Determination of the Copy Number of Porcine Endogenous Retroviruses (PERV) in Auckland Island Pigs Repeatedly Used for Clinical Xenotransplantation and Elimination of PERV-C.

Auckland Island pigs represent an inbred population of feral pigs isolated on the sub-Antarctic island for over 100 years. The animals have been maintained under pathogen-free conditions in New Zealand; they are well characterized virologically and have been used as donor sources in first clinical trials of porcine neonatal islet cell transplantation for the treatment of human diabetes patients. The animals do not carry any of the xenotransplantation-relevant viruses, and in the first clinical trials, no porcine viruses, including porcine endogenous retroviruses (PERVs) were transmitted to the human recipients.

Transmitted drug resistance and subtype patterns of viruses from reported new HIV diagnoses in Germany, 2017-2020.

Background: The transmission of resistant HIV variants jeopardizes the effective use of antiretrovirals for therapy and prophylaxis. Molecular surveillance of new HIV diagnoses with a focus on prevalence and type of resistance associated mutations and the subtype of circulating viruses is mandatory.

Method: From 2017 to 2020, 11,527 new HIV diagnoses were reported in Germany to the Robert Koch Institute (RKI).

Comparison of a Genotype 1 and a Genotype 2 Macaque Foamy Virus Gene Indicates Distinct Infectivity and Cell-Cell Fusion but Similar Tropism and Restriction of Cell Entry by Interferon-Induced Transmembrane Proteins.

Foamy viruses (FVs) are naturally found in many different animals and also in primates with the notable exception of humans, but zoonotic infections are common. In several species, two different envelope () gene sequence clades or genotypes exist. We constructed a simian FV (SFV) clone containing a reporter gene cassette.

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.

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

Background: 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.

Transmission of Porcine Circovirus 3 (PCV3) by Xenotransplantation of Pig Hearts into Baboons.

Porcine circovirus 3 (PCV3) is a newly described member of the virus family Circoviridae. PCV3 is highly distributed among pigs and wild boars worldwide. A sudden introduction of PCV3 was recently observed in a herd of triple genetically modified pigs generated for xenotransplantation.

Human SAMHD1 restricts the xenotransplantation relevant porcine endogenous retrovirus (PERV) in non-dividing cells.

The release of porcine endogenous retrovirus (PERV) particles from pig cells is a potential risk factor during xenotransplantation by way of productively infecting the human transplant recipient. Potential countermeasures against PERV replication are restriction factors that block retroviral replication. SAMHD1 is a triphosphohydrolase that depletes the cellular pool of dNTPs in non-cycling cells starving retroviral reverse transcription.

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.

Porcine endogenous retroviruses: Quantification of the copy number in cell lines, pig breeds, and organs.

Background: Porcine endogenous retroviruses (PERVs) may pose a risk of xenotransplantation using porcine cells, tissues, or organs. PERVs are integrated in the genome of all pigs, and some can infect certain human cells. The copy number of PERVs in different pig breeds has been determined by using different methods, with varying results.

Distribution of Porcine Cytomegalovirus in Infected Donor Pigs and in Baboon Recipients of Pig Heart Transplantation.

The porcine cytomegalovirus (PCMV) is a herpesvirus that may pose a risk for xenotransplantation using pig cells, tissues, or organs. Here, three orthotopic pig heart transplantations into baboons were studied. To detect PCMV, a real-time PCR and a Western blot assay based on four PCMV protein sequences, including two tegument proteins, were used.

Antibody Cross-Reactivity between Porcine Cytomegalovirus (PCMV) and Human Herpesvirus-6 (HHV-6).

Porcine cytomegalovirus (PCMV) infection is widely prevalent among pigs, and PCMV is one of the viruses which may be transmitted during xenotransplantation using pig cells, tissues, or organs. While human cytomegalovirus (HCMV) is a major risk factor for allotransplantation, it is still unclear whether PCMV is able to infect human cells or pose a risk for xenotransplantation. Previously, it was shown that transmission of PCMV after pig kidney to non-human primate transplantations resulted in a significantly reduced survival time of the transplanted organ.

Organization and ELISA-Based Results of the First Proficiency Testing to Evaluate the Ability of European Union Laboratories to Detect Staphylococcal Enterotoxin Type B (SEB) in Buffer and Milk.

The aim of this work was to organize the first proficiency test (PT) dedicated to staphylococcal enterotoxin B (SEB) detection in milk and buffer solutions. This paper describes the organization of the PT trial according to EN ISO 17043 requirements. Characterization of the SEB stock solution was performed using SDS-PAGE and SE-specific ELISA, and amino acid analysis was used to assign its protein concentration.

Detection of koala retrovirus subgroup B (KoRV-B) in animals housed at European zoos.

Many koalas carry an endogenous retrovirus, KoRV-A, in their genome. Recently, a second retrovirus, KoRV-B, was detected in koalas in Japanese and U.S.

Generation and Characterization of Six Recombinant Botulinum Neurotoxins as Reference Material to Serve in an International Proficiency Test.

