Early diagnosis of neonatal sepsis: the role of 16S rRNA gene sequence analysis

Early diagnosis and treatment of newborn infants with suspected sepsis are required to reduce sepsis associated mortality and to prevent severe life-threatening complications. Clinical diagnosis is difficult and, blood culture, is often negative in the face of strong clinical indicators of septicemia, on the other hand, molecular diagnostics have proved to be a valuable adjunct for detection of neonatal bacteremia. The purpose of this study was to evaluate the role of l6S rRNA gene sequencing for rapid and sensitive diagnosis of clinically septic neonates with pathogen identification to allow for early and specific treatment of neonatal infections. A prospective study conducted in both NICUs of Kasr Al Aini hospital and Children's hospital, Cairo University over a period of 3 months from April 2008 to June 2008, Fifty neonates undergoing sepsis evaluation were included in this study. Risk factors, clinical manifestations and hematological findings suggestive of sepsis were recorded. Bloodculture and broad range 16S rRNA polymerase chain reaction [PCR] were collected from clinically septic neonates to confirm diagnosis of sepsis. Genotypic identification of bacteria by sequence analysis of the 16S rRNA gene was performed on all samples positive in PCR. Fifty neonates suspected clinically of having sepsis were included in the study. 61 samples were collected as a part of evaluation of infection, 50 samples upon admission and 11 samples as follow-up due to persistence of clinical manifestations. The rate of culture proven sepsis was 19.7% [12/61]. With the molecular method of broad range 16S rRNA PCR the detection of bacteria improved to 29.5% [18/61]. PCR revealed sensitivity, specificity, positive predictive value and negative predictive value of 91.7%, 85.4%. 61% and 97.6% respectively, while the accuracy of this test was 86.7%. so, compared to culture, the 16S rRNA PCR demonstrated a high negative predictive value for ruling out neonatal sepsis. According to the PCR results, hyperthermia, feeding intolerance and abdominal distension were found to be significant among neonates with positive PCR. As for hematological manifestations, we found a significant association between increasing number of abnormal hematological findings and PCR positivity [p=0.006, p for ordinal correlation =0.01]. Odds ratio for having 2 or more abnormal hematological findings and PCR positivity was 3.77 [95% CI=l.2-12.3]. Sequence analysis was done for further identification of organism on all samples positive in PCR. The sequenced PCR was in accordance with blood culture results in 10 samples while one sample showed conflicting results [91% agreement], Seven samples showed no growth by culture but were identified by sequence analysis. The sequencing-based identification of these isolates included: Enterobacter cloacae. Staphylococcus carnosus, Proteus mirabilis, Haemophilus influeuzae, Streptococcus pyogenes and Klebsiella sp. One sample yielded insufficient DNA to be analyzed. Broad range 16S rRNAPCR can he used to rule out sepsis. Both PCR and sequence analysis can provide additional diagnostic data that cannot he obtained with the use of broad-range PCR or routine laboratory tests alone. In neonates with clinically suspected sepsis, a primary decision could be made when the PCR results are completed, and within few hours, genotypic identification from the sequencing could be available to allow for early treatment

value of routine screening of TT virus in blood transfusion practice

To clarify the prevalence of TT virus [TTV] infection in blood donors as well as recipients to predict the value of its routine screening and to minimize the incidence of transfusion associated hepatitis [TAH], the present study included 180 subjects divided into three groups. Group A included 50 consecutive healthy blood donors with elevated ALT at the time of donation and group B included 100 consecutive healthy volunteer blood donors with normal ALT; both groups were negative for the serological markers for hepatitis A, hepatitis B and hepatitis C viruses. Group C included 30 blood product recipients in whom post-transfusion follow up revealed elevated ALT and negative hepatitis A-C markers. All subjects were subjected to laboratory evaluation in the form of serum ALT as well as a panel of A-C hepatitis viruses studies which was carried out using ELISA II technique to detect HbsAG, anti HCV antibodies and anti HAV IgM. TTV DNA was amplified and detected using polymerase chain reaction, followed by gel electrophoresis

Secretory otitis media and local treatment with mucolytics

Thirty-one patients suffering from secretory otitis media for more than 3 months have been locally treated with mucolytic injections. The treatment was in the form of 0.1-0.3 cc of bromhexine [Bisolvon] injection into the middle ear cavity. The course of treatment was 3-5 injections at 3 days interval. Clinically and audiologically, at the end of the course were satisfactory. It clarified that this line of medication is effective against the disease. The improved cases were followed up for 6 months with no recurrence in almost all cases. The advantages of this simple therapy have been mentioned, and its success rate was compared with the surgical treatment by ventilating tubes