Here, we report the whole-genome sequences of two ovine-pathogenic Corynebacterium pseudotuberculosis isolates: strain 3/99-5, which represents the first C. pseudotuberculosis genome originating from the United Kingdom, and 42/02-A, the second from Australia. These genome sequences will contribute to the objective of determining the global pan-genome of this bacterium.

Corynebacterium pseudotuberculosis causes disease in several animal species, although distinct biovars exist that appear to be restricted to specific hosts. In order to facilitate a better understanding of the differences between biovars, we report here the complete genome sequence of the equine pathogen Corynebacterium pseudotuberculosis strain 1/06-A.

Vibrio cholerae is the causal organism of the cholera epidemic, which is mostly prevalent in developing and underdeveloped countries. However, incidences of cholera in developed countries are also alarming. Because of the emergence of new drug-resistant strains, even though several generic drugs and vaccines have been developed over time, Vibrio infections remain a global health problem that appeals for the development of novel drugs and vaccines against the pathogen. Here, applying comparative proteomic and reverse vaccinology approaches to the exoproteome and secretome of the pathogen, we have identified three candidate targets (ompU, uppP and yajC) for most of the pathogenic Vibrio strains. Two targets (uppP and yajC) are novel to Vibrio, and two targets (uppP and ompU) can be used to develop both drugs and vaccines (dual targets) against broad spectrum Vibrio serotypes. Using our novel computational approach, we have identified three peptide vaccine candidates that have high potential to induce both B- and T-cell-mediated immune responses from our identified two dual targets. These two targets were modeled and subjected to virtual screening against natural compounds derived from Piper betel. Seven compounds were identified first time from Piper betel to be highly effective to render the function of these targets to identify them as emerging potential drugs against Vibrio. Our preliminary validation suggests that these identified peptide vaccines and betel compounds are highly effective against Vibrio cholerae. Currently we are exhaustively validating these targets, candidate peptide vaccines, and betel derived lead compounds against a number of Vibrio species.

Corynebacterium pseudotuberculosis is a facultative intracellular pathogen and the causative agent of several infectious and contagious chronic diseases, including caseous lymphadenitis, ulcerative lymphangitis, mastitis, and edematous skin disease, in a broad spectrum of hosts. In addition, Corynebacterium pseudotuberculosis infections pose a rising worldwide economic problem in ruminants. The complete genome sequences of 15 C. pseudotuberculosis strains isolated from different hosts and countries were comparatively analyzed using a pan-genomic strategy. Phylogenomic, pan-genomic, core genomic, and singleton analyses revealed close relationships among pathogenic corynebacteria, the clonal-like behavior of C. pseudotuberculosis and slow increases in the sizes of pan-genomes. According to extrapolations based on the pan-genomes, core genomes and singletons, the C. pseudotuberculosis biovar ovis shows a more clonal-like behavior than the C. pseudotuberculosis biovar equi. Most of the variable genes of the biovar ovis strains were acquired in a block through horizontal gene transfer and are highly conserved, whereas the biovar equi strains contain great variability, both intra- and inter-biovar, in the 16 detected pathogenicity islands (PAIs). With respect to the gene content of the PAIs, the most interesting finding is the high similarity of the pilus genes in the biovar ovis strains compared with the great variability of these genes in the biovar equi strains. Concluding, the polymerization of complete pilus structures in biovar ovis could be responsible for a remarkable ability of these strains to spread throughout host tissues and penetrate cells to live intracellularly, in contrast with the biovar equi, which rarely attacks visceral organs. Intracellularly, the biovar ovis strains are expected to have less contact with other organisms than the biovar equi strains, thereby explaining the significant clonal-like behavior of the biovar ovis strains.

In this work we report the genome of Corynebacterium pseudotuberculosis strain 267, isolated from a llama. This pathogen is of great veterinary and economic importance, as it is the cause of caseous lymphadenitis in several livestock species around the world and causes significant losses due to the high cost of treatment.

