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Tuesday, April 21, 2020

Genome sequence analysis of 91 Salmonella Enteritidis isolates from mice caught on poultry farms in the mid 1990s.

A total of 91 draft genome sequences were used to analyze isolates of Salmonella enterica serovar Enteritidis obtained from feral mice caught on poultry farms in Pennsylvania. One objective was to find mutations disrupting open reading frames (ORFs) and another was to determine if ORF-disruptive mutations were present in isolates obtained from other sources. A total of 83 mice were obtained between 1995-1998. Isolates separated into two genomic clades and 12 subgroups due to 742 mutations. Nineteen ORF-disruptive mutations were found, and in addition, bigA had exceptional heterogeneity requiring additional evaluation. The TRAMS algorithm detected only 6 ORF disruptions. The…

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Monday, March 30, 2020

AGBT Conference: Automated de novo genome assemblies and bacterial epigenomes using PacBio sequencing

In this AGBT plenary talk, Jonas Korlach presented a number of collaborative studies between PacBio and other institutions to make use of highly accurate, long-read sequence data, which has led to a revival of finished genomes. Examples from the infectious disease or pathogen realm included Pertussis, Salmonella, and Listeria, all of which now have closed genomes from PacBio-generated data. Korlach also reported on epigenomic information in Salmonella and Listeria, indicating potential new forms of DNA modifications.

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Monday, March 30, 2020

ASM PacBio Workshop: Genomics in food security – 100k pathogen genome project

UC Davis’s Bart Weimer describes foodborne pathogens and their proclivity for rapid genome rearrangement. The 100K Pathogen Genome Project he leads is using PacBio long-read sequencing to close genomes and analyze methylation; Weimer reports that his team has already discovered new epigenetic modifications in Salmonella and Listeria with the technology.

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Monday, March 30, 2020

AGBT Conference: Automated, non-hybrid de novo genome assemblies and epigenomes of bacterial pathogens

Jonas Korlach, CSO of PacBio, discusses the revival of finished genomes the microbial community will see with long read data, emphasizing that for certain organisms such as rapidly evolving microbes, having a de novo finished genome will be more useful than creating a draft based on a previous related reference genome. Korlach describes two bioinformatic methods from PacBio, a hierarchical genome assembly process (HGAP) and an consensus caller (Quiver), which are used to generate finished genomes from just long-read PacBio data, with final genome sequence accuracies over 99.999%. Korlach demonstrates the ability of PacBio data to generate closed, high-quality de…

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Monday, March 30, 2020

AGBT Virtual Poster: Interspecies interation amoung meat spoilage-related lactic acid bacteria

In this AGBT 2017 poster, the University of Helsinki’s Petri Auevinen reports on efforts to understand bacteria that grow on, and subsequently spoil, food. This analysis monitored DNA modifications and transcriptomic changes in three species of lactic acid bacteria. Scientists discovered that the organisms’ metabolic profiles change substantially when grown together compared to those cultured individually, and are now studying how Cas protein activity changes under these conditions too.

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Wednesday, February 26, 2020

Comparative genomics of Shiga toxin-producing Escherichia coli O145:H28 strains associated with the 2007 Belgium and 2010 US outbreaks.

Shiga toxin-producing Escherichia coli (STEC) is an emerging pathogen. Recently there has been a global in the number of outbreaks caused by non-O157 STECs, typically involving six serogroups O26, O45, 0103, 0111, and 0145. STEC O145:H28 has been associated with severe human disease including hemolytic-uremic syndrome (HUS), and is demonstrated by the 2007 Belgian ice-cream-associated outbreak and 2010 US lettuce-associated outbreak, with over 10% of patients developing HUS in each. The goal of this work was to do comparative genomics of strains, clinical and environmental, to investigate genome diversity and virulence evolution of this important foodborne pathogen.

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Wednesday, February 26, 2020

New discoveries from closing Salmonella genomes using Pacific Biosciences continuous long reads.

The newer hierarchical genome assembly process (HGAP) performs de novo assembly using data from a single PacBio long insert library. To assess the benefits of this method, DNA from several Salmonella enterica serovars was isolated from a pure culture. Genome sequencing was performed using Pacific Biosciences RS sequencing technology. The HGAP process enabled us to close sixteen Salmonella subsp. enterica genomes and their associated mobile elements: The ten serotypes include: Salmonella enterica subsp. enterica serovar Enteritidis (S. Enteritidis) S. Bareilly, S. Heidelberg, S. Cubana, S. Javiana and S. Typhimurium, S. Newport, S. Montevideo, S. Agona, and S. Tennessee. In addition,…

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Sunday, September 22, 2019

Using PacBio long-read high-throughput microbial gene amplicon sequencing to evaluate infant formula safety.

