Alteration in gut microbial characteristics of patients with acromegaly.

PURPOSE: Studies on intestinal microbiota in acromegaly are scant. This study aimed to characterize the gut microbiome in patients with acromegaly. METHOD: Stool samples were collected from 11 patients newly diagnosed with acromegaly and 12 healthy controls matched for body mass index (BMI) and age after three days on a standard diet. Clinical and gut microbial composition assessments were performed for the two participant groups using 16S rRNA gene amplicon sequencing. RESULTS: There was no difference in the alpha diversity of the microbiota between the samples from patients with acromegaly and those from the healthy controls. Based on beta diversity measurements, differences in microbial community structures were found to be significant only when compared using the Jaccard similarity index. The corresponding Firmicutes/Bacteroidota ratio tended to be higher in individuals with acromegaly than in healthy controls. The mean relative abundance of Actinobacteriota was 2.3 times higher in the acromegaly patient group than in the control group. Eggerthellaceae, Christensenellaceae, and Bacteroidaceae were among the significantly abundant bacterial families in the samples from the acromegaly patient group, while Butyricicoccaceae and Tannerellaceae were decreased. At the level of the genus, the most discriminative features were the abundance of Prevotella 7, Bacteroides, Senegalimassilia, Enterohabdus, the Family XIII AD3011 group, Howardella, and Hungatella in the samples from the acromegaly patient group. In contrast, the Butyrivibrio and the Eubacterium eligens group were the most discriminative genera for the healthy controls and were significantly less abundant in patients with acromegaly. While there were no significantly differentiated taxa between the diabetic and non-diabetic subgroups, Prevotella_7 was significantly enriched in the osteoarthritis (OA) subgroup. No significant association was found between individual genera and growth hormone (GH) levels and insulin-like growth factor-1 (IGF-1) levels as well as the upper limit of normal (ULN). CONCLUSION: Although alpha and beta diversity were mainly similar between the two groups, significant differences were observed between the acromegaly group and the control group at the family and genus levels. These results suggest that the differences between the microbial communities in patients with acromegaly and those in healthy individuals consist primarily of compositional differences independent of abundance. Prospective studies are needed to further explore the clinical implications of gut microbiome dysbiosis in patients with acromegaly.

Inulin protects against the harmful effects of dietary emulsifiers on mice gut microbiome.

Background: The prevalence of inflammatory bowel diseases is increasing, especially in developing countries, with adoption of Western-style diet. This study aimed to investigate the effects of two emulsifiers including lecithin and carboxymethyl cellulose (CMC) on the gut microbiota, intestinal inflammation and the potential of inulin as a means to protect against the harmful effects of emulsifiers. Methods: In this study, male C57Bl/6 mice were divided into five groups (n:6/group) (control, CMC, lecithin, CMC+inulin, and lecithin+inulin). Lecithin and CMC were diluted in drinking water (1% w/v) and inulin was administered daily at 5 g/kg for 12 weeks. Histological examination of the ileum and colon, serum IL-10, IL-6, and fecal lipocalin-2 levels were analyzed. 16S rRNA gene V3-V4 region amplicon sequencing was performed on stool samples. Results: In the CMC and lecithin groups, shortening of the villus and a decrease in goblet cells were observed in the ileum and colon, whereas inulin reversed this effect. The lipocalin level, which was 9.7 ± 3.29 ng in the CMC group, decreased to 4.1 ± 2.98 ng with the administration of inulin. Bifidobacteria and Akkermansia were lower in the CMC group than the control, while they were higher in the CMC+inulin group. In conclusion, emulsifiers affect intestinal health negatively by disrupting the epithelial integrity and altering the composition of the microbiota. Inulin is protective on their harmful effects. In addition, it was found that CMC was more detrimental to microbiota composition than lecithin.

Crohn's Disease Prediction Using Sequence Based Machine Learning Analysis of Human Microbiome.

