Biosensor for Multimodal Characterization of an Essential ABC Transporter for Next-Generation Antibiotic Research
ACS Appl Mater Interfaces. 2023 Mar 3. doi: 10.1021/acsami.2c21556. Online ahead of print.
ABSTRACT
As the threat of antibiotic resistance increases, there is a particular focus on developing antimicrobials against pathogenic bacteria whose multidrug resistance is especially entrenched and concerning. One such target for novel antimicrobials is the ATP-binding cassette (ABC) transporter MsbA that is present in the plasma membrane of Gram-negative pathogenic bacteria where it is fundamental to the survival of these bacteria. Supported lipid bilayers (SLBs) are useful in monitoring membrane protein structure and function since they can be integrated with a variety of optical, biochemical, and electrochemical techniques. Here, we form SLBs containing Escherichia coli MsbA and use atomic force microscopy (AFM) and structured illumination microscopy (SIM) as high-resolution microscopy techniques to study the integrity of the SLBs and incorporated MsbA proteins. We then integrate these SLBs on microelectrode arrays (MEA) based on the conducting polymer poly(3,4-ethylenedioxy-thiophene) poly(styrene sulfonate) (PEDOT:PSS) using electrochemical impedance spectroscopy (EIS) to monitor ion flow through MsbA proteins in response to ATP hydrolysis. These EIS measurements can be correlated with the biochemical detection of MsbA-ATPase activity. To show the potential of this SLB approach, we observe not only the activity of wild-type MsbA but also the activity of two previously characterized mutants along with quinoline-based MsbA inhibitor G907 to show that EIS systems can detect changes in ABC transporter activity. Our work combines a multitude of techniques to thoroughly investigate MsbA in lipid bilayers as well as the effects of potential inhibitors of this protein. We envisage that this platform will facilitate the development of next-generation antimicrobials that inhibit MsbA or other essential membrane transporters in microorganisms.
PMID:36866935 | DOI:10.1021/acsami.2c21556
Is it realistic to use microbial photosynthesis to produce electricity directly?
PLoS Biol. 2023 Mar 2;21(3):e3001970. doi: 10.1371/journal.pbio.3001970. eCollection 2023 Mar.
ABSTRACT
It is possible to generate small amounts of electrical power directly from photosynthetic microorganisms-arguably the greenest of green energy. But will it have useful applications, and what are the hurdles if so?
PMID:36862663 | DOI:10.1371/journal.pbio.3001970
Associations between alcohol and cigarette use and type 1 and 2 myocardial infarction among people with HIV
HIV Med. 2023 Feb 28. doi: 10.1111/hiv.13466. Online ahead of print.
ABSTRACT
OBJECTIVES: People with HIV have a higher risk of myocardial infarction (MI) than the general population, with a greater proportion of type 2 MI (T2MI) due to oxygen demand-supply mismatch compared with type 1 (T1MI) resulting from atherothrombotic plaque disruption. People living with HIV report a greater prevalence of cigarette and alcohol use than do the general population. Alcohol use and smoking as risk factors for MI by type are not well studied among people living with HIV. We examined longitudinal associations between smoking and alcohol use patterns and MI by type among people living with HIV.
DESIGN AND METHODS: Using longitudinal data from the Centers for AIDS Research Network of Integrated Clinical Systems cohort, we conducted time-updated Cox proportional hazards models to determine the impact of smoking and alcohol consumption on adjudicated T1MI and T2MI.
RESULTS: Among 13 506 people living with HIV, with a median 4 years of follow-up, we observed 177 T1MI and 141 T2MI. Current smoking was associated with a 60% increase in risk of both T1MI and T2MI. In addition, every cigarette smoked per day was associated with a 4% increase in risk of T1MI, with a suggestive, but not significant, 2% increase for T2MI. Cigarette use had a greater impact on T1MI for men than for women and on T2MI for women than for men. Increasing alcohol use was associated with a lower risk of T1MI but not T2MI. Frequency of heavy episodic alcohol use was not associated with MI.
CONCLUSIONS: Our findings reinforce the prioritization of smoking reduction, even without cessation, and cessation among people living with HIV for MI prevention and highlight the different impacts on MI type by gender.
PMID:36855253 | DOI:10.1111/hiv.13466
An open label trial of nemiralisib, an inhaled PI3 kinase delta inhibitor for the treatment of Activated PI3 kinase Delta Syndrome
Pulm Pharmacol Ther. 2023 Feb 23:102201. doi: 10.1016/j.pupt.2023.102201. Online ahead of print.
