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Effect of emamectin benzoate undersemi-field and field conditions on keypredatory biological control agentsused in vegetable greenhousesFermin Amor a , Pilar Medina a , Paloma Bengochea a , MónicaCánovas b , Pedro Vega b , Rui Correia b , Federico García c ,Manuel Gómez c , Flor Budia a , Elisa Viñuela a & Juan AntonioLópez ba Unidad de Protección de Cultivos, Escuela Técnica Superior deIngenieros Agrónomos, Universidad Politécnica de Madrid, CiudadUniversitaria s/n, Madrid, Spainb Syngenta Agro S.A, Ribera del Loira, 8-10, 3°, Madrid, Spainc Syngenta Agro S.A. – Bioline, Jazminero, 1, Edificio Guay, Ofic.3, Aguadulce, 04720, Almería, Spain

Available online: 20 Dec 2011

To cite this article: Fermin Amor, Pilar Medina, Paloma Bengochea, Mónica Cánovas, Pedro Vega,Rui Correia, Federico García, Manuel Gómez, Flor Budia, Elisa Viñuela & Juan Antonio López (2012):Effect of emamectin benzoate under semi-field and field conditions on key predatory biologicalcontrol agents used in vegetable greenhouses, Biocontrol Science and Technology, 22:2, 219-232

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RESEARCH ARTICLE

Effect of emamectin benzoate under semi-field and field conditions onkey predatory biological control agents used in vegetable greenhouses

Fermin Amora, Pilar Medinaa*, Paloma Bengocheaa, Monica Canovasb,

Pedro Vegab, Rui Correiab, Federico Garcıac, Manuel Gomezc, Flor Budiaa,

Elisa Vinuelaa and Juan Antonio Lopezb

aUnidad de Proteccion de Cultivos, Escuela Tecnica Superior de Ingenieros Agronomos,Universidad Politecnica de Madrid, Ciudad Universitaria s/n, Madrid, Spain; bSyngenta AgroS.A., Ribera del Loira, 8-10, 38, Madrid, Spain; cSyngenta Agro S.A. � Bioline, Jazminero, 1,

Edificio Guay, Ofic. 3, 04720 Aguadulce, Almerıa, Spain

(Received 11 October 2011; final version received 13 December 2011)

Predatory arthropods are commonly used as biological control agents (BCAs).They are released in commercial vegetable greenhouses as primary elements ofintegrated pest management programmes for some of the most devastating pestson pepper and tomato in southeastern Spain. Emamectin benzoate, a macro-cyclic lactone insecticide derived from the avermectin family of natural products,is being developed for the control of Lepidoptera pests on a variety of crops inEurope including vegetables. The compatibility of emamectin benzoate with thepredatory BCAs Amblyseius swirskii Athias-Henriot and Orius laevigatus (Fieber)in field trials (direct spray and aged residues) and Macrolophus pygmaeus(Rambur) and Chrysoperla carnea (Stephens) in semi-field studies was studied.Emamectin benzoate at the highest recommended concentration (14.25 mg L�1)was compatible with A. swirskii and O. laevigatus when applied 3 days beforethe introduction of the arthropods, but it was toxic when directly sprayed.M. pygmaeus and C. carnea adults survived to direct spray applications.

Keywords: emamectin benzoate; Amblyseius swirskii; Orius laevigatus;Macrolophus pygmaeus; Chrysoperla carnea; greenhouse crops

Introduction

Spain is the most important producer and exporter of fresh greenhouse vegetables in

Europe with approximately 59,000 ha dedicated to these crops. The main production

area is located in the southeastern (SE) regions of Almeria and Murcia, where

peppers (Capsicum annuum L.) and tomatoes (Solanum lycopersicum L.) under

greenhouses are the major crops [Ministerio de Medio Ambiente, Rural y Marino

(MARM) 2010a]. Currently, the most devastating pests affecting these two crops are

the western flower thrips Frankliniella occidentalis (Pergande) (Thysanoptera:

Thripidae), the tobacco whitefly Bemisia tabaci (Gennadius) (Homoptera: Aleyr-

odidae) and the tomato borer Tuta absoluta (Meyrick) (Lepidoptera: Gelechiidae),

which were recently introduced (Urbaneja, Mouton, and Molla 2008). These pests

are commonly managed through integrated pest management (IPM) programmes

primarily based on the release of commercial arthropods as biological control agents

