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Clostridium difficileinfection (CDI) is a leading cause of infectious diarrhea in solid organ transplant recipients (SOT). We aimed to assess incidence, risk factors, and outcome of CDIwithin the Swiss Transplant Cohort Study (STCS). We performed a case‐control study of SOTrecipients in the STCSdiagnosed with CDIbetween May 2008 and August 2013. We matched 2 control subjects per case by age at transplantation, sex, and transplanted organ. A multivariable analysis was performed using conditional logistic regression to identify risk factors and evaluate outcome of CDI. Two thousand one hundred fifty‐eight SOTrecipients, comprising 87 cases of CDIand 174 matched controls were included. The overall CDIrate per 10 000 patient days was 0.47 (95% confidence interval ([CI] 0.38‐0.58), with the highest rate in lung (1.48, 95% CI0.93‐2.24). In multivariable analysis, proven infections (hazard ratio [HR] 2.82, 95% CI1.29‐6.19) and antibiotic treatments (HR 4.51, 95% CI2.03‐10.0) during the preceding 3 months were independently associated with the development of CDI. Despite mild clinical presentations, recipients acquiring CDIposttransplantation had an increased risk of graft loss (HR2.24, 95% CI1.15‐4.37; P= .02). These findings may help to improve the management of SOTrecipients. The authors demonstrate that despite mild clinical presentations and good clinical responses, Clostridium difficileinfections are associated with an increased risk of graft loss in the Swiss Transplant Cohort Study.

BKpolyomavirus (BKPyV) causes premature kidney transplant (KT) failure in 1–15% of patients. Because antivirals are lacking, most programs screen for BKPyV‐viremia and, if positive, reduce immunosuppression. To evaluate the relationship of viremia and BKPyV‐specific immunity, we examined prospectively cryopreserved plasma and peripheral blood mononuclear cells at the time of transplantation (T0) and at 6 mo (T6) and 12 mo (T12) after transplant from 28 viremic KTpatients and 68 nonviremic controls matched for the transplantation period. BKPyVIgG seroprevalence was comparable between cases (89.3%) and controls (91.2%; p = 0.8635), but cases had lower antibody levels (p = 0.022) at T0. Antibody levels increased at T6 and T12 but were not correlated with viremia clearance. BKPyV‐specific T cell responses to pools of overlapping 15mers (15mer peptide pool [15mP]) or immunodominant CD8 9mers (9mer peptide pool [9mP]) from the early viral gene region were not different between cases and controls at T0; however, clearance of viremia was associated with stronger 9mP responses at T6 (p = 0.042) and T12 (p = 0.048), whereas 15mP responses were not informative (T6 p = 0.359; T12 p = 0.856). BKPyV‐specific T cells could be expanded in vitrofrom all patients after transplant, permitting identification of 78 immunodominant 9mer epitopes including 50 new ones across different HLAclass I. Thus, 9mP‐responses may be a novel marker of reconstituting CD8 T cell function that warrants further study as a complement of plasma BKPyVloads for guiding immunosuppression reduction. Interferon‐γ ELISpot responses of CD8 T cells to immunodominant 9mer epitopes from the BK polyomavirus early viral gene region significantly increased in BK viremic patients but not in nonviremic controls, and are associated with viremia clearance posttransplantation, while standard 15mer T cell responses or antibody levels were uninformative.

We assessed the impact of antiviral preventive strategies on the incidence of herpes simplex virus (HSV) and varicella‐zoster virus (VZV) infections in a nationwide cohort of transplant recipients. Risk factors for the development of HSVor VZVinfection were assessed by Cox proportional hazards regression. We included 2781 patients (56% kidney, 20% liver, 10% lung, 7.3% heart, 6.7% others). Overall, 1264 (45%) patients received antiviral prophylaxis (ganciclovir or valganciclovir, n = 1145; acyclovir or valacyclovir, n = 138). Incidence of HSVand VZVinfections was 28.9 and 12.1 cases, respectively, per 1000 person‐years. Incidence of HSVand VZVinfections at 1 year after transplant was 4.6% (95% confidence interval [CI]3.5–5.8) in patients receiving antiviral prophylaxis versus 12.3% (95% CI10.7–14) in patients without prophylaxis; this was observed particularly for HSVinfections (3% [95% CI2.2–4] versus 9.8% [95% CI8.4–11.4], respectively). A lower rate of HSVand VZVinfections was also seen in donor or recipient cytomegalovirus‐positive patients receiving ganciclovir or valganciclovir prophylaxis compared with a preemptive approach. Female sex (hazard ratio [HR]1.663, p = 0.001), HSVseropositivity (HR5.198, p < 0.001), previous episodes of rejection (HR1.95, p = 0.004), and use of a preemptive approach (HR2.841, p = 0.017) were significantly associated with a higher risk of HSVinfection. Although HSVand VZVinfections were common after transplantation, antiviral prophylaxis significantly reduced symptomatic HSVinfections. In this national cohort of solid organ transplant recipients, patients receiving antiviral prophylaxis either with (val)ganciclovir or (val)acyclovir have a lower incidence of herpes simplex virus infections compared to patients managed with a preemptive approach for cytomegalovirus infection or not receiving antiherpes prophylaxis.

