Biological Sciences
Baculovirus
Baculovirus is a type of virus that infects insects, particularly caterpillars. It is widely used in biotechnology as a vector for gene expression in insect cells. Baculoviruses have a unique life cycle, with two distinct forms: occlusion-derived virus (ODV) and budded virus (BV). They are of interest for their potential applications in pest control and as tools for protein production.
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eBook - PDFViral Genomes
Molecular Structure, Diversity, Gene Expression Mechanisms and Host-Virus Interactions
- Maria Laura Garcia, Victor Romanowski, Maria Laura Garcia, Victor Romanowski(Authors)
- 2012(Publication Date)
- IntechOpen(Publisher)
Part 1 Virus Genomes Organization and Functions 1 The Baculoviral Genome M. Leticia Ferrelli 1 , Marcelo F. Berretta 2 , Mariano N. Belaich 3 , P. Daniel Ghiringhelli 3 , Alicia Sciocco-Cap 2 and Víctor Romanowski 1 1 Instituto de Biotecnología y Biología Molecular, Facultad de Ciencias Exactas, Universidad Nacional de La Plata, CONICET 2 Laboratorio de Ingeniería Genética y Biología Celular y Molecular - Area Virosis de Insectos, Departamento de Ciencia y Tecnología, Universidad Nacional de Quilmes 3 Instituto de Microbiología y Zoología Agrícola, INTA Castelar Argentina 1. Introduction The molecular biology of Baculoviruses has drawn a great deal of interest due to the variety of applications of these viruses as: 1) agents for biological control of insect pests (Szewczyk et al., 2006); 2) vectors for expression of recombinant proteins in insect cells (Kost et al., 2005); 3) vehicles for gene transduction of mammal cells (Hu, 2006, 2008); and 4) display systems of recombinant epitopes (Makela et al., 2010). Baculoviridae is a family of insect-specific viruses, with more than 600 reported species, mainly isolated from Lepidoptera (butterflies and moths) and in some cases from Hymenoptera (sawflies) and Diptera (mosquitoes). Baculoviruses have circular, double-stranded DNA genomes ranging in size from approximately 80 to 180 kbp, depending on the species, that are predicted to encode for up to 180 genes. The viral genome associates with proteins forming a nucleocapsid. This structure is surrounded by a membrane envelope to form a rod-shaped virion (hence, the name of the Family: baculum is Latin for rod or stick).
eBook - PDFInsect Pharmacology
Channels, Receptors, Toxins and Enzymes
- Lawrence I. Gilbert, Sarjeet S. Gill(Authors)
- 2010(Publication Date)
- Academic Press(Publisher)
5 Baculoviruses: Biology, Biochemistry, and Molecular Biology B C Bonning , Iowa State University, Ames, IA, USA ß 2005, Elsevier BV. All Rights Reserved. 5.1. Introduction 125 5.1.1. Taxonomy 125 5.1.2. Nomenclature 126 5.2. Structure 126 5.2.1. Nucleocapsids 127 5.2.2. Budded Virus 128 5.2.3. Occlusion-Derived Virus 130 5.2.4. Occlusion Bodies 131 5.3. Life Cycle 132 5.3.1. Infection 132 5.3.2. Dissemination within the Host 135 5.3.3. Dissemination from the Host 137 5.4. Virus Replication 138 5.4.1. Early Gene Expression 138 5.4.2. DNA Replication 139 5.4.3. Late and Very Late Gene Expression 140 5.5. Effects on the Host 141 5.5.1. Virulence 141 5.5.2. Host Range 142 5.5.3. Survival Time and Yield 144 5.5.4. Sublethal Effects and Latent Infections 144 5.5.5. Resistance 145 5.6. Baculovirus Genomics 145 5.6.1. General Properties of Baculovirus Genomes 145 5.6.2. Baculovirus Evolution 146 5.6.3. Identification of Genes Involved in Virus–Insect Interactions 147 5.6.4. Identification of Genes that Have Undergone Adaptive Molecular Evolution 147 5.7. Conclusion 148 5.1. Introduction Initial interest in Baculoviruses stemmed from their potential use in insect pest management (Moscardi, 1999). Modern baculovirology is driven by genetic enhancement of their insecticidal properties (van Beek and Hughes, 1998; Bonning et al ., 2002), their use for the study of fundamental biological processes (Clem, 2001; Manji and Friesen, 2001), for protein expression (Jarvis, 1997), and for gene therapy (Ghosh et al ., 2002; Kost and Condreay, 2002). Addressing their limitations for these purposes allows for increased understanding of Baculovirus biology. As a result of the various applications of Baculoviruses, they represent the best studied of the invertebrate viruses.
