Aging of Copper Nanoparticles in the Marine Environment Regulates Toxicity for a Coastal Phytoplankton Species.

Environmental conditions in aquatic ecosystems transform toxic chemicals over time, influencing their bioavailability and toxicity. Using an environmentally relevant methodology, we tested how exposure to seawater for 1-15 weeks influenced the accumulation and toxicity of copper nanoparticles (nano-Cu) in a marine phytoplankton species. Nano-Cu rapidly agglomerated in seawater and then decreased in size due to Cu dissolution. Dissolution rates declined during weeks 1-4 and remained low until 15 weeks, when the large agglomerates that had formed began to rapidly dissolve again. Marine phytoplankton species were exposed for 5-day periods to nano-Cu aged from 1 to 15 weeks at concentrations from 0.01 to 20 ppm. Toxicity to phytoplankton, measured as change in population growth rate, decreased significantly with particle aging from 0 to 4 weeks but increased substantially in the 15-week treatment due apparently to elevated Cu dissolution of reagglomerated particles. Results indicate that the transformation, fate, and toxicity of nano-Cu in marine ecosystems are influenced by a highly dynamic physicochemical aging process.

Modeled and measured SARS-CoV-2 virus in septic tank systems for wastewater surveillance.

SARS-CoV-2 wastewater surveillance (WWS) at wastewater treatment plants (WWTPs) can reveal sewered community COVID-19 prevalence. For unsewered areas using septic tank systems (STSs) or holding tanks, how to conduct WWS remains unexplored. Here, two large STSs serving Zuma Beach (Malibu, CA) were studied. Supernatant and sludge SARS-CoV-2 concentrations from the directly-sampled STSs parameterized a dynamic solid-liquid separation, mass balance-based model for estimating the infection rate of users. Pumped septage before hauling and upon WWTP disposal was also sampled and assessed. Most (96%) STS sludge samples contained SARS-CoV-2 N1 and N2 genes, with concentrations exceeding the supernatant and increasing with depth while correlating with total suspended solids (TSS). The trucked septage contained N1 and N2 genes which decayed (coefficients: 0.09-0.29 h-1) but remained detectable. Over approximately 5 months starting in December 2020, modeled COVID-19 prevalence estimations among users ranged from 8 to 18%, mirroring a larger metropolitan area for the first 2 months. The approaches herein can inform public health intervention and augment conventional WWS in that: (1) user infection rates for communal holding tanks are estimable and (2) pumped and hauled septage can be assayed to infer where disease is spreading in unsewered areas.

Mechanistic Insights to Combating NDM- and CTX-M-Coproducing Klebsiella pneumoniae by Targeting Cell Wall Synthesis and Outer Membrane Integrity.

Metallo-ß-lactamase (MBL)-producing Gram-negative bacteria cause infections associated with high rates of morbidity and mortality. Currently, a leading regimen to treat infections caused by MBL-producing bacteria is aztreonam combined with ceftazidime-avibactam. The purpose of the present study was to evaluate and rationally optimize the combination of aztreonam and ceftazidime-avibactam with and without polymyxin B against a clinical Klebsiella pneumoniae isolate producing NDM-1 and CTX-M by use of the hollow fiber infection model (HFIM). A novel de-escalation approach to polymyxin B dosing was also explored, whereby a standard 0-h loading dose was followed by maintenance doses that were 50% of the typical clinical regimen. In the HFIM, the addition of polymyxin B to aztreonam plus ceftazidime-avibactam significantly improved bacterial killing, leading to eradication, including for the novel de-escalation dosing strategy. Serial samples from the growth control and monotherapies were explored in a Galleria mellonella virulence model to assess virulence changes. Weibull regression showed that low-level ceftazidime resistance and treatment with monotherapy resulted in increased G. mellonella mortality (P < 0.05). A neutropenic rabbit pneumonia model demonstrated that aztreonam plus ceftazidime-avibactam with or without polymyxin B resulted in similar bacterial killing, and these combination therapies were statistically significantly better than monotherapies (P < 0.05). However, only the polymyxin B-containing combination therapy produced a statistically significant decrease in lung weights (P < 0.05), indicating a decreased inflammatory process. Altogether, adding polymyxin B to the combination of aztreonam plus ceftazidime-avibactam for NDM- and CTX-M-producing K. pneumoniae improved bacterial killing effects, reduced lung inflammation, suppressed resistance amplification, and limited virulence changes.

