Virological assessment of hospitalized patients with COVID-2019.

Coronavirus disease 2019 (COVID-19) is an acute infection of the respiratory tract that emerged in late 20191,2. Initial outbreaks in China involved 13.8% of cases with severe courses, and 6.1% of cases with critical courses3. This severe presentation may result from the virus using a virus receptor that is expressed predominantly in the lung2,4; the same receptor tropism is thought to have determined the pathogenicity-but also aided in the control-of severe acute respiratory syndrome (SARS) in 20035. However, there are reports of cases of COVID-19 in which the patient shows mild upper respiratory tract symptoms, which suggests the potential for pre- or oligosymptomatic transmission6-8. There is an urgent need for information on virus replication, immunity and infectivity in specific sites of the body. Here we report a detailed virological analysis of nine cases of COVID-19 that provides proof of active virus replication in tissues of the upper respiratory tract. Pharyngeal virus shedding was very high during the first week of symptoms, with a peak at 7.11 × 108 RNA copies per throat swab on day 4. Infectious virus was readily isolated from samples derived from the throat or lung, but not from stool samples-in spite of high concentrations of virus RNA. Blood and urine samples never yielded virus. Active replication in the throat was confirmed by the presence of viral replicative RNA intermediates in the throat samples. We consistently detected sequence-distinct virus populations in throat and lung samples from one patient, proving independent replication. The shedding of viral RNA from sputum outlasted the end of symptoms. Seroconversion occurred after 7 days in 50% of patients (and by day 14 in all patients), but was not followed by a rapid decline in viral load. COVID-19 can present as a mild illness of the upper respiratory tract. The confirmation of active virus replication in the upper respiratory tract has implications for the containment of COVID-19.

Osteomyelitis caused by Burkholderia pseudomallei: relapse six years after pulmonary infection.

BACKGROUND: Burkholderia pseudomallei is highly endemic in Southeast Asia, whereas in Europe usually only few imported cases of melioidosis occur. CASE REPORT: In 2006, a 52-year-old male patient had been admitted to hospital with pneumonia after returning from a trip to Thailand. A blood culture isolate had been identified as Pseudomonas fluorescens and the patient had been treated with Piperacillin according to the antibiogram. Six years later the patient developed osteomyelitis of the leg and Burkholderia pseudomallei was identified as the causative agent. CONCLUSIONS: Misidentification of the cultural isolate in 2006 had led to inadequate therapy and to an unusually late relapse of melioidosis six years later.

Active immunization with amyloid-beta 1-42 impairs memory performance through TLR2/4-dependent activation of the innate immune system.

Active immunization with amyloid-ß (Aß) peptide 1-42 reverses amyloid plaque deposition in the CNS of patients with Alzheimer's disease and in amyloid precursor protein transgenic mice. However, this treatment may also cause severe, life-threatening meningoencephalitis. Physiological responses to immunization with Aß(1-42) are poorly understood. In this study, we characterized cognitive and immunological consequences of Aß(1-42)/CFA immunization in C57BL/6 mice. In contrast to mice immunized with myelin oligodendrocyte glycoprotein (MOG)(35-55)/CFA or CFA alone, Aß(1-42)/CFA immunization resulted in impaired exploratory activity, habituation learning, and spatial-learning abilities in the open field. As morphological substrate of this neurocognitive phenotype, we identified a disseminated, nonfocal immune cell infiltrate in the CNS of Aß(1-42)/CFA-immunized animals. In contrast to MOG(35-55)/CFA and PBS/CFA controls, the majority of infiltrating cells in Aß(1-42)/CFA-immunized mice were CD11b(+)CD14(+) and CD45(high), indicating their blood-borne monocyte/macrophage origin. Immunization with Aß(1-42)/CFA was significantly more potent than immunization with MOG(35-55)/CFA or CFA alone in activating macrophages in the secondary lymphoid compartment and peripheral tissues. Studies with TLR2/4-deficient mice revealed that the TLR2/4 pathway mediated the Aß(1-42)-dependent proinflammatory cytokine release from cells of the innate immune system. In line with this, TLR2/4 knockout mice were protected from cognitive impairment upon immunization with Aß(1-42)/CFA. Thus, this study identifies adjuvant effects of Aß(1-42), which result in a clinically relevant neurocognitive phenotype highlighting potential risks of Aß immunotherapy.

Alteration of T cell cytokine production in PLPp-139-151-induced EAE in SJL mice by an immunostimulatory CpG oligonucleotide.

