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Rabies Vaccine Production from Different Viral Infected Cell Line and Vaccine Production Praveen Gaurav Singh and S. K. Singh ABSTRACT- Bullet-shaped rabies virus is classified as Lyssavirus genus in Rhabdoviridae family which transferred by animals like Dogs, Raccoons, Skunks, Foxes, Coyotes, Bat to Human. Virus has a single-stranded, negative-sense RNA. Among of 7 member of Lyssavirus genus, only serotype 1 infects humans. Human rabies have two form; furious and Paralytic rabies. The first rabies vaccine is developed by two French scientists, Louis pasture and Emile Roux in 1885. Various types of rabies virus strain i.e. CVS, HEP, PV, ERA, PM, CPZ, BOLIVAR etc. are grown and isolated from cultured cell i.e. Pig kidney cells, Chick embryo fibroblasts, Dog kidney cells, BHK-21 cells, Human diploid cells etc. The preparation of rabies vaccines involve production of seed virus followed by preparation of final vaccine that include live attenuated vaccine and inactivated vaccine (Vaccines containing animal brain tissues, Avian vaccines, and Cell-cultured vaccines).These strains are inactivated by acetylethylenimine, ethylenimine, β-propiolactone or ultraviolet irradiation. The potency of inactivated vaccine is determined by single radial immunodiffusion and NIH potency test. In dog inactivated PV strain by ethylenimine have higher antigenic value and potency. BHK-1 is frequently used for the production of vaccine using suspension culture in bioreactor ranging from 20 to 1000 lt. and potency rate is 6 IU for 20 lt. In Europe and USA, human fibroblast is used in Human diploid cell culture vaccine (HDCV). Key words: Rabies virus; Rabies vaccine; Rabies strain; Rabies infection; Cell lines; NIH potency test; Vaccine production. Introduction
Single-stranded negative-sense RNA of approximately 12 kb rabies virus has 12 viral species (RABV; geno-type I) belongs to the genus Lyssavirus in the family Rhabdoviridae. These are responsible for greater than 55,000 human deaths annually throughout the world. Humans are infected by RABV through the saliva of infected carnivores and bat in America. In Latin America, common vampire bat (Desmodusrotundus) and several other Neotropical bat species are major cause of rabies infection [5], [48], [49]. In Ontario, Canada, more than 54,800 animals were infected with rabies, from 1954 to 2006 (no including clinical cases). Red foxes (Vulpesvulpes; 46%) and striped skunks (Mephitis mephitis; 20%) are infected with an Arctic variant of rabies virus from the majority (66%) cases [36]. • Author Praveen Gaurav Singh is currently pursuing master of technology program in biotechnology engineering in Lovely Professional University, Chiheru, Kapurthala, Punjab, India, PH-9041382361. E-mail: singhpraveengaurav@gmail.com • Co-Author S. K. Singh is currently Assistance Professor in School of Biosciences and Biotechnology in Lovely Professional University, Chiheru, Kapurthala, Punjab, India, PH-01123456789. E-mail: author_name@mail.com
In China surpass 3,000 human rabies cases per year reported in the recent year [51]. In United States and Puerto Rico, 6,292 cases of wildlife rabies were reported in 2004. Coyotes (Canislatrans), gray fox (Urocyoncinereoargenteus), red fox (Vulpesvulpes), and skunks (Mephitis spp. and Spirogolaputorius) raccoons (Procylon lo-tor), bats (Lasionycterus spp., Myotis spp., and Pipstrellus spp.) are involved in these rabies infection [35]. Mostly deaths due to rabies infection were reported for near about 40% death of children aged below 15 years or 15 years in Asia [52]. In eastern Ontario, 135 cases are found with raccoon rabies infection after the implementation of oral virus-rabies glycoprotein (VRG) vaccine between 1999 and 2005 [40]. Rabies infected dogs are responsible for more than 90% of human death [47]. The World Health Organization (WHO) estimated that 40,000-70,000 death every year in all over world due to the rabies virus [31]. 2 Rabies Virus Structure and Infection 2.1 Structure In Rhabdoviridae, such as RV and vesicular stomatitis virus (VSV) the nonsegmented negativestranded RNA is enclosed within bullet-shaped envelope structure [10], [14]. The 12-kb genome encodes only five different type protein closely
