Antibiotic capture by bacterial lipocalins uncovers an extracellular mechanism of intrinsic antibiotic resistance

15 p.-6 fig.The potential for microbes to overcome antibiotics of different classes before they reach bacterial cells is largely unexplored. Here we show that a soluble bacterial lipocalin produced by Burkholderia cenocepacia upon exposure to sublethal antibiotic concentrations increases resistance to diverse antibiotics in vitro and in vivo. These phenotypes were recapitulated by heterologous expression in B. cenocepacia of lipocalin genes from Pseudomonas aeruginosa, Mycobacterium tuberculosis,and methicillin-resistant Staphylococcus aureus. Purified lipocalin bound different classes of bactericidal antibiotics and contributed to bacterial survival in vivo. Experimental and X-ray crystal structure-guided computational studies revealed that lipocalins counteract antibiotic action by capturing antibiotics in the extracellular space. We also demonstrated that fat-soluble vitamins prevent antibiotic capture by binding bacterial lipocalin with higher affinity than antibiotics. Therefore, bacterial lipocalins contribute to antimicrobial resistance by capturing diverse antibiotics in the extracellular space at the site of infection, which can be counteracted by known vitamins.This work was funded by grants from Cystic Fibrosis Canada, the European Commission,a Marie Curie Career Integration grant (projects 618095, NONANTIRES), and the Infection and Immunity Translational Research Group, Northern Ireland HSC to M.A.V.;the Spanish Ministry for Economy and Competitiveness (MINECO CTQ2011-22724 and CTQ2014-57141-R), European Commission Marie Curie grants GLYCOPHARM FP7-PITNGA-2012-317297 and TOLLerant H2020-MSC-ETN-642157 to S.M.S.; and Canadian Institutes of Health research grant MOP-49597 and a grant from Cystic Fibrosis Canada to M.E.P.M.Peer reviewe

Localization of ABCG5 and ABCG8 proteins in human liver, gall bladder and intestine

BACKGROUND: The molecular mechanisms that regulate the entry of dietary sterols into the body and their removal via hepatobiliary secretion are now beginning to be defined. These processes are specifically disrupted in the rare autosomal recessive disease, Sitosterolemia (MIM 210250). Mutations in either, but not both, of two genes ABCG5 or ABCG8, comprising the STSL locus, are now known to cause this disease and their protein products are proposed to function as heterodimers. Under normal circumstances cholesterol, but not non-cholesterol sterols, is preferentially absorbed from the diet. Additionally, any small amounts of non-cholesterol sterols that are absorbed are rapidly taken up by the liver and preferentially excreted into bile. Based upon the defects in sitosterolemia, ABCG5 and ABCG8 serve specifically to exclude non-cholesterol sterol entry at the intestinal level and are involved in sterol excretion at the hepatobiliary level. METHODS: Here we report the biochemical and immuno-localization of ABCG5 and ABCG8 in human liver, gallbladder and intestine using cell fractionation and immunohistochemical analyses. RESULTS: We raised peptide antibodies against ABCG5 and ABCG8 proteins. Using human liver samples, cell fractionation studies showed both proteins are found in membrane fractions, but they did not co-localize with caveolin-rafts, ER, Golgi or mitochondrial markers. Although their distribution in the sub-fractions was similar, they were not completely contiguous. Immunohistochemical analyses showed that while both proteins were readily detectable in the liver, ABCG5 was found predominately lining canalicular membranes, whereas ABCG8 was found in association with bile duct epithelia. At the cellular level, ABCG5 appeared to be apically expressed, whereas ABCG8 had a more diffuse expression pattern. Both ABCG5 and ABCG8 appeared to localize apically as shown by co-localization with MRP2. The distribution patterns of ABCG5 and ABCG8 in the gallbladder were very similar to each other. In the small intestine both ABCG5 and ABCG8 appear to line the brush border. However, at the level of the enterocyte, the cellular distribution patterns of ABCG5 and ABCG8 differed, such that ABCG5 was more diffuse, but ABCG8 was principally apical. Using standard deglycosylation methods, ABCG5 and ABCG8 do not appear to be glycosylated, suggesting a difference between human and mouse proteins. CONCLUSION: We report the distribution patterns of ABCG5 and ABCG8 in human tissues. Cell fractionation studies showed that both proteins co-fractionated in general, but could also be found independent of each other. As predicted, they are expressed apically in both intestine and liver, although their intracellular expression patterns are not completely congruent. These studies support the concept of heterodimerization of ABCG5 and ABCG8, but also support the notion that these proteins may have an independent function

Combined field inoculations of pseudomonas bacteria, arbuscular mycorrhizal fungi, and entomopathogenic nematodes and their effects on wheat performance

