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ARTICLES: Girija Natarajan, Yvette R. Johnson, Fan Zhang, Kang Mei Chen, and Maria J. Worsham Real-Time Polymerase Chain Reaction for the Rapid Detection of Group B Streptococcal Colonization in Neonates Pediatrics 2006; 118: 14-22 [Abstract] [Full text] [PDF]

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RT- Polymerase Chain Reaction Method for the Prevention of Early Onset Group B Streptococcal Disease

Marcello Lanari, Laura Serra, Francesca Cavrini, Giovanna Liguori, Elisa Storni, Vittorio Sambri   (2 September 2006)

RT- Polymerase Chain Reaction Method for the Prevention of Early Onset Group B Streptococcal Disease 2 September 2006
Marcello Lanari, MD, PhD Department of Pediatrics Santa Maria della Scaletta Hospital 4,via Montericco,40026 Imola (BO), Laura Serra, Francesca Cavrini, Giovanna Liguori, Elisa Storni, Vittorio Sambri Send letter to journal: Re: RT- Polymerase Chain Reaction Method for the Prevention of Early Onset Group B Streptococcal Disease m.lanari{at}ausl.imola.bo.it Marcello Lanari, et al.	To the Editor.- --We read with great interest the recent article by Natarajan et al regarding the real-time Polymerase Chain Reaction (RT-PCR) method for the detection of Streptococcus agalactiae (Group B streptococcus: GBS) colonization in neonates (1). The report focused on the question of the management of asymptomatic newborns from women with unknown GBS carriage status at delivery. --The 2002 Centers for Disease Control and Prevention’s guidelines provide a very complete and useful schedule based principally on a universal cultural screening to be performed on vaginal and rectal swabs obtained during the 35th-37th week of pregnancy and on an intrapartum antibiotic prophylaxis (IAP) for all the women identified as colonized by GBS. This approach has been proved efficacious and have conduced to a considerable reduction in the number of early onset GBS neonatal (EOGBS) disease (2 ). --Nevertheless, as stated by Natarajan and co-workers, the management of neonates born to women with unknown GBS colonization status at delivery is “vague”, such as in the event that IAP is not given or is “uncompleted” (i.e. one dose of antibiotic < 4 hours before delivery), despite an indication for the IAP (such as GBS proven colonization and/or other obstetric risk factors). It is still under debate if asymptomatic infants born to mothers with one or more risk factors in the absence of a known maternal GBS colonization status, should receive antibiotic prophylaxis; as an alternative, only neonates showing microbiological evidence of sepsis should be treated with specific anti-microbial therapy (3). Although a positive blood culture constitutes the microbiological diagnostic criterion for sepsis, it is well know that blood culture has a quite low sensitivity, in particular in newborns (4). In addition the mean time to achieve a positive result for blood culture is about 48 hours and this does not provide a fast diagnosis. The sensitivity, specificity and predictive values of others laboratory tests (i.e. C-reactive protein and white blood cell count) are variable and often quite low. The dilemma posed by these asymptomatic neonates is that the delay in initiating an antibiotic treatment, when it is needed, may significantly increase neonatal morbidity and mortality; on the other hand a widely prophylactic use of antibiotics immediately after birth will result in a “over treatment” of many of such infants who are not infected. At present sufficient data are not available to recommend a single management strategy . - In the Department of Pediatrics in Imola, we adopted the universal cultural screening-based strategy. as proposed by the CDC’s 2002 guidelines. From January 2003 to December 2005 we enrolled 3050 pregnant women and their children, recording notices about risk factors for infections during the pregnancy, labour and GBS colonization status. The percentage of GBS colonization in our population was 16% as detected by standard microbiologic culture, performed by using the Todd-Hewitt broth on vaginal and rectal swabs. Although widely efforts to enhance compliance to the CDC’s guidelines and implementation by clinical audits between medical and nursing staff responsible for prenatal and intrapartum care, GBS maternal carriage at delivery was unknown in 15% of the women entering the hospital for delivery. Among the group of women resulted eligible for IAP due to a positive GBS colonization , and/or as a consequence of other obstetric risk factors, only 64% received a “complete prophylaxis”. For asymptomatic term newborns from uncompleted or untreated GBS colonized mothers we used selective antibiotic treatment after the detection of a C- reactive protein increase, as a marker suggesting neonatal infection; instead a prophylactic antibiotic treatment is given to babies born to unknown GBS status mothers with obstetric risk factors. No case of EOGBS disease were identified in our cohort since the beginning of this management protocol. However more than 7% of the newborns underwent diagnostic evaluations and/or antibiotic treatment. -- Therefore, there is a great need for a rapid test to identify between at risk neonates those who are really infected and consequently those that must be properly and immediately treated. In addition, such a rapid test could limit the number of patients under clinical observation and investigation only to the “at documented risk” babies. - Recently a systematic review to determine the accuracy and rapidity of various intrapartum GBS colonization tests (5) concluded that RT-PCR and optical immunoassay (OIA) are candidate for rapid intrapartum GBS testing, in order to determine the need for IAP. RT-PCR is a fast test (less than 60 minutes) to be performed and the OIA is even faster (30 minutes). These methods used at the onset of labour allow to identify the GBS carriage status and to treat a high percentage of women that