Development and characterisation of highly antibiotic resistant Bartonella bacilliformis mutants

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.

Carrion's Disease: the Sound of Silence

“…continues to be highly susceptible to antibiotics (334). Antimicrobial resistance levels have been established in a few clinical or collection isolates of B. bacilliformis (123,305,314,(335)(336)(337), and it should be taken into account that in vitro susceptibility does not consistently correlate with the in vivo outcomes for patients (330). In addition to the constitutive nalidixic acid resistance and related diminished fluoroquinolone susceptibility described previously (305,338), in a recent report 26% of isolates presented resistance to ciprofloxacin (314).…”

“…These flagellated nonmotile variants can be bound to erythrocytes, promoting surface contact interaction. The optimal binding of bacteria to the surface of erythrocytes requires B. bacilliformis energy, most likely proton motive force-dependent motility (123,140), with the whole in vitro process taking approximately 6 h (161).…”

“…To our knowledge only four in vitro studies have been developed with B. bacilliformis, selecting in vitro resistance to different antimicrobial agents, including chloramphenicol, ciprofloxacin, coumermycin A1, erythromycin, and rifampin (123,330,341,342). In these studies, ciprofloxacin resistance ranked among the most easily selected, while chloramphenicol resistance ranked among the most difficult to obtain (123). Additionally, 3 reports have also analyzed the presence of antibiotic resistance-related mutations in clinical isolates of B. bacilliformis (337,343,344), but 1 of these studies did not include the MICs of the antibiotic (344) ( Table 6).…”

Design of Multiplex Lateral Flow Tests: A Case Study for Simultaneous Detection of Three Antibiotics

“…However, improper use of antibiotics has provoked massive resistance to them of many microorganisms. Such resistance to tetracycline is described for strains of Pseudomonas aeruginosa, Proteus, Staphylococcus aureus (MRSA) and Streptococcus pneumoniae [14], to chloramphenicol -for Bartonella and Staphylococcus [15], to lincomycin -for Kocuria kristinae, Sphingomonas paucimobilis, Pantoea, Staphylococcus vitulinus, Clostridium, etc. [16,17].…”

Subtractive Genomic Analysis for Identification of Novel Drug Targets and Vaccine Candidates against Bartonella bacilliformis subsp. Ver097

“…Though studies suggested that most of Bartonella species including B. bacilliformis are highly susceptible to mostof antibiotics, such as beta-lactams, aminoglycosides, chloramphenicol, rifampin, macrolides, tetracyclines, cotrimoxazole, fluoroquinolones[12] and gentamicin[1], but there is a potential risk of the spontaneous selection of successful antibiotic resistant B. bacilliformis clone in a clinical setting. For examples, few studies developed in vitro has showed the ability of B. bacilliformis to acquire chromosomal mutations leading to high levels of antibiotic resistance [13,14]. Lack of comprehensive analysis of B. bacilliformis genomic structure is an obstacle in developing species-specific new therapeutic compounds and vaccine candidates [5].…”