Publications by authors named "Marirosa Mora"

26 Publications

  • Page 1 of 1

The Human Pathogen Streptococcus pyogenes Releases Lipoproteins as Lipoprotein-rich Membrane Vesicles.

Mol Cell Proteomics 2015 Aug 27;14(8):2138-49. Epub 2015 May 27.

From the ‡Novartis Vaccines and Diagnostics (a GSK company), Via Fiorentiina 1, 53100 Siena, Itlay;

Bacterial lipoproteins are attractive vaccine candidates because they represent a major class of cell surface-exposed proteins in many bacteria and are considered as potential pathogen-associated molecular patterns sensed by Toll-like receptors with built-in adjuvanticity. Although Gram-negative lipoproteins have been extensively characterized, little is known about Gram-positive lipoproteins. We isolated from Streptococcus pyogenes a large amount of lipoproteins organized in vesicles. These vesicles were obtained by weakening the bacterial cell wall with a sublethal concentration of penicillin. Lipid and proteomic analysis of the vesicles revealed that they were enriched in phosphatidylglycerol and almost exclusively composed of lipoproteins. In association with lipoproteins, a few hypothetical proteins, penicillin-binding proteins, and several members of the ExPortal, a membrane microdomain responsible for the maturation of secreted proteins, were identified. The typical lipidic moiety was apparently not necessary for lipoprotein insertion in the vesicle bilayer because they were also recovered from the isogenic diacylglyceryl transferase deletion mutant. The vesicles were not able to activate specific Toll-like receptor 2, indicating that lipoproteins organized in these vesicular structures do not act as pathogen-associated molecular patterns. In light of these findings, we propose to name these new structures Lipoprotein-rich Membrane Vesicles.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1074/mcp.M114.045880DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4528243PMC
August 2015

SslE elicits functional antibodies that impair in vitro mucinase activity and in vivo colonization by both intestinal and extraintestinal Escherichia coli strains.

PLoS Pathog 2014 May 8;10(5):e1004124. Epub 2014 May 8.

Novartis Vaccines and Diagnostics Srl, Siena, Italy.

SslE, the Secreted and surface-associated lipoprotein from Escherichia coli, has recently been associated to the M60-like extracellular zinc-metalloprotease sub-family which is implicated in glycan recognition and processing. SslE can be divided into two main variants and we recently proposed it as a potential vaccine candidate. By applying a number of in vitro bioassays and comparing wild type, knockout mutant and complemented strains, we have now demonstrated that SslE specifically contributes to degradation of mucin substrates, typically present in the intestine and bladder. Mutation of the zinc metallopeptidase motif of SslE dramatically impaired E. coli mucinase activity, confirming the specificity of the phenotype observed. Moreover, antibodies raised against variant I SslE, cloned from strain IHE3034 (SslEIHE3034), are able to inhibit translocation of E. coli strains expressing different variants through a mucin-based matrix, suggesting that SslE induces cross-reactive functional antibodies that affect the metallopeptidase activity. To test this hypothesis, we used well-established animal models and demonstrated that immunization with SslEIHE3034 significantly reduced gut, kidney and spleen colonization by strains producing variant II SslE and belonging to different pathotypes. Taken together, these data strongly support the importance of SslE in E. coli colonization of mucosal surfaces and reinforce the use of this antigen as a component of a broadly protective vaccine against pathogenic E. coli species.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1371/journal.ppat.1004124DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4014459PMC
May 2014

Neisseria adhesin A variation and revised nomenclature scheme.

Clin Vaccine Immunol 2014 Jul 7;21(7):966-71. Epub 2014 May 7.

Department of Zoology, University of Oxford, Oxford, United Kingdom.

