For several decades, the use of antibiotic growth promoters (AGPs) in the livestock sector enhanced growth performance, intestinal health, feed efficiency, and mitigation of the stress caused by environmental and management factors in intensive poultry farming (1). However, with the long-term and extensive use of AGP, public concerns have augmented with respect to resistant bacteria and antibiotic drug residues (2). For this, many countries such as the European Union (3) and, recently, China have completely banned the utilize of AGP as growth promoters in animal diets. Given the high demand for high-quality poultry products, the development of sustainable, safe, practical, and effective feed additives that can both maintain broiler health and enhance productive potential is imperative. Presently, the use of herbal plants and their derivatives is gaining more attention globally for poultry production that could support in maintaining health status and growth performance (4, 5). Development of abundant plant resource for use as feedstuff or additives is one of the most important strategies to support sustainable animal production.

Lasia spinosa Thw. (LST) is a perennial herb, which grows extensively in numerous moist areas in tropical and subtropical regions (6). LST contains high content of fiber and a range of health-enhancing molecules such as flavonoids, phenolic compounds, and carotenoids, which have antioxidant, anti-inflammatory, and antihyperlipidemic properties; therefore, it has been used as a medicine in folk for many years to cure various diseases such as cholic, intestinal disorders, and rheumatism and even humans in the Bodo communities used LST as the cheapest source of nutrition to supplement vitamins and minerals (7), representing beneficial potency in poultry diets.

The functional properties of LST have already been applied to animal production. In some earlier studies, dietary inclusion of LST meal has been described to have boosted the performance production and feed efficiency in fish and buffalo (8, 9). This may attribute to the improvement of intestinal health in fish by LST supplementation (8). Based on the evidences stressing out a significant ability of LST to promote growth performance of animals, we designed this study to evaluate the effects of LST dietary inclusion on the growth performance, blood metabolites, antioxidant profile, intestinal morphology, and cecal microbiota in broilers, aiming at increasing knowledge and providing innovative information on the usage of LST in chicken feed as a brand new natural additive to support the sustainable and better development of livestock industry.

This study was conducted on the Experiment Farm of Guangxi University and the Guangxi Fufeng Agriculture and Animal Husbandry Corporation, Ltd (Nanning, PR China). The bird maintenance and experimental protocols in this study were conducted following the guidelines approved by the Institutional Animal Care and Use Committee of Guangxi University (GXU2018-003).

In the last decades, dietary manipulations with innovative feed supplements, such as medicinal plants and their derivatives, are among the common advanced strategies (3). Medicinal plants are reflected as a cornerstone of medicine since earliest times and have been employed as promoters of antioxidant capacity, immunity, and growth in animals (12). LST has been used medicinally since ancient times for its diverse biological activities such as antibacterial and antiseptic properties (7).

In this study, dietary LST powder supplementation significantly boosted the growth performance of the chickens involved, produced a remarkable increase in ADG and final BW, and caused a statistical decrease in FCR when compared to the control diet. These results are in accordance with the findings that the inclusion of LST in fish diets significantly improved the growth performance and feed utilization (8). In addition, this study displayed no significant difference in mortality rate was noticed in groups that received LST compared with the control, which agreed with a study by Munglue et al. (8) who perceived no significant alterations concerning survival rate in fish as a response to the dietary LST. In parallel, the supplementation of LST in buffalo had a positive impact on the growth indices (9). However, the underlying mechanisms of the growth-promoting effect of LST are still not understood. Phesatcha et al. (13) found that the inferior FCR and boosted growth may relate to enhanced appetite and digestion in beef cattle fed with diets enriched with LST (60–90 g/day/head). The authors suggested that this could be related to the constructive role of LST in improving the FI, apparent digestibility, and efficiency of microbial N synthesis in cattle (13). Similarly, Abolfathi et al. (14) found that elecampane (Inula helenium L.) rhizome showed positive effects on growth performance and this improvement in growth parameters was accompanied by significant increase in apparent ileal digestibility of dry matter, organic matter, ether extract, and gross energy. Thus, the improved digestibility may be one of the motives in this study.

