Abstract Background Amuc_1100, the most abundant outer membrane protein of Akkermansia muciniphila, alleviates high-fat diet (HFD)-induced hepatic lipid accumulation and modulates gut microbiota in fish; however, its mechanism and mediators remain unknown. Using zebrafish model, this study aims to determine the mechanism by which Amuc_1100 reduces HFD-induced hepatic lipid accumulation through modulation of gut microbiota. Methods In main study, 1-month-old zebrafish were fed a low-fat diet (LFD), HFD, or HFD supplemented with 0.01% Amuc_1100 (AM0.01) for 4 weeks. Body weight gain, hepatic lipid accumulation, microbial translocation, and gut microbiota composition were evaluated. In parallel, larvae at 5 d post-fertilization were fed the same diets for 7 d and analyzed by Oil Red O staining. In validation experiments, germ-free (GF) zebrafish received microbiota transplants from donor fish fed HFD or AM0.01. Antibiotics (ABS)-treated zebrafish were fed LFD, HFD, or AM0.01 for 4 weeks. Intestinal protein interacting with Amuc_1100 was identified via pull-down and co-immunoprecipitation, and its role was confirmed using protein-protein interaction (PPI) inhibitor BV02 and gene knockdown. Data were analyzed by Student’s t-test or one-way ANOVA. Results Compared with HFD group, zebrafish in AM0.01 group showed lower body weight gain, reduced hepatic lipid accumulation, and decreased microbial translocation (P < 0.05). AM0.01 feeding increased Bacillus abundance while reducing Acinetobacter, Plesiomonas and Aeromonas abundances relative to HFD (P < 0.05). GF zebrafish colonized with microbiota from AM0.01-fed donors showed less hepatic lipid accumulation than those receiving microbiota from HFD-fed donors (P < 0.05). In contrast, ABS-treated zebrafish showed no significant difference in hepatic triacylglycerol content between HFD and AM0.01 groups (P > 0.05). Using pull-down assays with intestinal proteins from LFD-fed zebrafish, we identified 14-3-3β/α-A as an interacting protein of Amuc_1100. When 14-3-3β/α-A PPI was inhibited by BV02, Amuc_1100 failed to alter the HFD-induced gut microbiota profile in 1-month-old zebrafish (P > 0.05). Moreover, either BV02 treatment or 14-3-3β/α-A knockdown abolished the protective effect of Amuc_1100 against hepatic lipid accumulation in conventional and GF zebrafish (P < 0.05). Conclusions Amuc_1100 reduces hepatic lipid accumulation by modulating gut microbiota through intestinal 14-3-3β/α-A, highlighting its potential as a therapeutic target.

Background: Abdominal Aortic Aneurysm (AAA) is a critical global health issue, affecting an estimated up to 8% of men over 65, with a complex etiology involving smoking, age and gender. However, the lack of specific drug treatments underscores the need for a robust in vitro model to advance our comprehension of AAA pathophysiology and serve as an ex vivo surrogate for drug testing. Methods: This study introduces an innovative AAA patient-derived organoid (PDO) model using a non-enzymatic procedure, a Matrigel-free system, and specialized organoid culture medium, leveraging 3-dimensional (3D) cultures to replicate the disease's microenvironment. The stability of this culture system was assessed through microscopic observation, H&E staining, immunohistochemistry (IHC), viability assays, and whole-genome sequencing (WES). Additionally, we conducted pharmacological assessments to explore the effects of drug treatments on AAA PDO. Results: Our model maintains aortic morphological integrity and pathological phenotypes, incorporates the immune microenvironment (validated by IHC markers for macrophages and lymphocytes), and adjacent tissues (loose connective tissue and vegetative blood vessels). The model demonstrates remarkable stability, confirmed by consistent genetic mutation sites throughout cultivation via WES, and cell survival after five weeks in vitro via live-cell staining. Preliminary pharmacological assessments show promising efficacy, with distinct responses to 1 μM metformin, 1 μM RU.521, or 1 μM STING-IN-2 treatments for 48 h via mass spectrometry. Conclusions: The AAA organoid model, to the best of our knowledge, is the first reported abdominal aortic aneurysm PDO model, and signifies a promising step towards therapeutic treatment options for AAA, potentially complementing existing surgical approaches.