eBook - ePub- Gordon D. O. Maramorosch(Author)
- 2018(Publication Date)
- CRC Press(Publisher)
Therefore, they have a minimal potential for damaging the environment. Yet, few of these viruses have been commercially developed for the control of pest insects of crop systems or forests. 6 - 8 One reason for the limited commercialization of Baculoviruses is that they are usually produced in living insects, which is a complex, expensive, and labor-intensive process. 6 The feasibility of automated scale-up of insect cell culture systems has been shown, which makes the in vitro production of viral pesticides possible. 6 - 24 Generating insect viruses in cell culture systems has the advantages over in vivo systems of being more highly controlled and reproducible, which therefore produces cleaner preparations. Increasing the purity of viral preparations generates higher activity/concentration ratios and minimizes the presence of contaminating biotic/abiotic agents capable of altering activity. 25 Since studies have shown that the Baculoviruses produced in vitro are comparable in virulence, morphology, and molecular structure to those produced in whole insects, 24, 26, 27 the optimization of cell culture systems will be an important accomplishment for the pest control industry. B Baculovirus Classification Baculoviruses have circular, covalently closed double-stranded DNA and are members of the Baculoviridae family. 2 - 5 This family is divided into two subfamilies, Eubaculovirinae and Nudibaculovirinae. 5 The Eubaculovirinae consist of the occluded Baculoviruses, which include the nuclear polyhedrosis virus (NPV) and granulosis virus (GV) genera. The NPV replicate within the nuclei of invertebrate cells and occlude virions (virus particles) within occlusion bodies (OB), also known as polyhedra. The virions, which are enveloped nucleo-capsids, can be packaged into the OB either as singly embedded viruses (SNPV) or multiply embedded viruses (MNPV), 2 with these two groups being considered subgenera
eBook - ePub- Opender Koul(Author)
- 2019(Publication Date)
- Academic Press(Publisher)
Bt plants to maintain an insect-susceptible population.Many researchers have found that Baculovirus can be used as an alternative biological control tool. For a number of reasons, Baculovirus biopesticide has successfully gained much interest over traditional pesticides (Inceoglu et al., 2006 ; Possee, 1997 ):- • Baculovirus is a natural pathogen.
- • It is highly target specific (their infectivity is restricted to only 600 insect species and closely related arthropods).
- • Vertebrates and other beneficial organisms are pathologically safe from Baculovirus infection.
- • Baculovirus can propagate in the insect host naturally and thus is easy to collect, harvest, and reapply.
- • Baculovirus be produced on a large scale for packaging, marketing, and selling purposes.
- • The Baculovirus-mediated pest management approach is much more environmentally friendly than that of chemical pesticides.