As a natural additive in animal feed, several phytohormones found in LST are proven to promote growth and feed utilization. Suthikrai et al. (9) found that LST contains considerable levels of natural phytoestrogens (10.76–14.55 pg/g DM) and phytoandrogens (0.15–0.92 pg/g DM). Therefore, it is reasonable to hypothesize that the production of growth hormone and insulin-like growth factor-1 may be modulated by phytohormones, resulting in the improvement of growth. Phytoandrogens may also enhance protein synthesis by activating the androgen receptor (AR)-mediated anabolism (15). In addition, combined with the findings of Buszczak et al. (16), the growth-promoting effect of LST may be due to modulating related genes expression related to protein synthesis and cell growth. Moreover, LST is excessively rich in alkaloids, flavonoids, terpenoids, phenolic compounds, steroids, phytosterols, saponins, coumarins, tannins, glycosides, and anthraquinone (7). Kikusato et al. (17) found that plant-derived isoquinoline alkaloids supplementation significantly improved the growth performance of chickens under heat stress conditions. Starčević et al. (18) reported that phenolic compounds boosted the growth performance and antioxidant profile of chickens. It has been reported that phytosterols could promote animal production and antioxidant capacity (19). Thus, these phytochemical constituents may synergistically contribute to the growth-promoting property of LST. However, due to various factors such as localities, sample processing, and the methods of analysis, not only the levels of bioactive substances, but also the composition may be different.

Blood variables are commonly reliable indicators of general health profile, as responses to external and internal stressors and stimuli. This study notices that LST supplements did not significantly affect the levels of blood metabolites (except for TBIL, TG, HDL-C, and LDL-C), showing that LST had no detrimental impacts or toxicity in the broilers. In this study, dietary inclusion of LST reduced the TG and LDL-C and boosted TBIL and HDL-C levels in broiler serum. The findings were consistent with prior studies in rats where the levels of TG and LDL-C were significantly decreased with the methanolic leaves extract of LST (200, 400, and 800 mg/kg BW) administration in comparison to the control group (20). Moreover, Kaewamatawong et al. (21) studied the acute to subchronic toxicity of LST in mice and found that LST significantly decreased the TG values, but did not induce any toxicological effects in the acute and subchronic term. It was reported that flavonoids from corn silk exhibited strong regulatory activity on several serum lipid values, reducing the levels of TG and LDL-C and increasing the level of HDL-C (22). Furthermore, scientists presented new evidence supporting a key role for Rhizoma coptidis alkaloids as a regulator of liver–gut axis in hyperlipidemic mice (23). Therefore, the presence of antihyperlipidemic compounds such as flavonoids and alkaloids present in LST could be responsible for a decrease of the serum TGs and LDL-C and increase of the HDL-C levels and, thus, avoiding cardiovascular ailment (7). Further studies should be conducted to validate the underlined mechanism of LST as a natural lipid-lowering agent for drug treatments in poultry. This study has shown that LST significantly improved the levels of Ca, Fe, and P in broiler serum as a response to 4% addition of LST in broiler diets. This enhancement could be due to the rich trace element content in LST (7). In addition, TBIL was significantly reduced only in the LST1 group and other biochemical parameters were similar among the groups, thus indicating that the LST powder consumption did not have a negative influence on the health status of the animals.

In the physiological process, free radicals and other oxygen-derived species are continuously produced in vivo, some of which can cause serious damage to biological molecules, especially to DNA, lipids, and proteins. Lipid peroxidation is a fundamental cellular degenerating process caused by free radicals and readily occurs in the tissues rich in highly oxidizable polyunsaturated fatty acids, which may contribute to cell death and disease in living organisms (24). The elimination of free radicals depends on the action of several enzymes such as SOD, GSH-Px, and CAT (25). Moreover, chicken meat is relatively rich in polyunsaturated fatty acids, making it extremely vulnerable to oxidation (26). Therefore, diet-derived antioxidants may be vital in reducing cumulative oxidative damage. Natural antioxidants are broadly found in medicinal herbs. 5 g/kg supplementation of ginger root powder enhanced the oxidative stability of broilers (27). Pulicaria gnaphalodes powder in broiler diets significantly increased the SOD and GSH-Px activities and decreased the MDA level in serum, liver, and thigh muscle (28). According to a large amount of literature, plant polyphenolic flavonoids was one of the major groups of compounds contributing mainly to the total antioxidant activity (29). The LST and its extracts have antioxidant properties due to its high content of flavonoids and phenolic compounds (7). It has been reported that the extracts of LST could scavenge the stable radical 1,1-diphenyl-2-picrylhydrazyl (DPPH), elucidating those polyphenols and ascorbic acid acted mainly as the primary antioxidant free-radical terminators of rhizomes (30). Besides, phytochemical constituents in LST such as flavonoids and phytosterols (19, 31) were proven to improve the antioxidant enzyme activity.

