ORIGINAL ARTICLE
Isolation of Serratia Marcescens from Blood Cultures in Paediatric Population at the Children’s Hospital Lahore MUHAMMAD ZUBAIR, AIZZA ZAFAR, GHULAM MURTAZA, NAIMA MEHDI, HUMAIRA JAVAID, HASAN EJAZ
ABSTRACT Objective: To determine the frequency of Serratia marcescens in positive blood cultures and its sensitivity pattern to various antibiotics in paediatric population. Methodology: The present study was conducted at The Children’s Hospital and The Institute of Child Health, Lahore. A total of 7680 blood cultures were processed for suspected cases of blood borne infections from different wards of The Children’s Hospital Lahore. The organisms were identified by routine laboratory tests including biochemical tests along with API 10S system. Antimicrobial susceptibility was determined by Kirby-Bauer disk diffusion method. Results: Out of total 7680 Blood cultures over a period of six months 1240 were positive blood cultures and the frequency of the Serratia marcescens was 17(1.4%) in positive samples. The gender distribution of S. marcescens was 13(76.5%) in male patients and in females 4(23.5%). These isolates were highly sensitive to amikacin, meropenem tazobactum/piperacillin and were resistant to amoxicillin-clavulanic acid and cephalosporins (ceftriaxone, cefuroxime, ceftazidime and cefotaxime). Conclusion: Although in this present study frequency of Serratia marcescens is low but these infections are a challenge for healthcare setting because of difficult treatment and multi drug resistance patterns to several antibiotics, thus physicians are left with a limited choice of antibiotics. Keywords: Serratia marcescens, blood borne infections, paediatric population
INTRODUCTION Hospital acquired infections caused by Serratia marcescens are an important cause of morbidity and mortality. They represent a major health problem and 1 an economic burden worldwide . S. marcescens is an important nosocomial pathogen, especially in neonatal intensive care units (NICUs). It may cause serious infections including septicemia and meningitis 2 in newborns . It is also associated with hospital acquired blood stream infections such as 3 endocarditis and pneumonia . After the application of preventive measures such as strict disinfection and sterilization procedures, the possibility of S. 4 marcescens infections has greatly decreased . S. marcescens potential virulence factors include various enzymes like protease, nuclease, lecithinase 5 and the hemolysins . Besides these enzymes, lipopoylsaccharide (LPS) located in the outer membrane of the organism is also an established virulence factor. S. marcescens produces several extracellular products including serrawettin, a wetting 6 agent that helps in the colonization of this organism . Serratia marcescens infections are not easy to treat because of the emergence of multiple drug resistant strains. S. marcescens is resistant to ampicillin, and first two generation of cephalosporins. ---------------------------------------------------------------------Department of Microbiology, The Children’s Hospital and Institute of Child Health, Lahore, Pakistan Correspondence to Dr. Hasan Ejaz, E-mail:
[email protected]
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It is also sensitive to aminoglycosides . Ciprofloxacin is the drug of choice and has proved to be a useful antibiotic in S. marcescens infected patients. Amikacin is suggested for all hospital acquired 8 infections of Serratia marcescens.
METHODOLOGY This study was performed at the Department of Microbiology, the Children’s Hospital and the Institute of Child Health, Lahore from June 2012 to November 2012. Paediatric patients with suspected cases of septicemia from both genders up to the age of 16 years were included in this study. A total number of 7680 Blood culture samples were processed to determine the frequency and sensitivity pattern of Serratia marcescens from positive blood cultures. A strict aseptic technique was applied so that the bacteria from the skin do not contaminate the Blood culture bottles. Blood culture bottles were incubated at 37°C for up to seven days. Bacterial growth was evidenced by turbidity, haemolysis, clot formation, gas production or pellicle formation. Blood culture bottles were sub-cultured on Blood agar and MacConkey agar. After 7 days, all negative blood culture bottles were discarded. Identification of bacteria was performed by colonial morphology, Gram staining and biochemical tests such as: Citrate test, Oxidase test, Triple Sugar Iron test, Urease test 9 and by API 10S system (bioMerieux) . On