needed to receive an IAP. In addition, the management of newborns can be related to the 2002 CDC’s guidelines for babies born to mothers with a known GBS colonization status. --Natarajan et al., on the contrary, evaluated the use of a RT-PCR based method in a cohort of 94 neonates of ³ 36 weeks’ gestation born to women with unknown GBS status at delivery. These Authors used the RT-PCR to detect GBS colonized newborns and in particular they focused on the detection of S.agalactiae in multiple surface and gastric sites. They report that RT- PCR had a higher detection rate, when compared with traditional cultures (51% vs 17%) and consequently they concluded that RT- PCR was a sensitive method for GBS detection on surface sites and it would be a useful clinical tool in the management of those infants born to mothers with unknown colonization status. --We substantially disagree with the conclusion drawn by Natarajan since the knowledge of the GBS surface colonization status in newborns is only predictive (as reported by the Author himself ) of a risk ranging from 1% to 3% to develop a S.agalactiae invasive disease and, moreover, the datum of GBS superficial colonization does not allow to identify, among the colonized babies those that will develop a EOGBS disease. The Authors detected in the studied population an incidence of GBS colonized newborns, by RT-PCR, of 51%. We express some doubt about the clinical usefulness of such a datum, since such a huge amount of GBS colonized children are prospectively under evaluation to receive a specific antibiotic prophylaxis or to undergo further laboratoristic or microbiological investigation, to identify those at risk for a GBS invasive disease. --Following the suggestion of the Authors, we believe that a higher, but not truly necessary, number of prophylaxis would be given to these babies, thus resulting in a general over treatment of the newborn population. We developed a “home made” RT-PCR test for detection of S.agalactiae. The group B streptococci-specific RT-PCR assay was developed on a LightCycler system (Roche Diagnostics GmbH, Penzberg, Germany) using GBS-specific primers (Sag59 5’-TTTCACCAGCTGTATTAGAAGTA-3’, Sag190 5’- GTTCCCTGAACATTATCTTTGAT-3’) targeted to the cfb gene encoding the CAMP factor and two labeled adjacent hybridization probes (5’- AAGCCCAGCAAATGGCTCAAA-fluoresceine-3’, 5’- LC Red 640- GCTTGATCAAGATAGCATTCAGTTGA-phosphate-3’) specific for the GBS amplicon as already described (6). Recently, we started a prospective study by our “home made” RT-PCR to assess: i) the sensitivity of this test for the rapid detection of GBS vaginal colonization at the time of onset of labour, in comparison with standard culture; ii) the correlation between the tests results on maternal GBS colonization (by traditional culture and by RT-PCR) at 35-37 week of gestation and at the time of onset of labour; iii) to detect the GBS infection in neonates born to mothers uncompleted or untreated by IAP, by evaluating the presence of S.agalactiae in blood samples at birth. Up to today a total of 92 pregnant women and their children has been evaluated in this study. --Our preliminary results regarding the identification of S.agalactiae in vaginal swab obtained during labour showed that 14/92 (15.2%) women are colonized when tested by RT-PCR, whereas only 9/92 (9.8%) were positive by culture. In 7 patients GBS was identified by both methods. One patient was detected as positive only by culture, being the RT-PCR result negative. These are only preliminary results obtained on a small population, but the sensitivity of the RT-PCR assays was 90% and the specificity 94% as compared with the culture results; the positive and negative predictive value were 64% and 99% respectively. Due to these results, we think that the RT-PCR is a promising and clinically useful screening technique for the identification of the GBS maternal colonization. In addition, we suggest that a rapid intrapartum test comparable or better in sensitivity to standard culture, as the RT- PCR developed in our Laboratory, might represent the gold standard for the evaluation of women with unknown GBS status at delivery and for the consequent management of babies born to these mothers. For the group of the asymptomatic babies from women with unknown GBS status or uncompleted/untreated with IAP we suggest that the RT-PCR method on neonatal blood sample collected at birth might represent the appropriate technique to perform a more targeted, timely and cost-effective approach to neonatal management. REFERENCES 1.Natarajan G, Johnson YR, Chen KM, Worsham MJ. Real-time polymerase chain reaction for the rapid detection of group B streptococcal colonization in neonates. Pediatrics 2006;118:14-22 2.Centers for Disease Control and Prevention. Prevention of Perinatal Group B Streptococcal Disease. MMWR 2002;51:1-18 3.Ungerer RLS, Ricetto O, McGuire W, Saloojee H, Gulmezoglu AM. Prophylactic versus selective antibiotics for term newborn infants of mothers with risk factors for neonatal infection . Cochrane Database of Systematic Review 2004;Issue 4. Art. No. : CD003957.DOI: 10.1002/14651858.CD003957.pub2. 4.Gerdes JS. Diagnosis and management of bacterial infections in the neonate. Pediatr Clin N Am 2004;51:939-959 5.Honest H. Rapid tests for Group B Streptococcus Colonization in Laboring Women: A systematic Review. Pediatrics 2006;117:1055-1066 6.Ke D, Menard C, Picard F, Boissinot M, Ouellette M, Roy PH, Bergeron MG. Development of conventional and Real-Time PCR assay for the rapid detection of group B Streptococci. Clinical Chemistry 2000;46(3):324-331 Marcello Lanari, MD, PhD Laura Serra, MD Department of Pediatrics Santa Maria della Scaletta Hospital 4,via Montericco 40026 Imola (BO) ITALY Francesca Cavrini,B.Sc. Giovanna Liguori, B.Sc. Elisa Storni, B.Sc. Vittorio Sambri, M.D., Ph.D. DMCSS - Section Microbiology University of Bologna St.Orsola Hospital 9, via Massarenti 40138 Bologna ITALY Conflict of Interest: None declared