Physiological selectivity of insecticides to adult of Doru luteipes

Doru luteipes (SCUDDER, 1876) is considered one of the best natural enemies of the fall armyworm Spodoptera frugiperda (J. E. Smith, 1797) (Lepidoptera: Noctuidae), feeding on their eggs and small caterpillars. For its conservation it is necessary to use selective insecticides to S. frugiperda and harmless to the predator. Therefore, objective of the present work was to evaluate the toxicity of insecticides registered to control of S. frugiperda. It was conducted bioassays with D. luteipes adults treated with insecticides directly by exposure to residues of compounds applied on glass plates and the consumption of eggs of S. frugiperda contaminated and offered each one, 24 and 48 hours after treatment. The insecticides were classified according to indices proposed by IOBC/WPRS. For adults D. luteipes treated directly with insecticides, triflumuron was harmless (class 1); chlorfenapyr and etofenprox as slightly harmful (class 2) teflubenzuron/ -cypermethrin and spinosad moderately harmful (class 3) and thiamethoxan/ -cyhalothrin harmful (class 4). In bioassay exposure of D. luteipes residues of insecticides applied to glass plates, all products were harmful to the predator, except triflumuron which was considered slightly harmful. The survival of adults after consumption of contaminated eggs was 46.7% for the insecticide tiametoxam/ -cialotrina considered slightly harmful to the predator. The other insecticides were innocuous. Due to the low toxicity presented by the triflumuron to the D. luteipes adults, this compound can be recommended in programs aimed at integrated pest management the preservation of this natural enemy. All other products must be evaluated in greenhouse and field to prove its toxicity.


INTRODUCTION
Among the several factors that affect maize (Zea mays L.) productivity, the fall armyworm Spodoptera frugiperda (J.E. SMITH, 1797) (Lepidoptera: Noctuidae) is considered the main pest of this crop in Brazil, and one of the most important in the Americas, which can cause losses of more than 50% (FIGUEIREDO; MARTINS-DIAS; CRUZ, 2006).The control of S. frugiperda in maize has been done mostly by using chemicals applied immediately after its detection in the field and generally causes biological unbalances and other negative environmental impacts (CRUZ; VIANA; WAQUIL, 2002).
In maize, natural enemies that act in the early stages of pest development, thus avoiding significant damages to plants, are the most important.This is the case, for example, of Doru luteipes (SCUDDER, 1876) (Dermaptera: Forficulidae).This predator has been considered of great potential as a biological control agent of maize pests, such as S. frugiperda, Helicoverpa zea (BODDIE, 1850) (Lepidoptera: Noctuidae) and aphids (REIS; OLIVEIRA; CRUZ, 1988;CRUZ;ALVARENGA;FIGUEIREDO, 1995).
The earwigs are considered voracious predators because they have high ability to attack and feed on different preys, particularly eggs and immature stages of insects of the orders Lepidoptera, Hemiptera, Coleoptera and Diptera.And the presence of D. luteipes in 80% of the maize plant is sufficient to maintain the fall armyworm under control, below the economic injury level (CRUZ; OLIVEIRA, 1997).Thus, is necessary to integrate the biological control of this predator and other control methods, as the chemical (PASINI; PARRA; LOPES, 2007).
The use of selective products minimizes exposure of natural enemies and at the same time, controls the pests species.It is necessary to integrate the biological control of this predator and other control methods, as the chemical.Since the use of chemicals must be used only when all the control alternatives had been extinguished and in an emergency way, by means of selective products (BOLLER et al., 2004).Reis, Oliveira and Cruz (1988), Faleiro et al. (1995) and Simões, Cruz and Salgado (1998) studied the selectivity of insecticides used in maize to control nymphs and adults of D. luteipes and concluded that the adult predator was more tolerant to the compounds than nymphs, mainly to the pyrethroid insecticides permethrin and deltamethrin.Zotti et al. (2010) observed that the survival of adults of Doru lineare (ESCHSCHOLTZ, 1822) (Dermaptera: Forficulidae) after ingestion eggs of S. frugiperda contaminated with insecticides was reduced to Certero ® with means of 50% and Engeo Pleno ® with 100% mortality.
In this context, the objective of this study was to evaluate the selectivity of six insecticides registered to the control of S. frugiperda in maize, in the higher doses recommended by manufacturers, for adults of the predator D. luteipes.
Eggs, nymphs and adults of D. luteipes were collected in an area with the 'BRS1031' maize, cultivated in an organic system and maintained in laboratory at 25 ± 2 °C, relative humidity 70 ± 10% and 12 hours photophase for creation and subsequent use in experiments.
For the creation of the predators in the laboratory couples of D. luteipes were placed in cages made from PVC pipe measuring 30 cm diameter and 50 cm high, which was capped at both ends with a 2 cm PVC ring closed with a nylon mesh 0.5 mm diameter.Inside each cage 10 central buds of maize (cartridges) were placed to be used as oviposition substrate and shelter for insects.The insects were fed ad libitum on unviable eggs of S. frugiperda and a diet based on cat food (CRUZ, 2009).
The compounds were diluted in 282 liters of water/ ha.After application of each product, pulverizer and spray nozzle were washed with water and then with acetone to remove residues of each compound.Insecticides were applied using a CO 2 pressurized sprayer with a fan type nozzle 80.03, adjusted to a pressure of 2.6 lb/pol 2 , coupled to a treadmill at constant speed of 6.2 km/h according to the methodology by Simões, Cruz e Salgado (1998).

