Staphylococcus aureus in Continuous Culture: A Tool for the Rational Design of Antibiotic Treatment Protocols

In vitro measures of the pharmacodynamics of antibiotics that account for the factors anticipated for bacteria in infected patients are central to the rational design of antibiotic treatment protocols. We consider whether or not continuous culture devices are a way to obtain these measures. Staphylococcus aureus PS80 in high-density continuous cultures were exposed to oxacillin, ciprofloxacin, vancomycin, gentamicin, daptomycin and linezolid. Contrary to results from low density retentostats as well as to predictions of traditional PK/MIC ratios, daily dosing with up to 100× MIC did not clear these cultures. The densities of S. aureus in these cultures oscillated with constant amplitude and never fell below 10(5) CFU per ml. Save for daptomycin "treated" populations, the densities of bacteria in these cultures remained significantly below that of similar antibiotic-free cultures. Although these antibiotics varied in their pharmacodynamic properties there were only modest differences in their mean densities. Mathematical models and experiments suggest that the dominant factor preventing clearance was wall-adhering subpopulations reseeding the planktonic population which can be estimated and corrected for. Continuous cultures provide a way to evaluate the potential efficacy of antibiotic treatment regimes in vitro under conditions that are more clinically realistic and comprehensive than traditional in vitro PK/PD indices.     

Origins and Evolution of Antibiotic Resistance

Summary: Antibiotics have always been considered one of the wonder discoveries of the 20th century. This is true, but the real wonder is the rise of antibiotic resistance in hospitals, communities, and the environment concomitant with their use. The extraordinary genetic capacities of microbes have benefitted from man''s overuse of antibiotics to exploit every source of resistance genes and every means of horizontal gene transmission to develop multiple mechanisms of resistance for each and every antibiotic introduced into practice clinically, agriculturally, or otherwise. This review presents the salient aspects of antibiotic resistance development over the past half-century, with the oft-restated conclusion that it is time to act. To achieve complete restitution of therapeutic applications of antibiotics, there is a need for more information on the role of environmental microbiomes in the rise of antibiotic resistance. In particular, creative approaches to the discovery of novel antibiotics and their expedited and controlled introduction to therapy are obligatory.     

Inhibiting the Evolution of Antibiotic Resistance

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基因的推理设计与改造—体外分子进化的捷径

体外分子进化是改造基因、获得新的功能蛋白质重要手段之一,已经取得了令人瞩目的成就。推理设计是利用序列和结构的比较信息创造新的基因和蛋白质,是研究改造蛋白质功能的有效方法。文章着重介绍了基因推理设计在基因体外分子进化中的应用,从简单实用的基因密码偏爱设计,结合多个不同性状相关功能基因信息的基因推理设计,关键功能区域的简并引物设计,位点直接蛋白质重组4个方面进行描述,并综述了该方法近年来的应用。     

The Antibiotic Dosage of Fastest Resistance Evolution: Gene Amplifications Underpinning the Inverted-U

To determine the dosage at which antibiotic resistance evolution is most rapid, we treated Escherichia coli in vitro, deploying the antibiotic erythromycin at dosages ranging from zero to high. Adaptation was fastest just below erythromycin’s minimal inhibitory concentration (MIC) and genotype-phenotype correlations determined from whole genome sequencing revealed the molecular basis: simultaneous selection for copy number variation in three resistance mechanisms which exhibited an “inverted-U” pattern of dose-dependence, as did several insertion sequences and an integron. Many genes did not conform to this pattern, however, reflecting changes in selection as dose increased: putative media adaptation polymorphisms at zero antibiotic dosage gave way to drug target (ribosomal RNA operon) amplification at mid dosages whereas prophage-mediated drug efflux amplifications dominated at the highest dosages. All treatments exhibited E. coli increases in the copy number of efflux operons acrAB and emrE at rates that correlated with increases in population density. For strains where the inverted-U was no longer observed following the genetic manipulation of acrAB, it could be recovered by prolonging the antibiotic treatment at subMIC dosages.     

A New Tool to Reveal Bacterial Signaling Mechanisms in Antibiotic Treatment and Resistance

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Highlights

• •Quantitative (phospho)proteome analysis of antibiotic treatment in E. coli.
• •Largest bacterial phosphorylation catalogue.
• •Specific phosphorylation motifs changes during resistance development.
• •Phosphorylation mediated signaling could be a potential target for drug design.

