community acquired uti and nosocomial infection are two types of urinary tract infections ( 2 ) . most of the cases of uti are commonly caused by gram negative bacteria ; escherichia coli is responsible for 80% of infections ( 3 ) followed by klebsiella pneumonia , acinetobacter spp and enterobacter spp ( 4 ) . the uropathogenic escherichia coli ( upec ) strains are the most important causes of utis ( 5 , 6 ) . however , the gram - positive bacteria , staphylococcus saprophyticus , can cause uti in young females ( 7 ) . increasing resistance to antibiotics by uti - causing organisms is a serious public health problem ( 8 , 9 ) . high levels of resistance to antibiotic agents that are frequently used to treat uti infections have been reported in uti - causing e. coli ( 10 , 11 ) . horizontal gene transfer ( hgt ) has an important role in spreading resistance genes among bacteria through mobile genetic elements such as , plasmids , transposons and integrons ( 12 ) . integrons are genetic elements that are capable of integration of resistance gene cassettes in their structure and subsequently denote resistance phenotype to their bacterial host ( 13 ) . integrons can be located within transposons or conjugated plasmids and transfer along with them , facilitating the dissemination of antibiotic resistance genes between bacteria ( 14 ) . structurally , integrons consist of two 5 ' conserved segments ( 5'-cs ) and 3 ' conserved segments ( 3'-cs ) that flank the central variable region or gene cassettes , which encode antibiotic resistance traits ( 15 ) . the 5'-cs of integrons encode the integrase ( inti ) gene , the recombination site ( atti ) and the common promoter ( pc ) , while the 3'-cs harbors the qace1 gene and sul1 gene that mediate resistance to certain detergents and sulphonamides , respectively ( 16 ) . in addition to encoding resistance genes by gene cassettes , they include a recombination site ( attc ) . recombination between the attl and attc sites leads to insertion of the gene cassette downstream of a resident promoter , which mediated by the inti gene ( 17 ) . based on the homology of the inti gene , four classes ( 1 - 4 ) of integrons have been identified ( 18 ) ; class 1 integrons are the most prevalent followed by class 2 integrons ( 19 ) . more than 194 gene cassettes encode antibiotic resistance to aminoglycoside , -lactams , chloramphenicol , quinolones and trimethoprim ( 20 ) . one or more gene cassette can be incorporated in integrons , however usually less than five cassettes with numerous combinations are carried by isolated bacteria ( 20 - 22 ) . hence , the presence of integrons among bacteria is associated with multiple drug resistance ( mdr ) , especially in enteric bacteria such as e. coli ( 23 ) . the mdr enterobacteriaceae serve as a major public health problem and a strong association between mdr and the presence of integrons has been shown in enterobacteriaceae , independent of species or strain origin ( 24 ) . many studies have investigated the presence of integrons in e. coli isolated from patient with uti , and they reported a significant association between antimicrobial resistance and existence of integron ( 23 - 28 ) . pattern of resistance to antibiotics change continuously in uti - causing organisms , therefore appropriate information about local and national antimicrobial resistance will be needed for empirical therapy of uti ( 28 ) . considering the importance of regional and local information about antibiotic resistance pattern among e. coli isolated from patients with utis , and the association between integrons and mdr among gram negative bacteria , the aim of this study was to determine the pattern of antibiotic resistance to commonly used antimicrobial agents , and to investigate the presence of class 1 , 2 and 3 integrons and their association with antibiotic resistance in e. coli isolated from patient with utis in yasuj , iran . in this cross - sectional study , midstream urine samples of 1820 patients diagnosed with utis were collected in sterile universal containers from february 2013 to november 2014 . exclusion criteria were having an indwelling urinary catheter , being pregnant , having genitourinary abnormalities and antibiotics