Infant formula (IF) requires a strict microbiological standard because of the high vulnerability of infants to foodborne diseases. The current study used the PacBio single molecule real-time (SMRT) sequencing platform to generate full-length 16S rRNA-based bacterial microbiota profiles of thirty Chinese domestic and imported IF samples. A total of 600 species were identified, dominated by Streptococcus thermophilus, Lactococcus lactis and Lactococcus piscium. Distinctive bacterial profiles were observed between the two sample groups, as confirmed with both principal coordinate analysis and multivariate analysis of variance. Moreover, the product whey protein nitrogen index (WPNI), representing the degree of preheating, negatively correlated with…

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Sunday, September 22, 2019

Evaluation of bacterial contamination in raw milk, ultra-high temperature milk and infant formula using single molecule, real-time sequencing technology.

The Pacific Biosciences (Menlo Park, CA) single molecule, real-time sequencing technology (SMRT) was reported to have some advantages in analyzing the bacterial profile of environmental samples. In this study, the presence of bacterial contaminants in raw milk, UHT milk, and infant formula was determined by SMRT sequencing of the full length 16S rRNA gene. The bacterial profiles obtained at different taxonomic levels revealed clear differences in bacterial community structure across the 16 analyzed dairy samples. No indicative pathogenic bacteria were found in any of these tested samples. However, some of the detected bacterial species (e.g., Bacillus cereus, Enterococcus casseliflavus, and…

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Sunday, September 22, 2019

Diverse antibiotic resistance genes in dairy cow manure.

Application of manure from antibiotic-treated animals to crops facilitates the dissemination of antibiotic resistance determinants into the environment. However, our knowledge of the identity, diversity, and patterns of distribution of these antibiotic resistance determinants remains limited. We used a new combination of methods to examine the resistome of dairy cow manure, a common soil amendment. Metagenomic libraries constructed with DNA extracted from manure were screened for resistance to beta-lactams, phenicols, aminoglycosides, and tetracyclines. Functional screening of fosmid and small-insert libraries identified 80 different antibiotic resistance genes whose deduced protein sequences were on average 50 to 60% identical to sequences deposited…

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Sunday, September 22, 2019

Early transmissible ampicillin resistance in zoonotic Salmonella enterica serotype Typhimurium in the late 1950s: a retrospective, whole-genome sequencing study.

Ampicillin, the first semi-synthetic penicillin active against Enterobacteriaceae, was released onto the market in 1961. The first outbreaks of disease caused by ampicillin-resistant strains of Salmonella enterica serotype Typhimurium were identified in the UK in 1962 and 1964. We aimed to date the emergence of this resistance in historical isolates of S enterica serotype Typhimurium.In this retrospective, whole-genome sequencing study, we analysed 288 S enterica serotype Typhimurium isolates collected between 1911 and 1969 from 31 countries on four continents and from various sources including human beings, animals, feed, and food. All isolates were tested for antimicrobial drug susceptibility with the…

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Sunday, September 22, 2019

Emergence of an extensively drug-resistant Salmonella enterica serovar Typhi clone harboring a promiscuous plasmid encoding resistance to fluoroquinolones and third-generation cephalosporins.

Antibiotic resistance is a major problem in Salmonella enterica serovar Typhi, the causative agent of typhoid. Multidrug-resistant (MDR) isolates are prevalent in parts of Asia and Africa and are often associated with the dominant H58 haplotype. Reduced susceptibility to fluoroquinolones is also widespread, and sporadic cases of resistance to third-generation cephalosporins or azithromycin have also been reported. Here, we report the first large-scale emergence and spread of a novel S. Typhi clone harboring resistance to three first-line drugs (chloramphenicol, ampicillin, and trimethoprim-sulfamethoxazole) as well as fluoroquinolones and third-generation cephalosporins in Sindh, Pakistan, which we classify as extensively drug resistant (XDR).…

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Sunday, September 22, 2019

A validation approach of an end-to-end whole genome sequencing workflow for source tracking of Listeria monocytogenes and Salmonella enterica.

Whole genome sequencing (WGS), using high throughput sequencing technology, reveals the complete sequence of the bacterial genome in a few days. WGS is increasingly being used for source tracking, pathogen surveillance and outbreak investigation due to its high discriminatory power. In the food industry, WGS used for source tracking is beneficial to support contamination investigations. Despite its increased use, no standards or guidelines are available today for the use of WGS in outbreak and/or trace-back investigations. Here we present a validation of our complete (end-to-end) WGS workflow for Listeria monocytogenes and Salmonella enterica including: subculture of isolates, DNA extraction, sequencing…

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Sunday, September 22, 2019

Complete genomic analysis of a Salmonella enterica Serovar Typhimurium isolate cultured from ready-to-eat pork in China carrying one large plasmid containing mcr-1.

One mcr-1-carrying ST34-type Salmonella Typhimurium WW012 was cultured from 3,200 ready-to-eat (RTE) pork samples in 2014 in China. Broth dilution method was applied to obtain the antimicrobial susceptibility of Salmonella Typhimurium WW012. Broth matting assays were carried out to detect transferability of this phenotype and whole-genome sequencing was performed to analyze its genomic characteristic. Thirty out of 3,200 RTE samples were positive for Salmonella and the three most frequent serotypes were identified as S. Derby (n = 8), S. Typhimurium (n = 6), and S. Enteritidis (n = 6). One S. Typhimurium isolate (S. Typhimurium WW012) cultured from RTE prepared…

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