Concept: Escherichia coli
The hospital environment is a potential reservoir of bacteria with plasmids conferring carbapenem resistance. Our Hospital Epidemiology Service routinely performs extensive sampling of high-touch surfaces, sinks, and other locations in the hospital. Over a 2-year period, additional sampling was conducted at a broader range of locations, including housekeeping closets, wastewater from hospital internal pipes, and external manholes. We compared these data with previously collected information from 5 years of patient clinical and surveillance isolates. Whole-genome sequencing and analysis of 108 isolates provided comprehensive characterization of blaKPC/blaNDM-positive isolates, enabling an in-depth genetic comparison. Strikingly, despite a very low prevalence of patient infections with blaKPC-positive organisms, all samples from the intensive care unit pipe wastewater and external manholes contained carbapenemase-producing organisms (CPOs), suggesting a vast, resilient reservoir. We observed a diverse set of species and plasmids, and we noted species and susceptibility profile differences between environmental and patient populations of CPOs. However, there were plasmid backbones common to both populations, highlighting a potential environmental reservoir of mobile elements that may contribute to the spread of resistance genes. Clear associations between patient and environmental isolates were uncommon based on sequence analysis and epidemiology, suggesting reasonable infection control compliance at our institution. Nonetheless, a probable nosocomial transmission of Leclercia sp. from the housekeeping environment to a patient was detected by this extensive surveillance. These data and analyses further our understanding of CPOs in the hospital environment and are broadly relevant to the design of infection control strategies in many infrastructure settings.IMPORTANCE Carbapenemase-producing organisms (CPOs) are a global concern because of the morbidity and mortality associated with these resistant Gram-negative bacteria. Horizontal plasmid transfer spreads the resistance mechanism to new bacteria, and understanding the plasmid ecology of the hospital environment can assist in the design of control strategies to prevent nosocomial infections. A 5-year genomic and epidemiological survey was undertaken to study the CPOs in the patient-accessible environment, as well as in the plumbing system removed from the patient. This comprehensive survey revealed a vast, unappreciated reservoir of CPOs in wastewater, which was in contrast to the low positivity rate in both the patient population and the patient-accessible environment. While there were few patient-environmental isolate associations, there were plasmid backbones common to both populations. These results are relevant to all hospitals for which CPO colonization may not yet be defined through extensive surveillance.
Misfolded alpha-synuclein (AS) and other neurodegenerative disorder proteins display prion-like transmission of protein aggregation. Factors responsible for the initiation of AS aggregation are unknown. To evaluate the role of amyloid proteins made by the microbiota we exposed aged rats and transgenic C. elegans to E. coli producing the extracellular bacterial amyloid protein curli. Rats exposed to curli-producing bacteria displayed increased neuronal AS deposition in both gut and brain and enhanced microgliosis and astrogliosis compared to rats exposed to either mutant bacteria unable to synthesize curli, or to vehicle alone. Animals exposed to curli producing bacteria also had more expression of TLR2, IL-6 and TNF in the brain than the other two groups. There were no differences among the rat groups in survival, body weight, inflammation in the mouth, retina, kidneys or gut epithelia, and circulating cytokine levels. AS-expressing C. elegans fed on curli-producing bacteria also had enhanced AS aggregation. These results suggest that bacterial amyloid functions as a trigger to initiate AS aggregation through cross-seeding and also primes responses of the innate immune system.
