Experimental mixtures of Phthorimaea operculella granulovirus isolates provide high biological efficacy on both Phthorimaea operculella and Tecia solanivora (Lepidoptera: Gelechiidae)

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Abstract

The Guatemalan potato moth Tecia solanivora (Povolny) recently invaded part of South America, colonizing zones where Phthorimaea operculella (Zeller), another potato moth species belonging to the same group, was previously established. T. solanivora is now the major insect pest of potato in this area encompassing Venezuela, Colombia and Ecuador. P. operculella granulovirus (PhopGV) (Betabaculovirus) is a biocontrol agent to be considered for the simultaneous management of these two potato pests, instead of classical chemical insecticides. In a previous work, five PhopGV isolates were isolated in Colombia from T. solanivora and were tested against larvae of the same species showing variable efficacies. Infections with mixtures of different genotypes of Baculoviruses had been carried out in a wide range of species and several showed interesting results. In the present study, the effect of sequential passages of PhopGV in P. operculella and T. solanivora larvae was analyzed through biological assays. Three different mixtures containing a Peruvian PhopGV isolate (Peru) adapted to P. operculella and a Colombian PhopGV isolate (VG003) adapted to T. solanivora were tested. A preliminary analysis of the correlation between the genotypic marker egt gene and the level of pathogenicity after a variable number of replication cycles was made. Mixtures of virus isolates showed a higher efficacy in both hosts compared to individual PhopGV isolates. This higher pathogenicity was maintained through passages. In P. operculella the mixtures were between 2.8 and 23.6-fold (from 7.15 OB/mm2 to 0.10 OB/mm2) more pathogenic than isolate Peru applied alone. In T. solanivora they were between 2.3 and 4.9-fold (from 12.29 OB/mm2 to 1.25 OB/mm2) more pathogenic than isolate VG003 alone. Viral biopesticide containing a mixture of selected genotypes active against each hosts seemed suitable for the development of a biopesticide aimed to simultaneously control P. operculella and T. solanivora.

Highlights

► The genotypic heterogeneity of mixtures of virus isolates determined their efficacy. ► All the mixtures tested had a great efficacy on both Phthorimaea operculella and Tecia solanivora. ► The mixtures always showed higher pathogenicity than each constitutive isolates alone. ► Successive passages on hosts did not affect efficacy of virus mixtures over time.

Introduction

Recently, the potato tuber moth Tecia solanivora Povolny (Lepidoptera: Gelechiidae), originating from Guatemala, became the major potato insect pest in Central America and in the northwest of South America i.e. Venezuela, Colombia and Ecuador (Barragán et al., 2004). T. solanivora entered the north of Colombia in 1985 through infested potato seeds imported from Venezuela and 11 years later, it was found in the southwest of the country, at the border with Ecuador (Puillandre et al., 2007). This species also invaded the Canary Islands (Spain) during the last decade (Carnero et al., 2008). It was then included by the European and Mediterranean Plant Protection Organization (EPPO) in its A2 action list of pests recommended for regulation as quarantine pest (EPPO/OEPP, 2005). During its invasion process, T. solanivora colonized many zones where the related species Phthorimaea operculella (Zeller) (Lepidoptera: Gelechiidae) was already established. The potato tuber moth P. operculella is an important pest for potato and is present worldwide under warm climates, both in field and in storage facilities (Sporleder et al., 2005). Nowadays, both species can be found in the same localities at different relative abundance according to local factors (Dangles et al., 2008). No specific chemical insecticides are available for the control of T. solanivora. Different molecules had been used but their efficacy has not been precisely quantified and they could cause adverse effects on beneficial organisms and environment (Arévalo and Castro, 2003). Furthermore, toxic residues can be found on potatoes (Benavides, 2000) and pesticides could generate resistance in insects. In this context, the use of biological control agents, and particularly insect viruses belonging to the family Baculoviridae (Baculoviruses), can be viewed as an interesting option to control these insects and to overcome the problems previously mentioned. Phthorimaea operculella granulovirus (PhopGV) (Betabaculovirus, Jehle et al., 2006) was first reported from P. operculella collected from Sri Lanka (Reed, 1969) but various isolates were found later in P. operculella populations from other countries (Vickers et al., 1991, Léry et al., 2005). PhopGV efficiently controls and prevents P. operculella proliferation under storage conditions without inducing any harm to beneficial organisms (Kroschel et al., 1996, Vickers et al., 1991). Several geographical graulovirus isolates were also found on T. solanivora. They appear to be genetically close to PhopGV (Espinel-Correal et al., 2010). Tests of some of these isolates on their original host species showed diverse efficacy levels (Espinel-Correal et al., 2010). Currently, the use of PhopGV-based insecticides is promoted by the International Potato Center (Lima-Peru) (CIP/IPC, 1992). We previously showed that the Colombian isolate VG003 PhopGV exhibited promising biological and technological characteristics for being used in the formulation of a viral biopesticide against T. solanivora (Chaparro et al., 2010). Furthermore, a significantly higher pathogenicity was observed in both P. operculella and T. solanivora when PhopGV isolates corresponding to a natural mixture of viral genotypes were used (Espinel-Correal et al., 2010), when compared to control (unmixed) isolates. Accordingly, virus isolates from either T. solanivora or P. operculella which exhibited apparent homogeneous genotypes appeared as more adapted to their respective original host as they had a lower efficacy on the other susceptible species (Espinel-Correal et al., 2010).

