title: Standardization and international multicenter validation of a PulseNet pulsed-field gel electrophoresis protocol for subtyping Shigella flexneri isolates.
abstract: Shigella flexneri is one of the agents most frequently linked to diarrheal illness in developing countries and often causes outbreaks in settings with poor hygiene or sanitary conditions. Travel is one of the means by which S. flexneri can be imported into developed countries, where this pathogen is not commonly seen. A robust and discriminatory subtyping method is needed for the surveillance of S. flexneri locally and regionally, and to aid in the detection and investigation of outbreaks. The PulseNet International network utilizes standardized pulsed-field gel electrophoresis (PFGE) protocols to carry out laboratory-based surveillance of foodborne pathogens in combination with epidemiologic data. A multicenter validation was carried out in nine PulseNet laboratories located in North and South America, Europe, and Asia, and it demonstrated that a new protocol is highly robust and reproducible for subtyping of S. flexneri. This protocol, already approved for PulseNet laboratories, applies NotI and XbaI as primary and secondary restriction enzymes, respectively, under electrophoresis conditions of initial switch time of 5 s to final switch time of 35 s, at 6 volts/cm.
authors.affiliation: Instituto Nacional de Enfermedades Infecciosas, ANLIS "Carlos G. Malbrán," Buenos Aires, Argentina. mpichel@anlis.gov.ar
authors.forename: Mariana
authors.initials: M
authors.lastname: Pichel
authors.forename: Silvina P
authors.initials: SP
authors.lastname: Brengi
authors.forename: Kara L F
authors.initials: KL
authors.lastname: Cooper
authors.forename: Efrain M
authors.initials: EM
authors.lastname: Ribot
authors.forename: Suleiman
authors.initials: S
authors.lastname: Al-Busaidy
authors.forename: Pamela
authors.initials: P
authors.lastname: Araya
authors.forename: Jorge
authors.initials: J
authors.lastname: Fernández
authors.forename: Tania Ibelli
authors.initials: TI
authors.lastname: Vaz
authors.forename: Kai Man
authors.initials: KM
authors.lastname: Kam
authors.forename: Myriam
authors.initials: M
authors.lastname: Morcos
authors.forename: Eva M
authors.initials: EM
authors.lastname: Nielsen
authors.forename: Celine
authors.initials: C
authors.lastname: Nadon
authors.forename: Guillermo
authors.initials: G
authors.lastname: Pimentel
authors.forename: Enrique
authors.initials: E
authors.lastname: Pérez-Gutiérrez
authors.forename: Peter
authors.initials: P
authors.lastname: Gerner-Smidt
authors.forename: Norma
authors.initials: N
authors.lastname: Binsztein
mesh_terms: D015373:Bacterial Typing Techniques / Q000592:standards; D004269:DNA, Bacterial / Q000737:chemistry / Q000378:metabolism*; D003718:Denmark; D015252:Deoxyribonucleases, Type II Site-Specific / Q000378:metabolism; D004405:Dysentery, Bacillary / Q000175:diagnosis / Q000382:microbiology; D016521:Electrophoresis, Gel, Pulsed-Field; D006723:Hong Kong; D008877:Middle East; D009656:North America; D011786:Quality Control; D015203:Reproducibility of Results; D012763:Shigella flexneri / Q000145:classification* / Q000302:isolation & purification / Q000378:metabolism; D013020:South America; D013997:Time Factors
chemical_list: D004269:DNA, Bacterial; C025580:endodeoxyribonuclease XBAI; D015252:Deoxyribonucleases, Type II Site-Specific; C038565:GCGGCCGC-specific type II deoxyribonucleases