The epidemiology of respiratory viruses (RVs) in lung transplant recipients (LTRs) and the relationship of RVs to lung function, acute rejection (AR) and opportunistic infections in these patients are not well known. We performed a prospective cohort study (2009–2014) by collecting nasopharyngeal swabs (NPSs) from asymptomatic LTRsduring seasonal changes and from LTRswith upper respiratory tract infectious disease (URTID), lower respiratory tract infectious disease (LRTID) and AR. NPSswere analyzed by multiplex polymerase chain reaction. Overall, 1094 NPSswere collected from 98 patients with a 23.6% positivity rate and mean follow‐up of 3.4 years (interquartile range 2.5–4.0 years). Approximately half of URTIDs(47 of 97, 48.5%) and tracheobronchitis cases (22 of 56, 39.3%) were caused by picornavirus, whereas pneumonia was caused mainly by paramyxovirus (four of nine, 44.4%) and influenza (two of nine, 22.2%). In LTRswith LRTID, lung function changed significantly at 1 mo (p = 0.03) and 3 mo (p = 0.04). In a nested case–control analysis, ARwas associated with RVs (hazard ratio [HR]6.54), Pseudomonas aeruginosawas associated with LRTID(HR8.54), and cytomegalovirus (CMV)replication or disease was associated with URTID(HR2.53) in the previous 3 mo. There was no association between RVs and Aspergillusspp. colonization or infection (HR0.71). In conclusion, we documented a high incidence of RVinfections in LTRs. LRTIDproduced significant lung function abnormalities. Associations were observed between ARand RVs, between P. aeruginosacolonization or infection and LRTID, and between CMVreplication or disease and URTID. A large prospective study of the epidemiology of respiratory viruses in lung transplant recipients using molecular assays demonstrates a very high incidence of respiratory viral infection and an association between respiratory virus infectious diseases, immediate allograft dysfunction, and the development of acute rejection and opportunistic infection.

Transplant patients are at increased risk of viral complications due to impaired control of viral replication, resulting from HLAmismatching between graft and host and the immunosuppression needed to avert alloimmune reactions. In the past decade, quantitative viral load measurements have become widely available to identify patients at risk and to inform treatment decisions with respect to immunosuppressive drugs and antiviral therapies. Because viral loads are viewed as the result of viral replication and virus‐specific immune control, virus‐specific T cell monitoring has been explored to optimize management of adenovirus, BKpolyomavirus and cytomegalovirus (“ABC”) in transplant patients. Although most studies are descriptive using different technologies, the overall results show that the quantity and quality of virus‐specific T cells inversely correlate with viral replication, whereby strong cellular immune responses are associated with containment of viral replication. The key obstacles to the introduction of assays for virus‐specific T cells into clinical practice is the definition of reliable cutoffs for clinical decision making, the poor negative predictive value of some assays, and the absence of interventional trials justifying changes of antiviral treatment or immunosuppression. More clinical research is needed using optimized assays and targets before standardization and commutability can be envisaged as achieved for viral load testing. This minireview summarizes current state‐of‐the‐art virus‐specific T cell responses, including assay principles, their role in controlling viral replication, and areas for clinical use in transplant patients.

Uncontrolled BK polyomavirus (BKPyV) replication in kidney transplant recipients (KTRs) causes polyomavirus‐associated nephropathy and allograft loss. Reducing immunosuppression is associated with clearing viremia and nephropathy and increasing BKPyV‐specific T cell responses in most patients; however, current immunoassays have limited sensitivity, target mostly CD4+T cells, and largely fail to predict onset and clearance of BKPyV replication. To characterize BKPyV‐specific CD8+T cells, bioinformatics were used to predict 9mer epitopes in the early viral gene region (EVGR) presented by 14 common HLAs in Europe and North America. Thirty‐nine EVGR epitopes were experimentally confirmed by interferon‐γ enzyme‐linked immunospot assays in at least 30% of BKPyV IgG–seropositive healthy participants. Most 9mers clustered in domains, and some were presented by more than one HLA class I, as typically seen for immunodominant epitopes. Specific T cell binding using MHC class I streptamers was demonstrated for 21 of 39 (54%) epitopes. In a prospective cohort of 118 pediatric KTRs, 19 patients protected or recovering from BKPyV viremia were experimentally tested, and 13 epitopes were validated. Single HLA mismatches were not associated with viremia, suggesting that failing immune control likely involves multiple factors including maintenance immunosuppression. Combining BKPyV load and T cell assays using immunodominant epitopes may help in evaluating risk and reducing immunosuppression and may lead to safe adoptive T cell transfer. To better characterize BK polyomavirus–specific CD8+ T cells, the authors identify 39 mostly novel immunodominant 9mer epitopes in the early viral gene region by ELISpot, CD107a, killing assay, and streptamer binding, and find that most 9mers cluster in domains, and that some are presented by more than one HLA class I.