     

TRAIL耐药产生的机制及逆转耐药的最新策略

肿瘤坏死因子相关凋亡诱导配体（tumor necrosis factor-related apoptosis inducing li-gand,TRAIL）是唯一能诱导癌细胞凋亡而对机体正常组织无明显损伤的内源性细胞因子,因而被认为是一种极具前景的抗癌药物。然而目前研究发现,许多恶性肿瘤细胞对TRAIL具有耐药性,使TRAIL在临床应用中遭遇瓶颈。越来越多的证据表明,一些关键信号通路可能与TRAIL耐药有关,且利用靶向基因治疗策略以及借助某些天然药物或小分子抑制剂能够部分恢复癌细胞对TRAIL的敏感性。该文主要描述了肿瘤细胞对TRAIL的耐药机制,并对如何有效克服和逆转TRAIL耐药的策略作了简要概括。     

Multi-scale Predictions of Drug Resistance Epidemiology Identify Design Principles for Rational Drug Design

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Prevalence of Antibiotic Resistance in Drinking Water Treatment and Distribution Systems

The occurrence and spread of antibiotic-resistant bacteria (ARB) are pressing public health problems worldwide, and aquatic ecosystems are a recognized reservoir for ARB. We used culture-dependent methods and quantitative molecular techniques to detect and quantify ARB and antibiotic resistance genes (ARGs) in source waters, drinking water treatment plants, and tap water from several cities in Michigan and Ohio. We found ARGs and heterotrophic ARB in all finished water and tap water tested, although the amounts were small. The quantities of most ARGs were greater in tap water than in finished water and source water. In general, the levels of bacteria were higher in source water than in tap water, and the levels of ARB were higher in tap water than in finished water, indicating that there was regrowth of bacteria in drinking water distribution systems. Elevated resistance to some antibiotics was observed during water treatment and in tap water. Water treatment might increase the antibiotic resistance of surviving bacteria, and water distribution systems may serve as an important reservoir for the spread of antibiotic resistance to opportunistic pathogens.The occurrence and spread of antibiotic-resistant bacteria (ARB) are pressing public health problems worldwide, and aquatic ecosystems are a recognized reservoir for ARB and antibiotic resistance genes (ARGs) (4, 6, 8, 11, 12, 15, 39). Naturally occurring ARB and ARGs in the aquatic environment are selected for and enriched for by antibiotics found in sewage and agricultural runoff, which result from the widespread and increased use of antibiotics (4, 11, 12, 15, 38). Historically, concerns about the microbial quality of drinking water have focused on the occurrence of pathogens in drinking water distribution systems (5, 34). However, the presence of trace levels of antibiotics and ARB in source water and finished drinking water may also greatly affect public health and is an emerging issue for the general public and the drinking water industry (3, 30). Although several studies have detected ARB in drinking water systems (2, 3, 20, 30, 38), most previous studies focused on cultivable bacteria and/or indicator organisms. Little is known about the fate of ARGs in drinking water systems, and it was recently proposed that ARGs are emerging contaminants (24).We used culture-dependent methods and molecular techniques to investigate the prevalence and dynamics of heterotrophic ARB and ARGs in a drinking water source (source RW-P) and treated drinking water (source DW-P) (see Materials and Methods in the supplemental material). We tested water from a drinking water plant located in Michigan and tap water from several small cities located in Michigan and Ohio (sources TW-1, TW-2, TW-3, and TW-4). Two independent samples were collected each time at each collection site at three different times, and we used four replicates from each sample for tests. We tested bacterial resistance to the following antibiotics: amoxicillin (amoxicilline), chloramphenicol, ciprofloxacin, gentamicin, rifampin (rifampicin), sulfisoxazole, and tetracycline. We also examined the presence of eight ARGs, including beta-lactam resistance genes (bla_TEM and bla_SHV), chloramphenicol resistance genes (cat and cmr), sulfonamide resistance genes (sulI and sulII), and tetracycline resistance genes (tetO and tetW).Total heterotrophic plate counts (HPC) were determined using R₂A agar without added antibiotics. The water treatment process reduced the total HPC from 9.9 × 10⁶ CFU/100 ml in source water to 68 CFU/100 ml in treated drinking water, indicating that there was efficient removal and/or deactivation of total HPC (Table ​(Table1).1). In contrast, the total 16S rRNA gene copy number decreased from 3.4 × 10⁷ copies/100 ml in source water to 1.6 × 10⁶ copies/100 ml in treated drinking water (Fig. ​(Fig.1).1). The discrepancy between the reduction in the HPC and the reduction in the total 16S rRNA gene copy number suggests that the final disinfection step effectively inactivated bacteria but most of the dead or damaged cells were still present in finished drinking water. The number of HPC in tap water ranged from 3.44 × 10² to 6.1 × 10⁴ CFU/100 ml water, values that are lower than those for source water but significantly higher than those for treated drinking water, indicating that there is regrowth of bacteria in drinking water distribution systems. The copy numbers of total 16S rRNA genes in tap water ranged from 2.45 × 10⁵ to 1.02 × 10⁷ copies/100 ml water. The higher levels suggested by the 16S rRNA data are consistent with results of previous studies demonstrating that only 5 to 10% and 1% of bacteria in wastewater and soil, respectively, can be cultivated or identified by culture-based methods (9, 37). A significant correlation (P < 0.05, R² = 0.78) was found between the 16S rRNA gene copy number and the total HPC if treated drinking water (DW-P) data were not included (Fig. ​(Fig.1).1). This suggests that cultivable bacteria in drinking water represent only a small portion of the total bacterial biomass. Including treated drinking water (DW-P) data resulted in a distorted correlation, suggesting that a large proportion of the 16S rRNA genes present came from dead and/or damaged cells. The levels of total heterotrophic bacteria were significantly higher in tap water (TW-1) than in treated drinking water (DW-P), indicating that there was bacterial regrowth in the water distribution system.Open in a separate windowFIG. 1.Heterotrophic bacteria and the 16S rRNA gene in different water samples. (A) Copy numbers of the 16S rRNA gene and numbers of heterotrophic bacteria (CFU) in 100 ml water. (B) Correlation (P < 0.05, R² = 0.78) between the copy number of the 16S rRNA gene and the number of heterotrophic bacteria in different water samples (without the data for DW-P). RW-P, source water from the drinking water treatment plant; DW-P, finished drinking water from the drinking water treatment plant; TW-1, tap water from the city where the drinking water treatment plant is located; TW-2, TW-3, and TW-4, tap water from three towns in Michigan and Ohio close to the city where the TW-1 drinking water treatment plant is located. The statistical analysis was done using six samples for each type of water sample. Lg, log₁₀.