The detection and identification of botulinum neurotoxins (BoNT) is complex due to the existence of seven serotypes, derived mosaic toxins and more than 40 subtypes. Expert laboratories currently use different technical approaches to detect, identify and quantify BoNT, but due to the lack of (certified) reference materials, analytical results can hardly be compared. In this study, the six BoNT/A1-F1 prototypes were successfully produced by recombinant techniques, facilitating handling, as well as improving purity, yield, reproducibility and biosafety.

Recommended Immunological Strategies to Screen for Botulinum Neurotoxin-Containing Samples.

Botulinum neurotoxins (BoNTs) cause the life-threatening neurological illness botulism in humans and animals and are divided into seven serotypes (BoNT/A-G), of which serotypes A, B, E, and F cause the disease in humans. BoNTs are classified as "category A" bioterrorism threat agents and are relevant in the context of the Biological Weapons Convention. An international proficiency test (PT) was conducted to evaluate detection, quantification and discrimination capabilities of 23 expert laboratories from the health, food and security areas.

Qualitative and Quantitative Detection of Botulinum Neurotoxins from Complex Matrices: Results of the First International Proficiency Test.

In the framework of the EU project EQuATox, a first international proficiency test (PT) on the detection and quantification of botulinum neurotoxins (BoNT) was conducted. Sample materials included BoNT serotypes A, B and E spiked into buffer, milk, meat extract and serum. Different methods were applied by the participants combining different principles of detection, identification and quantification.

Suppressing active replication of a live attenuated simian immunodeficiency virus vaccine does not abrogate protection from challenge.

Although safety concerns preclude the use of live attenuated HIV vaccines in humans, they provide a useful system for identifying the elusive correlates of protective immunity in the SIV/macaque animal model. However, a number of pieces of evidence suggest that protection may result from prior occupancy of susceptible target cells by the vaccine virus rather than the immune response. To address this, we developed a Nef-deletion variant of an RT-SHIV whose active replication could be shut off by treatment with RT-inhibitors.

Induction of neutralizing antibodies specific for the envelope proteins of the koala retrovirus by immunization with recombinant proteins or with DNA.

Background: The koala retrovirus (KoRV) is the result of a transspecies transmission of a gammaretrovirus with fatal consequences for the new host. Like many retroviruses, KoRV induces lymphoma, leukemia and an immunodeficiency that is associated with opportunistic infections in the virus-infected animals. We recently reported the induction of neutralizing antibodies by immunization with the recombinant ectodomain of the transmembrane envelope protein p15E of KoRV.

Lack of antiviral antibody response in koalas infected with koala retroviruses (KoRV).

Many wild koalas are infected with the koala retrovirus, KoRV, some of which suffer from lymphoma and chlamydial disease. Three subgroups, KoRV-A, KoRV-B and KoRV-J, have so far been described. It is well known that other closely related gammaretroviruses can induce tumours and severe immunodeficiencies in their respective hosts and a possible role for KoRV infection in lymphoma and chlamydial disease in koalas has been suggested.

A novel small animal model to study the replication of simian foamy virus in vivo.

Preclinical evaluation in a small animal model would help the development of gene therapies and vaccines based on foamy virus vectors. The establishment of persistent, non-pathogenic infection with the prototype foamy virus in mice and rabbits has been described previously. To extend this spectrum of available animal models, hamsters were inoculated with infectious cell supernatant or bioballistically with a foamy virus plasmid.

Generation of neutralising antibodies against porcine endogenous retroviruses (PERVs).

Antibodies neutralising porcine endogenous retroviruses (PERVs) were induced in different animal species by immunisation with the transmembrane envelope protein p15E. These antibodies recognised epitopes, designated E1, in the fusion peptide proximal region (FPPR) of p15E, and E2 in the membrane proximal external region (MPER). E2 is localised in a position similar to that of an epitope in the transmembrane envelope protein gp41 of the human immunodeficiency virus-1 (HIV-1), recognised by the monoclonal antibody 4E10 that is broadly neutralising.

Novel functions of prototype foamy virus Gag glycine- arginine-rich boxes in reverse transcription and particle morphogenesis.

Prototype foamy virus (PFV) Gag lacks the characteristic orthoretroviral Cys-His motifs that are essential for various steps of the orthoretroviral replication cycle, such as RNA packaging, reverse transcription, infectivity, integration, and viral assembly. Instead, it contains three glycine-arginine-rich boxes (GR boxes) in its C terminus that putatively represent a functional equivalent. We used a four-plasmid replication-deficient PFV vector system, with uncoupled RNA genome packaging and structural protein translation, to analyze the effects of deletion and various substitution mutations within each GR box on particle release, particle-associated protein composition, RNA packaging, DNA content, infectivity, particle morphology, and intracellular localization.

Increased neutralizing antibody response after simultaneous immunization with leucogen and the feline leukemia virus transmembrane protein.

To develop improved vaccination strategies against feline leukemia virus (FeLV), rats were immunized with the transmembrane envelope protein p15E of FeLV alone or in combination with the commercial vaccine Leucogen® comprising the nonglycosylated FeLV surface envelope protein. Binding and neutralizing antibodies were induced in both groups and in the group immunized with Leucogen alone. Higher titers of antibodies neutralizing FeLV were induced by simultaneous immunization with Leucogen and p15E compared to the responses using Leucogen or p15E alone, suggesting that combination vaccines should be used in the future.