Nigerian Veterinary Journal 39(3)
Total Page：16
File Type：pdf, Size：1020Kb
Nigerian Veterinary Journal 39(3). 2018 Asambe et al. NIGERIAN VETERINARY JOURNAL ISSN 0331-3026 Nig. Vet. J., September 2018 Vol 39 (3): 199 -208. https://dx.doi.org/10.4314/nvj.v39i3.3 ORIGINAL ARTICLE In Vitro Comparative Activity of Ciprofloxacin and Enrofloxacin against Clinical Isolates from Chickens in Benue State, Nigeria Asambe, A.1*; Babashani, M2. and Salisu, U. S.1 ¹.Federal University Dutsinma, Katsina State. 2.Ahmadu Bello University Zaria. *Corresponding author: Email: [email protected]; Tel No:+2348063103254 SUMMARY This study compares the in vitro activities of enrofloxacin and its main metabolite ciprofloxacin against clinical Escherichia coli and non-lactose fermenting enterobacteria isolates from chickens. Ten (10) Escherichia coli and 8 non lactose fermenting enterobacteriaceae species isolated from a pool of clinical cases at the Microbiology Laboratory of the Veterinary Teaching Hospital, University of Agriculture Makurdi were used in this study. Ten-fold serial dilution of 10 varying concentrations (0.1-50μg/mL) of enrofloxacin and ciprofloxacin were tested against the isolates in vitro by Bauer’s disc-diffusion method to determine and compare their antimicrobial activities against the isolates. The 18 isolates tested were susceptible to both enrofloxacin and ciprofloxacin, and their mean values in the susceptibility of Escherichia coli and non-lactose fermenters were significantly different (p < 0.01). The study concluded that the clinical isolates are susceptible to both enrofloxacin and ciprofloxacin though ciprofloxacin exhibit higher activity. Comparatively, ciprofloxacin was found to be more potent than enrofloxacin and the difference statistically significant. Ciprofloxacin was recommended as a better choice in the treatment of bacterial infections of chicken in this area compared to enrofloxacin. It was also recommended that proper steps should be taken in the administration of antimicrobials so as to reduce the incidences of bacterial resistance. Key words: Escherichia coli, Non-lactose fermenter, Antimicrobials, Ciprofloxacin. Enrofloxacin, Chicken. INTRODUCTION treatment of chronic respiratory and Fluoroquinolones are among the most gastrointestinal infections (Jelena et al., commonly used antibacterial drugs in 2006; Devreese et al., 2014; Vanni et al., general veterinary practice (Escher et al., 2014 Ruennarong et al., 2016). They also 2011; Kireewan and Suanpairintr, 2017). exhibit rapid bactericidal action against a Enrofloxacin and ciprofloxacin are synthetic wide variety of clinically important fluoroquinolones which acts by inhibiting microorganisms of human and animal origin the DNA gyrase enzyme and commonly (Jelena et al., 2006; Wayne et al., 2011). indicated in intensive poultry farming for the Though their usage in poultry have been 199 Nigerian Veterinary Journal 39(2). 2018 Asambe et al. restricted in some countries to avoid the problem to the poultry industry. The development of antimicrobial-resistant indiscriminate use of antimicrobials without bacteria, they are still of importance in the recourse to susceptibility testing is often treatment of human infections (Ferrari et al., attributed to be the major cause (Ramanan et 2015). However, ciprofloxacin usage al., 2013). particularly has persisted with arguments for The aims of this study therefore was to asses its use in animals intensified on the basis of and compare the antimicrobial activities of results of bacterial culture and antimicrobial ciprofloxacin and enrofloxacin against sensitivity testing demonstrating resistance clinical isolates to ascertaining the most to enrofloxacin but susceptibility to appropriate and desirable amongst the two ciprofloxacin (Sumano and Sunamo, 2001; thereby limiting the development of resistant Boothe et al., 2006). strains. Ciprofloxacin is a major metabolite of enrofloxacin used in human medicine MATERIAL AND METHODS (EMEA, 1998), but is only metabolised to a Escherichia coli and non-lactose limited range of 5–10 % in broiler chickens fermenting enterobacteria isolates (Redman, 2007; Slana et al., 2014). It also Ten Avian Escherichia coli and 8 non has similar spectrum of activity with lactose fermenting enterobacteriaceae enrofloxacin but with no reported effect species isolates were tested. The isolates against gram-positive bacteria (Slana et al., were collected from a pool of clinical cases 2014). from Veterinary Microbiology Laboratory of Ciprofloxacin and enrofloxacin are widely the Veterinary Teaching Hospital, used in Nigeria in the management and University of Agriculture, Makurdi. Proper treatment of poultry diseases. Several history of each flock including management reports point to the fact that the (mis)use of practices