[01] Narmin S. Merza, Scientific Research Center, Duhok University, Duhok, Iraq. [02] Jaladet M. S. Jubrael, Scientific Research Center, Duhok University, Duhok, Iraq.

A total of 364 clinical isolates of Escherichia coli were collected and characterized from different hospitals in Kurdistan region. Two hundred ninety six of these were isolated from female accounting (81%) whereas 68(19%) of them were recovered from male. One hundred fifty isolates of UPEC (Fifty from each province) were randomly selected and confirmed as E. coli isolates at molecular level by PCR using uidA primer as a species-specific primer producing 670bp DNA fragment for uidA gene. They were assigned into one of the major phylogenetic groups (A, B1, B2 and D). This is based on the presence or absence of genetic markers including: two genes (chuA and yjaA) and a DNA fragment (TSPE4.C2) as proposed by Clermont et al., 2000. It has been found that 85 (56.7%) of these isolates belonged to phylogenetic group B2, 31 (20.7%) belonged to group D, 27 isolates were phylo-group (A) a counting (18%), and only 7 isolates (4.7%) belonged to group B1. Further classification has been done in order to increase the discrimination power of E. coli population analyses by classification these phylogroups into seven subgroups including, A0, A1, B1, B22, B23, D1and D2. Phylogenetic group B2 showed two types of gene patterns. Seven isolates belonged to B22 and 77 isolates designated as B23. In addition to unusual strain (-++) which assigned to phylogroup B2. Phylogenetic group D revealed two gene patterns; it has been found that 11 of them were belonging to subgroup D1 and 20 isolates belonged to sub group D2. In the third phylogroup A, only 2 isolates belonged to subgroup A0 and 25 isolates belonged to subgroup A1. Out of 7 isolates which belonged to Phylogroup B1, 2 isolates had unique pattern and assigned as B1a. From these results it have been found that B23 (+++) was the most predominant among all other subgroups accounting (77/150) 51.3%, while subgroup A0 (---) was the least prevalent among all others accounting (2/150) 1.3%.

Phylogenetic Grouping, Triplex PCR, Escherichia coli

[01] Adamus-Bialek, W.; Wojtasik, A.; Majchrzak, M.; Sosnowski, M., and Parniewski, P. (2009). (CGG) 4-Based PCR as a Novel Tool for Discrimination of Uropathogenic Escherichia coli Strains: Comparison with Enterobacterial Repetitive Intergenic Consensus-PCR. Journal of Clinical Microbiology, 47(12): 3937–3944. [02] Bashir, S.; Haque, A.; Sarwar, Y.; Ali, A., and Anwar, M. I. (2012). Virulence profile of different phylogenetic groups of locally isolated community acquired uropathogenic E. coli from Faisalabad region of Pakistan. Annals of Clinical Microbiology and Antimicrobials, 11(23):1-6. [03] Bergthorsson, U., and Ochman, H. (1998). Distribution of chromosome length variation in natural isolates of Escherichia coli. Mol Biol Evol, 15 (1): 6-16. [04] Carlos, C.; Pires, M. M; Stoppe, N. C.; Hachich, E. M.; Sato, M. I.; Gomes, T. A.; Amaral, L. A., and Ottoboni, L. M. (2010). Escherichia coli phylogenetic group determination and its application in the identification of the major animal source of fecal contamination, BMC Microbiology, 10:161. [05] Cheesbrough Monica (2006) District Laboratory Practice in Tropical Countries,. part2, 2nd Ed. Cambridge University, ISBN-13 978-0-511-34842-6. [06] Clermont, O.; Bonacorsi, S., and