Muhammad Zubair, Aizza Zafar, Ghulam Murtaza et al
MacConkey agar colonies appeared as non-lactose 10 fermenting . Serratia marcescens appeared as a metallic sheen on blood agar. A four digit numerical profile was obtained and was compared with API 10 S identification book/software to confirm S. marcescens. Antimicrobial Susceptibility test was performed by Kirby-Bauer disk diffusion technique according to CLSI, 2011. Suspensions of the bacterial colonies were prepared against McFarland 0.5 turbidity standard. Sterilized wire loop was dipped in bacterial suspension and inoculated on the centre of the Mueller Hinton agar plate. Then with a sterile cotton swab the colonies were streaked on the plate starting from the centre. The appropriate antimicrobial discs amoxicillin, amikacin, ceftazidime, ciprofloxacin, ceftriaxone, cefotaxime, cefuroxime, meropenem, sulbactam, tazobactum/piperacillin were placed and o the plate was incubated at 37 C for overnight. After overnight incubation the diameter of each zone of inhibition was measured in mm. The endpoint of inhibition was where the growth started. The interpretation of zone sizes of each antimicrobial disc was made, as Sensitive, Intermediate Sensitive or Table 2: Antimicrobial Susceptibility of Serratia marcescens Antibiotics Sensitive Amikacin 15(88.2%) Meropenem 12(70.58%) Tazobactum/Piperacillin 11(64.7%) Ciprofloxacin 10(58.8%) Sulbactam/Cefoparazone 7(41.2%) Ceftazidime 2(11.8%) Ceftriaxone 2(11.8%) Cefotaxime 2(11.8%) Cefuroxime 2(11.8%) Amoxicillin-Clavulanic Acid 1(5.9%)
DISCUSSION Serratia marcescens has become an important opportunistic pathogen in hospitalized individuals worldwide, particularly in neonatal intensive care units. Patients who are hospitalized for extended periods are frequently colonized by this organism and are at high risk of developing infections. Due to possession of multiple virulence factors and resistance to several therapeutic antibiotics, it is very difficult to treat patients infected with S. marcescens. According to the present study, 7680 samples of Blood cultures were collected from various wards of the Children’s Hospital Lahore, out of which 1240 (16.1%) were positive and 6440 (83.8%) were negative for bacterial growth. S. marcescens was isolated in 17 (1.4%) samples whereas 1223(98.6%) samples contain organisms other than S.
Resistant using interpretation chart of zone sizes of CLSI guidelines.
RESULTS During the study period 1240 cases were positive cultures, out of which 17 were Serratia marcescens. The frequency of Serratia marcescens in positive blood cultures was 1.4% (Table 1). There were 13 (76.5%) male and 4(23.5%) female patients. Serratia marcescens was found to be highly sensitive to amikacin (88.2%), meropenem (70.6%), ciprofloxacillin (58.8%) and tazobactum-piperacillin (64.7%). The sensitivity pattern to other antimicrobial discs was sulbactam (41.2%), ceftriaxone (11.8%), ceftazidime (11.8%), cefuroxime (11.8%), cefotaxime (11.8%) and amoxicillin-clavulanic acid (5.9%) (Table 2). Table 1: Frequency of Serratia marcescens (n=1240) Organism Frequency %age Serratia marcescens 17 1.4 Bacteria other than S. 1223 98.6 marcescens
Intermediate sensitive 1(5.9%) 2(11.8%) 0(0%) 1(5.9%) 5(29.4%) 0(0%) 1(5.9%) 1(5.9%) 0(0%) 0(0%)
Resistant 1(5.9%) 3(17.6%) 6(35.3%) 6(35.3%) 5(29.4%) 15(88.2%) 14(82.4%) 14(82.4%) 15(88.2%) 16(94.1%)
marcescens. The highest proportion of S. marcescens was found in Neonatal Nursery Unit from which 6 (35.3%) isolates were followed by Medical ward and Medical Intensive Care Unit each with 4 (23.5%) isolates, Nephrology ward, Cardiac Intensive Care Unit and Gastroenterology ward each with 1 (5.9%) isolate. Serratia marcescens is one of the important nosocomial pathogens that cause infections like septicemia, wound infections, sinusitis, septic arthritis, peritonitis, conjunctivitis and meningitis in the hospitalized patients of various severities. Infections caused by S. marcescens may be difficult to treat because of resistance to variety of antibiotics, including ampicillin, first and second generation cephalosporins. Aminoglycosides have good activity against S. marcescens, but resistant strains have also been reported.
