Safety and efficacy of a biodegradable implant releasing tenofovir alafenamide for vaginal protection in a macaque model.

OBJECTIVES: To advance the initiative of ending the global epidemic, long-lasting HIV protection is needed through sustained release of antiretroviral drugs for months to years. We investigated in macaques the safety and efficacy of biodegradable polycaprolactone implants releasing tenofovir alafenamide for HIV pre-exposure prophylaxis (PrEP). METHODS: Implants were administered subcutaneously in the arm using a contraceptive trocar. Efficacy against vaginal simian-HIV (SHIV) infection was investigated in six pigtailed macaques that received two tenofovir alafenamide implants (0.35 mg/day), one in each arm, for a total release rate of tenofovir alafenamide at 0.7 mg/day. Macaques were exposed to SHIV twice weekly for 6 weeks. Statistical analyses were used to compare outcome with eight untreated controls. Histological assessments were performed on skin biopsies collected near implantation sites. RESULTS: Median (range) tenofovir diphosphate level in PBMCs was 1519 (1068-1898) fmol/106 cells. All macaques with tenofovir alafenamide implants were protected against vaginal SHIV infection. In contrast, 7/8 controls were infected after a median of 4 SHIV exposures (P = 0.0047). Histological assessment of tissues near tenofovir alafenamide implant sites showed inflammation and necrosis in 5/6 animals, which were not evident by visual inspection. CONCLUSIONS: We demonstrated complete protection against vaginal SHIV infection with two implants releasing a total of 0.7 mg of tenofovir alafenamide per day. We also identified tenofovir diphosphate concentrations in PBMCs associated with complete vaginal protection. Consistent with previous findings, we observed adverse local toxicity and necrosis near the tenofovir alafenamide implant site. Improved tenofovir alafenamide implants that are safe and maintain high efficacy have the potential to provide long-lasting protection against vaginal HIV infection.

Human infection with foamy viruses.

Virtually all nonhuman primate species investigated thus far including prosimians, New World and Old World monkeys and apes all harbor distinct and species-specific clades of simian foamy virus (SFV). However, evidence supporting the existence of a human-specific foamy virus (FV) is not yet available. Early reports describing widespread infection of healthy and sick humans with FV could not be confirmed. In contrast, all FV infections documented in humans are of zoonotic origin and are identified in persons occupationally exposed to nonhuman primates. The introduction of SFV into humans raises several public health questions regarding disease outcomes and potential for human-to-human transmissibility. The available data from a very limited number of SFV-infected humans suggest that these infections are nonpathogenic and are not easily transmissible. Additional studies are needed to better define the prevalence and natural history of SFV in humans.

Susceptibility of human T cell leukemia virus type 1 to reverse-transcriptase inhibitors: evidence for resistance to lamivudine.

Nucleoside reverse-transcriptase (RT) inhibitors (NRTIs), including lamivudine (3TC) and zidovudine (Zdv), are being evaluated for the treatment of human T cell lymphotropic virus type 1 (HTLV-1)-associated disease. However, information on the susceptibility of HTLV-1 to these drugs is limited. The activity of 5 NRTIs on HTLV-1 RT was evaluated. IC(50) values for Zdv, zalcitabine (ddC), didanosine (ddI), 3TC, and stavudine (d4T) were determined, using an enzymatic assay, for 5 HTLV-1 isolates and for reference wild-type and NRTI-resistant human immunodeficiency virus type 1 (HIV-1). Both HTLV-1 and wild-type HIV-1 were equally susceptible to Zdv, ddC, ddI, and d4T. In contrast, high-level resistance to 3TC was found in all HTLV-1 isolates. The findings support the clinical use of Zdv, ddC, ddI, and d4T but not of 3TC for the antiretroviral treatment of HTLV-1-associated disease.

Lack of cross-species transmission of porcine endogenous retrovirus infection to nonhuman primate recipients of porcine cells, tissues, or organs.