BK polyomavirus (BKPyV) replication causes nephropathy and premature kidney transplant failure. Insufficient BKPyV‐specific T cell control is regarded as a key mechanism, but direct effects of immunosuppressive drugs on BKPyV replication might play an additional role. We compared the effects of mammalian target of rapamycin (mTOR)‐ and calcineurin‐inhibitors on BKPyV replication in primary human renal tubular epithelial cells. Sirolimus impaired BKPyV replication with a 90% inhibitory concentration of 4 ng/mL by interfering with mTOR–SP6‐kinase activation. Sirolimus inhibition was rapid and effective up to 24 h postinfection during viral early gene expression, but not thereafter, during viral late gene expression. The mTORC‐1 kinase inhibitor torin‐1 showed a similar inhibition profile, supporting the notion that early steps of BKPyV replication depend on mTOR activity. Cyclosporine A also inhibited BKPyV replication, while tacrolimus activated BKPyV replication and reversed sirolimus inhibition. FK binding protein 12kda (FKBP‐12) siRNA knockdown abrogated sirolimus inhibition and increased BKPyV replication similar to adding tacrolimus. Thus, sirolimus and tacrolimus exert opposite effects on BKPyV replication in renal tubular epithelial cells by a mechanism involving FKBP‐12 as common target. Immunosuppressive drugs may therefore contribute directly to the risk of BKPyV replication and nephropathy besides suppressing T cell functions. The data provide rationales for clinical trials aiming at reducing the risk of BKPyV replication and disease in kidney transplantation. Comparing direct effects of immunosuppressive drugs on BK polyomavirus replication in primary human renal proximal tubular epithelial cells, this study demonstrates that the virus is inhibited by sirolimus but activated by tacrolimus through a mechanism involving competitive binding to the small host cell protein FKBP‐12.

BK polyomavirus (BKPyV) infection represents a major problem in transplantation, particularly for renal recipients developing polyomavirus‐associated nephropathy (PyVAN). The possibility that BKPyV may also be oncogenic is not routinely considered. Twenty high‐grade renourinary tumors expressing polyomavirus large T antigen in the entirety of the neoplasm in 19 cases, including the metastases in six, have been reported in transplant recipients with a history of PyVAN or evidence of BKPyV infection. Morphological and phenotypical features consistent with inactivation of the tumor suppressors pRB and p53 were found in the bladder tumors, suggesting a carcinogenesis mechanism involving the BKPyV large tumor oncoprotein/antigen. The pathogenesis of these tumors is unclear, but given the generally long interval between transplantation and tumor development, the risk for neoplasms after BKPyV infections may well be multifactorial. Other elements potentially implicated include exposure to additional exogenous carcinogens, further viral mutations, and cell genomic instability secondary to viral integration, as occurs with the Merkel cell PyV–associated carcinoma. The still scarce but increasingly reported association between longstanding PyVAN and renourinary neoplasms requires a concerted effort from the transplant community to better understand, diagnose, and treat the putative association between the BKPyV and these neoplasms.

In a pediatric kidney transplant recipient, retrospective serology studies indicate primary JC polyomavirus infection that remained undetected by the current screening recommendations for BK polyomavirus and progressed to polyomavirus‐associated nephropathy.

The authors characterize BK virus–specific cellular and humoral immunity in immunocompetent controls across a wide age range and in renal transplant recipients, and show that the level of immunity reflects phases of active viral replication either after primary infection in childhood or during reactivation after transplantation.

In this nationwide cohort of solid organ transplant recipients, the authors find that the incidence of cytomegalovirus disease is similar irrespective of the antiviral preventive strategy used (preemptive vs. prophylaxis), although patients who received antiviral prophylaxis have better graft failure‐free survival.

This study analyzes the histological features associated with the resolution of polyomavirus nephropathy, and shows that resolution is regularly associated with a significant increase in intraepithelial lymphocytes and interstitial infl ammation, morphologically indistinguishable from interstitial rejection. See editorial by Randhawa on page 1379.