Experimental autoimmune encephalomyelitis (EAE) is--in certain aspects--regarded as an animal model of the human CNS autoimmune disease multiple sclerosis (MS). While in EAE CNS-autoantigen-specific immunity is induced in a defined way, the initial processes leading to CNS autoimmunity in humans are so far unknown. Despite essential restrictions, which exist regarding the interpretation of EAE data towards MS, EAE might be a useful model to study certain basic aspects of CNS autoimmunity. Studies in MS have demonstrated that established autoimmune pathology can be critically influenced by environmental factors, in particular viral and bacterial infections. To investigate this interaction, EAE as an instrument to study CNS autoimmunity under defined conditions appears to be a suitable experimental tool. For this reason, we here investigated the influence of the Toll-like-receptor (TLR) ligand CpG oligonucleotide (CpG) on already established CNS autoimmunity in murine proteolipid protein (PLP)-induced EAE in SJL mice. CpG were found to co-stimulate PLPp-specific IFN-γ production in the peripheral immune system and in the CNS. However, CpG induced Interleukin (IL)-17 production in the inflamed CNS both alone and in combination with additional PLPp stimulation. These findings might indicate a mechanism by which systemic infections and the microbial stimuli associated with them may influence already existing CNS autoimmune pathology.

IL-6 controls Th17 immunity in vivo by inhibiting the conversion of conventional T cells into Foxp3+ regulatory T cells.

The conditions leading to the induction of adaptive Foxp3(+) regulatory T cells (T-regs) from peripheral T cells in vivo are incompletely understood. Here, we show that unresponsiveness of T cells to IL-6 by T cell-selective deletion of gp130 or immunization of wild-type mice with antigen in incomplete Freund's adjuvant (IFA), which fails to induce IL-6, promotes the conversion of peripheral CD4(+) T cells into adaptive Foxp3(+) T-regs. Thus, both T cell-conditional gp130 knockout (KO) mice immunized with MOG35-55 in complete Freund's adjuvant (CFA) and wild-type mice immunized with MOG35-55 in IFA develop overwhelming antigen-specific T-reg responses and are protected from experimental autoimmune encephalomyelitis (EAE). Depletion of T-regs restores T helper (Th)17 responses and clinical EAE in MOG/CFA-immunized T cell-conditional gp130 KO mice, but not in MOG/IFA-immunized wild-type mice. We conclude that in the absence of T-regs, IL-6 signaling is dispensable for the induction of Th17 cells, and alternative pathways exist to induce Th17 cells and EAE in the absence of IL-6 signaling. However, IL-6 signaling is dominant in inhibiting the conversion of conventional T cells into Foxp3(+) T-regs in vivo, and in the absence of IL-6 signaling, no other cytokine can substitute in inhibiting T-reg conversion. These data identify IL-6 as an important target to modulate autoimmune responses and chronic inflammation.

Embodiment of sadness and depression--gait patterns associated with dysphoric mood.

OBJECTIVE: To analyze gait patterns associated with sadness and depression. Embodiment theories suggest a reciprocal relationship between bodily expression and the way in which emotions are processed. METHODS: In Study 1, the gait patterns of 14 inpatients suffering from major depression were compared with those of matched never-depressed participants. In Study 2, we employed musical mood induction to induce sad and positive mood in a sample of 23 undergraduates. A Fourier-based description of walking data served as the basis for the computation of linear classifiers and for the analysis of gait parameters. RESULTS: Gait patterns associated with sadness and depression are characterized by reduced walking speed, arm swing, and vertical head movements. Moreover, depressed and sad walkers displayed larger lateral swaying movements of the upper body and a more slumped posture. CONCLUSION: The results of the present study indicate that a specific gait pattern characterizes individuals in dysphoric mood.

The antidepressant venlafaxine ameliorates murine experimental autoimmune encephalomyelitis by suppression of pro-inflammatory cytokines.

Antidepressants are known to impact on the immune system. In this study, we examined the immunomodulatory properties of venlafaxine, a selective serotonin/norepinephrine reuptake inhibitor (SNRI), in murine experimental autoimmune encephalomyelitis (EAE), a T-cell-mediated CNS demyelinating disease model of multiple sclerosis. EAE was induced in SJL/J mice by adoptive transfer of myelin-specific T cells. Mice received different doses of venlafaxine before induction and after onset of disease. Sustained daily oral treatment with 6, 20 and 60 mg/kg significantly ameliorated the clinical symptoms of the disease compared to vehicle during both preventive and therapeutic intervention. Venlafaxine suppressed the generation of pro-inflammatory cytokines IL-12 p40, TNF-alpha and IFN-gamma in encephalitogenic T-cell clones, splenocytes and peritoneal macrophages in vitro. It also diminished mRNA expression of a number of inflammatory genes in the inflamed CNS tissue, among them CD3, CD8, Granzyme B, IL-12 p40, IFN-gamma, TNF-alpha and the chemokines Ccl2 and RANTES, whereas the expression of brain-derived neurotrophic factor was increased. These findings demonstrate the strong immunomodulatory property of the selective SNRI venlafaxine. Further studies are warranted to clarify whether venlafaxine may exert similar effects in humans.

