608,811 results on '"ESCHERICHIA coli"'
Search Results
2. Multiple-drug resistant Shiga toxin-producing E. coli in raw milk of dairy bovine
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Ullah, Safir, Khan, Saeed Ul Hassan, Khan, Muhammad Jamil, Khattak, Baharullah, Fozia, Fozia, Ahmad, Ijaz, Wadaan, Mohammad Ahmad, Khan, Muhammad Farooq, Baabbad, Almohannad, and Goyal, Sagar M
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- 2024
3. Characteristics of pathogenic Escherichia coli associated with diarrhea in children under five years in northwestern Ethiopia
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Mulu, Berihun Mossie, Belete, Mequanint Addisu, Bialfew Demlie, Tiliksew, Tassew, Habtamu, and Tessema, Tesfaye Sisay
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- 2024
4. Rationally seeded computational protein design of ɑ-helical barrels.
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Albanese, Katherine, Petrenas, Rokas, Pirro, Fabio, Naudin, Elise, Borucu, Ufuk, Dawson, William, Scott, D, Leggett, Graham, Weiner, Orion, Oliver, Thomas, and Woolfson, Derek
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Escherichia coli ,Proteins ,Protein Conformation ,alpha-Helical ,Protein Engineering ,Models ,Molecular ,Peptides ,Computational Biology ,Amino Acid Sequence ,Protein Folding - Abstract
Computational protein design is advancing rapidly. Here we describe efficient routes starting from validated parallel and antiparallel peptide assemblies to design two families of α-helical barrel proteins with central channels that bind small molecules. Computational designs are seeded by the sequences and structures of defined de novo oligomeric barrel-forming peptides, and adjacent helices are connected by loop building. For targets with antiparallel helices, short loops are sufficient. However, targets with parallel helices require longer connectors; namely, an outer layer of helix-turn-helix-turn-helix motifs that are packed onto the barrels. Throughout these computational pipelines, residues that define open states of the barrels are maintained. This minimizes sequence sampling, accelerating the design process. For each of six targets, just two to six synthetic genes are made for expression in Escherichia coli. On average, 70% of these genes express to give soluble monomeric proteins that are fully characterized, including high-resolution structures for most targets that match the design models with high accuracy.
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- 2024
5. Guide RNA structure design enables combinatorial CRISPRa programs for biosynthetic profiling.
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Fontana, Jason, Sparkman-Yager, David, Faulkner, Ian, Cardiff, Ryan, Kiattisewee, Cholpisit, Walls, Aria, Primo, Tommy, Kinnunen, Patrick, Garcia Martin, Hector, Zalatan, Jesse, and Carothers, James
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Escherichia coli ,RNA ,Guide ,CRISPR-Cas Systems ,CRISPR-Cas Systems ,Metabolic Engineering ,Biosynthetic Pathways ,Promoter Regions ,Genetic ,Humans ,Gene Expression Regulation ,Bacterial ,RNA Folding - Abstract
Engineering metabolism to efficiently produce chemicals from multi-step pathways requires optimizing multi-gene expression programs to achieve enzyme balance. CRISPR-Cas transcriptional control systems are emerging as important tools for programming multi-gene expression, but poor predictability of guide RNA folding can disrupt expression control. Here, we correlate efficacy of modified guide RNAs (scRNAs) for CRISPR activation (CRISPRa) in E. coli with a computational kinetic parameter describing scRNA folding rate into the active structure (rS = 0.8). This parameter also enables forward design of scRNAs, allowing us to design a system of three synthetic CRISPRa promoters that can orthogonally activate (>35-fold) expression of chosen outputs. Through combinatorial activation tuning, we profile a three-dimensional design space expressing two different biosynthetic pathways, demonstrating variable production of pteridine and human milk oligosaccharide products. This RNA design approach aids combinatorial optimization of metabolic pathways and may accelerate routine design of effective multi-gene regulation programs in bacterial hosts.
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- 2024
6. The hallmarks of a tradeoff in transcriptomes that balances stress and growth functions.
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Dalldorf, Christopher, Rychel, Kevin, Szubin, Richard, Hefner, Ying, Patel, Arjun, Zielinski, Daniel, and Palsson, Bernhard
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Escherichia coli ,ribosomes ,sigma factors ,transcriptional regulation ,Transcriptome ,Escherichia coli ,DNA-Directed RNA Polymerases ,Mutation ,Stress ,Physiological ,Gene Expression Regulation ,Bacterial ,Escherichia coli Proteins ,Sigma Factor ,Adaptation ,Physiological - Abstract
Fast growth phenotypes are achieved through optimal transcriptomic allocation, in which cells must balance tradeoffs in resource allocation between diverse functions. One such balance between stress readiness and unbridled growth in E. coli has been termed the fear versus greed (f/g) tradeoff. Two specific RNA polymerase (RNAP) mutations observed in adaptation to fast growth have been previously shown to affect the f/g tradeoff, suggesting that genetic adaptations may be primed to control f/g resource allocation. Here, we conduct a greatly expanded study of the genetic control of the f/g tradeoff across diverse conditions. We introduced 12 RNA polymerase (RNAP) mutations commonly acquired during adaptive laboratory evolution (ALE) and obtained expression profiles of each. We found that these single RNAP mutation strains resulted in large shifts in the f/g tradeoff primarily in the RpoS regulon and ribosomal genes, likely through modifying RNAP-DNA interactions. Two of these mutations additionally caused condition-specific transcriptional adaptations. While this tradeoff was previously characterized by the RpoS regulon and ribosomal expression, we find that the GAD regulon plays an important role in stress readiness and ppGpp in translation activity, expanding the scope of the tradeoff. A phylogenetic analysis found the greed-related genes of the tradeoff present in numerous bacterial species. The results suggest that the f/g tradeoff represents a general principle of transcriptome allocation in bacteria where small genetic changes can result in large phenotypic adaptations to growth conditions.IMPORTANCETo increase growth, E. coli must raise ribosomal content at the expense of non-growth functions. Previous studies have linked RNAP mutations to this transcriptional shift and increased growth but were focused on only two mutations found in the proteins central region. RNAP mutations, however, commonly occur over a large structural range. To explore RNAP mutations impact, we have introduced 12 RNAP mutations found in laboratory evolution experiments and obtained expression profiles of each. The mutations nearly universally increased growth rates by adjusting said tradeoff away from non-growth functions. In addition to this shift, a few caused condition-specific adaptations. We explored the prevalence of this tradeoff across phylogeny and found it to be a widespread and conserved trend among bacteria.
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- 2024
7. Molecular Engineering of Functional SiRNA Agents
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Batra, Neelu, Tu, Mei-Juan, and Yu, Ai-Ming
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Biological Sciences ,Industrial Biotechnology ,Biotechnology ,Bioengineering ,Genetics ,1.3 Chemical and physical sciences ,Underpinning research ,RNA ,Small Interfering ,Humans ,Synthetic Biology ,RNA ,Transfer ,Proto-Oncogene Proteins c-bcl-2 ,Escherichia coli ,Genetic Engineering ,Green Fluorescent Proteins ,MicroRNAs ,RNA engineering ,small interfering RNA ,RNAinterference ,therapy ,RNA interference ,Medicinal and Biomolecular Chemistry ,Biochemistry and Cell Biology ,Biomedical Engineering ,Biochemistry and cell biology ,Bioinformatics and computational biology - Abstract
Synthetic biology constitutes a scientific domain focused on intentional redesign of organisms to confer novel functionalities or create new products through strategic engineering of their genetic makeup. Leveraging the inherent capabilities of nature, one may address challenges across diverse sectors including medicine. Inspired by this concept, we have developed an innovative bioengineering platform, enabling high-yield and large-scale production of biological small interfering RNA (BioRNA/siRNA) agents via bacterial fermentation. Herein, we show that with the use of a new tRNA fused pre-miRNA carrier, we can produce various forms of BioRNA/siRNA agents within living host cells. We report a high-level overexpression of nine target BioRNA/siRNA molecules at 100% success rate, yielding 3-10 mg of BioRNA/siRNA per 0.25 L of bacterial culture with high purity (>98%) and low endotoxin (
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- 2024
8. Proteome allocation is linked to transcriptional regulation through a modularized transcriptome.
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Patel, Arjun, McGrosso, Dominic, Hefner, Ying, Campeau, Anaamika, Sastry, Anand, Maurya, Svetlana, Rychel, Kevin, Gonzalez, David, and Palsson, Bernhard
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Proteome ,Transcriptome ,Gene Expression Regulation ,Bacterial ,Bacterial Proteins ,Transcription ,Genetic ,Bacteria ,Gene Expression Profiling ,Escherichia coli ,Proteomics - Abstract
It has proved challenging to quantitatively relate the proteome to the transcriptome on a per-gene basis. Recent advances in data analytics have enabled a biologically meaningful modularization of the bacterial transcriptome. We thus investigate whether matched datasets of transcriptomes and proteomes from bacteria under diverse conditions can be modularized in the same way to reveal novel relationships between their compositions. We find that; (1) the modules of the proteome and the transcriptome are comprised of a similar list of gene products, (2) the modules in the proteome often represent combinations of modules from the transcriptome, (3) known transcriptional and post-translational regulation is reflected in differences between two sets of modules, allowing for knowledge-mapping when interpreting module functions, and (4) through statistical modeling, absolute proteome allocation can be inferred from the transcriptome alone. Quantitative and knowledge-based relationships can thus be found at the genome-scale between the proteome and transcriptome in bacteria.
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- 2024
9. WGS of intrauterine E. coli from cows with early postpartum uterine infection reveals a non-uterine specific genotype and virulence factors.
