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The Sterne and Ames strains of Bacillus anthracis carry chromosomal genes bla1 and bla2 , which confer β-lactam resistance when expressed in Escherichia coli . MIC measurements and steady-state kinetic analyses indicate that Bla1 possesses penicillinase activity while Bla2 possesses penicillinase, cephalosporinase, and carbapenem-hydrolyzing activities.

Carbapenemase detection has become a major problem in hospitals that encounter outbreaks of infections caused by carbapenem-resistant Gram-negative bacteria. Rapid detection systems have been reported using multiplex PCR analyses and DNA microarray assays. Major carbapenemases that are detected by these systems include the KPC and OXA serine carbapenemases, and the IMP, VIM and NDM families of metallo-β-lactamases. However, increasing numbers of the SME serine carbapenemase are being reported from Serratia marcescens, especially from North and South America. These organisms differ from many of the other carbapenemase-producing pathogens in that they are generally susceptible to the expanded-spectrum cephalosporins ceftazidime and cefepime while retaining resistance to almost all other β-lactam antibiotics. Thus, multiplex PCR assays or DNA microarray testing of carbapenem-resistant S. marcescens isolates should include analyses for production of the SME carbapenemase. Confirmation of the presence of this enzyme may provide reassurance that oxyimino-cephalosporins can be considered for treatment of infections caused by these carbapenem-resistant pathogens.

New antibiotic agents are desperately needed to treat the multidrug-resistant pathogens that continue to emerge at alarming rates. Many of the agents that have entered full clinical development since 1995 have been members of previously accepted classes of antibiotics. Among these are a new aminoglycoside (plazomicin), anti-MRSA cephalosporins (ceftobiprole and ceftaroline), a monocyclic β-lactam (BAL30072), the β-lactamase inhibitor combination of tazobactam with the anti-pseudomonal cephalosporin ceftolozane, β-lactam combinations with new non-β-lactam inhibitors (MK-7655 with imipenem, and avibactam with ceftazidime and ceftaroline), new macrolides (cethromycin and solithromycin), oxazolidinones (tedizolid phosphate and radezolid), and quinolones (delafloxacin, nemonoxacin and JNJ-Q2). Resistance and safety issues have been circumvented by some of these new agents that have well-established mechanisms of action and defined pathways leading toward regulatory approval.

The metallo-beta-lactamase gene, ccrA, has been cloned from three clinical isolates of Bacteroides fragilis, TAL3636, QMCN3, and QMCN4. Although all three isolates harbored a gene encoding a potent beta-lactamase, the MICs of benzylpenicillin, piperacillin, cefotaxime, ceftazidime, imipenem, and biapenem for the three isolates varied from 4- to > 128-fold. QMCN4 was the most susceptible of the three isolates, followed by QMCN3. TAL3636 was resistant to all of the beta-lactams. Previous DNA sequence analysis of the three ccrA genes revealed that the enzymes differed at 5 amino acid residues (B. A. Rasmussen, Y. Gluzman, and F. P. Tally, Mol. Microbiol. 5:1211-1219, 1991). Biochemical characterization of the three enzymes revealed only small differences in kcat and Km values for the majority of beta-lactams tested. Thus, the 5 amino acid substitutions affected the hydrolyzing activity of the enzymes only modestly. Crypticity differences between the three isolates showed that QMCN4 was the least permeable of the isolates to cephaloridine, followed by TAL3636, and that QMCN3 was highly permeable to cephaloridine. Therefore, neither catalytic activity nor permeability was a major contributor to the dramatic differences in the MICs. Instead, microbiological susceptibility was closely related to the level of metallo-beta-lactamase present in each isolate. Both biochemical and physical studies indicated that TAL3636 produced 5- to 10-fold and 50- to 100-fold more metallo-beta-lactamase than QMCN3 and QMCN4, respectively. Therefore, the level of CcrA enzyme production is the dominant contributing factor to high-level resistance among strains harboring a ccrA gene.