*Corresponding author. Email: pilar.medina@upm.es

Biocontrol Science and Technology,

Vol. 22, No. 2, February 2012, 219�232

ISSN 0958-3157 print/ISSN 1360-0478 online

# 2012 Taylor & Francis

http://dx.doi.org/10.1080/09583157.2011.650152

http://www.tandfonline.com

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(BCAs). These include several major predatory organisms, such as the phytoseiid

Amblyseius swirskii Athias-Henriot and the hemipterans Orius laevigatus (Fieber),

Macrolophus pygmaeus (Rambur) [formerly cited and commercialised as

Macrolophus caliginosus (Wagner) (�Macrolophus melanotoma (Costa)) according

to Martınez-Cascales, Cenis, Cassis, and Sanchez (2006)] and Nesidiocoris tenuis

(Reuter). The phytoseiid mite is particularly useful against the whitefly B. tabaci and

can also feed on first instar thrips. O. laevigatus controls F. occidentalis and the othertrue bugs are being studied to regulate T. absoluta populations (Gabarra, Zapata,

Castane, Riudavets, and Arno 2006; Gabarra, Arno, and Riudavets 2008; Van der

Blom 2008; Van der Blom, Robledo, Torres, Sanchez, and Contreras 2008). The

green lacewing Chrysoperla carnea (Stephens) (Neuroptera: Chrysopidae) is also

considered a key generalist BCA in vegetables greenhouses (Gabarra et al. 2008; Van

der Blom 2008). However, because effective BCAs are not available for controlling all

pests, such as the noctuid caterpillars Helicoverpa armigera (Hubner) and Spodoptera

exigua (Hubner) and because they are not always able to maintain populations of

foliar piercing-sucking arthropods under economic thresholds, pesticides remain an

important management tool in greenhouse IPM to control key pests.

Emamectin benzoate (4ƒ-deoxy-4 ƒ-methylamino-4 ƒ-epiavermectin B1 benzoate)

is a new macro-cyclic lactone insecticide derived from the avermectin family of

natural products. This product has been developed for the control of Lepidoptera

pests on a variety of vegetable crops worldwide (Liguori, Cestari, Serrati, and

Fusarini 2008), such as T. absoluta, H. armigera and S. exigua (Liguori et al. 2010;Lopez et al. 2010). The compound is not systemic, but it exhibits translaminar

activity (Willis and McDowell 1989). Its main physiological mode of action is to

stimulate the release of the neurotransmitter g-aminobutyric acid (GABA), thus

causing a continuous flow of chloride ions into muscle cells resulting in a suppression

of contraction and paralysis (Ishaaya, Kontsedalov, and Horowitz 2002). Emamectin

benzoate acts via ingestion and, to a lesser extent, by contact (Dybas and Babu

1988). It is rapidly taken up by plants and metabolised on the surface by photo-

oxidation to non-toxic levels which favours its selectivity for biological control

organisms (Ishaaya et al. 2002). This insecticide is currently being evaluated for

inclusion in Annex I of the European directive 91/414/EEC which regulates the

Registration of Plant Protection Products and was recently approved for its use on

vegetables in Spain (MARM 2010b).

Knowledge of pesticide selectivity to BCAs is necessary for a successful

implementation of biological and chemical control methods in IPM programmes.

Fresh residues of emamectin benzoate have been shown to be harmful to a related

species of Orius, Orius insidiosus, in laboratory and extended laboratory tests(Studebaker and Kring 2003a,Studebaker and Kring 2003b). Under field conditions,

aged residues were found to be harmless to other phytoseiids, Euseius alatus DeLeon

and Euseius citrifolius Denmark and Muma in citrus (Reis, Neto, Franco, and

Teodoro 2004). Thus, for A. swirskii and O. laevigatus, field trials have been

implemented because they are the most important BCAs used widely in commercial

pepper greenhouses in SE Spain. Given the preliminary information on these two

BCAs, the effects of direct spray on established populations and of aged residues on

inoculative introductions have been studied to determine whether the product is

compatible with existing BCA populations. Alternatively, in case of negative effects

of direct spraying on BCAs, the optimal re-entry interval must be determined. For

220 F. Amor et al.

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C. carnea and M. pygmaeus, less used in SE Spain in comparison with the former

BCAs, semi-field assays have been performed to evaluate the level of compatibility of

fresh residues of the insecticide.

Therefore, the objective of this study is to determine the level of selectivity of the

insecticide emamectin benzoate for predatory BCAs commonly used in greenhouse

vegetable production in SE Spain.

Materials and methods

Field trials

In field tests, the compounds were tested under typical conditions in a commercial

crop. European Plant Protection Organization (EPPO) guideline PP1/151(2), ‘Side-

effects on Phytoseiulus persimilis’ (EPPO 2004) was considered for the general

aspects in the establishment of these trials.