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Garzon, Adriana, Basbas, Carl, Schlesener, Cory, Silva-Del-Rio, Noelia, Karle, Betsy, Lima, Fabio, Weimer, Bart, and Pereira, Richard
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antibiotic resistance ,cattle ,mutation ,uterine infection ,whole-genome sequencing ,Cattle ,Animals ,Female ,Virulence Factors ,Escherichia coli Infections ,Escherichia coli ,Genotype ,Cattle Diseases ,Postpartum Period ,Cross-Sectional Studies ,Whole Genome Sequencing ,Uterine Diseases ,Genome ,Bacterial ,Uterus ,Anti-Bacterial Agents ,Genome-Wide Association Study ,Drug Resistance ,Bacterial - Abstract
Escherichia coli has been attributed to playing a major role in a cascade of events that affect the prevalence and severity of uterine disease in cattle. The objectives of this project were to (i) define the association between the prevalence of specific antimicrobial resistance and virulence factor genes in E. coli with the clinical status related to uterine infection, (ii) identify the genetic relationship between E. coli isolates from cows with diarrhea, with mastitis, and with and without metritis, and (iii) determine the association between the phenotypic and genotypic antimicrobial resistance identified on the E. coli isolated from postpartum cattle. Bacterial isolates (n = 148) were obtained from a larger cross-sectional study. Cows were categorized into one of three clinical groups before enrollment: metritis, cows with purulent discharge, and control cows. For genomic comparison, public genomes (n = 130) from cows with diarrhea, mastitis, and metritis were included in a genome-wide association study, to evaluate differences between the drug classes or the virulence factor category among clinical groups. A distinct E. coli genotype associated with metritis could not be identified. Instead, a high genetic diversity among the isolates from uterine sources was present. A virulence factor previously associated with metritis (fimH) using PCR was not associated with metritis. There was moderate accuracy for whole-genome sequencing to predict phenotypic resistance, which varied depending on the antimicrobial tested. Findings from this study contradict the traditional pathotype classification and the unique intrauterine E. coli genotype associated with metritis in dairy cows.IMPORTANCEMetritis is a common infectious disease in dairy cattle and the second most common reason for treating a cow with antimicrobials. The pathophysiology of the disease is complex and is not completely understood. Specific endometrial pathogenic Escherichia coli have been reported to be adapted to the endometrium and sometimes lead to uterine disease. Unfortunately, the specific genomic details of the endometrial-adapted isolates have not been investigated using enough genomes to represent the genomic diversity of this organism to identify specific virulence genes that are consistently associated with disease development and severity. Results from this study provide key microbial ecological advances by elucidating and challenging accepted concepts for the role of Intrauterine E. coli in metritis in dairy cattle, especially contradicting the existence of a unique intrauterine E. coli genotype associated with metritis in dairy cows, which was not found in our study.
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- 2024
10. Draft genome sequence of Escherichia coli MTR_GS_S1457 strain isolated from a soil sample of a vegetable garden in Gazipur, Bangladesh.
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Islam, Md Saiful, Pramanik, Pritom, Rana, Md, Ullah, Md, Neloy, Fahim, Ramasamy, Srinivasan, Schreinemachers, Pepijn, Oliva, Ricardo, and Rahman, Md
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Bangladesh ,Escherichia coli ,environment ,soil ,vegetable gardening system ,whole-genome sequencing - Abstract
We announce the sequence of the Escherichia coli MTR_GS_S1457 strain isolated from a soil sample of a vegetable gardening system for the first time in Bangladesh. With a length of 4,918,647 bp, this strain contained one plasmid, two CRISPR arrays, 54 predicted antibiotic resistance genes, and 81 predicted virulence factor genes.
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- 2024
11. Addressing Fecal Contamination in Rural Kenyan Households: The Roles of Environmental Interventions and Animal Ownership.
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Swarthout, Jenna, Mureithi, Maryanne, Mboya, John, Arnold, Benjamin, Wolfe, Marlene, Dentz, Holly, Lin, Audrie, Arnold, Charles, Rao, Gouthami, Stewart, Christine, Clasen, Thomas, Colford, John, Null, Clair, and Pickering, Amy
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Escherichia coli ,WASH ,diarrhea ,handwashing ,sanitation ,stunting ,transmission pathway ,water ,Feces ,Animals ,Kenya ,Humans ,Family Characteristics ,Escherichia coli ,Rural Population ,Drinking Water ,Sanitation ,Hand Disinfection ,Water Microbiology ,Ownership ,Diarrhea - Abstract
Combined water, sanitation, and handwashing (WSH) interventions could reduce fecal contamination along more transmission pathways than single interventions alone. We measured Escherichia coli levels in 3909 drinking water samples, 2691 child hand rinses, and 2422 toy ball rinses collected from households enrolled in a 2-year cluster-randomized controlled trial evaluating single and combined WSH interventions. Water treatment with chlorine reduced E. coli in drinking water. A combined WSH intervention improved water quality by the same magnitude but did not affect E. coli levels on hands or toys. One potential explanation for the limited impact of the sanitation intervention (upgraded latrines) is failure to address dog and livestock fecal contamination. Small ruminant (goat or sheep) ownership was associated with increased E. coli levels in stored water and on child hands. Cattle and poultry ownership was protective against child stunting, and domesticated animal ownership was not associated with child diarrhea. Our findings do not support restricting household animal ownership to prevent child diarrheal disease or stunting but do support calls for WSH infrastructure that can more effectively reduce household fecal contamination.
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- 2024
12. New and old lessons from a devastating case of neonatal E coli meningitis.
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Saleh, Tawny, Kamau, Edwin, and Rathe, Jennifer
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Acridine orange stain ,CSF sterility ,Neonatal E coli meningitis ,Neonatal sepsis ,Humans ,Infant ,Newborn ,Anti-Bacterial Agents ,Infant ,Premature ,Infant ,Premature ,Diseases ,Meningitis ,Escherichia coli - Abstract
BACKGROUND: Neonatal Escherichia coli (E coli) meningitis results in significant morbidity and mortality. We present a case of a premature infant with extensive central nervous system (CNS) injury from recurrent E coli infection and the non-traditional methods necessary to identify and clear the infection. CASE PRESENTATION: The infant was transferred to our institutions pediatric intensive care unit (PICU) after recurrence of E coli CNS infection requiring neurosurgical intervention. He had been treated for early onset sepsis (EOS) with ampicillin and gentamicin for 10 days followed by rapid development of ampicillin-resistant E coli septic shock and meningitis after discontinuation of antibiotics. Sterility of the CNS was not confirmed at the end of 21 days of cefepime therapy and was subsequently followed by recurrent ampicillin-resistant E coli septic shock and CNS infection. Despite 6 weeks of appropriate therapy with sterility of CSF by traditional methods, he suffered from intractable seizures with worsening hydrocephalus. Transferred to our institution, he underwent endoscopic 3rd ventriculostomy with cyst fenestration revealing purulent fluid and significant pleocytosis. An additional 3 weeks of systemic and intraventricular antibiotics with cefepime and tobramycin were given but a significant CNS neutrophil-predominant pleocytosis persisted (average of ∼ 21,000 cells/mm3). Repeated gram stains, cultures, polymerase chain reaction (PCR) testing, and metagenomic next generation sequencing (NGS) testing of CSF were negative for pathogens but acridine orange stain (AO) revealed numerous intact rod-shaped bacteria. After the addition of ciprofloxacin, sterility and resolution of CSF pleocytosis was finally achieved. CONCLUSION: Neonatal E coli meningitis is a well-known entity but unlike other bacterial infections, it has not proven amenable to shorter, more narrow-spectrum antibiotic courses or limiting invasive procedures such as lumbar punctures. Further, microbiologic techniques to determine CSF sterility suffer from poorly understood limitations leading to premature discontinuation of antibiotics risking further neurologic damage in vulnerable hosts.
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- 2024
13. Proteome partitioning constraints in long-term laboratory evolution.
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Mori, Matteo, Patsalo, Vadim, Euler, Christian, Williamson, James, and Scott, Matthew
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Proteome ,Escherichia coli ,Directed Molecular Evolution ,Glucose ,Adaptation ,Physiological ,Gene Expression Regulation ,Bacterial ,Escherichia coli Proteins ,Glycolysis - Abstract
Adaptive laboratory evolution experiments provide a controlled context in which the dynamics of selection and adaptation can be followed in real-time at the single-nucleotide level. And yet this precision introduces hundreds of degrees-of-freedom as genetic changes accrue in parallel lineages over generations. On short timescales, physiological constraints have been leveraged to provide a coarse-grained view of bacterial gene expression characterized by a small set of phenomenological parameters. Here, we ask whether this same framework, operating at a level between genotype and fitness, informs physiological changes that occur on evolutionary timescales. Using a strain adapted to growth in glucose minimal medium, we find that the proteome is substantially remodeled over 40 000 generations. The most striking change is an apparent increase in enzyme efficiency, particularly in the enzymes of lower-glycolysis. We propose that deletion of metabolic flux-sensing regulation early in the adaptation results in increased enzyme saturation and can account for the observed proteome remodeling.
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- 2024
14. Engineering an Escherichia coli strain for production of long single-stranded DNA
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Shen, Konlin, Flood, Jake J, Zhang, Zhihuizi, Ha, Alvin, Shy, Brian R, Dueber, John E, and Douglas, Shawn M
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Biological Sciences ,Industrial Biotechnology ,Human Genome ,Biotechnology ,Genetics ,DNA ,Single-Stranded ,Escherichia coli ,Genetic Engineering ,Plasmids ,Environmental Sciences ,Information and Computing Sciences ,Developmental Biology ,Biological sciences ,Chemical sciences ,Environmental sciences - Abstract
Long single-stranded DNA (ssDNA) is a versatile molecular reagent with applications including RNA-guided genome engineering and DNA nanotechnology, yet its production is typically resource-intensive. We introduce a novel method utilizing an engineered Escherichia coli 'helper' strain and phagemid system that simplifies long ssDNA generation to a straightforward transformation and purification procedure. Our method obviates the need for helper plasmids and their associated contamination by integrating M13mp18 genes directly into the E. coli chromosome. We achieved ssDNA lengths ranging from 504 to 20 724 nt with titers up to 250 μg/l following alkaline lysis purification. The efficacy of our system was confirmed through its application in primary T-cell genome modifications and DNA origami folding. The reliability, scalability and ease of our approach promise to unlock new experimental applications requiring large quantities of long ssDNA.