β-Lactamases represent some of the most widely studied examples of protein evolution, with mutations and new enzyme forms closely related to the introduction and use of novel β-lactam antibiotics. Early antibiotic literature described the presence of species-specific penicillinases and cephalosporinases. However, the recognition of plasmid-encoded β-lactamases expanded the list of enzymes to include hundreds of broad-spectrum enzymes that could be transferred among species. In spite of our best efforts to predict the evolutionary course for these enzymes, surprises have frequently occurred with the previously unnoticed occurrence of highly mutable plasmid-encoded β-lactamases from environmental sources, such as the now widely disseminated CTX-M extended-spectrum β-lactamases and the KPC family of serine carbapenemases. This unexplored natural reservoir as a source for new β-lactamases limits our ability to forecast the future evolutionary path(s) for these structurally diverse but functionally similar families of enzymes.

The development and spread of antibiotic resistance in bacteria is a universal threat to both humans and animals that is generally not preventable but can nevertheless be controlled, and it must be tackled in the most effective ways possible. To explore how the problem of antibiotic resistance might best be addressed, a group of 30 scientists from academia and industry gathered at the Banbury Conference Centre in Cold Spring Harbor, New York, USA, from 16 to 18 May 2011. From these discussions there emerged a priority list of steps that need to be taken to resolve this global crisis.

β-Lactamase evolution presents to the infectious disease community a major challenge in the treatment of infections caused by multidrug-resistant gram-negative bacteria. Because over 1,000 of these naturally occurring β-lactamases exist, attempts to correlate structure and function have become daunting. Although new enzymes in the extended-spectrum β-lactamase (ESBL) families are frequently identified, the older CTX-M-14 and CTX-M-15 enzymes have become the most prevalent ESBLs in global surveillance. Carbapenemases with either serine-based or zinc-facilitated hydrolysis mechanisms are posing some of the most critical problems. Most geographical regions now report KPC serine carbapenemases and the metallo-β-lactamases VIM, IMP, and NDM-1, even though NDM-1 was only recently identified. The rapid emergence of these newer enzymes, with multiple β-lactamases appearing in a single organism, makes the design of new β-lactamase inactivators or β-lactamase-stable β-lactams all the more difficult. Combination therapy will likely be required to counteract the continuing evolution of these insidious enzymes in multidrug-resistant pathogens.

Antibiotic resistance issues necessitate the continued discovery and development of new antibacterial agents. Efforts are ongoing in two approaches to find new compounds that are effective against antibiotic-resistant pathogens. These efforts involve modification of existing classes including fluoroquinolones, tetracyclines, aminoglycosides, and β-lactams and identification of inhibitors against previously unexploited antibacterial targets. Examples of both approaches are described here with emphasis on compounds in late pre-clinical or clinical stages of development.

Ceftazidime-resistant isolates of Escherichia coli and Klebsiella pneumoniae produced a plasmid-mediated beta-lactamase with a pI of 5.6 with biochemical characteristics comparable to those of the TEM-10 beta-lactamase. Plasmids from the two strains were nonidentical. Both TEM-10 sequences differed from TEM-1 by substitutions of Ser-162 and Lys-237. The nucleotide sequences of the two genes were identical except for three silent nucleotide substitutions corresponding to the nucleotide differences in the Tn2 TEM-1 or Tn3 TEM-1 genes. The original TEM-10 plasmid was identical to that found in the E. coli isolate and coded for a gene that corresponded to the TEM-10 beta-lactamase from Tn2.

Tazobactam was shown to be a potent inhibitor of group 1, 2a, 2b, and 2b' beta-lactamases. Extended kinetic studies with class A and C serine beta-lactamases showed that the PC1, TEM-2, and P99 enzymes all were reversibly inhibited prior to inactivation of the enzymes. The CcrA metallo-beta-lactamase was less well inhibited, with a 50% inhibitory concentration at least 3 orders of magnitude less favorable than those for most serine beta-lactamases. The numbers of hydrolytic turnovers of tazobactam before inactivation were 2 for PC1, 125 for TEM-2, 50 for P99, and 4,000 for the CcrA enzyme. In spectral studies, transient intermediates were formed after reaction of tazobactam with the PC1, TEM-2, and CcrA beta-lactamases, corresponding to enzyme-associated intermediates responsible for hydrolysis of tazobactam. Chromophores absorbing at 270 nm (CcrA) and 288 nm (TEM-2 and PC1) were observed for these reaction intermediates. The P99 cephalosporinase formed a stable complex with a UV maximum at 295 nm. Incubation of tazobactam with all of the enzymes resulted in accumulation of a tazobactam reaction product with a short-wavelength absorbance. This product has characteristics similar to those of the major eucaryotic metabolite of tazobactam. Possible reaction mechanisms are presented to explain the findings. In conclusion, both serine-based and metallo-beta-lactamases were irreversibly inactivated by tazobactam following an initial transient inhibition phase.