Genetic sequence data evidence that human faecal-associated HF183 sequences are on human skin and in urine.

AIMS: The DNA marker HF183 is a partial 16S rRNA gene sequence highly specific to human-associated Bacteroides including Bacteroides dorei. While HF183 is used to assess human faecal contamination in aquatic environments worldwide, little is known about the existence of HF183 and B. dorei in human microbiomes outside of the human gastrointestinal tract and faeces. METHODS AND RESULTS: Previously published human skin and urine microbiome data sets from five independent human body skin studies, the Human Microbiome Project (HMP) and three independent human urine studies were analysed. The HF183 gene sequence was detected in all skin data sets, with the ratios of positive samples ranging from 0.5% to 36.3%. Popliteal fossa (knee), volar forearm and inguinal (groin) creases were identified as hot spots. HF183 was detected in two of three urine data sets, with ratios of positive samples ranging from 0% to 37.5%. All HF183-containing sequences from these data sets were classified as associated with B. dorei. CONCLUSIONS: HF183 is widespread on human skin and present in urine. SIGNIFICANCE AND IMPACT OF STUDY: Skin and urine microbiomes could be sources of HF183 to environmental waters. Such non-faecal sources of HF183 might explain low concentrations of HF183 in recreational waters when swimmers are present.

SARS-CoV-2 Wastewater Surveillance for Public Health Action.

Wastewater surveillance for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has garnered extensive public attention during the coronavirus disease pandemic as a proposed complement to existing disease surveillance systems. Over the past year, methods for detection and quantification of SARS-CoV-2 viral RNA in untreated sewage have advanced, and concentrations in wastewater have been shown to correlate with trends in reported cases. Despite the promise of wastewater surveillance, for these measurements to translate into useful public health tools, bridging the communication and knowledge gaps between researchers and public health responders is needed. We describe the key uses, barriers, and applicability of SARS-CoV-2 wastewater surveillance for supporting public health decisions and actions, including establishing ethics consideration for monitoring. Although wastewater surveillance to assess community infections is not a new idea, the coronavirus disease pandemic might be the initiating event to make this emerging public health tool a sustainable nationwide surveillance system, provided that these barriers are addressed.

Limited Bacterial Removal in Full-Scale Stormwater Biofilters as Evidenced by Community Sequencing Analysis.

In urban areas, untreated stormwater runoff can pollute downstream surface waters. To intercept and treat runoff, low-impact or "green infrastructure" approaches such as using biofilters are adopted. Yet, actual biofilter pollutant removal is poorly understood; removal is often studied in laboratory columns, with variable removal of viable and culturable microbial cell numbers including pathogens. Here, to assess bacterial pollutant removal in full-scale planted biofilters, stormwater was applied, unspiked or spiked with untreated sewage, in simulated storm events under transient flow conditions, during which biofilter influents versus effluents were compared. Based on microbial biomass, sequences of bacterial community genes encoding 16S rRNA, and gene copies of the human fecal marker HF183 and of the Enterococcus spp. marker Entero1A, removal of bacterial pollutants in biofilters was limited. Dominant bacterial taxa were similar for influent versus effluent aqueous samples within each inflow treatment of either spiked or unspiked stormwater. Bacterial pollutants in soil were gradually washed out, albeit incompletely, during simulated storm flushing events. In post-storm biofilter soil cores, retained influent bacteria were concentrated in the top layers (0-10 cm), indicating that the removal of bacterial pollutants was spatially limited to surface soils. To the extent that plant-associated processes are responsible for this spatial pattern, treatment performance might be enhanced by biofilter designs that maximize influent contact with the rhizosphere.

University Stormwater Management within Urban Environmental Regulatory Regimes: Barriers to Progressivity or Opportunities to Innovate?

