Effect of probiotic Lysinibacillus macroides on the innate immune system of the common carp, Cyprinus carpio, utilizing a machine learning approach

Shree Rama Mani; Sudharsanan Radhakrishnan; Gopirajan Punnivakotti Varadharajan; Premalatha Kandhasamy; Sivakumar Kandhasamy

doi:10.3153/AR26015

EN

Effect of probiotic Lysinibacillus macroides on the innate immune system of the common carp, Cyprinus carpio, utilizing a machine learning approach

Abstract

The present study aimed to examine the effects of the probiotic Lysinibacillus macroides (LM) on the innate immune parameters of common carp (Cyprinus carpio) using a machine learning (ML) approach. C. carpio were fed diets supplemented with 0% LM (control), 2% LM, 4% LM, 6% LM, and 8% LM for 60 days. As indicators of innate immunity, neutrophil adherence, serum lysozyme activity, and nitroblue tetrazolium activity were measured. Using parameter optimization and evaluation techniques based on a regression tree approach, the best predictive models for the innate immunity of C. carpio under different feeding regimes were determined through an ML methodology. Nitroblue Tetrazolium (NBT) activity, neutrophil adherence, and serum lysozyme were significantly enhanced by the supplemented diet. Lysinibacillus macroides was administered in a concentration-dependent manner. Gradient Boosting models predicted immunological changes with R2 values exceeding 0.85. We conclude that L. macroides could be a promising probiotic agent to enhance the innate immunity of common carp, and that ML technologies offer constructive explanations for the mechanisms underlying probiotic-mediated immunomodulation.

Keywords

Ethical Statement

The conducted research is approved by Karpaga Vinayaga Institute of Medical Sciences and Research Institute, Tamil Nadu, as per the guidelines of the Committee for Control and Supervision of Experiments on Animals (CCSEA), Department of Animal Husbandry and Dairying, Ministry of Fisheries, Animal Husbandry and Dairying, Government of India.

Thanks

The authors are thankful to the Management, Principal, and Head of the Department of Biotechnology, Karpaga Vinayaga College of Engineering and Technology, India, for providing the necessary resources to carry out the work.

References

Ahmadifar, E., Moghadam, M. S., Dawood, M.A., Hoseinifar, S.H. (2019). Lactobacillus fermentum and/or ferulic acid improved the immune responses, antioxidative defence and resistance against Aeromonas hydrophila in common carp (Cyprinus carpio) fingerlings. Fish and Shellfish Immunology, 94, 916–923. https://doi.org/10.1016/j.fsi.2019.10.019
Ai, Q., Xu, H., Mai, K., Xu, W., Wang, J., Zhang, W. (2011). Effects of dietary supplementation of Bacillus subtilis and fructooligosaccharide on growth performance, survival, non-specific immune response and disease resistance of juvenile large yellow croaker, Larimichthys crocea. Aquaculture, 317(1–4), 155–161. https://doi.org/10.1016/j.aquaculture.2011.04.036
Alcorn, S.W., Murray, A.L., Pascho, R.J. (2002). Effects of rearing temperature on immune functions in sockeye salmon. Fish and Shellfish Immunology, 12(4), 303–334. https://doi.org/10.1006/fsim.2001.0373
Aleman, R.S., Yadav, A. (2023). Systematic Review of probiotics and their potential for developing functional nondairy foods. Applied Microbiology, 4(1), 47–69. https://doi.org/10.3390/applmicrobiol4010004
Biller, J.D., Polycarpo, G.D.V., Moromizato, B.S., Sidekerskis, A.P.D., Da Silva, T.D., Reis, I.C.D., Fierro-Castro, C. (2021). Lysozyme activity as an indicator of innate immunity of tilapia (Oreochromis niloticus) when challenged with LPS and Streptococcus agalactiae. Revista Brasileira De Zootecnia, 50. https://doi.org/10.37496/rbz5020210053
Calduch-Giner, J., Holhorea, P.G., Ferrer, M.Á., Naya-Català, F., Rosell-Moll, E., García, C.V., Prunet, P., Espmark, Å., Leguen, I., Kolarevic, J., Vega, A., Kerneis, T., Goardon, L., Afonso, J.M., Pérez-Sánchez, J. (2022). Revising the impact and prospects of activity and ventilation rate bio-loggers for tracking welfare and fish-environment interactions in salmonids and Mediterranean farmed fish. Frontiers in Marine Science, 9. https://doi.org/10.3389/fmars.2022.854888
Chen, T., Guestrin, C. (2016). XGBoost. XGBoost: A Scalable Tree Boosting System. Proceedings of the 22nd ACM SIGKDD International Conference on Knowledge Discovery and Data Mining. 785 – 794. https://doi.org/10.1145/2939672.2939785
Dennis, R.N., Keerthana, D.N. (2024). Performance analysis of voting regression-based ensemble learning methods for food demand forecasting. International Journal of Data Informatics and Intelligent Computing, 3(2), 34–41. https://doi.org/10.59461/ijdiic.v3i2.114

