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While the use of long-term macrolide therapy to prevent exacerbations in chronic respiratory diseases is widespread, its impact on the oropharyngeal microbiota and macrolide resistance, and the potential for onward transmission of resistance to close contacts are poorly understood. We determined the effects of long-term exposure to azithromycin or erythromycin on phenotypic and genotypic macrolide resistance within the oropharyngeal microbiome of healthy adults and their close contacts in a randomized, single-blinded, parallel-group trial of 4 weeks of twice-daily oral 400 mg erythromycin ethylsuccinate or twice-daily oral 125 mg azithromycin. Using oropharyngeal swabs collected from 20 index healthy adults and 20 paired close contacts, the oropharyngeal microbial composition and macrolide resistance in streptococci were assessed by 16S rRNA sequencing and antibiotic susceptibility testing of oropharyngeal cultures, respectively, at baseline and weeks 4 and 8 (washout). Targeted quantitative PCR of antibiotic resistance genes was performed to evaluate paired changes in resistance gene levels in index patients and close contacts and to relate the potential transmission of antibiotic resistance. Neither azithromycin nor erythromycin altered oropharyngeal microbiota characteristics significantly. Proportional macrolide resistance in oropharyngeal streptococci increased with both erythromycin and azithromycin, remaining above baseline levels for the azithromycin group at washout. Levels of resistance genes increased significantly with azithromycin[ erm (B) and mef ] and erythromycin ( mef ), returning to baseline levels at washout only for the erythromycin group. We found no evidence of onward transmission of resistance to close contacts, as indicated by the lack of concomitant changes in resistance gene levels detected in close contacts. (This study has been registered with the Australian and New Zealand Clinical Trials Registry under identifier ACTRN12617000278336.).

Antimicrobial resistance in campylobacters has been described worldwide. The emergence of multiresistant isolates, particularly among Campylobacter coli isolates, is concerning. New resistance mechanisms appear frequently, and DNA-sequence-based methods such as whole-genome sequencing (WGS) have become useful tools to monitor their emergence. The genomes of 51 multiresistant French Campylobacter sp. clinical strains from 2018 to 2019 were analyzed to identify associated resistance mechanisms. Analyses of erythromycin-resistant strains revealed 23S rRNA mutations among most of them and two different methyltransferases in 4 strains: Erm(B) and a novel methyltransferase, named Erm(N) here. The erm (B) gene was found in multidrug-resistant genomic islands, whereas erm (N) was inserted within CRISPR arrays of the CRISPR- cas9 operon. Moreover, using PCR screening in erythromycin-resistant strains from our collection, we show that erm (N) was already present in 3 French clinical strains 2 years before its first report in 2018 in Quebec, Canada. Bacterial transformations confirmed that the insertion of erm (N) into a CRISPR- cas9 operon can confer macrolide resistance. Campylobacter species are easily able to adapt to their environment and acquire new resistance mechanisms, and the emergence of methyltransferases in campylobacters in France is a matter of concern in the coming years.

Amphotericin B (AmB) is the antifungal with the strongest fungicidal activity, but its use has several limitations, mainly associated with its toxicity. Although some lipidic and liposomal formulations that present reduced toxicity are available, their price limits their application in developing countries. Flucytosine (5FC) has shown synergistic effect with AmB for treatment of some fungal infections, such as cryptococcosis, but again, its price is a limitation for its use in many regions. In the present work, we aimed to identify new drugs that have a minor effect on Cryptococcus neoformans , reducing its growth in the presence of subinhibitory concentrations of AmB. In the initial screening, we found fourteen drugs that had this pattern. Later, checkerboard assays of selected compounds, such as erythromycin, riluzole, nortriptyline, chenodiol, nisoldipine, promazine, chlorcyclizine, cloperastine, and glimepiride, were performed and all of them confirmed for their synergistic effect (fractional inhibitory concentration index [FICI] < 0.5). Additionally, toxicity of these drugs in combination with AmB was tested in mammalian cells and in zebrafish embryos. Harmless compounds, such as the antibiotic erythromycin, were found to have synergic activity with AmB, not only against C. neoformans but also against some Candida spp., in particular against Candida albicans In parallel, we identified drugs that had antifungal activity against C. neoformans and found 43 drugs that completely inhibited the growth of this fungus, such as ciclopirox and auranofin. Our results expand our knowledge about antifungal compounds and open new perspectives in the treatment of invasive mycosis based on repurposing off-patent drugs., (Copyright © 2020 American Society for Microbiology.)

Although macrolides are known as excellent antibacterials, their medical use has been significantly limited due to the spread of bacterial drug resistance. Therefore, it is necessary to develop new potent macrolides to combat the emergence of drug-resistant pathogens. One of the key steps in rational drug design is the identification of chemical groups that mediate binding of the drug to its target and their subsequent derivatization to strengthen drug-target interactions. In the case of macrolides, a few groups are known to be important for drug binding to the ribosome, such as desosamine. Search for new chemical moieties that improve the interactions of a macrolide with the 70S ribosome might be of crucial importance for the invention of new macrolides. For this purpose, here we studied a classic macrolide, dirithromycin, which has an extended (2-methoxyethoxy)-methyl side chain attached to the C-9/C-11 atoms of the macrolactone ring that can account for strong binding of dirithromycin to the 70S ribosome. By solving the crystal structure of the 70S ribosome in complex with dirithromycin, we found that its side chain interacts with the wall of the nascent peptide exit tunnel in an idiosyncratic fashion: its side chain forms a lone pair-π stacking interaction with the aromatic imidazole ring of the His69 residue in ribosomal protein uL4. To our knowledge, the ability of this side chain to form a contact in the macrolide binding pocket has not been reported previously and potentially can open new avenues for further exploration by medicinal chemists developing next-generation macrolide antibiotics active against resistant pathogens., (Copyright © 2019 American Society for Microbiology.)

The objective of this study was to determine the phenotypic patterns of antibiotic resistance and the epidemiology of drug-resistant Campylobacter spp. from a low-resource setting. A birth cohort of 303 patients was followed until 5 years of age. Stool samples from asymptomatic children ( n = 10,008) and those with diarrhea ( n = 3,175) were cultured for Campylobacter Disk diffusion for ciprofloxacin (CIP), nalidixic acid (NAL), erythromycin (ERY), azithromycin (AZM), tetracycline (TE), gentamicin (GM), ampicillin (AMP), amoxicillin and clavulanic acid (AMC), ceftriaxone (CRO), chloramphenicol (C), and trimethoprim-sulfamethoxazole (TMS) was determined. Antibiotic resistances in Campylobacter jejuni and non- C. jejuni isolates from surveillance and diarrhea samples were compared, and the association between personal macrolide exposure and subsequent occurrence of a macrolide-resistant Campylobacter spp. was assessed. Of 917 Campylobacter isolates, 77.4% of C. jejuni isolates and 79.8% of non- C. jejuni isolates were resistant to ciprofloxacin, while 4.9% of C. jejuni isolates and 24.8% of non- C. jejuni isolates were not susceptible to azithromycin. Of the 303 children, 33.1% had been diagnosed with a Campylobacter strain nonsusceptible to both azithromycin and ciprofloxacin. Personal macrolide exposure did not affect the risk of macrolide-resistant Campylobacter Amoxicillin and clavulanic acid (94.0%) was one of the antibiotics with the highest rates of susceptibility. There is a high incidence of quinolone- and macrolide-resistant Campylobacter infections in infants under 24 months of age. Given the lack of association between personal exposure to macrolides and a subsequent Campylobacter infection resistant to macrolides, there is a need to evaluate the source of multidrug-resistant (MDR) Campylobacter This study provides compelling evidence to propose amoxicillin/clavulanic acid as a treatment for campylobacteriosis., (Copyright © 2019 Schiaffino et al.)

