The International Scientific Association for Probiotics and Prebiotics defines probiotics as “live microorganisms that, when given in sufficient amounts, have a beneficial effect on the health of the host” ^[1][73]. They protect the intestinal barrier, attenuate pathogen overgrowth, decrease bacterial translocation, reduce serum pro-inflammatory cytokine concentrations while increasing the serum anti-inflammatory cytokine concentrations, and induce host immunomodulation to prevent infection ^[2][3][4][5][74,75,76,77]. In addition, probiotics also act through pharmacokinetics. For example, gut Actinobacterium Eggerthella lenta could affect the pharmacokinetic of digoxin and reduce its toxicity in treating congestive heart failure ^[6][78]. Moreover, the probiotic E. coli strain Nissle 1917 influences the pharmacokinetics of the antiarrhythmic amiodarone and increases drug absorption ^[7][79].

Several probiotics play a role in adult intensive care ^[8][80]. Probiotic therapy significantly reduces the incidence of diarrhea, acquired infections, and VAP in critically ill patients ^[9][10][11][24,26,81]. In sepsis-induced, severe ALI, Akkermansia muciniphila (A. muciniphila) was significantly negatively correlated with TNF-α, IL-1β and IL-6, suggesting that Gut A. muciniphila plays an important role in ALI and that supplementation with A. muciniphila may be a possible therapy for ALI ^[12][82]. In addition, the combination of probiotics Bifidobacterium longum, Lactobacillus bulgaricus and Streptococcus thermophilus was more effective as an adjuvant therapy for severe and critically ill patients with COVID-19, shortening the nucleic acid conversion time, and reducing the inflammatory index such as procalcitonin and C-reactive protein ^[13][83]. Another probiotic, L. reuteri., can reduce lung inflammation and mortality of ARDS ^[14][30]. In uremic dialysis patients, oral administration of Lactobacillus acidophilus led to a decrease in serum dimethylamine, a potential uremic toxin ^[15][84]. The administration of probiotics (e.g., Bifidobacterium bifidum, Bifidobacterium catenulatum, Bifidobacterium longum, Lactobacillus plantarum) can also significantly reduce serum proinflammatory endotoxin, decrease cytokine levels, and improve life quality ^[16][85]. A double-blind clinical study has shown that the probiotics Lactobacillus rhamnosus GG or a combination application of probiotics (including streptococcus thermophiles, lactobacillus acidophilus, lactobacillus delbrueckii ssp. bulgaricus, lactobacillus paracasei, lactobacillus plantarum, Bifidobacterium longum, Bifidobacterium infantis, and Bifidobacterium breve) has beneficial effects on heart failure caused by adverse cardiac remodeling after myocardial infarction (MI). These probiotics play roles through decreasing the levels of intestinal metabolite trimethylamine N-oxide (TMAO) ^[17][18][34,86].

Prebiotics are substrates that can be selectively utilized by host microbes to maintain gut homeostasis and improve health outcomes. Dietary fiber (DF) is a powerful beneficial prebiotic that promotes the production of SCFAs ^[19][87].

Several prospective cohort studies have found that the use of DF in critically ill patients is effective in enhancing intestinal barrier function, reducing the systemic inflammatory response, modulating gut microbiota: increasing the abundance of the SCFAs-producing bacteria, and decreasing the level of potentially pathogenic microbiota ^[20][35]. The use of DF also improves the clinical outcomes, shortening hospital days, and reducing morbidity and mortality in critically ill patients ^[20][35]. Researchers have also found that prebiotics may ameliorate the prognosis of COVID-19 by offering anti-inflammatory nutrition, improving malnutrition, and enhancing immunity through the gut–lung microbial axis ^[21][22][23][36,37,88]. More importantly, DF-fermented SCFAs increase the production of CD103⁺DCs, promote the differentiation of activated CD8⁺T cells to effector cells with a memory phenotype, and improve the outcomes of anti-PD-1 immune checkpoint inhibitor therapy in patients with melanoma ^[24][25][89,90]. Similarly, the SCFAs produced by DF provide energy to the gut microbiota and promote the amino acids to reach the colon for absorption into the body rather than fermenting into uremic solutes ^[26][38].