Effects of insecticides directly applied on adults of D. luteipes
Couples were placed in Petri dishes with 18 cm diameter and 1.5 cm high and submitted to insecticides spraying in a treadmill, as previously mentioned.Each treated couple was placed in a plastic cup inserted on Styrofoam tray which was maintained in laboratory at 25 ± 2 °C, relative humidity 70 ± 10% and 12 hours photophase.The insects were fed ad libitum on unviable eggs of S. frugiperda.The number of dead insects was evaluated every 24 hours for 15 days after contamination of the adults with insecticides.Five replications with 24 insects were done for each treatment.

Effect of residual contact of insecticides on adults of D. luteipes
Insecticides applications were done in arenas comprised by capped Petri dishes measuring 18 cm diameter and 1.5 cm high under treadmill, which was maintained in laboratory at 25 ± 2 °C, relative humidity 70 ± 10% and 12 hours photophase.After elimination of the excess of moisture couples of the predator were released into the arenas, being in constant contact with the dry film of each insecticide throughout the evaluation period.The number of dead adults was recorded every 24 hours during the 15 days of exposition of the insects to the residues of compounds.Oviposition capacity and egg viability of surviving couples were assessed.The insects were fed ad libitum on unviable eggs of S. frugiperda.Each treatment consisted of five replications with six couples.

Effect of the ingestion of food contaminated by insecticides on adults of D. luteipes
Ovipositions of about 50 eggs of S. frugiperda 24 hours old were placed on Petri dishes covered with aluminum paper and maintained under refrigeration at 8 o C for 48 hours to make eggs unfeasible.After this period, each oviposition was fixed through an entomological pin on a styrofoam plate 36 cm long by 24 cm wide and subjected to the spraying of insecticides as earlier mentioned.After the applications, part of the S. frugiperda eggs unfeasible after one hour was offered to adults of the predator previously individualized in 50 mL plastic cups fixed in styrofoam trays.Other offerings were done at 24 and 48 hours after pulverization.
Survival of adults up to 15 days (360 hours) after the beginning of exposure to S. frugiperda eggs treated with insecticides was evaluated.Oviposition capacity and egg viability of surviving couples were assessed.Each treatment comprised five replications with six couples.
Data from these experiments were analyzed by one-way Analysis of Variance through the SISVAR (FERREIRA, 2007) and treatment means were compared with the Scott-Knott test (P = 0.05) (SCOTT; KNOTT, 1974); Burr-Foster Q and Shapiro-Wilk W tests were used to test equality of variance and normality of the data, following description found in Anderson and McLean (1974).

