Zuzana Machovcova
DVM, PhD ADDICQO GROUP s.r.o.
Salmonella is a common cause of alimentary disease in humans. Within the EU, salmonellosis is the second most common zoonotic disease, with up to 90 000 cases recorded annually. Salmonella Enteritidis is the most common causative agent of reported cases, with poultry being the main source of infection. Contaminated food and undercooked poultry meat are a risk for the spread of Salmonelfa. The Czech Republic’s national Sa/monelia control programme seeks to ensure effective measures in laying hens, breeding flocks and broiler breeds with a maximum prevalence of 1-2% of S. Enteritidis or S. Typhimurium positive flocks.
Limiting the spread of Safmonella Enteritidis
Salmonella spp. is a Gram-negative, facultatively anaerobic, non-sporulating motile bacterium belonging to the Enterobacteriaceae family. There are about 2700 serovars and 200 of them are confirmed in poultry. According to EFSA, the most common serovar in laying hens is S. Enteritidis and in broilers S. Infantis.
Salmonella spreads by oral route, colonising the digestive tract and penetrating other tissues such as the liver and spleen. To cause gastroenteritis, the pathogen must enter the distal part of the small intestine and the appendix, where it must overcome various defence mechanisms. The microbiome in healthy birds is able to prevent Salmonella from colonising the intestinal cells. The infectious load depends on several factors including general health, host immunity, presence of other infections, environment and stress load. The infectious dose is estimated to be 10° CFU/ml, but during the early stages of infection, not all infected birds may always show obvious clinical signs. Clinical signs in poultry include diarrhoea, difficulty in breathing, reduced feed intake, weight loss and others. Younger birds are more susceptible to the spread of infection, so it is important to take precautions against the spread of salmonellosis in poultry farms from day-old chicks onwards.
The use of antimicrobial therapy is a major strategy to reduce the spread of Salmonella and increase performance in food-producing animals. Due to increasing antibiotic resistance, the possibility of residues and intestinal dysbiosis, current research is focusing on alternative methods to control Salmonella infections in poultry. Available non-antibiotic solutions to reduce the incidence of salmonellosis include the use of fatty acid esters, prebiotics, probiotics, postbiotics, phytobiotics, bacteriophages or vaccination.
Short- and medium-chain fatty acid esters play an important role in reducing the number of pathogenic bacteria and preventing zoonoses in the digestive tract of animals.
Mechanism of action of fatty acid esters
a) Pathogen reduction – ester molecules disrupt the structure and function of the cell membrane, whose permeability increases, resulting in the destruction of the bacterial cell. Further, fatty acid esters act in the bacterial cell to lower the pH of the cytosol, leading to disruption of osmotic balance and lysis of the cell. This is a complex effect with a very low risk of resistance.
b) Increase in production of antimicrobial peptides — increase in gene expression for their synthesis.
c) Anti-inflammatory effect – modulation of the release of pro-inflammatory and anti-inflammatory cytokines.
d) Immunity support – fatty acid esters promote the development of the mucosal immune system, reduce inadequate immune response and optimize the immune response to vaccination.
Supporting a beneficial microbiome and strengthening the integrity of the gut are other integral parts of addressing the negative impact of intestinal pathogens. Tight junctions between epithelial cells and their nutrition are essential for a functional intestinal barrier and prevention of leaky gut syndrome. If the intestinal barrier is compromised, undesirable agents (bacteria and toxins) enter the bloodstream, creating an energy-consuming pro-inflammatory state for the animal, and performance begins to decline. Intestinal health is therefore essential in young animals where the digestive tract is still developing, but also for older animals at a time of recovery and increased demands on the body. The aim of the research by ADDICOO GROUP s.r.o. was to verify the effect of feed additives based on short- and medium-chain fatty acid esters in Fortibac® on the spread of Salmonella Enteritidis and on intestinal integrity.
Effect of fatty acid esters on the spread of Salmonella Enteritidis
In 2023, in cooperation with the Research Institute of Veterinary Medicine in Brno (VUVeL), a test was conducted on the effect of short- and medium-chain fatty acid esters on the spread of Salmonella. This project was co-financed from the state budget by the Technology agency of the Czech Republic under the Programme National Competence Centres (www.tacr.cz). The testing lasted 23 days and a total of 40 chickens of ISA brown genetics were divided into two groups – control and experimental with Fortibac®. On the fourth day, infection was induced in 7 chickens (SB – seeder birds) by oral inoculation with S. Enteritidis at a dose of 10′ CFU/ml and 13 chickens (CB – contact birds) remained in contact with the infected chickens in the group. All chickens were slaughtered consecutively on days 4, 18 and 19 post-infection (DPI) to assess the microbial load of S. Enteritidis from caecum content samples.
The results of the culture in contact birds (CB) on the fourth day after infection showed a positive effect of fatty acid esters on reducing the rate of spread of Sa/monelia Enteritidis infection in the flack by 50% (Table) and reducing appendix colonization by 86% compared to the control group (Figure 1). On the 18th day after infection, a similar trend was observed in contact chickens (CB) with a 43% reduction in the rate of spread of Safmonelia in the flock and a 40% reduction in the infectious load.
The results show that on the 19th day after infection, infected chickens (SB) showed a 28% elimination of infection in the flock and up to 65% reduction in appendix colonization compared to the control. The observed findings support the hypothesis that short- and medium-chain fatty acid esters exhibit antimicrobial activity against Salmonella Enteritidis in poultry.
Table: Effect on the spread of S. Enteritidis
Figure 1: Effect on S. Enteritidis colonization in the cecum (log CFU/g)
Effect of fatty acid esters on intestinal integrity
Monitoring of the effect of fatty acid esters on intestinal integrity was carried out in 2023 at VUVeL in Brno, where biomarkers of intestinal integrity were tested in ISA brown chickens at the age of eight days. A total of 38 chickens were included in the testing, divided into two groups – control and experimental with Fortibac®. This project was also co-financed from the state budget by the Technology agency of the Czech Republic under the Programme National Competence Centres (www.tacr.cz).
The experimental group showed higher expression of tight-junction (TJ) protein synthesis – Occludin, Zonula occludens, Claudin (Figure 2), confirming better tight junction between intestinal epithelial cells and prevention of leaky gut syndrome. Higher values were also confirmed for the production of antimicrobial peptides, specifically Cathelicidin (Figure 3), which is responsible for the control and prevention of infectious diseases. Antimicrobial peptides are able to destroy lipoprotein membranes of microbes (bacteria and viruses), modulate the host immune response and promote regeneration of the intestinal mucosa. The results suggest that fatty acid esters have a positive effect on intestinal health and integrity.
Figure 2: Tight-junction protein synthesis gene expression jejunum)
Figure 3: Expression of an antimicobial peptide syntheis gene – Cathelicidin
Conclusion
Performance parameters such as feed conversion, gain and mortality are influenced by the health of the digestive tract, pathogen load in the litter and in the house, the level of immune response and the integrity of the intestinal epithelium. The optimal combination of short and medium chain fatty acid esters in the Fortibac® range has a positive effect on animal health, immunity and gut integrity. It offers an effective non-antibiotic solution to reduce the spread and colonisation of Salmonella and improve farm economics.