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associated with nucleoprotein or nucleoocapsid proteinn (N), matriix protein (M)), phosphoprootein (P), virall polymerasee (V) and the single s transmem mbrane proteinn 0], [12], [47].T There are ninee or glycoprrotein (G) [10 moter, bind w with the sugarr nucleotide per N prom NA by virall phosphate backbone of the RN nucleocapssidprotein (N).. This N-RNA template havee important role for repliication and trranscription off R RNA and are catalyzed by b the viral RNA-dependent t L). Polymerrase cofactorr RNA poolymerase (L (phosphoprotein (P)) hellped in the binnding of RNA-ymerase (L) to N-RNA A dependent RNA poly wn as internnal core orr template.thhis is know ribonucleooprotien (RNP)) complex andd formed thee e compponent, virall RV virionn with the external lipoproteinn envelope whiich is composeed of M and G protein[4], [12], [47]. Th he RV M and G protein aree budding and assemblyy and M aree essential for f n (NC). The G surroundinng a helical nucleocapsid protein plaayed important role in the inteeraction with a d pH-dependeent membranee cellular reeceptor(s) and hen the amounnt of virions iss fusion. If G is deleted th aree G reduced about 30-fold. Spike G-proteins ve at least one subunit of thee amphiphiliic proteins hav oligomericc protein. This is inserted iinto the lipidd bilayer off the viral env velope anchorring the spikee The spike G-proteein involved inn to the membrane. m the stimulaation of virus--neutralizing aantibody as ann Rabies viruss antigen [10], [12], [14], [55].R he ERA strainiis a 505 aminoo glycoproteein (RGP) of th acid type I membrane gllycoprotein thaat oligomerizess mers and that contains three A Asn-X-Ser/Thrr into trimm potential N-glycosylatio on sites (seqquons) in thee extracellular domain [25]]
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2.2 Structure S of N-RNA N Coplex
Fig.2. Ribbon Diiagram of N-R RNA of Rabies Virus [50]. In thhis ribbon diagrram of N-RNA A, RNA is shoown as blackk coil with different color oof N-protomerr. The A ring structuure as 11-nuucleotide oliggomer N-RNA wed from the boottom in Ribbon diagram [50]] view
Fig.33. The helical ssecondary struccture of N-prottomer D are representted by (Ribbbon Diagram). NTD and CTD dark and light blue color respectivvely [50].
Fig.1. Show w the rabies virus v envelope with Matrix (M) proteinn, Glycoproteiin (G), Nucleeocap protein and lipid [54].
NTD D (N-terminall core domainn) have 32 too 233 residdues of aminoo acid. Foldeed helical strructure consiist 6 helices annd connected bby a large loopss.CTD (C-teerminal core doomain) have 2336 to 356 aminno acid residdues. CTD is ccomposed of 111 helices joinned by loop tighter than NT TD [50].
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2.3 Infection Rabies viruses can cause the fatal diseasedue to the replication in central nervous system, so known as neurotropic virus. RV invades the central nervous system (CNS) from peripheral sites, due to this RV known as neuroinvasivenes. This process expressed thepathogenicity of rabies virus [47]. Rabies or Rabies related virus caused hindlimb paralysis in immunocompetent in mice and hamsters [15].
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reeze-dried fixed-virus strain Flury LEP grown in cultures of chicken fibroblasts. Both vaccines induced the Induces neutralizing antibodies and cellular immunity. Imovax are stored at 35° to 46°F. Do not freeze and RabAvert are stored at 36° to 46°F. Protect from light [55].
Many types of commercial product of rabies vaccine are available.
2.4 Virus Survival
References
Rabies virus are survived at 5 degrees C for 114 hours, when spread in a thin layer onto surface like metal, leaves or metal. Rabies virus is infected the mammalian for 24 hours at 240C when spread o glass and leaves and 48 hours on metal. But due to exposure of sunlight virus are inactivated within 1.5 hours at 300C [54]
[1] R. Fischman et al. “Recovery from Experimental Rabies by Adoptive Transfer of Immune Cell”. J. gen. Virol. (1981), 56, 25-3l.
