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Mentula S.,Finnish National Institute for Health and Welfare | Laakso S.,Mobidiag Ltd. | Lyytikainen O.,Finnish National Institute for Health and Welfare | Kirveskari J.,Mobidiag Ltd.
Expert Review of Molecular Diagnostics | Year: 2015

Objective: Hypervirulent Clostridium difficile clade has been shown to include several lineages of ribotype 027 and also other ribotypes. We present data on additional non-027 strains, identified as presumptive 027 by two commercial molecular C. difficile assays. Methods: The tested clinical isolates were selected from the national reference laboratory collection on the basis of toxin gene profile similarities with ribotype 027 and tested with Xpert®C. difficile/Epi and Amplidiag™ C. difficile+027 assay. Result: Xpert misclassified five ribotypes (016, 019, 080, 176 and variant of type 046) as presumptive 027 and Amplidiag two ribotypes (016, 176). The misclassified strains were rare, covering 1.6% of reference laboratory strain collection. Conclusion: Our findings confirm the concept that there are closely related outliers to hypervirulent 027 clones that can be misclassified as 027, and that these comprise numerous ribotypes, including previously reported four ribotypes (198, 176, 244, 019), and additional three (016, v046, 080) identified in the present study. © 2015 © Informa UK, Ltd.


Allen E.K.,University of Virginia | Pitkaranta A.,University of Helsinki | Maki M.,Mobidiag Ltd | Hendley J.O.,University of Virginia | And 3 more authors.
International Forum of Allergy and Rhinology | Year: 2013

Background: Patients  with  viral respiratory infections/viral rhinitis/common colds are often treated with antibiotic; however, there is little information on whether or how bacterial microbiota in the nose and nasopharynx might influence the course of viral illnesses. Methods: To initiate investigation of possible interaction between viral respiratory illness and microbiota of the nose/nasopharynx, we used microarray technology to examine 100 nasal lavage fluid (NLF) samples for bacterial species and recorded the bacterial titer of culturable bacteria. Rhinovirus illnesses were induced by self-inoculation using the "finger to nose or eye natural transmission route" in 10 otherwise healthy young adults. NLF samples were collected during wellness and at specific time points following experimental rhinovirus inoculation. Results: The rhinovirus infection rate was 70%. There were no consistent changes in the prevalence of different bacterial species determined by microarray and bacterial titer by culture methods during rhinovirus infection. The bacterial profile in NLF samples showed high variability between volunteers but low variability in multiple NLFs obtained before and following infection from the same volunteer. Streptococcus epidermidis/coagulase-negative staphylococcus (CNS) were identified in all 10 subjects. One or more bacterial sinus/otitis pathogens were identified by microarray in 6 of the 10 volunteers. The microarray identified a few bacteria not included in traditional bacterial cultures. Conclusion: Our pilot study showed that each of the 10 volunteers had a unique bacterial profile in the nose by microarray analysis and that bacterial load did not change during experimental rhinovirus colds. Larger scale studies are warranted. © 2013 The Authors. International Forum of Allergy & Rhinology published by Wiley Periodicals, Inc., on behalf of ARS-AAOA, LLC.


Tapiovaara L.,University of Helsinki | Lehtoranta L.,University of Helsinki | Swanljung E.,University of Helsinki | Makivuokko H.,Valio Ltd | And 4 more authors.
International Journal of Pediatric Otorhinolaryngology | Year: 2014

Objective: Probiotics may have potency in reducing upper respiratory infections, in particular in children. We studied findings from middle ear effusion (MEE) samples after randomized, placebo-controlled 3-week oral administration of probiotic Lactobacillus rhamnosus GG (L. GG). Methods: 40 children referred to tympanostomy were randomized to receive either L. GG or placebo (1:1) for 3 weeks before surgery. MEE samples were collected from 13 children (in total, 25 samples, 19 from the L. GG group and 6 from the placebo group) and analyzed for L. GG and pathogenic bacterial and viral findings. Results: L. GG was present in 5 of the 25 MEE samples (4 from the L. GG group). Haemophilus infuenzae was the most prominent pathogen in 12 samples (10 from the L. GG group). Rhinovirus was present in 12 samples (10 from the L. GG group) and enterovirus in 1 sample (L. GG group). Conclusions: L. GG was present in the middle ear of children suffering from otitis media with effusion, but did not reduce the presence of pathogenic bacteria or viruses. © 2014 Elsevier Ireland Ltd.


