Pig Cytomegalovirus Research Articles

Virus Safety of Xenotransplantation.

The practice of xenotransplantation using pig islet cells or organs is under development to alleviate the shortage of human donor islet cells or organs for the treatment of diabetes or organ failure. Multiple genetically modified pigs were generated to prevent rejection. Xenotransplantation may be associated with the transmission of potentially zoonotic porcine viruses. In order to prevent this, we developed highly sensitive PCR-based, immunologicals and other methods for the detection of numerous xenotransplantation-relevant viruses. These methods were used for the screening of donor pigs and xenotransplant recipients. Of special interest are the porcine endogenous retroviruses (PERVs) that are integrated in the genome of all pigs, which are able to infect human cells, and that cannot be eliminated by methods that other viruses can. We showed, using droplet digital PCR, that the number of PERV proviruses is different in different pigs (usually around 60). Furthermore, the copy number is different in different organs of a single pig, indicating that PERVs are active in the living animals. We showed that in the first clinical trials treating diabetic patients with pig islet cells, no porcine viruses were transmitted. However, in preclinical trials transplanting pig hearts orthotopically into baboons, porcine cytomegalovirus (PCMV), a porcine roseolovirus (PCMV/PRV), and porcine circovirus 3 (PCV3), but no PERVs, were transmitted. PCMV/PRV transmission resulted in a significant reduction of the survival time of the xenotransplant. PCMV/PRV was also transmitted in the first pig heart transplantation to a human patient and possibly contributed to the death of the patient. Transmission means that the virus was detected in the recipient, however it remains unclear whether it can infect primate cells, including human cells. We showed previously that PCMV/PRV can be eliminated from donor pigs by early weaning. PERVs were also not transmitted by inoculation of human cell-adapted PERV into small animals, rhesus monkey, baboons and cynomolgus monkeys, even when pharmaceutical immunosuppression was applied. Since PERVs were not transmitted in clinical, preclinical, or infection experiments, it remains unclear whether they should be inactivated in the pig genome by CRISPR/Cas. In summary, by using our sensitive methods, the safety of xenotransplantation can be ensured.

Porcine cytomegalovirus in xenotransplantation: The new frontier in human transplantation?

Porcine cytomegalovirus in xenotransplantation: The new frontier in human transplantation?

The first clinical pig heart transplant: Was IVIg or pig cytomegalovirus detrimental to the outcome?

The clinical course of the first patient to receive a gene-edited pig heart transplant was recently reported by the University of Maryland team. Although the pig heart functioned well for >40 days, serum anti-pig antibodies then increased, and the patient sadly died after 60 days. Because of his debilitated pre-transplant state, the patient never thrived despite excellent graft function for several weeks, and the cause of his demise continues to be uncertain. A few days before an increase in anti-pig antibodies was observed, the patient had received intravenous human immunoglobulin (IVIg), and whether this played a role in his cardiac deterioration has been discussed. Furthermore, mcfDNA testing indicated an increase in pig cytomegalovirus (CMV), and its possible role in the development of cardiac dysfunction has also been considered. On the basis of the limited data provided in the publication and on our previous investigations into whether IVIg contains anti-TKO pig antibodies and therefore might be deleterious to TKO pig organ xenografts, we suggest that the steady rise in anti-pig antibody titer was more consistent with the failure of the immunosuppressive regimen to prevent elicited anti-TKO pig antibody production, rather than from the passive transfusion of IVIg or the presence of pig CMV in the graft. Although the outcome of the Maryland experience was disappointing, valuable lessons were learned. Our attention was drawn to the potential risks of heart transplantation in a "deconditioned" patient, the administration of IVIg, the transmission of pig CMV, and of the difficulties in interpreting myocardial biopsy findings.

Porcine cytomegalovirus: A very unwelcome stowaway.

