Characterization of highly pathogenic avian influenza virus in retail dairy products in the US.

In March 2024, clade 2.3.4.4b H5N1 highly pathogenic avian influenza virus (HPAIV) was detected in dairy cattle in the US, and it was discovered that the virus could be detected in raw milk. Although affected cow's milk is diverted from human consumption and current pasteurization requirements are expected to reduce or eliminate infectious HPAIV from the milk supply, a study was conducted to characterize whether the virus could be detected by quantitative real-time RT-PCR (qrRT-PCR) in pasteurized retail dairy products and, if detected, to determine whether the virus was viable. From 18 April to 22 April 2024, a total of 297 samples of Grade A pasteurized retail milk products (23 product types) were collected from 17 US states that represented products from 132 processors in 38 states. Viral RNA was detected in 60 samples (20.2%), with qrRT-PCR-based quantity estimates (non-infectious) of up to 5.4log1050% egg infectious doses per mL, with a mean and median of 3.0log10/mL and 2.9log10/mL, respectively. Samples that were positive for type A influenza by qrRT-PCR were confirmed to be clade 2.3.4.4 H5 HPAIV by qrRT-PCR. No infectious virus was detected in any of the qrRT-PCR-positive samples in embryonating chicken eggs. Further studies are needed to monitor the milk supply, but these results provide evidence that the infectious virus did not enter the US pasteurized milk supply before control measures for HPAIV were implemented in dairy cattle.IMPORTANCEHighly pathogenic avian influenza virus (HPAIV) infections in US dairy cattle were first confirmed in March 2024. Because the virus could be detected in raw milk, a study was conducted to determine whether it had entered the retail food supply. Pasteurized dairy products were collected from 17 states in April 2024. Viral RNA was detected in one in five samples, but infectious virus was not detected. This provides a snapshot of HPAIV in milk products early in the event and reinforces that with current safety measures, infectious viruses in milk are unlikely to enter the food supply.

Detection of echinostomatid trematode eggs at the forest-oil palm interface in Sabah, Malaysia.

In this study, we report the occurrence of echinostomatid eggs in feces of wildlife, domestic animals and humans frequenting the forest­oil palm plantation interface in the Kinabatangan (Sabah, Malaysia), and discuss potential implications for public health. Using microscopy, we detected echinostomatid eggs in six host species, including Asian palm civets (Paradoxurus hermaphroditus [13/18]), leopard cats (Prionailurus bengalensis [3/4]), long-tailed macaques (Macaca fascicularis [1/10]), domestic dogs [3/5] and cats [1/1], and humans [7/9]. Molecular analysis revealed a close genetic proximity of civet echinostomatids to Artyfechinostomum malayanum, a zoonotic parasite of public health relevance. The intermediate hosts for A. malayanum have been reported in at least 3 districts in Sabah, suggesting that all the necessary elements required for the completion of the parasite's life cycle are present. Our findings point at the presence of zoonotic trematodes in an area with high human­wildlife interaction and highlight the potential public and animal health concern of zoonotic trematode infection in the context of Southeast Asia's rapidly changing ecosystems.

Bacterial adhesion inhibition by microalgal EPSs from Cylindrotheca closterium and Tetraselmis suecica biofilms.

