Gut microbiota alteration and modulation in hepatitis B virus-related fibrosis and complications: Molecular mechanisms and therapeutic inventions

Table 1 Gut microbiota alteration and additional findings in hepatitis B virus-related fibrosis

Ref.	Population (n)	Detection method	Gut microbiota alteration	Additional findings
Lu et al[30]	Healthy volunteers (n = 32); HBV carriers (n = 30); CHB (n = 31); Decompensated HBV-LC (n = 31)	qPCR	Phylum	Copies of operons that code for virulence factors markedly increased. Fecal sIgA and TNF-α in decompensated HBV-LC patients were higher than other groups
			Bacteroidetes ↓
			Firmicutes ↓
			Family
			Bifidobacteria/Enterobacteriaceae ↓
Xu et al[142]	Healthy volunteers (n = 15); CHB (n = 16); HBV-LC (n = 16)	qPCR	Species	B. dentium, which was considered to be an opportunistic pathogen, increased in HBV-LC patients. Species composition of Bifidobacterium shifted from beneficial to pathogenic
			(Bifidobacterium specific)
			B. catenulatum ↓
			B. longum ↓
			B. dentinum ↑
Wu et al[143]	Healthy volunteers (n = 38); Decompensated HBV-LC (n = 61); HBV-LT (after LC) (n =74)	qPCR	Species (Lactobacilli specific)	Less complex fecal lactobacilli composition was found especially in decompensated HBV-LC patients
			L. rhamnosus ↓
			L. fermentus ↓
Wei et al[38]	Healthy volunteers (n = 120); HBV-LC (n = 120): CTP-A (n = 40); CTP-B (n = 40); CTP-C (n = 40)	Solexa sequencing	Phylum	A negative correlation was observed between the Child-Turcotte-Pugh scores and Bacteroidetes (P < 0.01)
			Bacteroidetes ↓
			Proteobacteria ↑
			Family
			Enterobacteriaceae ↑
			Genera
			Veillonella ↑
Wang et al[23]	Healthy volunteers (n = 22); CHB (n = 85): CP-A (n = 76); CP-B (n = 9)	16S rRNA sequencing	Family	Streptococcus, Veillonella, Streptococcus and Haemophilus had strong correlations with liver function indices and serum metabolites. They were significantly higher in patients with higher Child-Pugh scores. The gut microbiota may be partially involved in the abnormal accumulation of serum metabolites
			Lachnospiraceae ↓
			Rikenellaceae, ↓
			Porphyromonadaceae ↓
			Ruminococcaceae ↓
			Veillonellaceae ↑
Deng et al[29]	Healthy volunteers (n = 20); HBV-LC (n = 80): CP-A (n = 30); CP-B (n = 31); CP-C (n = 19)	16S rRNA sequencing	Phylum	Gut microbiota alteration mentioned on the left were all independent risk or protective factors for HBV-LC. Serum endotoxin increased in patients with higher CP classes (P = 0.000)
			Firmicutes/Bacteroidetes ↑
			Genera
			Megamonas ↓
			Veillonella ↓
Zeng et al[140]	Healthy volunteers (n = 15); CHB (n = 21); HBV-LC (n = 25); HBV-HCC (n = 21)	16S rRNA sequencing	Phylum	Higher Bacteroidetes/firmicutes ratio represented for higher LPS exposure
			Proteobacteria ↑
			Bacteroidetes ↑
			Firmicutes ↓
			Family
			Bifidobacteria/Enterobacteriaceae ↓
Wang et al[59]	Healthy volunteers (n = 21); CHB (n = 69); F0-1 (n = 25); F2-4 (n = 44)	16S rRNA sequencing	Genera	Genera responsible for bile acid metabolism decreased in CHB fibrosis patients
			Prevotella ↑
			Bacteroides ↓
			Ruminococcus ↓
Chen et al[28]	Healthy volunteers (n = 21); HBV carriers (n = 23); CHB (n = 28); HBV-LC (n = 25)	16S rRNA sequencing	Phylum	HBV-LC patients had higher bacterial network complexity with lower abundance of potential beneficial bacterial taxa
			Actinobacteria ↑
			Bacteroidetes ↓
			Firmicutes ↓
			Proteobacteria ↑
Yang et al[27]	Healthy volunteers (n = 31); HBV carriers (n = 24); CHB (n = 56); HBV-LC (n = 54); HBV-ACLF (n = 52)	16S rRNA sequencing	There are fluctuations in the changes	HBV carriers might be the most suitable donors for FMT for higher α diversity and abundance of potential beneficial bacteria
Wang et al[37]	Healthy volunteers (n = 877); CHB (n = 252); HBV-LC (n = 162); HBV-ACLF (n = 212)	16S rRNA sequencing; metagenomic sequencing	Species	High abundance of Enterococcus is associated with progression while that of Faecalibacterium is associated with regression of HBV-ACLF
			Enterococcus faecium ↑

HBV: Hepatitis B virus; CHB: Chronic hepatitis B; ACLF: Acute-on-chronic liver failure; CP: Child-Pugh scores; CTP, Child-Turcotte-Pugh scores; FMT: Faecal microbiota transplantation; LC: Liver cirrhosis; LT: Liver transplant; qPCR: Quantitative polymerase chain reaction.