The objective of this study was to determine the prevalence of Rhodococcus equi strains resistant to macrolides and rifampin over time in clinical samples from foals submitted to diagnostic laboratories in central Kentucky. We performed a retrospective observational study of all clinical samples from foals that were submitted to veterinary diagnostic laboratories in Kentucky between January 1995 and December 2017. Samples were included if the R. equi bacterium was cultured and tested for in vitro susceptibility to erythromycin or rifampin. In vitro susceptibility testing to erythromycin was available for 2,169 isolates of R. equi , while susceptibility testing to both erythromycin and rifampin was available for 1,681 isolates. Rifampin resistance was first detected in 2000, and erythromycin resistance was first detected in 2004. Between 1995 and 2006, the proportion of resistant isolates of R. equi was 0.7% for erythromycin and 2.3% for rifampin. There was a significant ( P < 0.001) increase in the proportion of resistant R. equi between 2007 and 2017, with 13.6% of isolates being resistant to erythromycin and 16.1% being resistant to rifampin. Between 2007 and 2017, isolates of R. equi resistant to erythromycin or rifampin were significantly less likely to be isolated from feces than from the respiratory tract, other soft tissues, or musculoskeletal infections. The considerable increase in the prevalence of isolates of R. equi resistant to macrolides and rifampin since 2007 is of concern for both human and animal health., (Copyright © 2018 American Society for Microbiology.)

The emergence of antimicrobial resistance (AMR) in foodborne bacteria is a growing concern worldwide. AMR surveillance is a key element in understanding the implications resulting from the use of antibiotics for therapeutic as well as prophylactic needs. The emergence and spread of AMR in foodborne human pathogens are indirect health hazards. This surveillance study reports the trend and pattern of AMR detected in Vibrio species isolated from molluscs harvested in Canada between 2006 and 2012 against 19 commonly used antibiotics. Five common antibiotics, ampicillin, cephalothin, erythromycin, kanamycin, and streptomycin, predominantly contributed to AMR, including multidrug resistance (MDR) in the molluscan Vibrio spp. isolated in 2006. A prospective follow-up analysis of these drugs showed a declining trend in the frequency of MDR/AMR Vibrio spp. in subsequent years until 2012. The observed decline appears to have been influenced by the specific downturn in resistance to the aminoglycosides, kanamycin, and streptomycin. Frequently observed MDR/AMR Vibrio spp. in seafood is a potential health concern associated with seafood consumption. Our surveillance study provides an indication of the antibiotics that challenged the marine bacteria, sourced to Canadian estuaries, during and/or prior to the study period., (© Crown copyright 2018.)

Non- diphtheriae Corynebacterium -associated disease has been increasingly observed and often presents a conundrum to the treating physician. Analysis of antibiotic susceptibility testing data for 1,970 clinical Corynebacterium isolates received between 2011 and 2016 revealed that empirical drug treatment options are limited to vancomycin and linezolid. Corynebacterium striatum was the most frequently observed species during this study period, along with C. amycolatum and C. pseudodiphtheriticum/C. propinquum Low levels of susceptibility to penicillin (14.5%), erythromycin (15.1%), and clindamycin (8.7%) were observed for non- diphtheriae Corynebacterium species, while 3.0% of isolates were not susceptible to daptomycin. Similarly, 26.9% and 38.1% of Corynebacterium isolates were susceptible to ciprofloxacin and trimethoprim-sulfamethoxazole, respectively. Our data show much lower susceptibility to penicillin than previously reported in the literature and an increasing number of isolates resistant to daptomycin, highlighting the need for continued antibiotic surveillance studies for appropriate patient management and treatment success., (© Crown copyright 2018.)

TetR21 controls the expression of Tet38 and LmrS efflux pumps. A tetR21 mutant, QT21, exhibited a 4-fold increase in the transcription level of lmrS Staphylococcus aureus lmrS overexpressor showed increases of 4-fold and 2-fold, respectively, in the MICs of chloramphenicol and erythromycin, while the MICs of lmrS mutant QT18 and lmrS-tetR21 mutant QT1821 remained similar to those of parental strain RN6390. TetR21 does not bind to the promoter of lmrS , suggesting indirect regulation of lmrS ., (Copyright © 2017 American Society for Microbiology.)

The tet39 tetracycline resistance determinant and the macrolide resistance genes msrE and mphE were found in an 18.2-kb plasmid, pS30-1, recovered from a global clone 2 (GC2) Acinetobacter baumannii isolate from Singapore, that conferred resistance to tetracycline and erythromycin. pS30-1 also contains mobA and mobC genes encoding MOB Q family proteins, but attempts to mobilize pS30-1 utilizing a coresident conjugative repAci6 plasmid were unsuccessful. Eight p dif sites, consisting of inversely oriented binding sites for the XerC and XerD recombinases separated by 6 bp, were detected in pS30-1. The tet39 determinant and the msrE-mphE gene pair are each surrounded by two p dif sites in inverse orientation. Identical regions in different contexts and many previously unnoticed p dif sites were found in a number of different plasmids in GenBank, showing that the tet39 and msrE-mphE dif modules are mobile. A putative toxin/antitoxin system, a gene encoding a serine recombinase, and open reading frames of unknown function were also part of dif modules in pS30-1. In general, modules with internal XerC or XerD sites alternate. Two copies of IS Ajo2-1 (94% identical to IS Ajo2 ) in pS30-1 were inserted 5 bp from a XerC site, and this appears to be the preferred insertion site for this insertion sequence (IS) group. Apparently, Acinetobacter plasmids exploit the Acinetobacter XerC-XerD recombinases to mobilize DNA units containing resistance and other genes, via an uncharacterized mechanism. The tet39 and msrE-mphE dif modules add to the oxa24 module and the oxa58 module redefined here, bringing the total of resistance gene-containing dif modules in Acinetobacter plasmids to four., (Copyright © 2017 American Society for Microbiology.)