The human BK polyomavirus (BKV) is the major cause of polyomavirus‐associated nephropathy (PyVAN) putting 1–15% of kidney transplant patients at risk of premature allograft failure, but is less common in other solid organ transplants. Because effective antiviral therapies are lacking, screening kidney transplant patients for BKV replication in urine and blood has become the key recommendation to guide the reduction of immunosuppression in patients with BKV viremia. This intervention allows for expanding BKV‐specific cellular immune responses, curtailing of BKV replication in the graft, and clearance of BKV viremia in 70–90% patients. Postintervention rejection episodes occur in 8–12%, most of which are corticosteroid responsive. Late diagnosis is faced with irreversible functional decline, poor treatment response, and graft loss. Adjunct therapies such as cidofovir, leflunomide and intravenous immunoglobulins have been used, but the benefit is not documented in trials. Retransplantation after PyVAN is largely successful, but requires close monitoring for recurrent BKV viremia.

Polyomavirus BK (BKV)‐associated nephropathy causes premature kidney transplant (KT) failure. BKV viruria and viremia are biomarkers of disease progression, but associated risk factors are controversial. A total of 682 KT patients receiving basiliximab, mycophenolic acid (MPA), corticosteroids were randomized 1:1 to cyclosporine (CsA) or tacrolimus (Tac). Risk factors were analyzed in 629 (92.2%) patients having at least 2 BKV measurements until month 12 posttransplant. Univariate analysis associated CsA‐MPA with lower rates of viremia than Tac‐MPA at month 6 (10.6% vs. 16.3%, p = 0.048) and 12 (4.8% vs. 12.1%, p = 0.004) and lower plasma BKV loads at month 12 (3.9 vs. 5.1 log10copies/mL; p = 0.028). In multivariate models, CsA‐MPA remained associated with less viremia than Tac‐MPA at month 6 (OR 0.60; 95% CI 0.36–0.99) and month 12 (OR 0.33; 95% CI 0.16–0.68). Viremia at month 6 was also independently associated with higher steroid exposure until month 3 (OR 1.19 per 1 g), and with male gender (OR 2.49) and recipient age (OR 1.14 per 10 years) at month 12. The data suggest a dynamic risk factor evolution of BKV viremia consisting of higher corticosteroids until month 3, Tac‐MPA compared to CsA‐MPA at month 6 and Tac‐MPA, older age, male gender at month 12 posttransplant.

ABSTRACTPCR-based molecular assays have a central role in polyomavirus diagnostics. To assure optimal performance, target sequences should be regularly updated according to newly available sequences. The aim of this study was to review our in-house polyomavirus BK (BKV) and JC (JCV) real-time PCR assays. Database analysis revealed variations in the BKV target region which might affect the assay performance, while no significant changes were found in the JCV target region. We compared two degenerate versions of our BKV primers which accommodated at least 95% of all published genetic variants. Dilutions of cloned viral genomic DNA and probit analysis indicated an analytical sensitivity of the updated BKV assay of 4.15 copies/reaction and that of the JCV assay was 3.37 copies/reaction. The specificity was assessed by testing JCV- and BKV-positive samples that showed no cross-reactivity. The performance of the original and updated BKV assay was compared in 101 urine and 200 plasma samples submitted to our routine diagnostic laboratory revealed similar quantitative results. We conclude that our JCV and updated BKV real-time PCR assays are robust and detect rare variants possibly encountered in the clinical routine.

Early detection of polyomavirus BK (BKV) viremia and reduction of immunosuppression is recommended for preventing polyomavirus‐associated nephropathy (PyVAN), but systematic histological evaluations were not performed in previous studies. We routinely screen for decoy cells and, if positive, measure plasma BKV‐loads. In a cohort of 203 consecutive renal transplantations performed from 2005–2008, 38 patients (19%) developed BKV‐viremia and were treated with reduction of immunosuppression. Based on subsequent allograft biopsy results and peak BKV‐viremia, patients were assigned to three groups: (i) definitive PyVAN (n = 13), (ii) presumptive PyVAN defined by plasma BKV‐loads of ≥4 log10copies/ml (n = 17) and (iii) low BKV‐viremia (n = 8). Clearance of BKV‐viremia was achieved in 35/38 patients (92%) and subsequent clinical rejection occurred in 3/35 patients (8.6%), both without any difference among the groups. Patients with definitive PyVAN had higher peak plasma BKV‐loads and required longer time for clearance (8.8 vs. 4.6 vs. 2.9 months; p = 0.001). However, allograft function remained stable from baseline to last follow‐up at 34 months (range 18–60) in all three groups with median serum creatinine of 1.6 mg/dl, 1.6 mg/dl and 1.3 mg/dl, respectively. We conclude that screening for BKV‐replication and reduction of immunosuppression is an effective strategy to preserve medium‐term allograft function even in patients developing definitive PyVAN.