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Isolation of Serratia Marcescens from Blood Cultures in paediatric population
Despite advances in supportive care and use of antibiotics, nosocomial sepsis is one of the most important causes of neonatal death with significant mortality and morbidity. The production of different enzymes by S. marcescens as virulence factors were also reported including chitinase, lipase, chloroperoxidase and an extracellular protein. Many other aspects of the pathogenicity and virulence of S. marcescens were also studied including adherence, hydrophobicity, lipopolysaccharide (LPS) and 6 extracellular products . A study was conducted to determine the S. marcescens bacteremia in a neonatal intensive care unit at the Yale New Haven Hospital London. The mortality rate was high in infected neonatal patients. Twenty five cases of S. marcescens bacteremia isolates were evaluated. Eighty six percent (86%) of S. marcescens isolates were resistant to ampicillin and none was resistant to gentamycin and third 11 generation cephalosporins . These studies support the results of present study. Another study was performed to determine the outbreaks of Serratia marcescens in a neonatal intensive care unit at the University Children’s Hospital, Zurich. Out of twenty patients eleven were found to colonize S. marcescens. It was found that S. marcescens is responsible for nosocomial infections in neonatal units with significant morbidity and 12 mortality . A research was conducted on neonatal sepsis caused by Gram negative bacteria in a neonatal intensive care unit, a six years analysis from 2002 to 2008. In this study there were 143 cases of neonatal sepsis caused by different Gram negative bacteria. Among these 143 cases, 81% had nosocomial neonatal sepsis and S. marcescens was the most common pathogen. Levofloxacin (97.4%), meropenem (97.1%), imipenem (95.6%), ciprofloxacin (95.4%) and amikacin (94.8%) were the 13 most sensitive . Another study was conducted for the treatment of S. marcescens meningitis with meropenem in patients of central nervous system disease. Almost 25-40% central nervous system infections were caused by Gram negative bacteria and S. marcescens infection of central nervous system was infrequent (2%). The typical sensitivity profile of S. marcescens revealed that it was resistant to ampicillin and cefazolin and sensitive to ciprofloxacin, levofloxacin, meropenem, and trimethoprim/ 14 sulfamethoxazole . A study on extended epidemics of hospital acquired infections caused by S. marcescens was conducted at the Chang Gung Memorial Hospital Taoyuan, Taiwan. This study was based on review of the laboratory and infection control record from 1991-
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2002 to determine the frequency of infections by S. marcescens. During 12 years 14,172 S. marcescens infections were reported. These were 100% resistance to ampicillin, cefuroxime, chloramphenicol, and tetracycline for S. marcescens isolates. Amikacin, ciprofloxacin and imipenem were 100% 15. sensitive to S. marcescens Another study was designed for the investigation of nosocomial outbreak of S. marcescens in a neonatal intensive care unit at King Hussain Medical Centre. Thirty-one cases of Serratia marcescens sepsis were identified, 19(61%) neonates were males and 12(39%) were females; the median age at presentation was six days. Neonates with Serratia marcescens sepsis were found to have low birth weight, low gestational age, and high rate of mechanical ventilation and central venous 16. catheterization with high overall mortality rate Between December 2000 and January 2001 Serratia marcescens was isolated from ten patients (eight from NICU and two from nursery). All isolates were from blood and were sensitive to imipenem, 17 meropenem, amikacin and ciprofloxacin . S. marcescens causes significant morbidity and mortality in preterm neonates. Cross-transmission via transient hand carriage appeared to be the probable route of transmission in NICU. Antimicrobial susceptibility patterns did not prove to be an accurate 18 predictor of strain relatedness for S. marcescens . Another study was designed to investigate the outbreak involved 210 patients, 61(29%) developed an infection; among them 16 were invasive infections. Multivariate analysis identified gestational age, arterial catheter use and antibiotic treatment as independent risk factors for colonization and infection with Serratia marcescens. Molecular typing was also 19 performed . Another study was performed to determine the Serratia marcescens infections outcomes. The most frequently observed infections were respiratory tract infection (32.5%). Seventythree (94.9%) patients were cured. Four deaths were observed; 3 of them were considered as attributed to the Serratia infection. In addition, antipseudomonal penicillins, as well as their combinations with betalactamase inhibitors, carbapenems, and fluoroquinolones exhibited high antimicrobial activity 20 against Serratia marcescens .
CONCLUSION Despite low frequency of S. marcescens, it is difficult to treat patients because of the resistance pattern to many available antibiotics. So proper infection control measures should be taken to keep its frequency low because the transmission of Serratia marcescens can occur through direct contact with infected
Muhammad Zubair, Aizza Zafar, Ghulam Murtaza et al
patients, fomites and with polluted hospital environment. This culture and susceptibility data will help the clinicians to choose the best antibiotics to treat the infections caused by S. marcescens.
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