Respiratory viruses in medicolegal autopsies during the winter season 2021/2022: observations after reduction of coronavirus disease-19 (COVID-19) pandemic restrictions.

In the context of the coronavirus disease (COVID-19) pandemic, measures were taken to protect the population from infection. These were almost completely lifted in several countries in the spring of 2022. To obtain an overview of the spectrum of respiratory viruses encountered in autoptical routine case work, and their infectivity, all autopsy cases at the Institute of Legal Medicine in Frankfurt/M. with flu-like symptoms (among others) were examined for at least 16 different viruses via multiplex PCR and cell culture. Out of 24 cases, 10 were virus-positive in PCR: specifically, 8 cases with severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), 1 with respiratory syncytial virus (RSV), and 1 with SARS-CoV-2 and the human coronavirus OC43 (HCoV-OC43), as a double infection. The RSV infection and one of the SARS-CoV-2 infections were only detected due to the autopsy. Two SARS-CoV-2 cases (postmortem interval of 8 and 10 days, respectively) showed infectious virus in cell culture; the 6 other cases did not show infectious virus. In the RSV case, virus isolation by cell culture was unsuccessful (Ct value of 23.15 for PCR on cryoconserved lung tissue). HCoV-OC43 was measured as non-infectious in cell culture, with a Ct value of 29.57. The detection of RSV and HCoV-OC43 infections may shed light on the relevance of respiratory viruses other than SARS-CoV-2 in postmortem settings; however, further, more extensive studies are needed for a robust assessment of the hazard potential due to infectious postmortem fluids and tissues in medicolegal autopsy settings.

Detection and infectivity of SARS-CoV-2 in exhumated corpses.

Postmortem detection of severe acute respiratory syndrome coronavirus type 2 (SARS-CoV-2) after the exhumation of a corpse can become important, e.g. in the case of subsequent medical malpractice allegations. To date, data on possible detection periods [e.g. by reverse transcription polymerase chain reaction (RT-PCR)] or on the potential infectivity of the virus after an exhumation are rare. In the present study, these parameters were examined in two cases with a time span of approximately 4 months between day of death and exhumation. Using SARS-CoV-2 RT-PCR on swabs of both lungs and the oropharynx detection was possible with cycle threshold (Ct) values of about 30 despite signs of beginning decay. RT-PCR testing of perioral and perinasal swabs and swabs collected from the inside of the body bag, taken to estimate the risk of infection of those involved in the exhumation, was negative. Cell culture-based infectivity testing was negative for both, lung and oropharyngeal swabs. In one case, RT-PCR testing at the day of death of an oropharyngeal swab showed almost identical Ct values as postmortem testing of an oropharyngeal swab, impressively demonstrating the stability of viral RNA in the intact corpse. However, favorable climatic conditions in the grave have to be taken into account, as it was wintertime with constant low temperatures. Nevertheless, it was possible to demonstrate successful postmortem detection of SARS-CoV-2 infection following exhumation even after months in an earth grave.

Infectivity of deceased COVID-19 patients.

The duration of infectivity of SARS-CoV-2 (severe acute respiratory syndrome coronavirus 2) in living patients has been demarcated. In contrast, a possible SARS-CoV-2 infectivity of corpses and subsequently its duration under post mortem circumstances remain to be elucidated. The aim of this study was to investigate the infectivity and its duration of deceased COVID-19 (coronavirus disease) patients. Four SARS-CoV-2 infected deceased patients were subjected to medicolegal autopsy. Post mortem intervals (PMI) of 1, 4, 9 and 17 days, respectively, were documented. During autopsy, swabs and organ samples were taken and examined by RT-qPCR (real-time reverse transcription-polymerase chain reaction) for the detection of SARS-CoV-2 ribonucleic acid (RNA). Determination of infectivity was performed by means of virus isolation in cell culture. In two cases, virus isolation was successful for swabs and tissue samples of the respiratory tract (PMI 4 and 17 days). The two infectious cases showed a shorter duration of COVID-19 until death than the two non-infectious cases (2 and 11 days, respectively, compared to > 19 days), which correlates with studies of living patients, in which infectivity could be narrowed to about 6 days before to 12 days after symptom onset. Most notably, infectivity was still present in one of the COVID-19 corpses after a post-mortem interval of 17 days and despite already visible signs of decomposition. To prevent SARS-CoV-2 infections in all professional groups involved in the handling and examination of COVID-19 corpses, adequate personal safety standards (reducing or avoiding aerosol formation and wearing FFP3 [filtering face piece class 3] masks) have to be enforced for routine procedures.

