{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,7,17]],"date-time":"2026-07-17T03:47:34Z","timestamp":1784260054079,"version":"3.55.0"},"reference-count":45,"publisher":"MDPI AG","issue":"5","license":[{"start":{"date-parts":[[2025,5,10]],"date-time":"2025-05-10T00:00:00Z","timestamp":1746835200000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"DOI":"10.13039\/100010269","name":"Wellcome Trust","doi-asserted-by":"publisher","award":["216268\/Z\/19\/Z"],"award-info":[{"award-number":["216268\/Z\/19\/Z"]}],"id":[{"id":"10.13039\/100010269","id-type":"DOI","asserted-by":"publisher"}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["Vaccines"],"abstract":"<jats:p>Background\/Objectives: Rising antibiotic resistance underscores the urgent need for effective vaccines against shigellosis. Our previous research identified the Shigella flexneri 2a truncated mutant (STM), a wzy gene knock-out strain cultivated in shake-flasks, as a promising broadly protective Shigella vaccine candidate. Expanding on this finding, our current study explores the feasibility of transitioning to a fermentor-grown STM as a vaccine candidate for further clinical development. Methods: The STM and STM-Cj, engineered to express the conserved Campylobacter jejuni N-glycan antigen, were grown in animal-free media, inactivated with formalin, and evaluated for key antigen retention and immunogenicity in mice. Results: The fermentor-grown STM exhibited significantly increased production yields and retained key antigens after inactivation. Immunization with the STM, particularly along with the double-mutant labile toxin (dmLT) adjuvant, induced robust immune responses to the conserved proteins IpaB, IpaC, and PSSP-1. Additionally, it provided protection against homologous and heterologous Shigella challenges in a mouse pulmonary model. The STM-Cj vaccine elicited antibody responses specific to the N-glycan while maintaining protective immune responses against Shigella. These findings underscore the potential of the fermentor-grown STM as a safe and immunogenic vaccine platform for combating shigellosis and possibly other gastrointestinal bacterial infections. Conclusions: Further process development to optimize growth and key antigen expression as well as expanded testing in additional animal models for the assessment of protection against Shigella and Campylobacter are needed to build on these encouraging initial results. Ultimately, clinical trials are essential to evaluate the efficacy and safety of STM-based vaccines in humans.<\/jats:p>","DOI":"10.3390\/vaccines13050506","type":"journal-article","created":{"date-parts":[[2025,5,12]],"date-time":"2025-05-12T10:58:07Z","timestamp":1747047487000},"page":"506","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":1,"title":["Shigella Mutant with Truncated O-Antigen as an Enteric Multi-Pathogen Vaccine Platform"],"prefix":"10.3390","volume":"13","author":[{"ORCID":"https:\/\/orcid.org\/0000-0002-4556-5553","authenticated-orcid":false,"given":"Jae-Ouk","family":"Kim","sequence":"first","affiliation":[{"name":"Science Unit, International Vaccine Institute, Seoul 08826, Republic of Korea"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-3542-7840","authenticated-orcid":false,"given":"Harald","family":"Nothaft","sequence":"additional","affiliation":[{"name":"VaxAlta Inc., Department of Medical Microbiology and Immunology, University of Alberta, Edmonton, AB T6G 2E1, Canada"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Younghye","family":"Moon","sequence":"additional","affiliation":[{"name":"Science Unit, International Vaccine Institute, Seoul 08826, Republic of Korea"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Seonghun","family":"Jeong","sequence":"additional","affiliation":[{"name":"Science Unit, International Vaccine Institute, Seoul 08826, Republic of Korea"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Anthony