Background

The use of food-grade Lactic Acid Bacteria (LAB) as DNA delivery vehicles represents an attractive strategy to deliver DNA vaccines at the mucosal surfaces as they are generally regarded as safe (GRAS). We previously showed that either native Lactococcus lactis (LL) or recombinant invasive LL expressing Fibronectin Binding Protein A of Staphylococcus aureus (LL-FnBPA+) or Internalin A of Listeria monocytogenes (LL-InlA+), were able to deliver and trigger DNA expression by epithelial cells, either in vitro or in vivo. InlA does not bind to its receptor, the murine E-cadherin, thus limiting the use of LL-InlA+ in in vivo murine models. Moreover, FnBPA binds to its receptors, integrins, via fibronectin introducing another limiting factor. In order to avoid the limitations of LL-InlA+ and LL-FnBPA+, a new L. lactis strain was engineered to produce a previously described mutated form of InlA (LL-mInlA+) allowing the binding of mInlA on murine E-cadherin.

Results

After showing the expression of mInLA at the surface of LL-mInlA+ strain, in vitro gentamycin survival assay in Caco-2 cells showed that LL-mInlA+ is 1000 times more invasive than LL. LL-mInlA+ invasivity was also validated by fluorescence microscopy. LL and LL-mInlA+ were transformed with pValacBLG, a plasmid containing the cDNA of bovine β-Lactoglobulin (BLG), resulting in strains LL-BLG and LL-mInlA+BLG. The plasmid transfer in vitro using LL-mInlA+BLG was increased 10 times compared to LL-BLG. Moreover, the number of mice producing BLG in isolated enterocytes after oral administration of LL-mInlA+BLG in vivo was slightly higher than after oral administration of LL-BLG.

Conclusions

We confirmed in this study that the production of mInlA at the surface of L. lactis is a promising strategy for plasmid transfer in vitro and in vivo.

The Actinobacteria, Corynebacterium pseudotuberculosis strain P54B96, a nonmotile, non-sporulating and a mesophile bacterium, was isolated from liver, lung and mediastinal lymph node lesions in an antelope from South Africa. This strain is interesting in the sense that it has been found together with non-tuberculous mycobacteria (NTMs) which could nevertheless play a role in the lesion formation. In this work, we describe a set of features of C. pseudotuberculosis P54B96, together with the details of the complete genome sequence and annotation. The genome comprises of 2.34 Mbp long, single circular genome with 2,084 protein-coding genes, 12 rRNA, 49 tRNA and 62 pseudogenes and a G+C content of 52.19%. The analysis of the genome sequence provides means to better understanding the molecular and genetic basis of virulence of this bacterium, enabling a detailed investigation of its pathogenesis.

Corynebacterium diphtheriae is one of the most prominent human pathogens and the causative agent of the communicable disease diphtheria. The genomes of 12 strains isolated from patients with classical diphtheria, endocarditis, and pneumonia were completely sequenced and annotated. Including the genome of C. diphtheriae NCTC 13129, we herewith present a comprehensive comparative analysis of 13 strains and the first characterization of the pangenome of the species C. diphtheriae. Comparative genomics showed extensive synteny and revealed a core genome consisting of 1,632 conserved genes. The pangenome currently comprises 4,786 protein-coding regions and increases at an average of 65 unique genes per newly sequenced strain. Analysis of prophages carrying the diphtheria toxin gene tox revealed that the toxoid vaccine producer C. diphtheriae Park-Williams no. 8 has been lysogenized by two copies of the ωtox+ phage, whereas C. diphtheriae 31A harbors a hitherto-unknown tox+ corynephage. DNA binding sites of the tox-controlling regulator DtxR were detected by genome-wide motif searches. Comparative content analysis showed that the DtxR regulons exhibit marked differences due to gene gain, gene loss, partial gene deletion, and DtxR binding site depletion. Most predicted pathogenicity islands of C. diphtheriae revealed characteristics of horizontal gene transfer. The majority of these islands encode subunits of adhesive pili, which can play important roles in adhesion of C. diphtheriae to different host tissues. All sequenced isolates contain at least two pilus gene clusters. It appears that variation in the distributed genome is a common strategy of C. diphtheriae to establish differences in host-pathogen interactions.