Honey bee pollination is a key ecosystem service to nature and agriculture. However, biosafety research on genetically modified crops rarely considers effects on nurse bees from intact colonies, even though they receive and primarily process the largest amount of pollen. The objective of this study was to analyze the response of nurse bees and their gut bacteria to pollen from Bt maize expressing three different insecticidal Cry proteins (Cry1A.105, Cry2Ab2, and Cry3Bb1). Naturally Cry proteins are produced by bacteria (Bacillus thuringiensis). Colonies of Apis mellifera carnica were kept during anthesis in flight cages on field plots with the Bt maize, two different conventionally bred maize varieties, and without cages, 1-km outside of the experimental maize field to allow ad libitum foraging to mixed pollen sources. During their 10-days life span, the consumption of Bt maize pollen had no effect on their survival rate, body weight and rates of pollen digestion compared to the conventional maize varieties. As indicated by ELISA-quantification of Cry1A.105 and Cry3Bb1, more than 98% of the recombinant proteins were degraded. Bacterial population sizes in the gut were not affected by the genetic modification. Bt-maize, conventional varieties and mixed pollen sources selected for significantly different bacterial communities which were, however, composed of the same dominant members, including Proteobacteria in the midgut and Lactobacillus sp. and Bifidobacterium sp. in the hindgut. Surprisingly, Cry proteins from natural sources, most likely B. thuringiensis, were detected in bees with no exposure to Bt maize. The natural occurrence of Cry proteins and the lack of detectable effects on nurse bees and their gut bacteria give no indication for harmful effects of this Bt maize on nurse honey bees.
Understanding of soil processes is essential for addressing the global issues of food security, disease transmission and climate change. However, techniques for observing soil biology are lacking. We present a heterogeneous, porous, transparent substrate for in situ 3D imaging of living plants and root-associated microorganisms using particles of the transparent polymer, Nafion, and a solution with matching optical properties. Minerals and fluorescent dyes were adsorbed onto the Nafion particles for nutrient supply and imaging of pore size and geometry. Plant growth in transparent soil was similar to that in soil. We imaged colonization of lettuce roots by the human bacterial pathogen Escherichia coli O157:H7 showing micro-colony development. Micro-colonies may contribute to bacterial survival in soil. Transparent soil has applications in root biology, crop genetics and soil microbiology.
- Proceedings of the National Academy of Sciences of the United States of America
- Published about 6 years ago
The large volumes of sequencing data required to sample deeply the microbial communities of complex environments pose new challenges to sequence analysis. De novo metagenomic assembly effectively reduces the total amount of data to be analyzed but requires substantial computational resources. We combine two preassembly filtering approaches-digital normalization and partitioning-to generate previously intractable large metagenome assemblies. Using a human-gut mock community dataset, we demonstrate that these methods result in assemblies nearly identical to assemblies from unprocessed data. We then assemble two large soil metagenomes totaling 398 billion bp (equivalent to 88,000 Escherichia coli genomes) from matched Iowa corn and native prairie soils. The resulting assembled contigs could be used to identify molecular interactions and reaction networks of known metabolic pathways using the Kyoto Encyclopedia of Genes and Genomes Orthology database. Nonetheless, more than 60% of predicted proteins in assemblies could not be annotated against known databases. Many of these unknown proteins were abundant in both corn and prairie soils, highlighting the benefits of assembly for the discovery and characterization of novelty in soil biodiversity. Moreover, 80% of the sequencing data could not be assembled because of low coverage, suggesting that considerably more sequencing data are needed to characterize the functional content of soil.
Whole genome sequencing (WGS) is becoming available as a routine tool for clinical microbiology. If applied directly on clinical samples this could further reduce diagnostic time and thereby improve control and treatment. A major bottle-neck is the availability of fast and reliable bioinformatics tools. This study was conducted to evaluate the applicability of WGS directly on clinical samples and to develop easy-to-use bioinformatics tools for analysis of the sequencing data. Thirty-five random urine samples from patients with suspected urinary tract infections were examined using conventional microbiology, WGS of isolated bacteria and by directly sequencing on pellets from the urine. A rapid method for analyzing the sequence data was developed. Bacteria were cultivated from 19 samples, but only in pure culture from 17. WGS improved the identification of the cultivated bacteria and almost complete agreement was observed between phenotypic and predicted antimicrobial susceptibility. Complete agreement was observed between species identification, multi-locus-sequence typing and phylogenetic relationship for the Escherichia coli and Enterococcus faecalis isolates when comparing the results of WGS of cultured isolates and directly from the urine samples. Sequencing directly from the urine enabled bacterial identification in polymicrobic samples. Additional putative pathogenic strains were observed in some culture negative samples. WGS directly on clinical samples can provide clinically relevant information and drastically reduce diagnostic time. This may prove very useful, but the need for data analysis is still a hurdle to clinical implementation. To overcome this problem a publicly available bioinformatics tool was developed in this study.
