Emergence Of Bacterial Antibiotic Resistance Research Articles

MiniBioReactor Array (MBRA) in vitro gut model: a reliable system to study microbiota-dependent response to antibiotic treatment.

BackgroundAntimicrobial drugs are mostly studied for their impact on emergence of bacterial antibiotic resistance, but their impact on the gut microbiota is also of tremendous interest. In vitro gut models are important tools to study such complex drug–microbiota interactions in humans.MethodsThe MiniBioReactor Array (MBRA) in vitro microbiota system; a single-stage continuous flow culture model, hosted in an anaerobic chamber; was used to evaluate the impact of three concentrations of a third-generation cephalosporin (ceftriaxone) on faecal microbiota from two healthy donors (treatment versus control: three replicates per condition). We conducted 16S microbiome profiling and analysed microbial richness, diversity and taxonomic changes. β-Lactamase activities were evaluated and correlated with the effects observed in the MBRA in vitro system.ResultsThe MBRA preserved each donor’s specificities, and differences between the donors were maintained through time. Before treatment, all faecal cultures belonging to the same donor were comparable in composition, richness, and diversity. Treatment with ceftriaxone was associated with a decrease in α-diversity, and an increase in β-diversity index, in a concentration-dependent manner. The maximum effect on diversity was observed after 72 h of treatment. Importantly, one donor had a stronger microbiota β-lactamase activity that was associated with a reduced impact of ceftriaxone on microbiota composition.ConclusionsMBRA can reliably mimic the intestinal microbiota and its modifications under antibiotic selective pressure. The impact of the treatment was donor- and concentration-dependent. We hypothesize these results could be explained, at least in part, by the differences in β-lactamase activity of the microbiota itself. Our results support the relevance and promise of the MBRA system to study drug–microbiota interactions.

Formulación y caracterización de liposomas encapsulados de ciprofloxacina: actividad antimicrobiana in vitro frente a Salmonella typhi multirresistente

Since decades antibiotics are used against bacterial infections, however for past few years world is fronting serious challenge due to emergence of bacterial antibiotic resistance for which new alternatives of cure are being sought. Liposomes are universally used nano-carriers designed to carry drug safely to action site. Emergence of antibiotic resistance in Salmonella typhi has left us with very few choices of cure. In current study liposomes were designed to carry ciprofloxacin and In vitro efficacy analyzed against multi drug resistant Salmonella typhi. Ciprofloxacin loaded liposomes prepared with Bx-DSPE-PEG(2000) (0.1%), total 100mg lipids, Phosphatidylcholine (PC) and Cholesterol (CH) at ratio of 5:3 had 93.95% encapsulation efficiency. Efficacy was observed through MIC and MBC by time kill assay. The encapsulation efficiency of cipro-loaded liposomes increased from 30% to 93.95% with increase cholesterol content. They showed slow to fast reléase, stability reduced from 93.95% to 71.9% over period of 9 months. Polydispersity index  was 0.7, particle size 95.27nm with -20.58mV zeta potential. As compared to drug alone MBC and MIC achieved for ciprofloxacin encapsulated liposomes was at lower drug concentrations i.e. 15 µg/ml, and 10 µg/ml. Conclusively, ciprofloxacin encapsulated liposomes prooved an effective in vitro drug delivery method against MDR Salmonella typhi.

Stereoselective metabolism of chloramphenicol by bacteria isolated from wastewater, and the importance of stereochemistry in environmental risk assessments for antibiotics

Wastewater treatment plants have been highlighted as a potential hotspot for the development and spread of antibiotic resistance. Although antibiotic resistant bacteria in wastewater present a public health threat, it is also possible that these bacteria play an important role in the bioremediation through the metabolism of antibiotics before they reach the wider environment. Here we address this possibility with a particular emphasis on stereochemistry using a combination of microbiology and analytical chemistry tools including the use of supercritical-fluid chromatography coupled with mass spectrometry for chiral analysis and high-resolution mass spectrometry to investigate metabolites. Due to the complexities around chiral analysis the antibiotic chloramphenicol was used as a proof of concept to demonstrate stereoselective metabolism due to its relatively simple chemical structure and availability over the counter in the U.K. The results presented here demonstrate the chloramphenicol can be stereoselectively transformed by the chloramphenicol acetyltransferase enzyme with the orientation around the first stereocentre being key for this process, meaning that accumulation of two isomers may occur within the environment with potential impacts on ecotoxicity and emergence of bacterial antibiotic resistance within the environment.