A cephalosporin based prodrug of doxorubicin has been synthesized which effciently releases doxorubicin in the presence of an immunoconjugate consisting of a β-lactamase-MAb.

Wild-type penicillin-binding protein (PBP) 2b from penicillin-susceptible Streptococcus pneumoniae had high affinity for ceftobiprole and penicillin (50% inhibitory concentrations [IC 50 s] of ≤0.15 μg/ml) but not ceftriaxone (IC 50 of >8 μg/ml). In clinical isolates, ceftobiprole and PBP 2b affinities were reduced 15- to 30-fold with a Thr-446-Ala substitution and further still with an additional Ala-619-Gly PBP 2b substitution. Ceftobiprole remained active (MICs of ≤1 μg/ml) against all strains tested and behaved more like penicillin than ceftriaxone with respect to PBP 2b binding.

The susceptibility of 513 clinical isolates to doripenem was determined by broth microdilution, agar dilution, and Etest. Overall agreements for Etest and agar dilution MIC values compared to reference broth microdilution at ±1 log 2 dilution were 88% and 94%, respectively. Etest MIC values demonstrated 98% agreement within ±2 log 2 dilutions compared to the reference broth microdilution method.

Oxazolidinone antibacterial agents, where the N-substituted piperazinyl group of eperezolid was replaced with a N-substituted piperidinyloxy moiety, were synthesized and shown to be active against a variety of resistant and susceptible Gram-positive organisms. The effect of ring size, positional isomerism, and fluorine substitution on antibacterial activity was examined.

Oxazolidinone antibacterial agents, where the morpholino group of linezolid was replaced with an N-substituted piperidinyloxy moiety, were synthesized and shown to be active against a variety of resistant and susceptible Gram-positive organisms. The functionality attached to the piperidine nitrogen was varied extensively to determine the SAR for this series. One of the most potent compounds, 11, showed in vivo efficacy upon subcutaneous administration in a Staphylococcus aureus Smith murine systemic infection.

JNJ-Q2, a novel fluorinated 4-quinolone, was evaluated for its antibacterial potency by broth and agar microdilution MIC methods in studies focused on skin and respiratory tract pathogens, including strains exhibiting contemporary fluoroquinolone resistance phenotypes. Against a set of 118 recent clinical isolates of Streptococcus pneumoniae , including fluoroquinolone-resistant variants bearing multiple DNA topoisomerase target mutations, an MIC 90 value for JNJ-Q2 of 0.12 μg/ml was determined, indicating that it was 32-fold more potent than moxifloxacin. Against a collection of 345 recently collected methicillin-resistant Staphylococcus aureus (MRSA) isolates, including 256 ciprofloxacin-resistant strains, the JNJ-Q2 MIC 90 value was 0.25 μg/ml, similarly indicating that it was 32-fold more potent than moxifloxacin. The activities of JNJ-Q2 against Gram-negative pathogens were generally comparable to those of moxifloxacin. In further studies, JNJ-Q2 exhibited bactericidal activities at 2× and 4× MIC levels against clinical isolates of S. pneumoniae and MRSA with various fluoroquinolone susceptibilities, and its activities were enhanced over those of moxifloxacin. In these studies, the activity exhibited against strains bearing gyrA , parC , or gyrA plus parC mutations was indicative of the relatively balanced (equipotent) activity of JNJ-Q2 against the DNA topoisomerase target enzymes. Finally, determination of the relative rates or frequencies of the spontaneous development of resistance to JNJ-Q2 at 2× and 4× MICs in S. pneumoniae , MRSA, and Escherichia coli were indicative of a lower potential for resistance development than that for current fluoroquinolones. In conclusion, JNJ-Q2 exhibits a range of antibacterial activities in vitro that is supportive of its further evaluation as a potential new agent for the treatment of skin and respiratory tract infections.

