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Aims: The present study aimed to detect the frequency of vancomycin resistance and virulence genes` profiles of multi-drug resistant (MDR) enterococcal isolates from different sources and to investigate the sequence heterogeneity between the esp genes of MDR and vancomycin-resistant E. faecalis isolates from chicken and human sources., Methods and Results: Conventional phenotypic methods identified 91 isolates (60.7%) as Enterococcus species and these isolates were retrieved from dairy (37/52), chicken (35/54), and human (19/44) origins. Enterococcal isolates were frequently resistant to rifampin (67%), and 38.5% of the isolates were MDR. Of the 22 vancomycin-resistant enterococci (VRE) detected isolates, 11 (50%), nine (41%), one (4.5%), and one (4.5%) isolate were identified as E. faecium, E. faecalis, E. casseliflavus, and un-specified Enterococcus spp., respectively. Moreover, 22 (100%) and 19 (86.4%) isolates harbored vanA and vanB genes, respectively. Of note, gelE and asa1 genes were more prevalent among the tested isolates (95.5% each), and the multi-virulence criteria were detected among 68.2% of the examined isolates. The sequences of esp genes of E. faecalis from the chicken breast meat and human urine samples were 100% identical with other esp genes and pathogenicity islands on GeneBank, which is undesirable., Conclusion: Our findings require strict hygienic measures during the processing of chickens, and their by-products to minimize the possibility of transmission of virulent enterococcal strains. Furthermore, the use of antimicrobials in poultry and animal production in developing countries should be controlled to minimize the prevalence of MDR and VRE isolates in humans., (© The Author(s) 2025. Published by Oxford University Press on behalf of Applied Microbiology International.)

The increasing recognition of the potential advantages beyond nanoencapsulation of probiotics gained great attention owing to effective properties. Hence, we provided the most in-depth look into the influence of nanoformulated multi strain probiotics; BLB-NPs comprising Bacillus subtilis ATCC19659, Lactobacillus plantarum ATCC8014 and Bifidobacterium bifidum ATCC29521 on growth performance, antioxidant status and intestinal immunity supporting the defense against Salmonella Typhimurium (S. Typhimurium) challenge in broilers chickens. A total of 2,800 one-day-old male Ross 308 boiler chicks were divided into 7 groups; 1 control without additives, 3 probiotics [fed control diets mixed with B. subtilis, L. plantarum and B. bifidum (BLB) at concentrations of 1 × 10 4 (BLBI), 1 × 10 6 (BLBII) and 1 × 10 8 (BLBIII) CFU /kg diet, respectively] and 3 nanoencapsulated probiotics [fed control diets supplemented with BLB loaded nanoparticles (BLB-NPs) at concentrations of 1 × 10 4 (BLB-NPsI), 1 × 10 6 (BLB-NPsII) and 1 × 10 8 (BLB-NPsIII) CFU /kg diet, respectively]. All previous groups were challenged at d 22 of age with S. Typhimurium. Birds fed BLB-NPs II and III exhibited better weight gain and FCR simultaneously with upregulation in nutrients transporters genes (LAT-1, PepT-1, CAT-1 and SGLT1) even after S. Typhimurium challenge. Upregulation of immmune related genes (IL-1β, IL-6, IL-8, MyD88, NF-kB, CCL20, CXCLi2, TLR-2, TLR-4 and SOCS1) was prominently subsided in BLB-NPsIII fed group. The strengthening ability of BLB-NPs for broilers' intestinal barriers was evidenced by augmented expression of JAM, MUC-2, occludin and FABP-2 genes, diminished S. Typhimurium counts and suppressed its virulence related genes (HilA and SopD) with restored histopathological pictures of cecum. Notably, post dietary inclusion of higher levels of BLB-NPsIII, the abundance of beneficial Biofidobacterium and Lactobacillus species was dominated over harmful E. coli ones. Birds fortified with BLB-NPs displayed potent antioxidant potential signified by boosting serum and intestinal antioxidant markers alongside reducing oxidative ones. Overall, the abovementioned positive outcomes of BLB-NPs encouraged their potential application in poultry feed to attain superior performance and elicit protective immunity against S. Typhimurium infection., Competing Interests: DISCLOSURES The authors declare no conflicts of interest., (Copyright © 2024 The Authors. Published by Elsevier Inc. All rights reserved.)