Neisseria adhesin A (NadA), involved in the adhesion and invasion of Neisseria meningitidis into host tissues, is one of the major components of Bexsero, a novel multicomponent vaccine licensed for protection against meningococcal serogroup B in Europe, Australia, and Canada. NadA has been identified in approximately 30% of clinical isolates and in a much lower proportion of carrier isolates. Three protein variants were originally identified in invasive meningococci and named NadA-1, NadA-2, and NadA-3, whereas most carrier isolates either lacked the gene or harbored a different variant, NadA-4. Further analysis of isolates belonging to the sequence type 213 (ST-213) clonal complex identified NadA-5, which was structurally similar to NadA-4, but more distantly related to NadA-1, -2, and -3. At the time of this writing, more than 89 distinct nadA allele sequences and 43 distinct peptides have been described. Here, we present a revised nomenclature system, taking into account the complete data set, which is compatible with previous classification schemes and is expandable. The main features of this new scheme include (i) the grouping of the previously named NadA-2 and NadA-3 variants into a single NadA-2/3 variant, (ii) the grouping of the previously assigned NadA-4 and NadA-5 variants into a single NadA-4/5 variant, (iii) the introduction of an additional variant (NadA-6), and (iv) the classification of the variants into two main groups, named groups I and II. To facilitate querying of the sequences and submission of new allele sequences, the nucleotide and amino acid sequences are available at http://pubmlst.org/neisseria/NadA/.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1128/CVI.00825-13DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4097447PMC
July 2014

SpyAD, a moonlighting protein of group A Streptococcus contributing to bacterial division and host cell adhesion.

Infect Immun 2014 Jul 28;82(7):2890-901. Epub 2014 Apr 28.

Novartis Vaccines and Diagnostics Srl, Siena, Italy.

Group A streptococcus (GAS) is a human pathogen causing a wide repertoire of mild and severe diseases for which no vaccine is yet available. We recently reported the identification of three protein antigens that in combination conferred wide protection against GAS infection in mice. Here we focused our attention on the characterization of one of these three antigens, Spy0269, a highly conserved, surface-exposed, and immunogenic protein of unknown function. Deletion of the spy0269 gene in a GAS M1 isolate resulted in very long bacterial chains, which is indicative of an impaired capacity of the knockout mutant to properly divide. Confocal microscopy and immunoprecipitation experiments demonstrated that the protein was mainly localized at the cell septum and could interact in vitro with the cell division protein FtsZ, leading us to hypothesize that Spy0269 is a member of the GAS divisome machinery. Predicted structural domains and sequence homologies with known streptococcal adhesins suggested that this antigen could also play a role in mediating GAS interaction with host cells. This hypothesis was confirmed by showing that recombinant Spy0269 could bind to mammalian epithelial cells in vitro and that Lactococcus lactis expressing Spy0269 on its cell surface could adhere to mammalian cells in vitro and to mice nasal mucosa in vivo. On the basis of these data, we believe that Spy0269 is involved both in bacterial cell division and in adhesion to host cells and we propose to rename this multifunctional moonlighting protein as SpyAD (Streptococcus pyogenes Adhesion and Division protein).
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1128/IAI.00064-14DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4097626PMC
July 2014

Conservation of meningococcal antigens in the genus Neisseria.

mBio 2013 Jun 11;4(3):e00163-13. Epub 2013 Jun 11.

Novartis Vaccines and Diagnostics, Siena, Italy.