Background

Pan-genomic studies aim, for instance, at defining the core, dispensable and unique genes within a species. A pan-genomics study for vaccine design tries to assess the best candidates for a vaccine against a specific pathogen. In this context, rather than studying genes predicted to be exported in a single genome, with pan-genomics it is possible to study genes present in different strains within the same species, such as virulence factors. The target organism of this pan-genomic work here presented is Corynebacterium pseudotuberculosis, the etiologic agent of caseous lymphadenitis (CLA) in goat and sheep, which causes significant economic losses in those herds around the world. Currently, only a few antigens against CLA are known as being the basis of commercial and still ineffective vaccines. In this regard, the here presented work analyses, in silico, five C. pseudotuberculosis genomes and gathers data to predict common exported proteins in all five genomes. These candidates were also compared to two recent C. pseudotuberculosis in vitro exoproteome results.

Results

The complete genome of five C. pseudotuberculosis strains (1002, C231, I19, FRC41 and PAT10) were submitted to pan-genomics analysis, yielding 306, 59 and 12 gene sets, respectively, representing the core, dispensable and unique in silico predicted exported pan-genomes. These sets bear 150 genes classified as secreted (SEC) and 227 as potentially surface exposed (PSE). Our findings suggest that the main C. pseudotuberculosis in vitro exoproteome could be greater, appended by a fraction of the 35 proteins formerly predicted as making part of the variant in vitro exoproteome. These genomes were manually curated for correct methionine initiation and redeposited with a total of 1885 homogenized genes.

Conclusions

The in silico prediction of exported proteins has allowed to define a list of putative vaccine candidate genes present in all five complete C. pseudotuberculosis genomes. Moreover, it has also been possible to define the in silico predicted dispensable and unique C. pseudotuberculosis exported proteins. These results provide in silico evidence to further guide experiments in the areas of vaccines, diagnosis and drugs. The work here presented is the first whole C. pseudotuberculosis in silico predicted pan-exoproteome completed till today.

Lactococci are noninvasive lactic acid bacteria frequently used as protein delivery vectors and, more recently, as DNA delivery vehicles. We previously showed that Lactococcus lactis (LL) expressing the Fibronectin-Binding Protein A of Staphylococcus aureus (LL-FnBPA+) showed higher internalization rates in vitro in Caco-2 cells than the native (wt) lactococci and were able to deliver a eukaryotic Green Fluorescent Protein (GFP) expression plasmid in 1% of human Caco-2 cells. Here, using the bovine beta-lactoglobulin (BLG), one of the major cow's milk allergen, and GFP we characterized the potential of LL-FnBPA+ as an in vivo DNA vaccine delivery vehicle. We first showed that the invasive strain LL-FnBPA+ carrying the plasmid pValac:BLG (LL-FnBPA+ BLG) was more invasive than LL-BLG and showed the same invasivity as LL-FnBPA+. Then we demonstrated that the Caco-2 cells, co-incubated with LL-FnBPA+ BLG produced up to 30 times more BLG than the Caco-2 cells co-incubated with the non invasive LL-BLG. Using two different gene reporters, BLG and GFP, and two different methods of detection, EIA and fluorescence microscopy, we showed in vivo that: i) in order to be effective, LL-FnBPA+ required a pre-coating with Fetal Calf Serum before oral administration; ii) plasmid transfer occurred in enterocytes without regard to the strains used (invasive or not); iii) the use of LL-FnBPA+ increased the number of mice producing BLG, but not the level of BLG produced. We thus confirmed the good potential of invasive recombinant lactic acid bacteria as DNA delivery vector in vivo.

