Thursday October 10, 2019 from 13:40 to 15:10
Sensitive detection systems for infectious agents
1Robert Koch Fellow, Robert Koch Institute, Berlin, Germany
Xenotransplantation using pig cells, tissues or organs may be associated with the transmission of various porcine agents, including porcine viruses, to the transplant recipient, potentially inducing a disease (xenozoonosis). To prevent such transmissions, sensitive and specific detection systems have been developed to screen the donor pigs and the recipients in preclinical and clinical trials. To detect DNA viruses such as porcine cytomegalovirus (PCMV), which was shown to reduce the survival time of pig transplants in non-human primates, porcine circoviruses (PCV) 1, 2 and 3, porcine lymphotropic herpesviruses (PLHV) 1, 2, and 3 and other DNA viruses, PCR-based methods are used. To detect RNA viruses such as the hepatitis E virus 3 (HEV3), a well-known zoonotic virus, and other RNA viruses, RT-PCR methods are used. Furthermore, immunological methods such as Western blot analysis or ELISA can be used to detect virus-specific antibodies, in which purified viruses, recombinant viral proteins or synthetic peptides are used as antigens. Detection of antiviral antibodies is a reliable and sensitive method to detect virus infections. Porcine endogenous retroviruses (PERVs) pose a special risk since they are integrated in the genome of all pigs (proviruses) and can be released as virus particles able to infect human cells. Integrated PERV proviruses are detected by PCR, their copy number is estimated by digital droplet PCR, their expression at the mRNA and protein levels are studied using RT-PCR and Western blot, and the formation of infectious particles is monitored using infection assays. Using these methods, numerous pig breeds were screened, among them Göttingen Minipigs, Aachen Minipigs, and multiple genetically modified pigs generated for xenotransplantation. In the first clinical trials of pig islet cell transplantation no porcine viruses were transmitted to human recipients. Absence of transmission was also observed when pig islet cells were transplanted into marmosets. In contrast, when orthotopic heart transplantations were performed in baboons using hearts from PCMV-positive animals, PCMV transmission was observed. However, when organs from PCMV-negative animals were used, significantly longer survival times were observed.
Conclusion: Sensitive and specific detection methods for numerous porcine infectious agents with known and unknown zoonotic potential are available for screening donor pigs and transplant recipients. These methods are also needed for elimination programs based on selection of virus-negative animals, vaccination, treatment, early weaning, colostrum deprivation, Caesarean delivery or embryo transfer.
Deutsche Forschungsgemeinschaft, TRR27.