Neisseria meningitidis, one of the major causes of bacterial meningitis and sepsis, is a member of the genus Neisseria, which includes species that colonize the mucosae of many animals. Three meningococcal proteins, factor H-binding protein (fHbp), neisserial heparin-binding antigen (NHBA), and N. meningitidis adhesin A (NadA), have been described as antigens protective against N. meningitidis of serogroup B, and they have been employed as vaccine components in preclinical and clinical studies. In the vaccine formulation, fHbp and NHBA were fused to the GNA2091 and GNA1030 proteins, respectively, to enhance protein stability and immunogenicity. To determine the possible impact of vaccination on commensal neisseriae, we determined the presence, distribution, and conservation of these antigens in the available genome sequences of the genus Neisseria, finding that fHbp, NHBA, and NadA were conserved only in species colonizing humans, while GNA1030 and GNA2091 were conserved in many human and nonhuman neisseriae. Sequence analysis showed that homologous recombination contributed to shape the evolution and distribution of both NHBA and fHbp, three major variants of which have been defined. fHbp variant 3 was probably the ancestral form of meningococcal fHbp, while fHbp variant 1 from N. cinerea was introduced into N. meningitidis by a recombination event. fHbp variant 2 was the result of a recombination event inserting a stretch of 483 bp from variant 1 into the variant 3 background. These data indicate that a high rate of exchange of genetic material between neisseriae that colonize the human upper respiratory tract exists. IMPORTANCE The upper respiratory tract of healthy individuals is a complex ecosystem colonized by many bacterial species. Among these, there are representatives of the genus Neisseria, including Neisseria meningitidis, a major cause of bacterial meningitis and sepsis. Given the close relationship between commensal and pathogenic species, a protein-based vaccine against N. meningitidis has the potential to impact the other commensal species of Neisseria. For this reason, we have studied the distribution and evolutionary history of the antigen components of a recombinant vaccine, 4CMenB, that recently received approval in Europe under the commercial name of Bexsero®. We found that fHbp, NHBA, and NadA can be found in some of the human commensal species and that the evolution of these antigens has been essentially shaped by the high rate of genetic exchange that occurs between strains of neisseriae that cocolonize the same environment.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1128/mBio.00163-13DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3685207PMC
June 2013

Multi high-throughput approach for highly selective identification of vaccine candidates: the Group A Streptococcus case.

Mol Cell Proteomics 2012 Jun 27;11(6):M111.015693. Epub 2012 Jan 27.

Novartis Vaccines and Diagnostics, Via Fiorentina 1, 53100 Siena, Italy.

We propose an experimental strategy for highly accurate selection of candidates for bacterial vaccines without using in vitro and/or in vivo protection assays. Starting from the observation that efficacious vaccines are constituted by conserved, surface-associated and/or secreted components, the strategy contemplates the parallel application of three high throughput technologies, i.e. mass spectrometry-based proteomics, protein array, and flow-cytometry analysis, to identify this category of proteins, and is based on the assumption that the antigens identified by all three technologies are the protective ones. When we tested this strategy for Group A Streptococcus, we selected a total of 40 proteins, of which only six identified by all three approaches. When the 40 proteins were tested in a mouse model, only six were found to be protective and five of these belonged to the group of antigens in common to the three technologies. Finally, a combination of three protective antigens conferred broad protection against a panel of four different Group A Streptococcus strains. This approach may find general application as an accelerated and highly accurate path to bacterial vaccine discovery.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1074/mcp.M111.015693DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3433891PMC
June 2012

Genome-based vaccine development: a short cut for the future.

Hum Vaccin 2008 May-Jun;4(3):184-8. Epub 2010 May 15.

Novartis Vaccines, Siena, Italy.

Bacterial infectious diseases remain a major cause of deaths and disabilities in the world. Although conventional vaccinology approaches were successful in conferring protection against several diseases, they failed in providing efficient vaccines against many others. Together to the sequencing of the first genome, a new chapter in the vaccinology history started to be written. Reverse vaccinology changed the way to think about vaccine development, using the information provided by the microorganisms' genome against themselves. Since then, reverse vaccinology has evolved and helped researchers to overcome the limits of the conventional vaccinology approaches and led to the discovery and development of novel vaccines concerning emerging diseases, like Neisseria meningitidis B and Streptococcus agalactiae. A lot of work must be done, but deciphering the information provided by genome sequences and using it to better understand the host-pathogen interactions has proved to be the key for protection.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.4161/hv.4.3.6313DOI Listing
November 2010

Genome-based approaches to vaccine development.

J Mol Med (Berl) 2010 Feb;88(2):143-7

Novartis Vaccines, Via Fiorentina 1, 53100 Siena, Italy.

The hundreds of bacterial genome sequences available together with advances in bioinformatics and the development of new experimental proteomic tools are revolutionizing the vaccinology field. The merge of stringent in silico criteria and different experimental approaches is allowing a more targeted strategy to obtain a restricted and prioritized list of potential antigens for testing in immunogenicity assays, reducing the time and the cost of novel protein vaccine formulations.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1007/s00109-009-0574-9DOI Listing
February 2010

Genome-based vaccine development: a short cut for the future.