To evaluate BCA population dynamics under commercial field conditions, four

trials were conducted during 2007 in Torre Pacheco, Murcia, Spain, under multiple

tunnel greenhouses of three extruded layer Indasol† (Solplast, Murcia) plastic at ex

CIFACITA S.L. facilities. Greenhouses were automatically acclimatised with Aco†

Vision Clima 727 equipment (Hortimax Growing Solutions S.L., Almeria, Spain).

Two trials on A. swirskii (direct spray and aged residues) and two trials on

O. laevigatus (direct spray and aged residues) were carried out. In all the trials, a fully

randomised design with 3 replicates and 160 m2 (272 plants) plots were used. To

avoid arthropod cross-contamination and spray drifts, plots were isolated both

laterally and on the top, with an anti-thrips net 10�14 thread cm�2 (Botanica

equipment S.L., Alcantarilla, Murcia) sealed between the sides (anchored to the

ground) and the top, resulting in cages of 8 m width, 20 m length and 3.5 m height,

separated and surrounded by 1 m corridors. The entrance to the plots consisted of

overlaps in the lateral nets (1.5 m overlap) closed with several wire loops.

Biological control arthropods were supplied by Syngenta Agro S.A. Bioline

(Aguadulce, Almeria) and released following the methodology provided by the

supplier. In the aged residue trials, the following use rates were employed: A. swirskii

(Swirskiiline† as in twin Gemini† sachets) at 250 mobiles per plant and O. laevigatus

(Oriline† activ) at 3.6 mobiles per plant. These release rates are considerably higher

than those commercially recommended (see below for the direct spray trials) to

ensure a faster development of the arthropod populations. In these trials, the

inoculative BCA introductions were performed 3, 7 and 14 days after insecticide

application (aged residues of 3, 7 and 14 days). On each plant, A. swirskii mites were

introduced by hanging a twin sachet (containing 250 mobiles) on a leaf petiole

approximately 20�25 cm from the ground forming an inverted V with the sides

containing the emergence holes inwards. For the inoculation of O. laevigatus, 2

bottles of 500 mobiles each were uniformly distributed over each plot, by tapping the

bottle and releasing about the same amount of the predators on the growing area of

every two plants. In the direct spray tests, populations of both BCAs, A. swirskii and

O. laevigatus, were already well established at the time of insecticide application

(Table 1) and came from inoculative releases performed on 13 April 2007 (31 days

before application) at commercially recommended rates, A. swirskii (Swirskiiline†

as) at 100 mobiles m�2 and O. laevigatus (Oriline† activ) at 3 mobiles m�2 and

Biocontrol Science and Technology 221

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Table 1. Average population density of Amblyseius swirskii and Orius laevigatus (number of mobile forms/plant) in the direct spray treatment of

emamectin benzoate over the different sampling dates. Field studies on greenhouse pepper.

Mobile forms per plant (9SE)

Treatment

Concentration

(mg a.i. L�1)

0 DBAA,C

(14 May 2007)

3 DAAA,B

(17 May 2007)

7 DAAA,C

(21 May 2007)

14 DAAA,B

(28 May 07)

21 DAAA,B

(4 June 2007)

Amblyseius swirskii

Control 12.83 (91.30)a 11.00 (91.80)a 9.17 (91.86)a 14.00 (91.00)a 14.33 (91.92)ab

Emamectin

benzoate

14.25 11.50 (91.61)a 2.83 (90.88)b 4.67 (90.73)a 1.50 (90.29)b 5.67 (91.69)a

Thiamethoxam 100.00 14.00 (93.25)a 4.67 (90.93)b 11.50 (92.29)a 11.00 (91.76)a 23.17 (95.43)b

Orius laevigatus

Control 2.02 (90.34)a 2.20 (90.29)a 1.22 (90.21)a 1.92 (90.17)a 2.30 (90.48)a

Emamectin

benzoate

14.25 1.93 (90.42)a 0.10 (90.03)b 0.08 (90.08)b 0b 0.05 (90.03)b

Thiamethoxam 100.00 2.23 (90.14)a 0.02 (90.02)b 0.07 (90.07)b 0b 0b

Note: DBA, days before application; DAA, days after application. Three replicates of 20 plants per replicate.AMeans within columns followed by the same letter are not significantly different (ANOVA, LSD; a�0.05) for A. swirskii.BMeans within columns followed by the same letter are not significantly different (KW, notched boxplot; a�0.05) for O. laevigatus.CMeans within columns followed by the same letter are not significantly different (ANOVA, LSD; a�0.05) for O. laevigatus.

