Microbiota of ascitic fluid and other biotopes in liver cirrhosis: a pilot study

Objective. To determine the composition of the microbiota of ascitic fluid in patients with liver cirrhosis and compare it with the intestinal microbiota and urobiota.

Materials and methods. A cross-sectional single-center study was conducted for 5 hospitalized patients with decompensated liver cirrhosis with ascites who, along with standard research methods, underwent metagenomic sequencing of ascitic fluid, urine and feces. High-throughput sequencing was performed using a MiSeq genetic analyzer (Illumina, USA) using a protocol based on the analysis of variable regions of the 16s rRNA gene. The assignment of taxonomic levels and quantitative assessment of the microbiome composition were performed using the Kraken 2 program (PlusPF database dated 06/05/2024). The study is registered in Clinicaltrials.gov (NCT05335213).

Results. A unique composition of the ascitic fluid microbiota represented by 29 types, 35 classes, 67 orders, 129 families, 231 genera, and 266 bacterial species has been determined in patients with liver cirrhosis with ascites in the absence of spontaneous bacterial peritonitis. The Pseudomonadota is the dominant type (95,4% [94,8; 95,9]), the Gammaproteobacteria is the dominant class (47,7% [45,9; 48,4]), the Stenotrophomonas is the dominant genera (47,6% [46,2; 48,0]) in the microbiota of ascitic fluid.

For the first time it has been shown that the similarity of the microbiota of ascitic fluid and urobiota is 68,3%, microbiota of ascitic fluid and gut – 69,0% at the type level.

Conclusion. The study examined the microbiota of ascitic fluid in patients with liver cirrhosis using metagenomic sequencing. Despite the negative results of cultural research methods, it has been shown that ascitic fluid is not a sterile medium and contains a large number of microorganisms, which in most cases have commonalities with the microbiota of the gut and urinary tract.

1. Yu X, Jiang W, Huang X, Lin J, Ye H, Liu B. rRNA Analysis Based on Long-Read High-Throughput Sequencing Reveals a More Accurate Diagnostic for the Bacterial Infection of Ascites. Biomed Res Int. 2021;2021:6287280. DOI: https://doi.org/10.1155/2021/6287280

2. Shi P, Liu J, Liang A, Zhu W, Fu J, Wu X, et al. Application of metagenomic next-generation sequencing in optimizing the diagnosis of ascitic infection in patients with liver cirrhosis. BMC Infectious Diseases. 2024;24(1):503. DOI: https://doi.org/10.1186/s12879-024-09396-9

3. Malaeva EG. Urinary tract infections and microbiota. Health and Ecology Issues. 2021;18(3):5-14. (In Russ.). DOI: https://doi.org/10.51523/2708-6011.2021-18-3-1

4. Feng Y, Chen CL, Chen TH, Liang YH, Chen HL, Lin CY, et al. Application of next-generation sequencing to study ascitic microbiome in cirrhotic patients with or without spontaneous bacterial peritonitis. J Microbiol Immunol Infect. 2015;48(5):504-509. DOI: https://doi.org/10.1016/j.jmii.2014.07.005

5. Wu HX, Wei FL, Zhang W, Han J, Guo S, Wang Z, et al. Clinical Evaluation of Metagenomic Next-Generation Sequencing Method for the Diagnosis of Suspected Ascitic Infection in Patients with Liver Cirrhosis in a Clinical Laboratory. Microbiol Spectr. 2023;11(1):e0294622. DOI: https://doi.org/10.1128/spectrum.02946-22

6. Zapater P, Francés R, González-Navajas JM, de la Hoz MA, Moreu R, Pascual S, et al. Serum and ascitic fluid bacterial DNA: a new independent prognostic factor in noninfected patients with cirrhosis. Hepatology. 2008;48(6):1924-1931. DOI: https://doi.org/10.1002/hep.22564

7. Santiago A, Pozuelo M, Poca M, Gely C, Nieto JC, Torras X, et al. Alteration of the serum microbiome composition in cirrhotic patients with ascites. Sci Rep. 2016;6:25001. DOI: https://doi.org/10.1038/srep25001

8. Efremova I, Maslennikov R, Medvedev O, Kudryavtseva A, Avdeeva A, Krasnov G, et al. Gut Microbiota and Biomarkers of Intestinal Barrier Damage in Cirrhosis. Microorganisms. 2024;12(3):463. DOI: https://doi.org/10.3390/microorganisms12030463