Human microbiota refers to the trillions of microorganisms that inhabit our bodies and have been discovered to have a substantial impact on human health and disease. By sampling the microbiota, it is possible to generate massive quantities of data for analysis using Machine Learning algorithms. In this study, we employed several modern Machine Learning techniques to predict Inflammatory Bowel Disease using raw sequence data. The dataset was obtained from NCBI preprocessed graph representations and converted into a structured form. Seven well-known Machine Learning frameworks, including Random Forest, Support Vector Machines, Extreme Gradient Boosting, Light Gradient Boosting Machine, Gaussian Naïve Bayes, Logistic Regression, and k-Nearest Neighbor, were used. Grid Search was employed for hyperparameter optimization. The performance of the Machine Learning models was evaluated using various metrics such as accuracy, precision, fscore, kappa, and area under the receiver operating characteristic curve. Additionally, Mc Nemar's test was conducted to assess the statistical significance of the experiment. The data was constructed using k-mer lengths of 3, 4 and 5. The Light Gradient Boosting Machine model overperformed over other models with 67.24%, 74.63% and 76.47% accuracy for k-mer lengths of 3, 4 and 5, respectively. The LightGBM model also demonstrated the best performance in each metric. The study showed promising results predicting disease from raw sequence data. Finally, Mc Nemar's test results found statistically significant differences between different Machine Learning approaches.

Synthesis and Evaluation of Novel Metacetamol Derivatives with Hydrazone Moiety as Anticancer and Antimicrobial Agents.

By exploiting the wide biological potential of the hydrazone scaffold, a series of hydrazone derivatives were synthesized, starting from N-(3-hydroxyphenyl)acetamide (metacetamol). The structures of the compounds were determined using IR, 1 H and 13 C-NMR, and mass spectroscopic methods. The obtained molecules (3 a-j) were evaluated for their anticancer potential against MDA-MB-231 and MCF-7 breast cancer cell lines. According to the CCK-8 assay, all tested compounds showed moderate to potent anticancer activity. Among them, N-(3-(2-(2-(4-nitrobenzylidene)hydrazinyl)-2-oxoethoxy)phenyl)acetamide (3 e) was found to be the most effective derivative with an IC50 value of 9.89 µM against MDA-MB-231 cell lines. This compound was further tested for its potential effects on the apoptotic pathway. Molecular docking studies was also carried out for 3 e in the colchicine binding pocket of tubulin. Additionally, compound 3 e also demonstrated effective antifungal activity, particularly against Candida krusei (MIC=8 µg/ml), indicating that nitro group at the 4th  position of the phenyl ring was the most preferable substituent for both cytotoxic and antimicrobial activity. Our preliminary findings suggest that compound 3 e could be exploited as a leading structure for further anticancer and antifungal drug development.

Gut Microbiome Alteration after Reboxetine Administration in Type-1 Diabetic Rats.

Antidepressants are drugs commonly used in clinical settings. However, there are very limited studies on the effects of these drugs on the gut microbiota. Herein, we evaluated the effect of reboxetine (RBX), a selective norepinephrine (noradrenaline) reuptake inhibitor (NRI), on gut microbiota in both diabetic and non-diabetic rats. This is the first report of relation between reboxetine use and the gut microbiota to our knowledge. In this study, type-1 diabetes induced by using streptozotocin (STZ) and RBX was administered to diabetic rats and healthy controls for 14 days. At the end of the treatment, stool samples were collected. Following DNA extraction, amplicon libraries for the V3-V4 region were prepared and sequenced with the Illumina Miseq platform. QIIME was used for preprocessing and analysis of the data. As a result, RBX had a significant effect on gut microbiota structure and composition in diabetic and healthy rats. For example, RBX exposure had a pronounced microbial signature in both groups, with a low Firmicutes/Bacteroidetes ratio and low Lactobacillus levels. While another abundance phylum after exposure to RBX was Proteabacteria, other notable taxa in the diabetic group included Flavobacterium, Desulfovibrionaceae, Helicobacteriaceae, Campylobacterales, and Pasteurellacae when compared to the untreated group.

Integrative proteomics and metabolomics approach to elucidate the antimicrobial effect of simvastatin on Escherichia coli.

Globally, simvastatin is one of the most commonly used statin drugs. Its antimicrobial properties have been investigated against various pathogens. However, its effect on biological processes in bacteria has been unclear. This study focused on altered biological and metabolic processes at protein and metabolite levels induced by simvastatin. MS-based proteomics and metabolomics were used to investigate the altered proteins and metabolites between experimental groups. Proteomics results showed that simvastatin induced various antimicrobial targets such as chaperon protein DnaK and cell division protein FtsZ. Metabolomics results revealed phenotypic changes in cells under simvastatin stress. Integrated proteomics and metabolomics result indicated that various metabolic processes were altered to adapt to stress conditions. Energy metabolism (glycolysis, tricarboxylic acid cycle, etc.), amino acid synthesis and ribosomal proteins, and purine and pyrimidine synthesis were induced by the effect of simvastatin. This study will contribute to the understanding of antimicrobial properties of statin drugs.