In addition, flavonoid is an important class of natural products widely found in fruits which belongs to a class of plant secondary metabolites ^[27][91]. Clinical trials have demonstrated that flavonoids can upregulate the abundance of probiotics, such as Bifidobaterium and Lactobacillus, while downregulating the abundance of some pathogenic bacteria such as Staphylococcus aureus and Clostridium histolyticum ^[28][29][92,93]. Moreover, flavonoids can also promote the production of SCFAs ^[30][94].

Synbiotics are mixtures of probiotics and prebiotics that exert beneficial effects on the host in two main ways: enhancing the viability of probiotic microorganisms and providing specific health effects ^[31][32][95,96]. Probiotics stimulated by prebiotics could regulate the metabolic activity of the gut, maintain the intestinal biostructure, and promote the growth and multiplication of probiotics and its resistance to reactive oxygen species and bile salts/acids ^[33][97]. In addition, synbiotic agents modulate the innate and adaptive immune systems to reduce systemic inflammation and promote extraintestinal organ function ^[33][34][71,97]. Synbiotics lead to lower concentrations of adverse metabolites which results in significantly increased SCFAs levels, which may contribute to a positive effect on the host health ^[31][95].

A double-blinded controlled clinical trial demonstrates that a combination of antibiotic intervention led to a significant reduction in pharyngeal aspiration in critically ill patients and an increased level of patient consciousness ^[35][98]. In addition, for hospital-acquired infections in critically ill patients, synbiotics may be a safer and more effective way to reduce endotoxin and inflammatory markers in serum and the complications of sepsis ^[4][11][76,81]. Studies have shown that prophylactic synbiotics (e.g., Bifidobacterium breve strain Yakult combined with Lactobacillus casei strain Shiorta, and galactooligosaccharides) increases the number of probiotics (e.g., Bifidobacterium, Lactobacillus) in fecal bacteria and intestinal SCFA levels, especially acetate. These may modulate the gut microbiota and environment, and have preventive effects on the incidence of enterocolitis and VAP in sepsis patients ^[36][25]. Moreover, synbiotics are used to maintain a stable intestinal microbiota after SIRS and major surgery including high-risk hepatectomy, colorectal resection surgery, Roux-en-Y gastric bypass (RYGB), and sepsis-associated encephalitis (SAE) ^[37][38][40,99]. Synbiotics could also reduce the incidence of diseases such as VAP and healthcare-associated pneumonia, and shorten ICU length of stay ^[39][40][100,101]. In the late stage of intestinal disorders, supplementation with synbiotics can accelerate the recovery of the microbiota, thus preventing the development of sepsis and the onset and progression of critical diseases such as ARDS to some extent ^[41][44]. Synbiotics intake has been demonstrated to reduce plasma levels of uremic toxin and may exert nephroprotective effects ^[15][84].

Based on the above literature, the researchers briefly summarize the different scenarios in which FMT, SDD, probiotics, prebiotics, and synbiotics act directly by regulating the gut microbiota (were summarized in Table 1)2.

123]. Furthermore, butyrate acid strengthens the intestinal biological barrier function. Elevated levels of butyrate acid lowers colonic PH and inhibits the growth and colonization of pathogenic bacteria ^[68][124]. It also prevents bacterial translocation across the intestinal barrier by promoting the antimicrobial activity of intestinal macrophages ^[69][125].

As a histone deacetylase inhibitor, it affects the gene expression profile of intestinal epithelial cells and immune cells through histone deacetylases ^[70][71][72][126,127,128]. It can alter the gene expression profile of immune cells such as Treg cells, intestinal macrophages, and bone marrow-derived macrophages through G protein-coupled receptors (GPCR), affecting their response to microbial stimulation ^[69][73][125,129]. Butyrate acid can relieve inflammation and clinical symptoms in critically ill patients by activating Treg cells ^[74][130]. In addition to directly affecting the function of the intestine itself, butyrate acid can also affect the organs via the gut–lung axis and gut–brain axis. Studies have shown that the GPCR overlap with each other in extraintestinal or intestinal organs ^[75][131]. For example, GPR109A viewed as GPCR common to both lung and intestine can be activated by butyrate ^[48][108]. Butyrate acid affects the intestinal epithelial barrier function and immune function regulation through GPR109A, with similar effects on lung tissue (Table 23). Intraperitoneal injection of sodium butyrate acid can decrease the expression of hypermobility protein 1 (HP-1), pro-inflammatory cytokines such as tumor necrosis factor-α (TNF-α) and interleukin-6 (IL-6), and inhibit the activation of the NF-κB signaling pathway in an ALI/ARDS mouse model ^[76][77][60,132].

