Neonatal outcomes following introduction of routine probiotic supplementation to very preterm infants.

AIM: To evaluate the combined outcome of death and/or severe grade necrotising enterocolitis (NEC) in very preterm infants admitted to Cork University Maternity Hospital, Ireland, before and after introduction of routine supplementation with Bifidobacterium bifidum and Lactobacillus acidophilus probiotics (Infloran®). METHODS: A retrospective study of infants <32 weeks gestation and < 1500 g surviving beyond 72 h of life was performed. Two 6-year epochs; pre-probiotics (Epoch 1: 2008-2013) and with probiotics (Epoch 2: 2015-2020), were evaluated. The primary outcome was defined as death after 72 h or NEC Bell stage 2a or greater. RESULTS: Seven-hundred-and-forty-four infants were included (Epoch 1: 391, Epoch 2: 353). The primary outcome occurred in 67 infants (Epoch 1: 37, Epoch 2: 30, p = 0.646). After adjustment, the difference was significant (OR [95% CI]: 0.53 [0.29 to 0.97], p = 0.038). Differences between epochs did not depend on gestational age group (<28 weeks; ≥28 weeks). CONCLUSION: There was an associated reduction of the composite outcome of severe grade NEC and/or death, after adjustment for confounding variables, with introduction of routine administration of a B. bifidum and L. acidophilus probiotic at our institution.

Altered oral microbiome in Sudanese Toombak smokeless tobacco users carries a newly emerging risk of squamous cell carcinoma development and progression.

There are an estimated 6-10 million smokeless tobacco (Toombak) users in Sudan, the majority being males. Toombak is known to be a carcinogenic product that is likely to modify the oral microbiome spatiality into a high-risk potential for the development and progression of oral cancer, but previous studies are lacking in this field. Here, we endeavour for the first time the exploration of the oral microbiome in key mucosal areas of the oral cavity and assess the microbiome variations in premalignant and oral squamous cell carcinoma (OSCC) samples from both users and non-users of Toombak. 16S rRNA sequencing was performed on DNA obtained from pooled saliva, oral mucosa and supragingival plaque from 78 Sudanese users and non-users of Toombak, aged between 20 and 70 years. In 32 of the pooled saliva samples, the mycobiome (fungal) environment was analysed through ITS sequencing. Then, 46 formalin-fixed paraffin-embedded samples of premalignant and OSCC samples were collected, and their associated microbiomes sequenced. The oral Sudanese microbiome was found to be enriched in Streptococcaceae, but Staphylococcaceae were significantly more abundant amongst Toombak users. Genera enriched in the oral cavity of Toombak users included Corynebacterium_1 and Cardiobacterium while in non-users, Prevotella, Lactobacillus and Bifidobacterium were prominent. Aspergillus was the most abundant fungus in the mouths of Toombak users with a marked loss of Candida. The genus Corynebacterium_1 was abundant in the buccal, floor of the mouth and saliva microbiomes as well as in oral cancer samples from Toombak users indicating a possible role for this genus in the early stages of oral cancer development. An oral cancer microbiome that favours poor survival and metastasis in those who use Toombak also emerged that includes the genera Stenotrophomonas and Schlegelella. Those utilising Toombak carry an altered oral microbiome that may be an additional risk factor for this products carcinogenicity to the oral structures. These significant microbiome modulations are a newly emerging key driving factor in oral cancer development and progression in Toombak users while it is also shown that Toombak users carry an oral cancer microbiome that may increase the potential for a poorer prognosis.

Cross-sectional observational study protocol: missing microbes in infants born by caesarean section (MiMIC): antenatal antibiotics and mode of delivery.

