Impact of pre-application treatment on municipal sludge composition, soil dynamics of antibiotic resistance genes, and abundance of antibiotic-resistance genes on vegetables at harvest.

In many jurisdictions sludge recovered from the sewage treatment process is a valued fertilizer for crop production. Pre-treatment of sewage sludge prior to land application offers the potential to abate enteric microorganisms that carry genes conferring resistance to antibiotics. Pre-treatment practices that accomplish this should have the desirable effect of reducing the risk of contamination of crops or adjacent water with antibiotic resistance genes carried in these materials. In the present study, we obtained municipal sludge that had been subjected to one of five treatments. There were, anaerobic-digestion or aerobic-digestion, in both instances with and without dewatering; and heat-treatment and pelletization. Each of the five types of biosolids was applied to an agricultural field at commercial rates, following which lettuce, carrots and radishes were planted. Based on qPCR, the estimated antibiotic gene loading rates were comparable with each of the five biosolids. However, the gene abundance in soil following application of the pelletized biosolids was anomalously lower than expected. Following application, the abundance of antibiotic resistance genes decreased in a generally coherent fashion, except sul1 which increased in abundance during the growing season in the soil fertilized with pelletized biosolids. Based on qPCR and high throughput sequencing evidence for transfer of antibiotic resistance genes from the biosolids to the vegetables at harvest was weak. Clostridia were more abundant in soils receiving any of the biosolids except the pelletized. Overall, the behavior of antibiotic resistance genes in soils receiving aerobically or anaerobically-digested biosolids was consistent and coherent with previous studies. However, dynamics of antibiotic resistance genes in soils receiving the heat treated pelletized biosolids were very different, and the underlying mechanisms merit investigation.

Persistence of antibiotic resistance and plasmid-associated genes in soil following application of sewage sludge and abundance on vegetables at harvest.

Sewage sludge recovered from wastewater treatment plants contains antibiotic residues and is rich in antibiotic resistance genes, selected for and enriched in the digestive tracts of human using antibiotics. The use of sewage sludge as a crop fertilizer constitutes a potential route of human exposure to antibiotic resistance genes through consumption of contaminated crops. Several gene targets associated with antibiotic resistance (catA1, catB3, ereA, ereB, erm(B), str(A), str(B), qnrD, sul1, and mphA), mobile genetic elements (int1, mobA, IncW repA, IncP1 groups -α, -ß, -δ, -γ, -ε), and bacterial 16S rRNA (rrnS) were quantified by qPCR from soil and vegetable samples obtained from unamended and sludge-amended plots at an experimental field in London, Ontario. The qPCR data reveals an increase in abundance of gene targets in the soil and vegetables samples, indicating that there is potential for additional crop exposure to antibiotic resistance genes carried within sewage sludge following field application. It is therefore advisable to allow an appropriate delay period before harvesting of vegetables for human consumption.

Impact of fertilizing with raw or anaerobically digested sewage sludge on the abundance of antibiotic-resistant coliforms, antibiotic resistance genes, and pathogenic bacteria in soil and on vegetables at harvest.

The consumption of crops fertilized with human waste represents a potential route of exposure to antibiotic-resistant fecal bacteria. The present study evaluated the abundance of bacteria and antibiotic resistance genes by using both culture-dependent and molecular methods. Various vegetables (lettuce, carrots, radish, and tomatoes) were sown into field plots fertilized inorganically or with class B biosolids or untreated municipal sewage sludge and harvested when of marketable quality. Analysis of viable pathogenic bacteria or antibiotic-resistant coliform bacteria by plate counts did not reveal significant treatment effects of fertilization with class B biosolids or untreated sewage sludge on the vegetables. Numerous targeted genes associated with antibiotic resistance and mobile genetic elements were detected by PCR in soil and on vegetables at harvest from plots that received no organic amendment. However, in the season of application, vegetables harvested from plots treated with either material carried gene targets not detected in the absence of amendment. Several gene targets evaluated by using quantitative PCR (qPCR) were considerably more abundant on vegetables harvested from sewage sludge-treated plots than on vegetables from control plots in the season of application, whereas vegetables harvested the following year revealed no treatment effect. Overall, the results of the present study suggest that producing vegetable crops in ground fertilized with human waste without appropriate delay or pretreatment will result in an additional burden of antibiotic resistance genes on harvested crops. Managing human exposure to antibiotic resistance genes carried in human waste must be undertaken through judicious agricultural practice.

