As far as we know, this is the first prospective study to simultaneously explore the effects of probiotics on gastrointestinal and vaginal adverse events as well as gut and vaginal microbiota in H. pylori eradication, and the first to investigate the gut-vaginal axis at the clinical level. In this study, we confirmed that probiotics could reduce the incidence of bloating, constipation, and excessive vaginal discharge during H. pylori eradication, effectively improving patients' treatment tolerability. High-throughput sequencing further revealed that probiotics positively regulates alterations in gut and vaginal microbiota induced by H. pylori eradication.
H. pylori not only elevates the risk of intragastric diseases like gastric cancer but is also closely associated with various extragastric diseases, and H. pylori eradication is the most effective way to prevent the development and progression of related diseases[5]. With the declining efficacy of the triple therapy, the standard quadruple therapy featuring high-dose antibiotics has emerged as the mainstream regimen for eradicating H. pylori, which leads to increasingly significant treatment-related adverse events[7, 25]. According to previous literature reports[7, 25], the incidence of H. pylori treatment-related adverse events can be as high as 20–40%, significantly affecting patients' treatment tolerability and potentially leading to treatment discontinuation, resulting in eradication failure. Although the role of probiotics in directly increasing the eradication rate of H. pylori is still debatable, there is general consensus on its efficacy in reducing adverse events[9, 17, 23]. Additionally, our previous study has demonstrated that L. plantarum can alleviate gastrointestinal symptoms by improving gut function[26]. Studies by Vladareanu et al. have confirmed the efficacy of oral L. plantarum in alleviating symptoms associated with vaginitis[18]. Interestingly, this study also revealed that during H. pylori eradication, L. plantarum MH-301 effectively decreases the occurrence of specific treatment-related gastrointestinal and vaginal adverse events, including bloating, constipation, and excessive vaginal discharge, thereby improving patients' treatment tolerability. Furthermore, PP analysis indicated that the rates of H. pylori eradication in both the probiotic group and the placebo group were above 90%, with no significant difference, which is consistent with other studies[9, 27].
The gut microbiota is a highly diverse ecosystem that not only influences energy balance and nutrient supply but also participates in physiological processes such as endocrine regulation, immune defense, and inhibition of pathogen colonization[28]. Gut microbiota dysbiosis has been related to a number of diseases, including obesity, diabetes, and inflammatory bowel disease[29]. Similar to previous studies[9, 10], we observed that H. pylori eradication results in decreased diversity and an altered structure of gut microbiota. Specifically, in this study, we observed a notable decline in gut microbiota diversity after H. pylori eradication, accompanied by a reduction in the abundance of the pivotal genus Prevotella and an increase in potentially pathogenic genera such as Streptococcus and Haemophilus. Streptococcus is a common bacterium in pyogenic cocci, mostly opportunistic pathogens, and its excessive proliferation in the gut can disrupt the balance of the microbiota[30]. Haemophilus is typically a common pathogen in the upper respiratory tract and oral cavity, less studied in the intestine, and possibly closely related to invasive infections and gut tumors[31, 32]. It is noteworthy that supplementation with L. plantarum MH-301 can increase the relative abundance of Megamonas, Prevotella, and Lactobacillus, while mitigating the increase of Streptococcus and Haemophilus. Megamonas play a crucial role in nutrition and metabolism by degrading different carbohydrates to produce propionic acid, acetic acid, and lactic acid, which may be associated with improved glucose homeostasis following H. pylori eradication[33, 34]. Prevotella has the ability to modulate immune responses and improve the prognosis of colorectal cancer[35]. Siew et al. further confirmed that Megamonas and Prevotella can reduce the incidence of adverse events such as bloating and constipation after COVID-19 vaccination[36]. Lactobacillus, possessing probiotic characteristics, serves as a beneficial colonizer in the gut, aiding not only in digestion and nutrient absorption but also in preventing Clostridium difficile infections and alleviating gut inflammation[37]. Through correlation analysis between the gut microbiota and gastrointestinal adverse events, we found that Lactobacillus, Dialister, and Prevotella are most closely associated with reducing gastrointestinal adverse events. Furthermore, the correlation between the gut microbiota suggests that Megamonas, Faecalibacterium, Sutterella, and Collinsella may also have a synergistic effect in reducing gastrointestinal adverse events. Thus, it can be seen that L. plantarum MH-301 in H. pylori eradication contributes to modulating the structure of the gut microbiota, which may be a potential mechanism for its ability to reduce gastrointestinal adverse events.