In the penicillins and cephalosporins, the β-lactam ring is fused to a five- and six-membered ring, respectively, but monocyclic β-lactam compounds also exist. There are six chemical skeletons other than penicillins or cephalosporins which support β-lactam-containing agents that have current therapeutic use. Although many of these novel β-lactam structures are based on natural products, classes of agents such as the penems and the oxacephems are purely synthetic molecules that have relied on previous knowledge of β-lactam properties to design agents with broad-spectrum antibacterial activity. Characteristics of each of the various classes are listed below

The penicillin class of antibiotics comprises a large group of bicyclic penam compounds ( pp. 226–227 ) differing in the nature of the acyl side chain attached to the fused β-lactam–thiazolidine ring system. Most are semisynthetic derivatives of the penicillin nucleus, 6-aminopenicillanic acid (6-APA), prepared synthetically by the addition of acyl side chains at the 6-amino group.

Three patients with bacteremic pneumonia caused by multidrug-resistant Streptococcus pneumoniae were treated unsuccessfully with azithromycin. One S. pneumoniae isolate carried a mef determinant for an efflux pump; a second isolate had an erm determinant. All 3 patients were successfully treated with levofloxacin, an antipneumococcal fluoroquinolone.

Two classification schemes for β-lactamases are currently in use. The molecular classification is based on the amino acid sequence and divides β-lactamases into class A, C, and D enzymes which utilize serine for β-lactam hydrolysis and class B metalloenzymes which require divalent zinc ions for substrate hydrolysis. The functional classification scheme updated herein is based on the 1995 proposal by Bush et al. (K. Bush, G. A. Jacoby, and A. A. Medeiros, Antimicrob. Agents Chemother. 39:1211-1233, 1995). It takes into account substrate and inhibitor profiles in an attempt to group the enzymes in ways that can be correlated with their phenotype in clinical isolates. Major groupings generally correlate with the more broadly based molecular classification. The updated system includes group 1 (class C) cephalosporinases; group 2 (classes A and D) broad-spectrum, inhibitor-resistant, and extended-spectrum β-lactamases and serine carbapenemases; and group 3 metallo-β-lactamases. Several new subgroups of each of the major groups are described, based on specific attributes of individual enzymes. A list of attributes is also suggested for the description of a new β-lactamase, including the requisite microbiological properties, substrate and inhibitor profiles, and molecular sequence data that provide an adequate characterization for a new β-lactam-hydrolyzing enzyme.

Ceftobiprole, a broad-spectrum cephalosporin with activity against methicillin-resistant Staphylococcus aureus (MRSA) (P. Hebeisen et al., Antimicrob. Agents Chemother. 45:825-836, 2001), was evaluated in a subcutaneous skin infection model with Staphylococcus aureus Smith OC 4172 (methicillin-susceptible S. aureus [MSSA]), S. aureus OC 8525 (MRSA), Pseudomonas aeruginosa OC 4351 (having an inducible AmpC β-lactamase), and P. aeruginosa OC 4354 (overproducing AmpC β-lactamase). In the MSSA and MRSA infection models, ceftobiprole, administered as the prodrug ceftobiprole medocaril, was more effective in reducing CFU/g skin ( P < 0.001) than were cefazolin, vancomycin, or linezolid based on the dose-response profiles. Skin lesion volumes in MSSA-infected animals treated with ceftobiprole were 19 to 29% lower than those for cefazolin-, vancomycin-, or linezolid-treated animals ( P < 0.001). In MRSA infections, lesion size in ceftobiprole-treated mice was 34% less than that with cefazolin or linezolid treatment ( P < 0.001). Against P. aeruginosa , ceftobiprole at similar doses was as effective as meropenem-cilastatin in reductions of CFU/g skin, despite 8- and 32-fold-lower MICs for meropenem; both treatments were more effective than was cefepime ( P < 0.001) against the inducible and overproducing AmpC β-lactamase strains of P. aeruginosa . Ceftobiprole was similar to meropenem-cilastatin and 47 to 54% more effective than cefepime ( P < 0.01) in reducing the size of the lesion caused by either strain of P. aeruginosa in this study. These studies indicate that ceftobiprole is effective in reducing both bacterial load and lesion volume associated with infections due to MSSA, MRSA, and P. aeruginosa in this murine model of skin and soft tissue infection.