Adv Exp Med Biol 2009 ;655:81-9

Novartis Vaccines, Via Fiorentina 1, 53100, Siena, Italy.

Bacterial infectious diseases remain a major cause of deaths and disabilities in the world. Although conventional vaccinology approaches were successful in conferring protection against several diseases, they failed in providing efficient vaccines against many others. Together to the sequencing of the first genome, a new chapter in the vaccinology history started to be written. Reverse vaccinology changed the way to think about vaccine development, using the information provided by the microorganisms' genome against themselves. Since then, reverse vaccinology has evolved and helped researchers to overcome the limits of the conventional vaccinology approaches and led to the discovery and development of novel vaccines concerning emerging diseases, like Neisseria meningitidis B and Streptococcus agalactiae. A lot of work must be done, but deciphering the information provided by genome sequences and using it to better understand the host-pathogen interactions has proved to be the key for protection.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1007/978-1-4419-1132-2_8DOI Listing
August 2010

Protein array profiling of tic patient sera reveals a broad range and enhanced immune response against Group A Streptococcus antigens.

PLoS One 2009 Jul 22;4(7):e6332. Epub 2009 Jul 22.

Novartis Vaccines and Diagnostics, Siena, Italy.

The human pathogen Group A Streptococcus (Streptococcus pyogenes, GAS) is widely recognized as a major cause of common pharyngitis as well as of severe invasive diseases and non-suppurative sequelae associated with the existence of GAS antigens eliciting host autoantibodies. It has been proposed that a subset of paediatric disorders characterized by tics and obsessive-compulsive symptoms would exacerbate in association with relapses of GAS-associated pharyngitis. This hypothesis is however still controversial. In the attempt to shed light on the contribution of GAS infections to the onset of neuropsychiatric or behavioral disorders affecting as many as 3% of children and adolescents, we tested the antibody response of tic patient sera to a representative panel of GAS antigens. In particular, 102 recombinant proteins were spotted on nitrocellulose-coated glass slides and probed against 61 sera collected from young patients with typical tic neuropsychiatric symptoms but with no overt GAS infection. Sera from 35 children with neither tic disorder nor overt GAS infection were also analyzed. The protein recognition patterns of these two sera groups were compared with those obtained using 239 sera from children with GAS-associated pharyngitis. This comparative analysis identified 25 antigens recognized by sera of the three patient groups and 21 antigens recognized by tic and pharyngitis sera, but poorly or not recognized by sera from children without tic. Interestingly, these antigens appeared to be, in quantitative terms, more immunogenic in tic than in pharyngitis patients. Additionally, a third group of antigens appeared to be preferentially and specifically recognized by tic sera. These findings provide the first evidence that tic patient sera exhibit immunological profiles typical of individuals who elicited a broad, specific and strong immune response against GAS. This may be relevant in the context of one of the hypothesis proposing that GAS antigen-dependent induction of autoantibodies in susceptible individuals may be involved the occurrence of tic disorders.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0006332PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2709431PMC
July 2009

Surfome analysis as a fast track to vaccine discovery: identification of a novel protective antigen for Group B Streptococcus hypervirulent strain COH1.

Mol Cell Proteomics 2009 Jul 28;8(7):1728-37. Epub 2009 Apr 28.

Research Centre, Novartis Vaccines and Diagnostics, 53100 Siena, Italy.

Safe recombinant vaccines, based on a small number of antigenic proteins, are emerging as the most attractive, cost-effective solution against infectious diseases. In the present work, we confirmed previous data from our laboratory showing that whole viable bacterial cell treatment with proteases followed by the identification of released peptides by mass spectrometry is the method of choice for the rapid and reliable identification of vaccine candidates in Gram-positive bacteria. When applied to the Group B Streptococcus COH1 strain, 43 surface-associated proteins were identified, including all the protective antigens described in the literature as well as a new protective antigen, the cell wall-anchored protein SAN_1485 belonging to the serine-rich repeat protein family. This strategy overcomes the difficulties so far encountered in the identification of novel vaccine candidates and speeds up the entire vaccine discovery process by reducing the number of recombinant proteins to be tested in the animal model.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1074/mcp.M800486-MCP200DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2709197PMC
July 2009

Preventing bacterial infections with pilus-based vaccines: the group B streptococcus paradigm.