3 Rabies Vaccines 3.1 OspA lipoprotein OspA lipoprotein is found on outer surface protein of Borrelia burgdorferi sensu stricto ZS7 and expressed by Escherichia coli. OspA lipoprotein contains lipoprotein OspA and use as vaccine against Lyme diease. Single polypeptide chain of OspA consist of 257 amino acids with lipds covalently bonded to N terminus and is conjugated with aluminum hydroxide as an adjuvant. The protein chemical formula of this vaccine is C1198H2012N322 - -O422S2 with average molecular weight of 27743.1000. In vitro condition, this vaccine has 1.2 hour half-time (Tested in mammalian reticulocytes). The hydrophobicity and isoelectric point of this protein is -0.65 and 6.72 respectively [53]. 3.2 Imovax (Human Diploid Cell) & RabAvert (Chicken Fibroblasts) Imovax- The imovax (trade name) are present in the form of powder for injection. PM-1503-3M strain of rabies virus (freeze-dried suspension) are cultured in human diploid cell culture for vaccine production and available at least 2.5 international unit rabies antigen/ml. RabAvert- The RabAvert are present in the form of powder for injection. RabAvert are produced by the
[2] Elizabeth Loza-Rubio et al. “Nucleocapsid of Rabies Virus Improve Immune Response of an Inactivated Avian Influenza Vaccine”. Vet Res Commun, DOI 10.1007/s11259-009-9206-7. [3] Celis et al. “Recognition of Rabies and RabiesRelated Viruses by T Cells Derived from Human Vaccine Recipients”. Journal of Virology, Sept. 1988, p. 3128-3134. [4] Ivan Ivanov et. al, “Structure of the Dimerization Domain of the Rabies Virus Phosphoprotein”. Journal of Virology, Apr. 2010, p. 3707–3710. [5] Rupprecht et al. “Oral Immunization and Protection of Raccoons (Procyon lotor) with a Vaccinia Rabies Glycoprotein Recombinant Virus Vaccine”. Proc. Nad. Acad. Sci. USA Vol. 83, pp. 7947-7950, October 1986 Medical Sciences. [6] Nunberg et al. “Interleukin 2 Acts as an Adjuvant to Increase the Potency of Inactivated Rabies Virus Vaccine”. Proc. Nati. Acad. Sci. USA Vol. 86, pp. 4240-4243, June 1989 Immunology. [7] Wall et al. “Monosynaptic Circuit Tracing in Vivo through Cre-dependent Targeting and Complementation of Modified Rabies Virus”. 21848–21853, PNAS, December 14, 2010, Vol. 107 no. 50. [8] H. Kallel et al. “Evaluation of Various Serum and Animal Protein Free Media for the Production of a Veterinary Rabies Vaccine in BHK-21 Cells”. Journal of Biotechnology 95 (2002) 195–204. [9] Li et al. “Postexposure Treatment with the LiveAttenuated Rabies Virus (RV) Vaccine TriGAS Triggers the Clearance of Wild-Type RV from the
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Central Nervous System (CNS) through the Rapid Induction of Genes Relevant to Adaptive Immunity in CNS Tissues”. J. Virol. 2012, 86(6):3200.
[19] Larghi et al.” Ethylenimine-Inactivated Rabies Vaccine of Tissue Culture Origin”. Journal of Clinical Microbiology, Jan. 1976, p. 26-33.
[10] Wirblich et al. “PPEY Motif within the Rabies Virus (RV) Matrix Protein Is Essential for Efficient Virion Release and RV Pathogenicity”. Journal of Virology, Oct. 2008, p. 9730–9738.
[20] Monique Lafon et al. “Detrimental Contribution of the Immuno-Inhibitor B7-H1 to Rabies Virus Encephalitis”. J Immunol 2008; 180:7506-7515.
[11] Brzozka et al. “Inhibition of Interferon Signaling by Rabies Virus Phosphoprotein P:Activation Dependent Binding of STAT1 and STAT2”. Journal of Virology, Mar. 2006, p. 2675–2683.
[21] M. V. Fernandes et al. “Endosymbiotic Relationship Between Animal Viruses and Host Cells”. Jem, December 1,1964, p. 1099-1122. [22] Cox et al. “Preparation and Characterization of
[12] Mcgettigan et al. “Second-Generation Rabies Virus-Based Vaccine Vectors Expressing Human Immunodeficiency Virus Type 1 Gag Have Greatly Reduced Pathogenicity but Are Highly Immunogenic.” Journal of Virology, Jan. 2003, p. 237–244.
