Rangmar Goelz

Cytomegalovirus (CMV) inactivation in breast milk: reassessment of pasteurization and freeze-thawing.

Breast-feeding mothers frequently transmit cytomegalovirus (CMV) to preterm infants of very low birth weight. Current recommendations for prevention of virus transmission are based on data published 20 y ago in the context of human milk banking. Two recent clinical trials examined storage of breast milk at −20°C to reduce virus transmission. However, in both studies, CMV transmission occurred. Using sensitive tools like quantitative PCR, CMV pp67 late mRNA assay, and a high-speed, centrifugation-based microculture assay for quantification of CMV infectivity, we reassessed the virological and biochemical characteristics of freeze-storing breast milk at −20°C, compared it with traditional Holder pasteurization (30 min at 62.5°C), and a new short-term pasteurization (5 s at 72°C) based on the generation of a milk film. Both heat treatment procedures were able to destroy viral infectivity and pp67 RNA completely. Preliminary results showed short-term heat inactivation below 72°C was less harmful in reducing the activity of marker enzymes than Holder pasteurization. Freezing breast milk preserved the biochemical and immunologic quality of the milk; however, late viral RNA and viral infectivity was also preserved. Compared with viral DNA, CMV-RNA more directly reflects infectious CMV in human milk samples. Further studies are necessary to evaluate short-term heat treatment below 72°C as an effective tool for prevention of CMV transmission.

Cytomegalovirus transmission to preterm infants during lactation.

Breastfeeding has a major impact on HCMV epidemiology. The incidence of postnatal HCMV reactivation during lactation equals the maternal seroprevalence. Infectious virus, viral DNA and RNA can be isolated easily from cell and fat-free milk whey. Early onset of viral DNAlactia and virolactia as well as high viral load in milk whey are maternal risk factors for virus transmission. The dynamics of HCMV reactivation can be described by unimodal kinetics with interindividual variation. Virus reactivation during lactation is a self-limiting local process in the absence of systemic HCMV infection. Preterm infants below 1000g birthweight and a gestational age below 30 weeks may be at high risk of acquiring a symptomatic HCMV infection. Several recent studies described low transmission rates and mostly asymptomatically infected neonates using frozen milk. Despite different freeze-storing procedures, HCMV transmissions occurred, and severe HCMV infections were observed. Few data exist on the long-term outcome of postnatally acquired HCMV infection via breast milk. To substantiate the international debate on the use of native or inactivated milk for feeding of preterm infants, additional data are necessary for better identification of mother-infant-pairs at risk for viral transmission and symptomatic infection early after birth.

Postnatally acquired cytomegalovirus infection via breast milk: effects on hearing and development in preterm infants

Background. In preterm infants there is a high risk of transmission of cytomegalovirus (CMV) via breast milk from seropositive mothers with reactivation of the virus during lactation. There is little information about the long term sequel of early postnatally acquired CMV infection in pre-term infants. This study aimed to investigate whether there was an increased frequency of impaired neurodevelopmental outcome and sensorineural hearing loss in preterm infants with postnatally acquired CMV infection through transmission by CMV-positive breast milk. Methods. Twenty-two preterm infants [median birth weight, 1020 g (range, 600 to 1870 g); median gestational age, 27.6 weeks (range, 23.6 to 32 weeks] with early postnatally acquired CMV infection by breast-feeding (onset of viruria between Days 23 and 190 postnatally) were compared with 22 CMV-negative preterm infants individually matched for gestational age, birth weight, gender, intracranial hemorrhage and duration of ventilation. At 2 to 4.5 years of age, follow-up assessments were conducted consisting of neurologic examination, neurodevelopmental assessment and detailed audiologic tests. Results. None of the children had sensorineural hearing loss. There was no difference between the groups with regard to neurologic, speech and language or motor development. Conclusion. The results of this study suggest that early postnatally acquired CMV infection via CMV-positive breast milk does not have a negative effect on neurodevelopment and hearing in this group of patients. Because we studied a small number of infants, further follow-up studies are warranted in preterm infants with early postnatally acquired CMV infection.

