Apramycin Overcomes the Inherent Lack of Antimicrobial Bactericidal Activity in Mycobacterium abscessus.

Antibiotic therapy of infections caused by the emerging pathogen Mycobacterium abscessus is challenging due to the organism's inherent resistance to clinically available antimicrobials. The low bactericidal potency of currently available treatment regimens is of concern and testifies to the poor therapeutic outcomes for pulmonary M. abscessus infections. Mechanistically, we demonstrate here that the acetyltransferase Eis2 is responsible for the lack of bactericidal activity of amikacin, the standard aminoglycoside used in combination treatment. In contrast, the aminoglycoside apramycin, with a distinct structure, is not modified by any of the pathogen's innate aminoglycoside resistance mechanisms and is not affected by the multidrug resistance regulator WhiB7. As a consequence, apramycin uniquely shows potent bactericidal activity against M. abscessus. This favorable feature of apramycin is reflected in a mouse model of pulmonary M. abscessus infection, which demonstrates superior activity, compared with amikacin. These findings encourage the development of apramycin for the treatment of M. abscessus infections and suggest that M. abscessus eradication in pulmonary disease may be within therapeutic reach.

Efficacy of Apramycin against Multidrug-Resistant Acinetobacter baumannii in the Murine Neutropenic Thigh Model.

Apramycin, an aminocyclitol aminoglycoside, was rapidly bactericidal against Acinetobacter baumannii In a neutropenic murine thigh infection model, treatment-associated A. baumannii CFU reductions of >4 log10 per thigh were observed for all exposures for which area under the curve (AUC)/MIC ratio was >50 and maximum concentration of drug in serum (Cmax)/MIC was ≈10 or higher. Based on these findings, we suggest that apramycin deserves further preclinical exploration as a repurposed therapeutic for multidrug-resistant Gram-negative pathogens, including A. baumannii.

In vivo efficacy of apramycin in murine infection models.

Apramycin is a unique aminoglycoside with a dissociation of antibacterial activity and ototoxicity. We assessed the antibacterial efficacy of apramycin in two murine models of infection, Mycobacterium tuberculosis aerosol infection and Staphylococcus aureus septicemia. In both infection models, the efficacy of apramycin was comparable to that of amikacin. These results suggest that apramycin has the potential to become a clinically useful agent against drug-resistant pathogens and support further development of this promising unique aminoglycoside.

Apramycin efficacy against carbapenem- and aminoglycoside-resistant Escherichia coli and Klebsiella pneumoniae in murine bloodstream infection models.

BACKGROUND: The aminoglycoside apramycin has been proposed as a drug candidate for the treatment of critical Gram-negative systemic infections. However, the potential of apramycin in the treatment of drug-resistant bloodstream infections (BSIs) has not yet been assessed. METHODS: The resistance gene annotations of 40 888 blood-culture isolates were analysed. In vitro profiling of apramycin comprised cell-free translation assays, broth microdilution, and frequency of resistance determination. The efficacy of apramycin was studied in a mouse peritonitis model for a total of nine Escherichia coli and Klebsiella pneumoniae isolates. RESULTS: Genotypic aminoglycoside resistance was identified in 87.8% of all 6973 carbapenem-resistant Enterobacterales blood-culture isolates, colistin resistance was shown in 46.4% and apramycin in 2.1%. Apramycin activity against methylated ribosomes was > 100-fold higher than that for other aminoglycosides. Frequencies of resistance were < 10-9 at 8 × minimum inhibitory concentration (MIC). Tentative epidemiological cut-offs (TECOFFs) were determined as 8 µg/mL for E. coli and 4 µg/mL for K. pneumoniae. A single dose of 5 to 13 mg/kg resulted in a 1-log colony-forming unit (CFU) reduction in the blood and peritoneum. Two doses of 80 mg/kg resulted in an exposure that resembles the AUC observed for a single 30 mg/kg dose in humans and led to complete eradication of carbapenem- and aminoglycoside-resistant bacteraemia. CONCLUSION: Encouraging coverage and potent in vivo efficacy against a selection of highly drug-resistant Enterobacterales isolates in the mouse peritonitis model warrants the conduct of clinical studies to validate apramycin as a drug candidate for the prophylaxis and treatment of BSI.

Antibacterial activity of apramycin at acidic pH warrants wide therapeutic window in the treatment of complicated urinary tract infections and acute pyelonephritis.