Bingen, E. (2000). Rapid and simple determination of the Escherichia coli phylogenetic group. Appl Environ Microbiol.; 66(10):4555-4558. [07] Clinical and Laboratory Standards Institute (2007). Performance standards for antimicrobial susceptibility testing; seventeenth informational supplement. M100-17. Clinical and Laboratory Standards Institute, Wayne, PA. [08] Daly, P.; Collier, T., and Doyle, S. (2002). PCR-ELISA detection of Escherichia coli in milk. Letters in Applied Microbiology, 34:222–226. [09] Dash, M.; Padhi, S.; Mohanty, I.; Panda, P., and Parida, B. (2013). Antimicrobial resistance in pathogens causing urinary tract infections in a rural community of Odisha, India, J Family Community Med., 20(1): 20–26. [10] Fadel Al- Deen, I. H. (2012). Molecular Characterization of Escherichia coli Isolated from UTI Patients Attending Azadi Hospital in Duhok City. M. Sc. Thesis, Collage of Education, Duhok University. Iraq. [11] Feng, P.; Lum, R., and Chang, G. W. (1991) Identification of uidA Gene Sequences in 3-D-Glucuronidase-Negative Escherichia coli. Applied and Environmental Microbiology, 57(1): 320-323. [12] Foxman, B. (2003). Epidemiology of urinary tract infections: incidence, morbidity, and economic costs, Disease a Month, 49: 53–70. [13] Giray, B.; Uçar, F. B., and Aydemir, S. Ş. (2012). Characterization of uropathogenic Escherichia coli strains obtained from urology outpatient clinic of Ege Medical Faculty in İzmir. Turk J Med Sc., 42 (1): 1328-1337. [14] Gordon David M., Clermont Olivier, Tolley Heather and Denamur Erick (2008) Assigning Escherichia coli strains to phylogenetic groups: multi-locus sequence typing versus the PCR triplex method. Environmental Microbiology, 10(10): 2484–2496. [15] Gupta, K.; Hooton, T. M.; Naber, K. G.; Wullt, B.; Colgan, R.; Miller, L. G.; Moran, G. J.; Nicolle, L. E.; Raz, R.; Schaeffer, A. J., and Soper, D. E. (2011). International clinical practice guidelines for the treatment of acute uncomplicated cystitis and pyelonephritis in women: A 2010 update by the Infectious Diseases Society of America and the European Society for Microbiology and Infectious Diseases. Clin Infect Dis., 52(5):103-120. [16] Hilbert, D. W. (2011). Antibiotic Resistance in Urinary Tract Infections: Current Issues and Future Solutions. Tenke Peter (2011) Urinary Tract Infections, chapter11, 193-208, ISBN: 978-953-307-757-4. [17] Hodiwala, A.; Dhoke, R., and Urhekar, A. D. (2013). Incidence of Metalo-Beta- Lactamse producing Pseudomonas, Acinetobacter and Enterobacterial isolates in Hospitalized Patients. IJPBS, (3) 1:79-83. [18] Jadhav, S.; Gandham, N.; Pual, A.; Misra,R. N., and Mania, K.R. (2012). Characterization of multiple drug resistant uropathogenic Escherichia coli(UPEC) special references to B2-O25B-ST131-CTX-15 India, Journal of Pharmaceutical and Biomedical Science, 23(25): ISSN. No. 2230-7885. [19] Lau, S. H.; Reddy, S.; Cheesbrough, J.; Bolton, F. J.; Willshaw, G.; Cheasty, T.; Fox, A. J., and Upton M. (2008). Major Uropathogenic Escherichia coli Strain Isolated in the Northwest of England Identified by Multilocus Sequence Typing. J Clin Microbiol; 46(3): 1076–1080. [20] Lecointre, G.; Rachdi, L.; Darlu, P., and Denamur. E. (1998). Escherichia coli molecular phylogeny using the incongruence length difference test. Mol Biol Evol 15: 1685–1695. [21] Levdansky E.; Romano, J.; Shadkchan Y.; Sharon, H.; Verstrepen, K. J.; Fink