Comparative Proteomic Analysis of Escherichia coli Under Ofloxacin Stress

Objectives: In the present study, proteomics was utilized to evaluate changes in Escherichia coli proteins in response to ofloxacin to understand the mechanism of action of ofloxacin and the mechanisms of ofloxacin resistance in E. coli. Materials and Methods: Proteomics analysis of E. coli was performed by using liquid chromatography quadrupole time-of-flight mass spectrometry followed by a data processing step using MaxQuant. Functional classification and pathway analysis showed a systematic effect of ofloxacin over E. coli proteome structure. Results: In total, 649 common proteins were identified in the untreated and ofloxacin-treated groups, while 98 proteins were significantly different in the ofloxacin-treated group. Functional classification and pathway analysis showed that ofloxacin has a systematic effect over ribosomal processes, energy pathways (tricarboxylic acid cycle and glycolysis), membrane proteins, microbial targets, and biofilm formation. Conclusion: The results showed that ofloxacin affected many cellular processes and pathways. In addition, proteomic analysis revealed that E. coli develops resistance mechanism with different biological processes.

Gut-Brain-Microbiota Axis: Antibiotics and Functional Gastrointestinal Disorders.

Gut microbiota composition and function are major areas of research for functional gastrointestinal disorders. There is a connection between gastrointestinal tract and central nervous system and this is mediated by neurotransmitters, inflammatory cytokines, the vagus nerve and the hypothalamic-pituitary-adrenal axis. Functional gastrointestinal disorders are prevalent diseases affecting more than one third of the population. The etiology of these disorders is not clarified. Visceral hyperalgesia is the main hypothesis for explaining clinical symptoms, however gut-brain axis disorder is a new terminology for functional disorders. In this review, microbiota-gut-brain axis connection pathways and related disorders are discussed. Antibiotics are widely used in developed countries and recent evidence indicates antibiotic-induced dysbiosis as an important factor for functional disorders. Antibiotics exert negative effects on gut microbiota composition and functions. Antibiotic-induced dysbiosis is a major factor for occurrence of post-infectious irritable bowel syndrome. Cognitive and mood disorders are also frequent in functional gastrointestinal disorders. Animal and human trials show strong evidence for the causal relationship between gut microbiota and brain functions. Therapeutic implications of these newly defined pathogenic pathways are also discussed.

Oxacillinase Gene Distribution, Antibiotic Resistance, and Their Correlation with Biofilm Formation in Acinetobacter baumannii Bloodstream Isolates.

Objectives: The limitations of treatment options in bloodstream infections caused by multidrug-resistant Acinetobacter baumannii (MDRAB) have been related to high morbidity and mortality. The aim of our present study was to determine antimicrobial susceptibility profiles, molecular resistance patterns, and biofilm properties of A. baumannii isolated from bloodstream infections. Materials and Methods: In the present study, a total of 44 A. baumannii bloodstream isolates were included. Antimicrobial susceptibility profiles and biofilm formation ability were assessed. The distribution of class D carbapenemases, ISAba1, ISAba1/blaOXA-23, blaNDM-1, mcr-1, and ompA was investigated by polymerase chain reaction (PCR). Arbitrarily primed-PCR (AP-PCR) was performed to evaluate clonal relationships. Results: A total of 32 isolates were MDRAB, whereas 6 isolates were also resistant to colistin without mcr-1 positivity. All isolates were harboring blaOXA-51 gene, whereas blaOXA-23 positivity was 63.6%. Fifty percent of the isolates had ISAba1. ISAba1 upstream of blaOXA-23 was determined in 18 isolates. None of the isolates were positive for blaNDM-1 gene. Majority of the strains were strong biofilm producers (86.8%). A total of 56.8% of the isolates were positive for ompA gene with no direct association with strong biofilm formation. However, blaOXA-51 + 23 genotype and trimethoprim-sulfamethoxazole resistance showed a significant relationship with biofilm formation. AP-PCR analysis revealed six distinct clusters of A. baumannii. Conclusions: Herein, majority of the A. baumannii blood isolates were characterized as blaOXA-51+OXA-23 carbapenemase genotype and were strong biofilm formers. None of the isolates were positive for blaNDM-1, which was promising. Resistant isolates were tended to form strong biofilms. Our results highlight the emergence of oxacillinase-producing MDRAB isolated from bloodstream with high biofilm formation ability.

Characterization of gut microbiota in polycystic ovary syndrome: Findings from a lean population.

