Free fetal DNA in maternal plasma in anembryonic pregnancies: confirmation that the origin is the trophoblast.

OBJECTIVE: To test the hypothesis that free fetal DNA (ffDNA) circulating in maternal plasma originates mainly from the placenta we studied ffDNA levels in anembryonic pregnancies. METHODS: Maternal blood samples were collected from 15 normal first-trimester pregnancies in which fetal sex was subsequently determined and nine patients with a diagnosis of anembryonic gestation (AG). The Y chromosome DYS14 gene was quantified by real-time quantitative PCR (RT-PCR) for the determination of fetal sex in both plasma and chorionic tissue samples. Fetal sex in chorionic tissue samples was also determined using quantitative fluorescence PCR (QF-PCR). RESULTS: The correct sex result was obtained from maternal plasma in all. Four AG pregnancies were female (DYS14 negative) results. In five of the AG cases, the chorionic tissue was found to be male (by both QF-PCR and RT-PCR which agreed) and positive male signal was found in maternal plasma by RT-PCR. There was no statistical difference between median free fetal DNA concentration in plasma between the AG male cases (148.3 GE/mL) and controls (145.8 GE/mL). CONCLUSION: Since ffDNA levels are normal in pregnancies without a fetus, the data support the hypothesis that the trophoblastic cells are the major source ffDNA in maternal plasma.

Genome sequence of Halobacterium species NRC-1.

We report the complete sequence of an extreme halophile, Halobacterium sp. NRC-1, harboring a dynamic 2,571,010-bp genome containing 91 insertion sequences representing 12 families and organized into a large chromosome and 2 related minichromosomes. The Halobacterium NRC-1 genome codes for 2,630 predicted proteins, 36% of which are unrelated to any previously reported. Analysis of the genome sequence shows the presence of pathways for uptake and utilization of amino acids, active sodium-proton antiporter and potassium uptake systems, sophisticated photosensory and signal transduction pathways, and DNA replication, transcription, and translation systems resembling more complex eukaryotic organisms. Whole proteome comparisons show the definite archaeal nature of this halophile with additional similarities to the Gram-positive Bacillus subtilis and other bacteria. The ease of culturing Halobacterium and the availability of methods for its genetic manipulation in the laboratory, including construction of gene knockouts and replacements, indicate this halophile can serve as an excellent model system among the archaea.

Evidence of genetic diversity underlying Rh D-, weak D (Du), and partial D phenotypes as determined by multiplex polymerase chain reaction analysis of the RHD gene.

The human blood group Rh antigens are expressed by proteins encoded by a pair of highly homologous genes located at chromosome 1p34-36. One of the genes (RHCE) encodes Rh CcEe antigens, while the other (RHD) the D antigen. Point mutations in the RHCE gene generate the C/c and E/e polymorphisms, while it has been shown that an RHD gene deletion can generate the D-negative phenotype. We have analyzed intron 4 of the RHCE and RHD genes and have defined the site of an RHD-specific deletion located in this intron. Using a multiplex RHD typing assay, which combines a reverse polymerase chain reaction (PCR) primer, which straddles this RHD-specific sequence, and a pair of primers located in exon 10 of the RHD gene, we have analyzed 357 different genomic DNA samples derived from individuals expressing D+, D-, weak D, and partial D phenotypes. Of these, we have noted a significant discordance with our multiplex PCR assay in the D- phenotypes dCcee and dccEe (which have been previously described) and weak D phenotypes. Our results suggest that in five serologically D- individuals we have identified an apparently intact RHD gene. Sequence analysis of transcripts obtained from one of these individuals (of phenotype dCCee) illustrates the presence of full-length RHD transcripts, which have a point mutation at nucleotide 121 (C --> T), which generates an in-frame stop codon (Gln41Stop). Thus, we describe a different molecular basis for generating the D- phenotype to the complete RHD gene deletion described previously. We also show that there are discordances with serotype and the multiplex assay in weak D and partial D phenotypes, indicating that the underlying molecular basis can be heterogeneous. Existing Rh D PCR assays assume the complete absence of the RHD gene in D- phenotypes. We describe a different molecular basis for generating the D- phenotype to the complete RHD gene deletion described previously.

Neuropsychological deficits following concussion.

The term 'concussion' has been applied to head injuries of varying severity. Most studies have examined subjects suffering concussion of a severity requiring hospital observation, usually as a consequence of motor vehicle accidents. Milder concussive injuries such as those resulting from contact sport are often not reported in hospital-based studies. In this study, subjects with mild concussive injuries were studied with the aim of determining if neuropsychological sequelae are detectable. The subjects received their injuries while playing Australian Rules Football. Baseline (pre-injury) measures on the Paced Auditory Serial Addition Test (PASAT), Digit Symbol Substitution Test (DSST) and Four-Choice Reaction time, involving measures of decision time (DT) and movement time (MT), were obtained in a sample of 130 players. Ten players subsequently concussed were re-tested at 5 days post-injury. A control group of age-matched umpires were assessed on two corresponding occasions. Analyses of covariance showed poorer performances following concussion on the DSST and DT measures. The results suggested that neuropsychological deficits are detectable after resolution of neurological symptoms in the early stages following mild concussive injury.

The assessment of orientation following concussion in athletes.

The purpose of this study was to investigate the sensitivity of orientation and recent memory questions in the diagnosis of concussion. In a prospective study over 7 consecutive years (1985-1991), all players at a professional Australian Rules Football club who sustained a concussive injury (n = 28) were administered a set of questions evaluating orientation and recent memory. Concussion was diagnosed independently on the basis of loss or disturbance of consciousness and clinical symptoms. A control group of age-matched nonconcussed players was administered the same set of questions. The results showed that items evaluating recently acquired information were more sensitive in the assessment of concussion than standard orientation items. The relative sensitivity of orientation questions must be considered when they are used in the clinical diagnosis of concussion in sport.

Hairy roots of Brassica napus: I. Applied glutamine overcomes the effect of phosphinothricin treatment.

Hairy roots of Brassica napus (rape cv. Giant) were produced by cocultivating leaf and cotyledon explants with Agrobacterium rhizogenes strain A4T. The hairy roots grew prolifically on solid and in liquid media. Incorporation of ammonium sulphate or phosphinothricin (PPT) into the media reduced growth. PPT treatment reduced glutamine synthetase (GS) activity and increased the ammonia content of the hairy roots. We have found that PPT treatment also induces a loss of glutamine from the roots and this may influence root growth. To test this we grew hairy roots in a liquid medium containing 10 mM glutamine. This glutamine treatment overcame the PPT induced suppression of growth but also significantly increased GS activity, reduced ammonia accumulation and increased the levels of glutamate and asparagine.

Clinical management of concussion.

Concussion is an injury that occurs frequently in the general community. Most cases are caused by sporting accidents, falls and assaults. This article reviews the presentation and management of this common problem. High velocity impacts as in motor vehicle accidents may result in concussion but are more likely to produce a serious brain injury.