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- 2024
15. FSHing for DNA Damage: Key Features of MutY Detection of 8-Oxoguanine:Adenine Mismatches.
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Majumdar, Chandrima, Demir, Merve, Merrill, Steven, Hashemian, Mohammad, and David, Sheila
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Humans ,DNA Repair ,Adenine ,Escherichia coli ,DNA Damage ,DNA ,Colorectal Neoplasms ,Follicle Stimulating Hormone ,Guanine - Abstract
ConspectusBase excision repair (BER) enzymes are genomic superheroes that stealthily and accurately identify and remove chemically modified DNA bases. DNA base modifications erode the informational content of DNA and underlie many disease phenotypes, most conspicuously, cancer. The OG of oxidative base damage, 8-oxo-7,8-dihydroguanine (OG), is particularly insidious due to its miscoding ability that leads to the formation of rare, pro-mutagenic OG:A mismatches. Thwarting mutagenesis relies on the capture of OG:A mismatches prior to DNA replication and removal of the mis-inserted adenine by MutY glycosylases to initiate BER. The threat of OG and the importance of its repair are underscored by the association between inherited dysfunctional variants of the MutY human homologue (MUTYH) and colorectal cancer, known as MUTYH-associated polyposis (MAP). Our functional studies of the two founder MUTYH variants revealed that both have compromised activity and a reduced affinity for OG:A mismatches. Indeed, these studies underscored the challenge of the recognition of OG:A mismatches that are only subtly structurally different than T:A base pairs. Since the original discovery of MAP, many MUTYH variants have been reported, with most considered to be variants of uncertain significance. To reveal features associated with damage recognition and adenine excision by MutY and MUTYH, we have developed a multipronged chemical biology approach combining enzyme kinetics, X-ray crystallography, single-molecule visualization, and cellular repair assays. In this review, we highlight recent work in our laboratory where we defined MutY structure-activity relationship (SAR) studies using synthetic analogs of OG and A in cellular and in vitro assays. Our studies revealed the 2-amino group of OG as the key distinguishing feature of OG:A mismatches. Indeed, the unique position of the 2-amino group in the major groove of OGsyn:Aanti mismatches provides a means for its rapid detection among a large excess of highly abundant and structurally similar canonical base pairs. Furthermore, site-directed mutagenesis and structural analysis showed that a conserved C-terminal domain β-hairpin FSH loop is critical for OG recognition with the His serving as the lesion detector. Notably, MUTYH variants located within and near the FSH loop have been associated with different forms of cancer. Uncovering the role(s) of this loop in lesion recognition provided a detailed understanding of the search and repair process of MutY. Such insights are also useful to identify mutational hotspots and pathogenic variants, which may improve the ability of physicians to diagnose the likelihood of disease onset and prognosis. The critical importance of the FSH loop in lesion detection suggests that it may serve as a unique locus for targeting probes or inhibitors of MutY/MUTYH to provide new chemical biology tools and avenues for therapeutic development.
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- 2024
16. Pathway Evolution Through a Bottlenecking-Debottlenecking Strategy and Machine Learning-Aided Flux Balancing.
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Deng, Huaxiang, Yu, Han, Deng, Yanwu, Qiu, Yulan, Li, Feifei, Wang, Xinran, He, Jiahui, Liang, Weiyue, Lan, Yunquan, Qiao, Longjiang, Zhang, Zhiyu, Zhang, Yunfeng, Keasling, Jay, and Luo, Xiaozhou
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biofoundry ,directed evolution ,machine learning ,pathway debottlenecking ,Escherichia coli ,Flavonoids ,Metabolic Networks and Pathways ,Machine Learning - Abstract
The evolution of pathway enzymes enhances the biosynthesis of high-value chemicals, crucial for pharmaceutical, and agrochemical applications. However, unpredictable evolutionary landscapes of pathway genes often hinder successful evolution. Here, the presence of complex epistasis is identifued within the representative naringenin biosynthetic pathway enzymes, hampering straightforward directed evolution. Subsequently, a biofoundry-assisted strategy is developed for pathway bottlenecking and debottlenecking, enabling the parallel evolution of all pathway enzymes along a predictable evolutionary trajectory in six weeks. This study then utilizes a machine learning model, ProEnsemble, to further balance the pathway by optimizing the transcription of individual genes. The broad applicability of this strategy is demonstrated by constructing an Escherichia coli chassis with evolved and balanced pathway genes, resulting in 3.65 g L-1 naringenin. The optimized naringenin chassis also demonstrates enhanced production of other flavonoids. This approach can be readily adapted for any given number of enzymes in the specific metabolic pathway, paving the way for automated chassis construction in contemporary biofoundries.
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- 2024
17. Protein-Based Rechargeable and Replaceable Antimicrobial and Antifouling Coatings on Hydrophobic Food-Contact Surfaces.
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Zou, Jiahan, Wong, Jody, Lee, Chih-Rong, Nitin, Nitin, Wang, Luxin, and Sun, Gang
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N-halamine ,biofilm ,foodborne pathogen ,fresh produce ,gelatin ,soy protein hydrolysate ,tannic acid ,Biofouling ,Polyethylene ,Anti-Infective Agents ,Povidone ,Escherichia coli ,Polyphenols - Abstract
The growing concerns regarding foodborne illnesses related to fresh produce accentuate the necessity for innovative material solutions, particularly on surfaces that come into close contact with foods. This study introduces a sustainable, efficient, and removable antimicrobial and antifouling coating ideally suited for hydrophobic food-contact surfaces such as low-density polyethylene (LDPE). Developed through a crosslinking reaction involving tannic acid, gelatin, and soy protein hydrolysate, these coatings exhibit proper stability in aqueous washing solutions and effectively combat bacterial contamination and prevent biofilm formation. The unique surface architecture promotes the formation of halamine structures, enhancing antimicrobial efficacy with a rapid contact killing effect and reducing microbial contamination by up to 5 log10 cfu·cm-2 against both Escherichia coli (Gram-negative) and Listeria innocua (Gram-positive). Notably, the coatings are designed for at least five recharging cycles under mild conditions (pH6, 20 ppm free active chlorine) and can be easily removed with hot water or steam to refresh the depositions. This removal process not only conveniently aligns with existing sanitation protocols in the fresh produce industry but also facilitates the complete eradication of potential developed biofilms, outperforming uncoated LDPE coupons. Overall, these coatings represent sustainable, cost-effective, and practical advancements in food safety and are promising candidates for widespread adoption in food processing environments.
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- 2024
18. Advancing the scale of synthetic biology via cross-species transfer of cellular functions enabled by iModulon engraftment.
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Olson, Connor, Szubin, Richard, Yang, Hannah, Sung, Jaemin, Palsson, Bernhard, Feist, Adam, and Choe, Donghui
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Synthetic Biology ,Escherichia coli ,Genes ,Bacterial ,Pseudomonas - Abstract
Machine learning applied to large compendia of transcriptomic data has enabled the decomposition of bacterial transcriptomes to identify independently modulated sets of genes, such iModulons represent specific cellular functions. The identification of iModulons enables accurate identification of genes necessary and sufficient for cross-species transfer of cellular functions. We demonstrate cross-species transfer of: 1) the biotransformation of vanillate to protocatechuate, 2) a malonate catabolic pathway, 3) a catabolic pathway for 2,3-butanediol, and 4) an antimicrobial resistance to ampicillin found in multiple Pseudomonas species to Escherichia coli. iModulon-based engineering is a transformative strategy as it includes all genes comprising the transferred cellular function, including genes without functional annotation. Adaptive laboratory evolution was deployed to optimize the cellular function transferred, revealing mutations in the host. Combining big data analytics and laboratory evolution thus enhances the level of understanding of systems biology, and synthetic biology for strain design and development.
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- 2024
19. Application of phylodynamics to identify spread of antimicrobial-resistant Escherichia coli between humans and canines in an urban environment
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Walas, Nikolina, Müller, Nicola F, Parker, Emily, Henderson, Abigail, Capone, Drew, Brown, Joe, Barker, Troy, and Graham, Jay P
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Medical Microbiology ,Biomedical and Clinical Sciences ,Antimicrobial Resistance ,Prevention ,Genetics ,Vaccine Related ,Emerging Infectious Diseases ,Biodefense ,2.2 Factors relating to the physical environment ,Aetiology ,Infection ,Animals ,Humans ,Dogs ,Escherichia coli ,Escherichia coli Infections ,Bayes Theorem ,Anti-Bacterial Agents ,Anti-Infective Agents ,Drug Resistance ,Bacterial ,Antimicrobial resistance ,Canines ,ESBL ,Environment ,Genomic epidemiology ,Phylodynamics ,Environmental Sciences - Abstract
The transmission of antimicrobial resistant bacteria in the urban environment is poorly understood. We utilized genomic sequencing and phylogenetics to characterize the transmission dynamics of antimicrobial resistant Escherichia coli (AMR-Ec) cultured from putative canine (caninep) and human feces present on urban sidewalks in San Francisco, California. We isolated a total of fifty-six AMR-Ec isolates from human (n = 20) and caninep (n = 36) fecal samples from the Tenderloin and South of Market (SoMa) neighborhoods of San Francisco. We then analyzed phenotypic and genotypic antimicrobial resistance (AMR) of the isolates, as well as clonal relationships based on cgMLST and single nucleotide polymorphisms (SNPs) of the core genomes. Using Bayesian inference, we reconstructed the transmission dynamics between humans and caninesp from multiple local outbreak clusters using the marginal structured coalescent approximation (MASCOT). Our results provide evidence for multiple sharing events of AMR-Ec between humans and caninesp. In particular, we found one instance of likely transmission from caninesp to humans as well as an additional local outbreak cluster consisting of one caninep and one human sample. Based on this analysis, it appears that non-human feces act as an important reservoir of clinically relevant AMR-Ec within the urban environment for this study population. This work showcases the utility of genomic epidemiology to reconstruct potential pathways by which antimicrobial resistance spreads.
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- 2024
20. Occurrence and molecular characteristics of antimicrobial resistance, virulence factors, and extended-spectrum β-lactamase (ESBL) producing Salmonella enterica and Escherichia coli isolated from the retail produce commodities in Bangkok, Thailand
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Saechue, Benjawan, Atwill, Edward R, and Jeamsripong, Saharuetai
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Biological Sciences ,Biomedical and Clinical Sciences ,Microbiology ,Clinical Sciences ,Medical Microbiology ,Antimicrobial Resistance ,Prevention ,Infectious Diseases ,Biodefense ,Foodborne Illness ,Vaccine Related ,Emerging Infectious Diseases ,Digestive Diseases ,2.2 Factors relating to the physical environment ,Aetiology ,Infection ,Antimicrobial resistance ,ESBL ,Escherichia coli ,Produce ,Salmonella enterica - Abstract
The incidence of antimicrobial resistance (AMR) in the environment is often overlooked and leads to serious health threats under the One Health paradigm. Infection with extended-spectrum β-lactamase (ESBL) producing bacteria in humans and animals has been widely examined, with the mode of transmission routes such as food, water, and contact with a contaminated environment. The purpose of this study was to determine the occurrence and molecular characteristics of resistant Salmonella enterica (S. enterica) (n = 59) and Escherichia coli (E. coli) (n = 392) isolated from produce commodities collected from fresh markets and supermarkets in Bangkok, Thailand. In this study, the S. enterica isolates exhibited the highest prevalence of resistance to tetracycline (11.9%) and streptomycin (8.5%), while the E. coli isolates were predominantly resistant to tetracycline (22.5%), ampicillin (21.4%), and sulfamethoxazole (11.5%). Among isolates of S. enterica (6.8%) and E. coli (15.3%) were determined as multidrug resistant (MDR). The prevalence of ESBL-producing isolates was 5.1% and 1.0% in S. enterica and E. coli, respectively. A minority of S. enterica isolates, where a single isolate exclusively carried blaCTX-M-55 (n = 1), and another isolate harbored both blaCTX-M-55 and blaTEM-1 (n = 1); similarly, a minority of E. coli isolates contained blaCTX-M-55 (n = 2) and blaCTX-M-15 (n = 1). QnrS (11.9%) and blaTEM (20.2%) were the most common resistant genes found in S. enterica and E. coli, respectively. Nine isolates resistant to ciprofloxacin contained point mutations in gyrA and parC. In addition, the odds of resistance to tetracycline among isolates of S. enterica were positively associated with the co-occurrence of ampicillin resistance and the presence of tetB (P = 0.001), while the E. coli isolates were positively associated with ampicillin resistance, streptomycin resistance, and the presence of tetA (P
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- 2024
21. Synthesis, insertion, and characterization of SARS-CoV-2 membrane protein within lipid bilayers
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Zhang, Yuanzhong, Anbir, Sara, McTiernan, Joseph, Li, Siyu, Worcester, Michael, Mishra, Pratyasha, Colvin, Michael E, Gopinathan, Ajay, Mohideen, Umar, Zandi, Roya, and Kuhlman, Thomas E
- Subjects
Biochemistry and Cell Biology ,Biological Sciences ,Infectious Diseases ,Biodefense ,Emerging Infectious Diseases ,Prevention ,Vaccine Related ,1.1 Normal biological development and functioning ,Underpinning research ,Generic health relevance ,Humans ,Lipid Bilayers ,SARS-CoV-2 ,Cryoelectron Microscopy ,COVID-19 ,Viral Matrix Proteins ,Membrane Proteins ,Escherichia coli - Abstract
Throughout history, coronaviruses have posed challenges to both public health and the global economy; nevertheless, methods to combat them remain rudimentary, primarily due to the absence of experiments to understand the function of various viral components. Among these, membrane (M) proteins are one of the most elusive because of their small size and challenges with expression. Here, we report the development of an expression system to produce tens to hundreds of milligrams of M protein per liter of Escherichia coli culture. These large yields render many previously inaccessible structural and biophysical experiments feasible. Using cryo-electron microscopy and atomic force microscopy, we image and characterize individual membrane-incorporated M protein dimers and discover membrane thinning in the vicinity, which we validated with molecular dynamics simulations. Our results suggest that the resulting line tension, along with predicted induction of local membrane curvature, could ultimately drive viral assembly and budding.