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following the methodology described earlier for the aged residue trials. In both cases,

the establishment and viability of the BCAs was ensured by maintaining an adequate

presence of their prey F. occidentalis from a natural infestation, although these

polyphagous predators can subsist feeding on other pests as well as on plant pollen.

Trials were conducted on pepper (C. annuum L.) var. Coyote in the direct spray

trials and var. Bilbo in the aged residue trials. Crops were grown on artificial media(coconut fibre Hidrosac†, Poliexmur, S.A., San Gines, Murcia) and arrayed at 1.5 m

row distance�0.4 m plant distance (1.7 plants m�2). Dates (planting and last

evaluation) and environmental conditions (Ta daily averages and relative humidity

[RH]) of the trials were as follows: direct spray trials (16 January 2007�14 June 2007):

14�25 8C, 67�89% RH; and aged residue trials (2 June 2007�24 October 2007): 19�30 8C, 70�81% RH.

The formulated insecticide emamectin benzoate 95 g kg�1 SG (Affirm†,

Syngenta Agro S.A., Madrid) was tested as an experimental product at the maximum

field-recommended concentration of the active ingredient, 14.25 mg L�1. Two other

insecticides were included in the study as references: thiamethoxam 400 g kg�1 WG

(Actara†, Syngenta Agro S.A., Madrid) in A. swirskii and O. laevigatus direct sprays

and lambda-cyhalothrin (100 g L�1 SC, Karate† Zeon†, Syngenta Agro S.A.,

Madrid) in aged residue trials. Thiamethoxam was chosen because neonicotinoids

have been shown to be harmful to many BCAs, in particular, to O. laevigatus

(Delbeke, Vercruysse, Tirry, De Clercq, and Degheele 1997; Van de Veire et al. 2002;Sterk, Heuts, Merck, and Bock 2003) as has imidacloprid (Angeli, Baldessari,

Maines, and Duso 2005). The use of separate references in the two trials is because

thiamethoxam was harmless to A. swirskiii in the direct spray trial, while pyrethroids

are generally known to be harmful to predatory mites (Aliniazee and Cranham 1980;

Bostanian and Belanger 1985; Bostanian and Racette 1997). Reference products were

applied at the recommended concentrations of 100 mg L�1 and 20 mg L�1 of active

ingredient, respectively (thiamethoxam and lambda-cyhalothrin). Insecticides were

applied as foliar spray until run off with a motorised knapsack sprayer (MS068,

Maruyama) and double cone nozzle gun (1.5 mm diameter), using a spray volume of

600 L ha�1 in the direct spray assay and 1000 L ha�1 in the aged residues assay. The

differences in volume are justified by the different growth stages, 61 and 75 BBCH,

respectively (Meier 2001), and size of the plants at application (50�60 and 80�90 cm

high, respectively). Output pressure was always 8 atm. The control plots were

sprayed with tap water under the same earlier conditions. In the direct spray trials,

application was performed on 14 May 2007, once the level of leaf colonisation by the

arthropods reached more than 75% (according to an evaluation made at 7 days

before application, on the 10 upper leaves, flowers included, of 20 plants per plotselected at random in the middle of the plots). In the aged residue trials, the products

were sprayed on 11 September, 7 September and 31 August 2007, which correspond

to 3, 7 and 14 days before the introduction of the arthropods (14 September 2007).

In all the trials, assessments were made on the 10 upper leaves, flowers included,

of 20 plants per plot selected at random in the middle of the plots. Leaves were

collected in plastic bags, kept in an ice-chest and transported to the laboratory.

Arthropods were identified and counted under a stereoscopic microscope such that

leaves and flowers were evaluated individually and the remaining arthropods in the

plastic bags were collected with a brush into a Petri dish, identified and reported as

number of mobile forms (nymphs and adults) per plant. Sampling in the direct spray

Biocontrol Science and Technology 223

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trials was performed at 0 days before application, and 3, 7, 14 and 21 days after

application. The first evaluation after treatment was made at 3 days after application

taking into consideration that, according to previous studies, several days are needed

for the insecticide to kill target insects (Ishaaya et al. 2002; Ishaaya, Barazani,Kontsedalov, and Horowitz 2007), so differences among earlier treatments would not

be expected. The rest of the evaluations were made at 7-day intervals after

application. In the aged residue trials, samplings were taken at 0 Days Before

BCA Introduction (DBI), and at 7, 17, 22, 29, 36 and 43 Days After Introduction

(DAI). The first evaluation after the introduction of the BCAs was made at 7 DAI to

allow populations to settle down uniformly across the untreated plots. Evaluations

were made following at approximately 7-day intervals after introduction.

