- Open Access
NERVE: New Enhanced Reverse Vaccinology Environment
BMC Biotechnology volume 6, Article number: 35 (2006)
Since a milestone work on Neisseria meningitidis B, Reverse Vaccinology has strongly enhanced the identification of vaccine candidates by replacing several experimental tasks using in silico prediction steps. These steps have allowed scientists to face the selection of antigens from the predicted proteome of pathogens, for which cell culture is difficult or impossible, saving time and money. However, this good example of bioinformatics-driven immunology can be further developed by improving in silico steps and implementing biologist-friendly tools.
We introduce NERVE (New Enhanced Reverse Vaccinology Environment), an user-friendly software environment for the in silico identification of the best vaccine candidates from whole proteomes of bacterial pathogens. The software integrates multiple robust and well-known algorithms for protein analysis and comparison. Vaccine candidates are ranked and presented in a html table showing relevant information and links to corresponding primary data. Information concerning all proteins of the analyzed proteome is not deleted along selection steps but rather flows into an SQL database for further mining and analyses.
After learning from recent years' works in this field and analysing a large dataset, NERVE has been implemented and tuned as the first available tool able to rank a restricted pool (~8–9% of the whole proteome) of vaccine candidates and to show high recall (~75–80%) of known protective antigens. These vaccine candidates are required to be "safe" (taking into account autoimmunity risk) and "easy" for further experimental, high-throughput screening (avoiding possibly not soluble antigens). NERVE is expected to help save time and money in vaccine design and is available as an additional file with this manuscript; updated versions will be available at http://www.bio.unipd.it/molbinfo.
Reverse Vaccinology (RV) is one of the best examples of how Bioinformatics can boost Molecular Immunology. The conventional approach to design vaccines requires pathogen's cultivation and dissection of its main components before testing their ability to elicit protective immunity. RV's novelty consists in starting the search for immunogenic antigens from in silico analyses of the pathogen's genome instead of culturing the microorganism . This allows scientists to save time and money while facing pathogens for which cell culture is difficult or impossible. RV potentially permits researchers to select, in addition to most in vivo expressed antigens (the easiest ones to purify), any protein encoded by the genome of a pathogen. RV is less helpful with eucaryotes due to complexities of cellular and tissue organization, and it is more effective with prokaryotes, extra- and intracellular. Indeed, the production of specific antibodies can boost immunity not only against extracellular pathogens, usually controlled by Th2-polarized responses, but also against either obligate or facultative intracellular ones, usually controlled by Th1-polarized responses. Even these latter pathogens are susceptible to humoral immunity during the extracellular phases of their infectious cycle and are made vulnerable by antibody cross-linking that modifies the intracellular milieu through signaling [2, 3].
The extracellular pathogen, Neisseria meningitidis serogroup B, stands as a milestone for RV. From its genome, Pizza and coworkers selected 570 out of 2158 predicted ORFs for protein expression as those meant to be new, surface-exposed antigens . 350 ORFs were successfully cloned, expressed in Escherichia coli and tested in a variety of assays. In the end, five proteins were found to satisfy criteria of surface exposure, conservation/expression in all strains as well as a significant titer in serum bactericidal assays. This work induced further research of other pathogens in the same way: Porphyromonas gingivalis , Streptococcus pneumoniae , Chlamydia pneumoniae , Bacillus anthracis  and group B streptococci . Enhancing such a genome-based approach with a trascriptome-based one – through the use of DNA microarrays – shows whether, when and to what extent antigens are expressed, thus identifying those highly expressed during infection processes . Finally, proteomics, although expensive and laborious, can verify in silico prediction of membrane composition .
