title: Profiling of Leptospira interrogans, L. santarosai, L. meyeri and L. borgpetersenii by SE-AFLP, PFGE and susceptibility testing--a continuous attempt at species and serovar differentiation.
abstract: Leptospirosis is a widespread systemic zoonosis, considered as reemerging in certain developing countries. Although the cross agglutinin absorption test is still considered the standard method for Leptospira identification, it presents several disadvantages. The aim of this study was to characterize Leptospira spp. isolated from various hosts by genotyping and broth microdilution susceptibility testing in an attempt to differentiate Leptospira species, serogroups and serovars. Forty-seven isolates were studied. They were previously serotyped, and species confirmation was performed by 16S rRNA sequencing. Single-enzyme amplified fragment length polymorphism (SE-AFLP) and pulsed-field gel electrophoresis (PFGE) analysis enabled the distinction of L. interrogans from L. santarosai, L. meyeri and L. borgpetersenii in two main clusters. Among L. interrogans, it was possible to differentiate into two new clusters the serogroup Icterohaemorrhagiae from the serogroups Canicola and Pomona. L. santarosai isolates presented higher genetic variation than the other species in both techniques. Interestingly, the minimum inhibitory concentration (MIC) cluster analysis also provided Leptospira serogroup differentiation. Further studies are necessary regarding serovar Bananal isolates, as they presented the highest MIC values for most of the antimicrobials tested. All studied techniques successfully distinguished Leptospira species and serogroups. Despite being library-dependent methods, these approaches are less labor intensive and more economically viable, particularly SE-AFLP, and can be implemented in most reference laboratories worldwide to enable faster Leptospira typing.
authors.affiliation: Laboratory of Molecular Epidemiology and Antimicrobial Resistance/Laboratory of Bacterial Zoonosis, School of Veterinary Medicine and Animal Science, University of São Paulo, São Paulo/SP 05508 270, Brazil.
authors.forename: Luisa Z
authors.initials: LZ
authors.lastname: Moreno
authors.affiliation: Laboratory of Molecular Epidemiology and Antimicrobial Resistance/Laboratory of Bacterial Zoonosis, School of Veterinary Medicine and Animal Science, University of São Paulo, São Paulo/SP 05508 270, Brazil.|Veterinary Bacteriology Laboratory, Department of Microbiology and Parasitology, Fluminense Federal University, Hernani Mello, 101 Niterói/RJ 24210 130, Brazil.
authors.forename: Fabiana
authors.initials: F
authors.lastname: Miraglia
authors.affiliation: Veterinary Bacteriology Laboratory, Department of Microbiology and Parasitology, Fluminense Federal University, Hernani Mello, 101 Niterói/RJ 24210 130, Brazil.
authors.forename: Walter
authors.initials: W
authors.lastname: Lilenbaum
authors.affiliation: Laboratory of Molecular Epidemiology and Antimicrobial Resistance/Laboratory of Bacterial Zoonosis, School of Veterinary Medicine and Animal Science, University of São Paulo, São Paulo/SP 05508 270, Brazil.
authors.forename: José S F
authors.initials: JS
authors.lastname: Neto
authors.affiliation: Laboratory of Animal Leptospirosis, Londrina State University, Rod Celso Garcia Cid, Londrina/PR 86057 970, Brazil.
authors.forename: Julio C
authors.initials: JC
authors.lastname: Freitas
authors.affiliation: Laboratory of Molecular Epidemiology and Antimicrobial Resistance/Laboratory of Bacterial Zoonosis, School of Veterinary Medicine and Animal Science, University of São Paulo, São Paulo/SP 05508 270, Brazil.
authors.forename: Zenaide M
authors.initials: ZM
authors.lastname: Morais
authors.affiliation: WHO/FAO/OIE and National Leptospirosis Reference Centre, KIT Biomedical Research, Royal Tropical Institute, Amsterdam 391105, Netherlands.
authors.forename: Rudy A
authors.initials: RA
authors.lastname: Hartskeerl
authors.affiliation: Laboratory of Molecular Epidemiology and Antimicrobial Resistance/Laboratory of Bacterial Zoonosis, School of Veterinary Medicine and Animal Science, University of São Paulo, São Paulo/SP 05508 270, Brazil.
authors.forename: Barbara L P
authors.initials: BL
authors.lastname: da Costa
authors.affiliation: Laboratory of Molecular Epidemiology and Antimicrobial Resistance/Laboratory of Bacterial Zoonosis, School of Veterinary Medicine and Animal Science, University of São Paulo, São Paulo/SP 05508 270, Brazil.
authors.forename: Silvio A
authors.initials: SA
authors.lastname: Vasconcellos
authors.affiliation: Laboratory of Molecular Epidemiology and Antimicrobial Resistance/Laboratory of Bacterial Zoonosis, School of Veterinary Medicine and Animal Science, University of São Paulo, São Paulo/SP 05508 270, Brazil.
authors.forename: Andrea M
authors.identifier: http://orcid.org/0000-0002-3290-566X
authors.initials: AM
authors.lastname: Moreno
mesh_terms: D054458:Amplified Fragment Length Polymorphism Analysis; D000900:Anti-Bacterial Agents / Q000494:pharmacology; D015373:Bacterial Typing Techniques; D016521:Electrophoresis, Gel, Pulsed-Field; D014644:Genetic Variation; D005838:Genotype; D007919:Leptospira / Q000145:classification* / Q000187:drug effects / Q000235:genetics* / Q000276:immunology; D007921:Leptospira interrogans / Q000145:classification / Q000235:genetics / Q000276:immunology; D008826:Microbial Sensitivity Tests; D011110:Polymorphism, Genetic; D012336:RNA, Ribosomal, 16S; D017423:Sequence Analysis, RNA; D065288:Serogroup; D012703:Serotyping
chemical_list: D000900:Anti-Bacterial Agents; D012336:RNA, Ribosomal, 16S