Uptake of pharmaceuticals, hormones and parabens into vegetables grown in soil fertilized with municipal biosolids.

Several recent greenhouse studies have established the potential for uptake of human pharmaceuticals from soil fertilized with municipal biosolids into a variety of crops. In the present study, a field experiment was undertaken to evaluate the uptake of organic micropollutants from soil fertilized with municipal biosolids at a regulated application rate into tomatoes, carrots, potatoes and sweet corn produced under normal farming conditions. The vegetables were grown according to farming practices mandated by the province of Ontario Canada, the key feature being a one-year offset between biosolid application and the harvest of crops for human consumption. Biosolids at application, and crop samples following harvest were analyzed for 118 pharmaceuticals and transformation products, 17 hormones or hormone transformation products, and 6 parabens. Analyte concentrations in the biosolids were consistent with those detected in other surveys. Eight of the 141 analytes were detected in one or two crop replicates at concentrations ranging from 0.33 to 6.25 ng/g dry weight, but no analytes were consistently detected above the detection limit in all triplicate treated plots. Overall, this study suggests that the potential for micropollutant uptake into crops under normal farming conditions is low.

Runoff of pharmaceuticals and personal care products following application of dewatered municipal biosolids to an agricultural field.

Municipal biosolids are a useful source of nutrients for crop production, and commonly used in agriculture. In this field study, we applied dewatered municipal biosolids at a commercial rate using broadcast application followed by incorporation. Precipitation was simulated at 1, 3, 7, 21 and 34 days following the application on 2 m(2) microplots to evaluate surface runoff of various pharmaceuticals and personal care products (PPCPs), namely atenolol, carbamazepine, cotinine, caffeine, gemfibrozil, naproxen, ibuprofen, acetaminophen, sulfamethoxazole, triclosan and triclocarban. There was little temporal coherence in the detection of PPCPs in runoff, various compounds being detected maximally on days 1, 3, 7 or 36. Maximum concentrations in runoff ranged from below detection limit (gemfibrozil) to 109.7 ng L(-1) (triclosan). Expressing the total mass exported as a percentage of that applied, some analytes revealed little transport potential (<1% exported; triclocarban, triclosan, sulfamethoxazole, ibuprofen, naproxen and gemfibrozil) whereas others were readily exported (>1% exported; acetaminophen, carbamazepine, caffeine, cotinine, atenolol). Those compounds with little transport potential had log K(ow) values of 3.18 or greater, whereas those that were readily mobilized had K(ow) values of 2.45 or less. Maximal concentrations of all analytes were below toxic concentrations using a variety of endpoints available in the literature. In summary, this study has quantified the transport potential in surface runoff of PPCPs from land receiving biosolids, identified that log K(ow) may be a determinant of runoff transport potential of these analytes, and found maximal concentrations of all chemicals tested to be below toxic concentrations using a variety of endpoints.

Runoff of pharmaceuticals and personal care products following application of biosolids to an agricultural field.

Municipal biosolids are a source of nutrients for crop production. Beneficial Management Practices (BMPs) can be used to minimize the risk of contamination of adjacent water resources with chemical or microbial agents that are of public or environmental health concern. In this field study, we applied biosolids slurry at a commercial rate using either subsurface injection or broadcast application followed by incorporation. Precipitation was simulated at 1, 3, 7, 22, 36 and 266 days post-application on 2 m(2) microplots to evaluate surface runoff of 9 model pharmaceuticals and personal care products (PPCPs), atenolol, carbamazepine, cotinine, gemfibrozil, naproxen, ibuprofen, acetaminophen, sulfamethoxazole and triclosan. In runoff from the injected plots, concentrations of the model PPCPs were generally below the limits of quantitation. In contrast, in the broadcast application treatment, the concentrations of atenolol, carbamazepine, cotinine, gemfibrozil, naproxen, sulfamethoxazole and triclosan on the day following application ranged from 70-1477 ng L(-1) in runoff and generally declined thereafter with first order kinetics. The total mass of PPCPs mobilized in surface runoff per m(2) of the field ranged from 0.63 microg for atenolol to 21.1 microg for ibuprofen. For ibuprofen and acetaminophen, concentrations in runoff first decreased and then increased, suggesting that these drugs were initially chemically or physically sequestered in the biosolids slurry, and subsequently released in the soil. Carbamazepine and triclosan were detected at low concentrations in a runoff event 266 days after broadcast application. Overall, this study showed that injection of biosolids slurry below the soil surface could effectively eliminate surface runoff of PPCPs.