BACKGROUND: The clinical-stage drug candidate EBL-1003 (apramycin) represents a distinct new subclass of aminoglycoside antibiotics for the treatment of drug-resistant infections. It has demonstrated best-in-class coverage of resistant isolates, and preclinical efficacy in lung infection models. However, preclinical evidence for its utility in other disease indications has yet to be provided. Here we studied the therapeutic potential of EBL-1003 in the treatment of complicated urinary tract infection and acute pyelonephritis (cUTI/AP). METHODS: A combination of data-base mining, antimicrobial susceptibility testing, time-kill experiments, and four murine infection models was used in a comprehensive assessment of the microbiological coverage and efficacy of EBL-1003 against Gram-negative uropathogens. The pharmacokinetics and renal toxicology of EBL-1003 in rats was studied to assess the therapeutic window of EBL-1003 in the treatment of cUTI/AP. FINDINGS: EBL-1003 demonstrated broad-spectrum activity and rapid multi-log CFU reduction against a phenotypic variety of bacterial uropathogens including aminoglycoside-resistant clinical isolates. The basicity of amines in the apramycin molecule suggested a higher increase in positive charge at urinary pH when compared to gentamicin or amikacin, resulting in sustained drug uptake and bactericidal activity, and consequently in potent efficacy in mouse infection models. Renal pharmacokinetics, biomarkers for toxicity, and kidney histopathology in adult rats all indicated a significantly lower nephrotoxicity of EBL-1003 than of gentamicin. INTERPRETATION: This study provides preclinical proof-of-concept for the efficacy of EBL-1003 in cUTI/AP. Similar efficacy but lower nephrotoxicity of EBL-1003 in comparison to gentamicin may thus translate into a higher safety margin and a wider therapeutic window in the treatment of cUTI/API. FUNDING: A full list of funding bodies that contributed to this study can be found in the Acknowledgements section.

Chitosan propolis nanocomposite alone or in combination with apramycin: an alternative therapy for multidrug-resistant Salmonella Typhimurium in rabbits: in vitro and in vivo study.

Introduction. The emergence of multidrug-resistant Salmonella Typhimurium strains has increased the need for safe, alternative therapies from natural sources with antibacterial properties.Hypothesis/Gap Statement. There are no published data regarding the use of chitosan propolis nanocomposite (CPNP) either alone or in combination with antibiotics as antimicrobials against S. Typhimurium, especially in Egypt.Aim. This study evaluated the antibacterial activities of five antimicrobials [apramycin, propolis, chitosan nanoparticles (CNPs), chitosan propolis nanocomposite (CPNP) and CPNP +apramycin] against ten virulent and multidrug-resistant (MDR) S. Typhimurium field strains recovered from diarrheic rabbits through in vitro and in vivo study.Methodology. The expression levels of three virulence genes of S. Typhimurium strains were determined by quantitative reverse-transcription PCR (RT-qPCR) after exposure to sub-inhibitory concentrations of apramycin, propolis, CNPs, CPNP alone, and CPNP +apramycin. Additionally, 90 New Zealand rabbits were divided into control and experimentally S. Typhimurium-infected groups. The infected rabbits were orally administered saline solution (infected-untreated); 10 mg apramycin/kg (infected-apramycin-treated); 50 mg propolis/kg (infected-propolis-treated); 15 mg CPNP/kg (infected-CPNP-treated) and 15 mg CPNP +10 mg apramycin/kg (infected-CPNP +apramycin-treated) for 5 days.Results. The RT-qPCR analysis revealed different degrees of downregulation of all screened genes. Furthermore, the treatment of infected rabbits with CPNP or CPNP +apramycin significantly improved performance parameters, and total bacterial and Salmonella species counts, while also modulating both oxidative stress and altered liver and kidney parameters.Conclusion. This work demonstrates the use of CPNP alone or in combination with apramycin in the treatment of S. Typhimurium in rabbits.

Effects of antibiotic regimens on the fecal shedding patterns of pigs infected with salmonella typhimurium.

