Successful Treatment of Postneurosurgical Ventriculitis Caused by Extensively Drug-resistant Acinetobacter baumannii in a Child: Case Report

Introduction: The incidence of postneurosurgical Acinetobacter baumannii ventriculitis/meningitis, primarily due to drug-resistant strains, has increased considerably in recent years. However, limited therapeutic options are available because most antibiotics poorly penetrate the blood-brain barrier, especially in pediatric patients. Case Presentation: A five-year-old boy developed ventriculitis due to extensively drug-resistant A. baumannii (XDRAB) after bilateral frontal external ventricular drainage for spontaneous intraventricular hemorrhage. The boy was safely and successfully treated with intraventricular (IVT)/intrathecal (ITH) polymyxin B together with intravenous tigecycline plus cefoperazone/sulbactam. Conclusions: In the present case, postneurosurgical XDRAB ventriculitis was closely associated with intraventricular hemorrhage and the placement of external ventricular drainage. IVT/ITH polymyxin B combined with intravenous tigecycline and cefoperazone sulbactam could be a therapeutic option against XDRAB ventriculitis in children.

Prednisolone and dexamethasone are systemically absorbed after topical application of ophthalmic suspensions in healthy dogs

OBJECTIVE To quantify plasma concentrations of prednisolone and dexamethasone (peripheral and jugular) and cortisol following topical ophthalmic application of 1% prednisolone acetate and 0.1% dexamethasone to healthy adult dogs. ANIMALS 12 purpose-bred Beagles. PROCEDURES Dogs received 1 drop of 1% prednisolone acetate (n = 6) or neomycin polymyxin B dexamethasone (ie, 0.1% dexamethasone; 6) ophthalmic suspension in both eyes every 6 hours for 14 days. Blood samples (peripheral and jugular) were collected on days 0, 1, 7, and 14 and analyzed for plasma prednisolone and dexamethasone concentrations. Plasma cortisol concentrations were measured at the beginning of the study and following topical drug administration. RESULTS Both drugs demonstrated systemic absorption. Prednisolone was detected on days 1, 7, and 14 (median plasma concentration, 24.80 ng/mL; range, 6.20 to 74.00 ng/mL), and dexamethasone was detected on days 1, 7, and 14 (2.30 ng/mL; 0 to 17.70 ng/mL). Neither prednisolone nor dexamethasone were detected in plasma samples on day 0 (baseline). Sampling from the jugular vein resulted in higher plasma drug concentrations than from a peripheral vein when samples from each day were combined. Plasma cortisol concentrations were significantly lower than baseline following 14 days of treatment with topical prednisolone acetate and dexamethasone. CLINICAL RELEVANCE Prednisolone and dexamethasone are detected in the plasma of healthy dogs following topical ophthalmic administration 4 times/d with prednisolone concentrations being close to a physiologic dose of orally administered prednisolone. Additional research is needed to evaluate the systemic absorption of these medications in dogs with ocular inflammation.

A method for increasing electroporation competency of Gram-negative clinical isolates by Polymyxin B nonapeptide

The study of clinically relevant bacterial pathogens relies on molecular and genetic approaches. However, the generally low transformation frequency among natural isolates poses technical hurdles to widely applying common methods in molecular biology, including transformation of large constructs, chromosomal genetic manipulation, and dense mutant library construction. Here we demonstrate that culturing clinical isolates in the presence of polymyxin B nonapeptide (PMBN) improves their transformation frequency via electroporation by up to 100-fold in a dose-dependent and reversible manner. The effect was observed for PMBN-binding uropathogenic Escherichia coli (UPEC) and Salmonella enterica strains but not naturally polymyxin resistant Proteus mirabilis. Using our PMBN electroporation method we show efficient delivery of large plasmid constructs into UPEC, which otherwise failed using a conventional electroporation protocol. Moreover, we show a 5-fold increase in the yield of engineered mutant colonies obtained in S. enterica with the widely used lambda-Red recombineering method, when cells are cultured in the presence of PMBN. Lastly, we demonstrate that PMBN treatment can enhance the delivery of DNA-transposase complexes into UPEC and increase transposon mutant yield by 8-fold when constructing Transposon Insertion Sequencing (TIS) libraries. Therefore, PMBN can be used as a powerful electropermeabilisation adjuvant to aid the delivery of DNA and DNA-protein complexes into clinically important bacteria.