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- 2024
22. Assessing changes in bacterial load and antibiotic resistance in the Legon sewage treatment plant between 2018 and 2023 in Accra, Ghana
- Author
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Adjei, Raymond Lovelace, Adomako, Lady Asantewah Boamah, Korang-Labi, Appiah, Avornyo, Franklin Kodzo, Timire, Collins, Larbi, Rita Ohene, Kubasari, Cletus, Ackon, Stephen ED, and Reid, Anthony
- Published
- 2023
23. StressME: Unified computing framework of Escherichia coli metabolism, gene expression, and stress responses.
- Author
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Zhao, Jiao, Chen, Ke, Yang, Laurence, and Palsson, Bernhard
- Subjects
Escherichia coli ,Escherichia coli Proteins ,Stress ,Physiological ,Oxidation-Reduction ,Heat-Shock Proteins ,Acids ,Gene Expression - Abstract
Generalist microbes have adapted to a multitude of environmental stresses through their integrated stress response system. Individual stress responses have been quantified by E. coli metabolism and expression (ME) models under thermal, oxidative and acid stress, respectively. However, the systematic quantification of cross-stress & cross-talk among these stress responses remains lacking. Here, we present StressME: the unified stress response model of E. coli combining thermal (FoldME), oxidative (OxidizeME) and acid (AcidifyME) stress responses. StressME is the most up to date ME model for E. coli and it reproduces all published single-stress ME models. Additionally, it includes refined rate constants to improve prediction accuracy for wild-type and stress-evolved strains. StressME revealed certain optimal proteome allocation strategies associated with cross-stress and cross-talk responses. These stress-optimal proteomes were shaped by trade-offs between protective vs. metabolic enzymes; cytoplasmic vs. periplasmic chaperones; and expression of stress-specific proteins. As StressME is tuned to compute metabolic and gene expression responses under mild acid, oxidative, and thermal stresses, it is useful for engineering and health applications. The modular design of our open-source package also facilitates model expansion (e.g., to new stress mechanisms) by the computational biology community.
- Published
- 2024
24. High fat intake sustains sorbitol intolerance after antibiotic-mediated Clostridia depletion from the gut microbiota
- Author
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Lee, Jee-Yon, Tiffany, Connor R, Mahan, Scott P, Kellom, Matthew, Rogers, Andrew WL, Nguyen, Henry, Stevens, Eric T, Masson, Hugo LP, Yamazaki, Kohei, Marco, Maria L, Eloe-Fadrosh, Emiley A, Turnbaugh, Peter J, and Bäumler, Andreas J
- Subjects
Microbiology ,Biological Sciences ,Biomedical and Clinical Sciences ,Prevention ,Complementary and Integrative Health ,Microbiome ,Nutrition ,Digestive Diseases ,Animals ,Mice ,Anti-Bacterial Agents ,Butyrates ,Carbohydrate Metabolism ,Inborn Errors ,Clostridium ,Escherichia coli ,Gastrointestinal Microbiome ,Sorbitol ,Clostridia ,antibiotics ,carbohydrate intolerance ,gut microbiota ,high-fat diet ,polyol ,sorbitol intolerance ,Medical and Health Sciences ,Developmental Biology ,Biological sciences ,Biomedical and clinical sciences - Abstract
Carbohydrate intolerance, commonly linked to the consumption of lactose, fructose, or sorbitol, affects up to 30% of the population in high-income countries. Although sorbitol intolerance is attributed to malabsorption, the underlying mechanism remains unresolved. Here, we show that a history of antibiotic exposure combined with high fat intake triggered long-lasting sorbitol intolerance in mice by reducing Clostridia abundance, which impaired microbial sorbitol catabolism. The restoration of sorbitol catabolism by inoculation with probiotic Escherichia coli protected mice against sorbitol intolerance but did not restore Clostridia abundance. Inoculation with the butyrate producer Anaerostipes caccae restored a normal Clostridia abundance, which protected mice against sorbitol-induced diarrhea even when the probiotic was cleared. Butyrate restored Clostridia abundance by stimulating epithelial peroxisome proliferator-activated receptor-gamma (PPAR-γ) signaling to restore epithelial hypoxia in the colon. Collectively, these mechanistic insights identify microbial sorbitol catabolism as a potential target for approaches for the diagnosis, treatment, and prevention of sorbitol intolerance.
- Published
- 2024
25. Characterization and Diversification of AraC/XylS Family Regulators Guided by Transposon Sequencing
- Author
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Pearson, Allison N, Incha, Matthew R, Ho, Cindy N, Schmidt, Matthias, Roberts, Jacob B, Nava, Alberto A, and Keasling, Jay D
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Biochemistry and Cell Biology ,Bioinformatics and Computational Biology ,Biological Sciences ,Genetics ,Escherichia coli ,Bacterial Proteins ,Promoter Regions ,Genetic ,Pseudomonas putida ,Plasmids ,Gene Expression Regulation ,Bacterial ,Escherichia coli Proteins ,AraC Transcription Factor ,Medicinal and Biomolecular Chemistry ,Biomedical Engineering ,Biochemistry and cell biology ,Bioinformatics and computational biology - Abstract
In this study, we explored the development of engineered inducible systems. Publicly available data from previous transposon sequencing assays were used to identify regulators of metabolism in Pseudomonas putida KT2440. For AraC family regulators (AFRs) represented in these data, we posited AFR/promoter/inducer groupings. Twelve promoters were characterized for a response to their proposed inducers in P. putida, and the resultant data were used to create and test nine two-plasmid sensor systems in Escherichia coli. Several of these were further developed into a palette of single-plasmid inducible systems. From these experiments, we observed an unreported inducer response from a previously characterized AFR, demonstrated that the addition of a P. putida transporter improved the sensor dynamics of an AFR in E. coli, and identified an uncharacterized AFR with a novel potential inducer specificity. Finally, targeted mutations in an AFR, informed by structural predictions, enabled the further diversification of these inducible plasmids.
- Published
- 2024
26. Association of metabolic dysfunction-associated fatty liver disease with gastrointestinal infections: insights from National Inpatient Sample Database.
- Author
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Patel, Jay, Sohal, Aalam, Bains, Kanwal, Chaudhry, Hunza, Kohli, Isha, Khanna, Tejasvini, Dukovic, Dino, and Roytman, Marina
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DIETARY - GASTROINTESTINAL INFECTIONS ,INFECTIOUS DISEASE ,INTESTINAL BACTERIA ,LIVER ,NONALCOHOLIC STEATOHEPATITIS ,Adult ,Humans ,Adolescent ,Inpatients ,Clostridioides difficile ,Escherichia coli ,Retrospective Studies ,Gastrointestinal Diseases ,Non-alcoholic Fatty Liver Disease - Abstract
OBJECTIVES: The study aimed to compare the risk of gastrointestinal infections among patients with and without metabolic dysfunction-associated fatty liver disease (MAFLD). METHODS: This was a population-based, retrospective, observational study using data from the National Inpatient Sample (NIS), the largest all-payer US inpatient care database. SETTING: Hospitalisation of adults aged ≥18 years old admitted in 2020 was identified using the NIS. Patients were stratified by the presence and absence of MAFLD. PARTICIPANTS: 26.4 million adults aged ≥18 years old were included in the study. Patients younger than 18 and those with missing demographic or mortality data were excluded. PRIMARY AND SECONDARY OUTCOMES: Primary outcome was to assess the overall risk of gastrointestinal infections in patients with and without MAFLD. Secondary outcomes were demographics and comorbidities stratified by the presence or absence of gastrointestinal infection, and the risk of specific gastrointestinal pathogens. RESULTS: Of 26.4 million patients admitted in 2020, 755 910 (2.85%) had the presence of MAFLD. There was a higher prevalence of bacterial gastrointestinal infections in patients with MAFLD than those without (1.6% vs 0.9%, p
- Published
- 2024
27. Draft genome sequences of five multidrug-resistant Escherichia coli strains isolated from vegetable samples in Bangladesh.
- Author
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Pramanik, Pritom, Rana, Md, Ramasamy, Srinivasan, Schreinemachers, Pepijn, Oliva, Ricardo, Rahman, Md, and Islam, Md Saiful
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Bangladesh ,Escherichia coli ,MDR ,antibiotic resistance genes ,gardening systems ,public health ,vegetables ,virulence factor genes ,whole-genome sequencing - Abstract
Reports indicate that vegetables are becoming a source of multidrug-resistant (MDR) bacteria, including Escherichia coli. Here, we present genome sequences of five MDR E. coli strains to assist future genomic analysis of this bacterium. These E. coli strains were isolated from vegetable samples of different gardening systems in Dhaka, Bangladesh.
