Processing-Associated Taxonomic Profiles of Microbiota in Liquid and Powdered Horse Milk Revealed by Full-Length 16S rRNA Sequencing
Abstract
Horse milk contains diverse microbial communities whose composition may be altered by processing and the addition of carrier ingredients. This study evaluated the effects of milk form and dextrin supplementation on the microbiota of horse milk prepared as pasteurized liquid milk (SKC), powdered milk supplemented with 15% dextrin (SKD), and powdered milk without dextrin (SKT). Microbial diversity and composition were analyzed using full-length 16S rRNA gene sequencing covering the V1–V9 regions. SKT showed the highest microbial richness, with 1185 observed taxa and a Shannon index of 3.43 ± 0.23, compared with 963 taxa and 2.40 ± 0.99 in SKD and 190 taxa and 2.02 ± 0.06 in SKC. SKC and SKD were dominated by lactic acid bacteria, particularly Lactobacillus gallinarum and Lactobacillus helveticus, whereas SKT contained higher proportions of stress-tolerant and spore-forming taxa such as Brevibacillus parabrevis (26%), Thalassoporum komareki (12%), and Lysinibacillus boronitolerans (10%). Sankey diagrams and principal component analysis further demonstrated distinct microbial community structures among the treatments. Overall, dextrin supplementation appeared to act as a protective carrier during drying, helping maintain the predominance of lactic acid bacteria, whereas powdered milk produced without dextrin was associated with a broader microbial profile.
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References
Abdul Hakim, B. N., Xuan, N. J., & Oslan, S. N. H. (2023). A comprehensive review of bioactive compounds from lactic acid bacteria: potential functions as functional food in dietetics and the food industry. Foods, 12(15), 2850. https://doi.org/10.3390/foods12152850
Bahram, M., Anslan, S., Hildebrand, F., Bork, P., & Tedersoo, L. (2019). Newly designed 16S rRNA metabarcoding primers amplify diverse and novel archaeal taxa from the environment. Environmental Microbiology Reports, 11(4), 487–494. https://doi.org/10.1111/1758-2229.12684
Bamidele, O. P., & Emmambux, M. N. (2021). Encapsulation of bioactive compounds by “extrusion” technologies: A review. Critical Reviews in Food Science and Nutrition, 61(18), 3100–3118. https://doi.org/10.1080/10408398.2020.1793724
Bao, W., He, Y., Yu, J., Yang, X., Liu, M., & Ji, R. (2022). Diversity analysis and gene function prediction of bacteria and fungi of Bactrian camel milk and naturally fermented camel milk from Alxa in Inner Mongolia. LWT, 169, 114001. https://doi.org/10.1016/j.lwt.2022.114001
Blanco-Doval, A., Barron, L. J. R., & Aldai, N. (2024). Nutritional quality and socio-ecological benefits of mare milk produced under grazing management. Foods, 13(9), 1412. https://doi.org/10.3390/foods13091412
Boranbayeva, T., Karahan, A. G., Toishimanov, M., Zhalelov, D., & Bolat, A. (2025). Effects of seasonal and regional variations on the bacterial and fungal biodiversity of mares’ milk and koumiss in the Almaty and Zhambyl regions of Kazakhstan. International Dairy Journal, 169, 106331. https://doi.org/10.1016/j.idairyj.2025.106331
Dash, K. K., Fayaz, U., Dar, A. H., Shams, R., Manzoor, S., Sundarsingh, A., Deka, P., & Khan, S. A. (2022). A comprehensive review on heat treatments and related impact on the quality and microbial safety of milk and milk-based products. Food Chemistry Advances, 1, 100041. https://doi.org/10.1016/j.focha.2022.100041
