Turkish Journal of Zoology Turk J Zool (2017) 41: 502-512 http://journals.tubitak.gov.tr/zoology/ © TÜBİTAK Research Article doi:10.3906/zoo-1603-50

Hymenoptera complex of oleae (Bernard) (: ) in Portugal

1, 1 2 3 4 1 Anabela NAVE *, Fátima GONÇALVES , Rita TEIXEIRA , Cristina AMARO COSTA , Mercedes CAMPOS , Laura TORRES 1 Centre for the Research and Technology of Agro-Environmental and Biological Sciences, University of Trás-os-Montes and Alto Douro, Vila Real, Portugal 2 Agrarian and Forestry Systems and Plant Health, National Institute of Agricultural and Veterinary Research, Oeiras, Portugal 3 Department of Ecology and Sustainable Agriculture, Agrarian School, Polytechnic Institute of Viseu, Viseu, Portugal 4 Department of Environmental Protection, Experimental Station Zaidín, Granada, Spain

Received: 23.03.2016 Accepted/Published Online: 29.11.2016 Final Version: 23.05.2017

Abstract: The , (Bernard) (Lepidoptera: Praydidae), is one of the most important pests of olive trees throughout the Mediterranean region, the Black Sea, the Middle East, and the Canary Islands. Thus, it is particularly important to develop alternative strategies to control this pest. Over a 4-year period, a survey was done in order to acquire knowledge about the complex of associated with this pest. Leaves, flowers, and fruit infested with larvae and pupae of P. oleae were collected from olive groves, conditioned in vials, and kept under laboratory conditions until the emergence of P. ol e ae adults or parasitoids. The abundance and richness of parasitoids as well as the rate of parasitism was estimated. parasitoids were found to be responsible for 43% of the mean mortality of the sampled individuals. From the 22 parasitoid recorded, three dominated in terms of the provoked parasitism rates: Ageniaspis fuscicollis (Dalman) (), Chelonus elaeaphilus Silvestri (), and flabellatus (Fonscolombe) (Elasmidae) (Hymenoptera). Additionally, several of the identified taxa were reported for the first time in Portugal as parasitoids of P. ol e ae . Moreover, a strong relationship between the species of parasitoids and the generation of the pest was found. Given the moth’s presence and its impact on olive tree production worldwide, the outcomes of this study show a rich complex of parasitoids that naturally occurs in association with P. ol eae , causing high mortality rates, and highlight the importance of these beneficial parasites in the control of this pest.

Key words: Olive, olive moth, parasitoids, Ageniaspis fuscicollis, Chelonus elaeaphilus, Elasmus flabellatus, Portugal

1. Introduction of the product. Crop protection is particularly affected by The olive tree has a genetic heritage of inestimable value these pressures, either by the costs associated with it or by in Mediterranean agriculture. In Portugal, there are more the undesirable side effects of pesticides on trophic chains than 350,000 ha of olive groves, and 36.6% of the country’s and biological balances. In Portugal, control of the olive olive grove area, mostly of traditional olive groves, is moth is usually achieved by spraying with pyrethroids located in the regions of Trás-os-Montes and Beira Interior (Mendes and Cavaco, 2009); however, it has been pointed (INE, 2014). out that, in general, the use of these products has adverse Prays oleae (Bernard) (Lepidoptera: Praydidae) is ecological, toxicological, and economic effects (Youssef et one of the most important pests of . This al., 2004). develops three generations per year. The overwintering Thus, it is particularly important to develop alternative generation (phyllophagous) develops in the leaves, the first strategies to control this pest. The natural control exerted generation (anthophagous) develops in inflorescences, by parasitoids seems especially promising. In fact, and the last generation (carpophagous) occurs in the these beneficial species constitute a large and relatively fruits. Bento et al. (2001) estimated losses at up to 368 diverse group, whose efficiency can reach high levels €/ha in the anthophagous generation and 535 €/ha in in some regions (Pralavorio and Arambourg, 1981; the carpophagous generation. In recent years, high Ramos et al., 1983). Studies conducted in Portugal have socioeconomic pressures have been forcing olive growers emphasized the frequency with which parasitoids are to reduce production costs while improving the quality observed in olive groves, especially those belonging to the * Correspondence: [email protected] 502 NAVE et al. / Turk J Zool

