Human Infections by Non-Rodent-Associated Hantaviruses in Africa.

Various hantaviruses have been discovered in unconventional hosts (shrews and bats) in Africa. Up to now, it was unknown whether these viruses pose a threat for human health. In this study, using newly established serological assays, we demonstrated evidence of shrew-borne hantavirus infections in humans from Côte d'Ivoire and Gabon.

Life-Threatening Sochi Virus Infections, Russia.

Sochi virus was recently identified as a new hantavirus genotype carried by the Black Sea field mouse, Apodemus ponticus. We evaluated 62 patients in Russia with Sochi virus infection. Most clinical cases were severe, and the case-fatality rate was as high as 14.5%.

Molekulare Identifikation von Hantaviren in neuen Wirten.

In addition to classical virus isolation in cell culture, the molecular detection of new virus variants by PCR techniques allows broader epidemiological insights into the world of viral pathogens. For the detection of hantaviruses-zoonotic viruses leading to fever and organ failure in humans-we developed a genus-wide nested RT-PCR format, which enables the discovery of new members within this virus genus. The methodological approach allowed the demonstration of first hantaviruses from Africa and revealed new hantavirus reservoir hosts, as shrews, moles, and bats.

The N terminus of Andes virus L protein suppresses mRNA and protein expression in mammalian cells.

Little is known about the structure and function of the 250-kDa L protein of hantaviruses, although it plays a central role in virus genome transcription and replication. When attempting to study Andes virus (ANDV) L protein in mammalian cells, we encountered difficulties. Even in a strong overexpression system, ANDV L protein could not be detected by immunoblotting. Deletion analysis revealed that the 534 N-terminal amino acid residues determine the low-expression phenotype. Inhibition of translation due to RNA secondary structures around the start codon, rapid proteasomal degradation, and reduced half-life time were excluded. However, ANDV L protein expression could be rescued upon mutation of the catalytic PD-E-K motif and further conserved residues of the putative endonuclease at the N terminus of the protein. In addition, wild-type ANDV L rather than expressible L mutants suppressed the level of L mRNA, as well as reporter mRNAs. Wild-type L protein also reduced the synthesis of cellular proteins in the high-molecular-weight range. Using expressible ANDV L mutants as a tool for localization studies, we show that L protein colocalizes with ANDV N and NSs but not Gc protein. A fraction of L protein also colocalized with the cellular processing (P) body component DCP1a. Overall, these data suggest that ANDV L protein possesses a highly active endonuclease at the N terminus suppressing the level of its own as well as heterologous mRNAs upon recombinant expression in mammalian cells.

Experimental Case Studies about Uniplanar SHS Joints with Full-Overlapped Top Connection.

Squared hollow steel profiles are commonly used in the construction of offshore structures or building facades. By welding two or more pipes, typical joints are created that are specific for different areas of applications. These joints are less resistant than straight pipes due to the geometrical heterogeneity and the complex stress behavior of the welding. Standards define these joints, but there are restrictions imposed regarding the material or geometry. This paper focused on full-overlapped joints with squared hollow section profiles and on-top connection, which are disregarded in current standards. The aim was to figure out the influence of the inclination angle on the resistance of the joint. In the analysis, experimental and numerical studies were performed. Four different inclination angles commonly used on construction sites were the focus. It was discovered that there is a total diminishment of 46% in the load bearing capacity between the steepest and the most obtuse angles. The structural behavior is non-linear and is influenced by the value of the angles. The second aspect is related to the influence of the steel profile, which is evaluated by a comparison between a squared profile and two circular profiles. It was discovered that the joint made with squared profiles has a higher bearing capacity than the one made with circular profiles, a statement valid for similar thicknesses of the elements.

Experimental and Numerical Case Studies about Two-Dimensional CHS Joints with a Symmetrical Y-Shape.

