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Journal article
Transformed Recombinant Enrichment Profiling Rapidly Identifies HMW1 as an Intracellular Invasion Locus in Haemophilus influenzae
PLoS pathogens, v 12(4), pe1005576
01 Apr 2016
PMID: 27124727
Featured in Collection : UN Sustainable Development Goals @ Drexel
Abstract
Many bacterial species actively take up and recombine homologous DNA into their genomes, called natural competence, a trait that offers a means to identify the genetic basis of naturally occurring phenotypic variation. Here, we describe "transformed recombinant enrichment profiling" (TREP), in which natural transformation is used to generate complex pools of recombinants, phenotypic selection is used to enrich for specific recombinants, and deep sequencing is used to survey for the genetic variation responsible. We applied TREP to investigate the genetic architecture of intracellular invasion by the human pathogen Haemophilus influenzae, a trait implicated in persistence during chronic infection. TREP identified the HMW1 adhesin as a crucial factor. Natural transformation of the hmw1 operon from a clinical isolate (86-028NP) into a laboratory isolate that lacks it (Rd KW20) resulted in similar to 1,000-fold increased invasion into airway epithelial cells. When a distinct recipient (Hi375, already possessing hmw1 and its paralog hmw2) was transformed by the same donor, allelic replacement of hmw2A(Hi375) by hmw1A(86-028NP) resulted in a similar to 100-fold increased intracellular invasion rate. The specific role of hmw1A(86-028NP) was confirmed by mutant and western blot analyses. Bacterial self-aggregation and adherence to airway cells were also increased in recombinants, suggesting that the high invasiveness induced by hmw1A(86-028NP) might be a consequence of these phenotypes. However, immunofluorescence results found that intracellular hmw1A(86-028NP) bacteria likely invaded as groups, instead of as individual bacterial cells, indicating an emergent invasion-specific consequence of hmw1A-mediated self-aggregation.
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Details
- Title
- Transformed Recombinant Enrichment Profiling Rapidly Identifies HMW1 as an Intracellular Invasion Locus in Haemophilus influenzae
- Creators
- Joshua Chang Mell - Drexel UniversityCristina Viadas - Universidad Publica de NavarraJavier Moleres - Universidad Publica de NavarraSunita Sinha - University of British ColumbiaAriadna Fernandez-Calvet - Universidad Publica de NavarraEric A. Porsch - Children's Hospital of PhiladelphiaJoseph W. St Geme - Children's Hospital of PhiladelphiaCorey Nislow - University of British ColumbiaRosemary J. Redfield - University of British ColumbiaJunkal Garmendia - Universidad Publica de Navarra
- Publication Details
- PLoS pathogens, v 12(4), pe1005576
- Publisher
- Public Library Science
- Number of pages
- 36
- Grant note
- F32AI084427 / NATIONAL INSTITUTE OF ALLERGY AND INFECTIOUS DISEASES; United States Department of Health & Human Services; National Institutes of Health (NIH) - USA; NIH National Institute of Allergy & Infectious Diseases (NIAID) SAF2012-31166; SAF2015-66520-R / Ministerio Economia y Competitividad-MINECO Canadian Institute of Health Research; Canadian Institutes of Health Research (CIHR) 359/2012 / Dpto. Salud Gobierno Navarra F32AI084427 / National Institutes of Health Ruth Kirschstein postdoctoral fellowship; United States Department of Health & Human Services; National Institutes of Health (NIH) - USA DC002873 / National Institutes of Heath R01 grant PRX12/00191 / Ministerio de Educacion; German Research Foundation (DFG) Faculty of Pharmaceutical Sciences, Canadian Foundation for Innovation R01DC002873 / NATIONAL INSTITUTE ON DEAFNESS AND OTHER COMMUNICATION DISORDERS; United States Department of Health & Human Services; National Institutes of Health (NIH) - USA; NIH National Institute on Deafness & Other Communication Disorders (NIDCD) SOF122 / Genome British Columbia
- Resource Type
- Journal article
- Language
- English
- Academic Unit
- Microbiology and Immunology
- Web of Science ID
- WOS:000378156900053
- Scopus ID
- 2-s2.0-84964845763
- Other Identifier
- 991019168457904721
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- Collaboration types
- Domestic collaboration
- International collaboration
- Web of Science research areas
- Microbiology
- Parasitology
- Virology