The Correlation And Mechanism Between Mycoplasma Gallisepticum Infection And Commensal Microbiota Disorder In Chickens

In this study,four experiments were conducted to investigate the correlation and mechanism between MG infection and commensal microbiota.Experiment 1: Mycoplasma gallisepticum(MG)is the pathogen that causes chronic respiratory disease(CRD)in chickens.Gut microbiota plays an important role in maintaining body health and resisting respiratory infection,but the correlationbetween gut microbiota and MG infection is still unknown.Therefore,in this study,the correlationbetween gut microbiota and MG infection was explored by disruptinggut microbiota in chickens with antibiotics cocktail.The results showed that the gut microbiota enhance the pulmonary immune response against MG infection via IL-17A/granulocyte–macrophage colony-stimulating factor(GM-CSF)signaling.In addition,the elimination of respiratory macrophages by administered liposome clodronate demonstrated that the gut microbiota resisted MG infection through respiratory macrophages.After gut microbiota dysbiosis,MG clearance in lung was impaired,and this could be restored by orally administrated toll-like receptor 2(TLR2)ligand.We also isolated a strain Lactobacillus salivarius that enhances pulmonary immunity against MG infection via IL-17A/GM-CSF signaling.Lactobacillus salivarius as feed additive improved growth performance and immune activityof chickens post-MG challenge.The present study revealed an important role of gut microbiota in defense against MG infection and isolated a strain of Lactobacillus salivarius which has the potential to prevent MG infection.Experiment 2: MG infection alone or in combination with other pathogens have brought huge economic losses to the poultry industry.The intestinal microbiota plays a critical role in host defence against respiratory infection.To explore the role of intestinal microbiota in MG-induced inflammation-mediated lung injury and secondary Escherichia coliinfection,MGinfection model and fecal microbiota transplantation model were developed.The results showed that MGinfection changed gut microbiota composition along with lung inflammation injury.Fecal microbiota transplantation from chickens infected with MG to antibiotics cocktail treated chickens decreased host defense against Escherichiacoli due to impaired intestinal mucosal barrier,downregulated the m RNA expression levels of host defense enzymes and blocked autophagic flux.Lactobacillus salivarius intake alleviated lung inflammation injury caused by MG infection and increased host defense against Escherichiacoli by improved gut microbiota composition.Experiment 3: Baicalin shows excellent protective effects against Mycoplasma gallisepticum(MG)induced inflammatory injury in our previous studies.However,the physiological effects of baicalin are in notable contrast to its low bioavailability,and the critical mechanism for protective effects of baicalin against MG infection is still unclear.By using an MG infection model,the results showed that baicalin treatment significantly reduced MG colonization and ameliorated the abnormal pathological changes in lung.Baicalin treatment also reduced the level of proinflammatory cytokinesand suppressedproinflammatory protein expression.Notably,MG infection changed gut microbiota compositions,however,these abnormal gut microbiota compositions were partially alleviated by baicalin treatment.Baicalin significantly enriched the commensal bacterium Bacteroides fragilis,and gavage with Bacteroides fragilis alleviated MG infection induced inflammatory injury in lung.Experiment 4: Commensal microbiota has been shown to play an important role in local infections.However,the correlation between host respiratory microbiota and Mycoplasma gallisepticum(MG)infectionis not well characterized.Here,the results of 16 S r RNA sequencingshowed that MG infection correlated with alteration in respiratory microbiota of chickenscharacterized by decreased richness and diversity.To explore whether respiratory microbiota contributed to MG infection,an antibiotics cocktail was used to deplete respiratory microbiota.It has beenfound that depletion of respiratory Gram-positive and Gram-negative bacteria promoted MG infection,as reflected in the form of increased MG colonization,pro-inflammatory cytokines and proteins expression,and severe lung damage compared to the control group.Importantly,depletion of Gram-negative bacteria in respiratory tractmitigated MG infection,which indicated that certain Gram-negative bacteria may promote MG infection.By reconstitution of individual cultivable respiratory tract bacteria in antibiotic-treated chickens,a respiratory commensal microbe Serratia marcescenswas identified to facilitate MG infection.We further found that Serratia marcescens may promote MG infection by downregulating Mucin 2(MUC2)and tight junction related gene m RNA expression levels in trachea and lung tissues.Together,our data demonstrated that MG infection induced disturbed respiratory microbiota andthe specific respiratory commensal bacterium Serratia marcescens could promote MG infection,and thus expand our understanding of the pathogenesis of MG infection.