ABSTRACT Background: Alcoholic liver disease (ALD) and metabolic-dysfunction associated steatotic liver disease (MASLD) are common, and gut microbiota (GM) is involved with both. Here we compared GM composition in animal models of MASLD and ALD to assess whether there are specific patterns for each disease. Methods: MASLD model- adult male Sprague Dawley rats, randomized into two groups: MASLD-control (n=10) fed a standard diet; MASLD-group (n=10) fed a high-fat-choline-deficient diet for 16 weeks. ALD model- adult male Wistar rats randomized: ALD-control (n=8) fed a standard diet and water+0.05% saccharin, ALD groups fed with sunflower seed and 10% ethanol+0.05% saccharin for 4 or 8 weeks (ALC4, n=8; ALC8, n=8). ALC4/8 on the last day received alcoholic binge (5g/kg of ethanol). Afterwards, animals were euthanized, and feces were collected for GM analysis. Results: Both experimental models induced typical histopathological features of the diseases. Alpha diversity was lower in MASLD compared with ALD (p<0.001), and structural pattern was different between them (P<0.001). Bacteroidetes (55.7%), Firmicutes (40.6%), and Proteobacteria (1.4%) were the most prevalent phyla in all samples, although differentially abundant among groups. ALC8 had a greater abundance of the phyla Cyanobacteria (5.3%) and Verrucomicrobiota (3.2%) in relation to the others. Differential abundance analysis identified Lactobacillaceae_unclassified, Lachnospiraceae_NK4A136_group, and Turicibacter associated with ALC4 and the Clostridia_UCG_014_ge and Gastranaerophilales_ge genera to ALC8. Conclusion: In this study, we demonstrated that the structural pattern of the GM differs significantly between MASLD and ALD models. Studies are needed to characterize the microbiota and metabolome in both clinical conditions to find new therapeutic strategies. Background (ALD metabolicdysfunction metabolic dysfunction (MASLD common (GM Methods model MASLDcontrol control n=10 n10 n 10 (n=10 MASLDgroup group highfatcholinedeficient high fat choline deficient 1 ALDcontrol n=8 n8 (n=8 water005 water 0 05 water+0.05 ethanol005 ethanol ethanol+0.05 ALC4, ALC (ALC4 n=8. . n=8) ALC48 ALC4/ 5g/kg 5gkg gkg 5g kg g ethanol. ethanol) Afterwards euthanized Results diseases p<0.001, p0001 p p<0.001 , 001 (p<0.001) P<0.001. P0001 P P<0.001 (P<0.001) 55.7%, 557 55.7% 55 7 (55.7%) 40.6%, 406 40.6% 40 6 (40.6%) 1.4% 14 (1.4% samples 5.3% 53 5 3 (5.3% 3.2% 32 2 (3.2% others Lactobacillaceaeunclassified Lactobacillaceae unclassified Lactobacillaceae_unclassified LachnospiraceaeNK4A136group LachnospiraceaeNKAgroup Lachnospiraceae NK4A136 NK A Lachnospiraceae_NK4A136_group ClostridiaUCG014ge ClostridiaUCGge Clostridia UCG 014 ge Gastranaerophilalesge Gastranaerophilales Conclusion study strategies n=1 n1 (n=1 n= (n= water00 water+0.0 ethanol00 ethanol+0.0 (ALC p000 p<0.00 00 (p<0.001 P000 P<0.00 (P<0.001 55.7 (55.7% 40.6 (40.6% 1.4 (1.4 5.3 (5.3 3.2 (3.2 LachnospiraceaeNK NKA NK4A13 ClostridiaUCG 01 (n water0 water+0. ethanol0 ethanol+0. p00 p<0.0 (p<0.00 P00 P<0.0 (P<0.00 55. (55.7 40. (40.6 1. (1. 5. (5. 3. (3. NK4A1 water+0 ethanol+0 p0 p<0. (p<0.0 P0 P<0. (P<0.0 (55. (40. (1 (5 (3 NK4A water+ ethanol+ p<0 (p<0. P<0 (P<0. (55 (40 ( p< (p<0 P< (P<0 (4 (p< (P< (p (P

RESUMO Contexto: A doença hepática alcoólica (DHA) e a doença hepática esteatótica associada à disfunção metabólica (MASLD) são comuns, e a microbiota intestinal (MI) está envolvida em ambas. Aqui, comparamos a composição da MI em modelos animais de MASLD e DHA para avaliar se existem padrões específicos para cada doença. Métodos: Modelo de MASLD - ratos machos adultos da linhagem Sprague Dawley, randomizados em dois grupos: MASLD-controle (n=10) alimentados com uma dieta padrão; grupo MASLD (n=10) alimentados com uma dieta rica em gordura e deficiente em colina por 16 semanas. Modelo de DHA - ratos machos adultos da linhagem Wistar randomizados: DHA-controle (n=8) alimentados com uma dieta padrão e água+0,05% de sacarina; grupos DHA alimentados com semente de girassol e 10% de etanol+0,05% de sacarina por 4 ou 8 semanas (DHA4, n=8; DHA8, n=8). DHA 4/8 no último dia receberam binge alcoólico (5 g/kg de etanol). Posteriormente, os animais foram sacrificados, e as fezes foram coletadas para análise da MI. Resultados: Ambos os modelos experimentais induziram características histopatológicas típicas das doenças. A diversidade alfa foi menor na MASLD em comparação com a DHA (P<0,001), e o padrão estrutural foi diferente entre elas (P<0,001). Bacteroidetes (55,7%), Firmicutes (40,6%) e Proteobactérias (1,4%) foram os filos mais