title: Separating Putative Pathogens from Background Contamination with Principal Orthogonal Decomposition: Evidence for Leptospira in the Ugandan Neonatal Septisome.
abstract: Neonatal sepsis (NS) is responsible for over 1 million yearly deaths worldwide. In the developing world, NS is often treated without an identified microbial pathogen. Amplicon sequencing of the bacterial 16S rRNA gene can be used to identify organisms that are difficult to detect by routine microbiological methods. However, contaminating bacteria are ubiquitous in both hospital settings and research reagents and must be accounted for to make effective use of these data. In this study, we sequenced the bacterial 16S rRNA gene obtained from blood and cerebrospinal fluid (CSF) of 80 neonates presenting with NS to the Mbarara Regional Hospital in Uganda. Assuming that patterns of background contamination would be independent of pathogenic microorganism DNA, we applied a novel quantitative approach using principal orthogonal decomposition to separate background contamination from potential pathogens in sequencing data. We designed our quantitative approach contrasting blood, CSF, and control specimens and employed a variety of statistical random matrix bootstrap hypotheses to estimate statistical significance. These analyses demonstrate that Leptospira appears present in some infants presenting within 48 h of birth, indicative of infection in utero, and up to 28 days of age, suggesting environmental exposure. This organism cannot be cultured in routine bacteriological settings and is enzootic in the cattle that often live in close proximity to the rural peoples of western Uganda. Our findings demonstrate that statistical approaches to remove background organisms common in 16S sequence data can reveal putative pathogens in small volume biological samples from newborns. This computational analysis thus reveals an important medical finding that has the potential to alter therapy and prevention efforts in a critically ill population.
authors.affiliation: Center for Neural Engineering, Pennsylvania State University, University Park, PA, USA; Department of Neurosurgery, Penn State College of Medicine, Hershey, PA, USA; Department of Engineering Science and Mechanics, Pennsylvania State University, University Park, PA, USA; Department of Physics, Pennsylvania State University, University Park, PA, USA.
authors.forename: Steven J
authors.initials: SJ
authors.lastname: Schiff
authors.affiliation: Department of Pediatrics, Mbarara University of Science and Technology , Mbarara , Uganda.
authors.forename: Julius
authors.initials: J
authors.lastname: Kiwanuka
authors.affiliation: Department of Biology, Pennsylvania State University, University Park, PA, USA; Department of Veterinary and Biomedical Sciences, Pennsylvania State University, University Park, PA, USA.
authors.forename: Gina
authors.initials: G
authors.lastname: Riggio
authors.affiliation: Department of Biology, Pennsylvania State University, University Park, PA, USA; Department of Veterinary and Biomedical Sciences, Pennsylvania State University, University Park, PA, USA; Department of Biochemistry and Molecular Biology, Institute for Personalized Medicine, Pennsylvania State University College of Medicine, Hershey, PA, USA.
authors.forename: Lan
authors.initials: L
authors.lastname: Nguyen
authors.affiliation: Department of Biology, Pennsylvania State University, University Park, PA, USA; Department of Veterinary and Biomedical Sciences, Pennsylvania State University, University Park, PA, USA.
authors.forename: Kevin
authors.initials: K
authors.lastname: Mu
authors.affiliation: Department of Biology, Pennsylvania State University, University Park, PA, USA; Department of Veterinary and Biomedical Sciences, Pennsylvania State University, University Park, PA, USA.
authors.forename: Emily
authors.initials: E
authors.lastname: Sproul
authors.affiliation: Department of Microbiology, Mbarara University of Science and Technology , Mbarara , Uganda.
authors.forename: Joel
authors.initials: J
authors.lastname: Bazira
authors.affiliation: Department of Pediatrics, Mbarara University of Science and Technology, Mbarara, Uganda; Epicentre Mbarara Research Centre, Mbarara, Uganda.
authors.forename: Juliet
authors.initials: J
authors.lastname: Mwanga-Amumpaire
authors.affiliation: Department of Pediatrics, Mbarara University of Science and Technology , Mbarara , Uganda.
authors.forename: Dickson
authors.initials: D
authors.lastname: Tumusiime
authors.affiliation: Department of Pediatrics, Mbarara University of Science and Technology , Mbarara , Uganda.
authors.forename: Eunice
authors.initials: E
authors.lastname: Nyesigire
authors.affiliation: Department of Microbiology, Mbarara University of Science and Technology , Mbarara , Uganda.
authors.forename: Nkangi
authors.initials: N
authors.lastname: Lwanga
authors.affiliation: Center for Neural Engineering, Pennsylvania State University, University Park, PA, USA; Schreyer's Honors College, Pennsylvania State University, University Park, PA, USA.
authors.forename: Kaleb T
authors.initials: KT
authors.lastname: Bogale
authors.affiliation: Department of Veterinary and Biomedical Sciences, Pennsylvania State University , University Park, PA , USA.
authors.forename: Vivek
authors.initials: V
authors.lastname: Kapur
authors.affiliation: Department of Biochemistry and Molecular Biology, Institute for Personalized Medicine, Pennsylvania State University College of Medicine , Hershey, PA , USA.
authors.forename: James R
authors.initials: JR
authors.lastname: Broach
authors.affiliation: Center for Neural Engineering, Pennsylvania State University, University Park, PA, USA; Harvard Neonatal-Perinatal Training Program, Children's Hospital Boston, Boston, MA, USA.
authors.forename: Sarah U
authors.initials: SU
authors.lastname: Morton
authors.affiliation: Department of Neurosurgery, Harvard Medical School, Boston Children's Hospital, Boston, MA, USA; Department of Global Health and Social Medicine, Harvard Medical School, Boston Children's Hospital, Boston, MA, USA; CURE Children's Hospital of Uganda, Mbale, Uganda.
authors.forename: Benjamin C
authors.initials: BC
authors.lastname: Warf
authors.affiliation: Department of Biology, Pennsylvania State University, University Park, PA, USA; Department of Veterinary and Biomedical Sciences, Pennsylvania State University, University Park, PA, USA.
authors.forename: Mary
authors.initials: M
authors.lastname: Poss
keywords: 16S rRNA; Leptospira; bacteria; neonatal sepsis; principal orthogonal decomposition; singular value decomposition