BACKGROUND: Nonhuman primates (NHPs) have been widely used in different porcine xenograft procedures inevitably resulting in exposure to porcine endogenous retrovirus (PERV). Surveillance for PERV infection in these NHPs may provide information on the risks of cross-species transmission of PERV, particularly for recipients of vascularized organ xenografts for whom data from human clinical trials is unavailable. METHODS: We tested 21 Old World and 2 New World primates exposed to a variety of porcine xenografts for evidence of PERV infection. These NHPs included six baboon recipients of pig hearts, six bonnet macaque recipients of transgenic pig skin grafts, and nine rhesus macaque and two capuchin recipients of encapsulated pig islet cells. Serologic screening for PERV antibody was done by a validated Western blot assay, and molecular detection of PERV sequences in peripheral blood mononuclear cells (PBMCs) and plasma was performed using sensitive polymerase chain reaction and reverse transcriptase-polymerase chain reaction assays, respectively. Spleen and lymph node tissues available from six bonnet macaques and three rhesus macaques were also tested for PERV sequences. RESULTS: All plasma samples were negative for PERV RNA suggesting the absence of viremia in these xenografted animals. Similarly, PERV sequences were not detectable in any PBMC and tissue samples, arguing for the lack of latent infection of these compartments. In addition, all plasma samples were negative for PERV antibodies. CONCLUSION: These data suggest the absence of PERV infection in all 23 NHPs despite exposure to vascularized porcine organs or tissue xenografts and the use of immunosuppressive therapies in some animals. These findings suggest that PERV is not easily transmitted to these NHP species through these types of xenografts.

Lack of evidence of endogenous avian leukosis virus and endogenous avian retrovirus transmission to measles, mumps, and rubella vaccine recipients.

The identification of endogenous avian leukosis virus (ALV) and endogenous avian retrovirus (EAV) in chick cell-derived measles and mumps vaccines in current use has raised concern about transmission of these retroviruses to vaccine recipients. We used serologic and molecular methods to analyze specimens from 206 recipients of measles, mumps, and rubella (MMR) vaccine for evidence of infection with ALV and EAV. A Western blot assay for detecting antibodies to endogenous ALV was developed and validated. All serum samples were negative for antibodies to endogenous ALV by Western blot analysis. Peripheral blood lymphocyte samples from 100 vaccinees were further tested by polymerase chain reaction for both ALV and EAV proviral sequences; all were negative. Matching serum samples were tested by reverse transcriptase polymerase chain reaction for ALV and EAV RNA, and all 100 samples were negative, providing no evidence of viremia. These findings do not indicate the presence of either ALV or EAV infection in MMR vaccine recipients and provide support for current immunization policies.

Characterization of endogenous avian leukosis viruses in chicken embryonic fibroblast substrates used in production of measles and mumps vaccines.

Previous findings of low levels of reverse transcriptase (RT) activity in chick cell-derived measles and mumps vaccines showed this activity to be associated with virus particles containing RNA of both subgroup E endogenous avian leukosis viruses (ALV-E) and endogenous avian viruses (EAV). These particles originate from chicken embryonic fibroblast (CEF) substrates used for propagating vaccine strains. To better characterize vaccine-associated ALV-E, we examined the endogenous ALV proviruses (ev loci) present in a White Leghorn CEF substrate pool by restriction fragment length polymorphism. Five ev loci were detected, ev-1, ev-3, ev-6, ev-18, andev-19. Both ev-18 and ev-19 can express infectious ALV-E, while ev-1, ev-3, and ev-6 are defective. We analyzed the full-length sequence of ev-1 and identified an adenosine insertion within the pol RT-beta region at position 5026, which results in a truncated RT-beta and integrase. We defined the 1,692-bp deletion in the gag-pol region of ev-3, and we found that in ev-6, sequences from the 5' long terminal repeat to the 5' pol region were absent. Based on the sequences of the ev loci, RT-PCR assays were developed to examine expression of ALV-E particles (EV) in CEF supernatants. Both ev-1- and ev-3-like RNA sequences were identified, as well as two other RNA sequences with intact pol regions, presumably of ev-18 and ev-19 origin. Inoculation of susceptible quail fibroblasts with CEF culture supernatants from both 5-azacytidine-induced and noninduced CEF led to ALV infection, confirming the presence of infectious ALV-E. Our data demonstrate that both defective and nondefective ev loci can be present in CEF vaccine substrates and suggest that both ev classes may contribute to the ALV present in vaccines.

Evidence of infection with simian type D retrovirus in persons occupationally exposed to nonhuman primates.