The announcement of the first pig-to-human heart transplantation early this year was the culmination of a very long and arduous path. The groundbreaking first life-sustaining pig-to man heart transplantation by the team of University of Maryland at Baltimore under the lead of Professor Griffith provided the long-awaited proof that it can work–even if survival was cut short by a hitherto still to be defined process. In a fascinating webinar, Professor Griffith described in detail the course of the patient.1 One potential element in the deterioration was the increase of porcine cytomegalovirus (PCMV) DNAemia, detected in the blood of the patient. The exact role of PCMV in the sequence of events is unclear and currently analyzed in detail, and will undoubtedly increase our knowledge. As a young researcher in the laboratory of Jay Fishman, I had the privilege to play a small role in early pig-to-baboon xenotransplantation experiments done in the Transplantation Biology Research Center at MGH, headed by Professor David Sachs. Organ- and recipient survival was short in the early 2000s, and reasons were explored on many different angles. From the experience in human-to-human solid organ transplantation, one avenue was to look for activation or primary infection of latent viruses, primarily from the herpesvirus family. PCMV had been identified as a potential zoonotic agent before.2 Using quantitative PCR assays, we demonstrated activation of PCMV and baboon cytomegalovirus (BCMV) in a series of pig-to primate organ kidney xenotransplantations. High copy numbers of PCMV DNAemia were detected in recipient baboon tissue and particularly in the transplanted porcine graft tissues.3, 4 In a model of primary porcine aortic endothelial cells infected in vitro with PCMV, an increase in porcine tissue factor was observed, and we hypothesized that this was a potential cofactor in the development of an often fatal consumptive coagulopathy observed in pig-to baboon kidney xenotransplantation.5 Using PCMV-free pig donors, which was achieved by early weaning of piglets, consumptive coagulopathy was prevented in pig-to-baboon heart transplantation, resulting in an improved graft survival.6 Given the many modifications in the protocols over time, it was not possible to establish PCMV as the sole cause of this coagulopathy. Nevertheless, at this point elimination of PCMV was the clear goal given the harmful potential of this virus.7 Eradication protocols were developed and successfully tested.8 This was all the more important, as ganciclovir failed to control PCMV activation.9 BCMV activation in the recipient baboons, on the other hand, by then was controlled by prophylactic use of ganciclovir in the majority of recipients and had become part of all protocols. The most comprehensive study on the role of PCMV was recently published by Denner et al. in 2020. In a series of orthotopic pig-to-baboon heart transplantations, transmission of PCMV was observed from PCMV-positive donor pigs in some but not all baboon recipients, whereas no transmission events occurred for a number of other potentially harmful latent porcine viruses tested, including porcine lymphotropic herpesviruses, and porcine endogenous retroviruses. In baboons transplanted with a heart from a PCMV-positive donor pig, survival was reduced, inflammatory cytokines such as IL-6 and TNF-α were increased and tPA-PAI-1 complexes circulated in high concentrations in the recipients, pointing to substantial alterations of the coagulation system. This detailed report provided additional evidence for the deleterious effects of PCMV, and reiterated the need for elimination of PCMV from the pig donor.10 Why then are PCMV-positive donor pigs still an issue? Breeding pathogen-free pigs was shown to be feasible, but the effort to achieve this is considerable. In a publication on virus prevalence in different minipig breeds in 2021, PCMV was still detected in some but not all breeds.11 Recent work did primarily focus on porcine endogenous retroviruses, as elimination of these viruses is incomparably more challenging than that of traditional latent viruses. Testing for elimination requires not only the use of PCR for detection PCMV DNAemia, but also by serology, as PCR can be negative in swabs from a healthy swine. Validated serological assays for PCMV are not readily available. Efforts did understandably concentrate on the genetic modifications of donor pigs. Given the potential unfavorable role in the first human case of orthotopic heart transplantation, the focus must be redirected to the elimination of PCMV. It is a feasible and necessary step. The amazing initial clinical course of the patient is a huge incentive to keep the momentum going and continue on this path.

Herpesvirus-associated inclusion body rhinitis and bacterial rhinitis in piglets.

Herpesvirus-associated inclusion body rhinitis and bacterial rhinitis in piglets.

Virological Characterization of Pigs with Erythema Multiforme.

