| Background Chronic osteomyelitis is a chronic intramedullary infection.Its characteristics are caused by pathogenic bacteria(microorganisms).Low inflammation,the formation of dead bone,with or without sinus tract formation.Chronic osteomyelitis is still one of themost challenging diseases in clinic due to its long duration,complicated treatment and high risk of recurrence.According to the etiology,chronic osteomyelitis can be divided into hematogenous osteomyelitis,traumatic osteomyelitis,and diabetic foot osteomyelitis.From the point of view of etiology,the pathogenic bacteria of hematogenous osteomyelitis and traumatic osteomyelitis aremainly of the golden grape fungus.And diabetic foot osteomyelitis ismainly with strange proteus.Microbiome refers to amicrobial community consisting of several species.There are various complex relationships between different communities,such asmutual benefit,symbiosis,coexistence and competition.It can be found that the gastrointestinal tract contains themost bacteria,which has important influence on humanmicrobiome.The gutmicrobiome is coevolutionary with their host.And it is strongly influenced by the host diet,mood,disease,medicine and so on.Therefore,we can reflect the health status and growth status of animal or human gutmicrobes.In the development of chronic osteomyelitis,both local and systemic immunity will change.Previous studies have shown that inflammation,antibiotic use,and long-term psychiatric anxiety are all linked to changes in gut flora.Chronic osteomyelitis inevitably accompanied by local or systemic inflammation,in the whole course of antibiotics use andmental factors change,the body's immune in the whole course of the disease at the same time also will play an important role.So in the process of the onset of chronic osteomyelitis,the composition of intestinal flora is changed,there is no related research reports,so we assumed that patients with chronic osteomyelitis of gutmicrobes through changes in its composition andmetabolites,affect the body's immune function,thus impact on disease outcome.Experiments in this paper,by constructing the chronic osteomyelitismodel to study its correlation with intestinalmicrobiome change and its clinical significance lies in the intestinal flora changes and chronic osteomyelitis correlation.Material andmethodsAnimal experiment A volume of 10 ?L of bacterial suspension was injected into the osseous cavity of the tibiae containing 106 CFU Staph.aureus/mL,of a clinical osteomyelitis isolate(n= 5).Animals in the control group were injected with the same volume of PBS(n = 5).Fecal sample collection and DNA extraction Fecal samples were collected at 6months after injection of the bacteria,Samples were collected directly from the animal upon defecation and immediately frozen at-80?,prior to DNA extraction.microbial genomic DNA was extracted from 200mg of each fecal sample using a TIANamp Stool DNA Kit(Spin Column,Cat.no.DP328).Amplification,sequencing of 16S rRNA gene,and data analysis The V4 hypervariable regions of 16S rRNA gene were amplified by PCR using the barcoded derivatives.The 16S rRNA gene amplicons were then sequenced using the paired-endmethod on the Illuminamiseq platform according tomanufacturer's instructions.MicroCT scan Themice were sacrificed via cervical dislocation and the right tibials were removed and fixed in 4%paraformaldehyde.All samples were scanned with amicroCT(Bruker Skyscan 1176,Kontiche,Belgium;9?m resolution).Identification of bacteria The mice were sacrificed via cervical dislocation and the right tibias were removed.The cavity content was taken and painted on the agar plates.After cultured at 37? for 18 h,sent it to a clinical laboratory at Nanfang Hospital for identification.Statistical analysis Histology scores obtained for each individual parameter were averaged to obtain a composite score for eachmouse.SPSS 20(IBM,USA)was used for the statistical analyses.The data were checked for homogeneity of variance using levene's test.Differences between groups were determined by independent-sample t test(Student's t test)for parametric two-tailed significance.The significance level was determined at P<0.05.Graphical representation of the data was performed in GraphPad Prism 7(GraphPad,USA).ResultIdentification of bacteria The automatic bacterial identification instrument in the clinical laboratory at Nanfang Hospital showed Staphylococcus aureus.MicroCT The cross-sectional images showed although the two groups had callus formation outside the cortex,destruction of the cortex dissolved occurred dramatically in the infection group.The BV and BV/TV in S.aureus group had a significant decrease compared to Control Group.ConclusionAmicemodel of chronic osteomyelitis has been successfully establishedThe gutmicrobiome of group S.aureus has a deduced ?-diversity without statistical significance.After T-test between the two groups,we found group S.aureus has high OTUs abundances of Lactobacillales,Lactobacillaceae and Lactobacillus.Then we performed themetaStat test,we found group S.aureus has a low OTUs abundance of Streptococcus thermophiles. |
PDF Full Text Request