In the food industry, successful bacterial pathogen colonization and persistence begin with their adhesion to a surface, followed by the spatial development of mature biofilm of public health concerns. Compromising bacterial settlement with natural inhibitors is a promising alternative to conventional anti-fouling treatments typically based on chemical biocides that contribute to the growing burden of antimicrobial resistance. In this study, three extracellular polymeric substance (EPS) fractions extracted from microalgae biofilms of Cylindrotheca closterium (fraction C) and Tetraselmis suecica (fraction Ta rich in insoluble scale structure and fraction Tb rich in soluble EPS) were screened for their anti-adhesive properties, against eight human food-borne pathogens belonging to Escherichia coli, Staphylococcus aureus, Salmonella enterica subsp. enterica, and Listeria monocytogenes species. The results showed that the fraction Ta was the most effective inducing statistically significant reduction for three strains of E. coli, S. aureus, and L. monocytogenes. Overall, EPSs coating on polystyrene surfaces of the different fractions increased the hydrophilic character of the support. Differences in bacterial adhesion on the different coated surfaces could be explained by several dissimilarities in the structural and physicochemical EPS compositions, according to HPLC and ATR-FTIR analysis. Interestingly, while fractions Ta and Tb were extracted from the same microalgal culture, distinct adhesion patterns were observed, highlighting the importance of the extraction process. Overall, the findings showed that EPS extracted from microalgal photosynthetic biofilms can exhibit anti-adhesive effects against food-borne pathogens and could help develop sustainable and non-toxic anti-adhesive surfaces for the food industry. KEY POINTS: •EPSs from a biofilm-based culture of C. closterium/T. suecica were characterized. •Microalgal EPS extracted from T. suecica biofilms showed bacterial anti-adhesive effects. •The anti-adhesive effect is strain-specific and affects both Gram - and Gram + bacteria.

Relapse of toxocariasis after completion of four-week treatment with albendazole.

Food-borne toxocariasis caused by the consumption of raw meat or liver has occasionally been reported from East Asia. We treated a 38-year-old Japanese man who was infected with Toxocara in China and underwent a four-week treatment with albendazole. The liver and lung lesions disappeared after the treatment, suggesting that the treatment was successful. One month after the end of the treatment, the patient relapsed, and albendazole was administered again for eight weeks. The patient has remained relapse-free for one year. Although toxocariasis can heal spontaneously, in some cases, such as the present case, the disease relapses even after long-term treatment. In conclusion, different durations of treatment are recommended by various guidelines, and the duration of treatment needs to be modified with each case, considering the response to the treatment.

Unveiling risks in healthy food: Vegetables and fruits are linked to the distribution chain of protozoan parasites.

Vegetable and fruit contamination is recognized as a significant parasite transmission route. This review presents the current state of vegetables ad fruits contamination with food-borne parasitic protozoa worldwide. We consider the methodologies and strategies for detecting parasitic stages developed in the last decade and the contamination data. Asia had the highest number of reports (94 studies), followed by Africa (74 studies). At the country level, with 41 studies, Iran had the most reports among other countries, followed by Nigeria (28 studies). According to the studies included in the current review, 41.22% of vegetables and fruits were contaminated with different species of protozoan parasites. Among different continents, Asia accounted for the highest contamination rate of protozoan parasites (57.12%). Giardia spp. (10%) had the highest contamination rate in vegetables and fruits, followed by Entamoeba coli (8%), E. histolytica/dispar (7%), and Cryptosporidium spp. (6%). This study provides essential data for health authorities to develop food safety programs. The presence of protozoan parasites in fruits and vegetables highlights the critical need for maintaining rigorous food safety measures across the entire production and distribution process, particularly in countries that are major producers and distributors of these food items.

Microscopic detection and molecular characterization of Sarcocystis miescheriana in wild boars (Sus scrofa): first report from Greece.

The genus Sarcocystis includes protozoan parasites with an indirect life cycle. Sarcocystis spp. can infect various animal species and humans, causing sarcocystosis, a parasitosis of economic importance and zoonotic concern. Wild boars can act as intermediate hosts for Sarcocystis miescheriana and the zoonotic Sarcocystis suihominis that infects humans by consumption of raw or undercooked infected swine meat. In the present study, the diaphragmatic muscle tissue of 123 wild boars hunted in Greece was examined to determine the frequency of Sarcocystis spp. The samples were examined by tissue compression and molecular techniques. Under light microscopy, 34 out of 123 (27.6%) wild boars tested positive for Sarcocystis spp., while a higher infection prevalence (75%) was revealed by multiplex PCR performed in 100 of the samples. The partial mtDNA cox1 gene (~ 1100 bp) of 20 samples tested positive for S. miescheriana by multiplex PCR was amplified and sequenced. Sarcocystis miescheriana was identified as the only species involved in these infections. This is the first study on the prevalence of Sarcocystis spp. in wild animals in Greece. Further, large-scale surveys are needed to assess the prevalence and species of this parasite in Greece and to design efficient control and preventive measures in a One Health perspective.