The SmeDEF pump of Stenotrophomonas maltophilia is negatively regulated by SmeT. In this study, strains KJΔT ( smeT deletion mutant) and KJT-D m (mutant with a defective SmeT-binding site) showed increased resistance to chloramphenicol/nalidixic acid/macrolides and susceptibility to aminoglycoside. Overexpression of the SmeDEF pump, in either KJΔT or KJT-D m , downregulated smeYZ expression, which is responsible for the reduced aminoglycoside resistance. Furthermore, the SmeRySy two-component regulatory system was downregulated in response to SmeDEF overexpression, which supports its involvement in the regulatory circuit., (Copyright © 2017 American Society for Microbiology.)

Monitoring the emergence of antibiotic resistance is a recent issue in the treatment of Legionnaires' disease. Macrolides are recommended as first-line therapy, but resistance mechanisms have not been studied in Legionella species. Our aim was to determine the molecular basis of macrolide resistance in L. pneumophila Twelve independent lineages from a common susceptible L. pneumophila ancestral strain were propagated under conditions of erythromycin or azithromycin pressure to produce high-level macrolide resistance. Whole-genome sequencing was performed on 12 selected clones, and we investigated mutations common to all lineages. We reconstructed the dynamics of mutation for each lineage and demonstrated their involvement in decreased susceptibility to macrolides. The resistant mutants were produced in a limited number of passages to obtain a 4,096-fold increase in erythromycin MICs. Mutations affected highly conserved 5-amino-acid regions of L4 and L22 ribosomal proteins and of domain V of 23S rRNA (G2057, A2058, A2059, and C2611 nucleotides). The early mechanisms mainly affected L4 and L22 proteins and induced a 32-fold increase in the MICs of the selector drug. Additional mutations related to 23S rRNA mostly occurred later and were responsible for a major increase of macrolide MICs, depending on the mutated nucleotide, the substitution, and the number of mutated genes among the three rrl copies. The major mechanisms of the decreased susceptibility to macrolides in L. pneumophila and their dynamics were determined. The results showed that macrolide resistance could be easily selected in L. pneumophila and warrant further investigations in both clinical and environmental settings., (Copyright © 2017 American Society for Microbiology.)

Lefamulin, an investigational pleuromutilin, was tested against a collection of 18 macrolide-susceptible and 42 macrolide-resistant Mycoplasma pneumoniae strains, and the results were compared with those of azithromycin, erythromycin, tetracycline, doxycycline, and moxifloxacin testing. Lefamulin was highly active against all strains tested, with all MICs at ≤0.008 μg/ml. The lefamulin MIC 90 (0.002 μg/ml) for macrolide-resistant strains was the lowest among all drugs tested. Minimum bactericidal concentrations were within 2 dilutions of the MIC values, indicating a bactericidal effect., (Copyright © 2017 American Society for Microbiology.)

In Gram-negative bacteria, a synergistic relationship between slow passive uptake of antibiotics across the outer membrane and active efflux transporters creates a permeability barrier, which efficiently reduces the effective concentrations of antibiotics in cells and, hence, their activities. To analyze the relative contributions of active efflux and the passive barrier to the activities of antibiotics, we constructed Escherichia coli strains with controllable permeability of the outer membrane. The strains expressed a large pore that does not discriminate between compounds on the basis of their hydrophilicity and sensitizes cells to a variety of antibacterial agents. We found that the efficacies of antibiotics in these strains were specifically affected by either active efflux or slow uptake, or both, and reflect differences in the properties of the outer membrane barrier, the repertoire of efflux pumps, and the inhibitory activities of antibiotics. Our results identify antibiotics which are the best candidates for the potentiation of activities through efflux inhibition and permeabilization of the outer membrane., (Copyright © 2016, American Society for Microbiology. All Rights Reserved.)

Members of the Erm methyltransferase family modify 23S rRNA of the bacterial ribosome and render cross-resistance to macrolides and multiple distantly related antibiotics. Previous studies have shown that the expression of erm is activated when a macrolide-bound ribosome stalls the translation of the leader peptide preceding the cotranscribed erm Ribosome stalling is thought to destabilize the inhibitory stem-loop mRNA structure and exposes the erm Shine-Dalgarno (SD) sequence for translational initiation. Paradoxically, mutations that abolish ribosome stalling are routinely found in hyper-resistant clinical isolates; however, the significance of the stalling-dead leader sequence is largely unknown. Here, we show that nonsense mutations in the Staphylococcus aureus ErmB leader peptide (ErmBL) lead to high basal and induced expression of downstream ErmB in the absence or presence of macrolide concomitantly with elevated ribosome methylation and resistance. The overexpression of ErmB is associated with the reduced turnover of the ermBL-ermB transcript, and the macrolide appears to mitigate mRNA cleavage at a site immediately downstream of the ermBL SD sequence. The stabilizing effect of antibiotics on mRNA is not limited to ermBL-ermB; cationic antibiotics representing a ribosome-stalling inducer and a noninducer increase the half-life of specific transcripts. These data unveil a new layer of ermB regulation and imply that ErmBL translation or ribosome stalling serves as a "tuner" to suppress aberrant production of ErmB because methylated ribosome may impose a fitness cost on the bacterium as a result of misregulated translation., (Copyright © 2016 Dzyubak and Yap.)

The envelope of Gram-negative bacteria constitutes an impenetrable barrier to numerous classes of antimicrobials. This intrinsic resistance, coupled with acquired multidrug resistance, has drastically limited the treatment options against Gram-negative pathogens. The aim of the present study was to develop and validate an assay for identifying compounds that increase envelope permeability, thereby conferring antimicrobial susceptibility by weakening of the cell envelope barrier in Gram-negative bacteria. A high-throughput whole-cell screening platform was developed to measure Escherichia coli envelope permeability to a β-galactosidase chromogenic substrate. The signal produced by cytoplasmic β-galactosidase-dependent cleavage of the chromogenic substrate was used to determine the degree of envelope permeabilization. The assay was optimized by using known envelope-permeabilizing compounds and E. coli gene deletion mutants with impaired envelope integrity. As a proof of concept, a compound library comprising 36 peptides and 45 peptidomimetics was screened, leading to identification of two peptides that substantially increased envelope permeability. Compound 79 reduced significantly (from 8- to 125-fold) the MICs of erythromycin, fusidic acid, novobiocin and rifampin and displayed synergy (fractional inhibitory concentration index, <0.2) with these antibiotics by checkerboard assays in two genetically distinct E. coli strains, including the high-risk multidrug-resistant, CTX-M-15-producing sequence type 131 clone. Notably, in the presence of 0.25 μM of this peptide, both strains were susceptible to rifampin according to the resistance breakpoints (R > 0.5 μg/ml) for Gram-positive bacterial pathogens. The high-throughput screening platform developed in this study can be applied to accelerate the discovery of antimicrobial helper drug candidates and targets that enhance the delivery of existing antibiotics by impairing envelope integrity in Gram-negative bacteria., (Copyright © 2016, American Society for Microbiology. All Rights Reserved.)