ABSTRACT
Activated PI3Kδ Syndrome (APDS) is a rare inherited inborn error of immunity caused by mutations that constitutively activate the p110 delta isoform of phosphoinositide 3-kinase (PI3Kδ), resulting in recurring pulmonary infections. Currently no licensed therapies are available. Here we report the results of an open-label trial in which five subjects were treated for 12 weeks with nemiralisib, an inhaled inhibitor of PI3Kδ, to determine safety, systemic exposure, together with lung and systemic biomarker profiles (Clinicaltrial.gov: NCT02593539). Induced sputum was captured to measure changes in phospholipids and inflammatory mediators, and blood samples were collected to assess pharmacokinetics of nemiralisib, and systemic biomarkers. Nemiralisib was shown to have an acceptable safety and tolerability profile, with cough being the most common adverse event, and no severe adverse events reported during the study. No meaningful changes in phosphatidylinositol (3,4,5)-trisphosphate (PIP3; the enzyme product of PI3Kδ) or downstream inflammatory markers in induced sputum, were observed following nemiralisib treatment. Similarly, there were no meaningful changes in blood inflammatory markers, or lymphocytes subsets. Systemic levels of nemiralisib were higher in subjects in this study compared to previous observations. While nemiralisib had an acceptable safety profile, there was no convincing evidence of target engagement in the lung following inhaled dosing and no downstream effects observed in either the lung or blood compartments. We speculate that this could be explained by nemiralisib not being retained in the lung for sufficient duration, suggested by the increased systemic exposure, perhaps due to pre-existing structural lung damage. In this study investigating a small number of subjects with APDS, nemiralisib appeared to be safe and well-tolerated. However, data from this study do not support the hypothesis that inhaled treatment with nemiralisib would benefit patients with APDS.
PMID:36841351 | DOI:10.1016/j.pupt.2023.102201
Actinobacillus pleuropneumoniae encodes multiple phase-variable DNA methyltransferases that control distinct phasevarions
Nucleic Acids Res. 2023 Feb 25:gkad091. doi: 10.1093/nar/gkad091. Online ahead of print.
ABSTRACT
Actinobacillus pleuropneumoniae is the cause of porcine pleuropneumonia, a severe respiratory tract infection that is responsible for major economic losses to the swine industry. Many host-adapted bacterial pathogens encode systems known as phasevarions (phase-variable regulons). Phasevarions result from variable expression of cytoplasmic DNA methyltransferases. Variable expression results in genome-wide methylation differences within a bacterial population, leading to altered expression of multiple genes via epigenetic mechanisms. Our examination of a diverse population of A. pleuropneumoniae strains determined that Type I and Type III DNA methyltransferases with the hallmarks of phase variation were present in this species. We demonstrate that phase variation is occurring in these methyltransferases, and show associations between particular Type III methyltransferase alleles and serovar. Using Pacific BioSciences Single-Molecule, Real-Time (SMRT) sequencing and Oxford Nanopore sequencing, we demonstrate the presence of the first ever characterised phase-variable, cytosine-specific Type III DNA methyltransferase. Phase variation of distinct Type III DNA methyltransferase in A. pleuropneumoniae results in the regulation of distinct phasevarions, and in multiple phenotypic differences relevant to pathobiology. Our characterisation of these newly described phasevarions in A. pleuropneumoniae will aid in the selection of stably expressed antigens, and direct and inform development of a rationally designed subunit vaccine against this major veterinary pathogen.
PMID:36840716 | DOI:10.1093/nar/gkad091
Machine-Learning Model for Prediction of Cefepime Susceptibility in Escherichia coli from Whole-Genome Sequencing Data
J Clin Microbiol. 2023 Feb 22:e0143122. doi: 10.1128/jcm.01431-22. Online ahead of print.
