coagulase - negative staphylococci ( cons ) , especially staphylococcus epidermidis , are considered as normal epithelial flora of every human in different parts of the body such as nares , head and axilla , with an essential role in maintaining the normal flora of healthy skin ( 1 ) .
s. epidermidis is also recognized as an important opportunistic pathogen in nosocomial infections , particularly in indwelling medical device users and contaminant agent of blood cultures .
nosocomial infection with s. epidermidis should be more complicated when bacteria are resistant to beta - lactam antibiotics such as methicillin .
these bacteria are common causes of nosocomial infections worldwide in immunocompromised patients or patients who carry indwelling devices ( 2 ) .    reduced affinity for beta - lactam antibiotics in methicillin - resistant staphylococci mediates by a penicillin - binding protein ( pbp2a ) encoded by meca gene located in staphylococcal chromosome cassette mec ( sccmec ) .
furthermore , it is believed that s. epidermidis acts as a reservoir for meca and transfers it to s. aureus by horizontal gene transfer .
recent data indicate that not only meca , but also other mobile genetic elements can be transferred from s. epidermidis to s. aureus ( 3 ) .
the aim of this study was to determine the prevalence of s. epidermidis as well as antibiotic susceptibility pattern and meca prevalence in s. epidermidis isolated from intensive care unit ( icu ) patients .
a cross - sectional study was conducted from september 2010 to september 2011 . a collection of 184 coagulase - negative staphylococci isolated from blood ( n = 135 ) , urine ( n = 17 ) , tracheal ( n = 19 ) , wound ( n = 6 ) , cerebrospinal fluid ( csf ) ( n = 2 ) , pleural fluid ( n = 2 ) , synovial fluid ( n = 2 ) , and peritoneal fluid ( ascitic fluid ) ( n = 1 ) , from three different teaching hospitals of tehran , iran , was included in this study
the sample was enriched in tryptic soy broth , and grown on mannitol salt agar , then catalase , tube coagulase and urease tests , and carbohydrate fermentation were performed .
s. epidermidis is catalase positive , urease positive , unable to ferment d - mannitol and d - trehalose , and able to ferment d - mannose and d - maltose ( 4 , 5 ) .
antibiotic susceptibility testing was performed for s. epidermidis isolates using disk diffusion method on mueller - hinton agar plates ( merck , germany ) and commercial antibiotic disks including : vancomycin ( 30 g ) , linezolid ( 30 g ) , rifampin ( 5 g ) , ciprofloxacin ( 5 g ) , trimethoprim - sulfamethoxazole ( 1.25/23.75 g ) , tetracycline ( 30 g ) , and erythromycin ( 15 g ) , ( mast , uk ) , according to the clinical and laboratory standards institute ( clsi ) guidelines ( 6 ) . dna of each s. epidermidis isolate was extracted from 1ml of overnight bacterial culture .
extraction was performed by adding te [ 10 mm tris , 1 mm edta ( ph = 8) ] buffer and lysostaphin , 95c for 5 minutes , then 0c for 5 minutes , and centrifuged in 10000 rpm for 5 minutes .
dna was measured using a biophotometer ( eppendorf , germany ) to determine the concentration and purity .
( 7 ) with the following specific primers : forward : 5-gtgaagatataccaagtgatt-3 reverse : 5-atgcgctatagattgaaaggat-3. amplification was performed in a mj mini .
u.s.a ; initial denaturation step at 94c for 5 minutes , followed by 30 cycles of 45 seconds at 94c,45 seconds at 55c , and 90 seconds at 72c , ending with a final extension step at 72c for 10 minutes , followed by a hold at 4c .
the amplicon size was 147 bp ( figure 1 ) . in the pcr reaction , s. epidermidis atcc 12228 and s. aureus atcc 33591
1 , marker , 100 bp ; 2 , s. aureus atcc 33591 ( 147 bp ) ; 3 - 5 , positive clinical strains .
confidence interval test was used to assess the statistical significance with confidence level of 95% (  = 0.05 ) .
a cross - sectional study was conducted from september 2010 to september 2011 . a collection of 184 coagulase - negative staphylococci isolated from blood ( n = 135 ) , urine ( n = 17 ) , tracheal ( n = 19 ) , wound ( n = 6 ) , cerebrospinal fluid ( csf ) ( n = 2 ) , pleural fluid ( n = 2 ) , synovial fluid ( n = 2 ) , and peritoneal fluid ( ascitic fluid ) ( n = 1 ) , from three different teaching hospitals of tehran , iran , was included in this study
the sample was enriched in tryptic soy broth , and grown on mannitol salt agar , then catalase , tube coagulase and urease tests , and carbohydrate fermentation were performed .
