Antibiotics in Chronic Liver Disease and Their Effects on Gut Microbiota

Impairments in liver function lead to different complications. As chronic liver disease progresses (CLD), hypoalbuminemia and alterations in bile acid compositions lead to changes in gut microbiota and, therefore, in the host–microbiome interaction, leading to a proinflammatory state. Alterations in gut microbiota composition and permeability, known as gut dysbiosis, have important implications in CLD; alterations in the gut–liver axis are a consequence of liver disease, but also a cause of CLD. Furthermore, gut dysbiosis plays an important role in the progression of liver cirrhosis and decompensation, particularly with complications such as hepatic encephalopathy and spontaneous bacterial peritonitis. In relation to this, antibiotics play an important role in treating CLD. While certain antibiotics have specific indications, others have been subjected to continued study to determine whether or not they have a modulatory effect on gut microbiota. In contrast, the rational use of antibiotics is important, not only because of their disrupting effects on gut microbiota, but also in the context of multidrug-resistant organisms. The aim of this review is to illustrate the role of gut microbiota alterations in CLD, the use and impact of antibiotics in liver cirrhosis, and their harmful and beneficial effects.


Introduction
The human microbiota refers to the living microorganisms that colonize our body.However, the microbiome refers not only to this collection of living microorganisms, but also to their genomes and products, such as structural elements and metabolites, and even environmental conditions.The microbiome colonizes our body from birth, and it undergoes a dynamic process of shaping and multiplication, with modifications in composition depending on genetic, nutritional, and environmental factors [1,2].The composition of the human microbiome varies from site to site; it is highly diverse and comprises trillions of microorganisms.The gut microbiome has the highest number of microorganisms and has been extensively studied because of its impacts on health and disease [3].Moreover, the interactions of the gut microbiome (GM) with different organs and systems each have a unique profile.Modifications in GM composition and function induce intestinal permeability, changes in digestion and metabolism, and immune responses.Misbalances in GM can lead to the onset and worsening of many diseases-not only gastrointestinal, but also metabolic, immunological, and neuropsychiatric [4][5][6].
In liver disease, interactions within the gut-liver axis are especially important, not only in relation to the decompensation and progression of liver cirrhosis, but, as stated before, in even in the absence of a previous condition when significant alterations in GM composition are caused by several factors.In fact, significant and constant changes in the diet or in the substances that are released in the gut will eventually alter GM composition and the way immune cells and microorganisms interact.For example, trimethylamine N-oxide (TMAO) is derived from the conversion of choline, mainly by Desulfovibrio desulfuricans and Escherichia coli.The increased synthesis of TMAO due to the overgrowth of these bacteria results in a lack of choline in the body, which, in turn, enhances oxidative stress in hepatocytes and increases liver inflammation and fibrosis [19,20].
In the context of chronic liver disease, patients have lower levels of Bacteroidetes and higher levels of Proteobacteria, Enterococcus, Veillonella, Megasphaera, Burkholderia, Prevotella and Fusobacteria.These changes in gut microbiota composition are, among other things, mainly related to alterations in BAs, lipid metabolism and the activation of inflammatory pathways.In fact, patients with liver fibrosis exhibit an altered BA profile that can change the composition of the gut microbiota and exacerbate fibrosis [21,22].
BAs have direct cytotoxicity and antibacterial activity, but also indirectly mediate the inhibition of microbial growth by regulating the expression of nitric oxide synthase and antimicrobial peptide genes.When BAs bind to FXR, antimicrobial peptides, such as angiogenin 1, are produced.These peptides can inhibit gut microbiota growth by increasing the intestinal epithelial cell potential to prevent bacterial uptake, improving gut barrier function.An increase in the harmful bacterial release of PAMPs into enterohepatic circulation in the context of a disrupted intestinal barrier induces the activation of immune cells in the liver.Chemokines, such as CC-chemokine ligand 2 (CCL2) and IL8, recruit immune cells, such as macrophages and neutrophils, to the liver.Another important cytokine is IL1β, which is induced by NF-kB following the activation of TLR4 by PAMPs such as LPS [23][24][25].
Furthermore, in patients with cirrhosis, this same mechanism can increase decompensation episodes, such as spontaneous SBP, and the risk of acute-on-chronic liver failure (ACLF) [25,26].
On the other hand, an altered, but not necessarily reversed, GM can mediate liver diseases.As stated before, the GM can influence the size and composition of the BA pool through the conversion of primary to secondary BAs, which act as signaling molecules affecting lipid and glucose metabolism, and predisposing individuals to metabolic diseases.In patients with MASLD, studies found that high levels of serum GCA and stool DCA are related to severe fibrosis and are positively correlated with Lachnospiraceae and negatively correlated with Bacteroidaceae levels.Another example of the impact of lipid metabolism is the reduction in SCFA-producing microbiota such as Bacteroidaceae, since SCFAs protect the intestinal barrier and prevent the development of MASLD by their effects on free fatty acid metabolism and visceral adipose tissues levels, reducing TNF expression and the activation of the NF-κB pathway [27,28].
Finally, patients with liver cirrhosis require different therapeutic options due to episodes of hepatic decompensation, which are particularly related to dysbiosis.This is the case of HE, wherein Rifaximin is the cornerstone treatment not only in acute events but also in preventing future episodes, since the overgrowth of ammonia-producing bacteria occurs in the gut.While this antibiotic seems to be more beneficial than harmful, there is no doubt that it has important effects on the GM [28,29].