R.","family":"Vortherms","sequence":"additional","affiliation":[{"name":"Walter Reed Army Institute for Research, Silver Spring, MD 20910, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Manki","family":"Song","sequence":"additional","affiliation":[{"name":"Science Unit, International Vaccine Institute, Seoul 08826, Republic of Korea"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Christine M.","family":"Szymanski","sequence":"additional","affiliation":[{"name":"VaxAlta Inc., Department of Medical Microbiology and Immunology, University of Alberta, Edmonton, AB T6G 2E1, Canada"},{"name":"Complex Carbohydrate Research Center, University of Georgia, Athens, GA 30602, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-1932-6345","authenticated-orcid":false,"given":"Jessica","family":"White","sequence":"additional","affiliation":[{"name":"PATH, Seattle, WA 98103, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Richard","family":"Walker","sequence":"additional","affiliation":[{"name":"PATH, Washington, DC 20001, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"1968","published-online":{"date-parts":[[2025,5,10]]},"reference":[{"key":"ref_1","doi-asserted-by":"crossref","unstructured":"MacLennan, C.A., and Steele, A.D. (2022). Frontiers in Shigella Vaccine Development. Vaccines, 10.","DOI":"10.3390\/vaccines10091536"},{"key":"ref_2","doi-asserted-by":"crossref","first-page":"1229","DOI":"10.1016\/S1473-3099(18)30475-4","article-title":"Morbidity and mortality due to shigella and enterotoxigenic Escherichia coli diarrhoea: The Global Burden of Disease Study 1990\u20132016","volume":"18","author":"Khalil","year":"2018","journal-title":"Lancet Infect. Dis."},{"key":"ref_3","doi-asserted-by":"crossref","first-page":"100368","DOI":"10.1016\/j.jvacx.2023.100368","article-title":"Exploring Shigella vaccine priorities and preferences: Results from a mixed-methods study in low- and middle-income settings","volume":"15","author":"Fleming","year":"2023","journal-title":"Vaccine X"},{"key":"ref_4","doi-asserted-by":"crossref","first-page":"S76","DOI":"10.1016\/j.vaccine.2022.12.037","article-title":"Vaccine value profile for Shigella","volume":"41","author":"Hausdorff","year":"2023","journal-title":"Vaccine"},{"key":"ref_5","unstructured":"World Health Organization (2021). WHO Preferred Product Characteristics for Vaccines Against Shigella, WHO."},{"key":"ref_6","doi-asserted-by":"crossref","first-page":"540","DOI":"10.1038\/nrmicro1662","article-title":"Clinical trials of Shigella vaccines: Two steps forward and one step back on a long, hard road","volume":"5","author":"Levine","year":"2007","journal-title":"Nat. Rev. Microbiol."},{"key":"ref_7","doi-asserted-by":"crossref","first-page":"933","DOI":"10.1093\/cid\/ciu468","article-title":"Shigella isolates from the global enteric multicenter study inform vaccine development","volume":"59","author":"Livio","year":"2014","journal-title":"Clin. Infect. Dis."},{"key":"ref_8","doi-asserted-by":"crossref","first-page":"4768","DOI":"10.1016\/j.vaccine.2019.02.070","article-title":"Vaccines Against Shigella and Enterotoxigenic Escherichia coli: A summary of the 2018 VASE Conference","volume":"37","author":"Barry","year":"2019","journal-title":"Vaccine"},{"key":"ref_9","doi-asserted-by":"crossref","first-page":"3735","DOI":"10.1016\/j.vaccine.2005.07.014","article-title":"Safety and immunogenicity of an oral, inactivated, whole-cell vaccine for Shigella sonnei: Preclinical studies and a Phase I trial","volume":"24","author":"McKenzie","year":"2006","journal-title":"Vaccine"},{"key":"ref_10","doi-asserted-by":"crossref","first-page":"315","DOI":"10.1128\/CVI.00608-15","article-title":"Evaluation of the Safety, Tolerability, and Immunogenicity of an Oral, Inactivated Whole-Cell Shigella flexneri 2a Vaccine in Healthy Adult Subjects","volume":"23","author":"Chakraborty","year":"2016","journal-title":"Clin. Vaccine Immunol."