Smart innovative nanocomposites based on active ingredients and metallic nanoparticles with effective wound healing and antifungal properties are efficient in overcoming the limitations of traditional therapeutic products. Open wounds provide an ideal niche for colonization by Candida albicans ( C. albicans ) which poses substantial global health issues owing to delayed wound healing and disordered healing mechanisms. Therefore, proficient innovative therapies that control C. albicans infection and promote wound healing are of imperative importance for the management of wounds and prevention of infection and possible complications. This study aims to design a novel nanocarrier platform based on a hydrogel loaded with silver nanoparticles (AgNPs) and doped with pomegranate peel extract (PPE) and hyaluronic acid (HA), offering an unprecedented opportunity to achieve skin repair and manage C. albicans colonization with an efficient wound healing process. Sprague-Dawley rats (n=100) were assigned to 5 groups and infected with C. albicans and distributed as follows: control positive (untreated) and four cutaneous wound-healing model groups treated topically with commercial cream and PPE-HA-AgNPs at full, 50%, and 25% concentrations for 15 days, respectively. Our findings revealed that the severity of clinical signs, C. albicans burden, and the expression of biofilm-related genes ALS1 , HYR1 , and PLB1 were diminished following treatment with PPE-HA-AgNPsIII. Notably, the formulated nanocomposite was very effective in extending the release of PPE-HA-AgNPs in infected wounds with retention percentages of 65.4% for PPE-HA-AgNPsIII. Topical administration of PPE-HA-AgNPsIII successfully alleviated the extensive inflammatory response and healed wounded skin via downregulation of tumor necrosis factor-alpha (TNF-α), interleukin-6 and IL-1 beta, and nitric oxide synthase (NOS) levels as shown by enzyme-linked immunosorbent (ELISA) and reverse transcription-quantitative polymerase chain reaction (RT-qPCR) assays. Interestingly, PPE-HA-AgNPsIII modulated angiogenic and wound healing markers as evidenced by the downregulation of MMP-9 and the upregulation of angiopoietin-1 (Ang-1), vascular endothelial growth factor (VEGF) (up to 10 days post-treatment), transforming growth factor-beta 1 (TGF-β1), bFGF , EGF, Ki- 67, and collagen I and III with efficient wound closure capability. This was evidenced by the lessening of histopathological severity, which accelerated the healing of the infected skin wounds post-treatment with PPE-HA-AgNPs. Overall, our formulated PPE-HA-AgNPs provide an effective innovative therapeutic strategy for the treatment of cutaneous wounds infected with C. albicans with maximized wound healing efficacy, indicating their potential in clinical practice., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2024 El-Hamid, Ibrahim, Abdelfattah-Hassan, Mohammed, Pet, Khalil, El-Badry, Metwally, Azouz, Elnegiry, Elnahriry, Ahmadi and Elazab.)

