Salmonella is a genus that commonly attacks the health of poultry, especially chickens. This genus can live in the digestive tract specifically in the intestines (Chen et al., 2025). Salmonella typhimurium is a species that attacks many birds with symptoms that are almost undetectable but can still reduce the immunity and health of poultry. S. typhimurium has a wide range of hosts with invasive but non-adaptive infectious biological activity to the host (Shaji et al., 2023).

One type of poultry that is widely attacked by S. typhimurium is chickens. Chicken farms in Indonesia are the sectors most affected by S. typhimurium infection, including domestic chicken farms (Daryono & Puspita, 2017). One of the domestic chickens that is quite commonly known in the community is Yudhistira Kampung Chicken. The AKY domestic chicken strain represents a locally adapted genetic line with limited immunophysiological characterization. Domestic chickens basically have a stronger character than broilers, namely being adaptable and more resistant to stress and disease (Badaruddin et al., 2025; (Daryono & Puspita, 2017)). This character does not rule out the possibility that domestic chickens can also be infected by S. typhimurium.

Salmonella typhimurium is an intracellular pathogenic bacterium capable of surviving and replicating within the host through various virulence mechanisms. It infects the host via multiple pathways, including directly into cells, receptor recognition, Salmonella Pathogenecity Island (SPI), and Lipopolysaccharide (LPS) (Barbosa et al., 2017).Lipopolysaccharides are molecules found in the cell membrane structure  of S. typhimurium that are responsible for pathogenicity and virulence to hosts called endotoxins (Hassan et al., 2020).

Lipopolysaccharide S. typhimurium can infect the host through binding to Toll-Like Receptor 4 (TLR4) found on the host cell membrane. This bond will give rise to an acute, local, and even systemic immune response of the host (Hassan et al., 2020). The initial or congenital immune response that arises is usually the occurrence of fever because cells will produce a considerable amount of cytokines, one of which is IL-6. Increased cytokines are directly proportional to the occurrence of fever in the host with a time span of one to 24 hours from the beginning of infection (Petes et al., 2018).

Handling Salmonella  infections has actually been carried out by many farmers and the government. Antibiotics have been routinely administered by farmers, but infections caused by Salmonella can still occur, reducing the quality of meat and production quantity. Vaccination is considered an effective preventive strategy to reduce Salmonella infection in poultry by stimulating protective immune responses. The vaccines currently available on the market are salmonellosis vaccines with inactivated vaccine manufacturing methods and attenuated vaccines(Khan et al., 2024)(Khan et al., 2025). These two vaccines are actually enough to reduce the problem  of Salmonella infection, but the weakness of these two vaccines is the potential for cross-contamination, is not suitable for DOC chickens, and there is a fear of reversal (Khan et al., 2025).

LPS derived from Salmonella typhimurium has been explored as a potential immunostimulatory component due to its ability to trigger host inflammatory responses without introducing live bacteria (Loor-Giler et al., 2025). An LPS-based approach may offer practical advantages, including relatively simple production and lower cost compared to conventional vaccine platforms. Therefore, investigating the physiological and early immune-related responses of domestic chickens to S. typhimurium-derived LPS represents an initial step toward understanding its potential relevance in future vaccine-oriented research.

This research was conducted in the biology laboratory, Faculty of Mathematics and Natural Sciences, University of Jember, from August to November 2025

Chicken body temperature measurements are carried out every hour for ten hours post-injection. There was a difference between temperatures in each group. The results of the temperature measurement of chicken breasts can be seen in Figure 1.

The results of the chicken body temperature measurement showed that treatment groups 1 and treatment 2 had different results from the control group. Treatment groups 1 and 2 had an increase in temperature at the 1st hour to the highest in the 3rd hour with a temperature range of 42.5°C to 42.6°C. In contrast to the control group, which has a static temperature with a range of 41°C to 41.8°C. These results can illustrate in simple terms that LPS+adjuvant injection as a vaccine model can increase the body temperature of chickens. The increase in the body temperature of chickens here means that chickens are able to respond to the presence of LPS in their bodies, triggering fever (Groves et al., 2015). The fever that occurs is still in a normal stage and does not cause severe effects on the health of chickens or even death. Broadly speaking, LPS+adjuvant is able to trigger an early innate immune response in chickens that reflects the activation of early defense mechanisms and potentially supports the formation of subsequent immune responses (Zhang et al., 2019). The results of the statistical analysis of chicken body temperature can be seen in Table 1 and Table 2.

The results of the LMM analysis showed that the treatment, time, and interaction between treatment and time had a significant impact on the body temperature of chickens. The treatment given in the form of LPS + adjuvant was able to trigger a real chicken immune response in the treatment group compared to the control group. This shows that the fever that occurs reflects the activity of the initial innate immune response in chickens that is active to overcome the presence of non-specific pathogens that enter from outside the body (Zhang et al., 2019).