Simian type D retrovirus (SRV) is enzootic in many populations of Asian monkeys of the genus Macaca and is associated with immunodeficiency diseases. However, the zoonotic potential of this agent has not been well defined. Screening for antibodies to SRV was performed as part of an ongoing study looking for evidence of infection with simian retroviruses among persons occupationally exposed to nonhuman primates (NHPs). Of 231 persons tested, 2 (0.9%) were found to be strongly seropositive, showing reactivity against multiple SRV antigens representing gag, pol, and env gene products by Western immunoblotting. Persistent long-standing seropositivity, as well as neutralizing antibody specific to SRV type 2, was documented in one individual (subject 1), while waning antibody with eventual seroreversion was observed in a second (subject 2). Repeated attempts to detect SRV by isolation in tissue culture and by using sensitive PCR assays for amplification of two SRV gene regions (gag and pol) were negative. Both individuals remain apparently healthy. We were also unable to transmit this seropositivity to an SRV-negative macaque by using inoculation of whole blood from subject 1. The results of this study provide evidence that occupational exposure to NHPs may increase the risk of infection with SRV and underscore the importance of both occupational safety practices and efforts to eliminate this virus from established macaque colonies.

Evidence of porcine endogenous retroviruses in porcine factor VIII and evaluation of transmission to recipients with hemophilia.

Since 1984, unheated porcine clotting factor VIII (Hyate:C) has been used to treat severe bleeding episodes in persons with hemophilia who have antibodies to human clotting factor. We document the presence of porcine endogenous retrovirus (PERV) in plasma samples of pigs and in clinical lots of Hyate:C. Both gag and pol PERV RNA sequences were detected by reverse-transcriptase (RT) polymerase chain reaction in 13 of 13 lots of Hyate:C tested. Among 10 of these lots, RT activity also was detected, which confirms the presence of retroviral particles. To assess the transmission of PERV to Hyate:C recipients, we tested serum specimens from 88 recipients of Hyate:C and 23 noninfused control subjects for anti-PERV antibodies by using a Western blot assay. None of the samples was positive. Our data document that PERV particles are a common contaminant of Hyate:C products and suggest that the risk of PERV transmission from these percutaneous exposures is very low.

Susceptibility of the porcine endogenous retrovirus to reverse transcriptase and protease inhibitors.

Porcine xenografts may offer a solution to the shortage of human donor allografts. However, all pigs contain the porcine endogenous retrovirus (PERV), raising concerns regarding the transmission of PERV and the possible development of disease in xenotransplant recipients. We evaluated 11 antiretroviral drugs licensed for human immunodeficiency virus type 1 (HIV-1) therapy for their activities against PERV to assess their potential for clinical use. Fifty and 90% inhibitory concentrations (IC(50)s and IC(90)s, respectively) of five nucleoside reverse transcriptase inhibitors (RTIs) were determined enzymatically for PERV and for wild-type (WT) and RTI-resistant HIV-1 reference isolates. In a comparison of IC(50)s, the susceptibilities of PERV RT to lamivudine, stavudine, didanosine, zalcitabine, and zidovudine were reduced >20-fold, 26-fold, 6-fold, 4-fold, and 3-fold, respectively, compared to those of WT HIV-1. PERV was also resistant to nevirapine. Tissue culture-based, single-round infection assays using replication-competent virus confirmed the relative sensitivity of PERV to zidovudine and its resistance to all other RTIs. A Gag polyprotein-processing inhibition assay was developed and used to assess the activities of protease inhibitors against PERV. No inhibition of PERV protease was seen with saquinavir, ritonavir, indinavir, nelfinavir, or amprenavir at concentrations >200-fold the IC(50)s for WT HIV-1. Thus, following screening of many antiretroviral agents, our findings support only the potential clinical use of zidovudine.

Absence of evidence of infection with divergent primate T-lymphotropic viruses in United States blood donors who have seroindeterminate HTLV test results.