[The virology of SARS-CoV-2]. / Die Virologie von SARS-CoV-2.

The novel severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has far-reaching effects on society, the economy and medical treatment. Therefore, it is all the more important to understand the characteristics of the virus and to utilize them in the diagnostics, treatment and epidemiology. This article firstly elucidates the medical significance of coronaviruses in general. Then angiotensin-converting enzyme 2 (ACE2) as the binding site of SARS-CoV­2 and the possible influence on disease susceptibility are explained. The gold standard for detection of an active SARS-CoV­2 infection is the direct detection of the pathogen with nucleic acid amplification techniques. At the onset of symptoms a swab of the upper airway is especially suitable due to the high viral load. At a later stage direct detection can be achieved in samples from the lower airway or in stool and anal swabs. Antibody tests cannot replace the direct detection of the pathogen; however, the detection of immunoglobulin G antibodies is of special interest for epidemiological questions (seroconversion time of sometimes several weeks). The plaque reduction neutralization test exclusively detects antibodies which neutralize viruses but the procedure is complicated and can only be carried out in secure laboratories (L3). In addition, the importance of these antibodies with respect to immunity against a second infection is uncertain. Thanks to modern techniques thousands of SARS-CoV­2 sequences are already available, which show a genomic variability. The D614G mutation in the S spikes seems to cause a higher infectiousness. Mutations can impair the diagnostics and treatment, which makes monitoring necessary.

[The virology of SARS-CoV-2]. / Die Virologie von SARS-CoV-2.

The new severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has far reaching effects on society, the economy and medical treatment. It is all the more important to understand the characteristics of the virus and to utilize them diagnostically, therapeutically and epidemiologically. This article firstly elucidates the medical importance of coronaviruses in general. Then angiotensin-converting enzyme 2 (ACE2) as the binding site of SARS-CoV­2 and the possible influence on the disease susceptibility are explained. The gold standard for detection of an active SARS-CoV­2 infection is the direct detection of the pathogen with nucleic acid amplification techniques. At the onset of symptoms, a swab of the upper airway is especially suitable due to the high viral load. At a later stage direct detection can be achieved in samples from the lower airway or a stool or anal swab. Antigen or antibody tests cannot replace the direct detection of the pathogen; however, the detection of immunoglobulin G antibodies are of special interest for epidemiological questions (seroconversion time of sometimes several weeks). The plaque reduction neutralization test exclusively detects antibodies which neutralize viruses but the procedure is complicated. In addition, the importance of these antibodies with respect to immunity against a second infection is uncertain. Thanks to modern techniques thousands of SARS-CoV­2 sequences are already available, which show a genomic variability. The D614G mutation in the S spikes seems to cause a higher infectiosity. Mutations can impair the diagnostics and treatment, which makes monitoring necessary.

[The virology of SARS-CoV-2]. / Die Virologie von SARS-CoV-2.

The new severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has far reaching effects on society, the economy and medical treatment. It is all the more important to understand the characteristics of the virus and to utilize them diagnostically, therapeutically and epidemiologically. This article firstly elucidates the medical importance of coronaviruses in general. Then angiotensin-converting enzyme 2 (ACE2) as the binding site of SARS-CoV­2 and the possible influence on the disease susceptibility are explained. The gold standard for detection of an active SARS-CoV­2 infection is the direct detection of the pathogen with nucleic acid amplification techniques. At the onset of symptoms, a swab of the upper airway is especially suitable due to the high viral burden. At a later stage direct detection can be achieved in samples from the lower airway or a stool or anal swab. Antigen or antibody tests cannot replace the direct detection of the pathogen; however, the detection of immunoglobulin G antibodies are of special interest for epidemiological questions (seroconversion time of sometimes several weeks). The plaque reduction neutralization test exclusively detects antibodies which neutralize viruses but the procedure is complicated. In addition, the importance of these antibodies with respect to immunity against a second infection is uncertain. Thanks to modern techniques thousands of SARS-CoV­2 sequences are already available, which show a genomic variability. The D614G mutation in the S spikes seems to cause a higher infectiosity. Mutations can impair the diagnostics and treatment, which makes monitoring necessary.

Efficacy and safety of sofosbuvir/ledipasvir for the treatment of patients with hepatitis C virus re-infection after liver transplantation.