Reevaluating the Therapeutic Use of Antibiotics in Liver Cirrhosis
Patients with cirrhosis are predisposed to bacterial infections; an example is the high prevalence of Spontaneous Bacterial Peritonitis (SBP).In addition, these patients exhibit small intestinal bacterial overgrowth, increased intestinal permeability and reduced intestinal motility that may be related to the severity and progression of liver disease.Moreover, Prado et al. [29] conducted a study aimed at determining whether rectal colonization by resistant bacteria increased the likelihood of subsequent infection by the same strain in critically ill patients with cirrhosis.The presence of resistant bacteria in the rectal flora was assessed through rectal swab samples.The findings suggest that rectal colonization serves as a reservoir for potential infections, particularly when the colonizing bacteria are resistant to antibiotics.This phenomenon is of particular concern in critically ill patients with cirrhosis, as their compromised immune system and impaired liver function contribute to increased vulnerability to infections [29,30] (Figure 1).by resistant bacteria increased the likelihood of subsequent infection by the same strain in critically ill patients with cirrhosis.The presence of resistant bacteria in the rectal flora was assessed through rectal swab samples.The findings suggest that rectal colonization serves as a reservoir for potential infections, particularly when the colonizing bacteria are resistant to antibiotics.This phenomenon is of particular concern in critically ill patients with cirrhosis, as their compromised immune system and impaired liver function contribute to increased vulnerability to infections [29,30] (Figure 1).In chronic liver diseases, dysbiosis and increased bacterial overgrowth provoke altered bile acid composition associated with reduced intestinal motility and the expression of tight junction proteins produces leaky gut.This leads to the increased passage of PAMPs, LPS, and bacterial products to the liver through the portal vein, with the consequent activation of inflammatory pathways.All of these alterations lead to increased ammonia production with consequent hepatic encephalopathy; the translocation of bacteria into the peritoneal fluid leading to spontaneous bacterial peritonitis; and the activation of proinflammatory and fibrotic pathways with increased hepatic vascular tone leading to portal hypertension.Furthermore, SBP, urinary tract infections, and pneumonia are the most common infections in patients with liver cirrhosis.Current evidence implies that about 48% of infections in liver cirrhosis are community-acquired, while 52% are related to nosocomial factors, and are healthcare-related.In this ma er, there is great concern regarding multidrugresistant organisms (MDROs) [31][32][33].Since antibiotics are frequently prescribed, a group of researchers conducted the ATTIRE trial to assess the impact of antibiotic therapy in patients with decompensated cirrhosis.Through a randomized controlled trial design, they compared the outcomes of patients who received early antibiotic treatment upon hospital admission to those who received antibiotics only if an infection was clinically In chronic liver diseases, dysbiosis and increased bacterial overgrowth provoke altered bile acid composition associated with reduced intestinal motility and the expression of tight junction proteins produces leaky gut.This leads to the increased passage of PAMPs, LPS, and bacterial products to the liver through the portal vein, with the consequent activation of inflammatory pathways.All of these alterations lead to increased ammonia production with consequent hepatic encephalopathy; the translocation of bacteria into the peritoneal fluid leading to spontaneous bacterial peritonitis; and the activation of proinflammatory and fibrotic pathways with increased hepatic vascular tone leading to portal hypertension.Furthermore, SBP, urinary tract infections, and pneumonia are the most common infections in patients with liver cirrhosis.Current evidence implies that about 48% of infections in liver cirrhosis are community-acquired, while 52% are related to nosocomial factors, and are healthcare-related.In this matter, there is great concern regarding multidrugresistant organisms (MDROs) [31][32][33].Since antibiotics are frequently prescribed, a group of researchers conducted the ATTIRE trial to assess the impact of antibiotic therapy in patients with decompensated cirrhosis.Through a randomized controlled trial design, they compared the outcomes of patients who received early antibiotic treatment upon hospital admission to those who received antibiotics only if an infection was clinically suspected.The results of the study revealed no significant difference in overall survival between the two groups, challenging the routine use of early antibiotic therapy in this patient population [33,34].Furthermore, Bajaj and colleagues found that prophylactic antibiotics may disrupt the natural phage-bacterial balance, leading to shifts in phage populations and potentially affecting microbial diversity [34].
Another common scenario of the interplay between dysbiosis and chronic use of antibiotics in CLD is HE.Historically, metronidazole, neomycin, and vancomycin have been used to treat HE, but these are currently no longer used due to their side effects and the growing prevalence of MDROs [35].In contrast, rifaximin is the preferred option for HE due to its proven safety and efficacy.Nevertheless, this drug is not exempt from the MDRO issue, exemplified by E. coli-resistant strains [36,37].Furthermore, while rifaximin is a classical positive modulator of GM, acting by maintaining gut microbiota diversity and composition and not changing the overall resistome, this continues to be questioned [38,39].
While antibiotic prophylaxis has proven beneficial, the careful consideration of individual patient characteristics is essential.Factors such as antibiotic resistance patterns, renal function, and the presence of comorbidities should be evaluated when selecting the appropriate prophylactic regimen.To further address the issue of the importance of antibiotics in the context of liver disease in contrast to their harmful effects, we summarize their clinical use below.