In eukaryotes, the classical form of programmed cell death (PCD) is apoptosis, which has as its specific characteristics DNA fragmentation and membrane depolarization. In Escherichia coli a different PCD system has been reported. It is mediated by the toxin-antitoxin system module mazEF. The E. coli mazEF module is one of the most thoroughly studied toxin-antitoxin systems. mazF encodes a stable toxin, MazF, and mazE encodes a labile antitoxin, MazE, which prevents the lethal effect of MazF. mazEF-mediated cell death is a population phenomenon requiring the quorum-sensing pentapeptide NNWNN designated Extracellular Death Factor (EDF). mazEF is triggered by several stressful conditions, including severe damage to the DNA. Here, using confocal microscopy and FACS analysis, we show that under conditions of severe DNA damage, the triggered mazEF-mediated cell death pathway leads to the inhibition of a second cell death pathway. The latter is an apoptotic-like death (ALD); ALD is mediated by recA and lexA. The mazEF-mediated pathway reduces recA mRNA levels. Based on these results, we offer a molecular model for the maintenance of an altruistic characteristic in cell populations. In our model, the ALD pathway is inhibited by the altruistic EDF-mazEF-mediated death pathway.
In 2015, scientists reported the emergence of the plasmid-encoded mcr-1 gene conferring bacterial resistance to the antibiotic colistin (1), signaling potential emergence of a pandrug-resistant bacterium. In May 2016, mcr-1-positive Escherichia coli was first isolated from a specimen from a U.S. patient (2) when a Pennsylvania woman was evaluated for a urinary tract infection. The urine culture and subsequent testing identified the gene in an extended-spectrum beta-lactamase (ESBL)-producing E. coli with reduced susceptibility to colistin. The patient had no international travel for approximately 1 year, no livestock exposure, and a limited role in meal preparation with store-bought groceries; however, she had multiple and repeated admissions to four medical facilities during 2016.
BACKGROUND: Beneficial mutations play an essential role in bacterial adaptation, yet little is known about their fitness effects across genetic backgrounds and environments. One prominent example of bacterial adaptation is antibiotic resistance. Until recently, the paradigm has been that antibiotic resistance is selected by the presence of antibiotics because resistant mutations confer fitness costs in antibiotic free environments. In this study we show that it is not always the case, documenting the selection and fixation of resistant mutations in populations of Escherichia coli B that had never been exposed to antibiotics but instead evolved for 2000 generations at high temperature (42.2[degree sign]C). RESULTS: We found parallel mutations within the rpoB gene encoding the beta subunit of RNA polymerase. These amino acid substitutions conferred different levels of rifampicin resistance. The resistant mutations typically appeared, and were fixed, early in the evolution experiment. We confirmed the high advantage of these mutations at 42.2[degree sign]C in glucose-limited medium. However, the rpoB mutations had different fitness effects across three genetic backgrounds and six environments. CONCLUSIONS: We describe resistance mutations that are not necessarily costly in the absence of antibiotics or compensatory mutations but are highly beneficial at high temperature and low glucose. Their fitness effects depend on the environment and the genetic background, providing glimpses into the prevalence of epistasis and pleiotropy.
The mcr-1 gene confers resistance to the polymyxins, including the antibiotic colistin, a medication of last resort for multidrug-resistant infections. The mcr-1 gene was first reported in 2015 in food, animal, and patient isolates from China (1) and is notable for being the first plasmid-mediated colistin resistance mechanism to be identified. Plasmids can be transferred between bacteria, potentially spreading the resistance gene to other bacterial species. Since its discovery, the mcr-1 gene has been reported from Africa, Asia, Europe, South America, and North America (2,3), including the United States, where it has been identified in Escherichia coli isolated from three patients and from two intestinal samples from pigs (2,4-6). In July 2016, the Pathogen Detection System at the National Center for Biotechnology Information (Bethesda, Maryland) identified mcr-1 in the whole genome sequence of an E. coli isolate from a Connecticut patient (7); this is the fourth isolate from a U.S. patient to contain the mcr-1 gene.