María Cernada

Microbial ecology and host-microbiota interactions during early life stages

The role of human microbiota has been redefined during recent years and its physiological role is now much more important than earlier understood. Intestinal microbial colonization is essential for the maturation of immune system and for the developmental regulation of the intestinal physiology. Alterations in this process of colonization have been shown to predispose and increase the risk to disease later in life. The first contact of neonates with microbes is provided by the maternal microbiota. Moreover, mode of delivery, type of infant feeding and other perinatal factors can influence the establishment of the infant microbiota. Taken into consideration all the available information it could be concluded that the exposure to the adequate microbes early in gestation and neonatal period seems to have a relevant role in health. Maternal microbial environment affects maternal and fetal immune physiology and, of relevance, this interaction with microbes at the fetal-maternal interface could be modulated by specific microbes administered to the pregnant mother. Indeed, probiotic interventions aiming to reduce the risk of immune-mediated diseases may appear effective during early life.

Factors influencing gastrointestinal tract and microbiota immune interaction in preterm infants

The role of microbial colonization is indispensable for keeping a balanced immune response in life. However, the events that regulate the establishment of the microbiota, their timing, and the way in which they interact with the host are not yet fully understood. Factors such as gestational age, mode of delivery, environment, hygienic measures, and diet influence the establishment of microbiota in the perinatal period. Environmental microbes constitute the most important group of exogenous stimuli in this critical time frame. However, the settlement of a stable gut microbiota in preterm infants is delayed compared to term infants. Preterm infants have an immature gastrointestinal tract and immune system which predisposes to infectious morbidity. Neonatal microbial dynamics and alterations in early gut microbiota may precede and/or predispose to diseases such as necrotizing enterocolitis (NEC), late-onset sepsis or others. During this critical period, nutrition is the principal contributor for immunological and metabolic development, and microbiological programming. Breast milk is a known source of molecules that act synergistically to protect the gut barrier and enhance the maturation of the gut-related immune response. Host-microbe interactions in preterm infants and the protective role of diet focused on breast milk impact are beginning to be unveiled.

Oxygen saturation after birth in preterm infants treated with continuous positive airway pressure and air: assessment of gender differences and comparison with a published nomogram

Aims The goal of the study was to compare preductal SpO2 in the first 10 min after birth in preterm infants treated with non-invasive continuous positive airway pressure (CPAP) and air with a published nomogram of preductal SpO2 in preterm infants who received no medical intervention, and to examine gender differences. Design Prospective observational study. Patients and methods We enrolled infants of ≤32 weeks gestation who were spontaneously breathing with heart rate >100 bpm, and treated with face mask CPAP and air during postnatal stabilisation. SpO2 limits were targeted at ≥75% at 5 min and ≥85% at 10 min and heart rate at >100 bpm. FIO2 was titrated against SpO2. Preductal SpO2, airway pressure and FIO2 were recorded with a data acquisition system from birth until stabilisation. Babies receiving supplemental oxygen (>21%), positive pressure ventilation, were intubated and/or received chest compressions or drugs were excluded. Results Measurements were obtained in 102 babies with median gestational age of 29 (range: 24–31) weeks. Median SpO2 was significantly higher in the observational group than in the reference range at 3 min (82% (CI 71% to 85%) vs 76% (CI 67% to 83%); p<0.05), at 4 min (87% (CI 81% to 90%) vs 81% (CI 72% to 88%); p<0.05), at 5 min (92% (CI 88% to 95%) vs 86% (CI 80% to 92%); p<0.05), at 6 min (94% (CI 90% to 97%) vs 90% (CI 81% to 95%); p<0.05), at 7 min (95% (CI 92% to 97%) vs 92% (CI 85% to 95%); p<0.05), at 8 min (96% (CI 93% to 98%) vs 92% (CI 87% to 96%); p<0.05) and at 9 min (97% (CI 92% to 99%) vs 93% (CI 87% to 96%); p<0.05). Female babies achieved targeted SpO2 significantly earlier than male babies. Conclusions Preterm babies receiving CPAP and air and especially female subjects achieve reference oxygen saturation more rapidly than spontaneously breathing preterm babies without respiratory aid.

