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Development and characterisation of highly antibiotic resistant Bartonella bacilliformis mutants

View Article: PubMed Central - PubMed

ABSTRACT

The objective was to develop and characterise in vitro Bartonella bacilliformis antibiotic resistant mutants. Three B. bacilliformis strains were plated 35 or 40 times with azithromycin, chloramphenicol, ciprofloxacin or rifampicin discs. Resistance-stability was assessed performing 5 serial passages without antibiotic pressure. MICs were determined with/without Phe-Arg-β-Napthylamide and artesunate. Target alterations were screened in the 23S rRNA, rplD, rplV, gyrA, gyrB, parC, parE and rpoB genes. Chloramphenicol and ciprofloxacin resistance were the most difficult and easiest (>37.3 and 10.6 passages) to be selected, respectively. All mutants but one selected with chloramphenicol achieved high resistance levels. All rifampicin, one azithromycin and one ciprofloxacin mutants did not totally revert when cultured without antibiotic pressure. Azithromycin resistance was related to L4 substitutions Gln-66 → Lys or Gly-70 → Arg; L4 deletion Δ62–65 (Lys-Met-Tyr-Lys) or L22 insertion 83::Val-Ser-Glu-Ala-His-Val-Gly-Lys-Ser; in two chloramphenicol-resistant mutants the 23S rRNA mutation G2372A was detected. GyrA Ala-91 → Val and Asp-95 → Gly and GyrB Glu474 → Lys were detected in ciprofloxacin-resistant mutants. RpoB substitutions Gln-527 → Arg, His-540 → Tyr and Ser-545 → Phe plus Ser-588 → Tyr were detected in rifampicin-resistant mutants. In 5 mutants the effect of efflux pumps on resistance was observed. Antibiotic resistance was mainly related to target mutations and overexpression of efflux pumps, which might underlie microbiological failures during treatments.

No MeSH data available.


Related in: MedlinePlus

Colony morphology.The photograph shows B. bacilliformis presenting T1 colony morphology20. The colony is characterised by a small, translucent round morphology, with a regular edge and a small halo. The colonies present a “bubble” in the center of the colony. The morphology was unstable and disappeared after reculture.
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f1: Colony morphology.The photograph shows B. bacilliformis presenting T1 colony morphology20. The colony is characterised by a small, translucent round morphology, with a regular edge and a small halo. The colonies present a “bubble” in the center of the colony. The morphology was unstable and disappeared after reculture.

Mentions: The time required for bacterial lyophilised reactivation varied from 5 weeks (strains 57.19 and 57.20) to 9 weeks (strain 57.18). Interestingly, strain 57.18 showed an initially different morphology, coinciding with the previously described T1 morphology20, although this reverted in the next passage (Fig. 1).


Development and characterisation of highly antibiotic resistant Bartonella bacilliformis mutants
Colony morphology.The photograph shows B. bacilliformis presenting T1 colony morphology20. The colony is characterised by a small, translucent round morphology, with a regular edge and a small halo. The colonies present a “bubble” in the center of the colony. The morphology was unstable and disappeared after reculture.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC5035977&req=5

f1: Colony morphology.The photograph shows B. bacilliformis presenting T1 colony morphology20. The colony is characterised by a small, translucent round morphology, with a regular edge and a small halo. The colonies present a “bubble” in the center of the colony. The morphology was unstable and disappeared after reculture.
Mentions: The time required for bacterial lyophilised reactivation varied from 5 weeks (strains 57.19 and 57.20) to 9 weeks (strain 57.18). Interestingly, strain 57.18 showed an initially different morphology, coinciding with the previously described T1 morphology20, although this reverted in the next passage (Fig. 1).

View Article: PubMed Central - PubMed

ABSTRACT

The objective was to develop and characterise in vitro Bartonella bacilliformis antibiotic resistant mutants. Three B. bacilliformis strains were plated 35 or 40 times with azithromycin, chloramphenicol, ciprofloxacin or rifampicin discs. Resistance-stability was assessed performing 5 serial passages without antibiotic pressure. MICs were determined with/without Phe-Arg-β-Napthylamide and artesunate. Target alterations were screened in the 23S rRNA, rplD, rplV, gyrA, gyrB, parC, parE and rpoB genes. Chloramphenicol and ciprofloxacin resistance were the most difficult and easiest (>37.3 and 10.6 passages) to be selected, respectively. All mutants but one selected with chloramphenicol achieved high resistance levels. All rifampicin, one azithromycin and one ciprofloxacin mutants did not totally revert when cultured without antibiotic pressure. Azithromycin resistance was related to L4 substitutions Gln-66 → Lys or Gly-70 → Arg; L4 deletion Δ62–65 (Lys-Met-Tyr-Lys) or L22 insertion 83::Val-Ser-Glu-Ala-His-Val-Gly-Lys-Ser; in two chloramphenicol-resistant mutants the 23S rRNA mutation G2372A was detected. GyrA Ala-91 → Val and Asp-95 → Gly and GyrB Glu474 → Lys were detected in ciprofloxacin-resistant mutants. RpoB substitutions Gln-527 → Arg, His-540 → Tyr and Ser-545 → Phe plus Ser-588 → Tyr were detected in rifampicin-resistant mutants. In 5 mutants the effect of efflux pumps on resistance was observed. Antibiotic resistance was mainly related to target mutations and overexpression of efflux pumps, which might underlie microbiological failures during treatments.

No MeSH data available.


Related in: MedlinePlus