EPA worst case water microcosms for testing phage biocontrol of Salmonella

A microplate method was developed as a tool to test phages for their ability to control Salmonella in aqueous environments. The method used EPA (U.S. Environmental Protection Agency) worst case water (WCW) in 96-well plates. The WCW provided a consistent and relatively simple defined turbid aqueous matrix, high in total organic carbon (TOC) and total dissolved salts (TDS), to simulate swine lagoon effluent, without the inconvenience of malodor and confounding effects from other biological factors. The WCW was originally defined to simulate high turbidity and organic matter in water for testing point-of-use filtration devices. Use of WCW to simulate lagoon effluent for phage testing is a new and innovative application of this matrix. Control of physical and chemical parameters (TOC, TDS, turbidity, temperature, and pH) allowed precise evaluation of microbiological parameters (Salmonella and phages). In a typical application, wells containing WCW were loaded with Salmonella enterica susp. enterica serovar Typhimurium (ATCC14028) and treated with phages alone and in cocktail combinations. Mean Salmonella inactivation rates (k, where the lower the value, the greater the inactivation) of phage treatments ranged from -0.32 to -1.60 versus -0.004 for Salmonella controls. Mean log(10) reductions (the lower the value, the greater the reduction) of Salmonella phage treatments were -1.60 for phage PR04-1, -2.14 for phage PR37-96, and -2.14 for both phages in a sequential cocktail, versus -0.08 for Salmonella controls. The WCW microcosm system was an effective tool for evaluating the biocontrol potential of Salmonella phages.     

A new sampler for stratified lagoon chemical and microbiological assessments

A sampler was needed for a spatial and temporal study of microbial and chemical stratification in a large swine manure lagoon that was known to contain zoonotic bacteria. Conventional samplers were limited to collections of surface water samples near the bank or required a manned boat. A new sampler was developed to allow simultaneous collection of multiple samples at different depths, up to 2.3 m, without a manned boat. The sampler was tethered for stability, used remote control (RC) for sample collection, and accommodated rapid replacement of sterile tubing modules and sample containers. The sampler comprised a PVC pontoon with acrylic deck and watertight enclosures, for a 12 VDC gearmotor, to operate the collection module, and vacuum system, to draw samples into reusable autoclavable tubing and 250-mL bottles. Although designed primarily for water samples, the sampler was easily modified to collect sludge. The sampler held a stable position during deployment, created minimal disturbance in the water column, and was readily cleaned and sanitized for transport. The sampler was field tested initially in a shallow fresh water lake and subsequently in a swine manure treatment lagoon. Analyses of water samples from the lagoon tests showed that chemical and bacterial levels, pH, and EC did not differ between 0.04, 0.47, and 1.0 m depths, but some chemical and bacterial levels differed between winter and spring collections. These results demonstrated the utility of the sampler and suggested that future manure lagoon studies employ fewer or different depths and more sampling dates.     

Factors Influencing Prymnesium parvum Population Dynamics During Bloom Initiation: Results from In-lake Mesocosm Experiments1

Roelke, Daniel L., Leslie Schwierzke, Bryan W. Brooks, James P. Grover, Reagan M. Errera, Theodore W. Valenti, Jr., and James L. Pinckney, 2010. Factors Influencing Prymnesium parvum Population Dynamics During Bloom Initiation: Results from In-Lake Mesocosm Experiments. Journal of the American Water Resources Association (JAWRA) 46(1):76-91. DOI: 10.1111/j.1752-1688.2009.00392.x Abstract: The alga Prymnesium parvum forms large fish-killing blooms in many Texas lakes. In some of these lakes, however, P. parvum occurs but does not develop blooms. In this study, we investigated factors that may influence bloom initiation by conducting a series of in-lake experiments involving mixing of waters from Lake Whitney, which has a history of P. parvum blooms, with waters from Lake Waco where no blooms have occurred. In all experiments, the addition of Lake Waco waters resulted in a poorer performance of P. parvum. Various experimental treatments and field data show that differences in grazing, pathogens, nutrients, and salts between the two lakes were not likely factors that contributed to this observation. Industrial and agricultural contaminants, allelochemicals and algicidal chemicals were not measured as a part of this research. However, anthropogenic contaminants other than nutrients were not observed at levels exceeding water quality standards in Lake Waco in recent years. On the other hand, nuisance cyanobacteria are common in Lake Waco, where Microcystis sp. and Anabaena sp. were abundant during the initiation of our experiments, both taxa are known to produce chemicals with allelopathic properties. In addition, the emergent field of algal-heterotrophic bacteria interactions suggests that chemicals produced by heterotrophic bacteria should not be overlooked. Further research focusing on the chemical interactions between cyanobacteria and P. parvum, as well as the potential role of algicidal bacteria, in the initiation of P. parvum blooms is necessary, as it may be important to the management of these blooms.     