Metso L.,University of Helsinki | Maki M.,Mobidiag Ltd. | Tissari P.,University of Helsinki | Remes V.,University of Helsinki | And 6 more authors.
Acta Orthopaedica | Year: 2014

Background and purpose-Polymerase chain reaction (PCR) methods enable detection and species identification of many pathogens. We assessed the efficacy of a new PCR and microarray-based platform for detection of bacteria in prosthetic joint infections (PJIs). Methods-This prospective study involved 61 suspected PJIs in hip and knee prostheses and 20 negative controls. 142 samples were analyzed by Prove-it Bone and Joint assay. The laboratory staff conducting the Prove-it analysis were not aware of the results of microbiological culture and clinical findings. The results of the analysis were compared with diagnosis of PJIs defined according to the Musculoskeletal Infection Society (MSIS) criteria and with the results of microbiological culture. Results-38 of 61 suspected PJIs met the definition of PJI according to the MSIS criteria. Of the 38 patients, the PCR detected bacteria in 31 whereas bacterial culture was positive in 28 patients. 15 of the PJI patients were undergoing antimicrobial treatment as the samples for analysis were obtained. When antimicrobial treatment had lasted 4 days or more, PCR detected bacteria in 6 of the 9 patients, but positive cultures were noted in only 2 of the 9 patients. All PCR results for the controls were negative. Of the 61 suspected PJIs, there were false-positive PCR results in 6 cases. Interpretation-The Prove-it assay was helpful in PJI diagnostics during ongoing antimicrobial treatment. Without preceding treatment with antimicrobials, PCR and microarray-based assay did not appear to give any additional information over culture.


Kanerva M.,University of Helsinki | Nissinen J.,University of Helsinki | Moilanen K.,Mobidiag Ltd. | Maki M.,Mobidiag Ltd. | And 2 more authors.
Otology and Neurotology | Year: 2013

Objective: Microbiologic causes of facial palsy in children were investigated. Study Design: Prospective clinical study. Setting: Tertiary referral center. Patients: Forty-six children aged 0 to 16 years with peripheral facial palsy. Interventions: Paired serum samples and cerebrospinal fluid were tested to find indications of microbes associated with facial palsy. The microbes tested were herpes simplex virus 1 and 2, varicella-zoster virus, human herpesvirus-6, Mycoplasma pneumoniae, Borrelia burgdorferi, influenza A and B virus, picorna, cytomegalovirus, parainfluenza virus, respiratory syncytial virus, coxsackie B5 virus, adenovirus, and enterovirus, Chlamydia psittaci, and Toxoplasma gondii. Besides the routine tests in clinical practice, serum and cerebrospinal fluid samples were tested with a highly sensitive microarray assay for DNA of herpes simplex virus 1 and 2; human herpes virus 6A, 6B, and 7; Epstein-Barr virus, cytomegalovirus, and varicella zoster virus. Results: Incidence for facial palsy was 8.6/100,000/children/year. Cause was highly plausible in 67% and probable in an additional 11% of cases. Borrelia burgdorferi caused facial palsy in 14 patients (30%), varicella zoster virus in 5 (11%) (one with concomitant adenovirus), influenza A in 3 (6%), herpes simplex virus 1 in 2 (4%) (one with concomitant enterovirus), otitis media in 2 (4%), and human herpesvirus 6 in 2 (4%). Mycoplasma pneumoniae, neurofibromatosis, and neonatal age facial palsy affected 1 child (2%) each. Conclusion: Microbiologic etiology association of pediatric facial palsy could frequently be confirmed. Borreliosis was the single most common cause; hence, cerebrospinal fluid sampling is recommended for all pediatric cases in endemic areas. Varicella zoster virus accounted for 11% of the cases, being the second most common factor. © 2013, Otology & Neurotology, Inc.


Viitala S.M.,University of Eastern Finland | Viitala S.M.,Biocenter Kuopio | Jaaskelainen A.J.,University of Helsinki | Kelo E.,Reagena Ltd | And 7 more authors.
Diagnostic Microbiology and Infectious Disease | Year: 2013

Microarrays are widely used in high-throughput DNA and RNA hybridization tests and recently adopted to protein and small molecule interaction studies in basic research and diagnostics. Parallel detection of serum antibodies and antigens has several potential applications in epidemiologic research, vaccine development, and in the diagnosis of allergies, autoimmunity, and infectious diseases. This study demonstrates an immobilization method for immunoassay-based microarray in conventional 96-well polystyrene plates for a serologic diagnostic method combined with quantitative C-reactive protein (CRP) assay. A synthetic peptide (HIV-1), a recombinant protein (Puumala hantavirus nucleocapsid), and purified virus preparations (Sindbis and adenoviruses) were used as antigens for virus-specific antibody detection and monoclonal anti-CRP antibody for antigen detection. The microarray was based on conventional enzyme immunoassays and densitometry from photographed results. Peptide and recombinant antigens functioned well, while whole virus antigens gave discrepant results in 1 out of 23 samples from the reference method, tested with human sera with various antibody responses. The CRP results were in concordance in the concentration range 0.5-150 mg/L with 2 commercially available CRP assays: ReaScan rapid test (R2 = 0.9975) and Cobas 6000 analyzer (R2 =0.9595). The results indicate that microtiter plates provide a promising platform for further development of microarrays for parallel antibody and antigen detection. © 2013 Elsevier Inc.


