In pursuit of pathogens: Using dPCR and sequencing for microbial surveillance
Join us for one or two 40-minute talks. Each presentation will have a 15-minute live Q&A at the end. To start, Dr. Maria A. Bautista will present her findings on using hybrid capture to sequence viral pathogens. Stay with us or join one hour after the start of the session for Dr. Aaron Bivins will compare dPCR systems for microbial tracking.
Talk 1: Unlocking viral diversity in wastewater: Using hybrid capture for enhanced human viral pathogen detection
Dr. Maria A. Bautista, University of Calgary
Time: 1 p.m. EDT/10 a.m. PDT
Duration: 1 hour
Wastewater surveillance is a promising tool for detecting and tracking viral pathogens in municipal wastewater, providing valuable insights into population health and disease risk. Despite this potential, genomic sequencing of pathogens is challenging due to their relatively low abundance and the high background of nucleic acid from humans, bacteria and other viruses. We have successfully used hybrid-capture panels to enhance the recovery of viral pathogenic sequences from wastewater. Here, I discuss increasing the output and detection of viruses of interest from wastewater using QIAGEN QIAseq xHYB hybrid capture panels in combination with ribosomal RNA removal.
Talk 2: Comparing dPCR systems for microbial source tracking in surface waters
Dr. Aaron Bivins, Louisiana State University
Time: 2 p.m. EDT/11 a.m. PDT
Duration: 1 hour
Wastewater surveillance has accelerated the adoption of digital PCR (dPCR) in various formats for quantifying molecular targets. Public health-related water microbiology requires analytical sensitivity, resilience to inhibition, and precise quantification, which makes dPCR an especially promising tool. Nonetheless, most standardized molecular methods for assessing water quality employ quantitative PCR (qPCR) for molecular measurements. We adapted qPCR assays from three U.S. EPA Methods (1696: HF183/BacR287; 1609.1 Entero1a; Draft Method C: EC23S857) to dPCR format and compared their performance for detecting and quantifying targets in positive controls (SRM 2917) and environmental samples (n=10) on two different dPCR platforms (QIAcuity [QIAGEN], Absolute Q [Themo Fischer Scientific]). We analyzed filter triplicates (x3) and dPCR technical replicates (x2) on both platforms for all environmental samples. The analytical sensitivity varied slightly between the platforms for each assay. Intra-class correlation coefficients (ICC) indicate excellent quantitative agreement (ICC > 0.96) between the QIAcuity and Absolute Q across positive control materials for all three assays. Overall, we found the QIAcuity and Absolute Q platforms yielded reasonably similar qualitative and quantitative results. However, we found there were important operational characteristics to consider when selecting a platform for routine water quality monitoring.
Talk 1: Unlocking viral diversity in wastewater: Using hybrid capture for enhanced human viral pathogen detection
Dr. Maria A. Bautista, University of Calgary
Time: 1 p.m. EDT/10 a.m. PDT
Duration: 1 hour
Wastewater surveillance is a promising tool for detecting and tracking viral pathogens in municipal wastewater, providing valuable insights into population health and disease risk. Despite this potential, genomic sequencing of pathogens is challenging due to their relatively low abundance and the high background of nucleic acid from humans, bacteria and other viruses. We have successfully used hybrid-capture panels to enhance the recovery of viral pathogenic sequences from wastewater. Here, I discuss increasing the output and detection of viruses of interest from wastewater using QIAGEN QIAseq xHYB hybrid capture panels in combination with ribosomal RNA removal.
Talk 2: Comparing dPCR systems for microbial source tracking in surface waters
Dr. Aaron Bivins, Louisiana State University
Time: 2 p.m. EDT/11 a.m. PDT
Duration: 1 hour
Wastewater surveillance has accelerated the adoption of digital PCR (dPCR) in various formats for quantifying molecular targets. Public health-related water microbiology requires analytical sensitivity, resilience to inhibition, and precise quantification, which makes dPCR an especially promising tool. Nonetheless, most standardized molecular methods for assessing water quality employ quantitative PCR (qPCR) for molecular measurements. We adapted qPCR assays from three U.S. EPA Methods (1696: HF183/BacR287; 1609.1 Entero1a; Draft Method C: EC23S857) to dPCR format and compared their performance for detecting and quantifying targets in positive controls (SRM 2917) and environmental samples (n=10) on two different dPCR platforms (QIAcuity [QIAGEN], Absolute Q [Themo Fischer Scientific]). We analyzed filter triplicates (x3) and dPCR technical replicates (x2) on both platforms for all environmental samples. The analytical sensitivity varied slightly between the platforms for each assay. Intra-class correlation coefficients (ICC) indicate excellent quantitative agreement (ICC > 0.96) between the QIAcuity and Absolute Q across positive control materials for all three assays. Overall, we found the QIAcuity and Absolute Q platforms yielded reasonably similar qualitative and quantitative results. However, we found there were important operational characteristics to consider when selecting a platform for routine water quality monitoring.
About the speaker
Dr. Maria A. Bautista, Senior Research Associate - Geomicrobiology
University of Calgary
Dr. Maria A. Bautista is an environmental microbiologist and virologist specializing in molecular methods for studying diverse microbial communities. She focuses on assessing microbial and viral diversity, community changes following disturbances and temporal/spatial comparisons.In her current role as a Senior Research Associate at the Geomicrobiology group at the University of Calgary, Maria serves as one of the technical leads for the Pan-Alberta wastewater monitoring program. Since 2020, she has been dedicated to testing, developing and applying methods for extracting, quantifying and sequencing viruses from municipal wastewater.Maria completed her PhD in Microbiology at the University of Illinois at Urbana-Champaign, where she studied host-virus interactions in hyperthermophilic archaea. Following her PhD she worked at the DNA Sequencing Laboratory, W.M. Keck Center for Comparative and Functional Genomics at the University of Illinois at Urbana-Champaign, where she further developed her interest in method development and optimization to generate high-quality sequencing output
Dr. Aaron Bivins, Assistant Professor
Louisiana State University
Dr. Aaron Bivins is a public health engineer leading research to mitigate infectious diseases via design and policy. His work is situated at the intersection of environmental microbiology, engineering and human health. He and his team use environmental microbiology techniques to observe microorganisms in built and natural environments. They use these observations along with quantitative techniques to characterize health risks associated with human exposures mediated by the environment. When integrated with design and policy decision-making, these techniques can be used to minimize the burden of infectious diseases on human health.In conjunction with his research, Dr. Bivins has spent significant time working across cultures, including projects in India (2017 Fulbright-Nehru Scholar), Mozambique, Nicaragua, Bolivia, and Jamaica. His research interests are informed by his professional practice designing various hydraulic infrastructures, such as water distribution, wastewater collection and stormwater systems. Dr. Bivins completed his PhD and Master’s degree in environmental engineering at the Georgia Institute of Technology, USA. Prior to graduate school, he worked as a consulting civil engineer in Savannah, GA, and is a licensed Professional Engineer and Board Certified Environmental Engineer. Dr. Bivins is currently an Assistant Professor of civil and environmental engineering at Louisiana State University in Baton Rouge, Louisiana.
Categories
Microbiology
Microbiome
dPCR
Next Generation Sequencing