Steel hollow section joints are mainly used for offshore structures, hall support or trusses. Current standards define different kinds of steel joints, but there are limitations regarding the geometry or load scenarios. Excluded joints are full-overlapped Y-joints with on-top connection. For these kinds of joints, there is no general design fundamental, so the design engineer has to build his/her own model. The aim of this paper is to figure out the resistance of this special undefined joint type and the influence of the inclination angle between the pipes. An experimental and numerical analysis of this joint was done. Due to this evaluation of the inclination angle, the design engineer could optimize the structure economically. Two different circular profile types were focused on. It was concluded that by increasing the inclination angle, the total applicable force decreased non-linearly. On one hand, the most economic design was achieved by choosing a steep angle. On the other hand, the resistance of the structure, regarding the maximum appliable force, could be increased. For the system in this paper, an increase up to 47% was achieved by choosing a profile with a thickness of 2.8 mm instead of 2.0 mm.

Numerical Case Studies about Two-Dimensional CHS Joints with Symmetrical Full-Overlapped Top-Connection.

Steel joints made out of circular hollow section profiles are used for many fields of applications, such as wide-span or representative halls for airports. By connecting two inclined pipes to a vertical third pipe, a ramified vertical column is created, where the node is the weakest point in the structure due to the geometrical heterogeneity. The current standards and Design Codes have limitations regarding the geometrical properties of hollow sections joints. However, the kind of steel joint presented in this paper is excluded in the current standards. This paper is about numerical FEA case studies of two-dimensional, circular hollow-section joints to figure out the resistance of atypical steel joints. In the first step, a small-scale model is generated to analyze the influence of the inclination. In the second step, the geometries of the different pipes are extended. The influence of the inclination angle and the stability of the joint are analyzed. It was discovered that the inclination angle between the three pipes has a large influence on the stresses and deflections at the node. By increasing the inclination angle, the maximum applied force can be increased. The extended members change the behavior and the stress distribution.

Sin Nombre hantavirus nucleocapsid protein exhibits a metal-dependent DNA-specific endonucleolytic activity.

We demonstrate that the nucleocapsid protein of Sin Nombre hantavirus (SNV-N) has a DNA-specific endonuclease activity. Upon incubation of SNV-N with DNA in the presence of magnesium or manganese, we observed DNA digestion in sequence-unspecific manner. In contrast, RNA was not affected under the same conditions. Moreover, pre-treatment of SNV-N with RNase before DNA cleavage increased the endonucleolytic activity. Structure-based protein fold prediction using known structures from the PDB database revealed that Asp residues in positions 88 and 103 of SNV-N show sequence similarity with the active site of the restriction endonuclease HindIII. Crystal structure of HindIII predicts that residues Asp93 and Asp108 are essential for coordination of the metal ions required for HindIII DNA cleavage. Therefore, we hypothesized that homologous residues in SNV-N, Asp88 and Asp103, may have a similar function. Replacing Asp88 and Asp103 by alanine led to an SNV-N protein almost completely abrogated for endonuclease activity.

Surveillance of Batai virus in bovines from Germany.

To estimate the veterinary importance of Batai virus (BATV), we investigated the presence of BATV-specific antibodies and BATV RNA in 548 bovines from southwest Germany, and we demonstrated that 3 cattle serum samples contained BATV-neutralizing antibodies, resulting in a seroprevalence of 0.55%. Thus, our results confirm local transmission and indicate cattle as potential hosts of BATV in southwest Germany.

Human Dobrava-Belgrade hantavirus infection, Kosovo.

Here we describe an acute Dobrava-Belgrade virus (DOBV) infection that presented as severe hemorrhagic fever with renal syndrome (HFRS) in an active-duty U.S. soldier. The infection was acquired in northern Kosovo in spring 2013. Amplification of DOBV genome segments directly from the patient's serum sample was successfully performed. Phylogenetic analysis demonstrated that the strain belong to DOBV genotype Dobrava and is closely related to strains circulating in Southeast Europe and Slovakia. Thus, our case confirms that DOBV genotype Dobrava is able to cause a severe form of HFRS, especially when compared to the other less pathogenic DOBV genotypes.

A 12-year molecular survey of clinical herpes simplex virus type 2 isolates demonstrates the circulation of clade A and B strains in Germany.