Antibiotic Effects on Portal Hypertension
The portal vein serves as a major conduit for nutrients, toxins, and microbial products from the gut to the liver.Disruption of the gut-liver axis can lead to dysbiosis, which has been implicated in the pathogenesis of CLD.Studies have shown that alterations in gut microbiota composition and function contribute to liver inflammation, fibrosis, and portal hypertension.The dysbiosis-induced increased intestinal permeability to gut microbial metabolites, such as LPS, secondary BAs, and TMAO, has been shown to influence hepatic vascular tone and contribute to portal hypertension [40].Moreover, evidence suggests that when those metabolites escape to the systemic circulation, they may induce systemic hypertension [41,42].
Recent research on factors influencing GM with regard to portal hypertension has opened new avenues for therapeutic interventions.Modulating the gut microbiota through strategies such as probiotics, prebiotics, antibiotics, and fecal microbiota transplantation might represent promising therapies to improve liver-related complications and reduce portal hypertension.Additionally, targeting gut microbial metabolites and their receptors may offer novel therapeutic options for the management of portal hypertension [40,43].In fact, bacterial-derived products may increase hyperdynamic circulation and intrahepatic vascular resistance, promoting a further increase in portal pressure and the risk of bleeding [44][45][46].
Regarding infections, when compared with controls, patients with liver cirrhosis and increased populations of Bacteroides, Escherichia, Shigella, and Prevotella have severe portal hypertension and high levels of IL-8 in their hepatic veins [47].Furthermore, it seems that patients with variceal bleeding have a higher rate of bacterial infections, and the administration of intravenous antibiotics, such as norfloxacin or ampicillin/sulbactam, may improve complications [45].
A recent study published by Mendoza et al [48].showed that the use of rifaximin or norfloxacin did not cause a significant reduction in hepatic venous pressure gradient (HPVG) in patients with cirrhosis, but the use of antibiotics for longer periods in association with non-selective beta blockers (NSBB) did decrease HPVG significantly [48].The use of rifaximin has been shown to reduce portal hypertension when associated with NSBB, compared to the use of propranolol alone [49] (Figure 2).However, norfloxacin did not perform better than the placebo in reducing HVPG [50].Moreover, the use of probiotic VSL#3 has been shown to improve the effect of propranolol in reducing HPVG [51].

