The human gut microbiota consists of bacteria, eukaryotes, archaea, and viruses, which live symbiotically in the gastrointestinal tract [1
]. The gut microbiota plays a key role in human health and disease. Alcohol-related liver disease is one of the most prevalent liver diseases worldwide. Alcohol-related liver disease progresses from simple steatosis to steatohepatitis, fibrosis, and cirrhosis. Alcoholic hepatitis is a severe form of alcohol-related liver disease and most commonly arises acutely on the background of cirrhosis [2
]. Treatment strategies are limited for patients with alcoholic hepatitis. Pronounced overgrowth of bacteria in the small intestine was found in patients with alcoholic hepatitis [3
]. Changes in the fecal bacteria were associated with disease severity in patients with alcoholic hepatitis [4
In addition to bacteria, intestinal mycobiota was also dysregulated in patients with alcoholic hepatitis as revealed by fungal-specific internal transcribed spacer (ITS) amplicon sequencing [5
]. Detailed taxonomic information could be revealed by this approach, however, the functional analysis of mycobiota is lacking. Shotgun metagenomics could provide information on the functional capacity of mycobiota. Although both bacteria and fungi are dysregulated in alcoholic hepatitis, previous efforts have been focused on either gut bacteria or fungi. The relationship between fungi and bacteria living in the same gastrointestinal tract is not well studied, although fungi and bacteria not only evolve together but also compete for resources.
Antibiotics are commonly used in alcoholic hepatitis patients for the treatment of infections. However, it is not clear whether fungi would overgrow when the gut bacteria are impacted by the antibiotic treatment. Although such studies in humans are lacking, commensal bacteria were reported to inhibit the colonization of Candida albicans
through the activation of hypoxia-inducible factor 1 α and antimicrobial peptide LL-37 in mice [6
]. This raises the question whether changes of bacteria induced by antibiotic treatment in alcoholic hepatitis patients would further influence the growth of fungi, which is important in clinical practice. In addition, human gut Bacteroidetes can utilize yeast mannan via the expression of a specific yeast α-mannan degrading system [7
]. Administration of Saccharomyces boulardii
altered the gut microbial composition [8
]. These findings provide evidence for bacteria–fungi interactions, which requires attention when developing gut microbiota-based therapeutic strategies, as targeting bacteria only or fungi only might also influence other parts of the intestinal microbiota in patients with alcoholic hepatitis.
Here, we sought to reveal the correlation between intestinal fungi and bacteria in patients with alcoholic hepatitis. Better understanding of the relationship between intestinal fungi and bacteria is helpful for the development of gut microbiota-based therapeutic strategies for alcoholic hepatitis patients, which not only take bacteria into consideration during the treatment but also fungi.
In the present study, we found an altered fungal pathway in patients with alcoholic hepatitis, predicted the diagnosis of alcoholic hepatitis using random forest model, and identified the association between fungi and bacteria in patients with alcoholic hepatitis. Superpathway of allantoin degradation in yeast was enriched in alcoholic hepatitis (Figure 2
c). Allantoin is a product of guanine and adenine catabolism, which is a product of uric acid oxidation and can serve as a sole nitrogen source and be degraded to ammonia [9
]. Allantoin can be used by both fungi and bacteria as carbon and nitrogen sources. However, the pathway for conversion of uric acid to allantoin has been lost in humans. Elevated uric acid has been reported to contribute to experimental and human alcohol-related liver disease [10
]. The elevation of uric acid might lead to the enrichment of allantoin degradation in the gut microbiota. However, the potential contribution of allantoin degradation to the pathogenesis of alcohol-related liver disease warrants further analysis.
Fungi and bacteria share micro-habitats and engage in complex communications within the microbial community. Such relationship cannot be easily predicted based on the knowledge of cultured microorganisms, especially in the disease condition of alcoholic hepatitis. The metabolic crosstalk develops interdependently, which leads to co-occurrence patterns of fungi and bacteria. The correlation network of fungi and bacteria in our study revealed these co-occurrence patterns in patients with alcoholic hepatitis. Especially, a negative association was found between fungus Cryptococcus
and bacterium Pseudomonas.
Species in Pseudomonas
such as Pseudomonas aeruginosa
are also responsible for life-threatening infections in immunocompromised persons [11
]. Since negative correlation exists between fungus Cryptococcus
and bacterium Pseudomonas,
treatment which targets the growth of Pseudomonas
might increase the growth opportunity for Cryptococcus.
Species within genus Cryptococcus
are responsible for invasive fungal infection, which cause Cryptococcosis and present substantial therapeutic challenges [12
]. Consistent with our study, Pseudomonas aeruginosa
has been reported to inhibit the growth of Cryptococcus
species by producing antifungal molecules pyocyanin and an extracellular quorum-sensing signal 2-heptyl-3,4-dihydroxyquinoline [13
]. Thus, such correlations need to be taken into consideration in clinical practice during the treatment for bacterial or fungal infections.
Age and BMI have influence on the gut microbiome. In present study, there were three bacteria genera (Pseudoflavonifractor, Oribacterium,
) positively correlated with age and one negatively correlated with age (Mitsuokella
) (Supplementary Table S1
). Two bacteria genera were positively correlated with BMI, Xanthomonas
, meanwhile four were negatively correlated with BMI, including Sphingomonas, Phascolarctobacterium, Holdemania,
(Supplementary Table S1
). The correlation between age, BMI, and fungal genera was reported in our previous study, where Altemaria
was found to be negatively correlated with age but no significant correlation was found between BMI and fungal genera [5
There are several limitations of this study. First, due to the limit of sample availability, ITS and 16S rRNA sequencing were not performed in all enrolled patients with alcoholic hepatitis. Second, patients with chronic alcoholic hepatitis were lacking in our patient cohort. Third, the sample size in our study is small and correlations need to be validated in a larger and independent patient cohort. Finally, it is noteworthy that this correlation approach may not be representative of causal relationship in microbial ecology.
In conclusion, this study highlights the correlation between the gut bacteria and fungi, which needs to be taken into consideration in clinical practice. Further mechanistic studies of fungal and bacterial ecological interactions in alcohol-related liver disease are required.