{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,10,26]],"date-time":"2026-10-26T19:48:32Z","timestamp":1793044112997,"version":"4.5.0"},"reference-count":52,"publisher":"MDPI AG","issue":"10","license":[{"start":{"date-parts":[[2025,10,6]],"date-time":"2025-10-06T00:00:00Z","timestamp":1759708800000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"name":"Ongoing Research Funding","award":["ORF-2025-833"],"award-info":[{"award-number":["ORF-2025-833"]}]}],"content-domain":{"domain":["www.mdpi.com"],"crossmark-restriction":true},"short-container-title":["Veterinary Sciences"],"abstract":"<jats:p>This study evaluated the antibacterial efficacy and growth-promoting potential of Moringa oleifera and Thymus vulgaris methanolic extracts in broiler chickens challenged with Escherichia coli O78. In vitro antibacterial screening using agar well diffusion and disc diffusion assays revealed that ciprofloxacin exhibited the strongest inhibitory effect, followed by tetracycline and enrofloxacin, whereas among plant extracts, T. vulgaris was more effective than M. oleifera. The optimal combination (M100T150; 100 mg M. oleifera + 150 mg T. vulgaris) produced the largest inhibition zones against E. coli strains. For the in vivo trial, 540 Ross-308 broiler chicks were distributed into six treatment groups in a completely randomized design and reared for 42 days. Parameters assessed included growth performance, carcass traits, gut pH, ileal microbial counts, and intestinal histomorphology. Results showed that E. coli challenge significantly reduced feed intake, weight gain, carcass yield, and villus integrity while increasing FCR and E. coli counts (p &lt; 0.05). Addition of plant extracts, particularly M100T150, significantly improved weight gain, FCR, Broiler Performance Efficiency Factor (BPEF), and Broiler Farm Economy Index (BFEI) compared to the positive control (p &lt; 0.05). Extracts reduced duodenal and jejunal pH (p &lt; 0.001), suppressed E. coli counts (p = 0.003), and enhanced Lactobacillus populations (p = 0.0004). Histological analysis revealed that extract-supplemented groups had greater villus height and surface area with shallower crypts than the positive control, indicating restoration of gut integrity. These findings suggest that methanolic extracts of M. oleifera and T. vulgaris, particularly in combination, can serve as natural alternatives to antibiotics in broiler production under pathogenic challenge.<\/jats:p>","DOI":"10.3390\/vetsci12100957","type":"journal-article","created":{"date-parts":[[2025,10,6]],"date-time":"2025-10-06T15:05:06Z","timestamp":1759763106000},"page":"957","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":3,"title":["In Vitro Screening of Antibacterial Efficacy of Moringa oleifera and Thymus vulgaris Methanolic Extracts Against Different Escherichia coli Strains and Their In Vivo Effects Against E. coli-Induced Infection in Broiler Chickens"],"prefix":"10.3390","volume":"12","author":[{"ORCID":"https:\/\/orcid.org\/0000-0001-9783-6688","authenticated-orcid":false,"given":"Majid","family":"Ali","sequence":"first","affiliation":[{"name":"Department of Poultry Science, Faculty of Animal Husbandry and Veterinary Sciences, The University of Agriculture, Peshawar 25130, Pakistan"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Naila","family":"Chand","sequence":"additional","affiliation":[{"name":"Department of Poultry Science, Faculty of Animal Husbandry and Veterinary Sciences, The University of Agriculture, Peshawar 25130, Pakistan"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Sarzamin","family":"Khan","sequence":"additional","affiliation":[{"name":"Department of Poultry Science, Faculty of Animal Husbandry and Veterinary Sciences, The University of Agriculture, Peshawar 25130, Pakistan"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-0924-0479","authenticated-orcid":false,"given":"Rifat Ullah","family":"Khan","sequence":"additional","affiliation":[{"name":"Physiology Lab, College of Veterinary Sciences, Faculty of Animal Husbandry and Veterinary Sciences, The University of Agriculture, Peshawar 25130, Pakistan"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-8197-3119","authenticated-orcid":false,"given":"Babar","family":"Maqbool","sequence":"additional","affiliation":[{"name":"Department of Veterinary