11.2.1 Structure and Life Cycle of Baculovirus
Baculovirus belongs to the family Baculoviridae, which includes the nuclear polyhedrosis viruses (NPVs) and granulosis viruses (GVs). It is a double-stranded DNA virus and is one of the most extensively studied insect pathogenic viruses (Inceoglu et al., 2006 ; Khurad et al., 2004 ) with most of the host belonging to the order Lepidoptera (500 species). But this virus can also be isolated from other insect orders such as Neuroptera, Trichoptera, Crustacea, Hymenoptera, Coleoptera, and Diptera (Possee, 1997 ). Fourth and fifth instar larvae are usually prone to Baculovirus infection (Fig. 11.1 ). The infection is polyorganotrophic, that is, at a time the virus can infect multiple tissues such as the epidermis, tracheal matrix, fat bodies, hemocytes, central nervous system cells, and pericardial cells (Adams and McClintock, 1991 ; Barrett et al., 1998 ; Torquato et al., 2006 ). The postinfection larval hemolymph becomes turbid and milky immediately due to the multiplication of the virus in large numbers (Chisti and Schaf, 1990 ). Both host protein and mRNA levels are completely shattered by 12–18 h postbaculoviral infection (hpi) with ultimate shutdown of the host protein synthesis and gene expression (Katsuma et al., 2005 ; Nobiron et al., 2003 ; Ooi and Miller, 1988
- R C Ray, O.P. Ward(Authors)
- 2008(Publication Date)
- CRC Press(Publisher)
Besides, these Baculoviruses are also inducer of dendritic cells' in vivo maturation and inflammatory cytokines' production. This study demonstrates a strong effect of Baculoviruses on the mammalian immune system (Hervas-Stubbs et al., 2007). In a different study, it has been shown that Baculovirus AcNPV is able to activate human monocyte-derived dendritic cells. This is one of the criteria in vaccination strategies targeting dendritic cells, since activation of dendritic cells is crucial for the initiation of an adaptive immune response (Schutz et al., 2006). Their effectiveness as powerful expression vectors stems from the fact that they have strong promoters driving high-level expression of foreign genes at a very late phase in the infection process in a eukaryotic environment. About 100 mg of protein per 10 9 cells can be obtained. Genes from a wide variety of sources ranging from viruses and bacteria to higher eukaryotes can be expressed using Baculovirus Expression Vector System (BEVS) technology. Being noninfectious for vertebrates, including human beings, their direct handling is safe. Even nontarget insects are protected from baculoviral infection, thus rendering Baculovirus an effective biopesticide in integrated pest management (IPM) programs where it is mandatory to conserve nontarget beneficial insects. Baculovirus BIOLOGY Baculoviruses ('baculum' meaning stick) are rod-shaped, extremely small (measuring 40-50 nm in diameter and 200-400 nm in length), double-stranded DNA viruses, which infect insects and other arthropods. In as early as 1527, during studies on 'jaundice disease' of silkworms, interest spurted in the direction of using Baculoviruses as pest control agents (Benz, 1986). These viruses are found to infect insects mostly of the order Lepidoptera, Hymenoptera and Coleóptera, and have also been found to infect members of the order Diptera, Thysanura, Trichoptera and from the class Crustacea.
eBook - PDF- Opender Koul, G. S. Dhaliwal(Authors)
- 2001(Publication Date)
- CRC Press(Publisher)
Baculoviruses may, in some cases, be the only effective viral- insect control agent available for controlling an insect species (Cunningham, 1988) and provide an avenue available to overcome specific problems, such as pesticide resistance. Even the use of two biologicals like viruses and Bt could lower the possibility for resistance development (Marrone, 1996). In addition to their safety to non-target organisms, the Baculoviruses are of particular interest because, (i) they cause mortality in the target insect population, (ii) geographically distinct populations of insects exhibit relatively uniform susceptibility to a virus, (iii) individual populations of insects do not acquire resistance readily upon continual virus pressure, (iv) can be formulated for easier application and long-term storage, (v) can be applied easily using methods similar to those employed in the application of chemical pesticides, and (vi) these are generally compatible with chemical pesticides. Collectively, these properties provide an overwhelming case for pursuing the development of Baculoviruses as biological (microbial) alternatives to chemical pest control (Miller, 1998). Taxonomic status of Baculoviruses Baculoviruses (Family: Baculoviridae) have only been isolated from invertebrates. Most examples have been found in insect species, but there are some reports of Baculoviruses, which are pathogenic to Crustacea. Baculovirus infections have been described in over 700 species of invertebrates including Lepidoptera, Hymenoptera, Diptera, Coleoptera, Trichoptera, Thysanura and Neuroptera besides Crustacea (Murphy et al., 1995). Until 1995, family Baculoviridae was subdivided into two subfamilies: Eubaculovirinae, which included the occluded nuclear polyhedrosis virus (NPV) and granulosis virus (GV); and Nudibaculovirinae, encompassing the non-occluded Baculoviruses.
