Showing papers by "Luis G.C. Pacheco published in 2011"

Evidence for reductive genome evolution and lateral acquisition of virulence functions in two Corynebacterium pseudotuberculosis strains.

Abstract: Background: Corynebacterium pseudotuberculosis, a Gram-positive, facultative intracellular pathogen, is the etiologic agent of the disease known as caseous lymphadenitis (CL). CL mainly affects small ruminants, such as goats and sheep; it also causes infections in humans, though rarely. This species is distributed worldwide, but it has the most serious economic impact in Oceania, Africa and South America. Although C. pseudotuberculosis causes major health and productivity problems for livestock, little is known about the molecular basis of its pathogenicity. Methodology and Findings: We characterized two C. pseudotuberculosis genomes (Cp1002, isolated from goats; and CpC231, isolated from sheep). Analysis of the predicted genomes showed high similarity in genomic architecture, gene content and genetic order. When C. pseudotuberculosis was compared with other Corynebacterium species, it became evident that this pathogenic species has lost numerous genes, resulting in one of the smallest genomes in the genus. Other differences that could be part of the adaptation to pathogenicity include a lower GC content, of about 52%, and a reduced gene repertoire. The C. pseudotuberculosis genome also includes seven putative pathogenicity islands, which contain several classical virulence factors, including genes for fimbrial subunits, adhesion factors, iron uptake and secreted toxins. Additionally, all of the virulence factors in the islands have characteristics that indicate horizontal transfer. Conclusions: These particular genome characteristics of C. pseudotuberculosis, as well as its acquired virulence factors in pathogenicity islands, provide evidence of its lifestyle and of the pathogenicity pathways used by this pathogen in the infection process. All genomes cited in this study are available in the NCBI Genbank database (http://www.ncbi.nlm.nih.gov/ genbank/) under accession numbers CP001809 and CP001829.

A combined approach for comparative exoproteome analysis of Corynebacterium pseudotuberculosis

Abstract: Bacterial exported proteins represent key components of the host-pathogen interplay. Hence, we sought to implement a combined approach for characterizing the entire exoproteome of the pathogenic bacterium Corynebacterium pseudotuberculosis, the etiological agent of caseous lymphadenitis (CLA) in sheep and goats. An optimized protocol of three-phase partitioning (TPP) was used to obtain the C. pseudotuberculosis exoproteins, and a newly introduced method of data-independent MS acquisition (LC-MSE) was employed for protein identification and label-free quantification. Additionally, the recently developed tool SurfG+ was used for in silico prediction of sub-cellular localization of the identified proteins. In total, 93 different extracellular proteins of C. pseudotuberculosis were identified with high confidence by this strategy; 44 proteins were commonly identified in two different strains, isolated from distinct hosts, then composing a core C. pseudotuberculosis exoproteome. Analysis with the SurfG+ tool showed that more than 75% (70/93) of the identified proteins could be predicted as containing signals for active exportation. Moreover, evidence could be found for probable non-classical export of most of the remaining proteins. Comparative analyses of the exoproteomes of two C. pseudotuberculosis strains, in addition to comparison with other experimentally determined corynebacterial exoproteomes, were helpful to gain novel insights into the contribution of the exported proteins in the virulence of this bacterium. The results presented here compose the most comprehensive coverage of the exoproteome of a corynebacterial species so far.

Similarity of rpoB gene sequences of sucrose-fermenting and non-fermenting Corynebacterium diphtheriae strains

Abstract: During the last decades, the majority of Brazilian Corynebacterium diphtheriae isolates were shown to be capable to metabolize sucrose, sometimes leading to erroneous identification as a non-diphtheric Corynebacterium species. The sequencing of the polymorphic region of the RNA polymerase beta subunit-encoding gene (rpoB) is an important taxonomic tool for identification of corynebacteria. The present study aimed to investigate the rpoB gene polymorphic features of sucrose-fermenting and non sucrose-fermenting strains. The results showed that sucrose-fermenting strains presented rpoB gene polymorphic regions with more than 98% similarity with the sequences deposited in the gene bank corresponding to non sucrose-fermenting strains. Data indicate that sucrose-fermenting isolates may act as a variant of C. diphtheriae biotype mitis. In addition we alert that sucrose-fermenting strains should not be discarded as contaminants mainly in countries where the possibility of isolation of this variant is higher.

Serological secretome analysis of Corynebacterium pseudotuberculosis

Abstract: Caseous lymphadenitis (CLA) is a chronic disease affecting small ruminants caused by Corynebacterium pseudotuberculosis and is responsible for significant economic losses. Previous studies have reported that a set of C. pseudotuberculosis secreted proteins react with sera from infected goats. Mapping of the secretome will shed light on the pathogenesis of CLA. We have hereby identified six immunoreactive secreted proteins of C. pseudotuberculosis by 2D Western blotting using sera from goats with CLA and characterized them by mass spectrometry. The information retrieved will support studies leading to the development of efficient vaccines and diagnostic kits.