Estimating the relative frequency of leukodystrophies and recommendations for carrier screening in the era of next‐generation sequencing

Leukodystrophies are a heterogeneous group of heritable disorders characterized by abnormal brain white matter signal on magnetic resonance imaging (MRI) and primary involvement of the cellular components of myelin. Previous estimates suggest the incidence of leukodystrophies as a whole to be 1 in 7,000 individuals, however the frequency of specific diagnoses relative to others has not been described. Next generation sequencing approaches offer the opportunity to redefine our understanding of the relative frequency of different leukodystrophies. We assessed the relative frequency of all 30 leukodystrophies (associated with 55 genes) in more than 49,000 exomes. We identified a relatively high frequency of disorders previously thought of as very rare, including Aicardi Goutières Syndrome, TUBB4A‐related leukodystrophy, Peroxisomal biogenesis disorders, POLR3‐related Leukodystrophy, Vanishing White Matter, and Pelizaeus‐Merzbacher Disease. Despite the relative frequency of these conditions, carrier‐screening laboratories regularly test only 20 of the 55 leukodystrophy‐related genes, and do not test at all, or test only one or a few, genes for some of the higher frequency disorders. Relative frequency of leukodystrophies previously considered very rare suggests these disorders may benefit from expanded carrier screening.     

Genetic basis for remating inDrosophila melanogaster. III. Correlated responses to selection for female remating speed

Two natural populations ofDrosophila melanogaster selected for fast and slow female remating (Gromko and Newport,Behav. Genet.18: 621–632, 1988) were analyzed for correlated responses to selection. Several of the correlated responses differed between the lines derived from the two different source populations. In particular, the degree to which female receptivity to remating was dependent on the number of sperm in storage ceptivity to remating was dependent on the number of sperm in storage remained high in one pair of fast and slow selected lines but not in the other. In the line which retained sperm dependence (JEFFERS), fast and slow remating was achieved by changes in the threshold at which remating occurred. In the other line (COMP) changes in the temporal pattern of sperm use were seen. Thus both the level of the receptivity threshold and the existence of a sperm effect appeared to be selectable features of this species. Mated female attractiveness and receptivity were shown to be genetically distinct from virgin female attractiveness and receptivity. Effects of the genotype of the female on copulation duration were found.This work was supported by NSF Grants 82-01183 and 84-18934.     

Immunoglobulin M antibody responses to Mycobacterium ulcerans allow discrimination between cases of active Buruli ulcer disease and matched family controls in areas where the disease is endemic

Buruli ulcer disease (BUD) is an emerging disease caused by Mycobacterium ulcerans. In the present study we have characterized the serological reactivities of sera from volunteer case patients with laboratory-confirmed BUD and controls living in three different regions of Ghana where the disease is endemic to determine if serology may be useful for disease confirmation. Our results showed highly reactive immunoglobulin G (IgG) responses among patients with laboratory-confirmed disease, healthy control family members of the case patients, and sera from patients with tuberculosis from areas where BUD is not endemic. These responses were represented by reactivities to multiple protein bands found in the M. ulcerans culture filtrate (CF). In contrast, patient IgM antibody responses to the M. ulcerans CF (MUCF) proteins were more distinct than those of healthy family members living in the same village. A total of 84.8% (56 of 66) of the BUD patients exhibited strong IgM antibody responses against MUCF proteins (30, 43 and 70 to 80 kDa), whereas only 4.5% (3 of 66) of the family controls exhibited such responses. The sensitivity of the total IgM response for the patients was 84.8% (95% confidence interval [CI], 74.3 to 91.6%), and the specificity determined with sera from family controls was 95.5% (95% CI, 87.5 to 98.4%). These studies suggest that the IgM responses of patients with BUD will be helpful in the identification and production of the M. ulcerans recombinant antigens required for the development of a sensitive and specific serological assay for the confirmation of active BUD.     

