To observe the application of alveolar lavage fluid second-generation sequencing in diagnosing and guiding the treatment of patients with severe pneumonia with unidentified pathogenic bacteria and to provide novel ideas and methods for the effective clinical treatment of this disease. The clinical data of 80 patients with severe pneumonia included in our intensive care unit from June 2020 to June 2022 were analyzed and all patients had undergone metagenomic next-generation sequencing and traditional cultures (alveolar lavage fluid culture, sputum culture) to analyze the advantages of metagenomic next-generation sequencing in detecting pathogens in patients with severe pneumonia. The positive rate of metagenomic next-generation sequencing pathogen detection was higher than that of conventional culture. In patients who were negative for conventional culture and positive for metagenomic next-generation sequencing, metagenomic nextgeneration sequencing was able to further identify multiple pathogenic infections. In terms of pathogen distribution, metagenomic next-generation sequencing detected 71 bacterial, 39 fungal and 3 viral strains. In mixed infections, metagenomic next-generation sequencing yielded a higher rate of positive diagnosis. In addition, metagenomic next-generation sequencing had a higher pathogen detection rate in patients with combined underlying diseases and metagenomic next-generation sequencing could identify specific pathogenic infections especially in patients with combined immunocompromised conditions. Metagenomic next-generation sequencing could improve the detection rate of pathogenic microorganisms in patients with severe pneumonia and could be used as a complementary test for patients negative for conventional cultures. Metagenomic next-generation sequencing has advantages in the diagnosis of mixed infections and can identify multiple pathogenic infections simultaneously. Clinically, early application of metagenomic next-generation sequencing in patients with severe pneumonia is recommended for increased clinical benefit.

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