| Background:With the popularization of computed tomography,more and more lung nodules(lung lesions smaller than 30 mm)are being detected,part of which are adenocarcinomas of the lung characterized by ground glass opacity(GGO-type lung adenocarcinoma).The high incidence of GGO-type lung adenocarcinoma not only places a heavy burden on the medical system,but also causes great emotional stress to patients.The treatment of lung adenocarcinoma characterized by GGO has also become a hot issue facing thoracic surgery,especially the treatment of GGO-type multiple primary lung adenocarcinoma,which poses a great challenge to thoracic surgeons.At present,the etiology of GGO-type lung adenocarcinoma,especially multiple primary lung adenocarcinoma,is still unclear.It has been suggested that "regional carcinogenesis" is closely related to the development of multiple primary lung cancer,while the distribution of microbiota in human body is regionalized,and many studies have shown that microbiota is closely related to the development of lung cancer.However,there is a lack of research on the microbiota in the lower respiratory tract of GGO-type single lung adenocarcinoma and GGO-type multiple primary lung adenocarcinoma and the differences in microbiota between the two groups using Metagenomics sequencing.Methods:Alveolar lavage fluid from patients with lung adenocarcinoma characterized by ground glass opacity who met the criteria and were seen in the Department of Thoracic Surgery I of the First Hospital of Jilin University from August 1,2022 to December 30,2022 were included for Metagenomics sequencing,depending on whether the lesions were single or multiple,they were divided into single lesion group and multiple lesions group.After performing Metagenomics sequencing,bioinformatics analysis of the sequencing data was performed.Species annotation was performed using the default parameters of Kraken2,while species-level abundance of Metagenomics samples was estimated by Bracken using Bayesian algorithm and Kraken classification results.For database selection,the Nt(202011)database was used.Functional annotation of non-redundant genes was generally performed using the BLASTP function of the software Diamond,the KEGG pathway database and the KEGG module database were included.The sample size was justified by plotting the species accumulation curve.The alpha diversity and beta diversity of the two data groups were calculated to determine the differences in macroscopic distribution of microbiota between the two groups.The LEf Se analysis was used to find biomarkers that were statistically different between groups.The enrichment results of KEGG pathway and KEGG module of microbiota in the two groups were tested statistically with Reporter Score.Results:A total of 19 patients were included in the study,and after Metagenomics sequencing,a total of 472 species were detected in both groups at the species level,406 species in the solitary-lesion group and 369 species in the multiple-lesion group.The distribution of the microbiota of the two groups at different levels was exhibited.The lower respiratory microbiota of patients in both the single-lesion group and multiple-lesion group were dominated by the Proteobacteria at the phylum level.There was no statistical difference in the alpha diversity and beta diversity of lower respiratory tract microbiota between the two groups.A total of 29 statistically significant differential species were detected in different taxonomic levels of microbiota in patients in the single-lesion group and multiple-lesion group,with species of Neisseria and its subordinate branches and species of Haemophilus and its subordinate branches being mainly more abundant in the multiple-lesion group,the difference was statistically significant.After KEGG database annotation,3108 functional genes were annotated for microbiota in the multiple-lesion group and 3494 functional genes were annotated for microbiota in the single-lesion group.The gene functions of lower respiratory tract microbiota in the two groups of patients were significantly different in 21 modules(KEGG module)and 1 pathway(KEGG pathway),and pathways related to biotin metabolism were enriched in the microbiota genes of the single-lesion group.The genes of the multiple-lesion group of microbiota are enriched in these aspects:Menaquinone biosynthesis,chorismate(+ polyprenyl-PP)=> menaquinol;Phylloquinone biosynthesis,chorismate(+ phytyl-PP)=> phylloquinol;Molybdenum cofactor biosynthesis,GTP => molybdenum cofactor;Assimilatory sulfate reduction,sulfate => H2S;C1-unit interconversion,eukaryotes;C1-unit interconversion,prokaryotes;3-Hydroxypropionate bi-cycle;Tetrahydrofolate biosynthesis,GTP =>THF;Heme biosynthesis,bacteria,glutamyl-t RNA => coproporphyrin III => heme;Thiamine biosynthesis,prokaryotes,AIR(+ DXP/glycine)=> TMP/TPP;Fatty acid biosynthesis,initiation.The genes of the single-lesion group of microbiota are enriched in these aspects: Proline biosynthesis,glutamate => proline;Pentose phosphate pathway,oxidative phase,glucose 6P => ribulose 5P;Lysine biosynthesis,DAP dehydrogenase pathway,aspartate => lysine;Pimeloyl-ACP biosynthesis,Bio CBio H pathway,malonyl-ACP => pimeloyl-ACP;Arginine biosynthesis,ornithine =>arginine;Urea cycle;UDP-N-acetyl-D-glucosamine biosynthesis,prokaryotes,glucose => UDP-Glc NAc;Entner-Doudoroff pathway,glucose-6P => glyceraldehyde-3P + pyruvate;Lysine biosynthesis,DAP aminotransferase pathway,aspartate =>lysine;Lysine biosynthesis,acetyl-DAP pathway,aspartate => lysine;NADH:quinone oxidoreductase,prokaryotes.Conclusions:1.Proteobacteria may be associated with the etiology of GGO-type lung adenocarcinoma.2.Neisseria and its subordinate branches and Haemophilus and its subordinate branches may be associated with the development of GGO-type multiple primary lung adenocarcinoma.3.Synthesis of tetrahydrofolate,synthesis of fatty acids,and production of hydrogen sulfide may be associated with the development of GGO-type multiple primary lung adenocarcinoma. |
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