We focused on the selection of the in silico vaccine candidates (VCs) from the list of predicted proteins to address two issues: the possibility (i) of automatizing the process with new criteria of analyses and (ii) of reducing the percentage of VCs to less than the 20–30% reported until now . Indeed, the real goal of in silico selection is to choose the minimal number of VCs sufficient to find protective antigens (PAs) during experimental phases, thus designing a vaccine that conserves time and money. Therefore, it has to be stressed that obtaining a high recall of PAs in a very small number of selected VCs is more convenient than trying to find all possible PAs. According to this approach it is more productive to select the PAs that will most likely be easily expressed. This reduces the risk of experimental troubles. The main cause of failed cloning and expression of 250 out of 600 VCs from Neisseria meningitidis B was the presence of more than one transmembrane spanning region (TM) . Thus, we decided to have no more than two predicted TMs as an a priori requirement. Of course, viable VCs can be missed because of such a filter; the extracellular loops of multispanning membrane proteins can actually be significant targets especially if reasonably large. Hence, one may focus on fragments likely to be exposed and accessible to antibodies rather than the entire protein in order to overcome eventual cloning and expression problems. Although such a "local" approach is reasonable, we designed the software to select entire antigens for experimental challenges rather than hazarding fine predictions that may not correspond to in vivo conditions.
Choosing such a conservative way to face vaccine design inevitably implies missing some PAs, but this is a small price to reach a valuable compromise. Even selecting only surface antigens may imply missing non-surface PAs. Yet discarding non-surface-exposed predicted antigens, thereby forwarding further bioinformatic analyses on selected ones, proved successful. However, to mimimize the risk of loosing good VCs, we stored up information concerning analytical steps for each sequence without distinction: the final output presents only selected VCs, but information concerning all other proteins is retained. VCs are presented with corresponding integrated analyses. In order to "rationalize" selection step, we took into account the importance of avoiding antigens that can potentially cause autoimmunity in man . Since Major Hystocompatibility Complex (MHC) ligands can be really short (as few as eight residues); this problem may rise also from antigens sharing weak global similarity with host proteins. Addressing such a question on a proteomic scale would make manual management of bioinformatic analyses impossible.
We report here on a new, fully automated RV system, developed to predict best VCs from bacterial proteomes (inferred by completely sequenced genomes and publicly available) and to manage and show data by user-friendly output.
NERVE software pipeline is presented in figure 1. NERVE is composed of eight Perl scripts. A script named "NERVE" leads the user through text-interface configuration. Once this is successfully completed, the script starts and manages the whole process that can be roughly divided in two parts: the first in which all data is produced and stored and the second in which a restricted part of this data is selected. The acquisition of data is performed by six different analytical steps, each forwarded by a special script and each in turn screens the whole proteome indistinctly. All the information produced is stored both in a MySQL table and in text files organized in special subdirectories. Once production and storage of data is completed, another script selects a restricted number of entries from the MySQL table (restriction) and ranks them in a user-friendly html table (ranking). Restriction is performed by four integrated filters and is based on values created by steps 1 to 4. Filters LOC and TOP select non cytoplasmic antigens containing no more than two TMs (NERVE a priori requirements). Subsequently, based on empirical threshold values, filters PAD and SHP discard antigens that show a low probability of being adhesins and/or a significant similarity to human proteins. To set these values, NERVE was tuned on ten entire proteomes containing known immunogens; we chose the pair of threshold values that gave the best compromise between restrictivness and immunogen recall. After restriction, NERVE uses values created by step 5 to rank the antigens extracted by the filters. An html table presents this restricted and ranked pool of VCs, showing in six columns the values created by the analytical steps. Entries and data are linked to the corresponding textfiles, thus providing the user with complete information.
Results and discussion
The system we created predicts the best VCs starting from the flatfile proteome of a prokaryotic pathogen. It forwards six proteome-wide analyses that mine and save data in text files and in an automatically generated MySQL table. The table will contain as many records of extracted information as the number of sequences in thirteen fields. The best VCs are finally selected and are shown in a user-friendly html table reporting seven out of the thirteen fields, linked to the textfile-information that they summarize.
