scholarly journals Metagenomic survey of agricultural water using long read sequencing: considerations for a successful analysis

2021 ◽  
Author(s):  
Meghan Maguire ◽  
Julie A. Kase ◽  
Eric W. Brown ◽  
Marc Allard ◽  
Steve Musser ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Magnetic Bead ◽  
The United States ◽  
Preliminary Evidence ◽  
Agricultural Water ◽  
Microbial Identification ◽  
Before And After ◽  
Specific Strain ◽  
Long Read ◽  
Microbiological Techniques

Abstract Background Leafy greens are responsible for nearly half of the produce-related Shiga toxin-producing Escherichia coli (STEC) outbreaks in the United States and recent investigations have implicated agricultural water as a potential source. Current FDA detection protocols require extensive analysis time. Development of methods for rapid detection of STECs in the field are imperative to maintain food safety. We aimed to use Oxford Nanopore rapid sequencing kits for an in-field determination of agricultural water microbiome and possible detection and characterization of STECs strain(s) in these samples. Results We tested the performance of the nanopore rapid sequencing kit (RAD004) for fast microbiome determination using the well characterized ZymoBIOMICS mock microbial community and the number of reads for each identified species was present in the expected proportion. Rapid sequencing kit (LRK001 and RAD004) library preparation of DNA extracted from agricultural water resulted in poor nanopore sequencing reactions, with low output (0.3–1.7 M reads), a high proportion of failed reads (50–60%), and highly sheared DNA before and after a magnetic bead clean up. To improve performance, we prepared a DNA library with the ligation kit (LSK109), which includes multiple cleaning steps, reducing inherent inhibitors and producing a better outcome (2.2 M reads, 15% failed reads). Metagenomic composition of four sample sites determined using the ligation kit, showed a highly diverse bacterial community for each site, with 11 total genera identified, including Synechococcus and Cyanobium. No definitive presence of STEC could be confirmed in any of the sites. Approximately 100 reads from each site (0.02% of total reads) were identified as Escherichia coli, but the specific strain or their virulence genes could not be detected. Sites 9, 10, and 12 were found to be positive for STEC presence by microbiological techniques after enrichment. Conclusions The rapid sequencing kits can be appropriate for genus or species level microbial identification, but we recommend the use of the ligation kit for increased sequencing depth and removal of contaminants in agricultural water. However, we were not able to identify any STEC strains in these nanopore microbiome samples, due to low initial concentrations. The results from this pilot study provide preliminary evidence that MinION sequencing of agricultural water using the ligation kit has the potential to be used for rapid microbiome determination in the field with optimal results for water quality surveillance.

scholarly journals Longitudinal Assessment of the Dynamics of Escherichia coli, Total Coliforms, Enterococcus spp., and Aeromonas spp. in Alternative Irrigation Water Sources: a CONSERVE Study

2020 ◽  
Vol 86 (20) ◽  
Author(s):  
Sultana Solaiman ◽  
Sarah M. Allard ◽  
Mary Theresa Callahan ◽  
Chengsheng Jiang ◽  
Eric Handy ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Food Safety ◽  
Irrigation Water ◽  
Reclaimed Water ◽  
The United States ◽  
Water Sources ◽  
Agricultural Water ◽  
Content Type ◽  
E Coli ◽  
Alternative Irrigation