El-Saadony, M.T., Alagawany, M., Patra, A.K., Kar, I., Tiwari, R., Dawood, M.A., Dhama, K., Abdel-Latif, H.M. (2021). The functionality of probiotics in aquaculture: An overview. Fish and Shellfish Immunology, 117, 36–52. https://doi.org/10.1016/j.fsi.2021.07.007
Faeed, M., Kermanshahi, R.K., PourKazemi, M., Ahmadnezhad, M. (2022). In vivo study on probiotic Lactobacillus brevis in Sander lucioperca and some of their nonspecific immune parameters, intestinal morphology and survival against Aeromonas hydrophila. Iranian Journal of Fishery Sciences, 21(2), 387–402.
Føre, M., Frank, K., Norton, T., Svendsen, E., Alfredsen, J. A., Dempster, T., Eguiraun, H., Watson, W., Stahl, A., Sunde, L.M., Schellewald, C., Skøien, K.R., Alver, M.O., Berckmans, D. (2017). Precision fish farming: A new framework to improve production in aquaculture. Biosystems Engineering, 173, 176–193. https://doi.org/10.1016/j.biosystemseng.2017.10.014
Friedman, J.H. (2001). Greedy function approximation: A gradient boosting machine. The Annals of Statistics, 29(5). https://doi.org/10.1214/aos/1013203451
Gao, X., Zhang, M., Li, X., Han, Y., Wu, F., Liu, Y. (2018). Effects of a probiotic (Bacillus licheniformis) on the growth, immunity, and disease resistance of Haliotis discus hannai Ino. Fish and Shellfish Immunology, 76, 143–152. https://doi.org/10.1016/j.fsi.2018.02.028
Gatesoupe, F. (2007). Live yeasts in the gut: Natural occurrence, dietary introduction, and their effects on fish health and development. Aquaculture, 267(1–4), 20–30. https://doi.org/10.1016/j.aquaculture.2007.01.005
Guo, G., Li, C., Xia, B., Jiang, S., Zhou, S., Men, X., Ren, Y. (2020). The efficacy of lactic acid bacteria usage in turbot Scophthalmus maximus on intestinal microbiota and expression of the immune related genes. Fish and Shellfish Immunology, 100, 90–97. https://doi.org/10.1016/j.fsi.2020.03.003
Hoseinifar, S.H., Ringø, E., Masouleh, A.S., Esteban, M.Á. (2014). Probiotic, prebiotic and synbiotic supplements in sturgeon aquaculture: a review. Reviews in Aquaculture, 8(1), 89–102. https://doi.org/10.1111/raq.12082
Irianto, A., & Austin, B. (2002). Probiotics in aquaculture. Journal of Fish Diseases, 25(11), 633–642. https://doi.org/10.1046/j.1365-2761.2002.00422.x
Kazuń, B., Małaczewska, J., Kazuń, K., Żylińska-Urban, J., Siwicki, A.K. (2018). Immune-enhancing activity of potential probiotic strains of Lactobacillus plantarum in the common carp (Cyprinus carpio) fingerling. Journal of Veterinary Research, 62(4), 485–492. https://doi.org/10.2478/jvetres-2018-0062
Kumar, S., Raman, R.P., Kumar, K., Pandey, P.K., Kumar, N., Mallesh, B., Mohanty, S., Kumar, A. (2012). Effect of azadirachtin on haematological and biochemical parameters of Argulus-infested goldfish Carassius auratus (Linn. 1758). Fish Physiology and Biochemistry, 39(4), 733–747. https://doi.org/10.1007/s10695-012-9736-8
Magnadóttir, B. (2005). Innate immunity of fish (overview). Fish Shellfish Immunology, 20(2), 137–151. https://doi.org/10.1016/j.fsi.2004.09.006