Semi-field tests

The methodology used in these tests was based on the semi-field test described for

O. laevigatus (Van de Veire et al. 2002), and modified to the different species and

conditions of the plants and facilities.

To evaluate the acute toxic effects of the direct spray application of emamectin

benzoate on adults of M. pygmaeus and C. carnea, two semi-field trials were

conducted on tomato plants in an experimental greenhouse equipped with coolingand heating systems under controlled environmental conditions (2595 8C; 4095%

RH) during May�July 2009 at the facilities of Escuela Tecnica Superior de

Ingenieros Agronomos (ETSIA), Universidad Politecnica de Madrid (UPM),

Madrid, Spain. A completely randomised design with 10 replicates (1 plant per

plot) was used.

Macrolophus pygmaeus (Macroline†) was supplied by Syngenta Agro S.A.

Bioline (Aguadulce, Almeria). C. carnea came from the colony reared at the Unidad

de Proteccion de Cultivos, ETSIA, UPM, Madrid. To ensure adequate quality, onthe day of the reception, M. pygmaeus adults were selected from the commercial

package and kept in groups of 30 (in plastic ventilated round cages with Ephestia

kuehniella Zeller and Artemia salina Leach eggs as ad libitum food) for 24 h, and

unhealthy stocks were discarded. C. carnea adults were selected 24 h after emergence.

In each of the replicates (1 plant), 15 individuals were used. In the M. pygmaeus

trials, a mix of E. kuehniella and A. salina eggs was supplied after the insecticide

application as ad libitum food in a small container hung on each plant. C. carnea was

provided with an artificial diet (Vogt et al. 2000).Bioassays were conducted on tomato (S. lycopersicum L.) var. Montfavet† 63/5

HF1 (Vilmorin Iberica S.A., Alicante), seeded in tray cells with sterilised peat

substrate (pH 6.5) and transplanted into individual pots (22 cm diameter) 6 weeks

later to obtain homogeneous pesticide-free plants. Each plant (ca. 20 cm height) was

surrounded by a plastic cylinder (21 cm diameter, 30 cm height) sealed to the surface

of the pot with foam and covered with a fine cloth mesh in the upper part to allow

ventilation. Cylinder cages had two lateral windows (15�15 cm2) covered with the

mesh.The formulated insecticide emamectin benzoate 95 g kg�1 SG (Affirm†,

Syngenta Agro S.A., Madrid) was tested as experimental product at the highest

recommended concentration of the active ingredient, 14.25 mg L�1. Imidacloprid at

200 g L�1 SC (Confidor†, Bayer CropSciences S.A., Valencia) was used, as a

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reference product, at the recommended concentration of 150 mg L�1 of active

ingredient, based on its harmfulness to the BCAs, M. pygmaeus (Tedeschi, Tirry, Van

de Veire, and de Clercq 2002; Van de Veire and Tirry 2003) and C. carnea (Huerta,

Medina, Smagghe, Castanera, and Vinuela 2003; Nasreen, Mustafa, and Ashafaq

2005).

Applications were made 1 h after the insect introduction on the tomato plants

until run off with a 1 L hand sprayer. Control plants were treated with distilled water

as described earlier.

Insect mortality (%) was assessed at 24, 48 and 72 h after treatment. Mortality in

the control treatments was 0% for C. carnea and 21% for M. pygmaeus, below the

25% recommended by the standards of the IOBC (Hassan 1985).

Data analysis

Data were subjected to an analysis of variance using Statgraphics† Plus v. 5.0 (STSC

1987), and differences among the means were determined with a LSD test performed

at a significance level of a�0.05. A non-parametric Kruskal�Wallis (KW) test and a

median discrimination by the notched boxplot were conducted at a�0.05 when the

normal distribution (Kolmogorov test) and/or homoscedasticity (Bartlett test)

hypothesis were not confirmed. Corrected data were calculated (Abbott 1925) and

according to the IOBC scheme (Sterk et al. 1999), the insecticides were classified into

one of the following categories: 1, harmless (B25%); 2, slightly harmful (25�50%); 3,

moderately harmful (51�75%) or 4, harmful (�75%).

