A Study On The Adaptation Of Lactobacillus Acidophilus During The Evolution Of Aminoglycoside Antibiotics

Lactobacillus acidophilus is a widely used probiotics in food industry.However,L.acidophilus is sensitive to antibiotics and easy to lose activity in practical application.Therefore,this research studied the change of resistance and phenotype characteristics of L.acidophilus under the adaptation of aminoglycoside antibiotics utilized whole genome sequencing technology,providing the basis for the combined use of L.acidophilus and aminoglycoside antibiotics.1.The research in antibiotic resistance of Lactobacillus acidophilus.The resistance of L.acidophilus to aminoglycoside antibiotics was observed by macro broth dilution method.The results showed that L.acidophilus ATCC4356 was sensitive to gentamicin,kanamycin,streptomycin and neomycin.Under the selection pressure of antibiotic,the MIC values of L.acidophilus to gentamicin,kanamycin,streptomycin and neomycin increased to 4?g/mL,128?g/mL,512?g/mL and 16?g/mL,respectively.Compared with the original strains,the resistance of L.acidophilus to gentamicin,kanamycin,streptomycin and neomycin increased by 32 times,64 times,512 times and 64 times,respectively.Under the removed selection pressure of antibiotic,except that the resistance to gentamicin remained constant,the resistance of L.acidophilus to kanamycin,streptomycin and neomycin were reduced to 64?g/mL,256?g/mL and 8?g/mL,respectively.Compared with the original strain,the antibiotic resistance was still higher.2.Phenotypic stability of L.acidophilus in the evolution of aminoglycoside antibiotics.The results showed that L.acidophilus could better maintain cell appearance,colony morphology and bacterial biomass,indicating that it has little effect on its biological characteristics.The acid-producing ability of L.acidophilus evolving in aminoglycoside antibiotics could be maintained well,but the ability to utilize fructose,lactose,cellobiose,maltose,and salicin was altered.L.acidophilus evolving in aminoglycoside antibiotics caused changes in its probiotic properties.L.acidophilus evolved under selection pressure of antibiotic had lower tolerance of acid and sodium salt than those under removed selection pressure.Under selection pressure of antibiotic,L.acidophilus showed the reduction of self-agglutination ability and cell hydrophobicity,and improvment of bile salt tolerance with little effect on phenol tolerance.3.Whole genome sequencing of L.acidophilus and safety evaluation.Whole genome sequencing was used to study the resistance mechanism of L.acidophilus to aminoglycoside antibiotics under selection pressure and removed selection pressure,and identified gene mutations associated with resistance and fermentation and probiotic properties.A total of 11 nonsynonymous mutations were identified related to antibiotic resistance,including 7 SNVs and 4 SVs.7 SNVs were the cell division protein FtsA,two-component sensor histidine kinase,BMP family ABC transporter substrate-binding protein,and PTS cellobiose transporter subunit IIC,hypothetical protein,hydrolase TatD and sugar transporter,and 4 SVs included hypothetical protein,uracil permease,ATP synthase alpha subunit and cardiolipin synthase.In aspect of fermentation characteristics and probiotic characteristics,5 gene mutations were detected,namely PTS cellobiose transporter subunit IIC,ECF transporter S component,and the ClC family H(+)/Cl(-)exchange transporter,branched chain amino acid aminotransferase and ABC transporter ATP-binding protein.After a preliminary analysis of the transfer of resistant genes,it was verified that none of them were on mobile elements,indicating that L.acidophilus was safe and providing a scientific basis for its future application.