The first proteome scannings assign each sequence three predictions: (i) subcellular localization, (ii) adhesin probability and (iii) topology, using the algorithms PSORTb 2.0 , SPAAN  and HMMTOP  respectively. Indeed, surface-exposed proteins such as outer membrane proteins and especially adhesins, are ideal targets for vaccine development [4, 17]. At the same time, the presence of more than one TM in many of these VCs, proved problematic in cloning and expression phases of RV (see introduction) . This clearly shows how predicting number of TMs is potentially a crucial step in saving time and money for subsequent experimental tests. The fourth step addresses the problem of sharing similarity regions between pathogen and host proteins. It is well known that in vaccine development this can either cause low immune response/tolerance or autoimmunity . We have chosen the algorithm BLAST to compare each pathogen's sequence as a query against the human proteome . Local alignment is suitable for this task because potential MHC ligands – the ones we search for to help predict potential interferences (tolerance or autoimmunity) with the immune system – can be very short (~9 residues) . Thus, the script scans alignments, extracting each sequence fragment (shared peptide, SP) that shows no more than three "positives" (compatible substitutions) and one "mismatch" (not compatible substitution) per nine-residues (minimal length required) window. In this way we not only take into account MHC II, commonly involved in presentation of exogenous antigens, but also MHC I, involved in cross-presentation [20, 21]. These settings can be changed at the start of the manager programme according to the user's preferences. Once an SP is found, corresponding bacterial and human sequences are reported with their position in the protein and the available description for the human entry in the MySQL table and the text file. Once this fourth step is complete, each pathogen's sequence is assigned to a file, named as its accession, and sent to four new fields in the MySQL database. The first two of these fields show query length and number of mined SPs. The third one reports four features for each SP: position, amino acid sequence, occurrence and "MHC ligand". This last feature is expressed as either "positive" or "negative" depending on the results from a screening of human MHC ligands derived from the database MHCPEP . These MHC ligands do not necessarily correspond to T-epitopes. It would be more advantageous to use a T-epitope prediction tool, but as of now there is no such tool available for standalone use. In addition, since the number of unknown MHC ligands is possibly high, "positive" is a "necessary-but-not-sufficient" condition, whereas "negative" stands as an absence of evidence rather than an evidence of absence. The fourth field reports the aggregate number of amino acid residues from all peptides.
The fifth step allows the user to compare the pathogen's proteome to any other from a different strain/serogroup. Indeed, inferring conservation of each sequence may be helpful in selecting the best VCs as the more an immunogen is conserved, the more protective the vaccine becomes. The way in which we compare two pathogen proteomes is the same process we use to compare pathogen versus human proteomes; all regions potentially capable of binding MHC ligands are searched for in an analogous way. Although this step is not compulsory, it allows NERVE to rank VCs from most to least represented in the compared prokaryotic proteome in the html table presenting the selection set (see below). When possible the sixth step assigns an homology-driven, function prediction to each VC. For instance, this allows one to focus on a VC that shows strong similarity to either known virulence factors or strong immunogens. NERVE uses BLASTp algorithm to compare each sequence to the Uniprot database. According to Ariel et al. (2003) , the script adopts these parameters: e-value ≤1e-10 and (alignment extension)/(query length) ≥ 0.8. This allows NERVE to attribute to the sequence the subject's most similar function as a query putative function, reported in a further special MySQL column – if no hits are found, the notice "no sufficient similarity found for function inference" is shown. The putative function is then saved in a special text file with the corresponding score and e-value. This last step completes the MySQL database, which in total contains thirteen fields complete with all the information produced along the six analyses. Although each type of information may be retrieved at any time, the final script presents the best VCs in a user-friendly html table. This displays accession numbers and six out of the eleven MySQL fields, linking them to text files containing summarized data (figure 2).