Results

Amblyseius swirskii

The analysis of the number of mobile forms per plant in the direct spray trial at 0

days before application revealed that there were no significant differences (F�0.32;

df �2,6; P�0.741) in populations among treatments. Compared with the control

treatment, the reference product thiamethoxam caused a significant reduction in the

population only at 3 days after application (moderately harmful) (F�10.35;

df �2,6; P�0.031) whereas emamectin benzoate caused significant reduction at 3

days after application (moderately harmful), and at 14 days after application

(harmful) (F�30.66; df �2,6; PB0.001). No statistical differences were shown at 7

days after application (F�3.92; df �2,6; P�0.081), and neither differed from the

control plots at 21 days after application; however, statistical differences were shown

between both compounds (F�1.75; df �2,6; PB0.001), as emamectin benzoate was

significantly more toxic than thiamethoxam in this evaluation (Table 1).

In the aged residue trial, no significant effect was seen among treatments 7 days

after the arthropod introduction (F�2.87; df �4,10; P�0.080). Lambda-cyhalo-

thrin 3 days residue caused significant population reduction from 17 DAI (harmful)

(F�5.22; df �4,10; P�0.016) until the last evaluation, 43 DAI (F�6.59; df �4,10;

P�0.006). None of the emamectin benzoate residues (3, 7 and 14) differed

significantly from the control plots in any of the assessments throughout the

experiment (7, 17, 22, 29, 36 and 43 DAI) (Table 2).

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Table 2. Average population density of Amblyseius swirskii and Orius laevigatus (number of mobile forms/plant) over the different sampling dates when

the natural enemies were exposed to aged residues (3, 7 and 14 days old) of emamectin benzoate. Field studies on greenhouse pepper.

Mobile forms per plant (9SE)

TreatmentConcentration(mg a.i. L�1)

0 DBI (14September

2007)

7 DAIA,C (21September

2007)17 DAIA,B (1October 2007)

22 DAIA,C (6October 2007)

29 DAIA,C (13October 07)

36 DAIA,C (20October 2007)

43 DAIA,C (27October 2007)

Amblyseius swirskii

Control 0 4.98 (91.98)a 28.67 (99.69)a 26.33 (912.87)ab 28.67 (98.38)a 42.33 (98.35)a 18.67 (93.84)aEmamectin benzoate 3 days (AD:

11 September 2007)14.25 0 4.25 (91.03)a 26.00 (92.56)a 17.25 (91.98)b 28.83 (95.87)a 28.17 (92.09)a 12.42 (90.74)a

Emamectin benzoate 7 days(AD:07 September 2007)

14.25 0 3.10 (90.73)a 28.00 (94.39)a 43.50 (93.97)a 13.67 (92.67)ab 31.92 (96.42)a 14.70 (92.06)a

Emamectin benzoate 14 days (AD:31 August 2007)

14.25 0 2.28 (91.03)a 36.33 (95.02)a 22.50 (95.01)ab 22.83 (91.59)a 29.17 (94.06)a 11.83 (91.01)a

Lambda-cyhalothrin 3 days (AD:11/09/07)

20.00 0 0.37 (90.07)a 3.00 (91.26)b 2.92 (90.58)c 5.67 (90.73)b 7.17 (90.93)b 4.17 (90.88)b

Orius laevigatus

Control 0 0.08 (90.08)a 2.17 (90.17)a 2.92 (90.65)ab 4.00 (90.52)a 2.92 (90.30)b 2.58 (90.51)bEmamectin benzoate 3 days (AD:

11 September 07)14.25 0 0.08 (90.08)a 1.83 (90.08)a 3.33 (90.60)a 2.50 (90.38)a 2.33 (90.27)b 1.92 (90.17)bc

Emamectin benzoate 7 days(AD:07 September 2007)

14.25 0 0.08 (90.08)a 2.17 (90.17)a 1.58 (90.36)b 3.25 (90.63)a 4.42 (90.30)a 3.08 (90.22)a

Emamectin benzoate 14 days (AD:31 August 2007)

14.25 0 0.33 (90.08)a 1.92 (90.16)a 3.17 (90.44)a 3.42 (90.58)a 3.92 (90.22)a 3.00 (90.25)a

Lambda-cyhalothrin 3 days (AD:11 September 07)

20.00 0 0.02 (90.02)a 0.08 (90.02)b 0.08 (90.08)c 0.09 (90.08)b 0.05 (90.05)c 0.13 (90.06)c

Note: AD, application date; DBI, dates before insects introduction; DAI, days after insects introduction. Three replicates of 20 plants per replicate.AMeans within columns followed by the same letter are not significantly different (ANOVA, LSD; a�0.05) for A. swirskii.BMeans within columns followed by the same letter are not significantly different (KW, notched boxplot; a�0.05) for O. laevigatus.CMeans within columns followed by the same letter are not significantly different (ANOVA, LSD; a�0.05) for O. laevigatus.