In agricultural ecosystems, pest insects, pathogens, and reduced soil fertility pose major challenges to crop productivity and are responsible for significant yield losses worldwide. Management of belowground pests and diseases remains particularly challenging due to the complex nature of the soil and the limited reach of conventional agrochemicals. Boosting the presence of beneficial rhizosphere organisms is a potentially sustainable alternative and may help to optimize crop health and productivity. Field application of single beneficial soil organisms has shown satisfactory results under optimal conditions. This might be further enhanced by combining multiple beneficial soil organisms, but this remains poorly investigated. Here, we inoculated wheat plots with combinations of three beneficial soil organisms that have different rhizosphere functions and studied their effects on crop performance. Plant beneficial Pseudomonas bacteria, arbuscular mycorrhizal fungi (AMF), and entomopathogenic nematodes (EPN), were inoculated individually or in combinations at seeding, and their effects on plant performance were evaluated throughout the season. We used traditional and molecular identification tools to monitor their persistence over the cropping season in augmented and control treatments, and to estimate the possible displacement of native populations. In three separate trials, beneficial soil organisms were successfully introduced into the native populations and readily survived the field conditions. Various Pseudornonas, mycorrhiza, and nematode treatments improved plant health and productivity, while their combinations provided no significant additive or synergistic benefits compared to when applied alone. EPN application temporarily displaced some of the native EPN, but had no significant long-term effect on the associated food web. The strongest positive effect on wheat survival was observed for Pseudomonas and AMF during a season with heavy natural infestation by the frit fly, Oscinella frit, a major pest of cereals. Hence, beneficial impacts differed between the beneficial soil organisms and were most evident for plants under biotic stress. Overall, our findings indicate that in wheat production under the test conditions the three beneficial soil organisms can establish nicely and are compatible, but their combined application provides no additional benefits. Further studies are required, also in other cropping systems, to fine-tune the functional interactions among beneficial soil organisms, crops, and the environment

Comparative assessment of gasification based coal power plants with various CO2 capture technologies producing electricity and hydrogen

Seven different types of gasification-based coal conversion processes for producing mainly electricity and in some cases hydrogen (H2), with and without carbon dioxide (CO2) capture, were compared on a consistent basis through simulation studies. The flowsheet for each process was developed in a chemical process simulation tool “Aspen Plus”. The pressure swing adsorption (PSA), physical absorption (Selexol), and chemical looping combustion (CLC) technologies were separately analyzed for processes with CO2 capture. The performances of the above three capture technologies were compared with respect to energetic and exergetic efficiencies, and the level of CO2 emission. The effect of air separation unit (ASU) and gas turbine (GT) integration on the power output of all the CO2 capture cases is assessed. Sensitivity analysis was carried out for the CLC process (electricity-only case) to examine the effect of temperature and water-cooling of the air reactor on the overall efficiency of the process. The results show that, when only electricity production in considered, the case using CLC technology has an electrical efficiency 1.3% and 2.3% higher than the PSA and Selexol based cases, respectively. The CLC based process achieves an overall CO2 capture efficiency of 99.9% in contrast to 89.9% for PSA and 93.5% for Selexol based processes. The overall efficiency of the CLC case for combined electricity and H2 production is marginally higher (by 0.3%) than Selexol and lower (by 0.6%) than PSA cases. The integration between the ASU and GT units benefits all three technologies in terms of electrical efficiency. Furthermore, our results suggest that it is favorable to operate the air reactor of the CLC process at higher temperatures with excess air supply in order to achieve higher power efficiency

ATP-Binding Cassette Transporter G5 and G8 Polymorphisms and Several Environmental Factors with Serum Lipid Levels