Human diphyllobothriasis in Argentina: assessing the epidemiological significance from historical records and reports of new cases.

Between 1898 and 1940, eight human cases of diphyllobothriasis were reported in Argentina, always in recently arrived European immigrants. In 1982, the first autochthonous case was detected, and since then, 33 other autochthonous cases have been reported, totaling 42 cases of human diphyllobothriasis in Argentina before the present study. Our aim is to update the information on diphyllobothriasis in Argentina by identifying specimens from new cases using morphometrical and/or molecular methods. We also aim to assess the epidemiological relevance of this food-borne disease in the country. Anamnestic data were obtained from patients or professionals, along with 26 worms identified using morphometrical (21 samples) and molecular techniques (5 samples). All the patients acquired the infection by consuming freshwater salmonids caught in Andean lakes in Northern Patagonia. Morphometrics and DNA markers of worms were compatible with Dibothriocephalus latus. In total, 68 human cases have been detected in Argentina, 60 of which were autochthonous. The human population living North-western Patagonia, whose lakes are inhabited by salmonids, is increasing. Similarly, the number of other definitive hosts for Dibothriocephalus dendriticus (gulls) and for D. latus (dogs) is also increasing. In addition, salmonid fishing and the habit of consuming home-prepared raw fish dishes are becoming widespread. Therefore, it is to be expected that diphyllobothriasis in Argentina will increase further.

Multinational collaboration in solving a European Salmonella Braenderup outbreak linked to imported melons, 2021.

A genomic cluster of Salmonella Braenderup ST22, a serovar of Salmonella enterica subsp. enterica which causes symptoms of gastrointestinal illness, was notified by Danish authorities to the European Centre for Disease Prevention and Control (ECDC) on 3 May 2021. By 6 July 2021, S. Braenderup outbreak cases (n = 348) had been reported from 12 countries in the European Union/European Economic Area (EU/EEA) and the United Kingdom (UK), including 68 hospitalised cases. With support from affected EU/EEA countries, and in partnership with the European Food Safety Authority (EFSA), ECDC established an international outbreak investigation team to rapidly identify the source and prevent outbreak spread. Consumption information was shared with affected countries through a standard line list, revealing that 124 of 197 cases (63%) reported having eaten (any) melons within 7 days prior to disease onset. The speed and completeness of the investigation, which identified the outbreak vehicle as galia melons imported from Honduras in June 2021, was a direct result of extensive collaboration and information sharing between countries' national food safety and public health authorities. This article describes the outbreak and the benefits, successes, and challenges of multi-country collaboration for consideration in future large foodborne outbreaks across Europe.

Vacation in Egypt associated with Shiga toxin-producing Escherichia coli infection in children and adolescents, northern Italy, 2023.

BackgroundHaemolytic uremic syndrome (HUS) is a severe complication of infection with Shiga toxin-producing Escherichia coli (STEC). Although the reservoirs of STEC are known, the source of the infection of sporadic cases is often unknown. In 2023, we observed several cases of bloody diarrhoea with STEC infection in children and adolescents returning from vacations.AimWe aimed to explore the association between travel and bloody diarrhoea with STEC infection in children and adolescents.MethodsWe included all children and adolescents with bloody diarrhoea with STEC infection identified in 2023 by the ItalKid-HUS Network surveillance system in northern Italy. We interviewed children's families and sent a questionnaire on recent travels abroad. The exposure time was between 3 days after arrival abroad and 5 days after return home. A self-controlled case series (SCCS) design was used in the analysis.ResultsOf the 43 cases, 11 developed HUS. Twenty-three cases did not travel abroad, while 20 had travelled to several destinations. The incidence rate ratio (IRR) associated with travel to Egypt was 88.6 (95% confidence interval (CI): 17.0-462). Serotype analysis excluded the possibility of a single strain causing the infections. We did not find the source of the infections.ConclusionThere is an elevated risk of acquiring STEC infection with bloody diarrhoea and HUS associated with travel to Egypt. Specific investigations to identify the source are needed to implement effective preventive measures.