ABSTRACT
The declining cost of performing bacterial whole-genome sequencing (WGS) coupled with the availability of large libraries of sequence data for well-characterized isolates have enabled the application of machine-learning (ML) methods to the development of nonlinear sequence-based predictive models. We tested the ML-based model developed by Next Gen Diagnostics for prediction of cefepime phenotypic susceptibility results in Escherichia coli. A cohort of 100 isolates of E. coli recovered from urine (n = 77) and blood (n = 23) cultures were used. The cefepime MIC was determined in triplicate by reference broth microdilution and classified as susceptible (MIC of ≤2 μg/mL) or not susceptible (MIC of ≥4 μg/mL) using the 2022 Clinical and Laboratory Standards Institute breakpoints. Five isolates generated both susceptible and not susceptible MIC results, yielding categorical agreement of 95% for the reference method to itself. Categorical agreement of ML to MIC interpretations was 97%, with 2 very major (false, susceptible) and 1 major (false, not susceptible) errors. One very major error occurred for an isolate with blaCTX-M-27 (MIC mode, ≥32 μg/mL) and one for an isolate with blaTEM-34 for which the MIC cefepime mode was 4 μg/mL. One major error was for an isolate with blaCTX-M-27 but with a MIC mode of 2 μg/mL. These preliminary data demonstrated performance of ML for a clinically important antimicrobial-species pair at a caliber similar to phenotypic methods, encouraging wider development of sequence-based susceptibility prediction and its validation and use in clinical practice.
PMID:36840604 | DOI:10.1128/jcm.01431-22
Prevalence of CFTR variants in PID patients with bronchiectasis - an important modifying co-factor
J Allergy Clin Immunol. 2023 Feb 22:S0091-6749(23)00217-8. doi: 10.1016/j.jaci.2023.01.035. Online ahead of print.
ABSTRACT
BACKGROUND: Cystic fibrosis (CF) is one of the most common life limiting autosomal recessive disorders and is caused by genetic defects in the cystic fibrosis transmembrane conductance regulator (CFTR) gene. Some of the features of this multisystem disease can be present in primary immunodeficiency (PID).
OBJECTIVE: We hypothesized that a 'carrier' CFTR status might be associated with worse outcome regarding structural lung disease in patients with PID.
METHODS: A within-cohort and population level statistical genomic analysis of a large European cohort of PID patients was performed using genome sequence data. Genomic analysis of variant pathogenicity was performed.
RESULTS: Compared to the general population, p.Phe508del carriage was enriched in lung-related PID. Additionally, carriage of several pathogenic CFTR gene variants were increased in PID associated with structural lung damage, compared to PID patients without the structural lung damage. We identified three additional biallelic cases including several variants not traditionally considered as CF-causing.
CONCLUSION: Genome sequencing identified cases of CFTR dysfunction in PID, driving an increased susceptibility to infection. Large national genomics services provide an opportunity for precision medicine by interpreting subtle features of genomic diversity when treating traditional Mendelian disorders.
PMID:36828084 | DOI:10.1016/j.jaci.2023.01.035
High-Throughput Mutagenesis Reveals a Role for Antimicrobial Resistance- and Virulence-Associated Mobile Genetic Elements in Staphylococcus aureus Host Adaptation
Microbiol Spectr. 2023 Feb 23:e0421322. doi: 10.1128/spectrum.04213-22. Online ahead of print.
ABSTRACT
Methicillin-resistant Staphylococcus aureus (MRSA) clonal-complex 398 (CC398) is the dominant livestock-associated (LA) MRSA lineage in European livestock and an increasing cause of difficult-to-treat human disease. LA-CC398 MRSA evolved from a diverse human-associated methicillin-sensitive population, and this transition from humans to livestock was associated with three mobile genetic elements (MGEs). In this study, we apply transposon-directed insertion site sequencing (TraDIS), a high-throughput transposon mutagenesis approach, to investigate genetic signatures that contribute to LA-CC398 causing disease in humans. We identified 26 genes associated with LA-CC398 survival in human blood and 47 genes in porcine blood. We carried out phylogenetic reconstruction on 1,180 CC398 isolates to investigate the genetic context of all identified genes. We found that all genes associated with survival in human blood were part of the CC398 core genome, while 2/47 genes essential for survival in porcine blood were located on MGEs. Gene SAPIG0966 was located on the previously identified Tn916 transposon carrying a tetracycline resistance gene, which has been shown to be stably inherited within LA-CC398. Gene SAPIG1525 was carried on a phage element, which in part, matched phiSa2wa_st1, a previously identified bacteriophage carrying the Panton-Valentine leucocidin (PVL) virulence factor. Gene deletion mutants constructed in two LA-CC398 strains confirmed that the SAPIG0966 carrying Tn916 and SAPIG1525 were important for CC398 survival in porcine blood. Our study shows that MGEs that carry antimicrobial resistance and virulence genes could have a secondary function in bacterial survival in blood and may be important for host adaptation. IMPORTANCE CC398 is the dominant type of methicillin-resistant Staphylococcus aureus (MRSA) in European livestock and a growing cause of human infections. Previous studies have suggested MRSA CC398 evolved from human-associated methicillin-sensitive Staphylococcus aureus and is capable of rapidly readapting to human hosts while maintaining antibiotic resistance. Using high-throughput transposon mutagenesis, our study identified 26 and 47 genes important for MRSA CC398 survival in human and porcine blood, respectively. Two of the genes important for MRSA CC398 survival in porcine blood were located on mobile genetic elements (MGEs) carrying resistance or virulence genes. Our study shows that these MGEs carrying antimicrobial resistance and virulence genes could have a secondary function in bacterial survival in blood and may be important for blood infection and host adaptation.