The flatworm Schistosoma mansoni is a blood fluke parasite that causes schistosomiasis, a debilitating disease that occurs throughout the developing world. Current schistosomiasis control strategies are mainly based on chemotherapy, but many researchers believe that the best long-term strategy to control schistosomiasis is through immunization with an antischistosomiasis vaccine combined with drug treatment. In the search for potential vaccine candidates, numerous tegument antigens have been assessed. As the major interface between parasite and mammalian host, the tegument plays crucial roles in the establishment and further course of schistosomiasis. Herein, we evaluated the potential of a GPI fraction, containing representative molecules located on the outer surface of adult worms, as vaccine candidate. Immunization of mice with GPI-anchored proteins induced a mixed Th1/Th2 type of immune response with production of IFN-γ and TNF-α, and low levels of IL-5 into the supernatant of splenocyte cultures. The protection engendered by this vaccination protocol was confirmed by 42% reduction in worm burden, 45% reduction in eggs per gram of hepatic tissue, 29% reduction in the number of granulomas per area, and 53% reduction in the granuloma fibrosis. Taken together, the data herein support the potential of surface-exposed GPI-anchored antigens from the S. mansoni tegument as vaccine candidate.

The enrichment analysis is a standard procedure to interpret ‘omics’ experiments that generate large gene lists as outputs, such as transcriptomics and protemics. However, despite the huge success of enrichment analysis in these classes of experiments, there is a surprising lack of application of this methodology to survey other categories of large-scale biological data available. Here, we report Kegg Orthology enrichMent-Online DetectiOn (KOMODO), a web tool to systematically investigate groups of monophyletic genomes in order to detect significantly enriched groups of homologous genes in one taxon when compared with another. The results are displayed in their proper biochemical roles in a visual, explorative way, allowing users to easily formulate and investigate biological hypotheses regarding the taxonomical distribution of genomic elements. We validated KOMODO by analyzing portions of central carbon metabolism in two taxa extensively studied regarding their carbon metabolism profile (Enterobacteriaceae family and Lactobacillales order). Most enzymatic activities significantly biased were related to known key metabolic traits in these taxa, such as the distinct fates of pyruvate (the known tendency of lactate production in Lactobacillales and its complete oxidation in Enterobacteriaceae), demonstrating that KOMODO could detect biologically meaningful differences in the frequencies of shared genomic elements among taxa. KOMODO is freely available at http://komodotool.org.

In this work, we report the whole-genome sequence of Corynebacterium pseudotuberculosis bv. equi strain CIP 52.97 (Collection Institut Pasteur), isolated in 1952 from a case of ulcerative lymphangitis in a Kenyan horse, which has evidently caused significant losses to agribusiness. Therefore, obtaining this genome will allow the detection of important targets for postgenomic studies, with the aim of minimizing problems caused by this microorganism.

In this work, we report the complete genome sequence of a Corynebacterium pseudotuberculosis PAT10 isolate, collected from a lung abscess in an Argentine sheep in Patagonia, whose pathogen also required an investigation of its pathogenesis. Thus, the analysis of the genome sequence offers a means to better understanding of the molecular and genetic basis of virulence of this bacterium.

Pathogenic intracellular bacteria can respond to antimicrobial mechanisms of the host cell through transient activation of stress-responsive genes by alternative sigma (σ) factors of the RNA polymerase. We evaluated the contribution of the extracytoplasmic function sigma factor σE for Corynebacterium pseudotuberculosis resistance to stress conditions resembling those found intracellularly during infection. A sigE-null mutant strain (ΔsigE) of this bacterium was more susceptible in vitro to acidic pH, cell surface stressors, and biologically relevant concentrations of nitric oxide (NO). The same mutant strain was unable to persist in C57BL/6 mice but remained infective in mice lacking inducible nitric oxide synthase (iNOS), confirming the significance of σE for resistance to nitric oxide/peroxide stress in vivo. High-throughput proteomic analysis identified NO-responsive extracellular proteins of C. pseudotuberculosis and demonstrated the participation of σE in composition of this bacterium’s exoproteome.