ABSTRACT As climate change continues to stress freshwater resources, we have a pressing need to identify alternative (nontraditional) sources of microbially safe water for irrigation of fresh produce. This study is part of the center CONSERVE, which aims to facilitate the adoption of adequate agricultural water sources. A 26-month longitudinal study was conducted at 11 sites to assess the prevalence of bacteria indicating water quality, fecal contamination, and crop contamination risk (Escherichia coli, total coliforms [TC], Enterococcus, and Aeromonas). Sites included nontidal freshwater rivers/creeks (NF), a tidal brackish river (TB), irrigation ponds (PW), and reclaimed water sites (RW). Water samples were filtered for bacterial quantification. E. coli, TC, enterococci (∼86%, 98%, and 90% positive, respectively; n = 333), and Aeromonas (∼98% positive; n = 133) were widespread in water samples tested. Highest E. coli counts were in rivers, TC counts in TB, and enterococci in rivers and ponds (P < 0.001 in all cases) compared to other water types. Aeromonas counts were consistent across sites. Seasonal dynamics were detected in NF and PW samples only. E. coli counts were higher in the vegetable crop-growing (May-October) than nongrowing (November-April) season in all water types (P < 0.05). Only one RW and both PW sites met the U.S. Food Safety Modernization Act water standards. However, implementation of recommended mitigation measures of allowing time for microbial die-off between irrigation and harvest would bring all other sites into compliance within 2 days. This study provides comprehensive microbial data on alternative irrigation water and serves as an important resource for food safety planning and policy setting. IMPORTANCE Increasing demands for fresh fruit and vegetables, a variable climate affecting agricultural water availability, and microbial food safety goals are pressing the need to identify new, safe, alternative sources of irrigation water. Our study generated microbial data collected over a 2-year period from potential sources of irrigation (rivers, ponds, and reclaimed water sites). Pond water was found to comply with Food Safety Modernization Act (FSMA) microbial standards for irrigation of fruit and vegetables. Bacterial counts in reclaimed water, a resource that is not universally allowed on fresh produce in the United States, generally met microbial standards or needed minimal mitigation. We detected the most seasonality and the highest microbial loads in river water, which emerged as the water type that would require the most mitigation to be compliant with established FSMA standards. This data set represents one of the most comprehensive, longitudinal analyses of alternative irrigation water sources in the United States.

scholarly journals Applying torque to the Escherichia coli flagellar motor using magnetic tweezers

2017 ◽  
Vol 7 (1) ◽  
Author(s):  
Maarten M. van Oene ◽  
Laura E. Dickinson ◽  
Bronwen Cross ◽  
Francesco Pedaci ◽  
Jan Lipfert ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
External Load ◽  
Magnetic Bead ◽  
Magnetic Tweezers ◽  
Flagellar Motor ◽  
Motor Speed ◽  
Motor Torque ◽  
Rotary Engine ◽  
Flagellar Motors ◽  
Before And After

Abstract The bacterial flagellar motor of Escherichia coli is a nanoscale rotary engine essential for bacterial propulsion. Studies on the power output of single motors rely on the measurement of motor torque and rotation under external load. Here, we investigate the use of magnetic tweezers, which in principle allow the application and active control of a calibrated load torque, to study single flagellar motors in Escherichia coli. We manipulate the external load on the motor by adjusting the magnetic field experienced by a magnetic bead linked to the motor, and we probe the motor’s response. A simple model describes the average motor speed over the entire range of applied fields. We extract the motor torque at stall and find it to be similar to the motor torque at drag-limited speed. In addition, use of the magnetic tweezers allows us to force motor rotation in both forward and backward directions. We monitor the motor’s performance before and after periods of forced rotation and observe no destructive effects on the motor. Our experiments show how magnetic tweezers can provide active and fast control of the external load while also exposing remaining challenges in calibration. Through their non-invasive character and straightforward parallelization, magnetic tweezers provide an attractive platform to study nanoscale rotary motors at the single-motor level.

Enumeration of Campylobacter spp., Escherichia coli, and Salmonella in Broiler Carcass Rinses before and after Simulated Transport in Artificial Ice for 24 Hours

2009 ◽  
Vol 72 (5) ◽  
pp. 1099-1101 ◽  
Author(s):  
NORMAN J. STERN ◽  
J. ERIC LINE
Keyword(s):  
United States ◽  
Escherichia Coli ◽  
Tap Water ◽  
Water Hardness ◽  
The United States ◽  
E Coli ◽  
Before And After ◽  
Peptone Water ◽  
Campylobacter Spp

The maintenance and survival of target pathogens during transport from the field collection site to the analytical laboratory is essential for obtaining accurate and reliable data. This study was conducted to compare the efficacy of sterile tap water (SW), buffered peptone water (BPW), and universal preenrichment broth (UP) for maintaining populations of Campylobacter spp., Salmonella, and Escherichia coli for 24 h under simulated transport conditions. Freshly processed broiler carcasses (n = 100) were rinsed in SW. The rinses were divided, and components were added to create equal volumes of rinse samples consisting of SW, BPW, and UP. The rinses were analyzed for the target organisms immediately and again after 24 h of simulated chilled transport conditions. The only meaningful difference between the different transport media was found for UP, which recovered fewer E. coli than did either SW or BPW. These findings support the conclusion that either SW or BPW should be used as a broiler carcass rinse and/or transport medium to accurately depict the levels or presence of these three target bacteria as a whole. Because potable water differs in pH and hardness across the United States, a follow-up study was conducted to investigate whether water hardness or pH within the ranges normally found across the United States would affect Campylobacter recovery from carcass rinses. No significant differences were detected.