Medicine, Faculty of Veterinary & Animal Sciences, The University of Agriculture, Dera Ismail Khan 29163, Pakistan"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Shabana","family":"Naz","sequence":"additional","affiliation":[{"name":"Department of Zoology, Government College University, Faisalabad 38000, Pakistan"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-7998-0344","authenticated-orcid":false,"given":"Ala","family":"Abudabos","sequence":"additional","affiliation":[{"name":"Department of Food and Animal Sciences, College of Agriculture, Tennessee State University, Nashville, TN 37209, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Abdul","family":"Hafeez","sequence":"additional","affiliation":[{"name":"Department of Poultry Science, Faculty of Animal Husbandry and Veterinary Sciences, The University of Agriculture, Peshawar 25130, Pakistan"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-2547-4395","authenticated-orcid":false,"given":"Ibrahim A.","family":"Alhidary","sequence":"additional","affiliation":[{"name":"Department of Animal Production, College of Food and Agriculture Science, King Saud University, Riyadh 11451, Saudi Arabia"}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"1968","published-online":{"date-parts":[[2025,10,6]]},"reference":[{"key":"ref_1","first-page":"2335810","article-title":"In vitro screening of antibacterial efficacy of stem waste of oyster mushrooms (Pleurotus ostreatus) against different Escherichia coli strains and in vivo effects against E. coli-induced infection in Japanese quails","volume":"10","author":"Nasir","year":"2024","journal-title":"Cogent Food Agric."},{"key":"ref_2","doi-asserted-by":"crossref","first-page":"105416","DOI":"10.1016\/j.livsci.2024.105416","article-title":"In vitro and in vivo effects of methanolic extract of dietary ginger (Zingiber officinale) and onion (Allium cepa) addition on growth performance and fecal microbiota in Escherichia coli-infected broiler chickens","volume":"281","author":"Kausar","year":"2024","journal-title":"Livest. Sci."},{"key":"ref_3","first-page":"39","article-title":"The effect of halquinol (HAL) on growth performance, carcass traits, and blood-lipid profile in broiler chickens","volume":"13","author":"Habib","year":"2024","journal-title":"Int. J. Agric. Biosci."},{"key":"ref_4","doi-asserted-by":"crossref","first-page":"1","DOI":"10.47278\/journal.abr\/2024.006","article-title":"Impact of probiotic Saccharomyces cerevisiae var. boulardii RC009 alone and in combination with a phytase in broiler chickens fed with antibiotic-free diets","volume":"16","author":"Magnoli","year":"2024","journal-title":"Agrobiol. Rec."},{"key":"ref_5","first-page":"1243","article-title":"Impact of Bacillus subtilis probiotic on growth performance, bone health, intestinal morphology, and cecal microbiota in Cobb broiler chicks","volume":"44","author":"Yu","year":"2024","journal-title":"Pak. Vet. J."},{"key":"ref_6","unstructured":"Ajmal, A.S., Hussain, Z., Jalees, M.M., Shafi, J., Manzoor, S., and Haq, A.U. (2023). Performance of broiler birds on feeding natural anti-stressors in summer during heat stress. Asian J. Agric. Biol., 2023."},{"key":"ref_7","first-page":"306","article-title":"Broiler product quality: The global scientific research landscape and implications for marketing performance","volume":"13","author":"Dinasarki","year":"2024","journal-title":"Int. J. Agric. Biosci."},{"key":"ref_8","first-page":"333","article-title":"Impacts of probiotic and dietary Lamtoro leaf meal on the growth performance, digestibility and small intestinal morphometry of Kampung chicken","volume":"13","author":"Fathanah","year":"2024","journal-title":"Int. J. Agric. Biosci."},{"key":"ref_9","doi-asserted-by":"crossref","first-page":"79","DOI":"10.1080\/09712119.2019.1573731","article-title":"Anticoccidial effect of garlic (Allium sativum) and ginger (Zingiber officinale) against experimentally induced coccidiosis in broiler chickens","volume":"47","author":"Ali","year":"2019","journal-title":"J. Appl. Anim. Res."},{"key":"ref_10","first-page":"352","article-title":"Could Paenibacillus xylanexedens MS58 be an eco-friendly antibiotic in poultry production? Impacts on performance, blood biochemistry, gut microbiota and meat quality","volume":"44","author":"Dabool","year":"2024","journal-title":"Pak. Vet. J."