Erythema multiforme in pigs is an acute, self-limiting disease characterized by red skin areas and often associated with anorexia, fever and respiratory problems. The cause of the disease remains unknown. In a recent study, animals of a commercial breeding herd in Greece were examined, and all animals were found seropositive for porcine reproductive and respiratory syndrome virus (PRRSV). However, neither PRRSV and porcine circovirus type 2 (PCV2) viremia nor antibodies against Aujeszky’s disease virus, African swine fever virus and classical swine fever virus were detected. Here, an extended examination of these pigs was performed on a wide range of porcine viruses using highly sensitive polymerase chain reaction (PCR)-based methods. Affected skin of five animals revealed the presence of porcine lymphotropic herpesvirus-1 (PLHV-1) in all cases, PLHV-2 in one animal and PLHV-3 in four animals. However, neither porcine cytomegalovirus (PCMV) nor porcine circoviruses (PCV1, PCV2, PCV3 and PCV4) were detected. In blood samples, PLHV-1 was present in two animals and PLHV-2, PCV2 and PCV3 in one individual, with PCMV, PCV1 and PCV4 in none of the animals. In one animal, four viruses were found in the blood (PLHV-1, PLHV-2, PCV2 and PCV3). A PRRSV viremia was also not detected. All animals carried porcine endogenous retrovirus C (PERV-C) in their genome, but recombinant PERV-A/C was not detected. The results suggest that porcine viruses may be involved in erythema multiforme in these animals and that further studies are needed to assess the role of these pathogens in the disease.

Virological and Parasitological Characterization of Mini-LEWE Minipigs Using Improved Screening Methods and an Overview of Data on Various Minipig Breeds.

Minipigs play an important role in biomedical research and have also been used as donor animals in xenotransplantation. To serve as a donor in xenotransplantation, the animals must be free of potential zoonotic viruses, bacteria and parasites. Porcine endogenous retroviruses (PERVs) are integrated in the genome of all pigs and cannot be eliminated as most of the other pig viruses can. PERV-A and PERV-B infect human cells in cell culture and are integrated in all pigs, whereas PERV-C infects only pig cells and it is found in many, but not all pigs. Minipigs are known for a high prevalence of recombinant PERV-A/C viruses able to infect human cells (Denner and Schuurman, Viruses, 2021;13:1869). Here, Mini-LEWE minipigs are screened for the first time for pig viruses including PERV. Peripheral blood mononuclear cells (PBMCs) from 10 animals were screened using PCR-based methods (PCR, RT-PCR, and real-time PCR). In comparison with our previous screening assays, numerous improvements were introduced, e.g., the usage of gene blocks as a PCR standard and foreign RNA to control reverse transcription in RT-PCR. Using these improved detection methods, Mini-LEWE pigs were found to be negative for porcine cytomegalovirus (PCMV), porcine lymphotropic herpesviruses (PLHV-1, -2 and -3), porcine circoviruses (PCV1, 2, 3 and 4), porcine parvovirus (PPV) and hepatitis E virus (HEV). All animals carried PERV-A, PERV-B and PERV-C in their genome. PERV-A/C was not found. In contrast to all other minipig breeds (Göttingen minipigs, Aachen minipigs, Yucatan micropig, Massachusetts General Hospital miniature pigs), Mini-LEWE minipigs have less viruses and no PERV-A/C. Parasitological screening showed that none of the Mini-LEWE minipigs harbored ecto- and gastrointestinal parasites, but at least one animal tested positive for anti-Toxoplasma gondii antibodies.

Porcine Lymphotropic Herpesviruses (PLHVs) and Xenotranplantation.

Porcine lymphotropic herpesviruses -1, -2 and -3 (PLHV-1, PLHV-2 and PLHV-3) are gammaherpesviruses which are widespread in pigs. They are closely related to the Epstein–Barr virus (EBV) and Kaposi sarcoma herpesvirus, both of which cause severe diseases in humans. PLHVs are also related to bovine and ovine gammaherpesviruses, which are apathogenic in the natural host, but cause severe diseases after transmission into other species. Until now, no association between PLHVs and any pig diseases had been described. However, PLHV-1 causes a post-transplantation lymphoproliferative disorder (PTLD) after experimental transplantations in minipigs. This disorder is similar to human PTLD, a serious complication of solid human organ transplantation linked to EBV. Xenotransplantation using pig cells, tissues and organs is under development in order to alleviate the shortage of human transplants. Meanwhile, remarkable survival times of pig xenotransplants in non-human primates have been achieved. In these preclinical trials, another pig herpesvirus, the porcine cytomegalovirus (PCMV), a roseolovirus, was shown to significantly reduce the survival time of pig xenotransplants in baboons and other non-human primates. Although PLHV-1 was found in genetically modified donor pigs used in preclinical xenotransplantation, it was, in contrast to PCMV, not transmitted to the recipient. Nevertheless, it seems important to use PLHV-free donor pigs in order to achieve safe xenotransplantation.