- Published
- 2024
28. Selection pressures on evolution of ribonuclease H explored with rigorous free-energy-based design.
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Hayes, Ryan, Nixon, Charlotte, Marqusee, Susan, and Brooks, Charles
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consensus sequence ,free energy ,protein folding ,ribonuclease H ,selection pressure ,Escherichia coli ,Phylogeny ,Computer Simulation ,Consensus Sequence ,Ribonuclease H - Abstract
Understanding natural protein evolution and designing novel proteins are motivating interest in development of high-throughput methods to explore large sequence spaces. In this work, we demonstrate the application of multisite λ dynamics (MSλD), a rigorous free energy simulation method, and chemical denaturation experiments to quantify evolutionary selection pressure from sequence-stability relationships and to address questions of design. This study examines a mesophilic phylogenetic clade of ribonuclease H (RNase H), furthering its extensive characterization in earlier studies, focusing on E. coli RNase H (ecRNH) and a more stable consensus sequence (AncCcons) differing at 15 positions. The stabilities of 32,768 chimeras between these two sequences were computed using the MSλD framework. The most stable and least stable chimeras were predicted and tested along with several other sequences, revealing a designed chimera with approximately the same stability increase as AncCcons, but requiring only half the mutations. Comparing the computed stabilities with experiment for 12 sequences reveals a Pearson correlation of 0.86 and root mean squared error of 1.18 kcal/mol, an unprecedented level of accuracy well beyond less rigorous computational design methods. We then quantified selection pressure using a simple evolutionary model in which sequences are selected according to the Boltzmann factor of their stability. Selection temperatures from 110 to 168 K are estimated in three ways by comparing experimental and computational results to evolutionary models. These estimates indicate selection pressure is high, which has implications for evolutionary dynamics and for the accuracy required for design, and suggests accurate high-throughput computational methods like MSλD may enable more effective protein design.
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- 2024
29. Mg2+ supplementation treats secretory diarrhea in mice by activating calcium-sensing receptor in intestinal epithelial cells.
- Author
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De Souza Goncalves, Livia, Chu, Tifany, Master, Riya, Chhetri, Parth, Gao, Qi, and Cil, Onur
- Subjects
Calcium ,Cell Biology ,Chloride channels ,Epithelial transport of ions and water ,Mice ,Humans ,Animals ,Receptors ,Calcium-Sensing ,Magnesium ,Cholera Toxin ,Cholera ,Calcium ,Escherichia coli ,Colforsin ,Intestinal Mucosa ,Diarrhea ,Epithelial Cells ,Dietary Supplements - Abstract
Cholera is a global health problem with no targeted therapies. The Ca2+-sensing receptor (CaSR) is a regulator of intestinal ion transport and a therapeutic target for diarrhea, and Ca2+ is considered its main agonist. We found that increasing extracellular Ca2+ had a minimal effect on forskolin-induced Cl- secretion in human intestinal epithelial T84 cells. However, extracellular Mg2+, an often-neglected CaSR agonist, suppressed forskolin-induced Cl- secretion in T84 cells by 65% at physiological levels seen in stool (10 mM). The effect of Mg2+ occurred via the CaSR/Gq signaling that led to cAMP hydrolysis. Mg2+ (10 mM) also suppressed Cl- secretion induced by cholera toxin, heat-stable E. coli enterotoxin, and vasoactive intestinal peptide by 50%. In mouse intestinal closed loops, luminal Mg2+ treatment (20 mM) inhibited cholera toxin-induced fluid accumulation by 40%. In a mouse intestinal perfusion model of cholera, addition of 10 mM Mg2+ to the perfusate reversed net fluid transport from secretion to absorption. These results suggest that Mg2+ is the key CaSR activator in mouse and human intestinal epithelia at physiological levels in stool. Since stool Mg2+ concentrations in patients with cholera are essentially zero, oral Mg2+ supplementation, alone or in an oral rehydration solution, could be a potential therapy for cholera and other cyclic nucleotide-mediated secretory diarrheas.
- Published
- 2024
30. IS26 drives the dissemination of bla CTX-M genes in an Ecuadorian community.
- Author
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Salinas, Liseth, Cárdenas, Paúl, Trueba, Gabriel, and Graham, Jay
- Subjects
Escherichia coli ,IS26 ,antimicrobial resistance ,bla CTX-M ,horizontal gene transfer - Abstract
The horizontal gene transfer events are the major contributors to the current spread of CTX-M-encoding genes, the most common extended-spectrum β-lactamase (ESBL), and many clinically crucial antimicrobial resistance (AMR) genes. This study presents evidence of the critical role of IS26 transposable element for the mobility of bla CTX-M gene among Escherichia coli isolates from children and domestic animals in the community. We suggest that the nucleotide sequences of IS26-bla CTX-M could be used to study bla CTX-M transmission between humans, domestic animals, and the environment, because understanding of the dissemination patterns of AMR genes is critical to implement effective measures to slow down the dissemination of these clinically important genes.
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- 2024
31. Chorioamnionitis accelerates granule cell and oligodendrocyte maturation in the cerebellum of preterm nonhuman primates.
- Author
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Newman, Josef, Tong, Xiaoying, Tan, April, Yeasky, Toni, De Paiva, Vanessa, Presicce, Pietro, Kannan, Paranthaman, Williams, Kevin, Damianos, Andreas, Tamase Newsam, Marione, Benny, Merline, Wu, Shu, Young, Karen, Miller, Lisa, Kallapur, Suhas, Chougnet, Claire, Jobe, Alan, Brambilla, Roberta, and Schmidt, Augusto
- Subjects
Cerebellum ,Chorioamnionitis ,Granule cell ,Maturation ,Oligodendrocyte ,Purkinje cell ,Infant ,Newborn ,Female ,Infant ,Animals ,Humans ,Pregnancy ,Hedgehog Proteins ,Macaca mulatta ,Chorioamnionitis ,Premature Birth ,Escherichia coli ,Infant ,Premature ,Cerebellum ,RNA ,Small Nuclear - Abstract
BACKGROUND: Preterm birth is often associated with chorioamnionitis and leads to increased risk of neurodevelopmental disorders, such as autism. Preterm birth can lead to cerebellar underdevelopment, but the mechanisms of disrupted cerebellar development in preterm infants are not well understood. The cerebellum is consistently affected in people with autism spectrum disorders, showing reduction of Purkinje cells, decreased cerebellar grey matter, and altered connectivity. METHODS: Preterm rhesus macaque fetuses were exposed to intra-amniotic LPS (1 mg, E. coli O55:B5) at 127 days (80%) gestation and delivered by c-section 5 days after injections. Maternal and fetal plasma were sampled for cytokine measurements. Chorio-decidua was analyzed for immune cell populations by flow cytometry. Fetal cerebellum was sampled for histology and molecular analysis by single-nuclei RNA-sequencing (snRNA-seq) on a 10× chromium platform. snRNA-seq data were analyzed for differences in cell populations, cell-type specific gene expression, and inferred cellular communications. RESULTS: We leveraged snRNA-seq of the cerebellum in a clinically relevant rhesus macaque model of chorioamnionitis and preterm birth, to show that chorioamnionitis leads to Purkinje cell loss and disrupted maturation of granule cells and oligodendrocytes in the fetal cerebellum at late gestation. Purkinje cell loss is accompanied by decreased sonic hedgehog signaling from Purkinje cells to granule cells, which show an accelerated maturation, and to oligodendrocytes, which show accelerated maturation from pre-oligodendrocytes into myelinating oligodendrocytes. CONCLUSION: These findings suggest a role of chorioamnionitis on disrupted cerebellar maturation associated with preterm birth and on the pathogenesis of neurodevelopmental disorders among preterm infants.
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- 2024
32. Effects of breastfeeding on childrens gut colonization with multidrug-resistant Enterobacterales in peri-urban Lima, Peru.
- Author
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Nadimpalli, Maya, Rojas Salvatierra, Luismarcelo, Chakraborty, Subhra, Swarthout, Jenna, Cabrera, Lilia, Calderon, Maritza, Saito, Mayuko, Gilman, Robert, Pajuelo, Monica, and Pickering, Amy
- Subjects
ESBL ,Peru ,antibiotic resistance ,breastfeeding ,human milk ,infant health ,urban informal settlement ,Child ,Female ,Humans ,Adolescent ,Infant ,Infant ,Newborn ,Breast Feeding ,Prospective Studies ,Peru ,Gastrointestinal Microbiome ,Escherichia coli ,Anti-Bacterial Agents - Abstract
Children living in low-resource settings are frequently gut-colonized with multidrug-resistant bacteria. We explored whether breastfeeding may protect against childrens incident gut colonization with extended-spectrum beta-lactamase-producing Escherichia coli (ESBL-Ec) and Klebsiella, Enterobacter, or Citrobacter spp. (ESBL-KEC). We screened 937 monthly stool samples collected from 112 children aged 1-16 months during a 2016-19 prospective cohort study of enteric infections in peri-urban Lima. We used 52,816 daily surveys to examine how exposures to breastfeeding in the 30 days prior to a stool sample were associated with childrens risks of incident gut-colonization, controlling for antibiotic use and other covariates. We sequenced 78 ESBL-Ec from 47 children to explore their diversity. Gut-colonization with ESBL-Ec was increasingly prevalent as children aged, approaching 75% by 16 months, while ESBL-KEC prevalence fluctuated between 18% and 36%. Through 6 months of age, exclusively providing human milk in the 30 days prior to a stool sample did not reduce childrens risk of incident gut-colonization with ESBL-Ec or ESBL-KEC. From 6 to 16 months of age, every 3 additional days of breastfeeding in the prior 30 days was associated with 6% lower risk of incident ESBL-Ec gut-colonization (95% CI: 0.90, 0.98, p = .003). No effects were observed on incident ESBL-KEC colonization. We detected highly diverse ESBL-Ec among children and few differences between children who were predominantly breastfed (mean age: 4.1 months) versus older children (10.8 months). Continued breastfeeding after 6 months conferred protection against childrens incident gut colonization with ESBL-Ec in this setting. Policies supporting continued breastfeeding should be considered in efforts to combat antibiotic resistance.