In this study, we measured the levels of antioxidant enzymes to determine the antioxidant status in serum, liver, and breast muscle. We found that GSH-Px, SOD, and CAT were significantly increased in the LST supplementation groups, while the levels of ROS and MDA, as indicators of OS, were significantly reduced in the LST supplementation groups. The increased antioxidant enzyme content is parallel with the decreased level of MDA and ROS in response to dietary LST inclusion, indicating that dietary LST supplements can promote the antioxidant capability of broilers by enhancing the enzymatic antioxidant scheme. As the improvement of the antioxidant enzyme would exert a positive influence on meat quality (19), so it is logical to speculate that the addition of LST can enhance the meat quality of the chickens involved in this experiment.

Results of this study indicated that the dietary LST powder addition affected the structure of the intestine in broilers. It is well-known that intestinal morphology can objectively reflect gut health and Vh, as well as Cd, can partially reflect the functional status of the intestine (11). A previous study demonstrated that the Vh was increased and the intestinal function was improved in fish after supplementation of LST extract in diet (8). Similarly, in this study, a statistical decrease in Cd and a statistical increase in Vh and the ratio of the two were observed in the intestine of chickens fed with LST powder compared with the control group. The increased Vh and Vh/Cd ratio for various gut fractions of chickens fed with LST powder were in accordance with appropriate growth performance and increased metabolizability of nutrients. In fact, the increase of Vh usually results in a higher total luminal villus absorptive area and subsequently leads to higher transport of nutrients at the villus surface (32). Moreover, the decrease in Cd usually indicates a slower turnover and healthier condition of the gut, which can reduce maintenance requirements and ultimately be beneficial to promote the growth of the animals (33). Rich fiber fractions in LST rhizome (7) may sustain the intestinal integrity through increasing the Vh and Vh/Cd and decreasing the Cd feature (34).

The gut microbiome is closely related to intestinal health, normal physiological functions, and poultry production and the composition of the microbial community could be altered upon the diet and over time (35). This study is setup necessarily to evaluate the effect of LST powder as a feed additive on the intestinal flora of Guangxi partridge broilers, which will contribute to developing nutrition intervention for optimal health, growth, and productivity in poultry. We found that the dominant phyla in the chicken are Firmicutes, Bacteroidetes, and Proteobacteria. Furthermore, the primary genera were Bacteroides, Faecalibacterium, and Oscillospira in this study, which was consistent with earlier works recognizing Bacteroidetes as the fundamental genus in the cecum (about 40% of sequences) (36).

Dietary LST powder addition also caused a significant alteration of the gut microbiota diversity in broilers. According to the α-diversity results, the 1% supplementation of LST powder significantly increased the abundance of caecal microbiota (PD whole-tree index, Chao1 index, observed species, and unique OTUs) compared with the other groups, while 2 and 4% supplementation of LST powder decreased the abundance (Chao1 index and observed species) of cecal microbiota. In addition, with respect to the β-diversity analysis, a clear separation of microbial community due to dietary LST powder inclusion was observed, which means microbial communities of each group were significantly different. Elevated levels of diversity mostly contribute to maintaining the stability of intestinal microbiota after exposure to environmental stress as well as resistance against potential invading pathogens (37). For these reasons, it is considered that a reduction of α- and β-diversity should be negative. This suggests that the structure of intestinal flora in the LST1 group was improved and, thus, better intestinal health. However, the LST2 and LST4 groups got a lower diversity and this may be due to the antibacterial activity of higher LST powder addition (7). However, no antibacterial substances were specified in these studies (7) and further investigation is required for isolating the specific bioactive constituents responsible for antibacterial properties.

