Abstract
A growing concern regarding the use of animal manure as fertilizer is the contamination of soil, plants, and the environment with a variety of antibiotic-resistant and pathogenic bacteria. This study quantified and characterized the antibiotic resistance profiles of Escherichia coli in soil before and after chicken litter application to determine the impact of manure on the soil resistome. Litter and soil samples were collected from a sugarcane field before and after litter application. E. coli was isolated and quantified using the Colilert®-18/Quanti-tray® 2000 and 10 randomly selected isolates from the positive wells of each Quanti-tray were putatively identified on eosin methylene blue agar. Real-time PCR was used to confirm the isolates by targeting the uidA gene. Antibiotic susceptibility test against 18 antibiotics was conducted using the disk diffusion method, and the multiple antibiotic resistance index was calculated. Soil amendment with chicken litter significantly increased the number of antibiotic-resistant E. coli in the soil. Among the 126 E. coli isolates purified from all the samples, 76% showed resistance to at least one antibiotic, of which 54.2% were multidrug-resistant (MDR). The highest percentage resistance was to tetracycline (78.1%), with the least percentage resistance (3.1%) to imipenem, tigecycline, and gentamicin. The isolates also showed resistance to chloramphenicol (63.5%), ampicillin (58.3%), trimethoprim-sulfamethoxazole (39.6%), cefotaxime (30.2%), ceftriaxone (26.0%), cephalexin (20.8%), cefepime (11.5%), amoxicillin-clavulanic acid (11.5%), cefoxitin (10.4%), Nalidixic acid (9.4%), amikacin (6.3%), and ciprofloxacin (4.2%). Of the 54.2% (52/96) MDR, the highest number was isolated from the litter-amended soil (61.5%) and the least isolates from soil samples collected before litter application (1.9%). The relatively higher mean MAR index of the litter-amended soil (0.14), compared to the soil before the amendment (0.04), suggests soil pollution with antibiotic-resistant E. coli from sources of high antibiotic use. E. coli could only be detected in the soil up to 42 days following manure application, making it a suitable short-term indicator of antibiotic resistance contamination. Notwithstanding its relatively short detectability/survival, the application of chicken litter appeared to transfer antibiotic-resistant E. coli to the soil, enhancing the soil resistome and highlighting the consequences of such agricultural practices on public health.
Highlights
The increasing prevalence of antibiotic-resistant bacteria (ARB) in the environment is a growing global threat to public health in the 21st century (Udikovic-Kolic et al, 2014; Wellcome Trust, 2018)
The present study show that chicken litter is a major reservoir of antibiotic-resistant E. coli that can be transferred to soil
The higher number of isolates with Multiple Antibiotic Resistance (MAR) index >0.2 in the litter-amended soil compared to the soil samples collected before the litter amendment indicates soil contamination with E. coli from sources with high use of antibiotics such as the chicken litter
Summary
The increasing prevalence of antibiotic-resistant bacteria (ARB) in the environment is a growing global threat to public health in the 21st century (Udikovic-Kolic et al, 2014; Wellcome Trust, 2018). A significant route by which ARB enter the environment and the food chain is through manure from antibiotic-treated animals applied to agricultural soil (Heuer et al, 2011; Marti et al, 2013). Chicken litter is often applied to agricultural soil as a substitute for inorganic fertilisers to meet the growing demand for crops and improve soil fertility, in organic farming (Jechalke et al, 2013; Marti et al, 2013; Atidégla et al, 2016). The application of chicken litter to agricultural soil as organic fertiliser is the cheapest means of disposing the large volumes of poultry waste generated from the rapidly growing poultry industry worldwide (Kyakuwaire et al, 2019). Chicken litter, a mixture of chicken faeces, waste feed, wood shavings, and other small invertebrates, is a major soil amendment that improves and maintains the chemical, physical, and biological soil properties (Brye et al, 2004)
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