BACKGROUND: Recently two different herpes simplex virus type 2 (HSV-2) clades (A and B) were described on DNA sequence data of the glycoprotein E (gE), G (gG) and I (gI) genes. OBJECTIVE: To type the circulating HSV-2 wild-type strains in Germany by a novel approach and to monitor potential changes in the molecular epidemiology between 1997 and 2008. STUDY DESIGN: A total of 64 clinical HSV-2 isolates were analyzed by a novel approach using the DNA sequences of the complete open reading frames of glycoprotein B (gB) and gG. Recombination analysis of the gB and gG gene sequences was performed to reveal intragenic recombinants. RESULTS: Based on the phylogenetic analysis of the gB coding DNA sequence 8 of 64 (12%) isolates were classified as clade A strains and 56 of 64 (88%) isolates were classified as clade B strains. Analysis of the gG coding DNA sequence classified 4 (6%) isolates as clade A strains and 60 (94%) isolates as clade B strains. In comparison, the 8 isolates classified as clade A strains using the gB sequence data were classified as clade B strains when using the gG coding DNA sequence, suggesting intergenic recombination events. Intragenic recombination events were not detected. CONCLUSION: The first molecular survey of clinical HSV-2 isolates from Germany demonstrated the circulation of clade A and B strains and of intergenic recombinants over a period of 12 years.

A 10-year molecular survey of herpes simplex virus type 1 in Germany demonstrates a stable and high prevalence of genotypes A and B.

BACKGROUND: Recently three different herpes simplex virus type 1 (HSV-1) genotypes (A, B and C) were described on DNA sequence data of the glycoprotein E (gE), G (gG) and I (gI) genes. OBJECTIVE: To type the circulating HSV-1 wild-type strains in Germany and to monitor potential changes in the molecular epidemiology over the past 10 years. STUDY DESIGN: A total of 569 clinical HSV-1 isolates from a 10-year survey in Germany were genotyped by a PCR-based restriction fragment length polymorphism analysis of gG and gI. Recombination analysis of the gE gene sequences was performed to reveal intragenic recombinants. RESULTS: Genotypes A and B strains represented 76% of all strains analyzed and showed a stable distribution within all age groups investigated, independently from the gender. Intergenic gG/gI and intragenic gE recombinants were demonstrated to be less prevalent. Interestingly, for one HIV infected patient a gG/gI genotype switch from A/A to C/A was observed within 3 years. CONCLUSION: The first molecular survey of clinical HSV-1 isolates from Germany demonstrated a stable distribution of two different genotypes and recombinants within age groups and over a period of 10 years. Moreover, homologous recombination seems to be an important feature in the evolution of the HSV-1 genome.

Novel approach to differentiate subclades of varicella-zoster virus genotypes E1 and E2 in Germany.

Varicella-zoster virus (VZV) is the causative agent of chicken pox (varicella) in children and reactivation of VZV in elderly or immunocompromised persons can cause shingles (zoster). A subclade differentiation of the most prevalent VZV genotypes E1 and E2 in Germany was not possible with the current genotyping methods in use, but is highly important to understand the VZV molecular evolution in more detail and especially to follow up the routes of infection. Therefore the objective of this study was to develop a simple PCR-based method for differentiation of E1 and E2 subclades. Viral DNA was isolated from vesicle fluid samples of six selected German zoster patients and used to amplify nine complete open reading frames (ORFs) of the VZV genome by different PCR assays. Phylogenetic analysis was performed by a Bayesian approach. Based on the analysis of a total of nine ORFs, a 7482 bp stretch consisting of ORFs 5, 37 and 62 contained informative sites for identification of novel subclades E1a, E2a and E2b for VZV genotypes E1 and E2. Specific single nucleotide polymorphisms (SNPs) were demonstrated for subclades E2a and E2b within the ORFs 5, 37 and 62, whereas a subclade E1a-specific SNP was found in ORF 56. The classification of E1 and E2 subclades may facilitate a more exact and in-depth monitoring of the molecular evolution of VZV in Germany in the future.