Antimicrobial Resistance patterns among nosocomial gram negative bacilli by E-test and disc diffusion methods in Sina and Imam Hospital

Introduction It has been reported that nosocomial infections concern 5 to 15% of hospitalized patients. It has been reported that nosocomial infections concern 5 to 15% of hospitalized patients. The patient in the Intensive care unit (ICU) has a 5 to 7 fold higher risk of nosocomial infection compared with the average patient. The patient in the Intensive care unit (ICU) has a 5 to 7 fold higher risk of nosocomial infection compared with the average patient. The frequent use of broad-spectrum antibiotics results in colonization with resistant gram- negative bacteria and consequently to serious infections. The frequent use of broad-spectrum antibiotics results in colonization with resistant gram- negative bacteria and consequently to serious infections.

Methods This was a cross-sectional study carried out in two university hospitals in Tehran, Iran (Sina and Imam-Khomeini Hospitals) This was a cross-sectional study carried out in two university hospitals in Tehran, Iran (Sina and Imam-Khomeini Hospitals) Gram-negative : Gram-negative : Pseudomonas Acinetobacter Klebsiella E.coli

Susceptibility testing was performed by E-test (AB Biodisk, Solna, Sweden) Susceptibility testing was performed by E-test (AB Biodisk, Solna, Sweden) Imipenem, Cefepime, Ceftazidime, Ciprofloxacin and Ceftriaxone Imipenem, Cefepime, Ceftazidime, Ciprofloxacin and Ceftriaxone

Quality control was assured by concurrent testing with American Type Culture Collection (ATCC) strains including E. coli ATCC and P.aeuroginosa ATCC After incubation at ambient air at 35 °C for 24 hours, MIC was interpreted as the point at which the inhibition ellipse intersected with the E-test strip edge. The testing procedures were validated in accordance with the guidelines of the National Committee for Clinical Laboratory Standards (NCCLS). Quality control was assured by concurrent testing with American Type Culture Collection (ATCC) strains including E. coli ATCC and P.aeuroginosa ATCC After incubation at ambient air at 35 °C for 24 hours, MIC was interpreted as the point at which the inhibition ellipse intersected with the E-test strip edge. The testing procedures were validated in accordance with the guidelines of the National Committee for Clinical Laboratory Standards (NCCLS).

Results

A total of 380 samples were obtained from 270 inpatients A total of 380 samples were obtained from 270 inpatients A total of studied patients: A total of studied patients: Male = 60.7% (164 cases) Male = 60.7% (164 cases) female 39.3% (106 cases ) female 39.3% (106 cases ) The mean age = 46.4 ± 21.6 years The mean age = 46.4 ± 21.6 years

Wards

Most common specimens

Most frequent Nosocomial Infections

The frequency of microbial agents

Table 2– Frequency of gram-negative isolates according to origins of nosocomial infections in the present study InfectionP.aeruginosa Number (%) E. coli Number (%) Acinetobacter spp.Number (%) Klebsiella spp. Number (%) Total Urinary tract infection 23 (36.5) 18 (28.6) 4 (6.3) 18 (28.6) 63 Pneumonia 25 (25.5) 11 (11.2) 25 (25.5) 37 (37.8) 98 Tracheitis 3 (33.3) 0 (0) 5 (55.6) 1 (11.1) 9 Meningitis 1 (16.7) 0 (0) 1 (16.7) 4 (66.7) 6 Central catheter infection 3 (13.6) 2 (9.1) 5 (22.7) 12 (54.5) 22 Bed sore 1 (33.3) 0 (0) 2 (66.7) 3 Surgical site infection 30(42.3) 9(12.7)12(16.9)20(28.2)71 Primary septicemia 21 (19.4) 7 (6.5) 23 (21.3) 57 (52.8) 108 Total 107 (28.2) 47 (12.4) 75 (19.7) 151 (39.7) 380

History of receiving antibiotics Total = 67.4% Total = 67.4% 60.43% had received antibiotics >7 days, 60.43% had received antibiotics >7 days, 25.3% had receved Antibiotics = 3-5 days 25.3% had receved Antibiotics = 3-5 days 15.3% had receved Antibiotics <3 days 15.3% had receved Antibiotics <3 days.

