The lung microbiome in patients with pneumocystosis.

BACKROUND: Pneumocystis jirovecii pneumonia (PCP) is an opportunistic fungal infection that is associated with a high morbidity and mortality in immunocompromised individuals. In this study, we analysed the microbiome of the lower respiratory tract from critically ill intensive care unit patients with and without pneumocystosis. METHODS: Broncho-alveolar fluids from 65 intubated and mechanically ventilated intensive care unit patients (34 PCP+ and 31 PCP- patients) were collected. Sequence analysis of bacterial 16S rRNA gene V3/V4 regions was performed to study the composition of the respiratory microbiome using the Illumina MiSeq platform. RESULTS: Differences in the microbial composition detected between PCP+ and PCP- patients were not statistically significant on class, order, family and genus level. In addition, alpha and beta diversity metrics did not reveal significant differences between PCP+ and PCP- patients. The composition of the lung microbiota was highly variable between PCP+ patients and comparable in its variety with the microbiota composition of the heterogeneous collective of PCP- patients. CONCLUSIONS: The lower respiratory tract microbiome in patients with pneumocystosis does not appear to be determined by a specific microbial composition or to be dominated by a single bacterial species.

Low sensitivity of SARS-CoV-2 rapid antigen self-tests under laboratory conditions.

SARS-CoV-2-antigen-testing has been proposed as a 'game-changing' tool to interrupt infection chains. Thereby European strategies focused on two pillars, namely rapid antigen tests conducted by health care experts and/or trained personal and so-called self-tests. Here, evidence is provided that these assays have a weak performance even under laboratory conditions.

Two cases of severe obstructive pneumonia associated with an HKU1-like coronavirus.

BACKGROUND: During the last few years a number of previously undescribed viruses, including human metapneumovirus, coronaviruses SARS, NL63 and HKU1, and bocavirus, were identified in nasopharyngeal samples from patients with signs of respiratory infections. These viruses may cause mild to life-threatening infections. OBJECTIVES: Nasopharyngeal samples from hospitalized pediatric patients with respiratory disease were analysed for the presence of coronaviruses and other well known and newly identified respiratory viruses. RESULTS: Two clinical cases of a severe obstructive pneumonia, which were associated with the presence of RNA of a novel variant (subtype) of HKU1 coronavirus in the nasopharyngeal aspirates, were identified. DISCUSSION: The detection of a HKU1-like coronavirus in pediatric patients in the current study complement the most recent independent finding of similar or closely related coronaviruses in patients with respiratory diseases in France (Vabret et al. 2006) and Norway (Jonassen et al., see accompanying manuscript). These observations indicate a wide dissemination of HKU1-like coronaviruses in Europe.

[Human metapneumovirus as a common cause of respiratory tract disease].

Human metapneumovirus (HMPV), the newly identified paramyxovirus, causes respiratory infections in children, immunosuppressed patients, and the elderly in different countries of the world. The epidemiology and clinical manifestations of HMPV infection are similar to those in human respiratory syncytial virus infection. The diagnosis of HMPV infection is based on the polymerase chain reaction detection of viral RNA or the recording of rising serum antibody titers. There are at least two genotypes and several subtypes of HMPV in the human population. The use of cell cultures and laboratory animals have provided new evidence for the pathogenesis of HMPV infection, the specific features of antiviral immunity and enabled recombinant HMPV vaccine candidates to be designed.

Fluctuation of the cytokine expression in the liver during the chronic woodchuck hepatitis virus (WHV) infection is not related to viral load.

The woodchuck together with the woodchuck hepatitis virus (WHV) is an excellent model to study the pathogenesis of hepadnaviral infections. Chronic WHV infection causes severe liver disease and hepatocellular carcinoma in woodchucks. The mechanism of viral clearance is not fully understood, interferons seem to play a major role in down-regulating viral replication prior to elimination of infected hepatocytes. We investigated on the pattern of cytokine and T-cell-marker expression in livers of woodchucks chronically infected with WHV. RNase-protection-assay (RPA) was used to determine mRNA of woodchuck specific genes (TNF-alpha, IFN-gamma, IL-15, CD3, CD4, CD8). Serial liver biopsies were performed daily or weekly in eight chronic WHV-carrier woodchucks. Cytokine/T-cell-marker expression differed significantly between the time points up to +/-50% within each woodchuck. The different expression patterns of cytokines or T-cell-markers did not correlate to the (weak) fluctuations in the viremia but may explain the observed fluctuations in the WHV/HBV-load in chronically infected individuals. Furthermore, we observed associations between cytokine and T-cell-marker expression. The marginal fluctuations in viremia during the chronic infection may indicate, that, once the chronic hepadnaviral infection is established, cytokines/interferons expressed endogenously (i.e. not vector-borne or injected) play only a minor role.