In Vitro Antibacterial Effect of the Methanolic Extract of the Korean Soybean Fermented Product Doenjang against Staphylococcus aureus.

Simple SummaryThe emergence of bacterial antibiotic resistance is a negative phenomenon occurring worldwide in both animals and humans. The EU banned the use of antibiotic growth promoters in animal production, as their administration to livestock is assumed to substantially contribute to the spread of bacterial resistance. Therefore, alternatives to antibiotic substances are needed to maintain the quality and quantity of animal products. Certain plant materials, such as fermented soybean products, can serve as a source of substances with potential to decrease the growth of resistant bacteria, such as Staphylococcus aureus. Fermented soybean products, including doenjang, are known to contain natural phytoestrogens called isoflavones, which are especially interesting due to their antimicrobial activity; these products can also be utilized in animal feed. Thus, the antibacterial activity of the methanolic extract of the Korean soybean fermented product doenjang was evaluated using standardized microbiological methods against nine strains of resistant and sensitive S. aureus, including those occurring in animals. The extract has been shown to be active at a concentration range of 2048–4096 µg/mL against all tested S. aureus strains and can therefore serve as a promising alternative to antibiotics in animal feed after additional testing in the laboratory and on living animals.Ultra-high performance liquid chromatography/mass spectrometry showed soyasaponin I and the isoflavones daidzein, genistein, and glycitein to be the main components of the methanolic extract of the Korean soybean fermented product doenjang, which is known to be a rich source of naturally occurring bioactive substances, at average contents of 515.40, 236.30, 131.23, and 29.00 ng/mg, respectively. The antimicrobial activity of the methanolic extract of doenjang against nine Staphylococcus aureus strains was determined in vitro by the broth microdilution method to investigate its potential to serve as an alternative antibacterial compound. The results suggest that the extract is an effective antistaphylococcal agent at concentrations of 2048–4096 µg/mL. Moreover, the tested extract also showed the ability to inhibit the growth of both methicillin-sensitive and methicillin-resistant animal and clinical S. aureus isolates. The growth kinetics of the chosen strains of S. aureus at the minimum inhibitory concentration of the methanolic extract of doenjang support the idea that the tested extract acts as an antibacterial compound. To the best of our knowledge, this is the first report on the antistaphylococcal action of the methanolic extract of doenjang thus, additional studies including in vivo testing are necessary to confirm this hypothesis.

Emerging resistance to carbapenem among Gram-negative bacteria in a tertiary care hospital

The emergence of bacterial antibiotic resistance is a cardinal concern in the health care system. The spread of resistance in Enterobacteriaceae and non-fermenters to the currently available drugs make the treatment of serious nosocomial infections troublesome. The purpose of the study is to find out the carbapenem resistance among Gram-negative bacilli in a tertiary care hospital. Antibiotic susceptibility pattern of 1913 aerobic Gram-negative bacilli isolated from clinical samples was made for a period of 6 months. All the isolates were tested for susceptibility to antibiotics by the Kirby-Bauer disc diffusion technique according to CLSI guidelines. Carbapenemase production was confirmed by the Modified Hodge Test (MHT). Minimum Inhibitory Concentration (MIC) by Epsilometer (E) test was performed (for Imipenem and Meropenem) for carbapenem-resistant strains. A total of 1731 clinical samples, 1913 Gram-negative bacilli were isolated. 1476 (77.1%) were Enterobacteriaceae and 433 (22.6%) were non-fermenters. 54 were carbapenemase-producing Gram-negative bacilli. Meropenem E test was done for carbapenemase-producing Gram-negative bacilli. The minimum inhibitory concentration for Meropenem ranged from 0.002μg/ml to 32μg/ml. To overcome the problem of emerging resistance, combined interaction and cooperation of microbiologists, clinicians and the infection control team is needed.