Molecular detection of Porcine cytomegalovirus (PCMV) in wild boars from Northeastern Patagonia, Argentina

Porcine cytomegalovirus (PCMV) is a recognized pathogen of domestic swine that is widely distributed around the world. PCMV is the etiological agent of inclusion body rhinitis and has also been associated with other diseases that cause substantial losses in swine production. Wild boar populations can act as reservoirs of numerous infectious agents that affect pig livestock, including PCMV. The aim of this work was to assess the circulation of this virus in free-living wild boars that inhabit Northeastern Patagonia (Buenos Aires and Río Negro Provinces), Argentina. Nested-PCR assays were conducted to evaluate the presence of PCMV in samples of tonsil tissue collected from 62 wild boar individuals. It was found that the overall rate of infection was about 56%, with significant higher values (almost 90%) in the age group corresponding to piglets (animals less than 6 months old). In addition, a seasonal variation was observed in the PCMV detection rate, with an increase during the transition from summer to autumn. In conclusion, this study confirmed that wild boars are major carriers and dispersal agents of PCMV in Northeastern Patagonia, which raises the necessity to evaluate the extent to which this virus affects local livestock production.

The origin of porcine endogenous retroviruses (PERVs).

Porcine endogenous retroviruses (PERVs) are integrated in the genome of all pigs, and they produce viral particles that are able to infect human cells and therefore pose a special risk for xenotransplantation. In contrast to other pig microorganisms that also pose a risk, such as porcine cytomegalovirus and hepatitis E virus, PERVs cannot be eliminated from pigs by vaccines, antiviral drugs, early weaning, or embryo transfer. Since PERVs are relevant for xenotransplantation, their biology and origin are of great interest. Recent studies have shown that PERVs are the result of a transspecies transmission of precursor retroviruses from different animals and further evolution in the pig genome. PERVs acquired different long terminal repeats (LTRs), and recombination took place. In parallel, it has been shown that the activity of the LTRs and recombination in the envelope are important for the transmissibility and pathogenesis of PERVs. Transspecies transmission of retroviruses is common, a well-known example being the transmission of precursor retroviruses from non-human primates to humans, resulting in human immunodeficiency virus (HIV). Here, recent findings concerning the origin of PERVs, their LTRs, and recombination events that occurred during evolution are reviewed and compared with other findings regarding transspecies transmission of retroviruses.

One-Tube Nested Real-Time PCR Assay for Rapid Screening of Porcine Cytomegalovirus in Clinical Samples.

Porcine cytomegalovirus (PCMV) is a pathogen that must be removed from pigs for use as organ donors in xenotransplantation. Recently, it has been found that when donor pigs are infected with PCMV, a pig-to-non-human-primate xenotransplantation lower transplant survival by 2–3 times. Therefore, highly sensitive methods are needed to maintain designated pathogen free (DPF) pig and screen for xenografts. The purpose of this study was to evaluate the performance of commercially available method with one-tube nested real-time PCR assay to quickly detect PCMV infection in clinical samples and compare the results with those of sequence analysis. Molecular diagnostic methods were used to evaluate 127 samples, including tissues and blood samples from pigs suspected of PCMV infection. The detection rate for positive PCMV was 38.6% (n = 49), 23.6% (n = 30), and 12.6% (n = 16) in one-tube nested real-time PCR, nested PCR, and conventional PCR methods, respectively. All PCMV-positive samples in conventional PCR or nested PCR methods were also positive in the one-tube nested real-time PCR assay. All the PCR products in the three methods were checked for amplification of PCMV gene by PCR and subsequent direct sequencing. The results of one-tube nested real-time PCR were found to be consistent with those of sequence analysis for all the samples and showed good agreement (κ = 1). Our study found that the one-tube nested real-time PCR assay is more sensitive than the other two methods. This assay required approximately 1.5 h for completion. Therefore, we concluded that one-tube nested real-time PCR assay is a fast and reliable method for the characterizing pathogen responsible for PCMV infection.