BACKGROUND: Recent identification of divergent simian or primate T-lymphotropic viruses (STLVs; PTLVs) in bonobos (formerly called pygmy chimpanzees; Pan paniscus; viruses: STLVpan-p and STLVpp1664) and a baboon (Papio hamadryas; viruses: STLVph969 or PTLV-L) have raised the possibility of human infection with these viruses. Divergent PTLV-infected primate sera show p24 bands on HTLV-I Western blots (WBs). It was investigated whether infection by divergent PTLV-like viruses could explain a subset of United States blood donors who reacted on HTLV-I EIAs and had indeterminate HTLV-I WBs with p24 bands. STUDY DESIGN AND METHODS: Epidemiologic characteristics of 1889 donors with HTLV-I-indeterminate WBs were compared to those of donors with confirmed retrovirus infections (393 with HIV, 201 with HTLV-I, 513 with HTLV-II) and 1.6 million donors with nonreactive screening tests. To directly probe for infection with divergent PTLVs, 2 HTLV-I-indeterminate donors born in Africa and 269 representative non-African-born donors with p24 bands on HTLV-I WBs (previously shown to be negative for HTLV-I and -II DNA by PCR) were selected for PTLV PCR analysis. DNA from peripheral blood MNC samples was tested for a proviral tax sequence by PCR using generic primers that amplify HTLV-I, HTLV-II, and the divergent PTLVs. Amplified tax sequences were detected by Southern blot hybridization to a (32)P-labeled generic PTLV probe. PCR-positive samples could then be typed by hybridization with virus-specific internal probes that differentiate HTLV-I, HTLV-II, PTLV-L, and STLVpan-p. RESULTS: In the epidemiologic analysis, HTLV-indeterminate status was independently associated with age of at least 25 years (OR = 2.19; 95% CI, 1.93-2.49), black (OR = 3.27; 95% CI, 2.90-3.67) or Hispanic (OR = 1.82; 95% CI, 1.52-2.16) race or ethnicity, and donation at one blood center (Baltimore) (OR = 1. 30; 95% CI, 1.11-1.53). None of the 271 HTLV-I WB-indeterminate samples tested positive by generic PTLV PCR analysis. CONCLUSION: Although the epidemiologic data suggest the possibility of undiagnosed HTLV-I, HTLV-II, or a cross-reactive virus such as PTLV among older, black, and Hispanic blood donors, the PCR data do not support the presence of divergent PTLV infection among US blood donors with HTLV-I-indeterminate results.

Simian foamy virus infection among zoo keepers.

We investigated 322 North American zoo workers in an anonymous serosurvey for antibodies to simian foamy viruses to establish the potential risk of zoonotic transmission by these retroviruses. 4 of 133 (3%) individuals who worked specifically with mammals including primates were seropositive, primarily with chimp-like viruses, indicating the importance of work practices to reduce exposure to these agents.

Ultrasensitive Detection of Reverse Transcriptase Activity by the Amp-RT Assay : Applications to the Measurement of Virus Loads and Phenotypic Drug Resistance.

Retroviruses are widely prevalent among vertebrates and are the causative agents of a variety of diseases in humans and animals including immunodeficiences, leukemias, and lymphomas (1). The retrovirus family is characterized by the presence of virion-associated reverse transcriptase (RT), an enzyme that transcribes the viral genomic RNA into a double-stranded DNA copy. This feature has led to studies of the unique enzymatic function of RT for two main applications. First, RT is a good diagnostic tool for the generic detection of the presence of retroviruses. Second, the RT enzyme constitutes a primary target for antiviral drug therapy (1,2).

Persistent zoonotic infection of a human with simian foamy virus in the absence of an intact orf-2 accessory gene.

Although foamy viruses (FVs) are endemic among nonhuman primates, FV infection among humans is rare. Recently, simian foamy virus (SFV) infection was reported in 4 of 231 individuals occupationally exposed to primates (1.8%). Secondary transmission to spouses has not been seen, suggesting that while FV is readily zoonotic, humans may represent dead-end hosts. Among different simian species, SFV demonstrates significant sequence diversity within the U3 region of the long terminal repeat (LTR) and 3' accessory open reading frames (ORFs). To examine if persistent human SFV infection and apparent lack of secondary transmission are associated with genetic adaptations in FV regulatory regions, we conducted sequence analysis of the LTR, internal promoter, ORF-1, and ORF-2 on a tissue culture isolate and peripheral blood mononuclear cell samples from a human infected with SFV of African green monkey origin (SFV-3). Compared to the prototype SFV-3 sequence, the LTR, internal promoter, and FV transactivator (ORF-1) showed sequence conservation, suggesting that FV zoonosis is not dependent on host-specific adaptation to these transcriptionally important regions. However, ORF-2 contains a number of deleterious mutations predicted to result in premature termination of protein synthesis. ORF-2 codes in part for the 60-kDa Bet fusion protein, proposed to be involved in the establishment of persistent cellular SFV infections. These results suggest that persistent human infection by SFV and reduced transmissibility may be influenced by the absence of a functional ORF-2.

