Rabies vaccine production from different viral infected cell line and vaccine production 04 feb

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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]

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3 Rabies Vaccines 3.1 OspA lipoprotein OspA lipoprotein is found on outer surface protein of Borrelia burgdorferi sensu stricto ZS7 and expressed

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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

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