Prophylactic Antibiotic Use for Cirrhosis
Current guidelines recommend antibiotic prophylaxis in specific situations.For patients with a history of SBP, long-term prophylaxis with oral norfloxacin or trimethoprimsulfamethoxazole is recommended to prevent recurrence.Additionally, short-term prophylaxis with intravenous antibiotics is advised for cirrhotic patients with gastrointestinal bleeding, as it reduces the risk of infections and improves survival rates [52,53].Regarding the la er, consensus guidelines recommend the prophylactic use of oral or intravenous antibiotics in this population.Furthermore, quinolones and beta-lactams, either alone or in combination, were effective in reducing rebleeding rates and hospital stay length in cirrhosis patients with gastrointestinal bleeding, according to a metanalysis.On the other hand, MDRO bacterial infections have reduced the efficacy of commonly used antibiotics, necessitating combined antibiotic therapy.Combination therapy with quinolones and beta-lactams has been associated with reduced mortality, rebleeding, and hospitalization lengths [53].
Patients with liver cirrhosis experience about 36% spontaneous infections, such as with SBP [54].When SBP is suspected, empiric antibiotics are used, with third-generation cephalosporins used commonly, except in the context of MDRO risk factors, where the first option is piperacillin/tazobactam.In the case of prophylaxis, norfloxacin and ciprofloxacin are the first options for both primary and secondary prevention, followed by