Large nontransplanted hepatocellular carcinoma in woodchucks: treatment with adenovirus-mediated delivery of interleukin 12/B7.1 genes.

BACKGROUND: Cytokine-based gene therapy strategies efficiently stimulate immune responses against many established transplanted tumors, leading to rejection of the tumor. In this study, we investigated the therapeutic potential of cancer immunotherapy in a clinically more relevant model, woodchucks with primary hepatocellular carcinomas induced by woodchuck hepatitis virus. METHODS: Large (2-5 cm), established intrahepatic tumors were given an injection once with 1 x 10(9) plaque-forming units of AdIL-12/B7.1, an adenovirus vector carrying genes for murine interleukin 12 and B7.1, or of AdEGFP, the control virus, and regression of the tumors was then monitored. Five animals were used in total. RESULTS: In four tumor-bearing animals, the antitumor response was assessed by autopsy and histologic analysis within 1-2 weeks after treatment. In all animals treated with AdIL-12/B7.1 therapy versus AdEGFP therapy, we observed substantial tumor regression (P =.006; two-sided unpaired Student's t test) accompanied by a massive infiltration of T lymphocytes. These tumors also contained increased levels of CD4(+) and CD8(+) T cells and interferon gamma (IFN gamma). In continuously growing tumor nodules given an injection of the control virus or in nontumoral liver, no such effects (i.e., tumor regression and increased levels of CD4(+) and CD8(+) T cells and IFN gamma) were detected. In the fifth animal, monitored for long-term antitumor efficacy by magnetic resonance imaging (MRI) after intratumoral vector administration by MRI guidance, the tumor was almost completely eliminated (> or = 95%) 7 weeks after treatment. CONCLUSION: Adenovirus vector-based immunotherapy appears to be an effective treatment of large nontransplanted (orthotopic) tumors that acquire malignant characteristics in a stepwise process, reflecting the real-world scenario of hepatocellular carcinoma in humans.

Serum antibody response to respiratory syncytial virus F and N proteins in two populations at high risk of infection: children and elderly.

Human respiratory syncytial virus (hRSV) is the main viral cause of severe respiratory infections in children and a common cause of morbidity in the elderly. The nucleocapsid (N) and fusion (F) proteins of hRSV were expressed in insect cells and used as antigens in two independent enzyme-linked immunosorbent assays (ELISAs) to measure the serum antibody response in two populations at high risk of hRSV infection, children and the elderly. Fifty-seven serum specimens from children aged from 1 to 10 years old and 91 sera from adults over 60 years old were tested. The ELISA results were compared with those obtained by an immunofluorescence assay (IFA) based on hRSV-infected cells, which was considered as the reference technique. Sensitivity and specificity were 94% and 85% for the N-ELISA and 86% and 81% for the F-ELISA, respectively. When the immune responses of the two groups of individuals were compared, it appeared that almost 100% of the elderly had antibodies against the N or F protein whereas only 50% of the sera from children had antibodies against either of the two viral proteins. In conclusion, the F and N ELISAs can be used successfully for detecting a specific antibody response to hRSV.

Viral infections in paediatric patients receiving conventional cancer chemotherapy.

