The impact of land use and land cover change on biodiversity within and adjacent to Kibasira Swamp in Kilombero Valley,Tanzania

Wetlands are crucial ecosystems with multiple values and functions to a range of different stakeholders. The future of wetlands depends both on the legacy of the past and how they are currently used. Using 48 vegetation survey plots (0.08 ha) combined with Landsat 5 and 7 TM imagery, we assessed the influence of long‐term (1990–2011) land use and land cover change on the biodiversity of the Kibasira Swamp. Information on perceptions of adjacent communities on historical changes and drivers for the changes were also collected. Results showed an increase in the area covered by open water by 1% and forest by 4% between 1990 and 1998 whilst Cyperus papyrus L and cultivated land area decreased by 8% and 3%, respectively on the same period. Between 1998 and 2011, there was a decrease in areas covered by water by 35% and forest by 9% whereas C. papyrus L increased by 40% and cultivated land increased by 8%. These changes have affected the biodiversity of the swamp and adjacent to it as numbers of mammals have declined. However, the Swamp still provides extensive habitat for plants and bird species despite the ongoing human pressure. Interventions may be necessary to maintain biodiversity in Kibasira Swamp to ensure sustainable ecosystem services.     

Antibodies define multiple proteins with epitopes exposed on the surface of live Babesia bigemina merozoites

Babesia bigemina is one of several tick-borne hemoparasitic diseases of cattle that are inadequately controlled and cause substantial livestock production losses in tropical and subtropical climates. Recovery from acute babesiosis is associated with development of protective immunity against subsequent challenge with both homologous and heterologous parasites. Viable and infectious merozoites, the intraerythrocytic stage of B. bigemina responsible for clinical disease, were separated from contaminating host cells by density gradient centrifugation. Monoclonal antibodies developed against gradient-separated merozoites were screened for surface reactivity against live merozoites in an immunofluorescent binding assay. Surface-reactive antibodies immunoprecipitated five major biosynthetically radiolabeled merozoite proteins with relative m.w. of 72,000, 58,000, 55,000, 45,000, and 36,000 in SDS-PAGE. Two additional proteins immunoprecipitated with the 45,000 m.w. protein were unreactive with monoclonal antibody in western blots and are apparently part of a membrane complex co-precipitated by this antibody. In contrast, additional proteins of m.w. of 36,000, 35,000, and 33,000, immunoprecipitated with the 58,000 protein, all contain the surface-exposed epitope bound by monoclonal antibody. Immune serum from an animal that had recovered from infection with a Mexico isolate of B. bigemina immunoprecipitated five radiolabeled proteins from the Mexico isolate that co-migrated in SDS-PAGE with major proteins precipitated by surface-reactive monoclonal antibodies. In addition, antibodies against a Kenya isolate of B. bigemina immunoprecipitated the same co-migrating proteins from radio-labeled Mexico isolate, demonstrating epitope conservation between surface proteins of geographically different isolates. The identification of proteins with epitopes exposed on the surface of live merozoites and accessible to antibody provides candidates to be tested as protective immunogens in cattle.     

Composite autologous-allogeneic skin replacement: development and clinical application

A major unsolved problem in skin restoration in severe burns is replacement of lost dermis. We report the development and clinical application of a composite grafting technique in which allogeneic skin is the source of dermis, and cultured autologous keratinocytes generate epidermis. Excised burn wounds are resurfaced with unmatched allograft. Immunosuppression from the burn and reduced immunoreactivity of the allograft permit successful allograft engraftment. Keratinocyte cultures are initiated from the patient. Allogeneic epidermis is removed, and the dermal bed is resurfaced with keratinocyte cultures. The allogeneic dermis promotes rapid (less than 7 days) stratification, maturation, and integration of the cultures and the synthesis of anchoring fibrils. One case followed 11 months has shown no evidence of rejection. We reason that removal of the epidermis from allograft eliminates the majority of cells constitutively expressing alloclass II antigens, leaving behind a viable allogeneic dermal bed that serves as an ideal substrate for engraftment and integration of keratinocyte cultures but does not initiate rejection.     

RNA structural elements for expression in Escherichia coli. Alpha 1-antitrypsin synthesis using translation control elements based on the cII ribosome-binding site of phage lambda

Analysis of a series of lambda cII::alpha 1-antitrypsin (alpha 1AT) gene fusions of different sizes showed that increased alpha 1AT expression correlated with the stabilisation of a particular computer-predicted RNA secondary structure. Moreover, significant synthesis of unfused alpha 1AT was achieved by reconstruction of this conformation to permit interaction between the upstream region of the ribosome-binding site and the first part of the alpha 1AT coding sequence. This high-level expression was dependent upon certain silent point mutations in the coding sequence, indicating that RNA primary and secondary structure determinants can operate in concert to dictate the efficiency of protein synthesis.     

