Prenatal genetic counseling for hemoglobinopathy carriers: a comparison of primary providers of prenatal care and professional genetic counselors.

Health personnel trained in medical genetics are insufficient to meet the demand for genetic services. Methods must be found to enable primary care providers to offer commonly needed genetic services themselves. In our recently reported community-wide prenatal screening program for hemoglobinopathies, 36% of women detected to have a hemoglobinopathy did not come to a tertiary center for counseling and thus may have not benefited from testing. To determine whether the efficiency of the program could be increased if counseling were provided by the prenatal care provider (obstetrician or family practitioner), we developed a pilot training program on the basis of our experience in offering such services and enlisted 68% of regional prenatal care providers to participate. The proportion of patients detected to have a hemoglobinopathy who received counseling was similar in the primary and tertiary provider groups: 59% versus 50%, respectively, for sickle trait, and 69% versus 66%, respectively, for beta-thalassemia trait. Knowledge after counseling was also similar for the primary and tertiary provider groups: 64% versus 66% (mean % correct), respectively, for sickle trait, and 79% versus 78%, respectively, for beta-thalassemia trait. However, the two provider groups significantly differed with regard to whether or not the patient had her partner tested. For sickle trait, it was 25% for the primary providers but 49% for the tertiary providers (P < .001). For beta-thalassemia trait, it was 47% for the primary providers but 78% for the tertiary providers (P < .001).(ABSTRACT TRUNCATED AT 250 WORDS)     

Calcium and the malaria parasite: parasite maturation and the loss of red cell deformability

In the studies reported here, we examined the role of calcium in the maturation of the human malaria parasite Plasmodium falciparum, and in the loss of red cell deformability associated with parasite maturation. P. falciparum alters the permeability of its host red cell, which normally maintains submicromolar cytoplasmic concentrations of calcium. Infection of the red cell and parasite maturation produce a 30-fold increase in calcium uptake. Both parasite maturation and the loss of red cell deformability are blocked by EGTA (by extracellular-free calcium concentrations less than or equal to 35 microM) and by other calcium antagonists. The loss of red cell deformability that occurs with parasite maturation is accompanied by alterations in the cytoskeletal proteins of parasitized red cells similar to those produced by the calcium ionophore A23187 (reductions in bands 2.1 [ankyrin], 4.1, and 5 [actin]). These results establish that parasite development and the loss of red cell deformability are calcium-dependent. They suggest that parasite-induced changes in the calcium permeability of the red cell activate endogenous transglutaminase activity by raising the free calcium concentration of the red cell cytoplasm.     

RecJ exonuclease: substrates, products and interaction with SSB

The RecJ exonuclease from Escherichia coli degrades single-stranded DNA (ssDNA) in the 5′–3′ direction and participates in homologous recombination and mismatch repair. The experiments described here address RecJ's substrate requirements and reaction products. RecJ complexes on a variety of 5′ single-strand tailed substrates were analyzed by electrophoretic mobility shift in the absence of Mg²⁺ ion required for substrate degradation. RecJ required single-stranded tails of 7 nt or greater for robust binding; addition of Mg²⁺ confirmed that substrates with 5′ tails of 6 nt or less were poor substrates for RecJ exonuclease. RecJ is a processive exonuclease, degrading ~1000 nt after a single binding event to single-strand DNA, and releases mononucleotide products. RecJ is capable of degrading a single-stranded tail up to a double-stranded junction, although products in such reactions were heterogeneous and RecJ showed a limited ability to penetrate the duplex region. RecJ exonuclease was equally potent on 5′ phosphorylated and unphosphorylated ends. Finally, DNA binding and nuclease activity of RecJ was specifically enhanced by the pre-addition of ssDNA-binding protein and we propose that this specific interaction may aid recruitment of RecJ.     

DNA repeat rearrangements mediated by DnaK-dependent replication fork repair

We propose that rearrangements between short tandem repeated sequences occur by errors made during a replication fork repair pathway involving a replication template switch. We provide evidence here that the DnaK chaperone of E. coli controls this template switch repair process. Mutants in dnaK are sensitive to replication fork damage and exhibit high expression of the SOS response, indicative of repair deficiency. Deletion and expansion of tandem repeats that occur by replication misalignment ("slippage") are also DnaK dependent. Because mutations in dnaX encoding the gamma and tau subunits of DNA polymerase III mimic dnaK phenotypes and are genetically epistatic, we propose that the DnaKJ chaperone remodels the replisome to facilitate repair. The fork remains largely intact because PriA or PriC restart proteins are not required. We also suggest that the poorly defined RAD6-RAD18-RAD5 mechanism of postreplication repair in eukaryotes occurs by an analogous mechanism to the DnaK template-switch pathway in prokaryotes.     

Effect of distraction osteogenesis of the mandible on upper airway volume and resistance in children with micrognathia

