Gulf of Mexico blue hole harbors high levels of novel microbial lineages.

Exploration of oxygen-depleted marine environments has consistently revealed novel microbial taxa and metabolic capabilities that expand our understanding of microbial evolution and ecology. Marine blue holes are shallow karst formations characterized by low oxygen and high organic matter content. They are logistically challenging to sample, and thus our understanding of their biogeochemistry and microbial ecology is limited. We present a metagenomic and geochemical characterization of Amberjack Hole on the Florida continental shelf (Gulf of Mexico). Dissolved oxygen became depleted at the hole's rim (32 m water depth), remained low but detectable in an intermediate hypoxic zone (40-75 m), and then increased to a secondary peak before falling below detection in the bottom layer (80-110 m), concomitant with increases in nutrients, dissolved iron, and a series of sequentially more reduced sulfur species. Microbial communities in the bottom layer contained heretofore undocumented levels of the recently discovered phylum Woesearchaeota (up to 58% of the community), along with lineages in the bacterial Candidate Phyla Radiation (CPR). Thirty-one high-quality metagenome-assembled genomes (MAGs) showed extensive biochemical capabilities for sulfur and nitrogen cycling, as well as for resisting and respiring arsenic. One uncharacterized gene associated with a CPR lineage differentiated hypoxic from anoxic zone communities. Overall, microbial communities and geochemical profiles were stable across two sampling dates in the spring and fall of 2019. The blue hole habitat is a natural marine laboratory that provides opportunities for sampling taxa with under-characterized but potentially important roles in redox-stratified microbial processes.

Shifting microbial communities sustain multiyear iron reduction and methanogenesis in ferruginous sediment incubations.

Reactive Fe(III) minerals can influence methane (CH4 ) emissions by inhibiting microbial methanogenesis or by stimulating anaerobic CH4 oxidation. The balance between Fe(III) reduction, methanogenesis, and CH4 oxidation in ferruginous Archean and Paleoproterozoic oceans would have controlled CH4 fluxes to the atmosphere, thereby regulating the capacity for CH4 to warm the early Earth under the Faint Young Sun. We studied CH4 and Fe cycling in anoxic incubations of ferruginous sediment from the ancient ocean analogue Lake Matano, Indonesia, over three successive transfers (500 days in total). Iron reduction, methanogenesis, CH4 oxidation, and microbial taxonomy were monitored in treatments amended with ferrihydrite or goethite. After three dilutions, Fe(III) reduction persisted only in bottles with ferrihydrite. Enhanced CH4 production was observed in the presence of goethite, highlighting the potential for reactive Fe(III) oxides to inhibit methanogenesis. Supplementing the media with hydrogen, nickel and selenium did not stimulate methanogenesis. There was limited evidence for Fe(III)-dependent CH4 oxidation, although some incubations displayed CH4 -stimulated Fe(III) reduction. 16S rRNA profiles continuously changed over the course of enrichment, with ultimate dominance of unclassified members of the order Desulfuromonadales in all treatments. Microbial diversity decreased markedly over the course of incubation, with subtle differences between ferrihydrite and goethite amendments. These results suggest that Fe(III) oxide mineralogy and availability of electron donors could have led to spatial separation of Fe(III)-reducing and methanogenic microbial communities in ferruginous marine sediments, potentially explaining the persistence of CH4 as a greenhouse gas throughout the first half of Earth history.

Metatranscriptomics reveal differences in in situ energy and nitrogen metabolism among hydrothermal vent snail symbionts.

Despite the ubiquity of chemoautotrophic symbioses at hydrothermal vents, our understanding of the influence of environmental chemistry on symbiont metabolism is limited. Transcriptomic analyses are useful for linking physiological poise to environmental conditions, but recovering samples from the deep sea is challenging, as the long recovery times can change expression profiles before preservation. Here, we present a novel, in situ RNA sampling and preservation device, which we used to compare the symbiont metatranscriptomes associated with Alviniconcha, a genus of vent snail, in which specific host-symbiont combinations are predictably distributed across a regional geochemical gradient. Metatranscriptomes of these symbionts reveal key differences in energy and nitrogen metabolism relating to both environmental chemistry (that is, the relative expression of genes) and symbiont phylogeny (that is, the specific pathways employed). Unexpectedly, dramatic differences in expression of transposases and flagellar genes suggest that different symbiont types may also have distinct life histories. These data further our understanding of these symbionts' metabolic capabilities and their expression in situ, and suggest an important role for symbionts in mediating their hosts' interaction with regional-scale differences in geochemistry.

The Calyptogena magnifica chemoautotrophic symbiont genome.

Chemoautotrophic endosymbionts are the metabolic cornerstone of hydrothermal vent communities, providing invertebrate hosts with nearly all of their nutrition. The Calyptogena magnifica (Bivalvia: Vesicomyidae) symbiont, Candidatus Ruthia magnifica, is the first intracellular sulfur-oxidizing endosymbiont to have its genome sequenced, revealing a suite of metabolic capabilities. The genome encodes major chemoautotrophic pathways as well as pathways for biosynthesis of vitamins, cofactors, and all 20 amino acids required by the clam.

