Tracing Antibiotic Resistance Genes along the Irrigation Water Chain to Chive: Does Tap or Surface Water Make a Difference?

Irrigation water is well known as potential source of pathogens in fresh produce. However, its role in transferring antibiotic resistance determinants is less well investigated. Therefore, we analyzed the contribution of surface and tap water to the resistome of overhead-irrigated chive plants. Field-grown chive was irrigated with either surface water (R-system) or tap water (D-system), from planting to harvest. Water along the two irrigation chains as well as the respective plants were repeatedly sampled and screened for 264 antibiotic resistance genes (ARGs) and mobile genetic elements (MGEs), using high-capacity qPCR. Differentially abundant (DA) ARGs were determined by comparing the two systems. On R-chive, β-lactam ARGs, multidrug-resistance (MDR) determinants, and MGEs were most abundant, while D-chive featured DA ARGs from the vancomycin class. Diversity and number of DA ARGs was the highest on young chives, strongly diminished at harvest, and increased again at the end of shelf life. Most ARGs highly enriched on R- compared to D-chive were also enriched in R- compared to D-sprinkler water, indicating that water played a major role in ARG enrichment. Of note, blaKPC was detected at high levels in surface water and chive. We conclude that water quality significantly affects the resistome of the irrigated produce

Transferable Extended-Spectrum β-Lactamase (ESBL) Plasmids in Enterobacteriaceae from Irrigation Water

Extended-spectrum β-lactamase (ESBL)-producing Enterobacteriaceae are classified as serious threats to human health by the U.S. Centers for Disease Control and Prevention. Water used for irrigation of fresh produce can transmit such resistant bacteria directly to edible plant parts. We screened ESBL-producing Escherichia coli, Enterobacter cloacae, and Citrobacter freundii isolated from irrigation water for their potential to transmit resistance to antibiotic-susceptible E. coli. All strains were genome-sequenced and tested in vitro for transmission of resistance to third-generation cephalosporins on solid agar as well as in liquid culture. Of the 19 screened isolates, five ESBL-producing E. coli were able to transfer resistance with different efficiency to susceptible recipient E. coli. Transconjugant strains were sequenced for detection of transferred antibiotic resistance genes (ARGs) and compared to the known ARG pattern of their respective donors. Additionally, phenotypic resistance patterns were obtained for both transconjugant and corresponding donor strains, confirming ESBL-producing phenotypes of all obtained transconjugants

Beyond the matrix-assisted laser desorption ionization (MALDI) biotyping workflow: in search of microorganism-specific tryptic peptides enabling discrimination of subspecies

A well-accepted method for identification of microorganisms uses matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) coupled to analysis software which identifies and classifies the organism according to its ribosomal protein spectral profile. The method, called MALDI biotyping, is widely used in clinical diagnostics and has partly replaced conventional microbiological techniques such as biochemical identification due to its shorter time to result (minutes for MALDI biotyping versus hours or days for classical phenotypic or genotypic identification). Besides its utility for identifying bacteria, MS-based identification has been shown to be applicable also to yeasts and molds. A limitation to this method, however, is that accurate identification is most reliably achieved on the species level on the basis of reference mass spectra, making further phylogenetic classification unreliable. Here, it is shown that combining tryptic digestion of the acid/organic solvent extracted (classical biotyping preparation) and resolubilized proteins, nano-liquid chromatography (nano-LC), and subsequent identification of the peptides by MALDI-tandem TOF (MALDI-TOF/TOF) mass spectrometry increases the discrimination power to the level of subspecies. As a proof of concept, using this targeted proteomics workflow, we have identified subspecies-specific biomarker peptides for three Salmonella subspecies, resulting in an extension of the mass range and type of proteins investigated compared to classical MALDI biotyping. This method therefore offers rapid and cost-effective identification and classification of microorganisms at a deeper taxonomic level

Long-Term Persistence of blaCTX-M-15 in Soil and Lettuce after Introducing Extended-Spectrum β-Lactamase (ESBL)-Producing Escherichia coli via Manure or Water

The number of environmental antibiotic-resistant bacteria (ARB) has increased dramatically since the start of antibiotic mass production for broad bacterial infection treatment in 1944. Nowadays, ARB and their resistance-determining genes (ARGs) are readily detected in all environments, including the human food chain. A highly relevant food group in this context is fresh produce, frequent raw consumption of which facilitates direct transfer of ARB and ARGs to the consumer. Here, we investigate the persistence of an extended-spectrum β-lactamase (ESBL)-producing Escherichia coli (E. coli) pEK499 and its clinically most important ARG (blaCTX-M-15), after introduction via irrigation water or manure into a lettuce-growing system. Culturable ESBL-producing E. coli persisted longest in soil and when introduced via manure (until 9 weeks after introduction), while being undetectable on lettuce beyond day 7. In contrast, qPCR detection of blaCTX-M-15 was much more frequent: introduction via water significantly increased blaCTX-M-15 on lettuce until week 4, as opposed to manure, which affected the soil in the long-term (9 weeks) while leading to blaCTX-M-15 detection on lettuce until day 7 only. Our findings demonstrate long-term persistence of undesired ARB and ARG after their introduction via both irrigation and amendment. Such an understanding of the persistence kinetics of an ESBL-producing E. coli and plasmid-encoded blaCTX-M-15 aids the determination of critical actions in order to mitigate their transfer to the consumer