Polyomavirus‐associated nephropathy (PyVAN) is rare in nonrenal solid organ transplantation and only limited information is available from single cases. We describe a 67‐year‐old female presenting with hypertension and progressive kidney failure due to PyVAN 60 months after lung transplantation. Plasma BK virus (BKV) loads were 4.85 log10copies/mL at diagnosis and cleared slowly over 14 months after switching from tacrolimus, mycophenolate and prednisone to low‐dose tacrolimus, sirolimus and leflunomide, the latter being discontinued for anemia and diarrhea. BKV‐ and JC virus‐specific immunoglobulins were detectable prior to transplantation. Only BKV‐specific IgG and IgM increased during follow‐up. BKV‐specific T cells were detectable in blood following in vitroexpansion, but cleared with reincreased sirolimus, yet BKV viremia remained undetectable. We identified eight other cases of PyVAN in nonrenal solid organ transplantation including lung (n = 1), heart (n = 6) and pancreas (n = 1). Overall, diagnosis was later than commonly seen in kidney transplants (median 18 months, interquartile range 10–29). Seven patients were male, five received triple immunosuppression consisting of tacrolimus, mycophenolate, prednisone. Immunosuppression was reduced in four cases and cidofovir and/or leflunomide administered in five and two cases, respectively. Renal function deteriorated in five requiring hemodialysis in four. We discuss mTOR inhibitors versus cidofovir and leflunomide as potential PyVAN rescue therapy.

The authors provide an introduction and overview of the Transplant Peer Review Network, a consortium of transplantation journals launched in January 2016.

Fast BK virus (BKV) replication in renal tubular epithelial cells drives polyomavirus-BK-associated nephropathy (PVAN) to premature kidney transplant (KT) failure. BKV also replicates in urothelial cells, but remains asymptomatic in two-thirds of affected KT patients. Comparing 518 day-matched plasma-urine samples from 223 KT patients, BKV loads were ?3000-fold higher in urine than in plasma (p < 0.000001). Molecular and quantitative parameters indicated that >95 of urine BKV loads resulted from urothelial replication and <5 from tubular epithelial replication. Fast BKV replication dynamics in plasma and urine with half-lives of <12 h accounted for daily urothelial and tubular epithelial cell loss of 4×107and 6×107, respectively. BKV dynamics in both sites were only partly linked, with full and partial discordance in 36 and 32, respectively. Viral expansion was best explained by models where BKV replication started in the kidney followed by urothelial amplification and tubular epithelial cell cross-feeding reaching a dynamic equilibrium after ?10 weeks. Curtailing intrarenal replication by 50 was ineffective and >80 was required for clearing viremia within 7 weeks, but viruria persisted for >14 weeks. Reductions >90 cleared viremia and viruria by 3 and 10 weeks, respectively. The model was clinically validated in prospectively monitored KT patients supporting >80 curtailing for optimal interventions.

The human polyomavirus BK (BKV) causes nephropathy and hemorrhagic cystitis in kidney and bone marrow transplant patients, respectively. The anti-viral cidofovir (CDV) has been used in small case series but the effects on BKV replication are unclear, since polyomaviruses do not encode viral DNA polymerases. We investigated the effects of CDV on BKV(Dunlop) replication in primary human renal proximal tubule epithelial cells (RPTECs). CDV inhibited the generation of viral progeny in a dose-dependent manner yielding a 90 reduction at 40 ?gmL. Early steps such as receptor binding and entry seemed unaffected. Initial large T-antigen transcription and expression were also unaffected, but subsequent intra-cellular BKV DNA replication was reduced by >90. Late viral mRNA and corresponding protein levels were also 90 reduced. In uninfected RPTECs, CDV 40 ?gmL reduced cellular DNA replication and metabolic activity by 7 and 11 in BrdU and WST-1 assays, respectively. BKV infection increased DNA replication to 142 and metabolic activity to 116, respectively, which were reduced by CDV 40 ?gmL to levels of uninfected untreated RPTECs. Our results show that CDV inhibits BKV DNA replication downstream of large T-antigen expression and involves significant host cell toxicity. This should be considered in current treatment and drug development.

