Pathogen identification by shotgun metagenomics of patients with necrotizing soft-tissue infections

Background Necrotizing soft-tissue infections (NSTIs) are life threatening, requiring broad-spectrum antibiotics. Their aetiological diagnosis can be limited by poor performance of cultures and administration of antibiotics before surgery. Objectives We aimed (i) to compare 16S-targeted metagenomics (TM) and unbiased semiquantitative panmicroorganism DNA- and RNA-based shotgun metagenomics (SM) with cultures, (ii) to identify patients who would best benefit from metagenomics approaches and (iii) to detect the microbial pathogens in surrounding non-necrotic 'healthy' tissues by SM-based methods. Methods A prospective observational study was performed to assess the analytical performance of standard cultures, TM and SM on tissues from 34 patients with NSTIs. Pathogen identification obtained with these three methods was compared. Results Thirty-four necrotic and 10 healthy tissues were collected from 34 patients. The performance of TM was inferior to that of the other methods (P < 0.05), whereas SM performed better than standard culture, although the result was not statistically significant (P = 0.08). SM was significantly more sensitive than TM for the detection of all bacteria (P = 0.02) and more sensitive than standard culture for the detection of anaerobic bacteria (P < 0.01). There was a strong correlation (r = 0.71, Spearman correlation coefficient) between the semi-quantitative abundance of bacteria in the culture and the bacteria to human sequence ratio in SM. Low amounts of bacterial DNA were found in healthy tissues, suggesting a bacterial continuum between macroscopically 'healthy' and necrotic tissue. Conclusions SM showed a significantly better ability to detect a broader range of pathogens than TM and identify strict anaerobes than standard culture. Patients with diabetes with NSTIs appeared to benefit most from SM. Finally, our results suggest a bacterial continuum between macroscopically 'healthy' non-necrotic areas and necrotic tissues.