Generalized Bacteriophage Transduction in Serratia marcescens

Shirshikova, Tatiana V.; Морозова, О. В.; Kamaletdinova, Leisan Kh.; Шарипова, М. Р.; Bogomolnaya, Lydia M.

doi:10.1007/s12668-016-0268-z

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“…All S. marcescens strains used in this study are listed in Table 3 . The Δ macAB mutant strain was generated by lambda red homologous recombination ( 74 , 75 ) in nuclease-deficient, restrictionless S. marcescens strain TT392 ( 76 ), and the mutation was then moved into S. marcescens SM6 by bacteriophage ΦOT8 transduction ( 77 , 78 ). Strains were routinely grown in LB broth (10 g/liter Bacto-Tryptone, 5 g/liter yeast extract, 5 g/liter NaCl) or Mueller-Hinton broth (BD Difco).…”

Section: Methodsmentioning

confidence: 99%

“…The resulting linear DNA fragment was used for lambda red-mediated homologous recombination to replace the chloramphenicol resistance cassette in the Δ macAB mutant strain (LMB28) with the full-length macAB coding sequence bearing FLAG tag and resistance to kanamycin. The mutant construct was moved into the S. marcescens SM6 strain background by bacteriophage ΦOT8 transduction ( 77 , 78 ). The resulting clones were confirmed by PCR.…”

Section: Methodsmentioning

confidence: 99%

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The ABC-Type Efflux Pump MacAB Is Involved in Protection of Serratia marcescens against Aminoglycoside Antibiotics, Polymyxins, and Oxidative Stress

Shirshikova

Sierra-Bakhshi

Kamaletdinova

et al. 2021

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Serratia marcescens is an emerging pathogen with increasing clinical importance due to its intrinsic resistance to several classes of antibiotics. The chromosomally encoded drug efflux pumps contribute to antibiotic resistance and represent a major challenge for the treatment of bacterial infections. The ABC-type efflux pump MacAB was previously linked to macrolide resistance in Escherichia coli and Salmonella enterica serovar Typhimurium. The role of the MacAB homolog in antibiotic resistance of S. marcescens is currently unknown. We found that an S. marcescens mutant lacking the MacAB pump did not show increased sensitivity to the macrolide antibiotic erythromycin but was significantly more sensitive to aminoglycoside antibiotics and polymyxins. We also showed that, in addition to its role in drug efflux, the MacAB efflux pump is required for swimming motility and biofilm formation. We propose that the motility defect of the ΔmacAB mutant is due, at least in part, to the loss of functional flagella on the bacterial surface. Furthermore, we found that the promoter of the MacAB efflux pump was active during the initial hours of growth in laboratory medium and that its activity was further elevated in the presence of hydrogen peroxide. Finally, we demonstrate a complete loss of ΔmacAB mutant viability in the presence of peroxide, which is fully restored by complementation. Thus, the S. marcescens MacAB efflux pump is essential for survival during oxidative stress and is involved in protection from polymyxins and aminoglycoside antibiotics. IMPORTANCE The opportunistic pathogen Serratia marcescens can cause urinary tract infections, respiratory infections, meningitis, and sepsis in immunocompromised individuals. These infections are challenging to treat due to the intrinsic resistance of S. marcescens to an extensive array of antibiotics. Efflux pumps play a crucial role in protection of bacteria from antimicrobials. The MacAB efflux pump, previously linked to efflux of macrolides in Escherichia coli and protection from oxidative stress in Salmonella enterica serovar Typhimurium, is not characterized in S. marcescens. We show the role of the MacAB efflux pump in S. marcescens protection from aminoglycoside antibiotics and polymyxins, modulation of bacterial motility, and biofilm formation, and we illustrate the essential role for this pump in bacterial survival during oxidative stress. Our findings make the MacAB efflux pump an attractive target for inhibition to gain control over S. marcescens infections.

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