Impact of porcine cytomegalovirus on long-term orthotopic cardiac xenotransplant survival

Xenotransplantation using pig organs has achieved survival times up to 195 days in pig orthotopic heart transplantation into baboons. Here we demonstrate that in addition to an improved immunosuppressive regimen, non-ischaemic preservation with continuous perfusion and control of post-transplantation growth of the transplant, prevention of transmission of the porcine cytomegalovirus (PCMV) plays an important role in achieving long survival times. For the first time we demonstrate that PCMV transmission in orthotopic pig heart xenotransplantation was associated with a reduced survival time of the transplant and increased levels of IL-6 and TNFα were found in the transplanted baboon. Furthermore, high levels of tPA-PAI-1 complexes were found, suggesting a complete loss of the pro-fibrinolytic properties of the endothelial cells. These data show that PCMV has an important impact on transplant survival and call for elimination of PCMV from donor pigs.

Objective pathogen monitoring in nursery and finisher pigs by monthly laboratory diagnostic testing

BackgroundInfectious diseases are of great economic importance in commercial pig production, causing both clinical and subclinical disease, with influence on welfare, productivity, and antibiotic use. The causes of these diseases are often multifactorial and laboratory diagnostics are seldom routinely performed. The aim of the present study was to explore the benefits of monthly pathogen monitoring in nursery and finisher herds and to examine association between laboratory results and observed clinical signs, including coughing and diarrhoea. Three monthly samplings were conducted in three different age groups in six nursery and four finisher production units. For each unit, two pens were randomly selected in each age group and evaluated for coughing and diarrhoea events. Furthermore, faecal sock and oral fluid samples were collected in the selected pens and analysed for 18 respiratory and enteric viral and bacterial pathogens using the high-throughput real-time PCR BioMark HD platform (Fluidigm, South San Francisco, USA).ResultsIn total, 174 pens were sampled in which eight coughing events and 77 diarrhoeic events were observed. The overall findings showed that swine influenza A virus, porcine circovirus 2, porcine cytomegalovirus, Brachyspira pilosicoli, Lawsonia intracellularis, Escherichia coli fimbria types F4 and F18 were found to be prevalent in several of the herds. Association between coughing events and the presence of swine influenza A virus, porcine cytomegalovirus (Cq ≤ 20) or a combination of these were found. Furthermore, an association between diarrhoeic events and the presence of L. intracellularis (Cq ≤ 24) or B. pilosicoli (Cq ≤ 26) was found.ConclusionsThe use of high-throughput real-time PCR analysis for continuous monitoring of pathogens and thereby dynamics of infections in a pig herd, provided the veterinarian and farmer with an objective knowledge on the distribution of pathogens in the herd. In addition, the use of a high-throughput method in combination with information about clinical signs, productivity, health status and antibiotic consumption, presents a new and innovative way of diagnosing and monitoring pig herds and even to a lower cost than the traditional method.

Pig-to-non-human primate heart transplantation: The final step toward clinical xenotransplantation?

The demand for donated human hearts far exceeds the number available. Xenotransplantation of genetically modified porcine organs provides an alternative. In 2000, an Advisory Board of the International Society for Heart and Lung Transplantation set the benchmark for commencing clinical cardiac xenotransplantation as consistent 60% survival of non-human primates after life-supporting porcine heart transplantations. Recently, we reported the stepwise optimization of pig-to-baboon orthotopic cardiac xenotransplantation finally resulting in consistent success, with 4 recipients surviving 90 (n = 2), 182, and 195 days. Here, we report on 4 additional recipients, supporting the efficacy of our procedure. The first 2 additional recipients succumbed to porcine cytomegalovirus (PCMV) infections on Days 15 and 27, respectively. In 2 further experiments, PCMV infections were successfully avoided, and 3-months survival was achieved. Throughout all the long-term experiments, heart, liver, and renal functions remained within normal ranges. Post-mortem cardiac diameters were slightly increased when compared with that at the time of transplantation but with no detrimental effect. There were no signs of thrombotic microangiopathy. The current regimen enabled the prolonged survival and function of orthotopic cardiac xenografts in altogether 6 of 8 baboons, of which 4 were now added. These results exceed the threshold set by the Advisory Board of the International Society for Heart and Lung Transplantation. The results of our current and previous experimental cardiac xenotransplantations together fulfill for the first time the pre-clinical efficacy suggestions. PCMV-positive donor animals must be avoided.

