Characterization of a Single-Nucleocapsid Nucleopolyhedrovirus of Thysanoplusia orichalcea L. (Lepidoptera: Noctuidae) from Indonesia

Thysanoplusia orichacea multicapsid nucleopolyhedrovirus (ThorMNPV) carrying an enhanced green fluorescent protein (EGFP) gene expression cassette (vThGFP) was used to study host-range mechanisms. Infection kinetics showed that vThGFP replication in Sf21 cells was too slow to suppress cell growth. Wide-host-range Autographa californica MNPV (AcMNPV) could speed up vThGFP infection and enhance the vThGFP infection rate in Sf21 cells. The enhancement was not due to recombination, as no recombinant virus was isolated from coinfection by plaque assay. No improvement of vThGFP infection in Sf21 was found by AcMNPV cosmid transactivation assay. However, culture medium from Sf21 cells infected with AcMNPV did enhance vThGFP replication in Sf21. Third-instar larvae of Spodoptera frugiperda, S. exigua and Helicoverpa zea were not killed by feeding with vThGFP polyhedra but were killed by intrahaemocoelic injection using budded viruses (BVs). This suggested that insufficient BVs were generated during the primary infection in the midgut. vThGFP infected haemocytes, tracheae and Malpighian tubules but not fat bodies of larvae of S. frugiperda, S. exigua and H. zea. Third-instar S. frugiperda larvae co-infected by injection with vThGFP and vAcDsRed2, an AcMNPV expressing a red fluorescent protein gene, showed EGFP expression in the fat body. This result suggests that vAcDsRed2 could help vThGFP to replicate in the fat body or trans-activate EGFP expression in the fat body. All these results suggested that slow cell infection, insufficient primary infection and inability to replicate in the fat body control the host range of ThorMNPV.

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“…Insects used in this test included P. includens, S. frugiperda, S. exigua and H. zea. Insects were reared according to Cheng & Carner (2000).…”

Section: Methodsmentioning

confidence: 99%

Slow cell infection, inefficient primary infection and inability to replicate in the fat body determine the host range of Thysanoplusia orichalcea nucleopolyhedrovirus

Wang

Salem

Lynn

et al. 2008

Journal of General Virology

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“…However, the images obtained by these authors do not differ from that obtained with the use of the technique three, described in this work, where the faces of the polyhedron appeared smooth, facilitating the morphologic descriptions. Similar methodology is described by Cheng et al (1998), Cheng & Carner (2000 and Cheng et al (2005), used to determine the morphology of SNPV and MNPV of Thysanoplusia orichalcea (L.) (Lepidoptera: Noctuidae).…”

Section: Resultsmentioning

confidence: 99%

Nucleopolyhedrovirus: scanning electron microscopy technique

Torquato¹,

Neto²,

Brancalhão³

et al. 2006

Neotrop. Entomol.

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Neotropical Entomology 35(6):787-790 (2006) Nucleopolyhedrovirus: Técnica de Microscopia Eletrônica de Varredura RESUMO -Um método simples foi desenvolvido para se estudar a forma geométrica do Bombyx mori Nucleopolyhedrovirus múltiplo, BmMNPV, para microscopia eletrônica de varredura. O vírus, pertencente à Família Baculoviridae, foi obtido do fi ltrado, do inóculo e da hemolinfa de lagartas de Bombyx mori (L.) (Lepidoptera: Bombycidae) infectadas. O material foi colocado diretamente sobre o suporte de varredura, onde previamente foi aderida uma fi ta de cobre. Após secagem e metalização com carbono e ouro, o vírus foi analisado ao microscópio de varredura. Os poliedros apresentaramse com as faces lisas, perfeitamente defi nidas, possibilitando a análise matemática que o identifi cou como um octaedro truncado. A forma do poliedro pode apresentar valor taxonômico, uma vez que ela é específi ca para cada linhagem viral. PALAVRAS-CHAVE: Baculovírus, Bombyx mori, BmMNPV, poliedro, octaedro truncado ABSTRACT -A simplifi ed methodology was developed to study the geometric form of multiple Bombyx mori Nucleopolyhedrovirus by scanning electron microscopy. The virus belongs to Baculoviridae family and was isolated from the silkworm Bombyx mori (L.) (Lepidoptera: Bombycidae). The polyhedra of Nucleopolyhedrovirus were obtained from the fi ltrate, inoculum and hemolymph of the silkworm experimentally infected with nuclear polyhedra. This material was placed on stubs, where a copper tape was previously adhered. After dry at room temperature the virus was covered with carbon and gold. Scanning electron microscopy analysis reveled a well defi ned morphology for the polyhedra of multiple Bombyx mori Nucleopolyhedrovirus, making possible the mathematical study that identifi ed it as a truncated octahedron. The form of the polyhedron can present taxonomic value, once it is specifi c for each viral lineage.KEY WORDS: Baculovirus, Bombyx mori, BmMNPV, polyhedra, truncated octahedron Nucleopolyhedrovirus (NPV) is an important entomopathogenic virus belonging to Baculoviridae family. It is made up of double stranded DNA molecule, associated with proteins of the capsid, forming the nucleocapsid. Two different subgroups are recognized for NPV: the occlusion derived virions and the budded virus. These virion subgroups have different functions in the infection cycle. The occluded subgroups could be divided into two subgroups: those containing one nucleocapsid per virion are referred to as single-capsid NPVs (SNPVs), whereas those containing multiple nucleocapsids per virion are known as multiple NPVs (MNPVs) (Murphy et al. 1995, Blissard 1996.Nucleocapsid and envelope constitutes the virion or infectious particle. In NPV several virions are contained in a protein structure, named occlusion body or polyhedron (Adams & McClintock 1991, Murphy et al. 1995, Blissard 1996. Polyhedra are stable structures that can persist for long time in the environment, where they are commonly found on the surface of the plants or in the soil. In the so...