Polyoma BK virus (BKV)-associated nephropathy (PVAN) is a relevant cause of poor renal allograft survival. In a prospective analysis, we monitored BKV DNA in blood and urine samples from 62 consecutive pediatric kidney recipients. In patients with BKV replication, we analyzed the impact of reduction of maintenance immunosuppression on viral load kinetics and PVAN in patients with BKV replication. BKV-specific immunity was concomitantly evaluated on blood samples of viremic patients, by measuring the frequency of BKV-specific interferon-?-producing and cytotoxic T cells, and BKV IgG antibody levels. At a median follow-up of 24 months, BK viruria was observed in 39 of 62 patients, while BK viremia developed in 13 patients (21). In all viremic patients, immunosuppression reduction resulted in the clearance of viremia, and prevented development of PVAN, without increasing the rate of acute rejection or causing graft dysfunction. As a consequence of immunosuppression adjustment, an expansion of BKV-specific cellular immunity was observed that coincided with viral clearance. We conclude that treating pediatric kidney transplant patients pre-emptively with immunosuppression reduction guided by BKV DNA in blood is safe and effective to prevent onset of PVAN. BKV-specific cellular immunity may be useful to guide this intervention.

Morbidität und Mortalität einer Virusinfektion wird maßgeblich bestimmt durch Determinanten des Patienten (z. B. die Qualität und Quantität der Immuneffektoren, Grunderkrankung, Alter), des Grafts (z. B. Organ, HLA-mismatching, Schädigung, Permissivität für Virusinfektion) und des Virus (z. B. Genus, Tropismus, Fitness, Virostatikaresistenz). Diese «ménage-à-trois» von Patient, Graft und Virus wird durch zum Teil dosisabhängige Faktoren moduliert wie Immunsuppression, lokales und systemisches Entzündungs- und Zytokinmuster, Gabe von Immunglobulinen und/oder Virostatika. Diese Konzepte sind bei Infektionen mit Zytomegalievirus (CMV) am besten untersucht. Drei Gründe sind dafür verantwortlich: 1. Klinisch-virologische Definitionen von Infektion und Erkrankung; 2. quantitative Testverfahren mit schneller Ergebniserstellung; 3. Verfügbarkeit wirksamer Virostatika. Aus der spezifischen Transplantationskonstellation ergibt sich der prophylaktische, präemptive oder therapeutische Einsatz von Virostatika und Immunglobulinen. Weitere Fortschritte könnten sich durch die bessere Charakterisierung der zellulären Immunität ergeben. Mit der breiten Verfügbarkeit von quantitativen molekularen Messverfahren sind nun ebenfalls Fortschritte bei anderen, transplantationsrelevanten Virusinfektionen zu erwarten wie zum Beispiel Epstein-Barr-Virus, Polyomavirus BK oder respiratorische Viren.

We review BK-virus nephropathy (BKN) as a new complication that increasingly affects renal allografts and causes dysfunction. Since starting in 1996, we have seen 11 cases. Currently, the prevalence of BKN is 3% in our graft biopsies. The diagnosis can only be made histologically. The virus affects tubular epithelial cells that show characteristic intranuclear inclusion bodies. The major reason for impaired graft function and a possible way for viral particles to gain access to the blood via peritubular capillaries is necrosis of infected epithelial cells. BK-virus DNA in the plasma, which can be detected by PCR, is closely associated with nephropathy. BK-virus does not stimulate tubular MHC-class II expression as judged by immunofluorescence double labelling. The inflammatory response is inconsistent and the frequency of rejection episodes is not increased during disease. Clinical manifestation of viral nephropathy evolves in several stages. (i) Initial, asymptomatic and reversible activation of the virus, judged by the presence of inclusion bearing cells in the urine. (ii) High dose immunosuppressive drug regimens, often including tacrolimus. (iii) Tubular injury and viraemia as additional promoting conditions. BKN nephropathy was associated with graft loss in 45% of our patients. The remaining patients with persistent viral nephropathy showed renal dysfunction (serum creatinine levels on average 150% above baseline readings). Currently, no established antiviral therapy is available. We discuss attempts to lower immunosuppression as a means to control viral replication. We propose a diagnostic algorithm for screening and monitoring the disease.

The biosynthesis and processing of the vacuolar (lysosomal) proteinase yscB was followed in vivo and in vitro. In vitro transcription–translation of the cloned proteinase yscB gene results in a high mol. wt precursor protein (HMr‐precursor) of Mr = 73,000. In vivo a precursor of identical mol. wt is found in sec6l mutant cells deficient in translocation of secretory protein precursors into the lumen of the endoplasmic reticulum (ER). In contrast to N‐glycosylated intermediate mol. wt forms of proteinase yscB, the HMr‐precursor of the enzyme is not N‐glycosylated, indicating its appearance outside the ER. sec18 Mutant cells, wild type for translocation of proteins into the ER but blocked in the delivery step of secretory proteins to the Golgi apparatus, accumulate a lower mol. wt precursor (LMr‐precursor) of proteinase yscB of Mr = 41,500. Processing of the HMr‐precursor of proteinase yscB to the LMr‐precursor requires proteolytic cleavage which is independent of the proteinase yscA gene product, a protein known to be involved in the processing event of the LMr‐precursor of proteinase yscB to the mature enzyme of Mr = 33,000. A LMr‐precursor of proteinase yscB of Mr = 42,000 accumulates in proteinase yscA‐deficient mutant cells (allele pep4‐3). This form is enzymatically active. Incubation in vitro of the LMr‐precursor of proteinase yscB with mature proteinase yscA leads to further activation and to processing of the enzyme yielding the mature 33,000 Mr protein.(ABSTRACT TRUNCATED AT 250 WORDS)