Braconidae, , Eulophidae, , paper new information on the occurrence of some species, and (Hymenoptera) (Herz et al., subfamilies, and families of parasitoids of P. ol e ae . 2005). The beneficial species reported as being the most important in the control of P. ol e ae populations 2. Materials and methods are Ageniaspis fuscicollis var. praysincola (Dalman) The experimental part of this study took place in traditional (Encyrtidae), Chelonus elaeaphilus Silvestri (Braconidae), olive groves, as characterized by Beaufoy (2001), located Apanteles xanthostigma (Haliday) (Braconidae), in the districts of Guarda (40°32′N, 7°15′W), Viseu agraules (Walker) (Eulophidae), Elasmus flabellatus (40°39vN, 7°54vW), Vila Real (41°17′N, 7°44′W), and (Fonscolombe) (Elasmidae), and Diadegma armillata Bragança (41°48vN, 06°45′W), in the central interior and (Gravenhorst) (Ichneumonidae) (Hymenoptera) (Bento et northeastern regions of Portugal (Figure 1). al., 2007). The age of the trees was variable, ranging from 10 In order to develop a successful conservation to over 100 years; the cultivars predominant in the biological control program against P. ol e ae , updated regions under study are Cobrançosa, Galega, Verdeal information is needed about these parasitoids. Therefore, Transmontana, and Madural. Groves were rain-watered studies were conducted to identify parasitoid species of and subject to phytosanitary treatments according to the pest, as well as to evaluate their relative importance integrated production and organic farming guidelines. in the natural control of the pest in the central interior The annual climatic characterization of Portugal, and northeastern regions of Portugal. We present in this temperature, and precipitation monthly mean for the

Figure 1. Map of Portugal showing the regions where parasitoid inventories were done.

503 NAVE et al. / Turk J Zool country for the 4 years in the study from 2009 to 2012 were variables (e.g., year, region, generation, temperature, obtained from official national records (IPMA, 2009, 2010, and precipitation) had a significant relationship with the 2011, 2012). parasitoid data. Statistical significance of the canonical The survey of parasitoids was carried from March axes was evaluated using the Monte Carlo permutation 2009 to September 2012. In each generation, samples of test (P < 0.05) (Šmilauer and Lepš, 2014). leaves, flowers, and fruits infested with larvae and pupae of P. oleae were collected by hand for the phyllophagous, 3. Results anthophagous, and carpophagous generations, respectively, During the 4 years, 9285 P. ol e ae larvae and pupae were with the exception of the anthophagous generation in 2010 collected; from these a total of 3186 parasitoid individuals and 2012. Leaves and flowers were collected from the trees emerged. From the remaining sample, 68% developed and fruits from among recently fallen fruits. For each into P. ol e ae and 32% died, probably due to abiotic generation, between 532 and 4962 samples were harvested, factors, sample handling, or diseases. depending on the observed attack in the olive groves, and The percentage of parasitism (Table 1), which differed in each sample olive grove, a minimum of 50 attacked between years and generations, was estimated at from organs were collected. 10.9% in the carpophagous generation in 2012 to 78.2% in In the laboratory, the collected leaves, flowers, and fruits the anthophagous in 2011. The greatest value by generation were enclosed singly into vials; maintained in a climate- was observed in the phyllophagous generation in 2012, controlled chamber at 25 ± 2 °C, 60 ± 5% relative humidity and in the anthophagous and carpophagous in 2011. (RH), and a photoperiod of 16 : 8 (L : D) h; and observed All parasitoids collected belonged to Hymenoptera, daily until adult emergence. Parasitoid adults obtained from which 78.7% were Chalcidoidea, and the remaining were identified on the basis of adult morphological criteria 21.3% were . The Chalcidoidea were (Medvedev and Sharma, 1988; Shaw and Huddleston, distributed into 6 families, namely Chalcididae, Elasmidae, 1991; Goulet and Huber, 1993). Encyrtidae, Eulophidae, , and Mymaridae, Parasitism was measured a posteriori by checking while within the Ichneumonoidea, both Braconidae and emergence and was calculated as the percentage of Ichneumonidae were found (Figure 2). parasitism [(number of P. ol e ae parasitized / total number From the 22 parasitoid taxa identified, the most of P. ol e ae ) × 100]. abundant were, in decreasing order, Ageniaspis fuscicollis The richness of parasitoids was calculated both by the with 64.9% of the total, Chelonus elaeaphilus with 11.5%, number of taxa (S) and by Margalef’s index: d = (S – 1) / Elasmus flabellatus with 10.2%, and Apanteles xanthostigma ln (N), where N is the number of individuals (Margalef, with 6.0% (Table 2), and with less representation Diadegma 1958). armillata, Bracon crassicornis Thomson (Braconidae), Differences in parasitoid communities’ structure elegans (Woldstedt) (Ichneumonidae), Pnigalio among P. ol e ae generations, as well as the relationship agraules, Brachymeria sp. (Figure 3A), between the studied taxa and the climate conditions, region, sp. (Figure 3B), albitarsis (Zetterstedt) (Figure and year, were analyzed with multivariate techniques 3C), sp. (Figure 3D), Hockeria sp. (Figure using CANOCO software version 5.04 (Biometris-Plant 3E), and sp. (Hymenoptera: Chalcididae) Research International, Wageningen, the Netherlands). (Figure 3F). Additionally, there were members of the Canonical correspondence analysis was performed families Mymaridae and Pteromalidae and subfamilies using log-transformed data (log [y + 1]) and based on Brachymeriinae, , Euderinae, Entedontinae, parasitoid data matrices. A forward selection procedure was Meteorinae, and Opiinae. Most of these specimens could performed to determine which explanatory environmental not be identified to the species level, either due to their

