Examinando por Autor "Sellek Cano, Ricela"

Mostrando 1 - 4 de 4

Acceso Abierto
OmpU as a biomarker for rapid discrimination between toxigenic and epidemic Vibrio cholerae O1/O139 and non-epidemic Vibrio cholerae in a modified MALDI-TOF MS assay
(Springer Nature Link, 2014-06-18) Paauw, Armand; Trip, Hein; Niemcewicz, Marcin; Sellek Cano, Ricela; Heng, Jonathan; Mars Groenendijk, Roos; De Jong, Ad; Majchrzykiewicz Koehorst, Joanna; Olsen, Jaran; Tsivtsivadze, Evgeni
Background Cholera is an acute diarrheal disease caused by Vibrio cholerae. Outbreaks are caused by a genetically homogenous group of strains from serogroup O1 or O139 that are able to produce the cholera toxin. Rapid detection and identification of these epidemic strains is essential for an effective response to cholera outbreaks. Results The use of ferulic acid as a matrix in a new MALDI-TOF MS assay increased the measurable mass range of existing MALDI-TOF MS protocols for bacterial identification. The assay enabled rapid discrimination between epidemic V. cholerae O1/O139 strains and other less pathogenic V. cholerae strains. OmpU, an outer membrane protein whose amino acid sequence is highly conserved among epidemic strains of V. cholerae, appeared as a discriminatory marker in the novel MALDI-TOF MS assay. Conclusions The extended mass range of MALDI-TOF MS measurements obtained by using ferulic acid improved the screening for biomarkers in complex protein mixtures. Differences in the mass of abundant homologous proteins due to variation in amino acid sequences can rapidly be examined in multiple samples. Here, a rapid MALDI-TOF MS assay was developed that could discriminate between epidemic O1/O139 strains and other less pathogenic V. cholerae strains based on differences in mass of the OmpU protein. It appeared that the amino acid sequence of OmpU from epidemic V. cholerae O1/O139 strains is unique and highly conserved.
Acceso Abierto
Phenotypic and genetic analyses of 111 clinical and environmental O1, O139, and non-O1/O139 Vibrio cholerae strains from different geographical areas
(Cambridge University Press, 2012-08-05) Sellek Cano, Ricela; Niemcewicz, Marcin; Olsen, Jaran; Bassy Alvarez, Olga; Lorenzo Lozano, P.; Martí, L.; Roszowiak, A.; Kocik, J.; Cabria Ramos, J. C.
A total of 111 clinical and environmental O1, O139 and non-O1/O139 Vibrio cholerae strains isolated between 1978 and 2008 from different geographical areas were typed using a combination of methods: antibiotic susceptibility, biochemical test, serogroup, serotype, biotype, sequences containing variable numbers of tandem repeats (VNTRs) and virulence genes ctxA and tcpA amplification. As a result of the performed typing work, the strains were organized into four clusters: cluster A1 included clinical O1 Ogawa and O139 serogroup strains (ctxA + and tcpA +); cluster A2 included clinical non-O1/O139 strains (ctxA − and tcpA −), as well as environmental O1 Inaba and non-O1/O139 strains (ctxA − and tcpA −/tcpA +); cluster B1 contained two clinical O1 strains and environmental non-O1/O139 strains (ctxA − and tcpA +/tcpA −); cluster B2 contained clinical O1 Inaba and Ogawa strains (ctxA + and tcpA +). The results of this work illustrate the advantage of combining several typing methods to discriminate between clinical and environmental V. cholerae strains.
Restringido
Preliminary validation of real-time PCR assays for the identification of Yersinia pestis
(De Gruyter Brill, 2008-07-21) Tomaso, Herbert; Jacob, Daniela; Eickhoff, Meike; Scholz, Holger; Al Dahouk, Sascha; Kattar, Mireille; Reischl, Udo; Plicka, Helga; Strand Olsen, Jaran; Nikkari, Simo; Matero, Pirjo; Beuret, Christian; Ciammaruconi, Andrea; Lista, Florigio; Gala, Jean Luc; Broll, Hermann; Appel, Bernd; Sellek Cano, Ricela; Ybarra, María del Carmen; Broekhuijsen, Martien; Indra, Alexander; Petersen, Roger; Neubauer, Heinrich
Background: Yersinia pestis (Y. pestis) is a zoonotic bacterium mainly circulating among rodents and their fleas. Transmission to humans can cause bubonic, pneumonic or septicemic plague with a high case-fatality rate. Therefore, rapid and reliable diagnostic tools are crucial. The objective of this study was to assess the inter-laboratory reproducibility of in-house developed real-time PCR assays for the identification of Y. pestis. Methods: A total of four samples of quantified Y. pestis DNA and two blank samples were sent blinded to 14 laboratories. To standardize the procedures, oligonucleotides were provided and the same instrument platform and a commercial mastermix were used. The participants were requested to report their results including cycle threshold and melting temperature values. Results: All participating laboratories were able to perform the real-time PCR assays according to the protocols provided and identified the samples containing Y. pestis DNA correctly. Significant differences between the reference laboratory and participating laboratories were observed in cycle threshold values and melting temperatures. This, however, did not adversely affect the interpretation of results. Conclusions: Our real-time PCR system proved to be highly reproducible and has the potential of complementing the diagnostic tools for rapid identification of Y. pestis isolates. Further steps of validation are needed to determine diagnostic accuracy and predictive values with clinical samples.
Restringido
Recovery of Francisella tularensis from soil samples by filtration and detection by real-time PCR and cELISA
(Royal Society of Chemistry, 2008-01-17) Sellek Cano, Ricela; Jiménez, Oscar; Aizpurua, Carmen; Fernández Frutos, Begoña; De León, Patricia; Camacho, Maite; Fernández Moreira, Daniel; Ybarra, Carmen; Cabria Ramos, J. C.
The aim of this study was to develop a specific and highly sensitive method able to detect very low concentrations of Francisella tularensis in soil samples by real-time PCR (qPCR) with SYBR Green I. tul4gene, which encodes the 17-kDa protein (TUL4) in F. tularensis strains, was amplified using a LightCycler (LC) device. We achieved a detection limit of 0.69 fg of genomic DNA from F. tularensis subp. holarctica live vaccine strain (LVS), corresponding to a value less than 3.4 genome equivalents per reaction. The qPCR was shown to be specific, highly sensitive and reproducible. In addition, we evaluated 2 new methods for recovering bacteria from soil based on 1-step filtration using glass fiber filters and PVDF filters. These filtration methods enabled us to recover F. tularensis efficiently from soil samples. As few as 50 CFU per 0.5 g of soil were detected by qPCR. Capture enzyme-linked immunosorbent assay (cELISA) allowed us to detect and quantify the amount of bacteria recovered from soil by an immunological method. Although qPCR was more sensitive than cELISA, we did not observe substantial differences in the amount of bacteria quantified by both methods.