BACKGROUND:Ovarian cancer, often diagnosed at advanced stages, has a 5-year survival rate below 50%, indicating a critical need for innovative treatments. The Zengmian Yiliu (ZMYL) formula, a Traditional Chinese Medicine (TCM) prescription, has shown potential in enhancing chemotherapy efficacy and improving patients' quality of life, PURPOSE: To investigate the effects of the ZMYL formula on ovarian cancer organoids, focusing on its impact on organoid phenotypes and underlying mechanisms, and to explore its potential as an immunotherapeutic agent. METHODS:Ovarian cancer organoids were established from surgical tissues and treated with the ZMYL formula at varying concentrations. Network pharmacology was utilized to predict the formula's therapeutic targets and pathways, and molecular docking was conducted to validate ingredient-target interactions. Phenotypic changes were monitored, and RNA sequencing was performed post-treatment to analyze gene expression alterations. RESULTS:A total of 34 overlapping targets of 10 compounds in the ZMYL formula and ovarian cancer were predicted by Network pharmacology analysis. The ZMYL formula induced dose-dependent morphological changes in organoids, including a reduction in size and structural sparsity at higher concentrations. RNA sequencing revealed significant modulation of cell cycle and immune response pathways, with a particular focus on immunomodulatory effects. The formula's treatment targeted key genes involved in these processes, reshaping the tumor's molecular landscape. CONCLUSIONS:This study establishes ZMYL's capacity to simultaneously target oncogenic drivers (e.g., cell cycle regulators) and immune checkpoints (e.g., CXCL10-mediated T cell recruitment) in ovarian cancer organoids. Unlike conventional monotherapy-focused approaches, ZMYL's multi-component mechanism offers a synergistic framework for integrating TCM with modern immunotherapies. These findings provide a foundation for future clinical evaluation of ZMYL as a precision medicine strategy to enhance treatment efficacy and mitigate chemoresistance in ovarian cancer.

In the last decade, organoid research has entered a golden era, signifying a pivotal shift in the biomedical landscape. The year 2023 marked a milestone with the publication of thousands of papers in this arena, reflecting exponential growth. However, amid this burgeoning expansion, a comprehensive and accurate overview of the field has been conspicuously absent. Our review is intended to bridge this gap, providing a panoramic view of the rapidly evolving organoid landscape. We meticulously analyze the organoid field from eight distinctive vantage points, harnessing our rich experience in academic research, industrial application, and clinical practice. We present a deep exploration of the advances in organoid technology, underpinned by our long-standing involvement in this arena. Our narrative traverses the historical genesis of organoids and their transformative impact across various biomedical sectors, including oncology, toxicology, and drug development. We delve into the synergy between organoids and avant-garde technologies such as synthetic biology and single-cell omics and discuss their pivotal role in tailoring personalized medicine, enhancing high-throughput drug screening, and constructing physiologically pertinent disease models. Our comprehensive analysis and reflective discourse provide a deep dive into the existing landscape and emerging trends in organoid technology. We spotlight technological innovations, methodological evolution, and the broadening spectrum of applications, emphasizing the revolutionary influence of organoids in personalized medicine, oncology, drug discovery, and other fields. Looking ahead, we cautiously anticipate future developments in the field of organoid research, especially its potential implications for personalized patient care, new avenues of drug discovery, and clinical research. We trust that our comprehensive review will be an asset for researchers, clinicians, and patients with keen interest in personalized medical strategies. We offer a broad view of the present and prospective capabilities of organoid technology, encompassing a wide range of current and future applications. In summary, in this review we attempt a comprehensive exploration of the organoid field. We offer reflections, summaries, and projections that might be useful for current researchers and clinicians, and we hope to contribute to shaping the evolving trajectory of this dynamic and rapidly advancing field.

