Simple DNA extraction method for dried blood spots and comparison of two PCR assays for diagnosis of vertical human immunodeficiency virus type 1 transmission in Rwanda

Dried blood spots (DBS) on filter paper facilitate the collection, transport, and storage of blood samples for laboratory use. A rapid and simple DNA extraction procedure from DBS was developed and evaluated for the diagnosis of human immunodeficiency virus type 1 (HIV-1) infection in children by an in-house nested-PCR assay on three genome regions and by the Amplicor HIV-1 DNA prototype assay version 1.5 (Roche Molecular Systems). A total of 150 samples from children born to HIV-1-infected mothers were collected in Kigali, Rwanda, in parallel as DBS and as peripheral blood mononuclear cell (PBMC) pellets. The results obtained on DBS by the two PCR assays were compared to the results of nested PCR on PBMCs. Of 150 PBMC samples, 10 were positive, 117 were negative, and 23 were indeterminate for HIV-1 infection. In DNA extracted from filter papers and amplified by using the in-house nested PCR, 9 of these 10 positive samples (90%) were found to be positive, and 1 was found to be indeterminate (only the pol region could be amplified). All of the negative samples and all of the 23 indeterminate samples tested negative for HIV-1 infection. When we used the Amplicor DNA test on DBS, all of the 10 PBMC-positive samples were found to be positive and all of the 23 indeterminate samples were found to be negative. Of the PBMC-negative samples, 115 were found to be negative and 2 were found to be indeterminate. We conclude that this simple rapid DNA extraction method on DBS in combination with both detection methods gave a reliable molecular diagnosis of HIV-1 infection in children born to HIV-infected mothers.     

Clinical and genetic analysis of Korean patients with Marfan syndrome: possible ethnic differences in clinical manifestation

Yoo E‐H, Woo H, Ki C‐S, Lee HJ, Kim D‐K, Kang I‐S, Park P, Sung K, Lee CS, Chung T‐Y, Moon JR, Han H, Lee S‐T, Kim J‐W. Clinical and genetic analysis of Korean patients with Marfan syndrome: possible ethnic differences in clinical manifestation. Marfan syndrome (MFS) is an autosomal dominant disorder of the fibrous connective tissue caused by mutations in the fibrillin‐1 (FBN1) gene. Although clinical and genetic analyses have been performed in various populations, there have been few studies in Korea. The aim of this study was to investigate the clinical characteristics and genetic background of Korean patients with MFS. In 39 Korean patients with MFS who met the Ghent criteria, the most common clinical finding was aortic dilatation and/or dissection (94.9%), whereas only 35.9% of patients had ectopia lentis. The majority of MFS patients had fewer than four of the skeletal findings required to fulfill the major skeletal Ghent criterion for MFS. Only 21% of Korean patients had major skeletal abnormalities and most cases showed only minor skeletal involvement. FBN1 gene mutations were detected in 35 out of 39 patients (89.7%), which is similar to rates presented in the previous reports. These results suggest that some clinical features in Korean patients with MFS differed from those reported in Western MFS patients.     

Flashlamp-excited dye laser therapy of idiopathic vulvodynia is safe and efficacious

Objective: The management of chronic vulvovaginal pain, not explicable on specific histologic grounds, presents a major problem in referral centers for lower genital tract diseases.Study design: This article reports on a two-step protocol in a sample of 175 medical nonresponders, drawn from a 2-year cohort of 725 women with vulvovaginal pain. The first maneuver was the use of a flashlamp-excited dye laser to selectively photocoagulate symptomatic subepithelial blood vessels in 168 women; the second was the microsurgical removal of chronically painful Bartholin's glands in 52 women not responsive or not suited to flashlamp-excited dye laser photothermolysis.Results: Dye laser response rates werer independent of whether patients manifested macroscopic focl of painful erythema (“vestibular adenitis”) or just colposcopically apparent hyperemia-ectasia of the individual blood vessels (“pruritic papillomatosis”) (55% vs 45% after a single surgical procedure; 76% vs 65% after serial retreatment; p not significant). Conversely, response rates were much lower among women in whom pressure on the Bartholin's glands produced sharp, lancinating pain (15% vs 66% after a single surgical procedure; 22% vs 93% after serial retreatment; p < 0.001). Forty-two (85%) of 50 patients with flashlamp-excited dye laser failure had deep pain; however, the impasse to progress was broken by gland removal. Final response rates were 92.5% (complete response 62%; partial response 30%) in the “surface-only” group and 80.3% in the “surface-plus-deep” group (χ² = 14.9; p < 0.001). The major complication was acute bacterial cellulitis, occurring in the first postoperative week. Modification of the treatment protocol to include topical antibiotics with an occlusive dressing reduced the cellulitis rate from 17.2% to 2.5%. In four women (1.8%) Koebner-like exophytic condylomas also developed within 1 month of flashlamp-excited dye laser surgery.Conclusion: The availability of a safe, efficacious, and relatively noninvasive treatment should reduce the need for resective surgery in most patients with idiopathic vulvodynia. 1995; 172; 1684–1701.)     