Effects of insecticides directly applied on adults of D. luteipes
For couples of D. luteipes directly treated with insecticides, triflumuron was the product responsible for lower mortality, causing the death of only 13.3% of males and 5% of females at 360 hours after start exposure (HASE), being considered harmless (Table 1).
The selectivity of triflumuron to the predator in the adult stage was possibly due to the mode of action, since belongs to the chemical group of benzoilureas, chitin synthesis inhibitors, which act mainly as ovicidal and larvicidal (OMOTO, 2000).The ovicidal and larvicidal actions were evaluated by Simões, Cruz e Salgado (1998), where eggs and nymphs of D. luteipes were treated through triflumuron pulverization (25 g a.i ha -1 ), and mortality of 78% of eggs and 69.6% of nymphs was observed.Tonet (1995) determined the impact of insecticides used on soybeans on D. lineare, triflumuron, the product was considered of low toxicity (mortality 21-40%) and was considered selective diflubenzuron (up to 20% mortality).
Mortality of males and females by thiamethoxan/cyhalothrin was 97.5% in the first 24 HASE, and 100% at 48 HASE proving to be harmful (class 4).Teflubenzuron/ -cypermethrin caused high mortality at 24 HASE, with 85% for males and 86.7% for females.At 360 HASE the means were 95% and 93.3%, respectively.Therefore, it was classified as moderately harmful (Table 1).
The high mortality observed in the present study at the first assessment for thiamethoxan/ -cyhalothrin and teflubenzuron/ -cypermethrin may have occurred because of the combination of two active ingredients with different mechanisms of action (neonicotinoid + pyrethroid and pyrethroid + benzoilureia, respectively).With different active ingredients and acting simultaneously on the same insect, the toxicity of the product can be significantly increased (RIGITANO; CARVALHO, 2001).
Chlorfenapyr caused mortality of about 15.8% for males and 16.7% for females in the first 24 HASE.At the next 48 HASE mortality increased to 31.7% and 24.2%, reaching 51.7% and 40% at 360 HASE for males and females, respectively, and was considered slightly harmful (class 2) (Table 1).
Few studies are known concerning the selectivity of chlorfenapyr to natural enemies, however Leite et al. (2010)  larvae and adults of the predator Cycloneda sanguinea (Linnaeus) (Coleoptera: Coccinellidae).Evaluations were accomplished at one, 12; 23 and 35 days after chlorfenapyr pulverization (240 SC -160 g a.i ha -1 ), and high toxicity was observed with 100% mortality of larvae and adults of this predator.This compound is an ATP synthesis inhibitor which can act by contact or ingestion, causing the uncoupling of oxidative phosphorylation reactions, so that mitochondria fails to produce ATP, stopping vital functions of the cell, causing insect death.Unlike neurotoxic insecticides, the action of chlorfenapyr is not immediate, and death occurs after the depletion of the energy reserves of the insect (OMOTO, 2000).
Etofenprox caused 37% mortality for males and 42.5% for females, reaching 57.5% and 53.3% for males and females, respectively, at 48 HASE, being classified as slightly harmful.Spinosad showed a gradual increase in mortality of D. luteipes, reaching 82.5% for males and 64.2 for females of the predator, being classified as moderately harmful to males and slightly harmful to females at 360 HASE (Table 1).
For insects directly treated through spraying of insecticides it was observed that 70.8% of females laid Decrease in number of ovipositions and viability of eggs and nymphs was also observed for chlorfenapyr, etofenproxi and spinosad, and after the treatment with spinosad only 45.8% of females laid eggs, which presented only 10.3% viability.Nymphs were also affected by chlorfenapyr; 70% survived the first instar and 63.3% reached the fourth instar.Etofenproxi presented the highest mortality throughout the development of nymphs, and 90% survived the first instar but only 50% reached the fourth instar.Due to the high toxicity of thiamethoxan/ -cyhalothrin and teflubenzuron/ -cypermethrin, no live insects enough to evaluate the sublethal effects were found (Table 2).
Negative effects on fecundity and fertility of some insect species caused by chemical compounds were also found by other authors.Ávila and Nakano (1999) evaluate the toxic effects of the growth regulator lufenurom (0.33 g a.i ha) on adult fecundity and egg viability of Diabrotica speciosa (Germar, 1824) (Coleoptera: Chrysomelidae) in the laboratory.The authors say the low number of eggs (177.5) as well as their low viability (19.8%) obtained from couples who have ingested the insecticide can be explained by trans ovarian transmission, affecting embryonic development and preventing larvae hatching.