Table 1. Percentage of larvae and pupae of P. oleae from which emerged parasitoids in each generation and year of study. The total numbers of individuals observed are in parentheses.

Generation / Year 2009 2010 2011 2012 Phyllophagous 24.2 (248) 29.1 (563) 26.7 (1999) 37.2 (541) Anthophagous 25.4 (614) * 78.2 (1529) * Carpophagous 64.2 (372) 68.5 (677) 76.3 (114) 10.9 (652)

* Data not available.

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Figure 2. Distribution of the parasitoids collected by family.

Table 2. Parasitoids of larvae and pupae of P. oleae collected in all groves and studied years by taxa, total number of individuals, and percentage (%) of the total.

Superfamily Family Subfamily Species Totals % Chalcidoidea Chalcididae Brachymeriinae** Brachymeria sp.* 2 0.06 Chalcidoidea Chalcididae Haltichellinae** Haltichella sp.* 16 0.50 Chalcidoidea Chalcididae Haltichellinae** Hockeria sp.* 2 0.06 Chalcidoidea Elasmidae Elasminae Elasmus flabellatus 326 10.23 Chalcidoidea Encyrtidae Encyrtinae Ageniaspis fuscicollis 2070 64.97 Chalcidoidea Eulophidae Entedontinae** *** 3 0.09 Chalcidoidea Eulophidae Entedontinae** Chrysocharis sp. * 2 0.06 Chalcidoidea Eulophidae Euderinae** *** 1 0.03 Chalcidoidea Eulophidae Euderinae** Euderus albitarsis* 1 0.03 Chalcidoidea Eulophidae Pnigalio agraules 39 1.22 Chalcidoidea Eulophidae Eulophinae *** 27 0.85 Chalcidoidea Eulophidae Eulophinae *** 11 0.35 Chalcidoidea Eulophidae Tetrastichus sp.* 4 0.13 Chalcidoidea Mymaridae **** **** **** 1 0.03 Chalcidoidea Pteromalidae**** **** **** 2 0.06 Ichneumonoidea Braconidae Braconinae Bracon crassicornis 90 2.82 Ichneumonoidea Braconidae Chelonus elaeaphilus 367 11.52 Ichneumonoidea Braconidae Meteorinae ** ** 1 0.03 Ichneumonoidea Braconidae Microgastrinae Apanteles xanthostigma 191 5.99 Ichneumonoidea Braconidae Opiinae** ** 6 0.19 Ichneumonoidea Ichneumonidae Campopleginae Diadegma armillata 6 0.19 Ichneumonoidea Ichneumonidae 13 0.41 Ichneumonoidea Ichneumonidae Pimplinae *** 5 0.16 * First record from the species as a P. oleae parasitoid in Portugal. ** First record from the subfamily as a P. oleae parasitoid in Portugal. *** Specimens in poor condition or sex obtained not used in key identification, but not previously cited as a P. oleae parasitoid in Portugal. **** First record of the family as a P. oleae parasitoid in Portugal; genus not identified.

