Multidrug-resistant Pseudomonas aeruginosa strains in Tehran Reference Burn Hospital , Tehran , Iran

Pseudomonas aeruginosa is one of the most common causes of burn wound infections. The aim of this study was to determine the frequency of antibiotic resistance burn morbidity during September 2005 to October 2007. The incidence of multi-drug resistant P. aeruginosa infection was indicated in 9238 patients admitted to Tehran Reference Burn Hospital. Specimens were collected after admitting the patients in the hospital wards. Susceptibility test were performed for all isolate. Specimens were collected based on hospital policy and cultures were performed on each patient. Isolated strains were identified by using standard bacteriological methods for isolation and identification of the causative agents. P. aeruginosa was identified in 3012 (70.5%), Staphylococcus aureus in 581 (13.6%), Acinetobacter 426 (9.9%), the rest organisms were 279 (6%). Frequencies resistant of applied antibiotics were tobramycin (82%), ceftazidim (78%), ceftizoxime (82%), ciprofloxacin (72%), amikacin (73%), gentamycin (80%), tetracyclin (60%), teazabactam (81%) and cotrimocxazol (98%), respectively for P. aeruginosa. Due to the high multi resistant rate in pseudomonas infection, there is need to immediately revise the hospital policy in control of nosocomial infection and treatment strategy such as more efficient antiseptic agents or new antibiotics used to reduce the rate of multidrug resistant.


INTRODUCTION
Resistance to antimicrobial agents is a global problem with the particular level of resistance.Yet, the problems posed by antimicrobial resistance in the burn population are not limited to a few microorganisms (Khosravani et al., 2008).Besides the Gram-positive microorganisms, a number of Gram negative bacteria are losing their susceptibility to mainstay antibiotics, as well (Rastegar 1998;Simor et al., 2002).Ceftazidime resistant pneumonia increased from 0 to 16.6%.Cefoperazone and ceftazidime resistant Enterobacteriacea were up to 22%.Also 13% of the P. aeruginosa were resistant to cefoperazone (Obritsch et al., 2005;Lee et al. 2006).As resistance develops, out breaks occur.Burn victims are obviously at high risk for nosocomial infection due to the *Corresponding author.E-mail: massoudhajia@yahoo.com.Fax: +982166728121.nature of the burn injury itself.Bacterial infections in burn patients are widely known.The time related changes in the predominant flora of the burn wound from Grampositive to Gram-negative recapitulate the history of burn wound infection.Selection and dissemination of intrinsic and acquired resistance mechanisms increase the probability of burn wound colonization by resistant species such as P. aeruginosa (Bonomo and Szabo, 2006;El'Garch, 2007;Lister et al., 2009).
However, everything from minor out-breaks to major epidemics of antibiotic resistant, P. aeruginosa remains frequently reported as a multi-resistant organisms (Hajia, et al., 2008;CLSI, 2006) with major cause of burn injury colonization and serious wound infections, which have been reported in Tehran Reference Burn Hospital as well as Burn Centers around the world (Hussein et al., 1989).The purpose of this study was to assess resistant pattern of applied antibiotics in admitted patients at Tehran Reference Burn (TRB) Hospital.

Studied group
A retrospective study was conducted during September 2005 to October 2007 on burn patients who were admitted in Tehran Reference Burn (TBR) Hospital.In the present study, a total 9238 admitted patients were examined for bacteriological investigation during study period.Those patients who had positive culture with P. aeruginosa was considered in this study.

TBR Hospital policy for admitting patients
The policy of our burn hospital is to admit male patients ≥ 20% total body surface area (TBSA) burns.These patients are mainly burned by different kinds of burning agents such as electrical and chemical contacts, or building firing.The first step for admitted patients is to apply Silver Sulphadiazine topically on the burnt patients after sampling which is the hospital policy for admitted patients at the burning ward.Dressing was also changed daily.Cephalothin and Amikacin were administered as first line of antibiotics from the first day of admission in-patients with ≥ 20% TBSA burns (II and III degrees).The wound was inspected daily during the dressing changes.The data gathered in the study was stored and processed by a computer database.

Specimens and sampling procedure
All wound and blood specimens were collected by sterile swabs from registered patients.

Culture and isolation
All samples were cultured on sheep blood agar and Eosin methylene blue.All isolated organisms were identified by using standard bacteriological protocols (Winn et al., 2006).