De Coster, W., D’Hert, S., Schultz, D. T., Cruts, M., & Van Broeckhoven, C. (2018). NanoPack: Visualizing and processing long-read sequencing data. Bioinformatics, 34(15), 2666–2669. https://doi.org/10.1093/bioinformatics/bty149
De Melo Pereira, G. V., De Carvalho Neto, D. P., Maske, B. L., De Dea Lindner, J., Vale, A. S., Favero, G. R., Viesser, J., De Carvalho, J. C., Góes-Neto, A., & Soccol, C. R. (2022). An updated review on bacterial community composition of traditional fermented milk products: What next-generation sequencing has revealed so far? Critical Reviews in Food Science and Nutrition, 62(7), 1870–1889. https://doi.org/10.1080/10408398.2020.1848787
Exarhopoulos, S., Karipoglou, D., Groztidou, O., Georgiou, D., Kalogianni, E. P., Goulas, A., & Dimitreli, G. (2025). Effect of drying aids on the quality properties of kefir powder. Dairy, 6(1), 9. https://doi.org/10.3390/dairy6010009
Gao, J., Li, X., Zhang, G., Sadiq, F. A., Simal‐Gandara, J., Xiao, J., & Sang, Y. (2021). Probiotics in the dairy industry—Advances and opportunities. Comprehensive Reviews in Food Science and Food Safety, 20(4), 3937–3982. https://doi.org/10.1111/1541-4337.12755
García, F. C., Clegg, T., O’Neill, D. B., Warfield, R., Pawar, S., & Yvon-Durocher, G. (2023). The temperature dependence of microbial community respiration is amplified by changes in species interactions. Nature Microbiology, 8(2), 272–283. https://doi.org/10.1038/s41564-022-01283-w
Ge, Z., Wang, D., Zhao, W., Wang, P., & Zhao, X. (2025). Advancements in the Structure, Spatial Configuration, Functional Properties, and Dairy Applications of Lactic Acid Bacteria Exopolysaccharides. Food Reviews International, 42(1), 1–30. https://doi.org/10.1080/87559129.2025.2467163
Grujović, M. Ž., Mladenović, K. G., Semedo‐Lemsaddek, T., Laranjo, M., Stefanović, O. D., & Kocić‐Tanackov, S. D. (2022). Advantages and disadvantages of non‐starter lactic acid bacteria from traditional fermented foods: Potential use as starters or probiotics. Comprehensive Reviews in Food Science and Food Safety, 21(2), 1537–1567. https://doi.org/10.1111/1541-4337.12897
Hamed, A. M., Galli, B., Hogan, S. A., Abdel‐Hamid, M., & Romeih, E. (2025). Adaptive and predictive approaches to mitigate the impact of milk seasonality on composition, processing technologies and quality of milk powders. International Journal of Dairy Technology, 78(1), e13148. https://doi.org/10.1111/1471-0307.13148
Hebishy, E., Yerlikaya, O., Mahony, J., Akpinar, A., & Saygili, D. (2023). Microbiological aspects and challenges of whey powders – I thermoduric, thermophilic and spore‐forming bacteria. International Journal of Dairy Technology, 76(4), 779–800. https://doi.org/10.1111/1471-0307.13006
İstanbullugil, F. R., Sanli, K., Ozturk, T., Keskin, B. C., Düyşöbayeva, A., Risvanli, A., Acaröz, U., Acaröz, D. A., Salykov, R., & Sahin, M. (2025). Koumiss microbiome: investigation of the microbial composition and functional potential of a unique beverage of fermented milk produced at Kyrgyz mountains. Probiotics and Antimicrobial Proteins, 1–17. https://doi.org/10.1007/s12602-025-10718-9
Kaur, H., Kaur, G., & Ali, S. A. (2022). Dairy-based probiotic-fermented functional foods: an update on their health-promoting properties. Fermentation, 8(9), 425. https://doi.org/10.3390/fermentation8090425