UHPLC-MS/MS and GC-MS Metabolic Profiling of a Medicinal Flourensia Fiebrigii Chemotype.

The comparative metabolic profiling and their biological properties of eight extracts obtained from diverse parts (leaves, flowers, roots) of the medicinal plant Flourensia fiebrigii S.F. Blake, a chemotype growing in highland areas (2750 m a.s.l.) of northwest Argentina, were investigated. The extracts were analysed by GC-MS and UHPLC-MS/MS. GC-MS analysis revealed the presence of encecalin (relative content: 24.86 %) in ethereal flower extract (EF) and this benzopyran (5.93 %) together sitosterol (11.35 %) in the bioactive ethereal leaf exudate (ELE). By UHPLC-MS/MS the main compounds identified in both samples were: limocitrin, (22.31 %), (2Z)-4,6-dihydroxy-2-[(4-hydroxy-3,5-dimethoxyphenyl)methylidene]-1-benzofuran-3-one (21.31 %), isobavachin (14.47 %), naringenin (13.50 %), and sternbin, (12.49 %). Phytocomplexes derived from aerial parts exhibited significant activity against biofilm production of Pseudomonas aeruginosa and Staphylococcus aureus, reaching inhibitions of 74.7-99.9 % with ELE (50 µg/mL). Notably, the extracts did not affect nutraceutical and environmental bacteria, suggesting a selective activity. ELE also showed the highest reactive species scavenging ability. This study provides valuable insights into the potential applications of this chemotype.

Exploring the potential of 1,8-cineole from cardamom oil against food-borne pathogens: Antibacterial mechanisms and its application in meat preservation.

Food-borne pathogenic bacteria are a major public health concern globally. Traditional control methods using antibiotics have limitations, leading to the exploration of alternative strategies. Essential oils such as cardamom possess antimicrobial properties and have shown efficacy against food-borne pathogenic bacteria. The utilization of essential oils and their bioactive constituents in food preservation is a viable strategy to prolong the shelf-life of food products while ensuring their quality and safety. To the best of our knowledge, there are no studies that have utilized 1,8-cineole (the main active constituent of cardamom essential oil) as a preservative in meat, so this study might be the first to utilize 1,8-cineole as an antibacterial agent in meat preservation. The application of 1,8-cineole had a significant suppressive impact on the growth rate of Listeria monocytogenes, Staphylococcus aureus, Escherichia coli, and Salmonella Typhimurium in meat samples stored for 7 days at 4 °C. Additionally, the surface color of the meat samples was not negatively impacted by the application of 1,8-cineole. The minimum inhibitory concentration was 12.5-25 mg/ml, and the minimum bactericidal concentration was 25-50.0 mg/ml. The bacterial cell membrane may be the target of cardamom, causing leakage of intracellular proteins, ATP, and DNA. The obtained data in this study may pave a new avenue for using 1,8-cineole as a new perspective for dealing with this problem of food-borne pathogens and food preservation, such as meat.

Green synthesis of zinc oxide nanoparticles using novel bacterium strain (Bacillus subtilis NH1-8) and their in vitro antibacterial and antibiofilm activities against Salmonellatyphimurium.