TABLE 1.

Prevalence of ARB HPC in source water, finished drinking water, and tap water from four towns^a

Gramicidin Peptide to Combat Antibiotic Resistance: A Review

International Journal of Peptide Research and Therapeutics - Emerging issues of Antibiotic drug resistance have increased the mortality cases all around the world. To deal with the situation...     

Antibiotic Resistance in Bacteria and Its Future for Novel Antibiotic Development

Since the first introduction of the sulfa drugs and penicillin into clinical use, large numbers of antibiotics have been developed and hence contributed to human health. But extensive use of antibiotics has raised a serious public health problem due to multiantibiotic resistant bacterial pathogens that inevitably develop resistance to every new drug launched in the clinic. Consequently, there is a pressing need to develop new antibiotics to keep pace with bacterial resistance. Recent advances in microbial genomics and X-ray crystallography provide opportunities to identify novel antibacterial targets for the development of new classes of antibiotics and to design more potent antimicrobial compounds derived from existing antibiotics respectively. To prevent and control infectious diseases caused by multiantibiotic resistant bacteria, we need to understand more about the molecular aspects of the pathogens’ physiology and to pursue ways to prolong the life of precious antibiotics.     

Fate of Antibiotic Resistant Bacteria and Genes during Wastewater Chlorination: Implication for Antibiotic Resistance Control

This study investigated fates of nine antibiotic-resistant bacteria as well as two series of antibiotic resistance genes in wastewater treated by various doses of chlorine (0, 15, 30, 60, 150 and 300 mg Cl₂ min/L). The results indicated that chlorination was effective in inactivating antibiotic-resistant bacteria. Most bacteria were inactivated completely at the lowest dose (15 mg Cl₂ min/L). By comparison, sulfadiazine- and erythromycin-resistant bacteria exhibited tolerance to low chlorine dose (up to 60 mg Cl₂ min/L). However, quantitative real-time PCRs revealed that chlorination decreased limited erythromycin or tetracycline resistance genes, with the removal levels of overall erythromycin and tetracycline resistance genes at 0.42 ± 0.12 log and 0.10 ± 0.02 log, respectively. About 40% of erythromycin-resistance genes and 80% of tetracycline resistance genes could not be removed by chlorination. Chlorination was considered not effective in controlling antimicrobial resistance. More concern needs to be paid to the potential risk of antibiotic resistance genes in the wastewater after chlorination.     

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.