Basophil recruitment and IL-4 production during human allergen-induced late asthma

BACKGROUND: Basophils represent an important source of inflammatory mediators and cytokines after IgE-dependent activation in human beings. OBJECTIVE: To assess the role of basophils in allergic asthma, we measured the number of basophils in the bronchial mucosa and their capacity to express IL-4 mRNA and protein during allergen-induced late asthmatic responses. METHODS: Fiberoptic bronchoscopic bronchial biopsies were obtained at 24 hours from sites of segmental bronchial allergen challenge and control sites in 19 patients with atopic asthma and 6 nonatopic healthy volunteers. Basophil numbers were assessed by immunohistochemistry through use of mAb 2D7. IL-4 mRNA--positive cells were detected through use of in situ hybridization and colocalized to basophils through use of sequential immunohistochemistry/in situ hybridization. IL-4 protein was detected and colocalized to basophils through use of dual immunohistochemistry. RESULTS: After allergen challenge, there was an increase in the median number of 2D7-positive basophils per square millimeter in the bronchial mucosa in patients with asthma (0.9 cells/mm(2) at baseline to 8.8 cells/mm(2) after challenge; P =.002), which also was significantly higher than what was seen in nonasthmatic controls (P =.01). Similarly, IL-4 mRNA--positive cells were increased at 24 hours in patients with asthma (1.4 to 14) in comparison with controls (0 to 0; P =.02). Colocalization studies revealed that 15% and 41% of the basophil population in patients with asthma after allergen-challenge expressed, respectively, IL-4 mRNA and protein. Conversely, 19% of IL-4 mRNA-positive cells and 72% of IL-4 protein--positive cells were accounted for by basophils. CONCLUSION: After allergen provocation in sensitive patients with atopic asthma, basophils are recruited to the bronchial mucosa and express IL-4 mRNA and protein, which might contribute to local IgE synthesis and/or tissue eosinophilia or other aspects of allergic inflammation during late responses and ongoing asthma.     

Antibody-mediated protection against Brucella abortus in BALB/c mice at successive periods after infection: variation between virulent strain 2308 and attenuated vaccine strain 19.

In BALB/c mice antibodies specific for the O polysaccharide (OPS) as well as T lymphocytes mediate protective immunity to Brucella abortus. We performed quantitative analyses of isotypes of OPS antibodies generated during primary infections, and tested the protective qualities of antisera at successive stages of infection against B. abortus strain 2308, representative of the wild type, and attenuated vaccine strain 19. IgM antibodies predominated during the first 3-4 weeks of infection. IgG3 antibodies increased slowly for the first 3 weeks but then rose rapidly and persisted at high levels (> 300 micrograms/ml). IgG1, IgG2a and IgG2b antibodies had increased slightly by week 4 and then remained at low to moderate levels (< 70 micrograms/ml). Week 2 serum pools (IgM high, IgG3 low or undetectable) transferred substantial protection against 2308 (> or = 1 log unit) which increased relatively little (to 1.2-1.5 log units) with later sera that were high in IgG antibodies. In contrast, week 2 sera conferred low levels of protection against 19 (< 0.6 log units), but protection was dramatically increased (to > or = 2.3 log units) with sera obtained 1 week later that had slightly increased IgG antibodies. Monoclonal IgM antibodies also provided better protection against 2308 than 19, while monoclonal IgG3 antibodies protected much better against 19. Strain 19 opsonized with antibodies taken at any stage of infection was killed within normal macrophages, whereas comparably opsonized 2308 underwent intracellular replication. Phagocytosis of 2308 was better than of 19 when brucellae were opsonized with either polyclonal IgM or IgG3 antibodies, and the difference between strains was more extreme following IgM opsonization. The data suggest an explanation for differences in the growth curves of 2308 and 19 in spleens of BALB/c mice. Higher numbers achieved by 19 at week 2 could result from extracellular replication owing to ineffectual opsonization by IgM antibodies, while the precipitous decline of 19 beginning at week 3 could be caused by the increase in more effective IgG3 opsonins that facilitate its rapid intracellular destruction.     

Antigen presentation is a function of all B cell subpopulations separated on the basis of size

Purified splenic B cells from nonimmune mice were separated by counterflow centrifugal elutriation into 6 subpopulations containing cells of discrete sizes ranging from 119 to 200 μm³. B cells of each subpopulation were competent to process and present a native globular protein antigen, cytochrome c, to a cytochrome c-specific T cell hybrid. In all cases, the B cells' antigen-presenting function was radiation sensitive and did not require T cells or T cell products, since B cells fixed with paraformaldehyde effectively presented a carboxyl-terminal peptide fragment of cytochrome c containing the T cell determinant. Furthermore, the antigen-presenting function of B cells of each subpopulation was augmented by treatment with submitogenic doses of the F(ab')₂ fragment of rabbit anti-mouse Ig antibodies, in that 10-30-fold fewer B cells were required and higher maximal T cell responses were achieved, indicating that B cells of all sizes are capable of being regulated in their antigen presentation function through their surface Ig. In addition, B cells of each subpopulation responded to soluble factors present in the supernatants of activated T cells as evidenced by an increase in volume and by the uptake of [³H]thymidine. These results indicate that B cells, regardless of size, are able to participate in at least two essential phases of T cell-dependent antibody responses, initiating the interaction by processing and presenting antigen to helper T cells and responding to soluble helper factors secreted by activated T cells.     

