Avian botulism toxicity is a common cause of death to water and shore birds that live near or migrate through Lake Michigan. The botulism neuro-toxin type E (bontE) gene is responsible for the production of botulinum neurotoxin type E. Quantitative Polymerase Chain Reaction (qPCR) was performed using a Step One Plus Thermocycler (Applied Biosystems) and protocol described in Getchell and others, 2011, Journal of Aquatic Animal Health. The assay was used to assess microbial community DNA obtained from environmental samples that were collected by Great Lakes Science Center and by National Park Service from 2011 to 2014 for the bontE gene. Samples were obtained by ponar grab or by divers and matrices collected included sediment, Cladophora, mussels, mussel micro habitat, and invertebrates. This data set is comprised of qPCR data, reported in gene copies per gram wet weight of material for each environmental matrix assessed. Ancillary information specific to sample collection is also included in the data-set, e.g., date, year, site, substrate type, depth, SER, "other description", MIBaRL ID, core depth in centimeter, matrix, collection agency, and copies per gram wet weight as an average reported value.
The terms in the "Remark Code I" column are defined as follows: the term "BDL" is used when the average reported value is below the limit of detection. The term "DNQ" is used when the value is higher than the BDL and lower than the Limit of Quantification (LoQ). The term "--" is used when the value is fully quantifiable (above the LoQ). The terms in the "Remark Code II" column are defined as follows: the symbol < indicates that the result is less than the detection limit. The letter E indicates that the value was estimated because it was higher than the upper range observed for the No Template Control (NTC) samples, but lower than the Limit of Quantification (LoQ). The letter Q indicates that the sample was fully quantifiable. Where no information was given for the sample, the term “Not Applicable”, (N/A) was used.
Quantitative Polymerase Chain Reaction (qPCR) and Trophic Pathways: Sediment samples collected at fixed and random sites during 2011-2014 were analyzed for the presence of the bontE gene (the C. botulinum gene that codes for botulinum type E toxin) using quantitative PCR (qPCR) using methods described in Getchell and others, 2011. Journal of Aquatic Animal Health. Quantitative PCR has been conducted on over 700 environmental samples collected during 2011-2014, including sediments, mussels and the mussel micro habitat, which is the immediate area surrounding mussel beds, attached and sloughed Cladophora, and several types of invertebrates from the waters near Sleeping Bear Dunes National Lakeshore. Not all samples collected are represented in this data table as some samples were used for testing purposes or rendered not tested. This effort has involved challenging sample collection and coordinated environmental and laboratory approaches and is the first such comprehensive analysis using modern gene-based approaches on multiple environmental matrices for this area of the Great Lakes. Preliminary results suggest that vegetative C. botulinum cells are widespread in sediments near SLBE and that seasonality may be a factor in C. botulinum proliferation. In addition, qPCR bontE detections are being evaluated with respect to their association with depth, relation to temperature, and relation to conditions at depth, including the nature of the bottom materials, and whether mats of dead Cladophora are present. When a unique identifier is seen more than one time, it indicates that the sample was run was run undiluted and diluted. The bontE qPCR Assay: The composited standard curve for this project has a slope of -3.546, an intercept of 42.5, an R2 value of .997, and an efficiency of 91%. The Limit of Detection (LoD)and Limit of Quantification (LoQ) were calculated as follows: LoD: Choose Dilution (gc/rxn): 250; Ct0LoD: 34; Std. Dev. (Ct0LoD): 1; Standard Curve eqn. and R2 for Lod: 235. LoQ: Ct 0LoQ: 32; Conc LoQ: Conc LoQ Mobio PowerSoil Kits were used to extract sample DNA. To test for inhibition, a subset of samples was spiked with standard DNA and Ct values were compared and ruled acceptable.