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Figure 3. Some of the parasitoids of P. ol e ae collected in this study, not yet identified in Portugal. poor condition or because the available keys do not allow generation in 2011, while no parasitism was found in the for the identification of the sex collected. phyllophagous generation in 2009. As expected, the relative abundance of each species of Elasmus flabellatus was the third most abundant parasitoid in samples changed between generations of P. parasitoid. The phyllophagous and anthophagous oleae as well as between years (Table 3). generations were found parasitizing P. ol e ae in 2009 and The richness of parasitoids per generation in the 4 2011, as well as the phyllophagous generation in 2012, years of study was between 6 and 7 in the phyllophagous, but it was not present in 2010 (Table 3). The parasitism 10 in the anthophagous, and between 5 and 6 in the levels ranged from 4% in 2011 to 30% in 2009, both in the carpophagous; these numbers correspond to a Margalef phyllophagous generation. index of 2.08 in the anthophagous generation, 1.46 in the Apanteles xanthostigma was also found in all generations phyllophagous, and 1.04 in the carpophagous. of P. ol e ae . The corresponding levels of parasitism ranged Ageniaspis fuscicollis was the most abundant in almost from 0.6% in the anthophagous generation in 2009 all samples collected, with a percentage of parasitism that and 19% in the carpophagous generation in 2010. The for the anthophagous generation in 2011 reached 78.7% of phyllophagous generation was found in 2 years, with levels the samples. of parasitism that ranged between 8.1% and 12.4% (Table In the second place of importance was C. elaeaphilus, 3). with a percentage of parasitism close to 50% of the Bracon crassicornis was found every year in the samples in the carpophagous generation in 2009 and carpophagous generation with levels of parasitism that 2012. C. elaeaphilus was present in every year and almost ranged from 2.5% in 2009 to 40.0% in 2011. Moreover, it always with a remarkable percentage of parasitism in the was observed parasitizing in the anthophagous generation carpophagous generation; however, the levels of parasitism in 2009, but was only 0.6% of the sample, while it was not were highly variable between generations and years. found in the phyllophagous generation. For example, it was lower than 1% in the anthophagous

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Table 3. Percentage (%) of larvae and pupae of P. ol e ae parasitized by each of the identified species of parasitoids in each year and generation.

2009 2010 2011 2012 Species generation generation generation generation Phy. . Car. Phy. Ant. Car. Phy. Ant. Car. Phy. Ant. Car. Ageniaspis fuscicollis 26.7 59.0 46.4 56.4 * 63.0 72.0 78.7 36.0 46.3 * 21.0 Apanteles xanthostigma 0.0 0.6 0.8 0.0 * 19.0 8.1 1.8 11.0 12.4 * 1.4 Brachymeria sp. 0.0 0.0 0.0 0.0 * 0.0 0.0 0.2 0.0 0.0 * 0.0 Bracon crassicornis 0.0 0.6 2.5 0.0 * 6.3 0.0 0.0 40.0 0.0 * 27.0 Chelonus elaeaphilus 0.0 3.8 48.1 39.9 * 12.0 16.0 0.8 6.9 1.0 * 49.0 Chrysocharis sp. 1.7 0.0 0.0 0.6 * 0.0 0.0 0.0 0.0 0.0 * 0.0 Diadegma armillata 0.0 2.6 0.0 0.0 * 0.0 0.0 0.0 2.3 0.0 * 0.0 Elasmus flabellatus 30.0 29.5 0.0 0.0 * 0.0 3.9 15.3 0.0 28.9 * 0.0 Euderus albitarsis 0.0 0.0 0.0 0.0 <0.1 0.0 0.0 0.0 0.0 0.0 * 0.0 Haltichella sp. 0.0 0.6 0.0 0.0 * 0.0 0.0 1.3 0.0 0.0 * 0.0 Hockeria sp. 0.0 0.0 0.0 0.0 * 0.0 0.0 0.2 0.0 0.0 * 0.0 Pnigalio agraules 11.7 0.0 0.0 0.6 * 0.2 0.2 1.9 3.4 1.0 * 1.4 Scambus elegans 1.7 0.6 0.0 1.2 * 0.0 0.0 0.2 0.0 0.0 * 0.0 Tetrastichus sp. 0.0 0.0 1.7 0.0 * 0.0 0.0 0.0 0.0 0.0 * 0.0 Entedontinae** 0.4 0.6 0.2 Euderinae** 0.6 Eulophinae 28.3 10.4 7.7 Mymaridae *** <0.1 Pteromalidae*** <0.1 Meteorinae ** 0.6 Opiinae** 1.9 <0.1 Pimplinae 0.42 Generation: Phy. – phyllophagous; Ant. – anthophagous; Car. – carpophagous. * Data not available. ** First record from the subfamily as a P. ol e ae parasitoid in Portugal; genus not identified. *** First record of the family as a P. ol e ae parasitoid in Portugal; genus not identified.

