in-vitro comparative study of cefoperazone, ceftazidime, ceftizoxime ...

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Dec 11, 2012 - members of third generation cephalosporins (Cefoperazone, Ceftazidime, Ceftizoxime, Cefotaxime, Ceftriaxone and Cefixime) were reviewed.
Humza Ahmad Ullah et al. IRJP 2013, 4 (1) INTERNATIONAL RESEARCH JOURNAL OF PHARMACY www.irjponline.com

ISSN 2230 – 8407 Research Article

IN-VITRO COMPARATIVE STUDY OF CEFOPERAZONE, CEFTAZIDIME, CEFTIZOXIME, CEFOTAXIME, CEFTRIAXONE AND CEFIXIME AGAINST PSEUDOMONAS AERUGINOSA Humza Ahmad Ullah1*, Iqbal Javeid1, Khan Khalid1, Hanif Muhammad 2, Jamil Sahrish 3 1Faculty of Pharmacy, Hamdard University, Karachi, Pakistan 2Department of Pharmacy, GC University, Faisalabad, Pakistan 3Faculty of Pharmacy, Federal Urdu University, Karachi, Pakistan Article Received on: 08/11/12 Revised on: 11/12/12 Approved for publication: 23/12/12

*Email: [email protected] ABSTRACT The prime intention of this study was the evaluation & accumulation of epidemiological data on the resistance of Pseudomonas aeruginosa, and to compare the activity of different third generation cephalosporins against Pseudomonas aeruginosa. For this purpose Modified Kirby-Bauer Method was used for the determination of sensitivity of antibacterial agents using strains of Pseudomonas aeruginosa ATCC 27853 as control. Total 250 isolates of Pseudomonas aeruginosa were collected from different public and private hospitals of Karachi, Pakistan. In-vitro qualities (i.e. sensitive, resistant and intermediate) of six members of third generation cephalosporins (Cefoperazone, Ceftazidime, Ceftizoxime, Cefotaxime, Ceftriaxone and Cefixime) were reviewed. Results showed that Cefoperazone was the most effective antibacterial agent (80% sensitive), while the second most effective antibacterial agent was Ceftazidime (70% sensitive). Cefotaxime and Ceftizoxime also showed intermediate activity. Cefixime and Ceftriaxone didn’t show any supportive activity i.e. 0% sensitive at all. KEY WORDS: Third generation cephalosporins, Pseudomonas aeruginosa, Susceptibility test.

INTRODUCTION Pseudomonas aeruginosa (Ps. aeruginosa) is termed as an opportunistic pathogen belonging to the family Pseudomonadaceae which have a large number of species and are able to cause opportunistic infections in human1. The Ps. aeruginosa is aerobic rods, non-fermentative, gramnegative motile and water soluble pigments producer. In laboratory, these characteristics are used for rapid diagnosis 2. Ps. aeruginosa can be isolated from the feces of healthy people and also found sporadically on the human body sites which are moist i.e. ear, axilla, throat, nasal mucosa. In hospitals; mops, sinks, respiratory equipment, food mixers and disinfectant solutions are reservoir of Ps. aeruginosa. It extensively causes nosocomial infections like nosocomial pneumonia, nosocomial urinary tract infections, nosocomial bacteremia and wounds3. The pattern of antibiotic resistance of Ps. aeruginosa isolates varied from hospital environment to geographical locations4. Several resistance mechanisms may be developed by Ps. aeruginosa against different antibiotics, beta-lactamase production is the major mechanism of resistance to beta-lactam antibiotics. Mostly, third generation cephalosporins are hydrolyzed by ESBL enzymes5. Also the defected methyl directed mismatching repair (MMR) system in the hypermutable strains of Ps. aeruginosa is being isolated frequently from cystic fibrosis (CF) patient’s lung1. In cystic fibrosis patients, it has been involved in the formation of biofilms, a unique characteristic to develop antibiotics resistance 2. Cephalosporins may be classified by their clinical pharmacology , chemical structure, antimicrobial spectrum or resistance to beta-lactamase, but the classification by generation is well accepted, very useful and based on basic features of antimicrobial activity6. Among all third generation cephalosporins, Cefoperazone and Ceftazidime are the most effective against Ps. aeruginosa and, they are also stable to breakdown effect of most beta-lactamases7. Ceftazidime is very effective to treat pediatric patients and nosocomial infections8. Ceftriaxone is unique because of its prolonged serum half-life, which permits once or twice daily dosing 9.