Comparison of selected nutrients and bacteria from common contiguous soils inside and outside swine lagoon effluent spray fields after long-term use

Swine (Sus scofa domestica) lagoon effluent is a valuable resource. In the U.S. Mid-South it is applied from April to September to fertilize grass hay in spray-irrigated fields. Lagoon levels of nutrients and bacteria, and soil levels of nutrients have been documented, but little was known of effluent bacterial levels in soil. The present study examined levels of selected effluent bacteria and nutrients in soils inside and outside spray fields after >15 yr of effluent irrigation. Samples were collected February to March 2009 from contiguous soils spanning adjacent irrigated and nonirrigated areas. Separate soil cores for bacterial and nutrient tests were collected in pairs <10 cm apart. Five cores each were collected at 15-m intervals and combined, respectively, to comprise inside and outside samples from each of 20 soils (four each from five farms/spray fields). Analyses of data combined across all soils showed higher pH and Mehlich-3-extracrable (M3-) P, Mg, K, Na, Cu, and Zn inside than outside spray fields, while total N, total C, M3-Ca, and M3-Mn did not differ. Bacterial levels were higher inside than outside spray fields for heterotrophic plate counts, thermotolerant coliforms, Staphylococcus spp., and Clostridium perfringens, but levels of Escherichia coli and Enterococcus spp. were not different. Cultural presence/absence tests for three pathogens (Listeria spp., Campylobacter spp., and Salmonella spp.) detected only Listeria spp., which did not differ inside (23% positive samples) and outside (28% positive). Molecular tests detected all three pathogens at low levels that were not different inside and outside. We found no evidence of cumulative buildup of Campylobacter spp., Listeria spp., or Salmonela s. in spray field soils.     

pH-Dependent fate and transport of NTA-complexed cobalt through undisturbed cores of fractured shale saprolite

The codisposal of toxic metals and radionuclides with organic chelating agents has been implicated in the facilitated transport of the inorganic contaminants away from primary waste disposal areas. We investigated the transport of Co(II)NTA through undisturbed cores of fractured shale saprolite. Experiments were conducted across the pH range 4 to 8 by collecting cores from different locations within the weathering profile. Aqueous complexation, adsorption, dissociation and oxidation reactions influenced Co(II)NTA transport. The suite of reaction products identified in column effluent varied with experimental pH. At low pH and in the presence of abundant exchangeable aluminum, Co transport occurred predominantly as the Co2+ ion. At higher pH, Co was transported primarily as Co(II)NTA and the Co(III) species Co(III)(HNTA)2 and Co(III)(IDA)2. The formation of the geochemical oxidation products (Co(III) species) has far reaching implications as these compounds are kinetically and thermodynamically stable, are transported more rapidly than Co(II)NTA, and are resistant to biodegradation. These results demonstrate that natural minerals, in the physical structure encountered naturally, can be more important in the formation of mobile, stable contaminant forms than they can be for the retardation and dissociation of the contaminants.     