Antimicrobial susceptibility test
The antimicrobial activity against P. aeruginosa and other organisms was tested with the disk diffusion method of Kirby-Bauer recommended by clinical laboratory standard institute (CLSI, 2006) using available paper disc (Hi-Media).Applied standard organisms: Following organisms were used as quality control strains.Organisms for quality control of disk diffusion method were E. coli (ATCC 25922), S. aureus (ATCC 25923), and P. aeruginosa (ATCC 27853).Standard organisms for checking the quality of the Mueller-Hinton medium were the above mentioned organisms plus Enterococcus faecalis (ATCC 29212) (CLSI, 2006).The used antibiotics were as follow: amikacin, gentamicin, tobramycin, ceftazidime, ceftizoxime, trimethoprim/sulfamethoxazole, ciprofloxacin, tetracycline, and tazabactam.

RESULTS
Among 9238 microbiologic samples, which were taken during the study period, bacterial strains were isolated and the frequency of P. aeruginosa (3012 strains) was found to be 70.5%.This was followed by Staphylococcus aureus (581 strains) 13.6%, Acinetobacter (426 strains) 9.9%, with other microorganisms(279 strains) 6%.P. aeruginosa were isolated from either wound or blood samples of all these patients.These patients were aged up to 72 years old with the mean 34.2 ± 20.18.
Quality control of disk diffusion were performed each time of runing the test to ensure relibility of applied antibiotics with E. coli (ATCC 25922), S. aureus (ATCC 25923), and P. aeruginosa (ATCC 27853) as it was mentioned in materials and methods.
The frequency of resistant P. aeruginosa strains was 60 to 98% for the examined antibiotics in 3012 isolates.82% of the isolates were resistant to tree and 80% was resistant to 5 or more antibiotics.

DISCUSSION
P. aeruginosa was the most common pathogen causing wound infection compatible with other reports, especially from developing countries (Hajia et al., 2008;Rahbar 2010).There was a significantly higher mortality rate in the Pseudomonas group (33% vs. 8%, p < 0.001) compared with other isolated organisms.Amount of blood products used, length of stay, number of surgical procedures and the cost of care were all significantly higher in the Pseudomonas (Armour et al., 2007).In our burn hospital the high incidence of P. aeruginosa infections and the widespread of high resistance to antibiotics, may be one of the most important influences on the mortality rate in burned patients.
Burns continue to be a major environmental factor responsible for significant morbidity and mortality in developing countries (Weber and Rutala, 1999).The key to control antibiotic -resistant pathogens in the burn hospital is rigorous adherence to infection control guidelines and prevention of antibiotic misuse.Antibiotic restriction policies clearly result in reduced drug costs.Prevention strategies are based on developing a program to prevent or reduce antimicrobial resistance.It indicates the necessity for urgent measures to be taken to restrict the spread of the species in the units and to limit administration of antimicrobial agents including; complete isolation of the contaminated patients, applying highly effective disinfectants and hygienic procedures.The Society for Health Care Epidemiology of America / Infectious Disease Society of America (SHEA / IDSA) guidelines have reviewed specific methods to implement antibiotic control policies.The goal is that, all patients to receive the most effective drug with least toxic effect, and least costly antibiotic for the precise period needed to cure or prevent infection (Weber and Rutala, 1999).
Factors affecting the increase and dissemination of antimicrobial resistance can be divided into transfer of resistance genes from one microbe to another, and mutation of existing genes to more resistant variants by the over-use and misuse of antimicrobial, increase infection control measures.Recently researchers have noted to Molecular typing methods to find out about resources of the nosocomial infections.Pulse field gel electrophoresis has been reported successfully for this purpose (Lambiase et al., 2009).
Besides, hospital staff might consider monitoring used antibiotic with its resistance pattern and then decreasing the use of specific antimicrobial agents (Wang et al., 2003;Winn et al., 2006).The careless health care workers are often the main vectors to promote the dissemination of nosocomial infections.

Conclusion
Analysis of results revealed high multi resistant rate in pseudomonas infection, although is lower than similar report from Iran.Therefore, it seems revision of hospital policy for more control of nosocomial infection and treatment strategy is still required as applying more efficient antiseptic agents or new antibiotics.