therapy within the previous two weeks . urine samples were inoculated on macconkey agar and blood agar plates using calibrated loops ( 0.001 ml ) and incubated at 37c for 18 to 24 hours . the number of colony forming units ( cfu ) was counted . a growth of > 10 colony forming units / ml of one type of organism was considered as significant bacteriuria . the isolates were identified to the species level using conventional biochemical tests , such as indole production , methyl red , voges - proskauer , simmons citrate , hydrogen sulfide , and urea hydrolysis ( 29 ) . all of the isolates were stored at 20c in trypticase soy broth ( mast group ltd . uk ) containing 15% glycerol ( 29 ) . disks containing antibiotics ( himedia , india ) were used to determine the susceptibility of e. coli isolates to the following antibiotics , according to the clinical and laboratory standards institute ( clsi ) guidelines ( 30 ) : amoxicillin ( 25 g ) , gentamycine ( 10 g ) , amikacin ( 30 g ) , co - trimoxazole ( 25 g ) , tetracycline ( 30 g ) , ciprofloxacin ( 5 g ) , nalidixic acid ( 30 g ) , imipenem ( 10 g ) , cephalothin ( 30 g ) , chloramphenicol ( 30 g ) , ceftazidime ( 30 g ) . this study was approved by the ethical committee of yasuj university of medical sciences ( ethical code number : 20.11.91 - 91011020 ) . briefly , three to five colonies of overnight culture of e. coli on brain heart infusion agar was suspended in 300 l of sterile distilled water and boiled at 95c for ten minutes then cell debris was removed by centrifugation for tenminutes at 13000 rpm , and 200 l of the supernatant was stored at 20 c for dna amplification . the inti1 , inti2 and inti3 genes were amplified by a multiplex - polymerase chain reaction ( pcr ) method using primers described by goldstein et al . ( 8) ( table 1 ) . the pcr amplification was performed in a total volume of 25 l containing 12.5 l of master mix ( cinnagen , iran ) , 25 pmol of each primer and 3 l of dna template . expected amplified products , 280 bp for integron class 1 , 233 bp for integron class 2 and 600 bp for integron class 3 were separated by electrophoresis on 1% agarose gel containing 0.5 g / ml ethidium bromide and photographed under uv illumination .   abbreviations : f , forward ; r , reverse . the spss software ( version 16 , chicago , il , usa ) the association between presence of integrons and antibiotic resistance was determined by  or fisher s exact test . a p value of < 0.05 was considered statistically significant . in this cross - sectional study , midstream urine samples of 1820 patients diagnosed with utis were collected in sterile universal containers from february 2013 to november 2014 . exclusion criteria were having an indwelling urinary catheter , being pregnant , having genitourinary abnormalities and antibiotics therapy within the previous two weeks . urine samples were inoculated on macconkey agar and blood agar plates using calibrated loops ( 0.001 ml ) and incubated at 37c for 18 to 24 hours . the number of colony forming units ( cfu ) was counted . a growth of > 10 colony forming units / ml of one type of organism was considered as significant bacteriuria . the isolates were identified to the species level using conventional biochemical tests , such as indole production , methyl red , voges - proskauer , simmons citrate , hydrogen sulfide , and urea hydrolysis ( 29 ) . all of the isolates were stored at 20c in trypticase soy broth ( mast group ltd . uk ) containing 15% glycerol ( 29 ) . disks containing antibiotics ( himedia , india ) were used to determine the susceptibility of e. coli isolates to the following antibiotics , according to the clinical and laboratory standards institute ( clsi ) guidelines ( 30 ) : amoxicillin ( 25 g ) , gentamycine ( 10 g ) , amikacin ( 30 g ) , co - trimoxazole ( 25 g ) , tetracycline ( 30 g ) , ciprofloxacin ( 5 g ) , nalidixic acid ( 30 g ) , imipenem ( 10 g ) , cephalothin ( 30 g ) , chloramphenicol ( 30 g ) , ceftazidime ( 30 g ) . this study was approved by the ethical committee of yasuj university of medical sciences ( ethical code number : 20.11.91 - 91011020 ) . , three to five colonies of overnight culture of e. coli on brain heart infusion agar was suspended in 300 l of sterile