Jim Grant

Long-Term Persistence and Leaching of Escherichia coli in Temperate Maritime Soils

ABSTRACT Enteropathogen contamination of groundwater, including potable water sources, is a global concern. The spreading on land of animal slurries and manures, which can contain a broad range of pathogenic microorganisms, is considered a major contributor to this contamination. Some of the pathogenic microorganisms applied to soil have been observed to leach through the soil into groundwater, which poses a risk to public health. There is a critical need, therefore, for characterization of pathogen movement through the vadose zone for assessment of the risk to groundwater quality due to agricultural activities. A lysimeter experiment was performed to investigate the effect of soil type and condition on the fate and transport of potential bacterial pathogens, using Escherichia coli as a marker, in four Irish soils (n = 9). Cattle slurry (34 tonnes per ha) was spread on intact soil monoliths (depth, 1 m; diameter, 0.6 m) in the spring and summer. No effect of treatment or the initial soil moisture on the E. coli that leached from the soil was observed. Leaching of E. coli was observed predominantly from one soil type (average, 1.11 ± 0.77 CFU ml−1), a poorly drained Luvic Stagnosol, under natural rainfall conditions, and preferential flow was an important transport mechanism. E. coli was found to have persisted in control soils for more than 9 years, indicating that autochthonous E. coli populations are capable of becoming naturalized in the low-temperature environments of temperate maritime soils and that they can move through soil. This may compromise the use of E. coli as an indicator of fecal pollution of waters in these regions.

Impact of chemical amendment of dairy cattle slurry on phosphorus, suspended sediment and metal loss to runoff from a grassland soil.

Emerging remediation technologies such as chemical amendment of dairy cattle slurry have the potential to reduce phosphorus (P) solubility and consequently reduce P losses arising from land application of dairy cattle slurry. The aim of this study was to determine the effectiveness of chemical amendment of slurry to reduce incidental losses of P and suspended sediment (SS) from grassland following application of dairy cattle slurry and to examine the effect of amendments on metal concentrations in runoff water. Intact grassed-soil samples were placed in two laboratory runoff boxes, each 200-cm-long by 22.5-cm-wide by 5-cm-deep, before being amended with dairy cattle slurry (the study control) and slurry amended with either: (i) alum, comprising 8% aluminium oxide (Al(2)O(3)) (1.11:1 aluminium (Al):total phosphorus (TP) of slurry) (ii) poly-aluminium chloride hydroxide (PAC) comprising 10% Al(2)O(3) (0.93:1 Al:TP) (iii) analytical grade ferric chloride (FeCl(2)) (2:1 Fe:TP), (iv) and lime (Ca(OH)(2)) (10:1 Ca:TP). When compared with the study control, PAC was the most effective amendment, reducing dissolved reactive phosphorus (DRP) by up to 86% while alum was most effective in reducing SS (88%), TP (94%), particulate phosphorus (PP) (95%), total dissolved phosphorus (TDP) (81%), and dissolved unreactive phosphorus (DUP) (86%). Chemical amendment of slurry did not appear to significantly increase losses of Al and Fe compared to the study control, while all amendments increased Ca loss compared to control and grass-only treatment. While chemical amendments were effective, the reductions in incidental P losses observed in this study were similar to those observed in other studies where the time from slurry application to the first rainfall event was increased. Timing of slurry application may therefore be a much more feasible way to reduce incidental P losses. Future work must examine the long-term effects of amendments on P loss to runoff and not only incidental losses.

Incidental phosphorus and nitrogen loss from grassland plots receiving chemically amended dairy cattle slurry