MICs of 25 Abiotrophia defectiva and 109 Granulicatella adiacens isolates were determined by broth microdilution. Using CLSI breakpoints, the susceptibilities of A. defectiva and G. adiacens isolates were, respectively, 24% and 34% to penicillin, 92% and 22% to ceftriaxone, 48% and 3% to cefepime, 72% and 87% to meropenem, 92% and 10% to cefotaxime, 100% and 97% to levofloxacin, 92% and 80% to clindamycin, and 24% and 50% to erythromycin. All isolates were susceptible to vancomycin. In the penicillin-susceptible subgroup, all A. defectiva isolates were susceptible to ceftriaxone; however, 62% of G. adiacens isolates were ceftriaxone nonsusceptible., (Copyright © 2016, American Society for Microbiology. All Rights Reserved.)

Efflux-mediated macrolide resistance due to mef(E) and mel, carried by the mega element, is common in Streptococcus pneumoniae, for which it was originally characterized, but it is rare in Streptococcus pyogenes In S. pyogenes, mega was previously found to be enclosed in Tn2009, a composite genetic element of the Tn916 family containing tet(M) and conferring erythromycin and tetracycline resistance. In this study, S. pyogenes isolates containing mef(E), apparently not associated with other resistance determinants, were examined to characterize the genetic context of mega. By whole-genome sequencing of one isolate, MB56Spyo009, we identified a novel composite integrative and conjugative element (ICE) carrying mega, designated ICESpy009, belonging to the ICESa2603 family. ICESpy009 was 55 kb long, contained 61 putative open reading frames (ORFs), and was found to be integrated into hylA, a novel integration site for the ICESa2603 family. The modular organization of the ICE was similar to that of members of the ICESa2603 family carried by different streptococcal species. In addition, a novel cluster of accessory resistance genes was found inside a region that encloses mega. PCR mapping targeting ICESpy009 revealed the presence of a similar ICE in five other isolates under study. While in three isolates the integration site was the same as that of ICESpy009, in two isolates the ICE was integrated into rplL, the typical integration site of the ICESa2603 family. ICESpy009 was able to transfer macrolide resistance by conjugation to both S. pyogenes and S. pneumoniae, showing the first evidence of the transferability of mega from S. pyogenes., (Copyright © 2016, American Society for Microbiology. All Rights Reserved.)

Lefamulin was evaluated against various Streptococcus pneumoniae serotypes that were collected from adults with lower respiratory tract infections. Lefamulin exhibited MIC50 and MIC90 values of 0.12 and 0.25 μg/ml, respectively, against the entire collection (n = 822). Similar results were obtained for lefamulin against each of the most common serotypes as well as against multidrug-resistant isolates and strains that are nonsusceptible to ceftriaxone or erythromycin. These data support the clinical development of lefamulin for the treatment of community-acquired respiratory tract infections., (Copyright © 2016, American Society for Microbiology. All Rights Reserved.)

We used classical mutagens in Gram-negative Escherichia coli to study synergies with different classes of antibiotics, test models of antibiotic mechanisms of action, and examine the basis of synergy. We used 4-nitroquinoline 1-oxide (4NQO), zebularine (ZEB), 5-azacytidine (5AZ), 2-aminopurine (2AP), and 5-bromodeoxyuridine (5BrdU) as mutagens (with bactericidal potency of 4NQO > ZEB > 5AZ > 2AP > 5BrdU) and vancomycin (VAN), ciprofloxacin (CPR), trimethoprim (TMP), gentamicin (GEN), tetracycline (TET), erythromycin (ERY), and chloramphenicol (CHL) as antibiotics. We detected the strongest synergies with 4NQO, an agent that oxidizes guanines and ultimately results in double-strand breaks when paired with the bactericidal antibiotics VAN, TMP, CPR, and GEN, but no synergies with the bacteriostatic antibiotics TET, ERY, and CHL. Each of the other mutagens displays synergies with the bactericidal antibiotics to various degrees that reflect their potencies, as well as with some of the other mutagens. The results support recent models showing that bactericidal antibiotics kill bacteria principally by ultimately generating more double-strand breaks than can be repaired. We discuss the synergies seen here and elsewhere as representing dose effects of not the proximal target damage but rather the ultimate resulting double-strand breaks. We also used the results of pairwise tests to place the classic mutagens into functional antibacterial categories within a previously defined drug interaction network., (Copyright © 2016, American Society for Microbiology. All Rights Reserved.)

Antibiotic resistance in Ureaplasma urealyticum/Ureaplasma parvum and Mycoplasma hominis is an issue of increasing importance. However, data regarding the susceptibility and, more importantly, the clonality of these organisms are limited. We analyzed 140 genital samples obtained in Bern, Switzerland, in 2014. Identification and antimicrobial susceptibility tests were performed by using the Mycoplasma IST 2 kit and sequencing of 16S rRNA genes. MICs for ciprofloxacin and azithromycin were obtained in broth microdilution assays. Clonality was analyzed with PCR-based subtyping and multilocus sequence typing (MLST), whereas quinolone resistance and macrolide resistance were studied by sequencing gyrA, gyrB, parC, and parE genes, as well as 23S rRNA genes and genes encoding L4/L22 ribosomal proteins. A total of 103 samples were confirmed as positive for U. urealyticum/U. parvum, whereas 21 were positive for both U. urealyticum/U. parvum and M. hominis. According to the IST 2 kit, the rates of nonsusceptibility were highest for ciprofloxacin (19.4%) and ofloxacin (9.7%), whereas low rates were observed for clarithromycin (4.9%), erythromycin (1.9%), and azithromycin (1%). However, inconsistent results between microdilution and IST 2 kit assays were recorded. Various sequence types (STs) observed previously in China (ST1, ST2, ST4, ST9, ST22, and ST47), as well as eight novel lineages, were detected. Only some quinolone-resistant isolates had amino acid substitutions in ParC (Ser83Leu in U. parvum of serovar 6) and ParE (Val417Thr in U. parvum of serovar 1 and the novel Thr417Val substitution in U. urealyticum). Isolates with mutations in 23S rRNA or substitutions in L4/L22 were not detected. This is the first study analyzing the susceptibility of U. urealyticum/U. parvum isolates in Switzerland and the clonality outside China. Resistance rates were low compared to those in other countries. We hypothesize that some hyperepidemic STs spread worldwide via sexual intercourse. Large combined microbiological and clinical studies should address this important issue., (Copyright © 2015, American Society for Microbiology. All Rights Reserved.)

