Microbiota Dysbiosis Of Tinea Pedis And Foot Commensal Bacteria Antagonize Pathogenic Fungi

Objective:Tinea pedis is a dermatophytic infection of the feet,including soles,interdigital clefts of toes,and nails.Recently,extentive studies have devoted to the chemical drug treatment,but the recurring pathology and underlying mechanism of tinea pedis are largely unclear.It is thought that Dermatophytic fungi,such as Trichophyton rubrum,are primarily responsible for these infections based on microbial culture technology.What’s more,studies find that skin microbiome dysbiosis affects the outcomes of several dermatological diseases,suggesting that foot microbiome profoundly influences the pathology of tinea pedis.However,the dysbiosis of microbial community associated with tinea pedis is largely unexplored.Therefore,it is urgent to investigate the relationship of tinea pedis and microbial community using deep-sequencing technique.This study attempted to explore alteration in the composition and function of microbiota associated with tinea pedis.Moreover,we elucidate the potential role of commensal bacteria in inhibiting pathogenic fungi of tinea pedis and provide an insight into the potential application of probiotics to tinea pedis disease.Methods:Part one:High-throughput sequencing technology was applied to analyze the composition and function of microboiome associated with tinea pedis.Including:1.Sample collection: the microbiota on foot surface from students of Fenyang College of Shanxi Medical University,tinea pedis patients and healthy people in Fenyang urban area were collected.Then samples were stored at-80 ℃ after recording.They would be sent to the company for high-throughput sequencing analysis after the samples were collected completely.2.The original data of samples were obtained through a series of processes including sample preparation,DNA extraction and detection,PCR amplification,product purification,database preparation and detection and Miseq computer sequencing.3.Effective data was used to cluster analysis after de-fitted,low-quality filtered and removed chimeric.Each cluster unit was an operational classification unit(OTU).The analysis of OTU was used to obtain the species distribution information of each sample.4.Based on the results of OTU analysis,α diversity analysis was prepared to obtain the species abundance and diversity information;The PCo A and Anoism analysis were performed to show the difference of community construction between samples and groups.5.PICRUSt functional prediction was performed to predict the metabolic function of microbial communities.Part two:1.A tinea-pedis-related pathogen was isolated from foot skin by extracorporeal fungal isolation technique,and was identified as A.alternata named as K1.The strain was used to screen the antagonism of probiotics.2.Total 32 strain of bacteria were separated from foot skin via microbial isolation technology and were identified by 16 S r DNA.3.Preliminary screening and rescreening the antagonists bacteria to K1: the bacteria with antagonistic effect to A.alternata K1 was isolated via medium confrontation,and was identified by morphology and molecular biology.The strain was named as Bacillus pumilus X1.4.A screen of antagonists against T.rubrum: the bacteria with antagonistic effect to the standard strain of T.rubrum was confirmed by medium confrontation.5.Inhibiting effects of B.pumilus X1 on the growth of mycelia and spore germination of A.alternata K1.6.Optimization of antagonistic conditions for B.pumilus X1.7.The effect of bacterial metabolites and A.alternata K1.8.Used in accordance with the antibacterial peptide lipid to the primer sequences amplification B.pumilus X1 gene,detected the secretion of fat antibacterial peptide.Results:1.Compared with healthy people,the microbial community structure of tinea pedis patients were changed,which was mainly reflected in the decreased abundance of Corynebacterium,Dermatococcus,short Bacillus and Aerococcus and increased abundance of Enterobacter,and the decreased abundance of Penicillium and Aspergillus and the increased abundance of Acremonium.Diversity analysis of species between groups showed that the microbial community of tines pedis was significantly different from that of healthy people(P<0.01).2.PICRUSt,COG and KEGG databases were used to predict microbial community functional map for sequencing data.The results showed that compared with the healthy group,cell motility,intracellular trafficking,secretion and environmental information processing were increased in the tinea pedis group.All results indicate that tinea pedis is accompanied with the dysbiosis of microbiota function.3.Total 32 strains bacteria were isolated from foot skin,among which X1 has a strong antagonistic effect against pathogenic fungus K1.X1 was identified as Bacillus pumilus by gene sequence analysis,and its sterile fermentation supernatant also has an inhibitory effect on K1.4.The strain X1 was proved to have a strong antagonistic effect to T.rubrum.5.Bacteria X1 would cause the abnormal phenomena such as disordered,twisted and even broken mycelia in K1,and decrease the number of spores through inhibiting the growth of spores.6.Under different p H,temperature and distance,the antifungal activity of strain X1 to K1 was different.7.B.pumilus X1 can inhibit A.alternata K1 independent of metabolite.Conclusion:Compared with healthy persons,the microbial community and function of tinea pedis were altered.B.pimilus X1 effectively inhibits A.alternata and T.rubrum that are associated with tinea pedis.Our studies reveal the relationship between tinea pedis diseases and microbiota dysbiosis,and screen the probiotics that inhibit the pathogenic fungi related to tinea pedis.These results provide an insight into the investigation of tinea pedis-associated microorganisms and a reliable potential resource for clinical on tinea pedis.