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- 2024
33. A review of the mechanisms that confer antibiotic resistance in pathotypes of E. coli.
- Author
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Nasrollahian, Sina, Graham, Jay, and Halaji, Mehrdad
- Subjects
ESBL ,Escherichia coli ,UPEC ,antibiotic resistance ,efflux pump ,Humans ,Escherichia coli ,Escherichia coli Infections ,Microbial Sensitivity Tests ,Anti-Bacterial Agents ,beta-Lactamases ,Drug Resistance ,Microbial - Abstract
The dissemination of antibiotic resistance in Escherichia coli poses a significant threat to public health worldwide. This review provides a comprehensive update on the diverse mechanisms employed by E. coli in developing resistance to antibiotics. We primarily focus on pathotypes of E. coli (e.g., uropathogenic E. coli) and investigate the genetic determinants and molecular pathways that confer resistance, shedding light on both well-characterized and recently discovered mechanisms. The most prevalent mechanism continues to be the acquisition of resistance genes through horizontal gene transfer, facilitated by mobile genetic elements such as plasmids and transposons. We discuss the role of extended-spectrum β-lactamases (ESBLs) and carbapenemases in conferring resistance to β-lactam antibiotics, which remain vital in clinical practice. The review covers the key resistant mechanisms, including: 1) Efflux pumps and porin mutations that mediate resistance to a broad spectrum of antibiotics, including fluoroquinolones and aminoglycosides; 2) adaptive strategies employed by E. coli, including biofilm formation, persister cell formation, and the activation of stress response systems, to withstand antibiotic pressure; and 3) the role of regulatory systems in coordinating resistance mechanisms, providing insights into potential targets for therapeutic interventions. Understanding the intricate network of antibiotic resistance mechanisms in E. coli is crucial for the development of effective strategies to combat this growing public health crisis. By clarifying these mechanisms, we aim to pave the way for the design of innovative therapeutic approaches and the implementation of prudent antibiotic stewardship practices to preserve the efficacy of current antibiotics and ensure a sustainable future for healthcare.
- Published
- 2024
34. Comprehensive Characterization of fucAO Operon Activation in Escherichia coli
- Author
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Zhang, Zhongge, Huo, Jialu, Velo, Juan, Zhou, Harry, Flaherty, Alex, and Saier, Milton H
- Subjects
Biochemistry and Cell Biology ,Biological Sciences ,Genetics ,fucAO operon ,operon promoter ,Crp ,FucR ,SrsR ,transcriptional activation ,transcriptionally silent region ,protein-protein interactions ,Escherichia coli ,Fucose ,Binding Sites ,Phosphorylation ,Operon ,fucAO operon ,Other Chemical Sciences ,Other Biological Sciences ,Chemical Physics ,Biochemistry and cell biology ,Microbiology ,Medicinal and biomolecular chemistry - Abstract
Wildtype Escherichia coli cells cannot grow on L-1,2-propanediol, as the fucAO operon within the fucose (fuc) regulon is thought to be silent in the absence of L-fucose. Little information is available concerning the transcriptional regulation of this operon. Here, we first confirm that fucAO operon expression is highly inducible by fucose and is primarily attributable to the upstream operon promoter, while the fucO promoter within the 3'-end of fucA is weak and uninducible. Using 5'RACE, we identify the actual transcriptional start site (TSS) of the main fucAO operon promoter, refuting the originally proposed TSS. Several lines of evidence are provided showing that the fucAO locus is within a transcriptionally repressed region on the chromosome. Operon activation is dependent on FucR and Crp but not SrsR. Two Crp-cAMP binding sites previously found in the regulatory region are validated, where the upstream site plays a more critical role than the downstream site in operon activation. Furthermore, two FucR binding sites are identified, where the downstream site near the first Crp site is more important than the upstream site. Operon transcription relies on Crp-cAMP to a greater degree than on FucR. Our data strongly suggest that FucR mainly functions to facilitate the binding of Crp to its upstream site, which in turn activates the fucAO promoter by efficiently recruiting RNA polymerase.
- Published
- 2024
35. Impact of zinc supplementation on phenotypic antimicrobial resistance of fecal commensal bacteria from pre-weaned dairy calves
- Author
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Lee, Katie Y, Atwill, Edward R, Li, Xunde, Feldmann, Hillary R, Williams, Deniece R, Weimer, Bart C, and Aly, Sharif S
- Subjects
Microbiology ,Biological Sciences ,Biomedical and Clinical Sciences ,Complementary and Integrative Health ,Clinical Research ,Antimicrobial Resistance ,Nutrition ,Clinical Trials and Supportive Activities ,Infection ,Animals ,Cattle ,Anti-Bacterial Agents ,Zinc ,Escherichia coli ,Drug Resistance ,Bacterial ,Anti-Infective Agents ,Enterococcus ,Diarrhea ,Organic Chemicals ,Dietary Supplements ,Ciprofloxacin - Abstract
The objective of this study was to evaluate the impact of dietary zinc supplementation in pre-weaned dairy calves on the phenotypic antimicrobial resistance (AMR) of fecal commensal bacteria. A repository of fecal specimens from a random sample of calves block-randomized into placebo (n = 39) and zinc sulfate (n = 28) groups collected over a zinc supplementation clinical trial at the onset of calf diarrhea, calf diarrheal cure, and the last day of 14 cumulative days of zinc or placebo treatment were analyzed. Antimicrobial susceptibility testing was conducted for Enterococcus spp. (n = 167) and E. coli (n = 44), with one representative isolate of each commensal bacteria tested per sample. Parametric survival interval regression models were constructed to evaluate the association between zinc treatment and phenotypic AMR, with exponentiated accelerated failure time (AFT) coefficients adapted for MIC instead of time representing the degree of change in AMR (MIC Ratio, MR). Findings from our study indicated that zinc supplementation did not significantly alter the MIC in Enterococcus spp. for 13 drugs: gentamicin, vancomycin, ciprofloxacin, erythromycin, penicillin, nitrofurantoin, linezolid, quinupristin/dalfopristin, tylosin tartrate, streptomycin, daptomycin, chloramphenicol, and tigecycline (MR = 0.96-2.94, p > 0.05). In E. coli, zinc supplementation was not associated with resistance to azithromycin (MR = 0.80, p > 0.05) and ceftriaxone (MR = 0.95, p > 0.05). However, a significant reduction in E. coli MIC values was observed for ciprofloxacin (MR = 0.17, 95% CI 0.03-0.97) and nalidixic acid (MR = 0.28, 95% CI 0.15-0.53) for zinc-treated compared to placebo-treated calves. Alongside predictions of MIC values generated from these 17 AFT models, findings from this study corroborate the influence of age and antimicrobial exposure on phenotypic AMR.
- Published
- 2024
36. Antimicrobial resistance in food-associated Escherichia coli in Mexico and Latin America.
- Author
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Babines-Orozco, Lorena, Balbuena-Alonso, María, Barrios-Villa, Edwin, Lozano-Zarain, Patricia, Martínez-Laguna, Ygnacio, Del Carmen Rocha-Gracia, Rosa, and Cortés-Cortés, Gerardo
- Subjects
Escherichia coli ,Latin America ,Mexico ,antimicrobial resistance ,food - Abstract
The World Health Organization (WHO) considers antimicrobial resistance to be one of the critical global public health priorities to address. Escherichia coli is a commensal bacterium of the gut microbiota in humans and animals; however, some strains cause infections and are resistant to antibiotics. One of the most common ways of acquiring pathogenic E. coli strains is through food. This review analyzes multidrug-resistant E. coli isolated from food, emphasizing Latin America and Mexico, and the mobile genetic elements (MGEs) responsible for spreading antibiotic resistance determinants among bacteria in different environments and hosts. We conducted a systematic search of the literature published from 2015 to 2022 in open access databases and electronic repositories. The prevalence of 11 E. coli pathotypes was described, with diarrheagenic E. coli pathotypes being the most frequently associated with foodborne illness in different Latin American countries, highlighting the presence of different antibiotic resistance genes mostly carried by IncF-type plasmids or class 1 integrons. Although the global incidence of foodborne illness is high, there have been few studies in Mexico and Latin America, which highlights the need to generate updated epidemiological data from the One Health approach, which allows monitoring of the multidrug-resistance phenomenon in E. coli from a common perspective in the interaction of human, veterinary, and environmental health.
- Published
- 2024
37. Statistical 3D morphology characterization of vaterite microspheres produced by engineered Escherichia coli
- Author
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Lin, Alex YW, Wu, Zong-Yen, Pattison, Alexander J, Müller, Isaak E, Yoshikuni, Yasuo, Theis, Wolfgang, and Ercius, Peter
- Subjects
Biological Sciences ,Engineering ,Physical Sciences ,Biomedical Engineering ,Regenerative Medicine ,Biotechnology ,Bioengineering ,Generic health relevance ,Calcium Carbonate ,Microscopy ,Electron ,Scanning ,Microspheres ,Escherichia coli ,Microscopy ,Electron ,Scanning Transmission ,Biocompatible Materials ,3D morphology ,Biomineralization ,MICP ,Scanning transmission electron microscopy ,Vaterite - Abstract
Hollow vaterite microspheres are important materials for biomedical applications such as drug delivery and regenerative medicine owing to their biocompatibility, high specific surface area, and ability to encapsulate a large number of bioactive molecules and compounds. We demonstrated that hollow vaterite microspheres are produced by an Escherichia coli strain engineered with a urease gene cluster from the ureolytic bacteria Sporosarcina pasteurii in the presence of bovine serum albumin. We characterized the 3D nanoscale morphology of five biogenic hollow vaterite microspheres using 3D high-angle annular dark field scanning transmission electron microscopy (HAADF-STEM) tomography. Using automated high-throughput HAADF-STEM imaging across several sample tilt orientations, we show that the microspheres evolved from a smaller more ellipsoidal shape to a larger more spherical shape while the internal hollow core increased in size and remained relatively spherical, indicating that the microspheres produced by this engineered strain likely do not contain the bacteria. The statistical 3D morphology information demonstrates the potential for using biogenic calcium carbonate mineralization to produce hollow vaterite microspheres with controlled morphologies. STATEMENT OF SIGNIFICANCE: The nanoscale 3D structures of biomaterials determine their physical, chemical, and biological properties, however significant efforts are required to obtain a statistical understanding of the internal 3D morphology of materials without damaging the structures. In this study, we developed a non-destructive, automated technique that allows us to understand the nanoscale 3D morphology of many unique hollow vaterite microspheres beyond the spectroscopy methods that lack local information and microscopy methods that cannot interrogate the full 3D structure. The method allowed us to quantitatively correlate the external diameters and aspect ratios of vaterite microspheres with their hollow internal structures at the nanoscale. This work demonstrates the opportunity to use automated transmission electron microscopy to characterize nanoscale 3D morphologies of many biomaterials and validate the chemical and biological functionality of these materials.
- Published
- 2024
38. Antimicrobial resistance, virulence profile, and genetic analysis of ESBL-producing Escherichia coli isolated from Nile tilapia in fresh markets and supermarkets in Thailand.