Effect of residual contact of insecticides on adults of D. luteipes
Triflumurom did not cause effect on predators in the first 24 hours, presenting mortality of 33% for males and 26.7% for females at 48 HASE.From 72 HASE, mortality remained at 46% for female but increased from 46% to 53.3% for males at 360 HASE and even so was considered slightly harmful (Table 3).
Teflubenzuron/ -cypermethrin and thiamethoxan/ -cyhalothrin presented toxicity of 66.7% and 60% for males and 56.6% and 66.7% for females, respectively, at 24 HASE.At 48 HASE, 100% of male was killed, so the products were harmful (class 4) to the predator.For females, the same products caused 100% and 93.3% mortality, respectively, considered harmful.Chlorfenapyr caused 100% mortality for males at 24 HASE.For females, mortality was 60% at 24 HASE, 70% at 48 HASE and 100% at 72 HASE.Etofenprox caused low mortality rates for males and females (3.3% and 0%, respectively) at 24 HASE, however, caused the deaths of 100% of males at 48 HASE and females at 72 HASE, and for this reason was in class 4 (harmful).Spinosad caused 93.3% mortality of males at 360 HASE and 100% of females at 72 HASE, considered slightly harmful (Table 3).
In general, all products were more toxic to the predator when applied to a surface rather than when applied directly to adults.This is due to the fact that the tarsal contact method where it is estimated a lethal time to the insects absorb insecticide throughout the test period.Rajashekar, Rao and Shivanandappa ( 2012) demonstrated experimentally that some insecticides are most effective when in contact with the tarsi of insects.
The insecticidal activity of some molecules is lost by hydrolysis, and various sugars, which modifies the toxic response, demonstrating that insecticidal activity can occur through sites located in tarsi.
According to Maia, Busoli and Delabie (2001), the longer the period of exposure of insects to chemicals, the greater the probability of intoxication of the target organism.Another possible explanation for the high mortality rates observed in the present study is related to the behavior of the Dermaptera, since they have the characteristic of constant cleaning of the body using the mouth parts (LANGSTON;POWELL, 1975), which increases even more the chances of contamination by the chemicals tested.The lower rate of penetration of insecticides when applied directly to adult insects may be due to the chemical composition and the greater thickness of the exoskeleton.But when the insect is exposed to the product for a long period, small amounts will penetrate the insect's body and can be lethal slowly in 360 HASE (LEITE et al., 1998).
It was not possible to evaluate the sublethal effects of thiamethoxan/ -cyhalothrin, teflubenzuron/ -cypermethrin, chlorfenapyr, etofenprox and spinosad due to the high toxicity presented.However, triflumuron was slightly harmful but no ovipositing females were observed in the period of evaluation.

Effect of the ingestion of food contaminated by insecticides on adults of D. luteipes
For couples that received eggs as food immediately after the chemical treatment the survival rate was higher than 80% in all treatments thus classified as class 1 (harmless), except thiamethoxan/ -cyhalothrin with means of survival for males 36.7% and 43.4 for females being included in class 2 (slightly toxic) (Table 4).
These results corroborate those found by Simões, Cruz and Salgado (1998), in which adults of D. luteipes were fed eggs of S. frugiperda treated with the insecticides triflumuron, permethrin, diflubenzuron and -cyhalothrin, with means of survival 94; 97; 97 and 93%, respectively.Similar results to the presented in this study were also reported by Bacci et al. (2001), where the pyrethroid permethrin was highly selective for adults of D. luteipes.Michereff Filho et al. (2002) evaluating the effect of deltamethrin on adults of this predator observed that the compound was harmless.
For insects that ingested eggs of S. frugiperda 24 and 48 hours after treatment with insecticides, no death was caused by triflumurom, differently of the other treatments (Table 4).Chlorfenapyr, etofenprox and spinosad were harmless, with mortalities of 10%, 0% and 10% for males and 10.1, 0 and 26.7% for females, respectively.Thiamethoxan/ -cyhalothrin was slightly harmful, presenting mortality rate of 53.3% and teflubenzuron/ -cypermethrin was harmless with mortality lower than 30% for males and females of D. luteipes that ingested eggs of S. frugiperda 48 hours after treated with insecticides (Table 4).
Different result was obtained by Farias et al. (2006), evaluating the effects at field of thiamethoxan + -cyhalothrin (Enge Max -21.2 g a.i ha -1 ) on the predators Doru lineare (ESCHSCHOLTZ, 1822) (Dermaptera: Forficulidae) and Chrysoperla sp.(Neuroptera: Chrysopidae) in soybean.The authors concluded that the product reduced the population of these enemies in more than 90%.Cisneros et al. (2002) evaluated the effect of the insecticide spinosad on Doru taeniatum (DOHRN, 1847) (Dermaptera: Forficulidae) and observed that mortality of the predator soon after consume treated larvae of S. frugiperda was 17% and at 72 hours reached 72%.
The divergence of results regarding action of some compounds on insects may be due to bioecological factors intrinsic to each species and also to   HASSAN et al., 1985) the differences in concentrations of the commercial product used and even the characteristic of the eggs of S. frugiperda, such as permeability of corium and adjacent layers, what may or not facilitate the penetration of the products (MANZONI et al., 2007).
In general, the survival of adults of the predator after ingestion of eggs of S. frugiperda at one, 24 and 48 hours after treatment was not affected by triflumuron, chlorfenapyr, etofenprox and spinosad, which were selective and classified as harmless.Males that ingested eggs of S. frugiperda 24 hours after treated with thiamethoxan/ -cyhalothrin presented survival rate of only 21.7% and females that consumed eggs treated with teflubenzuron/ -cypermethrin showed survival of 73.3%.The high survival rates of adults of the predator observed in this study can be due to the action of insecticides in the body of insects, since the ingested product can undergo the action of several enzymes in the digestive system and Malpighi tubules, making the molecule non-toxic or less toxic to the body (YU, 2002).
Insecticide metabolism can be complex and occurs in all insects, regardless of insecticide resistance status, and is likely to involve a multistep pathway.For many insecticides, different metabolic breakdown products are possible.Although there is evidence of insecticide metabolism in other tissues there is building evidence that the key tissues for the metabolism of most compounds are the midgut, the Malpighian tubules and the fat body.Recent large-scale transcript sequencing projects and microarray studies identify a large number of detoxification genes expressed in these tissues (PERRY; BATTERHAM; DABERR, 2011).
For couples who ingested eggs of S. frugiperda one and 24 hours after treatment with triflumuron, chlorfenapyr and etofenprox the percentage of ovipositing females were 25, 25, 29 and 20, 25, 37%, respectively.Females that fed eggs of S. frugiperda 48 hours after treatment presented similar results to the females that fed eggs one hour after treatment.For treatments with thiamethoxan/ -cyhalothrin, teflubenzuron/ -cypermethrin and spinosad no ovipositing females were observed (Table 5).
Despite low mortality of D. luteipes after ingestion of treated eggs of S. frugiperda, it was observed that insecticides negatively affected oviposition and egg viability.Many eggs were dry and with dark color after two days, without formation of embryo.Among the evaluated compounds, only triflumuron allowed the birth of nymphs, with mean of 7.6% (Table 5).
The ovicide mode of action of lufenuron to Podisus nigrispinus (Heteroptera: Pentatomidae) was reported by Evangelista Júnior et al. (2002).These authors observed that the eggs after three to four days of oviposition were reddish due to embryonic development, but being interrupted by the insecticide they become dark and withered with the death of the embryo.
The transovarian action of tebufenozide (84 g a.i ha -1 ) was reported by Pratissoli et al. (2003) who verified that adults of S. frugiperda presented significant reduction of 60% of laid eggs and 46.8% of viability after ingestion of honey + tebufenozide (a growth regulator from diacil-hidrazine group), but without decrease in longevity of adults.The authors observed that the growth regulator lufenuron did not affect fecundity or longevity when ingested by females of S. frugiperda, but reduced the average viability of eggs in 93%.
evaluated its action on third instar A. C. M. Redoan et al.