Aims: This study aimed to explore the effectiveness of dietary citronellol, thymol, and trans-cinnamaldehyde (CTC) essential oils blend on broilers' growth performance, immunity, intestinal microbial count, gut integrity, and resistance against Clostridium perfringens utilizing the necrotic enteritis (NE) challenge model., Methods and Results: A total of 200 Ross 308 male broiler chicks received either a control diet or diet supplemented with three graded levels of CTC blend, including 300, 600, and 900 mg of CTC blend/kg diet and experimentally infected with C. perfringens strain at 23 days of age. Herein, dietary CTC blend fortifications significantly improved the broilers' growth performance, which was supported by upregulating the expression levels of MUC-2, occludin, and JAM-2 genes. Moreover, dietary CTC blend inclusion significantly enhanced the levels of blood phagocytic percentage and serum IgA, IgG, and MPO, and reduced the values of serum CRP, and NO at 5 days pre-infection, 10-, and 15 days post-infection (dpi) with C. perfringens. At 15 dpi, CTC blend inclusion significantly reduced the intestinal digesta pH, coliforms and C. perfringens loads, and the expression levels of genes related to C. perfringens virulence (cpe, cnaA, and nanI), proinflammatory cytokines (IL-1β and TNF-α), and chemokines (CCL20), in addition to increasing the count of beneficial total Lactobacillus and total aerobic bacteria, and the expression levels of genes related to anti-inflammatory cytokines (IL-10) and chemokines (AvBD6 and AvBD612)., Conclusion: Our results point to the growth-provoking, immunostimulant, antibacterial, anti-inflammatory, and antivirulence characteristics of the CTC blend, which improves the broilers' resistance to C. perfringens and ameliorates the negative impacts of NE., (© The Author(s) 2024. Published by Oxford University Press on behalf of Applied Microbiology International.)

Encapsulated phytochemicals with augmented therapeutic and nutritional characteristics have become promising alternatives to antimicrobials in the poultry industry. Hence, our key target was to explore the efficacy of liposomal encapsulation, as a novel carrier, for essential oils (LEOs) on growth, digestibility, intestinal microbiota, and bacterial metabolites of broiler chickens. Moreover, the impact of encapsulated EOs on transcription mechanisms targeting the genes encoding digestive enzymes, gut barrier functions and antioxidant potential of broiler chickens was evidenced. Four equal broiler groups were fed 4 basal diets fortified with LEOs (oregano, cinnamon, and clove) at the levels of 0, 200, 300, and 400 mg/kg diet. Our findings revealed significant improvement in body weight gain and feed conversion ratio of birds fed higher levels of LEOs. These results came concurrently with increasing the activities of digestive enzymes at both serum and molecular levels and consequently nutrient digestibility (dry matter, ether extract, crude protein, and crude fiber) in these groups. Remarkably, the abundance of beneficial bacteria as well as the bacterial metabolites (valeric acid, butyric acid, propionic acid, acetic acid, and total short-chain fatty acids) was increased, while that of pathogenic ones was reduced following dietary inclusion of LEOs. Of note, the mRNA expression of genes encoding antioxidant stability [catalase (CAT), superoxide dismutase 1 (SOD-1), glutathione peroxidase 1 (GPX-1), nuclear factor erythroid 2-related factor 2 (NRF2), NAD(P)H dehydrogenase quinone 1 (NQO1), and heme oxygenase-1 (HO-1)] as well as barrier functions [mucin-2 (MUC-2)] and tight junction proteins, TJP [junctional adhesion molecule-2 (JAM-2) and occludin] were noticeably upregulated in broilers fortified with 400 mg/kg diet of LEOs. Overall, the present work recommended dietary inclusion of LEOs as beneficial additives for attaining targeted performance, gut health and antioxidant stability in poultry farming., Competing Interests: DISCLOSURES The authors declare no conflicts of interest., (Copyright © 2023 The Authors. Published by Elsevier Inc. All rights reserved.)

Probiotics as novel antibiotics' substitutes are verified to provide barriers for hindering the colonization of enteric bacterial pathogens with nutritional benefits. For enhancement of the probiotics' effectiveness, their integration within nanomaterials is a paramount tool to support the progress of new compounds with functional features. Therefore, we addressed the impact of effective delivery of probiotics ( Bacillus amyloliquefaciens ) loaded nanoparticles (BNPs) on performance and Campylobacter jejuni ( C. jejuni ) shedding and colonization in poultry. Two hundred Ross broiler chickens were divided into four groups fed various BNP levels: BNPs I, BNPs II, BNPs III, and BNPs-free diets for 35 days. Nanoparticles delivery of probiotics within broiler diets improved growth performance as reflected by higher body weight gain and superior feed conversion ratio, especially in BNPs II- and BNPs III-fed groups. In parallel, the mRNA expression levels of digestive enzymes encoding genes ( AMY2a , PNLIP , CELA1 , and CCK ) achieved their peaks in BNPs III-fed group (1.69, 1.49, 1.33, and 1.29-fold change, respectively) versus the control one. Notably, with increasing the levels of BNPs, the abundance of beneficial microbiota, such as Bifidobacterium and Lactobacillus species, was favored over harmful ones, including Clostridium species and Enterobacteriaceae . Birds fed higher levels of BNPs displayed significant improvement in the expression of barrier functions-linked genes including DEFB1 , FABP-2 , and MUC-2 alongside substantial reduction in cecal colonization and fecal shedding of C. jejuni . From the aforementioned positive effects of BNPs, we concluded their potential roles as growth promoters and effective preventive aids for C. jejuni infection in poultry.

