β-Lactams and β-Lactamase Inhibitors: An Overview.

Disease-processes experienced by critically ill patients commonly cause unique pharmacokinetic (PK) changes, that may result in drug concentrations that are either sub-therapeutic or toxic when dosing is based on PK data obtained from non-critically ill patients. Ensuring therapeutic dosing of beta-lactam antibiotics in critically ill patient is of increasing concern, given the difficult-to-predict PK in this population, frequent use of this antibiotic class in treating severe infections and the importance of optimal antibiotic therapy for reducing mortality rates. Therapeutic drug monitoring (TDM), a strategy frequently used in dosing drugs with complex PK, has demonstrated potential clinical advantages for improving dosing of beta-lactams in the critically ill. However, the practice of beta-lactam TDM and dose optimisation is still limited by a number of factors. Large-scale data on the adequacy of current dosing regimens and optimal PK/pharmacodynamic (PK/PD) targets associated with maximal clinical outcome is still limited. In particular, currently available data on beta-lactam TDM have not adequately quantified the potentially profound impact of hypoalbuminaemia, common among the critically ill, on the PK of the pharmacologically active unbound concentration of beta-lactams.This Thesis aims to describe the application of a beta-lactam TDM-guided dosing program in critically ill patients, with the utilization of directly-measured unbound drug concentration data. Specifically, this work will first characterise the impact of altered protein binding in critically ill patients on the accuracy of commonly used methods for calculation of unbound beta-lactam concentrations (from measured total concentrations). Secondly, this work will describe the achievement of PK/PD targets in relation to directly-measured unbound beta-lactam concentrations in critically ill patients receiving empiric dosed and TDM-guided beta-lactam antibiotic treatments. Finally, the association of beta-lactam concentrations with clinical outcome in critically ill patients with blood stream infections will also be characterised.This Thesis comprises of seven chapters. Chapter one introduces relevant PK/PD and beta-lactam dosing concepts for this Thesis.Chapter two includes a published review article that systemically analyses the current literature at the time on the risk of inadequate drug exposure in critically ill patients and discusses the role of TDM for different class of antibiotics in this setting. This chapter specifically addresses available evidence relating to practices of antibiotic TDM, and the practical aspects of applying TDM to potentially improve outcomes from severe infections in the critically ill.Chapter three is an original research article that reports findings of a multicentre survey on the various approaches used for beta-lactam TDM in intensive care units (ICUs) worldwide.Chapter four is an original research article that describes findings of a single-centre prospective study on unbound beta-lactam concentrations in the critically ill determined by two assays. The aim of this study was to compare the measured unbound concentrations with the unbound concentrations conventionally predicted from published protein binding values for seven beta-lactam antibiotics commonly used in the critically ill. Chapter five describes findings of a single-centre prospective study on attainment of predefined PK/PD targets in critically ill patients who received empirically dosed and TDM-guide (using direct-measured unbound drug concentrations) beta-lactam antibiotics. Furthermore, the study also aimed to describe patient and treatment factors associated with attainment of target concentrations.Chapter six describes findings of association between beta-lactam exposures, using PK/PD indices, and clinical outcomes from a retrospective analysis of prospective studies database. Furthermore, the study also aimed to describe patient and treatment factors associated with patient outcome.Chapter seven includes an overall discussion and summary of the findings of this Thesis, and explores on potential future research in this area.