Pseudomonas aeruginosa Oligoribonuclease Controls Susceptibility to Polymyxin B by Regulating Pel Exopolysaccharide Production

Polymyxins are considered as the last resort antibiotics to treat infections caused by multidrug-resistant Gram negative pathogens. Pseudomonas aeruginosa is an opportunistic pathogen that causes various infections in humans. Proteins involved in lipopolysaccharide modification and maintaining inner and outer membrane integrities have been found to contribute to the bacterial resistance to polymyxins. Oligoribonuclease (Orn) is an exonuclease that regulates the homeostasis of intracellular (3'-5')-cyclic dimeric guanosine monophosphate (c-di-GMP), thereby regulating the production of extracellular polysaccharide in P. aeruginosa . Previously, we demonstrated that Orn affects the bacterial resistance to fluoroquinolone, β-lactam and aminoglycoside antibiotics. In this study, we found that mutation of orn increased the bacterial survival following polymyxin B treatment in a wild type P. aeruginosa strain PA14. Overexpression of c-di-GMP degradation enzymes in the orn mutant reduced the bacterial survival. By using a fluorescence labeled polymyxin B, we found that mutation of orn increased the bacterial surface bound polymyxin B. Deletion of the Pel synthesis genes or treatment with a Pel hydrolase reduced the surface bound polymyxin B and bacterial survival. We further demonstrated that Pel binds to extracellular DNA (eDNA), which traps polymyxin B and thus protects the bacterial cells. Collectively, our results revealed a novel defense mechanism against polymyxin in P. aeruginosa .

Atypical and dual biotypes variant of virulent SA-NAG-Vibrio cholerae: an evidence of emerging/evolving patho-significant strain in municipal domestic water sources

Introduction and purpose The recent cholera spread, new cases, and fatality continue to arouse concern in public health systems; however, interventions on control is at its peak yet statistics show continuous report. This study characterized atypical and patho-significant environmental Vibrio cholerae retrieved from ground/surface/domestic water in rural-urban-sub-urban locations of Amathole District municipality and Chris Hani District municipality, Eastern Cape Province, South Africa. Methods Domestic/surface water was sampled and 759 presumptive V. cholerae isolates were retrieved using standard microbiological methods. Virulence phenotypic test: toxin co-regulated pili (tcp), choleragen red, protease production, lecithinase production, and lipase test were conducted. Serotyping using polyvalent antisera (Bengal and Ogawa/Inaba/Hikojima) and molecular typing: 16SrRNA, OmpW, serogroup (Vc-O1/O139), biotype (tcpAClas/El Tor, HlyAClas/El Tor, rstRClas/El Tor, RS1, rtxA, rtxC), and virulence (ctxA, ctxB, zot, ace, cep, prt, toxR, hlyA) genes were targeted. Result Result of 16SrRNA typing confirmed 508 (66.9%) while OmpW detected/confirmed 61 (12.01%) V. cholerae strains. Phenotypic-biotyping scheme showed positive test to polymyxin B (68.9%), Voges proskauer (6.6%), and Bengal serology (11.5%). Whereas Vc-O1/O139 was negative, yet two of the isolates harbored the cholera toxin with a gene-type ctxB and hlyAClas: 2/61, revealing atypical/unusual/dual biotype phenotypic/genotypic features. Other potential atypical genotypes detected include rstR: 7/61, Cep: 15/61, ace: 20/61, hlyAElTor: 53/61, rtxA: 30/61, rtxC: 11/61, and prtV: 15/61 respectively. Conclusion Although additional patho-significant/virulent genotypes associated with epidemic/sporadic cholera cases were detected, an advanced, bioinformatics, and post-molecular evaluation is necessary. Such stride possesses potential to adequately minimize future cholera cases associated with dynamic/atypical environmental V. cholerae strains.