Rabies Virus Hemagglutinin”. Immunity, Nov. 1980, p. 572-577.
[13] James P. Mcgettigan et al. “Rabies Virus-Based Vectors Expressing Human Immunodeficiency Virus Type 1 (HIV-1) Envelope Protein Induce a Strong, Cross-Reactive Cytotoxic T-Lymphocyte Response against Envelope Proteins from Different HIV-1 Isolates”. Journal of Virology, May 2001, p. 4430– 4434.
[24] Fehlner-gardiner et al. “Era Vaccine-Derived Cases of Rabies in Wildlife and Domestic Animals in Ontario, Canada, 1989–2004”. Journal of Wildlife Diseases, 44(1), 2008, pp. 71–85.
[14] Thoulouze et al. “The Neural Cell Adhesion Molecule Is a Receptor for Rabies Virus”. Journal of Virology, Sept. 1998, p. 7181–7190. [15] Masami Sugamata et al. “Paralysis of Street Rabies Virus-Infected Mice Is Dependent on T Lymphocytes”. Journal of Virology, Feb. 1992, p. 1252-1260. [16] Donald L. Lodmell et al. “Raccoon Poxvirus Recombinants Expressing the Rabies Virus Nucleoprotein Protect Mice against Lethal Rabies Virus Infection”. Journal of Virology, June 1991, p. 3400-3405. [17] Dietzschold et al. “Differences in Cell-to-Cell Spread of Pathogenic and Apathogenic Rabies Virus in Vivo and In Vitro”. Journal of Virology, Oct. 1985, p. 12-18. [18] Larghi and Nebel. “Rabies Virus Inactivation by Binary Ethylenimine: New Method for Inactivated Vaccine Production”. Journal of Clinical Microbiology, Feb. 1980. p. 120-122.
Infection
and
[23] Wiktor and Clark.” Chronic Rabies Virus Infection of Cell Cultures”. Infection and Immunity, Dec. 1972, p. 988-995.
[25] B.Wojczyk et al. “The Role of Site-Specific Nglycosylation in Secretion of Soluble Forms of Rabies Virus Glycoprotein”. Glycobiology vol. 8 no. 2 pp. 121–130, 1998. [26] Cox and Schneider. “Prophylactic Immunization of Humans Against Rabies by Intradermal Inoculation of Human Diploid Cell Culture Vaccine”. Journal of Clinical Microbiology, Feb. 1976, p. 96101. [27] Deborah A. Grosenbaugh et al. “Rabies Challenge of Captive Striped Skunks (Mephitis mephitis) following Oral Administration of a Live Vaccinia-Vectored Rabies Vaccine”. Journal of Wildlife Diseases, 43(1), 2007, pp. 124–128. [28] P. Coulon et al. “Molecular Basis of Rabies Virus Virulence. I. Selection of Avirulent Mutants of the CVS Strain with Anti G Monoelonal Antibodies”. J. gen. Virol. (1982), 61, 97-100. [29] C. Prehaud et al. “Characterization of a New Temperature-sensitive and Avirulent Mutant of the Rabies Virus”. J. gen. Viral. (1989), 70, 133-143. [30] Francoise Bussereau et al. “Isolation and Study of Temperature-sensitive Mutants of Rabies Virus”. J. gen. Virol. (1982), 60, 153-158.
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[31] Kaur et al. “Rabies DNA Vaccine Encoding Lysosome-targeted Glycoprotein Supplemented with Emulsigen-DConfers CompleteProtection in Preexposure and Postexposure studies in BALB/c Mice”. The FASEB Journal, 176 Vol. 24 January 2010. [32] Morag Ferguson and G.C. Schild. “A SingleRadial-lmmunodiffusion Technique for the Assay of Rabies Glycoprotein Antigen: Application for Potency Tests of Vaccines Against Rabies”. J. gen. Virol. (1982) 59, 197-201. [33] Perry et al. “A Study of Techniques for the Distribution of Oral Rabies Vaccine to Wild Raccoon Populations”. Journal of Wildlife Diseases, 25(2), 1989, pp. 206-217. [34] A. Coku. “Molecular Farming of Vaccines from Transgenic Plants”. MMG 445 Basic Biotechnology eJournal 2007 3: 110-116.