Tumors obtained from v-Ha-ras-transformed PB-3c cells are characterized by autocrine interleukin-3 (IL3) expression, which occurs either without (class I tumors) or with enhanced transcription (class II tumors). To address possible post-transcriptional mechanisms of IL3 expression, IL3 mRNA stability was examined in both tumor classes. Increased stability of IL3 mRNA was detected in class I tumor lines (t1/2 > 3 h), whereas rapid decay of IL3 transcripts (t1/2 < 0.5 h) was found in class II tumor lines. In both tumor classes, the c-myc and interleukin-6 transcripts were short-lived. Transcripts of a constitutively expressed IL3 reporter gene were stable in class I tumor cells but unstable in class II tumor cells, suggesting that IL3 mRNA stabilization involved a trans-acting mechanism. Rapid decay of IL3 reporter transcripts was observed in untransformed PB-3c as well as in v-Ha-ras expressing precursor cells linking transcript stabilization to the tumor stage. Reporter transcript stabilization in class I tumor cells correlated with increased IL3 production. Deletion of the AU-rich element from the IL3 reporter gene further augmented IL3 mRNA levels as well as IL3 production, suggesting that the stabilizing mechanism in class I tumor cells is not equivalent to AU-rich element deletion.

The biosynthesis and processing of the vacuolar (lysosomal) proteinase yscB was followed in vivo and in vitro. In vitro transcription–translation of the cloned proteinase yscB gene results in a high mol. wt precursor protein (HMr‐precursor) of Mr = 73,000. In vivo a precursor of identical mol. wt is found in sec6l mutant cells deficient in translocation of secretory protein precursors into the lumen of the endoplasmic reticulum (ER). In contrast to N‐glycosylated intermediate mol. wt forms of proteinase yscB, the HMr‐precursor of the enzyme is not N‐glycosylated, indicating its appearance outside the ER. sec18 Mutant cells, wild type for translocation of proteins into the ER but blocked in the delivery step of secretory proteins to the Golgi apparatus, accumulate a lower mol. wt precursor (LMr‐precursor) of proteinase yscB of Mr = 41,500. Processing of the HMr‐precursor of proteinase yscB to the LMr‐precursor requires proteolytic cleavage which is independent of the proteinase yscA gene product, a protein known to be involved in the processing event of the LMr‐precursor of proteinase yscB to the mature enzyme of Mr = 33,000. A LMr‐precursor of proteinase yscB of Mr = 42,000 accumulates in proteinase yscA‐deficient mutant cells (allele pep4‐3). This form is enzymatically active. Incubation in vitro of the LMr‐precursor of proteinase yscB with mature proteinase yscA leads to further activation and to processing of the enzyme yielding the mature 33,000 Mr protein.(ABSTRACT TRUNCATED AT 250 WORDS)

After v-H-ras expression, the interleukin 3 (IL-3)-dependent PB-3c mast cells progress in vivo to two different classes of IL-3 autocrine tumors. Class I tumors show a germline configuration of the IL-3 gene and represent more than 90% of tumors analyzed so far. Somatic cell fusion of class I tumor lines with the nontumorigenic parental PB-3c resulted in loss of oncogenic IL-3 expression by a posttranscriptional mechanism with concomitant tumor suppression. Class II tumors arise rarely and contain an insertion in one IL-3 allele. This alteration was linked to enhanced IL-3 gene transcription. For one tumor, the insertion was shown to be an endogenous retroviral element (intracisternal A-particle). Cell hybrids of class II tumors with PB-3c remained IL-3 independent, expressed IL-3, and formed tumors rapidly. These results suggest that the v-H-ras oncogene synergizes with a recessive and a dominant lesion in class I and II tumors, respectively, both of which lead to the autocrine production of IL-3.