Oxidants were shown to contribute to the lethality of bactericidal antibiotics in different bacterial species, including the laboratory strain Streptococcus pneumoniae R6. Resistance to penicillin among S. pneumoniae R6 mutants was further shown to protect against the induction of oxidants upon exposure to unrelated bactericidal compounds. In the work described here, we expanded on these results by studying the accumulation of reactive oxygen species in the context of antibiotic sensitivity and resistance by including S. pneumoniae clinical isolates. In S. pneumoniae R6, penicillin, ciprofloxacin, and kanamycin but not the bacteriostatic linezolid, erythromycin, or tetracycline induced the accumulation of reactive oxygen species. For the three bactericidal compounds, resistance to a single molecule prevented the accumulation of oxidants upon exposure to unrelated bactericidal antibiotics, and this was accompanied by a reduced lethality. This phenomenon does not involve target site mutations but most likely implicates additional mutations occurring early during the selection of resistance to increase survival while more efficient resistance mechanisms are being selected or acquired. Bactericidal antibiotics also induced oxidants in sensitive S. pneumoniae clinical isolates. The importance of oxidants in the lethality of bactericidal antibiotics was less clear than for S. pneumoniae R6, however, since ciprofloxacin induced oxidants even in ciprofloxacin-resistant S. pneumoniae clinical isolates. Our results provide a clear example of the complex nature of the mode of action of antibiotics. The adaptive approach to oxidative stress of S. pneumoniae is peculiar, and a better understanding of the mechanism implicated in response to oxidative injury should also help clarify the role of oxidants induced by antibiotics., (Copyright © 2015, American Society for Microbiology. All Rights Reserved.)

The macrolide resistance gene erm(T) was identified for the first time in a porcine Erysipelothrix rhusiopathiae isolate from swine in China. The novel 3,749-bp small plasmid pER29, which carries erm(T), had a G+C content of 31% and four distinct open reading frames. The presence of pER29 increased by at least 128-fold the MICs of clindamycin and erythromycin for E. rhusiopathiae. The fitness cost of pER29 could be responsible for the low frequency of erm(T) in E. rhusiopathiae., (Copyright © 2015, American Society for Microbiology. All Rights Reserved.)

Mycoplasma pneumoniae is a major pathogen causing community-acquired pneumoniae (CAP), which is generally treated with macrolides. In recent years, however, although macrolide-resistant M. pneumoniae has been reported frequently, particularly in China, very little is known about the prevalence of macrolide-resistant M. pneumoniae infection in adults. In this study, we survey the macrolide-resistant M. pneumoniae in adults in Zhejiang province and characterize the mechanisms of resistance to macrolide. Six hundred fifty throat swab samples were collected from adult patients with CAP from January 2012 to August 2014. These samples were assayed by nested PCR and then cultivated for M. pneumoniae. All isolates were sequenced to determine the mutation in domain V of the 23S rRNA gene. The activities of 10 antibiotics against macrolide-resistant M. pneumoniae isolates were also investigated in vitro. Moreover, restriction fragment length polymorphism (RFLP) analysis of the amplified P1 gene was used to type 50 resistant strains. One hundred percent (71/71) of M. pneumoniae strains isolated from adults with CAP were resistant to erythromycin (MIC=128 to >256 μg/ml), clarithromycin (MIC=128 to >256 μg/ml), and azithromycin (MIC=32 to >64 μg/ml). Furthermore, all macrolide-resistant M. pneumoniae strains identified had an A2063G mutation in domain V of the 23S rRNA gene. Forty-six resistant strains (92.0%) were classified into type I strain on the basis of P1 gene PCR-RFLP analysis. According to these findings, it is suggested that macrolide-resistant M. pneumoniae infection is very prevalence among adults in Zhejiang province. Thus, there is necessary to perform the epidemiological monitoring of macrolide-resistant M. pneumoniae in the future., (Copyright © 2015, American Society for Microbiology. All Rights Reserved.)

We studied the molecular mechanisms of linezolid resistance in 9 isolates of toxigenic Clostridium difficile with high linezolid MICs. The activity of linezolid was determined against 891 clinical isolates of toxigenic C. difficile. The MIC50 and MIC90 of linezolid were 0.75 μg/ml and 1.5 μg/ml, respectively. Nine strains (1%) showed high linezolid MICs (6 μg/ml to 16 μg/ml) and also were resistant to clindamycin, erythromycin, and chloramphenicol. These strains were selected for molecular studies: sequencing of domain V of the 23 rRNA gene, detection of the cfr methyltransferase gene, and sequencing of the ribosomal protein genes rplC and rplD. Molecular relatedness between strains was assessed using PCR ribotyping and MLVA (multilocus variable-number tandem-repeat analysis) typing. The strains belonged to ribotypes 001 (2/9), 017 (6/9), and 078 (1/9). MLVA showed that strains of ribotype 001 and 017 belonged to the same clonal complex in each ribotype. We did not detect mutations in the 23S rRNA gene. The cfr gene was detected in 7 of 9 strains. Sequencing of cfr amplicons revealed a similarity of 100% to a fragment of transposon Tn6218 of C. difficile, which was annotated as a putative chloramphenicol/florfenicol resistance protein. We were unable to detect mechanisms of resistance to linezolid in the 2 strains belonging to ribotype 001. While the relevance of our results lies in the detection of the cfr gene as a possible mechanism of resistance to linezolid in C. difficile, our findings should be assessed by further investigations to characterize these possible cfr genes and their contribution to linezolid resistance., (Copyright © 2015, American Society for Microbiology. All Rights Reserved.)

Acinetobacter baumannii contains RND-family efflux systems AdeABC and AdeIJK, which pump out a wide range of antimicrobial compounds, as judged from the MIC changes occurring upon deletion of the responsible genes. However, these studies may miss changes because of the high backgrounds generated by the remaining pumps and by β-lactamases, and it is unclear how the activities of these pumps compare quantitatively with those of the well-studied AcrAB-TolC system of Escherichia coli. We expressed adeABC and adeIJK of A. baumannii, as well as E. coli acrAB, in an E. coli host from which acrAB was deleted. The A. baumannii pumps were functional in E. coli, and the MIC changes that were observed largely confirmed the substrate range already reported, with important differences. Thus, the AdeABC system pumped out all β-lactams, an activity that was often missed in deletion studies. When the expression level of the pump genes was adjusted to a similar level for a comparison with AcrAB-TolC, we found that both A. baumannii efflux systems pumped out a wide range of compounds, but AdeABC was less effective than AcrAB-TolC in the extrusion of lipophilic β-lactams, novobiocin, and ethidium bromide, although it was more effective at tetracycline efflux. AdeIJK was remarkably more effective than a similar level of AcrAB-TolC in the efflux of β-lactams, novobiocin, and ethidium bromide, although it was less so in the efflux of erythromycin. These results thus allow us to compare these efflux systems on a quantitative basis, if we can assume that the heterologous systems are fully functional in the E. coli host., (Copyright © 2014, American Society for Microbiology. All Rights Reserved.)