INTRODUCTION: The intestinal microbiome in early life plays a major role in infant health and development. Factors like antibiotic exposure, breast/formula feeding and mode of delivery are known to affect the microbiome. The increasing occurrence of caesarean section (C-section) deliveries and antibiotic exposure warrants further insight into the potential missing microbes in those infants. The study objective is to study the effect of maternal antibiotic administration during pregnancy and/or C-section mode of delivery on the development of the infant's intestinal microbiome until the age of 2 years. METHODS AND ANALYSIS: A single site, cross-sectional observational study of C-section and vaginally delivered infants being either exposed to maternal antibiotic treatment or not during the third trimester of pregnancy. Throughout the nine visits, stool, urine, saliva, hair, breast milk and vaginal swabs will be collected from either mother and/or infant for microbiome and metabolomic analysis. ETHICS AND DISSEMINATION: The protocol was approved by the Clinical Research Ethics Committee of the Cork Teaching Hospitals. The trial has been registered at ClinicalTrials.gov.The findings from this study will be disseminated in peer-reviewed journals, during scientific conferences, and directly to the study participants. Sequencing data will be deposited in public databases. TRIAL REGISTRATION NUMBER: NCT04134819.

Dynamic Changes in the Human Milk Metabolome Over 25 Weeks of Lactation.

Human milk (HM) provides essential nutrition for ensuring optimal infant growth and development postpartum. Metabolomics offers insight into the dynamic composition of HM. Studies have reported the impact of lactation stage, maternal genotype, and gestational age on HM metabolome. However, the majority of the studies have considered changes within the first month of lactation or sampled with large intervals. This leaves a gap in the knowledge of progressing variation in HM composition beyond the first month of lactation. The objective of this study was to investigate whether the HM metabolome from mothers with term deliveries varies beyond 1 month of lactation, during the period in which HM is considered fully mature. Human milk samples (n = 101) from 59 mothers were collected at weeks 1-2, 3-5, 7-9, and 20-25 postpartum and analyzed using 1H nuclear magnetic resonance spectroscopy. Several metabolites varied over lactation and exhibited dynamic changes between multiple time points. Higher levels of HM oligosaccharides, cis-aconitate, O-phosphocholine, O-acetylcarnitine, gluconate, and citric acid were observed in early lactation, whereas later in lactation, levels of lactose, 3-fucosyllactose, glutamine, glutamate, and short- and medium-chain fatty acids were increased. Notably, we demonstrate that the HM metabolome is dynamic during the period of maturity.

The human milk microbiome aligns with lactation stage and not birth mode.

We analysed the human milk microbiome in a cohort of 80 lactating women and followed the dynamics in taxa over the course of lactation from birth to 6 months. Two hundred and thirty one milk samples were collected from full-term lactating women at 1, 4, 8 and 24 weeks following birth and analysed for microbiota composition using 16S rRNA sequencing. A significant decrease in milk microbiota diversity was observed throughout the first 6 months of lactation, with the greatest difference seen between week 8 and week 24. Nine genera predominated in milk over lactation from week 1 to week 24, comprising of Staphylococcus, Streptococcus, Pseudomonas, Acinetobacter, Bifidobacterium, Mesorhizobium, Brevundimonas, Flavobacterium, and Rhodococcus; however, fluctuations in these core genera were apparent over time. There was a significant effect of stage of lactation on the microbiome, while no effect of birth mode, infant sex and maternal BMI was observed throughout lactation. Streptococcus had the highest mean relative abundance at week 1 and 24 (17.3% and 24% respectively), whereas Pseudomonas predominated at week 4 (22%) and week 8 (19%). Bifidobacterium and Lactobacillus had the highest mean relative abundance at week 4 (5% and 1.4% respectively), and occurred at a relative abundance of ≤ 1% at all other time points. A decrease in milk microbiota diversity throughout lactation was also observed. This study concluded that lactation stage was the primary driving factor in milk microbiota compositional changes over lactation from birth to 6 months, while mode of delivery was not a factor driving compositional changes throughout human lactation.

First encounters of the microbial kind: perinatal factors direct infant gut microbiome establishment.