J Infect Dis 2009 Jan;199(1):108-15

Novartis Vaccines, Siena, Italy.

We recently described the presence of 3 pilus variants in the human pathogen group B streptococcus (GBS; also known as Streptococcus agalactiae), each encoded by a distinct pathogenicity island, as well as the ability of pilus components to elicit protection in mice against homologous challenge. To determine whether a vaccine containing a combination of proteins from the 3 pilus types could provide broad protection, we analyzed pili distribution and conservation in 289 clinical isolates. We found that pilus sequences in each island are conserved, all strains carried at least 1 of the 3 islands, and a combination of the 3 pilus components conferred protection against all tested GBS challenge strains. These data are the first to indicate that a vaccine exclusively constituted by pilus components can be effective in preventing infections caused by GBS, and they pave the way for the use of a similar approach against other pathogenic streptococci.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1086/595564DOI Listing
January 2009

Sequence variation in group A Streptococcus pili and association of pilus backbone types with lancefield T serotypes.

J Infect Dis 2008 Dec;198(12):1834-41

Novartis Vaccines, Siena, Italy.

Background: We previously reported that group A Streptococcus (GAS) pili are the T antigens described by Rebecca Lancefield. We also showed that these pili, constituted by backbone, ancillary 1, and ancillary 2 proteins, confer protection against GAS challenge in a mouse model.

Methods: We evaluated pilus distribution and conservation by sequencing the subunits of 39 new GAS isolates and used immunoblot analysis and agglutination assays to define the specificity of T sera to pilus subunits.

Results: GAS pili are encoded by 9 different islands within which backbone protein, ancillary protein 1, and ancillary protein 2 cluster in 15, 16, and 5 variants, respectively. Immunoblot and agglutination assays revealed that T type is determined by the backbone variant. This observation enabled us to set up a simple polymerase chain reaction assay to define the T type of GAS isolates.

Conclusions: We propose the use of a tee gene sequence typing, analogous to the emm gene typing, as a valuable molecular tool that could substitute for the serological T classification of GAS strains. From our sequence analysis and from recent epidemiological data, we estimate that a vaccine comprising a combination of 12 backbone variants would protect against > 90% of currently circulating strains.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1086/593176DOI Listing
December 2008

Streptococcus pyogenes pili promote pharyngeal cell adhesion and biofilm formation.

Mol Microbiol 2007 May;64(4):968-83

Novartis Vaccines and Diagnostics, Via Fiorentina 1, 53100, Siena, Italy.

Group A Streptococcus (GAS, Streptococcus pyogenes) is a Gram-positive human pathogen responsible for several acute diseases and autoimmune sequelae that account for half a million deaths worldwide every year. GAS infections require the capacity of the pathogen to adhere to host tissues and assemble in cell aggregates. Furthermore, a role for biofilms in GAS pathogenesis has recently been proposed. Here we investigated the role of GAS pili in biofilm formation. We demonstrated that GAS pilus-negative mutants, in which the genes encoding either the pilus backbone structural protein or the sortase C1 have been deleted, showed an impaired capacity to attach to a pharyngeal cell line. The same mutants were much less efficient in forming cellular aggregates in liquid culture and microcolonies on human cells. Furthermore, mutant strains were incapable of producing the typical three-dimensional layer with bacterial microcolonies embedded in a carbohydrate polymeric matrix. Complemented mutants had an adhesion and aggregation phenotype similar to the wild-type strain. Finally, in vivo expression of pili was indirectly confirmed by demonstrating that most of the sera from human patients affected by GAS-mediated pharyngitis recognized recombinant pili proteins. These data support the role of pili in GAS adherence and colonization and suggest a general role of pili in all pathogenic streptococci.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1111/j.1365-2958.2007.05704.xDOI Listing
May 2007

Microbial genomes and vaccine design: refinements to the classical reverse vaccinology approach.

