Lipopolysaccharides (LPSs) or endotoxin are the major constituent of the bacterial Gram-negative outer membrane including Salmonella. It is an immunodominant molecule that is important for the virulence and pathogenesis of Salmonella typhimurium.
Aim: this study was aimed to extract LPS of Salmonella typhimurium and evaluate its purity by using HPLC. Then assessment of its effect on RNA expression of the immune-related genes; interferon-γ (IFN-γ) and interleukin 1β (IL-1β).
Material and methods: One hundred Samples were collected from 79 farms (broiler and layers) showing symptoms of Salmonellosis between 2017 - 2019. Samples for each flock (cecum, Liver and spleen) were collected, labelled, transported and pooled together as one Sample. Salmonella typhimurium was isolated and identified by biochemical and serological test and confirmed by using conventional PCR Using STM4495 gene of S. typhimurium. Extraction and purification of LPS from isolated S. typhimurium was done by using isopropanol and sodium hydroxide mixture. its purity was assessed by HPLC. To assess its effect on poultry immunity, different concentrations of extracted LPS (50, 100, 150 and 200 μg/kg) were injected subcutaneously in 21-day old Specific Pathogen Free (SPF) Chicks of the broiler breeds. After 3 hours of injection, spleens were collected for quantitative measurement of interleukin 1β (IL-1β), interferon-γ (IFN-γ) gene expression.
Results: Two Salmonella typhimurium were confirmed out of 25 isolated Salmonella. HPLC showed highly purified extracted LPS compared with standard commercial LPS. quantitative measurement of RNA expression of the immune-related genes showed significant upregulation of IFN-γ and low downregulation of IL-1β.
Conclusions: Extraction of LPS by using isopropanol and sodium hydroxide mixture overcome the presence of nucleic acids of extracted Salmonella in the final product and yielded highly purified LPS. Its injection in 21-day old chicks revealed significant upregulation of IFN-γ and low downregulation of IL-1β of the immune-related genes |