Short-course therapy for diarrhea-predominant irritable bowel syndrome: understanding the mechanism, impact on gut microbiota, and safety and tolerability of rifaximin.

Irritable bowel syndrome (IBS) is a common gastrointestinal (GI) disorder characterized by abdominal pain that occurs with defecation or alterations in bowel habits. Further classification is based on the predominant bowel habit: constipation-predominant IBS, diarrhea-predominant IBS (IBS-D), or mixed IBS. The pathogenesis of IBS is unclear and is considered multifactorial in nature. GI dysbiosis, thought to play a role in IBS pathophysiology, has been observed in patients with IBS. Alterations in the gut microbiota are observed in patients with small intestinal bacterial overgrowth, and overgrowth may occur in a subset of patients with IBS. The management of IBS includes therapies targeting the putative factors involved in the pathogenesis of the condition. However, many of these interventions (eg, eluxadoline and alosetron) require long-term, daily administration and have important safety considerations. Agents thought to modulate the gut microbiota (eg, antibiotics and probiotics) have shown potential benefits in clinical studies. However, conventional antibiotics (eg, neomycin) are associated with several adverse events and/or the risk of bacterial antibiotic resistance, and probiotics lack uniformity in composition and consistency of response in patients. Rifaximin, a nonsystemic antibiotic administered as a 2-week course of therapy, has been shown to be safe and efficacious for the treatment of IBS-D. Rifaximin exhibits a favorable benefit-to-harm ratio when compared with daily therapies for IBS-D (eg, alosetron and tricyclic antidepressants), and rifaximin was not associated with the emergence of bacterial antibiotic resistance. Thus, short-course therapy with rifaximin is an appropriate treatment option for IBS-D.

Antagonistic Coevolution Drives Whack-a-Mole Sensitivity in Gene Regulatory Networks.

Robustness, defined as tolerance to perturbations such as mutations and environmental fluctuations, is pervasive in biological systems. However, robustness often coexists with its counterpart, evolvability—the ability of perturbations to generate new phenotypes. Previous models of gene regulatory network evolution have shown that robustness evolves under stabilizing selection, but it is unclear how robustness and evolvability will emerge in common coevolutionary scenarios. We consider a two-species model of coevolution involving one host and one parasite population. By using two interacting species, key model parameters that determine the fitness landscapes become emergent properties of the model, avoiding the need to impose these parameters externally. In our study, parasites are modeled on species such as cuckoos where mimicry of the host phenotype confers high fitness to the parasite but lower fitness to the host. Here, frequent phenotype changes are favored as each population continually adapts to the other population. Sensitivity evolves at the network level such that point mutations can induce large phenotype changes. Crucially, the sensitive points of the network are broadly distributed throughout the network and continually relocate. Each time sensitive points in the network are mutated, new ones appear to take their place. We have therefore named this phenomenon “whack-a-mole” sensitivity, after a popular fun park game. We predict that this type of sensitivity will evolve under conditions of strong directional selection, an observation that helps interpret existing experimental evidence, for example, during the emergence of bacterial antibiotic resistance.

Steering Evolution with Sequential Therapy to Prevent the Emergence of Bacterial Antibiotic Resistance.

The increasing rate of antibiotic resistance and slowing discovery of novel antibiotic treatments presents a growing threat to public health. Here, we consider a simple model of evolution in asexually reproducing populations which considers adaptation as a biased random walk on a fitness landscape. This model associates the global properties of the fitness landscape with the algebraic properties of a Markov chain transition matrix and allows us to derive general results on the non-commutativity and irreversibility of natural selection as well as antibiotic cycling strategies. Using this formalism, we analyze 15 empirical fitness landscapes of E. coli under selection by different β-lactam antibiotics and demonstrate that the emergence of resistance to a given antibiotic can be either hindered or promoted by different sequences of drug application. Specifically, we demonstrate that the majority, approximately 70%, of sequential drug treatments with 2–4 drugs promote resistance to the final antibiotic. Further, we derive optimal drug application sequences with which we can probabilistically ‘steer’ the population through genotype space to avoid the emergence of resistance. This suggests a new strategy in the war against antibiotic–resistant organisms: drug sequencing to shepherd evolution through genotype space to states from which resistance cannot emerge and by which to maximize the chance of successful therapy.