9. Martinez-Gili L, Pechlivanis A, McDonald JAK, Begum S, Badrock J, et al. Bacterial and metabolic phenotypes associated with inadequate response to ursodeoxycholic acid treatment in primary biliary cholangitis. Gut Microbes. 2023;15(1):2208501. DOI: https://doi.org/10.1080/19490976.2023.2208501

10. Fernández J, Bert F, Nicolas-Chanoine MH. The challenges of multi-drug-resistance in hepatology. J Hepatol. 2016;65(5):1043-1054. DOI: https://doi.org/10.1016/j.jhep.2016.08.006

11. Ferstl PG, Müller M, Filmann N, Hogardt M, Kempf VA, et al. Noninvasive screening identifies patients at risk for spontaneous bacterial peritonitis caused by multidrug-resistant organisms. Infect Drug Resist. 2018;11:2047-2061. DOI: https://doi.org/10.2147/IDR.S172587

12. Ahrens AP, Culpepper T, Saldivar B, Anton S, Stoll S, Handberg EM, et al. A Six-Day, Lifestyle-Based Immersion Program Mitigates Cardiovascular Risk Factors and Induces Shifts in Gut Microbiota, Specifically Lachnospiraceae, Ruminococcaceae, Faecalibacterium prausnitzii: A Pilot Study. Nutrients. 2021;13(10):3459. DOI: https://doi.org/10.3390/nu13103459

13. Zhang LJ, Huang WQ, Zhang Y, Zhou YL, Xu HM, et al. Exploring the diagnostic potential of immunoglobulin A-microbiota interplay in liver cirrhosis and spontaneous bacterial peritonitis. Kaohsiung J Med Sci. 2024;40(9):837-851. DOI: https://doi.org/10.1002/kjm2.12876

14. Rodríguez-Negrete EV, Gálvez-Martínez M, Sánchez-Reyes K, Fajardo-Felix CF, Pérez-Reséndiz KE, Madrigal-Santillán EO, et al. Liver Cirrhosis: The Immunocompromised State. J Clin Med. 2024;13(18):5582. DOI: https://doi.org/10.3390/jcm13185582

15. Vu PQ, Thiriveedi M, Patel S, Gopal K. Spontaneous Bacterial Peritonitis: A Rare Incidence by Achromobacter xylosidans. Cureus. 2024;16(8):e67855. DOI: https://doi.org/10.7759/cureus.67855

16. Pan S, Moming Z, Awuti A, Zhou K, Tuerxun M, Chong Y, et al. Clinical Insights into Brucella Peritonitis: A Comprehensive Analysis of Four Cases. J Epidemiol Glob Health. 2024;14(3):1300-1304. DOI: https://doi.org/10.1007/s44197-024-00287-5

17. Goelz H, Wetzel S, Mehrbarzin N, Utzolino S, Häcker G, Badr MT. Next- and Third-Generation Sequencing Outperforms Culture-Based Methods in the Diagnosis of Ascitic Fluid Bacterial Infections of ICU Patients. Cells. 2021;10(11):3226. DOI: https://doi.org/10.3390/cells10113226

18. He T, Luo N, Kang J, Ling N, Zhang D. Use of metagenomic next-generation sequencing for diagnosis of peritonitis in end-stage liver disease. Int J Med Sci. 2023;20(13):1698-1704. DOI: https://doi.org/10.7150/ijms.89242

19. Liu B, Zhou Z, Jin Y, Lu J, Feng D, Peng R, et al. Hepatic stellate cell activation and senescence induced by intrahepatic microbiota disturbances drive progression of liver cirrhosis toward hepatocellular carcinoma. J Immunother Cancer. 2022;10(1):e003069. DOI: https://doi.org/10.1136/jitc-2021-003069

20. Gromova GG, Verizhnikova LN, Karpin VA. The role of intestinal dysbiosis in the occurrence of urinary tract infection. Bulletin of SurGU. Medicine. 2019;2 (40):86-89. (In Russ.).

21. Iqbal ZS, Halkjær SI, Ghathian KSA, Heintz JE, Petersen AM. The Role of the Gut Microbiome in Urinary Tract Infections: A Narrative Review. Nutrients. 2024;16(21):3615. DOI: https://doi.org/10.3390/nu16213615