},{"key":"ref_11","doi-asserted-by":"crossref","first-page":"366","DOI":"10.1128\/CVI.00683-13","article-title":"Development and preclinical evaluation of a trivalent, formalin-inactivated Shigella whole-cell vaccine","volume":"21","author":"Kaminski","year":"2014","journal-title":"Clin. Vaccine Immunol."},{"key":"ref_12","doi-asserted-by":"crossref","first-page":"1581","DOI":"10.1016\/j.vaccine.2006.11.012","article-title":"Vaccine potential for inactivated shigellae","volume":"25","author":"Osorio","year":"2007","journal-title":"Vaccine"},{"key":"ref_13","doi-asserted-by":"crossref","unstructured":"Kantele, A., Riekkinen, M., Jokiranta, T.S., Pakkanen, S.H., Pietil\u00e4, J.P., Patjas, A., Eriksson, M., Khawaja, T., Klemets, P., and Marttinen, K. (2023). Safety and immunogenicity of ETVAX\u00ae, an oral inactivated vaccine against enterotoxigenic Escherichia coli diarrhoea: A double-blinded, randomized, placebo-controlled trial amongst Finnish travellers to Benin, West Africa. J. Travel. Med., 30.","DOI":"10.1093\/jtm\/taad045"},{"key":"ref_14","doi-asserted-by":"crossref","first-page":"514","DOI":"10.1016\/S1473-3099(23)00742-9","article-title":"Effectiveness of one dose of killed oral cholera vaccine in an endemic community in the Democratic Republic of the Congo: A matched case-control study","volume":"24","author":"Malembaka","year":"2024","journal-title":"Lancet Infect. Dis."},{"key":"ref_15","doi-asserted-by":"crossref","first-page":"546","DOI":"10.1128\/CVI.00538-10","article-title":"Characterization of a mutant Escherichia coli heat-labile toxin, LT(R192G\/L211A), as a safe and effective oral adjuvant","volume":"18","author":"Norton","year":"2011","journal-title":"Clin. Vaccine Immunol."},{"key":"ref_16","doi-asserted-by":"crossref","first-page":"S821","DOI":"10.1093\/infdis\/jiab475","article-title":"Mucosal Immune Responses Against an Oral Enterotoxigenic Escherichia coli Vaccine Evaluated in Clinical Trials","volume":"224","author":"Svennerholm","year":"2021","journal-title":"J. Infect. Dis."},{"key":"ref_17","doi-asserted-by":"crossref","first-page":"6884","DOI":"10.1016\/j.vaccine.2023.09.052","article-title":"Safety, tolerability, and immunogenicity of an oral inactivated ETEC vaccine (ETVAX(R)) with dmLT adjuvant in healthy adults and children in Zambia: An age descending randomised, placebo-controlled trial","volume":"41","author":"Sukwa","year":"2023","journal-title":"Vaccine"},{"key":"ref_18","doi-asserted-by":"crossref","unstructured":"Muthuramalingam, M., Whittier, S.K., Picking, W.L., and Picking, W.D. (2021). The Shigella Type III Secretion System: An Overview from Top to Bottom. Microorganisms, 9.","DOI":"10.3390\/microorganisms9020451"},{"key":"ref_19","doi-asserted-by":"crossref","first-page":"1222","DOI":"10.1128\/IAI.06174-11","article-title":"Broadly protective Shigella vaccine based on type III secretion apparatus proteins","volume":"80","author":"Kissmann","year":"2012","journal-title":"Infect. Immun."},{"key":"ref_20","doi-asserted-by":"crossref","unstructured":"Turbyfill, K.R., Clarkson, K.A., Oaks, E.V., and Kaminski, R.W. (2022). From Concept to Clinical Product: A Brief History of the Novel Shigella Invaplex Vaccine\u2019s Refinement and Evolution. Vaccines, 10.","DOI":"10.3390\/vaccines10040548"},{"key":"ref_21","doi-asserted-by":"crossref","first-page":"4470","DOI":"10.1128\/IAI.00859-13","article-title":"Characterization of a novel fusion protein from IpaB and IpaD of Shigella spp. and its potential as a pan-Shigella vaccine","volume":"81","author":"Chen","year":"2013","journal-title":"Infect. Immun."},{"key":"ref_22","doi-asserted-by":"crossref","first-page":"381","DOI":"10.1128\/CVI.00661-14","article-title":"Shigella outer membrane protein PSSP-1 is broadly protective against Shigella infection","volume":"22","author":"Kim","year":"2015","journal-title":"Clin. Vaccine Immunol."