Bacterial antibiotic resistance studies using in vitro dynamic models: Population analysis vs. susceptibility testing as endpoints of mutant enrichment

Emergence of bacterial antibiotic resistance is usually characterised either by population analysis or susceptibility testing. To compare these endpoints in their ability to demonstrate clear relationships with the ratio of 24-h area under the concentration–time curve (AUC24) to the minimum inhibitory concentration (MIC), enrichment of ciprofloxacin-resistant mutants of four clinical isolates of Pseudomonas aeruginosa was studied in an in vitro dynamic model that simulates mono-exponential pharmacokinetics of ciprofloxacin over a wide range of the AUC24/MIC ratios. Each organism was exposed to twice-daily ciprofloxacin for 3 days. Amplification of resistant mutants was monitored by plating on media with 2×, 4×, 8× and 16× MIC of ciprofloxacin. Population analysis data were expressed by the area under the bacterial mutant concentration–time curve (AUBCM). Changes in P. aeruginosa susceptibility were examined by daily MIC determinations. To account for the different susceptibilities of P. aeruginosa strains, post-exposure MICs (MICfinal) were related to the MICs determined with the starting inoculum (MICinitial). For each organism, AUC24/MIC relationships both with AUBCM and MICfinal/MICinitial were bell-shaped, but the latter were more strain-specific than the former. Using combined data on all four isolates, AUBCM showed a better correlation than MICfinal/MICinitial (r2=0.75 vs. r2=0.53). The shift of MICfinal/MICinitial relative to AUBCM vs. AUC24/MIC curves resulted in a weak correlation between AUBCM and MICfinal/MICinitial (r2=0.41). These data suggest that population analysis is preferable to susceptibility testing in bacterial resistance studies and that these endpoints should not be considered interchangeable.

Acceleration of Emergence of Bacterial Antibiotic Resistance in Connected Microenvironments

The emergence of bacterial antibiotic resistance is a growing problem, yet the variables that influence the rate of emergence of resistance are not well understood. In a microfluidic device designed to mimic naturally occurring bacterial niches, resistance of Escherichia coli to the antibiotic ciprofloxacin developed within 10 hours. Resistance emerged with as few as 100 bacteria in the initial inoculation. Whole-genome sequencing of the resistant organisms revealed that four functional single-nucleotide polymorphisms attained fixation. Knowledge about the rapid emergence of antibiotic resistance in the heterogeneous conditions within the mammalian body may be helpful in understanding the emergence of drug resistance during cancer chemotherapy.

Pattern and antibiogram of urinary tract infection at the University of Port Harcourt Teaching Hospital

Objective To explore the prevalence, pathogenicity, and antibiotic susceptibility patterns of urinary tract infections at the University of Port Harcourt Teaching Hospital. Methods Samples from 400 patients with a presumptive diagnosis of urinary tract infection including 250 non pregnant females and 150 males were used for this study. They were distributed into two groups: children aged 2 to 17 (Group A) and adults aged 18 to 75 (Group B). The standard wire loop and agar diffusion technique were employed for culture and susceptibility testing, respectively. Data obtained were analysed using SPSS, version 14. Results 30.0% of Group A and 41.0% of Group B had significant bacteriuria with 66.7% and 79.3% as females, respectively. The commonest isolates cultured were Escherichia coli (32.8%), Staphylococcus aureus (17.2%), and Klebsiella spp. (16.4%). About 76.6% of isolates were sensitive to the fluorinated quinolones, 31.2% to the aminoglycosides, and 22.7% to the urinary antiseptic, nitrofurantoin. The isolates were non-sensitive to tetracycline (93.8%), cotrimoxazole (92.2%), and nalidixic acid (86.7%). Most isolates showed non-uniform sensitivity patterns to the cephaloporins (cefuroxime and ceftazidime). Pseudomonas spp. isolates were generally resistant to the fluorinated quinolones. Conclusion Though the fluorinated quinolones are still largely effective for empirical therapy in urinary tract infections, the importance of prior sensitivity testing in checking the emergence of bacterial antibiotic resistance can not be overemphasized.