Interleukin 3 (IL-3)-dependent PB-3c mouse mastocytes can be transformed by the v-Ha-ras oncogene to generate autocrine IL-3-producing mastocytomas. Hybrid cell lines were constructed by fusing an IL-3-producing mastocytoma cell line with its IL-3-dependent normal parental cell. Unlike the mastocytoma parent cell line, hybrid cell lines required growth factor for in vitro proliferation, indicating that the IL-3-dependent phenotype is dominant. IL-3 mRNA, expressed at high levels in the tumor cells, appeared down-regulated in the cell hybrids. In contrast, p21v-Ha-ras levels were not reduced in the hybrids. The hybrid lines generated tumors in vivo with drastically prolonged latency times when compared to the tumor parent (10 versus 2 weeks). We propose that down-regulation of IL-3 mRNA production after cell fusion is responsible for the loss of growth autonomy in the hybrids and is likely to play a role in the partial suppression of tumor formation in vivo. Our data are consistent with the hypothesis that a tumor suppressor, present in PB-3c cells, acts as a negative regulator of IL-3 expression.

Summary Plasma separation was carried out using a new cellulose diacetate hollow-fibre module. The pore size of the membranes used was 0.2 µm and the cut-off at approximately 3 million dalton. Five litres of filtrate was separated in 80 min. The protein content of the filtrate was 60% of that in the blood plasma. When plasma and filtrate were compared, there was little difference in the proportions of the immunoglobulins. At the end of treatment, the immunoglobulin content of the plasma had been reduced to 40% of its initial concentration.

We analyzed the effect of rapamycin on autocrine mast cell tumor lines with abnormally stable interleukin-3 (IL-3) transcripts due to a defect in mRNA degradation. Rapamycin inhibited IL-3 mRNA expression specifically, while transcripts of IL-4 and IL-6 were not affected. As indicated by the use of the transcriptional inhibitor actinomycin D or by reporter constructs, inhibition was posttranscriptional and resulted from destabilization of the mRNA. Transcripts from transgenes lacking the AU-rich 3' untranslated region were refractory to drug-induced degradation, suggesting that these 3' sequences contain the target of the rapamycin effect. Rapamycin did not promote IL-3 mRNA degradation in cells of a tumor variant lacking expression of FKBP12, the binding protein of rapamycin. Experiments with wortmannin indicated that rapamycin does not act via p70S6 kinase. FK-506, another ligand of FKBP12 affecting the phosphatase calcineurin, did not antagonize but shared the effect of rapamycin. Our data fit a model whereby both FKBP12 and calcineurin target an unknown regulator of IL-3 mRNA turnover.

There is a maximum green density to which pure copper can be compacted and still exhibit no growth upon sintering. This limits the maximum density and thereby the maximum conductivity and strength that can be achieved with sintered copper bodies. The swelling of a part pressed above this critical compacting pressure is caused by premature closure of the outer pores of the compact, thus sealing off the egress of internally generated gases. In this study a number of carefully selected chemical compounds were added to the as-pressed compacts using two methods in an effort to find a way of keeping these outer pores temporarily open until all internal gases are eliminated. These pores are then sealed off in the later stages of sintering, resulting in a higher density body with improved electrical conductivity throughout. Two compounds proved to be most effective in accomplishing this without leaving a residue deleterious to conductivity. Two commercial copper powders were die pressed into thick discs and then impregnated with an aqueous salt solution, the preferred treatment. After a special sintering procedure, the best conductivity obtained in an 18 mm diameter × 10 mm thick disc using the most responsive powder and treatment was 93·9% IACS at the centre and 94·2% IACS at the surface. Untreated, the same powder showed a maximum centre conductivity of 89% IACS.

Die Wirkung des Berberins richtet sich hauptsächlich gegen die gelben Fermente. Im Vordergrund der Wirkung steht die Blockierung der Atmungskette durch Hemmung der Diaphorasen. Die besondere Empfindlichkeit des Tumorgewebes gegen Berberin ist auf den bei Tumoren bestehenden Mangel an gelben Fermenten zurückzuführen.

Zusammenfassung Mit Hilfe der Isotopentechnik (51Cr) wurde das Schicksal der Thrombocyten beim extrakorporalen Kreisverlauf verfolgt. Der beim EKK auftretende Thrombocytenverlust beruht z.T. auf Thrombocytenniederschlägen an der HLM, z.T. auf einer Zerstörung der Thrombocyten im Kreislauf des Patienten. Die Befunde sprechen dafür, daß dieser vermehrte Abbau über die Bildung von Thrombocytenaggregaten erfolgt. Ein Teil dieser Aggregate ist reversibel, da es nach Beendigung der Perfusion zu einem vorübergehenden Anstieg der Thrombocytenzahl im peripheren Blut kommt.

Zusammenfassung 1. In Blutkonserven kommt es in Abhängigkeit von der Lagerungszeit zu einem Anstieg des Siebungsdruckes und der Aktivität des plättchenagglutinierenden Faktors.