Whole-genome sequencing in the investigation of recurrent invasive group A streptococcus outbreaks in a maternity unit.

BACKGROUND: The clinical manifestations of group A streptococcus (GAS) (Streptococcus pyogenes) are diverse, ranging from asymptomatic colonization to devastating invasive disease. Maternity-related clusters of invasive GAS (iGAS) infection are complex to investigate and control, especially if recurrent. AIM: To investigate three episodes of emm 75 GAS/iGAS infection in maternity patients at one hospital site over a four-year period (two with monophyletic ancestry). METHODS: The episodes are described, together with whole-genome sequence (WGS) isolate analyses. Single nucleotide polymorphism differences were compared with contemporaneous emm 75 genomes. FINDINGS: Over the four-year study period, seven mothers had emm 75 GAS/iGAS and one mother had emm 3 iGAS (in year 4) (subsequently discounted as linked). Three (clinical/screening samples) of the seven babies of emm-75-positive mothers and three screened healthcare workers were positive for emm 75 GAS. WGS similarity suggested a shared ancestral lineage and a common source transmission, but directionality of transmission cannot be inferred. However, the findings indicate that persistence of a particular clone in a given setting may be long term. CONCLUSIONS: Occupational health procedures were enhanced, staff were screened, and antibiotic therapy was provided to GAS-positive staff and patients. The definitive source of infection could not be identified, although staff-patient transmission was the most likely route. The pattern of clonal GAS transmission over the four-year study period suggests that long-term persistence of GAS may have occurred.

Description of a method for inducing fetal growth restriction in the spiny mouse.

Intrauterine or fetal growth restriction (IUGR) is a major complication of pregnancy and leads to significant perinatal morbidities and mortality. Typically, induction of IUGR in animals involves the complete occlusion or ablation of vessels to the uterus or placenta, acutely impairing blood flow and fetal growth, usually with high fetal loss. We aimed to produce a model of reduced fetal growth in the spiny mouse with minimal fetal loss. At 27 days gestational age (term is 38-39 days), a piece of silastic tubing was placed around the left uterine artery to prevent the further increase of uterine blood flow with advancing gestation to induce IUGR (occluded). Controls were generated from sham surgeries without placement of the tubing. Dams were humanely euthanized at 37 days gestational age and all fetuses and placentas were weighed and collected. Of the 17 dams that underwent surgery, 15 carried their pregnancies to 37 days gestational age and 95% of fetuses survived to this time. The difference in fetal body weight between occluded and control was ~21% for fetuses in the left uterus side: there were no differences for fetuses in the right uterus side. Offspring from the occluded group had significantly lower brain, liver, lung, kidney and carcass weights compared with shams. Preventing the gestation-related increase of uterine blood flow induced significant growth restriction in the fetal spiny mouse, with minimal fetal loss. This technique could be readily adapted for other small animal.

Review: Sexual dimorphism in the formation, function and adaptation of the placenta.

Exposure of the embryo or fetus to perturbations in utero can result in intrauterine growth restriction, a primary risk factor for the development of adult disease. However, despite similar exposures, males and females often have altered disease susceptibility or progression from different stages of life. Fetal growth is largely mediated by the placenta, which, like the fetus is genetically XX or XY. The placenta and its associated trophoblast lineages originate from the trophectoderm (TE) of the early embryo. Rodent models (rat, mouse, spiny mouse), have been used extensively to examine placenta development and these have demonstrated the growth trajectory of the placenta in females is generally slower compared to males, and also shows altered adaptive responses to stressful environments. These placental adaptations are likely to depend on the type of stressor, duration, severity and the window of exposure during development. Here we describe the divergent developmental pathways between the male and female placenta contributing to altered differentiation of the TE derived trophoblast subtypes, placental growth, and formation of the placental architecture. Our focus is primarily genetic or environmental perturbations in rodent models which show altered placental responsiveness between sexes. We suggest that perturbations during early placental development may have greater impact on viability and growth of the female fetus whilst those occurring later in gestation may preferentially affect the male fetus. This may be of great relevance to human pregnancies which result from assisted reproductive technologies or complications such as pre-eclampsia and diabetes.

