Antimicrobial Activity of Gallium Protoporphyrin IX against Acinetobacter baumannii Strains Displaying Different Antibiotic Resistance Phenotypes

Abstract

A paucity of effective, currently available antibiotics and a lull in antibiotic development pose significant challenges for treatment of patients with multidrug-resistant (MDR) Acinetobacter baumannii infections. Thus, novel therapeutic strategies must be evaluated to meet the demands of treatment of these often life-threatening infections. Accordingly, we examined the antibiotic activity of gallium protoporphyrin IX (Ga-PPIX) against a collection of A. baumannii strains, including nonmilitary and military strains and strains representing different clonal lineages and isolates classified as susceptible or MDR. Susceptibility testing demonstrated that Ga-PPIX inhibits the growth of all tested strains when cultured in cation-adjusted Mueller-Hinton broth, with a MIC of 20 g/ml. This concentration significantly reduced bacterial viability, while 40 g/ml killed all cells of the A. baumannii ATCC 19606 T and ACICU MDR isolate after 24-h incubation. Recovery of ATCC 19606 T and ACICU strains from infected A549 human alveolar epithelial monolayers was also decreased when the medium was supplemented with Ga-PPIX, particularly at a 40-g/ml concentration. Similarly, the coinjection of bacteria with Ga-PPIX increased the survival of Galleria mellonella larvae infected with ATCC 19606 T or ACICU. Ga-PPIX was cytotoxic only when monolayers or larvae were exposed to concentrations 16-fold and 1,250-fold higher than those showing antibacterial activity, respectively. These results indicate that Ga-PPIX could be a viable therapeutic option for treatment of recalcitrant A. baumannii infections regardless of the resistance phenotype, clone lineage, time and site of isolation of strains causing these infections and their iron uptake phenotypes or the iron content of the media. Immediately following the advent of antibiotics as therapeutic agents, resistance to these drugs emerged among pathogenic bacteria. Drug-resistant bacterial strains are selected for immediately after initiation of antibiotic regimens, and as a consequence of continued selective pressure, very few treatment options now exist for infections caused by resistant strains of some pathogens (1, 2). The continuing battle with resistance has led to the initial emergence of pathogens displaying multidrug-resistant (MDR) phenotypes, which has since been followed by the emergence of extremely drug-resistant (XDR) or totally drug-resistant (TDR) strains, an outcome that has recreated the preantibiotic era (3, 4). This crisis has involved major Gram-positive and Gram-negative pathogens, including Enterococcus spp., Staphylococcus aureus, members of the family Enterobacteriaceae, Neisseria gonorrhoeae, Pseudomonas aeruginosa, and Acinetobacter spp. (4). The emergence of MDR strains of each of these microorganisms has contributed to increased morbidity and mortality among patients, leading to extended lengths of hospital stay and exorbitant health care financial burdens that often go unremitted. Compounding this continuum of resistance evolution and treatment failures is t...

Antimicrobial Activity of Gallium Compounds on ESKAPE Pathogens

“…On the other hand, the HemT and Isd heme-uptake systems, which are present in A. baumannii and S. aureus , respectively (Ascenzi et al, 2015 ), appear by themselves unable to confer GaPPIX susceptibility in RPMI-HS (but not in MHB and DMHB), probably because these two systems cannot efficiently extract HSA-bound GaPPIX for transport into the cell. These observations are in line with a previous report showing that the addition of 10% HS to MHB caused a 3-fold increase of the GaPPIX MIC for A. baumannii (Arivett et al, 2015 ). Intriguingly, the activity of GaPPIX against S. aureus and A. baumannii was independent of the iron content of the medium, given that: (i) it was similar in MHB and DMHB media, irrespective of their different iron content, and, (ii) amendment of MHB with an exceedingly high FeCl 3 concentration did not neutralize the antibacterial activity of GaPPIX (Figure 2 ), in agreement with previous reports (Stojiljkovic et al, 1999 ; Arivett et al, 2015 ).…”

“…These observations are in line with a previous report showing that the addition of 10% HS to MHB caused a 3-fold increase of the GaPPIX MIC for A. baumannii (Arivett et al, 2015 ). Intriguingly, the activity of GaPPIX against S. aureus and A. baumannii was independent of the iron content of the medium, given that: (i) it was similar in MHB and DMHB media, irrespective of their different iron content, and, (ii) amendment of MHB with an exceedingly high FeCl 3 concentration did not neutralize the antibacterial activity of GaPPIX (Figure 2 ), in agreement with previous reports (Stojiljkovic et al, 1999 ; Arivett et al, 2015 ). Interestingly, the addition of hemin partially reversed the antibacterial activity of GaPPIX in MHB, albeit in one S. aureus and in all A. baumannii strains tested not even a molar excess of hemin completely abrogated the effect of GaPPIX (Figure 2 ).…”

“…A bactericidal activity has previously been reported for GaN and GaPPIX on P. aeruginosa and A. baumannii , respectively. However, previous killing assays were conducted in rich laboratory media, which do not mimic the condition encountered in vivo by infecting pathogens (Kaneko et al, 2007 ; Arivett et al, 2015 ). For this reason, we devised to assess the bactericidal activity of the three Ga(III) compounds in RPMI-HS, i.e., under conditions that trend to mimic biological fluids.…”

Acinetobacter baumannii Biofilm Formation in Human Serum and Disruption by Gallium

“…Given the importance of iron in host-pathogen interactions, drugs interfering with microbial iron metabolism represent viable candidates for the development of new antibacterials (21). In this work, we provide evidence that gallium, which behaves as an iron-mimetic metal (22), is endowed with potent antibiofilm properties in addition to its previously reported ability to inhibit A. baumannii planktonic growth (7,23). The antimicrobial activity of GaN is dependent on the growth medium as this activity is invariably abrogated in an iron-rich medium, such as MH broth (7,8,22).…”

Miltefosine Reduces the Cytolytic Activity and Virulence of Acinetobacter baumannii

“…The A. baumannii strains used in this work were routinely stored as Luria-Bertani (LB) broth glycerol stocks at Ϫ80°C (28) and are listed in Table 1. The A. baumannii ATCC 19606 T strain was used as the model organism throughout this study due to it being the A. baumannii type strain as well as its multidrug-resistant phenotype, as determined using the European Committee on Antimicrobial Susceptibility Testing (EUCAST) standards, which demonstrated this isolate's resistance to three antibiotic classes clinically used to treat A. baumannii: (i) penicillins, (ii) cephalosporins, and (iii) aminoglycosides (29). A. baumannii strains were subcultured from LB agar into Chelex 100-treated Trypticase soy broth dialysate (TSBD) and grown for 24 h at 37°C with shaking at 200 rpm (30).…”