The Use and Misuse of Oxygen During the Neonatal Period

This article describes aerobic metabolism, oxygen free radicals, antioxidant defenses, oxidative stress, inflammatory response and redox signaling, the fetal to neonatal transition, arterial oxygen saturation, oxygen administration in the delivery room, oxygen during neonatal care in the NICU, evolving oxygen needs in the first few weeks of life, and complications that can occur when infants go home from the hospital on oxygen.

Minimally invasive surfactant therapy with a gastric tube is as effective as the intubation, surfactant, and extubation technique in preterm babies

Preterm infants requiring surfactant replacement have been treated using the INSURE technique, which requires sedation and comprises tracheal intubation, surfactant instillation and extubation. However, minimally invasive surfactant therapy (MIST) does not require sedation, minimises airway injury and avoids placing positive pressure ventilation on an immature lung. This study compared the feasibility of the two techniques and the outcomes in preterm babies with respiratory distress syndrome (RDS).

Cord blood interleukin-6 as a predictor of early-onset neonatal sepsis.

Aim:  To compare diagnostic accuracy in cord blood of interleukin‐6 (IL‐6) with C‐reactive protein (CRP) as predictors of early‐onset neonatal sepsis (EOS) in newborns with prenatal risk factors for infection.

Ventilator-associated pneumonia in newborn infants diagnosed with an invasive bronchoalveolar lavage technique: a prospective observational study.

Objectives: To establish the incidence, etiology, risk factors, and outcomes associated with ventilator-associated pneumonia using an invasive sampling technique to avoid contamination. Patients: Eligible patients were intubated neonates treated with mechanical ventilation who followed the criteria of the Centers for Disease Control and Prevention/National Nosocomial Infection Surveillance. Bronchoalveolar lavage samples were collected using a blind-protected catheter to avoid contamination of upper respiratory microorganisms. Isolation of >103 colony-forming unit/mL was required for diagnosis. Measurements and Main Results: In 198 neonates intubated for >48 hrs, a total of 18 episodes of ventilator-associated pneumonia in 16 infants representing a prevalence of 8.1 were diagnosed. The pooled mean ventilator-associated pneumonia rate was 10.9/1,000 ventilator days. The mean age at diagnosis of ventilator-associated pneumonia was 29±15 days after a mean of 21±16 days of mechanical ventilation. Gram-negative bacteria were the most commonly isolated pathogens and Pseudomonas aeruginosa was the most frequent causative agent. Hospital length of stay was significantly longer for ventilator-associated pneumonia patients; however, no significant differences in mortality were found. Univariate analysis comparing patients with and without ventilator-associated pneumonia showed that days of mechanical ventilation, days of oxygen, number of reintubations, number of transfusions, bloodstream infection, and enteral feeding were all significantly associated with ventilator-associated pneumonia. However, in multivariate analysis the unique independent risk factor was days of mechanical ventilation (odds ratio 1.12, confidence interval 95% 1.07–1.17). Conclusions: Ventilator-associated pneumonia is a frequent nosocomial infection in newborns. Only duration of mechanical ventilation has been identified as an independent risk factor for ventilator-associated pneumonia. The use of a blind invasive sampling technique seems to diminish sample contamination.

