The complete cDNA and polypeptide sequences of human erythroid alpha-spectrin.

Overlapping human erythroid alpha-spectrin cDNA clones were isolated from lambda gt11 libraries constructed from cDNAs of human fetal liver and erythroid bone marrow. The composite 8001-base pair (bp) cDNA nucleotide sequence contains 187-bp 5'- and 528-bp 3'-untranslated regions and has a single long open reading frame of 7287 bp that encodes a polypeptide of 2429 residues. As previously described (Speicher, D. W., and Marchesi, V. T. (1984) Nature 311, 177-180), spectrin is composed largely of homologous 106-amino acid repeat units. From the amino acid sequence deduced from the cDNA, alpha-spectrin can be divided into 22 segments. Segments 1-9 and 12-19 are homologous and can therefore be considered repeats; the average number of identical residues in pairwise comparisons of these repeats is 22 out of 106, or 21%. Of these 17 repeats, 11 are exactly 106 amino acids in length, whereas five others differ from this length by a single residue. Segments 11, 20, and 21, although less homologous, appear to be related to the more highly conserved repeat units. The very N-terminal 22 residues, segment 10, which is atypical both in length and sequence, and the C-terminal 150 residues in segment 22 appear to be unrelated to the conserved repeat units. The sequence of the erythroid alpha-spectrin polypeptide chain is compared to that of human alpha-fodrin and chicken alpha-actinin to which it is related. alpha-Spectrin is more distantly related to dystrophin.     

Rat-atouille: A Mixed Method Study to Characterize Rodent Hunting and Consumption in the Context of Lassa Fever

Lassa fever is a zoonotic hemorrhagic illness predominant in areas across Nigeria, Sierra Leone, Guinea, Liberia, and southern Mali. The reservoir of Lassa virus is the multimammate mouse (Mastomys natalensis), a highly commensal species in West Africa. Primary transmission to humans occurs through direct or indirect contact with rodent body fluids such as urine, feces, saliva, or blood. Our research draws together qualitative and quantitative methods to provide a fuller and more nuanced perspective on these varied points of human–animal contact. In this article, we focus on the hunting, preparation, and consumption of rodents as possible routes of exposure in Bo, Sierra Leone. We found that the consumption of rodents, including the reservoir species, is widespread and does not neatly tally against generational or gender lines. Further, we found that the reasons for rodent consumption are multifactorial, including taste preferences, food security, and opportunistic behavior. We argue that on certain topics, such as rodent consumption, establishing trust with communities, and using qualitative research methods, is key to investigate sensitive issues and situate them in their wider context. To conclude, we recommend ways to refine sensitization campaigns to account for these socio-cultural contexts.     

Three novel mtDNA restriction site polymorphisms allow exploration of population affinities of African Americans

To develop informative tools for the study of population affinities in African Americans, we sequenced the hypervariable segments I and II (HVS I and HVS II) of mitochondrial DNA (mtDNA) from 96 Sierra Leoneans; European Americans; rural, Gullah-speaking African Americans; urban African Americans living in Charleston, South Carolina; and Jamaicans. We identified single nucleotide polymorphisms (SNPs) exhibiting ethnic affinities, and developed restriction endonuclease tools to screen these SNPs. Here we show that three HVS restriction site polymorphisms (RSPs), EcoRV, FokI, and MfeI, exhibit appreciable differences in frequency (average delta = 0.4165) between putative African American parental populations (i.e., extant Africans living in Sierra Leone and European Americans). Estimates of European American mtDNA admixture, calculated from haplotypes composed of these three novel RSPs, show a cline of increasing admixture from Gullah-speaking African American (m = 0.0300) to urban Charleston African American (m = 0.0689) to West Coast African American (m = 0.1769) populations. This haplotype admixture in the Gullahs is the lowest recorded to date among African Americans, consistent with previous studies using autosomal markers. These RSPs may become valuable new tools in the study of ancestral affinities and admixture dynamics of African Americans.     