Glioblastoma (GBM) is a highly aggressive brain tumor characterized by rapid growth and high heterogeneity, posing challenges for fundamental research and personalized drug screening due to the lack of suitable models. GBM organoids serve as an innovative research tool, providing a valuable model for studying the biological characteristics of GBM. In this study, we successfully generated 4 GBM organoids and employed enzymatic digestion and mechanical fragmentation techniques for subsequent cultivation. Through continuous observation, pathological assessment, and RNA sequencing (RNA-seq), we observed that all the organoids generated through both methods demonstrated good growth characteristics. The organoids derived from mechanical fragmentation not only achieved a two-dimensional (2D) area of ~ 1.5 mm2 but also exhibited distinct vascular structures. The organoids derived from enzymatic digestion achieved a 2D area of approximately 0.8 mm2. Furthermore, RNA-seq analysis has revealed that organoids cultured using two distinct methods exhibit a heterogeneous cellular composition, comprising a total of 20 cell types (endothelial, immune cells ...). Our studies show that both methods successfully maintained the essential characteristics of GBM, encompassing its distinctive tissue structure and gene expression patterns. Each method exhibits its own attributes, contributing to the understanding of GBM organoids.

Ovarian cancer, a common gynecologic tumor, is associated with a high mortality, due to challenges in early detection within the reproductive system. According to our previous research, cultivating patient-specific organoids from mechanically sheared tissues can be utilized for drug response evaluation but has limitations for high-throughput screening efficiency due to their inconsistent size. In this research, we focused on organoids developed from single-cell suspensions to address the critical requirement for uniformity in organoid size. By the day 3 of culture, single-cell suspensions rapidly and spontaneously aggregated into spherical structures with a more consistent size. Notably, the organoids of sample OVA-37 were ten times larger after 8 days of culture. Transcriptomic analysis was used to compare the two organoid culture techniques, demonstrating that the variations between different organoid culture methods were minimal, with higher variability observed among patients. Gene set enrichment analysis (GSEA) revealed only minor discrepancies in specific pathways, such as TGF-u03B2 and tight junctions. Furthermore, treatment with carboplatin in a 96-well plate setup resulted in reproducible drug responses, as evidenced by coefficients of variation lower than 40%. This finding suggests that single-cell suspension-cultured organoids can be employed for reproducible high-throughput drug screening. This approach holds potential for personalized drug screening in ovarian cancer and may contribute to the development of novel therapeutic strategies.

Ovarian cancer is high recurrence and mortality malignant tumor. The most common ovarian cancer was High-Grade Serous Ovarian Cancer. However, High-Grade Serous Ovarian Cancer organoid is rare, which organoid with patient immune microenvironment and blood vessels even absence. Here, we report a novel High-Grade Serous Ovarian Cancer organoid system derived from patient ovarian cancer samples. These organoids recapitulate High-Grade Serous Ovarian Cancer organoids' histological and molecular heterogeneity while preserving the critical immune microenvironment and blood vessels, as evidenced by the presence of CD34 + endothelial cells. Whole exome sequencing identifies key mutations (CSMD3, TP53, GABRA6). Organoids show promise in testing cisplatin sensitivity for patients resistant to carboplatin and paclitaxel, with notable responses in cancer proteoglycans and p53 (TP53) signaling, like ACTG/ACTB1/AKT2 genes and BBC3/MDM2/PERP. Integration of immune microenvironment and blood vessels enhances potential for novel therapies like immunotherapies and angiogenesis inhibitors. Our work may provide a new detection system and theoretical basis for ovarian cancer research and individual therapy.