For final selection of VCs, NERVE uses feature-based, a priori requirements to discard proteins that are likely to waste time and money during the experimental tasks (such as those with > 2 TMs) and to exclude non-surface antigens such as cytoplasmic/inner membrane proteins (see introduction for rationale). To further improve final selection efficiency, NERVE considers two characteristics for empirical tuning: the probability that a sequence is an adhesin and the presence of regions common to human proteins. Variation of threshold values for such features along iterative analysis of a known data set (ten proteomes, from both Gram+ and Gram-, both extra- and intracellular bacteria) allowed NERVE to define restricted VC pools (average size: 8.17% proteome sequences, figure 3) including a high number (33/42 = 78.6%) of the described PAs that fulfill NERVE a priori requirements (see table 1: selected PAs are underlined; PAs not fulfilling NERVE a priori requirements are not shown). Settings consistently confirmed themselves as reliably effective – in terms of both PA recall and selection restrictiveness – when tested on six further proteomes (from both Gram+ and Gram-, both extra- and intracellular bacteria). In fact, most (22/29 = 75.9%) of the described PAs fulfilling NERVE a priori requirements (see table 2: same PAs representation criteria as in table 1) were included in 9.32% whole proteomes average size pools (figure 4).
RV stands as a turning stone in Vaccinology. It shows how powerful and useful Bioinformatics can be in the post-genomic era. Creating a tool specifically designed to automatize in silico steps not only makes RV really available but also time and cost efficient. Indeed, NERVE was conceived to combine automation with an exhaustive treatment of VCs selection task by implementing and integrating six different kinds of analyses. Its modular structure allows further development of new, additional steps as well as the refinement of existing modules. This would improve the compromise between VC selection restrictivness and PA recall. Another goal was to avoid loosing information, thereby giving the user the chance to recover all data mined by NERVE for further investigation. For instance, recovered data regarding shared similarity regions between VCs and human proteins may be of help when taking into account possible occurrences of either tolerance or autoimmunity.
Finally, the NERVE prediction system proved – on a large number of bacterial proteomes – to be reliably effective in selecting very restricted VC pools that are characterized by a high recall (75–80%) of PAs. In particular these results narrow VC pool restriction from the reported 20–30% to an improved 8–9% of proteome sequences. NERVE's attempt to save time and money is also mediated by selection criteria, meant to facilitate crucial experimental steps, the protein cloning and expression phases. Last but not least, NERVE's user-friendly format and easily interpretable output should further aid researchers in designing subunit vaccines against bacterial pathogens.
Availability and requirements
Project name: NERVE
Home page: http://www.bio.unipd.it/molbinfo
This software is also available as an additional file [See Additional file 1]
Operating system: Linux (tested distribution: Debian)
Programming language: Perl
Other requirements: Perl 5.8.7 or higher, PSORTb 2.0, SPAAN
License: GNU GPL
Any restrictions to use by non-academics: licence needed
= major hystocompatibility complex
= open reading frame
= protective antigen
= Reverse Vaccinology
= shared peptide
= transmembrane helix
= vaccine candidate.
Rappuoli R: Reverse vaccinology. Curr Opin Microbiol. 2000, 3: 445-450. 10.1016/S1369-5274(00)00119-3.
Casadevall A: Antibody-Mediated Immunity against Intracellular Pathogens: Two-Dimensional Thinking Comes Full Circle. Infect Immun. 2003, 71: 4225-4228. 10.1128/IAI.71.8.4225-4228.2003.
Montigiani S, Falugi F, Scarselli M, Finco O, Petracca R, Galli G, Mariani M, Manetti R, Agnusdei M, Cevenini R, Donati M, Nogarotto R, Norais N, Garaguso I, Nuti S, Saletti G, Rosa D, Ratti G, Grandi G: Genomic Approach for Analysis of Surface Proteins in Chlamydia pneumoniae. Infect Immun. 2002, 70: 368-379. 10.1128/IAI.70.1.368-379.2002.