Chemical amendment of dairy cattle slurry has been shown to effectively reduce incidental phosphorus (P) losses in runoff; however, the effects of amendments on incidental nitrogen (N) losses are not as well documented. This study examined P and N losses in runoff during three simulated rainfall events 2, 10 and 28 days after a single application of unamended/chemically amended dairy cattle slurry. Twenty-five hydraulically isolated plots, each measuring 0.9 m by 0.4 m and instrumented with runoff collection channels, were randomly assigned the following treatments: (i) grass-only, (ii) slurry-only (the study-control), (iii) slurry amended with industrial grade liquid alum comprising 8% Al₂O₃, (iv) slurry amended with industrial grade liquid poly-aluminum chloride (PAC) comprising 10% Al₂O₃, and (v) slurry amended with lime. During the first rainfall event, lime was ineffective but alum and PAC effectively reduced dissolved reactive P (DRP) (by 95 and 98%, respectively) and total P (TP) flow-weighted-mean-concentrations (by 82 and 93%, respectively) in runoff compared to the study-control. However, flow-weighted-mean-concentrations of ammonium-N (NH₄--N) in runoff were increased with alum- (81%) and lime-treated (11%) slurry compared to the study-control whereas PAC reduced the NH₄--N by 82%. Amendments were not observed to have a significant effect on NO₃--N losses during this study. Slurry amendments reduced P losses for the duration of the study, whereas the effect of amendments on N losses was not significant following the first event. Antecedent volumetric water content of the soil or slope of the plots did not appear to affect runoff volume. However, runoff volumes (and consequently loads of P and N) were observed to increase for the chemically amended plots compared to the control and soil-only plots. This work highlights the importance of considering both P and N losses when implementing a specific nutrient mitigation measure.

Evaluation of headspace equilibration methods for quantifying greenhouse gases in groundwater.

The objective of the study was to evaluate the different headspace equilibration methods for the quantification of dissolved greenhouse gases in groundwater. Groundwater samples were collected from wells with contrasting hydrogeochemical properties and degassed using the headspace equilibration method. One hundred samples from each well were randomly selected, treatments were applied and headspace gases analysed by gas chromatography. Headspace equilibration treatments varied helium (He):water ratio, shaking time and standing time. Mean groundwater N(2)O, CO(2) and CH(4) concentrations were 0.024 mg N L(-1), 13.71 mg C L(-1) and 1.63 μg C L(-1), respectively. All treatments were found to significantly influence dissolved gas concentrations. Considerable differences in the optimal He:water ratio and standing time were observed between the three gases. For N(2)O, CO(2) and CH(4) the optimum operating points for He:water ratio was 4.4:1, 3:1 and 3.4:1; shaking time was 13, 12 and 13 min; and standing time was 63, 17 and 108 min, respectively. The headspace equilibration method needs to be harmonised to ensure comparability between studies. The experiment reveals that He:water ratio 3:1 and shaking time 13 min give better estimation of dissolved gases than any lower or higher ratios and shaking times. The standing time 63, 17 and 108 min should be applied for N(2)O, CO(2) and CH(4), respectively.

Consequences of varied soil hydraulic and meteorological complexity on unsaturated zone time lag estimates.

The true efficacy of a programme of agricultural mitigation measures within a catchment to improve water quality can be determined only after a certain hydrologic time lag period (subsequent to implementation) has elapsed. As the biophysical response to policy is not synchronous, accurate estimates of total time lag (unsaturated and saturated) become critical to manage the expectations of policy makers. The estimation of the vertical unsaturated zone component of time lag is vital as it indicates early trends (initial breakthrough), bulk (centre of mass) and total (Exit) travel times. Typically, estimation of time lag through the unsaturated zone is poor, due to the lack of site specific soil physical data, or by assuming saturated conditions. Numerical models (e.g. Hydrus 1D) enable estimates of time lag with varied levels of input data. The current study examines the consequences of varied soil hydraulic and meteorological complexity on unsaturated zone time lag estimates using simulated and actual soil profiles. Results indicated that: greater temporal resolution (from daily to hourly) of meteorological data was more critical as the saturated hydraulic conductivity of the soil decreased; high clay content soils failed to converge reflecting prevalence of lateral component as a contaminant pathway; elucidation of soil hydraulic properties was influenced by the complexity of soil physical data employed (textural menu, ROSETTA, full and partial soil water characteristic curves), which consequently affected time lag ranges; as the importance of the unsaturated zone increases with respect to total travel times the requirements for high complexity/resolution input data become greater. The methodology presented herein demonstrates that decisions made regarding input data and landscape position will have consequences for the estimated range of vertical travel times. Insufficiencies or inaccuracies regarding such input data can therefore mislead policy makers regarding the achievability of water quality targets.