Children with craniofacial anomalies often have compromise of the upper airway, a condition with potential for morbidity and mortality. In children with microretrognathia, the diminutive size and retruded position of the mandible reduces the size of the oropharynx, thereby predisposing to glossoptosis and airway obstruction. Although several authors have reported successful use of mandibular distraction osteogenesis to alleviate this type of upper airway obstruction, the physiologic relationship between changes in mandibular shape, size, and position and upper airway dynamics remains undefined. The purpose of this study was to develop methodologies to quantitatively evaluate upper airway dynamics in children with micrognathia both before and after mandibular distraction osteogenesis. The patient population consisted of four children with micrognathia who had successfully undergone upper airway stabilization by bilateral mandibular distraction osteogenesis. The data used were digitally archived computed tomographic scan data from high-resolution, thin-slice head computed tomographic scans obtained before and after mandibular distraction. Upper airway evaluation was performed in two ways: static and dynamic. Static analysis consisted of computer quantification of predistraction and postdistraction mandibular and upper airway volumes using Analyze imaging software. Dynamic analysis consisted of fabrication of rigid stereolithographic hollow cast models of the upper airway produced from computed tomographic scan data. Models were used for characterization of upper airway resistance and flow patterns as related to respiration. After distraction osteogenesis, mandibular total volume increased 32, 32, 18, and 25 percent (mean, 27 percent) and upper airway volume increased by 20, 31, 23, and 71 percent (mean, 37 percent). A significant decrease in flow resistance, both inspiratory and expiratory, was observed in the patient with the greatest upper airway volume increase (71 percent) after distraction. After distraction, the inspiratory resistance was diminished by 51 percent and the expiratory resistance diminished by 85 percent. However, the three patients with more modest upper airway volume increases of 20 to 31 percent demonstrated no statistically significant change in flow resistance after distraction. Results of this study support the conclusion that distraction osteogenesis of the micrognathic mandible increases the volume of the upper airway, roughly paralleling the increase in mandibular volume. In the biomechanical airway model studied, upper airway volume expansion has been shown to be able to decrease the flow resistance over the length of the airway, presumably secondary to an increase in the average cross-sectional area. The artificial rigidity of the stereolithographic "airway" compared with the elasticity of the human upper airway may account for the insensitivity of this model to smaller but clinically significant airway changes.     

Acceptor specificity of the human leukocyte alpha3 fucosyltransferase: role of FucT-VII in the generation of selectin ligands

The alpha3 fucosyltransferase, FucT-VII, is one of the key glycosyltransferases involved in the biosynthesis of the sialyl Lewis X (sLex) antigen on human leukocytes. The sialyl Lewis X antigen (NeuAcalpha(2-3)Galbeta(1-4)[Fucalpha(1-3)]GlcNAc-R) is an essential component of the recruitment of leukocytes to sites of inflammation, mediating the primary interaction between circulating leukocytes and activated endothelium. In order to characterize the enzymatic properties of the leukocyte alpha3 fucosyltransferase FucT-VII, the enzyme has been expressed in Trichoplusia ni insect cells. The enzyme is capable of synthesizing both sLexand sialyl-dimeric-Lexstructures in vitro , from 3'-sialyl-lacNAc and VIM-2 structures, respectively, with only low levels of fucose transfer observed to neutral or 3'-sulfated acceptors. Studies using fucosylated NeuAcalpha(2-3)-(Galbeta(1- 4)GlcNAc)3-Me acceptors demonstrate that FucT-VII is able to synthesize both di-fucosylated and tri-fucosylated structures from mono- fucosylated precursors, but preferentially fucosylates the distal GlcNAc within a polylactosamine chain. Furthermore, the rate of fucosylation of the internal GlcNAc residues is reduced once fucose has been added to the distal GlcNAc. These results indicate that FucT-VII is capable of generating complex selectin ligands, in vitro , however the order of fucose addition to the lactosamine chain affects the rate of selectin ligand synthesis.      

An Examination of the Association between 5-HTTLPR,Combat Exposure,and PTSD Diagnosis among U.S. Veterans

Objective

To examine the association between the 5-HTTLPR polymorphism of the serotonin transporter (SLC6A4) gene, combat exposure, and posttraumatic stress disorder (PTSD) diagnosis and among two samples of combat-exposed veterans.

Method

The first sample included 550 non-Hispanic Black (NHB) combat-exposed veterans. The second sample included 555 non-Hispanic White (NHW) combat-exposed veterans. Participants were genotyped for the 5-HTTLPR/rs25531 variants of the SLC6A4 gene. A structured clinical interview was used to diagnose PTSD. Combat and civilian trauma exposure were assessed with validated self-report instruments. Logistic regression was used to test for main effects of 5-HTTLPR on PTSD diagnosis as well as gene x environment (GxE) interactions after adjusting for sex, ancestry proportion scores, civilian trauma exposure, and combat exposure.

Results

Within the NHB sample, a significant additive effect was observed for 5-HTTLPR (OR = 1.502, p = .0025), such that the odds of having a current diagnosis of PTSD increased by 1.502 for each additional S’ allele. No evidence for an association between 5-HTTLPR and PTSD was observed in the NHW sample. In addition, no evidence for combat x 5-HTTLPR effects were observed in either sample.

Conclusion

The present study suggests that there may be an association between 5-HTTLPR genotype and PTSD diagnosis among NHB veterans; however, no evidence for the hypothesized 5-HTTLPR x combat interaction was found.     

Sequential colonization by river periphyton analysed by microscopy and molecular fingerprinting

1. An artificial glass substratum was incubated in the River Danube for a period of 28 days in order to detect the sequential colonization of microorganisms.
2. Light and fluorescent microscopy showed that microalgae and the picoalgal fraction on the slides increased rapidly over the first 2 weeks of colonization. Diatoms were numerically the most abundant component of the periphyton and their species richness and diversity increased rapidly in the early phase of colonization whereas diversity subsequently increased moderately.
3. Evenness of the diatom community was initially high, lower in the intermediate phase and again higher later on. Succession involving early, intermediate and late colonizer species was observed. Community composition during the first 5 days of colonization was very different from later stages whereas there were only minor changes subsequently.
4. Molecular community analysis by means of terminal restriction fragment length polymorphism analysis of PCR amplified 16S rRNA and 18S rRNA genes pointed to even larger differences between the composition of samples obtained early and late in the period.
5. The number of 18S rRNA and 16S rRNA terminal restriction fragments (T-RF-s) was variable over the colonization period and the fragment patterns of both the bacterial and eukaryotic portion of the microbial community were variable, with most T-RF-s unique to a single sample, suggesting a wide diversity and dynamic properties of periphytic organisms.