Kim, D., Song, L., Breitwieser, F. P., & Salzberg, S. L. (2016). Centrifuge: Rapid and sensitive classification of metagenomic sequences. Genome Research, 26(12), 1721–1729. https://doi.org/10.1101/gr.210641.116
Kong, F., Zhao, Q., Wang, S., Mu, G., & Wu, X. (2025). Comparative study on the physical and chemical properties influenced by variations in fermentation bacteria groups: inoculating different fermented Mare’s milk into cow’s milk. Foods, 14(8), 1328. https://doi.org/10.3390/foods14081328
Kossaliyeva, G., Rysbekuly, K., Zhaparkulova, K., Kozykan, S., Li, J., Serikbayeva, A., Shynykul, Z., Zhaparkulova, M., & Yessimsiitova, Z. (2025). Chemical composition, physical properties, and immunomodulating study of mare’s milk of the Adaev horse breed from Kazakhstan. Frontiers in Nutrition, 12, 1443031. https://doi.org/10.3389/fnut.2025.1443031
Kudaibergenova, A., Begdildayeva, N., Amirkhanova, A., Nurgazina, A., Ospanova, A., Kondybayev, A., Jumagaziyeva, A., Iskakbayeva, Z., Jumabayeva, S., & Akhmetsadykova, S. (2025). Probiotic characterization of lactic acid bacteria isolated from traditional fermented camel and mare’s milk products. ES Food and Agroforestry, 21, 1763. https://doi.org/10.30919/faf1763
Liu, X., Mao, B., Tang, X., Zhang, Q., Zhao, J., Chen, W., & Cui, S. (2025). Bacterial viability retention in probiotic foods: A review. Critical Reviews in Food Science and Nutrition, 65(32), 7964–7986. https://doi.org/10.1080/10408398.2025.2488228
Ljubojević Pelić, D., Lazić, S., & Živkov Baloš, M. (2024). Chemical contaminants in donkey milk: A review of literature on sources, routes and pathways of contamination, regulatory framework, health risks, and preventive measures. Heliyon, 10(21), e39999. https://doi.org/10.1016/j.heliyon.2024.e39999
Lohita, B., & Srijaya, M. (2024). Novel technologies for shelf-life extension of food products as a competitive advantage: a review. In R. Chakraborty, P. Mathur, & S. Roy (Eds.), Food Production, Diversity, and Safety Under Climate Change (pp. 285–306). Springer Nature Switzerland. https://doi.org/10.1007/978-3-031-51647-4_24
Manyi-Loh, C. E., & Lues, R. (2025). Listeria monocytogenes and Listeriosis: The Global Enigma. Foods, 14(7), 1266. https://doi.org/10.3390/foods14071266
Medjahdi, K., Didouh, N., Mathot, A.-G., Leguerinel, I., & Boudjemaa, B. M. (2025). Diversity, spoilage potential and biofilm formation assessment of spore-forming bacteria in post-pasteurization dairy environments. International Dairy Journal, 168, 106290. https://doi.org/10.1016/j.idairyj.2025.106290
Monareng, N. J., Ncube, K. T., Van Rooi, C., Modiba, M. C., & Mtileni, B. (2025a). A systematic review on microbial profiling techniques in goat milk: implications for probiotics and shelf-life. International Journal of Molecular Sciences, 26(12), 5551. https://doi.org/10.3390/ijms26125551
Nam, N., Do, H., Loan Trinh, K., & Lee, N. (2023). Metagenomics: an effective approach for exploring microbial diversity and functions. Foods, 12(11), 2140. https://doi.org/10.3390/foods12112140
Nygaard, A. B., Tunsjø, H. S., Meisal, R., & Charnock, C. (2020). A preliminary study on the potential of Nanopore MinION and Illumina MiSeq 16S rRNA gene sequencing to characterize building-dust microbiomes. Scientific Reports, 10(1). https://doi.org/10.1038/s41598-020-59771-0
PT Genetika Science Indonesia. (2023). Full-length sequence of 16S rRNA gene (V1-V9 regions) using Oxford Nanopore Technology.