scholarly journals The mcr-9 Gene of Salmonella and Escherichia coli Is Not Associated with Colistin Resistance in the United States

2020 ◽  
Vol 64 (8) ◽  
Author(s):  
Gregory H. Tyson ◽  
Cong Li ◽  
Chih-Hao Hsu ◽  
Sherry Ayers ◽  
Stacey Borenstein ◽  
...  
Keyword(s):  
United States ◽  
Escherichia Coli ◽  
Antimicrobial Resistance ◽  
Resistance Genes ◽  
Clinical Relevance ◽  
The United States ◽  
Colistin Resistance ◽  
Content Type ◽  
Long Read ◽  
Genetic Context

ABSTRACT Reports of transmissible colistin resistance show the importance of comprehensive colistin resistance surveillance. Recently, a new allele of the mobile colistin resistance (mcr) gene family designated mcr-9, which shows variation in genetic context and colistin susceptibility, was reported. We tested over 100 Salmonella enterica and Escherichia coli isolates with mcr-9 from the National Antimicrobial Resistance Monitoring System (NARMS) in the United States for their susceptibility to colistin and found that every isolate was susceptible, with an MIC of ≤1 μg/ml. Long-read sequencing of 12 isolates revealed mcr-9 on IncHI plasmids that were either independent or integrated into the chromosome. Overall, these results demonstrate that caution is necessary when determining the clinical relevance of new resistance genes.

Changes in Perceptions of Filipino Immigrants about Themselves and of Americans Before and After Coming to the United States

2019 ◽  
Author(s):  
Gremil Alessandro Naz
Keyword(s):  
United States ◽  
Symbolic Interaction ◽  
The United States ◽  
The Philippines ◽  
Appraisal Theory ◽  
American Values ◽  
The Us ◽  
Before And After ◽  
Symbolic Interaction Theory ◽  
Filipino Immigrants

<p>This paper examines the changes in Filipino immigrants’ perceptions about themselves and of Americans before and after coming to the United States. Filipinos have a general perception of themselves as an ethnic group. They also have perceptions about Americans whose media products regularly reach the Philippines. Eleven Filipinos who have permanently migrated to the US were interviewed about their perceptions of Filipinos and Americans. Before coming to the US, they saw themselves as hardworking, family-oriented, poor, shy, corrupt, proud, adaptable, fatalistic, humble, adventurous, persevering, gossipmonger, and happy. They described Americans as rich, arrogant, educated, workaholic, proud, powerful, spoiled, helpful, boastful, materialistic, individualistic, talented, domineering, friendly, accommodating, helpful, clean, and kind. Most of the respondents changed their perceptions of Filipinos and of Americans after coming to the US. They now view Filipinos as having acquired American values or “Americanized.” On the other hand, they stopped perceiving Americans as a homogenous group possessing the same values after they got into direct contact with them. The findings validate social perception and appraisal theory, and symbolic interaction theory.</p>

scholarly journals Health Insurance Coverage before and after the Affordable Care Act in the USA

Sci ◽  
2021 ◽  
Vol 3 (2) ◽  
pp. 25
Author(s):  
Jesse Patrick ◽  
Philip Q. Yang
Keyword(s):  
Health Insurance ◽  
National Health ◽  
Affordable Care Act ◽  
The United States ◽  
Insurance Status ◽  
Health Insurance Status ◽  
Full Implementation ◽  
Before And After ◽  
The Affordable Care Act ◽  
Partial Implementation

The Affordable Care Act (ACA) is at the crossroads. It is important to evaluate the effectiveness of the ACA in order to make rational decisions about the ongoing healthcare reform, but existing research into its effect on health insurance status in the United States is insufficient and descriptive. Using data from the National Health Interview Surveys from 2009 to 2015, this study examines changes in health insurance status and its determinants before the ACA in 2009, during its partial implementation in 2010–2013, and after its full implementation in 2014 and 2015. The results of trend analysis indicate a significant increase in national health insurance rate from 82.2% in 2009 to 89.4% in 2015. Logistic regression analyses confirm the similar impact of age, gender, race, marital status, nativity, citizenship, education, and poverty on health insurance status before and after the ACA. Despite similar effects across years, controlling for other variables, youth aged 26 or below, the foreign-born, Asians, and other races had a greater probability of gaining health insurance after the ACA than before the ACA; however, the odds of obtaining health insurance for Hispanics and the impoverished rose slightly during the partial implementation of the ACA, but somewhat declined after the full implementation of the ACA starting in 2014. These findings should be taken into account by the U.S. Government in deciding the fate of the ACA.