s. epidermidis is catalase positive , urease positive , unable to ferment d - mannitol and d - trehalose , and able to ferment d - mannose and d - maltose ( 4 , 5 ) .
antibiotic susceptibility testing was performed for s. epidermidis isolates using disk diffusion method on mueller - hinton agar plates ( merck , germany ) and commercial antibiotic disks including : vancomycin ( 30 g ) , linezolid ( 30 g ) , rifampin ( 5 g ) , ciprofloxacin ( 5 g ) , trimethoprim - sulfamethoxazole ( 1.25/23.75 g ) , tetracycline ( 30 g ) , and erythromycin ( 15 g ) , ( mast , uk ) , according to the clinical and laboratory standards institute ( clsi ) guidelines ( 6 ) .
extraction was performed by adding te [ 10 mm tris , 1 mm edta ( ph = 8) ] buffer and lysostaphin , 95c for 5 minutes , then 0c for 5 minutes , and centrifuged in 10000 rpm for 5 minutes .
dna was measured using a biophotometer ( eppendorf , germany ) to determine the concentration and purity .
pcr was performed as described by zhang et al . ( 7 ) with the following specific primers : forward : 5-gtgaagatataccaagtgatt-3 reverse : 5-atgcgctatagattgaaaggat-3. amplification was performed in a mj mini .
u.s.a ; initial denaturation step at 94c for 5 minutes , followed by 30 cycles of 45 seconds at 94c,45 seconds at 55c , and 90 seconds at 72c , ending with a final extension step at 72c for 10 minutes , followed by a hold at 4c .
the amplicon size was 147 bp ( figure 1 ) . in the pcr reaction ,
s. epidermidis atcc 12228 and s. aureus atcc 33591 were used as negative and positive controls , respectively ( 7 ) .
1 , marker , 100 bp ; 2 , s. aureus atcc 33591 ( 147 bp ) ; 3 - 5 , positive clinical strains .
confidence interval test was used to assess the statistical significance with confidence level of 95% (  = 0.05 ) .
in this study , frequency of s. epidermidis was 34.8% ( 95% ci : 46.5 - 23.1 ) .
of the 64 s. epidermidis isolates , 52 ( 81.3% ) were from blood , 6 ( 9.4% ) tracheal , 4 ( 6.2% ) urine , and 2 ( 3.1% ) wound samples .
meca was detected in 92.2% ( 95%ci : 98.8 - 85.6 ) of s. epidermidis isolates .
resistance to rifampin and ciprofloxacin were 9.4% ( 95%ci : 16.5 - 2.3 ) and 23.4% ( 95%ci : 33.8 - 13 ) , respectively .
resistance to trimethoprim - sulfamethoxazole was observed in 51.6% ( 95% ci : 63.8 - 39.4 ) of s. epidermidis isolates , while 46.9% ( 95% ci : 59.1 - 34.7 ) were susceptible to trimethoprim - sulfamethoxazole .
susceptibility to tetracycline was observed in 42.1% ( 95% ci : 54.2 - 30 ) of s. epidermidis isolates , while resistance was observed in 57.8% ( 95%ci : 69.9 - 45.7 ) and 59.4% ( 95%ci : 71.4 - 47.4 ) to tetracycline and erythromycin , respectively
abbreviations : cip , ciprofloxacin ; e , erythromycin ; meca , presence of meca ; rp , rifampin ; sxt , trimethoprim - sulfamethoxazole ; te , tetracycline ,   three of the 64 isolates ( 4.7% ) were resistance to six antibiotics , 4 ( 6.3% ) to five antibiotics , 20 ( 31.3% ) to four antibiotics , 9 ( 14% ) to three antibiotics , 13 ( 20.3% ) to two antibiotics , 14 ( 21.9% ) to one antibiotic , and 1 ( 1.7% ) was susceptible to all the antibiotics .
the most common resistance pattern was trimethoprim - sulfamethoxazole , tetracycline , erythromycin , and methicillin resistance , found in 15 isolates ( 23.4% ) , followed by resistance to methicillin only , the second - most common resistance pattern , observed in13 ( 20.3% ) of the isolates .
in the recent years , s. epidermidis has been isolated as an infective agent in nosocomial infections .
some reasons for such infections increasing are increased number of immune - deficient patients , use of indwelling medical devices , and use of antibiotics and disinfectants ( 8) .
methicillin resistance was observed in 75 - 90% of isolated s. epidermidis from nosocomial infections , which was higher than the resistance rate for s. aureus isolates ( 40 - 60% ) .
noticing the presence of s. epidermidis as a human commensal flora , it is assumed to be carrier and reservoir for different genes such as antimicrobial resistance gene ( 2 ) .  
frequency of s. epidermidis isolates in this study was significantly lower than reports of okee et al .