The significant influence of time shows that the temperature changes that occur at any given time are dynamic, which means that they change at any time. This characteristic is typical of the innate immune response that changes over time according to the physiological conditions that occur (Groves et al., 2015)(Zhang et al., 2019). The significant interaction of treatment and time reinforced that each group had different temperatures throughout the observation time. These differences include the speed of the increase in temperature, the time of the highest fever peak, etc. This difference illustrates that the injection of the LPS vaccine model is influential in triggering an innate immune response (Zhang et al., 2019).

The results of the above statistics show that in the control group when compared to the two treatment groups the treatment differs significantly. The two treatment groups had higher temperatures significantly different from the control group. This shows that the LPS-based vaccine model can trigger an innate immune response in the form of fever, while in the control group that was only injected with PBS, the temperature tended to be normal without fever. This study is in line with the research of (Xie et al., 2000), that broiler chickens injected with LPS had a higher temperature than the control group that only injected NaCl.

In both treatment groups, there was also a significant difference with temperatures at a concentration of 1mg/kg having higher temperatures and fever peaks. The increase in body temperature observed following LPS administration is associated with the activation of innate immune responses. LPS is recognized by Toll-like receptor 4 (TLR4), leading to the release of pro-inflammatory cytokines such as IL-1β and IL-6, which act on the hypothalamic thermoregulatory center and induce fever as part of the systemic inflammatory response (Santacroce et al., 2023). Therefore, body temperature in this study was interpreted as a physiological indicator of systemic inflammatory response rather than direct evidence of pathogen-specific immunity. In this case, the administration of LPS with a higher concentration triggers a higher fever than a lower concentration.

Fever is the body's normal response in overcoming the presence of external pathogens that enter the body. LPS binds to TLR4 receptors which will then trigger the formation of cytokines including IL-10, IL-6, INF, etc. (Santacroce et al., 2023)(Windoloski et al., 2022). The formation of these cytokines is a stage of the innate immune response in response to nonspecific pathogens. In responding to this pathogen, the body will emit the same response, namely the occurrence of fever.

The occurrence of fever in a normal way shows a good indicator because it means that the body is able to respond to the presence of pathogens. Fever can also occur due to a response from immune cells, namely macrophages that induce the production of cytokine IL-1, mast cells that also secrete histamine (Santacroce et al., 2023). This histamine production by the brain will cause an increase in body temperature until a fever occurs (Blomqvist, 2024).

The research conducted also had the same results as the research of (Xie et al., 2000) and (Groves et al., 2015), namely chickens injected with LPS had a higher body temperature than chickens in the control group. The occurrence of fever in treatment groups 1 and 2 showed positive results that the vaccine model can trigger a chicken-innate immune response. The response caused was not severe, which shows that the LPS vaccine model has the potential to be developed in the future (Groves et al., 2015). The response of the chicken's body also looks good by showing a regular fever with a time span that tends to be stable.

Although a significant febrile response was observed following crude LPS administration, fever alone does not fully represent protective innate immune activation. The present study did not evaluate cytokine expression, leukocyte dynamics, or molecular inflammatory markers. Therefore, further studies are required to confirm the immunostimulatory profile of the LPS preparation at the cellular and molecular levels.

This study confirms the pyrogenic effect of crude LPS derived from Salmonella typhimurium in domestic chickens, as reflected by a dose-dependent increase in body temperature. While these findings indicate systemic inflammatory activation, additional immunological assessments are necessary to determine its relevance for vaccine-related applications.

Acknowledgements and funding statements: The authors gratefully acknowledge to BIMA 2026 grant via Dr. Asmoro Lelono for providing financial support for this research. The authors also thank the Laboratory of Biology, Faculty of Mathematics and Natural Sciences, University of Jember, for technical assistance and facilities provided during the experiment.

Competing of interest: The authors declare that there are no financial, professional, or personal relationships that could be perceived as influencing the research outcomes. The authors declare no competing interests.

Author’s contributions: Conceptualization: Rizky Surya Wijaya, Dr. Asmoro Lelono, Dr. Rike Oktarianti; Methodology: Rizky Surya Wijaya, Dr. Asmoro Lelono, Dr. Rike Oktarianti; Investigation and Data Collection: Rizky Surya Wijaya; Formal Analysis: Rizky Surya Wijaya; Funding Acquisition: Dr. Asmoro Lelono; Supervision: Dr. Asmoro Lelono; Writing – Original Draft Preparation: Rizky Surya Wijaya; Writing – Review & Editing: Dr. Asmoro Lelono, Dr. Rike Oktarianti.

Generative AI: No generative artificial intelligence tools were used in the preparation of this manuscript.

Data availability: The data supporting the findings of this study are available from the corresponding author upon reasonable request.