Yeast as a tool for characterizing mono-ADP-ribosyltransferase toxins

The emergence of bacterial antibiotic resistance poses a significant challenge in the pursuit of novel therapeutics, making new strategies for drug discovery imperative. We have developed a yeast growth-defect phenotypic screen to help solve this current dilemma. This approach facilitates the identification and characterization of a new diphtheria toxin (DT) group, ADP-ribosyltransferase toxins from pathogenic bacteria. In addition, this assay utilizes Saccharomyces cerevisiae, a reliable model for bacterial toxin expression, to streamline the identification and characterization of new inhibitors against this group of bacterial toxins that may be useful for antimicrobial therapies. We show that a mutant of the elongation factor 2 target protein in yeast, G701R, confers resistance to all DT group toxins and recovers the growth-defect phenotype in yeast. We also demonstrate the ability of a potent small-molecule toxin inhibitor, 1,8-naphthalimide (NAP), to alleviate the growth defect caused by toxin expression in yeast. Moreover, we determined the crystal structure of the NAP inhibitor-toxin complex at near-atomic resolution to provide insight into the inhibitory mechanism. Finally, the NAP inhibitor shows therapeutic protective effects against toxin invasion of mammalian cells, including human lung cells.

Characterization of Antibiotic Resistance Among Veterinary Bacterial Pathogens

Since the introduction of antimicrobials into veterinary medicine some 45 years ago, animal health and productivity has improved significantly.1,4,10,12 Despite considerable use, and some misuse, many antimicrobials continue to remain effective today. However, loss of efficacy through emergence of bacterial antibiotic resistance is always an ever present risk.1,2,3,6,14,15 Antibiotic resistant bacterial pathogens in animals not only pose a problem with respect to animal health but are a growing concern regarding possible transmission to humans as foodborne pathogens.7,9,15,18 The dilemma of antibiotic resistance is worsened by the growing number of bacterial pathogens resistant to multiple, structurally unrelated drugs, and to the fact that few veterinary antimicrobials are likely to be available before the end of the decade.4,8,12 Accordingly, more attention is now being paid to the ease at which resistance can develop to both single and multiple antimicrobials among bacterial pathogens. If current trends continue, we may encounter bacterial pathogens which are resistant to all known antimicrobials. This situation is being addressed by both FDA and USDA which are currently implementing strategies to head off this potential threat.

Natural genetic engineering in evolution.

The results of molecular genetics have frequently been difficult to explain by conventional evolutionary theory. New findings about the genetic conservation of protein structure and function across very broad taxonomic boundaries, the mosaic structure of genomes and genetic loci, and the molecular mechanisms of genetic change all point to a view of evolution as involving the rearrangement of basic genetic motifs. A more detailed examination of how living cells restructure their genomes reveals a wide variety of sophisticated biochemical systems responsive to elaborate regulatory networks. In some cases, we know that cells are able to accomplish extensive genome reorganization within one or a few cell generations. The emergence of bacterial antibiotic resistance is a contemporary example of evolutionary change; molecular analysis of this phenomenon has shown that it occurs by the addition rearrangement of resistance determinants and genetic mobility systems rather than by gradual modification of pre-existing cellular genomes. In addition, bacteria and other organisms have intricate repair systems to prevent genetic change by sporadic physicochemical damage or errors of the replication machinery. In their ensemble, these results show that living cells have (and use) the biochemical apparatus to evolve by a genetic engineering process. Future research will reveal how well the regulatory systems integrate genomic change into basic life processes during evolution.