Tehran University of Medical Sciences Iranian Journal of Microbiology 2008-3289 18 4 2026 08 04 574 583 EN Asad Ghalib Prof. Dr. Ikram-ul-Haq Institute of Industrial Biotechnology, Government College University, Lahore, Pakistan Sajjad Hussain Veterinary Research Institute, Livestock and Dairy Development Department, Government of the Punjab, Lahore, Pakistan Waseem Shahzad Veterinary Research Institute, Livestock and Dairy Development Department, Government of the Punjab, Lahore, Pakistan Rashid Mir Prince Fahd Bin Sultan Research Chair for Biomedical Research, Faculty of Applied Medical Sciences, University of Tabuk, Tabuk, Kingdom of Saudi Arabia; Department of Medical Laboratory Technology, University of Tabuk, Tabuk, Kingdom of Saudi Arabia Bushra Muneer Prof. Dr. Ikram-ul-Haq Institute of Industrial Biotechnology, Government College University, Lahore, Pakistan 2026 05 06 2026 07 08 Background and Objectives: Salmonella gallinarum, causes fowl typhoid, continues to be a significant limitation to poultry production in developing nations, with continuously rising antimicrobial resistance. The current research was design and test an indigenous S. gallinarum ghost vaccine and to measure its immunogenicity and protective effect on layer birds. Materials and Methods: Bacteriological culture of suspected clinical cases of fowl typhoid was performed, and isolates were verified by PCR against the 16S rRNA gene, after which they were sequenced, and phylogenetic analysis was performed. To obtain bacterial ghost cells, a representative native isolate was subjected to a detergent-based chemical lysis procedure, followed up by scanning electron microscopy. 120-day-old layer chicks were divided into six groups randomly and immunized at the age of seven days using oral, intramuscular, air-spray, intramuscular with oil adjuvant or eye-drop routes. There was a 1st dose at 26 days of age in all vaccinated groups and the challenge of virulent S. gallinarum at 42 and 93 days of age. ELISA was used to test humoral immune response at different time points. Results: The highest antibody titers were found in vaccinated groups compared to controls (p < 0.05), and the intramuscular route generated the best and most enduring response in systemic immunity. Mucosal pathways elicited not high but rather significant antibody responses and provided protection against clinical disease. Conclusion: The research contributes to the feasibility of ghost-based vaccination in the role of a sustainable alternative to the disease control measures that are highly reliant on antibiotics in endemic poultry production systems. https://ijm.tums.ac.ir/index.php/ijm/article/view/6181 https://ijm.tums.ac.ir/index.php/ijm/article/download/6181/1917