History of receiving antibiotics

Table 1– Antimicrobial drug susceptibility of the five antibiotics we studied" according to National Committee for Clinical Laboratory Standards (NCCLS) Antimicrobial agent Sensitivity(µg/ml) Intermediate (µg/ml) Resistance(µg/ml) Ciprofloxacin ≤ 1 ≤ 2 4 Imipenem≤4≤816 Cefepime ≤ 8 ≤ Ceftriaxone ≤ 8 ≤ Ceftazidime ≤ 8 ≤ 16 32

Table 3 – Resistance pattern of the studied antimicrobial agents to bacteria of nosocomial origin Antimicrobial agent Susceptible Number (%) Intermediate ResistantNumber (%, 95% CI) Imipenem319(83.9)12(3.2)49 (12.9; ) Cefepime99(26.1)29(7.6)252 (66.3; ) Ciprofloxacin98(25.1)34(8.9)248 (65.3; ) Ceftazidime75(19.1)13(3.4)292 (76.8; ) Ceftriaxone20(10.1)7(3.5)171 (86.4; )

Table 4 – Comparison of resistance pattern of isolated bacteria of nosocomial origin Antimicrobial agent/ organism Susceptible Number (%) Intermediate Number (%) Resistant Number (%) Ceftazidime P. aeruginosa P. aeruginosa 34 (31.8) 7 (6.5) 66 (61.7) E. coli E. coli 10 (21.3) 4 (8.5) 33 (70.2) Acinetobacter spp. Acinetobacter spp. 12 (16) 2 (2.7) 61 (81.3) Klebsiella spp. Klebsiella spp. 19 (12.6) (87.4) Total Total 75 (19.7) 13 (3.4) 292 (76.8)

Table 4 – Comparison of resistance pattern of isolated bacteria of nosocomial origin Antimicrobial agent/ organism Susceptible Number (%) Intermediate Number (%) Resistant Number (%) Imipenem P. aeruginosa P. aeruginosa 80 (74.8) 6 (5.6) 21 (19.6) E. coli E. coli 43 (91.5) 0 4 (8.5) Acinetobacter spp. Acinetobacter spp. 58 (77.3) 4 (5.3) 3 (17.3) Klebsiella spp. Klebsiella spp. 138 (91.4) 2 (1.3) 11 (7.3) Total Total 319 (83.9) 12 (3.2) 49 (12.9)

Table 4 – Comparison of resistance pattern of isolated bacteria of nosocomial origin Antimicrobial agent/ organism Susceptible Number (%) Intermediate Number (%) Resistant Number (%) Ciprofloxacin P. aeruginosa P. aeruginosa 42 (39.3) 5 (4.7) 60 (56.1) E. coli E. coli 8 (17) 3 (6.4) 36 (76.6) Acinetobacter Acinetobacter 16 (21.3) 3 (4) 56 (74.7) Klebsiella spp Klebsiella spp 32 (21.2) 23 (15.2) 96 (63.6) Total Total 98 (25.8) 34 (8.9) 248 (65.3)

Table 4 – Comparison of resistance pattern of isolated bacteria of nosocomial origin Antimicrobial agent/ organism Susceptible Number (%) Intermediate Number (%) Resistant Number (%) Ceftriaxone E. coli E. coli 10 (21.3) 0 37 (78.7) Klebsiella Klebsiella 10 (6.6) 7 (4.6) 134 (88.7) Total Total 20 (10.1) 7 (3.5) 171 (86.4)

Table 4 – Comparison of resistance pattern of isolated bacteria of nosocomial origin Antimicrobial agent/ organism Susceptible Number (%) Intermediate Number (%) Resistant Number (%) Cefepime P. aeruginosa P. aeruginosa 39 (36.4) 4 (3.7) 64 (59.8) E. Coli E. Coli 9 (19.1) 3 (6.4) 35 (74.5) Acinetobacter Acinetobacter 13 (17.3) 10 (12.3) 52 (69.3) Klebsiella spp Klebsiella spp 38 (25.2) 12 (7.9) 101 (66.9) Total Total 99 (26.1) 29 (7.6) 252 (66.3)

Table 5– Comparison of resistance pattern of gram- negative bacteria to antimicrobial agents in different studies Antimicrobial agent/ organism Oma n (9) (%) Turkey (3) (%) Japan (13) (%) US (18) (%) Belgium (6) (%) Turkey (19) (%) Present study on Iran Number (%, 95% CI) Ceftazidime----7 P. Aeruginosa P. Aeruginosa <-- 66 (61.7; 51/7-70.8) E. Coli E. Coli (70.2; ) Acinetobacter Acinetobacter > (81.3; ) Klebsiella Klebsiella (87.4; )

