Metagenomics analysis of foods has the potential to provide comprehensive data on the presence and prevalence of antimicrobial resistance (AMR) genes in the microbiome of foods. However, AMR genes ...are generally present in low abundance compared to other bacterial genes in the food microbiome and consequently require multiple rounds of in-depth sequencing for detection. Here, a metagenomics approach, using bait-capture probes targeting antimicrobial resistance and plasmid genes, is used to characterize the resistome and plasmidome of retail beef, chicken, oyster, shrimp, and veal enrichment cultures (
= 15). Compared to total shotgun metagenomics, bait-capture required approximately 40-fold fewer sequence reads to detect twice the number of AMR gene classes, AMR gene families, and plasmid genes across all sample types. For the detection of critically important extended spectrum beta-lactamase (ESBL) genes the bait capture method had a higher overall positivity rate (44%) compared to shotgun metagenomics (26%), and a culture-based method (29%). Overall, the results support the use of bait-capture for the identification of low abundance genes such as AMR genes from food samples.
Sequence-based methods for the detection of bacteria such as 16S rRNA amplicon sequencing and metagenomics can provide a comprehensive view of the bacterial microbiome of food. These methods rely on ...the detection of gene sequences to indicate the presence of viable bacteria. This indirect form of detection can be prone to experimental artefacts. Sample handling and processing are key sources of variation that require standard approaches. Extracting sufficient quantities of high quality DNA from food matrices is challenging because target bacterial species are usually minor components of the microbiota and foods contain an array of compounds that are inhibitory to downstream DNA applications. Here, three DNA extraction methods are compared for their ability to extract high quality bacterial DNA from retail chicken breast rinses, with or without enrichment. Method performance was assessed by comparing ease of use, DNA yield, DNA quality, PCR amplicon yield, and the detection of bacterial taxa by 16S rRNA amplicon sequencing. All three DNA extraction methods yielded DNA of sufficient quantity and quality to perform quantitative PCR and 16S rRNA amplicon sequencing. The extraction methods differed in ease of use, with the two commercial kits (PowerFood, PowerSoil) offering considerable time and cost savings over a hybrid method that used laboratory reagents for lysis and commercial column based kits for further purification. Bacterial richness as determined by 16S rRNA amplicon sequencing was similar across the three DNA extraction methods. However, differences were noted in the relative abundance of bacterial taxa, with significantly higher abundance of Gram-positive genera detected in the DNA samples prepared using the PowerFood DNA extraction kit. The choice of DNA extraction method can affect the detection of bacterial taxa by 16S rRNA amplicon sequencing in chicken meat rinses. Investigators should be aware of this procedural bias and select methods that are fit for the purposes of their investigation.
Between November 2018 and May 2019, Canada experienced a nationwide salmonellosis outbreak linked to the presence of Salmonella enterica ser. Enteritidis in frozen profiteroles. Analysis of the ...implicated food products revealed low levels of Salmonella ranging from 0.2 to 0.7 MPN/100g. Water activity and pH of the food samples ranged from 0.9479 to 0.9867 and 4.6–6.8 respectively indicating conditions conducive to bacterial growth. Higher levels of the hygiene indicators Enterobacteriaceae and coliforms were associated with Salmonella positive samples compared to Salmonella negative samples. Investigation of the relationship between storage conditions, temperature, and pathogen levels during thawing revealed that the profiteroles reached temperatures permissive to pathogen growth (≥5 °C) much sooner than pathogen growth was observed and that the composition of the food matrix can influence bacterial levels upon thawing. Collectively these data can be used to inform guidance to minimize the risk of infection from the consumption of contaminated cream-filled frozen desserts.
•Frozen profiteroles and eclairs were implicated in a national salmonellosis outbreak.•MPN analysis indicated low level contamination of a few individual profiteroles.•Water activity and pH of frozen profiteroles were permissive for Salmonella growth.•Salmonella positive samples had higher levels of Enterobacteriaceae and coliforms.•Improper storage of frozen cream-filled desserts can lead to pathogen growth.
Over the past 15 years, multiple foodborne outbreaks have occurred in Canada due to the presence of Salmonella enterica in frozen breaded chicken products. These chicken products were raw and ...required cooking in conventional household ovens to inactivate any pathogens that they may have been harboring. During the course of food safety investigations associated with these outbreaks, many consumers reported using alternative household appliances such as air fryers to cook these products. The effectiveness of these appliances for the inactivation of pathogens in food is not known. Here, we compare the ability of a toaster oven, air fryer, deep fryer, and conventional oven to inactivate a cocktail of Salmonella Enteritidis in frozen breaded chicken strips. Deep frying was the most effective cooking method, demonstrating a median 7-log reduction; the conventional oven was next with a median 6-log reduction. Both the air fryer and toaster oven performed poorly, with respective median 4- and 3-log reductions. Overall, the results of this study suggest the revision of cooking instructions is required for the safe household use of toaster ovens and air fryers.