( 9 ) , eftekhar and raei ( 10 ) , bouchami et al . ( 11 ) , barbier et al .
( 12 ) , ruppe et al . ( 13 ) , and mombach pinheiro machado ab et al . ( 14 ) . in the present study ,
all the s. epidermidis isolates were susceptible to vancomycin , which was in accordance with the result of mendes et al .
( 15 ) , eftekhar and raei ( 10 ) , abd el hafez et al .
( 19 ) . in our study , all the s. epidermidis isolates were susceptible to linezolid , which was in accordance with the results of hellmark et al .
( 18 ) , but significantly higher than delgado et al . ( 19 ) report .   in the present study , resistance to rifampin
( 17 ) , and bouchami et al . ( 11 ) reports , and it also was significantly higher than abd el hafez et al .
( 17 ) , bouchami et al . ( 11 ) , abd el hafez et al .
, resistant to trimethoprim - sulfamethoxazole was significantly lower than report of delgado et al .
( 17 ) , but was significantly higher than the report of haque et al .
( 16 ) report . in this study , susceptibility to tetracycline was significantly lower than the report of mendes et al .
( 19 ) ; also , resistance to tetracycline was significantly higher than the report of bouchami et al . ( 11 ) . in this study , 92.2% of s. epidermidis
rohde et al . ( 21 ) reported that 87.5% of s. epidermidis isolates harbored meca .
our data did not show significant difference with these studies , but was significantly higher than lduma et al .
( 9 ) results , which reported only 10% meca distribution , and mendes et al .
( 15 ) study , which reported 73.2% methicillin resistant . in our study , 56.3% of s. epidermidis isolates were resistant to three antibiotics or more .
considering the presence of s. epidermidis in icu patients , multidrug - resistant bacteria can cause infection and would be more complicated in treatment . in conclusion ,
presence rate of meca and susceptibility to linezolid and vancomycin did not show significant differences with other investigations , while susceptibility to trimethoprim - sulfamethoxazole was significantly higher , and susceptibility to tetracycline was significantly lower than other investigations .
presence of meca - positive s. epidermidis isolates in icu patients , especially in ones with compromised immune systems , may cause infection and would be more complicated in the case of antibiotic resistance .
 background : coagulase - negative staphylococci ( cons ) , especially staphylococcus epidermidis , are considered as normal flora of human epithelia and also important opportunistic pathogens for nosocomial infections . 
 s. epidermidis can also act as a reservoir for meca , responsible for high - level resistance to methicillin and transferring it to s. aureus.objectives:the aim of this study was to determine the prevalence of s. epidermidis as well as antibiotic susceptibility pattern and meca prevalence in s. epidermidis isolated from intensive care unit ( icu ) patients.materials and methods : a cross - sectional study was conducted from september 2010 to september 2011 and 184 coagulase - negative staphylococci were collected from different clinical samples in three hospitals . 
 s. epidermidis was identified by conventional bacteriological tests . 
 antibiotic susceptibility testing was performed using disk diffusion method . 
 frequency of meca was detected by specific pcr.results:frequency of s. epidermidis was 34.8% , the most susceptibility was seen to linezolid and vancomycin , and the least susceptibility was seen to tetracycline.majority of the s. epidermidis isolates carried meca ( 92.2% ) . 
 the most common resistant pattern was trimethoprim - sulfamethoxazole , tetracycline , erythromycin , and methicillin resistance , found in 23.4% of the isolates , followed by resistance to methicillin as the second - most common resistant pattern , observed in 20.3% of the isolates.conclusions:frequency of s. epidermidis was significantly lower , compared to other studies . 
 presence rate of meca and susceptibility to linezolid and vancomycin did not show significant differences with other investigations , while resistant to trimethoprim - sulfamethoxazole was significantly lower compared to other investigations , and resistance to tetracycline was significantly higher in comparison to other investigations . 
 presence of methicillin - resistant s. epidermidis in icu patients , especially in individuals with compromised immune systems , may cause infection and would be more complicated in the case of antibiotic resistance . 
1. Background
2. Objectives
3. Materials and Methods
3.1. Bacterial Isolates
3.2. Antibiotic Susceptibility Testing
3.3. DNA Extraction
3.4. Detection of mecA
3.5. Statistical Analysis
4. Results
5. Discussion