INTERNATIONAL JOURNAL OF BIOASSAYS ISSN: 2278-778X CODEN: IJBNHY ORIGINAL RESEARCH ARTICLE OPEN ACCESS Prevalence of multidrug resistance in Escherichia coli strains isolated from river Yamuna, Delhi stretch Richa Bhardwaj, Anshu Gupta and J. K. Garg* University School of Environment Management, Guru Gobind Singh Indraprsatha University, Sector-16-C, Dwarka-110078, New Delhi, India. Received for publication: September 13, 2015; Revised: September 27, 2015; Accepted: October 10, 2015 Abstract: River water can act as a sink of multidrug resistant strains that may have serious public health implications. The aim of the study was to assess the antibiotic susceptibility and multidrug resistance character of twenty-eight Escherichia coli strains that have been isolated from water samples collected from the River Yamuna in Delhi stretch, India. The isolates were subjected to antibiotic sensitivity test by Kirby Bauer disc diffusion method as per Clinical and Laboratory Standard Institute (CLSI) guidelines using 24 different antibiotics belonging to three different modes of action namely β-lactams, aminoglycosides, phenicols, tetracyclines and quinolones. Most evident results of the study were that none of antibiotics used in the study was 100% effective. Findings have revealed that 100% of the isolates exhibited multi drug resistance (MDR) character and all the isolates were having a very high multiple antibiotic-resistance (MAR) index, suggesting the origin of the isolates to be of high antibiotic usage. Presence of multi drug resistant E. coli in river water can act as a medium to spread antibiotic resistance to other bacteria. The current attempt towards identification of existence of multidrug resistant bacteria isolated from the river Yamuna is a rather timely event which may find utility in the development of strategies to counteract this problem by incorporating the use of antibiotic resistance as a bacteriological water quality parameter. Key words: River Yamuna; Coliforms; Escherichia coli; Antibiotics; Multidrug resistance INTRODUCTION Antibiotics are the biologically active compounds them ideal indicator organisms to test environmental of natural or synthetic origin, which are widely used to conditions for fecal contamination. The presence of E. coli in prevent or treat infections in humans, animals and food- water is an indicator of fecal contamination and also producing insects and plants43. However, the emergence and represents the existence of all possible types of pathogenic spread of antibiotic resistance has emerged as an issue of bacteria transmittable through feces22. major concern worldwide29 as the development of resistance for antibiotics in bacteria will make the use of these It must be noted that commensal E. coli in human antibiotics ineffective. A wide range of biochemical and gut can act as reservoirs of antibiotic resistance genes which physiological mechanisms may be responsible for resistance. might be rapidly transferred to other commensal or The abuse of antibiotics in human medicines, animal pathogenic bacteria42, 9. During the fecal contamination of treatment and agriculture combined with inadequate water, antibiotic resistant bacteria harboring resistance genes wastewater treatment has led to the presence of antibiotics present a potent threat to human health. Therefore, E. coli is and antibiotic resistant bacteria in the environment a useful indicator of the spread of acquired antibiotic particularly in the surface waters20, 7. Subsequently, it has led resistance genes in both commensal as well as in the to the development of multiple drug resistance in many environmental bacterial communities11. bacterial species15, 38. Several studies have reported the growing Bacterial contamination of surface waters has long antibiotics resistance in clinical as well as environmental been a major water quality issue due to potential for disease isolates of E. coli in both developed and developing transmission. River water acts as a medium where bacteria countries19, 2, 46, 18. These studies have indicated that E. coli (both pathogenic and non-pathogenic) from different exhibits high rate of resistance as well as MDR against sources like human, animal and environment (wastewater clinically significant antibiotics. The studies also suggested plants, urban or industrial effluents, agricultural run-off) mix that the MDR is not congenital feature of E. coli but high together and this may result in exchange of antibiotic and uncontrolled use is closely related to resistance resistance genes among them. Furthermore, it may act as a development. Some studies have also provided the source of dissemination of antibiotic resistant mechanisms of development of antibiotic resistance21, 1, 28, 5. microorganisms