eBook - PDF- Karl Maramorosch(Author)
- 2012(Publication Date)
- Academic Press(Publisher)
IV. Physical, Biological and Chemical Characteristics This page intentionally left blank THE STRUCTURE AND PHYSICAL CHARACTERISTICS OF BaculovirusES D. C. KELLY Natural Environment Research Council Institute of Virology Mansfield Road> Oxford Baculoviruses are large structurally complex viruses. Their very complexity has long intrigued electron microscopists and most evidence concerning the structure of the viruses de-rives from electron microscope studies. Other biophysical techniques have been applied to study Baculovirus architecture and, in some instances, these approaches have proved signifi-cant insights into the structural organization of the viruses. Baculovirus particles are, in most cases, packaged into large protein crystals variously termed as polyhedra, capsules, granules, whole inclusion bodies or polyhedral inclusion bodies. The occlusion or packaging of virus particles ap-pears to be a genetically defined trait with some virus types containing just one virus particle per crystal whereas other types package many virus particles within a protein crystal. Baculoviruses have been classified according to their structure into nuclear polyhedrosis viruses, granulosis viruses, and nonoccluded Baculoviruses (Matthews, 1982). Non-occluded Baculoviruses, typified by a Baculovirus from Oryctes rhinoceros (Payne, 1974) fail to synthesize polyhedra (Huger, 1966; Kelly, 1975) and, consequently, never become packaged in protein crystals. Granulosis viruses (GVs) typically com-prise small crystals (capsules or granules—hence the term granulosis) containing a solitary virus particle. Nuclear polyhedrosis viruses (NPVs), on the other hand, contain numer-ous virus particles within a given polyhedron. They take VIRAL INSECTICIDES Copyright © 1985 by Academic Press, Inc. FOR BIOLOGICAL CONTROL 4 6 9 All rights of reproduction in any form reserved. ISBN 0-12-470295-3 470 D. C. KELLY their name from the presence of polyhedra in an infected cell nucleus.
eBook - PDF- Marc H.V. van Regenmortel, Brian W.J. Mahy(Authors)
- 2010(Publication Date)
- Academic Press(Publisher)
This article focuses primarily on the lepidopteran NPVs (the proposed alphaba-culoviruses). Lepidopteran NPVs are the most widely studied of the Baculoviridae , primarily due to the availability of insect cell culture systems that are permissive for infection. A number of lepidopteran NPVs have been developed as bioinsecticides, and in addition lepidopteran NPVs are used widely as protein expression vectors and more recently as vectors for mammalian cell transduction. Also, there is a great deal of recent interest in developing certain lepidop-teran NPVs as mammalian gene therapy vectors. Life Cycle Like all Baculoviruses the lepidopteran NPVs contain genomes of double-stranded circular DNA, with genomes 434 Baculoviruses: Molecular Biology of Nucleopolyhedroviruses ranging in size from 111 to 168 kbp. Two distinct types of virions are produced in the life cycle: the budded virus (BV) and the occlusion-derived virus (ODV) ( Figure 1 ). BVs generally contain a single nucleocapsid and obtain an envelope by budding from the plasma membrane of the infected cell. In contrast, ODVs are formed in the nucleus, acquire a membrane that is derived from the inner nuclear membrane, and virions consist of either single (S) or multiple (M) nucleocapsids per envelope. The S and M designation (SNPV and MNPV) does not appear to hold any taxonomic significance above the species level but this characteristic is clearly associated with certain viral species. As the infection progresses, ODVs are assembled in the nucleus and become embedded in a paracrystalline proteinaceous matrix to form occlusion bodies (OBs) (also known as polyhedra) that can range in size from 0.15 to 15 m m. A single matrix protein called polyhedrin makes up the majority of the mass of the polyhedra or OB. The OB as a whole is surrounded by a carbohydrate-protein layer known as the polyhedral envelope.