Development of a TaqMan Based Real-Time Fluorescent Quantitative PCR Assay for Detection of Porcine Cytomegalovirus in Semen.

This study described a TaqMan based real-time fluorescent quantitative PCR (qPCR) method to detect porcine cytomegalovirus (PCMV) infection, targeting the conserved region of the DNA polymerase (DPOL) gene. The standard curve showed a linear regression relationship with a coefficient of 0.999 and a slope of y = −3.249x + 38.958 corresponding to the amplification efficiency at 99.8%. The limit of the qPCR method was 51.9 copies/μl. The established qPCR method showed excellent specificity, with no cross-reaction observed with common porcine pathogens. The coefficient of variation for intra-assay and interassay variability ranged up to 1.51% and 2.24%, respectively. PCMV positive signals can be found in semen using this qPCR method, which suggested that we should pay more attention to PCMV contamination in semen in order to eliminate PCMV infection in artificial insemination and xenotransplantation.

A Comprehensive Strategy for Screening for Xenotransplantation-Relevant Viruses in a Second Isolated Population of Göttingen Minipigs

Xenotransplantation using pig tissues and organs is under development in order to alleviate the increasing shortage of human transplants. Since xenotransplantation may be associated with the transmission of porcine microorganisms to the human recipient, the donor pigs should be carefully analyzed, especially for the presence of potentially zoonotic viruses. Göttingen Minipigs (GöMP) are potential donors of islet cells for the treatment of diabetes. Despite the fact that all animals produced at Ellegaard Göttingen Minipigs A/S carry porcine endogenous retroviruses (PERVs) in their genome and that very few animals were infected with porcine cytomegalovirus (PCMV), hepatitis E virus (HEV) and porcine lymphotropic herpesvirus (PLHV), no transmission of these viruses was observed in a preclinical trial transplanting GöMP islet cells into cynomolgus monkeys. Using a new comprehensive strategy, we then analyzed an isolated subpopulation of Göttingen Minipigs which remained at the University of Göttingen. We concentrated on 11 xenotransplantation-relevant viruses and combined co-incubation assays with susceptible human target cells and molecular biological methods to evaluate the risk posed by PERV. All animals in Göttingen carry PERV-A, PERV-B, and PERV-C in their genome but they are not infected with PCMV, PLHV and HEV. The difference may be explained by selection of negative animals and/or de novo infection. The PERV copy number was established using ddPCR (93 copies) and a human-tropic PERV-A/C was found released from PBMCs of one animal with a high expression of PERV-C.

Development of a one-run real-time PCR detection system for pathogens associated with porcine respiratory diseases.

The etiology of Porcine respiratory disease complex is complicated by infections with multiple pathogens, and multiple infections increase the difficulty in identifying the causal pathogen. In this present study, we developed a detection system of microbes from porcine respiratory by using TaqMan real-time PCR (referred to as Dempo-PCR) to screen a broad range of pathogens associated with porcine respiratory diseases in a single run. We selected 17 porcine respiratory pathogens (Actinobacillus pleuropneumoniae, Boldetella bronchiseptica, Haemophilus parasuis, Pasteurella multocida, Pasteurella multocida toxin, Streptococcus suis, Mycoplasma hyopneumoniae, Mycoplasma hyorhinis, Mycoplasma hyosynovie, porcine circovirus 2, pseudorabies virus, porcine cytomegalovirus, swine influenza A virus, porcine reproductive and respiratory virus US strain, EU strain, porcine respiratory coronavirus and porcine hemagglutinating encephalomyelitis virus) as detection targets and designed novel specific primer-probe sets for seven of them. In sensitivity test by using standard curves from synthesized DNA, all primer-probe sets showed high sensitivity. However, porcine reproductive and respiratory virus is known to have a high frequency of genetic mutations, and the primer and probe sequences will need to be checked at a considerable frequency when performing Dempo-PCR from field samples. A total of 30 lung samples from swine showing respiratory symptoms on six farms were tested by the Dempo-PCR to validate the assay’s clinical performance. As the results, 12 pathogens (5 virus and 7 bacteria) were detected and porcine reproductive and respiratory virus US strain, Mycoplasma hyorhinis, Haemophilus parasuis, and porcine cytomegalovirus were detected at high frequency. These results suggest that Dempo-PCR assay can be applied as a screening system with wide detection targets.