BACKGROUND: Hepatitis C virus (HCV) infection is associated with a particularly poor outcome after liver transplantation. In December 2014, sofosbuvir/ledipasvir (SOF/LDV) fixed-dose combination (FDC) was approved for HCV genotype 1 and 4 in Europe. In orthotopic liver transplantation (OLT) recipients, the interferon-free treatment of HCV re-infection with novel direct-acting antivirals has been demonstrated to be safe and effective in clinical trials, but real-world data are missing. The aim of this study was to investigate the safety and efficacy of SOF/LDV FDC in OLT recipients in the real-life setting. METHODS: All consecutive OLT patients started on SOF/LDV FDC for 12 or 24 weeks at the University Medical Center Hamburg-Eppendorf and Medical School Hannover between October 2014 and August 2015 were retrospectively analyzed (n = 30). The primary efficacy endpoint was sustained virological response (SVR), i.e., absence of viremia 12 weeks after end of treatment (SVR 12). Liver function tests, creatinine, blood count, and HCV RNA (by polymerase chain reaction assay) were determined at each visit. RESULTS: SVR was achieved in 29/30 patients (96.67%) treated with SOF/LDV ± ribavirin (RBV) for 12 (n = 4) or 24 weeks (n = 25). Twenty-five patients (86.2%) received RBV. However, in 15 of the 25 patients, RBV administration had to be discontinued because of severe anemia (57.7%). One RBV-treated patient died of a myocardial infarction during antiviral therapy; this event was most likely not directly related to SOF/LDV. Aside from RBV-associated anemia, no severe side effects of the antiviral regimen were observed. CONCLUSION: Antiviral treatment with SOF/LDV is highly effective, safe, and well tolerated in OLT recipients. The addition of RBV often results in severe anemia, requiring dose reduction or discontinuation.

Hepatitis C virus core antigen testing in liver and kidney transplant recipients.

HCV RNA levels correlate with the long-term outcome of hepatitis C in liver transplant recipients. Nucleic acid testing (NAT) is usually used to confirm HCV reinfection and to examine viral loads after liver transplantation. HCV core antigen (HCVcoreAg) testing could be an alternative to NAT with some potential advantages including very low intra- and interassay variabilities and lower costs. The performance of HCVcoreAg testing in organ transplant recipients is unknown. We prospectively studied 1011 sera for HCV RNA and HCVcoreAg in a routine real-world setting including 222 samples obtained from patients after liver or kidney transplantation. HCV RNA and HCVcoreAg test results showed a consistency of 98% with a very good correlation in transplanted patients (r > 0.85). The correlation between HCV RNA and HCVcoreAg was higher in sera with high viral loads and in samples from patients with low biochemical disease. Patients treated with tacrolimus showed a better correlation between both parameters than individuals receiving cyclosporine A. HCV RNA/HCVcoreAg ratios did not differ between transplanted and nontransplanted patients, and HCV RNA and HCVcoreAg kinetics were almost identical during the first days after liver transplantation. HCVcoreAg testing can be used to monitor HCV viral loads in patients after organ transplantation. However, the assay is not recommended to monitor antiviral therapies.

Immunosuppression, liver injury and post-transplant HCV recurrence.

Hepatitis C virus (HCV) infection is a major cause for liver transplantation worldwide. Still, HCV re-infection of the graft occurs in almost all cases. Most liver transplant recipients experience episodes of graft hepatitis associated with fibrosis progression and graft failure. Clinical management of graft hepatitis can be challenging as in addition to rejection and HCV-induced hepatitis various other factors might be involved including toxic liver injury, steatohepatitis, ischaemic bile duct lesions or infections with other pathogens. Treatment options are often contradictory for different causes of graft hepatitis, and the role of distinct immunosuppressive drugs has been discussed controversially. Corticosteroids increase the infectivity of HCV by altering expression levels of entry factors and other immunosuppressive agents may have diverse effects on HCV replication and fibrosis progression. Interferon alpha-therapy of hepatitis C shows limited efficacy and tolerability in liver transplant recipients and may also cause rejection. In this review we summarize the current knowledge on mechanisms of liver injury in post-transplant hepatitis C, discuss the pros and cons of immunosuppressive agents in this specific setting and describe potential novel approaches to prevent HCV reinfection.

Hepatitis C virus enters human peripheral neuroblastoma cells - evidence for extra-hepatic cells sustaining hepatitis C virus penetration.

