Issues‎ > ‎vol17n3‎ > ‎

Antibiotic Resistance Pattern and Detection of Some ESBL Genes amongDiarrheagenic E. coli O157:H7.

Sivan Tariq Faiq1 & Zirak Faqe Ahmed Abdulrahman1

1Department of Biology, College of Education, Salahaddin University – Hawler, Iraq

During the period of 20th of May to1st of November 2013, twenty five isolatesof E. coli
O157:H7 were identified among 200 samples taken from children under ten years old
suffer from diarrhea (68 males and 132 females) admitted to Rapareen Pediatric hospitals
in Erbil City. Isolates were identified from stool sample by using cultural, morphological,
biochemical characteristics and serological test. Antibiotic sensitivity testing was done
for all isolates by using fourteen antibiotic types The resistance rate of the isolates
toward these antibiotics were 100 % for AMP, 96% for MET, 76% for CTX, 72% for TE
and KF, 68% for TMP, 56% for FEP, 52% for C and SXT, 48% for CRO, 28% for AK,
20% for CIP, while the lowest percent 16% was for CN, and all isolates were sensitive
for IPM. Extended spectrum ß-lactamase (ESBL) production was done for all isolates
.Out of 25
E. coli O157:H7isolates, 8(32%) were ESBL producers, while17(68%) were
non ESBL producers. The plasmid profile of 25 isolates conducted by using gel
the results revealed different bands with molecular weight ranging
between (2kbp- more than 10kbp) in size. The transformation process conducted
successfully for (E1 and E15) and the results showed that the resistance genes for
ampicillin, Cephalothin, Cefepime, metronidazole and trimethoprimwere located on
plasmid DNA for E1 isolate, while for E15 isolate, the ampicillin, Cephalothin,
Cefepime, ceftriaxone, Cefotaxime, metronidazole and trimethoprimlocated on plasmid.
The results of transformation confirmed by gel electrophoresis, and showed that one
plasmids of E1 and two plasmid E15 had been transformed successfully with molecular
weight of more than 10 Kbp. All
E. coli O157:H7isolates were screened for the presence
of ESBL genes including (
blaCTX-M - blaSHV-and blaTEM) on plasmid DNA using
polymerase chain reaction (PCR) assay. Results showed that all isolates were positive for
the existence of (
blaSHV-and blaTEM), while 88% of the isolates contained( blaCTXM) . 

Key Words: E. coli O157:H7 antibiotic resistance plasmid profile transformation ESBLs genes 