G. R., and Osherov N. (2007). Coding Tandem Repeats Generate Diversity in Aspergillus fumigatus Genes. Eukaryotic Cell, 6(8): 1380–1391 [22] Maheux, A. F.; PicardFrancois, J.; Boissinot, M.; Bissonnette, L.; Paradis, S., Bergeron, M. G. (2009). Analytical comparison of nine PCR primer sets designed to detect the presence of Escherichia coli/Shigella in water samples. Water Resaerch 43(12): 3019-3028. (Abstract). [23] Marrs, C. F.; Zhang, L., and Foxman, B. (2005). Escherichia coli mediated urinary tract infections: Are there distinct uropathogenic E. coli (UPEC) pathotypes? FEMS Microbiology Letters, 252:183–190. [24] Mendonça, N.; Calhau, V.; Lin, T.; Boaventura, L.; Ribeiro, G., and Da-Silva, G. J. (2011). Unusual Genotype of a Uropathogenic Escherichia coli Strain Assigned to the B2 Phylogenetic Group. Journal of Clinical Microbiology, 49(8): 3105–3106. [25] Mukherjee, M.; Basu, S.; Mukherjee, S. K., and Globe, M. M. (2013). Multidrug-Resistance and Extended Spectrum Beta-Lactamase Production in Uropathogenic E. coli which were Isolated from Hospitalized Patients in Kolkata, India. Journal of Clinical and Diagnostic Research, 7(3): 449-453. [26] Navidinia, M.; Peerayeh, S. N.; Fallah, F.; Bakhshi, B.; Adabian, S.; Alimehr, S.; Gholinejad, Z., and Malekan, M. A. (2012). Phylogenic Typing of Urinary E coli isolates and commensal E. coli in patients with UTI in mofid children hospital in Iran. Arch. Dis. child., 97 (Abstract). [27] Nazik, H.; Öngen, B.; Yildirim, E. E., and Ermi, F. (2011). High prevalence of CTX-M-type beta-lactamase in Escherichia coli isolates producing extended-spectrum beta-lactamase (ESBL) and displaying antibiotic co-resistance. African Journal of Microbiology Research. 5(1): 44-49. [28] Nicolle, L. E. (2005). Complicated urinary tract infection in adults. Can J Infect Dis Med Microbiol. 16(6): 349–360. [29] Nordstrom, L.; Liu, C. M., and Price, L. B. (2013) Food borne urinary tract infections: a new paradigm for anti-microbial-resistant food borne illness Frontiers in Microbiology. 4( 29): 1-6 [30] Parveen, S.; Lukasik, J.; Scott, T. M.; Tamplin, M. L.; Portier, K. M.; Sheperd, S.; Braun, K., and Farrah, S. R. (2004). Geographical variation in antibiotic resistance profiles of Escherichia coli isolated from swine, poultry, beef and dairy cattle farm water retention ponds in Florida. Journal of Applied Microbiology. 100(1):50-7. [31] Paterson, D. L. (2006). Resistance in gram-negative bacteria: Enterobacteriaceae, J. AJI C, (34):5, 20-28. [32] Prescott, L. M.; Harley, J. P., and Klein, D. A. (1993). Microbiology, 2nd ed. Wm. C. Brown Communication, Inc., Boston, MA. [33] Rezaee, M. A.; Sheikhalizadeh,V., and Hasani, A. (2011). Detection of integrons among multi-drug resistant (MDR) Escherichia coli strains isolated from clinical specimens in northern west of Iran. Braz. J. Microbiol. 42 (4):1308-1313. [34] Rijavec, M.; Starčič Erjavec, M.; Ambrožič Avguštin, J.; Reissbrodt, R.; Fruth, A.; Križan- Hergouth, V. & Žgur-Bertok, D. (2006). High prevalence of multidrug resistance and random distribution of mobile genetic elements among uropathogenic Escherichia coli (UPEC) of the four major phylogenetic groups. Current Microbiology, 53(2): 158-162. [35] Ruiz del Castillo, Belén; Ocampo-Sosa, Alain A.; Martínez-Martínez, Luis (2011) Detection of phylogenetic group B1 Escherichia coli by multiplex PCR: Description of a new