Prophylactic Antibiotic Use for Cirrhosis
Current guidelines recommend antibiotic prophylaxis in specific situations.For patients with a history of SBP, long-term prophylaxis with oral norfloxacin or trimethoprimsulfamethoxazole is recommended to prevent recurrence.Additionally, short-term prophylaxis with intravenous antibiotics is advised for cirrhotic patients with gastrointestinal bleeding, as it reduces the risk of infections and improves survival rates [52,53].Regarding the latter, consensus guidelines recommend the prophylactic use of oral or intravenous antibiotics in this population.Furthermore, quinolones and beta-lactams, either alone or in combination, were effective in reducing rebleeding rates and hospital stay length in cirrhosis patients with gastrointestinal bleeding, according to a metanalysis.On the other hand, MDRO bacterial infections have reduced the efficacy of commonly used antibiotics, necessitating combined antibiotic therapy.Combination therapy with quinolones and beta-lactams has been associated with reduced mortality, rebleeding, and hospitalization lengths [53].
Patients with liver cirrhosis experience about 36% spontaneous infections, such as with SBP [54].When SBP is suspected, empiric antibiotics are used, with third-generation cephalosporins used commonly, except in the context of MDRO risk factors, where the first option is piperacillin/tazobactam.In the case of prophylaxis, norfloxacin and ciprofloxacin are the first options for both primary and secondary prevention, followed by trimethoprimsulfamethoxazole [55][56][57][58].The empirical antibiotics discussed above seem to exert similar effects against SBP, but response-guided therapy, by performing a second paracentesis at 48 h to assess antibiotic response, should be considered [56].The use of prophylactic norfloxacin might increase the risk of MDR bacterial infections, and practitioners should be aware of this after the first month of liver transplantation [59].Hence, MDR bacterial infection remains controversial, so norfloxacin prophylaxis should be indicated in carefully selected patients [60].
Another novel strategy is selective digestive decontamination (SDD), which consists of the combination of topical nonabsorbable antibiotics or antifungal agents applied to the upper gastrointestinal tract with a short course of intravenous antibiotics.Its use began in patients with neutropenia, and it is a topic of interest in critically ill patients despite controversial evidence [61,62].In cirrhosis, SDD was used to treat both gastrointestinal bleeding and SBP, at first with oral nonabsorbable antibiotics such as polymyxin, neomycin, gentamycin and colistin, and then with trimethoprim-sulfamethoxazole and fluroquinolones.Still, the disrupting effects of antibiotics in GM may be linked to the asymptomatic colonization of the gut by MDROs.This colonization not only represents a potential source of infection for the affected patient, but also contributes to the transmission of MDRO infections within healthcare settings.Consequently, until comprehensive studies have been conducted across multiple centers, investigating the impact of SDD on rates of multidrug resistance at both the individual and population levels, the use of SDD should be restricted to cirrhosis patients who face the highest risk of developing an infection [63].To address this issue, the use of rifaximin is proposed; this non-absorbable antibiotic possesses distinctive effects on the gut microbiota [58].However, the results of a recent study found that, overall, systemic antibiotic prophylaxis is more effective than rifaximin in SBP prevention and should be the standard of care for patients with advanced cirrhosis and a high risk of SBP [64].
Finally, rifaximin, in combination with lactulose or L-ornithine L-aspartate, is employed for the purpose of preventing the recurrence of HE [65,66].According to research findings, it seems that rifaximin enhances the population of beneficial intestinal bacteria, such as Bifidobacterium, Atopobium, and Faecalibacterium prausnitzii.Meanwhile, it does not significantly alter the overall composition of the gut microbiota, including the lactobacilli.Additionally, rifaximin contributes to the restoration of the intestinal barrier, potentially mitigating bacterial translocation and systemic endotoxemia in individuals with cirrhosis.This effect may be attributed to the inhibition of NF-kB activation via the pregnane X receptor (PXR) and a reduction in interleukins and TNFα expression [39,59,67] (Table 1).Bacterial infections represent one of the leading causes of hospitalization, morbidity, and mortality in cirrhotic patients.The most frequent infections are urinary infections, pneumonia, and spontaneous bacterial peritonitis, with an increasing incidence of MDROs [31].
Owing to the increasing use of broad antibiotics in cirrhotic patients, multidrugresistant bacterial infections have been rising; in particular, patients who received prophylactic norfloxacin for SBP experience higher risks of MDRO infection [68].Hence, this assertion remains controversial; in a study performed by Marciano et al., they found that norfloxacin exerts a beneficial effect on SBP prophylaxis, with no increased incidence of MDRO infections [60].To address the uncertainty as to whether antibiotic prophylaxis is beneficial or not, more clinical trials should be performed to test long-term antibiotics [69].Furthermore, in a multicenter study in Europe, it was found that about 30% of positive cultures from infections in patients with liver cirrhosis were caused by MDROs.The most frequently isolated MDROs in this series were extended-spectrum beta-lactamaseproducing Enterobacteriaceae.In that same study, in a second series of patients it was revealed that the prevalence of MDROs was 23% (392 infections out of 2587 patients), and among culture-positive infections, it was 38%.A slight increase in the rate of carbapenemresistant Enterobacteriaceae was observed in this series [77].In general, a global prevalence of 34% MDR bacterial infection is estimated in liver cirrhosis [32].Antibiotic resistance is as-sociated with poor prognosis and the failure of antibiotic strategies, particularly those based on third-generation cephalosporins or quinolones [78].Furthermore, the main risk factors for MDRO infections in patients with cirrhosis are long-term norfloxacin prophylaxis, recent infection by multi-resistant bacteria, and the recent use of β-lactams [79].
It is important to consider the spectrum of infectious pathogens from Gram-negative bacteria in community-acquired infections compared with Gram-positive bacteria in hospital-acquired infections [80].
Antibiotics may also predispose individuals to other infections, such as invasive fungal infections.Fungal infections are much less frequent; they are usually nosocomial and associated with extremely high short-term mortality.In patients with cirrhosis, invasive fungal infections occur in approximately 3-7% of culture-positive infected individuals, and they are more commonly observed as secondary or nosocomial infections during the course of acute-on-chronic liver failure (ACLF).Among them, invasive candidiasis, or candidemia, is the most frequent, accounting for 70-90% of cases, followed by invasive aspergillosis.
Invasive fungal infections in patients with decompensated cirrhosis are generally associated with an extremely poor prognosis.Candidemia and other invasive candidiasis infections are accompanied by 28-day mortality rates ranging from 45% to 60%.ACLF complicated by IA has an even worse prognosis, with only rare cases of survival despite receiving appropriate antifungal treatment [81].
Using a targeted metagenomics approach, Delavy et al. [82] observed a high degree of interindividual diversity in healthy gut microbiota.They found that the prevalence of C. albicans was much higher than previously reported, with all subjects except one carrying C. albicans, albeit at varying levels.The administration of third-generation cephalosporins significantly altered the composition of the microbiota, and the fungal load was increased both in the short and the long term.The variations in C. albicans levels in response to third-generation cephalosporin treatment could be partially explained by changes in the levels of endogenous fecal β-lactamase activity.Subjects with higher β-lactamase activity showed lower C. albicans levels [82].This suggests that the use of a particular antibiotic treatment may change the specific types of microorganisms, either fungal or bacterial, in the GM [83].