eBook - ePubAnimal Cell Technology
From Biopharmaceuticals to Gene Therapy
- Leda Castilho, Angela Moraes, Elisabeth Augusto, Mike Butler(Authors)
- 2008(Publication Date)
- Taylor & Francis(Publisher)
Another aspect to be considered is the need for increased production. At the moment, the production of the bioinsecticide is accomplished by harvesting infected caterpillars in the areas infected or by growing the caterpillars in a laboratory. In the first case, there is great variability in productivity from year to year, since the production depends on the insect abundance during each harvest, which varies with multiple factors. However, there has been considerable progress in the production of the virus under laboratory-controlled conditions (Moscardi and Santos, 2005). In March of 2005 EMBRAPA finished the construction of a pilot plant with the capacity to inoculate about 30 000 caterpillars per day, and another institution, the COODETEC (Cascavel, PR), is enlarging its production capacity to 600 000 caterpillars per day. However, the caterpillars have to be fed with artificial diets and the cost of production of a dose of the biopesticide, based on Baculovirus anticarsia, using raw material produced locally, is approximately 90% higher than that obtained by direct harvesting from the field. The use of alternative formulations, such as substituting agar for carrageen as a gelling agent, has reduced production cost, making it more economically feasible to produce the bioinsecticide in the laboratory (Santos, 2003). However, this type of process has high labor demands. Large-scale virus production in cell culture in bioreactors is a desirable development, since it allows virus multiplication in a smaller area and it reduces labor requirement, in comparison with in vivo production in the laboratory.19.2 Baculovirus as a bioinsecticide: mechanism of action
Baculovirus belongs to the Baculoviridae family, which has two different genera: Nucleopolyhedrovirus (NPV) and Granulovirus (GV). Baculo- virus exists as two phenotypes designated extracellular virus or budded virus (BV) and the occlusion bodies, also known as OBs, COPs, GDP, or polyhedra (Figure 19.1). The occlusion bodies are structures that are formed inside the nucleus of the infected insect cells by Baculovirus (Friesen and Miller, 2001). These are highly resistant and able to stay viable in the environment for several years under different climatic conditions. The protected viral particles (virions) are known as ODVs (occlusion- derived virus) (Harrap, 1972).The protein contained in the GV OBs is called granulin and that in the NPV is called polyhedrin (Rohrmann, 1999; Winstanley and O’Reilly, 1999). GVs are smaller than NPVs and possess rounded OBs with just one virion occluded (Winstanley and O’Reilly, 1999). NPVs present a polyhedral form and they can be multiple type (MNPV) (Figure 19.1A) or simple type (SNPV) (Figure 19.1B), depending on the number of capsids per virion (Rohrmann, 1999). Since most of the Baculovirus used as biopesticidas are NPVs, in this chapter the OBs will be referred as polyhedra.