Valores en la educación infantil : un año en la clase de las Jirafas

Resumen basado en el de la publicaciónSe pretende mostrar los dinamismos de la educación en valores que se dan día a día en una clase de Educación Infantil. Durante un curso escolar, se acompaña a un grupo de alumnos y a su profesora para ver cómo en la clase de las jirafas se cuidan las prácticas de gestión del conocimiento, se fomentan los espacios de dialogo y deliberación y se construyen lazos que no dejan indiferentes a ninguno de sus protagonistas. Y es que, la educación en valores en una comunidad escolar es una tarea dinámica, una experiencia que impregna el clima y la atmosfera de una institución y que permite a sus protagonistas vivir experiencias de autonomía y solidaridad en primera persona.CataluñaBiblioteca de Educación del Ministerio de Educación, Cultura y Deporte; Calle San Agustín, 5 - 3 planta; 28014 Madrid; Tel. +34917748000; [email protected]

Bacillus cereus Group-Type Strain-Specific Diagnostic Peptides

The Bacillus cereus group consists of eight very closely related species and comprises both harmless and human pathogenic species such as Bacillus anthracis, Bacillus cereus, and Bacillus cytotoxicus. Numerous efforts have been undertaken to allow presumptive differentiation of B. cereus group species from one another. However, methods to rapidly and accurately distinguish these species are currently lacking. We confirmed that classical matrix-assisted laser desorption/ionization-time-of-flight (MALDI-TOF) biotyping cannot achieve reliable identification of each type strain. We therefore assigned type strain-specific diagnostic peptides to the B. cereus group based on comparisons of their proteomic profiles. The number of diagnostic peptides varied remarkably in a type strain-dependent manner. The accuracy of the reference database was crucial to validate candidate diagnostic peptides and led to a noteworthy reduction of verified diagnostic peptides. Diagnostic peptides ranged from one for B. weihenstephanensis to 62 for B. pseudomycoides and were associated with proteins involved in diverse biological processes, e.g. amino acid biosynthesis, cell envelope, cellular processes, energy metabolism, and transport processes. However, 45.6% of all diagnostic peptides comprised currently unclassified proteins or proteins of unknown function. In addition, a phylogenetic tree based on clustering of theoretical precursor masses deduced from in silico-generated tryptic peptides was reconstructed

Dynamics of culturable mesophilic bacterial communities of three fresh herbs and their production environment

Aim Investigate dynamics of culturable mesophilic bacteria and selected food-contaminating bacteria from three herbs and their production environment. Methods and Results Marjoram, basil and thyme were investigated during one growing season by sampling plants, organic fertilizers, soil, irrigation water and marketed products. Mesophilic bacteria and selected food-contaminating bacteria (Escherichia coli, Enterococcus spp., Bacillus cereus group) were cultured and identified by MALDI biotyping. Culturable mesophilic bacteria on marjoram and basil plants decreased over time by two orders of magnitude starting at above 106 colony forming units per gram (CFU per g), while they remained constant on thyme (~104 CFU per g). Compared to the last field sample, mesophilic bacteria were increased on all market-ready products by one order of magnitude. Marjoram and basil were dominated by B. cereus group, Enterobacter spp. and Pseudomonas spp., thyme by Bacillus spp. and Pseudomonas spp. All selected food-contaminating bacteria were detected in soil and reservoir-sourced irrigation water, whereas in municipal water, only B. cereus group and rarely Enterococcus spp. were found. Escherichia coli was detected only on young marjoram and basil plants (5 × 102 and 5 × 101 CFU per g, respectively), whereas Enterococcus spp. and B. cereus group were consistently detected on these two herbs. Thyme plants only contained B. cereus group consistently (above 103 CFU per g). Marketed marjoram and thyme contained Enterococcus spp. (5 × 102 and 104 CFU per g) and B. cereus group (~5 × 102 CFU per g), while no selected food-contaminating bacteria were found on marketed basil. Conclusions Overall, culturable mesophilic bacteria were dominated by Pseudomonas spp. and Bacillus spp., with increased numbers on market-ready products. Selected food-contaminating bacteria were readily detectable, however, only the B. cereus group was found throughout in all systems. Significance and Impact of the Study Insight into composition and development of mesophilic bacterial communities and selected food-contaminating bacteria of fresh herbs contributes to estimating consumer exposure.ISSN:1364-5072ISSN:1365-267

<i>Bacillus cereus</i> Group-Type Strain-Specific Diagnostic Peptides

The <i>Bacillus cereus</i> group consists of eight very closely related species and comprises both harmless and human pathogenic species such as <i>Bacillus anthracis, Bacillus cereus</i>, and <i>Bacillus cytotoxicus</i>. Numerous efforts have been undertaken to allow presumptive differentiation of <i>B. cereus</i> group species from one another. However, methods to rapidly and accurately distinguish these species are currently lacking. We confirmed that classical matrix-assisted laser desorption/ionization-time-of-flight (MALDI-TOF) biotyping cannot achieve reliable identification of each type strain. We therefore assigned type strain-specific diagnostic peptides to the <i>B. cereus</i> group based on comparisons of their proteomic profiles. The number of diagnostic peptides varied remarkably in a type strain-dependent manner. The accuracy of the reference database was crucial to validate candidate diagnostic peptides and led to a noteworthy reduction of verified diagnostic peptides. Diagnostic peptides ranged from one for <i>B. weihenstephanensis</i> to 62 for <i>B. pseudomycoides</i> and were associated with proteins involved in diverse biological processes, e.g. amino acid biosynthesis, cell envelope, cellular processes, energy metabolism, and transport processes. However, 45.6% of all diagnostic peptides comprised currently unclassified proteins or proteins of unknown function. In addition, a phylogenetic tree based on clustering of theoretical precursor masses deduced from <i>in silico</i>-generated tryptic peptides was reconstructed