Molecular Engineering of Polymyxin B for Imaging and Treatment of Bacterial Infections

The emergence of multi-drug resistant bacteria and the lack of novel antibiotics to combat them have led to the revival of polymyxin B, a previously abandoned antibiotic due to its potential nephrotoxicity and neurotoxicity. To facilitate its widely clinical applications, increasing effort has been devoted to molecularly engineer polymyxin B for the targeted imaging and effective treatment of bacterial infections. Herein, the molecular engineering strategies will be summarized in this mini review, with selected recent advances for illustration. Perspective of the challenges and trends in this exciting and eagerly anticipated research area will also be provided in the end. We hope this mini review will inspire researchers from diverse fields to bring forward the next wave of exploiting molecular engineering approaches to propel the “old” polymyxin B to “new” clinical significance in combating bacterial infections.

Pharmacokinetic/Pharmacodynamic Based Breakpoints of Polymyxin B for Bloodstream Infections Caused by Multidrug-Resistant Gram-Negative Pathogens

The latest PK/PD findings have demonstrated negligible efficacy of intravenous polymyxins against pulmonary infections. We investigated pharmacokinetic/pharmacodynamic (PK/PD)-based breakpoints of polymyxin B for bloodstream infections and the rationality of the recent withdrawal of polymyxin susceptibility breakpoints by the CLSI. Polymyxin B pharmacokinetic data were obtained from a phase I clinical trial in healthy Chinese subjects and population pharmacokinetic parameters were employed to determine the exposure of polymyxin B at steady state. MICs of 1,431 recent clinical isolates of Pseudomonas aeruginosa, Acinetobacter baumannii and Klebsiella pneumoniae collected from across China were determined. Monte-Carlo simulations were performed for various dosing regimens (0.42–1.5 mg/kg/12 h via 1 or 2-h infusion). The probability of target attainment, PK/PD breakpoints and cumulative fraction of response were determined for each bacterial species. MIC90 of polymyxin B was 1 mg/L for P. aeruginosa and 0.5 mg/L for A. baumannii and K. pneumoniae. With the recommended polymyxin B dose of 1.5–2.5 mg/kg/day, the PK/PD susceptible breakpoints for P. aeruginosa, A. baumannii and K. pneumoniae were 2, 1 and 1 mg/L respectively for bloodstream infection. For Chinese patients, polymyxin B dosing regimens of 0.75–1.5 mg/kg/12 h for P. aeruginosa and 1–1.5 mg/kg/12 h for A. baumannii and K. pneumoniae were appropriate. Breakpoint determination should consider the antimicrobial PK/PD at infection site and delivery route. The recent withdrawal of polymyxin susceptible breakpoint by CLSI primarily based on poor efficacy against lung infections needs to be reconsidered for bloodstream infections.

Identification and antimicrobial susceptibility of microbial agents of otitis externa in dogs

