zahra salahi - Department of Microbiology, Faculty of Medicine, Shahed University, Tehran, Iran.
Mojtaba Nofeli - Razi Vaccine and Serum Research Institute (RVSRI), Agricultural Research, Education and Extension Organization (AREEO), Karaj, Iran.
Mohammad mehdi Ranjbar - Razi Vaccine and Serum Research Institute (RVSRI), Agricultural Research, Education and Extension Organization (AREEO), Karaj, Iran.
Masoumeh Bagheri - Razi Vaccine and Serum Research Institute (RVSRI), Agricultural Research, Education and Extension Organization (AREEO), Karaj, Iran.
Majid Esmaelizad - Razi Vaccine and Serum Research Institute (RVSRI), Agricultural Research, Education and Extension Organization (AREEO), Karaj, Iran.
Mohammad Niakan - Department of Microbiology, Faculty of Medicine, Shahed University, Tehran, Iran.

Traditionally, diphtheria-tetanus-pertussis (DTwP or DTaP) as pediatric vaccines are produced from the corresponding inactivated toxins or whole cell pathogenic bacteria of Corynebacterium diphtheria toxin (Tox), Clostridium tetani toxin (TetX) and Bordetella pertussis. There are major concerns in the classic or acellular DTP (DTaP) vaccine production processes from native live sources of bacteria as it may raise concerns on adverse effects and safety issues, complexity of the purifications for each agent as well as costs. Here, we designated recombinant multi-epitope candidate vaccine by vaccino-informatics study to address mentioned issues and to develop a single trivalent fusion protein as potent recombinant DTP vaccine. To follow these goals, stages of immune-bioinformatics were retrieved by means of proteinaceous toxins sequences, predicting secondary/tertiary structure and transmembrane topology, energy minimization and models validation. Then, conformational and linear B cell epitope prediction by several different servers, mapping consensus linear/discontinuous immunogenic regions and constructing synthetic fusion vaccine candidate in respect to optimal immunogenic, physico-chemical properties and highly expression in prokaryote host were achieved. Finally, reverse translation, codon optimization, addition of cloning tags for pet ۲۸a vector and optimizations of physico-chemical characteristics of synthetic trivalent fusion protein were performed. By different hybrid immune-informatics and structural bioinformatics analysis predicted and experimental epitopes finally, ۱۲ new consensus highly immunogenic linear and discontinuous epitopes in Tox, TetX and PTXa proteins were selected. Peptide sequence of these immunogenic regions were as follow; PTXA (AA۳۴-۶۴, AA۱۸۴-۲۵۶ and AA۹۸-۱۱۶), Tox (AA۴۷-۷۶, AA۱۱۷-۱۵۹, AA۵۱۵-۵۵۷ and AA۲۴۵-۲۶۵) and TetX (AA۲۲۶-۲۴۹, AA۸۱۹-۸۴۴, AA۹۲۳-۹۶۷, AA۱۰۰۹-۱۰۶۷ and AA۱۲۲۵-۱۳۱۵). Moreover, the characteristics of recombinant trivalent fusion construct were; ۵۴۶ residue length, solvable (Grand average of hydropathicity (GRAVY) was -۰.۴۷۵), estimated half-life was >۱۰ hours in Escherichia coli, pI ۵.۹۴ (a little acidic), stable protein (The instability index (II) ۳۵.۵۸) as well as thermal stable (Aliphatic index (AI) ۷۱.۶۷). The putative antigenic epitopes from different organisms in a single protein, as in the current study, possibly will improve the protective effectiveness as novel potent, safe, cheap and broad-spectrum vaccines for better prevention of diphtheria, tetanus and pertussis infections in future.

Bordetella pertussis, Corynebacterium diphtheria, Clostridium tetani, Epitopes, Recombinant vaccine