[42] Y. Honda et al. “Persistent Infection of Rabies Virus (HEP-Flury Strain) in Human Neuroblastoma Cells Capable of Producing Interferon”. J. gen. Virol. (1985), 66, 957-967. [43] R. Marcovistz et al. “Distribution of Rabies Virus, Interferon and Interferon-mediated Enzymes in the Brains of Virus-infected Rats”. J. gen. Virol. (1984), 65, 995-997. [44] Shuichi Cho et al. “Murine T Cell Clones Directed to Rabies Virus: Isolation and Their Properties”. J. gen. Virol. (1987), 68, 1115-1123. [45] Antoine P. Maillard and Yves Gaudin. “Rabies Virus Glycoprotein can Fold in Two Alternative, Antigenically Distinct Conformations Depending on Membrane-anchor Type”. Journal of General Virology (2002), 83, 1465–1476.
[35] Shwiff et al. “Direct and Indirect Costs oF Rabies Exposure: A Retrospective Study in Southern California (1998–2002)”. Journal of Wildlife Diseases, 43(2), 2007, pp. 251–257.
[46] Hans Bunschoten et al. “Rabies Virus-specific Human T Cell Clones Provide Help for an in vitro Antibody Response against Neutralizing Antibodyinducing Determinants of the Viral Glycoprotein”. J. gen. Virol. (1989), 70, 1513-1521.
[36] Rosatte et al. “Aerial Distribution of Onrabh® Baits as A Tactic to Control Rabies in Raccoons and Striped Skunks in Ontario, Canada”. Journal of Wildlife Diseases, 45(2), 2009, pp. 363–374.
[47] M. Fekadu et al. “An Immune Stimulating Complex (ISCOM) Subunit Rabies Vaccine Protects Dogs and Mice Against Street Rabies Challenge”. Vaccine, Vol. 10, Issue 3, 1992.
[37] Donald L. Lodmell et al. “Interferon Induced within the Central Nervous System during Infection is Inconsequential as a Mechanism Responsible for Murine Resistance to Street Rabies Virus”. J. gen. ViroL (1989), 70, 473-478.
[48] Amy T. Gilbert et al. “Evidence of Rabies Virus Exposure among Humans in the Peruvian Amazon”. Am. J. Trop. Med. Hyg., 87(2), 2012, pp. 206–215.
[38] H. Tsiang et al. “Inhibition of Rabies Virus in vitro by the Ammonium-5-Tungsto-2-Antimoniate”. J. gen. Virol. 0978), 4o, 665-668. [39] Frederick F. Knowlton et al. “Serological Responses of Coyotes to Two Commercial Rabies Vaccines”. Journal of Wildlife Diseases, 37(4), 2001, pp. 798–802. [40] Sobeys et al. “Field Evaluation of an Inactivated Vaccine to Control Raccoon Rabies in Ontario, Canada”. Journal of Wildlife Diseases, 46(3), 2010, pp. 818–831.
[49] Guoqing Zhang and Zhen F. Fu. “Complete Genome Sequence of a Street Rabies Virus from Mexico”. J. Virol. 2012, 86(19):10892. [50] Aurélie A. V. Albertini et al. “Crystal Structure of the Rabies Virus Nucleoprotein-RNA Complex”. Science 313, 360 (2006). [51] Fulai Yu et al. “Complete Genome Sequence of a Street Rabies Virus Isolated from a Rabid Dog in China”. J. Virol. 2012, 86(19):10890. [52] Henry Wilde et al, “Rabies Update for Travel Medicine Advisors”. CID 2003:37 (1 July) • Travel Medicine.
[41] Jason R. Boulanger et al. “Comparison of Suburban Vaccine Distribution Strategies to Control Raccoon Rabies”. Journal of Wildlife Diseases, 44(4), 2008, pp. 1014–1023.
[53]Drugbank, http://www.drugbank.ca/drugs/DB000 45#propertie
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[54]http://www.wormsandgermsblog.com/2012/07/ar ticles/animals/dogs/rabies-virus-survival/ [55] http://www.drugs.com/ppa/rabies-vaccine.html
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