The revival of fosfomycin

Chapter 15 - Emerging New Therapeutics Against Key Gram-Negative Pathogens

To learn the safety profile of carbapenems and compare suspected adverse drug reactions (ADRs) among carbapenem classes by data mining the FDA adverse event reporting system (FAERS) database. This retrospective study described the general characteristics of adverse drug event (ADE) reports related to carbapenems in the FAERS during 2015 - 2018. The 95% confidence intervals (CIs) of proportional reporting ratio (PRR), the reporting odds ratio (ROR), and information component (IC) of Bayesian confidence propagation neural network (BCPNN) were calculated to identify potential safety signals. A total number of 5,899 reports associated with carbapenems were submitted to the FAERS from January 1, 2015 to December 31, 2018. The most frequently reported ADE associated with carbapenems was drug ineffective (10.51%). Serious ADEs and death associated with carbapenems were reported in 41.24and 25.12%, respectively. Infections and infestations was the strongest signal detected in both meropenem and imipenem. Nervous system disorders and psychiatric disorders were strongly detected in ertapenem. Hepatobiliary disorders were common in doripenem patients. Carbapenem resistance is alarming in the United States, and carbapenem is more likely to be associated with serious and fatal ADEs among β-lactam antibiotics. Both differences and similarities exist in the safety profile among carbapenems classes. Close attention should be paid to patients with special disease when administrated carbapenems.

Structural Determinants Of Antibiotic And β-lactamase Diffusion Through Bacterial Porins

Methicillin-resistant Staphylococcus aureus (MRSA) is a global bacterial scourge that has become resistant to many classes of antibiotics, and treatment options for MRSA infections are limited. The cause of MRSA resistance to all commercially available beta-lactam antibiotics is the acquisition of the gene mecA, which encodes penicillin-binding protein 2a (PBP 2a). PBP 2a is a transpeptidase, which in contrast to the other transpeptidases of S. aureus does not experience inhibition by beta-lactam antibiotics. The lack of inhibition is due to a closed conformation for the active site for PBP 2a, which opens up only in the course of the catalytic function of the protein. Here we show that two new anti-MRSA antibiotics now undergoing clinical trials, ceftaroline and ME1036, are able to inhibit PBP 2a effectively, a process that is enhanced in the presence of a cell wall structural surrogate. It is likely that in the course of bacterial growth the occupancy of the allosteric site for the cell wall is co-opted by these antibiotics, and under these conditions the second-order rate constant for the encounter of the antibiotic and PBP 2a approaches the clinically useful value of 10(4)-10(5) M-1 s-1. These compounds are potent inhibitors of PBP 2a as well as PBPs from other species, and have potential as therapeutic agents for treatment of serious infections by MRSA and other resistant bacterial pathogens.

Gram-negative bacteria shield themselves from antibiotics by producing an outer membrane (OM) that forms a formidable permeability barrier. Multidrug resistance among these organisms is a particularly acute problem that is exacerbated by the OM. The poor penetrance of many available antibiotics prevents their clinical use, and efforts to discover novel classes of antibiotics against Gram-negative bacteria have been unsuccessful for almost 50 years. Recent insights into how the OM is built offer new hope. Several essential multiprotein molecular machines (Bam, Lpt, and Lol) work in concert to assemble the barrier and offer a swathe of new targets for novel therapeutic development. Murepavadin has been at the vanguard of these efforts, but its recently reported phase III clinical trial toxicity has tempered the anticipation of imminent new clinical options. Nonetheless, the many concerted efforts aimed at breaking down the OM barrier provide a source of ongoing optimism for what may soon come through the development pipeline. We will review the current state of drug development against the OM assembly targets, highlighting insightful new discovery approaches and strategies.

