ENZYMATIC IDENTFICATION OF Glyptapanteles sp. (INSECTA: ) FROM MADEIRA ISLAND

L. OLIVEIRA, V. VIEIRA, J. TAVARES & P. GAKCIA

OLIVEIRA,L., V. VIEIRA, J. TAVARES & P. GARCIA. 3001. Enzymatic identification of Glyptaparzteles sp. (Insecta: Hymenoptera) from Madeira Island. Arquipélago. Life and Marine Science. Supplement 2 (Part B): 39-42. Ponta Delgada. ISSN 0873-4704.

Duririg a scientific expedition carried out in Madeira Island in September 1997, Pseudaletia (=Mytlzinzrza)ui~iliilricta (Lepidoptera: Noctuidae) larvae were collected in maize fields and pastures. These larvae wcre parasitized by a braconid belonging to Glyptaparzteles . This population was characterised biochemically to identify the species. Seven enzyme systems studied by electrophoresis were analysed: aldehyde oxidase (AO), a- glycerophosphate dehydrogenase (a-GPD). tetrazolium oxidase (TO), malate dehydrogenase (MDH), glucose-6-dehydrogenase (GóPD), malic enzyme (ME). isocitrate dehydrogenase (IDH). AI1 systems showed only one band, with two exceptions: a-GPD and ME which had two bands which corresponded to different loci. No polymorphic enzymes were detected. Comparing Lhis results with those obtained from G. nzilitaris collected in Azores Islands and in Quebec-Canada we can suggest that the population of Glyptapaiiteles collected in Madeira Island belongs to G. militaris species.

L. Olil~ira(e-r?lail: [email protected]), V. Vieira, J. lavares & P. Gnrcia, Universidade dos Açores, Departaniento de Biologia, Rua da Mãe de Deus 58, PT - 9501-801 Poiita Delgada, Port~lgul.

INTRODUCTION (PINTUREAIJ& VOEGELÉ1980; PINTUREAU1987; POWELL& WALTON1989). ln the Azores, P. urzipuncra has few species of Pseudaletia (=Mythiriiria) uilipurzcta from Hymenoptera (TAVARES1989; (Lepidoptera: Noctuidae) is a cosmopolitan OLIVEIRA1996). with the braconid G. militaris species, considered as the most important pest in being the main native biological control agent. Azorean gramineous fields (TAVARES 1989). Therefore, a larger survey for P. irnipuncta Severa1 studies concerning the establishment of biological control agents was expanded to other biological control programs against this pest have Macaronesian Islands, such as Madeira. been carried out in our laboratory (TAVARES During a scientific expedition carried out in 1989; VIEIRA 1992; OLIVEIRA1996). In this Madeira Island in September 1997, P. unipuncta sense, it is important to survey the pest natural larvae were collected in maize fields and pastures. enemies and preserve their characteristics during These larvae were parasitized by a braconid the severa1 steps of biological control programs, belonging to Glyptapanteles genus, as in Azores such as, the mass rearing processes and field Islands. releases. However, some problems may arise Since electrophoretic studies have been used when such programs are under development such to identify and distinguish between sibling as species mis-identification (especially for species (PINTUREAU & VOEGELÉ 1980; sibling species) leading to a loss of effort, time PINTUREAU1987; PINTUREAUet al. 1990) we and money (POWELL & WALTON 1989). analyzed seven enzyme systems by gel Electrophoretic study, as a too1 for identification, electrophoresis to compare the parasitoids from may help to overcome some of such errors Madeira with the Azorean population of G. militaris. This study allowed an increase on the Tetrazolium oxidase. Malate dehydrogenase, records of Madeira native species and the genetic Glucose-6-dehydrogenase and Isocitrate characterization of the 's populations, dehydrogenase exhibited a single band, for each contributing to the proper development of of these five systems, therefore only one locus biological control programs. was found. However, a-Glycerophosphate dehydrogenase and Malic enzyme exhibited two MATERIAL AND METHODS bands, corresponding two different loci. Table 1 Adult parasitoids were obtained from larvae of P. Electrophoretic conditions used to study seven unipuncta, randomly collected at Madeira Island enzymes systems in G. militaris. Miguel Island only in pastures. The encountered P. unipuflcta larvae, more than 100, Were Buffers Mi~ration Revelation P"'"i" Bridgr Grl iitir voltnpe Lim. suIulioo individually isolated in plastic cages (4.5~3em), (Minutrs) (Vollrl (Minutcs) and kept in the laboratory at 22rr 1"C, 7545% RH Trir/HCI T,~SWCI TrirWCI and L:D 16:8, for daily observation of ~~~;~~d;Ao) 4 ?4 7 ',E 30 ISO 30 Bçnzeldridr pH 6 7 pH 8 9 90 300 rmmtcmp NBT emergence. Twenty-eight of these larvae were PMS parasitized by G. militaris, producing an average TrisMCI a-Glycrropliosliiiair U- of 32 cocoons per host. After parasitoid adult D,L,Glyçrrnplinsplia~c drhydroprnnsr (u-GPD) 40 NAD emergence, the were stored frozen at -20 Triraroliuni 37°C NBT oxid~~r(TO) PMS "C until subsequent electrophoresis (frozen period MTT