Transfer of sterile filtrates from donor feces, rather than fecal microbiota, has been shown to be sufficient to restore normal bowel habits and eliminate symptoms ^[46][106]. Therefore, researchers speculate that gut metabolites may contribute to critical illness and dysbiosis. There is observational data that correlates critical illness, dysbiosis, and altered gut metabolites, including SCFAs, flavonoids, indole derivations, amines, bile acids, etc. ^[47][107]. In this section, several microbiota-derived metabolites and proteins therapies will be summarized.

Typically, undigested DF, as well as proteins and peptides, can be fermented by gut bacteria in the cecum and colon. The main products of these fermentative reactions are SCFAs, consisting of groups of fatty acids with less than six carbons. SCFAs include formic acid (C1), acetic acid (C2), propionic acid (C3), butyric acid (C4), and valeric acid (C5). The major SCFAs in the gut are C2, C3, and C4, accounting for more than 95% of all SCFAs ^[48][108]. SCFAs are key mediators in the regulation of myocardial tissue repair by gut microbiota ^[49][20]. Decreased microbiota abundance has been shown to alter immune cell responses to infectious damage, usually resulting in a pro-inflammatory phenotype ^[50][109], which further aggravates disease progression. Normal levels of SCFAs support the activity of innate lymphocytes, T cells, and B cells in the gut, thereby improving immune tolerance in the gut, strengthening the gut immune barrier, and enhancing its ability to clear pathogens ^[51][52][14,110]. Furthermore, a large proportion of gut-derived SCFAs are transported out of the gut to affect other organs through the gut–lung axis, gut–brain axis, gut–liver axis, gut–kidney axis, gut–bone axis, gut–skin axis, gut–fat axis, gut–heart axis, and so on ^{[53][54][55][56][57][58]}[111,112,113,114,115,116]. Available studies have shown that gut microbiota dysbiosis and a lack of SCAFs are significantly associated with the severity of COVID-19 ^[59][45]. Therefore, maintaining healthy gut microbiota and normal levels of SCFAs contribute to critical illness prevention and prognosis. However, little is known about their therapeutic mechanism in critically ill patients, except for mouse models, so verifying the therapeutic mechanism of SCFAs in clinical trials may be a possible direction in subsequent research. In the following sections, the researchers introduce the beneficial role that major SCFAs play in critical illness.

Previous work on mouse models has shown that butyrate, a specific type of SCFAs readily produced from fiber-rich diets through microbial fermentation, is critical for the maintenance of intestinal homeostasis ^[60][117]. Thus, the researchers speculate that butyrate acid plays a crucial role in critically ill patients. 

In a mouse model of sepsis-associated encephalitis, butyrate is a major metabolite of intestinal microbiota and may have a neuroprotective effect in the process of sepsis through the GPR109A/Nrf2/HO-1 pathway ^[61][118].

Butyrate acid promotes the proliferation and differentiation of intestinal epithelial cells (IECs) and the synthesis of intestinal epithelial tight junction protein, such as the increased expression of Zo-1 and Occludin, reduces cell apoptosis, and inhibits intestinal permeability, resulting in enhanced intestinal mucosa mechanical barrier function ^[62][63][119,120]. In addition, butyrate acid enhances the intestinal mucosal immune barrier. It can maintain immune homeostasis by restoring the IRF3 signaling pathway ^[64][50]. Moreover, butyrate acid promotes the production of anti-microbial peptides (AMPs) ^[65][121]. AMPs are small molecular peptides produced by IECs with broad-spectrum antimicrobial activities, such as butyrate acid which promotes RegIIIγ and β-defensins in a GPR43-dependent manner, which play important roles in limiting bacteria and manipulating species composition ^[66][67][122,

Propionate is mainly produced by Bacteroides spp. and is used as a gluconeogenic substrate in the liver and intestinal to provide energy to the body ^[68][124]. Propionic acid is an anti-inflammatory cytokine, its level in serum may predict the severity and prognosis in critically ill patients and may be a cytokine regulatory marker for critical illnesses such as sepsis ^[89][144].

