Impact Of Macrolide Resistance On The Fitness In Streptococcus Suis

As an important zoonotic pathogen,Streptococcus suis have a worldwide distribution since reported.Macrolides is the effective drug to treat the Swine streptococosis initially,but more and more macrolide resistant Streptococcus suis appear in any country where used macrolides extensively.The spread of drug-resistant bacteria in environment is determined by the effect of resistance mechanisms on the fitness of host bacteria.To understand the development and fitness of macrolide resistance in Streptococcus suis,in our study,we studied the influence of resistant mutations and resistant genes on the fitness of host bacteria.The results of the study could provide ideas for developping effective measures to control and prevent macrolide resistant Streptococcus suis.Firstly,the growth curve and relative fitness of the isolates which carrying mefA/msrD have compare with that of the susceptible strains.To further study the fitness cost of the macrolide resistance gene msrD,the thermo sensitive suicide vector pSET4s was used to construct the msrD gene deleted Streptococcus suis NJ-2 mutants.The macrolide resistance gene mefA which in the appropriate vector were transferred into E.coli,then determine the influence of mefA on the resistance and adaptability of host bacteria.The the in vitro fitness has no significant difference between resistant strains and sensitive strains.The fitness of the E.coli which mefA had been transformed into has no significant differences compared to the parental strain.After the pSET4s::msrDL-msrDR plasmid constructed by fution-PCR electro-transformed into Streptococcus suis strain NJ-2,there was no positive site mutated strain was found in our selection.It implied that msrD may be important for the fitness of Streptococcus suis NJ-2.To understand the development and fitness cost imposed by macrolide resistance gene ermB,we made a primary research on the influence of resistant genes on the fitness of host bacteria.The growth curve and relative fitness of the isolates which carrying ermB are compare with that of the susceptible strains.To further study the fitness cost of the macrolide resistance gene ermB,the macrolide resistance gene ermB which in the appropriate vector were transferred into E.coli,then determine the influence of ermB on the resistance and adaptability of host bacteria.The in vitro fitness has no significant difference between resistant strains HB1004,HB1010 and SC05017 and two sensitive strains,but the fitness of JS07002 was lower than the sensitive strains.There is a decline in the fitness of the macrolide resistant E.coli which were mediated by ermB gene compared to the parental strain.To examine the development of macrolide resistance in Streptococcus suis and assess the fitness of the macrolide resistant mutants,four macrolide susceptible Streptococcus suis strains from different geographic areas were exposed to increasing concentrations of erythromycin,azithromycin and tylosin in vitro.Mutation in 23 S rRNA gene and genes encoding ribosomal protein L4 and L22 of highly resistant strains selected by macrolide passage in vitro was sequenced.The resistant strains with mutation in target genes were subject to fitness assay in vitro and in vivo.The results showed that all induced Streptococcus suis mutants that highly resistant to macrolide has mutations in 23S rRNA gene,some strains mutated at genes encoding ribosomal protein L4 and L22.All of the mutants harbor significant fitness cost compared to the parental strain in vivo and in vitro except mutants of the strains mutated at position of A2059 in 23 S rRNA gene.The above results indicate that the relative fitness of the isolates which carrying mefA/msrD was compared to that of the susceptible strains has no significant difference.The finess of S.suis type 2 strain JS07002 which carrying ermB was lower than that of the susceptible strains.The fitness of the E.coli which mefA had been transformed into has no significant differences compared to the parental strain,but there is a decline in the fitness of the macrolide resistant E.coli that were mediated by ermB gene compared to the parental strain.the mutation at the position of A2059 in 23SrRNA have imposed no fitness cost in host strain,or even raises the in vivo fitness of host strain in ICR mice;the mutation at the position of G2057 and A2062 in 23SrRNA sequence may harbor high fitness cost.