A review of fundamental principles for animal models of DOHaD research: an Australian perspective.

Epidemiology formed the basis of 'the Barker hypothesis', the concept of 'developmental programming' and today's discipline of the Developmental Origins of Health and Disease (DOHaD). Animal experimentation provided proof of the underlying concepts, and continues to generate knowledge of underlying mechanisms. Interventions in humans, based on DOHaD principles, will be informed by experiments in animals. As knowledge in this discipline has accumulated, from studies of humans and other animals, the complexity of interactions between genome, environment and epigenetics, has been revealed. The vast nature of programming stimuli and breadth of effects is becoming known. As a result of our accumulating knowledge we now appreciate the impact of many variables that contribute to programmed outcomes. To guide further animal research in this field, the Australia and New Zealand DOHaD society (ANZ DOHaD) Animals Models of DOHaD Research Working Group convened at the 2nd Annual ANZ DOHaD Congress in Melbourne, Australia in April 2015. This review summarizes the contributions of animal research to the understanding of DOHaD, and makes recommendations for the design and conduct of animal experiments to maximize relevance, reproducibility and translation of knowledge into improving health and well-being.

Maternal creatine in pregnancy: a retrospective cohort study.

OBJECTIVE: To estimate creatine concentrations in maternal plasma and urine, and establish relationships with maternal characteristics, diet and fetal growth. DESIGN: Retrospective cohort study. SETTING: Lyell McEwin Hospital, Adelaide, Australia. POPULATION: A biobank of plasma and urine samples collected at 13, 18, 30 and 36 weeks' gestation from 287 pregnant women from a prospective cohort of asthmatic and non-asthmatic women. METHODS: Creatine was measured by enzymatic analysis. Change in creatine over pregnancy was assessed using the Friedman test. Linear mixed models regression was used to determine associations between maternal factors and diet with creatine across pregnancy and between creatine with indices of fetal growth at birth. MAIN OUTCOME MEASURES: Maternal creatine concentrations, associations between maternal factors and creatine and between creatine and fetal growth parameters. RESULTS: Maternal smoking, body mass index, asthma and socio-economic status were positively and parity negatively associated with maternal plasma and/or urine creatine. Maternal urine creatine concentration was positively associated with birthweight centile and birth length. After adjustment, each µmol/l increase in maternal urinary creatine was associated with a 1.23 (95% CI 0.44-2.02) unit increase in birthweight centile and a 0.11-cm (95% CI 0.03-0.2) increase in birth length. CONCLUSIONS: Maternal factors and fetal growth measures are associated with maternal plasma and urine creatine concentrations. TWEETABLE ABSTRACT: Maternal creatine is altered by pregnancy; fetal growth measures are associated with maternal creatine concentrations.

The establishment of DOHaD working groups in Australia and New Zealand.

The evidence underpinning the developmental origins of health and disease (DOHaD) is overwhelming. As the emphasis shifts more towards interventions and the translational strategies for disease prevention, it is important to capitalize on collaboration and knowledge sharing to maximize opportunities for discovery and replication. DOHaD meetings are facilitating this interaction. However, strategies to perpetuate focussed discussions and collaborations around and between conferences are more likely to facilitate the development of DOHaD research. For this reason, the DOHaD Society of Australia and New Zealand (DOHaD ANZ) has initiated themed Working Groups, which convened at the 2014-2015 conferences. This report introduces the DOHaD ANZ Working Groups and summarizes their plans and activities. One of the first Working Groups to form was the ActEarly birth cohort group, which is moving towards more translational goals. Reflecting growing emphasis on the impact of early life biodiversity - even before birth - we also have a Working Group titled Infection, inflammation and the microbiome. We have several Working Groups exploring other major non-cancerous disease outcomes over the lifespan, including Brain, behaviour and development and Obesity, cardiovascular and metabolic health. The Epigenetics and Animal Models Working Groups cut across all these areas and seeks to ensure interaction between researchers. Finally, we have a group focussed on 'Translation, policy and communication' which focusses on how we can best take the evidence we produce into the community to effect change. By coordinating and perpetuating DOHaD discussions in this way we aim to enhance DOHaD research in our region.