The human gut microbiome harbors a diverse range of microbes that play a fundamental role in the health and well-being of their host. The early-life microbiome has a major influence on human development and long-term health. Perinatal factors such as maternal nutrition, antibiotic use, gestational age and mode of delivery influence the initial colonization, development, and function of the neonatal gut microbiome. The perturbed early-life gut microbiome predisposes infants to diseases in early and later life. Understanding how perinatal factors guide and shape the composition of the early-life microbiome is essential to improving infant health. The following review provides a synopsis of perinatal factors with the most decisive influences on initial microbial colonization of the infant gut.

Vertical transfer of antibiotics and antibiotic resistant strains across the mother/baby axis.

Antibiotic resistance is a health and socioeconomic crisis recognized as a serious threat affecting humans worldwide. Overuse of antibiotics enhances the spread of multidrug-resistant bacteria, causing drug-resistant infections which can be difficult to treat. This resistance, mostly of the acquired type, is thus a major clinical issue. Acquired resistance can occur by horizontal transfer of genes between bacteria (community settings), by vertical transmission that can occur between mother and her offspring at birth and during lactation, or spontaneously due to antibiotic exposure. While there have been multiple studies about the horizontal transfer of antibiotic-resistance genes, not many studies have been conducted to study their vertical transmission. Vertical transmission is of importance as the early bacterial colonization of infants has an impact on their health and immune programming throughout life. This review discusses some possible mechanisms of mother-to-infant transmission of antibiotics and antibiotic-resistant strains and addresses the knowledge gaps for further studies.

Clinical implications of preterm infant gut microbiome development.

Perturbations to the infant gut microbiome during the first weeks to months of life affect growth, development and health. In particular, assembly of an altered intestinal microbiota during infant development results in an increased risk of immune and metabolic diseases that can persist into childhood and potentially into adulthood. Most research into gut microbiome development has focused on full-term babies, but health-related outcomes are also important for preterm babies. The systemic physiological immaturity of very preterm gestation babies (born earlier than 32 weeks gestation) results in numerous other microbiome-organ interactions, the mechanisms of which have yet to be fully elucidated or in some cases even considered. In this Perspective, we compare assembly of the intestinal microbiome in preterm and term infants. We focus in particular on the clinical implications of preterm infant gut microbiome composition and discuss the prospects for microbiome diagnostics and interventions to improve the health of preterm babies.

Probiotics, Prebiotics, and Synbiotics for the Prevention of Necrotizing Enterocolitis.

Necrotizing enterocolitis (NEC) is a major cause of morbidity and mortality in preterm infants. The exact mechanism by which NEC develops is poorly understood however there is growing evidence to suggest that perturbations in the early-life gut microbiota composition increase the risk for NEC. Modulation of the gut microbiota with probiotics, prebiotics, or in combination (synbiotics) is an area which has attracted intense interest in recent years. In this narrative review, we present an overview of the role of the gut microbiota in the pathogenesis of NEC. We also examine the evidence currently available from randomized controlled trials, observational studies, systematic reviews, and meta-analysis examining the role of probiotics, prebiotics, and synbiotics in reducing the risk of or preventing NEC. Current clinical practice guidelines with recommendations on the routine administration of probiotics to preterm infants for NEC are also explored.

The ultra-structural, metabolomic and metagenomic characterisation of the sudanese smokeless tobacco 'Toombak'.

Toombak is a smokeless tobacco produced from the Nicotiana rustica tobacco plant from Sudan. Pre-prepared and ready to buy Toombak samples were analysed using mass spectrometry (heavy metals), gas and liquid chromatography (metabolomics), 16S rRNA metagenomic sequencing (microbiome) and Phylogenetic Investigation of Communities by Reconstruction of Unobserved States (PICRUSt), scanning electron microscopy with energy dispersive X-ray spectroscopy (SEM-EDX) and pH analysis. Chromium, cobalt, and copper were high in the pre-prepared form of Toombak while iron, tobacco specific nitrosamines (TSNAs), formaldehyde and acetaldehyde were high in both types. Firmicutes and Actinobacteria dominated Toombak. Samples of ready to buy Toombak showed inter-variational differences depending on place of purchase. We found Virgibacillus were increased in the pre-prepared form while Corynebacterium casei, Atopococus tabaci, Atopostipes suicloacalis, Oceanobacillus chironomi and Staphylococcus gallinarum were the most abundant species in the ready to buy forms. PICRUSt analysis highlighted increased activity of metal transport systems in the ready to buy samples as well as an antibiotic transport system. SEM-EDX highlighted large non-homogenous, irregular particles with increased sodium, while pH of samples was in the alkaline range. The final composition of Toombak is affected by its method of preparation and the end product has the potential to impart many negative consequences on the health of its users. TSNA levels observed in Toombak were some of the highest in the world while the micro-environment of Toombak supports a distinct microbiota profile.