BACKGROUND: Limited available animal and human data suggest an association between dysbiosis of gut microbiota and PCOS. We aimed to determine whether gut microbiota in lean women with PCOS shows any alterations compared to healthy women. MATERIALS AND METHODS: Twenty-four lean patients with PCOS phenotype A according to the Rotterdam 2003 diagnostic criteria and 22 BMI-matched healthy women were included in this study. Anthropometric, hormonal and biochemical measurements were carried out in all participants. 16S rRNA gene V3-V4 region amplicon sequencing was performed on stool samples. Preprocessing of the raw data was performed using QIIME, and both QIIME and R packages were used for microbiome analysis. RESULTS: Bacterial richness and diversity did not show a significant difference between patients and controls. Beta diversity was similar between the groups. However, Erysipelotrichaceae, Proteobacteria, Gammaproteobacteria, Enterobacteriaceae, Planococcaceae, Gemmules and Bacillales were significantly abundant in PCOS group according to LEfSe analysis. Clostridium cluster XVII showed increased abundance in patient group, while Clostridium sensustricto and Roseburia were decreased compared to controls. Random forest prediction analysis revealed Clostridium cluster XIVb as the most discriminative feature of patient group and Roseburia for healthy controls. Testosterone and androstenedione were negatively correlated with alpha and phylogenetic diversity. CONCLUSIONS: Our results suggest that gut microbiome of lean PCOS patients with full phenotype shows compositional alterations with similar bacterial richness and diversity compared to controls and that hyperandrogenism is associated with dysbiosis.

Effects of endothelin receptor blockade and COX inhibition on intestinal I/R injury in a rat model: Experimental research.

BACKGROUND: Intestinal ischemia is a highly morbid and mortal condition with no specific treatment. The present study aimed to investigate the effects of cyclooxygenase (COX) inhibition synchronized with nitric oxide (NO) release and endothelin (ET) receptor blockade on oxidative stress, inflammation, vasoconstriction, and bacterial translocation which occur during ischemia-reperfusion (I/R) injury in in-vivo rat intestinal I/R model. MATERIALS AND METHODS: 36 male Wistar rats were randomly divided into six groups (n = 6). Superior mesenteric artery blood flow (SMABF) was recorded; SMA was occluded for 30 min; SMABF was re-recorded at the beginning of the reperfusion phase. Rats were sacrificed after the reperfusion period of 60 min. Blood and tissue samples were obtained. Acetylsalicylic acid (ASA), NO-ASA, flurbiprofen (FLUR), and Tezosentan (TS) were administered 15 min after ischemia. Histopathological examination, bacterial translocation, and biochemical analysis were performed in plasma and tissue samples. RESULTS: SMABF difference, mean Chiu's score and bacterial translocation were increased in the I/R group and decreased in the treatment groups. Plasma LDH, transaminases, intestinal fatty acid-binding protein (I-FABP), TNF-α, ICAM-1, interferon-gamma (IFN-Ɣ) and proinflammatory cytokine panel; tissue lipid peroxidation, MPO, xanthine oxidase (XO), NO, NF-kB levels and the expression of TNF-α were significantly elevated in the I/R group and markedly decreased in the treatment groups. The tissue antioxidant status was decreased in the I/R group and increased in the treatment groups. CONCLUSION: It is suggested that NO-ASA, TS, and FLUR can be introduced as promising therapeutics to improve intestinal I/R injury. INSTITUTIONAL PROTOCOL NO: 2018-29-05 (Animal Experimentations Ethics Committee, Hacettepe University).

1,3-Disubstituted urea derivatives: Synthesis, antimicrobial activity evaluation and in silico studies.

The development of new antimicrobial compounds is in high demand to overcome the emerging drug resistance against infectious microbial pathogens. In the present study, we carried out the extensive antimicrobial screening of disubstituted urea derivatives. In addition to the classical synthesis of urea compounds by the reaction of amines and isocyanates, we also applied a new route including bromination, oxidation and azidination reactions, respectively, to convert 2-amino-3-methylpyridine to 1,3-disubstituted urea derivatives using various amines. The evaluation of antimicrobial activities against various bacterial strains, Candida albicans as well as Mycobacterium tuberculosis resulted in the discovery of new active molecules. Among them, two compounds, which have the lowest MIC values on Pseudomonas aeruginosa, were further evaluated for their inhibition capacities of biofilm formation. In order to evaluate their potential mechanism of biofilm inhibition, these two compounds were docked into the active site of LasR, which is the transcriptional regulator of bacterial signaling mechanism known as quorum sensing. Finally, the theoretical parameters of the bioactive molecules were calculated to establish their drug-likeness properties.