Improvements in genome sequencing techniques have resulted in generation of huge volumes of data. As a consequence of this progress, the genome assembly stage demands even more computational power, since the incoming sequence files contain large amounts of data. To speed up the process, it is often necessary to distribute the workload among a group of machines. However, this requires hardware and software solutions specially configured for this purpose. Grid computing try to simplify this process of aggregate resources, but do not always offer the best performance possible due to heterogeneity and decentralized management of its resources. Thus, it is necessary to develop software that takes into account these peculiarities. In order to achieve this purpose, we developed an algorithm aimed to optimize the functionality of de novo assembly software ABySS in order to optimize its operation in grids. We run ABySS with and without the algorithm we developed in the grid simulator SimGrid. Tests showed that our algorithm is viable, flexible, and scalable even on a heterogeneous environment, which improved the genome assembly time in computational grids without changing its quality.

The adaptability of pathogenic bacteria to hosts is influenced by the genomic plasticity of the bacteria, which can be increased by such mechanisms as horizontal gene transfer. Pathogenicity islands play a major role in this type of gene transfer because they are large, horizontally acquired regions that harbor clusters of virulence genes that mediate the adhesion, colonization, invasion, immune system evasion, and toxigenic properties of the acceptor organism. Currently, pathogenicity islands are mainly identified in silico based on various characteristic features: (1) deviations in codon usage, G+C content or dinucleotide frequency and (2) insertion sequences and/or tRNA genetic flanking regions together with transposase coding genes. Several computational techniques for identifying pathogenicity islands exist. However, most of these techniques are only directed at the detection of horizontally transferred genes and/or the absence of certain genomic regions of the pathogenic bacterium in closely related non-pathogenic species. Here, we present a novel software suite designed for the prediction of pathogenicity islands (pathogenicity island prediction software, or PIPS). In contrast to other existing tools, our approach is capable of utilizing multiple features for pathogenicity island detection in an integrative manner. We show that PIPS provides better accuracy than other available software packages. As an example, we used PIPS to study the veterinary pathogen Corynebacterium pseudotuberculosis, in which we identified seven putative pathogenicity islands.

Modern genomic sequencing technologies produce a large amount of data with reduced cost per base; however, this data consists of short reads. This reduction in the size of the reads, compared to those obtained with previous methodologies, presents new challenges, including a need for efficient algorithms for the assembly of genomes from short reads and for resolving repetitions. Additionally after abinitio assembly, curation of the hundreds or thousands of contigs generated by assemblers demands considerable time and computational resources. We developed Simplifier, a stand-alone software that selectively eliminates redundant sequences from the collection of contigs generated by ab initio assembly of genomes. Application of Simplifier to data generated by assembly of the genome of Corynebacterium pseudotuberculosis strain 258 reduced the number of contigs generated by ab initio methods from 8,004 to 5,272, a reduction of 34.14%; in addition, N50 increased from 1 kb to 1.5 kb. Processing the contigs of Escherichia coli DH10B with Simplifier reduced the mate-paired library 17.47% and the fragment library 23.91%. Simplifier removed redundant sequences from datasets produced by assemblers, thereby reducing the effort required for finalization of genome assembly in tests with data from Prokaryotic organisms.

Availability

Simplifier is available at http://www.genoma.ufpa.br/rramos/softwares/simplifier.xhtmlIt requires Sun jdk 6 or higher.

Background

Singular value decomposition (SVD) is a powerful technique for information retrieval; it helps uncover relationships between elements that are not prima facie related. SVD was initially developed to reduce the time needed for information retrieval and analysis of very large data sets in the complex internet environment. Since information retrieval from large-scale genome and proteome data sets has a similar level of complexity, SVD-based methods could also facilitate data analysis in this research area.

Results

We found that SVD applied to amino acid sequences demonstrates relationships and provides a basis for producing clusters and cladograms, demonstrating evolutionary relatedness of species that correlates well with Linnaean taxonomy. The choice of a reasonable number of singular values is crucial for SVD-based studies. We found that fewer singular values are needed to produce biologically significant clusters when SVD is employed. Subsequently, we developed a method to determine the lowest number of singular values and fewest clusters needed to guarantee biological significance; this system was developed and validated by comparison with Linnaean taxonomic classification.