Pizza M, Scarlato V, Masignani V, Giuliani MM, Arico B, Comanducci M, Jennings GT, Baldi L, Bartolini E, Capecchi B, Galeotti CL, Luzzi E, Manetti R, Marchetti E, Mora M, Nuti S, Ratti G, Santini L, Savino S, Scarselli M, Storni E, Zuo P, Broeker M, Hundt E, Knapp B, Blair E, Mason T, Tettelin H, Hood DW, Jeffries AC, Saunders NJ, Granoff DM, Venter JV, Moxon ER, Grandi G, Rappuoli R: Identification of vaccine candidates against serogroup B meningococcus by whole-genome sequencing. Science. 2000, 287: 1816-1820. 10.1126/science.287.5459.1816.
Ross BC, Czajkowski L, Hocking D, Margetts M, Webb E, Rothel L, Patterson M, Agius C, Camuglia S, Reynolds E, Littlejohn T, Gaeta B, Ng A, Kuczek ES, Mattick JS, Gearing D, Barr IG: Identification of vaccine candidate antigens from a genomic analysis of Porphyromonas gingivalis. Vaccine. 2001, 19: 4135-4142. 10.1016/S0264-410X(01)00173-6.
Wizemann TM, Heinrichs JH, Adamou JE, Erwin AL, Kunsch C, Choi GH, Barash SC, Rosen CA, Masure HR, Tuomanen E, Gayle A, Brewah YA, Walsh W, Barren P, Lathigra R, Hanson M, Langermann S, Johnson S, Koenig S: Use of a whole genome approach to identify vaccine molecules affording protection against Streptococcus pneumoniae infection. Infect Immun. 2001, 69: 1593-1598. 10.1128/IAI.69.3.1593-1598.2001.
Ariel N, Zvi A, Makarova KS, Chitlaru T, Elhanany E, Velan B, Cohen S, Friedlander AM, Shafferman A: Genome-based bioinformatic selection of chromosomal bacillus anthracis putative vaccine candidates coupled with proteomic identification of surface-associated antigens. Infect Immun. 2003, 71: 4563-4579. 10.1128/IAI.71.8.4563-4579.2003.
Maione D, Margarit I, Rinaudo CD, Masignani V, Mora M, Scarselli M, Tettelin H, Brettoni C, Iacobini ET, Rosini R, D'Agostino N, Miorin L, Buccato S, Mariani M, Galli G, Nogarotto R, Nardi Dei V, Vegni F, Fraser C, Mancuso G, Teti G, Madoff LC, Paoletti LC, Rappuoli R, Kasper DL, Telford JL, Grandi G: Identification of a universal Group B streptococcus vaccine by multiple genome screen. Science. 2005, 309: 148-150. 10.1126/science.1109869.
Grifantini R, Bartolini E, Muzzi A, Draghi M, Frigimelica E, Berger J, Ratti JG, Petracca R, Galli G, Agnusdei M, Giuliani MM, Santini L, Brunelli N, Tettelin H, Rappuoli R, Randazzo F, Grandi G: Previously unrecognized vaccine candidates against group B meningococcus identified by DNA microarrays. Nature Biotechnol. 2002, 20: 914-921. 10.1038/nbt728.
Grandi G: Rational antibacterial vaccine design through genomic technologies. Int J Parasitol. 2003, 33: 615-620. 10.1016/S0020-7519(03)00059-6.
Grandi G: Bioinformatics, DNA Microarrays and Proteomics in Vaccine Discovery: Competing or Complementary Technologies?. Genomics, Proteomics and Vaccines. Edited by: Grandi G. 2004, John Wiley & Sons Ltd, 23-41.
Serruto D, Rappuoli R, Pizza M: Meningococcus B: from Genome to Vaccine. Genomics, Proteomics and Vaccines. Edited by: Grandi G. 2004, John Wiley & Sons Ltd, 185-201.
Oldstone MB: Molecular mimicry, microbial infection, and autoimmune disease: evolution of the concept. Curr Top Microbiol Immunol. 2005, 296: 1-17. full_text.