Table 2 Gut microbiota-related treatment toward hepatitis B virus-related fibrosis and complications (studies in animal models)

Ref.	Study populations (n)	Treatment and grouping (n)	Conclusions
Antiviral therapy
Li et al[96]	AAV-mediated persistent HBV infection (AAV-HBV) mice (n = 10)	35 d after HBV infection, 4 wk of daily ETV treatment. ETV (n = 5)	Gut microbiota dysbiosis of the AAV-HBV mice was reversed by ETV treatment with restored α diversity and changed proportion of Akkermansia, Lacnospiracea and Marvinbryantia
Rifaximin
Kang et al[105]	Germ-free mice (n = 16)	15 d of rifaximin 100 mg/(kg·d), or humanized with stools from a HCV-induced cirrhotic patient with MHE. Rifaximin (n = 4); Humanized (n = 4); Rifaximin + humanized (n = 4)	Rifaximin beneficially altered intestinal ammonia generation by regulating intestinal glutaminase expression independent of gut microbiota. MHE-associated fecal colonization resulted in intestinal and systemic inflammation. It was ameliorated with rifaximin
Engineered probiotics
Nicaise et al[120]	Ornithine transcarbamoylase-deficient Sparse-fur mice; Carbon tetrachloride rats; Thioacetamide-induced acute liver failure mice	NCIMB8826 (wild-type strain Lactobacillus plantarum), or EV101 (engineered Lactobacillus plantarum, LDH-/AlaD+) oral and intrarectal administration	EV101 administration was effective in controlling hyperammonemia in constitutive animal models with a significant effect on survival, which might be involved with direct ammonia consumption in the gut
Kurtz et al[121]	Ornithine transcarbamylase-deficient spfash mice; Thioacetamide-induced acute liver failure mice; Healthy volunteers (n = 52)	Non-modified Escherichia coli Nissle 1917 (EcN), SYNB1020 (engineered EcN, ΔargR, ΔthyA, malEK:PfnrS-argAfbr) administration	SYNB1020 converted NH3 to l-arginine in vitro, and reduced systemic hyperammonemia, improved survival in mouse models. SYNB1020 was well tolerated in healthy volunteers
Ochoa-Sanchez et al[122]	Bile-duct ligated rats	Non-modified EcN, S-ARG, or S-ARG + BUT administration	S-ARG converted ammonia to arginine, it was further modified to additionally synthesize butyrate, which had the potential to prevent HE
FMT
Liu et al[134]	Germ-free mice	Sterile supernatant or entire stool from pre-FMT and post-FMT cirrhotic patients with HE was transplanted to Germ-free mice	Transferred microbiota mediated neuroinflammation. Cirrhosis-associated dysregulation of gut microbiota was related with frontal cortical inflammation

AAV: Adeno-associated virus; HBV: Hepatitis B virus; ETV: Entecavir; HCV: Hepatitis C virus; MHE: Minimal hepatic encephalopathy; HE: Hepatic encephalopathy; FMT: Fecal microbiota transplantation.

Table 3 Gut microbiota-related treatment toward hepatitis B virus-related fibrosis and complications (studies in human)