In this study, global intra- and extracellular metabolic profiles were exploited to investigate the impact of antibiotic compounds with different cellular targets on the metabolome of Staphylococcus aureus HG001. Primary metabolism was largely covered, yet uncommon staphylococcal metabolites were detected in the cytosol of S. aureus, including sedoheptulose-1,7-bisphosphate and the UDP-MurNAc-pentapeptide with an alanine-seryl residue. By comparing the metabolic profiles of unstressed and stressed staphylococcal cells in a time-dependent manner, we found far-ranging effects within the metabolome. For each antibiotic compound, accumulation as well as depletion of metabolites was detected, often comprising whole biosynthetic pathways, such as central carbon and amino acid metabolism and peptidoglycan, purine, and pyrimidine synthesis. Ciprofloxacin altered the pool of (deoxy)nucleotides as well as peptidoglycan precursors, thus linking stalled DNA and cell wall synthesis. Erythromycin tended to increase the amounts of intermediates of the pentose phosphate pathway and lysine. Fosfomycin inhibited the first enzymatic step of peptidoglycan synthesis, which was followed by decreased levels of peptidoglycan precursors but enhanced levels of substrates such as UDP-GlcNAc and alanine-alanine. In contrast, vancomycin and ampicillin inhibited the last stage of peptidoglycan construction on the outer cell surface. As a result, the amounts of UDP-MurNAc-peptides drastically increased, resulting in morphological alterations in the septal region and in an overall decrease in central metabolite levels. Moreover, each antibiotic affected intracellular levels of tricarboxylic acid cycle intermediates., (Copyright © 2014, American Society for Microbiology. All Rights Reserved.)

Among 1,827 group B Streptococcus (GBS) strains collected between 2006 and 2013 by the French National Reference Center for Streptococci, 490 (26.8%) strains were erythromycin resistant. The erm(T) resistance gene was found in six strains belonging to capsular polysaccharides Ia, III, and V and was carried by the same mobilizable plasmid, which could be efficiently transferred by mobilization to GBS and Enterococcus faecalis recipients, thus promoting a broad dissemination of erm(T)., (Copyright © 2014, American Society for Microbiology. All Rights Reserved.)

Strains of Neisseria gonorrhoeae with mosaic penA genes bearing novel point mutations in penA have been isolated from ceftriaxone treatment failures. Such isolates exhibit significantly higher MIC values to third-generation cephalosporins. Here we report the in vitro isolation of two mutants with elevated MICs to cephalosporins. The first possesses a point mutation in the transpeptidase region of the mosaic penA gene, and the second contains an insertion mutation in pilQ., (Copyright © 2014, American Society for Microbiology. All Rights Reserved.)

Ketolides, the third generation of expanded-spectrum macrolides, have in the last years become a successful weapon in the endless war against macrolide-resistant pathogens. Ketolides are semisynthetic derivatives of the naturally produced macrolide erythromycin, displaying not only improved activity against some erythromycin-resistant strains but also increased bactericidal activity as well as inhibitory effects at lower drug concentrations. In this study, we present a series of novel ketolides carrying alkyl-aryl side chains at the C-6 position of the lactone ring and, additionally, one or two fluorine atoms attached either directly to the lactone ring at the C-2 position or indirectly via the C-13 position. According to our genetic and biochemical studies, these novel ketolides occupy the known macrolide binding site at the entrance of the ribosomal tunnel and exhibit lower MIC values against wild-type or mutant strains than erythromycin. In most cases, the ketolides display activities comparable to or better than the clinically used ketolide telithromycin. Chemical protection experiments using Escherichia coli ribosomes bearing U2609C or U754A mutations in 23S rRNA suggest that the alkyl-aryl side chain establishes an interaction with the U2609-A752 base pair, analogous to that observed with telithromycin but unlike the interactions formed by cethromycin. These findings reemphasize the versatility of the alkyl-aryl side chains with respect to species specificity, which will be important for future design of improved antimicrobial agents.

Streptococcus pneumoniae isolates (6,958) were collected from patients at 163 U.S. medical centers during 2009 through 2012. Isolates were evaluated for multidrug resistance (MDR) to penicillin, ceftriaxone, erythromycin, tetracycline, trimethoprim-sulfamethoxazole, and levofloxacin. Ceftaroline was 16-fold more potent than ceftriaxone (MIC50/MIC90, ≤0.25/2 μg/ml) against all isolates. For MDR isolates (35.2% of tested strains), ceftaroline (MIC50/MIC90, 0.06/0.25 μg/ml; 100.0% susceptible) was the most active agent tested, being 8-fold more potent than ceftriaxone (MIC50/MIC90, 0.5/2 μg/ml) and 16-fold more potent than penicillin (MIC50/MIC90, 1/4 μg/ml).

The in vitro antibacterial activity of solithromycin (CEM-101) against macrolide-resistant isolates (n=62) of Streptococcus agalactiae (group B streptococcus [GBS]) was determined. Phenotypic characterization of macrolide-resistant strains was performed by double-disc diffusion testing. A multiplex PCR was used to identify the erm(B), erm(TR), and mef(A/E) genes, capsular genotypes, and alpha-like (Alp) protein genes from the GBS strains. Determination of MIC was carried out using the microdilution broth method. The Etest method was used for penicillin, azithromycin, clarithromycin, and erythromycin. Solithromycin had a MIC50 of ≤0.008 μg/ml and a MIC90 of 0.015 μg/ml against macrolide-susceptible S. agalactiae. These MICs were lower than those displayed by penicillin (MIC50 of 0.032 μg/ml and MIC90 of 0.047 μg/ml), the antibiotic agent of choice for prophylaxis and treatment of GBS infections. Against macrolide-resistant S. agalactiae, solithromycin had a MIC50 of 0.03 μg/ml and a MIC90 of 0.125 μg/ml. Against erm(B) strains, solithromycin had a MIC50 of 0.03 μg/ml and a MIC90 of 0.06 μg/ml, while against mef(A) strains, it had a MIC50 of 0.03 μg/ml and a MIC90 of 0.125 μg/ml. Most erythromycin-resistant GBS strains were of serotype V (64.5%) and associated significantly with alp2-3. Moreover, a statistically significant association was observed between the constitutive macrolide-lincosamide-streptogramin B resistance (cMLSB) phenotype and the erm(B) gene-carrying strains, the alp2-3 gene and the M phenotype, and the mef(A/E) gene and epsilon. Overall, our results show that solithromycin had lower or similar MICs than penicillin and potent activity against macrolide-resistant strains independent of their genotype or phenotype, representing a valid therapeutic alternative where β-lactams cannot be used.