PMID:36815781 | DOI:10.1128/spectrum.04213-22
Identifying ways of producing pigs more sustainably: tradeoffs and co-benefits in land and antimicrobial use
Sci Rep. 2023 Feb 17;13(1):2840. doi: 10.1038/s41598-023-29480-5.
ABSTRACT
Pork accounts for the largest proportion of meat consumed globally and demand is growing rapidly. Two important externalities of pig farming are land use and antimicrobial resistance (AMR) driven by antimicrobial use (AMU). Land use and AMU are commonly perceived to be negatively related across different production systems, so those with smaller land footprints pose greater risk to human health. However, the relationship between land use and AMU has never been systematically evaluated. We addressed this by measuring both outcomes for 74 highly diverse pig production systems. We found weak evidence of an AMU/land use tradeoff. We also found several systems characterized by low externality costs in both domains. These potentially promising systems were spread across different label and husbandry types and indeed no type was a reliable indicator of low-cost systems in both externalities. Our findings highlight the importance of using empirical evidence in decision-making, rather than relying on assumptions.
PMID:36804956 | DOI:10.1038/s41598-023-29480-5
Genome-wide subcellular protein map for the flagellate parasite Trypanosoma brucei
Nat Microbiol. 2023 Feb 20. doi: 10.1038/s41564-022-01295-6. Online ahead of print.
ABSTRACT
Trypanosoma brucei is a model trypanosomatid, an important group of human, animal and plant unicellular parasites. Understanding their complex cell architecture and life cycle is challenging because, as with most eukaryotic microbes, ~50% of genome-encoded proteins have completely unknown functions. Here, using fluorescence microscopy and cell lines expressing endogenously tagged proteins, we mapped the subcellular localization of 89% of the T. brucei proteome, a resource we call TrypTag. We provide clues to function and define lineage-specific organelle adaptations for parasitism, mapping the ultraconserved cellular architecture of eukaryotes, including the first comprehensive 'cartographic' analysis of the eukaryotic flagellum, which is vital for morphogenesis and pathology. To demonstrate the power of this resource, we identify novel organelle subdomains and changes in molecular composition through the cell cycle. TrypTag is a transformative resource, important for hypothesis generation for both eukaryotic evolutionary molecular cell biology and fundamental parasite cell biology.
PMID:36804636 | DOI:10.1038/s41564-022-01295-6
Evidence of widespread endemic populations of highly multidrug resistant Klebsiella pneumoniae in hospital settings in Hanoi, Vietnam: a prospective cohort study
Lancet Microbe. 2023 Feb 16:S2666-5247(22)00338-X. doi: 10.1016/S2666-5247(22)00338-X. Online ahead of print.
ABSTRACT
BACKGROUND: Patients with prolonged hospitalisation have a significant risk of carriage of and subsequent infection with extended spectrum β-lactamase (ESBL)-producing and carbapenemase-producing Klebsiella pneumoniae. However, the distinctive roles of the community and hospital environments in the transmission of ESBL-producing or carbapenemase-producing K pneumoniae remain elusive. We aimed to investigate the prevalence and transmission of K pneumoniae within and between the two tertiary hospitals in Hanoi, Viet Nam, using whole-genome sequencing.
