Congenital Cytomegalovirus Infection : Association between Dried Blood Spots Viral Load Determined by Real-Time PCR and Long-Term Outcome

BACKGROUND: Cytomegalovirus (CMV) is the leading cause of congenital infection in humans and a major cause of sensorineural hearing loss and neurologic impairment in children. Several studies have been performed to identify predictive factors of short- and long-term outcome in congenitally CMV infected infants. A high viral load, as measured by real-time quantitative polymerase chain reaction (qPCR), either in fetal amniotic fluid or in neonatal blood and urine, has been reported to correlate with future damage in the fetus and in the child. OBJECTIVE: To define the prognostic value of qPCR performed on dried blood spots (DBS) collected on filter paper (Guthrie-card) in congenitally CMV infected infants. DESIGN/METHODS: CMV infected infants were followed-up for at least 3 years: virologic tests, clinical, psychometric, ophthalmologic and audiologic evaluations were performed at birth, at 3-6-12-18 months of age, and annually thereafter. Guthrie-cards performed in the first days of life for neonatal screening of genetic and metabolic disorders were subsequently obtained from the regional screening laboratory for all the study subjects and tested for CMV by qualitative and quantitative PCR (nested and qPCR). RESULTS: A cohort of 55 CMV infected infants were studied (mean GA 37,4 wks, mean BW 2750 g). 16/55 (29%) were symptomatic at birth; 7 of them (43,7%) developed sequelae. 4/39 (10,2%) asymptomatic infants developed sequelae. Nested PCR was positive in 98,2% (54/55) and qPCR in 94,5% (52/55) of all infants studied. The viral load determined by qPCR was 65 103 copies/mL in 66,1% of infants with symptoms at birth vs 34,9% of those without symptoms (p < 0,005) and in 61,5% of infants who developed sequelae vs 34,2% of those who did not (p < 0,05) (\u3c72 test). CONCLUSIONS: DBS test using real-time qPCR seems to be a useful method to identify infants at high risk of sequelae. The ability to identify children who are at increased risk for neurologic impairment should provide a basis for more timely and appropriate counseling for parents, permit careful monitoring and aid in formulation of interventional strategie

Cytomegalovirus (CMV) Infection Acquired through Breast Milk in Preterm Infants : Short- and Long-Term Outcome

BACKGROUND: Shedding of CMV into breast milk is the main source of CMV postnatal infection in early life. Many studies have shown a higher risk for premature infants, compared to term infants, of acquiring a symptomatic infection via this route. However, few studies have investigated the long-term neurological outcome of preterm infants with breast milk-acquired CMV infection. OBJECTIVE: To evaluate short- and long-term outcomes of CMV infection acquired by breast milk in preterm infants. DESIGN/METHODS: During a 36-month period we assessed maternal CMV serological status after preterm delivery 64 34 wks; vaginal swabs were taken on the day of delivery. We collected saliva and urine samples from all breastfed infants at 3 days of life, weekly until discharge and then monthly until babies were breastfed. At the same time breast milk samples were taken from the mothers. All samples were tested for CMV by \u201cshell vial\u201d culture (SVC) and by nested PCR. Children's blood was tested by PCR on day 1 of life to identify congenital CMV infection. CMV infected infants were included in a follow-up study until age 6 years. RESULTS: 78/94 (83%) women were CMV-seropositive; vaginal swabs were positive in 10/94 (10.6%) women who underwent caesarean section. CMV was detected in milk samples by PCR in 47 (66.2%) and by SVC in 43 (60.6%) of the 71 seropositive breastfeeding mothers. No infant born to CMV-seronegative mothers became infected. 96 neonates were born to the 71 CMV-seropositive lactating mothers: median GA 31 wks, median BW 1240 g. Congenital infection was diagnosed in 1 newborn. The mother to child transmission rate by breast milk was 29.8% (14 of 47 mothers), resulting in postnatal infection in 17/62 (27.5%) infants who received CMV-positive milk. In these 17 infants viral DNA or virus was detected in saliva or urine samples at a mean age of 49.7 days. Four (23.5%) of the 17 infants had clinical signs of infection: sepsis-like symptoms in 2 babies and sensorineural hearing loss at follow-up evaluation in the other 2 babies. CONCLUSIONS: In our study, the proportion of infants who acquired a CMV infection by breast milk was high and the incidence of neurological sequelae was far from negligible. It could be recommended to treat human milk from seropositive women to prevent CMV transmission to extremely small preterm infants

