The Characterization Of Airway Microbiome In Non Small Cell Lung Cancer Patients With Different Clinicopathology

Background:The incidence and mortality rate of lung cancer ranked first in all tumors.Different biological models and different molecular biological patterns exist in different pathological types and different stages of lung cancer.Previous studies have confirmed that compared with healthy control,lung cancer patients showed alteration of lung microbiome,mainly manifested in the significant change of microbiome diversity and abundance of some genera.However,few studies had investigated the correlation between lung microbiome and different pathological types and metastasis stage of lung cancer.In addition,bronchoalveolar washing fluid(BWF)and sputum are commonly used samples to study the lung microbiome of patients with advanced lung cancer.Up to now,there is no study comparing the difference between the two sample types.Objective:To characterize lung microbiome in squamous cell lung carcinoma with(SCC_M1)or without distant metastasis(SCC_M0)and lung adenocarcinoma with(AD_M1)or without distant metastasis(AD_M0).To investigate the difference between BWF sample and spontaneous sputum sample of lung cancer.Methods:We collected BWF samples from 40 newly diagnosed lung cancer patients(LC)and 7 patients with benign lung disease(SIS),and 52 spontaneous sputum samples from patients with newly diagnosed LC.Five blank BWF tubes were collected as negative controls.Bacterial species of BWF samples and sputum samples were sequenced via 16S rRNA sequencing with V3-V4 region.Bioinformatics and statistical methods were used to compare the lung microbiome differences between each two groups.We compared the lung microbiome between BWF samples and sputum samples,BWF samples of LC and BWF samples of SIS,SCC_M0 and AD_M0,SCC_M1 and AD_M1,SCC_M0 and SCC_M1,AD_M0 and AD_M1.Results:The predominant phylum of BWF samples from LC patients were Firmicutes(32%),Proteobacteria(30%),Bacteroides(25%).The predominant genera were Prevotella(20%),Streptococcus(12%),Veillonella(8%),Neisseria(5%).While the predominant phylun of sputum samples in LC patients were Firmicutes(43%),Bacteroidetes(27%),Proteobacteria(17%)and Streptococcus(23%),Prevotella(20%),Neisseria(11%),Porphyromonas(6%),Veillonella(5%)for the predominant genera.Phylum Proteobacteria in BWF samples were significantly higher than sputum samples(Mann-Whitney U test,P=0.003)and genus Streptococcus was significantly lower(Mann-Whitney U test,P=0.0002).At phylum level,the microbiome of BWF samples was more similar to tihat of lung cancer tissues reported in the previous literature.We found that Streptococcus was the most important differential genus between BWF and sputum sramples(LDA score=4.8).What's more,the relative abundance of Streptococcus in sputum of peripheral lung adenocarcinoma was significantly higher than that of central lung adenocarcinoma(Mann-Whitney U test,P=0.0158),indicated that some bacterium may have breeded in sputum.The a diversity(Chaol,P=0.354;Shannon index,P?0.43 ? Simpson index,P?0.14)and ? diversity(PERMONOVA,P? 0.14)between BWF samples from LC patients and SIS patients were similar.LEFse analysis showed that Proteobacteria was significantly increased in patients with LC.The diversity of lung microbiome was limilar among different pathological types and different metastatic states of lung cancer.In BWF group and sputum group,The a diversity(Chaol,Shannon index and Simpson index,P>0 05)and p diversity(PERMONOVA,P>005)between SCC_M0 and AD_M0,SCC_M1 and AD_M1,SCC_M0 and SCC_M1,AD_M0 and AD_M1 were similar.In different metastatic states,differential genera between different pathological types are different.LEFse analysis showed that in BWF group,tgenera Veillonell,Megasphaera,Actinomyces and Arthrobacter in AD_M0 were significantly higher than those in SCC_M0,and genera Capnocytophaga and Rothia in AD_M1 was significantly lower than that in SCC_M1.In different pathological types,differential genera between different metastatic states are different.LEFse analysis showed that in BWF group,compared with AD_M0,genus Streptococcus of AD_M1 was significantly lower,and genera Veillonella and Rothia in SCC_M1 was significantly higher than that in SCC_M1.Conclusions:Our study suggests that BWF samples may better reflect the lung microbiome of lung cancer tissues than sputum samples.Compared with SIS patients,Proteobacteria was significantly increased in BWF samplesfrom LC patients.Our results showed complex interaction between lung microbiome and histology type and metastatic stage of lung cancer.The difference of microbiome between squamous cell carcinoma and lung adenocarcinoma existed and was related with different distant metastatic states.While the difference of microbiome between different distant metastatic states of lung cancer was related with different pathological types.