Metagenomic analysis of mother-infant gut microbiome reveals global distinct and shared microbial signatures.

Emerging evidence indicates maternal microbiota as one major reservoir for pioneering microbes in infants. However, the global distinct and identical features of mother-infant gut microbiota at various taxonomic resolutions and metabolic functions across cohorts and potential of infant microbial prediction based on their paired mother's gut microbiota remain unclear. Here, we analyzed 376 mother-infant dyads (468 mother and 1024 infant samples) of eight studies from six countries and observed higher diversity at species and strain levels in maternal gut microbiota but not their metabolic functions. A number of 290 species were shared in at least one mother-infant dyad, with 26 species (five at strain level) observed across cohorts. The profile of mother-infant shared species and strains was further influenced by delivery mode and feeding regimen. The mother-sourced species in infants exhibited similar strain heterogeneity but more metabolic functions compared to other-sourced species, suggesting the comparable stability and fitness of shared and non-shared species and the potential role of shared species in the early gut microbial community, respectively. Predictive models showed moderate performance accuracy for shared species and strains occurrences in infants. These generalized mother-infant shared species and strains may be considered as the primary targets for future work toward infant microbiome development and probiotics exploration.

Protein levels and protease activity in milk from mothers of pre-term infants: A prospective longitudinal study of human milk macronutrient composition.

BACKGROUND AND AIMS: The composition and enzymology of human milk changes throughout the lactation period, and differ for mothers who give birth prematurely compared to those who deliver at full-term. Understanding the composition of milk from mothers of very low birth weight premature infants is of great significance, and the objective of this study was to evaluate the composition, protein profile and plasmin activity of milk from mothers who delivered infants at different gestational ages. METHODS: Samples of human milk were donated by women (n = 74) in the Cork, Ireland, area who gave birth to full-term (>37 weeks gestation, FT), pre-term (32-37 weeks, PT) and very pre-term (≤32 weeks, VPT) infants. FT milk was collected at 1, 3, 6 and 10 weeks post-partum (PP), while PT and VPT milk was collected weekly until the FT due date of the infant and subsequently followed the FT protocol. RESULTS: Gestational age did not significantly affect lactose or fat content or total energy content of milk. However, protein content, and levels of some individual proteins, were significantly affected by both gestational age at birth and duration of lactation, with significantly higher protein levels in PT or VPT milk samples at 0-7 days and 1-2 months, respectively. Plasmin activity was significantly higher in VPT milk, indicating differences in proteolytic processing in milk. CONCLUSION: Compositional differences between the milk of mothers of term and pre-term infants were greatest in terms of the protein profile, which showed both qualitative and quantitative differences, as well as difference in proteolytic activity.

The Sporobiota of the Human Gut.

The human gut microbiome is a diverse and complex ecosystem that plays a critical role in health and disease. The composition of the gut microbiome has been well studied across all stages of life. In recent years, studies have investigated the production of endospores by specific members of the gut microbiome. An endospore is a tough, dormant structure formed by members of the Firmicutes phylum, which allows for greater resistance to otherwise inhospitable conditions. This innate resistance has consequences for human health and disease, as well as in biotechnology. In particular, the formation of endospores is strongly linked to antibiotic resistance and the spread of antibiotic resistance genes, also known as the resistome. The term sporobiota has been used to define the spore-forming cohort of a microbial community. In this review, we present an overview of the current knowledge of the sporobiota in the human gut. We discuss the development of the sporobiota in the infant gut and the perinatal factors that may have an effect on vertical transmission from mother to infant. Finally, we examine the sporobiota of critically important food sources for the developing infant, breast milk and powdered infant formula.