},{"key":"ref_11","unstructured":"Nguyen, D.H., Ta, N.A.T., Van, H.G., Chu, D.T., Nguyen, T.S., Can, V.M., Nguyen, Q.U., and Vinh, H.V. (2024). Effects of the multi-strain probiotic preparation LabMix on some immune indices and intestinal microbiota in an antibiotic-associated diarrhea rat model. Asian J. Agric. Biol., 2024."},{"key":"ref_12","first-page":"744","article-title":"Valorization and optimization of protein in fermented palm kernel cake: Influence on broiler chicks growth","volume":"13","author":"Othman","year":"2024","journal-title":"Int. J. Agric. Biosci."},{"key":"ref_13","first-page":"222","article-title":"Phenolics of botanical origin for the control of coccidiosis in poultry","volume":"44","author":"Aljohani","year":"2024","journal-title":"Pak. Vet. J."},{"key":"ref_14","unstructured":"Malik, S., Sial, N., Shahzad, M.I., Anjum, S., Javid, A., and Rivera, G. (2024). Characterization and identification of bioactive natural products in the ethanol extracts of Acacia nilotica, Melia azedarach, and Euphorbia hirta from Cholistan desert, Pakistan. Asian J. Agric. Biol., 2024."},{"key":"ref_15","first-page":"89","article-title":"Phytogenic substances as safe growth promoters in poultry nutrition","volume":"12","author":"Mohamed","year":"2023","journal-title":"Int. J. Vet. Sci."},{"key":"ref_16","first-page":"75","article-title":"Performance and carcass characteristics of broiler chickens kept on heated soybean meal","volume":"17","author":"Nouri","year":"2024","journal-title":"Agrobiol. Rec."},{"key":"ref_17","doi-asserted-by":"crossref","first-page":"498","DOI":"10.47278\/journal.ijvs\/2022.216","article-title":"Metabolic response and meat quality of goats fed Artocarpus heterophyllus and Moringa oleifera","volume":"12","author":"Utari","year":"2023","journal-title":"Int. J. Vet. Sci."},{"key":"ref_18","unstructured":"Liang-liang, L., Kuan-hui, L., Shi-hui, X., Ting, L., Guang-shuang, Z., Almutairi, M.H., Zahid, M., Gondal, M.A., Qudratullah Mehmood, K., and Wu, Y. (2023). Immune enhancing activity of garlic polysaccharide, selenizing Codonopsis pilosula compounds in chickens. Asian J. Agric. Biol., 2023."},{"key":"ref_19","first-page":"757","article-title":"Evaluation of anticoccidial activity of ethanolic extract of clove in broiler chicken","volume":"44","author":"Aljohani","year":"2024","journal-title":"Pak. Vet. J."},{"key":"ref_20","first-page":"175","article-title":"Sweet orange (Citrus sinensis) peel powder with xylanase addition improved growth performance, antioxidant status, and immunity of broiler chickens","volume":"12","author":"Mohamed","year":"2023","journal-title":"Int. J. Vet. Sci."},{"key":"ref_21","unstructured":"Shaukat, N., Farooq, U., Akram, K., Shafi, A., Hayat, Z., Naz, A., Hakim, A., Hayat, K., Naseem, S., and Khan, M.Z. (2023). Antimicrobial potential of banana peel: A natural preservative to improve food safety. Asian J. Agric. Biol., 2023."},{"key":"ref_22","first-page":"1477","article-title":"Effect of different combinations of methanolic extract of Moringa oleifera and Moringa oleifera on production performance, gut morphology, hematology and nutrient digestibility in broilers","volume":"56","author":"Ali","year":"2023","journal-title":"Pak. J. Zool."},{"key":"ref_23","first-page":"333","article-title":"Spermatozoa quality of Kintamani dogs in coconut water-egg yolk diluent with addition of Moringa leaves and carrot extract","volume":"12","author":"Bebas","year":"2023","journal-title":"Int. J. Vet. Sci."},{"key":"ref_24","first-page":"980","article-title":"The impact of dietary Moringa oleifera leaf addition on stress markers, immune responses, and productivity in heat-stressed broilers","volume":"13","author":"Kamel","year":"2024","journal-title":"Int. J. Vet. Sci."},{"key":"ref_25","doi-asserted-by":"crossref","unstructured":"Khan, R.U., Khan, A., Naz, S., Ullah, Q., Laudadio, V., Tufarelli, V., and Ragni, M. (2021). Potential applications of Moringa oleifera in poultry health and production as alternative to antibiotics: A review. Antibiotics, 10.","DOI":"10.3390\/antibiotics10121540"},{"key":"ref_26","doi-asserted-by":"crossref","first-page":"1353","DOI":"10.1080\/10495398.2021.1899936","article-title":"Role of dietary Moringa oleifera leaf extract on productive parameters, humoral immunity and lipid peroxidation in broiler chicks","volume":"33","author":"Khan","year":"2022","journal-title":"Anim. Biotechnol."