METHODS: We did a prospective cohort study of 69 patients in intensive care units (ICUs) from two hospitals in Hanoi, Viet Nam. Patients were included if they were aged 18 years or older, admitted for longer than the mean length of stay in their ICU, and cultured K pneumoniae from their clinical samples. Longitudinally collected samples from patients (collected weekly) and the ICU environment (collected monthly) were cultured on selective media, and whole-genome sequences from K pneumoniae colonies analysed. We did phylogenetic analyses and correlated phenotypic antimicrobial susceptibility testing with genotypic features of K pneumoniae isolates. We constructed transmission networks of patient samples, relating ICU admission times and locations with genetic similarity of infecting K pneumoniae.
FINDINGS: Between June 1, 2017, and Jan 31, 2018, 69 patients were in the ICUs and eligible for inclusion, and a total of 357 K pneumoniae isolates were cultured and successfully sequenced. 228 (64%) of K pneumoniae isolates carried between two and four different ESBL-encoding and carbapenemase-encoding genes, with 164 (46%) isolates carrying genes encoding both, with high minimum inhibitory concentrations. We found a novel co-occurrence of blaKPC-2 and blaNDM-1 in 46·6% of samples from the globally successful ST15 lineage. Despite being physically and clinically separated, the two hospitals shared closely related strains carrying the same array of antimicrobial resistance genes.
INTERPRETATION: These results highlight the high prevalence of ESBL-positive carbapenem-resistant K pneumoniae in ICUs in Viet Nam. Through studying K pneumoniae ST15 in detail, we showed how important resistance genes are contained within these strains that are carried broadly by patients entering the two hospitals directly or through referral.
FUNDING: Medical Research Council Newton Fund, Ministry of Science and Technology, Wellcome Trust, Academy of Medical Sciences, Health Foundation, and National Institute for Health and Care Research Cambridge Biomedical Research Centre.
PMID:36801013 | DOI:10.1016/S2666-5247(22)00338-X
A systematic review of economic evaluations of whole-genome sequencing for the surveillance of bacterial pathogens
Microb Genom. 2023 Feb;9(2). doi: 10.1099/mgen.0.000947.
ABSTRACT
Whole-genome sequencing (WGS) has unparalleled ability to distinguish between bacteria, with many public health applications. The generation and analysis of WGS data require significant financial investment. We describe a systematic review summarizing economic analyses of genomic surveillance of bacterial pathogens, reviewing the evidence for economic viability. The protocol was registered on PROSPERO (CRD42021289030). Six databases were searched on 8 November 2021 using terms related to 'WGS', 'population surveillance' and 'economic analysis'. Quality was assessed with the Drummond-Jefferson checklist. Following data extraction, a narrative synthesis approach was taken. Six hundred and eighty-one articles were identified, of which 49 proceeded to full-text screening, with 9 selected for inclusion. All had been published since 2019. Heterogeneity was high. Five studies assessed WGS for hospital surveillance and four analysed foodborne pathogens. Four were cost-benefit analyses, one was a cost-utility analysis, one was a cost-effectiveness analysis, one was a combined cost-effectiveness and cost-utility analysis, one combined cost-effectiveness and cost-benefit analyses and one was a partial analysis. All studies supported the use of WGS as a surveillance tool on economic grounds. The available evidence supports the use of WGS for pathogen surveillance but is limited by marked heterogeneity. Further work should include analysis relevant to low- and middle-income countries and should use real-world effectiveness data.
PMID:36790430 | DOI:10.1099/mgen.0.000947
DNA nanostructures as a tool for targeted antimicrobial delivery
Biophys J. 2023 Feb 10;122(3S1):550a. doi: 10.1016/j.bpj.2022.11.2910.
NO ABSTRACT
PMID:36784859 | DOI:10.1016/j.bpj.2022.11.2910
Author Correction: Tracking SARS-COV-2 variants using Nanopore sequencing in Ukraine in 2021
Sci Rep. 2023 Feb 13;13(1):2555. doi: 10.1038/s41598-023-29749-9.
NO ABSTRACT
PMID:36781923 | DOI:10.1038/s41598-023-29749-9
Immunoinformatics design of multivalent epitope vaccine against monkeypox virus and its variants using membrane-bound, enveloped, and extracellular proteins as targets
Front Immunol. 2023 Jan 26;14:1091941. doi: 10.3389/fimmu.2023.1091941. eCollection 2023.
ABSTRACT
INTRODUCTION: The current monkeypox (MPX) outbreak, caused by the monkeypox virus (MPXV), has turned into a global concern, with over 59,000 infection cases and 23 deaths worldwide.