Developing potentially better practices to prevent neonatal nosocomial bloodstream infections

BACKGROUND: Nosocomial infections constitute a leading cause of morbility and mortality in Neonatal Intensive Care Units (NICUs). Late-onset sepsis (occurring after 3 days of age) is frequent in NICUs, particularly in very low birth weight infants (VLBWI). A systematic approach using quality improvement techniques can reduce the incidence of nosocomial sepsis. OBJECTIVE: 1)To evaluate the usefulness of \u201cPotentially Better Practices\u201d to reduce nosocomial sepsis in NICUs 2)To evaluate health-care workers (HCWs) compliance with new protocols. DESIGN/METHODS: Phase 1(June 2007-May 2008). Two \u201cPotentially Better Practices\u201d were evaluated: a) hand hygiene, promoting handrubbing with an alcohol based solution b) proper management of central lines, implementing hub care and reducing the duration of central venous catheter use. Education programs and promotion campaigns were performed to improve HCWs compliance. Phase 2(June 2008-May 2009). Application of the \u201cPotentially Better Practices\u201d developed. Adherence of HCWs to new protocols was evaluated by trained internal observers:1) proper handwashing before entering the ward by cameras 2) proper handwashing during patient care 3)bacterial colony counts on the right hand before patient care 4) proper central lines management 5) duration of central lines use. RESULTS: During the phase 1 we enrolled 597 neonates (mean GA 34 wks, mean BW 1910 g; VLBWI 120) and during the phase 2 574 neonates (mean GA 33 wks, mean BW 1895 g; VLBWI 116). The incidence of nosocomial sepsis decreased globally from 10% (phase 1) to 7.7% (phase 2). In VLBWI, the incidence of late-onset sepsis was 25.8% in phase 1 and 19.8% in phase 2. Late-onset sepsis decreased significantly in neonates with BW 500-749 g (93.3% vs 60%, p<0,02). The incidence of sepsis sustained by S. aureus, K. pneumoniae and E. faecalis/faecium decreased from 22.6 to 8.7%, 16.1 to 0, 12.9 to 4.3 respectively (p<0,01). HCWs adherence to new protocols gradually improved during the phase 2, reaching values above 80% for all the indicators considered. CONCLUSIONS: According to our results, the application of \u201cPotentially Better Practices\u201d is effective in reducing nosocomial sepsis in NICUs. The staff training is essential to improve HCWs adherence with infection control measures, resulting in a reduction of nosocomial infection

Distribution and high frequency of novel alleles at NF1 polymorphic markers in the Italian population

Segregation analysis of Neurofibromatosis type 1 (NF1) intragenic polymorphisms is a useful diagnostic tool for linkage analysis in familial cases and for the exclusion/detection of deletion in sporadic patients. We performed a segregation analysis of intragenic NF1 polymorphic markers in an Italian NF1 population consisting of 17 familial and 41 sporadic cases, for a total of 79 affected and 105 unaffected individuals. The haplotype in linkage with the mutation could be identified in all of the familial cases. Furthermore, an intragenic deletion was found in one sporadic case and confirmed by means of FISH using an NF1 IVS27 specific probe generated by a novel PCR procedure. In order to determine the allele frequencies at four NF1 polymorphisms in the Italian population, the unaffected family members and 25 unrelated Italian individuals were genotyped. Allele frequencies were found to be statistically different from those in the literature for markers IVS27AC28.4 and IVS38GT53.0. In addition four novel alleles were found in four unrelated subjects, and we observed a mutation during paternal gametogenesis in one case. These data suggest that NF1 polymorphic intragenic loci are unstable. It is unclear whether or not their marked instability may enhance the high mutation rate of the NF1 gen