Co-infection or mixed infections caused by different genotypes of a single viral species or even by different species of viruses are frequently observed in insect–pathogen interactions. As a consequence, antagonism or synergism can occur between these genotypes or viral species. These interactions can modify biological parameters such as pathogenicity, virulence or virus yield, among others. In some occasions, the mixture of viral genotypes caused an increase of efficacy (Frank, 2003, López-Ferber et al., 2003, Shapiro and Shepard, 2006, Simon et al., 2005) but in other cases, virulence was reduced (Arends et al., 2005, Muñoz et al., 1998, Muñoz and Caballero, 2000).

Considering these results, our study examines the comparative efficacy of experimental mixtures of two PhopGV isolates (Peru and VG003) adapted to the two hosts P. operculella and T. solanivora, respectively. We also followed the evolution of pathogenicity through successive passages of various mixture ratios. Previous works (Léry et al., 2005, Léry et al., 2008) showed that some regions of the PhopGV genome exhibited significant sequence variation between isolates. Thus, we analyzed the evolution of the genetic composition of mixed viral populations through replication cycles using the genotypic marker egt gene. The correlation between the allelic frequency of this gene and efficacy was examined. The potential of mixture of PhopGV isolates as active ingredients of a biopesticide for the potato moths and others Gelechiidae pests is discussed.

Section snippets

Insect and viruses

P. operculella larvae were obtained from a laboratory colony maintained during more than 10 years in the research facility of the Institut de Recherche pour le Développement (IRD) in Saint-Christol-lez-Alès, France. The colony, originated from Egypt, was reared continuously on potato tubers (v. Agata) at 27 ± 1 °C, 70 ± 5% RH and 18L: 6D h photoperiod. T. solanivora larvae were multiplied in a laboratory colony of the Center of Biotechnology and Bioindustry of the Colombian Corporation for

Biological activity of virus mixtures in P. operculella and T. solanivora larvae

For P. operculella (Table 1), the probit analysis indicated a significantly higher pathogenicity of the three experimental populations (10A:90B, 50A:50B and 90A:10B) at passage 0 when compared to isolate Peru (χ2 = 83.9; df = 6; P < 0.05) and VG003 (χ2 = 167; df = 6; P < 0.05), individually applied to the same insect species. The decreases in LC50 of the experimental virus populations were between 2.8 and 23.6-fold compared to the isolate Peru which is considered as well adapted to this host species. Most

Conclusions

Based on biological assays, we conclude that the genotypic heterogeneity in PhopGV experimentally mixed isolates determined their efficacy. Moreover, the three experimental populations showed a higher pathogenicity than isolates tested separately. Apparently, virus passages did not seem to affect the higher efficacy of mixtures over time, at least in the first 3 (T. solanivora) or 5 (P. operculella) passages. The possibility relying on a viral population active against both insect hosts

Acknowledgments

The authors acknowledge the technical assistance of Mr. John Gallo for the rearing of T. solanivora. This work was partially funded by the Ecole des Mines d’Alès and IRD in France, and CORPOICA in Colombia. C. Espinel-Correal received a PhD fellowship from the Ecole des Mines d’Alès.

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