The adult phenotype in Costello syndrome.

We report clinical findings in 17 adults with Costello syndrome ranging in age from 16 to 40 years. Two patients in this series have had bladder carcinoma, the only malignancy reported to affect adults with Costello syndrome. Benign tumors included multiple ductal papillomata in two women, and a fourth ventricle mass in one man, thought to be a choroid plexus papilloma. Endocrine problems in this series were osteoporosis, central hypogonadism, and delayed puberty. Other health problems were symptomatic Chiari malformations in three patients. Four patients had adult-onset gastro-esophageal reflux, three of whom had Chiari malformations. Fourteen adults had mild to moderate intellectual disability with three individuals having severe intellectual disability; 15 individuals attained some reading and writing skills and 14 showed ongoing acquisition of new skills into adulthood. On the basis of this data, we recommend that neuro-imaging be considered in adults with Costello syndrome if they develop symptoms suggestive of a Chiari malformation. In the event of pubertal delay, endocrine investigations are indicated and hormone treatment may be required. Bone density assessments should be performed in adults with Costello syndrome, particularly in those with pubertal abnormalities. Screening for microscopic hematuria as a marker for bladder carcinoma may be indicated, although this requires further evaluation.

Methyl-specific DNA binding by McrBC, a modification-dependent restriction enzyme.

McrBC, a GTP-requiring, modification-dependent endonuclease of Escherichia coli K-12, specifically recognizes DNA sites of the form 5' R(m)C 3'. DNA cleavage normally requires translocation-mediated coordination between two such recognition elements at distinct sites. We have investigated assembly of the cleavage-competent complex with gel-shift and DNase I footprint analysis. In the gel-shift system, McrB(L) binding resulted in a fast-migrating specific shifted band, in a manner requiring both GTP and Mg(2+). The binding was specific for methylated DNA and responded to local sequence changes in the same way that cleavage does. Single-stranded DNA competed for McrB(L)-binding in a modification and sequence-specific fashion. A supershifted species was formed in the presence of McrC and GTPgammaS. DNase I footprint analysis showed modest cooperativity in binding to two sites, and a two-site substrate displayed protection in non-specific spacer DNA in addition to the recognition elements. The addition of McrC did not affect the footprint obtained. We propose that McrC effects a conformational change in the complex rather than a reorganization of the DNA:protein interface.

A simple in vitro Tn7-based transposition system with low target site selectivity for genome and gene analysis.

A robust Tn7-based in vitro transposition system is described that displays little target site selectivity, allowing the efficient recovery of many different transposon insertions in target DNAs ranging from small plasmids to cosmids to whole genomes. Two miniTn7 derivatives are described that are useful for the analysis of genes: one a derivative for making translational and transcriptional target gene fusions and the other a derivative that can generate 15 bp (5 amino acid) insertions in target DNAs (proteins).

Early amniocentesis: effect of removing a reduced volume of amniotic fluid on pregnancy outcome.

In mid-trimester amniocentesis (MTA), 12-15 ml of amniotic fluid is aspirated for cytogenetic analysis. When a similar volume of amniotic fluid is removed by early amniocentesis (EA), it represents a significant proportion of the total amniotic fluid volume in the first trimester. The fluid depletion, which may persist for 7 to 10 days, is considered to impair development of fetal lungs and extremities and, possibly, contribute towards procedure-related congenital abnormalities and miscarriages. By only removing 7 ml of amniotic fluid, we have demonstrated a total miscarriage rate (3.8 per cent) comparable with previous large studies (Table V), a low incidence of respiratory difficulties at birth (2.7 per cent) and a low incidence of fixed flexion deformities (1.6 per cent), at the expense of a small increase in the incidence of culture failure (2.2 per cent).

Dependence of McrBC cleavage on distance between recognition elements.

DNA cleavage by the modification-dependent restriction enzyme McrBC requires the presence of two suitably modified recognition elements appropriately spaced in the substrate. To characterize the spacing requirement in more detail, we have constructed a plasmid with a single McrBC cleavage site, in which the distance between recognition elements could be systematically varied while preserving the local sequence surrounding the recognition elements. Optimal separation between elements was 55-103 basepairs, with detectable cleavage observed at spacing of 32 bp to 2 kb; no cleavage was seen with spacing of 22 bp or less or with 3 kb between elements. Changing the spacing by 4 basepairs within the optimal range had little effect on the efficiency of cleavage, suggesting that the recognition elements need not lie on the same face of the DNA helix.

Plantar lipomatosis, unusual facial phenotype and developmental delay: a new MCA/MR syndrome.

