Dear Jan,

besides target modification by methylation, drug efflux or enzymatic inactivation of macrolides might cause a resistant pheotype.

in addtion to these well-known mechanisms, mutations in the 23SrRNA operons (e.g. at postions 2058/2059, E. coli numbering) or in ribosomal proteins L4 or L22 might cause macrolide and/or clindamycin resistance, at least in diverse Gram-positive or Gram-negative pathogens. This can be checked by sequencing those genes (23S rDNA, genes for L4/L22).

Kind regards

Wolf Splettstoesser

Hi Jan,

I am currently studying a mutant variant of the mdeA gene in S. aureus. According to the original publication (Huang, J; et al. Antimcrob Agents Chemother. 2004; 48(3):909-17. Pubmed PMID: 14982783). This gene when expressed at sufficiently high levels is capable of conferring resistance to clindamycin. Have you created msrA knockouts in S. aureus? I am currently in the process of creating an mdeA knockout. Let me know if you are interested in collaborating.

Thank you,

Zach Tackett

Dear Jan,

According to chapter 23 of the book antibiogram [by Patrice Courvalin, Roland LeClercq, Louis B. Rice] , "Mutant strains have been essentially isolated in the context of cystic fibrosis". I think you should keep that in mind since you are working on CF patients.

I agree with Wolf about the first two candidate mutants A2058 and A2059 because they may confer resistance to both erythromycin and clindamycin. However, mutants of protein L4 are usually susceptible to clindamycin and protein L22 mutants are hypersusceptible to this antibiotic.

You can also try erm(Y) and erm(33) genes [Please check Matsuoka et al, FEMS Microbiol Lett. 1998, PMID 9809423 for more information].

Regards,

Emran Askari

Thanks to all of you.

Sorry Zach I´m not working with mrsA mutants, I'm just studying prevalence of resistance mechanisms.

Kind regards

Jan

Update. Indeed the cause of the resistance in my isolates was mutations in the target site of MLSB antibiotics at the ribosome.  In the majority, it was the A2058G mutation, and more rarely mutations in L4 protein. Interestingly mutations were found in half of MLSB resistant isolates. And the association of mutations with the azithromycin treatment of the cystic fibrosis patients was proved.