Acetate is mainly derived from gut microbes and is a metabolite released into the intestinal lumen by anaerobic bacteria from the gut microbiota, which is then absorbed by IEC and distributed to peripheral capillaries ^[90][145]. In recent years, studies in mice have found that acetate inhibits the permeability of the alveolar–capillary barrier, reduces pulmonary edema, inhibits oxidative stress, suppresses inflammatory cell recruitment and inflammatory mediator production, and regulates MAPK pathway activation; thus leading to amelioration of ALI and ARDS ^[91][146]. Elevated levels of acetate may prevent IEC translocation by inhibiting endotoxin and increasing claudin, thereby reducing the incidence of sepsis ^[34][36][92][25,71,147].

Researchers have found that flavonoids ingested with food enter the circulatory system through the intestines to exert their beneficial effects ^[93][148]. Generally, flavonoids are absorbed as metabolites, and gut microbiota participate in this metabolism ^[94][95][149,150]. Flavonoid metabolites can shape the gut microbiota by inhibiting the growth of various pathogens and increasing beneficial genera; these could reduce the endotoxin, maintain gut immune homeostasis, and promote nutrients absorption ^[96][97][151,152]. Flavonoids metabolites have anti-inflammatory effects and play a part in local and systemic immunity. In mouse models, flavonoid metabolites can improve intestinal barrier function by reducing intestinal mucosal inflammation and maintaining the intestinal tight junction barrier and structure  ^{[98][99][100]}[46,153,154]. In addition, flavonoid metabolites regulate inflammatory mediators, such as through inhibiting endothelial activation, NLRP3 inflammasome, toll-like receptors (TLRs), or bromodomain-containing protein 4 (BRD4), as well as activating nuclear factor erythroid-derived 2-related factor 2 (Nrf2), thereby restoring cytokine storm in critical illness including SARS-CoV-2 infection ^[101][47]. DAT, a kind of flavonoid metabolite, has been shown to treat upper respiratory tract infections in mice and protect mice from bacterial endotoxin-induced septic shock ^[102][103][155,156]. Therefore, the researchers suggest that flavonoid metabolites produced by intestinal microbial metabolism play important roles when critically ill patients and hosts are infected with viruses and lethal bacterial infections.

Other than SCFAs and flavonoid metabolites, other microbiota-derived metabolites and proteins also play a critical role in maintaining the balance of intestinal mucosa and contributing to the treatment of critical illness. Indole-3-propionic acid (IPA) could modulate gut microbiota in normal mice, increase the levels of some probiotics (e.g., Akkermansiaceae, Bifidobacteriaceae), strengthen the mucus barrier, and attenuate LPS-induced inflammatory factors in sepsis by increasing mucins and goblet cell secretion products ^[104][157]. Studies on mouse models of sepsis suggest that the anti-inflammatory activity of IPA may associated with the increased abundance of Bifidobacteriaceae and inhibited expansion of Enterobacteriaceae, contributing to an improvement in mortality in sepsis ^[105][158]. Aromatic microbial metabolites (AMMs), such as phenyl lactic and 4-hydroxyphenyllactic acids, have been observed at a much higher level than normal in the serum of septic patients ^[106][159]. A prospective observational pilot study found that high level of AMMs were associated with severity and mortality in critically ill patients, and may become a possible direction to improve the prognosis of critical illness ^[107][160]. TMAO has been found to be higher in individuals with heart failure than in controls, suggesting that TMAO is a new and novel risk factor in heart failure development and can lead to cardiac hypertrophy and cardiac fibrosis ^[108][161]. Beta-lactamase, produced primarily by extended-spectrum β-lactam Enterobacteriaceae, has been shown to reduce the jejunal concentration of antibiotics and to prevent antibiotics from reaching the colon, thus alleviating the effect of antibiotics on gut microbiota disturbance ^[109][110][162,163].