Conclusions
The use of antibiotics, mainly rifaximin, can be beneficial in reducing inflammation and liver fibrosis, thus modifying the gut microbiota, and could exert a reducing effect on portal hypertension when associated with NSBB.The use of norfloxacin for the primary or secondary prophylaxis of SBP is controversial and should be enforced on a case-by-case basis, but it could have favorable effects on survival and SBP incidence and recurrence rates.Nonetheless, the excessive growth of MDROs should be considered by physicians to inform rational use.Rifaximin has shown several beneficial effects, including reducing HPVG when associated with NSBB, reducing ammonia-producing bacteria (thus improving hepatic encephalopathy), and reducing intestinal permeability and dysbiosis; therefore, reducing the passage of PAMPs decreases liver inflammation and probably liver fibrosis, in turn reducing SBP incidence.For these reasons, the use of antibiotics in patients with cirrhosis should aim to reduce the incidence of MDROs.

Figure 1 .
Figure1.In chronic liver diseases, dysbiosis and increased bacterial overgrowth provoke altered bile acid composition associated with reduced intestinal motility and the expression of tight junction proteins produces leaky gut.This leads to the increased passage of PAMPs, LPS, and bacterial products to the liver through the portal vein, with the consequent activation of inflammatory pathways.All of these alterations lead to increased ammonia production with consequent hepatic encephalopathy; the translocation of bacteria into the peritoneal fluid leading to spontaneous bacterial peritonitis; and the activation of proinflammatory and fibrotic pathways with increased hepatic vascular tone leading to portal hypertension.

Figure 1 .
Figure 1.In chronic liver diseases, dysbiosis and increased bacterial overgrowth provoke altered bile acid composition associated with reduced intestinal motility and the expression of tight junction proteins produces leaky gut.This leads to the increased passage of PAMPs, LPS, and bacterial products to the liver through the portal vein, with the consequent activation of inflammatory pathways.All of these alterations lead to increased ammonia production with consequent hepatic encephalopathy; the translocation of bacteria into the peritoneal fluid leading to spontaneous bacterial peritonitis; and the activation of proinflammatory and fibrotic pathways with increased hepatic vascular tone leading to portal hypertension.

Figure 2 .
Figure 2. Dysbiosis enhances the secretion of PAMPs, secondary bile acids, TMAO, and ammonia, and activates TLR4 and NFkB pathways.This results in proinflammatory cytokine and chemokyne secretion, with increases in TNFα and IL8 that lead to portal hypertension.Antibiotics such as rifaximin seem to exert beneficial effects on multiple dysbiosis-reducing targets, IL8-producing bacteria, and the passage of bacterial products, with consequent proinflammatory pathway activation.

Figure 2 .
Figure 2. Dysbiosis enhances the secretion of PAMPs, secondary bile acids, TMAO, and ammonia, and activates TLR4 and NFkB pathways.This results in proinflammatory cytokine and chemokyne secretion, with increases in TNFα and IL8 that lead to portal hypertension.Antibiotics such as rifaximin seem to exert beneficial effects on multiple dysbiosis-reducing targets, IL8-producing bacteria, and the passage of bacterial products, with consequent proinflammatory pathway activation.

Table 1 .
Antibiotics used in chronic liver disease clinical trials and meta-analysis results.
3.3.Multidrug-Resistant Bacterial Infections in Patients with Cirrhosis and the Role of Gut Microbiota