ABSTRACT

Cattle dung is used as organic fertilizer and alternative source of fuel or biogas. However, there is a public health concern as cattle dung could be a source of antibiotic resistant genes in the environment. Also microbiological and physicochemical characteristics of cattle dung might influence the properties of the receiving soils. This study isolated, identified and assessed antibiotic susceptibility pattern of bacteria from cattle dung and its contaminated soil as well as the physicochemical properties.

Sixty (60) samples of fresh cattle dung, cattle dung manure and cattle dung contaminated soil were collected from four (4) different cattle settlements/ markets located at Federal Staff road (location A), Dumez (location B), Enyaen (location C) and Upper Mission Road (location D) in Benin City, Edo State, Nigeria for this study. Control soil was collected outside the cattle settlement area. Physicochemical and heavy metals analyses of all samples were determined using standard methods. Bacteria isolation and identification were based on standard techniques while hemolytic activity was used to confirm potential pathogenic bacteria. Antibiotic susceptibility pattern of isolated pathogens were assayed by disk diffusion method and resistant genes qnrS, bla_TEM and bla_Zwere determined on resistant isolates using Polymerase Chain Reaction gel electrophoresis.

Physicochemical analyses showed significant difference in pH range (4.24±0.74to 4.74±0.12), manganese (35.99±2.08to 295.52±132.52) and copper (56.63±3.31 to 297.11±115.01) in cattle dung for various sites (P < 0.05). Moisture, pH and nitrate of dung manure were significantly different among sampling sites and the range were 34.58±11.25to 71.55±7.32, 4.07±0.23 to 4.86±0.20 and 30.51±4.32 to 69.63±9.86respectively. Conversely, moisture, pH, conductivity and phosphorus level of dung contaminated soil were significantly different among sampling sites (P < 0.05). High level of nitrate, phosphorus, ammonium, and potassium were recorded in all three samples. Apart from nickel and copper, heavy metal analyses from the three samples were within WHO range for soils. The highest total heterotrophic bacteria count was recorded in fresh cattle dung (2.0±0.3 x10⁶cfu/g) while the least count (0.23±0.04x10⁶) was recorded in cattle dung contaminated soil. Staphylococcus spp, Staphylococcus aureus, Bacillus spp, Escherichia coli, Pseudomonas spp, and Salmonella spp were isolated from all samples while Micrococcus spp, Nocardia spp, Neiseria spp and Serratia marcersens were only exclusive to cattle dung contaminated soil. Among isolated bacteria, Staphylococcus spp had highest occurrence of 23.8 % while Micrococcus spp was the least at 1.3 %. Hemolytic bacteria isolates were Staphylococcus aureus (16.5 %), Bacillus spp (17.4 %), Nocardia spp (4.6 %), Escherichia coli (29.4 %), Pseudomonas spp (13.8 %), Serratia marcersens (2.8 %) and Salmonella spp (15.6 %). High resistance (100 %) against Ampiclox (30 µg) was observed in all Staphylococcus aureus and Bacillus spp isolates while Pseudomonas aeruginosa isolates showed 100 % resistance to Ofloxacin (30 µg). Most Gram-positive bacterial isolates were majorly resistant to Beta lactams while Gram negative bacteria were resistant to Fluoroquinolones antibiotics. Multiple antibiotics resistant index (MARI) was measured at greater than 0.2, and was observed in 71.5 % of the hemolytic pathogens. Isolates from cattle dung samples were used for molecular analysis, Pseudomonas aeruginosa and Escherichia coli harbored quinolone resistance gene (qnrS), Staphylococcus aureus harbored bla_Zgene while Salmonella spp and Bacillus spp did not harbor qnrS and bla_TEMrespectively. Antibiotics resistance in hemolytic bacterial pathogens from this study is indicative of environmental sources of antibiotic resistance and possible adverse effects on human health. Horizontal gene transfer increases the risk of the spread of Antibiotic Resistant Gene (ARGs) from soil microorganisms to human pathogens.