Conclusions

By using SVD, we can reduce uncertainty concerning the appropriate rank value necessary to perform accurate information retrieval analyses. In tests, clusters that we developed with SVD perfectly matched what was expected based on Linnaean taxonomy.

Brucella abortus is a facultative intracellular bacterial pathogen that causes abortion in domestic animals and undulant fever in humans. IFN-γ, IL-12, and CD8+ T lymphocytes are important components of host immune responses against B. abortus. Herein, IFN-γ and IL-12/β2-microglobulin (β2-m) knockout mice were used to determine whether CD8+ T cells and IL-12-dependent IFN-γ deficiency would be more critical to control B. abortus infection compared to the lack of endogenous IFN-γ. At 1 week after infection, IFN-γ KO and IL-12/β2-m KO mice showed increased numbers of bacterial load in spleens; however, at 3 weeks postinfection (p.i.), only IFN-γ KO succumbed to Brucella. All IFN-γ KO had died at 16 days p.i. whereas death within the IL-12/β2-m KO group was delayed and occurred at 32 days until 47 days postinfection. Susceptibility of IL-12/β2-m KO animals to Brucella was associated to undetectable levels of IFN-γ in mouse splenocytes and inability of these cells to lyse Brucella-infected macrophages. However, the lack of endogenous IFN-γ was found to be more important to control brucellosis than CD8+ T cells and IL-12-dependent IFN-γ deficiencies.

Post-genomic analysis techniques such as next-generation sequencing have produced vast amounts of data about micro organisms including genetic sequences, their functional annotations and gene regulatory interactions. The latter are genetic mechanisms that control a cell's characteristics, for instance, pathogenicity as well as survival and reproduction strategies. CoryneRegNet is the reference database and analysis platform for corynebacterial gene regulatory networks. In this article we introduce the updated version 6.0 of CoryneRegNet and describe the updated database content which includes, 6352 corynebacterial regulatory interactions compared with 4928 interactions in release 5.0 and 3235 regulations in release 4.0, respectively. We also demonstrate how we support the community by integrating analysis and visualization features for transiently imported custom data, such as gene regulatory interactions. Furthermore, with release 6.0, we provide easy-to-use functions that allow the user to submit data for persistent storage with the CoryneRegNet database. Thus, it offers important options to its users in terms of community demands. CoryneRegNet is publicly available at http://www.coryneregnet.de.

Background

S. aureus is one of the main pathogens involved in ruminant mastitis worldwide. The severity of staphylococcal infection is highly variable, ranging from subclinical to gangrenous mastitis. This work represents an in-depth characterization of S. aureus mastitis isolates to identify bacterial factors involved in severity of mastitis infection.

Methodology/Principal Findings

We employed genomic, transcriptomic and proteomic approaches to comprehensively compare two clonally related S. aureus strains that reproducibly induce severe (strain O11) and milder (strain O46) mastitis in ewes. Variation in the content of mobile genetic elements, iron acquisition and metabolism, transcriptional regulation and exoprotein production was observed. In particular, O11 produced relatively high levels of exoproteins, including toxins and proteases known to be important in virulence. A characteristic we observed in other S. aureus strains isolated from clinical mastitis cases.

Conclusions/Significance

Our data are consistent with a dose-dependant role of some staphylococcal factors in the hypervirulence of strains isolated from severe mastitis. Mobile genetic elements, transcriptional regulators, exoproteins and iron acquisition pathways constitute good targets for further research to define the underlying mechanisms of mastitis severity.

Background

Caseous lymphadenitis (CLA), caused by Corynebacterium pseudotuberculosis, is one of the most important diseases of sheep and goats, causing considerable economic losses for herd owners.