A novel series of 7-(1,2,3,4-tetrahydropyrrolo[1,2- a ]pyrazin-7-yl) quinolones has been designed and synthesized in which the heterocyclic side chain is attached to the quinolone core through a carbon–carbon linkage. The antibacterial activity of the compounds was determined against a panel of Gram-positive and Gram-negative pathogens. Compounds 1b and 1e , bearing an 8-methoxy group as well as unsubstituted and (3 S )-methyl substituted 1,2,3,4-tetrahydropyrrolo[1,2- a ]pyrazin-7-yl side chains, respectively, demonstrated notable activity against ciprofloxacin-resistant clinical isolates of Streptococcus pneumoniae .

MurF catalyzes the last cytoplasmic step of bacterial cell wall synthesis and is essential for bacterial survival. Our previous studies used a pharmacophore model of a MurF inhibitor to identify additional inhibitors with improved properties. We now present the characterization of two such inhibitors, the diarylquinolines DQ1 and DQ2. DQ1 inhibited Escherichia coli MurF (50% inhibitory concentration, 24 μM) and had modest activity (MICs, 8 to 16 μg/ml) against lipopolysaccharide (LPS)-defective E. coli and wild-type E. coli rendered permeable with polymyxin B nonapeptide. DQ2 additionally displayed activity against gram-positive bacteria (MICs, 8 to 16 μg/ml), including methicillin (meticillin)-susceptible and -resistant Staphylococcus aureus isolates and vancomycin-susceptible and -resistant Enterococcus faecalis and Enterococcus faecium isolates. Treatment of LPS-defective E. coli cells with ≥2× MIC of DQ1 resulted in a 75-fold-greater accumulation of the MurF substrate compared to the control, a 70% decline in the amount of the MurF product, and eventual cell lysis, consistent with the inhibition of MurF within bacteria. DQ2 treatment of S. aureus resulted in similar effects on the MurF substrate and product quantities. At lower levels of DQ1 (≤1× MIC), the level of accumulation of the substrate was less pronounced (15-fold greater compared to the amount for the control). However, a 50% increase in the amount of the MurF product compared to the control was reproducibly observed, consistent with the possible upregulation of muropeptide biosynthesis upon partial inhibition of this pathway. The overexpression of cloned MurF appeared to partly alleviate the DQ1-mediated inhibition of muropeptide synthesis. The identification of MurF inhibitors such as DQ1 and DQ2 that disrupt cell wall biosynthesis suggests that MurF remains a viable target for an antibacterial agent.

RWJ-416457 is an investigational pyrrolopyrazolyl-substituted oxazolidinone with activity against antibiotic-susceptible and -resistant gram-positive pathogens. Efficacies of RWJ-416457, linezolid, and vancomycin against methicillin-susceptible Staphylococcus aureus (MSSA) and community-associated methicillin-resistant S. aureus (CA-MRSA) in murine skin and systemic infections were compared, as were efficacies against Streptococcus pneumoniae in a lower respiratory infection. In staphylococcal systemic infections, RWJ-416457 was equipotent with to twofold more potent than linezolid, with 50% effective dose values ranging from 1.5 to 5 mg/kg of body weight/day. RWJ-416457 was two- to fourfold less potent than vancomycin against MSSA but up to fourfold more potent than vancomycin against CA-MRSA. In MSSA and CA-MRSA skin infections, RWJ-416457 demonstrated an efficacy similar to that of linezolid, reducing CFU/g skin approximately 1.0 log 10 at all doses tested; vancomycin yielded greater reductions than the oxazolidinones, with decreases in CFU/g skin of 3 log 10 (MSSA) and 2 log 10 (CA-MRSA). In the pneumococcal model, RWJ-416457 was two- to fourfold more potent than linezolid. The free-drug area under the concentration-time curves at 24 h ( f AUC 24 ) were similar for RWJ-416457 and linezolid. The half-life of RWJ-416457 was up to threefold longer than that of linezolid for all routes of administration. The f AUC 24 /MIC ratio, the pharmacodynamic parameter considered predictive of oxazolidinone efficacy, was approximately twofold greater for RWJ-416457 than for linezolid. Since the f AUC values were similar for both compounds, the higher f AUC/MIC ratios of RWJ-416457 appear to result from its greater in vitro potency. These results demonstrate that RWJ-416457 is a promising new oxazolidinone with efficacy in S. aureus or S. pneumoniae mouse infection models.