Response of Ceriodaphnia dubia to ionic silver: discrepancies among model predictions, measured concentrations and mortality

Silver thiosulfate, often a waste product of photoprocessing, is less bioavailable or toxic to aquatic organisms than is ionic silver. We conducted duplicate 48-h Ceriodaphnia dubia tests in reconstituted laboratory water using treatments of 92.7 nM Ag+ with various concentrations of thiosulfate. Expected Ag+ concentrations were generated for thiosulfate treatment levels using MINEQL + chemical equilibrium modeling. Ag+ concentrations in treatments were determined using a novel silicon-based sensor. Based on predicted Ag+ and published 48-h LC50 values for C. dubia, we did not expect to observe adverse effects. Yet, 100% mortality was observed at low thiosulfate treatments, whereas > 85% and > 95% survival was observed at higher thiosulfate treatment levels, respectively. Our results indicate that biotic responses match the sensor-based Ag+ concentrations. However, there is a discrepancy between these empirical results and responses expected to occur with Ag+ concentrations as predicted by MINEQL + chemical modeling. By correlating silicon sensor data with toxicity results obtained from our laboratory, our work clearly relates a specific chemical form (Ag+) to toxicity results.     

Correlation of human olfactory responses to airborne concentrations of malodorous volatile organic compounds emitted from swine effluent

Direct multicomponent analysis of malodorous volatile organic compounds (VOCs) present in ambient air samples from 29 swine (Sus scrofa) production facilities was used to develop a 19-component artificial swine odor solution that simulated olfactory properties of swine effluent. Analyses employing either a human panel consisting of 14 subjects or gas chromatography were performed on the air stream from an emission chamber to assess human olfactory responses or odorant concentration, respectively. Analysis of the olfactory responses using Fisher's LSD statistics showed that the subjects were sensitive to changes in air concentration of the VOC standard across dilutions differing by approximately 16%. The effect of chemical synergisms and antagonisms on human olfactory response magnitudes was assessed by altering the individual concentration of nine compounds in artificial swine odor over a twofold concentration range while maintaining the other 18 components at a constant concentration. A synergistic olfactory response was observed when the air concentration of acetic acid was increased relative to the concentration of other VOC odorants in the standard. An antagonistic olfactory response was observed when the air concentration of 4-ethyl phenol was increased relative to the other VOC odorants in the standard. The collective odorant responses for nine major VOCs associated with swine odor were used to develop an olfactory prediction model to estimate human odor response magnitudes to swine manure odorants through measured air concentrations of indicator VOCs. The results of this study show that direct multicomponent analysis of VOCs emitted from swine effluent can be applied toward estimating perceived odor intensity.     

Vapor flammability above aqueous solutions of flammable liquids

The flammability of vapors above aqueous solutions of ethanol and acetonitrile was studied experimentally in a 20-L combustion apparatus. No liquid was present in the apparatus, but the vapor concentrations were adjusted to correspond to the vapor in equilibrium with a specified aqueous solution. The experimental results for these two systems show that

• As water is added to the vapor, the lower boundary of the flammability zone decreases. For ethanol, the lower flammability limits (LFL) decreases from 3.7% for pure vapor to 3.2% with saturated water vapor. For acetonitrile, the decrease is from 4.2% to 3.8%. Thus, to a good approximation, the water vapor can be treated as an inert, enabling the data to be displayed on a single flammability triangle diagram. This provides a very simplified method for estimating the flammable behavior for aqueous solutions.

• The upper boundary of the flammability zone is unchanged with the addition of water.

• The limiting oxygen concentration (LOC) is essentially constant for all concentrations of aqueous solutions. The LOC for the pure solvent may be used as a universal LOC for all solvent concentrations.

• The vapor mixture above the aqueous solution is not flammable below a certain liquid mol fraction of flammable. The flammable concentration at which this occurs can be called the maximum safe solvent concentration (MSSC). A method is presented to determine the MSSC from experimental flammability data.

• The oxygen concentration defining the flammable boundary for the vapor decreases rapidly from the MSSC and then increases as the liquid solvent concentration increases.

The calculated adiabatic flame temperature (CAFT) method qualitatively predicts the same behavior as the experimental data.