U.S. public university campuses are held directly responsible for compliance with many of the same federal- and state-level environmental regulations as cities, including stormwater management. While operating as 'cities within cities' in many respects, campuses face unique constraints in achieving stormwater regulatory compliance. To compare the abilities of campuses to comply with stormwater regulations to municipalities, we conduct mixed-methods research using primary data from five University of California (UC) campuses. Public universities constituted over 20% of California's "nontraditional" permittees under the municipal separate storm sewer system (MS4) regulation regime in 2013. We utilize semi-structured interviews with campus and regulatory officials, a survey of campus students and staff around support and willingness to pay for innovative stormwater management, and content analysis of campus stormwater management documents to examine challenges to public university stormwater compliance. We find that, despite their progressive environmental practices in other areas like energy and water conservation, even as compared to cities, stormwater management practices on the evaluated campuses are constrained by several factors: infrastructure financing limitations, lack of transparent and coordinated decision-making, a lack of campus resident involvement, and regulatory inflexibility. Our study provides new insights, both for understanding campuses as sustainable 'cities within cities' and more broadly for urban environmental compliance regimes globally.

Physical Properties of Carbon Nanomaterials and Nanoceria Affect Pathways Important to the Nodulation Competitiveness of the Symbiotic N2 -Fixing Bacterium Bradyrhizobium diazoefficiens.

The pathogenicity and antimicrobial properties of engineered nanomaterials (ENMs) are relatively well studied. However, less is known regarding the interactions of ENMs and agriculturally beneficial microorganisms that affect food security. Nanoceria (CeO2 nanoparticles (NPs)), multiwall carbon nanotubes (MWCNTs), graphene nanoplatelets (GNPs), and carbon black (CB) have been previously shown to inhibit symbiotic N2 fixation in soybeans, but direct rhizobial susceptibility is uncertain. Here, Bradyrhizobium diazoefficiens associated with symbiotic N2 fixation in soybeans is assessed, evaluating the role of soybean root exudates (RE) on ENM-bacterial interactions and the effects of CeO2 NPs, MWCNTs, GNPs, and CB on bacterial growth and gene expression. Although bacterial growth is inhibited by 50 mg L-1 CeO2 NPs, MWCNTs, and CB, all ENMs at 0.1 and 10 mg L-1 cause a global transcriptomic response that is mitigated by RE. ENMs may interfere with plant-bacterial signaling, as evidenced by suppressed upregulation of genes induced by RE, and downregulation of genes encoding transport RNA, which facilitates nodulation signaling. MWCNTs and CeO2 NPs inhibit the expression of genes conferring B. diazoefficiens nodulation competitiveness. Surprisingly, the transcriptomic effects on B. diazoefficiens are similar for these two ENMs, indicating that physical, not chemical, ENM properties explain the observed effects.

Determining the optimal dosing of a novel combination regimen of ceftazidime/avibactam with aztreonam against NDM-1-producing Enterobacteriaceae using a hollow-fibre infection model.

BACKGROUND: MBL-producing strains of Enterobacteriaceae are a major public health concern. We sought to define optimal combination regimens of ceftazidime/avibactam with aztreonam in a hollow-fibre infection model (HFIM) of MBL-producing strains of Escherichia coli and Klebsiella pneumoniae. METHODS: E. coli ARLG-1013 (blaNDM-1, blaCTX-M, blaCMY, blaTEM) and K. pneumoniae ARLG-1002 (blaNDM-1, blaCTXM-15, blaDHA, blaSHV, blaTEM) were studied in the HFIM using simulated human dosing regimens of ceftazidime/avibactam and aztreonam. Experiments were designed to evaluate the effect of staggered versus simultaneous administration, infusion duration and aztreonam daily dose (6 g/day versus 8 g/day) on bacterial killing and resistance suppression. Prospective validation experiments for the most active combination regimens were performed in triplicate to ensure reproducibility. RESULTS: Staggered administration of the combination (ceftazidime/avibactam followed by aztreonam) was found to be inferior to simultaneous administration. Longer infusion durations (2 h and continuous infusion) also resulted in enhanced bacterial killing relative to 30 min infusions. The rate of killing was more pronounced with 8 g/day versus 6 g/day aztreonam combination regimens for both tested strains. In the prospective validation experiments, ceftazidime/avibactam with aztreonam dosed every 8 and 6 h, respectively (ceftazidime/avibactam 2/0.5 g every 8 h + aztreonam 2 g every 6 h), or ceftazidime/avibactam with aztreonam as continuous infusions resulted in maximal bacterial killing and resistance suppression over 7 days. CONCLUSIONS: Simultaneous administration of aztreonam 8 g/day given as a continuous or 2 h infusion with ceftazidime/avibactam resulted in complete bacterial eradication and resistance suppression. Further study of this combination is needed with additional MBL-producing Gram-negative pathogens. The safety of this double ß-lactam strategy also warrants further study in Phase 1 clinical trials.

