The present study involved in vivo evaluation of the growth promoting effects of thymol and thymol nanoemulsion and their protection against Salmonella Typhimurium infection in broilers. One-day old 2400 chicks were randomly divided into eight groups; negative and positive control groups fed basal diet without additives and thymol and thymol nanoemulsion groups (0.25, 0.5 and 1% each). At d 23, all chicks except negative control were challenged with S. Typhimurium. Over the total growing period, birds fed 1% thymol nanoemulsion showed better growth performance even after S. Typhimurium challenge, which came parallel with upregulation of digestive enzyme genes (AMY2A, PNLIP and CCK). Additionally, higher levels of thymol nanoemulsion upregulated the expression of MUC-2, FABP2, IL-10, IgA and tight junction proteins genes and downregulated IL-2 and IL-6 genes expression. Moreover, 1% thymol nanoemulsion, and to lesser extent 0.5% thymol nanoemulsion and 1% thymol, corrected the histological alterations of cecum and liver postinfection. Finally, supplementation of 1% thymol, 0.5 and 1% thymol nanoemulsion led to increased Lactobacilli counts and decreased S. Typhimurium populations and downregulated invA gene expression postinfection. This first report of supplying thymol nanoemulsion in broiler diets proved that 1% nano-thymol is a potential growth promoting and antibacterial agent.

Multidrug resistant (MDR) methicillin-resistant Staphylococcus aureus (MRSA) is a superbug pathogen that causes serious diseases. One of the main reasons for the lack of the effectiveness of antibiotic therapy against infections caused by this resistant pathogen is the recalcitrant nature of MRSA biofilms, which results in an increasingly serious situation worldwide. Consequently, the development of innovative biofilm inhibitors is urgently needed to control the biofilm formation by this pathogen. In this work, we thus sought to evaluate the biofilm inhibiting ability of some promising antibiofilm agents such as zinc oxide nanoparticles (Zno NPs), proteinase K, and hamamelitannin (HAM) in managing the MRSA biofilms. Different phenotypic and genotypic methods were used to identify the biofilm producing MDR MRSA isolates and the antibiofilm/antimicrobial activities of the used promising agents. Our study demonstrated strong antibiofilm activities of ZnO NPs, proteinase K, and HAM against MRSA biofilms along with their transcriptional modulation of biofilm (intercellular adhesion A, icaA) and quorum sensing (QS) (agr) genes. Interestingly, only ZnO NPs showed a powerful antimicrobial activity against this pathogen. Collectively, we observed overall positive correlations between the biofilm production and the antimicrobial resistance/agr genotypes II and IV. Meanwhile, there was no significant correlation between the toxin genes and the biofilm production. The ZnO NPs were recommended to be used alone as potent antimicrobial and antibiofilm agents against MDR MRSA and their biofilm-associated diseases. On the other hand, proteinase-K and HAM can be co-administrated with other antimicrobial agents to manage such types of infections.

Foodborne pathogens have been associated with severe and complicated diseases. Therefore, these types of infections are a concern for public health officials and food and dairy industries. Regarding the wide-spread multidrug resistant (MDR) and extensively drug resistant (XDR) foodborne pathogens such as Salmonella Enteritidis (S. Enteritidis), new and alternative therapeutic approaches are urgently needed. Therefore, we investigated the antimicrobial, anti-virulence, and immunostimulant activities of a stable formulation of thymol as thymol nanoemulsion in an in vivo approach. Notably, treatment with 2.25% thymol nanoemulsion led to a pronounced improvement in the body weight gain and feed conversion ratio in addition to decreases in the severity of clinical findings and mortality percentages of challenged chickens with XDR S. Enteritidis confirming its pronounced antimicrobial activities. Moreover, thymol nanoemulsion, at this dose, had protective effects through up-regulation of the protective cytokines and down-regulation of XDR S. Enteritidis sopB virulence gene and interleukins (IL)-4 and IL-10 cytokines as those hinder the host defenses. Furthermore, it enhanced the growth of gut Bifidobacteria species, which increases the strength of the immune system. For that, we suggested the therapeutic use of thymol nanoemulsion against resistant foodborne pathogens. Finally, we recommended the use of 2.25% thymol nanoemulsion as a feed additive for immunocompromised individuals as well as in the veterinary fields.