Polymerase chain reaction assays for the diagnosis of infection with the porcine endogenous retrovirus and the detection of pig cells in human and nonhuman recipients of pig xenografts.

BACKGROUND: Pigs offer an unlimited source of xenografts for humans. However, recipients of pig xenografts are inevitably exposed to the porcine endogenous retrovirus (PERV), which is carried in the pig germline. The ability of PERV to infect human cells in vitro has heightened safety concerns regarding the transmission of PERV to pig xenograft recipients. METHODS: In response to the need to establish laboratory tests for the surveillance of PERV infection, we have developed polymerase chain reaction (PCR) assays to detect PERV pol and gag sequences by using conserved primers and probes. In addition, we have developed a PCR assay to detect pig-specific mitochondrial DNA (mtDNA) sequences as a marker of pig cells. RESULTS: Analysis of assay sensitivities using cloned target copies in a background of human DNA demonstrated a detection threshold of 1, 5, and 1 copy for the PERV gag, pol, and pig mtDNA PCR assays, respectively. All three PCR assays gave negative results on peripheral blood lymphocyte samples from 69 humans, as well as 6 baboons and 6 macaques, demonstrating 100% specificity. The PERV and pig mtDNA assays were integrated into a simple testing algorithm that allows the differentiation between pig cell microchimerism and true xenogeneic infection. To allow for monitoring of PERV expression, a reverse transcriptase-PCR assay was also developed to detect cell-free PERV RNA. CONCLUSION: The use of the diagnostic tests described here will help define the risks of PERV transmission associated with the use of pig xenografts in humans and nonhuman primates.

Search for cross-species transmission of porcine endogenous retrovirus in patients treated with living pig tissue. The XEN 111 Study Group.

Pig organs may offer a solution to the shortage of human donor organs for transplantation, but concerns remain about possible cross-species transmission of porcine endogenous retrovirus (PERV). Samples were collected from 160 patients who had been treated with various living pig tissues up to 12 years earlier. Reverse transcription-polymerase chain reaction (RT-PCR) and protein immunoblot analyses were performed on serum from all 160 patients. No viremia was detected in any patient. Peripheral blood mononuclear cells from 159 of the patients were analyzed by PCR using PERV-specific primers. No PERV infection was detected in any of the patients from whom sufficient DNA was extracted to allow complete PCR analysis (97 percent of the patients). Persistent microchimerism (presence of donor cells in the recipient) was observed in 23 patients for up to 8.5 years.

Evidence of avian leukosis virus subgroup E and endogenous avian virus in measles and mumps vaccines derived from chicken cells: investigation of transmission to vaccine recipients.

Reverse transcriptase (RT) activity has been detected recently in all chicken cell-derived measles and mumps vaccines. A study of a vaccine manufactured in Europe indicated that the RT is associated with particles containing endogenous avian retrovirus (EAV-0) RNA and originates from the chicken embryonic fibroblasts (CEF) used as a substrate for propagation of the vaccine. We investigated the origin of RT in measles and mumps vaccines from a U.S. manufacturer and confirm the presence of RT and EAV RNA. Additionally, we provide new evidence for the presence of avian leukosis virus (ALV) in both CEF supernatants and vaccines. ALV pol sequences were first identified in particle-associated RNA by amplification with degenerate retroviral pol primers. ALV RNA sequences from both the gag and env regions were also detected. Analysis of hypervariable region 2 of env revealed a subgroup E sequence, an endogenous-type ALV. Both CEF- and vaccine-derived RT activity could be blocked by antibodies to ALV RT. Release of ALV-like virus particles from uninoculated CEF was also documented by electron microscopy. Nonetheless, infectivity studies on susceptible 15B1 chicken cells gave no evidence of infectious ALV, which is consistent with the phenotypes of the ev loci identified in the CEF. PCR analysis of ALV and EAV proviral sequences in peripheral blood mononuclear cells from 33 children after measles and mumps vaccination yielded negative results. Our data indicate that the sources of RT activity in all RT-positive measles and mumps vaccines may not be similar and depend on the particular endogenous retroviral loci present in the chicken cell substrate used. The present data do not support transmission of either ALV or EAV to recipients of the U.S.-made vaccine and provide reassurance for current immunization policies.