IFPA meeting 2015 workshop report II: mechanistic role of the placenta in fetal programming; biomarkers of placental function and complications of pregnancy.

Workshops are an integral component of the annual International Federation of Placenta Association (IFPA) meeting, allowing for networking and focused discussion related to specialized topics on the placenta. At the 2015 IFPA meeting (Brisbane, Australia) twelve themed workshops were held, three of which are summarized in this report. These workshops focused on various aspects of placental function, particularly in cases of placenta-mediated disease. Collectively, these inter-connected workshops highlighted the role of the placenta in fetal programming, the use of various biomarkers to monitor placental function across pregnancy, and the clinical impact of novel diagnostic and surveillance modalities in instances of late onset fetal growth restriction (FGR).

Evaluating a systematic voiding programme for patients with urinary incontinence after stroke in secondary care using soft systems analysis and Normalisation Process Theory: findings from the ICONS case study phase.

BACKGROUND: Urinary incontinence (UI) affects between 40 and 60% of people in hospital after stroke, but is often poorly managed in stroke units. OBJECTIVES: To inform an exploratory trial by three methods: identifying the organisational context for embedding the SVP; exploring health professionals' views around embedding the SVP and measuring presence/absence of UI and frequency of UI episodes at baseline and six weeks post-stroke. DESIGN: A mixed methods single case study included analysis of organisational context using interviews with clinical leaders analysed with soft systems methodology, a process evaluation using interviews with staff delivering the intervention and analysed with Normalisation Process Theory, and outcome evaluation using data from patients receiving the SVP and analysed using descriptive statistics. SETTING: An 18 bed acute stroke unit in a large Foundation Trust (a 'not for profit' privately controlled entity not accountable to the UK Department of Health) serving a population of 370,000. PARTICIPANTS: Health professionals and clinical leaders with a role in either delivering the SVP or linking with it in any capacity were recruited following informed consent. Patients were recruited meeting the following inclusion criteria: aged 18 or over with a diagnosis of stroke; urinary incontinence (UI) as defined by the International Continence Society; conscious; medically stable as judged by the clinical team and with incontinence classified as stress, urge, mixed or 'functional'. All patients admitted to the unit during the intervention period were screened for eligibility; informed consent to collect baseline and outcome data was sought from all eligible patients. RESULTS: Organisational context: 18 health professionals took part in four group interviews. Findings suggest an environment not conducive to therapeutic continence management and a focus on containment of UI. Embedding the SVP into practice: 21 nursing staff took part in six group interviews. Initial confusion gave way to embedding of processes facilitated by new routines and procedures. Patient outcome: 43 patients were recruited; 28 of these commenced the SVP. Of these, 6/28 (21%) were continent at six weeks post-stroke or discharge. CONCLUSION: It was possible to embed the SVP into practice despite an organisational context not conducive to therapeutic continence care. Recommendations are made for introducing the SVP in a trial context.

Treatment of pregnant spiny mice at mid gestation with a synthetic glucocorticoid has sex-dependent effects on placental glycogen stores.