prevalentes em todas as amostras, embora com abundâncias diferenciadas entre os grupos. DHA8 teve uma maior abundância dos filos Cyanobacteria (5,3%) e Verrucomicrobiota (3,2%) em relação aos outros. A análise de abundância diferencial identificou Lactobacillaceae_unclassified, Lachnospiraceae_NK4A136 e Turicibacter associados ao grupo DHA4, e os gêneros Clostridia_UCG_014_ge e Gastranaerophilales_ge associados ao DHA8. Conclusão: Neste estudo, demonstramos que o padrão estrutural da MI difere significativamente entre os modelos de MASLD e DHA. Estudos são necessários para caracterizar a microbiota e os metabólitos ativos em ambas as condições clínicas, a fim de encontrar novas estratégias terapêuticas. Contexto (DHA (MASLD comuns (MI Aqui Métodos Dawley MASLDcontrole controle n=10 n10 n 10 (n=10 1 DHAcontrole n=8 n8 (n=8 água005 água 0 05 água+0,05 etanol005 etanol etanol+0,05 DHA4 (DHA4 n=8. . n=8) 48 4/ 5 ( gkg g kg etanol. etanol) Posteriormente sacrificados Resultados doenças P<0,001, P0001 P P<0,001 , 001 (P<0,001) P<0,001. 55,7%, 557 55,7% 55 7 (55,7%) 40,6% 406 40 6 (40,6% 1,4% 14 (1,4% amostras 5,3% 53 3 (5,3% 3,2% 32 2 (3,2% outros Lactobacillaceaeunclassified Lactobacillaceae unclassified Lactobacillaceae_unclassified LachnospiraceaeNK4A136 LachnospiraceaeNKA Lachnospiraceae NK4A136 NK Lachnospiraceae_NK4A13 ClostridiaUCG014ge ClostridiaUCGge Clostridia UCG 014 ge Gastranaerophilalesge Gastranaerophilales Conclusão estudo clínicas terapêuticas n=1 n1 (n=1 n= (n= água00 água+0,0 etanol00 etanol+0,0 P000 P<0,00 00 (P<0,001 55,7 (55,7% 40,6 (40,6 1,4 (1,4 5,3 (5,3 3,2 (3,2 LachnospiraceaeNK LachnospiraceaeNK4A13 NKA NK4A13 Lachnospiraceae_NK4A1 ClostridiaUCG 01 (n água0 água+0, etanol0 etanol+0, P00 P<0,0 (P<0,00 55, (55,7 40, (40, 1, (1, 5, (5, 3, (3, LachnospiraceaeNK4A1 NK4A1 Lachnospiraceae_NK4A água+0 etanol+0 P0 P<0, (P<0,0 (55, (40 (1 (3 LachnospiraceaeNK4A NK4A água+ etanol+ P<0 (P<0, (55 (4 P< (P<0 (P< (P

Abstract Background: Some evidence suggests an association between gut dysbiosis and cirrhosis progression. The authors investigated Gut Microbiome (GM) influence on 90-day mortality and hospitalization/rehospitalization rates in cirrhotic patients. Methods: Compensated/decompensated outpatients and decompensated inpatients were prospectively included and compared to healthy controls. Clinical, laboratory, GM, and two ratios between phyla were evaluated. Patients were followed up for 90 days for hospitalization/rehospitalization and mortality. Results: 165 individuals were included (50 compensated, 49 decompensated outpatients; 36 decompensated inpatients; 30 healthy), 48.5 % female, mean age was 61, main cirrhosis etiology was hepatitis C (27.3 %), and mostly Child-Pugh (CP) B patients, median MELD of 13. As liver disease progressed, microbiota diversity decreased between the groups (p = 0.05; p < 0.004). There were 9 deaths and 22 hospitalizations or rehospitalizations. GM composition had correlation with norfloxacin (p = 0.36, p = 0.04), encephalopathy (p = 0.31, p = 0.01), lactulose (p = 0.26, p = 0.01), 90-day mortality (p = 0.22, p = 0.04), CP (p = 0.17, p = 0.01), previous 6-month antibiotic use (p = 0.16, p = 0.01), MELD (p = 0.145, p = 0.01), ALBI (p = 0.1, p = 0.04) and 90-day hospitalization/rehospitalization (p = 0.08, p = 0.03). Firmicutes/Bacteroidetes (F/B) and Firmicutes/Proteobacteria (F/P) ratios were progressively lower and more significant and had an association with 90-day mortality (p < 0.001). Three MELD set-points (≥ 15, 18 and 20) were significantly associated with both ratios, with similar accuracies. Conclusions: GM dysbiosis was associated with higher CP, MELD, 90-day mortality and hospitalization/rehospitalization. F/B and F/P ratios were associated with 90-day mortality. Background progression (GM 90day day hospitalizationrehospitalization hospitalization rehospitalization patients Methods Compensateddecompensated Compensated controls Clinical laboratory evaluated Results 16 50 (5 compensated 4 3 healthy, , healthy) 485 48 5 48. female 61 27.3 273 27 (27. %, %) ChildPugh Child Pugh (CP 13 progressed 0.05 005 0 05 0.004. 0004 0.004 . 004 0.004) 2 rehospitalizations 036 0.36 0.04, 0.04 04 031 31 0.31 0.01, 001 0.01 01 0.01) 026 26 0.26 022 0.22 017 17 0.17 6month month 6 016 0.16 0145 145 0.145 1 0.1 008 08 0.08 0.03. 003 0.03 03 0.03) FirmicutesBacteroidetes Firmicutes Bacteroidetes FB F (F/B FirmicutesProteobacteria Proteobacteria FP P (F/P 0.001. 0001 0.001 0.001) setpoints set points ≥ ( 15 20 accuracies Conclusions 27. (27 0.0 00 000 0.00 0.3 02 0.2 014 14 0.14 0. (2