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Download BST_results120612 - Huron River Watershed Council
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Results Information 28044 James Dr. Warren, MI 48092 Phone: 586-806-4243 Fax: 313- 577-8616 Email: [email protected] Customer ID: Huron River Watershed Project #: P-173B Sample Date Received: 6/12/12 Date of Analysis: 6/14/12 – 6/19/12 Sample Type: Water Analysis Type: MST Procedure: Submitted water samples from Honey Creek were tested for the presence of source specific Bacteroides bacteria using target DNA markers. Samples received were from June 5, 2012 collection. Submitted water samples had the following sample designation: HC1 HC4 HC7 HC9 HC12 HC14 HC15 HC17 Samples HC1, HC4, HC7, HC9, HC12, HC15 and HC17 were analyzed using MST 100 ml of each water sample tested was suctioned filtered through a 0.45 micron filter membrane that proceeded to capture any bacteria present in the water samples. Each individual filter membrane containing captured bacteria was removed from each individual filter column following filtration and used to grow a pre-enrichment bacterial culture on media selective for Bacteroides at 37 degrees C for 72 hours. After incubation, bacterial DNA was extracted from the filter membranes used in the pre-enrichment cultures. Bacterial DNA extractions were then used as DNA template for qualitative PCR analysis. PCR analysis incorporated oligonucleotide primers that target specific marker DNA sequences unique to source specific Bacteroides bacteria. Appropriate positive and negative control samples were included in PCR experimentation. Following the PCR procedure, the PCR products were run on a 1.5% agarose gel at 90V for 60 minutes, after which the agarose gel containing the PCR products was visualized under UV light and examined for PCR amplification products that represent the amplification of the target DNA markers. Replicates of each sample were run for quality assurance purposes and to confirm the observed results. Results Results of the qualitative PCR testing indicate the presence (positive) or absence (negative) of source specific DNA markers through PCR amplification of source specific DNA marker sequences. A sample testing positive for a source specific DNA marker indicates that the marker DNA sequence representing the specific bacteria was amplified, thus confirming the absence of that source specific DNA marker in the sample. Sample Source DNA Marker Result HC1 HC1 HC1 HC1 HC1 HC1 General Bacteroides Bovine Canine Goose Equine Human + + + + - HC4 HC4 HC4 HC4 HC4 HC4 General Bacteroides Bovine Canine Goose Equine Human + + + + + + HC7 HC7 HC7 HC7 HC7 HC7 General Bacteroides Bovine Canine Goose Equine Human + + + + - HC9 HC9 HC9 HC9 HC9 HC9 General Bacteroides Bovine Canine Goose Equine Human + + + + + - HC12 HC12 HC12 HC12 HC12 HC12 General Bacteroides Bovine Canine Goose Equine Human + + + + + HC15 HC15 HC15 HC15 HC15 HC15 General Bacteroides Bovine Canine Goose Equine Human + + + + HC17 HC17 HC17 HC17 HC17 HC17 General Bacteroides Bovine Canine Goose Equine Human + + + + A positive result for a source specific DNA marker at a collection station implies that source is contributing to fecal contamination that may have been determined at the collection station during the collection period. A negative result for a source specific DNA marker at a collection station implies that source is not contributing to fecal contamination that may have been determined at the at the collection station during the collection period. These results were verified by Tom Prychitko, Laboratory Director for Helix Biological Laboratory.