distilled water and boiled at 95c for ten minutes then cell debris was removed by centrifugation for tenminutes at 13000 rpm , and 200 l of the supernatant was stored at 20 c for dna amplification . the inti1 , inti2 and inti3 genes were amplified by a multiplex - polymerase chain reaction ( pcr ) method using primers described by goldstein et al . the pcr amplification was performed in a total volume of 25 l containing 12.5 l of master mix ( cinnagen , iran ) , 25 pmol of each primer and 3 l of dna template . expected amplified products , 280 bp for integron class 1 , 233 bp for integron class 2 and 600 bp for integron class 3 were separated by electrophoresis on 1% agarose gel containing 0.5 g / ml ethidium bromide and photographed under uv illumination .   abbreviations : f , forward ; r , reverse . the spss software ( version 16 , chicago , il , usa ) was used for statistical analysis . the association between presence of integrons and antibiotic resistance was determined by  or fisher s exact test . a p value of < 0.05 was considered statistically significant . among the 200 uti e. coli strains , 144 ( 72% ) strains were isolated from females and 56 ( 28% ) strains were isolated from males . high frequencies of resistance were observed toward cephalothin ( 99% ) and amoxicillin ( 76% ) while only two ( 1% ) isolates showed resistance to imipenem . the e. coli isolates were also resistant to co - trimoxazole ( 62% ) , tetracycline ( 50% ) , nalidixicacid ( 48.5% ) , ceftazidime ( 40.5% ) , ciprofloxacin ( 29% ) , gentamicin ( 15.5% ) , chloramphenicol ( 13% ) and amikacin ( 3% ) . of the 200 tested e. coli isolates , 158 ( 79% ) were mdr and were resistant to three or more antibiotics . none of the isolates were fully susceptible or resistant to all of the tested antibiotics . sixty - six antibiotic resistance patterns were observed in e. coli isolates , yet 30 resistance phenotypes were presented by only one isolate . resistance to cephalothin was the most common phenotype , which was detected in 27 isolates . the second resistance phenotype was towards 13 isolates , including resistance to cephalothin , amoxicillin , co - trimoxazole and tetracycline . class 1 and 2 integrons were detected in 104 ( 52% ) and 5 ( 2.5% ) isolates , respectively , while none of isolates were positive for class 3 integrons . ninety - three of the isolates ( 46.5% ) did not contain any classes of integrons . among the 158 mdr e. coli , 101 ( 63.9% ) harbored class 1 or class 2 integrons yet only 5 ( 11.9% ) of the 42 non - mdr isolates had integrons . the carriage of class 1 integron was found to be significantly higher in co - trimoxazole ( p < 0.001 ) , nalidixic acid ( p < 0.001 ) , ciprofloxacin ( p < 0.015 ) , amoxicillin ( p < 0.001 ) , ceftazidime ( p < 0.003 ) and tetracycline ( p < 0.012 ) resistant isolates ( table 3).there was no significant association between resistance to chloramphenicol , amikacin and gentamicin with the presence of class 1 integrons .   abbreviation : na , not available . consistent with previous reports , in this study class 1 integrons were the most prevalent compared to the other tested integrons . in accordance with our study , chang et al . from taiwan and li et al . from china detected class 1 integrons in 64% and 66.5% of e. coli isolated from different clinical specimens and blood stream infections , respectively ( 31 , 32 ) . many reports have shown the prevalence rate of integrons to be between 6.25% and 54.6% of clinical isolates ( 33 - 38 ) . there have been various studies from different cities of iran : fallah et al . from tehran detected class1 integrons in 50.3% of isolates ( 37 ) while rezaee et al . from tabriz , and japoni et al . and farshad et al . from shiraz identified this class of integrons in 22.05% , 33.34% and 6.25% of e. coli , respectively ( 33 - 35 ) . detected integrons in only 6.25% of e. coli isolated from children with uti and suggested that antibiotic resistance cassettes may be carried on transposable elements or other plasmids rather than integrons . class 2 integrons were detected in 2.5% of isolates in this study , which is lower than that reported by other studies , such as fallah et al . similar to previous studies we could