Table 1 -
(HASSAN et al., 1985)of adults (males and females) of Doru luteipes (Dermaptera: Forficulidae) after being treated with insecticides at different time intervals and classes of toxicity of the compounds evaluated 1 Mean followed by the same letter, in uppercase or lowercase column lines for each insecticide, do not differ by Scott-Knott test (SCOTT;KNOTT, 1974)at 5% significance level. 2 Classe toxicity of insecticides second IOBC / WPRS(HASSAN et al., 1985)6% of eggs were viable and 60% of nymphs from these couples reached the fourth instar (Table2).

Table 2 -
Percentage of female Doru luteipes (Dermaptera: Forficulidae ) postures, egg viability (%) and survival of nymphs from directly couples treated with insecticides 1 Mean followed by the same letter, in uppercase or lowercase column lines for each insecticide, do not differ by Scott-Knott test (SCOTT; KNOTT, 1974) at 5% significance.*No live insects for evaluation of effect residual contact of insecticides on adults of D. luteipes

Table 3 -
Cumulative mortality (%) (±SE) of adult Doru luteipes (Dermaptera: Forficulidae ) after initial exposure of insects to residues of the compounds at different time intervals and classes of toxicity of the insecticides evaluated

Table 4 -
KNOTT, 1974)ival of adult Doru luteipes (Dermaptera: Forficulidae) after the consumption of eggs of Spodoptera frugiperda (Lepidoptera: Noctuidae) were treated with insecticides and provided to the predator after one, 24 and 48 hours of classes and toxicity of the compounds evaluated Mean followed by the same letter, in uppercase or lowercase column lines for each insecticide, do not differ by Scott-Knott test (SCOTT;KNOTT, 1974)at 5% significance. 2Classe toxicity of insecticides second IOBC / WPRS (

Table 5 -
Percentage of female Dorus luteipes (Dermaptera: Forficulidae) posture and egg viability (%) from couples who consumed eggs of Spodoptera frugiperda (Lepidoptera: Noctuidae) treated with insecticides and provided at time intervals to the predator