We report the complete nucleotide sequence and analysis of pETBTY825, a Staphylococcus aureus TY825 plasmid encoding exfoliative toxin B (ETB). S. aureus TY825 is a clinical isolate obtained from an impetigo patient in 2002. The size of pETBTY825, 60.6 kbp, was unexpectedly larger than that of the archetype pETBTY4 (∼30 kbp). Genomic comparison of the plasmids shows that pETBTY825 has the archetype pETBTY4 as the backbone and has a single large extra DNA region of 22.4 kbp. The extra DNA region contains genes for resistance to aminoglycoside [aac(6')/aph(2″)], macrolide (msrA), and penicillin (blaZ). A plasmid deletion experiment indicated that these three resistance elements were functionally active. We retrospectively examined the resistance profile of the clinical ETB-producing S. aureus strains isolated in 1977 to 2007 using a MIC determination with gentamicin (GM), arbekacin (ABK), and erythromycin (EM) and by PCR analyses for aac(6')/aph(2″) and msrA using purified plasmid preparations. The ETB-producing S. aureus strains began to display high resistance to GM, which was parallel with the detection of aac(6')/aph(2″) and mecA, after 1990. Conversely, there was no significant change in the ABK MIC during the testing period, although it had a tendency to slightly increase. After 2001, isolates resistant to EM significantly increased; however, msrA was hardly detected in ETB-producing S. aureus strains, and only five isolates were positive for both aac(6')/aph(2″) and msrA. In this study, we report the emergence of a fusion plasmid carrying the toxin gene etb and drug resistance genes. Prevalence of the pETBTY825 carrier may further increase the clinical threat, since ETB-producing S. aureus is closely related to more severe impetigo or staphylococcal scalded-skin syndrome (SSSS), which requires a general antimicrobial treatment.

The CmeABC efflux pump in Campylobacter jejuni confers resistance to structurally divergent antimicrobials, and inhibition of CmeABC represents a promising strategy to control antibiotic-resistant Campylobacter. Antisense peptide nucleic acids (PNAs) targeting the three components of CmeABC were evaluated for inhibition of CmeABC expression. The result revealed a synergistic effect of the PNAs targeting CmeA and CmeB on sensitizing C. jejuni to antibiotics. This finding further demonstrates the feasibility of using PNAs to potentiate antibiotics against antibiotic-resistant Campylobacter.

We conducted nationwide surveillance to investigate regional differences in macrolide-resistant (MR) Mycoplasma pneumoniae strains in Japan. The prevalence of MR M. pneumoniae in pediatric patients gradually increased between 2008 and 2012. Although regional differences were observed, high levels of MR genes were detected in all seven surveillance areas throughout Japan and ranged in prevalence from 50% to 93%. These regional differences were closely related to the previous administration of macrolides.

A total of 146 group B streptococcus isolates from 8 cities across China belonged to four serotypes. Serotype Ia was more common in children. A high prevalence of resistance was observed for levofloxacin (37.7%), erythromycin (71.2%), clindamycin, (53.4%), and tetracycline (81.5%). The levofloxacin and clindamycin resistances among the 4 serotypes differed significantly. Eighty percent of fluoroquinolone-resistant isolates belonged to the sequence type 19 (ST19)/serotype III clone, with GyrA-ParC-ParE triple substitutions. This clone carried the erm(B), mef(E), and tet(M) genes.

Macrolide antibiotics are important for clinical treatment of infections caused by Campylobacter jejuni. Development of resistance to this class of antibiotics in Campylobacter is a complex process, and the dynamic molecular changes involved in this process remain poorly defined. Multiple lineages of macrolide-resistant mutants were selected by stepwise exposure of C. jejuni to escalating doses of erythromycin or tylosin. Mutations in target genes were determined by DNA sequencing, and the dynamic changes in the expression of antibiotic efflux transporters and the transcriptome of C. jejuni were examined by real-time reverse transcription-PCR, immunoblotting, and DNA microarray analysis. Multiple types of mutations in ribosomal proteins L4 and L22 occurred early during stepwise selection. On the contrary, the mutations in the 23S rRNA gene, mediating high resistance to macrolides, were observed only in the late-stage mutants. Upregulation of antibiotic efflux genes was observed in the intermediately resistant mutants, and the magnitude of upregulation declined with the occurrence of mutations in the 23S rRNA gene. DNA microarray analysis revealed the differential expression of 265 genes, most of which occurred in the intermediate mutant, including the upregulation of genes encoding ribosomal proteins and the downregulation of genes involved in energy metabolism and motility. These results indicate (i) that mutations in L4 and L22 along with temporal overexpression of antibiotic efflux genes precede and may facilitate the development of high-level macrolide resistance and (ii) that the development of macrolide resistance affects the pathways important for physiology and metabolism in C. jejuni, providing an explanation for the reduced fitness of macrolide-resistant Campylobacter.

Helicobacter pylori is a globally important and genetically diverse gastric pathogen that infects most people in developing countries. Eradication efforts are complicated by antibiotic resistance, which varies in frequency geographically. There are very few data on resistance in African strains. Sixty-four Gambian H. pylori strains were tested for antibiotic susceptibility. The role of rdxA in metronidazole (Mtz) susceptibility was tested by DNA transformation and sequencing; RdxA protein variants were interpreted in terms of RdxA structure. Forty-four strains (69%) were resistant to at least 8 μg of Mtz/ml. All six strains from infants, but only 24% of strains from adults, were sensitive (P = 0.0031). Representative Mtz-resistant (Mtz(r)) strains were rendered Mtz susceptible (Mtz(s)) by transformation with a functional rdxA gene; conversely, Mtz(s) strains were rendered Mtz(r) by rdxA inactivation. Many mutations were found by Gambian H. pylori rdxA sequencing; mutations that probably inactivated rdxA in Mtz(r) strains were identified and explained using RdxA protein's structure. All of the strains were sensitive to clarithromycin and erythromycin. Amoxicillin and tetracycline resistance was rare. Sequence analysis indicated that most tetracycline resistance, when found, was not due to 16S rRNA gene mutations. These data suggest caution in the use of Mtz-based therapies in The Gambia. The increasing use of macrolides against respiratory infections in The Gambia calls for continued antibiotic susceptibility monitoring. The rich variety of rdxA mutations that we found will be useful in further structure-function studies of RdxA, the enzyme responsible for Mtz susceptibility in this important pathogen.

We have synthesized new derivatives of the macrolide antibiotics erythromycin and azithromycin. Novel deoxysugar moieties were attached to these standard antibiotics by biotransformation using a heterologous host. The resulting compounds were tested against several standard laboratory and clinically isolated bacterial strains. In addition, they were also tested in vitro against standard and drug-resistant strains of human malaria parasites (Plasmodium falciparum) and the liver stages of the rodent malaria parasite (Plasmodium berghei). Antibacterial activity of modified erythromycin and azithromycin showed no improvement over the unmodified macrolides, but the modified compounds showed a 10-fold increase in effectiveness after a short-term exposure against blood stages of malaria. The new compounds also remained active against azithromycin-resistant strains of P. falciparum and inhibited growth of liver-stage parasites at concentrations similar to those used for primaquine. Our findings show that malaria parasites have two distinct responses to macrolide antibiotics, one reflecting the prokaryotic origin of the apicoplast and a second, as-yet uncharacterized response that we attribute to the eukaryotic nature of the parasite. This is the first report for macrolides that target two different functions in the Plasmodium parasites.

ICESp1116, responsible for erm(B)-mediated, inducible erythromycin resistance in Streptococcus pyogenes, was comprehensively characterized, and its chromosomal integration site was determined. It displayed a unique mosaic organization consisting of a scaffold, related to TnGallo1 from Streptococcus gallolyticus, with two inserted fragments separated by IS1216. One fragment, containing erm(B), displayed high-level identity to a portion of the S. pyogenes plasmid pSM19035; the other, containing a truncated tet(M) gene, displayed high-level identity to the right-hand portion of Clostridium difficile Tn5397.