Magrone, N.T., Jirillo, N.E. (2012). Mechanisms of neutrophil-mediated disease: innovative therapeutic interventions. Current Pharmaceutical Design, 18(12), 1609–1619. https://doi.org/10.2174/138161212799958512
Mani, S.R., Vijayan, K., Jacob, J.P., Vijayakumar, S., Kandhasamy, S. (2021). Evaluation of probiotic properties of Lysinibacillus macroides under in vitro conditions and culture of Cyprinus carpio on growth parameters. Archives of Microbiology, 203(7), 4705–4714. https://doi.org/10.1007/s00203-021-02452-x
Marcos-Zambrano, L.J., Karaduzovic-Hadziabdic, K., Turukalo, T.L., Przymus, P., Trajkovik, V., Aasmets, O., Berland, M., Gruca, A., Hasic, J., Hron, K., Klammsteiner, T., Kolev, M., Lahti, L., Lopes, M.B., Moreno, V., Naskinova, I., Org, E., Paciência, I., Papoutsoglou, G., Shigdel, R., Stres, B., Vilne, B., Yousef, M., Zdravevski, E., Tsamardinos, I., de Santa Pau, E.C., Claesson, M.J., Moreno-Indias, I.J., Truu, J. (2021). Applications of Machine Learning in Human Microbiome Studies: A Review on Feature Selection, Biomarker Identification, Disease Prediction and Treatment. Frontiers in Microbiology, 12. https://doi.org/10.3389/fmicb.2021.634511
Melo-Bolívar, J.F., Ruiz-Pardo, R.Y., Hume, M.E., Sidjabat, H. E., Villamil-Díaz, L.M. (2020). Probiotics for cultured freshwater fish. Microbiology Australia, 41(2), 105. https://doi.org/10.1071/MA20026
Merrifield, D.L., Dimitroglou, A., Foey, A., Davies, S.J., Baker, R.T., Bøgwald, J., Castex, M., Ringø, E. (2010). The current status and future focus of probiotic and prebiotic applications for salmonids. Aquaculture, 302(1–2), 1–18. https://doi.org/10.1016/j.aquaculture.2010.02.007
Michael, E.T., Amos, S.O., Hussaini, L.T. (2014). A Review on probiotics application in aquaculture. Fisheries and Aquaculture Journal, 05(04). https://doi.org/10.4172/2150-3508.10000111
Mustafa, S.A., Al-Rudainy, A.J., Salman, N.M. (2024). Effect of environmental pollutants on fish health: An overview. The Egyptian Journal of Aquatic Research, 50(2), 225–233. https://doi.org/10.1016/j.ejar.2024.02.006
Nakharuthai, C., Boonanuntanasarn, S., Kaewda, J., Manassila, P. (2023). Isolation of Potential Probiotic Bacillus spp. from the Intestine of Nile Tilapia to Construct Recombinant Probiotic Expressing CC Chemokine and Its Effectiveness on Innate Immune Responses in Nile Tilapia. Animals, 13(6), 986. https://doi.org/10.3390/ani13060986 Natekin, A., Knoll, A. (2013). Gradient boosting machines, a tutorial. Frontiers in Neurorobotics, 7. https://doi.org/10.3389/fnbot.2013.00021
Neumann, N.F., Stafford, J.L., Barreda, D., Ainsworth, A, Belosevic, M. (2001). Antimicrobial mechanisms of fish phagocytes and their role in host defense. Developmental and Comparative Immunology, 25(8–9), 807–825. https://doi.org/10.1016/s0145-305x(01)00037-4
Newaj‐Fyzul, A., Austin, B. (2014). Probiotics, immunostimulants, plant products and oral vaccines, and their role as feed supplements in the control of bacterial fish diseases. Journal of Fish Diseases, 38(11), 937–955. https://doi.org/10.1111/jfd.12313
Rahman, A., Tasnim, S. (2014). Application of machine learning techniques in aquaculture. International Journal of Computer Trends and Technology, 10(4), 214–215. https://doi.org/10.14445/22312803/ijctt-v10p137