Crimean-Congo haemorrhagic fever virus in Hyalomma impeltatum ticks from North Kordofan, the Sudan.

An evidence for Crimean-Congo hemorrhagic fever virus (CCHFV) was found in Hyalomma impeltatum ticks collected from sheep in North Kordofan in the Sudan. Based on sequencing of the partial segment S, the detected virus belongs to lineage I with closest similarity to CCHFV strains from Senegal. So far, this lineage is unknown in the Sudan.

Implications of antiinflammatory properties of the anticonvulsant drug levetiracetam in astrocytes.

There is accumulating evidence that epileptic activity is accompanied by inflammatory processes. In the present study, we evaluated the effect of levetiracetam (Keppra), an anticonvulsant drug with decisive antiepileptic features, with regard to its putative antiinflammatory potential. We previously established an in vitro cell culture model to mimic inflammatory conditions: Primary astrocytic cultures of newborn rats were cocultured with 30% (M30) microglial cells. Alternatively, cocultures containing 5% microglia (M5) were incubated with the proinflammatory mediator, the cytokine interleukin-1beta (IL-1beta), and lipopolysaccharide (LPS), a potent bacterial activator of the immune system. For the M30 cocultures, we observed reduced expression of connexin 43 (Cx43), the predominant gap junction protein. Impaired functional dye coupling and depolarized membrane resting potential (MRP) were monitored in M30 cocultures as well as in M5 cocultures treated with IL-1beta and LPS. We could show that the Cx43 expression, the coupling property, and the membrane resting potential on which we focused our inflammatory coculture model were normalized to noninflammatory level under treatment with levetiracetam (Keppra). Cumulatively, our results provide evidence for antiinflammatory properties of levetiracetam in seizure treatment.

Two Distinct Clinical Courses of Human Cowpox, Germany, 2015.

Here we present two cases of human infection with cowpox virus with distinct clinical courses. A series of clinical photographs documents lesion progression over time. In the first case-an unvaccinated young veterinary assistant-a pustule was treated locally with cortisone. The lesion turned into a large ulcer accompanied by severe lymphadenitis. Based on her close contact to a sick stray cat, infection with cowpox virus was assumed and confirmed by virus isolation, PCR, and serology. The clinical course took up to eleven months until healing of the wound was complete. Transmission of cowpox virus from the cat was likely because a skin swab was PCR-positive and the cat had a high titer of anti-orthopoxvirus antibodies. In contrast, a rather mild clinical course of cowpox was confirmed in a 49-year-old male farmer vaccinated against smallpox. Only a small eschar developed, and wound closure was complete after 6 weeks.

[Brucellosis, an overview and current aspects]. / Brucellose.

Although brucellosis, a zoonosis mostly associated with sheep, goats and cattle, is not endemic in Germany, it is a relevant imported infectious disease. If patients suffer from fever of unknown origin after a stay in highly endemic countries like the Mediterranean area or the Arab world, it is mandatory to formally exclude brucellosis. Cultural methods are the diagnostic gold standard, but due to special methodical and infrastructural requirements it is essential to inform the laboratory at suspicion of infection. The treatment of brucellosis is challenging and usually based on a long-term combination regime using doxycycline and rifampin for at least 6 weeks.

Hantavirus cardiopulmonary syndrome due to Puumala virus in Germany.

In Germany Puumala virus (PUUV), known to cause mild forms of hemorrhagic fever with renal syndrome (HFRS), is the predominating endemic hantavirus. We herein describe an unusually severe case of a PUUV infection that occurred in summer 2015 in South Eastern Germany in a region known to be endemic for PUUV since over ten years. A 54-year-old female gardener was admitted to hospital with fever, cough and dyspnea. Within 48hours the patient developed a rapid progressive adult respiratory distress syndrome (ARDS) with circulatory failure and required ECMO (extracorporeal membrane oxygenation) treatment. Serological and molecular biological examinations of serum samples confirmed an infection with PUUV. Partial sequences of the S- and M-segment clustered to a strain previously described in South Eastern Germany. Our reported case highlights, that in rare incidents PUUV can cause hantavirus cardiopulmonary syndrome, a syndrome that is usually found after infections with New World hantaviruses, and neurological symptoms.

Complete genome sequence of a chikungunya virus imported from bali to Germany.