Gardy JL, Laird MR, Chen F, Rey S, Walsh CJ, Ester M, Brinkman FSL: PSORTb v.2.0: expanded prediction of bacterial protein subcellular localization and insights gained from comparative proteome analysis. Bioinformatics. 2005, 21: 617-623. 10.1093/bioinformatics/bti057.
Sachdeva G, Kumar K, Preti J, Ramachandran S: SPAAN: a software program for prediction of adhesins and adhesin-like proteins using neural networks. Bioinformatics. 2004, 21: 483-491. 10.1093/bioinformatics/bti028.
Tusnady GE, Simon I: The HMMTOP transmembrane topology prediction server. Bioinformatics. 2001, 17: 849-850. 10.1093/bioinformatics/17.9.849.
Wizemann TM, Adamou JE, Langermann S: Adhesins as targets for vaccine development. Emerg Infect Dis. 1999, 5: 395-403. 10.3201/eid0503.990310.
Altschul FS, Madden TL, Schäffer AA, Zhang J, Zhang Z, Miller W, Lipman DJ: Gapped BLAST and PSI-BLAST: A new generation of protein database search programs. Nucleic Acids Res. 1997, 25: 3389-3402. 10.1093/nar/25.17.3389.
Reche PA, Glutting JP, Zhang H, Reinherz EL: Enhancement to the RANKPEP resource for the prediction of peptide binding to MHC molecules using profiles. Immunogenetics. 2004, 56: 405-419. 10.1007/s00251-004-0709-7.
Lehner PJ, Cresswell P: Recent developments in MHC-class-I-mediated antigen presentation. Curr Opin Immunol. 2004, 16: 82-89. 10.1016/j.coi.2003.11.012.
Ackerman AL, Kyristis C, Tampé R, Cresswell P: Access of soluble antigens to the endoplasmic reticulum can explain cross-presentation by dendritic cells. Nat Immunol. 2005, 6: 107-113. 10.1038/ni1147.
Brusic V, Rudy G, Kyne AP, Harrison LC: MHCPEP, a database of MHC-binding peptides: update 1997. Nucleic Acids Res. 1998, 26: 368-371. 10.1093/nar/26.1.368.
Mourez M, Lacy DB, Cunningham K, Legmann R, Sellman BR, Mogridge J, Collier RJ: 2001: a year of major advances in anthrax toxin research. Trends Microbiol. 2002, 10: 287-293. 10.1016/S0966-842X(02)02369-7.
Von Specht BU, Gabelsberger J, Knapp B, Hundt E, Schmidt-Pilger H, Bauernsachs S, Lenz U, Domdey H: Immunogenic efficacy of differently produced recombinant vaccines candidates against Pseudomonas aeruginosa infections. J Biotechnol. 2000, 83: 3-12. 10.1016/S0168-1656(00)00311-4.
Worgall S, Krause A, Rivara M, Hee KK, Vintayen EV, Hackett NR, Roelvink PW, Bruder JT, Wickham TJ, Kovesdi I, Crystal RG: Protection against P. aeruginosa with an adenovirus vector containing an OprF epitope in the capsid. J Clin Invest. 2005, 115: 1281-1289. 10.1172/JCI200523135.
DiGiandomenico A, Rao J, Goldberg JB: Oral vaccination of BALB/c mice with Salmonella enterica serovar Typhimurium expressing Pseudomonas aeruginosa O antigen promotes increased survival in an acute fatal pneumonia model. Infect Immun. 2004, 72: 7012-7021. 10.1128/IAI.72.12.7012-7021.2004.
Jones T, Adamovicz JJ, Cyr SL, Bolt CR, Bellerose N, Pitt LM, Lowell GH, Burt DS: Intranasal Protollin/F1-V vaccine elicits respiratory and serum antibody responses and protects mice against lethal aerosolized plague infection. Vaccine. 2006, 24: 1625-1632. 10.1016/j.vaccine.2005.09.052.