A single early-in-life antibiotic course increases susceptibility to DSS-induced colitis.

BACKGROUND: There is increasing evidence that the intestinal microbiota plays a crucial role in the maturation of the immune system and the prevention of diseases during childhood. Early-life short-course antibiotic use may affect the progression of subsequent disease conditions by changing both host microbiota and immunologic development. Epidemiologic studies provide evidence that early-life antibiotic exposures predispose to inflammatory bowel disease (IBD). METHODS: By using a murine model of dextran sodium sulfate (DSS)-induced colitis, we evaluated the effect on disease outcomes of early-life pulsed antibiotic treatment (PAT) using tylosin, a macrolide and amoxicillin, a beta-lactam. We evaluated microbiota effects at the 16S rRNA gene level, and intestinal T cells by flow cytometry. Antibiotic-perturbed or control microbiota were transferred to pups that then were challenged with DSS. RESULTS: A single PAT course early-in-life exacerbated later DSS-induced colitis by both perturbing the microbial community and altering mucosal immune cell composition. By conventionalizing germ-free mice with either antibiotic-perturbed or control microbiota obtained 40 days after the challenge ended, we showed the transferrable and direct effect of the still-perturbed microbiota on colitis severity in the DSS model. CONCLUSIONS: The findings in this experimental model provide evidence that early-life microbiota perturbation may increase risk of colitis later in life.

Development of besifloxacin HCl loaded nanofibrous ocular inserts for the treatment of bacterial keratitis: In vitro, ex vivo and in vivo evaluation.

Novel drug delivery systems have emerged to treat bacterial keratitis, an acute infection of the cornea. In this study, besifloxacin HCl loaded insert formulations were designed and investigated in vitro, ex vivo and in vivo for the treatment of bacterial keratitis. Besifloxacin HCl (BH) or BH-hydroxypropyl-beta-cyclodextrin (HP-ß-CD) complex containing poly(caprolactone)/polyethylene glycol (PLC/PEG) fibrous inserts were prepared with an electrospinning method. These fibrous inserts were coated with mucoadhesive polymers such as sodium alginate (SA) or thiolated sodium alginate (TSA). Developed inserts compared to commercially available drug and it was found that coating of the insert surfaces with SA and TSA, increases bioadhesion of the formulations. Insert formulations showed a burst release in the first 2 days followed by a slow-release profile. Ex vivo transport studies showed that HP-ß-CD possessed a drug delivery level close to the commercial drug. Both TSA coated inserts as well as inserts containing HP-ß-CD-drug complex were effectively reducing bacterial keratitis in rabbit eyes upon single-dose application compared to multiple dosing with the commercial drug. Consequently, TSA coated inserts as well as the inserts containing HP-ß-CD-drug complex, may be potential alternatives to conventional market product by reducing the application frequency in the clinic leading to increased patient compliance.

Islamic fasting leads to an increased abundance of Akkermansia muciniphila and Bacteroides fragilis group: A preliminary study on intermittent fasting.

BACKGROUND/AIMS: It has been largely accepted that dietary habits affect intestinal microbiota composition. In this pilot study, we hypothesized that time-restricted feeding, which can be regarded as a type of intermittent fasting, may have a distinct effect on intestinal microbiota. Ramadan fasting is an excellent model to understand how time-restricted feeding affect the microbiota. MATERIALS AND METHODS: A total of nine subjects were included in this study during Ramadan, consisting of 17 h of fasting/day during a 29-day period. Stool samples were collected at baseline and the day of the end of Ramadan. 16S rRNA qPCR assay has been performed for quantification of Akkermansia muciniphila, Faecalibacterium prausnitzii, Bifidobacterium spp., Lactobacillus spp., Bacteroides fragilis group, and Enterobacteriaceae. Blood samples were also collected to test for metabolic and nutritional parameters. RESULTS: A significantly increased abundance of A. muciniphila and B. fragilis group was observed in all subjects after Islamic fasting when compared with the baseline levels (p=0.004 and 0.008, respectively). Serum fasting glucose and total cholesterol levels were also significantly reduced in all of the subjects (p<0.01 and p=0.009, respectively). CONCLUSION: Islamic fasting, which represents intermittent fasting, leads to an increase in A. muciniphila and B. fragilis group, which were considered as healthy gut microbiota members. Although this is a pilot study, which should be tested with larger sample size, there are a very limited number of studies in the literature on fasting and microbiota in human subjects. Thus, our present findings may contribute to the understanding of fasting-gut microbiota interaction.