In severely immunocompromised patients, the diagnosis of viral infections relies on PCR/RT-PCR based methods. The availability of these modern diagnostic tools facilitates timely diagnosis and contributes to our increasing knowledge of the epidemiology and clinical spectrum of common and emerging viral pathogens in this highly susceptible population. Viral infections may result in life threatening disease in paediatric cancer patients after stem cell transplantation and also during conventional chemotherapy. Often, clinical symptoms are a consequence of endogenous reactivation of latent viral infection. Many of these viruses are easily transmitted between patients, relatives and health care workers. As prolonged symptomatic and asymptomatic viral shedding is a common feature in paediatric cancer patients, it is necessary to implement strategies for the prevention and control of these communicable pathogens in the hospital setting and in the outpatient clinic. Although no randomised controlled studies on paediatric cancer patients are available, physicians should be aware of potential treatment options since early treatment may prevent a complicated or fatal outcome and shorten the period of contagiosity.

[Human Metapneumovirus in hospitalized children - a review]. / Das humane Metapneumovirus als Erreger von Atemwegsinfektionen bei hospitalisierten Kindern - eine Ubersicht.

The human Metapneumovirus (HMPV) has been discovered by von den Hoogen et al. in 2001 and seems to play an important role as etiologic agent in childhood respiratory tract infections in particular involving infants after the 6th month of life and toddlers. Duly considering the hitherto published studies and retrospective analysis of two HMPV seasons (2002-2004) at our institution this review focuses on children, who had to be hospitalized due to HMPV infection. The analysis confirmed, that among those patients there is a high proportion of children with pre-existing risk factors for a complicated clinical course, a high proportion of children with bronchiolitis or pneumonia and a relevant proportion of children with HMPV related apnoeas, most prevalent in the prematurely born. Although the first HMPV infection takes place somewhat later in infancy, the data do not show that HMPV infection is in general milder than RSV infection in hospitalized children. Clinical symptoms and radiological signs do not permit tentative conclusions on the causative agent. This underlines the necessity of specific diagnostic efforts (in case of HMPV with PCR). HMPV may cause lobar or segmental pneumonias difficult to distinguish from bacterial lower respiratory tract infection. Children admitted to the hospital with an acute exacerbation of asthma bronchiale or cystic fibrosis should not only be tested for RSV but also for HMPV. Prospective studies investigating specific therapeutic interventions or describing the impact and prevention of nosocomial HMPV in fection are awaited for. There has been one report of a meningoencephalitis possibly related to HMPV. Thus, liquor samples in such cases should be tested for HMPV too.

[The human bocavirus: pathogen in airway infections?]. / Das humane Bocavirus: Erreger von Atemwegsinfektionen?

The human Bocavirus (HBoV), the second member of the parvovirus family, which displays pathogenicity in humans, has been described in 2005 by Allander et al.. It seems to be distributed worldwide and has been isolated mainly in infants and children with respiratory tract infection. This review covers all studies published on HBoV to February 2007 and discusses this emerging viral pathogen from the perspective of inpatient medical treatment centers.

Identification of a glycosylation site in the woodchuck hepatitis virus preS2 protein and its role in protein trafficking.

The middle surface antigen (M-sAg) of hepadnaviruses is one of three envelope proteins that share a common C-terminal S domain. M-sAg contains the preS2 domain in addition to the S region. The preS2 region of woodchuck hepatitis virus (WHV) contains a potential glycosylation site Asn-Gln-Thr at amino acid (aa) positions 3-5. In this study, we mutated this site by site-directed mutagenesis and confirmed that glycosylation occurs here. In in vitro translation assays, the mutation Thr to Asn at aa 5 significantly impaired glycosylation of M-sAg. The mutated M-sAg formed abnormal clustered structures in transfected cells as determined by immunofluorescent staining. Confocal microscopic analysis showed that an enrichment of this glycosylation-deficient protein in the Golgi apparatus occurred, which is not typical for the wild-type protein. These results are consistent with earlier findings that incorrect glycosylation of viral proteins may interfere with virus assembly.

Determinants of pathogenicity of echovirus 9 in men: significance of a functional RGD-motif.

In this study, we investigated nine independent echovirus 9 isolates obtained from sick children in 1995. It is discovered that these isolates differ in respect to their pathogenicity for newborn mice indicating that the degree of human pathogenicity of an echovirus 9 variant does not necessarily correlate with mouse pathogenicity. Nevertheless, all virus variants are found to code for an RGD-motif within their VP1 protein. Hence, the RGD-motif and its highly conserved flanking regions are the conditio sine qua non, but, as expected, not sufficient for the mouse-pathogenic character.