparative acaricide susceptibility and detoxifying enzyme activities in Þeld-collected resistant and susceptible strains of Tetranychus urticae. Pest Manage. Sci. 61: 499 Ð 507. Van Leeuwen, T., S. Van Pottelberge, and L. Tirry. 2006a. Biochemical analysis of a chlorfenapyr-selected resistant
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strain of Tetranychus urticae Koch. Pest Manage. Sci. 62: 425Ð 433. Van Leeuwen, T., L. Tirry, and R. Nauen. 2006b. Complete maternal inheritance of bifenazate resistance in Tetranychus urticae Koch (Acari: Tetranychidae) and its implications in mode of action considerations. Insect Biochem. Mol. Biol. 36: 869 Ð 877. Van Leeuwen, T., S. Van Pottelberge, R. Nauen, and L. Tirry. 2007. Organophosphate insecticides and acaricides antagonise bifenazate toxicity through esterase inhibition in Tetranychus urticae. Pest Manage. Sci. 63: 1172Ð1177. Van Leeuwen, T., J. Vontas, A. Tsagkarakou, W. Dermauw, and L. Tirry. 2010. Acaricide resistance mechanisms in the two-spotted spider mite Tetranychus urticae and other important Acari: a review. Insect Biochem. Mol. Biol. 40: 563Ð572. Van Pottelberge, S., T. Van Leeuwen, R. Nauen, and L. Tirry. 2009a. Resistance mechanisms to mitochondrial electron transport inhibitors in a Þeld-collected strain of Tetranychus urticae Koch (Acari: Tetranychidae). Bull. Entomol. Res. 99: 23Ð31. Van Pottelberge, S., T. Van Leeuwen, J. Khajehali, and L. Tirry. 2009b. Genetic and biochemical analysis of a laboratory-selected spirodiclofen-resistant strain of Tetranychus urticae Koch (Acari: Tetranychidae). Pest Manage. Sci. 65: 358 Ð366. Vassiliou, V. A., and P. Kitsis. 2013. Acaricide resistance in Tetranychus urticae (Acari: Tetranychidae) populations from Cyprus. J. Econ. Entomol. 106: 1848 Ð1854. Yang, N. N., W. Xie, C. M. Jones, C. Bass, X. G. Jiao, X. Yang, B. M. Liu, R. M. Li, and Y. J. Zhang. 2013. Transcriptome proÞling of the whiteßy Bemisia tabaci reveals stagespeciÞc gene expression signatures for thiamethoxam resistance. Insect Mol. Biol. 22: 485Ð 496. Zhao, W. D., K. Y. Wang, X. Y. Jiang, and M. Q. Yi. 2001. The monitoring of resistance of Tetranychus urticae Koch to several insecticides. Chinese J. Pestic. Sci. 3: 86 Ð 88. Zhu, L., Z. J. Kang, S. J. Wei, Y. J. Gong, and B. C. Shi. 2013. Investigation of occurrence of Tetranychus urticae Koch in different areas of Beijing in 2012. North. Hortic. 4: 120 Ð123. Received 20 February 2014; accepted 19 July 2014.