Development of Cerebral Blood Flow Volume in Preterm Neonates during the First Two Weeks of Life

To investigate the postnatal development of cerebral perfusion in preterm neonates with normal brains over the first 2 wk of life, a prospective longitudinal study was designed. Quantitative measurement of cerebral blood flow (CBF) volume was performed using ultrasound flowmetry of the extracranial, brain-feeding arteries in 32 preterm infants of 28–35 wk gestational age. Measurements were done in the internal carotid and vertebral arteries of both sides on d 1, 2, 3, 7, and 14 after birth. A 10.0-MHz linear transducer of a computed sonography system (Acuson 128/XP10) was used. Intravascular flow volumes were calculated as the product of angle-corrected time-averaged flow velocity and the cross-sectional area of the vessel. Mean CBF volume increased markedly over the first 2 wk. One-third of this rise already occurred from the first to the second postnatal day, thereafter there was a continuous increase from d 2 to d 14 of life. Whereas the absolute level of CBF volume was primarily determined by postmenstrual age, the pattern of postnatal changes in CBF volume was found to be independent of gestational age. Arterial carbon dioxide tension, mean arterial blood pressure, and hematocrit had no influence on the development of CBF volume. The pronounced increase of CBF volume from d 1 to d 2 is likely to represent a normal adaptive response of the cerebral circulation to postnatal life. The data presented here may serve as the basis for further studies to investigate whether deviations from this adaptive response are associated with an increased risk of brain injury.

Topical timolol for small hemangiomas of infancy.

Abstract:  Propranolol has become the treatment of choice of large and complicated infantile hemangiomas. There is a controversy concerning the safety of systemic propranolol. Here we show that topical use of the beta‐blocker timolol can also inhibit the growth and promote regression of infantile hemangiomas. In this case series we treated 11 infantile hemangiomas in nine children including six preterm babies with the nonselective betablocker timolol. A timolol containing gel was manufactured from an ophthalmic formulation of timolol 0.5% eyedrops. This gel was applied using a standardized occlusive dressing (Finn‐Chambers) containing approximately 0.25 mg of timolol. In all infants topical timolol was associated with growth arrest, a reduction in redness and thickness within the first 2 weeks. Seven hemangiomas showed almost complete resolution, and four became much paler and thinner. No data are available on the transdermal absorption of timolol. Even supposing complete absorption of timolol from the occlusive dressing, a maximum dose of 0.25 mg of timolol would result per day and hemangioma. Regression of infantile hemangiomas treated using 0.5% timolol gel in this case series occurred earlier than spontaneous regression which is generally not observed before the age of 9–12 months. The promising results need to be verified in prospective randomized trials on topical beta blocker administration for infantile hemangiomas which should address dose, duration, and mode of application.

Long-term outcome in preterm children with human cytomegalovirus infection transmitted via breast milk.

Aim:  To investigate neurodevelopmental outcome and hearing in preterm children with breast milk transmitted human cytomegalovirus (HCMV) infection.

Rapid detection and quantification of cell free cytomegalovirus by a high-speed centrifugation-based microculture assay: comparison to longitudinally analyzed viral DNA load and pp67 late transcript during lactation.

BACKGROUND Human cytomegalovirus (HCMV) is reactivated in nearly every seropositive breastfeeding mother during lactation [Lancet 357 (2001) 513]. Conventional tissue culture (TC) and low-speed centrifugation-enhanced microtiter culture methods are not able to detect HCMV from milk during all stages of lactation. OBJECTIVES Development of a sensitive and quantitative microculture technique to describe the dynamics of HCMV reactivation in different milk compartments during lactation. STUDY DESIGN Milk samples were collected longitudinally from seropositive breastfeeding mothers of preterm infants. Native milk samples were separated into fraction 1 (aqueous extract of milk fat), fraction 2 (cell and fat free milk whey) and fraction 3 (milk cells). Each of these fractions was screened qualitatively (TC, nPCR, pp67 late mRNA) and quantitatively (high-speed centrifugation-based microculture, quantitative PCR). RESULTS Prior to low-speed centrifugation-enhanced inoculation, virus particles were concentrated by high-speed centrifugation (60 min at 50,000 x g, 4 degrees C). Using fraction 2 we were able to describe the dynamics of viral reactivation during lactation. We present the course of the quantitative virolactia and DNAlactia and qualitative detection of HCMV pp67 late mRNA in milk whey of four mothers (three transmitters and one non-transmitter). In all these cases virolactia described an unimodal and self limited course. Peak levels of virolactia for transmitters (T1: day 44; T2: day 43; T3: day 50) were closely related the onset of viruria of the corresponding preterm infants (U1: day 39; U2a/U2b: day 44/57; U3: day 60). The courses of viral load coincidence with the courses of DNA load. CONCLUSIONS We present a rapid and highly sensitive microculture method for the quantification of cell free HCMV from milk whey and aqueous extracts from milk fat. Viral reactivation during lactation describes an unimodal course. Our findings have strong implications for quality control of any virus inactivation procedure.