Structure, function, and motility of vacuoles in filamentous fungi

Current information on the structure and function of motile tubular vacuoles in Pisolithus tinctorius and other fungi is reviewed. The use of fluorochromes to label the vacuole lumen is evaluated and observations on the structure and motility of vacuoles in P. tinctorius are differentiated from possible artifacts. The styryl dyes FM4-64 and MDY-64, used in yeast to demonstrate endocytosis, show little or no labeling of internal membranes in undamaged P. tinctorius cells. This agrees with our data showing that other probes for endocytosis such as Lucifer yellow CH are not taken up by hyphal tip cells. Overall, the observations do not support endocytosis in hyphal tips. It has been suggested that tubular vacuole systems carry out longitudinal transport, and evidence in favor of this hypothesis is evaluated. New data are presented to show that many of the large vacuoles in subapical cells are attached to the plasma membrane and are relatively immobile, while video sequences show movement of fluorochrome in pulses along a series of several large vacuoles, all interconnected via tubules. Tubular vacuoles from thick sections of hyphae processed under anhydrous conditions are shown by X-ray microanalysis to contain relatively high levels of P and K, as seen previously in the larger vacuoles. These results provide further evidence for a role of the tubular vacuoles in longitudinal transport of P. Copyright 1998 Academic Press.     

Rapid and sensitive GC/MS characterization of glycolipid released Galalpha1,3Gal-terminated oligosaccharides from small organ specimens of a single pig

Pig to human xenotransplantation is considered a possible solution to the prevailing chronic lack of human donor organs for allotransplantation. The Galalpha1,3Gal determinant is the major porcine xenogeneic epitope causing hyperacute rejection following human antibody binding and complement activation. In order to characterize the tissue distribution of Galalpha1,3Gal-containing and blood group- type glycosphingolipids in pig, acid and nonacid glycosphingolipids were isolated from the kidney, small intestine, spleen, salivary gland, liver, and heart of a single pig obtained from a semi-inbred strain homozygous at the SLA locus. Glycolipids were analyzed by thin-layer immunostaining using monoclonal antibodies, and following ceramide glycanase cleavage as permethylated oligosaccharides by gas chromatography, gas chromatography-mass spectrometry, and matrix- assisted laser desorption/ionization mass spectrometry. The kidney contained large amounts of Galalpha1,3Gal-containing penta- and hexasaccharides having carbohydrate sequences consistent with the Galalpha1,3nLc4and Galalpha1,3Lexstructures, respectively. The former structure was tentatively identified in all organs by GC/MS. The presence of extended Galalpha1,3Gal-terminated structures in the kidney and heart was suggested by antibody binding, and GC/MS indicated the presence of a Galalpha1,3nLc6structure in the heart. The kidney, spleen, and heart contained blood group H pentaglycosylceramides based on type 1 (H-5-1) and type 2 (H-5-2) chains, and H hexaglycosylceramides based on the type 4 chain (H-6-4). In the intestine H-5-1 and H-6-4 were expressed, in the salivary gland H-5-1 and H-5-2, whereas only the H-5-1 structure was identified in the liver. Blood group A structures were identified in the salivary gland and the heart by antibody binding and GC/MS, indicating an organ- specific expression of blood group AH antigens in the pig.      

Detection and quantification of Lyme spirochetes using sensitive and specific molecular beacon probes

Background  

Lyme disease, caused by Borrelia burgdorferi, affects a large number of people in both the USA and Europe. The mouse is a natural host for this spirochete and is widely used as a model system to study Lyme pathogenesis mechanisms. Since disease manifestations often depend upon the spirochete burden in a particular tissue, it is critical to accurately measure the bacterial number in infected tissues. The current methods either lack sensitivity and specificity (SYBR Green), or require independent analysis of samples in parallel to quantitate host and bacterial DNA (TaqMan). We have developed a novel molecular beacon-based convenient multiplex real-time quantitative PCR assay to identify and detect small numbers of B. burgdorferi in infected mouse tissues.