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Orius laevigatus

In the direct spray trial, there were no significant differences (F�0.23; df �2,6;

P�0.798) in populations among treatments at 0 days before application. Compared

with the control treatment, both emamectin benzoate and the reference product,

thiamethoxam, caused significant reduction of the population at 3 days after

application (KW �6.94; P�0.031), 7 days after application (F�23.70; df �2,6;

P�0.001), 14 days after application (KW �6.72; P�0.035) and 21 days afterapplication (KW �6.76; P�0.034), and both were classified both as harmful in all

the cases (Table 1).

In the aged residue trial, no significant effect was seen among treatments at 7

DAI (F�2.68; df �4,10; P�0.094). Lambda-cyhalothrin 3 days residue caused

significant population reduction (harmful) from 17 DAI (KW �9.78; P�0.044)

until the last evaluation, 43 DAI (F�18.47; df �4,10; PB0.001). None of the

emamectin benzoate residues (3, 7 and 14) differed significantly from the control

plots in assessments 7, 17, 22, 29 DAI. Numbers of O. laevigatus found were evensignificantly higher in comparison with the control in the evaluations 36 and 43 DAI

(Table 2).

Macrolophus pygmaeus

A direct spray application of emamectin benzoate at the highest recommended rate

(14.25 mg L�1) in the semi-field direct spray trial in tomato plants was notsignificantly different from the control at 24 and 48 h but showed significantly higher

mortality at 72 h (F�77.8; df �2,27; PB0.001) that was however classified as

harmless (16% corrected mortality). The reference product, the neonicotinoid

insecticide imidacloprid (150 mg L�1), produced significantly higher mortality at

24 h (F�336.79; df �2,27; PB0.001), 48 h (F�192.58; df �2,27; PB0.001) and

72 h (F�77.8; df �2,27; PB0.001) after application and was classified as harmful

in all cases (Table 3).

Chrysoperla carnea

The effect of emamectin benzoate application in tomato plants on C. carnea at the

highest recommended rate (14.25 mg L�1) in the semi-field direct spray trial was not

significantly different from the control at 24, 48 or 72 h after application. In contrast,

the reference insecticide, imidacloprid (150 mg L�1), produced a significantly highermortality at 24 h (58%) (KW �26.11; PB0.001), 48 h (67.33%) (KW �26.08;

PB0.001) and 72 h (69.33%) (KW �26.07; PB0.001) after application and was

classified as moderately harmful in all cases (Table 3).

Discussion

Emamectin benzoate is less toxic to a variety of beneficial organisms than it is to its

main targets, according to our results.

Amblyseius swirskii is a relatively new BCA and few data exist on its susceptibility

to pesticides, and no studies assessing the effects of emamectin benzoate were found.

In our study, conducted in a commercial greenhouse, emamectin benzoate was

Biocontrol Science and Technology 227

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harmful to A. swirskii on peppers up to 14 days after application when sprayed

directly on an established population. Previous studies on the predatory mite Euseius

victoriensis Womersley (Acari: Phytoseiidae) using a ‘worst case scenario’ direct

overspray assays showed that emamectin benzoate is highly toxic after a single-use

application at the highest recommended rate, 1000 L ha�1 (Bernard et al. 2010),

suggesting that it may not be completely safe for phytoseiids as demonstrated in our

test field. Abamectin, the first avermectin of commercial interest, is known for its

unprecedented potency against a broad spectrum of phytophagous mites, members

of Tetranychidae, Eriophyidae and Tarsonemidae. It is markedly less toxic to other

phytophagous mites, such as Panonychus ulmi (Koch), and its toxicity to insects is

more variable (Jansson and Dybas 1998). In contrast, emamectin benzoate is highly

potent against a broad spectrum of Lepidoptera, but not comparable to abamectin as

acaricide. The toxicity of emamectin benzoate on mites depends more on the species,

and field studies in citrus carried out showed no effect of fresh emamectin benzoate

residues (20 g ha�1) on E. alatus and E. citrifolius (Reis et al. 2004).