Table 5– Comparison of resistance pattern of gram- negative bacteria to antimicrobial agents in different studies Antimicrobial agent/ organism Oman (9) (%) Turkey (3) (%) Japan (13) (%) US (18) (%) Belgium (6) (%) Turkey (19) (%) Present study on Iran Number (%, 95% CI) Imipenem P. Aeruginosa P. Aeruginosa <- 21 (19.6; ) E. Coli E. Coli (8.5; ) Acinetobacter spp. Acinetobacter spp < (17.3; ) Klebsiella spp Klebsiella spp (7.3; )

Table 5– Comparison of resistance pattern of gram- negative bacteria to antimicrobial agents in different studies Antimicrobial agent/ organism Oman (9) (%) Turkey (3) (%) Japan (13) (%) US (18) (%) Belgium (6) (%) Turkey (19) (%) Present study on IranNumber (%, 95% CI) Ciprofloxacin P.aeruginosa P.aeruginosa <- 60 (56.1; ) E. coli E. coli (76.6; ) Acinetobacter spp. Acinetobacter spp > (74.7; ) Klebsiella spp Klebsiella spp (63.6; )

Table 5– Comparison of resistance pattern of gram- negative bacteria to antimicrobial agents in different studies Antimicrobial agent/ organism Oman (9) (%) Turkey (3) (%) Japan (13) (%) US (18) (%) Belgium (6) (%) Turkey (19) (%) Present study on IranNumber (%, 95% CI) Ceftriaxone E. coli E. coli (78.7; ) Klebsiella spp Klebsiella spp (88.7; )

Table 5– Comparison of resistance pattern of gram- negative bacteria to antimicrobial agents in different studies Antimicrobial agent/ organism Oman (9) (%) Turkey (3) (%) Japan (13) (%) US (18) (%) Belgium (6) (%) Turkey (19) (%) Present study on IranNumber (%, 95% CI) Cefepime P. Aeruginosa P. Aeruginosa <-- 64 (59.8; 49-84) E. Coli E. Coli (74.5; ) Acinetobacter spp. Acinetobacter spp (69.3; ) Klebsiella spp. Klebsiella spp (66.9; )

The comparison of the resistance status of microorganisms by both disc diffusion and E-test methods showed a clinically noticeable agreement between these two tests. The comparison of the resistance status of microorganisms by both disc diffusion and E-test methods showed a clinically noticeable agreement between these two tests.

Discussion The findings of the present study are indicative of high resistance rate in case of most microorganisms. The findings of the present study are indicative of high resistance rate in case of most microorganisms. The rate of receiving imipenem (which is not administrated on an out-patients basis) was low and the corresponding resistance rate was low as well. The rate of receiving imipenem (which is not administrated on an out-patients basis) was low and the corresponding resistance rate was low as well.

Discussion Cefepime despite its low administration rate in hospital setting had a high resistance rate in our study which might be due to its cross-resistancy with other beta- lactams. Cefepime despite its low administration rate in hospital setting had a high resistance rate in our study which might be due to its cross-resistancy with other beta- lactams. Even ciprofloxacin which is widely used for gram-negative infections on an outpatient basis was found to have a high resistance rate. Even ciprofloxacin which is widely used for gram-negative infections on an outpatient basis was found to have a high resistance rate.

Discussion Susceptibility to quinolones has decreased more in recent years in comparison with other antibiotics and this might be a consequence of wide usage of these antibiotics. Susceptibility to quinolones has decreased more in recent years in comparison with other antibiotics and this might be a consequence of wide usage of these antibiotics. Quinolone-resistant strains might therefore spread more easily than the strains resistant to other agents. Quinolone-resistant strains might therefore spread more easily than the strains resistant to other agents.

Conclusion Awareness of antimicrobial resistance pattern particularly is crucial for choosing an appropriate antimicrobial treatment and consequently minimizing hospitalization period, morbidity and mortality along with economic burden of nosocomial infection. Awareness of antimicrobial resistance pattern particularly is crucial for choosing an appropriate antimicrobial treatment and consequently minimizing hospitalization period, morbidity and mortality along with economic burden of nosocomial infection.

Conclusion Antimicrobial resistance is an increasingly emerging worldwide problem especially in ICUs, so identifying the resistance pattern of microorganisms in every hospital is therefore a key to success in appropriate treatment of patients. Antimicrobial resistance is an increasingly emerging worldwide problem especially in ICUs, so identifying the resistance pattern of microorganisms in every hospital is therefore a key to success in appropriate treatment of patients.

Dr.Hadadi Dr.Hadadi Dr.Rasoulinejad Dr.Rasoulinejad Dr.Yonesian Dr.Yonesian Dr.Maleki Dr.Maleki