**5. Conclusions**

We can conclude that the addition of 0.5% HS to laying hen feed had a positive effect on laying rate, daily egg mass, egg weight, feed conversion, and eggshell quality and also had an immunostimulatory effect manifested by the increasing activity of the phagocytes and the B cell immune response. On the other hand, there was an observed increase in the number of enterobacteria in the intestinal contents, and the proportion of T lymphocytes in the blood was significantly reduced. The presented results confirm that HS can be used for the improvement of egg production and quality and for activation of phagocytosis and specific antibody immunity, but their influence on the intestinal microbiota will need to be further studied with respect to a wider range of microbial species inhabiting the digestive tract of laying hens.

**Author Contributions:** D.M., investigation, formal analysis, writing; V.K., investigation, formal analysis, writing; B.S., investigation, writing; P.N. and J.K., investigation; M.B., animal experiment managemen<sup>t</sup> and investigation; A.M. and L.B., investigation; J.N. and J.M., supervision; S.M., conceptualization and resources. All authors have read and agreed to the published version of the manuscript.

**Funding:** This research was funded by the Operational Programme Integrated Infrastructure within the project Demand-driven research for the sustainable and innovative food, Drive4SIFood 313011V336, co-financed by the European Regional Development Fund.

**Institutional Review Board Statement:** The study was conducted according to the European directive 2010/63/EU on the protection of animals used for scientific purposes and was approved by the Ethical Committee of the University of Veterinary Medicine and Pharmacy in Košice and the State Veterinary and Food Administration of the Slovak Republic, approved by the experimental protocol number 3040/14-221 in December 2014.

**Informed Consent Statement:** Not applicable.

**Data Availability Statement:** The data presented in this study are available upon request from the corresponding author.

**Acknowledgments:** The authors gratefully acknowledge Humac Ltd., Košice, Slovakia, for providing humic substances for the experiment. A special thanks to Salonee Udani, an English native speaker from London, who is a student at the University of Veterinary Medicine and Pharmacy in Košice (Slovakia), for English proofreading.

**Conflicts of Interest:** The authors state that there are no conflicts of interest regarding the publication of this article.