Zinc oxide nanoparticles (ZnO NPs) are used in a range of applications, including food packaging, preservation, and storage. In the current investigation, extracellular green synthesis of ZnO NPs through an simple, eco-friendly, and rapid approach using a novel bacterial strain (Bacillus subtilis NH1-8) was studied. To assess the morphological, optical, and structural properties of ZnO NPs, transmission electron microscopy (TEM), energy-dispersive X-ray (EDX), scanning electron microscopy (SEM), fourier transform infrared (FTIR) spectroscopy, ultraviolet-visible (UV-vis) spectroscopy, and X-ray diffraction (XRD) techniques were carried out. In addition, disk diffusion, minimum bactericidal concentration (MBC), and minimum inhibitory concentration (MIC) methods were performed to determine the antibacterial activity of ZnO NPs. The average size of biosynthesized ZnO NPs was 39 nm, exhibiting semi-spherical, which was confirmed by TEM analyses. The UV-vis spectroscopy exhibited the absorption peak at 200-800nm. The ZnO NPs have shown effective antimicrobial and antibiofilm activities against S. typhimurium. Thus, biosynthesized ZnO NPs could be exploited as a breakthrough technology in the surface coating of food containers and cans to minimize contamination by S. typhimurium.

Invasive listeriosis in Finland: surveillance and cluster investigations, 2011-2021.

Foodborne pathogen Listeria monocytogenes may cause serious, life-threatening disease in susceptible persons. We combined data from Finnish national listeriosis surveillance, patient interview responses, and laboratory data of patient samples and compared them to listeria findings from food and food production plants collected as part of outbreak investigations during 2011-2021. The incidence of invasive listeriosis in Finland (1.3/100000 in 2021) is higher than the EU average (0.5/100000 in 2021), and most cases are observed in the elderly with a predisposing condition. Many cases reported consuming high-risk foods as well as improper food storage. Since ongoing patient interviews and whole genome sequencing were introduced, several listeriosis outbreaks were detected and food sources identified. Recommendations about high-risk foods for listeriosis and proper food storage should be better communicated to susceptible people. In Finland, patient interviews and typing and comparing listeria isolates in foods and patient samples are crucial in solving outbreaks and determining measures to control invasive listeriosis.

Retrospective evaluation of an integrated molecular-epidemiological approach to cyclosporiasis outbreak investigations - United States, 2021.

Cyclosporiasis results from an infection of the small intestine by Cyclospora parasites after ingestion of contaminated food or water, often leading to gastrointestinal distress. Recent developments in temporally linking genetically related Cyclospora isolates demonstrated effectiveness in supporting epidemiological investigations. We used 'temporal-genetic clusters' (TGCs) to investigate reported cyclosporiasis cases in the United States during the 2021 peak-period (1 May - 31 August 2021). Our approach split 655 genotyped isolates into 55 genetic clusters and 31 TGCs. We linked two large multi-state epidemiological clusters (Epidemiologic Cluster 1 [n = 136 cases, 54 genotyped] and Epidemiologic Cluster 2 [n = 42 cases, 15 genotyped]) to consumption of lettuce varieties; however, product traceback did not identify a specific product for either cluster due to the lack of detailed product information. To evaluate the utility of TGCs, we performed a retrospective case study comparing investigation outcomes of outbreaks first detected using epidemiological methods with those of the same outbreaks had TGCs been used to first detect them. Our study results indicate that adjustments to routine epidemiological approaches could link additional cases to epidemiological clusters of cyclosporiasis. Overall, we show that CDC's integrated genotyping and epidemiological investigations provide valuable insights into cyclosporiasis outbreaks in the United States.

Fermenting a place in history: The first outbreak of Escherichia coli O157 associated with kimchi in Canada.