Pnigalio agraules was the sixth most abundant Haltichella sp., Brachymeria sp., Hockeria sp., and E. parasitoid species identified, corresponding to 1.2% of albitarsis were found only in P. ol e ae in the anthophagous the total registered parasitism (Table 2). This species was generation. Haltichella sp. was found in 2009 and 2011, collected in all generations with parasitism rates ranging with levels of parasitism that ranged from 0.6% in 2009 to from 0.2% in the carpophagous generation in 2010 to 1.3% in 2011; Brachymeria sp. and Hockeria sp. were found 11.7% in the anthophagous generation in 2009 (Table 3). in 2011 both at 0.2%; and only one E. albitarsis individual Diadegma armillata was found only in the was found in 2010 (Table 3). anthophagous generation in 2009 and in the carpophagous Chrysocharis sp. was present in 2009 and 2010, in the generation in 2011, parasitizing respectively 2.6% and phyllophagous generation in both years; Tetrastichus sp. 2.3% of the sample (Table 3). was present only in 2009 in the carpophagous generation.

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The specimens identified only to the family and is significant (P = 0.018; F = 1.9) and explains 12.7% of subfamily level totaled 52 individuals distributed as parasitoid variation, and the first two axes explain 43.57% follows: Opiinae (6), Meteorinae (1), Euderinae (1), of the variance in the parasitoid species (Figure 4). Entedontinae (3), Eulophinae (38), Mymaridae (1), and There was a very strong relation between the species Pteromalidae (2). and the generation (pseudocanonical correlation = 0.94 on The Opiinae were collected from the anthophagous the first axis) (Figure 4). The first two axes explain 56.13% generation of P. ol e ae in 2009 and 2010, while the of the variance in parasitoid species. The carpophagous Meteorinae were also collected from this generation in generation is significant (P = 0.002; F = 2.5) and explains 2009. The Euderinae were collected in the phyllophagous 16.1% of parasitoid variation; the anthophagous generation generation in 2010; the Entedontinae in the carpophagous is significant (P = 0.008; F = 2.4), explaining 15.8% of generation in 2009 and in the phyllophagous generation parasitoid variation; and the phyllophagous generation in 2010 and 2011; the Eulophinae in the phyllophagous is significant (P = 0.04; F = 1.6) and explains 16.1% of generation in 2009 and 2012; the Mymaridae in the parasitoid variation. The carpophagous generation is phyllophagous generation in 2009; and the Pteromalidae positively related with Bracon crassicornis and Tetrastichus in the anthophagous generation in 2010. sp.; the anthophagous generation is positively related with There was a strong relationship between the species and Haltichella sp., Brachymeria sp., Hockeria sp., E. albitarsis, the variables under study (pseudocanonical correlation Meteorinae, Opiinae, Pimplinae, and Pteromalidae (Figure = 0.89 on the first axis), wherein the generation variable 4).

Figure 4. Canonical correspondence analysis between parasitoids and pest generations.