Effects of carbonaceous nanomaterials on soil-grown soybeans under combined heat and insect stresses.

Because carbonaceous nanomaterials (CNMs) are expected to enter soils, the exposure implications to crop plants and plant-microbe interactions should be understood. Most investigations have been under ideal growth conditions, yet crops commonly experience abiotic and biotic stresses. Little is known how co-exposure to these environmental stresses and CNMs would cause combined effects on plants. We investigated the effects of 1000 mg kg-1 multiwalled carbon nanotubes (CNTs), graphene nanoplatelets (GNPs) and industrial carbon black (CB) on soybeans grown to the bean production stage in soil. Following seed sowing, plants became stressed by heat and infested with an insect (thrips). Consequently, all plants had similarly stunted growth, leaf damage, reduced final biomasses and fewer root nodules compared with healthy control soybeans previously grown without heat and thrips stresses. Thus, CNMs did not significantly influence the growth and yield of stressed soybeans, and the previously reported nodulation inhibition by CNMs was not specifically observed here. However, CNMs did significantly alter two leaf health indicators: the leaf chlorophyll a/b ratio, which was higher in the GNP treatment than in either the control (by 15 %) or CB treatment (by 14 %), and leaf lipid peroxidation, which was elevated in the CNT treatment compared with either the control (by 47 %) or GNP treatment (by 66 %). Overall, these results show that, while severe environmental stresses may impair plant production, CNMs (including CNTs and GNPs) in soil could additionally affect foliar health of an agriculturally important legume.

Azithromycin Pharmacodynamics against Persistent Haemophilus influenzae in Chronic Obstructive Pulmonary Disease.

The pharmacodynamic profile of azithromycin against persistent strains of nontypeable Haemophilus influenzae (NTHi) from chronic obstructive pulmonary disease (COPD) patients was characterized. Azithromycin displayed differential concentration-dependent activities (R2 ≥ 0.988); the pharmacodynamic response was attenuated when we compared the "first" and "last" strains of NTHi that persisted in the airways of the same patient for 819 days (the 50% effective concentration [EC50] increased more than 50 times [0.0821 mg/liter versus 4.23 mg/liter]). In the hollow-fiber infection model, NTHi viability was maintained throughout simulated azithromycin (Zithromax) Z-Pak regimens over 10 days.

Carbonaceous Nanomaterials Have Higher Effects on Soybean Rhizosphere Prokaryotic Communities During the Reproductive Growth Phase than During Vegetative Growth.

Carbonaceous nanomaterials (CNMs) can affect agricultural soil prokaryotic communities, but how the effects vary with the crop growth stage is unknown. To investigate this, soybean plants were cultivated in soils amended with 0, 0.1, 100, or 1000 mg kg-1 of carbon black, multiwalled carbon nanotubes (MWCNTs), or graphene. Soil prokaryotic communities were analyzed by Illumina sequencing at day 0 and at the soybean vegetative and reproductive stages. The sequencing data were functionally annotated using the functional annotation of prokaryotic taxa (FAPROTAX) database. The prokaryotic communities were unaffected at day 0 and were altered at the plant vegetative stage only by 0.1 mg kg-1 MWCNTs. However, at the reproductive stage, when pods were filling, most treatments (except 1000 mg kg-1 MWCNTs) altered the prokaryotic community composition, including functional groups associated with C, N, and S cycling. The lower doses of CNMs, which were previously shown to be less agglomerated and thus more bioavailable in soil relative to the higher doses, were more effective toward both overall communities and individual functional groups. Taken together, prokaryotic communities in the soybean rhizosphere can be significantly phylogenetically and functionally altered in response to bioavailable CNMs, especially when soybean plants are actively directing resources to seed production.