The association of ATP-binding cassette (ABC) transporter single nucleotide polymorphisms (SNPs) and serum lipid profiles is inconsistent. The present study was undertaken to detect the association of ABCG5/G8 SNPs and several environmental factors with serum lipid levels.Genotyping of the ABCG5 (rs4131229 and rs6720173) and ABCG8 (rs3806471 and rs4148211) SNPs was performed in 719 unrelated subjects of Mulao nationality and 782 participants of Han nationality. There were no differences in the genotypic and allelic frequencies of four SNPs between the two ethnic groups besides the genotypic frequencies of rs4131229 SNP in Han. The levels of triglyceride (TG), apolipoprotein (Apo) A1, and ApoA1/ApoB ratio (rs4131229); low-density lipoprotein cholesterol (LDL-C) and ApoB (rs6720173); high-density lipoprotein cholesterol (HDL-C), ApoA1, ApoB, and ApoA1/ApoB ratio (rs3806471); and HDL-C, ApoA1, and ApoA1/ApoB ratio (rs4148211) in Han were different among their genotypes (P<0.05-0.001). The levels of LDL-C (rs6720173) and ApoA1 (rs3806471) in Mulao were also different among their genotypes (P<0.05 for each). The levels of TC, TG, HDL-C, ApoA1, and ApoA1/ApoB ratio (rs4131229); LDL-C and ApoB (rs6720173); HDL-C, ApoA1, and ApoA1/ApoB ratio (rs3806471); and TG, HDL-C, ApoA1, and ApoA1/ApoB ratio (rs4148211) in Han males; and ApoA1/ApoB ratio (rs4131229); LDL-C, ApoB, and ApoA1/ApoB ratio (rs3806471); HDL-C, ApoA1, and ApoA1/ApoB ratio (rs4148211) in Han females were different between the genotypes (P<0.05-0.001). The levels of LDL-C in Mulao females were also different between GG and GC/CC genotypes of rs6720173 (P<0.05). The correlation between serum lipid parameters and genotypes of four SNPs was observed in Han, especially in Han males. Serum lipid parameters were also correlated with several environmental factors.The associations of four ABCG5/G8 SNPs and serum lipid levels are different between the Mulao and Han populations, or between males and females, suggesting that there may be a racial/ethnic- and/or sex-specific association between ABCG5/G8 SNPs and some serum lipid parameters

Neuropsychological effects of chronic low-dose exposure to polychlorinated biphenyls (PCBs): A cross-sectional study

BACKGROUND: Exposure to indoor air of private or public buildings contaminated with polychlorinated biphenyls (PCBs) has raised health concerns in long-term users. This exploratory neuropsychological group study investigated the potential adverse effects of chronic low-dose exposure to specific air-borne low chlorinated PCBs on well-being and behavioral measures in adult humans. METHODS: Thirty employees exposed to indoor air contaminated with PCBs from elastic sealants in a school building were compared to 30 non-exposed controls matched for education and age, controlling for gender (age range 37–61 years). PCB exposure was verified by external exposure data and biological monitoring (PCB 28, 101, 138, 153, 180). Subjective complaints, learning and memory, executive function, and visual-spatial function was assessed by standardized neuropsychological testing. Since exposure status depended on the use of contaminated rooms, an objectively exposed subgroup (N = 16; PCB 28 = 0.20 μg/l; weighted exposure duration 17.9 ± 7 years) was identified and compared with 16 paired controls. RESULTS: Blood analyses indicated a moderate exposure effect size (d) relative to expected background exposure for total PCB (4.45 ± 2.44 μg/l; d = 0.4). A significant exposure effect was found for the low chlorinated PCBs 28 (0.28 ± 0.25 μg/l; d = 1.5) and 101 (0.07 ± 0.09 μg/l; d = 0.7). Although no neuropsychological effects exceeded the adjusted significance level, estimation statistics showed elevated effect sizes for several variables. The objectively exposed subgroup showed a trend towards increased subjective attentional and emotional complaints (tiredness and slowing of practical activities, emotional state) as well as attenuated attentional performance (response shifting and alertness in a cued reaction task). CONCLUSION: Chronic inhalation of low chlorinated PCBs that involved elevated blood levels was associated with a subtle attenuation of emotional well-being and attentional function. Extended research is needed to replicate the potential long-term low PCB effects in a larger sample

Whole metagenome sequencing of chlorinated drinking water distribution systems

There has been an explosion of research into the microorganisms present within drinking water distribution systems (DWDS). However, previous studies have focused mainly on the taxonomic composition and little is known about the actual genes composing the metagenomes of DWDS and their function or whether such information could be used for genetic profiling and monitoring processes taking place in DWDS. We use here for the first time whole metagenome shotgun sequencing to characterise microbial communities from both biofilm and bulk water samples from operational, chlorinated DWDS. Gene content analysis revealed habitat-specific (biofilm vs. water) differences in terms of organisms as well as gene functions, suggesting adaptation to specific environments. In addition, several resistance mechanisms were identified preferentially within biofilms, including genes associated with the prevention and repair of disinfectant radical-induced damage and antibiotic resistance. This research highlights the potential of such information to help protect drinking water quality and safety in the future following further research and wider application

Global Retinoblastoma Presentation and Analysis by National Income Level.