An experiment was conducted to determine (i) the effects of antibiotic regimens on the shedding patterns of pigs infected with Salmonella Typhimurium and (ii) whether antibiotic resistance increases the incidence of pathogen shedding. The experiment involved 48 50-day-old pigs challenged with Salmonella Typhimurium and receiving one of four antibiotic regimens including (i) intramuscular injection of ceftiofur sodium followed by inclusion of oxytetracycline in the feed; (ii) apramycin in the feed for 14 days followed by oxytetracycline; (iii) carbadox in the feed until pigs reached 35 kg followed by oxytetracycline; (iv) no antibiotics (control). Fecal samples were collected preinoculation, 2 and 4 days postinoculation (DPI) and at weekly and biweekly intervals thereafter to determine shedding patterns. Salmonella Typhimurium isolates from 2, 4, 7, 21, 42, and 70 DPI were analyzed for antibiotic resistance. A time effect (P < 0.05) was observed, indicating that the proportion of isolates resistant to at least one antibiotic varied over time. Overall resistance was determined to be 46% at 2 DPI and increased significantly (P < .05) thereafter. Treatment x time and antibiotic x time interactions were also observed (P < 0.05) as the percentage of isolates resistant to each test antibiotic increased over time. In no case did the development of antibiotic resistance result in an increased incidence of shedding of the original inoculate. The incidence of shedding was reduced in pigs receiving the apramycin-oxytetracycline treatment, when compared to control pigs; however, no differences were observed between antibiotic treatments.

Effects of antibiotic use in sows on resistance of E. coli and Salmonella enterica Typhimurium in their offspring.

To determine effects of exposure of parental animals to antibiotics on antibiotic resistance in bacteria of offspring, sows were either treated or not treated with oxytetracycline prior to farrowing and their pigs were challenged with Salmonella enterica Typhimurium and treated or not treated with oxytetracycline and apramycin. Fecal Escherichia coli were obtained from sows, and E. coli and salmonella were recovered from pigs. Antibiotic resistance patterns of isolates were determined using a minimum inhibitory concentration (MIC) analysis. Polymerase chain reaction (PCR) and electroporation were used to characterize the genetic basis for the resistance and to determine the location of resistance genes. Treatments had little effect on resistance of the salmonella challenge organism. The greatest resistance to apramycin occurred in E. coli from pigs treated with apramycin and whose sows had earlier exposure to oxytetracycline. Resistance to oxytetracycline was consistently high throughout the study in isolates from all pigs and sows; however, greater resistance was noted in pigs nursing sows that had previous exposure to that drug. The aac(3)-IV gene, responsible for apramycin resistance, was found in approximately 90% of apramycin-resistant isolates and its location was determined to be on plasmids. Several resistant E. coli bio-types were found to contain the resistance gene. These results indicate that resistance to apramycin and oxytetracycline in E. coli of pigs is affected by previous use of oxytetracycline in sows.

An evaluation of apramycin soluble powder for the treatment of naturally acquired Escherichia coli infections in broilers.

During 1983, a series of trials was carried out in Greece, Italy and Jordan, to test the efficacy of the aminocyclitol antibiotic apramycin as a treatment for naturally acquired Escherichia coli infections in broilers. The trials involved a total of 40,389 broilers, 27-39 days of age, in twelve houses at eight different production sites. Three levels of apramycin medication were evaluated: 125, 250 and 500 mg activity/litre drinking water, administered for five consecutive days. Treatment was only initiated after disease had been confirmed by laboratory examination of dead birds. Not all treatments were evaluated in every house. However, each house contained a group of unmedicated birds as controls, while the remaining birds were allocated to one or more apramycin treatment groups. There was a reduction in mortality and an improvement in the final weight and the economic feed conversion ratio in all three treatment groups. The data presented provide support for the use of apramycin sulphate administered in the drinking water for the treatment of E. coli infections in broilers.

[Isolation and identification of the antibiotics amicetin and nebramycin formed by a Streptomyces coeruleoaurantiacus 4009 culture]. / Vydelenie i identifikatsiia antibiotikov amitsetina i nebramitsina, obrazuemykh kul'turoi Streptomyces coeruleoaurantiacus 4009.

It was found that Str. coeruleoaurantiacus, strain 4009 produced antibiotics 4009-A and 4009-B belonging to different groups. Antibiotic 4009-A was identified as amicetin belonging to the group of pyrimidine bases and antibiotic 4009-B as the nebramycin complex belonging to the group of aminoglycosides. The identity of the antibiotics was confirmed by the physico-chemical constants, spectral and chromatographic characteristics and their chemotherapeutic activity.