Plant-derived bioactive compounds with promising nutritional and therapeutic attributes (phytogenics) are among the top priorities in the aquaculture sector. Therefore, the impact of thymol (Thy) and/or thymoquinone (ThQ) on the growth, immune response antioxidant capacity, and Aeromonas sobria ( A. sobria ) resistance of Nile tilapia was investigated. Four fish groups were fed a control diet and three basal diets supplemented with 200 mg/kg diet of Thy or ThQ and a blend of both Thy and ThQ at a level of 200 mg/kg diet each. At the end of the feeding trial (12 weeks), the tilapias were challenged intraperitoneally with virulent A. sobria (2.5 × 10 8 CFU/mL) harboring aerolysin ( aero ) and hemolysin ( hly ) genes. The results revealed that tilapias fed diets fortified with a combination of Thy and ThQ displayed significantly enhanced growth rate and feed conversion ratio. Notably, the expression of the genes encoding digestive enzymes (pepsinogen, chymotrypsinogen, α-amylase and lipase) and muscle and intestinal antioxidant enzymes (glutathione peroxidase, catalase and superoxide dismutase) was significantly upregulated in Thy/ThQ-fed fish. An excessive inflammatory response was subsided more prominently in the group administrated Thy/ThQ as supported by the downregulation of il-β , il-6 and il-8 genes and in contrast, the upregulation of the anti-inflammatory il-10 gene. Remarkably, dietary inclusion of Thy/ThQ augmented the expression of autophagy-related genes, whilst it downregulated that of mtor gene improving the autophagy process. Furthermore, Thy/ThQ protective effect against A. sobria was evidenced via downregulating the expression of its aero and hly virulence genes with higher fish survival rates. Overall, the current study encouraged the inclusion of Thy/ThQ in fish diets to boost their growth rates, promote digestive and antioxidant genes expression, improve their immune responses and provide defense against A. sorbia infections with great 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, H 2 O 2 , 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 log 10 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., Competing Interests: The authors declare no conflict of interest.

Weaning is the most crucial period associated with increased stress and susceptibility to diseases in rabbits. Methicillin-resistant Staphylococcus aureus (MRSA), a historic emergent pathogen related to post weaning stressors, adversely affects rabbit's growth rate and productive cycle. Since MRSA is rapidly evolving antibiotics resistance, natural products are desperately required to tackle the public health threats posed by antimicrobial resistance. Thus, this study aimed to screen the iin vitro antibacterial activity of Nigella sativa extract (NSE) and its interactions with antibiotics against MRSA isolates. Moreover, 200 weaned rabbits were divided into 4 groups to investigate the iin vivo superiority of NSE graded levels towards growth performance, tight junction integrity, immune responsiveness and resistance against MRSA. Herein, NSE showed promising antimicrobial activities against MRSA isolates from animal (77.8%) and human (64.3%) origins. Additionally, MRSA isolates exposed to NSE became sensitive to all antimicrobials to which they were previously resistant. Our results described that the growth-promoting functions of NSE, especially at higher levels, were supported by elevated activities of digestive linked enzymes. Post-NSE feeding, rabbits' sera mediated bactericidal activities against MRSA. Notably, upregulated expression of occludin, CLDN-1 , MUC-2 and JAM-2 genes was noted post NSE supplementation with maximum transcriptional levels in 500 mg/kg NSE fed group. Our data described that NSE constitutively motivated rabbits' immune responses and protected them against MRSA-induced experimental infection. Our results suggest the antimicrobial, growth stimulating and immunomodulation activities of NSE to maximize the capability of rabbits for disease response.