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“…Penelitian ini bertujuan untuk mengetahui karakteristik molekuler melalui urutan nukleotida isolat NPV yang berasal dari H. talaca di Perkebunan teh Gunung Mas Bogor dengan gen lef-8. (Cheng, 1998).…”

Section: Pendahuluanunclassified

Karakterisasi Molekuler Nucleaopolyhedrovirus (Npv) Hyposidra Talaca Wlk. Di Perkebunan Teh Gunung Mas Bogor

Kusumah¹,

Revi²,

Kurniawati³

2017

JURNAL HAMA DAN PENYAKIT TUMBUHAN TROPIKA

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Characterize molecular the Nucleopolyhedrovirus (NPV) Hyposidra talaca Wlk. of tea plantation at Gunung Mas Bogor. Hyposidra talaca is one of the most important pest in tea plantation, and generally attacks of leaves and shoots. This pest cause yield loss up to 40-100%. NPV can be pathogenic to the pest of H. talaca and can be developed as an alternative measure to control H. talaca in tea plantations and based management appears to be more ecofriendly and effective. However, information regarding characterisation molecular of HytaNPV is limited. The study conducted to characterize molecular the NPV of H. talaca by restriction nucleotide and amino acid, by using gens lef-8. Molecular identification used Polymerase Chain Reaction (PCR) consisted of DNA extraction, DNA amplication, and DNA electrophoresis. DNA amplication using gen lef-8 showed positif result with approximately 770 bp. Gen lef-8 can identified HytaNPV. DNA sequance showed that isolate HytaNPV Bogor had high homology of pathogonic NPV of genus Helicoverpa from Brazil, Australia, Spanyol and Netherland with homology nucleotide and amino acid reached 98% and 100%. Based on philogeny tree of HytaNPV was one group with pathogenic NPV of genus Helicoverpa. Key words: characterize NPV, gens lef-8, Hyposidra talaca, NPV ABSTRAKKarakterisasi molekuler nucleopolyhedrovirus (NPV) Hyposidra talaca wlk. di perkebunan teh Gunung Mas Bogor. Larva Hyposidra talaca termasuk hama penting pada tanaman teh yang menyerang daun yang masih muda atau pucuk daun, hama ini dapat menyebabkan kerugian mencapai 40-100% jika tidak dilakukan pengendalian. Salah satu teknik pengendalian H. talaca yang sejalan dengan prinsip PHT yaitu memanfaatkan agen hayati seperti nucleopolyhedrovirus (NPV) yang memiliki inang spesifik, tidak membahayakan lingkungan, dapat mengatasi masalah keresistansian hama terhadap insektisida, dan selaras dengan komponen PHT lainnya. Namun, informasi mengenai karakterisasi molekuler HytaNPV masih sangat terbatas. Tujuan penelitian ini adalah mengetahui karakteristik molekuler melalui urutan nukleotida dan asam amino isolat NPV yang berasal dari H. talaca dengan menggunakan gen lef-8. Identifikasi molekuler dengan PCR terdiri beberapa tahap yaitu ekstraksi, amplifikasi dan elektroforesis DNA. Amplifikasi DNA HytaNPV menggunakan primer lef-8 berhasil mengisolasi gen lef-8 HytaNPV dengan ukuran fragmen DNA sekitar 770 bp. Analisis urutan DNA berdasarkan gen lef-8 menunjukkan bahwa isolat HytaNPV Bogor tergolong ke dalam spesies virus yang memiliki hubungan kekerabatan yang dekat dengan isolat NPV yang menyerang genus Helicoverpa yang berasal dari Brazil, Australia, Spanyol dan Belanda dengan nilai homologi nukleotida dan asam amino sebesar 98% dan 100%. Berdasarkan analisis filogeni, isolat HytaNPV masuk dalam grup yang sama dengan NPV yang menyerang genus Helicoverpa. Angka produksi tersebut masih tergolong rendah dibandingkan dengan produksi negara penghasil teh lainnya. Turunnya produksi sangat mungkin disebabkan oleh turunnya produktivitas tanama...

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Characterization of a Single-Nucleocapsid Nucleopolyhedrovirus of Thysanoplusia orichalcea L. (Lepidoptera: Noctuidae) from Indonesia

Cited by 18 publications

References 19 publications

Slow cell infection, inefficient primary infection and inability to replicate in the fat body determine the host range of Thysanoplusia orichalcea nucleopolyhedrovirus

Slow cell infection, inefficient primary infection and inability to replicate in the fat body determine the host range of Thysanoplusia orichalcea nucleopolyhedrovirus

Nucleopolyhedrovirus: scanning electron microscopy technique

Karakterisasi Molekuler Nucleaopolyhedrovirus (Npv) Hyposidra Talaca Wlk. Di Perkebunan Teh Gunung Mas Bogor

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