VIRUS ENTOMOPATOGEN By Irda Safni Pendahuluan Virus berasal
VIRUS ENTOMOPATOGEN By Irda Safni
Pendahuluan
“Virus” berasal dari bahasa Latin, yang artinya racun dan selalu berhubungan dengan penyakit dan kematian. Definisi virus (modern): biosistem paling dasar yang menunjukkan keragaman makromolekuler yang mampu berkembang biak dan membelah sendiri, tapi kurang mampu bereaksi terhadap faktor lingkungan, tidak dapat bergerak sendiri, dan tidak dapat menggunakan sumber energi metaboliknya sendiri bertolak belakang dengan definisi makhluk hidup
Struktur Virus 1. Mengandung DNA atau RNA 2. Mengandung protein coat (capsid) 3. Reseptor – pada capsid yang menentukan sel inang apa yang dapat diinfeksi dan bagaimana virus menginfeksi sel.
Struktur Virus ØInner core nucleic acid Mengandung bahan genetik (DNA atau RNA) ØOuter core nucleic acid Mengelilingi virus (Capsid) Mengandung reseptor
Struktur yang tidak dijumpai pada semua virus Amplop Tail, end plate, tail fibres • mengelilingi beberapa virus hewan • Terbuat dari membran inang • Hanya pada Bacteriophage
Siklus Hidup Virus • Untuk dapat bertahan hidup, virus harus mampu melakukan sebagai berikut: ▫ 1. Menemukan sel inang yang di dalamnya virus dapat bereplikasi ▫ 2. Mengikat pada sel ▫ 3. Memasuki sel ▫ 4. Melepas genomnya agar dapat bereplikasi ▫ 5. Mereplikasi genomnya ▫ 6. Transkripsi dan translasi protein virusnya ▫ 7. Membungkus genom dan proteinnya ▫ 8. keluar dari sel
Siklus Hidup Virus
Siklus Hidup Virus yang tidak memiliki Amplop
Klasifikasi Virus Apa tujuan klasifikasi? Untuk membuat susunan struktural untuk perbandingan yang mudah. Untuk berkomunikasi untuk keputusan taksonomi bagi komunitas Internasional ahli virologi. Untuk memudahkan prediksi properti virus-virus baru. Kemungkinan hubungan evolusioner
Properti yang digunakan pada Taxonomi Virus Properti Virion Morfologi properti virion Ukuran; bentuk; ada tidaknya amplop atau peplomer; simetri capsomer & struktur Properti fisik virus Massa Molekuler Buoyant density Sedimentation coefficient Kestabilan p. H, kestabilan solvent, kestabilan radiasi, kestabilan detergen Kestabilan kation (Mg 2+, Mn 2+, Ca 2+)
Properti yang digunakan pada taksonomi virus Properti genom Jenis asam nukleat Strandedness of n/acid Linear atau circular Sense: +ve or –ve or ambisense Jumlah segmen Ukuran genom atau segmen genom Presence or absence and type of 5’ terminal cap Presence o or absence of 5’terminal covalently- linked polypeptide Presence o or absence of 3’terminal poly (A) tract (or other specific tract) Perbandingan sekuensing nukleotida
Properti protein Number; size; functional activities and Amino acid sequence comparisons Lipids Presence or absence; Nature Carbohydrates Presence or absence; Nature Genome organization and replication Genome organization Strategy of replication of nucleic acid Characteristics of transcription Characteristics of translation and post translational processing Sites of accumulation of virion proteins, site of assembly site of maturation and release Cytopathology inclusion body formation
Antigenic properties Serological relationship Mapping epitopes Biological properties Host range, natural & experimental; pathogenicity, etiology Tissue tropism, pathology, histopathology, Mode of transmission in nature; vector relationship; Geological distribution
Criteria demarcating different virus taxa Order Common properties between several families including Biochemical composition Viurs replication strategy Particle structure General genome organization Family Common properties between several families including Biochemical composition Virus replication strategy Nature of particle structure Genome organization
Criteria demarcating different virus taxa Genus Common properties with a genus including: Virus replication strategy Genome size, organization and/ or number of segments Sequence homologies Vector transmission Species Common properties within a including: Genome rearrangement Sequence homologies Serological relationship Vector transmission Host range Pathogenicity Tissue tropism Geographical distribution
International Committee on Taxonomy of Viruses (ICTV) telah menyetujui: 3 ordo, 73 famili, 9 subfamili, 287 genus, dan 1950 spesies virus termasuk viroid, virusoid, (satelit virus), dan prion Ordo diakhiri dengan “ales” Famili diakhiri dengan “viridae” Subfamili diakhiri dengan “virinae” Virus bakteri disebut “Bacteriophage atau phage” (dengan beberapa pengecualian)
Virus Classification I - the Baltimore classification Based on genetic contents and replication strategies of viruses. According to the Baltimore classification, viruses are divided into the following seven classes: 1. ds. DNA viruses 2. ss. DNA viruses 3. ds. RNA viruses 4. (+) sense ss. RNA viruses (codes directly for protein) 5. (-) sense ss. RNA viruses 6. RNA reverse transcribing viruses 7. DNA reverse transcribing viruses where "ds" represents "double strand" and "ss" denotes "single strand".