Department of Biology, Indiana University, Bloomington, IN, USA; Lahey Clinic, Burlington, MA, USA; University of L’Aquila, L’Aquila, Italy; Louis Stokes Cleveland Department of Veteran Affairs Medical Center, Cleveland, OH, USA; Infectious Disease Discovery Research, Wyeth Research, Pearl River, NY, USA; Department of Pathology, University of Verona, Italy; Hospital Universitari Mutua de Terrassa, Barcelona, Spain; Athens Medical School, Athens, Greece; Assistance Publique-Hopitaux de Paris, France; Service of Microbiology, University Hospital Marques de Valdecilla, Santander, Spain; Laboratory of Bacteriology, Hellenic Pasteur Institute, Athens, Greece; Department of Pharmacology, Baylor College of Medicine, Houston, TX, USA; School of Medicine, Sevilla, Spain; Hospital Universitario Virgen Macarena, Seville, Spain; Dipartimento di Biologia Molecolare, Universita di Siena, Siena, Italy; Laboratoire de Bacteriologie LRMA, Faculte de Medecine UPMC, Paris, France; Department of Microbiology, National School of Public Health, Athens, Greece

By site-directed mutagenesis, TEM-1 beta-lactamase was altered to contain single amino acid changes of E104K, R164S, and E240K, in addition to double changes of E104K/R164S or R164S/E240K and the triple change of E104K/R164S/E240K. Hydrolysis rates for cephaloridine and benzylpenicillin were lowered at least 1 order of magnitude for all enzymes containing R164S substitutions. All mutant enzymes exhibited increased kcat values for beta-lactam antibiotics containing an aminothiazole oxime side chain. Hydrolysis of ceftazidime was most affected, with kcat values increased 3-4 orders of magnitude in all enzymes with the substituted R164S moiety. Km values decreased for all substrates except ceftazidime in the enzymes with multiple mutations. Aztreonam was most affected, with Km values lowered 23-56-fold in the enzymes bearing multiple mutations. When the crystal structures of aztreonam and related monobactams were studied and projected into an active-site model of the PC1 beta-lactamase, it became apparent that the two lysine residues might serve equivalent roles by interacting with the carboxylate of the aminothiazole oxime side chain. Hydrogen-bonding interactions involving the oxime and N7 of the lysine, particularly Lys-104, may also be important in some antibiotics. Ser-164 apparently serves an indirect role, since it is somewhat distant from the active-site cleft.

Seven-valent pneumococcal conjugate vaccine (PCV7) provides protection against invasive pneumococcal disease that extends to unvaccinated populations, such as elderly or immunocompromised adults. PCV7 also reduces incidence of pneumococcal penicillin resistance. In this study, the potential impact of PCV7 on pneumococcal fluoroquinolone resistance was examined.U.S. levofloxacin-resistant isolates (264) from TRUST surveillance studies (1999-2004) were serotyped and quinolone resistance-determining region of parC/E and gyrA/B sequenced. Changes in prevalence of vaccine/nonvaccine serotypes during 2000-2004 and 1999-2004 were analyzed by regression analyses and chi-square trend test.The introduction of PCV7 (2000-2004) did not affect fluoroquinolone resistance prevalence, but mutants with vaccine serotypes declined linearly at -6.6 +/- 0.8% per year (p = 0.003), with concomitant replacement by nonvaccine serotypes; vaccine-related serotypes (6A, 9N, 19A, and 23N) increased (p = 0.04). Differential selection between vaccine and nonvaccine serotypes occurred for mutants containing amino acid substitutions at either ParC Ser79 (p = 0.01) or both ParC Ser79 and GyrA Ser81 (p = 0.04). Among mutants with ParC Ser79 substitutions, vaccine serotypes declined linearly (p = 0.02), whereas nonvaccine serotypes increased (p = 0.04). Additionally, a vaccine-independent effect became apparent during 1999-2004, as the incidence of ParC Ser79 and Asp83 mutations declined in fluoroquinolone-resistant strains, suggesting that these substitutions conferred decreased fitness.PCV7 has led to extensive replacement of vaccine serotypes by nonvaccine serotypes among levofloxacin-resistant pneumococci.