Effect of storage, temperature, and extraction kit on the phylogenetic composition detected in the human milk microbiota.

Human milk is considered the optimum feeding regime for newborns and is a source of bacteria for the developing infant gastrointestinal tract. However, as with all low biomass samples, standardization across variabilities such as sample collection, storage, and extraction methods is needed to eliminate discrepancies in microbial composition across studies. The aim of this study was to investigate how different storage methods, temperatures, preservatives, and extraction kits influence the human milk microbiome, compared to fresh samples. Breast milk samples were processed via six different methods: fresh (Method 1), frozen at -80°C (Method 2), treated with RNAlater and stored at 4°C or -80°C (Methods 3 and 4), and treated with Milk Preservation Solution at room temperature (Methods 5 and 6). Methods 1-5 were extracted using PowerFoodTM Microbial DNA Isolation kit (Mobio), and Method 6 was extracted using Milk DNA Preservation and Isolation kit (Norgen BioTek). At genus level, the most abundant genera were shared across Methods 1-5. Samples frozen at -80°C had fewest significant changes while samples treated and extracted using Milk Preservation and Isolation kit had the most significant changes when compared to fresh samples. Diversity analysis indicated that variation in microbiota composition was related to the method and extraction kit used. This study highlighted that, when extraction from fresh milk samples is not an option, freezing at -80°C is the next best option to preserve the integrity of the milk microbiome. Furthermore, our results demonstrate that choice of extraction kit had a profound impact on the microbiota populations detected in milk.

The Role of the Microbiome in Oral Squamous Cell Carcinoma with Insight into the Microbiome-Treatment Axis.

Oral squamous cell carcinoma (OSCC) is one of the leading presentations of head and neck cancer (HNC). The first part of this review will describe the highlights of the oral microbiome in health and normal development while demonstrating how both the oral and gut microbiome can map OSCC development, progression, treatment and the potential side effects associated with its management. We then scope the dynamics of the various microorganisms of the oral cavity, including bacteria, mycoplasma, fungi, archaea and viruses, and describe the characteristic roles they may play in OSCC development. We also highlight how the human immunodeficiency viruses (HIV) may impinge on the host microbiome and increase the burden of oral premalignant lesions and OSCC in patients with HIV. Finally, we summarise current insights into the microbiome-treatment axis pertaining to OSCC, and show how the microbiome is affected by radiotherapy, chemotherapy, immunotherapy and also how these therapies are affected by the state of the microbiome, potentially determining the success or failure of some of these treatments.

A good start in life is important-perinatal factors dictate early microbiota development and longer term maturation.

Maternal health status is vital for the development of the offspring of humans, including physiological health and psychological functions. The complex and diverse microbial ecosystem residing within humans contributes critically to these intergenerational impacts. Perinatal factors, including maternal nutrition, antibiotic use and maternal stress, alter the maternal gut microbiota during pregnancy, which can be transmitted to the offspring. In addition, gestational age at birth and mode of delivery are indicated frequently to modulate the acquisition and development of gut microbiota in early life. The early-life gut microbiota engages in a range of host biological processes, particularly immunity, cognitive neurodevelopment and metabolism. The perturbed early-life gut microbiota increases the risk for disease in early and later life, highlighting the importance of understanding relationships of perinatal factors with early-life microbial composition and functions. In this review, we present an overview of the crucial perinatal factors and summarise updated knowledge of early-life microbiota, as well as how the perinatal factors shape gut microbiota in short and long terms. We further discuss the clinical consequences of perturbations of early-life gut microbiota and potential therapeutic interventions with probiotics/live biotherapeutics.

