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Huhtamo E.,Haartman Institute | Hasu E.,Haartman Institute | Uzcategui N.Y.,Haartman Institute | Erra E.,University of Helsinki | And 5 more authors.
Journal of Clinical Virology | Year: 2010

Background: The increased traveling to dengue endemic regions and the numerous epidemics have led to a rise in imported dengue. The laboratory diagnosis of acute dengue requires several types of tests and often paired samples are needed for obtaining reliable results. Although several diagnostic methods are available, proper comparative data on their performance are lacking. Objectives: To compare the performance of novel methods including a novel pan-DENV real-time RT-PCR and a commercially available NS1 capture-EIA in regard to IgM detection for optimizing the early diagnosis of DENV in travelers. Study design: A panel of 99 selected early phase serum samples of dengue patients was studied by real-time RT-PCR, NS1 antigen ELISA, IgM-EIA, IgG-IFA and cell culture virus isolation. Results: The novel real-time RT-PCR was shown specific and sensitive for detection of DENV-1-4 RNA and suitable for diagnostic use. The diagnostic rate using combination of RNA and IgM detection was 99% and using NS1 and IgM detection 95.9%. The results of RNA and NS1 antigen detection disagreed in 15.5% of samples that had only RNA or NS1 antigen detected. Conclusions: The diagnostic rates of early samples are higher when either RNA or NS1 antigen detection is combined with IgM detection. Besides the differences in the RNA and NS1 detection assays, the observed discrepancy of results could suggest individual variation or differences in timing of these markers in patient serum. © 2009 Elsevier B.V. All rights reserved. Source

Molsa M.,Centres for Military Medicine and for Biological Threat Preparedness | Hemmila H.,Centres for Military Medicine and for Biological Threat Preparedness | Katz A.,Centres for Military Medicine and for Biological Threat Preparedness | Niemimaa J.,Natural Resources Institute Finland | And 7 more authors.
Journal of Microbiological Methods | Year: 2015

In the event of suspected releases or natural outbreaks of contagious pathogens, rapid identification of the infectious agent is essential for appropriate medical intervention and disease containment. The purpose of this study was to compare the performance of a novel portable real-time PCR thermocycler, PikoReal™, to the standard real-time PCR thermocycler, Applied Biosystems® 7300 (ABI 7300), for the detection of three high-risk biothreat bacterial pathogens: Francisella tularensis, Bacillus anthracis and Yersinia pestis. In addition, a novel confirmatory real-time PCR assay for the detection of F. tularensis is presented and validated. The results show that sensitivity of the assays, based on a dilution series, for the three infectious agents ranged from 1 to 100. fg of target DNA with both instruments. No cross-reactivity was revealed in specificity testing. Duration of the assays with the PikoReal and ABI 7300 systems were 50 and 100. min, respectively. In field testing for F. tularensis, results were obtained with the PikoReal system in 95. min, as the pre-PCR preparation, including DNA extraction, required an additional 45. min. We conclude that the PikoReal system enables highly sensitive and rapid on-site detection of biothreat agents under field conditions, and may be a more efficient alternative to conventional diagnostic methods. © 2015 Elsevier B.V. Source

Molsa M.,Centres for Military Medicine and for Biological Threat Preparedness | Hemmila H.,Centres for Military Medicine and for Biological Threat Preparedness | Ronkko E.,Finnish National Institute for Health and Welfare | Virkki M.,Paijat Hame Social and Health Care Group | And 3 more authors.
Journal of Medical Virology | Year: 2016

Although adenoviruses were identified as important respiratory pathogens many years ago, little information is available concerning the prevalence of different adenovirus serotypes, which are circulating and causing epidemics in Finnish military training centers. Over a period of five years from 2008 to 2012, 3577 respiratory specimens were collected from military conscripts presenting with symptoms compatible with acute respiratory tract infection. Upon initial testing for certain respiratory viruses by real-time PCR, 837 of these specimens were identified as adenovirus-positive. For 672 of these specimens, the serotype of the adenovirus responsible was successfully determined by DNA sequencing. Serotypes 1, 2, 3, and 4 were detected in 1, 3, 181, and 487 samples, respectively. Adenovirus epidemics were observed during each year of this study. Based on these findings, adenovirus vaccination should be considered for military conscripts in the Finnish Defence Forces. © 2015 Wiley Periodicals, Inc. Source

Molsa M.,Centres for Military Medicine and for Biological Threat Preparedness | Kalin-Manttari L.,Finnish National Institute for Health and Welfare | Tonteri E.,Centres for Military Medicine and for Biological Threat Preparedness | Hemmila H.,Centres for Military Medicine and for Biological Threat Preparedness | Nikkari S.,Centres for Military Medicine and for Biological Threat Preparedness
Journal of Microbiological Methods | Year: 2016

Bacillus spp. include human pathogens such as Bacillus anthracis, the causative agent of anthrax and a biothreat agent. Bacillus spp. form spores that are physically highly resistant and may remain active over sample handling. We tested four commercial DNA extraction kits (QIAamp DNA Mini Kit, RTP Pathogen Kit, ZR Fungal/Bacterial DNA MiniPrep, and genesig Easy DNA/RNA Extraction kit) for sample inactivation and DNA recovery from two powders (icing sugar and potato flour) spiked with Bacillus thuringiensis spores. The DNA was analysed using a B. thuringiensis-specific real-time PCR assay. The detection limit was 3 × 101 CFU of spiked B. thuringiensis spores with the QIAamp DNA Mini, RTP Pathogen, and genesig Easy DNA/RNA Extraction kits, and 3 × 103 CFU with the ZR Fungal/Bacterial DNA MiniPrep kit. The results showed that manual extraction kits are effective and safe for fast and easy DNA extraction from powder samples even in field conditions. Adding a DNA filtration step to the extraction protocol ensures the removal of Bacillus spp. spores from DNA samples without affecting sensitivity. © 2016 Elsevier B.V. Source

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