and previous treatment were noted. fluoroquinolone in chickens have resulted in Liver, spleen, kidney, lungs and bile higher incidences of bacteria resistance samples were collected from either (Abu-Basha et al., 2012; Devreese et al., moribund or dead birds during post-mortem 2014; Vanni et al., 2014). According to the examination and labelled individually. European Food Safety Authority (EFSA), The isolates were identified on the basis of (2014), Salmonella spp., Campylobacter culture, morphological and biochemical jejuni, Campylobacter coli and E. coli characteristics. On the basis of microscopic derived from domestic chickens’ resistance examination, morphology of bacteria was to ciprofloxacin stood at 37.3 %, 44.1 %, noted as rod, spiral or filament. It was 78.4 % and 57.6 %, respectively. differentiated by Biochemical Resistant bacteria from food animals can characterization as per Reynolds (2005). On spread to humans directly or indirectly cultural basis, MacConkey agar and Eosin- (Adenipekun et al., 2015; Rugumisa et al., methylene blue agar (EMBA) were used to 2016). This microbial resistance as well as confirm the identity of the E. coli isolates. the spread to human population are growing Swabs collected were directly inoculated and the outlook for the use of antimicrobial onto blood agar and MacConkey agar in drug in the near future is uncertain. This is a duplicates for every sample inoculum and significant public health concern when incubated at 37ºC for 24 hours. Similar animal husbandry practices promote colonies from growth observed were resistance to medically important antibiotics “Gram” stained and examined on the basis (Anderson et al., 2003; Rugumisa et al., of size, morphology and staining 2016). Several reports of outbreaks of characteristics. The Gram negative bacterial diseases of poultry are of public coccobacilli colony types were further health concern and have posed enormous characterized. 200 Nigerian Veterinary Journal 39(2). 2018 Asambe et al. On MacConkey agar only lactose susceptibility to the antimicrobials while fermenting (LF+) pink coloured colonies growth around the disc implies resistance. were isolated and sub cultured for further The diameters of the zones of inhibition as characterization to check whether the judged by an unaided eye were measured to bacteria are E. coli (i.e., there are other the nearest whole millimetre (mm) using a lactose fermenters like: Klebsiella and calibrated scale. The average diameters of Enterobacter). The LF+ colonies were the zones of inhibition were calculated and reinoculated on EMB agar for presence of result interpreted for each antibiotic by metallic sheen characteristics of E. coli, comparing to the standard chart which while non-lactose fermenting (LF-) represents the National Committee for colourless colonies were isolated and sub Clinical Laboratory Standards (NCCLS) cultured on Muller Hinton agar to obtain subcommittee’s recommendation for the pure cultures of non-lactose fermenting particular bacteria of interest. However, as enterobacteriaceae. Pure cultures of both the study was not designed to assess the isolates grown in nutrient broth were mixed incidence of resistance to the antimicrobial with sterile glycerol 1:1 and stored at agents, any isolate that was not sensitive to −20°C. an antimicrobial in the concentration range tested was deemed resistant and excluded Preparation of antibacterial drug stock from the analyses. solutions and dilution trays Standard ciprofloxacin and enrofloxacin Data analyses with 99 % purity were both sourced from The means were determined standard error Sigma-Aldriech, USA. The serial dilutions of mean (SEM). Mean difference between of the antimicrobial agents were prepared groups were compared using one-way from a stock solution of 10 varying analysis of variance (ANOVA) followed by concentrations (50 – 0.1 μg/ml) using Tukey’s post hoc test, while t-test was appropriate solvents with positive growth applied to compare the effect of the two control tubes without an antimicrobial agent fluoroquinolones at varying concentrations (Andrews, 2001). on all isolates at 5 % significant level (p ≤ 0.05) using the Statistical Package for Social Disc diffusion test Sciences (SPSS) version 20.0. The isolates were tested by the Kirby- Bauer’s disk-diffusion method as described RESULTS by Bauer et al. (1966). A lawn culture was The results from the determination of zones prepared using the primary inoculums by of inhibition by disc diffusion test showed spreading the inoculums onto the agar that Escherichia coli was susceptible to the surface nicely using a sterile glass spreader two antimicrobials at concentrations of (sterilized by 70 % alcohol). (12.50 - 50.00) μg/mL exhibiting larger After 15 minutes, ciprofloxacin and zones of inhibition