Study Of Vitreous Humoral Flora And Pathogenic Bacteria In Exogenous Endophthalmitis Patients Based On Culture And High-Throughput Sequencing

Objective:This study is aim to analysis the ocular microbial of endophthalmitis patients via16 S r DNA High-throughput squencing technology and culture with vitreous humor,describe the abundance and diversity of the microbial flora,explore the main pathogenic bacteria,so that provide some clinical and theoretical proofs to the application of microbiome in ophthalmology.Methods:Endophthalmitis patients admitted to Shanxi Eye Hospital from January 2021 to May 2022 were selected as the subjects.The control group were patients with retinal detachment or idiopathic macular hole who underwent vitrectomy surgery during the same period.Collect 0.5ml of vitreous fluid of these patients,then examine via High-throughput squencing technology and culture.The intraocular lavage fluid used during the operation was used as a blank control.Extracte total bacterial DNA from the specimens,perform the PCR amplifications,build the gene library,and sequence the V3-V4 region of the 16 S r DNA gene with the Illumina Nova Seq 6000.Compare endophthalmitis group with control group of Tags number,OTUs,diversity analysis and species difference analysis.Compare the sequencing results with the conventional cultures results.Analyze the classification composition,distribution characteristics and the relative content of each pathogen in the specimen.Results:The analysis of Tags,OTUs,diversity analysis and species difference analysis between the endophthalmitis group and the control group found that some of the endophthalmitis group samples showed significant similarity with the control group.These samples(9 cases)were separately set as Low Bioinformation endophthalmitis group,referred to as LB group,and the remaining endophthalmitis samples(18 cases)were set as High Bioinformation endophthalmitis group,referred to as HB group.There was no significant difference between the LB group samples and the control group in the tags,OTUs,α diversity and β diversity analysis.LB group samples and control group share the dominant bacterial genus: Bacteroides,Faecalibacterium,Lactobacillus.The top 5 bacterial genera with the highest relative abundance in the HB group are :Staphylococcus,Streptococcus,Pseudomonas,Brevundimonas,Moraxella.The flora abundance and diversity of the HB group were significantly different from the control group.Significant differences were observed at the phylum and genus levels between the HB and control dominant samples: More distribution of Firmicutes in the HB group compared to the control group(P<0.001);The mean relative abundance of Streptococcus and Staphylococcus genera increased significantly in the HB group(P <0.05);Positive bacterial cultures were obtained in 12 of 27 samples(44.4%);There were 11 cases consistent with the dominant bacteria,including 6 Staphylococcus,3 Streptococcus,1Brevundimonas and 1 Pseudomonas.In the culture results,the main dominant bacteria was Staphylococcus epidermidis.Nine samples were positive for only one bacterium.Three samples tested positive for two or more bacteria.Seven cases containing two or more bacteria with an average relative abundance greater than 10% were obtained by the sequencing method.The absolute proportion of metallic lesions(wire,nail / sheet / chip)in traumatic endophthalmitis(70.37%).Conclusion:The bacteria causing exogenous endophthalmitis were mainly Staphylococcus species and Streptococcus species.Two or more dominant bacteria may be included in the vitreous fluid of patients with exogenous endophthalmitis.In the identification of infectious endophthalmitis pathogens,high-throughput sequencing technology may be complementary when applied together with traditional culture.