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Tularemia is a rare but potentially life threatening zoonotic disease caused by Francisella tularensis. F. novicida, previously considered a subspecies of F. tularensis, is currently considered a separate species. Human infections related to F. novicida are exceedingly rare but can cause morbidity and mortality in debilitated or immunocompromised individuals.A 42-year-old male presented at the hospital with vomiting, dehydration, constipation and pain in the right iliac fossa. He was first diagnosed with pancreatitis and admitted for further analysis. Chest computerized tomography scan showed the presence of parenchymal consolidation in the left upper and lower lobes of the lung with pleural effusion. Blood cultures isolated a Gram-negative coccobacillus, that was at first identified by MALDI-TOF as Francisella tularensis. Serological analysis for the detection of total antibodies against F. tularensis and Real-Time PCR targeting the gene coding for 23 kDa, resulting negative. Subsequently, PCR targeting helicases and tul4 genes, and the Regions of Difference RD1 and RD6 were performed allowing the identification of F. novicida. The isolate was further genetically characterized by whole genome sequencing (WGS).This is the first reported case of human infection caused by F. novicida in Italy.Given the rarity of human cases and the lack of specific symptoms, this pathogen is difficult to identify and the diagnosis can be extremely challenging. In this case report, despite the lack of amplification of the gene encoding for 23 kDa protein, the identification of Francisella species was achieved with the amplification of different genes and characterized by WGS.

Francisella tularensis can cause severe disease in humans via the respiratory or cutaneous routes and a case fatality ratio of up to 10 % is reported due to lack of proper antibiotic treatment, while F. novicida causes disease in severely immunocompromised individuals. Efforts are needed to develop effective vaccine candidates against Francisella species. Thus, in this study, a systematic computational work frame was used to deeply investigate the whole proteome of Francisella novicida containing 1728 proteins to develop vaccine against F. tularensis and related species. Whole-proteome analysis revealed that four proteins including (A0Q492) (A0Q7Y4), (A0Q4N4), and (A0Q5D9) are the suitable vaccine targets after the removal of homologous, paralogous and prediction of subcellular localization. These proteins were used to predict the T cell, B cell, and HTL epitopes which were joined together through suitable linkers to construct a multi-epitopes vaccine (MEVC). The MEVC was found to be highly immunogenic and non-allergenic while the physiochemical properties revealed the feasible expression and purification. Moreover, the molecular interaction of MEVC with TLR2, molecular simulation, and binding free energy analyses further validated the immune potential of the construct. According to Jcat analysis, the refined sequence demonstrates GC contents of 41.48 % and a CAI value of 1. The in-silico cloning and optimization process ensured compatibility with host codon usage, thereby facilitating efficient expression. Computational immune simulation studies underscored the capacity of MEVC to induce both primary and secondary immune responses. The conservation analysis further revealed that the selected epitopes exhibit 100 % conservation across different species and thus provides wider protection against Francisella. • Francisella novicida can cause severe disease in humans via the respiratory or cutaneous routes. • Investigation of the whole-proteome revealed four proteins as suitable vaccine targets. • We designed a multi-epitopes vaccine (MEVC) from T cell, B cell, and HTL epitopes. • Molecular docking, simulation, in silico cloning and immune simulation confirmed the potential of our MEVC. • Our MEVC could also provide cross-protection against the other species of Francisella. [ABSTRACT FROM AUTHOR]

Tularemia is a rare but potentially life threatening zoonotic disease caused by Francisella tularensis . F. novicida , previously considered a subspecies of F. tularensis , is currently considered a separate species. Human infections related to F. novicida are exceedingly rare but can cause morbidity and mortality in debilitated or immunocompromised individuals.A 42-year-old male presented at the hospital with vomiting, dehydration, constipation and pain in the right iliac fossa. He was first diagnosed with pancreatitis and admitted for further analysis. Chest computerized tomography scan showed the presence of parenchymal consolidation in the left upper and lower lobes of the lung with pleural effusion. Blood cultures isolated a Gram-negative coccobacillus, that was at first identified by MALDI-TOF as  Francisella tularensis.  Serological analysis for the detection of total antibodies against  F. tularensis  and Real-Time PCR targeting the gene coding for 23 kDa, resulting negative. Subsequently, PCR targeting  helicases  and  tul4  genes, and the Regions of Difference  RD1  and  RD6  were performed allowing the identification of  F. novicida . The isolate was further genetically characterized by whole genome sequencing (WGS).This is the first reported case of human infection caused by  F. novicida  in Italy.Given the rarity of human cases and the lack of specific symptoms, this pathogen is difficult to identify and the diagnosis can be extremely challenging. In this case report, despite the lack of amplification of the gene encoding for 23 kDa protein, the identification of  Francisella  species was achieved with the amplification of different genes and characterized by WGS., Competing Interests: None., (© 2024 The Authors. Published by Elsevier Ltd.)