Comparative studies of Yersinia pestis outer membrane isolation techniques and their potential use in plague epidemiology

Abath, Frederico G. C.; Ferreira, Luís Carlos de Sousa

doi:10.1590/s0036-46651990000200002

Cited by 2 publications

References 19 publications

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Taxonomy of Yersinia pestis

Zhang

Cui

Zhang

2016

Advances in Experimental Medicine and Biology

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This chapter summarized the taxonomy and typing works of Yersinia pestis since it's firstly identified in Hong Kong in 1894. Phenotyping methods that based on phenotypic characteristics, including biotyping, serotyping, antibiogram analysis, bacteriocin typing, phage typing, and plasmid typing, were firstly applied in classification of Y. pestis in subspecies level. And then, with the advancement of molecular biological technology, the methods based on outer membrane protein profiles, fatty acid composition, and bacterial mass fingerprinting were also used to identify the populations within Y. pestis. However, Y. pestis is a highly homogenous species; therefore, the above typing methods could only provide low resolution, e.g., only one serotype and one phage type were observed for the whole species. Since the 1990s, molecular typing based on DNA variations, including single-nucleotide polymorphism, gene gain/loss, variable-number tandem repeats, clustered regularly interspaced short palindromic repeat, etc., was introduced and improved the resolution and robust of typing result. Especially in recent years, genotyping-based whole-genome-wide variations were successfully employed in Y. pestis, which built the "gold standard" of typing scheme of the species and could distinguish the samples under the strain level. The taxonomy and typing works leaved us enormous polymorphism data; therefore, a comprehensive fingerprint database of Y. pestis was needed to collect and standardize these data, for facilitating future works on evolution, plague surveillance and control, anti-bioterrorism, and microbial forensic researches.

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Taxonomy of Yersinia pestis

Zhang

Cui

Zhang

2016

Advances in Experimental Medicine and Biology

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A Surface-Focused Biotinylation Procedure Identifies the Yersinia pestis Catalase KatY as a Membrane-Associated but Non-Surface-Located Protein

Myers-Morales

Cowan

Gray

et al. 2007

Appl Environ Microbiol

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This study identified major surface proteins of the plague bacterium Yersinia pestis. We applied a novel surface biotinylation method, followed by NeutrAvidin (NA) bead capture, on-bead digestion, and identification by liquid chromatography-tandem mass spectrometry (LC-MS-MS). The use of stachyose during biotinylation focused the reaction to the surface. Coupled with NA pulldown and immunoblot analysis, this method determined whether a protein was accessible to the surface. We applied the method to test the hypothesis that the catalase KatY is a surface protein of the plague bacterium Y. pestis. A rabbit serum recognized the catalase KatY as a major putative outer membrane-associated antigen expressed by Y. pestis cells grown at 37°C. Similar findings by other groups had led to speculations that this protein might be exposed to the surface and might be a candidate for evaluation as a protective antigen for an improved plague vaccine. KatY was obtained only in the total membrane fraction, and stachyose greatly reduced its biotinylation as well as that of the periplasmic maltose binding protein, indicating that KatY is not on the bacterial surface. LC-MS-MS analysis of on-bead digests representing ca. 10 9 cells identified highly abundant species, including KatY, Pal, and OmpA, as well as the lipoprotein Pcp, all of which bound in a biotin-specific manner. Pla, Lpp, and OmpX (Ail) bound to the NA beads in a non-biotin-specific manner. There was no contamination from abundant cytoplasmic proteins. We hypothesize that OmpX and Pcp are highly abundant and likely to be important for the Y. pestis pathogenic process. We speculate that a portion of KatY associates with the outer membrane in intact cells but that it is located on the periplasmic side. Consistent with this idea, it did not protect C57BL/6 mice against bubonic plague.

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Comparative studies of Yersinia pestis outer membrane isolation techniques and their potential use in plague epidemiology

Cited by 2 publications

References 19 publications

Taxonomy of Yersinia pestis

Taxonomy of Yersinia pestis

A Surface-Focused Biotinylation Procedure Identifies the Yersinia pestis Catalase KatY as a Membrane-Associated but Non-Surface-Located Protein

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