FILE INVENTORY Dataset citation: (dataset) Van Winkle, Taylor; Ponce, Marco A.; Quellhorst, Hannah E.; Bruce, Alexander; Albin, Chloe E.; Kim, Tania N.; Zhu, Kun Yan; Morrison, William R. (2021). Data from: Microbial volatile organic compounds mediate attraction by a primary but not secondary stored product insect pest in wheat. Ag Data Commons. https://doi.org/10.15482/USDA.ADC/1520888. /////// Title: GC-MS/Headspace Data File name: tvw_final_gc_ms_data.csv Description: Volatile Collection Assay. Headspace collection was performed in order to determine the volatiles released by the various moisture treatments (no grain as a negative control, or 20 g of untempered grain control, or grain tempered to 12, 15, or 19%) and incubated at different intervals (9, 18, 27 d). First, central air was scrubbed using an activated charcoal filter, then pushed through the remaining apparatus. The airflow was restricted to 1 L/min using a flow meter (Volatile Collection Systems, Gainesville, FL, USA) placed directly prior to the sample collection in headspace chambers (10.2 × 12.7 cm D:H) with an inlet for air and an outlet for a volatile collection trap (VCT). The headspace volatiles from the grain were collected for 3 h onto a VCT consisting of a drip tip borosilicate glass tube packed with 20 mg of absorbent Porapak-Q™ (Volatile Collection Systems, Gainesville, FL, USA) to adsorb volatiles with a stainless steel screen (No. 316) on one side, and held in place with a borosilicate glass wool plug followed by a PTFE Teflon compression seal. The volatiles on the traps were eluted with 300 µl of dichloromethane (Millipore, Billerica, MA, USA) by pushing the solvent through with inert N2 gas into 2-mL glass vials with caps containing Teflon-lined septa and then stored at -20C until GC-MS analysis (Agilent Technologies, Inc., Santa Clara, CA, USA). Volatile collection traps were reused after they were washed three times with 700 µl of dichloromethane. Subsequently, to quantify the samples, 1 µl of tetradecane (190.5 ng, 99% purity, GC analytical grade, Millipore, Billerica, MA, USA) was added as an internal standard using a microsyringe (2 ul capacity syringe, Hamilton Co., Reno, NV, USA). A total of n = 4–6 replicate collections were done for each combination of moisture and incubation treatment. Gas Chromatography Coupled with Mass Spectrometry. All headspace collection sample extracts were run on an Agilent 7890B gas chromatograph (GC) equipped with an Agilent Durabond HP-5 column (30 m length, 0.250 mm diameter and 0.25 um film thickness) with He as the carrier gas at a constant 1.2 mL/min flow and 40 cm/s velocity, which was coupled with a single-quadrupole Agilent 5997B mass spectrometer (MS). The compounds were separated by auto-injecting 1 ul of each sample under split mode into the GC-MS at room temperature (approximately 23C). The flow was split in a 15:1 ratio with a split flow rate of 18 ml/min. The program consisted of 40C for 1 min followed by 10C/min increases to 300C and then held for 26.5 min. After a solvent delay of 3 min, mass ranges between 50 and 550 atomic mass units were scanned. Compounds were tentatively identified by comparison of spectral data with those from the NIST 17 library and by GC retention index (Adams 2009). Using the ratio of the peak area for the internal standard to the peak area for the other compounds in the headspace, the emission rates of samples in ng of volatile per g of grain, per μl of solvent, and per h of collection were calculated. /////// Title: Microbial damage on wheat evaluated with near-infrared spectroscopy File name: tvw_nearinfrared_sorting_damaged_grain_fungal_exp.csv Description: Near-Infrared Spectroscopy of Grain. A multispectral sorting device was used to assess the proportion of grain kernels in each treatment that were infested with fungi and other microbes, which manifested in color or reflectance changes (Pearson et al. 2013). Prior work has found that this sorter can correctly identify 90% of grain kernels damaged with the fungal disease, Fusarium head blight, from undamaged kernels (Pearson et al. 2013). The sorter used three visible and three near-infrared light-emitting diodes with peak emission wavelengths of 470, 527, 624, 850, 940, and 1070 nm by rapidly blinking each LED (~12 kHz) in quick succession, and measuring the reflected light from the wheat kernels using a miniature 16 mm lens (V-4316-2.0, Marshall Electronics, Inc., CA, USA) as kernels dropped off a feeder chute at a 45˚ angle with a speed of 1.5 m/s. The device employed a microcontroller (ATmega328P, Atmel Corp., San Jose, CA) that directs the LED pulses, digitizes the analog signal from a photodiode (PDB-C171SM, Advanced Photonix, Inc., Ann Arbor, MI, USA), performs signal processing, and applies classifications. The device sorted grain at 20 kernels/s through the activation of an air valve that diverted the wheat kernels based on a prior calibration. Calibration of the machine occurred by training the machine to recognize damaged grain by using 