scholarly journals Multidrug Resistance Dissemination in Escherichia coli Isolated from Wild Animals: Bacterial Clones and Plasmid Complicity

2021 ◽  
Vol 12 (1) ◽  
pp. 123-137
Author(s):  
Carolina Sabença ◽  
Gilberto Igrejas ◽  
Patrícia Poeta ◽  
Frédéric Robin ◽  
Richard Bonnet ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Epidemiological Data ◽  
Generation Cephalosporin ◽  
Wild Animals ◽  
Variable Regions ◽  
E Coli ◽  
Wild Fauna ◽  
Long Read ◽  
Sequence Types ◽  
Animal Isolates

Objectives. Epidemiological data concerning third-generation cephalosporin (3GC) resistance in wild fauna are scarce. The aim of this study was to characterize the resistance genes, their genetic context, and clonal relatedness in 17 Escherichia coli resistant to 3GC isolated from wild animals. Methods. The isolates were characterized by short-read whole genome sequencing, and long-read sequencing was used for the hybrid assembly of plasmid sequences. Results. The 3GC resistance gene most identified in the isolates was the extended-spectrum β-lactamases (ESBL)-encoding gene blaCTX-M-1 (82.3%), followed by blaCTX-M-32 (5.9%), blaCTX-M-14 (5.9%), and blaSHV-12 (5.9%). E. coli isolates mainly belonged to the sequence types (STs) rarely reported from humans. The single nucleotide polymorphism (SNP)-based typing showed that most E. coli genomes from wild animals (wild boars, birds of prey, and buzzards) formed clonal clusters (<5 SNPs), showing a clonal dissemination crossing species boundaries. blaCTX-M-1-harboring IncI1-ST3 plasmid was the predominant ESBL-encoding plasmid (76.4%) in wild animal isolates. Plasmid comparison revealed a 110-kb self-transferable plasmid consisting of a conserved backbone and two variable regions involved in antimicrobial resistance and in interaction with recipient cells during conjugation. Conclusion. Our results highlighted the unexpected clonal dissemination of blaCTX-M-1-encoding clones and the complicity of IncI1-ST3 plasmid in the spread of blaCTX-M-1 within wild fauna.

College Student Commitment and Outdoor Orientation Programming

2020 ◽  
Vol 43 (3) ◽  
pp. 299-316
Author(s):  
Curt Davidson ◽  
Alan Ewert
Keyword(s):  
College Student ◽  
Undergraduate Students ◽  
Emotional Reactivity ◽  
Social Activity ◽  
The United States ◽  
College Life ◽  
Self Confidence ◽  
Before And After ◽  
Reactivity Study ◽  
Outdoor Orientation

Background: Increasingly colleges and universities are utilizing Outdoor Orientation Programs (OOPs) to help incoming students assimilate into college life. These programs have shown promise in recent analyses for enhancing desired outcomes with particular consideration shown to pro-social behavior and retention outcomes. Purpose: To examine how effective OOPs are in preparing students for a successful college student experience, particularly with variables known to influence student success and commitment to college. Methodology/Approach: Data were collected from four universities across the United States. Participants in this study were 205 undergraduate students from 17 to 25 years old who self-enrolled in their respective institutions OOP. This study used the College Student Readiness Inventory to generate a hypothesis concerning the possible effects of an OOP experience concerning predictive and outcomes variables relative to college student commitment. Findings/Conclusions: Using SmartPLS, the main effects of the OOP indicated predictive relationships between Commitment to College and Goal Striving, Communication Skills, Social Activity, Emotional Reactivity, Study Skills, and Social Connection. Academic Self-Discipline, Academic Self-Confidence, and Self-Determination on Commitment to College. Implications: Study findings suggest specific connections between predicting college student commitment before and after an OOP.

scholarly journals Trends in Suicidality 1 Year Before and After Birth Among Commercially Insured Childbearing Individuals in the United States, 2006-2017

2020 ◽  
Author(s):  
Lindsay K. Admon ◽  
Vanessa K. Dalton ◽  
Giselle E. Kolenic ◽  
Susan L. Ettner ◽  
Anca Tilea ◽  
...  
Keyword(s):  
United States ◽  
The United States ◽  
Before And After
Sign in / Sign up

Export Citation Format

Share Document