Matson JS, Durick KA, Bradley DS, Nilles ML: Immunization of mice with YscF provides protection from Yersinia pestis infections. BMC Microbiol. 2005, 5: 38-10.1186/1471-2180-5-38.
Shin EA, Lee JY, Kim TG, Park YK, Langridge WH: Synthesis and assembly of an adjuvanted Porphyromonas gingivalis fimbrial antigen fusion protein in plants. Protein Expr Purif. 2006, 47: 99-109. 10.1016/j.pep.2005.09.005.
Zhang P, Yang QB, Balkovetz DF, Lewis JP, Clements JD, Michalek SM, Katz J: Effectiveness of the B subunit of cholera toxin in potentiating immune responses to the recombinant hemagglutinin/adhesin domain of the gingipain Kgp from Porphyromonas gingivalis. Vaccine. 2005, 23: 4734-4744. 10.1016/j.vaccine.2005.05.004.
Yang QB, Martin M, Michalek SM, Katz J: Mechanisms of monophosphoryl lipid A augmentation of host responses to recombinant HagB from Porphyromonas gingivalis. Infect Immun. 2002, 70: 3557-3565. 10.1128/IAI.70.7.3557-3565.2002.
Ross BC, Czajkowski L, Vandenberg KL, Camuglia S, Woods J, Agius C, Paolini R, Reynolds E, Barr IG: Characterization of two outer membrane protein antigens of Porphyromonas gingivalis that are protective in a murine lesion model. Oral Microbiol Immunol. 2004, 19: 6-15. 10.1046/j.0902-0055.2003.00096.x.
Yoder A, Wang X, Ma Y, Philipp MT, Heilbrun M, Weis JH, Kirschning CJ, Wooten RM, Weis JJ: Tripalmitoyl-S-glyceryl-cysteine-dependent OspA vaccination of toll-like receptor 2-deficient mice results in effective protection from Borrelia burgdorferi challenge. Infect Immun. 2003, 71: 3894-3900. 10.1128/IAI.71.7.3894-3900.2003.
Scheiblhofer S, Weiss R, Durnberger H, Mostbock S, Breitenbach M, Livey I, Thalhamer J: A DNA vaccine encoding the outer surface protein C from Borrelia burgdorferi is able to induce protective immune responses. Microbes Infect. 2003, 5: 939-946. 10.1016/S1286-4579(03)00182-5.
Labandeira-Rey M, Baker EA, Skare JT: VraA (BBI16) protein of Borrelia burgdorferi is a surface-exposed antigen with a repetitive motif that confers partial protection against experimental Lyme borreliosis. Infect Immun. 2001, 69: 1409-1419. 10.1128/IAI.69.3.1409-1419.2001.
Eko FO, He Q, Brown T, McMillan L, Ifere GO, Ananaba GA, Lyn D, Lubitz W, Kellar KL, Black CM, Igietseme JU: A novel recombinant multisubunit vaccine against Chlamydia. J Immunol. 2004, 173: 3375-3382.
Palmqvist N, Silverman GJ, Josefsson E, Tarkowski A: Bacterial cell wall-expressed protein A triggers supraclonal B-cell responses upon in vivo infection with Staphylococcus aureus. Microbes Infect. 2005, 7: 1501-1511. 10.1016/j.micinf.2005.05.008.
Senna JP, Roth DM, Oliveira JS, Machado DC, Santos DS: Protective immune response against methicillin resistant Staphylococcus aureus in a murine model using a DNA vaccine approach. Vaccine. 2003, 21: 2661-2666. 10.1016/S0264-410X(02)00738-7.
Josefsson E, Hartford O, O'Brien L, Patti JM, Foster T: Protection against experimental Staphylococcus aureus arthritis by vaccination with clumping factor A, a novel virulence determinant. J Infect Dis. 2001, 184: 1572-1580. 10.1086/324430.