Long-Term Effects of Early-Life Antibiotic Exposure on Resistance to Subsequent Bacterial Infection.

Early-life antibiotic exposure may provoke long-lasting microbiota perturbation. Since a healthy gut microbiota confers resistance to enteric pathogens, we hypothesized that early-life antibiotic exposure would worsen the effects of a bacterial infection encountered as an adult. To test this hypothesis, C57BL/6 mice received a 5-day course of tylosin (macrolide), amoxicillin (ß-lactam), or neither (control) early in life and were challenged with Citrobacter rodentium up to 80 days thereafter. The early-life antibiotic course led to persistent alterations in the intestinal microbiota and even with pathogen challenge 80 days later worsened the subsequent colitis. Compared to exposure to amoxicillin, exposure to tylosin led to greater disease severity and microbiota perturbation. Transferring the antibiotic-perturbed microbiota to germfree animals led to worsened colitis, indicating that the perturbed microbiota was sufficient for the increased disease susceptibility. These experiments highlight the long-term effects of early-life antibiotic exposure on susceptibility to acquired pathogens.IMPORTANCE The gastrointestinal microbiota protects hosts from enteric infections; while antibiotics, by altering the microbiota, may diminish this protection. We show that after early-life exposure to antibiotics host susceptibility to enhanced Citrobacter rodentium-induced colitis is persistent and that this enhanced disease susceptibility is transferable by the antibiotic-altered microbiota. These results strongly suggest that early-life antibiotics have long-term consequences on the gut microbiota and enteropathogen infection susceptibility.

Synthesis of Disulfide-Bridged N-Phenyl-N'-(alkyl/aryl/heteroaryl)urea Derivatives and Evaluation of Their Antimicrobial Activities.

The discovery of new antimicrobial agents is extremely needed to overcome multidrug-resistant bacterial and tuberculosis infections. In the present study, eight novel substituted urea derivatives (10a-10h) containing disulfide bond were designed, synthesized and screened for their in vitro antimicrobial activities on standard strains of Gram-positive and Gram-negative bacteria as well as on Mycobacterium tuberculosis. According to the obtained results, antibacterial effects of the compounds were found to be considerably better than their antimycobacterial activities along with their weak cytotoxic effects. Molecular docking studies were performed to gain insights into the antibacterial activity mechanism of the synthesized compounds. The interactions and the orientation of compound 10a (1,1'-((disulfanediylbis(methylene))bis(2,1-phenylene))bis(3-phenylurea)) were found to be highly similar to the original ligand within the binding pocket E. faecalis ß-ketoacyl acyl carrier protein synthase III (FabH). Finally, a theoretical study was established to predict the physicochemical properties of the compounds.

Determination of certain bacterial groups in gut microbiota and endotoxin levels in patients with nonalcoholic steatohepatitis.

BACKGROUND/AIMS: In recent years, the role of the gut microbiota has emerged in several diseases. Herein we aimed to determine the fecal microbiota, endotoxin levels, and inflammation markers in patients with nonalcoholic steatohepatitis (NASH) and healthy controls. MATERIALS AND METHODS: A total of 46 NASH patients and 38 healthy controls were included. NASH patients were diagnosed according to the steatosis, activity, and fibrosis score/fatty liver inhibition of progression algorithm. 16S rRNA gene-targeted specific primers were used for quantification of certain bacterial groups, and a plasmid library was constructed and sequenced in order to determine dominant Lactobacillus and Bifidobacterium members in patients and controls. RESULTS: Significantly decreased Akkermansia muciniphila and increased Enterobacteriaceae levels were determined in patients compared to healthy controls even after adjusting for the body mass index (BMI) and age. Patients with ≥F2 fibrosis had significantly higher Enterobacteriaceae levels compared to F0-F1 fibrosis. Serum endotoxin and high-sensitivity C-reactive protein levels were significantly higher in patients group. According to the sequencing results, L. reuteri, which was one of the dominant Lactobacillus species in the patient group, could not be detected in healthy controls. Bifidobacterium infantis was found in the patients' feces but not in the controls. CONCLUSION: We demonstrated a BMI and age-independent association between the presence of NASH and levels of A. muciniphila and Enterobacteriaceae as well as increased endotoxin levels. L. reuteri was abundant in the patient group, suggesting that dominant Lactobacillus species should be considered before probiotic treatments.