OBJECTIVES: Herein, we aimed to exploit robust immunoinformatics approach, targeting membrane-bound, enveloped, and extracellular proteins of MPXV to formulate a chimeric antigen. Such a strategy could similarly be applied for identifying immunodominant epitopes and designing multi-epitope vaccine ensembles in other pathogens responsible for chronic pathologies that are difficult to intervene against.
METHODS: A reverse vaccinology pipeline was used to select 11 potential vaccine candidates, which were screened and mapped to predict immunodominant B-cell and T-cell epitopes. The finalized epitopes were merged with the aid of suitable linkers, an adjuvant (Resuscitation-promoting factor), a PADRE sequence (13 aa), and an HIV TAT sequence (11 aa) to formulate a multivalent epitope vaccine. Bioinformatics tools were employed to carry out codon adaptation and computational cloning. The tertiary structure of the chimeric vaccine construct was modeled via I-TASSER, and its interaction with Toll-like receptor 4 (TLR4) was evaluated using molecular docking and molecular dynamics simulation. C-ImmSim server was implemented to examine the immune response against the designed multi-epitope antigen.
RESULTS AND DISCUSSION: The designed chimeric vaccine construct included 21 immunodominant epitopes (six B-cell, eight cytotoxic T lymphocyte, and seven helper T-lymphocyte) and is predicted non-allergen, antigenic, soluble, with suitable physicochemical features, that can promote cross-protection among the MPXV strains. The selected epitopes indicated a wide global population coverage (93.62%). Most finalized epitopes have 70%-100% sequence similarity with the experimentally validated immune epitopes of the vaccinia virus, which can be helpful in the speedy progression of vaccine design. Lastly, molecular docking and molecular dynamics simulation computed stable and energetically favourable interaction between the putative antigen and TLR4.
CONCLUSION: Our results show that the multi-epitope vaccine might elicit cellular and humoral immune responses and could be a potential vaccine candidate against the MPXV infection. Further experimental testing of the proposed vaccine is warranted to validate its safety and efficacy profile.
PMID:36776835 | PMC:PMC9908764 | DOI:10.3389/fimmu.2023.1091941
Induction of aphid resistance in tobacco by the cucumber mosaic virus CMV∆2b mutant is jasmonate-dependent
Mol Plant Pathol. 2023 Feb 12. doi: 10.1111/mpp.13305. Online ahead of print.
ABSTRACT
Cucumber mosaic virus (CMV) is vectored by aphids, including Myzus persicae. Tobacco (Nicotiana tabacum 'Xanthi') plants infected with a mutant of the Fny strain of CMV (Fny-CMVΔ2b, which cannot express the CMV 2b protein) exhibit strong resistance against M. persicae, which is manifested by decreased survival and reproduction of aphids confined on the plants. Previously, we found that the Fny-CMV 1a replication protein elicits aphid resistance in plants infected with Fny-CMVΔ2b, whereas in plants infected with wild-type Fny-CMV this is counteracted by the CMV 2b protein, a counterdefence protein that, among other things, inhibits jasmonic acid (JA)-dependent immune signalling. We noted that in nontransformed cv. Petit Havana SR1 tobacco plants aphid resistance was not induced by Fny-CMVΔ2b, suggesting that not all tobacco varieties possess the factor(s) with which the 1a protein interacts. To determine if 1a protein-induced aphid resistance is JA-dependent in Xanthi tobacco, transgenic plants were made that expressed an RNA silencing construct to diminish expression of the JA co-receptor CORONATINE-INSENSITIVE 1. Fny-CMVΔ2b did not induce resistance to M. persicae in these transgenic plants. Thus, aphid resistance induction by the 1a protein requires JA-dependent defensive signalling, which is countered by the CMV 2b protein.
PMID:36775660 | DOI:10.1111/mpp.13305
The human proton pump inhibitors inhibit <em>Mycobacterium tuberculosis</em> rifampicin efflux and macrophage-induced rifampicin tolerance
Proc Natl Acad Sci U S A. 2023 Feb 14;120(7):e2215512120. doi: 10.1073/pnas.2215512120. Epub 2023 Feb 10.