[1]Afset, J. E.; Bergh, K. and Bevanger, L. High prevalence of a typical enteropathogenic Escherichia coli (EPEC) in
Norwegian children with diarrhea. J. Med. Microbiol. 52:1015-1019, (2003).
[2] Tortora, G.J.; B.R. Funki and C.L. Case. Microbjology. 8th Ed., Pearson Education, Inc publishing as Benjamin
Cummings. (2004).
[3] Schlundt, J. Emerging food-borne pathogens. Biomed.Enviro. Sci. 14 (2): 44-52, (2001).
[4] Nataro J.P. and Kaper J.B. Diarrheagenic Escherichia coli.Clinical Microbiology Reviews, 11, 142–201, (1998).
[5] Levine, M. M., and R. Edelman. Enteropathogenic Escherichia coli of classic serotypes associated with infant
diarrhea:epidemiology and pathogenesis. Epidemiol.Rev. 6:31-51, (1984).
[6] Adamus- Bialek , W.; Zajac, E.; Parniewski, P. and Kaca, W. Comparison of antibiotic resistance patterns in
collections of Escherichia coli and Proteusmirabilisuropathogenic strains. MolBiol Rep., 40:3429–3435, (2013).
[7] Al-Faisal, A. Genetic engineering. Dar Al-shrooq for publishing Amman. (in Arabic) (1999).
[8] Hamad, I. S. Distribution of Conjugative Plasmids among Multiple Antibiotic Resistant E .coli Strains Isolated
from Patients of Teaching Hospital in Sulaimani City. J. Duhok Univ.12(1): 282-287, (2009).
[9] Silva, G.J. and Menduca, N. Laboratory of Microbiology and Center of Pharmaceutical Studies; Faculty of
Pharmacy; University of Coimbra; Coimbra, Portugal.Open Conference Proceedings Journal. Virulence 3(1):18–28,
[10] Jawetz, E. ; Melnick, J. L.; Adelberg, E. A.; Brooks, G. F. ; Butel, J. S. and Morse, S. A.Medical Microbiology .
23rd ed. McGraw-Hill, USA. (2004).
[11] Bali, E.B.; Acik, L. and Sultan N. Phenotypic and molecular characterization of SHV, TEM, CTX-M and
extended-spectrum β-lactamase produced by Escherichia coli, Acinetobacter baumanii and Klebsiella isolates in a
Turkish hospital. Afr. J. Microbiol. Res.; 4(8): 650–654, 2010.
[12] Lal, K.; Kapil, A.; Das, B.K.andSood S. Occurrence of TEM & SHV gene in extended spectrum β-lactamases
(ESBLs) producing Klebsiella sp. Isolated from tertiary care hospital. Indian J Med Res.; 125: 173–178, (2007).
[13] Wistreich, G.A. and Lechtman, M.D. Laboratory exercises in Microbiology 4th ed., Glencoe publishing
Co.,Inc.New York. (1980).
[14] Wayne, P.A. Clinical and laboratory standard institute. Performance standard for antimicrobial susceptibility
testing. 15 th Ed. Information supplement. CLSI/NCCLS M 100-S15.(2005).
[15] Collins, C.H.; Lyne, P.M.; Grange, J.M.; and Flalkinham, J.O. Collins &Lyne’s microbiological methods. 8th Ed.
London: Arnold. 95-183, (2004).
[16] Sambrook, J. and Russella, D.W. (2001). Molecular cloning: a laboratory manual.3rd Ed. Cold spring, Herbour lab.
New York.
[17] Casali, N. and Preston, A. E. coli plasmid vectors: methods and applications. Totwa, N.J., Huumana press. (2003).
[18] Jemima, S.A. and Vergese, S. Multiplex PCR for blaTEM & blaSHV in the extended spectrum beta lactamase
(ESBL) producing Gram-negative isolates. Indian J Med Res.; 128: 313–317, (2008).
[19] Eftekhar, F.; Hosseini- Mazinani, S. M.; Ghandili, S.; Hamraz, M. and Zamani, S.PCR detection of plasmid
mediated TEM, SHV and AmpC β-lactamases in community and nosocomial urinary isolates of Escherichiacoli.
Iranian J Biotech.; 3(1): 48-54, (2005).
[20] Sharma, S.; Bhat, G.K.; Shenoy, S. Virulence factors and drug resistance in Escherichia coli isolated from
extraintestinal infections. Indian J. Med. Microbiol. 25(4):369-373, (2007).
[21] Shamki, J.A. ;Al-Charrakh, A.H. and. Al-Khafaji, J.K. Detection of ESBLs in Enteropathogenic E. coli (EPEC)
Isolates Associated with Infantile Diarrhea in Kut City. University of Babylon, Hilla, Iraq. Medical Journal of
Babylon, 9( 2):10-15, (2012).