200 fungal-damaged kernels and 200 undamaged kernels. Prior to data collection, we evaluated the accuracy the sorter by calibrating the device based on training with the 12, 15, and 19% grain moisture incubated at 27 d, or for maximally infested grain (as described above) in comparison to clean grain held at 10.8% moisture in each case. The 19% grain moisture calibration was used for data collection based on its high accuracy (see results below), to obtain a conservative estimate of microbial damage, and to ensure consistency with incubation methodology compared to all other treatments. To evaluate damage, 100 g of grain, approximately 1,900 individual kernels, was sorted for each combination of grain moisture and incubation period, including the undamaged grain (control). /////// Title: Release-Recapture Datasets with LGB & RFB File name: tvw_rr_lgb_rfb_microbial_cues.csv Description: Release-Recapture Trapping Assay. To assess how fungal volatiles may improve capture of T. castaneum and R. dominica adults in monitoring traps, a trapping assay was utilized (Fig. 1B). In each replicate, 20 mixed-sex adults were settled on clean cardboard slats (8 × 8 cm) and then placed in one corner of a plastic bin (86.3 × 30.5 × 39.4 cm L:H:W). In the opposite corner, diagonally across from the release point in the bin, a pitfall trap (Dome Trap™, Trécé, Inc., Adair, OK, USA; Campbell 2012) was deployed. The base was rough along the ascending edges to create traction for the beetles; however, the crest was concave and smooth to prevent subsequent escapes. Semiochemical stimuli were placed within the concave portion of the trap. Each trap contained either no grain (negative control), or 5 g of one of the treatments: untempered grain (positive control), grain tempered to either 12%, 15%, or 19% grain moisture, or 950 µl of Trécé- produced StorgardTM Oil (a known semiochemical kairomone and food cue, which is a standard lure used in commercial facilities [Campbell 2012], and acted as the positive control; Trécé, Inc., Adair, OK, USA). All grain in this study was either incubated for 9, 18, or 27 d, and trapping assays were initiated on the same days as the wind tunnel assays described above. The bins were located in a large (5 × 6 × 2 m, L:W:H) walk-in environmental chamber (Percival Instruments, Dallas County, IA, USA) under constant conditions (25°C, 65% RH, 14:10 L:D). The adults were given 24 h to respond to the stimuli. Afterwards, the traps were collected, the adults were sieved (No. 10, W.S Tyler Inc., Mentor, OH), and the number of beetles in each trap was recorded. On a given round of release, two replicates of each treatment listed above were performed simultaneously, for a total of 12 bins set up containing 240 adults total. In total, there were n = 8 replicates per species and combination of treatment and incubation, including the controls. /////// Title: Wind tunnel response by RGB & LGB File name: tvw_wt_lgb_rfb_data_microbial_cues.csv Description: Wind Tunnel Assay. A wind tunnel assay was used to better understand the taxis and attraction of T. castaneum and R. dominica to fungal volatiles from the grain. In the wind tunnel, air was pushed through a turbine (diameter: 36.5 cm), an activated carbon filter to purify the air, and two slatted-metal sieves (73  85 cm) to create a laminar airflow, with an average airspeed of 0.38 m/s. A total of 20 g of each odor treatment described above was placed 13.5 cm upwind of the stimulus edge of the test arena and 5 cm from the last sieve. Each odor treatment was held in a plastic petri dish (100 × 15 mm D:H). The adults were placed individually in the release point at the center of an arena consisting of a 21.6 × 27.9 cm sheet of paper. In each round of sampling, mixed-sex adult beetles were collected from their colonies using a sieve (No. 25, W.S Tyler Inc., Mentor, Ohio). Each adult was given a maximum of 2 min to respond, and the edge of the arena on which the adult exited was recorded as either the stimulus edge (upwind edge of the arena), or a non-stimulus edge (one of the other three edges) (Fig. 1A). In addition, the time to decision was recorded for each individual. Adults that did not respond within the timeframe were excluded from the final statistical analysis. Each adult was considered a replicate and was never used more than once. Odor flow from the petri dish to the release arena was confirmed with a smoke test. In total, there were n = 60 replicates per each species and treatment combination of grain moisture and incubation interval. /////// Title: GC/MS Grain MVOC Headspace Data File name: taylor_headspace_final_data_peer_reviewed_ag_commons.csv Description: These data were collected from grain that had been tempered and then incubated to different moistures and periods. Changes in volatiles represent activity of microbial activity. Compounds were tentatively identified using the NIST17 library, but this information was not included, because it was not confirmed and tangential to the main goal of the study. ///////