A Canadian outbreak investigation was initiated in January 2022 after a cluster of cases of Shiga-toxin-producing Escherichia coli (STEC) O157 was identified through whole genome sequencing (WGS). Exposure information was collected through case interviews. Traceback investigations were conducted, and samples from case homes, retail, and the manufacturer were tested for STEC O157. Fourteen cases were identified in two provinces in Western Canada, with isolates related by 0-5 whole genome multi-locus sequence typing allele differences. Symptom onset dates ranged from 11 December 2021 to 7 January 2022. The median age of cases was 29.5 (range 0-61); 64% were female. No hospitalisations or deaths were reported. Of 11 cases with information available on fermented vegetable exposures, 91% (10/11) reported consuming Kimchi Brand A during their exposure period. The traceback investigation identified Manufacturer A in Western Canada as the producer. One open and one closed sample of Kimchi Brand A tested positive for STEC O157, with isolates considered genetically related by WGS to the outbreak strain. Napa cabbage within the kimchi product was hypothesised as the most likely source of contamination. This paper summarises the investigation into this STEC O157 outbreak associated with kimchi, the first reported outside of East Asia.

Characteristics associated with successful foodborne outbreak investigations involving United States retail food establishments (2014-2016).

This study examined relationships between foodborne outbreak investigation characteristics, such as the epidemiological methods used, and the success of the investigation, as determined by whether the investigation identified an outbreak agent (i.e. pathogen), food item and contributing factor. This study used data from the Centers for Disease Control and Prevention's (CDC) National Outbreak Reporting System and National Environmental Assessment Reporting System to identify outbreak investigation characteristics associated with outbreak investigation success. We identified investigation characteristics that increase the probability of successful outbreak investigations: a rigorous epidemiology investigation method; a thorough environmental assessment, as measured by number of visits to complete the assessment; and the collection of clinical samples. This research highlights the importance of a comprehensive outbreak investigation, which includes epidemiology, environmental health and laboratory personnel working together to solve the outbreak.

A scoping review of factors associated with antimicrobial-resistant Campylobacter species infections in humans.

Human infection with antimicrobial-resistant Campylobacter species is an important public health concern due to the potentially increased severity of illness and risk of death. Our objective was to synthesise the knowledge of factors associated with human infections with antimicrobial-resistant strains of Campylobacter. This scoping review followed systematic methods, including a protocol developed a priori. Comprehensive literature searches were developed in consultation with a research librarian and performed in five primary and three grey literature databases. Criteria for inclusion were analytical and English-language publications investigating human infections with an antimicrobial-resistant (macrolides, tetracyclines, fluoroquinolones, and/or quinolones) Campylobacter that reported factors potentially linked with the infection. The primary and secondary screening were completed by two independent reviewers using Distiller SR®. The search identified 8,527 unique articles and included 27 articles in the review. Factors were broadly categorised into animal contact, prior antimicrobial use, participant characteristics, food consumption and handling, travel, underlying health conditions, and water consumption/exposure. Important factors linked to an increased risk of infection with a fluoroquinolone-resistant strain included foreign travel and prior antimicrobial use. Identifying consistent risk factors was challenging due to the heterogeneity of results, inconsistent analysis, and the lack of data in low- and middle-income countries, highlighting the need for future research.

Catering of high-risk foods and potential of stored food menu data for timely outbreak investigations in healthcare facilities, Italy and Germany.

Healthcare-associated foodborne outbreaks (HA-FBOs) can cause significant morbidity and mortality, affecting particularly vulnerable hospital populations. Electronic records of food served in healthcare facilities (HCFs) could be useful for timely investigations of HA-FBOs. We explored the availability and usability of electronic food menu data to support investigations of HA-FBOs through a survey among 35 HCFs in Germany (n = 13) and in Italy (n = 22). Large variability was reported in the storage time of menu data (from no storage up to 10 years) and their formats, including paper, electronic (PDF, Word, Excel), or fully searchable databases (15/22 in Italian HCFs, 3/13 in German HCFs). Food products that may present a risk to vulnerable persons - including deli salads, raw/fermented sausage products, soft cheese, smoked fish or frozen berries - were offered on the menu of all HCFs in Germany, and one-third of the Italian HCFs. The usability of electronic food menu data for the prevention or investigation of HA-FBOs may be suboptimal in a large number of HCFs in Germany, as well as in some HCFs in Italy. Standardised collection for use of electronic food menu data might help discover the association between illnesses and food eaten during outbreak investigations. Hospital hygienists, food safety and public health authorities should collaborate to increase implementation of food safety guidelines.