Results

We assessed the seroprevalence of infection with C. pseudotuberculosis in 805 sheep from 23 sheep farms that supply slaughterhouses in the state of Minas Gerais; we also analyzed management practices that could be associated with CLA occurrence, used on these and nearby farms that also supplied animals to the slaughterhouse (n = 60). The serum samples for assaying CLA infection were taken at the slaughterhouse. Frequency of infection with C. pseudotuberculosis was estimated at 43.7%, and farm frequency was estimated at 100%. Management practices were analyzed through a questionnaire. All farmers (60/60) had extensive/semi-extensive rearing system; 70.0% (42/60) identified sheep individually; 11.7% (7/60) had periodical technical assistance; 41.7% (25/60) disinfected the facilities; 86.7% (52/60) used barbed wire fences and did not implement adequate CLA control measures; only 11.7% (7/60) of breeders reported vaccination against C. pseudotuberculosis; 13.3% (8/60) took note of animals with clinical signs of CLA; 1.7% (1/60) opened and sanitized abscesses, and isolated the infected animals; 10.0% (6/60) knew the zoonotic potential of this disease and 1.7% (1/60) of the farmers culled animals in case of recurrence of abscesses.

Conclusions

It can be concluded that C. pseudotuberculosis infection is widely spread in sheep flocks in Minas Gerais state in Brazil and that there is a lack of good management measures and vaccination, allowing transmission of this infectious agent throughout the production network.

The flatworm Schistosoma mansoni is a blood fluke parasite that causes schistosomiasis, a debilitating disease that occurs throughout the developing world. Current schistosomiasis control strategies are mainly based on chemotherapy, but many researchers believe that the best long-term strategy to control schistosomiasis is through immunization with an antischistosomiasis vaccine combined with drug treatment. Several papers on Schistosoma mansoni vaccine and drug development have been published in the past few years, representing an important field of study. The advent of technologies that allow large-scale studies of genes and proteins had a remarkable impact on the screening of new and potential vaccine candidates in schistosomiasis. In this postgenomic scenario, bioinformatic technologies have emerged as important tools to mine transcriptomic, genomic, and proteomic databases. These new perspectives are leading to a new round of rational vaccine development. Herein, we discuss different strategies to identify potential S. mansoni vaccine candidates using computational vaccinology.

Background

Corynebacterium ulcerans has been detected as a commensal in domestic and wild animals that may serve as reservoirs for zoonotic infections. During the last decade, the frequency and severity of human infections associated with C. ulcerans appear to be increasing in various countries. As the knowledge of genes contributing to the virulence of this bacterium was very limited, the complete genome sequences of two C. ulcerans strains detected in the metropolitan area of Rio de Janeiro were determined and characterized by comparative genomics: C. ulcerans 809 was initially isolated from an elderly woman with fatal pulmonary infection and C. ulcerans BR-AD22 was recovered from a nasal sample of an asymptomatic dog.

Results

The circular chromosome of C. ulcerans 809 has a total size of 2,502,095 bp and encodes 2,182 predicted proteins, whereas the genome of C. ulcerans BR-AD22 is 104,279 bp larger and comprises 2,338 protein-coding regions. The minor difference in size of the two genomes is mainly caused by additional prophage-like elements in the C. ulcerans BR-AD22 chromosome. Both genomes show a highly similar order of orthologous coding regions; and both strains share a common set of 2,076 genes, demonstrating their very close relationship. A screening for prominent virulence factors revealed the presence of phospholipase D (Pld), neuraminidase H (NanH), endoglycosidase E (EndoE), and subunits of adhesive pili of the SpaDEF type that are encoded in both C. ulcerans genomes. The rbp gene coding for a putative ribosome-binding protein with striking structural similarity to Shiga-like toxins was additionally detected in the genome of the human isolate C. ulcerans 809.

Conclusions

The molecular data deduced from the complete genome sequences provides considerable knowledge of virulence factors in C. ulcerans that is increasingly recognized as an emerging pathogen. This bacterium is apparently equipped with a broad and varying set of virulence factors, including a novel type of a ribosome-binding protein. Whether the respective protein contributes to the severity of human infections (and a fatal outcome) remains to be elucidated by genetic experiments with defined bacterial mutants and host model systems.