Ovarian cancer (OC), particularly high-grade serous cancer (HGSC), is the leading cause of mortality among gynecological cancers owing to the treatment difficulty and high recurrence probability. As therapeutic drugs approved for OC, poly ADP-ribose polymerase inhibitors (PARPi) lead to synthetic lethality by inhibiting single-strand DNA repair, particularly in homologous recombination-deficient cancers. However, even PARPi have distinct efficacies and are prone to have drug resistance, the molecular mechanisms underlying the PARPi resistance in OC remain unclear. A patient-derived organoid platform was generated and treated with a PARPi to understand the factors associated with PARPi resistance. PARPi significantly inhibits organoid growth. After 72 h of treatment, both the size of organoids and the numbers of adherent cells decreased. Moreover, immunofluorescence results showed that the proportion of Ki67 positive cells significantly reduced. When the PARPi concentration reached 200 nM, the percentage of Ki67 + /4′,6-diamidino-2-phenylindole (DAPI) cells decreased approximately 50%. PARPi treatment also affected the expression of genes involved in base excision repair and cell cycle. Functional assays revealed that PARPi inhibits cell growth by upregulating early apoptosis. The expression levels of several key genes were validated. In addition to previously reported genes, some promising genes FEN1 and POLA2, were also be founded. The results demonstrate the complex effects of PARPi treatment on changes in potential genes relevant to PARPi resistance, and provide perspectives for further research on the PARPi resistance mechanisms.

OBJECTIVE:Liver cancer (LC) is a frequently occurring lethal malignancy worldwide, yet the molecular mechanisms of carcinogenesis and their development remain uncharacterized. In this study, bioinformatics methods were used to find candidate hub genes for prognosis assessment and clinical treatment of LC.METHODS:Differential analysis was carried out based on the evidence of gene expression profiling in LC on The Cancer Genome Atlas (TCGA). The differentially expressed genes (DEGs) were constructed into co-expression networks and divided into modules by virtue of weighted gene co-expression network analysis (WGCNA). Based on the Gene Ontology (GO) and the Kyoto Encyclopedia of Genes and Genomes (KEGG), the module genes were subjected to functional enrichment analysis. The LC microarray (GSE105130) in the Gene Expression Omnibus was selected to verify the hub genes' expression profiles. The validity of the hub genes was verified via survival analysis, as well as expression correlation with the clinicopathological features. Thereafter, gene set variation analysis (GSVA) and single-sample gene set enrichment analysis (GSEA) were applied to investigate the possible biological functions of the hub genes.RESULTS:In total, 3780 DEGs and 17 co-expression modules were obtained. The blue module had the strongest correlation with the tumour stage and the module genes were principally enriched in tumour-associated GO terms, as well as pathways such as Ras protein signal transduction, ERK1/2 cascade, Ras signal pathway, and ECM-receptor interaction. RASAL1, which is highly expressed in LC, was identified as a hub gene for LC progression. Its high expression suggested unfavorable patient prognosis and was correlated with T stage, gender and tumour stage. Further analysis identified that the overexpression of RASAL1 was substantially enriched in cancer-associated gene sets.CONCLUSION:RASAL1 is a hub gene that influences LC progression, constituting a novel biomarker and molecular target in the future diagnosis and therapy of LC.