The nitrification inhibitor dicyandiamide increases mineralization–immobilization turnover in slurry-amended grassland soil

Nitrification inhibitors are used in agriculture for the purpose of decreasing nitrogen (N) losses, by limiting the microbially mediated oxidation of ammonium (NH 4 + ) to nitrate (NO 3 − ). Successful inhibition of nitrification has been shown in numerous studies, but the extent to which inhibitors affect other N transformations in soil is largely unknown. In the present study, cattle slurry was applied to microcosms of three different grassland soils, with or without the nitrification inhibitor dicyandiamide (DCD). A solution containing NH 4 + and NO 3 − , labelled with 15 N either on the NH 4 + or the NO 3 − part, was mixed with the slurry before application. Gross N transformation rates were estimated using a 15 N tracing model. In all three soils, DCD significantly inhibited gross autotrophic nitrification, by 79–90%. Gross mineralization of recalcitrant organic N increased significantly with DCD addition in two soils, whereas gross heterotrophic nitrification from the same pool decreased with DCD addition in two soils. Fungal to bacterial ratios were not significantly affected by DCD addition. Total gross mineralization and immobilization increased significantly across the three soils when DCD was used, which suggests that DCD can cause non-target effects on soil N mineralization–immobilization turnover.

Insights into the low-temperature adaptation and nutritional flexibility of a soil-persistent Escherichia coli

An understanding of the survival capacity of Escherichia coli in soil is critical for the evaluation of its role as a faecal indicator. Recent reports that E. coli can become long-term residents in maritime temperate soils have raised the question of how the organism survives and competes for niche space in the suboptimal soil environment. The ability of an environmental isolate to utilize 380 substrates was assessed together with that of a reference laboratory strain (E. coli K12) at both 15 and 37 °C. At 15 °C, the environmental strain could utilize 161 substrates, with only 67 utilizable by the reference strain, while at 37 °C, 239 and 223 substrates could be utilized by each strain respectively. An investigation into the cold response of the strains revealed that E. coli K12 was found to reduce the expression of biosynthetic proteins at 15 °C, while the environmental isolate seemed to switch on proteins involved in stress response, suggesting low-temperature adaptation in the latter. Taken together, the results indicate that the environmentally persistent E. coli strain is well adapted to use a wide range of nutrient sources at 15 °C and to direct its protein expression to maintain a relatively fast growth rate at low temperature.

Groundwater nitrate reduction versus dissolved gas production: A tale of two catchments

At the catchment scale, a complex mosaic of environmental, hydrogeological and physicochemical characteristics combine to regulate the distribution of groundwater and stream nitrate (NO3-). The efficiency of NO3- removal (via denitrification) versus the ratio of accumulated reaction products, dinitrogen (excess N2) & nitrous oxide (N2O), remains poorly understood. Groundwater was investigated in two well drained agricultural catchments (10km2) in Ireland with contrasting subsurface lithologies (sandstone vs. slate) and landuse. Denitrification capacity was assessed by measuring concentration and distribution patterns of nitrogen (N) species, aquifer hydrogeochemistry, stable isotope signatures and aquifer hydraulic properties. A hierarchy of scale whereby physical factors including agronomy, water table elevation and permeability determined the hydrogeochemical signature of the aquifers was observed. This hydrogeochemical signature acted as the dominant control on denitrification reaction progress. High permeability, aerobic conditions and a lack of bacterial energy sources in the slate catchment resulted in low denitrification reaction progress (0-32%), high NO3- and comparatively low N2O emission factors (EF5g1). In the sandstone catchment denitrification progress ranged from 4 to 94% and was highly dependent on permeability, water table elevation, dissolved oxygen concentration solid phase bacterial energy sources. Denitrification of NO3- to N2 occurred in anaerobic conditions, while at intermediate dissolved oxygen; N2O was the dominant reaction product. EF5g1 (mean: 0.0018) in the denitrifying sandstone catchment was 32% less than the IPCC default. The denitrification observations across catchments were supported by stable isotope signatures. Stream NO3- occurrence was 32% lower in the sandstone catchment even though N loading was substantially higher than the slate catchment.

Effect of feeding sodium butyrate in the late finishing period on Salmonella carriage, seroprevalence, and growth of finishing pigs.

