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Xie et al. | Journal of Clinical and Translational Research 2024; 10(3): 180-190 185

A that LDA >3 indicates a significant difference. However, given
the large variety of bacteria in the bile, gallbladder mucosa,
and fecal samples from the gallstone and control groups,
LDA >4 was adopted as the threshold value for screening
characteristic bacteria (Figure 6). We found that in the gallstone
group, Prevotella had LDA >4 in the fecal samples; Gamma
proteobacteria, Pseudomonadales, Moraxellaceae, and
Acinetobacter had LDA >4 in the bile sample; Proteobacteria,
Betaproteobacteria, and Burkholderiales had LDA value >4
in the gallbladder mucosa sample. None of the bacteria had
B LDA >4 in gallstone samples, while the representative Bacilli,
Lactobacillales, Enterococcus, and Enterococcaceae had LDA
>3. Bacteria species with LDA >4 in the feces of the control
group included Bacteroidia, Bacteroidales, Bacteroidaceae,
Bacteroides, Clostridia, Clostridiales, Firmicutes, and
Ruminococcaceae (Figure 6).

4. Discussion
Gut microbiota studies have gained momentum in recent
years. The human intestine is colonized by over 100 trillion
bacteria, involved in many body activities. Intestinal dysbiosis
has been associated with various human diseases, such as
kidney stones, obesity, diabetes, osteoporosis, and polycystic
Figure 2. Relative abundance of bacterial flora at the (A) phylum ovary syndrome [19]. The relationship between gallstones and
and (B) genus levels between gallstone patients’ feces and healthy gut microbiota has gradually become a research hotspot. Given
subjects’ feces specimens. Blue: healthy subjects’ feces; red: gallstone that the gut microbiota is subject to many potential influencing
patients’ feces; *P < 0.05. factors, strict inclusion and exclusion criteria were established
for this study. For instance, we excluded patients with severe
A bacteremia, sepsis, and a history of antibiotic or probiotic use
in the past 3 months [20]. Moreover, we excluded patients with
serious comorbidities (e.g., metabolic diseases) [21], prior use of
somatostatin and other drugs affecting gallstone formation [22],
history of intestinal surgery [23], and pregnant women or long-
term contraceptive users. Due to the strict exclusion criteria,
the number of samples enrolled in this study was limited. In
this study, we studied the composition of bacterial communities
in gallstones, bile, gallbladder mucosa, and intestinal samples
from 21 gallstone patients, as well as the gut of 20 normal
B individuals. High-throughput sequencing was used to sequence
V3-V4 fragments of the bacterial 16S rRNA gene. The total
number of sequences obtained was 10 429 883 from 72 samples
for subsequent statistical analysis.
This study found no significant difference in the
abundance and diversity of gut microbiota between patients
in the gallstone and control groups, consistent with previous
literature [18]. However, studies have reported a decrease in
intestinal microbial diversity and the abundance of the genus
Roseburia [24]. This discrepancy in findings warrants further
multicenter studies with larger samples to validate and evaluate
the robustness of our findings. In comparison to the control
group, the gallstone group reported a decreased abundance of
Figure 3. Relative abundance of bacterial flora at the (A) phylum and Achromobacter, Faecalibacterium, and Lachnospira and an
(B) genus levels between the biliary and intestinal tracts of gallstone increased abundance of Enterococcus. Research has found that
patients. Blue: gallbladder mucosa; green: bile; orange: gallstone; Enterococcus can shorten the nucleation time of cholesterol
red: gallstone patients’ feces; *P < 0.05. crystals in simulated bile and promote nuclear activity [25],

DOI: https://doi.org/10.36922/jctr.23.00118