Reply.

We would like to thank Zhang et al for their interest in our article and for highlighting the importance of mechanistic studies to shed light on functional properties of native microbiota and its molecules mediating virus colonization.

We found that impaired short-chain fatty acids (SCFAs) and l-isoleucine biosynthesis in the gut microbiome correlated with coronavirus disease 2019 (COVID-19) severity as well as increased plasma concentrations of C-X-C motif chemokine ligand 10 (CXCL-10), N-terminal prohormone of brain natriuretic peptide (NT-proBNP), and C-reactive protein (CRP). Among other cytokines assessed, including interleukin (IL) 10, IL12, IL1b, IL6, and tumor necrosis factor (TNF)-α and chemokines such as CXCL-8 and C-C motif chemokine ligand 2 (CCL-2), we found that only increased plasma levels of IL10 significantly associated with more severe symptoms in patients with COVID-19.

It is likely that elevated endogenous systemic IL10 stimulates inflammatory cytokine production and directly activates and promotes effector cluster of differentiation 8-positive T-cell proliferation, which may play a pathologic role in COVID-19 severity. Intriguingly, fecal butyrate level in patients with COVID-19 showed a significantly negative correlation with plasma IL10, suggesting that microbiota-derived butyrate may be involved in preventing over-expression of IL10 in COVID-19.

To this end, emerging studies have provided new insights into the relationship between the gut microbiome, host immunity, and disease severity in COVID-19. In a separate cohort of 100 hospitalized patient with COVID-19, we found that disrupted gut microbiota were associated with higher levels of TNF-α, IL10, and CXCL10. Others have reported that Enterococcus faecalis was negatively correlated with cluster of differentiation 8-positive T cells and IL4, and Eubacterium ramulus was negatively correlated with IL6 in patients with COVID-19. These cytokines and chemokines are involved in interferon-driven T helper type 1 (Th1) response, implying that the gut microbiota may regulate Th1 response in severe acute respiratory syndrome coronavirus 2 (SARS-COV-2) infection, while more cytokines and proteins associated with immune cell activation should be determined to support this notion.

We agree with the authors that relationships between plasma markers, clinical measures, and disease severity deserve in-depth exploration. In our study, we detected clinical measurements, including blood counts (platelet count, white cell count, neutrophil count) and the plasma concentrations of lactate dehydrogenase (LDH), CRP, albumin, hemoglobin, alkaline phosphatase, and aspartate aminotransferase, alanine aminotransferase (ALT), total bilirubin, and creatinine, and elucidate their relationship with disease severity and microbial functions. We found increased levels of LDH, CRP, and ALT and decreased levels of platelet count, albumin, and hemoglobin (MaAsLin2 [R Foundation for Statistical Computing, Vienna, Austria] false discovery rate corrected P < .05) significantly associated with more severe symptoms in patients with COVID-19.

LDH, ALT, and albumin are well-known markers of liver or kidney dysfunction, highlighting host tissue damage in patients with severe COVID-19. Importantly, the fecal level of butyric acid positively correlated with the plasma level of albumin, indicating microbiota-derived butyrate may have the potential to prevent tissue damage caused by SARS-COVID-2 infection. We also evaluated blood urea nitrogen level mentioned by Zhang et al in their letter in patients with COVID-19 and found patients with severe symptoms showed a significantly higher blood urea nitrogen level than those patients with mild symptoms. This may be associated with higher serum concentrations of urea and disruption of urea cycle functions during COVID-19 infection, , highlighting kidney dysfunction and abnormal amino acid catabolism in patients infected with SARS-COVID-2.

In summary, current evidence supports the notion that the gut microbiota may contribute to disease severity in COVID-19 via regulation of Th1 response, and proof-of-concept studies dissecting how microbiota-derived molecules mediate host immune response in patients with COVID-19 and disease severity are desperately needed to provide more mechanistic insights, and this will be of benefit to exploiting microbial-based therapy.