- Sadettin Ozturk, Wei-Shou Hu, Sadettin Ozturk, Wei-Shou Hu(Authors)
- 2005(Publication Date)
- CRC Press(Publisher)
Environmental conditions, such as solar radiation and rainfall can rapidly inactivate Baculoviruses even in their occluded form. To pre- vent such a problem, various materials that act as UV-shielding agents, including lig- nin derivatives, starch, and fluorescent brighteners, have been employed in product formulations to further protect the virions (260,261). Temperature, application tim- ing, leaf distribution, plant architecture, and virulence differences due to genetic variability must also be considered for proper performance of Baculoviruses in the field (261). r-Baculoviruses One of the main limitations of Baculovirus pesticides is their slow lethal action, which can be delayed between 5 and 15 days postinfection before killing the insect pest. During such time, larvae can continue feeding and consequently cause substan- tial damage to crops (6,8,275). A solution to such a problem, initially attempted by Carbonell et al. (277), has been the development of genetically modified baculo- viruses. In r-Baculoviruses, genes that exert a deleterious effect on the insect are placed under the control of the polh promoter or others. To date, various genes have been tested (81,256,275,278,279). These include those for toxins of several scorpions (Buthus eupeus and Androctonus australis), mite (Pyemotes tritici), wasp, and bacteria (Bacillus thuringiensis delta-endotoxin), as well as hormones (Manduca sexta diuretic hormone), enzymes (H. virescens juvenile hormone esterase and ecdysteroid UDP– glucosyl transferase), and other proteins (viral enhancing factor). Depending on the gene selected, diverse effects on the larvae are obtained, including reduction of hemolymph volume, feeding cessation, blackening, paralysis, tremors, dorsal arch- ing, premature melanization, and low weight gain (6).
eBook - ePub- S.C. Makrides(Author)
- 2003(Publication Date)
- Elsevier Science(Publisher)
However, it is a disappointment for those investigators that would like to exploit the inherent replication deficiency of the virus and use it for in vivo gene delivery applications. It has been shown that Baculovirus is capable of delivering genes in complement deficient animals, or in situations where the virus is protected from exposure to complement. Thus, an active area of investigation in this field is to find methods to modify the physical properties of the virus to render it resistant to complement inactivation, and turn the vector into an in vivo gene delivery tool [ 18, 22, 23 ]. 6 Applications of the Baculovirus mammalian gene delivery vector Several advantages of the Baculovirus system are listed in Table 1. The items listed in the right column are general characteristics of the system that make Baculoviruses very desirable for gene expression using either insect or mammalian cells. In the column on the left are characteristics of the virus that are advantageous for its application as a mammalian cell gene delivery vector. The versatility provided by the wide variety of cell types that are transduced by the virus, as well as its utility for transient or stable gene expression, have already been discussed. The level of protein expression obtained by Baculovirus transduction can be modulated not only by chemical means (i.e., butyrate or Trichostatin A), but also by the multiplicity of virus used to treat a cell population [ 12, 13, 24 ]. Additionally, treatment with the virus, even at very high multiplicities, does not produce overt cytotoxicity in transduced cultures [ 18 ], thus providing a gentle method to introduce recombinant genes into mammalian cells
eBook - ePub- Fernando E. Vega, Harry K. Kaya(Authors)
- 2011(Publication Date)
- Academic Press(Publisher)
As a consequence, secondary pest outbreaks and primary pest rebounds that are consequences of the use and abuse of chemical insecticides are avoided (Grzywacz et al., 2010). The narrow host range also makes Baculoviruses a good choice for controlling a pest when it is desired to conserve other non-target lepidopteran species (Reardon et al., 2009). However, the Baculovirus narrow host range makes it unappealing for situations where more than one pest species needs to be controlled. If additional pest species are not sufficiently controlled by other factors, then additional pesticides would be need to be purchased. The low sales potential of a pesticide that targets one or two pests is expected to discourage investment in development of Baculovirus-based pesticides. As a consequence, the Baculoviruses that have been registered and sold as pesticides are viruses that efficiently infect and kill pest species that by themselves are of great economic significance, such as C. pomonella, and not against pests that are part of a pest complex or that are pests only on an intermittent basis. Broad host-range Baculoviruses such as AcMNPV would appear to offer a solution to the problems posed by host range, but while such viruses are able to infect a wide variety of species in laboratory bioassays, only a few of these species are susceptible at concentrations that translate to economically feasible application rates in the field (Black et al., 1997). Survival Time Baculovirus-infected larvae live for days to weeks after infection and continue to feed and cause crop damage during this time. This issue has been resolved through the development of fast-killing recombinant Baculoviruses (see Section 4.4.3), but recombinant Baculoviruses currently are not in use in pest control programs
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