Secombes, C. (1996). The nonspecific immune system: Cellular defenses. In Fish physiology (pp. 63–103). https://doi.org/10.1016/s1546-5098(08)60272-1
Shandilya, S., Kumar, S., Jha, N.K., Kesari, K.K., Ruokolainen, J. (2021). Interplay of gut microbiota and oxidative stress: Perspective on neurodegeneration and neuroprotection. Journal of Advanced Research, 38, 223–244. https://doi.org/10.1016/j.jare.2021.09.005
Shree Rama, M., Karthikeyan, V., Jacob, J.P., Sekar, V., Sivakumar K. (2021). Evaluation of probiotic properties of Lysinibacillus macroides under in vitro conditions and culture of Cyprinus carpio on growth parameters. Archives of Microbiology. 203, 4705–4714. https://doi.org/10.1007/s00203-021-02452-x
Shree Rama M. (2022). In vitro study of Lysinibacillus macroides probiotic properties and its effect on growth, survival and immunological parameters of Cyprinus carpio. PhD thesis, Anna University, Tamil Nadu. 1-118.
Szydłowska, A., Sionek, B. (2022). Probiotics and postbiotics as the functional food components affecting the immune response. Microorganisms, 11(1), 104. https://doi.org/10.3390/microorganisms11010104
Tannock, G.W., Munro, K., Harmsen, H.J.M., Welling, G.W., Smart, J., Gopal, P.K. (2000). Analysis of the fecal microflora of human subjects consuming a probiotic product containing Lactobacillus rhamnosus DR20. Applied and Environmental Microbiology, 66(6), 2578–2588. https://doi.org/10.1128/aem.66.6.2578-2588.2000
Tovar-Ramírez, D., Mazurais, D., Gatesoupe, J., Quazuguel, P., Cahu, C., Zambonino-Infante, J. (2009). Dietary probiotic live yeast modulates antioxidant enzyme activities and gene expression of sea bass (Dicentrarchus labrax) larvae. Aquaculture, 300(1–4), 142–147. https://doi.org/10.1016/j.aquaculture.2009.12.015
Xia, Y., Wang, M., Gao, F., Lu, M., Chen, G. (2019). Effects of dietary probiotic supplementation on the growth, gut health and disease resistance of juvenile Nile tilapia (Oreochromis niloticus). Animal Nutrition, 6(1), 69–79. https://doi.org/10.1016/j.aninu.2019.07.002
Xiong, J., Nie, L., Chen, J. (2019). Current understanding on the roles of gut microbiota in fish disease and immunity. Zoological Research, 40(2), 70–76. https://doi.org/10.24272/j.issn.2095-8137.2018.069
Yao, Y., Wang, X., Lin, X., Wu, J., Wang, P., Zhu, C., Yan, Q. (2024). Isolation and characterization of probiotic Lysinibacillus species from the gastrointestinal tract of large yellow croaker (Larimichthys crocea). Frontiers in Marine Science, 11. https://doi.org/10.3389/fmars.2024.1408979
Yildiz, H.Y., Guzey, İ.M., Ergonul, M.B. (2009). Changes of non-specific immune parameters in rainbow trout, Oncorhynchus mykiss, after exposure to antimicrobial agents used in aquaculture. Journal of Applied Aquaculture, 21(3), 139–150. https://doi.org/10.1080/10454430903113529
Zhao, S., Zhang, S., Liu, J., Wang, H., Zhu, J., Li, D., Zhao, R. (2021). Application of machine learning in intelligent fish aquaculture: A review. Aquaculture, 540, 736724. https://doi.org/10.1016/j.aquaculture.2021.736724