ABSTRACT
Tuberculosis treatment requires months-long combination chemotherapy with multiple drugs, with shorter treatments leading to relapses. A major impediment to shortening treatment is that Mycobacterium tuberculosis becomes tolerant to the administered drugs, starting early after infection and within days of infecting macrophages. Multiple lines of evidence suggest that macrophage-induced drug tolerance is mediated by mycobacterial drug efflux pumps. Here, using assays to directly measure drug efflux, we find that M. tuberculosis transports the first-line antitubercular drug rifampicin through a proton gradient-dependent mechanism. We show that verapamil, a known efflux pump inhibitor, which inhibits macrophage-induced rifampicin tolerance, also inhibits M.tuberculosis rifampicin efflux. As with macrophage-induced tolerance, the calcium channel-inhibiting property of verapamil is not required for its inhibition of rifampicin efflux. By testing verapamil analogs, we show that verapamil directly inhibits M. tuberculosis drug efflux pumps through its human P-glycoprotein (PGP)-like inhibitory activity. Screening commonly used drugs with incidental PGP inhibitory activity, we find many inhibit rifampicin efflux, including the proton pump inhibitors (PPIs) such as omeprazole. Like verapamil, the PPIs inhibit macrophage-induced rifampicin tolerance as well as intramacrophage growth, which has also been linked to mycobacterial efflux pump activity. Our assays provide a facile screening platform for M. tuberculosis efflux pump inhibitors that inhibit in vivo drug tolerance and growth.
PMID:36763530 | DOI:10.1073/pnas.2215512120
30 years of Acta D
Acta Crystallogr D Struct Biol. 2023 Feb 1;79(Pt 2):95-97. doi: 10.1107/S2059798323001006. Epub 2023 Feb 8.
NO ABSTRACT
PMID:36762854 | DOI:10.1107/S2059798323001006
Coughs, colds and "freshers' flu" survey in the University of Cambridge, 2007-2008
Epidemics. 2022 Dec 23;42:100659. doi: 10.1016/j.epidem.2022.100659. Online ahead of print.
ABSTRACT
Universities provide many opportunities for the spread of infectious respiratory illnesses. Students are brought together into close proximity from all across the world and interact with one another in their accommodation, through lectures and small group teaching and in social settings. The COVID-19 global pandemic has highlighted the need for sufficient data to help determine which of these factors are important for infectious disease transmission in universities and hence control university morbidity as well as community spillover. We describe the data from a previously unpublished self-reported university survey of coughs, colds and influenza-like symptoms collected in Cambridge, UK, during winter 2007-2008. The online survey collected information on symptoms and socio-demographic, academic and lifestyle factors. There were 1076 responses, 97% from University of Cambridge students (5.7% of the total university student population), 3% from staff and <1% from other participants, reporting onset of symptoms between September 2007 and March 2008. Undergraduates are seen to report symptoms earlier in the term than postgraduates; differences in reported date of symptoms are also seen between subjects and accommodation types, although these descriptive results could be confounded by survey biases. Despite the historical and exploratory nature of the study, this is one of few recent detailed datasets of influenza-like infection in a university context and is especially valuable to share now to improve understanding of potential transmission dynamics in universities during the current COVID-19 pandemic.
PMID:36758342 | DOI:10.1016/j.epidem.2022.100659
Chemical Validation of Mycobacterium tuberculosis Phosphopantetheine Adenylyltransferase Using Fragment Linking and CRISPR Interference
Angew Chem Int Ed Engl. 2023 Feb 9. doi: 10.1002/anie.202300221. Online ahead of print.
ABSTRACT
The coenzyme A (CoA) biosynthesis pathway has attracted attention as a potential target for much-needed novel antimicrobial drugs, including for the treatment of tuberculosis (TB), the lethal disease caused by Mycobacterium tuberculosis (Mtb). Seeking to identify inhibitors of Mtb phosphopantetheine adenylyltransferase (MtbPPAT), the enzyme that catalyses the penultimate step in CoA biosynthesis, we performed a fragment screen. In doing so, we discovered three series of fragments that occupy distinct regions of the MtbPPAT active site, presenting a unique opportunity for fragment linking. Here we show how, guided by X-ray crystal structures, we could link weakly-binding fragments to produce an active site binder with a KD < 20 µM and on-target anti-Mtb activity, as demonstrated using CRISPR interference. This study represents a big step toward validating MtbPPAT as a potential drug target and designing a MtbPPAT-targeting anti-TB drug.
PMID:36757665 | DOI:10.1002/anie.202300221