Influence of rhlR and lasR on Polymyxin Pharmacodynamics in Pseudomonas aeruginosa and Implications for Quorum Sensing Inhibition with Azithromycin.

The impact of quorum sensing on polymyxin and azithromycin pharmacodynamics was assessed in Pseudomonas aeruginosa PAO1 and an isogenic rhlR/lasR double knockout. For polymyxin B, greater killing against the rhlR/lasR knockout than against PAO1 was observed at 108 CFU/ml (polymyxin B half-maximal effective concentration [EC50], 5.61 versus 12.5 mg/liter, respectively; P < 0.005). Polymyxin B combined with azithromycin (256 mg/liter) was synergistic against each strain, significantly reducing the respective polymyxin B EC50 compared to those with monotherapy (P < 0.005), and is a promising strategy by which to combat P. aeruginosa.

New Polymyxin B Dosing Strategies To Fortify Old Allies in the War against KPC-2-Producing Klebsiella pneumoniae.

Pharmacodynamics of a polymyxin B, meropenem, and rifampin triple combination were examined against Klebsiella pneumoniae carbapenemase-producing Klebsiella pneumoniae (KPC-Kp) ST258. In time-kill experiments against three KPC-Kp isolates, triple combination generated 8.14, 8.19, and 8.29 log10 CFU/ml reductions within 24 h. In the hollow-fiber infection model, the triple combination caused maximal killing of 5.16 log10 CFU/ml at 78 h and the time required for regrowth was more than doubled versus the 2-drug combinations. Remarkably, combinations with a high single-dose polymyxin B burst plus rifampin preserved KPC-Kp polymyxin susceptibility (MIC240 h = 0.5 mg/liter) versus the same combination with traditionally dosed polymyxin B, where resistance was amplified (MIC240 h = 32 mg/liter).

Pharmacodynamics of dose-escalated 'front-loading' polymyxin B regimens against polymyxin-resistant mcr-1-harbouring Escherichia coli.

Objectives: Gram-negative bacteria harbouring the mcr-1 plasmid are resistant to the 'last-line' polymyxins and have been reported worldwide. Our objective was to define the impact of increasing the initial polymyxin B dose intensity against an mcr-1 -harbouring strain to delineate the impact of plasmid-mediated polymyxin resistance on the dynamics of bacterial killing and resistance. Methods: A hollow fibre infection model (HFIM) was used to simulate polymyxin B regimens against an mcr-1 -harbouring Escherichia coli (MIC 8 mg/L) over 10 days. Four escalating polymyxin B 'front-loading' regimens (3.33, 6.66, 13.3 or 26.6 mg/kg for one dose followed by 1.43 mg/kg every 12 h starting 12 h later) simulating human pharmacokinetics were utilized in the HFIM. A mechanism-based, mathematical model was developed using S-ADAPT to characterize bacterial killing. Results: The 3.33 mg/kg 'front-loading' regimen resulted in regrowth mirroring the growth control. The 6.66, 13.3 and 26.6 mg/kg 'front-loading' regimens resulted in maximal bacterial reductions of 1.91, 3.79 and 6.14 log 10 cfu/mL, respectively. Irrespective of the early polymyxin B exposure (24 h AUC), population analysis profiles showed similar growth of polymyxin B-resistant subpopulations. The HFIM data were well described by the mechanism-based model integrating three subpopulations (susceptible, intermediate and resistant). Compared with the susceptible subpopulation of mcr-1 -harbouring E. coli , the resistant subpopulation had an approximately 10-fold lower rate of killing due to polymyxin B treatment. Conclusions: Manipulating initial dose intensity of polymyxin B was not able to overcome plasmid-mediated resistance due to mcr-1 in E. coli . This reinforces the need to develop new combinatorial strategies to combat these highly resistant Gram-negative bacteria.