Laakso S.,Mobidiag Ltd | Kirveskari J.,University of Helsinki | Tissari P.,University of Helsinki | Maki M.,Mobidiag Ltd
PLoS ONE | Year: 2011

Background: High incidence of septic patients increases the pressure of faster and more reliable bacterial identification methods to adapt patient management towards focused and effective treatment options. The aim of this study was to assess two automated DNA extraction solutions with the PCR and microarray-based assay to enable rapid and reliable detection and speciation of causative agents in the diagnosis of sepsis. Methodology/Principal Findings: We evaluated two automated DNA instruments NucliSENS® easyMAG® and NorDiag Arrow for the preparation of blood culture samples. A set of 91 samples flagged as positive during incubation was analyzed prospectively with the high-throughput generation of Prove-it™ Sepsis assay designed to identify over 60 Gram-negative and Gram-positive bacterial species as well as methicillin resistance marker from a blood culture. Bacterial findings were accurately reported from 77 blood culture samples, whereas 14 samples were reported as negative, containing bacteria not belonging to the pathogen panel of the assay. No difference was observed between the performance of NorDiag Arrow or NucliSENS® easyMAG® with regard to the result reporting of Prove-it™ Sepsis. In addition, we also assessed the quality and quantity of DNA extracted from the clinical Escherichia coli isolate with DNA extraction instruments. We observed only minor differences between the two instruments. Conclusions: Use of automated and standardized sample preparation methods together with rapid, multiplex pathogen detection offers a strategy to speed up reliably the diagnostics of septic patients. Both tested DNA extraction devices were shown to be feasible for blood culture samples and the Prove-it™ Sepsis assay, providing an accurate identification of pathogen within 4,5 hours when the detected pathogen was in the repertoire of the test. © 2011 Laakso et al.


Aittakorpi A.,Mobidiag Ltd. | Kuusela P.,University of Helsinki | Koukila-Kahkola P.,University of Helsinki | Vaara M.,University of Helsinki | And 3 more authors.
Journal of Clinical Microbiology | Year: 2012

The rapid identification of microbes responsible for bloodstream infections (BSIs) allows more focused and effective therapies and outcomes. DNA sequence-based methods offer an opportunity for faster, accurate diagnosis and for effective therapy. As our objective of the study, the ability of the Prove-it Sepsis platform, already proven as a rapid PCR- and microarray-based assay for the majority of sepsis-causing bacteria, was extended to also rapidly identify clinically relevant yeasts in blood culture. The performance characteristics of this extended platform are described. We found that the extended diagnostic Prove-it Sepsis platform was found to be highly accurate when analyzing primary isolates, spiked blood cultures, nucleic acid extracts from a retrospective blood culture data set, and primary blood cultures. Comparison of the blood culture results from the Prove-it Sepsis platform with those from conventional culture-based methods or by gene sequencing demonstrated a sensitivity of 99% and a specificity of 98% for fungal targets (based on analysis of a total of 388 specimens). Total assay time was 3 h from DNA extraction to BSI diagnosis. These results extend the performance characteristics of the Prove-it platform for bacteria to the easy, rapid, and accurate detection and species identification of yeasts in positive blood cultures. Incorporation of this extended and rapid diagnostic platform into the tools for clinical patient management would allow possibly faster identification and more focused therapies for BSIs. Copyright © 2012, American Society for Microbiology. All Rights Reserved.


Liedert R.,VTT Technical Research Center of Finland | Amundsen L.K.,VTT Technical Research Center of Finland | Hokkanen A.,VTT Technical Research Center of Finland | Maki M.,Mobidiag Ltd. | And 10 more authors.
Lab on a Chip - Miniaturisation for Chemistry and Biology | Year: 2012

We present a high-throughput roll-to-roll (R2R) manufacturing process for foil-based polymethyl methacrylate (PMMA) chips of excellent optical quality. These disposable, R2R hot embossed microfluidic chips are used for the identification of the antibiotic resistance gene mecA in Staphylococcus epidermidis. R2R hot embossing is an emerging manufacturing technology for polymer microfluidic devices. It is based on continuous feeding of a thermoplastic foil through a pressurized area between a heated embossing cylinder and a blank counter cylinder. Although mass fabrication of foil-based microfluidic chips and their use for biological applications were foreseen already some years ago, no such studies have been published previously. © 2012 The Royal Society of Chemistry.

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