- Author
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Hinthong, Woranich, Thaotumpitak, Varangkana, Sripradite, Jarukorn, Indrawattana, Nitaya, Srisook, Thassanee, Kongngoen, Thida, R Atwill, Edward, and Jeamsripong, Saharuetai
- Subjects
Animals ,Escherichia coli ,Anti-Bacterial Agents ,Escherichia coli Infections ,Virulence ,Cichlids ,Thailand ,Supermarkets ,beta-Lactamases ,Drug Resistance ,Bacterial ,Ampicillin ,Tetracycline - Abstract
This study investigated the prevalence and antimicrobial resistance (AMR) of Escherichia coli (E. coli) in Nile tilapia from fresh markets and supermarkets. A total of samples (n = 828) were collected from Nile tilapia including fish flesh (n = 276), liver and kidney (n = 276), and intestine (n = 276). Overall prevalence of fecal coliforms (61.6%) and E. coli (53.0%) were observed. High prevalence of E. coli was found in the intestine (71.4%), followed by the liver and kidney (45.7%). The highest prevalence of resistance was commonly found against tetracycline (78.5%), ampicillin (72.8%), and sulfamethoxazole (45.6%) with resistance to only tetracycline (15.2%) as the most common antibiogram. The prevalence of multidrug resistance (MDR) (54.4%) and Extended-spectrum beta-lactamases (ESBLs) (5.7%) were examined. The predominant virulence genes (n = 158) were st (14.6%), followed by eaeA (0.6%). The blaTEM (73.4%), tetA (65.2%), and qnrS (57.6%). There is statistical significance between Nile tilapia from fresh markets and supermarkets. Based on logistic regression analysis, ampicillin-resistant E. coli was statistically associated with the phenotypic resistance to tetracycline and trimethoprim, and the presence of blaTEM and tetA (p < 0.05). Further investigation of AMR transference and their mechanisms is needed for AMR control.
- Published
- 2024
39. Synthesis and antibacterial activity of phenolipid methyl dihydroxystearate-gallic acid.
- Author
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Firdaus, Radhinal Zikri, Handayani, Sri, Wukirsari, Tuti, and Hudiyono, Sumi
- Subjects
- *
ANTIBACTERIAL agents , *ESCHERICHIA coli , *GALLATES , *ACID derivatives , *SULFURIC acid , *FORMIC acid - Abstract
Diseases caused by bacteria are still a serious health problem in the world. In many studies, oleic acid derivatives have bioactivity such as antibacterial activity. In this research, oleic acid was first esterified to form methyl oleate using methanol in the presence of sulphuric acid as the catalyst at 60°C for 4 hours. Further modification of methyl oleate structure was conducted by oxidation of C=C using a mixture of H2O2 and formic acid at 40 °C for 24 hours. The hydroxyl group from the oxidation reaction was converted into an ester by using gallic acid. Steglich esterification was employed and the molar ratio of diol and gallic acid was 1:2. The ester was characterized using FTIR and its antibacterial activity against Escherichia coli and Staphylococcus aureus was examined. FTIR spectrum showed the presence of ester and aromatic ring at 1750 cm−1 to 1600 cm−1. The antibacterial assay showed that the inhibitory zone of ester was 7 mm against E. coli and 6.8 mm against S. aureus which showed a slight improvement in antibacterial activity. [ABSTRACT FROM AUTHOR]
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- 2024
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40. Novel chalcone synthesis from phloroglucinol and its activity against multidrug resistance strain of S. aureus and E. coli.
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Sepvianti, Wiwit, Rahman, Aulia, Kusumaningrum, Serafica Btari Christiyani, and Sari, Arum
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ESCHERICHIA coli , *MULTIDRUG resistance , *CHALCONE , *PHLOROGLUCINOL , *PATHOGENIC bacteria , *DRUG resistance in bacteria - Abstract
Antibiotic resistance was still a problem in curing infectious diseases, especially infections by pathogenic bacteria. Various studies to improve the quality of antibiotics by minimizing the minimum inhibitory concentration (MIC) value was still developed. One of the natural compounds that has antibiotic ability and has been widely developed was chalcone. This study aims to evaluate the combination effects of chalcone derivatives compounds that The compunds that have beeb synthesis were (E)-1-(2-hydroxy-4,6dimethoxyphenyl)-3-phenylprop-2-en-1-one(3a); (E)-1-(2-hydroxy-4,6-dimethoxyphenyl)-3-(4methoxyphenyl) prop-2-en-2-one(3b); and (E)-3-(3,4-dimethoxyphenyl)-1-(2-hydroxy-4,6-dimethoxyphenyl) prop-2en-1-one (3c) with antibiotics (ciprofloxacin, ampicillin, and doxycycline) by reducing the MIC value in Staphylococcus aureus and Escherichia coli. The results of this study indicate that the combination of chalcone derivatives and antibiotics could significantly reduce the MIC value. [ABSTRACT FROM AUTHOR]
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- 2024
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41. Synthesis and antibacterial activity test of aminoalkylated eugenol compounds in vitro and in silico.
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Syahri, Jufrizal, Nurlaili, Rahim, Anisa Aulia, Dhony, Roma, Zulya, Shinta Okka, and Wahyuningsih, Sri
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- *
ANTIBACTERIAL agents , *EUGENOL , *MORPHOLINE , *BACTERIAL proteins , *ESCHERICHIA coli , *MOLECULES - Abstract
Antimicrobial resistance is one of the world's greatest health challenges. In 2015, WHO reported that at least 700,000 people died from antimicrobial resistance. One of the natural compounds found in Indonesia and reported to have antimicrobial activity is eugenol. However, the activity of the resulting compound is still very weak, so it is necessary to modify the functional group to increase its antimicrobial activity by adding an amine group. In this study, the synthesis of Aminoalkylated eugenol compounds with the addition of morpholine (compound 1) and dimethylamine (compound 2) groups was carried out based on In Vitro and In Silico studies. In Silico method using Discovery Studio software on sensitive Staphylococcus aureus bacteria with protein code 3VOB.pdb. and resistant Staphylococcus aureus bacteria with protein code 2X3F.pdb. In addition, antibacterial activity was also tested against sensitive Eschericia coli bacteria with the protein code 2JOW.pdb. and resistant Eschericia coli bacteria with protein code 7C7O.pdb. To understand the interactions formed by the structure of the compounds confirmed by spectrophotometric analysis using MS, 1H-NMR and 13C-NMR. Eugenol without the addition of an amine group in S. aureus showed a zone of 12.2 mm and the addition of morpholine and dimethylamine significantly increased the antibacterial activity to 15.2 mm and 18.6 mm, respectively, while eugenol without the addition of an amine group in E. coli bacteria. showed a zone of 11.5 mm and the addition of morpholine and dimethylamine could significantly increase the antibacterial activity to 14.3 mm and 20 mm, respectively. The molecular docking of these compounds showed strong hydrogen bond interactions with SER186, ARG188, ASP151, HIS35, LYS150, LYS185, PHE147, with CDOCKER interaction energy of-48.84 kcal/mol. [ABSTRACT FROM AUTHOR]
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- 2024
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42. The crosstalk between microbial sensors ELMO1 and NOD2 shape intestinal immune responses
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Sharma, Aditi, Achi, Sajan Chandrangadhan, Ibeawuchi, Stella-Rita, Anandachar, Mahitha Shree, Gementera, Hobie, Chaudhury, Uddeep, Usmani, Fatima, Vega, Kevin, Sayed, Ibrahim M, and Das, Soumita
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Microbiology ,Biochemistry and Cell Biology ,Biomedical and Clinical Sciences ,Biological Sciences ,Autoimmune Disease ,Digestive Diseases ,Inflammatory Bowel Disease ,Crohn's Disease ,1.1 Normal biological development and functioning ,2.1 Biological and endogenous factors ,Aetiology ,Underpinning research ,Inflammatory and immune system ,Infection ,Oral and gastrointestinal ,Animals ,Mice ,Adaptor Proteins ,Signal Transducing ,Crohn Disease ,Escherichia coli ,Immunity ,Intestines ,Macrophages ,Microbial sensors ,NOD2 ,bacterial engulfment ,AIEC-LF82 ,epithelial cells and macrophages ,3D-organoid ,ELMO-1 ,AIEC-LF82 ,epithelial cells and macrophages ,Ecological Applications ,Medical Microbiology ,Medical microbiology - Abstract
Microbial sensors play an essential role in maintaining cellular homoeostasis. Our knowledge is limited on how microbial sensing helps in differential immune response and its link to inflammatory diseases. Recently we have confirmed that ELMO1 (Engulfment and Cell Motility Protein-1) present in cytosol is involved in pathogen sensing, engulfment, and intestinal inflammation. Here, we show that ELMO1 interacts with another sensor, NOD2 (Nucleotide-binding oligomerization domain-containing protein 2), that recognizes bacterial cell wall component muramyl dipeptide (MDP). The polymorphism of NOD2 is linked to Crohn's disease (CD) pathogenesis. Interestingly, we found that overexpression of ELMO1 and mutant NOD2 (L1007fs) were not able to clear the CD-associated adherent invasive E. coli (AIEC-LF82). The functional implications of ELMO1-NOD2 interaction in epithelial cells were evaluated by using enteroid-derived monolayers (EDMs) from ELMO1 and NOD2 KO mice. Subsequently we also assessed the immune response in J774 macrophages depleted of either ELMO1 or NOD2 or both. The infection of murine EDMs with AIEC-LF82 showed higher bacterial load in ELMO1-KO, NOD2 KO EDMs, and ELMO1 KO EDMs treated with NOD2 inhibitors. The murine macrophage cells showed that the downregulation of ELMO1 and NOD2 is associated with impaired bacterial clearance that is linked to reduce pro-inflammatory cytokines and reactive oxygen species. Our results indicated that the crosstalk between microbial sensors in enteric infection and inflammatory diseases impacts the fate of the bacterial load and disease pathogenesis.
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- 2023
43. Characterization of the Membrane-Associated Electron-Bifurcating Flavoenzyme EtfABCX from the Hyperthermophilic Bacterium Thermotoga maritima.