Pork is an important source of human salmonellosis and low-cost on-farm control measures may provide a useful element in reducing the prevalence of this pathogen in food. This study investigated the effectiveness of dietary supplementation with sodium butyrate administered to finisher pigs for ∼4-weeks prior to slaughter to control Salmonella shedding on highly contaminated farms. Two trials (A and B) were conducted on two commercial pig farms, which had a history of high Salmonella seroprevalence. In both trials, pens (14 pens of 12 pigs/pen in Trial A and 12 pens of 12-17 pigs/pen in Trial B) were randomly assigned to a control (finisher feed without additive) or a treatment group (the same feed with 3kg sodium butyrate/t) for 24-28days, depending on the trial. Faeces were collected from each pig on days 0, 12 and 24/28, and blood, caecal digesta and ileocaecal/mesenteric lymph nodes were collected from the slaughterhouse. Pigs were weighed at the start and end of the trials, feed intake was recorded, and carcass quality parameters were recorded at slaughter. In Trial A, Salmonella shedding was reduced in the treatment compared to the control group at the end of the trial (30% versus 57% probability of detecting Salmonella in faeces, respectively; p<0.001). This reflected the serology results, with detection of a lower seroprevalence in the treatment compared to the control group using the 20% optical density cut-off (69.5% versus 89%; p=0.001). However, no effect on faecal shedding or seroprevalance was observed in Trial B, which may be explained by the detection of a concomitant infection with Lawsonia intracellularis. No significant differences in Salmonella recovery rates were observed in the caecal digesta or lymph nodes in either trial. Furthermore, feed intake, weight gain, and feed conversion efficiency (FCE) did not differ between groups (p>0.05) in either trial. Numerical improvements in weight gain and FCE were found with sodium butyrate treatment, which gave a cost benefit of €0.04/kg of live-weight gain. Overall, results suggest that strategic feeding of sodium butyrate, at 3kg/t of feed, to finishing pigs for 24-28days prior to slaughter was effective in reducing Salmonella shedding and seroprevalance but perhaps only in the absence of co-infection with other pathogens. However, sodium butyrate supplementation at this rate did not influence intestinal carriage, nor did it reduce seroprevalence to below the cut-off used for the high Salmonella risk category in Ireland (50%), or significantly improve growth performance.

The in vitro rumen methane output of perennial grass species and white clover varieties, and associative effects for their binary mixtures, evaluated using a batch-culture technique

Grassland swards containing white clover varieties (WCV) may result in lower enteric methane (CH4) output from grazing ruminants than swards of only perennial grass species (PGS) due to differences in their chemical composition and rumen fermentation dynamics. The objectives of the present study were to compare the chemical composition, in vitro rumen fermentation variables and CH4 output per unit of feed for a range of common PGS and WCV harvested in May from simulated grazing regimes, and to determine the effects of binary mixtures of the PGS and WCV on in vitro rumen fermentation variables and CH4 output, using a batch-culture technique. Four PGS (perennial ryegrass, cocksfoot, meadow fescue and timothy) and three WCV (Aran, Chieftain and Crusader) were incubated as sole substrates or as part of binary mixtures (PGS : WCV ratios of 0 : 1, 0.25 : 0.75, 0.5 : 0.5, 0.75 : 0.25 and 1 : 0) for 24 h at 39°C with buffered rumen fluid. All WCV had lower (P < 0.001) CH4 output per unit of apparent DM disappeared (aDMD; mean value across WCV (s.e.m.) of 27.0 (1.35) mL/g aDMD v. 36.1 (0.90) mL/g aDMD across PGS) during the in vitro rumen incubation than did all PGS. The WCV also had lower CH4 output per unit of total volatile fatty acids (corresponding values of 0.147 (0.0090) v. 0.199 (0.0073) mmol/mmol total volatile fatty acid output) and per unit of total gas produced (0.118 (0.0022) v. 0.153 (0.0024) mmol/mmol total gas produced) than did the PGS. In addition, Aran and Crusader had lower (P < 0.001) CH4 output per unit of feed DM incubated than did all PGS. There were synergistic associative effects (i.e. where the response was greater for the mixtures than the arithmetic calculation using the responses for PGS and WCV alone; P < 0.05) of mixing the PGS and WCV in binary combinations on all CH4 output variables.