Curr Opin Microbiol 2006 Oct 4;9(5):532-6. Epub 2006 Aug 4.

Novartis Vaccines, Via Fiorentina, 53100 Siena, Italy.

The advent of whole-genome sequencing of bacteria and advances in bioinformatics have revolutionized the study of bacterial pathogenesis, enabling the targeting of possible vaccine candidates starting from genomic information. Nowadays, the availability of hundreds of bacterial genomes enables identification of the genetic differences across several genomes from the same species. The unexpected degree of intra-species diversity suggests that a single genome sequence is not entirely representative and does not offer a complete picture of the genetic variability of a species. The practical consequence is that, in many cases, a universal vaccine is possible only by including a combination of antigens and this combination must take into account the pathogen population structure.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.mib.2006.07.003DOI Listing
October 2006

Identification of novel genomic islands coding for antigenic pilus-like structures in Streptococcus agalactiae.

Mol Microbiol 2006 Jul;61(1):126-41

Chiron Vaccines, Via Fiorentina 1, 53100, Siena, Italy.

We have recently reported the presence of covalently linked pilus-like structures in the human pathogen, Group B Streptococcus (GBS). The pilus operon codes for three proteins which contain the conserved amino acid motif, LPXTG, associated with cell wall-anchored proteins together with two genes coding for sortase enzymes. Analysis of the eight sequenced genomes of GBS has led to the identification of a second, related genomic island of which there are two variants, each containing genes coding for proteins with LPXTG motifs and sortases. Here we show that both variant islands also code for pilus-like structures. Furthermore, we provide a thorough description and characterization of the genomic organization of the islands and the role of each protein in the assembly of the pili. For each pilus, polymerization of one of the three component proteins is essential for incorporation of the other two proteins into the pilus structure. In addition, two sortases are required for complete pilus assembly, each with specificity for one of the pilus components. A component protein of one of the newly identified pili is also a previously identified protective antigen and a second component of this pilus is shown to confer protection against GBS challenge. We propose that pilus-like structures are important virulence factors and potential vaccine candidates.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1111/j.1365-2958.2006.05225.xDOI Listing
July 2006

Characterization and identification of vaccine candidate proteins through analysis of the group A Streptococcus surface proteome.

Nat Biotechnol 2006 Feb 15;24(2):191-7. Epub 2006 Jan 15.

Chiron Vaccines, Via Fiorentina, 1 53100 Siena, Italy.

We describe a proteomic approach for identifying bacterial surface-exposed proteins quickly and reliably for their use as vaccine candidates. Whole cells are treated with proteases to selectively digest protruding proteins that are subsequently identified by mass spectrometry analysis of the released peptides. When applied to the sequenced M1_SF370 group A Streptococcus strain, 68 PSORT-predicted surface-associated proteins were identified, including most of the protective antigens described in the literature. The number of surface-exposed proteins varied from strain to strain, most likely as a consequence of different capsule content. The surface-exposed proteins of the highly virulent M23_DSM2071 strain included 17 proteins, 15 in common with M1_SF370. When 14 of the 17 proteins were expressed in E. coli and tested in the mouse for their capacity to confer protection against a lethal dose of M23_DSM2071, one new protective antigen (Spy0416) was identified. This strategy overcomes the difficulties so far encountered in surface protein characterization and has great potential in vaccine discovery.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1038/nbt1179DOI Listing
February 2006

Group A Streptococcus produce pilus-like structures containing protective antigens and Lancefield T antigens.

Proc Natl Acad Sci U S A 2005 Oct 13;102(43):15641-6. Epub 2005 Oct 13.

Chiron Vaccines, Via Fiorentina 1, 53100 Siena, Italy.