Ovarian cancers are the major cause of mortality for women worldwide. This study was aimed to elucidate the biological activities of CCDC106 in the proliferation and invasion of mutant p53 and of wild-type p53 ovarian cancer cells. CAOV3 (mutant p53) cells showed high expression levels of CCDC106, but it was expressed at low levels in SKOV3 (mutant p53) and in A2780 (wild-type p53) cells. The overexpression of CCDC106 promoted the expression of proliferation markers (cyclin family members), invasion and Epithelial-to-mesenchymal transition (EMT) markers (claudin-1, claudin-4, N-cadherin, snail, slug) while the knockdown of CCDC106 inhibited their expression in mutant p53 cells but not in wild-type p53 cells. Treatment with a CK2 inhibitor blocked the translocation of CCDC106 into the nuclei of mutant p53 cells. Immunoprecipitation assays confirmed that ATF4 is a potential binding partner of CCDC106. The overexpression of CCDC106 reduced p21 and p27 protein expression levels while treatment with an ATF4 siRNA rescued their expression. The overexpression of CCDC106 promoted colony formation and invasion of mutant p53 cells, which was suppressed by treatment with an ATF4 siRNA. Immunohistochemistry results showed that CCDC106 and ATF4 are expressed at high levels but p21 is expressed at low levels in FIGO III-IV stage and in mutant p53 ovarian cancer samples. A significant association between poor overall survival and high CCDC106 and ATF4 expression levels was observed in human ovarian cancer samples. In conclusion, CCDC106 promotes proliferation, invasion and EMT of mutant p53 ovarian cancer cells via the ATF4 mediated inhibition of p21.

Purpose Macrophages play crucial roles as early responders to bacterial pathogens and promote/ or impede chronic inflammation in various tissues. Periodontal macrophage-induced pyroptosis results in physiological and pathological inflammatory responses. The transcription factor Dec2 is involved in regulating immune function and inflammatory processes. To characterize the potential unknown role of Dec2 in the innate immune system, we sought to elucidate the mechanism that may alleviate macrophage pyroptosis in periodontal inflammation. Methods Porphyromonas gingivalis lipopolysaccharide (LPS) was used to induce pyroptosis in RAW 264.7 macrophages. Subsequently, we established an LPS-stimulated Dec2 overexpression cellular model in macrophages. Human chronic periodontitis tissues were employed to evaluate potential changes in inflammatory marker expression and pyroptosis. Finally, the effects of Dec2 deficiency on inflammation and pyroptosis were characterized in a P. gingivalis-treated experimental periodontitis Dec2-knockout mouse model. Results Macrophages treated with LPS revealed significantly increased messenger RNA expression levels of Dec2 and interleukin (IL)-1β. Dec2 overexpression reduced IL-1β expression in macrophages treated with LPS. Overexpression of Dec2 also repressed the cleavage of gasdermin D (GSDMD), and the expression of caspase-11 was concurrently reduced in macrophages treated with LPS. Human chronic periodontitis tissues showed significantly higher gingival inflammation and pyroptosis-related protein expression than non-periodontitis tissues. In vivo, P. gingivalis-challenged mice exhibited a significant augmentation of F4/80, tumor necrosis factor-α, and IL-1β. Dec2 deficiency markedly induced GSDMD expression in the periodontal ligament of P. gingivalis-challenged mice. Conclusions Our findings indicate that Dec2 deficiency exacerbated P. gingivalis LPS-induced periodontal inflammation and GSDMD-mediated pyroptosis. Collectively, our results present novel insights into the molecular functions of macrophage pyroptosis and document an unforeseen role of Dec2 in pyroptosis.

α-Galactosidase is a debranching enzyme widely used in the food, feed, paper, and pharmaceuticals industries and plays an important role in hemicellulose degradation. Here, T26, an aerobic bacterial strain with thermostable α-galactosidase activity, was isolated from laboratory-preserved lignocellulolytic microbial consortium TMC7, and identified as Parageobacillus thermoglucosidasius. The α-galactosidase, called T26GAL and derived from the T26 culture supernatant, exhibited a maximum enzyme activity of 0.4976 IU/ml when cultured at 60°C and 180 rpm for 2 days. Bioinformatics analysis revealed that the α-galactosidase T26GAL belongs to the GH36 family. Subsequently, the pET-26 vector was used for the heterologous expression of the T26 α-galactosidase gene in Escherichia coli BL21 (DE3). The optimum pH for α-galactosidase T26GAL was determined to be 8.0, while the optimum temperature was 60°C. In addition, T26GAL demonstrated a remarkable thermostability with more than 93% enzyme activity, even at a high temperature of 90°C. Furthermore, Ca2+ and Mg2+ promoted the activity of T26GAL while Zn2+ and Cu2+ inhibited it. The substrate specificity studies revealed that T26GAL efficiently degraded raffinose, stachyose, and guar gum, but not locust bean gum. This study thus facilitated the discovery of an effective heat-resistant α-galactosidase with potent industrial application. Meanwhile, as part of our research on lignocellulose degradation by a microbial consortium, the present work provides an important basis for encouraging further investigation into this enzyme complex.