High-intensity meropenem combinations with polymyxin B: new strategies to overcome carbapenem resistance in Acinetobacter baumannii.

OBJECTIVES: The pharmacodynamics of polymyxin/carbapenem combinations against carbapenem-resistant Acinetobacter baumannii (CRAB) are largely unknown. Our objective was to determine whether intensified meropenem regimens in combination with polymyxin B enhance killing and resistance suppression of CRAB. METHODS: Time-kill experiments for meropenem and polymyxin B combinations were conducted against three polymyxin B-susceptible (MIC of polymyxin B = 0.5 mg/L) CRAB strains with varying meropenem MICs (ATCC 19606, N16870 and 03-149-1; MIC of meropenem = 4, 16 and 64 mg/L, respectively) at 108 cfu/mL. A hollow-fibre infection model was then used to simulate humanized regimens of polymyxin B and meropenem (2, 4, 6 and 8 g prolonged infusions every 8 h) versus N16870 at 108 cfu/mL over 14 days. New mathematical mechanism-based models were developed using S-ADAPT. RESULTS: Time-kill experiments were well described by the mathematical mechanism-based models, with the presence of polymyxin B drastically decreasing the meropenem concentration needed for half-maximal activity against meropenem-resistant populations from 438 to 82.1 (ATCC 19606), 158 to 93.6 (N16870) and 433 to 76.0 mg/L (03-149-1). The maximum killing effect of combination treatment was similar among all three strains despite divergent meropenem MIC values (Emax = 2.13, 2.08 and 2.15; MIC of meropenem = 4, 16 and 64 mg/L, respectively). Escalating the dose of meropenem in hollow-fibre combination regimens from 2 g every 8 h to 8 g every 8 h resulted in killing that progressed from a >2.5 log10 cfu/mL reduction with regrowth by 72 h (2 g every 8 h) to complete eradication by 336 h (8 g every 8 h). CONCLUSION: Intensified meropenem dosing in combination with polymyxin B may offer a unique strategy to kill CRAB irrespective of the meropenem MIC.

Host-Symbiont Interaction Model Explains Non-monotonic Response of Soybean Growth and Seed Production to Nano-CeO2 Exposure.

Recent nanotoxicity studies have demonstrated non-monotonic dose-response mechanisms for planted soybean that have a symbiotic relationship with bacteroids in their root nodules: reduction of growth and seed production was greater for low, as compared to high, exposures. To investigate mechanistic underpinnings of the observed patterns, we formulated an energy budget model coupled to a toxicokinetic module describing bioaccumulation, and two toxicodynamic modules describing toxic effects on host plant and symbionts. By fitting data on plants exposed to engineered CeO2 nanoparticles to the newly formulated model, we show that the non-monotonic patterns can be explained as the interaction of two, individually monotonic, dose-response processes: one for the plant and the other for the symbiont. We further validate the newly formulated model by showing that, without the need for additional parameters, the model successfully predicts changes in dinitrogen fixation potential as a function of exposure (dinitrogen fixation potential data not used in model fitting). The symbiont buffers overall toxicity only when, in the absence of exposure to a toxicant, it has a parasitic interaction with the host plant. If the interaction is mutualistic or commensal, there is no buffering and only monotonic toxic responses are possible. Because the model is based on general biological principles, we expect it to be applicable to other similar symbiotic systems, especially other nodule-forming legumes.

Spatial Models of Sewer Pipe Leakage Predict the Occurrence of Wastewater Indicators in Shallow Urban Groundwater.