Importance: Early diagnosis of retinoblastoma, the most common intraocular cancer, can save both a child's life and vision. However, anecdotal evidence suggests that many children across the world are diagnosed late. To our knowledge, the clinical presentation of retinoblastoma has never been assessed on a global scale. Objectives: To report the retinoblastoma stage at diagnosis in patients across the world during a single year, to investigate associations between clinical variables and national income level, and to investigate risk factors for advanced disease at diagnosis. Design, Setting, and Participants: A total of 278 retinoblastoma treatment centers were recruited from June 2017 through December 2018 to participate in a cross-sectional analysis of treatment-naive patients with retinoblastoma who were diagnosed in 2017. Main Outcomes and Measures: Age at presentation, proportion of familial history of retinoblastoma, and tumor stage and metastasis. Results: The cohort included 4351 new patients from 153 countries; the median age at diagnosis was 30.5 (interquartile range, 18.3-45.9) months, and 1976 patients (45.4%) were female. Most patients (n = 3685 [84.7%]) were from low- and middle-income countries (LMICs). Globally, the most common indication for referral was leukocoria (n = 2638 [62.8%]), followed by strabismus (n = 429 [10.2%]) and proptosis (n = 309 [7.4%]). Patients from high-income countries (HICs) were diagnosed at a median age of 14.1 months, with 656 of 666 (98.5%) patients having intraocular retinoblastoma and 2 (0.3%) having metastasis. Patients from low-income countries were diagnosed at a median age of 30.5 months, with 256 of 521 (49.1%) having extraocular retinoblastoma and 94 of 498 (18.9%) having metastasis. Lower national income level was associated with older presentation age, higher proportion of locally advanced disease and distant metastasis, and smaller proportion of familial history of retinoblastoma. Advanced disease at diagnosis was more common in LMICs even after adjusting for age (odds ratio for low-income countries vs upper-middle-income countries and HICs, 17.92 [95% CI, 12.94-24.80], and for lower-middle-income countries vs upper-middle-income countries and HICs, 5.74 [95% CI, 4.30-7.68]). Conclusions and Relevance: This study is estimated to have included more than half of all new retinoblastoma cases worldwide in 2017. Children from LMICs, where the main global retinoblastoma burden lies, presented at an older age with more advanced disease and demonstrated a smaller proportion of familial history of retinoblastoma, likely because many do not reach a childbearing age. Given that retinoblastoma is curable, these data are concerning and mandate intervention at national and international levels. Further studies are needed to investigate factors, other than age at presentation, that may be associated with advanced disease in LMICs

Global Retinoblastoma Presentation and Analysis by National Income Level

Importance: Early diagnosis of retinoblastoma, the most common intraocular cancer, can save both a child's life and vision. However, anecdotal evidence suggests that many children across the world are diagnosed late. To our knowledge, the clinical presentation of retinoblastoma has never been assessed on a global scale. Objectives: To report the retinoblastoma stage at diagnosis in patients across the world during a single year, to investigate associations between clinical variables and national income level, and to investigate risk factors for advanced disease at diagnosis. Design, Setting, and Participants: A total of 278 retinoblastoma treatment centers were recruited from June 2017 through December 2018 to participate in a cross-sectional analysis of treatment-naive patients with retinoblastoma who were diagnosed in 2017. Main Outcomes and Measures: Age at presentation, proportion of familial history of retinoblastoma, and tumor stage and metastasis. Results: The cohort included 4351 new patients from 153 countries; the median age at diagnosis was 30.5 (interquartile range, 18.3-45.9) months, and 1976 patients (45.4%) were female. Most patients (n = 3685 [84.7%]) were from low- A nd middle-income countries (LMICs). Globally, the most common indication for referral was leukocoria (n = 2638 [62.8%]), followed by strabismus (n = 429 [10.2%]) and proptosis (n = 309 [7.4%]). Patients from high-income countries (HICs) were diagnosed at a median age of 14.1 months, with 656 of 666 (98.5%) patients having intraocular retinoblastoma and 2 (0.3%) having metastasis. Patients from low-income countries were diagnosed at a median age of 30.5 months, with 256 of 521 (49.1%) having extraocular retinoblastoma and 94 of 498 (18.9%) having metastasis. Lower national income level was associated with older presentation age, higher proportion of locally advanced disease and distant metastasis, and smaller proportion of familial history of retinoblastoma. Advanced disease at diagnosis was more common in LMICs even after adjusting for age (odds ratio for low-income countries vs upper-middle-income countries and HICs, 17.92 [95% CI, 12.94-24.80], and for lower-middle-income countries vs upper-middle-income countries and HICs, 5.74 [95% CI, 4.30-7.68]). Conclusions and Relevance: This study is estimated to have included more than half of all new retinoblastoma cases worldwide in 2017. Children from LMICs, where the main global retinoblastoma burden lies, presented at an older age with more advanced disease and demonstrated a smaller proportion of familial history of retinoblastoma, likely because many do not reach a childbearing age. Given that retinoblastoma is curable, these data are concerning and mandate intervention at national and international levels. Further studies are needed to investigate factors, other than age at presentation, that may be associated with advanced disease in LMICs