Immunization with DNA Plasmids Coding for Crimean-Congo Hemorrhagic Fever Virus Capsid and Envelope Proteins and/or Virus-Like Particles Induces Protection and Survival in Challenged Mice.

Crimean-Congo hemorrhagic fever virus (CCHFV) is a bunyavirus causing severe hemorrhagic fever disease in humans, with high mortality rates. The requirement of a high-containment laboratory and the lack of an animal model hampered the study of the immune response and protection of vaccine candidates. Using the recently developed interferon alpha receptor knockout (IFNAR) mouse model, which replicates human disease, we investigated the immunogenicity and protection of two novel CCHFV vaccine candidates: a DNA vaccine encoding a ubiquitin-linked version of CCHFV Gc, Gn, and N and one using transcriptionally competent virus-like particles (tc-VLPs).

Rapid Bedside Inactivation of Ebola Virus for Safe Nucleic Acid Tests.

Rapid bedside inactivation of Ebola virus would be a solution for the safety of medical and technical staff, risk containment, sample transport, and high-throughput or rapid diagnostic testing during an outbreak. We show that the commercially available Magna Pure lysis/binding buffer used for nucleic acid extraction inactivates Ebola virus. A rapid bedside inactivation method for nucleic acid tests is obtained by simply adding Magna Pure lysis/binding buffer directly into vacuum blood collection EDTA tubes using a thin needle and syringe prior to sampling.

One-step real-time RT-PCR assays for serotyping dengue virus in clinical samples.

Background: Dengue is one of the leading causes of morbidity in tropical and subtropical regions and infection with any of the four dengue virus serotypes (DENV1-4) result in a wide range of clinical manifestations. Given the geographic expansion of DENV1-4, assays for serotyping are needed to be able to perform surveillance and epidemiological studies. In this study, we describe the design and validation of one-step real-time serotype-specific DENV RT-PCR assays.

Molecular and serological findings in suspected patients with Crimean-Congo hemorrhagic fever virus in Iran.

Crimean-Congo hemorrhagic fever (CCHF) is an arthropod-borne disease of humans associated with a severe clinical picture, including hemorrhagic syndrome and a high mortality rate. CCHF virus is widely distributed throughout large areas of the world. To characterize the serological status in CCHF patients, paired clinical samples were collected from suspected CCHF patients and analyzed by microbiological and other laboratory analyses with the aim of: determining the presence of neutralizing antibodies against CCHF virus; investigating the cross-reactivity of these neutralizing antibodies against virus isolated from the same outbreak and against other available laboratory strain; and studying the relationship between the isolated virus with other virus by whole genome sequencing.

Multiplex nucleic acid suspension bead arrays for detection and subtyping of filoviruses.

Here we describe multiplex suspension bead array systems that allow fast and reliable detection of reverse transcriptase (RT) PCR amplified filovirus genomes and also enable subtyping of Ebola virus species and Marburg virus strains. These systems have an analytical sensitivity equivalent to that of RT-PCR.

Universal single-probe RT-PCR assay for diagnosis of dengue virus infections.

Background: Dengue is a mosquito-borne viral disease that has become more prevalent in the last few decades. Most patients are viremic when they present with symptoms, and early diagnosis of dengue is important in preventing severe clinical complications associated with this disease and also represents a key factor in differential diagnosis. Here, we designed and validated a hydrolysis-probe-based one-step real-time RT-PCR assay that targets the genomes of dengue virus serotypes 1-4.

Colorimetric nucleic acid testing assay for RNA virus detection based on circle-to-circle amplification of padlock probes.

We developed a molecular diagnostic method for detection of RNA virus based on padlock probes and colorimetric readout. The feasibility of our approach was demonstrated by using detection of Crimean-Congo hemorrhagic fever (CCHF) virus as a model. Compared with conventional PCR-based methods, our approach does not require advanced equipment, involves easier assay design, and has a sensitivity of 10(3) viral copies/ml.

Quantitative analysis of particles, genomes and infectious particles in supernatants of haemorrhagic fever virus cell cultures.