As effective as antibiotics have been since they were first discovered, antibiotic resistance proves to be a worldwide threat, endangering their efficacy. Over the years, there has been a significant decline in development of novel class antibiotics. These two factors have made once easily treated infections difficult to manage. Carbapenem‐resistant Enterobacteriaceae (CRE) are an emerging threat due to their high levels of antibiotic resistance. CRE have become resistant to almost all antibiotics, and cause 23,000 infections per year in the United States. Metals are a vital nutrient in all living organisms including bacteria and, have thus been targeted by antimicrobial agents. Bacteria, like all living organisms, must regulate their metal homeostasis in order to survive and replicate. Therefore, mechanisms able to diminish the availability of the metals to bacterial pathogens can be used to develop new therapeutic drugs.An example of a mechanism of bacterial resistance to antibiotics is the production of inactivating enzymes, such as carbapenemases, an example of which is seen in Klebsiella pneumoniae. Carbapenemases play an important role against b‐lactam antibiotics such as imipenem in Enterobacteriaceae. Carbapenemases are categorized based on their functional and molecular properties. Class B carbapenemases are metallo‐b‐lactamases (MBL) which require zinc in their active site, and are therefore considered metalloenzymes. MBL can hydrolyze all b‐lactams and are not inactivated by b‐lactamase inhibitors.In this study we analyzed antimicrobial susceptibility of K. pneumoniae isolates, KPC 2146, KPC 1705 and KPC 1706 which contain class B, class A and no carbapenemase, respectively. Metal homeostasis of isolates was disrupted by inducing zinc deficiency using TPEN, a metal chelator with high affinity for zinc, and preparing a zinc deficient minimal media. Carbapenemase activity was assessed using CARBA‐NP test and LC‐MS. Bacterial susceptibility of zinc‐deficient isolates to carbapenems was analyzed using spot plating, growth curve, minimum inhibitory concentration and ring of inhibition.Our findings demonstrate zinc dependency of class B carbapenemase in the KPC 2146 isolate, while class A carbapenemase in the KPC 1705 isolate was not affected. Spot plating and growth curve analyses demonstrated that zinc deficiency lowers bacterial growth in KPC 2146 in the presence of imipenem. Zinc deficiency via TPEN addition lowered imipenem, doripenem, ertapenem and meropenem MIC in KPC 2146. Ring of inhibition of KPC 2146 with imipenem was greatly affected by zinc chelation. In conclusion, we demonstrate that disrupting zinc homeostasis affects class B carbapenemase activity, which in turn increases bacterial susceptibility to carbapenems.Support or Funding InformationNika Padyab was supported by funds from the Biomedical Sciences Program at MWU. Drs. Veltri and Hernandez were supported by MWU intramural funds.

The extensive use and misuse of antibiotics during the last seven decades has led to the evolution and global spread of a variety of resistance mechanisms in bacteria. Of high medical importance are β-lactamases, a group of enzymes inactivating β-lactam antibiotics. Metallo-β-lactamases (MBLs) are particularly problematic because of their ability to act on virtually all classes of β-lactam antibiotics. An engineered MBL (evMBL9) characterized by low level activity with several β-lactam antibiotics was constructed and employed as a parental MBL in an experiment to examine how an enzyme can evolve toward increased activity with a variety of β-lactam antibiotics. We designed and synthesized a mutant library in which the substrate activity profile was varied by randomizing six active site amino acid residues. The library was expressed in Salmonella typhimurium, clones with increased resistance against seven different β-lactam antibiotics (penicillin G, ampicillin, cephalothin, cefaclor, cefuroxime, cefoperazone, and cefotaxime) were isolated, and the MBL variants were characterized. For the majority of the mutants, bacterial resistance was significantly increased despite marked reductions in both mRNA and protein levels relative to those of parental evMBL9, indicating that the catalytic activities of these mutant MBLs were highly increased. Multivariate analysis showed that the majority of the mutant enzymes were generalists, conferring increased resistance against most of the examined β-lactams.

Since the discovery of penicillin, the forerunner of the most widely used class of antibiotics (i.e. β-lactams), natural compounds and their derivatives represented a major source of antibacterial therapeutic products whose availability enabled modern medical practices (invasive surgery, organ transplant, etc.). However, the relentless emergence of resistant bacteria is challenging the long-term efficacy of antibiotics, also decreasing their economic attractiveness for big pharma, leading to a significant decay in antibacterial development in the 21st century and an increased use of last-resort drugs such as carbapenems or colistin. Indeed, bacteria evolved an arsenal of resistance mechanisms, leading to the emergence of totally-drug resistant isolates, already sporadically isolated among Gram-negative bacterial species. To face this deadly peril, it is fundamental to explore new ground-breaking approaches. In view of the significance of both β-lactam antibiotics and the production of one or more β-lactamases as the major resistance mechanism (especially in Gram-negative bacteria), we implemented an original approach to selectively deliver antibacterial zidovudine (AZT) exploiting the β-lactamase-mediated hydrolysis of a β-lactam-conjugate prodrug. The synthesis of the targeted pronucleosides was performed in 5-7 steps and based on an original Pd-catalyzed cross-coupling reaction. Enzymatic and microbiological evaluations were performed to evaluate the synthesized pronucleosides, yielding new insights into molecular recognition of β-lactamase enzymes. This approach would potentially allow a targeted and selective eradication of antibiotic-resistant β-lactamase-producing (opportunistic) pathogens, as the inactive prodrug is unable to harm the commensal microbial flora.