Linear discriminant analysis effect size analysis revealed that the relative abundance of Anaerofilum was significantly increased in the LST1 group compared with other groups, which has been proposed to boost the antioxidant capability and absorption of energy as previously reported by Guo et al. (38). Interestingly, Odoribacter (39, 40) and Sutterella (41), known as short-chain fatty acid (SCFA)-producing bacteria, were also overrepresented in the LST1 group. SCFAs, as the main energy source of colorectal cells, play an essential role in regulating the absorption of several nutrients, digestive and hormonal secretions in the intestine, and participating in energy metabolism widely (39, 41). Further investigation will be focused on measuring the composition and levels of SCFAs in broilers supplemented with LST powder. Notably, Sutterella, a potential probiotic, could contribute to improving the growth rate and the feed conversion ratio of chickens (42) and its deficiency might lead to the destruction of the normal function of colonic epithelium and the induction of inflammatory bowel disease, suggesting its vital role in maintaining intestinal health (43). In addition, Bacillus has been widely proven to promote growth performance (41, 44). In this study, we found the enrichment of Bacillus in the LST1 group, while the best growth performance was found in the LST1 group. Therefore, the increase of relative abundance in Anaerofilum, Odoribacter, Sutterella, and Bacillus in the LST1 groups in this study suggests the efficacy of 1% LST powder in promoting beneficial bacteria, which would eventually contribute to improved performance and health. These findings suggest that there is a close association between intestinal flora and physiological performance.

However, no reports have verified the associations of growth performance, serum metabolites, antioxidant status, and intestinal morphology with changed gut microbiome structure in Guangxi partridge broilers fed with LST powder. By using the Spearman's correlation analysis, we revealed that the profiles of taxonomic composition at the genus level in cecal microbiota were significantly associated with growth performance, serum metabolites, antioxidant status, and intestinal morphology. Consistently, the relative abundance of Odoribacter and Bacillus was positively correlated with growth performance, which showed that 1% addition level of LST powder could improve ADG and decrease FCR potentially by increasing the abundance of Odoribacter and Bacillus as detected at day 42. In this study, we also demonstrated that Bacillus, which has been reported to improve intestinal morphology (45, 46), was positively related to the Vh and Vh/Cd of the duodenum. Furthermore, the abundance of Bacillus was closely correlated to the concentrations of CAT, SOD, and GSH-Px and many studies have reported that Bacillus could participate in boosting of the antioxidant scheme (47–49). Actually, intestinal flora can regulate the well-being, immunity, and disease of host by determining the biological value of the diet (50). In this study, we found that serum metabolites including TG, TBIL, Ca, Mg, and LDL-C affected by dietary treatment of LST powder were faithfully connected with the dynamic fluctuations of broiler gut microbiome structure.

Overall, the diversity of gut microbiota is reshaped by LST supplements and further contributes to the improvement of growth performance, intestinal morphology, antioxidant capacity, Ca and Mg, and reduction of TG, LDL-C, and TBIL.

Our results indicate that LST powder contributed to the improvement of growth performance, intestinal morphology, antioxidant capacity, Ca, Mg, Fe and P, and reduction of blood lipids. Especially, 1% supplementation of LST significantly increased the diversity of gut microbiota, which further promoted physiological performance. Of note, during the antibiotic-free era, LST has a great potential application in broiler feeding when the importance of nutrition, feed processing, management, and biosafety strategies is highly recognized.

The datasets presented in this study can be found in online repositories. The names of the repository/repositories and accession number(s) can be found in the article/Supplementary Material.

The animal study was reviewed and approved by the Institutional Animal Care and Use Committee of Guangxi University.

YH and LZ contribute to the investigation, formal analysis, and writing—original draft. YLi contributes to the project administration. KT and HS contribute to the investigation. XL contributes to the writing—review and editing. YT contributes to the resources. AH and SA contribute to the writing—original draft. WS, KS, and TT contribute to the conceptualization. YLu contributes to the writing—review and editing and funding acquisition. All authors contributed to the article and approved the submitted version.

This study was jointly supported by the National Natural Science Foundation of China (No. 31960157), the Guangxi Pioneer of Animal Science and Technology (No. 202109-02), and the Guangxi Special Fund for Invited Expert.

YT was employed by Guangxi Fufeng Agriculture and Animal Husbandry Corporation Ltd. The remaining 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.

All claims expressed in this article are solely those of the authors and do not necessarily represent those of their affiliated organizations, or those of the publisher, the editors and the reviewers. Any product that may be evaluated in this article, or claim that may be made by its manufacturer, is not guaranteed or endorsed by the publisher.