Comparative Analysis of Roseoloviruses in Humans, Pigs, Mice, and Other Species.

Viruses of the genus Roseolovirus belong to the subfamily Betaherpesvirinae, family Herpesviridae. Roseoloviruses have been studied in humans, mice and pigs, but they are likely also present in other species. This is the first comparative analysis of roseoloviruses in humans and animals. The human roseoloviruses human herpesvirus 6A (HHV-6A), 6B (HHV-6B), and 7 (HHV-7) are relatively well characterized. In contrast, little is known about the murine roseolovirus (MRV), also known as murine thymic virus (MTV) or murine thymic lymphotrophic virus (MTLV), and the porcine roseolovirus (PRV), initially incorrectly named porcine cytomegalovirus (PCMV). Human roseoloviruses have gained attention because they can cause severe diseases including encephalitis in immunocompromised transplant and AIDS patients and febrile seizures in infants. They have been linked to a number of neurological diseases in the immunocompetent including multiple sclerosis (MS) and Alzheimer’s. However, to prove the causality in the latter disease associations is challenging due to the high prevalence of these viruses in the human population. PCMV/PRV has attracted attention because it may be transmitted and pose a risk in xenotransplantation, e.g., the transplantation of pig organs into humans. Most importantly, all roseoloviruses are immunosuppressive, the humoral and cellular immune responses against these viruses are not well studied and vaccines as well as effective antivirals are not available.

Virological aspects of non-human primates or swine-to human xenotransplantation

There are a number of human diseases, which can lead to organ failure. The consequence is often the need for a transplant. The number of performed operations is very low due to the shortage of organs for transplantation. As a consequence, the number of people waiting for transplant is still growing. The solution to this situation may be xenotransplantation. Xenotransplantation word comes from the Greek xenos meaning stranger, the other. It is defined as any procedure that involves the transplantation, implantation or infusion of tissues or zoonotic organs into a human recipient, but also human body fluids, cells, tissues, organs (or fragments) that have ex vivo contact with zoonotic cells, tissues or organs. One of the obstacles of the xenograft transplantation is the risk of animal pathogens transmission to the humans. Viruses that pose risk in the non-human primates-to-human xenotransplantation includes: the human immunodeficiency virus - HIV and the Marburg virus described in this paper. In addition viruses, which is a problem in pig-to-human xenotransplantation have also been described, including: porcine endogenous retrovirus - PERV, porcine cytomegalovirus - PCMV, porcine lymphotropic herpesvirus - PLHV and hepatitis E virus - E - HEV. This review of literature is the latest knowledge of the microbiological safety of xenotransplantation.

Early weaning completely eliminates porcine cytomegalovirus from a newly established pig donor facility for xenotransplantation.

For clinical xenotransplantation, transplants must be free of porcine cytomegalovirus (PCMV). Piglets become infected primarily in the perinatal period by the mother sow. While individual donor animals can be protected from infection by isolation husbandry, success is not guaranteed and this strategy poses the risk of undetected infections and raises animal welfare questions. Here, we present the establishment of a completely PCMV-negative pig herd for breeding donor animals for xenotransplantation. Eleven pregnant DanAvl Basic hybrid sows were purchased from a designated pathogen-free (DPF), PCMV-positive colony and transferred to a new pig facility at the Centre for Innovative Medical Models (CiMM) 4weeks prior to farrowing. At the age of 24hours, piglets were early-weaned and transferred to a commercially available Rescue Deck system dedicated to motherless rearing of piglets. Sows were removed from the facility. The PCMV status of F1-generation animals was determined at regular intervals over a period of 14months by a sensitive real-time PCR-based detection method testing blood, nasal swabs and cultured peripheral blood mononuclear cells (PBMCs). F1 sows were used as recipients of genetically modified embryos to generate a xenotransplant donor herd. Offspring were tested for PCMV accordingly. All offspring have remained PCMV negative over the whole observation period of 14months. A completely PCMV-negative pig herd for xenotransplantation has thus been successfully established.