amplification pattern, Enferm Infecc Microbiol Clin;29(10):778–786. [36] Russo, T. A., and Johnson, J. R. (2000). Proposal for a New Inclusive Designation for Extraintestinal Pathogenic Isolates of Escherichia coli: ExPEC. The Journal of Infectious Diseases, 181:1753–1754. [37] Sahoo K. C.; Tamhankar, A. J.; Sahoo, S.; Sahu, P. S.; Klintz, S. R., and Lundborg, C. S. (2012). Geographical Variation in Antibiotic-Resistant Escherichia coli Isolates from Stool, Cow-Dung and Drinking Water. Int J Environ Res Public Health. 9(3): 746–759. [38] Scholes, D.; Hooton, T. M.; Roberts, P. L.; Stapleton, A. E.; Gupta, K., and Stamm, W. E. (2000). Risk factors for recurrent urinary tract infection in young women. J Infect Dis., 182: 1177-1182 (Cited by: Katouli, M., and Vollmerhausen, T. L. (2010). Population structure of gut Escherichia coli and its role in development of extra-intestinal infections. Journal of Iranian Microbiology, 2 (2): 59-72. [39] Siripattanapipong, S.; Mungthinn, M.; Tan-ariya, P., and Leelayoova, S. (2010). Multilocuse sequence typing (MLST) and its aplications in epidemiological studies for parasitic infections. In. parasitology research trend. Ed. De Bruyn O. and Peeters, 99-117. [40] Smith, J.; Fratamico, P. M., and Gunther, N. W. (2007). Extraintestinal Pathogenic Escherichia coli, Food Pathogens and Disease, 4(2): 87. [41] Tang, Y. W.; Procop, G. W., and Persing, D. H. (1997). Molecular diagnostics of infectious diseases. Clinical Chemistry 43:112021–2038. [42] Totsika, M.; Moriel, D. G.; Idris, A.; Rogers, B. A.; Wurpel, D. J.; Phan, M.; Paterson, D. L., and Schembri, M. A. (2012). Uropathogenic Escherichia coli Mediated Urinary Tract Infection. Current Drug Targets. 13(11):1386-1399. [43] Van de, S. N.; Grundmann, H.; Verloo, D.; Tiemersma, E.; Monen, J.; Goossens, H.; Ferech, M.; European Antimicrobial Resistance Surveillance System Group, and European Surveillance of Antimicrobial Consumption Project Group. (2008). Antimicrobial drug use and resistance in Europe. Emerg Infect Dis, 14: 1722-1730 (Cited by: Molina-López, J.; Aparicio-Ozores, G.; Ribas-Aparicio, R. M.; Gavilanes-Parra, S.; Chávez-Berrocal, M. E.; Hernández-Castro, R. H., and Manjarrez-Hernández, Á. (2011). Drug resistance, serotypes, and phylogenetic groups among uropathogenic Escherichia coli including O25-ST131 in Mexico City. J Infect Dev Ctries., 5(12):840-849). [44] Warren, J. W.; Abrutyn, E.; Hebel, J. R.; James, R. J.; Anthony, J. S., and Stamm, W. E. (1999). Guidelines for Antimicrobial Treatment of Uncomplicated Acute Bacterial Cystitis and Acute Pyelonephritis in Women, Clinical Infectious Diseases; 29: 745–58. [45] Wiles, T. J.; Kulesus, R. R., and Mulvey, M. A. (2008). Origins and virulence mechanisms of uropathogenic Escherichia coli. Experimental and Molecular Pathology, 85:11–19. [46] Yamamoto, S.; Tsukamoto, T.; Terai, A.; Kurazono, H.; Takeda, Y., and et al., (1997) Genetic Evidence Supporting the Fecal-Perineal-Urethral Hypothesis in Cystitis Caused by Escherichia Coli. The Journal of Urology, 157 (3): 1127-1129. [47] Yasufuku, T.; Shigemura, K.; Shirakawa, T.; Matsumoto, M.; Nakano, Y.; Tanaka, K.; Arakawa, S.; Kinoshita, S.; Kawabata, M., and Fujisawa, M. (2011). Correlation of overexpression of efflux pump genes with antibiotic resistance in Escherichia coli Strains clinically isolated from urinary tract infection patients. J Clin Microbiol. 49(1):189-94.