Ref.	Study populations (n)	Treatment and grouping (n)	Conclusions
Antiviral therapy
Lu et al[97]	Healthy volunteers (n = 30); CHB (n = 30)	8 wk of daily ETV treatment. ETV (n = 30)	After ETV treatment, gut microbiota abundance increased markedly, blood biochemical, immunological and virological responses improved significantly
Lu et al[98]	Healthy volunteers (n = 30); CHB patients (n = 60)	8 wk of daily ETV treatment, or with additional CB. ETV (n = 30); ETV + CB (n = 30)	Additional CB fail to improve blood biochemical, immunological and virological responses, but affects the gut microbiota in the CHB patients treated with ETV
Rifaximin
Bajaj et al[104]	Decompensated LC patients with MHE (n = 20):CHB (NM)	8 wk of rifaximin 550-mg BD. Rifaximin (n = 20)	Rifaximin affected little on gut microbiota, there was just a modest decrease in Veillonellaceae and increase in Eubacteriaceae. Rifaximin significantly improved cognition and endotoxemia, it increased increase in serum saturated and unsaturated fatty acids post-rifaximin
Lutz et al[144]	Decompensated LC patients with ascites (n = 152): Viral hepatitis (n = 35)	Prophylactic antibiotic treatment before the time of paracentesis. Rifaximin (n = 27); Other systemic antibiotics (n = 17)	Prophylactic rifaximin did not reduce SBP occurrence. Prophylactic rifaximin was associated with the dominant bacteria in ascites: Escherichia coli and enterococci were dominant of patients without prophylaxis, klebsiella species were mostly recovered from the rifaximin group
Kimer et al[102]	Decompensated LC patients (n = 54): CHB (NM)	4 wk of rifaximin 550-mg BD or placebo BD. Rifaximin (n = 36); Placebo (n = 18)	Rifaximin had minor effects on bacteria translocation and gut microbiota. Rifaximin showed little impact on the inflammatory state (reflected as the level of TNF-α, IL-6, IL-10, IL-18, SDF-1α, TGF-1β, CRP)
Kaji et al[103]	Decompensated LC patients (n = 30): CHB (n = 4)	4 wk of rifaximin 1200 mg/d. Rifaximin (n = 30)	Rifaximin alleviated HE and endotoxemia with improved intestinal hyperpermeability, and it is involved in a gut microbial change. Rifaximin didn’t affect serum proinflammatory cytokine levels (TNF-α, IL-6, IFN-γ, IL-10)
Probiotics
Agrawal et al[117]	LC patients with recovered HE (n = 235): CHB (n = 49)	3 mo of lactulose 30–60 mL/d, or 3 capsules of probiotics per day, which contained 4 strains of Lactobacillus. Lactulose (n = 80); Probiotics (n = 77)	Lactulose and probiotics were effective for secondary prophylaxis of HE in cirrhotic patients
Ziada et al[115]	Decompensated LC patients with MHE (n = 90): CHB (NM)	4 wk of lactulose 30–60 mL/d, or 3 capsules of probiotics per day, which contained Lactobacillus acidophilus. Lactulose (n = 30); Probiotics (n = 30)	Probiotic was better tolerated than lactulose. Both of them can improve blood ammonia and psychometric tests and reduce the risk of developing overt HE. Magnetic resonance spectroscopy showed more improvement in the levels of brain neurometabolites in the probiotic group
Xia et al[116]	Decompensated HBV-LC patients with MHE (n = 67)	3 mo of probiotics 1500-mg TD, which contained Clostridium butyricum combined with Bifidobacterium infantis. Probiotics (n = 30)	After probiotics treatment, the therapeutic bacteria were significantly enriched, while Enterococcus and Enterobacteriaceae were decreased. Probiotics contributed to the improved cognition and the decreased ammonia levels
FMT
Ren et al[132]	CHB with positive HBeAg, received over 3 yr of antiviral treatment (n = 18)	FMT was performed by gastroscope every 4 wk until HBeAg clearance. FMT (n = 5)	FMT was effective on HBeAg-positive CHB, especially in patients who could not cease the oral antiviral treatment even after long-term treatment
Bajaj et al[135]	Decompensated LC patients with recurrent HE (n = 20). CHB (NM)	After 5 d of antibiotics, FMT was performed by enema, or standard of care (SOC, rifaximin/lactulose) was applied. FMT (n = 10); SOC (n = 10)	FMT increased diversity and beneficial taxa of gut microbiota, improved cognition and showed good tolerance, other than SOC
Bajaj et al[136]	Decompensated LC patients with recurrent HE (n = 20). CHB (NM)	FMT was performed by enema, or standard of care (SOC, rifaximin/lactulose) was applied. FMT (n = 10); SOC (n = 10)	Oral FMT capsules are safe and well tolerated. Post-FMT, duodenal mucosal diversity increased with higher Ruminococcaceae and Bifidobacteriaceae and lower Streptococcaceae and Veillonellaceae. Reduction in Veillonellaceae were noted post-FMT in sigmoid and stool
Chauhan et al[133]	CHB with positive HBeAg, received over 1 years of antiviral treatment (n = 18)	6 FMTs were performed by gastroscope every 4 wk FMT (n = 12)	FMT appeared to be safe and effective on HBeAg-positive CHB in viral suppression and HBeAg clearance

CHB: Chronic hepatitis B; CB: Clostridium butyricum; CRP: C-reactive protein; EcN: Escherichia coli Nissle 1917; ETV: Entecavir; HBeAg: Hepatitis B e antigen; HE: Hepatic encephalopathy; IFN: Interferon; IL: Interleukin; LC: Liver cirrhosis; MHE: Minimal hepatic encephalopathy; NM: Not mentioned; SBP: Spontaneous bacterial peritonitis; SDF-1α: Stromal cell-derived factor 1-α; TDF: Tenofovir disoproxil fumarate; TGF-1β: Transforming growth factor β-1; TNF: Tumor necrosis factor; FMT: Faecal microbiota transplantation.