We describe two boys with global developmental delay and a phenotype of microcephaly, midface hypoplasia, enlarged fleshy ears, depressed nasal bridge, anteverted nostrils, central palatal ridge, and high forehead. Bilateral congenital fat pads are present anteromedial to the heels. Fetal finger and toe pads are present and palmar and plantar grooves are deeper than normal with "pillowing" of the areas between the grooves. No patients with similar clinical findings have been located, but these two children have a remarkably similar clinical presentation which we consider a "new" syndrome.

Rural and urban female secondary school students' attitudes towards and use of primary care services.

In light of the current and ongoing threats to young women's health of unplanned pregnancy and sexually transmissible diseases (including HIV/AIDS), assured access to appropriate and sensitive health services is of paramount importance. In this article, the results of a recent Victorian study involving rural and urban female secondary school students on issues relating to sexual health are reported. Drawing upon the findings of an extensive self-report questionnaire, young women's attitudes towards and use of health services are presented with a particular focus upon the variations that exist between the rural and urban populations. Apart from issues associated with privacy, which were found to be more crucial to rural young women's decisions to seek medical care, this study found young women's concern about the attitudes of medical and non-medical staff alike, the atmosphere of surgery or clinic, and gender of doctor all to be important factors in their general use of health services, regardless of location.

Autopsy findings in the Wolcott-Rallison syndrome.

Wolcott-Rallison syndrome is a rare autosomal recessive condition characterized by diabetes mellitus arising in early infancy and multiple epiphyseal dysplasia. To date, nine cases have been described in the world literature. We report an affected girl who died at the age of 4 years and on whom a full autopsy was performed. In addition to neonatal diabetes mellitus and epiphyseal dysplasia, this child had mental retardation and recurrent episodes of self-limiting hepatic failure. Autopsy revealed severe pancreatic hypoplasia and markedly abnormal pancreatic histology, while histology of the bone was consistent with epiphyseal dysplasia. There was laryngeal stenosis and pulmonary hypoplasia. The heart was enlarged with mitral value dysplasia and stenosis, left atrial dilatation, left ventricular hypertrophy, and endocardial fibroelastosis. Examination of the central nervous system showed arrhinencephaly and cerebellar cortical dysplasia. The liver showed minor histological abnormalities but no features were present to account for the recurrent hepatic failure. In addition to Wolcott-Rallison syndrome this child had a deletion at 15q11-12 in 65% of her cells.

An autosomal dominant syndrome of acromegaloid facial appearance and generalised hypertrichosis terminalis.

We report a family in which a phenotype of acromegaloid facial appearance (AFA) and generalised hypertrichosis terminalis segregates through three generations. Congenital hypertrichosis terminalis and AFA have been previously reported as independent autosomal dominant traits. This is the first report to delineate an autosomal dominant transmission of the combined phenotype.

Focal dermal hypoplasia (Goltz syndrome) associated with intestinal malrotation and mediastinal dextroposition.

Focal dermal hypoplasia (Goltz syndrome) is a rare syndrome comprising developmental anomalies of tissues and organs of mesoectodermal derivation. We report on a characteristic case of focal dermal hypoplasia with the previously unreported association of mediastinal dextroposition and intestinal malrotation.

Wolcott-Rallison syndrome associated with congenital malformations and a mosaic deletion 15q 11-12.

Wolcott-Rallison syndrome is a rare autosomal recessive condition first described in 1972. It is characterised by diabetes mellitus which arises in early infancy and multiple epiphyseal dysplasia. We describe an affected girl who had recurrent episodes of hepatic failure for which no obvious cause was found. Post-mortem examination revealed abnormal pancreatic histology and congenital abnormalities of the central nervous and cardio-respiratory systems which have not been previously described in this condition. She also demonstrated a deletion at 15q 11-12 in 65% of her cells.

Automated sequencing detects all mutations in Northern Irish patients with phenylketonuria and mild hyperphenylalaninaemia.

In the first phase of the Northern Ireland PKU Study, we used automated sequencing to identify the spectrum of mutations in a random group of 32 unrelated phenylketonuria (PKU) families. We also investigated 7 Northern Irish patients with mild hyperphenylalaninaemia not requiring dietary intervention (MHP, previously referred to as non-PKU HPA). Disease-causing mutations were identified on all 78 investigated chromosomes. We found 23 different mutations, including 20 missense, 1 nonsense and 2 splice site mutations. All mutations were located within exons or at intron-exon boundaries of the phenylalanine hydroxylase gene. Seven mutations occurred at CpG sites, confirming these sites as mutation hot-spots in PKU. Mutations R408W and I65T are the two commonest PKU mutations in the Northern Irish population. Two mutations (T380M and V245A) can be characterized as MHP mutations; they are quasi dominant markers for MHP since they cause mild hyperphenylalaninaemia even when occurring in conjunction with the most severe PKU mutations. The results have proven valuable for the development of a routine PKU mutation analysis system in Northern Ireland.