Although pili have long been recognized in Gram-negative pathogens as important virulence factors involved in adhesion and invasion, very little is known about extended surface organelles in Gram-positive pathogens. Here we report that Group A Streptococcus (GAS), a Gram-positive human-specific pathogen that causes pharyngitis, impetigo, invasive disease, necrotizing fasciitis, and autoimmune sequelae has long, surface-exposed, pilus-like structures composed of members of a family of extracellular matrix-binding proteins. We describe four variant pili and show that each is recognized by a specific serum of the Lancefield T-typing system, which has been used for over five decades to characterize GAS isolates. Furthermore, we show that immunization of mice with a combination of recombinant pilus proteins confers protection against mucosal challenge with virulent GAS bacteria. The data indicate that induction of a protective immune response against these structures may be a useful strategy for development of a vaccine against disease caused by GAS infection.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1073/pnas.0507808102DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC1253647PMC
October 2005

Genome analysis of multiple pathogenic isolates of Streptococcus agalactiae: implications for the microbial "pan-genome".

Proc Natl Acad Sci U S A 2005 Sep 19;102(39):13950-5. Epub 2005 Sep 19.

Institute for Genomic Research, 9712 Medical Center Drive, Rockville, MD 20850, USA.

The development of efficient and inexpensive genome sequencing methods has revolutionized the study of human bacterial pathogens and improved vaccine design. Unfortunately, the sequence of a single genome does not reflect how genetic variability drives pathogenesis within a bacterial species and also limits genome-wide screens for vaccine candidates or for antimicrobial targets. We have generated the genomic sequence of six strains representing the five major disease-causing serotypes of Streptococcus agalactiae, the main cause of neonatal infection in humans. Analysis of these genomes and those available in databases showed that the S. agalactiae species can be described by a pan-genome consisting of a core genome shared by all isolates, accounting for approximately 80% of any single genome, plus a dispensable genome consisting of partially shared and strain-specific genes. Mathematical extrapolation of the data suggests that the gene reservoir available for inclusion in the S. agalactiae pan-genome is vast and that unique genes will continue to be identified even after sequencing hundreds of genomes.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1073/pnas.0506758102DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC1216834PMC
September 2005

Identification of a universal Group B streptococcus vaccine by multiple genome screen.

Science 2005 Jul;309(5731):148-50

Chiron srl, Via Fiorentina 1, 53100 Siena, Italy.

Group B Streptococcus (GBS) is a multiserotype bacterial pathogen representing a major cause of life-threatening infections in newborns. To develop a broadly protective vaccine, we analyzed the genome sequences of eight GBS isolates and cloned and tested 312 surface proteins as vaccines. Four proteins elicited protection in mice, and their combination proved highly protective against a large panel of strains, including all circulating serotypes. Protection also correlated with antigen accessibility on the bacterial surface and with the induction of opsonophagocytic antibodies. Multigenome analysis and screening described here represent a powerful strategy for identifying potential vaccine candidates against highly variable pathogens.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1126/science.1109869DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC1351092PMC
July 2005

Genome analysis reveals pili in Group B Streptococcus.

Science 2005 Jul;309(5731):105

Chiron Srl, Via Fiorentina 1, 53100 Siena, Italy.

Pili are essential virulence factors in many Gram-negative bacteria; however, they have not been described in most important Gram-positive pathogens. While screening the sequence of multiple genomes of Group B Streptococcus, we identified protective antigens that formed high molecular weight polymers. Immunogold electron microscopy revealed that the structures have a pilus-like form. These large structures have gone unrecognized in decades of studies of Group B Streptococcus.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1126/science.1111563DOI Listing
July 2005

NadA diversity and carriage in Neisseria meningitidis.

Infect Immun 2004 Jul;72(7):4217-23

IRIS, Chiron Vaccines, via Fiorentina 1, 53100 Siena, Italy.

