In this study, we found that there were differences in the gut microorganisms between in healthy people HCA, HT1A with normal thyroid function in HT, and HT2A patients with hypothyroidism in HT, and these differences were enhanced as the severity of HT increased, indicating that HT and the consequent thyroid dysfunction have an important impact on the gut microbial composition. Studies have also found that gender has a strong impact on gut microbes. In healthy people, gender becomes the most important factor affecting gut microbial composition, and although hormonal effects gradually replace the role of gender with the development of HT, the impact of HT on gut microbes in different genders remains, suggesting that the impact caused by gender-based factors must be considered in the process of continuing in-depth study as well as in future gut microbiota-based interventions, so as to achieve more precise treatment and intervention.
Differences in Gut microbiota between healthy people and patients with Hashimoto’s thyroiditis and their mediated functional correlations
This study found that Lachnoclostridium, Holdemania, Eggerthella, andBilophila were enriched in the HCA group; Akkermansia, Lachnoclostridium, and Holdemania were enriched in HT1A; and Akkermansia, Acetitomaculum, Shuttleworthia, Flavobacteriaceae, unclassified_c__Clostridia, Lachnospiraceae_NC2004_group, and unclassified_o__Oscillospirales were enriched in HT1A. A previous study found that Lachnoclostridium was significantly enriched in rectal adenomas and could be used as a new biomarker for the non-invasive diagnosis of colorectal adenomas . In addition, it was found to be significantly enriched in the guts of patients with cholestatic liver disease  and atherosclerosis . The above results suggest that this bacterium maybe a biomarker that has a strong correlation with diseases, but it is interesting that this bacterium is enriched in HCA, and its abundance is higher than that in HT patients. However, the HT1A group still maintained a relatively high abundance, and the abundance of this bacteria was reduced in the HT2A group. While Holdemania, which was found to be enriched in HCA and HT1A patients, was found to be an important target of the Chinese herbal medicine known as Erchendecoction (ECD) and to enhance insulin sensitivity in previous studies , it was significantly decreased in the HT2A group. Additionally, it was found that Bilophila, a lipopolysaccharide-producing bacterium, exacerbates inflammation and leads to metabolic disturbances in HFD-mice on a high-fat diet in adulthood  and that improvements in body weight and glycolipid metabolism are inversely correlated with the relative abundance of Bilophila. These bacteria were also enriched in HCA, but decreased in HT2A. Akkermansia is a normal bacterium in the human gut and is a Mucin-degrading bacterium that is negatively correlated with obesity, diabetes, cardiovascular disease, and low-grade inflammation. It is a class of recently discovered star probiotics that has shown great potential for a number of applications  and has been found to be enriched in HT1A and HT2A. Likewise, Lachnospiraceae, which was enriched in HT1A and HT2A, were identified as being inversely associated with colitis and was able to protect mice from colitis-induced death . Shuttleworthia was found to be enriched in systemic lupus erythematosus (SLE) gut and was significantly reduced after treatment . The SLE is a typical autoimmune disease characterized by chronic inflammation and multiple organ damage, the patient’s self-tolerance is reduced, and autoantibodies continue to be produced. This disease and HT are both autoimmune diseases, thus the Shuttleworthia may possess the potential to be a biomarker of autoimmune diseases. In addition, gut microbes and their metabolites were found to play a key role in the development of hepatocellular carcinoma (HCC), and Clostridia was the dominant genus comprising the gut microbes in the HCC group, suggesting bacteria of this genus negatively regulate health .
Interestingly, some bacterial groups are strongly associated with disease, such as Lachnoclostridium and Bilophila, both of which were enriched in the HCA group, but decreased in the HT2A group; however, some probiotics, such as Akkermansia and Lachnospiraceae, were enriched in the HT2A group instead. Some disease-associated bacteria, such as Shuttleworthia and Clostridia, among others, were enriched in the HT2A group. The results suggest that although the gut microbiota of HT patients has changed, is the change in the direction of becoming healthier? Does it help the host to fight the disease, or does it promote the development of the disease? Conclusions cannot be drawn from the current data. In this study, genera were identified by high-throughput sequencing based on 16S rDNA. Previous studies have found that there may be great differences in the functions between different strains of the same strain , which is also the reason for the current situation. Therefore, the relationship between these strains and host health and disease needs further study.
