Spatial and Temporal Pattern of Norovirus Dispersal in an Oyster Growing Region in the Northeast Pacific.

Contamination of Pacific oysters, Crassostrea gigas, by human norovirus (HuNoV) is a major constraint to sustainable shellfish farming in coastal waters of the Northeast Pacific. HuNoV is not a marine virus and must originate from a human source. A barrier to effective management is a paucity of data regarding HuNoV dispersal in the marine environment. The main objective of this study was to identify the spatial distribution and persistence of HuNoV in an active shellfish farming region in the Northeast Pacific. Market-size C. gigas were sequentially deployed for two-week intervals at 12 sites during the 2020 winter risk period from January to April. Detection of HuNoV quantification was performed by reverse transcription real-time PCR (RTqPCR) according to method ISO 15216-1:2017, with modifications. RTqPCR did not detect GI HuNoV. The estimated prevalence of GII HuNoV in oyster digestive tissue was 0.8 ± 0.2%. Spatiotemporal analysis revealed that contamination of oysters with GII HuNoV changed through time and space during the surveillance period. A single cluster of oysters contaminated with GII.2 HuNoV was detected in a small craft harbor on 23 April. There was no significant increase in the proportion of positive pools in the next nearest sampling station, indicating that HuNoV is likely to disperse less than 7 km from this non-point source of contamination. Results from this study indicate that HuNoV contamination of coastal waters from non-point sources, such as small craft harbors and urban settings, can pose a significant localised risk to shellfish farming operations in the region.

Measurement of Diacetyl and Related Compounds in Coffee Roasteries and Breweries.

α-Diketones such as diacetyl (2,3-butanedione) and 2,3-pentanedione are generated during the roasting and fermentation of foods and are also used as flavoring compounds. Exposure to these compounds has been associated with obliterative bronchiolitis in workers. We report indoor air concentrations of diacetyl and 2,3-pentanedione, as well as acetoin (3-hydroxy-2-butanone), in several small coffee roasteries and breweries using standard integrated air sampling sorbent tubes followed by gas chromatography tandem mass spectrometry as well as the first use of on-site continuous real-time proton-transfer reaction time-of-flight mass spectrometry (PTR-ToF-MS). Diacetyl and 2,3-pentanedione were detected in most of the sorbent samples at concentrations between 0.02 and 8 ppbv, and in general were higher in coffee roasteries compared with breweries. Three integrated air samples, all from the barista area at one facility, exceeded the NIOSH recommended exposure limit (REL) of 5 ppbv for diacetyl. 2,3-Pentanedione concentrations in these three samples were greater than 50% of its REL, but did not exceed it. Acetoin, a precursor to diacetyl, was also detected at concentrations between 0.03 and 5 ppbv in most sorbent tube samples, with concentrations generally higher in breweries. PTR-ToF-MS measurements exhibited similar trends and provided continuous real-time volatile organic compound data that showed episodic excursions with peak concentrations of diacetyl and 2,3-pentanedione between 15 and 20 ppbv. Examination of the time series data identified specific activities associated with peak diketone emissions, including transfer of freshly roasted coffee beans to the cooling tray, or the opening of a brew kettle. Additional indoor air quality parameters including CO2, NO2, and PM2.5 were also assessed on-site. Airway inflammation was assessed in 19 workers before and after each work shift using online measurements of fractional exhaled nitric oxide (FENO). The pre-shift mean FENO was 3.7 (95% confidence interval: -3.6, 11.0) ppbv higher and the post-shift FENO was 7.1 (-1.9, 16.1) ppbv higher for workers at coffee roasteries compared with breweries. The cross-shift change in FENO was 3.4 (-2.8, 9.6) ppbv higher for workers at coffee roasteries compared with breweries. However, none of these differences were statistically significant, and the cross-shift change in FENO was not statistically different from zero for either group of workers. The findings from this pilot study demonstrate that α-diketones and related compounds are present in the indoor air of both breweries and coffee roasteries and may exceed health protective guidelines in coffee roasteries. Additional studies are required to fully characterize worker exposures in these settings and to identify specific work activities and processes associated with high exposures. Engineering controls, including targeted exhaust ventilation and the use of low-cost sensors, are recommended as an approach to protect workers from exposure to hazardous levels of α-diketones.

Structural Basis of Transcription Inhibition by CBR Hydroxamidines and CBR Pyrazoles.

CBR hydroxamidines are small-molecule inhibitors of bacterial RNA polymerase (RNAP) discovered through high-throughput screening of synthetic-compound libraries. CBR pyrazoles are structurally related RNAP inhibitors discovered through scaffold hopping from CBR hydroxamidines. CBR hydroxamidines and pyrazoles selectively inhibit Gram-negative bacterial RNAP and exhibit selective antibacterial activity against Gram-negative bacteria. Here, we report crystal structures of the prototype CBR hydroxamidine, CBR703, and a CBR pyrazole in complex with E. coli RNAP holoenzyme. In addition, we define the full resistance determinant for CBR703, show that the binding site and resistance determinant for CBR703 do not overlap the binding sites and resistance determinants of other characterized RNAP inhibitors, show that CBR703 exhibits no or minimal cross-resistance with other characterized RNAP inhibitors, and show that co-administration of CBR703 with other RNAP inhibitors results in additive antibacterial activities. The results set the stage for structure-based optimization of CBR inhibitors as antibacterial drugs.