[22] Nanakaly, Zh. Gh. A. Characterization of plasmid DNA content and antimicrobial effect of bile salt on E.coli
isolated from urinary tract infection of pregnant women. These of Higher Diploma.College of Education.Salahaddin
University.Erbil.Iraqi, (2010).
[23] Salah H. F. Effect of some medicinal plant extacts on antibiotic resistance by plasmids of Escherichia coli isolated
from different sources in Erbil City. M.Sc. thesis, College of Science Education, Salahaddin University, Erbil,
[24] Al-Hilali, S. Occurrence and molecular characterization of enteropathogenEscherichia coli (EPEC) serotype
isolates from children with diarrhea in Najaf. M.Sc.Thesis, , College of Medicine. Kufa University.Iraq, (2010).
[25] Woodford, N. and Johnson, A.P. Molecular Bacteriology. Protocols and clinical applications. Humana press Inc.
[26] Decre' D.; Verdet C.; Raskine L.; Blanchard H.; Brughoffer B.; Philippon A.; Sanson-Le-Pors M. J.; Petit J. C.;
and Arlet G. Characterization of CMY-type β-lactamases in clinical strains of Proteus mirabilis and Klebsiella
pneumonia isolated in four hospitals in the Paris area. J AntimicrobChemother., 50: 681-688, (2002).
[27] Bekele, T. ;Zewde, G.; Tefera,G.; Feleke, A. and Zerom, K. Escherichia coli O157:H7 in Raw Meat in Addis
Ababa, Ethiopia: Prevalence at an Abattoir and Retailers and Antimicrobial Susceptibility. nternational Journal of
Food Contamination, 1(4):45-55, (2014)
[28] Rice, L.B.; Marshal, S.H. andCarias, L.L. Tn5381, a conjugative transposon identifiable as a circular from in
Enterococcus feacalis. J. Bacterio., 174:7308-7315, .(1992).
[29] Elder, J.S. Urologic disorders in infants and children. In: Behrman RE, Kliegman RM, Jenson HB and Stanton BE.
editors. Nelson Textbook of Pediatrics.17th ed. Philadelphia. Saunders. PP: 1621-1788, (2004).
[30] Jalal, K.C.A.; NurFatin, U.T.; Mardiana, M.A.; Akbar John, B.; Kamaruzzaman, Y.B.; Shahbudin, S. and Omar,
M. N. Antibiotic resistance microbes in tropical mangrove sediments in east coast peninsular, Malaysia. African
Journal of Microbiology Research; 4 (8): 640-46, (2010).
[31] Akata, F.; Tatman- Otcum, M.; Ozkan, E.; Tansel, O.; Otkum, M. and Tugrul, M. Prevalence by extended
spectrum beta lactamses produced by nosocomial isolates of enterobacteriaceae in Trakata University Hospital.
Turkey. New Microbial; 26:257-62, (2003).
[32] Sharma, J.; Sharma, M. and Ray, P. Detection of TEM & SHV genes in Escherichia coli&Klebsiella pneumoniae
isolates in a tertiary care hospital from India. Indian J Med Res ,132: 332-336, .(2010).
[33] Hassan, W.M.; Hashim, A. and Domany, R. Plasmid mediated quinolone resistance determinants qnr, aac(6′)-Ib-cr,
and qep in ESBL-producing Escherichia coli clinical isolates from Egypt. Indian J Med Microbiol ,30:442-447,
[34] Aibinu, I.E.; Peters, R. F.;Amisu, K.O. ; Adesida, S. A.; Ojo, M.O. and Odugbemi,T. Multidrug Resistance in
E.coli 0157 Strains and the Public Health Implication, Journal of American Science, 3(3): 34-42, (2007).
[35] Mahesh, E.; Ramesh, D.; Indumathi, V.A.; Punith, K.; Kirthi, R. and Anupama, H A. Complicated urinary tract
infection in a Tertiary Care Center in South India. Al Ameen J Med Sci; 3 (2):120 -27, (2010).
[36] Ramesh, N.; Sumathi, C.S.; Balasurbramanian, V.B.; Palaniappan, K.R. and Kannan, V.R. Urinary tract infection
and antimicrobial susceptibility pattern of extended spectrum beta lactamase producing clinical isolates. Advances
in Biological Research; 2 (5-6):78-82, (2008).
[37] Kjerulf, A.; Hansen, D.S.; Sandvang, D.; Hansen, F.; Frimodt-Møller, N. The prevalence of ESBL-producing E.
coli and Klebsiella strains in the Copenhagen area of Denmark. APMIS;116:118-24, .(2008).
[38] Wallensten, A.; Hernandez, J.; Ardiles, K.; DVM3, Gonza ́ lez-Acun ̃ a,D.; Drobni, M. and Olsen, B. : Extended
spectrum beta-lactamases detected in Escherichia coli from gulls in Stockholm, Sweden. Infection Ecology and
Epidemiology, 1(7):4-9,(2011).
[39] Nanakaly, Zh. Gh.A.. Molecular and Genetic study of uropathogenicEscherichia coli (UPEC) isolated from urinary
tract infection among children in Erbil City. M.Sc. Thesis, College of Medicine, Hawler Medical
University.Erbil.Iraqi. (2013).
[40] Kiratisin, P.; Apisarnthanarak , A.; Laesripa, C. and Saifon, P. Molecular characterization and epidemiology of
extended-spectrum β-lactamase producing Escherichia coli and Klebsiella pneumoniaeisolates causing health care-
associated infections in Thailand, where the CTX-M family is endemic. J AntimicrobChemother., 52(8): 2818-24,(2008).
[41] Singhal, S.; Mathur, T.; Khan, S.; Upadhyay, D.; Chugh, S. and Gaind, R. Evaluation of the methods for AmpC β-
lactamase in gram negative clinical isolates from tertiary care hospitals. Indian J Med Microbiol., 23(2):120-24,

[42] Bonnet, R. Growing group of extended spectrum ß-lactamases: the CTX-M enzymes. J Antimicrob and
Chemother., 48: 1-14, (2004).
[43] Mirelis, R.; Navarro, M.E.; Mesa, R.J.; Coll, P. and Parts, G.Community transmission of extended spectrum beta
lactamase. Emerg Infect Dis; 9:1024-25, .(2003).
[44] Al-Hamdani, M. A. and Hamad, I. G. Study of plasmid profile, susceptibility patterns of clinical Staphylococcus
aureus isolated from patients with otits media in Basrah. Journal of Basrah Researches Sciences; 38 (1) : 79- 89,(2012).
[45] Khan A.U., Sultan A, Tyagi A, Zahoor S , Akram M , KaurS.Amplification of mecA gene in multi-drug resistant
Staphylococcus aureus strains from hospital personnel. J Infect Developing Countries; 1(3): 289-295, .(2007).
[46] Rasmussent, J. W.; Jonsen, A.H. and Hoiby, N. Terminal truncations Amps ß-lactamase from a clinical isolate of
P.aeroginosa. Eur.J. Biochem., 263:473-485, (1999).
[47] Al-Alosi, A.N.H. Molecular study on some virulence factors produced by Gram-negative bacteria.M.Sc.thesis.
Genetic Enginnering and biotechnology Institute for post graduate studies .University of Baghdad. Baghdad. Iraq,(2004).
[48] Robinson, J. B. and Tuovinen, O. H. Microbiol. Rev. 48, 95-124, (1984).
[49] Clowes, R. molecular structure of bacterial plasmids.Bacteriology Review, 36:361, (1972).
[50] Al-Sorchee S. M. A. Causative agents of diarrhea in Erbil children and the effect of some plant extracts on
bacterial isolates.Ph.D. Thesis, College of Education, Ibn Al-Haitham University of Baghdad, Iraq, (2009).
[51] Yang, Y.; Bhachech, N.; Bradford, P. A.; Jett, B. D.; Sahm, D.F. and Bush, K. Ceftazidime-resistant Klebsiella
pneumoniae and E. coli isolates producing TEM-10 and TEM-43 b-lactamases from St. Louis. Antimicrobial Agents
and Chemotherapy, 42(12): 1671-1676, (1998).