Information on the replication of viral haemorrhagic fever viruses is not readily available and has never been analysed in a comparative approach. Here, we compared the cell culture growth characteristics of haemorrhagic fever viruses (HFV), of the Arenaviridae, Filoviridae, Bunyaviridae, and Flavivridae virus families by performing quantitative analysis of cell culture supernatants by (i) electron microscopy for the quantification of virus particles, (ii) quantitative real time PCR for the quantification of genomes, and (iii) determination of focus forming units by coating fluorescent antibodies to infected cell monolayers for the quantification of virus infectivity.The comparative analysis revealed that filovirus and RVFV replication results in a surplus of genomes but varying degrees of packaging efficiency and infectious particles.

Comparison of antiviral activity of recombinant and natural interferons against crimean-congo hemorrhagic Fever virus.

As a first line of defence against a virus infection, mammalian cells elicit an innate immune response, characterized by secretion of type I interferons (IFN) and up-regulation of interferon stimulated genes (ISGs). We have previously included Crimean Congo Hemorrhagic Fever Virus (CCHFV) in the list of type I IFN-sensitive viruses. In this in vitro study, we have compared the antiviral activity of two recombinant IFN-alpha preparations (Roferon A and Intron A) with a natural IFN-alpha produced in human leukocytes (Multiferon).

Crimean-Congo hemorrhagic fever virus infection is lethal for adult type I interferon receptor-knockout mice.

Crimean-Congo hemorrhagic fever virus (CCHFV) poses a great threat to public health due to its high mortality, transmission and geographical distribution. To date, there is no vaccine or specific treatment available and the knowledge regarding its pathogenesis is highly limited. Using a small-animal model system, this study showed that adult mice missing the type I interferon (IFN) receptor (IFNAR(-/-)) were susceptible to CCHFV and developed an acute disease with fatal outcome.

Migrating birds and tickborne encephalitis virus.

During spring and autumn 2001, we screened 13,260 migrating birds at Ottenby Bird Observatory, Sweden, and found 3.4% were infested with ticks. Four birds, each a different passerine species, carried tickborne encephalitis virus (TBEV)-infected ticks (Ixodes ricinus).

Fatal subarachnoidal haemorrhage in a Norwegian traveller with dengue virus infection.

We present a Norwegian female in her thirties who acquired dengue fever caused by dengue virus serotype 2 while travelling to Mexico. When hospitalised 3 days after symptom onset, the patient had severe headache, fever, rash and a positive tourniquet test, but did not fulfil the criteria of dengue haemorrhagic fever (DHF). Five days later she developed a fatal subarachnoidal haemorrhage.

DNA microarray technique for detection and identification of seven flaviviruses pathogenic for man.

A flavivirus microarray was developed for detection and identification of yellow fever (YF), West Nile, Japanese encephalitis (JE), and the dengue 1-4 viruses, which are causing severe human disease all over the world. The microarray was based on 500-nucleotide probe fragments from five different parts of the seven viral genomes. A low-stringent amplification method targeting the corresponding regions of the viral genomic RNA was developed and combined with hybridization to the microarray for detection and identification.

Optimized diagnosis of acute dengue fever in Swedish travelers by a combination of reverse transcription-PCR and immunoglobulin M detection.

The dengue viruses (genus Flavivirus, family Flaviviridae) are mosquito borne and cause 100 million cases of dengue fever each year in most tropical and subtropical areas of the world. Increased global travel has been accompanied by an increased import not only of dengue but also of severe fevers of unknown origin to Sweden. Fifty-seven Swedish travelers to dengue epidemic areas, with clinical and serologically diagnosed dengue fever, were included in this study.

Hantaviruses in Estonia.

Human serum samples collected from healthy individuals in 14 counties were screened by ELISA in order to investigate the presence of hantavirus infections in Estonia. Out of 1,234 serum samples, 124 were found positive for hantavirus-specific IgG and were subsequently serotyped by a focus reduction neutralization test. A total of 112 samples neutralized at least one of the examined hantaviruses-Puumala (PUUV), Saaremaa (SAAV), Dobrava (DOBV), Hantaan, and Seoul viruses-and thereby, the focus reduction neutralization test confirmed the overall hantavirus seroprevalence rate in Estonia to be 9.