Chieko Taguchi, Fuyuki Sato, Chen Wang, Shigeru Nakamura, Kosuke Oikawa, Ujjal Kumar Bhawal, Hiroyuki Okada, and Kazumune Arikawa. Oral Biol Res 2022;46:141-9. https://doi.org/10.21851/obr.46.04.202212.141

The cold-shock protein Y-box-binding protein (YB)-1 regulates the expression of various chemokines and their receptors at the transcriptional level. Expression of the orphan chemokine CXCL14 is repressed by EGF induced signaling. The possible links between EGF-mediated YB-1 and CXCL14 as well as the functions of critical kinase pathways in the progression of prostate cancer have remained unexplored. Here we examined the correlation between YB-1 and CXCL14, and the ERK/AKT/mTOR pathways in prostate cancer. Knockdown of YB-1 decreased cyclinD1 expression with an upregulation of cleaved-PARP in human prostate cancer cells. EGF treatment upregulated phospho-YB-1 expression in a time-dependent manner, while treatment with an ERK inhibitor completely silenced its expression in prostate cancer cells. EGF treatment stimulates CyclinD1 and YB-1 phosphorylation in an ERK-dependent pathway. Positive and negative regulation of YB-1 and CXCL14 was observed after EGF treatment in prostate cancer cells, respectively. EGF rescues cell cycle and apoptosis via the AKT and ERK pathways. Furthermore, YB-1 silencing induces G1 arrest and apoptosis, while knockdown of CXCL14 facilitates cell growth and inhibits apoptosis in prostate cancer cells. YB-1 and CXCL14 were inversely correlated in prostate cancer cells and tissues. A significant association between poor overall survival and High YB-1 expression was observed in human prostate cancer patients. In conclusion, our data reveal the functional relationship between YB-1 and CXCL14 in EGF mediated ERK signaling, and YB-1 expression is a significant prognostic marker to predict prostate cancer.

Ingestion of excess systemic fluoride results in physiological and pathological disturbances of bone homeostasis. We and others have established that treatment with 50 μM sodium fluoride (NaF) is safe and effective for bone remodeling in cellular and animal models. This study aimed to study the effects of treatment with 50 μM NaF on macrophage-driven osteoclastogenesis and to characterize the function of 50 μM NaF in alveolar bone resorption during aging. Murine RAW264.7 monocytic cells were treated with RANKL in the presence or absence of 50 μM NaF. The mRNA expression levels of Cathepsin K and nuclear factor-activated T-cell cytoplasmic 1 (NFATc1), which are involved in the mechanism of osteoclast induction, were measured using quantitative real time RT-PCR. An experimental 50 μM NaF-treated aging mouse model was used to confirm alveolar bone resorption. Micro-CT was used to assess bone loss and immunohistochemistry was performed to detect the protein expression levels of RANKL and Cathepsin K in periodontal tissues. Treatment of RAW264.7 cells with 50 μM NaF repressed osteoclastogenesis. Micro-CT results confirmed that treatment with 50 μM NaF alleviated alveolar bone resorption in aging. Immunohistochemical analysis revealed decreased expression levels of RANKL and cathepsin K in 50 μM NaF-treated mice during aging. Taken together, these results contribute fascinating experimental clues that 50 μM NaF has the potential to function as an anti-resorptive agent during alveolar bone aging.