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4. Discussion (Bento, 1999; Teixeira et al., 2000; Soares et al., 2005), The results of this study show that a rich complex of individuals from the Brachymeriinae and Haltichellinae parasitoids that naturally occurs parasitizes P. ol e ae . (Chalcididae), Euderinae and Entedontinae (Eulophidae), Not only the composition of this complex but also the Meteorinae (Ichneumonidae), and Opiinae (Braconidae) relative abundance of each species of parasitoid in the were reported for the first time in Portugal as parasitoids sample changed both between generations of P. ol e ae of P. ol e ae in this work. Due to its constant presence and and years. Parasitoid generations are suggested to be abundance, Ageniaspis fuscicollis is considered by many positively associated with pest generations. The richness authors as the most important parasitoid of the three P. of parasitoids was higher in the anthophagous generation, oleae generations (Arambourg, 1964; Carmona and Alvim, corresponding to a generation with greater in 1966; Martelli, 1967; Campos, 1976; Bento, 1999). In this the sampled community, and lower in the carpophagous study, A. fuscicollis was found in each P. ol e ae generation, generation. Teixeira et al. (2000) reported three periods of being more abundant in 2011, the warmest year of the higher intensity of Hymenoptera captures in olive groves, studied period (Table 4). As for the maximum annual which coincided with the occurrence of the last larval temperature, the values recorded puts 2011 right between stages and pupae of each of the three generations of the the three hottest years since 1931, with the total annual olive moth. precipitation below the normal value (IPMA, 2011). Our study shows that, consistently throughout the The parasitism caused by C. elaeaphilus, close to 50%, 4 years (from 2009 to 2012), Hymenoptera parasitoids is in agreement with that recorded by Pelekassis (1962), were responsible for 43% of the mean mortality of the Arambourg (1964), and Bellido (1977) and close to that sampled individuals. Twenty-two parasitoid species were presented by Herz et al. (2005), who reported levels of recorded. Notably, three appear to have a dominant role in mortality for 2004 caused by this parasitoid higher than the control of P. ol e ae due to the percentage of parasitism those caused by A. fuscicollis. Chelonus elaeaphilus was observed, namely A. fuscicollis, C. elaeaphilus, and E. present in the majority of samples. However, this species flabellatus. While A. fuscicollis, C. elaeaphilus, E. flabellatus, seems to be associated with warm, dry weather conditions, A. xanthostigma, D. armillata, B. crassicornis, S. elegans, as occurred in 2009 and 2012 when its higher rates of and P. agraules had already been reported in Portugal parasitism were found. In 2009, the mean values for the

Table 4. Climatic characterization of the 4 years under study: temperature (Temp.) and precipitation (Precipit.).

Year 2009 2010 2011 2012 Temp. Precipit. Temp. Precipit. Temp. Precipit. Temp. Precipit. January = ↑ ↓ ↑ = = = ↓ February = = ↓ ↑# = = § »↓ March ↑≈ ↓ ↓* ↑# = = ↑≈ »↓ April = ↓ ↑ ↓ ↑+ ≈ = =  May ↑≈ ↓ ↑≈ ↓ ↑ +≈≈ = ↑≈ = June ≈ ↑ = ↓ ↑ = = = July = ↓ ↑≈ ≈ ↓ - = = = = August ↑≈ ↓ ↑≈ ↓ - = = = September ≈ ↓ ↑ ↓ - ↑ = ↑≈ = October ↑≈ ↓ ↓ ↑ ↑+≈≈ = = = November = ↑ ↓ ↑ = ↑ =  December = ↑ ↓ ↑ = ↓ = = Maximum Rainiest Below the Synthesis of the Normal to Hottest Lower and mean High mean of the last normal Dry year dry since 1931 middle high decade value = Normal + Value greater at least in terms decade  Snow ≈ Heat wave  Highest maximum monthly since 1931 » Extreme drought ↑ Higher than the mean value (1971–2000) - Driest in 22 years § Cold snap ↓ Below the mean value * Coldest in 24 years # Wettest in 24 years

