Multiplex detection of meningitis pathogens by a vertical flow paper microarray and signal enhancement suitable for low-resource settings: Proof of concept
Pedro Réu,
Giulia Gaudenzi,
Deborah Nanjebe,
Gustav Svedberg,
Dan Nyehangane,
Miren Urrutia Iturritza,
Phuthumani Mlotshwa,
Chris Hadjineophytou,
Jens Karlsson,
Jesper Gantelius,
Juliet Mwanga-Amumpaire,
Edmund Loh,
Helene Andersson Svahn,
Elias Kumbakumba,
Tobias Alfvén,
Yap Boum II,
Aman Russom
Affiliations
Pedro Réu
Division of Nanobiotechnology, Department of Protein Science, Science for Life Laboratory, KTH Royal Institute of Technology, Solna, Sweden
Giulia Gaudenzi
Division of Nanobiotechnology, Department of Protein Science, Science for Life Laboratory, KTH Royal Institute of Technology, Solna, Sweden; Department of Global Public Health, Karolinska Institutet, Stockholm, Sweden; Corresponding author at: Tomtebodavägen 18A, Solna, Sweden.
Deborah Nanjebe
MSF Epicentre Mbarara Research Centre, Mbarara, Uganda
Gustav Svedberg
Division of Nanobiotechnology, Department of Protein Science, Science for Life Laboratory, KTH Royal Institute of Technology, Solna, Sweden
Dan Nyehangane
MSF Epicentre Mbarara Research Centre, Mbarara, Uganda
Miren Urrutia Iturritza
Division of Nanobiotechnology, Department of Protein Science, Science for Life Laboratory, KTH Royal Institute of Technology, Solna, Sweden
Phuthumani Mlotshwa
Department of Global Public Health, Karolinska Institutet, Stockholm, Sweden
Chris Hadjineophytou
Department of Microbiology, Tumor and Cell Biology, Karolinska Institutet, Stockholm, Sweden
Jens Karlsson
Department of Microbiology, Tumor and Cell Biology, Karolinska Institutet, Stockholm, Sweden
Jesper Gantelius
Division of Nanobiotechnology, Department of Protein Science, Science for Life Laboratory, KTH Royal Institute of Technology, Solna, Sweden
Juliet Mwanga-Amumpaire
MSF Epicentre Mbarara Research Centre, Mbarara, Uganda; Department of Paediatrics and Child Health, Faculty of Medicine, Mbarara University of Science and Technology, Mbarara, Uganda
Edmund Loh
Department of Microbiology, Tumor and Cell Biology, Karolinska Institutet, Stockholm, Sweden
Helene Andersson Svahn
Division of Nanobiotechnology, Department of Protein Science, Science for Life Laboratory, KTH Royal Institute of Technology, Solna, Sweden
Elias Kumbakumba
Department of Paediatrics and Child Health, Faculty of Medicine, Mbarara University of Science and Technology, Mbarara, Uganda
Tobias Alfvén
Department of Global Public Health, Karolinska Institutet, Stockholm, Sweden; Sachs' Children & Youth Hospital, South General Hospital, Stockholm, Sweden
Yap Boum II
Institut Pasteur of Bangui, Bangui, Central Africa Republic
Aman Russom
Division of Nanobiotechnology, Department of Protein Science, Science for Life Laboratory, KTH Royal Institute of Technology, Solna, Sweden; AIMES Center for the Advancement of Integrated Medical and Engineering Sciences at Karolinska Institutet and KTH Royal Institute of Technology, Stockholm, Sweden
Objectives: Meningitis is a medical emergency, and it is crucial to diagnose it accurately and promptly in order to manage patients effectively. It would, therefore, be essential to introduce and have fast, accurate, and user-friendly methods to determine the cause of these infections. This study aimed to demonstrate a potentially cost-effective new approach for detecting meningitis using a paper-based vertical flow microarray, which could be useful in settings with limited resources. Methods: We describe a multiplex paper microarray for detecting Neisseria meningitidis, Haemophilus influenzae, Streptococcus pneumoniae, and Salmonella spp. by the passive vertical flow of PCR-amplified clinical samples. A multibiotinylated amplicon was obtained as a product of PCR in the presence of both a biotinylated primer and biotin-11-dUTP. An enhancement step based on an enzyme-free gold enhancement protocol was also used to facilitate visual detection. Results: This study showed that the vertical flow microarray (previously evaluated for one pathogen) can discriminately detect the amplification results down to the 102 copies of DNA limit for four meningitis pathogens in a multiplexed set-up. The study further demonstrated the ability of this device and setup to detect three of the four pathogens from clinical biosamples. Discussion: This study demonstrated the capacity of a vertical flow microarray device to detect amplification products for four prevalent meningitis pathogens in a multiplex format. The vertical flow microarray demonstrated consistent visualization of the expected gene amplification results; however, indicating limitations in the pre- and amplification steps. This study highlights the potential of this multiplexing method for diagnosing meningitis and other syndromic diseases caused by various pathogens, especially in resource-limited areas.