NadA is a novel vaccine candidate recently identified in Neisseria meningitidis and involved in adhesion to host tissues. The nadA gene has been found in approximately 50% of the strains isolated from patients and in three of the four hypervirulent lineages of non-serogroup A strains. Here we investigated the presence of the nadA gene in 154 meningococcal strains isolated from healthy people (carrier strains). Only 25 (16.2%) of the 154 carrier isolates harbored the nadA gene. The commensal species Neisseria lactamica was also found not to have the nadA gene. Eighteen of the carrier strains belonged to the ET-5 and ET-37 hypervirulent clusters, indicating that only the 5.1% of the genuine carrier population actually harbored nadA (7 of 136 strains). Five of the seven strains harbored a novel allele of the nadA gene that was designated nadA4. The NadA4 protein was present on the bacterial surface as heat-stable high-molecular-weight oligomers. Antibodies against the recombinant NadA4 protein were bactericidal against homologous strains, whereas the activity against other NadA alleles was weak. In conclusion, the nadA gene segregates differently in the population of strains isolated from healthy individuals and in the population of strains isolated from patients. The presence of NadA can therefore be used as a tool to study the dynamics of meningococcal infections and understand why this bacterium, which is mostly a commensal, can become a severe pathogen.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1128/IAI.72.7.4217-4223.2004DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC427459PMC
July 2004

Reverse vaccinology.

Drug Discov Today 2003 May;8(10):459-64

IRIS, Chiron S.r.l., via Fiorentina 1, 53100 Siena, Italy.

Whole-genome sequencing of bacteria and advances in bioinformatics have revolutionized the vaccinology field, leading to the identification of potential vaccine candidates without the need for cultivating the pathogen. This approach, termed "reverse vaccinology", reduces the time and cost required for the identification of candidate vaccines and provides new solutions for those diseases for which conventional approaches have failed. The first example of the potential of reverse vaccinology has been the identification of novel antigens of meningococcus B as potential candidates for a novel and effective vaccine. The same approach has been successfully applied to other important human pathogens, demonstrating the feasibility to develop vaccines against any infectious disease. This review focuses on some recent advances in the identification of vaccine candidates by mining the genomic sequences of pathogenic bacteria.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/s1359-6446(03)02689-8DOI Listing
May 2003

Complete genome sequence and comparative genomic analysis of an emerging human pathogen, serotype V Streptococcus agalactiae.

Proc Natl Acad Sci U S A 2002 Sep 28;99(19):12391-6. Epub 2002 Aug 28.

The Institute for Genomic Research, 9712 Medical Center Drive, Rockville, MD 20850, USA.

The 2,160,267 bp genome sequence of Streptococcus agalactiae, the leading cause of bacterial sepsis, pneumonia, and meningitis in neonates in the U.S. and Europe, is predicted to encode 2,175 genes. Genome comparisons among S. agalactiae, Streptococcus pneumoniae, Streptococcus pyogenes, and the other completely sequenced genomes identified genes specific to the streptococci and to S. agalactiae. These in silico analyses, combined with comparative genome hybridization experiments between the sequenced serotype V strain 2603 V/R and 19 S. agalactiae strains from several serotypes using whole-genome microarrays, revealed the genetic heterogeneity among S. agalactiae strains, even of the same serotype, and provided insights into the evolution of virulence mechanisms.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1073/pnas.182380799DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC129455PMC
September 2002

NadA, a novel vaccine candidate of Neisseria meningitidis.

J Exp Med 2002 Jun;195(11):1445-54

Immunological Research Institute Siena, Chiron S.p.A., via Fiorentina 1, 53100 Siena, Italy.

Neisseria meningitidis is a human pathogen, which, in spite of antibiotic therapy, is still a major cause of mortality due to sepsis and meningitis. Here we describe NadA, a novel surface antigen of N. meningitidis that is present in 52 out of 53 strains of hypervirulent lineages electrophoretic types (ET) ET37, ET5, and cluster A4. The gene is absent in the hypervirulent lineage III, in N. gonorrhoeae and in the commensal species N. lactamica and N. cinerea. The guanine/cytosine content, lower than the chromosome, suggests acquisition by horizontal gene transfer and subsequent limited evolution to generate three well-conserved alleles. NadA has a predicted molecular structure strikingly similar to a novel class of adhesins (YadA and UspA2), forms high molecular weight oligomers, and binds to epithelial cells in vitro supporting the hypothesis that NadA is important for host cell interaction. NadA induces strong bactericidal antibodies and is protective in the infant rat model suggesting that this protein may represent a novel antigen for a vaccine able to control meningococcal disease caused by three hypervirulent lineages.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2193550PMC
http://dx.doi.org/10.1084/jem.20020407DOI Listing
June 2002