Twentieth century municipal wastewater infrastructure greatly improved U.S. urban public health and water quality. However, sewer pipes deteriorate, and their accumulated structural defects may release untreated wastewater to the environment via acute breaks or insidious exfiltration. Exfiltrated wastewater constitutes a loss of potentially reusable water and delivers a complex and variable mix of contaminants to urban shallow groundwater. Yet, predicting where deteriorated sewers impinge on shallow groundwater has been challenging. Here we develop and test a spatially explicit model of exfiltration probability based on pipe attributes and groundwater elevation without prior knowledge of exfiltrating defect locations. We find that models of exfiltration probability can predict the probable occurrence in underlying shallow groundwater of established wastewater indicators including the artificial sweetener acesulfame, tryptophan-like fluorescent dissolved organic matter, nitrate, and a stable isotope of water (δ18O). The strength of the association between exfiltration probability and indicators of wastewater increased when multiple pipe attributes, distance weighting, and groundwater flow direction were considered in the model. The results prove that available sanitary sewer databases and groundwater digital elevation data can be analyzed to predict where pipes are likely leaking and contaminating groundwater. Such understanding could direct sewer infrastructure reinvestment toward water resource protection.

Effect of Fluoroquinolones and Macrolides on Eradication and Resistance of Haemophilus influenzae in Chronic Obstructive Pulmonary Disease.

Little is known about the effect of antibiotics on eradication of carriage and development of resistance in Haemophilus influenzae in individuals with chronic obstructive pulmonary disease (COPD). Our goals were to assess antibiotic susceptibilities, prevalence of resistance genes, and development of resistance in H. influenzae and to evaluate the effect of macrolide and fluoroquinolone administration on H. influenzae eradication. Data were from a 15-year longitudinal study of COPD. Genome sequence data were used to determine genotype and identify resistance genes. MICs of antibiotics were determined by reference broth microdilution. Generalized linear mixed models were used to evaluate associations between antibiotic use and H. influenzae eradication. We examined 267 H. influenzae isolates from 77 individuals. All newly acquired H. influenzae isolates were susceptible to azithromycin. Five of 27 (19%) strains developed 4-fold increases in azithromycin MICs and reached or exceeded the susceptibility breakpoint (≤4 µg/ml) during exposure. H. influenzae isolates were uniformly susceptible to ciprofloxacin, levofloxacin, and moxifloxacin (MIC90s of 0.015, 0.015, and 0.06, respectively); there were no mutations in quinolone resistance-determining regions. Fluoroquinolone administration was associated with increased H. influenzae eradication compared to macrolides (odds ratio [OR], 16.67; 95% confidence interval [CI], 2.67 to 104.09). There was no difference in H. influenzae eradication when comparing macrolide administration to no antibiotic (OR, 1.89; 95% CI, 0.43 to 8.30). Fluoroquinolones are effective in eradicating H. influenzae in individuals with COPD. Macrolides are ineffective in eradicating H. influenzae, and their use in COPD patients may lead to decreased macrolide susceptibility and resistance.

Optimization of Polymyxin B in Combination with Doripenem To Combat Mutator Pseudomonas aeruginosa.

Development of spontaneous mutations in Pseudomonas aeruginosa has been associated with antibiotic failure, leading to high rates of morbidity and mortality. Our objective was to evaluate the pharmacodynamics of polymyxin B combinations against rapidly evolving P. aeruginosa mutator strains and to characterize the time course of bacterial killing and resistance via mechanism-based mathematical models. Polymyxin B or doripenem alone and in combination were evaluated against six P. aeruginosa strains: wild-type PAO1, mismatch repair (MMR)-deficient (mutS and mutL) strains, and 7,8-dihydro-8-oxo-deoxyguanosine system (GO) base excision repair (BER)-deficient (mutM, mutT, and mutY) strains over 48 h. Pharmacodynamic modeling was performed using S-ADAPT and facilitated by SADAPT-TRAN. Mutator strains displayed higher mutation frequencies than the wild type (>600-fold). Exposure to monotherapy was followed by regrowth, even at high polymyxin B concentrations of up to 16 mg/liter. Polymyxin B and doripenem combinations displayed enhanced killing activity against all strains where complete eradication was achieved for polymyxin B concentrations of >4 mg/liter and doripenem concentrations of 8 mg/liter. Modeling suggested that the proportion of preexisting polymyxin B-resistant subpopulations influenced the pharmacodynamic profiles for each strain uniquely (fraction of resistance values are -8.81 log10 for the wild type, -4.71 for the mutS mutant, and -7.40 log10 for the mutM mutant). Our findings provide insight into the optimization of polymyxin B and doripenem combinations against P. aeruginosa mutator strains.