Various methods have been described to isolate third generation cephalosporin (3GC) resistant Enterobacteriaceae from foods, but it is not known how comparable they are between studies. Here, the ...performance of five enrichment broths and two selective agars are compared for their ability to isolate 3GC resistant Enterobacteriaceae from retail chicken, beef, pork, and veal samples. The results showed equivalence between Enterobacteriaceae enrichment broth (EE), lauryl sulfate broth (LST), and modified typtone soy broth (mTSB). Lower isolation rates were observed when LST and mTSB were supplemented with the 3GC antibiotic cefotaxime. The overall performance of MacConkey agar supplemented with cefotaxime and a proprietary selective agar (ESBL CHROMagar) was equivalent, although differences linked to the microbiota of specific meat commodities were noted. Regardless of the isolation method, further screening was required to confirm the taxonomy and resistance of the presumptive positive strains. Approximately 40% of confirmed 3GC resistant foodborne Enterobacteriaceae strains tested positive for extended spectrum beta-lactamase (ESBL) activity. Strains that were resistant to ceftriaxone and susceptible to cefoxitin were more likely to test positive for ESBL activity, as were strains that possessed either of two ESBL genes (blaSHV or blaTEM). Based on our results, we recommend using an antibiotic-free enrichment broth, two selective agars, and an isolate screening strategy to isolate 3GC resistant Enterobacteriaceae from retail meats. Antibiotic susceptibility testing and/or PCR screening for blaSHV or blaTEM can then be used to identify ESBL producing strains among the 3GC resistant meat isolates. The adoption of this approach by the research community will enable more effective monitoring of antibiotic resistance rates and trends among foodborne Enterobacteriaceae over time and across jurisdictions.
•Comparison of media to isolate 3GC resistant Enterobaceriaceae from retail meats.•Antibiotic-free enrichment improved recovery of 3GC resistant Enterobaceriaceae.•Cefotaxime and cefoxitin resistance testing can indicate ESBL activity.•Presence of blaTEM and blaSHV was associated with ESBL activity.•Harmonized isolation methods will improve AMR monitoring in foods.
Whole genome sequencing (WGS) is rapidly replacing other molecular techniques for identifying and subtyping bacterial isolates. The resolution or discrimination offered by WGS is significantly higher ...than that offered by other molecular techniques, and WGS readily allows infrequent differences that occur between 2 closely related strains to be found. In this investigation, WGS was used to identify the changes that occurred in the genomes of 13 strains of bacterial foodborne pathogens after 100 serial subcultures. Pure cultures of Shiga-toxin-producing Escherichia coli, Salmonella enterica, Listeria monocytogenes, and Vibrio parahaemolyticus were subcultured daily for 100 successive days. The 1st and 100th subcultures were whole-genome sequenced using short-read sequencing. Single nucleotide polymorphisms (SNPs) were identified between the 1st and final culture using 2 different approaches, and multilocus sequence typing of the whole genome was also performed to detect any changes at the allelic level. The number of observed genomic changes varied by strain, species, and the SNP caller used. This study provides insight into the genomic variation that can be detected using next-generation sequencing and analysis methods after repeated subculturing of 4 important bacterial pathogens.
Waste generation is becoming increasingly prominent in the environmental arena due to the increase in population and living standards of life. Dioxin and Dioxin-related compounds are a set of ...hazardous chemicals that are ubiquitously distributed. Polychlorinated dioxins are introduced into our surroundings by both spontaneous and induced activities like combustion, incineration of waste, recycling of e-waste, and paper and pesticide manufacturing. They are chloroaromatic compounds that are found to be lethal and possess carcinogenic properties and are one of the primary examples of persistent environmental pollutants (POP). Removal of these compounds from the environment is very challenging due to their recalcitrant nature. An alternative technique is the use of microbial technology which includes the use of bacteria and fungi to detoxify the dioxins that are considered to be a more effective, economical, and environmentally sustainable alternative. Different microbial interactions were studied for their degradation potential. Polychlorinated dibenzo-p-dioxin and furans (PCDD/F) are found to be degraded by bacteria by adopting either aerobic or anaerobic pathways and the details regarding the diversity, distribution, bioremediation potential, metabolic pathway have been analyzed. This review provides an overview of the source of contamination, its potential toxicity assessment, and various bioremediation techniques that are employed are discussed in detail. It also highlights the nanoremediation technique - a promising tool in which nanoparticles are used in the treatment of toxic organic pollutants.
How does culture matter for development? Do certain societies have cultures which condemn them to poverty? Led by Arjun Appadurai, Mary Douglas, and Amartya Sen, the anthropologists and economists in ...this volume contend that culture is central to development, and that cultural processes are neither inherently good nor bad and never static. Rather, they are contested and evolving, and can be a source of profound social and economic transformation through their influence on aspirations and collective action; yet they can also be exploitative, exclusionary, and can lead to inequality.Culture and Public Action includes case studies from Africa, Asia, and Latin America, which examine the role of culture in community-based development, ethnic conflict, famine relief, gender discrimination, and HIV-AIDS policy. The editors conclude by proposing how a “cultural lens” can better inform future research and public policy on development. Accessible, balanced, and engaging, this book will be of interest to anyone concerned with the relationship between culture and economics, and the design and implementation of development policy.
Circular RNAs are a class of regulatory RNA transcripts, which are ubiquitously expressed in eukaryotes. In the current study, we evaluate the function of a novel circRNA derived from the β-catenin ...gene locus, circβ-catenin.
Circβ-catenin is predominantly localized in the cytoplasm and displays resistance to RNase-R treatment. We find that circβ-catenin is highly expressed in liver cancer tissues when compared to adjacent normal tissues. Silencing of circβ-catenin significantly suppresses malignant phenotypes in vitro and in vivo, and knockdown of this circRNA reduces the protein level of β-catenin without affecting its mRNA level. We show that circβ-catenin affects a wide spectrum of Wnt pathway-related genes, and furthermore, circβ-catenin produces a novel 370-amino acid β-catenin isoform that uses the start codon as the linear β-catenin mRNA transcript and translation is terminated at a new stop codon created by circularization. We find that this novel isoform can stabilize full-length β-catenin by antagonizing GSK3β-induced β-catenin phosphorylation and degradation, leading to activation of the Wnt pathway.
Our findings illustrate a non-canonical function of circRNA in modulating liver cancer cell growth through the Wnt pathway, which can provide novel mechanistic insights into the underlying mechanisms of hepatocellular carcinoma.
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