This study was designed to determine the In-vitro susceptibility of Ps. aeruginosa isolated from clinical specimens in Karachi, Pakistan for the analysis of third generation cephalosporins. Modified Kirby-Bauer disk diffusion technique was used to determine susceptibility of isolates10. Objective The prime intention of this study was to evaluate and accumulate the epidemiological data on the resistance of Ps. aeruginosa within the community and to compare the activity of third generation cephalosporins against Ps. aeruginosa. MATERIALS & METHODS Three Biological culture media were used i.e. Mueller-Hinton Agar (Batch No. CM0337, IVD, Oxoid, England), MuellerHinton Broth (Batch No. CM0405, IVD, Oxoid, England), and Blood Agar (Oxoid, England). Antibiotics discs used in this experiment were: Cefoperazone (CFP) 75µg, Cefixime (CFM) 5µg, Ceftazidime (CAZ), Ceftizoxime (ZOX), Cefotaxime (CTX), and Ceftriaxone (CRO) each were of 30µg. These discs were commercially purchased from Oxoid Ltd, England. Bacterial Cultures Isolates of Ps. aeruginosa were collected from central laboratories of different public and private hospitals of Karachi, Pakistan. They were isolated on nutrient agar slant and transported under cooled condition. Sub culturing of isolates were done on Media (Mueller Hinton Broth). Identification of Bacterial Isolates The identification of isolates was done on the basis of cultural characteristics, Gram staining and biochemical tests11. Susceptibility Test The test was performed using modified Kirby-Bauer disc diffusion method according to the guidance of the Clinical and Laboratory Standards Institute12 10. Mueller-Hinton agar was used as the growth medium in disk susceptibility test while inoculum was compared with 0.5 McFarland standard turbidity to adjust inoculum density13. Page 135

Humza Ahmad Ullah et al. IRJP 2013, 4 (1) Reading and Interpretation The diameter of each zone of inhibition was measured by a ruler, and results were reported as susceptible, intermediate and resistant to the agents that were tested. Data was S.No.

Antibiotics

1 2 3 4 5 6

Cefoperazone Ceftazidime Cefotaxime Ceftizoxime Ceftriaxone Cefixime

interpreted according to CLSI (Table 2B-1) Ver. 201012. Diameters of Zones of Inhibition for each antibiotic were also obtained for the control strains Ps. aeruginosa ATCC 27853 to ensure that the method performed correctly.

Table 1. Measured Zone of Inhibition against Ps.aeruginosa Code Resistance Intermediate (R) (I) CFP 10% 10% CAZ 11% 19% CTX 3.3% 80% ZOX 20% 60% CRO 57% 43% CFM 100% 0% (w.r.t percentages Resistance, Intermediate & Sensitivity)

Sensitive (S) 80% 70% 16.8% 20% 0% 0%

Image: I. CTX

Image: II. CFP

Image: III. CRO

Image: IV. CAZ

CFP= Cefoperazone; CAZ=Ceftazidime; CTX= Cefotaxime; CRO= Ceftriaxone;

RESULTS & DISCUSSION Zone of Inhibition was measured by disk diffusion method, Cefoperazone (CFP) was found to be the most effective, having 80% antibacterial activity against Ps. aeruginosa, due to highest sensitivity in 200 isolates while Cefoperazone (CFP) was also found to be 10% sensitive as shown in table 1. Similar results were also reported by Farida Anjum et al., in 2010 after studying Cefoperazone against Ps. aeruginosa, they reported 60% antibacterial activity5. The second most effective antibacterial agent against Ps. aeruginosa was Ceftazidime (CAZ) which showed 70% effect in 175 isolates. Ceftazidime was also studied by Saleem Hafeez et al., in 2000 and reported 80 % sensitivity against Ps. aeruginosa14. Similar findings were also studied by Tahira et al., in 2009 and reported 89% sensitivity8 while Farida et al., in 2010 reported 62% sensitivity against the same pathogen5. Results revealed that Cefotaxime (CTX) was the third choice of drug and found to be effective in 42 isolates i.e., 16.8% effective against Ps. aeruginosa as shown in table.1. Similar

antimicrobial activities was also studied by J Puri et al, in 1996 regarding CFP and CAZ15. The least choice of drug was Ceftizoxime (ZOX) which showed 20% effectiveness while Hafeez et al., in 2000 reported 16% activity against Ps. aeruginosa (14). Cefixime (CFM) and Ceftriaxone (CRO) were not effective against all isolates but lateral was considered 21% effective by Tahira et al., in 20098. Ps. aeruginosa showed resistance against Cefixime and Ceftriaxone. Hence frequent measures should be taken to control the problem of resistance. Continually updated and validated antimicrobial susceptibility profiles data are necessary to ensure the provision of effective and safe empiric therapy. Antibiotic resistance is considered due to many biochemical mechanisms like presence of beta-lactamase resistance in Ps. aeruginosa which is generally initiated by a chromosomal class I beta lactamase or a plasmid mediated beta-lactamase. The process of resistance to the cephalosporins (third generations) usually originated through a mutation that Page 136

