Tina B. Bech

Transport and Distribution of Salmonella enterica Serovar Typhimurium in Loamy and Sandy Soil Monoliths with Applied Liquid Manure

ABSTRACT A leaching experiment, where liquid manure spiked with Salmonella enterica serovar Typhimurium (Tet+) DSM554 was applied to soil surfaces, was conducted on intact soil monoliths (60 cm in diameter and 100 cm long). A total of 6.5 × 1010 CFU was applied to each column. We found that Salmonella serovar Typhimurium could be transported to a 1-m depth in loamy soil at concentrations reaching 1.3 × 105 CFU/ml of leachate. The test strain was found in concentrations ranging from 300 to 1.35 cells/ml in loamy soil throughout the 27 days of the experiment, while concentrations below 20 cells/ml were sporadically detected in the leachates from sandy monoliths. Real-time PCR targeting invA DNA showed a clear correspondence between the total and culturable numbers of cells in the leachate, indicating that most cells leached were viable. On day 28, distribution of Salmonella serovar Typhimurium at five depths in the four monoliths was determined. The highest recovery rate, ranging from 1.5% to 3.8% of the total applied inoculum, was found in the top 0.2 m.

Leaching of Cryptosporidium parvum Oocysts, Escherichia coli, and a Salmonella enterica Serovar Typhimurium Bacteriophage through Intact Soil Cores following Surface Application and Injection of Slurry

ABSTRACT Increasing amounts of livestock manure are being applied to agricultural soil, but it is unknown to what extent this may be associated with contamination of aquatic recipients and groundwater if microorganisms are transported through the soil under natural weather conditions. The objective of this study was therefore to evaluate how injection and surface application of pig slurry on intact sandy clay loam soil cores influenced the leaching of Salmonella enterica serovar Typhimurium bacteriophage 28B, Escherichia coli, and Cryptosporidium parvum oocysts. All three microbial tracers were detected in the leachate on day 1, and the highest relative concentration was detected on the fourth day (0.1 pore volume). Although the concentration of the phage 28B declined over time, the phage was still found in leachate at day 148. C. parvum oocysts and chloride had an additional rise in the relative concentration at a 0.5 pore volume, corresponding to the exchange of the total pore volume. The leaching of E. coli was delayed compared with that of the added microbial tracers, indicating a stronger attachment to slurry particles, but E. coli could be detected up to 3 months. Significantly enhanced leaching of phage 28B and oocysts by the injection method was seen, whereas leaching of the indigenous E. coli was not affected by the application method. Preferential flow was the primary transport vehicle, and the diameter of the fractures in the intact soil cores facilitated transport of all sizes of microbial tracers under natural weather conditions.

Leaching of Salmonella enterica in Clay Columns Comparing Two Manure Application Methods

Transfer of zoonotic bacterial pathogens through intact soil columns was monitored in an outdoor lysimeter over 36 d. Manure spiked with Salmonella enterica serovar Senftenberg was applied to either the soil surface or injected 0.08 m into the soil to compare leaching associated with the two manure application methods. The highest concentrations of S. enterica (up to 60,000 S. enterica CFU/mL) were detected on Day 1 in the first drainage samples, with measurable but declining concentrations persisting for 10 to 36 d depending on replicate columns. The total recovery of leached S. enterica in drainage samples ranged from 0.08% to 13.8%. When comparing the two application methods, there was no statistically significant difference in the leaching concentration of S. enterica at each sampling time during the study period. In addition, comparison of enumerations by selective plating and real-time polymerase chain reaction yielded similar concentrations of S. enterica, indicating that mainly viable and culturable cells were leached from the columns. When the experiment was terminated, the fluorescent dye Acid Yellow was applied to four selected columns and the distribution of dye and size of active (dye-stained) pores were measured with a digital camera and visualization software. The profiles showed that the area covered by active pores ranged from 0.1% to 3.6%. The relatively small fraction of active pores in the soil profile was consistent with the evidence of rapid transport of S. enterica and chloride in the columns.