Saha, N., Sharma, A., & Bora, P. (2025). Expanding the functional landscape of microbial entomopathogens in agriculture beyond pest management. Folia Microbiologica, 70(2), 343–357. https://doi.org/10.1007/s12223-025-01251-x
Sari, Y., Utama, A., Irawan, A., Andiana, P., Faradila, R., Manab, A., & Radiati, L. (2025). Optimizing dextrin levels for enhanced physicochemical properties and microstructure of sumbawa mare’s milk powder. Advances in Animal and Veterinary Sciences, 13(2), 431–439. https://doi.org/10.17582/journal.aavs/2025/13.2.431.439
Selmi, H., Presutto, E., Capozzi, V., Bonassisa, L., Drider, D., Abidi, F., Spano, G., de Chiara, M. L. V., & Fragasso, M. (2026). Optimizing fruit and vegetable by-products/wastes exploitation: Diversity, safety and biotechnological innovations based on lactic acid bacteria. Critical Reviews in Food Science and Nutrition, 1–37. https://doi.org/10.1080/10408398.2026.2624473
Shalini, G., & Gurunathan, N. (2025). Microbiological hazards in milk production, processing and products development. In Rana, T. (Ed.), Handbook of milk production, quality and nutrition (pp. 173–180). Academic Press. https://doi.org/10.1016/B978-0-443-24820-7.00039-0
Shori, A. B. (2024). Comparative analysis of Lactobacillus starter cultures in fermented camel milk: effects on viability, antioxidant properties, and sensory characteristics. Foods, 13(22), 3711. https://doi.org/10.3390/foods13223711
Shymialevich, D., Wójcicki, M., & Sokołowska, B. (2024). The novel concept of synergically combining: high hydrostatic pressure and lytic bacteriophages to eliminate vegetative and spore-forming bacteria in food products. Foods, 13(16), 2519. https://doi.org/10.3390/foods13162519
Sibanda, T., Marole, T. A., Thomashoff, U. L., Thantsha, M. S., & Buys, E. M. (2024). Bifidobacterium species viability in dairy-based probiotic foods: Challenges and innovative approaches for accurate viability determination and monitoring of probiotic functionality. Frontiers in Microbiology, 15, 1327010. https://doi.org/10.3389/fmicb.2024.1327010
Supamri, S., Radiati, L. E., & Susilo, A. (2025a). Microbial diversity and food safety in duck meat preservation: Effects of nanoemulsion nutmeg seed extract with focus on proteobacteria and lactococcus. CyTA - Journal of Food, 23(1), 2453531. https://doi.org/10.1080/19476337.2025.2453531
Tan, Y.-S., Zhang, R.-K., Liu, Z.-H., Li, B.-Z., & Yuan, Y.-J. (2022). Microbial adaptation to enhance stress tolerance. Frontiers in Microbiology, 13, 888746. https://doi.org/10.3389/fmicb.2022.888746
Tița, O., Constantinescu, M. A., Rusu, L., & Tița, M. A. (2024). Natural polymers as carriers for encapsulation of volatile oils: applications and perspectives in food products. Polymers, 16(8), 1026. https://doi.org/10.3390/polym16081026
Tong, T. A. N., Cao, T. L., & Pham, B. N. (2025). Antimicrobial strategies in food processing and post-processing. In D. Li (Ed.), Antimicrobial Strategies in the Food System: Updates, Opportunities, Challenges (pp. 365–447). Springer Nature Switzerland. https://doi.org/10.1007/978-3-031-95056-8_11
Toor, M. D., Ur Rehman, M., Abid, J., Nath, D., Ullah, I., Basit, A., Ud Din, M. M., & Mohamed, H. I. (2024). Microbial ecosystems as guardians of food security and water resources in the era of climate change. Water, Air, & Soil Pollution, 235(11), 741. https://doi.org/10.1007/s11270-024-07533-3
Tuesta-Popolizio, D. A., Velázquez-Fernández, J. B., Rodriguez-Campos, J., & Contreras-Ramos, S. M. (2021). Thalassobacillus, a genus of extreme to moderate environmental halophiles with biotechnological potential. World Journal of Microbiology and Biotechnology, 37(9), 147. https://doi.org/10.1007/s11274-021-03116-0
Wei, W., Hu, X., Hou, Z., Wang, Y., & Zhu, L. (2021). Microbial community structure and diversity in different types of non-bovine milk. Current Opinion in Food Science, 40, 51–57. https://doi.org/10.1016/j.cofs.2021.01.008
Wick, R. R., Judd, L. M., & Holt, K. E. (2019). Performance of neural network basecalling tools for Oxford Nanopore sequencing. Genome Biology, 20(1), 129. https://doi.org/10.1186/s13059-019-1727-y
Wu, X., Zhang, J., Yan, X., Wu, X., Zhang, Q., & Luan, M. (2024). An In-depth overview of the structural properties, health benefits, and applications of resistant dextrin. Journal of Food Processing and Preservation, 2024, 1–14. https://doi.org/10.1155/2024/8055063
Xiong, Y., Xu, J., Guo, L., Chen, F., Jiang, D., Lin, Y., Guo, C., Li, X., Chen, Y., Ni, K., & Yang, F. (2022). Exploring the effects of different bacteria additives on fermentation quality, microbial community and in vitro gas production of forage oat silage. Animals, 12(9), 1122. https://doi.org/10.3390/ani12091122
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