The vaginal microbiota is an essential component of vaginal microbial ecology and significantly influences the health and diseases of the female reproductive system[38]. Lactobacillus predominates in the normal vaginal microbiota, maintaining the acidic environment of the vagina and inhibiting the growth of pathogenic bacteria[39]. Disruption of the vaginal microbiota often leads to vaginal infections, such as aerobic vaginitis, bacterial vaginosis, or vulvovaginal candidiasis[40]. While numerous studies have clearly demonstrated the significant impact of H. pylori eradication on the gut microbiota, research regarding its influence on the vaginal microbiota remains limited. We employed high-throughput sequencing technology, for the first time, to examine the vaginal microbiota following H. pylori eradication, revealing a significant decrease in diversity and Lactobacillus abundance, accompanied by an increase in the pathogenic genus Gardnerella. Fortunately, L. plantarum MH-301 was found to alleviate the impact on Lactobacillus and restrict the enrichment of Gardnerella. Lactobacillus plays a crucial role by adhering to the surface of the vaginal epithelium, competitively inhibiting the colonization of pathogens, producing antimicrobial substances such as bacteriocins and hydrogen peroxide, and significantly contributing to maintaining the stability of the vaginal microenvironment, enhancing immune defenses, and halting the progression of cervical cancer[39, 41]. Gardnerella is often closely associated with disruptions in the vaginal microbiota, considered a primary pathogen leading to the occurrence of bacterial vaginosis, and is also associated with increased risks of pelvic inflammatory disease, preterm birth, and human immunodeficiency virus infection[42]. Furthermore, through correlation analysis of the vaginal microbiota and vaginal adverse events, we found that the increase in Lactobacillus abundance in the vagina plays a crucial role in reducing vaginal adverse events, while Gardnerella plays the opposite role. The correlation among vaginal microbiota suggests a negative correlation between Lactobacillus and most other genera in the vagina, indicating that Lactobacillus, as a dominant genus in maintaining vaginal microbiota homeostasis, has a role in inhibiting the overgrowth of other vaginal microbiota. In summary, L. plantarum MH-301 can mitigate the effects of H. pylori eradication on the vaginal microbiota, potentially elucidating its mechanism of action in the vagina.
Currently, there is no evidence indicating gender differences in H. pylori infection, with females comprising half of the total infected population. In clinical practice, it is essential to consider the impact of H. pylori eradication on the vaginal microbiota in females, to monitor the vaginal microbiota, and to take appropriate preventive and therapeutic measures as necessary. Fortunately, we found that during H. pylori eradication, L. plantarum MH-301 not only regulates the gut microbiota but also exerts a similarly positive regulatory effect on the vaginal microbiota. Correlation analysis revealed a positive correlation between Lactobacillus in gut microbiota and Lactobacillus in vaginal microbiota, as well as a positive correlation between gastrointestinal and vaginal adverse events. In addition, research[18] has demonstrated that administering L. plantarum orally can enhance the colonization of Lactobacillus in the vaginal microbiota via "cross-contamination." The emerging concept of the gut-vagina axis in recent years has offered new perspectives for developing therapeutic approaches aimed at promoting vaginal health in women[19, 43]. Our study is also the first to demonstrate at the clinical level that orally administered L. plantarum MH-301 may act in the vagina through the gut-vagina axis after entering the gastrointestinal tract. However, further research is still needed to clarify the mechanisms underlying the gut-vaginal axis and its implications in disease.
This study also has several limitations. Firstly, we did not rigorously control the symptoms of patients at baseline, and defined H. pylori treatment-related adverse events as the emergence of new symptoms or worsening of pre-existing symptoms based on previous research, inevitably leading to potential false positives. Secondly, we only performed high-throughput sequencing analysis on the bacterial community and did not explore other microorganisms such as fungi and viruses. Additionally, we did not conduct further gynecological examinations, which limited our evaluation of other relevant vaginal microbial indicators such as vaginal cleanliness, pH, and hydrogen peroxide. Lastly, we collected samples only before and after H. pylori eradication, without sampling over a longer time span, which limited our ability to further explore the mechanisms of gut and vaginal microbiota.
Dashed lines indicate statistically insignificant differences. The Figure was partly drawn using Servier Medical Art licensed under a Creative Commons Attribution 4.0 Unported License (https://creativecommons.org/licenses/by/4.0/).