Evaluation of land use and water quality in an agricultural watershed in the USA indicates multiple sources of bacterial impairment.

Pathogens are the number one cause of impairments of assessed rivers and streams in the USA and pose a significant human health hazard. The Dry Run Creek Watershed in Northeast Iowa has been designated as impaired by the State of Iowa because of high levels of Escherichia coli bacteria. To investigate the nature of this impairment, land use and stream bank assessments were coupled with comprehensive water quality monitoring. Physical, chemical, and biological parameters were measured at 13 different sites in the watershed, including pH, temperature, conductivity, dissolved oxygen, turbidity, total Kjeldahl nitrogen, ammonia-N, nitrate + nitrite-N, total phosphorus, and E. coli. In addition, benthic macroinvertebrate communities were analyzed at seven sites, and optical brightener tests were performed late in the season. Results identified segments of the watershed that were more prominent contributors of E. coli, and correlations were observed between levels of E. coli and several chemical parameters, including ammonia-N, total Kjeldahl nitrogen, and total phosphorus. Interestingly, distinct sites emerged as more prominent contributors of these elements during rain vs. non-rain events, suggesting different types of sources. Both the amount of rainfall and the time elapsed between the rain event and the sampling influenced E. coli levels during wet weather conditions. Nitrate + nitrite-N displayed a unique response to rain events compared with the other parameters, suggesting a different delivery route. Analyses of benthic macroinvertebrate communities were consistent with pollution trends. Collectively, these data suggest distinct agriculturally related E. coli contributions, as well as specific areas and practices for water quality improvement strategies. This study can serve as a resource for evaluating agricultural watersheds that are impaired for bacteria.

OmpR regulation of the uropathogenic Escherichia coli fimB gene in an acidic/high osmolality environment.

Uropathogenic Escherichia coli (UPEC) causes more than 90 % of all human urinary tract infections through type 1 piliated UPEC cells binding to bladder epithelial cells. The FimB and FimE site-specific recombinases orient the fimS element containing the fimA structural gene promoter. Regulation of fimB and fimE depends on environmental pH and osmolality. The EnvZ/OmpR two-component system affects osmoregulation in E. coli. To ascertain if OmpR directly regulated the fimB gene promoters, gel mobility shift and DNase I footprinting experiments were performed using OmpR or phosphorylated OmpR (OmpR-P) mixed with the fimB promoter regions of UPEC strain NU149. Both OmpR-P and OmpR bound weakly to one fimB promoter. Because there was weak binding to one fimB promoter, strain NU149 was grown in different pH and osmolality environments, and total RNAs were extracted from each population and converted to cDNAs. Quantitative reverse-transcriptase PCR showed no differences in ompR transcription among the different growth conditions. Conversely, Western blots showed a significant increase in OmpR protein in UPEC cells grown in a combined low pH/high osmolality environment versus a neutral pH/high osmolality environment. In a high osmolality environment, the ompR mutant expressed more fimB transcripts and Phase-ON positioning of the fimS element as well as higher type 1 pili levels than wild-type cells. Together these results suggest that OmpR may be post-transcriptionally regulated in UPEC cells growing in a low pH/high osmolality environment, which regulates fimB in UPEC.

Genetic determinants of biofilm development of opaque and translucent Vibrio parahaemolyticus.

Vibrio parahaemolyticus isolates display variation in colony morphology, alternating between opaque (OP) and translucent (TR) cell types. Phase variation is the consequence of genetic alterations in the locus encoding the quorum sensing output regulator OpaR. Here, we show that both cell types form stable, but distinguishable biofilms that differ with respect to attachment and detachment profiles to polystyrene, pellicle formation and stability at the air/medium interface, and submerged biofilm architecture and dispersion at a solid/liquid interface. The pellicle, which is a cohesive mat of cells, was exploited to identify mutants having altered or defective biofilm formation. Transposon insertion mutants were obtained with defects in genes affecting multiple cell surface characteristics, including extracellular polysaccharide, mannose-sensitive haemagglutinin type 4 pili and polar (but not lateral) flagella. Other insertions disrupted genes coding for potential secreted proteins or transporters of secreted proteins, specifically haemolysin co-regulated protein and an RTX toxin-like membrane fusion transporter, as well as potential modifiers of cell surface molecules (nagAC operon). The pellicle screen also identified mutants with lesions in regulatory genes encoding H-NS, a CsgD-like repressor and an AraC-like protein. This work initiates the characterization of V. parahaemolyticus biofilm formation in the OP and TR cell types and identifies a diverse repertoire of cell surface elements that participate in determining multicellular architecture.