Chikungunya virus (CHIKV) strain DH130003 was isolated from a traveler with Chikungunya fever returning from Bali to Germany. Although strains of the east-central/south African lineage bearing the A226V mutation have predominated in most parts of Asia since 2005, CHIKV DH130003 belongs to the Asian lineage.

18F-FDG PET detects inflammatory infiltrates in spinal cord experimental autoimmune encephalomyelitis lesions.

UNLABELLED: Multiple sclerosis (MS) is a heterogeneous disease with respect to lesion pathology, course of disease, and treatment response. Imaging modalities are needed that allow better definition of MS lesions in vivo. The aim of this study was to establish an MRI- and PET/CT-based imaging modality and to evaluate approved and promising PET tracers in experimental autoimmune encephalomyelitis (EAE), the animal model of MS. METHODS: MRI and PET/CT scans were obtained in Dark agouti rats with EAE and healthy control rats. The PET tracers 2-(18)F-fluoro-2-deoxy-d-glucose ((18)F-FDG), 3'-deoxy-3'-(18)F-fluorothymidine ((18)F-FLT), and O-(2-(18)F-fluoro-ethyl)-l-tyrosine ((18)F-FET) were used as surrogate markers of glucose utilization, proliferative activity, and amino acid transport and protein biosynthesis. Immediately after the PET/CT scan, animals were sacrificed for autoradiography, histologic work-up, or RNA expression analysis. RESULTS: EAE lesions were predominantly located in the spinal cord. With MRI, we were able to detect inflammatory lesions in diseased rats, which correlated well with inflammatory infiltrates as determined by histology. Increased (18)F-FDG uptake was observed in spinal cord lesions in all diseased rats. Further investigation by volume-of-interest analysis demonstrated a correlation between the density of histologically proven cellular infiltrates and the (18)F-FDG signal intensity in PET (F(DF=3) = 5.9, P = 0.001) and autoradiography (F(DF=3) = 4.2, P = 0.008). With (18)F-FET and (18)F-FLT, no definite uptake could be observed on PET scans, whereas autoradiography showed slight radiotracer accumulation in some lesions. CONCLUSION: Spinal cord inflammatory lesions in the EAE model can be noninvasively visualized in vivo using MRI and (18)F-FDG PET/CT. Localized (18)F-FDG uptake correlates better with a histologically proven abundance of inflammatory cells as a critical marker of disease activity than MRI. Neither (18)F-FET nor (18)F-FLT seems to be a suitable marker for the in vivo detection of inflammatory lesions.

The combination of interferon-beta and HMG-CoA reductase inhibition in multiple sclerosis: enthusiasm lost too soon?

Recent studies support the notion that statins, widely prescribed cholesterol-lowering agents, may target key elements in the immunological cascade leading to inflammation and tissue damage in the pathogenesis of multiple sclerosis (MS). Compelling experimental and observational clinical studies highlighted the possibility that statins may also exert immunomodulatory synergy with approved MS drugs, resulting in several randomized clinical trials testing statins in combination with interferon-beta (IFN-ß). Some data, however, suggest that this particular combination may not be clinically beneficial, and might actually have a negative effect on the disease course in some patients with MS. In this regard, a small North American trial indicated that atorvastatin administered in combination with IFN-ß may increase disease activity in relapsing-remitting MS. Although other trials did not confirm this finding, the enthusiasm for studies with statins dwindled. This review aims to provide a comprehensive overview of the completed clinical trials and reports of the interim analyses evaluating the combination of IFN-ß and statins in MS. Moreover, we try to address the evident question whether usage of this combination routinely requires caution, since the number of IFN-ß-treated MS patients receiving statins for lowering of cholesterol is expected to grow.

Venlafaxine exhibits an anti-inflammatory effect in an inflammatory co-culture model.

Growing evidence indicates immunoregulatory effects of various antidepressants. Through the interaction of the nervous and immune systems, the norepinephrine-serotonin system was shown to modulate inflammatory CNS diseases. Thus, we examined the norepinephrine-serotonin reuptake inhibitor venlafaxine in an astroglia-microglia co-culture model which allows mimicking of an inflammatory milieu by increasing the cultured microglial fraction. Astrocytic membrane resting potential and intercellular coupling, two markers becoming severely impaired under inflammation, were assessed with the patch-clamp technique. We measured IL-6, IL-10, IFN-gamma and TGF-beta concentrations and analysed phenotypic changes of microglia. We found (i) a reversal of the inflammation-induced depolarization effect on the membrane resting potential, (ii) an augmentation of TGF-beta release with a concomitant reduction in the secretion of pro-inflammatory IL-6 and IFN-gamma, and (iii) a significant change of microglial phenotype from activated to resting morphology. Our data clearly indicate anti-inflammatory properties of venlafaxine which might be a result of monoamine-mediated immunomodulation.