In this study, a total of 277 unmedicated dogs with otitis externa were used. Overall, 413 agents were isolated from 277 ear swab samples; 52.7% of the cases were mono-infections (146 cases), and 37.1% of the cases were poly-infections (103 cases). In 10.1% (28) of the cases, neither bacteria nor yeasts were isolated. Coagulase-positive Staphylococcus spp. were the most frequently isolated bacteria and were found in 90 (21.8%) of the samples. Fifty-eight samples, (14%) were positive for Staphylococcus aureus, 51 (12.3%) for Pseudomonas aeruginosa, 27 (6.5%) for Proteus mirabilis, 27 (6.5%) for Malassezia pachydermatis, 21 (5%) for Corynebacterium spp., 21 (5%) for β-haemolytic Streptococcus spp., 15 (3.6%) for Staphylococcus pseudointermedius, 12 (2.9%) for Proteus spp., 12 (2.9%) for Escherichia coli, 9 (2.1%) for Acinetobacter calcoaceticus, 7 (1.6%) for Trichophyton mentagrophytes, 5 (1.2%) for Staphylococcus auricularis, and 46 (11.1%) for different bacteria and yeasts. A total of 14 different bacteria and yeasts were isolated and identified. Kirby-Bauer antibiotic susceptibility testing was carried out for 10 different antibiotics. The bacterial isolates were found to be resistant to amoxicillin-clavulanic acid (45%), gentamycin (28%), ampicillin/cloxacillin (69%), tobramycin (28%), amikacin (23%), enrofloxacin (47%), chloramphenicol (58%), doxycycline (65%), lincomycin/spectinomycin (58%) and polymyxin B (62%). In conclusion, it is important to test the antimicrobial sensitivity of aetiological agents of otitis externa before treatment so as to prevent the development of antibiotic resistance in bacteria and yeasts.

Bacterial Mixology: Combining Pharmacodynamic Models to Predict In Vitro Competition of MCR-1-Harboring E. coli

The emergence of mobile colistin resistance (mcr)-mediated polymyxin resistance has resulted in a significant detriment to the utility of the polymyxins in the clinical setting. Though the risk for horizontal transfer of an mcr-containing plasmid is a major component of the transmissibility, selection of polymyxin resistant subpopulations is still a major risk factor for developing polymyxin-resistant infections. Using static time-kills over 24 h (h), we performed competition studies by mixing known inocula of isogenic Escherichia coli strains (wildtype [WT] and mcr-1-harboring) and treating with a concentration array of polymyxin B. These results were then compared to a priori predictions of bacterial-killing effects by polymyxin B on a mixed population of E. coli cells using a previously published mechanism-based model. The data showed that both selective pressure between WT and mcr-1-harboring strains as well as underlying polymyxin B heteroresistance within each of the two strains contributed to bacterial regrowth despite treatment with high concentration polymyxin B. Moreover, the simulations showed that when mcr-1-harboring cells were 1% or 10% of the total population, regrowth by 24 h was still observed in ≥50% of the simulated subjects for both a 106 and 108 inoculum. These results indicate that at lower inoculums with a low proportion of mcr-1-harboring cells, selective pressure from a pharmacokinetic-optimized regimen of polymyxin B still results in regrowth and selection of polymyxin-resistant cells.

Unveiling the Membrane and Cell Wall Action of Antimicrobial Cyclic Lipopeptides: Modulation of the Spectrum of Activity

Antibiotic resistance is a major public health challenge, and Gram-negative multidrug-resistant bacteria are particularly dangerous. The threat of running out of active molecules is accelerated by the extensive use of antibiotics in the context of the COVID-19 pandemic, and new antibiotics are urgently needed. Colistin and polymyxin B are natural antibiotics considered as last resort drugs for multi-resistant infections, but their use is limited because of neuro- and nephrotoxicity. We previously reported a series of synthetic analogues inspired in natural polymyxins with a flexible scaffold that allows multiple modifications to improve activity and reduce toxicity. In this work, we focus on modifications in the hydrophobic domains, describing analogues that broaden or narrow the spectrum of activity including both Gram-positive and Gram-negative bacteria, with MICs in the low µM range and low hemolytic activity. Using biophysical methods, we explore the interaction of the new molecules with model membranes that mimic the bacterial inner and outer membranes, finding a selective effect on anionic membranes and a mechanism of action based on the alteration of membrane function. Transmission electron microscopy observation confirms that polymyxin analogues kill microbial cells primarily by damaging membrane integrity. Redistribution of the hydrophobicity within the polymyxin molecule seems a plausible approach for the design and development of safer and more selective antibiotics.