P1548 Rapid molecular detection of bacterial species and resistance determination in paediatric blood cultures

S434 Methods: Our prospective study included all neonates with suspected sepsis born in our Department of neonatology in a two year’s period (2004 and 2005). Microbiological identification and susceptibility were performed with routine methods. Results: A total of 8,356 neonates were evaluated during this study period and 167 patients fulfilled the eligible criteria. Three most frequent bacterial strains were Staphylococcus epidermidis, Escherichia coli and Group B streptococci. Less frequent were Staphylococcus aureus, Klebsiella pneumoniae and Pseudomonas aeruginosa. In 21/167 (12.57%) patients initial antibiotic treatment was inappropriate, and the most resistant strains were S. aureus (8) followed by K. pneumoniae (7), E. coli (4) and P. aeruginosa (2). Discussion: the initial use of appropriate antibiotics, before the detection of bacteria, is very useful, it reduces the time of therapy, length of stay and costs of treatment. Conclusion: It is recommended for each neonatal department to determine their own spectrum of bacteria and to adjust the initial antibiotic treatment to the findings which are typical for the unit. But, also repeated surveillance is necessary in order to follow the longitudinal changes of the bacterial spectra due to the resistance acquired. P1548 Rapid molecular detection of bacterial species and resistance determination in paediatric blood cultures D. Menichella, P. Bernaschi, B. Lucignano, C. Manfredini, C. Russo (Rome, IT) Bacteraemia is one of the most serious and potentially life-threatening infection disease in children, specially in case of septic shock. Signs or symptoms of bacteraemia are highly variable in childhood and diagnosis is routinely supported by blood culture. In our study, we evaluated a new molecular genetic assay (GenoType BC Gram-positive® and GenoType BC Gram-negative® – Hain Lifescience) as rapid tool for identification of most of bacteria involved in bloodstream infections in children suspected to have bacteraemia, hospitalised at “Bambino Ges`u” Children Hospital (Rome, Italy) and to improve the work-flow TAT in a microbiology laboratory. At instrumental positive time (BACTEC 9240 Becton Dikinson) all the bottles (PEDS PLUS/F) were evaluated with microscopy Gram stain and subcultured in agar standard media. Phenotypic identification was obtained on colonies grown after 24 hours of incubation by using the VITEK® 2 (bioM´erieux) automated system. For molecular identification 20 mL of blood positive culture were spotted on GENO-CARD® and after a drying step, a punch of 1 mm in diameter was transferred into the PCR mix followed by thermal-cycling amplification. Hybridisation protocol was performed as recommended using a shaking incubator and results were obtained by comparison of the hybridisation pattern present on the test strip with the ones of the provided evaluation sheet. 148 positive blood culture bottles representing 106 Gram-positive cocci, 28 Gram-negative bacilli and 14 mixed infections were included in this study. At start time we performed all of the proceduring steps as manufacturing instructions and the GenoType BC system correctly amplified and identified 41/49 Gram-positive cocci (sensitivity 83%); 8 Staphylococci were not amplified. 17/17 Gram-negative bacilli were correctly amplified and identified (sensitivity 100%). In the effort to optimised the GenoType assay we improved by in house steps the direct DNA extraction. Finally all of 53 Gram-positive cocci, 11 Gram-negative bacilli were correctly amplified, 4 K. kristinae were correctly not amplified (sensitivity and specificity 100%). About 14 mixed cultures were correctly identified and for 1 of them the GenoType BC assay recognized a double infection (P. aeruginosa and S. maltophilia) demonstrated only after this result by a new subculturing. GenoType BC assay is very helpful in the rapid diagnosis of paediatric blood stream infections in effort to prevent severe sepsis and mortality.

17th ECCMID / 25th ICC, Posters P1549 Genetic factors in the development of unfavourable effects of intrauterine infections on the course of neonatal periods in neonates N. Gorovenko, Z. Rossokha, S. Podolskaya (Kiev, UA) Objectives: Intrauterine infection in neonates is an important factor of perinatal risk in the development of complications during neonatal period. The latent forms of these infections are not always when effecting neonates. In a great number of studies the genes deletion polymorphism of glutathione-S-transferases was associated with higher sensitivity to the influence of many factors. The aim of this research was to study the prevalence of intrauterine infections and GSTT1 genes allelic polymorphism in neonates. Methods: The prevalence of Toxoplasma gondii, Chlamydia trachomatis, Mycoplasma hominis, Herpesvirus I/II types, Cytomegalovirus in the blood of neonates with perinatal pathologies and neonates from the comparative group (clinical healthy neonates) was tested by polymerase chain reaction (PCR) method. The allelic polymorphism of GSTT1 genes was investigated by means of multiplex PCR in neonates. Differences in these groups were assessed by the Pearson chi-square analyses. Results: There were detected latent forms of intrauterine infections in 74 of 117 neonates with perinatal pathologies (63.24%) as compared to the comparative group of the neonates where the frequency of latent forms accounted for 9 of 70 neonates (12.85%). In our investigation we have observed an increased frequency of GSTT1 null genotypes among infected neonates (31.32%; 26 of 83 individuals were homozygous for GSTT1 deletion genotype) as compared to uninfected neonates (16.98%; 18 of 104 individuals were homozygous for GSTT1 deletion genotype). The frequency of deletion polymorphism of GSTT1 genes was significantly increased among infected neonates of both groups (4.59, P < 0.05), i.e. GSTT1 null genotype was associated with the development of intrauterine infections. The GSTT1 frequency of deletion polymorphisms was significantly (6.66, P < 0.01) increased among infected neonates with perinatal pathologies (33.78%; 25 of 74 individuals were homozygous for GSTT1 deletion genotype) as compared to uninfected clinical healthy neonates (13.11%; 8 of 61 individuals were homozygous for GSTT1 deletion genotype). Conclusion: Thus, the deletion polymorphism of GSTT1 genes in neonates stipulates the predisposition to the intrauterine infections causes its effect with further development of perinatal pathologies.

Paediatric viral infections P1550 Features of norovirus and rotavirus infections in small children I. Narkeviciute, I. Tamusauskaite, G. Bernatoniene (Vilnius, LT) Objectives: No comparison studies of clinical presentation of norovirus and rotavirus infection in small children were ever made. We investigated norovirus infection features in children under three year’s age and compared the results with rotavirus infection in the children of the same age. Methods: The random selection and retrospective analysis of 70 norovirus and 70 rotavirus-infected children‘s case notes were done. All children were treated in Vilnius University Children Hospital in 2005. The norovirus antigen was assayed using ELISA, rotavirus – using immunochromatography diagnostic assay. Results: In small children norovirus infection manifested as vomiting (94.3%), diarrhoea (81.4%) and pyrexia (65.7%). It manifested as either gastroenteritis with pyrexia (47%) or gastroenteritis alone (30% of all cases) while 18.6% cases were without diarrhoea. Granulocytosis was found in 72.7% of blood results. Pyrexia was present in 97.1% case of rotavirus infection and 80.9% were >38ºC. However, in norovirus infection accordingly 65.7% and 47.8% (p < 0.0001). Diarrhoea (4 times/d) more frequently appeared in children with rotavirus infection than with norovirus (p < 0.0001). Repeated vomiting (4 times/d) has been more common for children with norovirus infection. As opposed to norovirus