How are viruses named? • Based on: - The disease they cause poliovirus, rabies virus - The type of disease murine leukemia virus - Geographic locations Sendai virus, Coxsackie virus - Their discovers Epstein-Barr virus - How they were originally thought to be contracted Tobacco mosaic virus, dengue virus (“evil spirit”), influenza virus (the “influence” of bad air) - Combinations of the above Rous Sarcoma virus
Latin binomials were proposed first by Holmes in 1939 Various other schemes proposed between 1940 and 1966 International Committee on the Nomenclature of Viruses (ICNV) (1966) Naming and cataloguing of viruses. Renamed: International Committee Taxonomy of Viruses (ICTV) in 1973.
v Virus menginfeksi manusia, hewan, tumbuhan, dan mikroorganisme, yang menimbulkan pengaruh yang merugikan. v Tetapi pengaruh yang merugikan juga dapat digunakan untuk keperluan manusia yang bermanfaat. v Beberapa virus dari famili Baculoviridae digunakan secara ekstensif untuk produksi protein yang berguna untuk keperluan penelitian, selain juga potensial digunakan di bidang industri pengobatan.
Virus Entomopatogen
Penyakit yang disebabkan virus entomopatogen mulai diketahui sejak abad ke-16. Penyakit yang disebut Jaundice o graserrie, sekarang diidentifikasi sebagai nucleopolyhedrosis, ditemukan pada ulat sutra (Bobyx mory) oleh Vida pada tahun 1524 dan kemudian juga diisolasi dari lebah madu (Apis mellifera). Pada tahun 1856, dua orang ahli Italia (Maestri dan Cornalia) menjelaskan occlusion bodies (OBs) ulat sutra nucleopolyhedrosis. Pada tahun 1926 Paillot mendeskripsikan granulovirus (GVs) pertama sekali. Pada tahun 1934 Ishimori menjelaskan jenis baru polyhedrosis di dalam ulat sutra OBs dibentuk didalam sitoplasma sel yang diinfeksi (bukan pada asam nukleat) sekarang dikenal dengan cypovirus.
Sejak tahun 1950 s/d 1970, Steinhaus dan koleganya menguji baculovirus sebagai agens hayati di lapangan dengan mengaplikasi nucleopolyhedrovirus (NPV) untuk mengendalikan ulat alfalfa (Colias eurytheme Boisduval; Lepidoptera: Pieridae). Bioinsektisida komersil berbahan aktif virus pertama dikembangkan pertama sekali pada tahun 1975 oleh Perusahaan Sandoz (dengan nama dagang Elcar) untuk mengendalikan Heliothis/Helicoverpa Lepidoptera: Noctuidae). Selama tahun 1979 s/d 1980, penemuan penting pada genetika virus entomopatogen, khususnya baculovirus. Hingga saat ini studi genetika virus entomopatogen difokuskan pada studi genom lengkap telah ada 29 sekuensing genom lengkap virus entomopatogen.
Virus yang Menginfeksi Invertebrata Virus DNA Double stranded DNA - Poxviridae - Iridoviridae - Baculoviridae: NPV & GV - Polydnaviridae Single stranded DNA - Parvoviridae Virus RNA Double stranded RNA - Rheoviridae: Cypovirus Single stranded RNA (-) - Rhabdoviridae - Bunyaviridae Single stranded RNA (+) - Picornaviridae, Togaviridae, Tetraviridae, Flaviridiae, Nodaviridae
Tabel 1. Kelompok Virus Entomopatogen FAMILY NUCLEIC ACID Baculoviridae ds. DNA Reoviridae ds. RNA Poxviridae ds. DNA Iridoviridae ds. DNA Parvoviridae ss. DNA Picornaviridae ss. RNA Ascoviridae ds. DNA Polydnaviridae ds. DNA Rhabdoviridae ss. RNA Nodaviridae ss. RNA Rhabdoviridae ss. RNA NON-CLASSIFIED RNA VIRUSESs Divided genome ss. RNA Nodaurelia ss. RNA Kelply group ss. RNA 5 -virus group ss. RNA Minivirus ss. RNA Ovoid virases ss. RNA Drosophila X Virus ds. RNA ds= double-stranded, ss= single-stranded. NUCLEOCAPSID SIMETRY Baciliform Isometric Ovoid Icosahedral Isometric Spherical Allantoid Ovoid Baciliform Icosahedral Baciliform OCCLUSION BODY + + + - Isometric Isometric Ovoid Isometric -
Klasifikasi Virus Entomopatogen v Klasifikasi virus entomopatogen dibuat sesuai dengan peraturan the International Commitee on Taxonomy of Viruses (ICTV). v Virus entomopatogen diklasifikasikan menjadi 12 famili. v Kriteria untuk menyusun klasifikasi disusun berdasarkan keragaman virus serangga, seperti: • Jenis bahan genetik (seperti singe- atau double-stranded DNA, singe- atau double-stranded RNA, positive atau negative strand) • Morfologi dan ukuran virion (seperti icosahedral, rodshaped, dll. ) • Kehadiran amplop di sekeliling virion. • Kehadiran occlusion body melingkupi virion. • Inang dan kisaran inang.
