Metagenomic Profiling of Settled Dust

Settled surface dust was collected from two different swine confinement facilities (housing 400–600 hogs), the storage facilities at two different grain elevators, and two different pet-free domestic homes as a control. The dust samples were obtained from surfaces approximately 3–5 feet above the floor to ensure the sampled dust had been airborne and potentially inhaled by a worker. Permission was granted by the owners of the swine confinement facilities, grain elevator facilities, and households to obtain samples in an anonymous manner. Total genomic DNA was isolated by bead beating following the manufacturer’s instructions (Mo Bio, PowerSoil Kit, Carlsbad, CA), then assayed using a Nanodrop ND-1000 UV spectrophotometer (NanoDrop Technologies, Wilmington, DE). Each DNA sample (3–5 µg) was used to prepare a shotgun pyrosequencing library using a kit for this purpose (Roche Applied Science, Indianapolis, IN) according to the manufacturer’s protocol. The multiple id, barcoded template DNAs were combined in equal amounts and titrated to obtain the optimal copies per bead (3 copies per bead). Emulsion PCR and pyrosequencing were performed with the Roche/454 Life Sciences’ Lib-L (LV) and XLR70 kits, respectively. Multiplexed shotgun metagenomic DNA pyrosequencing was performed by the Core for Applied Genomics and Ecology Laboratory at the University of Nebraska, Lincoln, using a Roche/454 Life Sciences’ GS FLX Titanium instrument (Branford, CT). GS FLX Off-Instrument Software was used to de-multiplex raw pyrosequencing reads into sample-specific bins. These six shotgun pyrosequencing metagenomic read datasets are publicly available at MG-RAST [DNAdustGrainTLV2011s (4465551.3), DNAdustGrainTLV2011e (4465547.3), DNAdustSwineTLV2011f (4465549.3), DNAdustSwineTLV2011n (4465550.3), DNAdustHouseTLV2011n (4465546.3), DNAdustHouseTLV2011p (4465548.3)].