title: Increased viability but decreased culturability of Mycobacterium avium subsp. paratuberculosis in macrophages from inflammatory bowel disease patients under Infliximab treatment.
abstract: Mycobacterium avium subsp. paratuberculosis (MAP) has long been implicated as a triggering agent in Crohn's disease (CD). In this study, we investigated the growth/persistence of both M. avium subsp. hominissuis (MAH) and MAP, in macrophages from healthy controls (HC), CD and ulcerative colitis patients. For viability assessment, both CFU counts and a pre16SrRNA RNA/DNA ratio assay (for MAP) were used. Phagolysosome fusion was evaluated by immunofluorescence, through analysis of LAMP-1 colocalization with MAP. IBD macrophages were more permissive to MAP survival than HC macrophages (a finding not evident with MAH), but did not support MAP active growth. The lower MAP CFU counts in macrophage cultures associated with Infliximab treatment were not due to increased killing, but possibly to elevation in the proportion of intracellular dormant non-culturable MAP forms, as MAP showed higher viability in those macrophages. Increased MAP viability was not related to lack of phagolysosome maturation. The predominant induction of MAP dormant forms by Infliximab treatment may explain the lack of MAP reactivation during anti-TNF therapy of CD but does not exclude the possibility of MAP recrudescence after termination of therapy.
authors.affiliation: FP-ENAS (UFP Energy, Environment and Health Research Unit), CEBIMED (Biomedical Research Centre), University Fernando Pessoa, Rua Carlos da Maia, 296, 4200-150, Porto, Portugal.
authors.forename: Nair
authors.initials: N
authors.lastname: Nazareth
authors.affiliation: Institute of Pharmacology and Therapeutics, Faculdade de Medicina, Universidade do Porto, Alameda Professor Hernâni Monteiro, 4200-319, Porto, Portugal.|Gastroenterology Department, Centro Hospitalar S. João, Alameda Professor Hernâni Monteiro, 4200-319, Porto, Portugal.|MedInUP - Center for Drug Discovery and Innovative Medicines, Universidade do Porto, Alameda Professor Hernâni Monteiro, 4200-319, Porto, Portugal.
authors.forename: Fernando
authors.initials: F
authors.lastname: Magro
authors.affiliation: Infection and Immunity Unit, IBMC - Instituto de Biologia Molecular e Celular, Universidade do Porto, Rua do Campo Alegre, 823, 4150-180, Porto, Portugal.|Department of Infectious Diseases, Centro Hospitalar S. João, Alameda Professor Hernâni Monteiro, 4200-319, Porto, Portugal.
authors.forename: Rui
authors.initials: R
authors.lastname: Appelberg
authors.affiliation: FP-ENAS (UFP Energy, Environment and Health Research Unit), CEBIMED (Biomedical Research Centre), University Fernando Pessoa, Rua Carlos da Maia, 296, 4200-150, Porto, Portugal.
authors.forename: Jani
authors.initials: J
authors.lastname: Silva
authors.affiliation: Institute of Pharmacology and Therapeutics, Faculdade de Medicina, Universidade do Porto, Alameda Professor Hernâni Monteiro, 4200-319, Porto, Portugal.|MedInUP - Center for Drug Discovery and Innovative Medicines, Universidade do Porto, Alameda Professor Hernâni Monteiro, 4200-319, Porto, Portugal.
authors.forename: Daniela
authors.initials: D
authors.lastname: Gracio
authors.affiliation: Gastroenterology Department, Centro Hospitalar S. João, Alameda Professor Hernâni Monteiro, 4200-319, Porto, Portugal.
authors.forename: Rosa
authors.initials: R
authors.lastname: Coelho
authors.affiliation: Institute of Pharmacology and Therapeutics, Faculdade de Medicina, Universidade do Porto, Alameda Professor Hernâni Monteiro, 4200-319, Porto, Portugal.|MedInUP - Center for Drug Discovery and Innovative Medicines, Universidade do Porto, Alameda Professor Hernâni Monteiro, 4200-319, Porto, Portugal.
authors.forename: José Miguel
authors.initials: JM
authors.lastname: Cabral
authors.affiliation: Department of Infectious Diseases, Centro Hospitalar S. João, Alameda Professor Hernâni Monteiro, 4200-319, Porto, Portugal.|Nephrology Research and Development Unit, Faculdade de Medicina, Universidade do Porto, Alameda Professor Hernâni Monteiro, 4200-319, Porto, Portugal.
authors.forename: Candida
authors.initials: C
authors.lastname: Abreu
authors.affiliation: Gastroenterology Department, Centro Hospitalar S. João, Alameda Professor Hernâni Monteiro, 4200-319, Porto, Portugal.
authors.forename: Guilherme
authors.initials: G
authors.lastname: Macedo
authors.affiliation: Infection and Immunity Research Institute, St George's University of London, Cranmer Terrace, London, SW17 0RE, UK.
authors.forename: Tim J
authors.initials: TJ
authors.lastname: Bull
authors.affiliation: FP-ENAS (UFP Energy, Environment and Health Research Unit), CEBIMED (Biomedical Research Centre), University Fernando Pessoa, Rua Carlos da Maia, 296, 4200-150, Porto, Portugal. assuncao@ufp.edu.pt.|Infection and Immunity Unit, IBMC - Instituto de Biologia Molecular e Celular, Universidade do Porto, Rua do Campo Alegre, 823, 4150-180, Porto, Portugal. assuncao@ufp.edu.pt.
authors.forename: Amélia
authors.initials: A
authors.lastname: Sarmento
mesh_terms: D000328:Adult; D000368:Aged; D058491:Bacterial Load; D016022:Case-Control Studies; D005260:Female; D006801:Humans; D015212:Inflammatory Bowel Diseases / Q000150:complications* / Q000188:drug therapy; D000069285:Infliximab / Q000009:adverse effects* / Q000627:therapeutic use; D052118:Lysosomal-Associated Membrane Protein 1 / Q000378:metabolism; D008264:Macrophages / Q000382:microbiology*; D008297:Male; D050296:Microbial Viability / Q000276:immunology; D008875:Middle Aged; D016927:Mycobacterium avium subsp. paratuberculosis / Q000235:genetics / Q000276:immunology*; D010283:Paratuberculosis / Q000209:etiology* / Q000382:microbiology*; D010587:Phagocytosis; D010588:Phagosomes / Q000276:immunology / Q000382:microbiology; D012336:RNA, Ribosomal, 16S / Q000235:genetics; D055815:Young Adult
chemical_list: D052118:Lysosomal-Associated Membrane Protein 1; D012336:RNA, Ribosomal, 16S; D000069285:Infliximab
keywords: Inflammatory bowel disease; Macrophages; Mycobacterium avium subsp. paratuberculosis; Phagosomal maturation