509 NAVE et al. / Turk J Zool maximum and mean air temperature registered in the the genus Brachymeria are parasitoids of insect larvae studied region were higher than the mean. The year was (Ghoneim, 2014). In Egypt, they are present in olive groves classified as a normal to dry year, with the spring the driest as parasitoids of Palpita unionalis Hübner (Lepidoptera: one registered since 1931 (Table 4) (IPMA, 2009). Crambidae) (El-Basha, 2002). Euderus albitarsis has In 2012 there was a meteorological drought, which been referenced as belonging to the parasitoid complex began in late 2011 and continued throughout most of of the genus Prays in the Mediterranean region (Moreno this year; thus, the winter of 2011/12 was the eighth driest et al., 1990), and Silvestri (1907) mentioned Hockeria year of the previous 82 years. Moreover, the annual mean sp. and Tetrastichus sp. as part of the parasitoid complex temperatures were lower than the mean value, which has associated with P. ol e ae . Chrysocharis sp. was present in not happened for several decades (IPMA, 2012). the phyllophagous generation, which is in accordance with The results observed for E. flabellatus, one of the most Agrò et al. (2009) and Hansson (1985, 1987). It should be abundant species recorded, are in accordance with those noted that Haltichella sp., Brachymeria sp., and Hockeria presented by Campos and Ramos (1981), Katsoyannos sp. were found only in 2011, characterized in climatic (1992), and Bento et al. (1998). Its occurrence seems terms as the hottest year since 1931, which is in accordance to have been related to low values of humidity and with its greater diversity in tropical areas (Askew, 1994). precipitation, an unrecorded condition in 2010 (Table 4). Braconidae are diverse in all areas (Wharton, High mean values of temperature and precipitation in 2010 1993), but they are highly abundant in cool temperate were registered, it being the wettest year in the last decade. regions (Quicke and Kruft, 1995). They have an important The first and fourth quarters were the main contributors role in the biological control of phytophagous , to the high level of annual precipitation, since July, August, particularly economically important pests (Ghahari et al., and September were the driest in the last 22 years. The 2006; Beyarslan et al., 2010; Ghahari and Fischer, 2011). spring was the seventeenth consecutive spring with mean In summary, our results not only prove that the P. temperature of the air above the standard temperature, oleae native parasitoid complex is rich but also point to and the second summer with the highest maximum and its importance in the establishment of a biological control mean temperature since 1931 (IPMA, 2010). program against this pest in olive groves. Considering the Apanteles xanthostigma, found in all generations of relationship determined in this study between parasitoid P. ol e ae , is considered a parasitoid of minor importance species and generations of the pest is of special importance in the phyllophagous generation (Campos and Ramos, in promoting the maintenance of functional biodiversity 1981), although this study has shown levels of parasitism at large. Carrying out this type of study relating several that ranged between 8.1% and 12.4%. variables like year, temperature, precipitation, generation, While the parasitism rate of B. crassicornis was generally and region, among many other possible variables, is of low compared with other species, it could occasionally be great importance, given the possibility of response tools important as observed in the carpophagous generation and current statistics, but sometimes the most obvious in 2011, similar to what was registered by Soares et al. solution is determined by the holistic aspect. In this (2005) in the Beira region of Portugal. Pnigalio agraules is context, the goal is increasing the action of this parasitoid a potentially important parasitoid for biological control of biodiversity by chemical reduction of pest control measures key pests (Schauff et al., 1998), such as lepidopterous leaf and creating a suitable ecological infrastructure to provide miners (Askew 1971, 1984; Yegorenkova and Yefremova, resources, which are mostly plant communities, with 2012). In olive groves, in addition to being a parasitoid essential food sources for the fitness of adult parasitoids. of P. ol e ae , it was also found parasitizing Bactrocera oleae (Rossi) (Diptera: Tephritidae) (Bento et al., 2007; Gebiola et Acknowledgments al., 2009; Gonçalves et al., 2012). Concerning D. armillata, This work was supported by FEDER Funds through the low levels of parasitism observed agree with data Programa Operacional Factores de Competitividade - from the literature (Bellido, 1977; Campos and Ramos, COMPETE and National Funds through FCT - Fundação 1981; Bento, 1999; Bento et al., 2007). Other taxonomic para a Ciência e Tecnologia, within project PTDC/AGR- groups reported as P. oleae parasitoids elsewhere, notably AAM/100979/2008 – “Increasing functional biodiversity in Greece, Egypt, and Italy (e.g., Stavraki, 1970; Nasr in olive groves to enhance conservation biological control et al., 2002; Agrò et al., 2009), but not in Portugal, were of insect pests”, as well as PhD grant SFRH/BD/34394/2008 Brachymeria sp., Chrysocharis sp., E. albitarsis, Haltichella awarded to the first author. This manuscript is part of the sp., Hockeria sp., and Tetrastichus sp., as well as members first author’s PhD dissertation. The authors are grateful to of the families Mymaridae and Pteromalidae and the olive grove owners who permitted access to the groves, subfamilies Brachymeriinae, Haltichellinae, Euderinae, with support from the Associação de Agricultores para Entedontinae, Meteorinae, and Opiinae. All species of Produção Integrada de Frutos de Montanha.

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