Reply to Radermacher et al. on “Serum Hydrogen Sulfide and Outcome Association in Pneumonia by the SARS-CoV-2 Coronavirus”

To the Editor:

We read with great interest the comments addressed by Radermacher et al. (1) on our publication regarding the importance of hydrogen sulfide (H₂S) for the prognosis and outcome of severe infection caused by the novel SARS-CoV-2 (also known as Covid-19) (2). Although serum H₂S levels as high as 249 μM and 580 μM have been demonstrated in patients with septic shock (3) and severe asthma (4), we agree that the elevated serum H₂S is an intriguing finding. We tried to deliver some answers that are based on: the performance of the used assay in healthy volunteers and in patients with other types of severe lung infection; and the reproducibility of the data by using another assay.

We measured levels of H₂S in 17 healthy volunteers and in 60 patients with ventilator-associated pneumonia (VAP). VAP was diagnosed according to standard definitions (5) and all patients had microbiological confirmation with one Gram-negative pathogen isolated in counts greater than 10⁵ colony-forming units/mL from the bronchoalveolar lavage by the culture technique already described (6). Isolated pathogens were Acinetobacter baumannii (n = 23), Pseudomonas aeruginosa (n = 19), and Klebsiella pneumoniae (n = 18). Blood samples were collected within the first 24 h from diagnosis of VAP and H₂S was measured by the monobromobimane derivatization assay followed by reverse phase HPLC separation (2). Results clearly showed that survivors from Covid-19 had H₂S levels significantly greater than healthy population and patients with VAP (Fig. 1). This elaborates the hypothesis that it is not the assay that leads to false-positive increased H₂S levels, but that H₂S increase may well be an intrinsic characteristic of Covid-19 described for the first time herein. H₂S of healthy was also within reported ranges (7).

Fig. 1.

Hydrogen sulfide (H₂S) levels in serum of (A) healthy volunteers; (B) patients with ventilator associated pneumonia (VAP) by P aeruginosa; K pneumoniae; A baumannii; and (C) patients with pneumonia by the SARS-CoV-2 coronavirus on day 1 after hospital admission measured by monobromobimane derivatization assay followed by reverse phase HPLC separation. Comparison by the Mann–Whitney U test; ns indicates non-significant, ^∗P < 0.05, ^∗∗P < 0.01, ^∗∗∗P < 0.001, ^∗∗∗∗P < 0.0001.

To strengthen the finding of increased H₂S in Covid-19 survivors, H₂S was measured in the same samples by a photometric methylene blue assay (8). Despite the lack of specificity of this assay leading to higher measurable levels, the interpretation of the findings was the same (Fig. 2).

Fig. 2.

Hydrogen sulfide (H₂S) levels in serum of (A) healthy volunteers; (B) survivors; and (C) non-survivors of pneumonia by the SARS-CoV-2 coronavirus on day 1 after hospital admission measured by a photometric methylene blue assay. Comparison by the Mann–Whitney U test; ns indicates non-significant, ^∗P < 0.05, ^∗∗P < 0.01, ^∗∗∗P < 0.001, ^∗∗∗∗P < 0.0001.

We feel that Covid-19 is a new territory of research where modulation of H₂S plays a major role and we wish to thank Rademacher et al. (1) for paving us the way to strengthen our data.