The effect of gender on the flora and its significance
Bifidobacterium is a bacterium that produces short-chain fatty acids (SCFAs), which degrade polysaccharides and dietary fiber. The relative abundance of Bifidobacterium is directly associated with improved body weight and glucose and lipid metabolism . This group of bacteria has long been considered to be positively related to health. In this study, the abundance of Bifidobacterium in healthy people was lower than that in HT patients, and the abundance in females was much higher than it was in males. However, in HT patients, the reversal occurred in both HT1A and HT2A, and the abundance in males was higher than it was in females. The total abundance increased with as the HT levels increased. The levels of Klebsiella bacteria were higher in males than in females and showed an initial increasing trend and then a decreasing trend as the HT level became more aggravated. In the HT2A group in particular, HT2AF was significantly reduced. Bifidobacterium and Klebsiella are two types of bacteria that are very variable in healthy people and in HT patients, but it was interesting that the Spearman correlation analysis showed that these two types of bacteria were only regulated according to sex, especially Bifidobacterium, which was found both in patients and in healthy people of both sexes. A reversal in the species abundances determining the diversity was observed. At the same time, the results show that HT not only leads to differences in the gut microbiota between males and females, but also suggests that HT may have different effects and symptoms in males and females, indicating that gender factors need to be considered in the process of clinical intervention and treatment. Another interesting phenomenon is that Bifidobacterium and Klebsiella are two types of very typical microbes found in the gut microbiota. Among them, Bifidobacterium is usually related to intestinal health, while Klebsiella is an opportunistic pathogen that is often accompanied by disturbances in the gut microbiota, many of which are caused by external conditions. Previous studies have found that Klebsiella is involved in the regulation of the gut microbiota–immune system–brain axis, and the overgrowth of Klebsiella in the gastrointestinal tract may be directly related to the increase in the number of specific immune cells and the occurrence of neurological damage observed in premature infants . In this study, it was found that the abundance of Bifidobacterium in the guts of HT patients increased with the development of the disease, while Klebsiella showed an initial increasing and then decreasing, especially in HT2F group was significantly lower than the healthy population. Is this the result of natural changes in the flora caused by hormone disorder? Or is it the active regulation taken by the body to protect itself and cope with the adverse effects of diseases, so as to use the immune system and other related metabolic systems? Or is it caused by the insufficient sample size of HT2A? The mechanism of this phenomenon deserves further study based on animal models.
Regulation and significance of hormones on bacterial flora
This study found that hormones can significantly affect the gut microbiota of HT patients. In the HT1A group, FT3 was significantly positively correlated with Romboutsia, Lachnoclostridium, and Monoglobus, but was significantly negatively correlated with Faecalibacterium. In the HT2A group, FT3 was significantly positively correlated with Haemophilus, Veillonella, and Streptococcus, and FT4 was significantly negatively correlated with norank_f__norank_o__Clostridia_UCG-014, Faecalibacterium, Lachnospiraceae_NK4A136_group, Eubacterium_ruminantium_group, and Lachnospiraceae_UCG-001. Previous studies have found that Romboutsia can regulate the behavior of mice through neuroactive metabolites, indicating that it is related to autism spectrum disorder (ASD). A positive correlation was observed, and as FT3 decreased, the abundance of the bacteria also decreased, suggesting that thyroiditis-mediated gut microbes may affect autistic behavior in the host . Haemophilus is negatively correlated with carotid atherosclerosis (CAS), and the abundance of this bacterium is decreased in the guts of CAS patients, suggesting that this bacterium may play an important role in regulating health . In HT2A, FT3 was positively correlated with Haemophilus, suggesting that asFT3 decreases, the abundance of this bacteria is decreased, and the composition of the gut microbiota of patients begins to develop in an unhealthy direction. On the other hand, Faecalibacterium is one of the most important bacteria in the human gut microbiota as well as an important producer of butyric acid, playing important anti-inflammatory roles and protecting the digestive system from intestinal pathogens . In the HT2A group, FT4 was negatively correlated with this bacterium. With the decreased of FT4, the abundance of Faecalibacterium increased, resulting in a protective effect on the intestinal tract. Lachnospiraceae, a class of probiotics that regulates colitis and helps the host to defend against disease , is negatively correlated with FT4, and with the decrease in FT4 in the HT2A group, the abundance of this bacteria also increased, resulting in a benefit to the host. Therefore, from the perspective of changes in gut microbiota regulated by hormones, some key probiotics are enriched in HT patients, while some bacteria associated with diseases are also enriched. The comprehensive result is that the function of gut microbiota changes in a beneficial or harmful direction, which still needs further study.
Significance of this study
The results of this study indicate that both the occurrence and development of HT have an impact on the gut microbiota. Whether it is an overall change or regulation based on sex or hormones, two opposite results were produced: on the one hand, in the intestinal tracts of HT patients, the abundances of some beneficial bacteria in the tract increase, and on the other hand, some microorganisms associated with the disease also increase. What impact will this change pattern in the gut microbiota have on host physiology and health? Whether it further exacerbates the disease or helps to delay and fight the disease is currently unknown. Of course, we know that, as the microbiota is a micro-ecosystem, it is impossible to change simple probiotics or pathogenic bacteria so that the gut composition changes in a single direction. These changes are complex and are influenced by both probiotics or pathogenic bacteria. Therefore, in the future, when we study the function of gut microbes, we cannot solely consider the function caused by the changes in a certain type of strain, but also need to use systematic thinking to make comprehensive judgments. On the other hand, through this study, we found that gender is an important regulator of gut microbes and responds to HT. Therefore, in the future interventions and treatments of thyroiditis, in addition to paying attention to the role of hormones, gender-mediated functional heterogeneity must also be considered in its entirety. Another thing worth noting is that, although we have collected samples for more than three years, but as we set strict standards for the included samples, many HT patients with thyroid dysfunction who have been treated are excluded, which leads to difficulties in sample collection, so the number of samples in this group is too small. Although the number of cases in this group is small, we also found some significant phenomena that distinguish other groups. From the experimental data, the data of HT2A group conforms to the trend of gradual development of a disease from HCA to HT1A, and then to HT2A. This is a very interesting phenomenon, and it's also logical. These phenomena have a good guiding significance for the subsequent collection of large samples for in-depth research. Of course, in order to be more rigorous, we should continue to increase the collection of samples in the future to avoid false estimation of the true changes of gut microorganisms caused by insufficient sample size.