No evidence for chromosomal mosaicism in multiple tissues of 10 patients with 45 XO Turner syndrome

Why the frequency of spontaneous abortions among monosomy X conceptuses is 98 % while the postnatal course of Turner syndrome is relatively benign has not been understood. One explanation could be that mosaicism for a euploid cell line confers viability and that those 2 % of 45, XO zygotes surviving in utero have some degree of mosaicism. We thus reasoned that if the non-mosaic 45, XO karyotype is lethal, a thorough study of living Turner syndrome patients might reveal a much higher frequency of mosaicism than the 30–40 % reported. Ten adult women with a 45 , XO leukocyte karyotype were investigated, looking at five tissue types from all three germ layers: buccal mucosa and hair from ectoderm, urinary epithelium from endoderm and ectoderm, and lymphocytes and skin fibroblasts from mesoderm. We were unable to confirm mosaicism in these patients, although in 2 out of 10 there was the suggestion of a small percentage of euploid cells in skin and blood karyotypes.     

Short-term effects of streptozotocin-induced diabetes on the electrocardiogram, physical activity and body temperature in rats

A variety of contractility defects have been reported in the streptozotocin (STZ)-induced diabetic rat heart including alterations to the amplitude and time course of cardiac muscle contraction. Transmitter devices were surgically implanted in the peritoneal cavity of young adult male Wistar rats. Electrodes from the transmitter were arranged in Einthoven bipolar lead II configuration. Electrocardiogram (ECG), physical activity and body temperature data were continuously recorded with a telemetry system before and following the administration of STZ (60 mg kg-1). Heart rate (HR), physical activity and body temperature declined rapidly 3-5 days after administration of STZ. The effects became more conspicuous with time and reached a new steady state approximately 10 days after STZ treatment when HR was 255+/-8 beats min-1 in diabetic rats compared to 348+/-17 beats min-1 in age-matched controls. Heart rate variability (HRV) was also significantly reduced after STZ treatment (18+/-3 beats min-1) compared to controls (36+/-3 beats min-1). Reduced physical activity and/or body temperature may partly underlie the reduction in HR and HRV. Reductions in power spectral density at higher frequencies (2.5-3.5 Hz) suggest that parasympathetic drive to the heart may be altered during the early stages of STZ-induced diabetes. Short-term diabetes-induced changes in vital signs can be effectively tracked by continuous recording using a telemetry system.     

Nitric oxide mediates intestinal hyperaemic responses to intraluminal bile-oleate

It has long been recognized that intestinal blood flow increases at mealtimes. Mesenteric hyperaemia is also evoked by activation of sensory peptidergic nerves. Our studies explored the possible role of endogenous nitric oxide (NO) in the rat intestinal vasodilator response to luminal instillation of an oleic acid plus bile mixture before and after acute intrajejunal instillation of capsaicin and after chronic pretreatment with capsaicin. In anaesthetized rats we measured jejunal blood flow (BF) with an ultrasonic Doppler flowmeter and systemic arterial pressure (AP) with a pressure transducer. Intestinal perfusion with 80 mM oleic acid in bile increased BF by 98±12%. Instillation of 4 mg of capsaicin into the jejunal lumen initially increased BF by 42±9% but was followed by vasoconstriction. Inhibition of NO synthase with 25 mg/kg i.v. N-nitro-L-arginine (L-NNA) decreased BF by 27±5% and increased AP by 37±11%. After treatment with L-NNA and after acute and chronic administration of capsaicin, the bile-oleate-induced maximal increases in BF above control levels were 42±7%, 65±12%, and 58±8%, respectively. The observed inhibitory effect of L-NNA on the intestinal hyperaemic response to the bile-oleate mixture was reversed by pretreatment with L-arginine (100 mg/kg i.V.). In capsaicin pretreated rats the subsequent bile-oleate-induced hyperaemia was reduced in magnitude but the inhibitory effects of L-NNA were proportionately the same as in animals not receiving capsaicin. These findings support the hypothesis that NO is involved with bile-oleate-induced mesenteric hyperaemia.