Respiratory diseases inflicted by Mycoplasma gallisepticum (MG) and Ornithobacterium rhinotracheale (ORT) cause severe economic losses and great burden to the poultry industry worldwide. Therefore, the current study was planned to assess the efficacy of aivlosin alone or in combination with zinc oxide nanoparticles (ZnO-NPs) in the treatment of experimental MG and/or ORT infections in broilers. Moreover, we also aimed to evaluate the role of ZnO-NPs on aivlosin residues in broiler tissues. A total of 1,440 Cobb chicks were allocated into 6 groups. At 14 d of age, chickens of groups 1 and 3 were experimentally infected with MG intratracheally and 6 d later, chickens of groups 2 and 3 were infected occulonasaly with ORT. Groups 1, 2, and 3 were divided into 4 subgroups; birds in subgroups 1, 2, and 3 were treated with aivlosin (A), ZnO-NPs (Z), and A/Z, respectively, while those in subgroups 4 was left without treatments. Moreover, groups 4 and 5 were kept noninfected and treated with aivlosin alone or in combination with ZnO-NPs, respectively. Finally, group 6 was kept as a negative control. The current results showed that the recovery from respiratory diseases caused by MG and/or ORT infections was most successful after treatment with A/Z in combination. Consequently, clinical signs, mortality rates, postmortem lesions of the respiratory organs, histopathological lesions of liver, trachea and lung and tracheal MG and ORT counts were significantly (P < 0.05) reduced following A/Z treatment. Taken together, high performance liquid chromatography analysis revealed that ZnO-NPs decreased the aivlosin residues in liver, muscle and skin of healthy and infected chickens. Based on these results, it could be concluded that aivlosin/ZnO-NPs therapy is a valuable approach for controlling MG and/or ORT infections in boilers., (Copyright © 2022 The Authors. Published by Elsevier Inc. All rights reserved.)

Recently, the concept of incorporating natural products into nanocarriers has been intended to promote fish growth and health via modulating their stability and bioavailability. In this concern, the potential role of reformulated quercetin into nanocarriers was examined, for the first time, on Nile tilapia's performance and immunity, flesh quality and antioxidant indices and disease resistance. Five hundred fish assigned into five experimental groups with formulated diets containing quercetin nanoparticles (QT-NPs) at levels of 0, 100, 200, 300 and 400 mg/kg were challenged with Aeromonas hydrophila (A. hydrophila) after 12 weeks feeding trial. Fish final body weight gain and feed efficiency were significantly maximized in groups enriched with 300 and 400 mg/kg of QT-NPs. Significant reduction in total saturated fatty acids and an elevation in polyunsaturated fatty acids' contents were noticed in fish fed higher QT-NPs doses. The levels of Hydrogen peroxide, reactive oxygen species and malondialdehyde, the markers of meat antioxidant capacity, were reduced by higher inclusion levels of QT-NPs. Accordingly, serum activities and transcriptional levels of GSH-Px, CAT and SOD genes were increased with elevated QT-NPs levels. Immune responses mediated by upregulation of IL-10 and TGF-β and downregulation of IL-1β, IL-8 and TNF-α mRNA levels were found to be positively affected by QT-NPs. Dietary QT-NPs downregulated the expression of ahyI and ahyR quorum sensing genes conferring protection against A. hydrophila challenge. This study concluded that supplementation of quercetin in encapsulated nanoparticles could improve its efficacy making it as a compelling approach to improve fish performance and as a promising drug candidate against A. hydrophila virulence., (Copyright © 2021 Elsevier Ltd. All rights reserved.)

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.