1. An outbreak of ESBL-producing Klebsiella pneumoniae blood stream infection in a neonatal intensive care unit at University Hospital in Egypt F. Amer, E. Elbehedy, H. Mohtady, R. Elbehedy*, A. Al-Hejin** Microbiology and Immunolgy Department, Pediatrics Departmetn*, Zagazig Faculty of Medicine, Zagazig, Egypt, Department of Biological Sciences, Faculty of Science**, King Abdulaziz University, Jedda, Kingdom of Saudi Arabia.

2. Background Aim of Study Materials and Methods Results Conclusions Contents

3. Background Infants in neonatal intensive care units (NICU) are subjected to multiple invasive procedures, and to excessive broad-spectrum antimicrobials. Such practices can lead to outbreaks of nosocomial infections.

4. Background In national and international research, special attention has been given to Neonatal bloodstream infections (BSIs). Of major concern are cases of BSIs caused by extended-spectrum ß-lactamase-producing Klebsiella pneumoniae (ESBL-KP). Markedly associated with treatment failure (Death). Nosocomial outbreaks  clonal or polyclonal spread. Risk factors for infections  determined to develop effective strategies aiming at curtailing outbreaks

5. Aim of Study To describe an outbreak of ESBL-producing K. pneumoniae blood stream infection in a neonatal intensive care unit. To conclude risk factors.

6. Methods Outbreak description: In September 2008, there was an increased number of cases of BSIs caused by ESBL-KP in the NICU of Zagazig University Hospital. This hospital is a 1030-bed, tertiary-care hospital in Zagazig City, Egypt. A case was defined as any neonate who acquired a BSI after at least three days of admission caused by ESBL-KP. Cases of blood stream infections were diagnosed according to CDC criteria for nosocomial infections. The outbreak period was between 5-31 September. The three months period before (June-August) were considered pre-outbreak period. Incidence of BSI during outbreak period was compared with the pre-outbreak period.

7. Methods Microbiological studies of patients and environment: samples cultured 1- Blood from NICU patients 2- Rectal swabs from neonates 3- 68 inanimate environmental samples. 4- hands of 43 HCWs (nurses, physicians, and housekeeping staff) All samples processed, and microorganisms identified by conventional methods and by API20E (biomeriux, Marcy l'Etoile, France).

8. Methods Antibiotic susceptibility of isolated K. pneumoniae was determined by the disk-diffusion methods. Antibiotics used (Oxoid, Basingstoke, UK): ampicillin, carbenicillin, chloramphenicol, cefotaxime, ceftriaxone, ceftazidime, aztreonam, cefepime, gentamicin, amikacin, sulphamethoxazole/trimethoprim, ciprofloxacin, meropenem, imipenem ESBL production was detected by the E-test strips (AB Biodisk, Solna, Sweden).

9. Methods Molecular typing : ESBL-KP  subjected to pulsed-field gel electrophoresis. K. pneumoniae ESBL ATCC700603 (control) Whole-cell DNA was extracted by commercially available kit. digested overnight by the restriction endonuclease Xba1. PFGE done in CHEF-DR III apparatus Lambda concatemers was used as molecular weight marker. PFGE Interpretation of DNA profiles visually by the criteria of Tenover et al. (1997).

10. Methods Case-control study: To determine possible risk factors for acquiring BSI with ESBL-producing Klebsiella pneumonia. Controls were defined as newborns who were admitted to the NICU during the same period but had no blood stream infection or BSI with organisms rather than ESBL-KP. For each case and control data about suspected risk factors were collected: birth weight, sex, gestational age, type of delivery, maternal premature rupture of membranes, place of delivery (our hospital, other hospitals or at home), instrumentation [endotracheal intubation, central venous catheter and umbilical catheter)], suction, mechanical ventilation, treatment with antibiotics, parenteral nutrition, NICU stay before bacteremia, severity of illness using the Severity of Neonatal Acute Physiology Scoring System.

11. Results Outbreak description: MonthJuneJulyAugustSeptember No of BSI cases67521 No of ESBL-KP. BSI1118 Organisms2 Candida, 2 E. coli, 1 CS 3 Candida, 2 E. coli, Psudomonas, 1 CS Candida, E. coli, K. pneumoniae, 2 CS Candida, E. coli, CS Admission no.25282026 Incidence24%25% 69.2% Table 1 Cases, admissions and incidence and aetiology of BSIs during the outbreak, and pre outbreak periods (CS: coagulase-negative Staphylococci) Pre-epidemic period Out-break period

12. Results Microbiological studies of patients and environmental: ESBL-producing K. pneumoniae isolated from All stools samples collected from neonates who had BSI. Hands of two nurses (N1 and N2 ). All strains isolated were susceptible only to ciprofloxacin, imipenem and meropenem.

13. Results Molecular typing: All ESBL-KP strains isolated were clonal

14. Results Case-control study: When the 18 case- and 36 control patients were compared, using univariate analysis, case-patients were more likely than controls to: 1- be borne from mothers who had premature rupture membrane, 2- to have endotracheal tube or central venous catheters, 3- to be exposed to suction procedures, 4- to receive aminoglycosides, 5- to stay in hospital for a long time, 6- to be cared by N1 or N2, When we include all the significant variables into a multivariate model, independent risk factors for acquiring ESBL-producing K. pneumoniae BSI were: Exposure to N1 (adjusted OR, 74.228; CI 95, 4.868-1131.892; P=0.002) Exposure to N2 (adjusted OR, 105.524; CI 95, 4.395-2533.713; P=0.004) Having endotracheal tube (adjusted OR, 18.901; CI 95, 1.442-247.772; P=0.025)

15. Conclusion The epidemic strain emerged as a result of a conversion of non ESBL-KP to ESBL-KP, based on the encountered wide, uncontrolled use of antibiotics.Then extensive colonization of the infants occurred. Clinically silent gastrointestinal colonization seemed to provide a reservoir for spread of ESBL-KP. Spread from baby to baby occurred by the hands of personnel. Using the PFGE technique, clonality among outbreak strains was demonstrated. All risk factors found could be related to nursing care either directly (exposure to N1 and N2), or indirectly [endotraheal tube (independent risk factor) and CVC, suction and length of stay (significant at a univariate level)].