[52] Brown, J.R.; Daniel G.; Julie A.; Ingraham B.K.; David J.H. and Stanhope M.J.Horizontal gene transfer of drug-
resistant amino-acyl-transfer- RNA synthetases of anthrax and Cram-positive. EMBO Reports, 4(7): 692-698, (2003).
[53] Nwanze, P. I.; Nwaru, L. M.; Oranusi, S.; Dimkpa, U.; Okwu, M. U.; Babatunde, B. B.; Anake, T. A.; Jatto,W. and
Asagwara, C. E. Urinary tract infection in Okada village: Prevalence and antimicrobial susceptibility pattern.
Scientific Research and Essay .2 (4).112-116, (2007).
[54] Noor, N.; Ajaz, M.; Rasool, S.A. and PirzadaZ.A.Uriary tract infections associated with multidrug resistant bacilli:
characterization and genetical studies. Pak.J. Bio. Sci., 17:115-123, (2004).
[55] Lennon, E. and DeCicco, B.T. Plasmids of Pseudomonas cepacia strains of diverse origin. Appl. Environ.
Microbiol. 57: 2345-2350, (1991).
[56] Al-Otrqchi, K.I. Molecular genetic studies of bacterial species isolated from patient with diarrhea. Ph.D. Thesis.
College of medicine-Hawler medical university-Erbil.Iraq, (2006).
[57] Ruppé, E.; Hem, S.; Lath, S.; Gautier, V.; Ariey, F.; Sarthou, J.; Monchy, D. and Arlet, G. CTX-M β-Lactamases
in Escherichia coli from Community-acquired urinary tract infections, Cambodia. Emerging Infectious Diseases.
15( 5):112-118, (2009).
[58] Suzuki, S.; Shibata1, N.; Yamane,K.; Jun-ichiWachino, J.; Ito,K. and Arakawa,Y. Change in the prevalence of
extended-spectrumb- lactamase-producing Escherichia coli in Japan by clonal spread. Journal of Antimicrobial
Chemotherapy (2009) 63, 72–79, .(2009)
[59] Shahid, M.; Sobia,F. ; Singh,A. and Khana, H. M. Concurrent Occurrence of blaampC Families and blaCTX-M
Genogroups and Association with Mobile Genetic Elements ISEcp1, IS26, ISCR1, and sul1-Type Class 1 Integrons
in Escherichia coli and Klebsiella pneumoniae Isolates Originating from India. Journal of Clinical Microbiology p.
1779–1782, (2012).
[60] Jacoby, A.G. and Munoz-Price, L.S. Mechanismsof disease the new β-lactamase. N Engl J Med.; 325: 380-91,(2005).
[61] Canton, R. and Coque, T.M. The CTX-M beta-lactamase pandemic.Curr.Opin.Microbiol., 9: 466-475, (2006).
[62] Naseer U, Sundsfjord A. The CTX-M conundrum: dissemination of plasmids and Escherichia coli clones. Microb
Drug Resist. Mar; 17(1):83-97, (2011).
[63] Goyal, A. ; Prasad, K. N.: Prasad, A.; Gupta, S. ; Ghosha, U. and Ayyagari, A. Extended spectrum β-lactamases in
Escherichia coli & Klebsiellapneumoniae & associated risk factors. Indian J Med Res. 2009; 129: 695–700, (2009).
[64] Kaftandzieva, A.; Trajkovska-Dokic, E.andPanovski, N. Prevalence and molecular characterization Of extended
spectrum beta-lactamases (ESBLs) producing Escherichia coli and Klebsilla pneumonia. Contributions, Sec. Biol.
Med. Sci., 8(2) :129–141, (2011).