Bacterial Source Tracking How One Small Environmental Lab
Bacterial Source Tracking How One Small Environmental Lab Developed an Impactful Molecular Biology Program Overall Goal of Bacterial Source Tracking Improve the water quality of the San Antonio River basin Identify the source of the bacteria in order to decrease the levels in the basin to the TCEQ /EPA recreational levels Reduce outsourcing of samples Added revenue stream for the lab 2 3
What is BST? Bacterial source tracking (BST) is a set of techniques used to determine the sources of fecal indicator bacteria in the environment. BST techniques attempt to determine whether fecal bacteria are being introduced into water bodies through human, wildlife, or domestic animal sources. 4
What is PCR? 5 SARAs Method Filtration DNA extraction qPCR Data analysis 6 Filtration Collection similar to MPN Hold time 8 hours Filter each sample
Analysis stopping point # 1 7 DNA Extraction Addition of lysis buffer to lyse cells Series of washes through a column Analysis stopping point # 2 8 qPCR Analysis Set up appropriate run on the instrument
Prep samples for analysis Load wells according to your run Set it and Forget it 45min 9 Designate a clean room Logistics Research and purchase equipment Lab set up Implementation of analysis
10 Clean Room Specific to BST analysis 3 separated rooms or areas Unidirectional workflow Designated supplies 11 Research and Purchase Equipment
Low, Medium, or High throughput? Industry standards Consider the space youre working with 12 Lab Set Up Install equipment
Order consumables Calibrate equipment 13 Implementation of Analysis QA/QC Measures DOCs Validation of paperwork Calibration & standardization SOPs and Bench sheets Training of analysts
Financials fee schedule 14 Training Life Technologies Principals of Quantitative PCR and Data Analysis 15 Lessons Learned Unusual equipment unusually long and or difficult delivery Always be on the lookout for training and conferences Helps to have a team member
who is familiar with molecular biology No standardized method be ready to improvise, Very detailed protocol 16 Resource allocation staff planning, research and development Expect the unexpected staff changes, projects, sample loads Networking with other labs Current Testing
SARA LAB E. coli OUTSOURCE LAB MPN/100 mL Human BST Human Bird Dog
Ruminant SAR @ Myrtle 2/9/2016 14:39 46 Present NA NA NA
NA SAR @ Myrtle 4/5/2016 14:39 55 Absent Absent Present (Trace)
Absent Not Detected SAR @ Pecan 2/9/2016 14:49 210 Present Present Not
Detected Not Detected Not Detected SAR @ Pecan 4/5/2016 14:20 230 Present
Present Present (Trace) Present (Trace) Not Detected SAMPLE 17
SAR Draining Results Sample # Sample Site MPN value (MPN/100mL) Present/Absent Human DNA AA99318-A Seep at Little Rhein
Results Add algorithm to Mining Environment Earth Science Data Sets Relationship analysis Coincidence searches Input for other algorithms Advanced Microwave Sounding Unit (AMSU-A)Data Calibration/ Limb Correction/ Converted to Tb Mining Environment Data Archive Result Results are placed on the web...
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