The resistance of methicillin-resistant Staphylococcus aureus (MRSA) to all β-lactam classes limits treatment options for serious infections involving this organism. Our goal is to discover new agents that restore the activity of β-lactams against MRSA, an approach that has led to the discovery of two classes of natural product antibiotics, a cyclic depsipeptide (krisynomycin) and a lipoglycopeptide (actinocarbasin), which potentiate the activity of imipenem against MRSA strain COL. We report here that these imipenem synergists are inhibitors of the bacterial type I signal peptidase SpsB, a serine protease that is required for the secretion of proteins that are exported through the Sec and Tat systems. A synthetic derivative of actinocarbasin, M131, synergized with imipenem both in vitro and in vivo with potent efficacy. The in vitro activity of M131 extends to clinical isolates of MRSA but not to a methicillin-sensitive strain. Synergy is restricted to β-lactam antibiotics and is not observed with other antibiotic classes. We propose that the SpsB inhibitors synergize with β-lactams by preventing the signal peptidase-mediated secretion of proteins required for β-lactam resistance. Combinations of SpsB inhibitors and β-lactams may expand the utility of these widely prescribed antibiotics to treat MRSA infections, analogous to β-lactamase inhibitors which restored the utility of this antibiotic class for the treatment of resistant Gram-negative infections.

Methicillin-resistant Staphylococcus aureus MRSA is a S. aureus that resistant to β-lactam antibiotics (e.g., Cefoxitin and Oxacillin). MRSA has a tremendous capacity to develop resistance to other classes of antibiotics and forming a real threat to patients. The process of exploring a new tactic of non-antibiotic treatments has become an urgent need. A bacteriophage is one of the possible treatments that strongly suggested. Bacteriophages are viruses that infect bacteria as a natural host with a bactericidal capability against multidrug-resistant bacteria that do not respond to conventional antibiotics. The current study investigates the lytic efficacy of phage-cocktail in vitro, specifically against S. aureus isolated from skin infections and find out the possible association of phage-antibiotic resistance. A total of 43 isolates of Methicillin-resistant staphylococcus aureus were isolated from skin infections. The isolates are distributed as (10 isolates of burn, 4 isolates of diabetic foot ulcer, 7 isolates of surgical wounds, 3 isolates of pressure ulcer, and 19 of skin and soft tissue infection). The isolates exhibited variant antibiotic susceptibility against 12 antibiotics (Cefoxitin FOX, Vancomycin VAN, Oxacillin OX, Rifampin RA, Chloramphenicol C, Nitrofurantoin F, Clindamycin DA, Azithromycin AZM, Amikacin AK, Trimethoprim-sulfamethoxazole SXT, Ciprofloxacin CIP, and Gentamicin CN). A bacteriophage cocktail was isolated using a phage-enrichment technique, high titer phage lysate (5*109 pfu/ml) was obtained and investigated against 43 MRSA isolates. The phage-cocktail showed high specificity to S. aureus but variable susceptibility to 43 MRSA isolates. It was observed that there was no association (p greater than 0.05) between phage and antibiotic resistance of (FOX, OX, VAN, RA, C, F, and DA) where the significant association was observed (p less thaN 0.05) with (AZM, AK, SXT, CIP, and CN). Significantly, the more antibiotic-resistant isolates exhibited more sensitivity to phage-cocktail, which represents a promising alternative to antibiotics that do not affect with increasing antibiotic resistance. How to cite this article: Hantoosh, S.M. and Jarallah, E.M. (2019). Isolation of Lytic Bacteriophage Cocktail Against Methicillin-Resistant Staphylococcus Aureus Isolated From Human Skin Infections. International Journal of Drug Delivery Technology, 9(4): 623-629. Source of support: Nil. Conflict of interest: None.