Details

Primary Language

English

Subjects

Aquaculture and Fisheries (Other)

Journal Section

Research Article

Authors

Shree Rama Mani
0000-0002-4492-6783
India

Sudharsanan Radhakrishnan
0000-0003-1799-426X
India

Gopirajan Punnivakotti Varadharajan
0000-0002-8653-880X
India

Premalatha Kandhasamy
0000-0002-5340-0265
India

Sivakumar Kandhasamy ^*
0000-0001-8971-869X
India

Early Pub Date

June 22, 2026

Publication Date

-

Submission Date

January 11, 2026

Acceptance Date

March 2, 2026

Published in Issue

Year 2026 Number: Advanced Online Publication

DOI

https://doi.org/10.3153/AR26015

IZ

https://izlik.org/JA86FM62TN

Cite

RIS / Bibtex

APA

Rama Mani, S., Radhakrishnan, S., Varadharajan, G. P., Kandhasamy, P., & Kandhasamy, S. (2026). Effect of probiotic Lysinibacillus macroides on the innate immune system of the common carp, Cyprinus carpio, utilizing a machine learning approach. Aquatic Research, Advanced Online Publication, 176-190. https://doi.org/10.3153/AR26015

AMA

1.Rama Mani S, Radhakrishnan S, Varadharajan GP, Kandhasamy P, Kandhasamy S. Effect of probiotic Lysinibacillus macroides on the innate immune system of the common carp, Cyprinus carpio, utilizing a machine learning approach. Aquat Res. 2026;(Advanced Online Publication):176-190. doi:10.3153/AR26015

Chicago

Rama Mani, Shree, Sudharsanan Radhakrishnan, Gopirajan Punnivakotti Varadharajan, Premalatha Kandhasamy, and Sivakumar Kandhasamy. 2026. “Effect of Probiotic Lysinibacillus Macroides on the Innate Immune System of the Common Carp, Cyprinus Carpio, Utilizing a Machine Learning Approach”. Aquatic Research, no. Advanced Online Publication: 176-90. https://doi.org/10.3153/AR26015.

EndNote

Rama Mani S, Radhakrishnan S, Varadharajan GP, Kandhasamy P, Kandhasamy S (June 1, 2026) Effect of probiotic Lysinibacillus macroides on the innate immune system of the common carp, Cyprinus carpio, utilizing a machine learning approach. Aquatic Research Advanced Online Publication 176–190.

IEEE

[1]S. Rama Mani, S. Radhakrishnan, G. P. Varadharajan, P. Kandhasamy, and S. Kandhasamy, “Effect of probiotic Lysinibacillus macroides on the innate immune system of the common carp, Cyprinus carpio, utilizing a machine learning approach”, Aquat Res, no. Advanced Online Publication, pp. 176–190, June 2026, doi: 10.3153/AR26015.

ISNAD

Rama Mani, Shree - Radhakrishnan, Sudharsanan - Varadharajan, Gopirajan Punnivakotti - Kandhasamy, Premalatha - Kandhasamy, Sivakumar. “Effect of Probiotic Lysinibacillus Macroides on the Innate Immune System of the Common Carp, Cyprinus Carpio, Utilizing a Machine Learning Approach”. Aquatic Research. Advanced Online Publication (June 1, 2026): 176-190. https://doi.org/10.3153/AR26015.

JAMA

1.Rama Mani S, Radhakrishnan S, Varadharajan GP, Kandhasamy P, Kandhasamy S. Effect of probiotic Lysinibacillus macroides on the innate immune system of the common carp, Cyprinus carpio, utilizing a machine learning approach. Aquat Res. 2026;:176–190.

MLA

Rama Mani, Shree, et al. “Effect of Probiotic Lysinibacillus Macroides on the Innate Immune System of the Common Carp, Cyprinus Carpio, Utilizing a Machine Learning Approach”. Aquatic Research, no. Advanced Online Publication, June 2026, pp. 176-90, doi:10.3153/AR26015.

Vancouver

1.Shree Rama Mani, Sudharsanan Radhakrishnan, Gopirajan Punnivakotti Varadharajan, Premalatha Kandhasamy, Sivakumar Kandhasamy. Effect of probiotic Lysinibacillus macroides on the innate immune system of the common carp, Cyprinus carpio, utilizing a machine learning approach. Aquat Res. 2026 Jun. 1;(Advanced Online Publication):176-90. doi:10.3153/AR26015