SUMMARY Carbapenemases are β-lactamases with versatile hydrolytic capacities. They have the ability to hydrolyze penicillins, cephalosporins, monobactams, and carbapenems. Bacteria producing these β-lactamases may cause serious infections in which the carbapenemase activity renders many β-lactams ineffective. Carbapenemases are members of the molecular class A, B, and D β-lactamases. Class A and D enzymes have a serine-based hydrolytic mechanism, while class B enzymes are metallo-β-lactamases that contain zinc in the active site. The class A carbapenemase group includes members of the SME, IMI, NMC, GES, and KPC families. Of these, the KPC carbapenemases are the most prevalent, found mostly on plasmids in Klebsiella pneumoniae . The class D carbapenemases consist of OXA-type β-lactamases frequently detected in Acinetobacter baumannii . The metallo-β-lactamases belong to the IMP, VIM, SPM, GIM, and SIM families and have been detected primarily in Pseudomonas aeruginosa ; however, there are increasing numbers of reports worldwide of this group of β-lactamases in the Enterobacteriaceae . This review updates the characteristics, epidemiology, and detection of the carbapenemases found in pathogenic bacteria.

Ceftobiprole is the first of the investigational beta-lactam antibiotics with in vitro activity against methicillin-resistant staphylococci to reach and complete Phase III therapeutic trials. Its antibacterial spectrum includes methicillin-resistant Staphylococcus aureus (MRSA), Enterococcus faecalis, penicillin-resistant streptococci and many Gram-negative pathogens. It has demonstrated in vivo activity against many experimental infections caused by these pathogens. Ceftobiprole has completed Phase III clinical trials for complicated skin and skin structure infections, is being studied in Phase III pneumonia trials and has demonstrated non-inferiority compared with vancomycin in a Phase III complicated skin and skin structure infections trial, resulting in > 90% clinical cures of infections caused by MRSA. Other anti-MRSA beta-lactams in therapeutic clinical trials include the carbapenem CS-023/RO-4908463 and the cephalosporin ceftaroline (PPI-0903). The future of all of these agents will depend on their clinical efficacy, safety and their ability to be accepted as beta-lactams for the reliable treatment of a broad spectrum of infections, including those caused by MRSA.

Because the treatment of inhalational anthrax cannot be studied in human clinical trials, it is necessary to conduct efficacy studies using a rhesus monkey model. However, the half-life of levofloxacin was approximately three times shorter in rhesus monkeys than in humans. Computer simulations to match plasma concentration profile, area under the concentration-time curve (AUC), and time above MIC for a human oral dose of 500 mg levofloxacin once a day identified a dosing regimen in rhesus monkeys that would most closely match human exposure: 15 mg/kg followed by 4 mg/kg administered 12 h later. Approximately 24 h following inhalational exposure to approximately 49 times the 50% lethal doses of Bacillus anthracis (Ames strain), monkeys were treated daily with vehicle, levofloxacin, or ciprofloxacin for 30 days. Ciprofloxacin was administered at 16 mg/kg twice a day. Following the 30-day treatment, monkeys were observed for 70 days. Nine of 10 control monkeys died within 9 days of exposure. No clinical signs were observed in fluoroquinolone-treated monkeys during the 30 treatment days. One monkey died 8 days after levofloxacin treatment, and two monkeys from the ciprofloxacin group died 27 and 36 days posttreatment, respectively. These deaths were probably related to the germination of residual spores. B. anthracis was positively cultured from several tissues from the three fluoroquinolone-treated monkeys that died. MICs of levofloxacin and ciprofloxacin from these cultures were comparable to those from the inoculating strain. These data demonstrate that a humanized dosing regimen of levofloxacin was effective in preventing morbidity and mortality from inhalational anthrax in rhesus monkeys and did not select for resistance.