Behavioural characteristics,prevalence of Chlamydia trachomatis and antibiotic susceptibility of Neisseria gonorrhoeae in men with urethral discharge in Thyolo,Malawi

A study was carried out in 2000/2001 in a rural district of Malawi among men presenting with urethral discharge, in order to (a) describe their health-seeking and sexual behaviour, (b) determine the prevalence of Neisseria gonorrhoeae and Chlamydia trachomatis, and (c) verify the antibiotic susceptibility of N. gonorrhoeae. A total of 114 patients were entered into the study; 61% reported having taken some form of medication before coming to the sexually transmitted infections clinic. The most frequent alternative source of care was traditional healers. Sixty-eight (60%) patients reported sexual encounters during the symptomatic period, the majority (84%) not using condoms. Using ligase chain reaction on urine, N. gonorrhoeae was detected in 91 (80%) and C. trachomatis in 2 (2%) urine specimens. Forty five of 47 N. gonorrhoeae isolates produced penicillinase, 89% showing multi-antimicrobial resistance. This study emphasizes the need to integrate alternative care providers and particularly traditional healers in control activities, and to encourage their role in promoting safer sexual behaviour. In patients presenting with urethral discharge in our rural setting, C. trachomatis was not found to be a major pathogen. Antimicrobial susceptibility surveillance of N. gonorrhoeae is essential in order to prevent treatment failures and control the spread of resistant strains.     

Human immunodeficiency virus dementia: evidence of a subcortical process from studies of fine finger movements

HIV-1 associated dementia is the major manifestation of HIV-1 within the central nervous system and a devastating disease which is characterized by cognitive, motor, and emotional deficits. HIV-1 associated minor motor deficits can manifest as psychomotor slowing and predict the later development of HIV-1 associated dementia, AIDS, and death. These minor motor deficits can be described, e.g., by electrophysiological assessment of basal ganglia motor function (frequency of most rapid alternating finger movements, reaction and contraction times of most rapid index finger extensions). Minor motor deficits quantified by contraction times can be subdivided into a more incipient and a more sustained type of deficit. Parallel examination of motor function and positron emission tomography, magnetic resonance spectroscopy of the basal ganglia, or SPECT helps to point to the basal ganglia as a pivotal point of HIV-1 associated CNS pathology.     

Inverse association of Pin1 and tau accumulation in Alzheimer's disease hippocampus

Neurofibrillary degeneration, one of the pathological hallmarks of Alzheimer's disease, is not ubiquitous to all brain regions or neurons. While a high degree of vulnerability has been documented for entorhinal cortex, hippocampal and neocortical pyramidal neurons other brain structures are largely spared. Even within highly vulnerable regions such as hippocampus neurons are affected to a variable extent. The molecular basis for this selective susceptibility remains unknown. Neurofibrillary degeneration involves hyperphosphorylation of tau which critically impairs its binding capacity to microtubule and, therefore, is believed to disrupt the axonal cytoskeleton. Recently, Lu et al. [Nature (1999) 399:784] described the ability of the peptidyl-prolyl cis-trans isomerase Pin1 to recover microtubule-binding affinity and microtubule stabilisation of phosphorylated tau. In the present study, we analysed the potential involvement of Pin1 in selective vulnerability of hippocampal neurons to neurofibrillary degeneration in Alzheimer's disease. Pin1 immunoreactivity appeared as cytoplasmic granules affecting hippocampal subfields to a different extent (CA2>subiculum>CA1>CA3/CA4). Since the main markers of granulovacuolar degeneration do not co-label Pin1-immunoreactive granules, we propose that these granules may represent a new lesion in Alzheimer's disease. Neurons containing Pin1 granules were devoid of neurofibrillary tangles. Granular accumulation of Pin1 may correspond to an absence of neurofibrillary lesions in these cells and might be associated with other mechanisms of neuronal degeneration.     

Effects of lesions of the caudal cerebellar vermis on cardiovascular regulation in awake cats

The vestibular system is known to participate in cardiovascular regulation during movement and postural alterations. The present study considered whether lesions of two regions of the posterior cerebellar vermis (the nodulus and uvula) that provide inputs to vestibular nucleus regions that affect control of blood pressure would alter cardiovascular responses during changes in posture. Blood pressure and heart rate were monitored in awake cats during nose-up tilts up to 60 degrees in amplitude before and following aspiration lesions of the nodulus or uvula; in most animals, cardiovascular responses were also recorded following the subsequent removal of vestibular inputs. Lesions of the nodulus or uvula did not affect baseline blood pressure or heart rate, although cardiovascular responses during nose-up tilts were altered. Increases in heart rate that typically occurred during 60 degrees nose-up tilt were attenuated in all three animals with lesions affecting both dorsal and ventral portions of the uvula; in contrast, the heart rate responses were augmented in the two animals with lesions mainly confined to the nodulus. Furthermore, following subsequent removal of vestibular inputs, uvulectomized animals, but not those with nodulus lesions, experienced more severe orthostatic hypotension than has previously been reported in cerebellum-intact animals with bilateral labyrinthectomies. These data suggest that the cerebellar nodulus and uvula modulate vestibulo-cardiovascular responses, although the two regions play different roles in cardiovascular regulation.