This study investigated, for the first time, the relationship between blood microbiota and LF in patients with severe obesity. The main finding of this study was that blood microbiota dysbiosis is associated with LF in two different populations.

There has been intense interest in the development of noninvasive methods as an alternative to biopsy for the identification of advanced fibrosis in patients with NAFLD. These include the NAFLD fibrosis score (NFS), enhanced liver fibrosis (ELF) panel, and transient elastography (TE). 39 The NFS is based on six readily available variables and has a receiver operating characteristic (ROC) score of 0.85 for predicting advanced fibrosis. The ELF panel, which consists in the dosage of plasma levels of three matrix turnover proteins, has a ROC score of 0.90. 29 TE, which measures liver stiffness noninvasively, has been successful in identifying advanced fibrosis in NAFLD. 40 However, it has a high failure rate in individuals with high BMI, and failure of liver stiffness measurement or unreliable results occur in 5%‐15% of patients of the general population, mainly attributed to obesity. The results presented here suggest that blood bacterial DNA measured by qPCR could be used effectively for the diagnosis of the presence or the absence of LF in patients with severe obesity. In the latter situation, patients would avoid unneeded liver biopsy.

PATHOPHYSIOLOGICAL IMPLICATIONS

The blood microbiome derives, at least partially (and probably mostly), from the gut microbiome as a result of several mechanisms of bacterial translocation41-43; however, as we previously published,35 the blood and gut microbiomes differ dramatically from each other, suggesting that the intestinal barrier, immune cells, and liver play a role of filter.43-45

The correlation between the increase in blood bacterial 16S rDNA and the presence of LF is consistent with the large body of evidence showing that, in liver cirrhosis, there is a disruption in the intestinal barrier, the immune defense, and an increase in the translocation of bacterial fragments from the intestine to the liver.9, 10, 15

Importantly, these results suggest that the function of firewall, normally played by the liver to clear commensal bacteria that have penetrated either intestinal or systemic vascular circuits, is impaired in patients with liver disease.45 Despite this difference of bacterial profiles between gut and blood, several bacterial taxa in fecal samples were correlated with other bacterial taxa present in blood, confirming a link between gut dysbiosis and blood dysbiosis.

The current study revealed that in Spanish patients with severe obesity, both a blood and a fecal bacterial profile associated with LF can be detected. Importantly, these profiles cannot be confirmed in the Italian cohort. This observation introduces the possibility of an interaction between bacterial communities and an environmental factor specific to the geographical area on LF in obese patients. In this respect, we observed in the Spanish cohort, but not in the Italian one (at least not at the early stages), that a deficiency in V. paradoxus DNA is associated with LF. V. paradoxus is a Gram‐negative bacterial species belonging to the Proteobacteria phylum.46 Unlike other bacteria, it is able to degrade complex organic compounds, including harmful chemical xenobiotics46 such as trichloroethylene and polychlorinated biphenyls, which contaminate human food and have been reported to induce LF, liver cirrhosis and liver autoimmune diseases.47, 48 An unequal distribution of xenobiotics depending on the geographical region could contribute to explain the discrepancies between Spanish and Italian cohorts in their bacterial correlation with fibrosis.

PICRUSt analysis revealed that, in the Spanish cohort, with the decrease of bacterial diversity, many blood microbiota functions, including xenobiotics biodegradation, are proportionally decreased in patients with fibrosis. On the other hand, the proportion of genes from other pathways, such as nitrogenase of the Nif family, is significantly increased in fibrosis. This observation is interesting given the observed link between LF and the metabolism of nitrogen compounds such as nitric oxide.49, 50

Bacteria are known to be implicated in bile salt metabolism in humans, including the conversion of primary bile acid to secondary bile acid.28

Liver cirrhosis correlates with gut microbiota modification and decrease of serum secondary/primary bile acid ratio.27 Interestingly, we observed a similar correlation with LF (including early stages) in the Spanish cohort, both in terms of taxa modified in gut microbiota (diminution of the Ruminococcaceae and Lachnospiraceae, for instance) and plasma bile acid profile (diminution of the ratio of the secondary/primary bile acid ratio). However, both PICRUSt analysis and manual verification of the metagenomic output (data not shown) showed that blood microbiota of both control and fibrotic patients contain virtually no taxa known to possess bile salt hydrolase (BSH).28 This enzyme, which catalyzes the initial step of bile acid metabolism, is, however, present in many taxa found in gut microbiota.28 The absence of BSH from the blood microbiota underlines the difference between blood and gut microbiota.

We have shown and then confirmed, for the first time, a relationship between LF in patients with severe obesity and blood bacterial burden. These results open two avenues in the field of diagnostics and the understanding of NAFLD‐associated LF. First, blood microbiota analysis provides biomarkers for the early detection of LF in patients with severe obesity. Second, the specific taxonomic profile and bacterial functions found to be associated with LF in Spanish patients, but not in the Italian patients, raises the possibility that bacterial communities present in blood and tissue in interaction with environmental factors play a causal role in LF in patients with severe obesity. Further studies that span all stages of LF are required to assess the predictive value of this candidate biomarker.