},{"key":"ref_27","first-page":"1329","article-title":"Effect of supplementing a blend of essential oils on the growth performance, carcass characteristics, meat quality, serological parameters and gut health in broiler chickens","volume":"44","author":"Yu","year":"2024","journal-title":"Pak. Vet. J."},{"key":"ref_28","first-page":"212","article-title":"Effects of oregano (Origanum vulgare L.) and rosemary (Rosmarinus officinalis L.) aqueous extracts on broiler performance, immune function and meat quality traits","volume":"15","author":"Franciosini","year":"2016","journal-title":"Ital. J. Anim. Sci."},{"key":"ref_29","first-page":"527","article-title":"Effect of three herbal extracts on growth performance, immune system, blood factors and intestinal selected bacterial population in broiler chickens","volume":"13","author":"Rahimi","year":"2011","journal-title":"J. Agric. Sci. Technol."},{"key":"ref_30","doi-asserted-by":"crossref","first-page":"585","DOI":"10.1515\/jbcpp-2014-0129","article-title":"Methanolic extract of Moringa oleifera leaves improves glucose tolerance, glycogen synthesis and lipid metabolism in alloxan-induced diabetic rats","volume":"26","author":"Olayaki","year":"2015","journal-title":"J. Basic Clin. Physiol. Pharmacol."},{"key":"ref_31","doi-asserted-by":"crossref","first-page":"181","DOI":"10.1016\/j.sajb.2019.05.005","article-title":"Effect of solvent evaporation method on phenolic compounds and the antioxidant activity of Moringa oleifera cultivated in Southern Tunisia","volume":"129","author":"Bennour","year":"2020","journal-title":"S. Afr. J. Bot."},{"key":"ref_32","doi-asserted-by":"crossref","first-page":"137","DOI":"10.5455\/egysebz.20180802111642","article-title":"In vitro evaluation of antibacterial activity of total protein extracts of some invertebrates against human pathogenic bacteria","volume":"14","author":"Mona","year":"2018","journal-title":"Egypt. J. Exp. Biol."},{"key":"ref_33","doi-asserted-by":"crossref","first-page":"280","DOI":"10.1086\/510431","article-title":"New consensus guidelines from the Clinical and Laboratory Standards Institute for antimicrobial susceptibility testing of infrequently isolated or fastidious bacteria","volume":"44","author":"Jorgensen","year":"2007","journal-title":"Clin. Infect. Dis."},{"key":"ref_34","doi-asserted-by":"crossref","first-page":"867","DOI":"10.1086\/511864","article-title":"Analysis and presentation of cumulative antibiograms: A new consensus guideline from the Clinical and Laboratory Standards Institute","volume":"44","author":"Hindler","year":"2007","journal-title":"Clin. Infect. Dis."},{"key":"ref_35","doi-asserted-by":"crossref","first-page":"339","DOI":"10.1590\/1806-9061-2016-0275","article-title":"Breed and sex differences in the gross anatomy, digesta pH and histomorphology of the gastrointestinal tract of Gallus gallus domesticus","volume":"19","author":"Mabelebele","year":"2017","journal-title":"Braz. J. Poult. Sci."},{"key":"ref_36","doi-asserted-by":"crossref","first-page":"21412","DOI":"10.1007\/s11356-017-9808-5","article-title":"The effect of corn distiller\u2019s dried grains with solubles (DDGS) fortified with enzyme on growth performance of broiler","volume":"24","author":"Abudabos","year":"2017","journal-title":"Environ. Sci. Pollut. Res."},{"key":"ref_37","first-page":"387","article-title":"Effect of dietary addition of Moringa oleifera leaves on growth performance, immune response and antioxidant status in broiler chickens","volume":"45","author":"Sultana","year":"2015","journal-title":"S. Afr. J. Anim. Sci."},{"key":"ref_38","doi-asserted-by":"crossref","first-page":"1","DOI":"10.1016\/j.anifeedsci.2010.03.007","article-title":"Essential oils in poultry nutrition: Main effects and modes of action","volume":"158","author":"Brenes","year":"2010","journal-title":"Anim. Feed. Sci. Technol."},{"key":"ref_39","doi-asserted-by":"crossref","first-page":"864","DOI":"10.1086\/381972","article-title":"Clinical relevance of bacteriostatic versus bactericidal mechanisms of action in the treatment of Gram-positive bacterial infections","volume":"38","author":"Pankey","year":"2004","journal-title":"Clin. Infect. Dis."},{"key":"ref_40","unstructured":"Saif, Y.M. (2008). Coccidiosis. Diseases of Poultry, Iowa State Press. [12th ed.]