Abstract
Human rabies, a neglected viral zoonosis, is preventable through domestic animals vaccination and post-exposure prophylaxis using inactivated rabies vaccines. During vaccine production, several mandatory in vivo quality control trials, such as potency, live virus, and safety, are responsible for the use of large numbers of laboratory animals. Over the years, global organizations encouraged the development of alternative methods to reduce, replace and refine the use of animals in the pharmaceutical industry. In this study we standardized an in vitro assay for determination of residual live virus combining viral isolation techniques with direct immunofluorescence detection and viral quantification by a molecular method. Standardization of viral recovery steps and quantification by RT-qPCR were performed and the combined method was shown to be 3 fold more sensitive than the in vivo assay. It was possible to identify viral suspensions cultures, which still had residual viable rabies virus particles, evidencing the importance to implement this method in quality control schemes of rabies vaccine production. In addition, this developed assay is more practical, inexpensive and less time consuming, producing results in just 4 days, which may allow greater agility in the internal quality control of the vaccine. The in vitro method may reduce 2/3rd of laboratory animals numbers used for this purpose, since it can be applied in the intermediate quality control of inactivated rabies vaccine production.
Highlights
Rabies is a viral disease caused by the rabies virus (RABV), which belongs to the Rhabdoviridae family [1]
In order to reduce the number of animals in this practice, in vitro methods for residual live virus detection need to be developed
This study shows that combining two simple and effective techniques may be a safer and more accurate method for the detection of residual rabies
Summary
Rabies is a viral disease caused by the rabies virus (RABV), which belongs to the Rhabdoviridae family [1]. African and Asian regions account for 95% of human rabies cases. In these regions, infected dogs whose saliva contains RABV are the main source of transmission through bites and scratches [2]. Rabies is considered a deadly disease, characterized as severe encephalitis, it is preventable through extensive vaccination of dogs and cats and pre/post-prophylaxis treatments in exposed humans along with public health education [5]. Vaccines used for human and veterinary rabies prevention are mostly inactivated vaccines, contrary to the wildlife oral vaccines that are produced with attenuated viruses [6]. The potency, inactivation, safety and pyrogenicity tests are responsible for the use of a large number of laboratory animals [7]
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