Measurement of volume of cerebral blood flow in healthy preterm and term neonates with ultrasound.

Changes in cerebral blood flow are important in the pathogenesis of ischaemic brain damage, but standard methods cannot measure volume of cerebral blood flow quantitatively in neonates. We used colour duplex sonography of the extracranial cerebral arteries to measure volume of global cerebral blood flow in 67 healthy preterm and term neonates. Volume of cerebral blood flow increased between the postmenstrual ages of 34 weeks (median 33 mL/min [range 23-43]) and 42 weeks (85 mL/min [57-104]). However, intersession and interobserver variability was quite large. This non-invasive method will allow quantitative bedside monitoring of global brain perfusion in preterm and term neonates with pathological disorders, and could also be used to monitor effects of neuroprotective measures.

Long-term cognitive and neurological outcome of preterm infants with postnatally acquired CMV infection through breast milk

Introduction Long-term follow-up data on preterm infants with breast milk–acquired postnatal cytomegalovirus (CMV) infection are sparse. Aim To systematically evaluate the long-term cognitive outcome and prevalence of cerebral palsy (CP) in patients after postnatal CMV infection. Patients and methods All surviving infants <1500 g born in our centre between 1 June 1995 and 1 June 2000, and with postnatal CMV infection acquired at up to 3 months of corrected age, were eligible for our study; this included neurological and neurocognitive assessment, using the Kaufman Assessment Battery for Children (K-ABC) at the age of >4 years. A blinded and controlled matched-pairs design was used with gestational age, gender and date of birth as matching criteria. Results Of 50 eligible children, 42 (84%) could be tested. There was no difference in the prevalence of cerebral palsy. Following CMV infection during their hospital stay, infants had significantly lower results in the simultaneous processing scale of the K-ABC (p=0.029) after correction for additional risk factors like socioeconomic status (SES). Results for the sequential and achievement scales were only slightly reduced (p>0.05). Conclusions It seems possible that breast milk-acquired CMV infection has a detrimental influence on cognitive development of preterm infants.

Evaluation of IL-8-Concentrations in Plasma and Lysed EDTA-Blood in Healthy Neonates and Those with Suspected Early Onset Bacterial Infection

Plasma IL-8 is a diagnostic parameter of early-onset bacterial infection (EOBI) in neonates but has a short half-life. The detergent-lysed whole-blood (DLWB) IL-8 consists of both extracellular and cell-bound IL-8. The objective of this study was to investigate kinetics of plasma and DLWB IL-8 in healthy newborns and those with suspected EOBI and to test the hypothesis that determination of DLWB IL-8 results in higher sensitivity for EOBI detection. Sixty-one neonates with clinical and serologic signs of EOBI composed the study group; 188 neonates with risk factors but without EOBI served as control subjects. IL-8 concentrations were determined in plasma and DLWB. In the control group, DLWB IL-8 concentrations were 280-fold higher (9599 pg/mL; SD 4433) up to 24 h post partum than corresponding plasma levels (34.2 pg/mL; SD 18.1). The sensitivity of DLWB versus plasma IL-8 for EOBI was 0.97 versus 0.71 after 6 h and 0.70 versus 0.32 after 24 h. Corresponding values for specificity were 0.95 versus 0.90 after 6 h and 0.92 versus 0.99 after 24 h. After 24 h, the negative predictive value for DLWB versus plasma IL-8 was 0.80 versus 0.66. DLWB IL-8 showed a higher sensitivity for EOBI within 6 h after first clinical suspicion than plasma IL-8. It also remained elevated longer. Our results suggest that DLWB IL-8 results in a higher sensitivity for EOBI.