."},{"key":"ref_41","doi-asserted-by":"crossref","first-page":"450","DOI":"10.1080\/0007166031000085508","article-title":"Effects of dietary essential oil components on growth performance, digestive enzymes and lipid metabolism in female broiler chickens","volume":"44","author":"Lee","year":"2003","journal-title":"Br. Poult. Sci."},{"key":"ref_42","doi-asserted-by":"crossref","first-page":"255","DOI":"10.1016\/j.foodchem.2013.07.059","article-title":"Fatty acid composition and oxidative stability of breast meat from broilers supplemented with Moringa oleifera leaf meal over a period of refrigeration","volume":"142","author":"Nkukwana","year":"2014","journal-title":"Food Chem."},{"key":"ref_43","doi-asserted-by":"crossref","first-page":"379","DOI":"10.3923\/ijps.2014.379.384","article-title":"Effect of addition of Moringa oleifera leaf meal in broiler chicken feed","volume":"13","author":"Gakuya","year":"2014","journal-title":"Int. J. Poult. Sci."},{"key":"ref_44","doi-asserted-by":"crossref","first-page":"169","DOI":"10.1093\/ps\/83.2.169","article-title":"Influence of two plant extracts on broilers performance, digestibility, and digestive organ size","volume":"83","author":"Hernandez","year":"2004","journal-title":"Poult. Sci."},{"key":"ref_45","doi-asserted-by":"crossref","first-page":"555","DOI":"10.3382\/japr.2006-00116","article-title":"Effect of formic acid and plant extracts on growth, nutrient digestibility, intestine mucosa morphology, and meat yield of broilers","volume":"16","author":"Garcia","year":"2008","journal-title":"J. Appl. Poult. Res."},{"key":"ref_46","doi-asserted-by":"crossref","first-page":"496","DOI":"10.1080\/00071660701463221","article-title":"The effects of dietary inclusion of herbs and their volatile oils on the performance of chickens","volume":"48","author":"Cross","year":"2007","journal-title":"Br. Poult. Sci."},{"key":"ref_47","doi-asserted-by":"crossref","first-page":"626","DOI":"10.1111\/j.1365-2672.1994.tb01661.x","article-title":"Factors that interact with the antibacterial action of thyme essential oil and its active constituents","volume":"76","author":"Juven","year":"1994","journal-title":"J. Appl. Bacteriol."},{"key":"ref_48","doi-asserted-by":"crossref","first-page":"485","DOI":"10.1080\/00071660500191056","article-title":"Use of active substances of plant origin in chicken diets based on maize and locally grown cereals","volume":"46","author":"Jamroz","year":"2005","journal-title":"Br. Poult. Sci."},{"key":"ref_49","first-page":"2139","article-title":"Herbal additives and organic acids as antibiotic alternatives in broiler chickens diet for organic production","volume":"11","author":"Saki","year":"2014","journal-title":"Afr. J. Biotechnol."},{"key":"ref_50","first-page":"908","article-title":"Effect of dietary addition with Moringa oleifera leaves and\/or probiotics on performance, carcass traits and blood chemistry of broiler chickens","volume":"100","author":"Ibrahim","year":"2016","journal-title":"J. Anim. Physiol. Anim. Nutr."},{"key":"ref_51","first-page":"379","article-title":"Effects of thyme and cinnamon essential oils on growth performance, carcass traits and blood parameters of broilers","volume":"2","author":"Kalantar","year":"2011","journal-title":"Ann. Biol. Res."},{"key":"ref_52","first-page":"1065","article-title":"Effects of different levels of thyme (Moringa oleifera) powder on carcass traits, meat quality and blood parameters of broiler chickens","volume":"101","author":"Pournazari","year":"2017","journal-title":"J. Anim. Physiol. Anim. Nutr."}],"container-title":["Veterinary Sciences"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/www.mdpi.com\/2306-7381\/12\/10\/957\/pdf","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2025,10,6]],"date-time":"2025-10-06T15:33:06Z","timestamp":1759764786000},"score":1,"resource":{"primary":{"URL":"https:\/\/www.mdpi.com\/2306-7381\/12\/10\/957"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2025,10,6]]},"references-count":52,"journal-issue":{"issue":"10","published-online":{"date-parts":[[2025,10]]}},"alternative-id":["vetsci12100957"],"URL":"https:\/\/doi.org\/10.3390\/vetsci12100957","relation":{},"ISSN":["2306-7381"],"issn-type":[{"value":"2306-7381","type":"electronic"}],"subject":[],"published":{"date-parts":[[2025,10,6]]}}}