The antibiotic resistome in Escherichia coli isolated from human, food, and animal sources.

AIMS: The aim of this study was to analyze and compare the prevalence and distribution of resistance genes in Escherichia coli genomes isolated from human clinical samples and animal-based foods worldwide. METHODS AND RESULTS: We download from NCBI Pathogen Detection Database the corresponding metadata of the 7,123 E. coli genome to access the information about the antimicrobial resistance gene content. The geographic location and the source of isolation were also obtained and compiled with the antimicrobial resistance gene for statistical analysis, results and discussion. Our criteria considered four groups for analyzing the antimicrobial resistance gene distribution. The first group of genomes from invasive clinical human (ICH) samples from countries with Human Development Index (HDI) ≥ 0.850; the second group of ICH from countries with an HDI ≤ 0.849; the third group of animal-based foods (ABF) from countries with HDI ≥ 0.850 and the fourth group of ABFs from countries with HDI ≤ 0.849. The most prevalent genes in the first group were blaCTX-M-134 (96.53%) and blaCTX-M-27 (86.35%). In the second group, ere(A) (95.96%), soxS (94.49%), qepA8 (90.81%), blaCTX-M-15 (85.66%), and fosA3 (80.88%). In the third group, the most frequently detected were aadA12 (98.5%), ant(3") (89.92%), and blaCARB-2 (87.2%). In the fourth group, aadA12 and aac(3)-IV were identified in 100% of the analyzed genomes. CONCLUSIONS: It was clear that the use of aminoglycosides in animal production is increasing the selective pressure on micro-organisms in both groups of countries since genes linked to aminoglycoside resistance are related to E. coli from ABF samples. The genomic profile of E. coli from HDI ≥ 0.850 countries indicates a selective pressure aimed at cephalosporins given the high prevalence in both sources.

Nigella sativa-chitosan nanoparticles: Novel intestinal mucosal immunomodulator controls and protects against Salmonella enterica serovar Enteritidis infection in broilers.

BACKGROUND: Salmonella Enteritidis (SE) propagates in chickens' gastrointestinal surfaces and is transmitted to humans, causing food poisoning. Oral supplementation with natural nanoparticles can overcome the harsh gastrointestinal conditions facing oral vaccines and requires no antibiotic administration to protect against microbial infection. This study was designed to study Nigella sativa-chitosan nanoparticles (CNP-NS) prophylactic immunomodulatory efficacy against SE infection in broiler chicks. The CNP-NS was prepared and characterized, and its in vivo immunomodulatory activities against an avian virulent-MDR SE-induced challenge in chicks were investigated. RESULT: To verify the immune-protective activities of the CNP-NS, colony forming units (CFU) in the liver and fecal droppings; intestinal histopathological alterations and immune cell recruitment; MUC-2, TLR-4, cecal cytokines, and specific IgA gene expression levels were assessed. On the 7th and 12th days after the SE challenge, the CNP-NS supplemented chicks showed complete clearance of SE CFU in livers and fecal droppings, as well as an improvement in food conversion rate compared to non-supplemented CNP-NS that revealed the presence of the challenge SE CFU on the same days. A prominent influx of antigen presenting cells and lymphoid aggregates into the intestinal wall, spleen, and liver was detected with improvements in the intestinal villi morphometry of the CNP-NS-supplemented chicks. The changes of INF-γ, IL-1ß, and IL-4 cecal cytokines, as well as TLR-4, MUC-2, and IgA mRNA expression levels, confirm CNP-NS immunomodulatory activities and provide a mechanism(s) for its protective actions against the induced SE challenge of the tested chickens. CONCLUSION: These findings suggest promising useful insights into CNP-NS supplementation as a safe food additive for poultry meat consumers' and a protective immunomodulator of the chickens' mucosal immune systems. It could be recommended for epidemiological purposes to reduce the risk of SE food poisoning and transmission to humans.