Comparison of the microbiome of sputum from HIV/AIDS patients using PacBio 16S rRNA sequencing and ddPCR.

backgroundPulmonary infections are prevalent opportunistic complications among HIV/AIDS patients, necessitating precise pathogen identification for targeted treatment. The traditional culture methods used for diagnosing these infections have inherent limitations when it comes to detecting the diverse microbial flora present in HIV/AIDS patients.

methodsIn this study, sputum samples were collected from 37 HIV/AIDS patients admitted to Mengchao Hepatobiliary Hospital of Fujian Medical University. A total of 45 samples were subjected to analysis using three different methods: traditional culture methods, third-generation long-read sequencing (PacBio), and droplet digital PCR (ddPCR). The study aimed to compare the diagnostic capabilities and accuracy of microorganism identification among these methods.

resultsOut of 45 sputum samples, the traditional culture method identified only 5 positive cases, whereas PacBio sequencing detected all samples positively. Among the 7 samples (including 5 traditional culture positive and 2 negative samples), PacBio sequencing was able to identify 51 species (77 strains), while ddPCR detected 15 species (64 strains). In contrast, traditional culture methods were only able to identify 3 bacterial species (6 strains). In samples that tested positive by culture, both PacBio sequencing and ddPCR were consistent in identifying the primary microorganisms, indicating the presence of multiple concurrent infections. In samples that were negative by culture, both PacBio and ddPCR were able to detect a broader range of microorganisms, showing a higher sensitivity compared to traditional methods. PacBio sequencing provided a comprehensive view of the microbial flora, whereas ddPCR enabled rapid and precise detection of common microorganisms and antibiotic resistance genes, compensating for PacBio's shortcomings in absolute quantification.

conclusionsThe study demonstrates that PacBio sequencing and ddPCR offer significant advantages over traditional culture methods in diagnosing pulmonary infections in HIV/AIDS patients. These advanced techniques not only detect a greater variety of pathogens but also provide a more detailed understanding of the microbial flora and resistance profiles, which is crucial for effective clinical management. The combination of PacBio's comprehensive profiling and ddPCR's rapid detection capabilities presents a powerful approach to overcoming the limitations of traditional diagnostic methods in this patient population.