},{"key":"ref_23","doi-asserted-by":"crossref","unstructured":"Ndungo, E., Randall, A., Hazen, T.H., Kania, D.A., Trappl-Kimmons, K., Liang, X., Barry, E.M., Kotloff, K.L., Chakraborty, S., and Mani, S. (2018). A Novel Shigella Proteome Microarray Discriminates Targets of Human Antibody Reactivity following Oral Vaccination and Experimental Challenge. mSphere, 3.","DOI":"10.1128\/mSphere.00260-18"},{"key":"ref_24","doi-asserted-by":"crossref","first-page":"641","DOI":"10.1038\/icb.2015.24","article-title":"Shigella IpaB and IpaD displayed on L. lactis bacterium-like particles induce protective immunity in adult and infant mice","volume":"93","author":"Heine","year":"2015","journal-title":"Immunol. Cell Biol."},{"key":"ref_25","doi-asserted-by":"crossref","unstructured":"Kim, M.J., Moon, Y.H., Kim, H., Rho, S., Shin, Y.K., Song, M., Walker, R., Czerkinsky, C., Kim, D.W., and Kim, J.O. (2018). Cross-Protective Shigella Whole-Cell Vaccine with a Truncated O-Polysaccharide Chain. Front. Microbiol., 9.","DOI":"10.3389\/fmicb.2018.02609"},{"key":"ref_26","doi-asserted-by":"crossref","first-page":"e0041724","DOI":"10.1128\/jb.00417-24","article-title":"Recent insights into Wzy polymerases and lipopolysaccharide O-antigen biosynthesis","volume":"207","author":"Ascari","year":"2025","journal-title":"J. Bacteriol."},{"key":"ref_27","doi-asserted-by":"crossref","unstructured":"Nothaft, H., Perez-Mu\u00f1oz, M.E., Yang, T., Murugan, A.V.M., Miller, M., Kolarich, D., Plastow, G.S., Walter, J., and Szymanski, C.M. (2021). Improving Chicken Responses to Glycoconjugate Vaccination Against Campylobacter jejuni. Front. Microbiol., 12.","DOI":"10.3389\/fmicb.2021.734526"},{"key":"ref_28","doi-asserted-by":"crossref","first-page":"287","DOI":"10.1039\/D0MO00032A","article-title":"Identifying the targets and functions of N-linked protein glycosylation in Campylobacter jejuni","volume":"16","author":"Cain","year":"2020","journal-title":"Mol. Omics"},{"key":"ref_29","doi-asserted-by":"crossref","unstructured":"Badjo, A.O.R., Kabore, N.F., Zongo, A., Gnada, K., Ouattara, A., Muhigwa, M., Ouangraoua, S., Poda, A., Some, S.A., and Schubert, G. (2024). Burden and epidemiology of Campylobacter species in acute enteritis cases in Burkina Faso. BMC Infect. Dis., 24.","DOI":"10.1186\/s12879-024-09709-y"},{"key":"ref_30","doi-asserted-by":"crossref","unstructured":"Zong Minko, O., Mabika Mabika, R., Moyen, R., Mounioko, F., Ondjiangui, L.F., and Yala, J.F. (2024). The Impact of Campylobacter, Salmonella, and Shigella in Diarrheal Infections in Central Africa (1998\u20132022): A Systematic Review. Int. J. Environ. Res. Public Health, 21.","DOI":"10.3390\/ijerph21121635"},{"key":"ref_31","doi-asserted-by":"crossref","first-page":"107","DOI":"10.1556\/1886.2023.00043","article-title":"Campylobacter in Africa\u2014A specific viewpoint","volume":"13","author":"Paintsil","year":"2023","journal-title":"Eur. J. Microbiol. Immunol."},{"key":"ref_32","doi-asserted-by":"crossref","first-page":"1171","DOI":"10.1093\/cid\/ciw542","article-title":"Epidemiology and Impact of Campylobacter Infection in Children in 8 Low-Resource Settings: Results from the MAL-ED Study","volume":"63","author":"Amour","year":"2016","journal-title":"Clin. Infect. Dis."},{"key":"ref_33","doi-asserted-by":"crossref","first-page":"2775","DOI":"10.1128\/IAI.71.5.2775-2786.2003","article-title":"Complete genome sequence and comparative genomics of Shigella flexneri serotype 2a strain 2457T","volume":"71","author":"Wei","year":"2003","journal-title":"Infect. Immun."},{"key":"ref_34","first-page":"249","article-title":"Immunochemical characteristics of Shigella sonnei and serotype 6 Shigella flexneri lipopolysaccharides and enterobacterial common antigen","volume":"26","author":"Romanowska","year":"1978","journal-title":"Arch. Immunol. Et Ther. Exp."