Biodegradation is the key process involved in natural lignocellulose biotransformation and utilization. Microbial consortia represent promising candidates for applications in lignocellulose conversion strategies for biofuel production; however, cooperation among the enzymes and the labor division of microbes in the microbial consortia remains unclear. In this study, metagenomic analysis was performed to reveal the community structure and extremozyme systems of a lignocellulolytic microbial consortium, TMC7. The taxonomic affiliation of TMC7 metagenome included members of the genera Ruminiclostridium (42.85%), Thermoanaerobacterium (18.41%), Geobacillus (10.44%), unclassified_f__Bacillaceae (7.48%), Aeribacillus (2.65%), Symbiobacterium (2.47%), Desulfotomaculum (2.33%), Caldibacillus (1.56%), Clostridium (1.26%), and others (10.55%). The carbohydrate-active enzyme annotation revealed that TMC7 encoded a broad array of enzymes responsible for cellulose and hemicellulose degradation. Ten glycoside hydrolases (GHs) endoglucanase, 4 GHs exoglucanase, and 6 GHs β-glucosidase were identified for cellulose degradation; 6 GHs endo-β-1,4-xylanase, 9 GHs β-xylosidase, and 3 GHs β-mannanase were identified for degradation of the hemicellulose main chain; 6 GHs arabinofuranosidase, 2 GHs α-mannosidase, 11 GHs galactosidase, 3 GHs α-rhamnosidase, and 4 GHs α-fucosidase were identified as xylan debranching enzymes. Furthermore, by introducing a factor named as the contribution coefficient, we found that Ruminiclostridium and Thermoanaerobacterium may be the dominant contributors, whereas Symbiobacterium and Desulfotomaculum may serve as "sugar cheaters" in lignocellulose degradation by TMC7. Our findings provide mechanistic profiles of an array of enzymes that degrade complex lignocellulosic biomass in the microbial consortium TMC7 and provide a promising approach for studying the potential contribution of microbes in microbial consortia.

The interaction between the microbial communities in aquatic animals and those in the ambient environment is important for both healthy aquatic animals and the ecological balance of aquatic environment. Crayfish ( Procambarus clarkii ), with their high commercial value, have become the highest-yield freshwater shrimp in China. The traditional cultivation in ponds (i.e., monoculture, MC) and emerging cultivation in rice co-culture fields (i.e., rice–crayfish co-culture, RC) are the two main breeding modes for crayfish, and the integrated RC is considered to be a successful rice-livestock integration practice in eco-agricultural systems. This study explored the ecological interactions between the microbial communities in crayfish intestine and the ambient environment, which have not been fully described to date. The bacterial communities in crayfish intestine, the surrounding water, and sediment in the two main crayfish breeding modes were analyzed with MiSeq sequencing and genetic networks. In total, 53 phyla and 1,206 genera were identified, among which Proteobacteria, Actinobacteria, Tenericutes, Firmicutes, Cyanobacteria, Chloroflexi, Bacteroidetes, Acidobacteria, RsaHF231, and Nitrospirae were the dominant phyla. The microbiota composition significantly differed between the water, sediment, and crayfish intestine, while it did not between the two breeding modes. We also generated a co-occurrence correlation network based on the high-confidence interactions with Spearman correlation ρ ≥ 0.75. In the genera co-correlation network, 95 nodes and 1,158 edges were identified, indicating significant genera interactions between crayfish intestine and the environment. Furthermore, the genera clustered into three modules, based on the different environments. Additionally, Candidatus_Bacilloplasma, g_norank_f_Steroidobacteraceae, Dinghuibacter, Hydrogenophaga, Methyloparacoccus , and Defluviicoccus had the highest betweenness centrality and might be important in the interaction between crayfish and the ambient environment. Overall, this study enhances our understanding of the characteristics of the microbiota in crayfish and their surrounding environment. Moreover, our findings provide insights into the microecological balance in crayfish eco-agricultural systems and theoretical reference for the development of such systems.