Redistribution and persistence of microorganisms and steroid hormones after soil-injection of swine slurry

The redistribution and fate of contaminants in pig slurry after direct injection were investigated at two field sites, Silstrup (sandy clay loam) and Estrup (sandy loam), in Denmark. Intact soil samples were collected for up to seven weeks after slurry injection and concentrations of Salmonella Typhimurium Bacteriophage 28B (phage 28B), Escherichia coli, steroid hormones and other slurry components (water, volatile solids, chloride and mineral N) determined in and around the injection slit. The two experiments at Silstrup and Estrup differed with respect to slurry solid content (6.3 vs. 0.8%), as well as soil clay content (27 vs. 15%) and differed considerably with respect to the initial redistribution of slurry-borne contaminants in soil. The transport of microorganisms from the slurry injection slit to the surrounding soil was much lower than that of mineral N and chloride due to attachment and entrapment. The redistribution of E. coli was more affected by site-specific conditions compared to phage 28B, possibly due to the larger cell size of E. coli. The overall recovery of phage 28B was 0.8-4%, and of E. coli 0.0-1.3% in different samples, by the end of the study. Nine different steroid hormones were detected in the slurry slit, and a slow redistribution to the surrounding soil was observed. Overall recovery of estrogens was 0.0 to 6.6% in different samples. The study showed that the combination of soil and slurry properties determined the initial spreading of contaminants, and hence the potential for subsequent leaching.

Bulk soil and maize rhizosphere resistance genes, mobile genetic elements and microbial communities are differently impacted by organic and inorganic fertilization

Abstract Organic soil fertilizers, such as livestock manure and biogas digestate, frequently contain bacteria carrying resistance genes (RGs) to antimicrobial substances and mobile genetic elements (MGEs). The effects of different fertilizers (inorganic, manure, digestate) on RG and MGE abundance and microbial community composition were investigated in a field plot experiment. The relative abundances of RGs [sul1, sul2, tet(A), tet(M), tet(Q), tet(W), qacE&Dgr;1/qacE] and MGEs [intI1, intI2, IncP‐1, IncP‐1&egr; and LowGC plasmids] in total community (TC)‐DNA from organic fertilizers, bulk soil and maize rhizosphere were quantified by qPCR before/after fertilization and prior to maize harvest. Microbial communities were analyzed via Illumina sequencing of 16S rRNA gene fragments amplified from TC‐DNA. Compared to inorganic fertilization, manure treatments increased relative abundances of all RGs analyzed, integrons and few genera affiliated to Bacteroidetes and Firmicutes in bulk soil, while digestate increased sul2, tet(W) and intI2. At harvest, treatment effects vanished in bulk soil. However, organic fertilizer effects were still detectable in the rhizosphere for RGs [manure: intI1, sul1; digestate: tet(W)] and Clostridium related sequences (digestates) with increased relative abundance. Our data indicated transient organic fertilizer effects on RGs, MGEs and microbial community composition in bulk soil with long‐term history of digestate or manure application. Figure. No Caption available.

Adhesion of Escherichia coli and Salmonella enterica to Soil in Runoff as Influenced by Polyacrylamide

Polyacrylamide (PAM) is used in agriculture to reduce soil erosion and has been reported to reduce turbidity, nutrients, and pollutants in surface runoff water. The objective of this work was to determine the effect of PAM on the concentration of enteric bacteria in surface runoff by comparing four enteric bacteria representing phenotypically different motility and hydrophobicity from three soils. Results demonstrated that bacterial surface runoff was differentially influenced by the PAM treatment. Polyacrylamide treatment increased surface runoff for adhered and planktonic cells from a clay soil; significantly decreased surface runoff of adhered bacteria, while no difference was observed for planktonic bacteria from the sandy loam; and significantly decreased the surface runoff of planktonic cells, while no difference was observed for adhered bacteria from the clay loam. Comparing strains from a final water sample collected after 48 h showed a greater loss of while serovar Poona was almost not detected. Thus, (i) the PAM efficiency in reducing the concentration of enteric bacteria in surface runoff was influenced by soil type and (ii) variation in the loss of enteric bacteria highlights the importance of strain-specific properties that may not be captured with general fecal indicator bacteria.