Zhu W, Thomas CE, Chen CJ, Van Dam CN, Johnston RE, Davis NL, Sparling PF: Comparison of immune responses to gonococcal PorB delivered as outer membrane vesicles, recombinant protein, or Venezuelan equine encephalitis virus replicon particles. Infect Immun. 2005, 73: 7558-7568. 10.1128/IAI.73.11.7558-7568.2005.
Price GA, Russell MW, Cornelissen CN: Intranasal administration of recombinant Neisseria gonorrhoeae transferrin binding proteins A and B conjugated to the cholera toxin B subunit induces systemic and vaginal antibodies in mice. Infect Immun. 2005, 73: 3945-3953. 10.1128/IAI.73.7.3945-3953.2005.
Rodriguez-Ortega MJ, Norais N, Bensi G, Liberatori S, Capo S, Mora M, Scarselli M, Doro F, Ferrari G, Garaguso I, Maggi T, Neumann A, Covre A, Telford JL, Grandi G: Characterization and identification of vaccine candidate proteins through analysis of the group A Streptococcus surface proteome. Nat Biotechnol. 24: 191-197. 10.1038/nbt1179.
Lee LH, Burg E, Baqar S, Bourgeois AL, Burr DH, Ewing CP, Trust TJ, Guerry P: Evaluation of a truncated recombinant flagellin subunit vaccine against Campylobacter jejuni. Infect Immun. 1999, 67: 5799-5805.
Wyszynska A, Raczko A, Lis M, Jagusztyn-Krynicka EK: Oral immunization of chickens with avirulent Salmonella vaccine strain carrying C. jejuni 72Dz/92 cjaA gene elicits specific humoral immune response associated with protection against challenge with wild-type Campylobacter. Vaccine. 2004, 22: 1379-1389. 10.1016/j.vaccine.2003.11.001.
Rossi G, Ruggiero P, Peppoloni S, Pancotto L, Fortuna D, Lauretti L, Volpini G, Mancianti S, Corazza M, Taccini E, Di Pisa F, Rappuoli R, Del Giudice G: Therapeutic vaccination against Helicobacter pylori in the beagle dog experimental model: safety, immunogenicity, and efficacy. Infect Immun. 2004, 72: 3252-3259. 10.1128/IAI.72.6.3252-3259.2004.
Capo S, Nuti S, Scarselli M, Tavarini S, Montigiani S, Mori E, Finco O, Abrignani S, Grandi G, Bensi G: Chlamydia pneumoniae genome sequence analysis and identification of HLA-A2-restricted CD8+ T cell epitopes recognized by infection-primed T cells. Vaccine. 2005, 23: 5028-5037. 10.1016/j.vaccine.2005.04.048.
Finco O, Bonci A, Agnusdei M, Scarselli M, Petracca R, Norais N, Ferrari G, Garaguso I, Donati M, Sambri V, Cevenini R, Ratti G, Grandi G: Identification of new potential vaccine candidates against Chlamydia pneumoniae by multiple screenings. Vaccine. 2005, 23: 1178-1188. 10.1016/j.vaccine.2004.07.045.
This work was supported by grant "Ricerca Nazionale – ex 60%" to FF. We thank Gabor Tusnady and Istvan Simon for providing HMMTOP algorithm, Massimo Facchinetti for addressing and managing intellectual property issues, Aaron Riedel for reviewing and editing the English and Dmitri Trakovsky for manuscript proofreading.
SV: conception, design, acquisition, analysis and interpretation of data, manuscript drafting; FB: design, manuscript critical reading; FF: conception, analysis and interpretation of data, manuscript drafting. All authors read and approved the final manuscript.
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Vivona, S., Bernante, F. & Filippini, F. NERVE: New Enhanced Reverse Vaccinology Environment. BMC Biotechnol 6, 35 (2006). https://doi.org/10.1186/1472-6750-6-35
- Vaccine Candidate
- Protective Antigen
- Neisseria Meningitidis
- Bacillus Anthracis
- Query Length