INTRODUCTION: Elevated maternal glucocorticoids during human pregnancy suppress fetal growth, more so if the fetus is male. The synthetic glucocorticoid dexamethasone (DEX) is known to affect placental glucose transport, but whether this also affects placental glycogen stores has not been investigated. METHOD: We examined the short and long term consequences of a single, 60 h exposure to DEX at mid gestation on the glycogen pathway in the placenta of the spiny mouse, with a focus on identifying sex-dependent differences in expression of genes involved in glycogen cell formation (PCDH12), and regulation of glycogen synthesis (GSK3B, GYS1, GBE1, FOXO1, UGP2). RESULTS: Placentas from female fetuses had increased amounts of glycogen on day 25 of gestation (term is 39 days) as identified by positive Periodic acid Schiff (PAS) reaction staining. DEX administration initially reduced expression of GSK3B, GYS1, GBE1, FOXO1, UGP2 in both male and female placentas, but reduced histologically detectable glycogen storage in placentas of female fetuses only. The DEX-induced reduction in expression of GSK3B and UGP2 persisted until day 37 of gestation, an effect that was significantly greater in the male placenta. DISCUSSION/CONCLUSION: We conclude that constitutive placental glycogen storage is regulated in pregnancy in a sex-dependant manner, and that glucocorticoids such as DEX induce sex-dependent changes in glycogen storage. Placental glycogen metabolism and its response to glucocorticoids may contribute to the different sensitivities of male and female fetuses to the effects of maternal illness and stress in utero.

Search for photon-linelike signatures from dark matter annihilations with H.E.S.S.

Gamma-ray line signatures can be expected in the very-high-energy (E(γ)>100 GeV) domain due to self-annihilation or decay of dark matter (DM) particles in space. Such a signal would be readily distinguishable from astrophysical γ-ray sources that in most cases produce continuous spectra that span over several orders of magnitude in energy. Using data collected with the H.E.S.S. γ-ray instrument, upper limits on linelike emission are obtained in the energy range between ∼ 500 GeV and ∼ 25 TeV for the central part of the Milky Way halo and for extragalactic observations, complementing recent limits obtained with the Fermi-LAT instrument at lower energies. No statistically significant signal could be found. For monochromatic γ-ray line emission, flux limits of (2 × 10(-7) -2 × 10(-5)) m(-2) s(-1) sr(-1) and (1 × 10(-8) -2 × 10(-6)) m(-2) s(-1)sr(-1) are obtained for the central part of the Milky Way halo and extragalactic observations, respectively. For a DM particle mass of 1 TeV, limits on the velocity-averaged DM annihilation cross section ⟨σv⟩(χχ → γγ) reach ∼ 10(-27) cm(3)s(-1), based on the Einasto parametrization of the Galactic DM halo density profile.

A novel method for efficient in vitro germination and tube growth of Arabidopsis thaliana pollen.

In addition to its importance in studies of plant reproduction and fertility, pollen is as widely employed as a model system of cell growth and development. This work demands robust, reproducible methods to induce pollen germination and morphologically normal growth of pollen tubes in vitro. Despite numerous advantages of Arabidopsis thaliana as a model plant, such experiments on pollen germination and pollen tube growth have often proved challenging. Our new method employs a physical cellulosic membrane, overlying an agarose substrate. By modulating the substrate composition, we provide important insights into the mechanisms promoting pollen growth both in vitro and in vivo. This effective new technical approach to A. thaliana pollen germination and tube growth results in swift, consistent and unprecedented levels of germination to over 90%. It can also promote rapid growth of long, morphologically normal pollen tubes. This technical development demonstrates that exogenous spermidine and a cellulosic substrate are key factors in stimulating germination. It has potential to greatly assist the study of reproduction in A. thaliana and its closest relatives, not only for the study of germination levels and pollen tube growth dynamics by microscopy, but also for biochemical and molecular analysis of germinating pollen.

Sexually dimorphic placental development throughout gestation in the spiny mouse (Acomys cahirinus).