< 1 month). TrisWCI L-Mslic acid Each adult was isolated and homogenized in Malaie 40 NAD ilchydruprnarc (MDH) 37°C NBT 15 microliter of "Trudgill" solution and PMS centrifuged for 5 minutes at 9 000 rpm, according TrisMCI to the methodology used by PINTUREAU(1987). Giyccrold-phi,rphaie NADP A vertical electrophoresis on polyacrilamide Gluci>rrd4ehydro~rnnsr 40 MgCI, (Gd~PD) 17°C NBT gel was performed on seven enzyme systems: PMS aldehyde oxidase (AO), a-glycerophosphate NBT TrisIHCI dehydrogenase (a-GPD), tetrazolium oxidase L-Maliç acid NADP (TO), malate dehydrogenase (MDH), glucose-6- Malic inzyrnc 15 MKIZ (ME) 3? C PMS dehydrogenase (GóPD), malic inzyme (ME) and MTT isocitrate dehydrogenase (IDH). The techniques Aga, 2 T used generally followed the methodology TriríHCI D.L-lsociuic acid described by PINTUREAU(1987), PINTUREAUet Iroçiirair drliydroprnnrr 60 NADP (IDH) 37°C MgCI2 al. (1991) and OLIVEIRA(1996), with specific PMS details in table 1. Eighteen individuals (each one from an individual parasitized host) collected in The two populations of G. militaris were Madeira Island were run for each enzymatic examined for nine allozyme loci, but a11 of them system. As control, two positively identified appeared entirely monomorphic. The enzymes specimens of G. militaris from São Miguel Island used in this study are useful to characterize (OLIVEIRA1996) were run for each enzymatic genetically C. militaris, but they constitute only a system. part of the species genome. Most species of Hymenoptera are reported to have low RESULTS AND DISCUSSION electrophoretic variation (PINTUREAU 1987; OMWEGA& OVERHOLT1996). Our data indicated The electrophoretic patterns observed in G. that G. militaris falls in the former group with militaris were similar for the populations of less variability, as Cotesia glomerata Madeira and Azores (Fig. 1). Aldehyde Oxidase, (unpublished data). The comparison of both populations suggests Glyptapanteles, Cotesia) is cited from Madeira, that the Glyptapanteles population collected in but the species were not identified. 'Therefore, the Madeira Island belong to G. militaris species. The species C. nlilitaris is the first time recorded for same bands were observed for a population of C. this island. militaris from Quebec-Canada (OLIVEIRA,1996). The presence of this parasitoid in Madeira The results of the morphological study carried on Island can be a good indicator of the natural both populations confirm the enzymatic control exerted by this wasp on the populations of identification of the species (OLNEIRA et the agricultura1 pest P. unipuncta. Furthermore, a1.1999). the population of C. militaris should be protected According to BAEZ (1993) and GRAHAM from the indiscriminate use of pesticides. (1986a, 1986b), the genus Apanteles (=

10.4

AO TO MD G-6-PD ID ME a4PD Madeira 18 18 18 18 18 18 18 S. Miguel 2 2 2 2 2 2 2 Fig. 1. Electrophoretic parttems of seven enzyme sistems observed in two populations of G. militaris from Madeira and Azores Islands; AO - Aldehyde oxidase; TO - Tetrazolium oxidase: MDH - Malate dehydrogenase; G-6-PD - Glucose-6-dehydrogenase; IDH - Isocitrate dehydrogenase; ME - Malic enzyme and (r-GPD - Glycerophosphate dehydrogenase.

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