Breast Milk, a Source of Beneficial Microbes and Associated Benefits for Infant Health.

Human breast milk is considered the optimum feeding regime for newborn infants due to its ability to provide complete nutrition and many bioactive health factors. Breast feeding is associated with improved infant health and immune development, less incidences of gastrointestinal disease and lower mortality rates than formula fed infants. As well as providing fundamental nutrients to the growing infant, breast milk is a source of commensal bacteria which further enhance infant health by preventing pathogen adhesion and promoting gut colonisation of beneficial microbes. While breast milk was initially considered a sterile fluid and microbes isolated were considered contaminants, it is now widely accepted that breast milk is home to its own unique microbiome. The origins of bacteria in breast milk have been subject to much debate, however, the possibility of an entero-mammary pathway allowing for transfer of microbes from maternal gut to the mammary gland is one potential pathway. Human milk derived strains can be regarded as potential probiotics; therefore, many studies have focused on isolating strains from milk for subsequent use in infant health and nutrition markets. This review aims to discuss mammary gland development in preparation for lactation as well as explore the microbial composition and origins of the human milk microbiota with a focus on probiotic development.

Maternal Vertical Transmission Affecting Early-life Microbiota Development.

The association of the human microbiome with health outcomes has attracted much interest toward its therapeutic manipulation. The likelihood of modulating the human microbiome in early life is high and offers great potential to exert profound effects on human development since the early microbiota shows more flexibility compared to that of adults. The human microbiota, being similar to human genetics, can be transmitted from mother to infant, providing insights into early microbiota acquisition, subsequent development, and potential opportunities for intervention. Here, we review adaptations of the maternal microbiota during pregnancy, birth, and infancy, the acquisition and succession of early-life microbiota, and highlight recent efforts to elucidate mother-to-infant microbiota transmission. We further discuss how the mother-to-infant microbial transmission is shaped; and finally we address potential directions for future studies to promote our understanding within this field.

Mining Google Trends Data for Health Information: The Case of the Irish "CervicalCheck" Screening Programme Revelations.

Background In April 2018, the Irish cervical smear screening programme, "CervicalCheck", came under intense scrutiny as the accuracy of hundreds of "negative" results were brought in question. Aim The goal of this brief report was to assess the impact of this real-life event on public information-seeking behaviour, using Google search anomalies as a proxy. Irish relative search volume data for several terms relating to cervical testing/cancer and human papillomavirus were extracted for a five-year period from February 2014 to January 2019 and analysed for the presence of anomalous spikes and shifts in the mean baseline. Results An unprecedented positive spike in searches relating to cervical testing/cancer was observed immediately after the CervicalCheck revelations, which remained anomalous for the month to follow (p < 0.05). This public interest preceded a mirroring increase in uptake of complimentary consultations offered by the Department of Health to the women concerned. Despite this service engagement and interest in cervical health, the relative search volumes for terms "human papillomavirus infection" and "HPV vaccine" were just 78 and 51% of their maximum search volume for the five-year period. Conclusions Anomaly analysis revealed an unprecedented spike in information-seeking behaviour following the CervicalCheck revelations. However, this was not associated with a comparable elevation in HPV interest. This suggests that more public education and promotion of the HPV vaccine is warranted, in the context of vastly reduced uptake in recent years. Finally, Google Trends data represents a free an open source means by which to assess information-seeking behaviour of the public in relation to health and disease.

Paediatrician's perspective of infant gut microbiome research: current status and challenges.

Due to its innately intriguing nature and recent genomic technological advances, gut microbiome research has been at the epicentre of medical research for over a decade now. Despite the degree of publicisation, a comprehensive understanding and, therefore, acceptance of the area as a whole may be somewhat lacking within the broader medical community. This paper summarises the main analytical techniques and tools currently applied to compositional microbiome research. In addition, we outline five major lessons learnt from a decade of infant microbiome research, along with the current research gaps. Finally, we aim to provide an introduction and general guidelines relating to infant gut microbiome research for the practising paediatrician.