Physicians often must select antibiotics for patients who are reported to have an antibiotic allergy. For penicillins, the sensitivity of penicillin skin testing for predicting serious allergic reactions is excellent. For other beta-lactam antibiotics, penicillin skin testing is useful for excluding the possibility of sensitivity to the beta-lactam ring. For other antibiotics, the patient history remains the most useful tool for determining whether a serious reaction is likely to occur with further drug exposure. The cross-reactivity between penicillins and second- or third-generation cephalosporins (excluding cefamandole) is probably no higher than is the cross-reactivity between penicillins and other classes of antibiotics. When a patient has a suspected immunoglobulin E-mediated antibiotic allergy, desensitization therapy should be considered, if the efficacy of alternate antibiotics is in doubt. For the treatment of serious infections, it is usually possible to safely administer the antibiotic of choice despite a history of possible antibiotic allergy.

Staphylococcus aureus ist ein ubiquitär vorkommender Erreger, der bei 15 - 40 % der Menschen zur normalen Haut- und Schleimhautflora gehört und bei Wundinfektionen eine ätiologisch bedeutende Rolle spielt. Zunehmende Bedeutung erlangten in den letzten Jahren die Methicillin-resistenten S. aureus-Stämme (MRSA), die sich durch eine multiple Resistenz gegenüber allen β-Laktamantibiotika und anderen Antibiotikasubstanzklassen auszeichnen. Besonders die zunehmende Bedeutung von MRSA-Stämmen, die gegenüber Glykopeptiden nur noch eingeschränkt empfindlich sind, verschärft die therapeutischen Optionen. Die Rate an MRSA aller in Deutschland isolierten S. aureus-Stämme beträgt ca. 15 % und liegt damit im europäischen Mittelmaß. Da der Hauptübertragungsweg dieser Stämme von Patient zu Patient über das medizinische Personal erfolgt, müssen bei dem Vorliegen einer MRSA-Kolonisation und/oder Infektion strikte hygienische Vorkehrungen sowie Sanierungsmaßnahmen getroffen werden, um eine epidemische Ausbreitung der Stämme zu verhindern. Eine antibiotische Therapie eines asymptomatischen MRSA-Trägers ist nicht indiziert. Bei manifesten Infektionen sollte primär lokal antiseptisch behandelt und sekundär erst eine systemische Antibiotikatherapie nach vorliegenden Antibiogrammen durchgeführt werden. Dabei sollten zuerst wirksame herkömmliche Antibiotika wie Vancomycin, Rifampicin, Fosfomycin und andere eingesetzt werden.

O objetivo do estudo é descrever as características clínico-epidemiológicas de pacientes internados com sepse, assim como, determinar o uso de drogas vasoativas, antimicrobianos e os principais desfechos nestes pacientes. Trata-se de um estudo descritivo, com abordagem quantitativa, realizado com 367 pacientes internados com diagnóstico de sepse no período de fevereiro de 2016 a julho de 2017 em um hospital privado do estado de Sergipe, Brasil, o qual realiza atendimentos ambulatoriais e de emergência. Após aprovação do Comitê de Ética em Pesquisa, os dados foram coletados por meio dos registros contidos em prontuários por um único pesquisador. A sepse foi responsável por 367 (2,2%) internações no período do estudo, a maioria dos pacientes eram idosos (78,5%), do sexo feminino (50,7%), com pelo menos três comorbidades (70,3%) e internados por sepse clínica (94,5%). A classe de antibióticos mais utilizada foi a dos β-lactâmicos (64,6%). O tempo de permanência e a mortalidade mostraram-se expressivamente maior nos pacientes que sofreram choque séptico em relação aos que não sofreram. O estudo evidenciou que a sepse acometeu majoritariamente idosos do sexo feminino e que as maiores taxas de mortalidade e o maior período de permanência foram observados nos pacientes com agravamento da sepse.