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Ge, Xiaoxuan, Schut, Gerrit, Tran, Jessica, Poole Ii, Farris, Niks, Dimitri, Menjivar, Kevin, Hille, Russ, and Adams, Michael
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Ferredoxins ,Thermotoga maritima ,NAD ,Electrons ,Flavin-Adenine Dinucleotide ,Escherichia coli ,Vitamin K 2 ,Bacteria ,Electron Transport ,Oxidation-Reduction ,Archaea - Abstract
Electron bifurcation is an energy-conservation mechanism in which a single enzyme couples an exergonic reaction with an endergonic one. Heterotetrameric EtfABCX drives the reduction of low-potential ferredoxin (E° ∼ -450 mV) by oxidation of the midpotential NADH (E° = -320 mV) by simultaneously coupling the reaction to reduction of the high-potential menaquinone (E° = -74 mV). Electron bifurcation occurs at the NADH-oxidizing bifurcating-flavin adenine dinucleotide (BF-FAD) in EtfA, which has extremely crossed half-potentials and passes the first, high-potential electron to an electron-transferring FAD and via two iron-sulfur clusters eventually to menaquinone. The low-potential electron on the BF-FAD semiquinone simultaneously reduces ferredoxin. We have expressed the genes encodingThermotoga maritimaEtfABCX in E. coli and purified the EtfABCX holoenzyme and the EtfAB subcomplex. The bifurcation activity of EtfABCX was demonstrated by using electron paramagnetic resonance (EPR) to follow accumulation of reduced ferredoxin. To elucidate structural factors that impart the bifurcating ability, EPR and NADH titrations monitored by visible spectroscopy and dye-linked enzyme assays have been employed to characterize four conserved residues, R38, P239, and V242 in EtfA and R140 in EtfB, in the immediate vicinity of the BF-FAD. The R38, P239, and V242 variants showed diminished but still significant bifurcation activity. Despite still being partially reduced by NADH, the R140 variant had no bifurcation activity, and electron transfer to its two [4Fe-4S] clusters was prevented. The role of R140 is discussed in terms of the bifurcation mechanism in EtfABCX and in the other three families of bifurcating enzymes.
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- 2023
44. A unified Watson-Crick geometry drives transcription of six-letter expanded DNA alphabets by E. coli RNA polymerase.
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Oh, Juntaek, Shan, Zelin, Hoshika, Shuichi, Xu, Jun, Chong, Jenny, Benner, Steven, Lyumkis, Dmitry, and Wang, Dong
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Escherichia coli ,DNA ,Base Pairing ,Nucleotides ,Hydrogen Bonding - Abstract
Artificially Expanded Genetic Information Systems (AEGIS) add independently replicable unnatural nucleotide pairs to the natural G:C and A:T/U pairs found in native DNA, joining the unnatural pairs through alternative modes of hydrogen bonding. Whether and how AEGIS pairs are recognized and processed by multi-subunit cellular RNA polymerases (RNAPs) remains unknown. Here, we show that E. coli RNAP selectively recognizes unnatural nucleobases in a six-letter expanded genetic system. High-resolution cryo-EM structures of three RNAP elongation complexes containing template-substrate UBPs reveal the shared principles behind the recognition of AEGIS and natural base pairs. In these structures, RNAPs are captured in an active state, poised to perform the chemistry step. At this point, the unnatural base pair adopts a Watson-Crick geometry, and the trigger loop is folded into an active conformation, indicating that the mechanistic principles underlying recognition and incorporation of natural base pairs also apply to AEGIS unnatural base pairs. These data validate the design philosophy of AEGIS unnatural basepairs. Further, we provide structural evidence supporting a long-standing hypothesis that pair mismatch during transcription occurs via tautomerization. Together, our work highlights the importance of Watson-Crick complementarity underlying the design principles of AEGIS base pair recognition.
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- 2023
45. Evaluating E. coli genome‐scale metabolic model accuracy with high‐throughput mutant fitness data
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Bernstein, David B, Akkas, Batu, Price, Morgan N, and Arkin, Adam P
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Biological Sciences ,Industrial Biotechnology ,Biotechnology ,Genetics ,Escherichia coli ,Genome ,Chromosome Mapping ,Carbon ,Models ,Biological ,flux balance analysis ,genome-scale metabolic model ,RB-TnSeq ,Biochemistry and Cell Biology ,Other Biological Sciences ,Bioinformatics ,Biochemistry and cell biology - Abstract
The Escherichia coli genome-scale metabolic model (GEM) is an exemplar systems biology model for the simulation of cellular metabolism. Experimental validation of model predictions is essential to pinpoint uncertainty and ensure continued development of accurate models. Here, we quantified the accuracy of four subsequent E. coli GEMs using published mutant fitness data across thousands of genes and 25 different carbon sources. This evaluation demonstrated the utility of the area under a precision-recall curve relative to alternative accuracy metrics. An analysis of errors in the latest (iML1515) model identified several vitamins/cofactors that are likely available to mutants despite being absent from the experimental growth medium and highlighted isoenzyme gene-protein-reaction mapping as a key source of inaccurate predictions. A machine learning approach further identified metabolic fluxes through hydrogen ion exchange and specific central metabolism branch points as important determinants of model accuracy. This work outlines improved practices for the assessment of GEM accuracy with high-throughput mutant fitness data and highlights promising areas for future model refinement in E. coli and beyond.
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- 2023
46. Sequence-based Functional Metagenomics Reveals Novel Natural Diversity of Functional CopA in Environmental Microbiomes.
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Li, Wenjun, Wang, Likun, Li, Xiaofang, Zheng, Xin, Cohen, Michael, and Liu, Yong-Xin
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CopA ,Cu resistance ,Evolutionary trace analysis ,Functional metagenomics ,Natural diversity ,Metagenomics ,Bacterial Proteins ,Phylogeny ,Microbiota ,Copper ,Metagenome ,Escherichia coli ,Genetic Variation - Abstract
Exploring the natural diversity of functional genes/proteins from environmental DNA in high throughput remains challenging. In this study, we developed a sequence-based functional metagenomics procedure for mining the diversity of copper (Cu) resistance gene copA in global microbiomes, by combining the metagenomic assembly technology, local BLAST, evolutionary trace analysis (ETA), chemical synthesis, and conventional functional genomics. In total, 87 metagenomes were collected from a public database and subjected to copA detection, resulting in 93,899 hits. Manual curation of 1214 hits of high confidence led to the retrieval of 517 unique CopA candidates, which were further subjected to ETA. Eventually, 175 novel copA sequences of high quality were discovered. Phylogenetic analysis showed that almost all these putative CopA proteins were distantly related to known CopA proteins, with 55 sequences from totally unknown species. Ten novel and three known copA genes were chemically synthesized for further functional genomic tests using the Cu-sensitive Escherichia coli (ΔcopA). The growth test and Cu uptake determination showed that five novel clones had positive effects on host Cu resistance and uptake. One recombinant harboring copA-like 15 (copAL15) successfully restored Cu resistance of the host with a substantially enhanced Cu uptake. Two novel copA genes were fused with the gfp gene and expressed in E. coli for microscopic observation. Imaging results showed that they were successfully expressed and their proteins were localized to the membrane. The results here greatly expand the diversity of known CopA proteins, and the sequence-based procedure developed overcomes biases in length, screening methods, and abundance of conventional functional metagenomics.
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- 2023
47. Contamination levels and phenotypic and genomic characterization of antimicrobial resistance in 'Escherichia coli' isolated from fresh salad vegetables in the United Arab Emirates
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Habib, Ihab, Al-Rifai, Rami H, Mohamed, Mohamed-Yousif Ibrahim, Ghazawi, Akela, Abdalla, Afra, Lakshmi, Glindya, Agamy, Neveen, and Khan, Mushtaq
- Published
- 2023
48. Electrospun statin-loaded nanofibers for treating diabetic wounds
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Pamu, Divya, Garikapati, Kusuma Kumari, Kuppusamy, Gowthamarajan, Krishnamurthy, Praveen Thaggikuppe, Ganesan, Srividhya, Naik, Mudavath Ravi, Karri, Veera Venkata Satyanarayana Reddy, Ponpandian, Nagamony, Alexiou, Athanasios, Antoniou, Sofia, Khan, Azmat Ali, Alanazi, Amer M., and Papadakis, Marios
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Streptozocin ,Statins ,Biological products ,Collagen ,Vascular endothelial growth factor ,Antibacterial agents ,Escherichia coli ,Engineering and manufacturing industries ,Science and technology - Abstract
This study aims to develop, characterize, and evaluate the best statin-loaded Collagen Silver nanoparticle-coated polycaprolactone nanofibers by electrospinning technique. The characterization and evaluation results reveal that nanofibers had shown an aligned Collagen Silver nanoparticle-coated polycaprolactone nanofibers distribution of fibers and aliment of collagen and silver nanoparticle coating material on the surface of the nanofibers. Furthermore, the electrospun nanofibers have shown no interaction between the drug and collagen and PCL polymeric material and have demonstrated controlled drug release for 360 h and significant degradation for 60 days of the study. In vitro, among the nanofiber treatment groups and the control groups, Pravastatin Collagen Silver nanoparticle-coated polycaprolactone nanofibers have shown cell proliferation and migration of NHDF cells and blood vessel development in the CAM assay and potential anti-bacterial activity against S. aureus and E. coli. In vivo, the Pravastatin Collagen Silver nanoparticle-coated polycaprolactone nanofibers have demonstrated improved wound recovery, reepithelization, fibroblast cell proliferation, angiogenesis, and ECM remodeling and enhanced VEGF-sprouting in the excised tissues of streptozotocin-induced diabetic rats for 7, 14, and 21 days of the study. Therefore, the developed Pravastatin Collagen Silver nanoparticle-coated polycaprolactone nanofibers could be a promising wound-dressing platform for effectively treating and managing diabetic wounds. Highlights * Nanofiber wound dressings provide numerous structural features in biomaterials. * Polycaprolactone can release the repurposed statin molecules into the wound site in a controlled manner. * In diabetic wounds, ECM tissue remodeling stimulates repair and vessel growth. * Pravastatin has been demonstrated to improve angiogenesis and ECM secretion for wound healing in diabetic conditions. KEYWORDS diabetic wounds, electrospinning, hypoxic microenvironment, nanofibers, pro-angiogenesis, re-epithelization, wound dressing, 1 | INTRODUCTION The most prevalent cause of death in diabetic individuals is diabetic wounds. Unfortunately, the bulk of the treatment options in the market do not entirely address how [...]
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- 2024
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49. Potentially Zoonotic Enteric Infections in Gorillas and Chimpanzees, Cameroon and Tanzania
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Strahan, Emily K., Witherbee, Jacob, Bergl, Richard, Lonsdorf, Elizabeth V., Mwacha, Dismas, Mjungu, Deus, Arandjelovic, Mimi, Ikfuingei, Romanus, Terio, Karen, Travis, Dominic A., and Gillespie, Thomas R.
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Chimpanzees ,Infection ,Phylogeny ,Zoonoses ,Resveratrol ,Disease susceptibility ,Adenoviruses ,Escherichia coli ,Health - Abstract
The close phylogenetic relationship between humans and great apes results in similarities in infection susceptibility and a high potential for pathogen exchange (2,2). Despite this zoonotic potential, previous studies of [...]
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- 2024
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50. Comparative study on the prevalence of Escherichia coli and it's antibiogram in Saum from three different storage places in household at Mizoram
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Debbarma, Binipi, Ralte, Lallawmzuali, Motina, E., Lalmuanpuia, Joy, Tolenkhomba, T.C., and Roychoudhury, Parimal
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- 2024
- Full Text
- View/download PDF
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