OBJECTIVES: It has been hypothesized that male fetuses down regulate placental growth during periods of accelerated fetal growth. We aimed to investigate this, and determine whether sexual dimorphism was apparent in the spiny mouse placenta. We hypothesized that expression of fetal growth promoters would be higher in placentas of males, whereas genes involved in placental structural development would be more highly expressed in placentas of females. METHODS: Spiny mouse dams, a precocial rodent with an in utero endocrine milieu dissimilar from other rodents, but akin to humans, were sacrificed at gestational ages 15-37 (term = 39 days). Placentas were collected and processed for histology or qPCR analysis of selected genes (GCM1, MAP2K1, SLC2A1, NR3C1, IGF1, IGF1R). RESULTS: Fetal and placental weights were similar for both sexes. Placentas of female fetuses had less spongy zone (P(SEX) < 0.0001), and more labyrinth (P(SEX) < 0.0001) than males. Early placenta and labyrinth expression of SLC2A1 was higher in males than females (P(SEX) < 0.05). Labyrinthine IGF1R remained constant until term in the female, compared with male where expression increased until term. Peak MAP2K1 expression occurred earlier in the male placenta than the female. Spongy zone SLC2A1 remained constant until term in the female, compared with male where expression increased until term. CONCLUSIONS: The spiny mouse is a species that exhibits sexually dimorphic placental development. We suggest that these sex differences in placental gene expression and structure may underlie or compound the male vulnerability to a sub-optimal in utero environment.

IFPA Meeting 2011 workshop report III: Placental immunology; epigenetic and microRNA-dependent gene regulation; comparative placentation; trophoblast differentiation; stem cells.

Workshops are an important part of the IFPA annual meeting as they allow for discussion of specialised topics. At IFPA meeting 2011 there were twelve themed workshops, five of which are summarized in this report. These workshops related to various aspects of placental biology: 1) immunology; 2) epigenetics; 3) comparative placentation; 4) trophoblast differentiation; 5) stem cells.

Sex specific changes in placental growth and MAPK following short term maternal dexamethasone exposure in the mouse.

OBJECTIVES: Maternal glucocorticoid (GC) exposure during pregnancy can alter fetal development and program the onset of disease in adult offspring. The placenta helps protect the fetus from excess GC exposure but is itself susceptible to maternal insults and may be involved in sex dependant regulation of fetal programming. This study aimed to investigate the effects of maternal GC exposure on the developing placenta. STUDY DESIGN AND MAIN OUTCOME MEASURES: Pregnant mice were treated with dexamethasone (DEX-1 µg/kg/h) or saline (SAL) for 60 h via minipump beginning at E12.5. Placentas were collected at E14.5 and E17.5 and the expression of growth factors and placental transporters examined by real-time PCR and/or Western blot. Histological analysis was performed to assess for morphological changes. RESULTS: At E14.5, DEX exposed male and female fetuses had a lower weight compared to SAL animals but placental weight was lower in females only. Hsd11b2 and Vegfa gene expression was increased and MAPK1 protein expression decreased in the placentas of females only. At E17.5 placental and fetal body weights were similar and differences in MAPK were no longer present although HSD11B2 protein was elevated in placentas of DEX females. Levels of glucose or amino acid transporters were unaffected. CONCLUSIONS: Results suggest sex specific responses to maternal GCs within the placenta. Decreased levels of MAPK protein in placentas of female fetuses suggest alterations in the MAPK pathway may contribute to the lower placental weights in this sex. This may contribute towards sex specific fetal programming of adult disease.

A maternal diet supplemented with creatine from mid-pregnancy protects the newborn spiny mouse brain from birth hypoxia.