Imipenem-resistant Serratia marcescens isolates were cultured from a lung transplant patient given multiple antibiotics over several months. The strains expressed SME-3, a β-lactamase of the rare SME carbapenem-hydrolyzing family. SME-3 differed from SME-1 by a single amino acid substitution of tyrosine for histidine at position 105, but the two β-lactamases displayed similar hydrolytic profiles.

A novel series of ketolides containing heteroaryl groups that are linked to the erythronolide ring via a C6-carbazate functionality has been successfully synthesized. Careful modulation of the heteroaryl groups, the length and degree of saturation of the C6-carbazate linker, and the substituents present on each of the carbazate nitrogens led to compounds with potent activity against key bacterial respiratory pathogens. The best analogs of this series had in vitro and in vivo (sc dosing) profiles that were comparable to telithromycin.

A series of pyrrolopyridine-substituted oxazolidinones containing various C-5 acetamide isosteres was synthesized and the structure-antibacterial activity relationships determined against a representative panel of susceptible and resistant Gram-positive bacteria.

The enzyme l , d -carboxypeptidase A is involved in the recycling of bacterial peptidoglycan and is essential in Escherichia coli during stationary phase. By high-throughput screening, we have identified a dithiazoline inhibitor of the enzyme with a 50% inhibitory concentration of 3 μM. The inhibitor appeared to cause lysis of E. coli during stationary phase, behavior that is similar to a previously described deletion mutant of l , d -carboxypeptidase A (M. F. Templin, A. Ursinus, and J.-V. Holtje, EMBO J. 18: 4108-4117, 1999). As much as a one-log drop in CFU in stationary phase was observed upon treatment of E. coli with the inhibitor, and the amount of intracellular tetrapeptide substrate increased by approximately 33%, consistent with inhibition of the enzyme within bacterial cells. Stationary-phase targets such as l , d -carboxypeptidase A are largely underrepresented as targets of the antibiotic armamentarium but provide potential opportunities to interfere with bacterial growth and persistence.

Antimicrobial-modifying resistance enzymes have traditionally been class specific, having coevolved with the antibiotics they inactivate. Fluoroquinolones, antimicrobial agents used extensively in medicine and agriculture, are synthetic and have been considered safe from naturally occurring antimicrobial-modifying enzymes. We describe reduced susceptibility to ciprofloxacin in clinical bacterial isolates conferred by a variant of the gene encoding aminoglycoside acetyltransferase AAC(6′)-Ib. This enzyme reduces the activity of ciprofloxacin by N-acetylation at the amino nitrogen on its piperazinyl substituent. Although approximately 30 variants of this gene have been reported since 1986, the two base-pair changes responsible for the ciprofloxacin modification phenotype are unique to this variant, first reported in 2003 and now widely disseminated. An intense increase in the medical use of ciprofloxacin seems to have been accompanied by a notable development: a single-function resistance enzyme has crossed class boundaries, and is now capable of enzymatically undermining two unrelated antimicrobial agents, one of them fully synthetic.

Carbapenemases are rare in the United States. This is the first report of a United States nosocomial outbreak of pan-resistant Pseudomonas aeruginosa infections due to an integron-borne metallo-β-lactamase, VIM-2. This emergence of carbapenemases on mobile genetic elements in the United States warrants surveillance.

An array of 15-amido substituted erythromycin A compounds was synthesized using a chemobiosynthesis approach. It was found that while the in vitro antibacterial activities of aryl amides were inferior to erythromycin A, substituted benzylamides showed equivalent and in some cases improved activity against the macrolide-resistant strains. The 15-amidoerythromycins represent a new class of antibacterial macrolides.

A new series of antibacterial ketolides is reported, which features the use of a C-6 carbamate for tethering the arylalkyl sidechain to the macrolide core. The best members of this series display in vitro and in vivo activity comparable to telithromycin. Partial epimerization at C-2, unobserved in previously reported ketolides, was noted for this series.

A novel series of oxazolidinones containing a pyrroloaryl substituent was synthesized and screened against a representative panel of susceptible and resistant Gram-positive bacteria. Several members of this series were found to have antibacterial activity comparable to or better than linezolid.