Genome-Wide Expression Profiles in Very Low Birth Weight Infants With Neonatal Sepsis

BACKGROUND: Bacterial sepsis is associated with high morbidity and mortality in preterm infants. However, diagnosis of sepsis and identification of the causative agent remains challenging. Our aim was to determine genome-wide expression profiles of very low birth weight (VLBW) infants with and without bacterial sepsis and assess differences. METHODS: This was a prospective observational double-cohort study conducted in VLBW (<1500 g) infants with culture-positive bacterial sepsis and non-septic matched controls. Blood samples were collected as soon as clinical signs of sepsis were identified and before antibiotics were initiated. Total RNA was processed for genome-wide expression analysis using Affymetrix gene arrays. RESULTS: During a 19-month period, 17 septic VLBW infants and 19 matched controls were enrolled. First, a three-dimensional unsupervised principal component analysis based on the entire genome (28 000 transcripts) identified 3 clusters of patients based on gene expression patterns: Gram-positive sepsis, Gram-negative sepsis, and noninfected control infants. Furthermore, these groups were confirmed by using analysis of variance, which identified a transcriptional signature of 554 of genes. These genes had a significantly different expression among the groups. Of the 554 identified genes, 66 belonged to the tumor necrosis factor and 56 to cytokine signaling. The most significantly overexpressed pathways in septic neonates related with innate immune and inflammatory responses and were validated by real-time reverse transcription polymerase chain reaction. CONCLUSIONS: Our preliminary results suggest that genome-wide expression profiles discriminate septic from nonseptic VLBW infants early in the neonatal period. Further studies are needed to confirm these findings.

Ultra high performance liquid chromatography coupled to tandem mass spectrometry determination of lipid peroxidation biomarkers in newborn serum samples

Byproducts of arachidonic (AA) and docosahexaenoic acid (DHA) oxidation are highly relevant for the study of free radical associated conditions in the perinatal period. Plasma metabolites can provide the clinician with a snapshot of the oxidant status of patients before and after specific clinical interventions (e.g.: supplementation with oxygen). We describe a new andreliable ultra-performance liquid mass spectrometry method to determine F2-isoprostanes and other byproducts (isoprostanes, isofurans, neuroprostanes, neurofurans) in newborn serum samples. Cord blood samples were obtained from severely depressed newborn infants (Apgar score 1 min < 3; arterial cord pH < 7.00), and aliquoted for serum determination and stored at -80 °C. A UHPLC-MS/MS method was employed. It has a series of technical advantages: simple sample treatment; reduced sample volume (100 μL) which is essential for preterm neonates with low circulating blood volume, high throughput of sample analysis (96 samples in less than 24 h) and high selectivity for different isoprostanes isomers. Excellent sensitivity was achieved within limits of detection between 0.06 and 4.2 nmol L(-1), which renders this method suitable to monitoranalyte concentration in newborn samples. The methods precision was satisfactory; with coefficients of variation around 5-12% (intra-day) and 7-17% (inter-day). The reliability of the described method was assessed by analysis of spiked serum samples obtaining recoveries between 70% and 120%. The proposed method has rendered suitable for serum determination for newborn babies at risk of oxygen free radical associated conditions.

Sepsis in preterm infants causes alterations in mucosal gene expression and microbiota profiles compared to non-septic twins

Sepsis is a life-threatening condition in preterm infants. Neonatal microbiota plays a pivotal role in the immune system maturation. Changes in gut microbiota have been associated to inflammatory disorders; however, a link with sepsis in the neonatal period has not yet been established. We aimed to analyze gut microbiota and mucosal gene expression using non-invasively obtained samples to provide with an integrative perspective of host-microbe interactions in neonatal sepsis. For this purpose, a prospective observational case-control study was conducted in septic preterm dizygotic twins and their non-septic twin controls. Fecal samples were used for both microbiota analysis and host genome-wide expression using exfoliated intestinal cells. Gene expression of exfoliated intestinal cells in septic preterm showed an induction of inflammatory and oxidative stress pathways in the gut and pro-oxidant profile that caused dysbiosis in the gut microbiota with predominance of Enterobacteria and reduction of Bacteroides and Bifidobacterium spp.in fecal samples, leading to a global reduction of beneficial anaerobic bacteria. Sepsis in preterm infants induced low-grade inflammation and oxidative stress in the gut mucosa, and also changes in the gut microbiota. This study highlights the role of inflammation and oxidative stress in neonatal sepsis on gut microbial profiles.