In aged residue trials, 3, 7 and 14 day residues of the insecticide were harmless to

the predators as it has been previously demonstrated for foliar residues of emamectin

benzoate. Within 1 day of application, and often within a few hours after application,

the residues were only slightly toxic (B20% mortality) to most beneficial insects,

including the honey bee, Apis mellifera L., and several predators and parasitoids

(Jansson and Dybas 1998). In the related insecticide, abamectin, 1 day residues (0.108

mg L�1) were harmless to A. swirskii in a field test on greenhouse cucumber showing

effects similar to the aged emamectin benzoate residues in our trial (Gradish, Scott-

Dupree, Shipp, Harrisa, and Ferguson 2011). On O. laevigatus, emamectin benzoate

is harmful up to 21 days after application in direct spray applications but 3, 7 and 14

days residue exposure is harmless. For another Orius species, O. insidiosus, it was

found in laboratory and extended laboratory tests that fresh residues of emamectin

benzoate, as well as abamectin, were moderately harmful, to harmful for nymphs and

Table 3. Average mortality of Macrolophus pygmaeus and Chrysoperla carnea adults after

direct spray of emamectin benzoate at 24, 48 and 72 h after application. Semi-field studies on

tomato plants.

Average mortality (%) (9SE)

Treatment

Concentration

(mg a.i. L�1) 24 h 48 h 72 h

Macrolophus pygmaeus

Control 8.0091.33a 16.6692.85a 21.3393.11a

Emamectin benzoate 14.25 10.6692.47a 18.6692.59a 34.0095.20b

Imidacloprid 150.00 86.6693.14b 86.6693.14b 86.6693.14c

Chrysoperla carnea

Control 0a 0a 0a

Emamectin benzoate 14.25 0.6790.67a 0.6790.67a 0.6790.67a

Imidacloprid 150.00 58.0093.72b 67.3393.77b 69.3394.12b

Note: Means within columns followed by the same letter are not significantly different (ANOVA, LSD;a�0.05 for M. pygmaeus and KW, notched boxplot, a�0.05 for C. carnea).

228 F. Amor et al.

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adults of the predator (Studebaker and Kring 2003a,Studebaker and Kring 2003b).

These results confirm our data from the direct spray trial and may be in accordance

with our aged residue results, considering that, unlike in a laboratory tests,

emamectin benzoate trial may be degraded on the leaf surface enough to prevent

any toxic effects after 3 days due to its fast degradation under field conditions by

photodegradation (Willis and McDowell 1989; Prabhu, Wehner, Egan, and Tway

1991; Wrzesinski et al. 1996).

Collectively, our field data demonstrated that emamectin benzoate provides, if

not physiological selectivity because of its direct toxicity, at least, ecological

selectivity for A. swirskii and O. laevigatus. This ecological selectivity is related to

the short half-life of the compound on foliage, and within an IPM programme, this

pesticide can be used 3 days before inoculative release of the natural enemies tested in

this study.

Under semi-field conditions, emamectin benzoate is harmless at the highest

recommended rate of 14.25 mg L�1 when sprayed directly onto M. pygmaeus adults

in tomato plants. Similarly, it was found under laboratory test conditions that

exposure to fresh residues of emamectin benzoate is harmless to nymphs of this

hemipteran predator (Tedeschi et al. 2002; Van de Veire and Tirry 2003). Previous

results obtained by the authors in residue bioassays under semi-field conditions

confirmed also the harmless effect on M. pygmaeus adults when exposed to fresh and

1 day residues of the insecticide (Lopez et al. 2011). Other naturally derived products,

abamectin and spinosad, are also harmless to nymphs after fresh residue exposure

(Van de Veire and Tirry 2003). On C. carnea, emamectin benzoate (14.25 mg L�1)

has been shown to be harmless in direct spray tests under semi-field conditions in

tomato plants up to 72 h. The harmless effects of two direct sprays of emamectin

benzoate at 13.5 g ha�1 on C. carnea larvae were observed in cotton under field

conditions (Sechser, Ayoub, and Monuir 2003). In the related species

C. externa (Hagen), no negative effects of topical or direct spray applications with

emamectin benzoate were found in laboratory or extended laboratory tests on L1, L2

and L3 larvae (Ferreira, Carvalho, Botton, and Lasmar 2006). Similarly, another

avermectin, abamectin, showed also no negative effects on adults when exposed to

fresh residues and was slightly harmful to L1 larvae (Giolo, Medina, Gruetzmacher,

and Vinuela 2009).

In conclusion, emamectin benzoate is compatible with A. swirskii and

O. laevigatus when applied 3 days before predator introduction, and with

M. pygmaeus and C. carnea in direct spray application. Taking into account these

considerations would complement biological control of the BCAs in pepper

greenhouses.

Acknowledgements

This work was partially supported by the Spanish Ministry of Education and Culture (projectAGL 2007-66399-C03-01 and AGL2010-22196-C02-02 to Elisa Vinuela). F. Amor andP. Bengochea acknowledge the Ministry of Education and Culture and UniversidadPolitecnica de Madrid (UPM), respectively, for doctoral fellowships.

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