},{"key":"ref_35","doi-asserted-by":"crossref","first-page":"1056","DOI":"10.1038\/ng.2369","article-title":"Shigella sonnei genome sequencing and phylogenetic analysis indicate recent global dissemination from Europe","volume":"44","author":"Holt","year":"2012","journal-title":"Nat. Genet."},{"key":"ref_36","doi-asserted-by":"crossref","first-page":"6640","DOI":"10.1073\/pnas.120163297","article-title":"One-step inactivation of chromosomal genes in Escherichia coli K-12 using PCR products","volume":"97","author":"Datsenko","year":"2000","journal-title":"Proc. Natl. Acad. Sci. USA"},{"key":"ref_37","doi-asserted-by":"crossref","first-page":"462","DOI":"10.1111\/j.1574-695X.2006.00118.x","article-title":"Developing live Shigella vaccines using \u03bb Red recombineering","volume":"47","author":"Ranallo","year":"2006","journal-title":"FEMS Immunol. Med. Microbiol."},{"key":"ref_38","doi-asserted-by":"crossref","first-page":"2690","DOI":"10.1021\/acschembio.1c00498","article-title":"Detecting Glucose Fluctuations in the Campylobacter jejuni N-Glycan Structure","volume":"16","author":"Nothaft","year":"2021","journal-title":"ACS Chem. Biol."},{"key":"ref_39","doi-asserted-by":"crossref","first-page":"9317","DOI":"10.1073\/pnas.85.23.9317","article-title":"Characterization of invasion plasmid antigen genes (ipaBCD) from Shigella flexneri","volume":"85","author":"Venkatesan","year":"1988","journal-title":"Proc. Natl. Acad. Sci. USA"},{"key":"ref_40","doi-asserted-by":"crossref","first-page":"1719","DOI":"10.1128\/jb.177.7.1719-1726.1995","article-title":"Cleavage of Shigella surface protein VirG occurs at a specific site, but the secretion is not essential for intracellular spreading","volume":"177","author":"Fukuda","year":"1995","journal-title":"J. Bacteriol."},{"key":"ref_41","doi-asserted-by":"crossref","first-page":"1203","DOI":"10.1074\/mcp.M112.021519","article-title":"Diversity in the protein N-glycosylation pathways within the Campylobacter genus","volume":"11","author":"Nothaft","year":"2012","journal-title":"Mol. Cell Proteom."},{"key":"ref_42","doi-asserted-by":"crossref","unstructured":"Tran, E.N., Doyle, M.T., and Morona, R. (2013). LPS unmasking of Shigella flexneri reveals preferential localisation of tagged outer membrane protease IcsP to septa and new poles. PLoS ONE, 8.","DOI":"10.1371\/journal.pone.0070508"},{"key":"ref_43","doi-asserted-by":"crossref","first-page":"111","DOI":"10.1016\/S1672-0229(07)60021-7","article-title":"Dynamic proteome changes of Shigella flexneri 2a during transition from exponential growth to stationary phase","volume":"5","author":"Zhu","year":"2007","journal-title":"Genom. Proteom. Bioinform."},{"key":"ref_44","doi-asserted-by":"crossref","unstructured":"Caboni, M., P\u00e9dron, T., Rossi, O., Goulding, D., Pickard, D., Citiulo, F., MacLennan, C.A., Dougan, G., Thomson, N.R., and Saul, A. (2015). An O antigen capsule modulates bacterial pathogenesis in Shigella sonnei. PLoS Pathog., 11.","DOI":"10.1371\/journal.ppat.1004749"},{"key":"ref_45","doi-asserted-by":"crossref","first-page":"374","DOI":"10.1093\/glycob\/cwz105","article-title":"Application of a gut-immune co-culture system for the study of N-glycan-dependent host-pathogen interactions of Campylobacter jejuni","volume":"30","author":"Zamora","year":"2020","journal-title":"Glycobiology"}],"container-title":["Vaccines"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/www.mdpi.com\/2076-393X\/13\/5\/506\/pdf","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2025,10,9]],"date-time":"2025-10-09T17:30:35Z","timestamp":1760031035000},"score":1,"resource":{"primary":{"URL":"https:\/\/www.mdpi.com\/2076-393X\/13\/5\/506"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2025,5,10]]},"references-count":45,"journal-issue":{"issue":"5","published-online":{"date-parts":[[2025,5]]}},"alternative-id":["vaccines13050506"],"URL":"https:\/\/doi.org\/10.3390\/vaccines13050506","relation":{},"ISSN":["2076-393X"],"issn-type":[{"value":"2076-393X","type":"electronic"}],"subject":[],"published":{"date-parts":[[2025,5,10]]}}}