Toward developing new tools for fighting resistance to antibiotics, we investigated the antibacterial properties of a new decanoyl-based oligo-acyl-lysyl (OAK) hexamer, aminododecanoyl-lysyl-[aminodecanoyl-lysyl](5) (α(12)-5α(10)). The OAK exhibited preferential activity against Gram-negative bacteria (GNB), as determined using 36 strains, including diverse species, with an MIC(90) of 6.2 μM. The OAK's bactericidal mode of action was associated with rapid membrane depolarization and cell permeabilization, suggesting that the inner membrane was the primary target, whereas the observed binding affinity to lipoteichoic acid suggested that inefficacy against Gram-positive species resulted from a cell wall interaction preventing α(12)-5α(10) from reaching internal targets. Interestingly, perturbation of the inner membrane structure and function was preserved at sub-MIC values. This prompted us to assess the OAK's effect on the proton motive force-dependent efflux pump AcrAB-TolC, implicated in the low sensitivity of GNB to various antibiotics, including erythromycin. We found that under sub-MIC conditions, wild-type Escherichia coli was significantly more sensitive to erythromycin (the MIC dropped by >10-fold), unlike its acr-deletion mutant. Collectively, the data suggest a useful approach for treating GNB infections through overcoming antibiotic efflux.

We report retrospective analysis of the clinical and antimicrobial susceptibility data of 140 Streptococcus pseudopneumoniae isolates. Strains were isolated mostly from respiratory tract samples from patients with underlying diseases. In the case of infection, pneumonia, mainly aspiration pneumonia, was the most frequent (27.1% of the patients). We documented high rates of decreased susceptibilities and resistance to erythromycin and tetracycline (57% and 43% of the isolates, respectively), as well as reduced susceptibility to penicillin in 21% of the isolates.

Gonorrhea may become untreatable, and new treatment options are essential. We investigated the in vitro activity of the first fluoroketolide, solithromycin. Clinical Neisseria gonorrhoeae isolates and reference strains (n = 246), including the two extensively drug-resistant strains H041 and F89 and additional isolates with clinical cephalosporin resistance and multidrug resistance, were examined. The activity of solithromycin was mainly superior to that of other antimicrobials (n = 10) currently or previously recommended for gonorrhea treatment. Solithromycin might be an effective treatment option for gonorrhea.

The objective of this study was to evaluate the rates of clindamycin and erythromycin resistance among group B Streptococcus (GBS)-positive isolates cultured from pregnant women in an upstate New York community hospital. All GBS-positive perinatal rectovaginal cultures obtained from January 2010 through October 2011 were tested for resistance to erythromycin and clindamycin. Among the 688 GBS-positive cultures, clindamycin resistance was found in 38.4% and erythromycin resistance was found in 50.7%. Rates of GBS resistance to clindamycin and erythromycin are much higher than reported in earlier U.S. studies, suggesting both increasing resistance and regional variation in resistance. These findings lend strong support to the CDC and American College of Obstetricians and Gynecologists (ACOG) recommendations that clindamycin use for intrapartum antibiotic prophylaxis be restricted to penicillin-allergic women at high risk of anaphylaxis and that GBS isolates be tested for antibiotic resistance prior to the use of clindamycin in these women.

The aim of this study was to examine macrolide resistance mutations in Campylobacter species. In 76 strains studied, point mutation A to G at position 2059 of the 23S rRNA gene was detected in 30 of the 33 erythromycin-resistant strains. An amino acid insertion in the ribosomal protein L22 was found in one resistant strain without a 23S rRNA mutation. The A2059G mutation is the main cause of macrolide resistance in Campylobacter species.

Antibiotic resistance of biofilm-grown bacteria contributes to chronic infections, such as marginal and periapical periodontitis, which are strongly associated with Porphyromonas gingivalis. Concurrent azithromycin (AZM) administration and mechanical debridement improve the clinical parameters of periodontal tissue in situ. We examined the in vitro efficacy of AZM against P. gingivalis biofilms. The susceptibilities of adherent P. gingivalis strains 381, HW24D1, 6/26, and W83 to AZM, erythromycin (ERY), ampicillin (AMP), ofloxacin (OFX), and gentamicin (GEN) were investigated using a static model. The optical densities of adherent P. gingivalis cells were significantly decreased by using AZM and ERY at sub-MIC levels compared with those of the controls in all the strains tested, except for the effect of ERY on strain W83. AMP and OFX inhibited P. gingivalis adherent cells at levels over their MICs, and GEN showed no inhibition in the static model. The effects of AZM and ERY against biofilm cells were investigated using a flow cell model. The ATP levels of P. gingivalis biofilms were significantly decreased by AZM at concentrations below the sub-MICs; however, ERY was not effective for inhibition of P. gingivalis biofilm cells at their sub-MICs. Furthermore, decreased density of P. gingivalis biofilms was observed three-dimensionally with sub-MIC AZM, using confocal laser scanning microscopy. These findings suggest that AZM is effective against P. gingivalis biofilms at sub-MIC levels and could have future clinical application for oral biofilm infections, such as chronic marginal and periapical periodontitis.

A pharmacokinetic-pharmacodynamic (PKPD) model that characterizes the full time course of in vitro time-kill curve experiments of antibacterial drugs was here evaluated in its capacity to predict the previously determined PK/PD indices. Six drugs (benzylpenicillin, cefuroxime, erythromycin, gentamicin, moxifloxacin, and vancomycin), representing a broad selection of mechanisms of action and PK and PD characteristics, were investigated. For each drug, a dose fractionation study was simulated, using a wide range of total daily doses given as intermittent doses (dosing intervals of 4, 8, 12, or 24 h) or as a constant drug exposure. The time course of the drug concentration (PK model) as well as the bacterial response to drug exposure (in vitro PKPD model) was predicted. Nonlinear least-squares regression analyses determined the PK/PD index (the maximal unbound drug concentration [fC(max)]/MIC, the area under the unbound drug concentration-time curve [fAUC]/MIC, or the percentage of a 24-h time period that the unbound drug concentration exceeds the MIC [fT(>MIC)]) that was most predictive of the effect. The in silico predictions based on the in vitro PKPD model identified the previously determined PK/PD indices, with fT(>MIC) being the best predictor of the effect for β-lactams and fAUC/MIC being the best predictor for the four remaining evaluated drugs. The selection and magnitude of the PK/PD index were, however, shown to be sensitive to differences in PK in subpopulations, uncertainty in MICs, and investigated dosing intervals. In comparison with the use of the PK/PD indices, a model-based approach, where the full time course of effect can be predicted, has a lower sensitivity to study design and allows for PK differences in subpopulations to be considered directly. This study supports the use of PKPD models built from in vitro time-kill curves in the development of optimal dosing regimens for antibacterial drugs.