Humza Ahmad Ullah et al. IRJP 2013, 4 (1) results in potentially depressed instead of inducible population. Cefoperazone is a broad spectrum third generation cephalosporin having potent activity against Ps. aeruginosa. It acts upon PBPs to stop peptidoglycan synthesis in bacteria (both gram-positive and gram-negative). The responsible factors for resistance to Cefoperazone are enzymatic hydrolysis, altered PBPs affinity and permeability barrier present in the membranes of gram negative bacteria15. Reported comparative studies of antibiotics have some limitations of short period of study designs, hence better results would be achieved by long term studies in collaboration of governmental and non-governmental organizations. In third world countries like Pakistan where the total health budget is less than 1% of GDP, cannot afford the parallel studies of antibiotic resistance problems like this16. CONCLUSION The results of this study provide useful and effective guidelines for choosing an appropriate antibacterial agent against infections caused by Pseudomonas aeruginosa. The drug of choice is Cefoperazone (CFP) while the second choice may be Ceftazidime (CAZ) while Cefotaxime (CTX) is also considered for the third line of therapy. ACKNOWLEDGEMENT We (authors) are very thankful to Faculty of Pharmacy, Hamdard University Karachi, Pakistan for providing us such facilities to conduct this research work. REFERENCES 1. S.G. Nadeem SAQ, F. Afaque, M. Saleem and S.T. Hakim. Comparison of the in vitro susceptibility of clinical isolates of Pseudomonas aeruginosa in a local hospital setting in karachi, Pakistan. BJMP 2009;2(4):35-39. 2. Luqman Satti SA, Tanveer Ahmed Qumar,Muhammad Shoaib Khan, and Zahid Ahmed Hashmi. In Vitro efficacy of Cefipime against multi-drug

resistant Pseudomonas aeruginosa- An alaraming situation in our setup. The Open Drug Resistence Journal 2011;1:12-16. 3. G. M. Rossolin EM. Treatment and control of severe infections caused by multiresistant Pseudomonas aeruginosa. Clin Microbiol Infect 2005;11(4):17-32. 4. Piyush Tripathi GB, Shivani Saxena, Mahendra Kumar Gupta, and P. W. Ramteke. Antibiotic resistance pattern of Pseudomonas aeruginosa isolated from patients of lower respiratory tract infection. African Journal of Microbiology Research 2011;5(19):2955-2959. 5. Farida Anjum AM. Susceptibility pattern of pseudomonas aeruginosa against various antibiotics. African Journal of Microbiology Research 2010;4(10):1005-1012. 6. Najma Sultana MSA. In Vitro activity of Cefadroxil, Cephalexin, Cefatrizine, and Cefpirome in presence of essential and trace elements. Pak. J. Pharm. Sci. 2007;20(4):305-310. 7. MARSH TD. Clinical pharmacology of Antibiotics: The Cephalosporin Antibiotic Agents: III. Third-generation Cephalosporins. Infection control 1985;6:78-83. 8. Tahira Mansoor MAM, Gulnaz Khalid, Mustafa Kamal. Pseudomonas aeruginosa in chronic suppurative otitis media: sensitivity spectrum against various antibiotics in karachi. J Ayub Med Coll Abbottabad 2009;21(2):120123. 9. Hakim Masood SBN, Nosheen Aslam. Cost effective analysis of different brands of ceftriaxone available in karachi, Pakistan. Pakistan Journal of Pharmacology 2008;25(2):13-19. 10. Performence standards for antimicrobial disk susceptibility tests; Approved standards. The Clinical and Laboratory Standards Institutes (CLSI), 10th edn. Jan 2009;29(1):M02-A10. 11. Barrow GIaF, R. K. A. Characters of Gram-negative Bacteria. In: Cowan & Steel Manual for Identification of Medical Bacteria. 2003;3rd edi. Cambridge, UK.(Ch 06.):130-131. 12. CLSI. Performing Standards for Antimicrobial Susceptibility Testing. Clinical Laboratory Standards Institute 2010 30:M100-S20. 13. Andrews JM. Determination of minimum inhibitory concentrations. Journal of Antimicrobial Chemotherapy 2001;48(S1):5-16. 14. Saleem Hafeez MI, Altaf Ahmed, Afia Zafar, Muhammad Naeem. In Vitro Antimicrobial Activity of Cefpirome:a new fourth generation Cephalosporin against clinically significant Bacteria. JPMA 2000;50(250). 15. J Puri GR, P Kundra, V Talwar. Activity of third generation cephalosporins against pseudomonas aeruginosa in high risk hospital units. Indian Journal of Medical Sciences 1996;50(7):239-243. 16. Khan MAFJ. Health Care Services and Government Spending in Pakistan, PIDE-Working Papers. Pakistan Institute of Development Economics. 2007;32:1-24.

Source of support: Nil, Conflict of interest: None Declared

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