Several food-poisoning outbreaks have been attributed to Clostridium perfringens (C. perfringens) worldwide. Despite that, this crisis was discussed in a few studies, and additional studies are urgently needed in this field. Therefore, we sought to highlight the prevalence, antimicrobial resistance, toxin profiles, and toxinotypes of C. perfringens isolates. In this study, 50 C. perfringens isolates obtained from 450 different animal origin samples (beef, chicken meat, and raw milk) were identified by phenotypic and genotypic methods. The antimicrobial susceptibility results were surprising, as most of the isolates (74%) showed multidrug-resistant (MDR) patterns. The phenotypic resistance to tetracycline, lincomycin, enrofloxacin, cefoxitin/ampicillin, and erythromycin was confirmed by the PCR detections of tet, lnu, qnr, bla, and erm(B) genes, respectively. In contrast to the toxinotypes C and E, toxinotype A prevailed (54%) among our isolates. Additionally, we found that the genes for C. perfringens enterotoxin (cpe) and C. perfringens beta2 toxin (cpb2) were distributed among the tested isolates with high prevalence rates (70 and 64%, respectively). Our findings confirmed that the C. perfringens foodborne crisis has been worsened by the evolution of MDR strains, which became the prominent phenotypes. Furthermore, we were not able to obtain a fixed association between the toxinotypes and antimicrobial resistance patterns.

Necrotic enteritis (NE) caused by Clostridium perfringens (C. perfringens) results in impaired bird growth performance and increased production costs. Nanotechnology application in the poultry industry to control NE outbreaks is still not completely clarified. Therefore, the efficacy of dietary garlic nano-hydrogel (G-NHG) on broilers growth performance, intestinal integrity, economic returns and its potency to alleviate C. perfringens levels using NE challenge model were addressed. A total of 1200 male broiler chicks (Ross 308) were assigned into six groups; four supplemented with 100, 200, 300 or 400 mg of G-NHG/kg diet and co-challenged with C. perfringens at 21, 22 and 23 d of age and two control groups fed basal diet with or without C. perfringens challenge. Over the total growing period, the 400 mg/kg G-NHG group had the most improved body weight gain and feed conversion efficiency regardless of challenge. Parallel with these results, the mRNA expression of genes encoding digestive enzymes (alpha 2A amylase (AMY2A), pancreatic lipase (PNLIP) and cholecystokinin (CCK)) and intestinal barriers (junctional adhesion molecule-2 (JAM-2), occludin and mucin-2 (Muc-2)) were increased in groups fed G-NHG at higher levels to be nearly similar to those in the unchallenged group. At 14 d post challenge, real-time PCR results revealed that inclusion of G-NHG led to a dose-dependently decrease in the C. perfringens population, thereby decreasing the birds’ intestinal lesion score and mortality rates. Using 400 mg/kg of G-NHG remarkably ameliorated the adverse effects of NE caused by C. perfringens challenge, which contributed to better growth performance of challenged birds with rational economic benefits.

The oxidative stress facing fish during intensive production brings about diseases and mortalities that negatively influence their performance. Along with that, the increased awareness of omega-3 polyunsaturated fatty acids (omega-3-PUFAs) health benefits has been triggered the introduction of alternative additives in aqua feed that cause not only modulation in fish immune response but also fortification of their fillet. In this context, the role of microalgae mix (NSS) containing Nannochloropsis oculate and Schizochytrium and Spirulina species, which were enriched with bioactive molecules, especially EPA and DHA, was assessed on Nile tilapia’s performance, fillet antioxidant stability, immune response, and disease resistance. Varying levels of NSS (0.75%, 1.5%, and 3%) were added to Nile tilapia’s diet for 12 weeks and then a challenge of fish with virulent Aeromonas hydrophila (A. hydrophila) was carried out. Results showed that groups fed NSS, especially at higher levels, showed an improved WG and FCR, which corresponded with enhanced digestive enzymes’ activities. Higher T-AOC was detected in muscle tissues of NSS_3.0% fed fish with remarkable reduction in ROS, H₂O₂, and MDA contents, which came in parallel with upregulation of GSH-Px, CAT, and SOD genes. Notably, the contents of EPA and DHA in fillet were significantly increased with increasing the NSS levels. The mean log₁₀ counts of pathogenic Vibrio and Staphylococcus species were reduced, and conversely, the populations of beneficial Lactobacillus and Bacillus species were increased more eminent after supplementation of NSS_3.0% and NSS_1.5%. Moreover, regulation of the immune response (lysozyme, IgM, ACH50, NO, and MPO), upregulation of IL-10, TGF-β, and IgM, and downregulation of IL-1β, TNF-α, HSP70, and COX-2 were observed following dietary higher NSS levels. After challenge, reduction in A. hydrophila counts was more prominent, especially in NSS_3.0% supplemented group. Taken together, the current study encourages the incorporation of such microalgae mix in Nile tilapia’s diet for targeting maximum performance, superior fillet quality, and protection against A. hydrophila.