The creatine-phosphocreatine shuttle is essential for the maintenance of cellular ATP, particularly under hypoxic conditions when respiration may become anaerobic. Using a model of intrapartum hypoxia in the precocial spiny mouse (Acomys cahirinus), the present study assessed the potential for maternal creatine supplementation during pregnancy to protect the developing brain from the effects of birth hypoxia. On day 38 of gestation (term is 39 days), the pregnant uterus was isolated and placed in a saline bath for 7.5 min, inducing global hypoxia. The pups were then removed, resuscitated, and cross-fostered to a nursing dam. Control offspring were delivered by caesarean section and recovered immediately after release from the uterus. At 24 h after birth hypoxia, the brains of offspring from dams fed a normal diet showed significant increases in lipid peroxidation as measured by the amount of malondialdehyde. In the cortical subplate, thalamus and piriform cortex there were significant increases in cellular expression of the pro-apoptotic protein BAX, cytoplasmic cytochrome c and caspase-3. When pregnant dams were fed the creatine supplemented diet, the increase in malondialdehyde, BAX, cytochrome c and caspase 3 were almost completely prevented, such that they were not different from control (caesarean-delivered) neonates. This study provides evidence that the neuroprotective capacity of creatine in the hypoxic perinatal brain involves abrogation of lipid peroxidation and apoptosis, possibly through the maintenance of mitochondrial function. Further investigation into these mechanisms of protection, and the long-term development and behavioural outcomes of such neonates is warranted.

Search for a dark matter annihilation signal from the galactic center halo with H.E.S.S.

A search for a very-high-energy (VHE; ≥100 GeV) γ-ray signal from self-annihilating particle dark matter (DM) is performed towards a region of projected distance r∼45-150 pc from the Galactic center. The background-subtracted γ-ray spectrum measured with the High Energy Stereoscopic System (H.E.S.S.) γ-ray instrument in the energy range between 300 GeV and 30 TeV shows no hint of a residual γ-ray flux. Assuming conventional Navarro-Frenk-White and Einasto density profiles, limits are derived on the velocity-weighted annihilation cross section (σv) as a function of the DM particle mass. These are among the best reported so far for this energy range and in particular differ only little between the chosen density profile parametrizations. In particular, for the DM particle mass of ∼1 TeV, values for (σv) above 3×10(-25) cm(3) s(-1) are excluded for the Einasto density profile.

MicroRNA misregulation: an overlooked factor generating somaclonal variation?

Somaclonal variation is an important phenomenon that can be observed at high levels in plant tissue culture. Although known to science since plant cell culture techniques were first developed, its origins remain mysterious. Here, we propose that misregulation of microRNAs and small RNA pathways can make a significant contribution to the phenomenon. For many reasons, microRNAs and related small RNAs appear ideal candidates. Their mode of action gives them disproportionate influence over the transcriptome, proteome and epigenome. They regulate important developmental and physiological events such as meristem formation, phase changes and hormone responses. However, the genomic locations of microRNA genes and their unique biogenesis might make them unusually susceptible to aberrant regulation in vitro.

Small RNA activity and function in angiosperm gametophytes.

Small non-coding RNAs are key post-transcriptional and transcriptional regulators of plant gene expression in angiosperm sporophytes. In recent years, gametophytic small RNAs have also been investigated, predominantly in Arabidopsis male gametophytes, revealing features in common with the sporophyte as well as some surprising differences. Transcriptomic and deep-sequencing studies confirm that multiple small RNA pathways operate in male gametophytes, with over 100 miRNAs detected throughout development. Trans-acting siRNA pathways that are associated with novel phased transcripts in pollen, and the nat-siRNA pathway have important roles in pollen maturation and gamete function. Moreover, a role for siRNA-triggered silencing of transposable elements in male and female germ cells has been established, a feature in common with the role of piRNAs in animal germlines. Current evidence supports an integral role for small RNAs in angiosperm gametophyte development and it can be anticipated that novel small RNAs with significant roles in germline development and genome integrity await discovery.

Plasma aminotransferase concentrations in preterm infants.

The aim of this study was to generate reference ranges for aspartate aminotransferase (AST) and alanine aminotransferase (ALT) in preterm infants by describing the observed plasma concentration of these enzymes in babies born between 22 and 36 weeks' gestation. A service evaluation was conducted in babies admitted to two large neonatal intensive care units in the UK. 7006 blood samples from 1860 infants admitted to the two units between 2004 and 2008 were included. Extremely premature infants had high plasma enzyme activities when compared to babies at a later corrected gestational age. This may be due to more severe illness immediately after birth.