Development and validation of a Western immunoblot assay for detection of antibodies to porcine endogenous retrovirus.

BACKGROUND: Reports that pig endogenous retrovirus (PERV) infects human cells in vitro have heightened the importance of molecular and serologic monitoring of xenograft recipients for evidence of infection with PERV. We report the development and validation of a PERV-specific Western immunoblot assay for the diagnostic testing of porcine xenografts recipients. This assay is based upon the serological cross-reactivity observed between PERV variants capable of infecting human cells in vitro and other mammalian C type retroviruses. METHODS AND RESULTS: Strong reactivity between PERV expressing embryonic pig kidney PK-15 cells and antisera raised against whole virus preparations of murine leukemia virus, gibbon ape leukemia virus (GALV), and simian sarcoma-associated virus was demonstrated by an immunofluorescence assay, suggesting specific antigenic cross-reactivity between this group of viruses and PERV. Western immunoblot analysis demonstrated that anti-GALV antisera reacted with three proteins in PK-15 cells having molecular masses of 30, 55, and 66 kDa. Antisera specific for the Gag proteins of either GALV or simian sarcoma-associated virus reacted with the 30-kDa (major) and 55-kDa (minor) proteins present in PK-15 cells and in PERV-infected 293 human kidney cells, likely representing reactivity to the processed and precursor forms of the PERV Gag protein, respectively. No reactivity was seen in uninfected 293 cells. Analysis of plasma samples from 200 United States blood donors and from 58 human immunodeficiency virus-1, 18 human immunodeficiency virus-2, 13 human T-cell lymphotrophic virus-I, 21 human T-cell lymphotrophic virus-II, and 15 cytomegalovirus infected controls were negative. CONCLUSIONS: As this assay is based on PERV antigen derived from infected human cells, it clearly has the capacity to detect a serologic response towards PERV variants that have zoonotic potential and will allow for the accurate determination of PERV-specific seroreactivity in porcine xenograft recipients.

No evidence of infection with porcine endogenous retrovirus in recipients of porcine islet-cell xenografts.

BACKGROUND: The study of whether porcine xenografts can lead to porcine endogenous retrovirus (PERV) infection of recipients is critical for evaluating the safety of pig-to-man xenotransplantation. PERV is carried in the pig germline, and all recipients of porcine tissues or organs will be exposed to the virus. METHODS: We studied 10 diabetic patients who had received porcine fetal islets between 1990 and 1993, looking for evidence of PERV infection by using PCR serology, PCR, and reverse transcriptase assays. Prolonged xenograft survival (up to a year) was confirmed in five patients by porcine C-peptide excretion and detection of pig mitochondrial DNA (mtDNA) in serum. FINDINGS: Despite the evidence for extended exposure to pig cells and despite concomitant immunosuppressive therapy, we were unable to detect markers of PERV infection in any patient. Screening for two PERV sequences in peripheral blood lymphocytes collected 4-7 years after the xenotransplantation was negative. Markers of PERV expression, including viral RNA and reverse transcriptase, were undetectable in sera from both early (day 3 to day 180) and late (4-7 years) time points. Western blot analysis for antibodies was consistently negative. INTERPRETATION: These results suggested the absence of PERV infection in these patients. Also this study establishes a minimum standard for post-transplant surveillance of patients given porcine xenografts.

Amplification of simian retroviral sequences from human recipients of baboon liver transplants.

Investigations into the use of baboons as organ donors for human transplant recipients, a procedure called xenotransplantation, have raised the specter of transmitting baboon viruses to humans and possibly establishing new human infectious diseases. Retrospective analysis of tissues from two human transplant recipients with end-stage hepatic disease who died 70 and 27 days after the transplantation of baboon livers revealed the presence of two simian retroviruses of baboon origin, simian foamy virus (SFV) and baboon endogenous virus (BaEV), in multiple tissue compartments. The presence of baboon mitochondrial DNA was also detected in these same tissues, suggesting that xenogeneic "passenger leukocytes" harboring latent or active viral infections had migrated from the xenografts to distant sites within the human recipients. The persistence of SFV and BaEV in human recipients throughout the posttransplant period underscores the potential infectious risks associated with xenotransplantation.