Klasifikasi Virus Entomopatogen v Kriteria utama sekuensing bahan genetik virus serangga bukan hanya menentukan diskriminasi antar spesies virus, akan tetapi juga membangun hubungan evolusioner antar virus dalam kelompok yang sama.
Penamaan Virus Entomopatogen Virus entomopatogen diberi nama dengan cara singkatan (akronim). Sesuai dengan inangnya, dan kelompok virusnya Contoh: Autographa californica multiple nucleopolyhedrovirus Ac. MNPV Semua nucleopolyhedroviruses dinamai NPV, granuloviruses dinamai GV, entomopoxviruses: EPV, iridoviruses: IV, dan cytoplasmic polyhedrosis viruses (cypoviruses) : CPV. Virus entomopatogen sangat tinggi keragamannya, tapi hanya beberapa kelompok saja yang dijumpai pada populasi serangga dan menunjukkan potensi sebagai agen pengendali hayati.
6 Baculoviruses Spodoptera littoralis 2 microns From Hunter-Fujita et al
7 Baculoviruses Mode of action From Hunter-Fujita et al
8 Baculoviruses Susceptibility of Alternative Hosts Found only in invertebrates No member of the family is known to infect plant or vertebrate Most have narrow host insect range, and infectivity is restricted to the original host genus or family
9 Baculoviruses Toxicity studies - mammals Toxicity test results from 1970 s/80 s of 29 NPVs indicated no toxicity or pathogenicity. Doses were generally 10 – 100 x the “per acre” (1 acre = 0. 45 ha) field rate equated to a 70 kg person. Heliothis zea NPV most extensively tested for toxicity in humans and led to registration of “Elcar” by Sandoz in USA.
10 Baculoviruses Toxicity studies – mammals cont. No effects of Hz. NPV found in: Acute toxicity-pathogenicity tests in mouse, rat, guinea pig, rabbit, monkey and man at 6 x 109 – 3 x 1012 OB / kg. Skin irritation sensitivity tests in guinea pigs, rabbits and man at 106 and 107 OB / mm 2 skin. Eye irritation tests in rabbits with 105 and 2 x 106 OB / eye Subacute toxicity-pathogenicity tests and subcutaneous injection into mice, rats, dogs and rhesus monkeys. Teratogenicity and carcenogenicity studies in rats and mice at 109 – 3. 5 x 1012 OB / kg. Similar but less extensive results for many other NPVs from the 1970 s/80 s
11 Baculoviruses Toxicity studies – wildlife Birds Able to pass NPV through the alimentary tract unaffected No deleterious effects Aquatic organisms No adverse effects Beneficial insects No direct effect on parasitoids, predators and pollinators Indirect effects on parasitoids resulting from host death
12 Baculoviruses Pathology studies Toxicity tests designed for testing effects of chemicals on vertebrates are insufficient Results reported in Gröner (1986) indicate no virus induced antibody production in test mammals and chicken. No cytogenetic effects of baculoviruses in mammalian cells either in vivo or in vitro.
13 Baculoviruses Virus-cell interactions in vitro Ac. NPV inoculated into vertebrate cells can be takenup and the degree of up-take depends on cell type, temperature, time and viral phenotype. BUT, none of the human and nonhuman vertebrate lines tested showed evidence of viral replication. NPVs unable to activate retroviruses in mammalian cell lines
15 Baculoviruses a list of the baculoviruses regulated as pesticide active ingredients by the US EPA Office of Pesticide Programs as of May 2005 Anagrapha falcifera NPV Cydia pomonella GV Douglas fir tussock moth NPV Gypsy moth NPV Helicoverpa zea NPV Indian meal moth GV Mamestra configurata NPV (pending) Spodoptera exigua NPV
16 Baculoviruses – US EPA fact sheet III. ASSESSING RISKS TO HUMAN HEALTH These viruses infect only the target insect larvae and closely related species. Toxicity tests show that the viruses pose no risk to the public. Workers wear protective clothing to prevent possible irritation from handling and applying the product. IV. ASSESSING RISKS TO THE ENVIRONMENT Tests show that the GV and NPVs that EPA has registered as pesticide active ingredients specifically infect only certain species of moth larvae. The viruses do not harm other organisms, including plants, beneficial insects, other wildlife, or the environment. These viruses occur naturally in their insect hosts.
17 Cypoviruses: Mode of action Polyhedra ingested and dissolved in larval midgut Virions released and attach to midgut columnar cells Viral core enters cell cytoplasm RNA transcription and replication RNA occluded in capsules Virus capsules occluded by virogenic stroma to form occlusion bodies
18 Cypoviruses (Rheoviridae) No CPV has been found infecting vertebrates or plants (Belloncik, 1989) Dendrolimus spectabilis CPV registered in Japan in 1974. Safety test results generally negative. – Katagiri, K. (1981) Pest control by cytoplasmic polyhedrosos viruses. In: Microbial control of pests and plant diseases 1970 -1980. (Ed Burges, H. D. ) Academic Press.
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