among human and animal populations, if contaminated water is used for drinking purpose17, 7. Occurrence and prevalence of antibiotics and multidrug resistant E. coli from various sources has been The microbiological safety of drinking water is investigated in India by a few researchers13, 45, 41, 40. However, generally assessed using microbial indicators called fecal resistance status of E. coli in the River Yamuna in Delhi coliform bacteria and in particular E. coli. E. coli is a Gram- stretch is inadequately studied so far. The present negative, rod-shaped bacterium that is commonly found in investigation was aimed to determine the antibiotic the lower intestine of warm-blooded organisms. It is also resistance pattern of the E. coli isolated from the River found in water, soil and vegetation. The genus is a member Yamuna, and to determine the existence of Multidrug of the Enterobacteriaceae family of the resistant strains based on the criteria established by Gammaproteobacteria13. E. coli cells are able to survive Magiorakos et al., 31. outside the body for a limited amount of time, which makes *Corresponding Author: Prof. J. K. Garg, Professor, University School of Environment Management, Guru Gobind Singh Indraprsatha University, Sector 16-C, Dwarka-110078, New Delhi, India. 4492 Richa Bhardwaj et al., International Journal of Bioassays 4.11 (2015): 4492-4498 MATERIALS AND METHODS 48 h. The positive tubes showing the formation of turbidity Study Area and gas were subjected to confirmative test. In confirmative The River Yamuna is the largest tributary of the test, the tubes showing positive results in presumptive test River Ganga that originates from Yamunotri glacier and has were inoculated into EC-MUG media followed by a total length of 1,376 kilometers till its confluence with the incubation at 44°C for 24 h in an incubator (Kuhner, River Ganga in Allahabad, India. Delhi stretch of the River Switzerland). All the tubes showing formation of blue color Yamuna is 22 km long and it is located between Wazirabad in EC-MUG media under UV light were recorded as MPN and Okhla Barrage. Central location of the river corresponds count of E. coli. Simultaneously, the cultures from positive to 28º36´ N and longitude 77º12´ E, at an altitude of 216 m lactose broth tubes were streaked onto Eosin Methylene above mean sea level12. The river is a major source of Blue Agar. Isolates having green metallic sheen were further potable water to Delhi, which unfortunately also receives, streaked and re-streaked onto MacConkey Agar till pure untreated and partially treated domestic, agricultural and colonies were obtained. One bacterial isolate per site and per industrial wastes from the mega-metropolis city of Delhi33. sampling was selected randomly from pure colonies growing on MacConkey Agar and was preserved on nutrient agar Sampling Sites slants at 4°C for further studies. Table 1: Description of study sites Site Code Site Name Location Description Identification of E. coli by morphological and Wazirabad This site is the entry A 28° 43’23” Latitude north and biochemical tests: The shape and color of each bacterial Barrage point of River Yamuna isolate was examined under the microscope after Gram 77°14’40” Longitude east in Delhi Najafgarh staining according to Bergey’s Manual of Systematic B 28° 42’40” Latitude north and It is the major polluting Drain Bacteriology16. Further confirmation of Escherichia species drain of River Yamuna (Outlet in the 77°13’55” Longitude east in Delhi was done using HiIMVic conventional biochemical test kit. river) This kit uses four conventional biochemical tests (Indole, Old Yamuna This is a place where C 28° 39’36” Latitude north and Iron Bridge there are many Methyl red, Voges Proskauer’s and Simmons citrate) and inhabitants by the eight carbohydrate tests (Glucuronidase, Nitrate reduction, riverbank and it reflects 77°14’59” Longitude east Ortho-Nitrophenyl-β-galactoside, Lysine utilization, Lactose, the impact of fecal Glucose, Sucrose, Sorbitol). discharge D ITO Barrage 28° 37’49” Latitude north and It is the intermediate point having high Description of Antibiotics: As per the Clinical 77°15’20” Longitude east amount of pollution and Laboratory Standards Institute (CLSI) standards, Nizamuddin This is a place where E 28° 36’20” Latitude north and twenty-four antibiotics belonging to ten different antibiotic Bridge there are few inhabitants with a visible presence of classes were used to check the susceptibility and resistance 77°15’44” Longitude east small houses pattern of E. coli. The antibiotics selected are those that are Okhla It is the exit point of F 28° 32’44” Latitude north and commonly used for medication against E. coli. Antibiotic Barrage River Yamuna from μg 77°18’57” Longitude