Patients with chronic hepatitis C virus (HCV) infection show an increased incidence of nervous system disorders such as chronic fatigue syndrome, depression and cognitive dysfunction. It is unclear whether this is because of HCV replication in the brain and in peripheral neuronal cells or to more indirect effects of HCV infection on the central or peripheral nervous system. The aim of this study was to investigate whether cells originating from these tissues are permissive for HCV cell entry, RNA replication and virus assembly. Among eight cell lines analysed, the human peripheral neuroblastoma cell line SKNMC expressed all HCV entry factors and was efficiently infected with HCV pseudoparticles (HCVpp) independent of the HCV genotype. All remaining cell types including human neuroblastoma and glioblastoma cell lines and microglial cells lacked expression of at least one host factor essential for HCV entry. When transfected with HCV luciferase reporter virus RNA, inoculated with HCV reporter viruses or challenged with high-titre cell culture-derived HCV, none of these cells supported detectable HCV RNA replication. Thus, in conclusion, this comprehensive screening did not reveal evidence directly strengthening the notion that HCV enters and replicates in the central nervous system. However, productive viral entry into the peripheral neuroblastoma cell line SKNMC indicates that HCV may penetrate into certain nonhepatic cell types which may serve as viral reservoirs and could modulate viral pathogenesis.

[Suspicion of a newly appearing hypertrophic obstructive cardiomyopathy in a patient treated for hepatocellular carcinoma]. / Verdacht auf neu aufgetretene hypertrophe obstruktive Kardiomyopathie bei hepatozellulärem Karzinom.

HISTORY AND ADMISSION FINDINGS: A 42-year-old women presented with shortness of breath, tachycardia and weakness to our department. Five years ago she had been diagnosed with hepatocellular carcinoma for which an extended hemihepatectomy had been performed. INVESTIGATIONS, DIAGNOSIS AND TREATMENT: The clinical examination revealed a systolic murmur over the artic region. Echocardiography showed an hypertrophed interventricular septum with signs like those in hypertrophic obstructive cardiomyopathy. Ultrasound demonstrated a cystic mass in the pelvis highly suspicious of a metastasis of a hepatocellular carcinoma. Fine needle biopsy confirmed the diagnosis of a metastatic lesion of hepatocellular carcinoma. Computed tomography demonstrated metastase in the lung and a space-occupying in the interventricular septum. The patients underwent resection of the lung and pelvic metastasis and died a few weeks later. CONCLUSION: This case demonstrates the rare occurence of a metastasis to the heart, imitating obstructive cardiac myopathy, in a patient with hepatocellular carcinoma.

Impaired TRAIL-dependent cytotoxicity of CD1c-positive dendritic cells in chronic hepatitis C virus infection.

Dendritic cells (DCs) play a central role in antiviral immunity. Conflicting data on DC function have been reported for hepatitis C virus (HCV) infection. In addition to antigen presentation and cytokine secretion, a subset of human DCs displays direct cytotoxic activity. It has been suggested that measles virus and human immunodeficiency virus (HIV) may enhance cytotoxicity of DCs potentially leading to apoptosis of activated T cells and subsequent down-regulation of antiviral immune responses. We demonstrate that CD1c-positive myeloid DCs, but not BDCA-4-positive plasmacytoid DCs, are able to kill different target cells mainly via tumour necrosis factor-related apoptosis-inducing ligand. The ability of CD1c+ DCs to lyze target cells was found to be completely impaired in patients with chronic hepatitis C (10 chronic HCV patients vs 10 healthy controls; P < 0.001) but not in patients with primary biliary cirrhosis. Successful antiviral therapy of chronic hepatitis C rescued the cytotoxicity of DCs. Myeloid DCs of HCV patients and healthy controls had a similar phenotype and endocytotic activity, however, the frequency of mDCs in the peripheral blood was lower (P = 0.004) and the allostimulatory function was weaker (P < 0.001) in chronic hepatitis C. Thus, in contrast to HIV and measles virus studies on monocyte-derived DCs, freshly isolated myeloid DCs of patients with hepatitis C do not show an increased but a completely abolished cytotoxic activity. The impaired DC cytotoxicity could represent a novel mechanism for the increased prevalence of autoimmunity in HCV infection.

Viral hepatitis after liver transplantation. Which immunosuppressive drugs should be recommended?