nt find class 3 integrons in the present study ( 31 - 39 ) . antibiotic resistance encoding gene cassettes for fluoroquinolones , -lactams , aminoglycosides , trimethoprim and chloramphenicol have been detected in integrons ( 20 ) . a significant association between presence of class 1 integrons and resistance to co - trimoxazole , nalidixic acid , ciprofloxacin , amoxicillin , ceftazidime and tetracycline was shown in this study . in accordance with our study , a significant association between presence of class 1 integrons and resistance tociprofloxacin , nalidixic acid and co - trimoxazole was reported by japoni et al , to co - trimoxazole , nalidixic acid and ceftazidime by fallah et al . and to nalidixic acid and tetracycline by mathai et al . while in many studies resistance to other antibiotics such as gentamicin , amikacin , cephalothin ( 35 ) and chloramphenicol ( 34 ) were correlated with class 1 integrons . in spite of many resistance gene cassettes including aada , aadb , aada7 , aada4 and aaca1 , which confer resistance to aminoglycosides like amikacin and gentamicin , in our study this association was not significant . this may be due to the low resistance rate to amikacin ( 2.5% ) , or other resistance mechanisms ( mutation in the rrna gene , efflux and permeability mechanisms ) to aminoglycosides rather than integrons by bacteria in this study . sincemost of the isolates ( 99% ) were resistant to cephalothin and only 1% were resistant to imipenem , no significant association was found between presence of integrons and resistance to these antibiotics , similar to many previous studies ( 33 , 34 ) . in this study , 79% of isolates were mdr , which is approximately similar to other reports from iran that showed 77% and 84.2% mdr in isolated e. coli ( 33 - 37 ) , while in the usa and slovenia this was 7.1% and 42% , respectively ( 39 , 40 ) . empirical therapy for mdr isolates is a major concern ; hence a periodical local study will be needed to assess the antibiotic susceptibility pattern . in the present study a high resistance rate was seen towards cephalothin ( 99% ) , which is higher than other studies that reported 77.8% , 60% and 40% ( 34 , 35 , 37 ) . . showed similar results ( 33 - 35 , 41 ) while , fallah et al . reported that 27.1% of their isolates were sensitive to imipenem ( 37 ) . although this antibiotic may be used as a drug of choice for treatment of uti caused by e. coli , yet it is only recommended for complicated urinary tract infections . resistance to ciprofloxacin and nalidixic acid was 29% and 48.5% in this study , which was lower than the studies of fallah et al . and rezaee et al . ( 34 , 37 ) , yet higher than other studies ( 35 , 42 , 43 ) . these two antibiotics are among the drugs of choice for treatment of uti , hence increased use of these antibiotics without knowledge about the pattern of antibiogram may gradually lead to a rise in antibiotic resistance . sixty - two percent of the isolates were resistant to co - trimoxazole , which was similar to the study of fallah et al . ( 67.7% ) but higher than the study of japoni et al . and lower than rezaee et al . and farshad et al . ( 33 - 35 , 37 ) . a large number of mdr isolates of e. coli were observed in this study , among which 62.6% had class 1 integrons.the significant association between class 1 integrons and resistance to ciprofloxacin , nalidixic acid , co - trimoxazole , amoxicillin , ceftazidime and tetracycline suggests that class 1 integrons have an important role in resistance to these antibiotics . we suggest that further research should be done on the association between antibiotic resistance and presence of integrons , integrase genes and gene cassettes . a large number of mdr isolates of e. coli were observed in this study , among which 62.6% had class 1 integrons.the significant association between class 1 integrons and resistance to ciprofloxacin , nalidixic acid , co - trimoxazole , amoxicillin , ceftazidime and tetracycline suggests that class 1 integrons have an important role in resistance to these antibiotics . we suggest that further research should be done on the association between antibiotic resistance and presence of integrons , integrase genes and gene cassettes .