Biology subjects

Prystajecky, N.

Publications and source records attributed to Prystajecky, N..

3 recordsLinked to original sources

bioRxiv2020

Improvements to the ARTIC multiplex PCR method for SARS-CoV-2 genome sequencing using nanopore

Genome sequencing has been widely deployed to study the evolution of SARS-CoV-2 with more than 90,000 genome sequences uploaded to the GISAID database. We published a method for SARS-CoV-2 genome sequencing (https://www.protocols.io/view/ncov-2019-sequencing-protocol-bbmuik6w) online on January 22, 2020. This approach has rapidly become the most popular method for sequencing SARS-CoV-2 due to its simplicity and cost-effectiveness. Here we present improvements to the original protocol: i) an updated primer scheme with 22 additional primers to improve genome coverage, ii) a streamlined library preparation workflow which improves demultiplexing rate for up to 96 samples and reduces hands-on time by several hours and iii) cost savings which bring the reagent cost down to {pound}10 per sample making it practical for individual labs to sequence thousands of SARS-CoV-2 genomes to support national and international genomic epidemiology efforts.

genomics↗

bioRxiv2019

Microbial community dynamics of surface water in British Columbia, Canada

Traditional methods for monitoring the microbiological quality of water focus on the detection of fecal indicator bacteria such as Escherichia coli, often tested as a weekly grab sample. To understand the stability of E.coli concentrations over time, we evaluated three approaches to measuring E. coli levels in water: microbial culture using Colilert, quantitative PCR for uidA and next-generation sequencing of the 16S rRNA gene. Two watersheds, one impacted by agricultural and the other by urban activities, were repeatedly sampled over a simultaneous ten-hour period during each of the four seasons. Based on 16S rRNA gene deep sequencing, each watershed showed different microbial community profiles. The bacterial microbiomes varied with season, but less so within each 10-hour sampling period. Enterobacteriaceae comprised only a small fraction (<1%) of the total community. The qPCR assay detected significantly higher quantities of E. coli compared to the Colilert assay and there was also variability in the Colilert measurements compared to Health Canadas recommendations for recreational water quality. From the 16S data, other bacteria such as Prevotella and Bacteroides showed promise as alternative indicators of fecal contamination. A better understanding of temporal changes in watershed microbiomes will be important in assessing the utility of current biomarkers of fecal contamination, determining the best timing for sample collection, as well as searching for additional microbial indicators of the health of a watershed.

microbiology↗

bioRxiv2017

Antibiotic resistance genes in agriculture and urban influenced watersheds in southwestern British Columbia

BackgroundThe dissemination of antibiotic resistance genes (ARGs) from anthropogenic activities into the environment poses an emerging public health threat. Water constitutes a major vehicle for transport of both biological material and chemical substances. The present study focused on putative antibiotic resistance and integrase genes present in the microbiome of agricultural, urban influenced and protected watersheds in southwestern British Columbia, Canada. A metagenomics approach and high throughput quantitative PCR (HT qPCR) were used to screen for elements of resistance including ARGs and integron-associated integrase genes (intI). Sequencing of bacterial genomic DNA was used to characterize the resistome of microbial communities present in watersheds over a one-year period.\n\nResultsData mining using CARD and Integrall databases enabled the identification of putative antibiotic resistance genes present in watershed samples. Antibiotic resistance genes presence in samples from various watershed locations was low relative to the microbial population (<1 %). Analysis of the metagenomic sequences detected a total of 78 ARGs and intI1 across all watershed locations. The relative abundance and richness of antibiotic resistance genes was found to be highest in agriculture impacted watersheds compared to protected and urban watersheds. Gene copy numbers (GCNs) from a subset of 21 different elements of antibiotic resistance were further estimated using HT qPCR. Most GCNs of ARGs were found to be variable over time. A downstream transport pattern was observed in the impacted watersheds (urban and agricultural) during dry months. Urban and agriculture impacted sites had a higher GCNs of ARGs compared to protected sites. Similar to other reports, this study found a strong association between intI1 and ARGs (e.g., sul1), an association which may be used as a proxy for anthropogenic activities. Chemical analysis of water samples for three major groups of antibiotics was negative. However, the high richness and GCNs of ARGs in impacted sites suggest effects of effluents on microbial communities are occurring even at low concentrations of antimicrobials in the water column.\n\nConclusionAntibiotic resistance and integrase genes in a year-long metagenomic study showed that ARGs were driven mainly by environmental factors from anthropogenized sites in agriculture and urban watersheds. Environmental factors accounted for almost 40% of the variability observed in watershed locations.

microbiology↗