Treatment of hepatitis C after liver transplantation can be challenging as graft rejection and graft hepatitis caused by hepatitis C virus (HCV) may be difficult to distinguish. Immunosuppressive medications may significantly alter the course of hepatitis C in liver transplant recipients. Moreover, single substances have been shown to display antiviral effects in vitro while others are believed to have antifibrotic properties. This review summarizes the modes of action of different classes of immunosuppressive drugs used after liver transplantation and discusses pros and cons of individual drugs in the setting of HCV infection. No definite recommendation for an optimal immunosuppressive regimen can be given at this stage. The most important lesson learned during the last two decades is that acute rejection episodes have to be avoided in particular in hepatitis C since these are associated with reduced graft and patient survival. Further trials are urgently needed to clarify the role of different immunosuppressive compounds in hepatitis C after transplantation.

[Retroperitoneal abscess in a man with severe necrotizing pancreatitis]. / Retroperitonealabszess bei schwerer nekrotisierender Pankreatitis.

HISTORY AND ADMISSION FINDINGS: A 65-year-old man was admitted to the emergency unit of another hospital with severe acute abdominal pain. Medial laparotomy revealed a tear in the lower pole of the capsule of the enlarged spleen. Results of some laboratory tests became available only after the procedure: Lipase and amylase levels were markedly elevated at 5097 U/l (normal range < 100 U/l) and 1776 U/l (normal range < 60 U/l), respectively. These findings suggested acute pancreatitis as cause of the acute abdominal pain. Conservative treatment was initiated, but because of progressive clinical deterioration with fever, dyspnea and tachyarrhythmia he was transferred to our hospital for further treatment. INVESTIGATIONS, DIAGNOSIS AND TREATMENT: Chest X-ray revealed bilateral pleural effusion. Contrast-enhanced abdominal CT scan revealed an acute peripancreatic fluid collection and extensive pancreatic necrosis extending into the perirenal space, hilum of the spleen and transverse mesocolon. Despite the placement of multiple percutaneous pigtail catheters clinical improvement of the severe necrotizing pancreatitis was only temporary and repeated episodes of fever and septicaemia occured. Supported by a multidisciplinary consensus an endoscopic ultrasound (EUS)-guided transgastric drainage of the pancreatic abscess was performed with placement of two double-pigtail catheters and a nasocystic drain for irrigation. After this intervention the patient}s condition rapidly improved, inflammatory parameters normalized and the retroperitoneal abscess gradually resolved. CONCLUSION: A multidisciplinary consensus and modern interventional approaches such as EUS-guided endoscopic therapy are mandatory to successfully approach severe necrotizing pancreatitis and its complications.

[Sequelae of organ transplantation]. / Folgeerkrankungen nach Organtransplantation.

Transplant medicine has significantly changed the prognosis of diseases leading to terminal organ failure. It has evolved from an experimental procedure to standard therapy for liver, kidney and cardio-vascular diseases. Transplant medicine combines operative organ replacement with the management of severely ill patients before transplantation, as well as life-long follow-up of organ graft recipients. Ten year survival rates of 65% to over 90% have led to a steady increase of transplanted patients seen by general medical care providers which represents a challenge for practicing internists. Apart from organ-specific conditions, infectious, immunosuppressant-associated and metabolic consequences determine long-term survival. These include virus reactivation, graft rejection, anastomotic problems but more importantly general mortality determining factors such as diabetes, renal insufficiency and hypertension, which are often a consequence of immunosuppressant administration. They directly impact long-term survival. The awareness and treatment of these secondary conditions of organ transplantation in routine medical practice contributes significantly to secure the long term success of transplant medicine.

Effects of cyclosporine on human dendritic cell subsets.

Cyclosporine is widely used as an immunosuppressive agent after solid organ transplantation. Limited data are available on the modulation of human dendritic cells by cyclosporine. We investigated the effects of cyclosporine on the phenotype and function of human dendritic cell (DC) subsets. DCs were isolated from peripheral blood using magnetic bead-conjugated antibodies. Cyclosporine did not alter the ability of myeloid and plasmacytoid dendritic cells to take up antigens. Expression of the co-stimulatory molecule CD80 but not CD86 increased on both DC subsets when stimulated with cyclosporine. The ability of cyclosporine treated myeloid DCs to stimulate proliferation of allogenic PBMC was significantly reduced. Similarly, stimulation of memory CD8+ T cells by dendritic cells was impaired by cyclosporine pretreatment. In conclusion, cyclosporine differentially alters function and phenotype of myeloid dendritic cells leading to a partially impaired capacity to stimulate allogenic and autologous T cells.