Latent Nonstructural Differentiation among Homologous Chromosomes at the Diploid Level: Chromosome 6B of AEGILOPS LONGISSIMA

Previous work has shown that chromosome pairing at metaphase I (MI) of wheat homologous chromosomes from different inbred lines (heterohomologous chromosomes) is reduced relative to that between homologous chromosomes within an inbred line (euhomologous chromosomes). In order to determine if a potential for this phenomenon exists in diploid species closely related to the wheat B genome, MI chromosome pairing was investigated between euhomologous and heterohomologous 6B^e (=6S^e) chromosomes, each from a different population of Aegilops longissima Schweinf. et Muschl. (2n = 2x = 14) substituted for chromosome 6B of Chinese Spring wheat (Triticum aestivum L., 2n = 6x = 42). Euhomologous and heterohomologous monotelodisomics, i.e., plants with one complete chromosome 6B^e and a telosome of either 6B^ep or 6B^eq, were constructed in the isogenic background of Chinese Spring. Pairing at MI of the Ae. longissima chromosomes was reduced in heterohomologous monotelodisomics compared to that in the corresponding euhomologous monotelodisomics. The remaining 20 pairs of Chinese Spring chromosomes paired equally well in the euhomologous and heterohomologous monotelodisomics. Thus, the cause of the reduced pairing must reside specifically in the Ae. longissima heterohomologues. In the hybrids between the Ae. longissima lines that contributed the substituted chromosomes, pairing between the heterohomologous chromosomes was normal and did not differ from that of the euhomologous chromosomes. These data provide evidence that a potential for reduced pairing between the heterohomologues is present in the diploid species, but is expressed only in the polyploid wheat genetic background. The reduction in heterohomologous chromosome pairing was greater in the p arm than in the q arm, exactly as in chromosome 6B of wheat. It is concluded that the reduced pairing between Ae. longissima heterohomologues has little to do with constitutive heterochromatin. The value of chromosome pairing as an unequivocal means of determining the origin of genomes in polyploid plants is questioned.     

Identification of a second mutation in the protein-coding sequence of the Z type alpha 1-antitrypsin gene

This study reports the entire nucleotide sequence of the protein coding region sequence of the alpha 1-antitrypsin (alpha 1AT) Z gene, a common form of the alpha 1AT gene associated with serum alpha 1AT deficiency. In addition to Glu342 to Lys342 mutation in exon V which has been previously identified by peptide analysis, another point mutation (GTG to GCG in exon III) in the gene sequence predicts a second amino acid substitution (Val213 to Ala213) in the Z protein. This Val213 to Ala213 mutation was confirmed to be a general finding in Z type alpha 1AT gene by evaluating genomic DNA from 40 Z haplotypes using synthetic oligonucleotide gene probes directed toward the mutated exon III sequences in the Z gene. Furthermore, the exon III Val213 to Ala213 mutation eliminates a BstEII restriction endonuclease site in the alpha 1AT Z gene, allowing rapid identification of this Val213 to Ala213 substitution at the genomic DNA level. Surprisingly, when genomic DNA samples from individuals thought to be homozygous for the M1 gene (the most common alpha 1AT normal haplotype) were evaluated with BstEII, 23% of the M1 haplotypes were BstEII site negative, thus identifying a new form of M1 (i.e. M1(Ala213], likely identical to M1 but with an isoelectric focusing "silent" amino acid substitution (Val213 to Ala213). Although the relative importance of the newly identified exon III Val213 to Ala213 mutation to the pathogenesis of the abnormalities associated with the Z gene is not known, it is likely that M1(Ala213) gene represents a common "normal" polymorphism of the alpha 1AT gene that served as an evolutionary intermediate between the M1(Val213) and Z genes.     

Light microscopic localization of glycosyltransferase activities in cells and tissues

We describe an assay for light microscopic visualization of specific glycosyltransferases on tissue sections or on cells. The assay uses a sequence of enzyme reactions that yields two moles of NADH for each mole of the uridine-5'-diphosphate (UDP) released during transfer of a monosaccharide from a UDP sugar to an acceptor. When diaphorase and tetrazolium salts are present in the incubation mixture, the tetrazolium salts are reduced to colored diformazans, which precipitate at the sites of glycosyltransferase activity. The validity of the assay was established by applying the technique to spermatozoa and liver, in which some glycosyltransferases have previously been localized. When suspensions of mouse spermatozoa were assayed for galactosyltransferase (GalTase) activity, diformazan precipitates appeared on the plasma membranes overlying the anterior heads of the spermatozoa, in agreement with immunochemical localizations. In mouse liver slices assayed with bilirubin as acceptor for glucuronyltransferase (GluTase) activity, dense diformazan deposits appeared on the hepatocytes but not on endothelial cells, also in agreement with immunochemical data. In the absence of acceptor or UDP sugar donor, diformazan deposits were minimal and random in all tissues tested. The assay's versatility was tested by incubating tissues with different sugar donors and acceptors to localize other sites of transferase activity. In mouse frozen liver sections, GalTase activity occurred in both hepatocytes and endothelial cells; in sections of rat submaxillary glands, GalTase activity was detected in mast cells. In liver sections, GlcuTase activity with o-aminophenol as acceptor was located primarily on the endothelial cells. With the appropriate sugar donor and acceptor, this assay should detect any transferase, other than the glucosyltransferases, that utilizes UDP sugars.     

Point mutations modifying the thrombin inhibition kinetics and antithrombotic activity in vivo of recombinant hirudin

The hirudin variant HV2 was modified by in vitro site-specific mutagenesis of HV2 cDNA to generate HV2(Asn-47----Lys), HV2(Asn-47----Arg) and HV2(Lys-35----Thr, Asn-47----Lys). Residues 35 and 47 are positioned respectively within the finger and prothrombin-like domains of hirudin, both of which have been suggested as thrombin binding sites. The modified polypeptides were synthesized in Saccharomyces cerevisiae using a secretion vector and purified from culture supernatants. By analysis of the human alpha-thrombin:hirudin inhibition reaction in steady-state conditions it was shown that the dissociation constants for HV2(Lys-47) and HV2(Arg-47) were 5- to 14-fold lower than for unmodified HV2, whereas mutation of Lys-35 did not significantly alter the inhibition kinetics. Furthermore, HV2(Lys-47), whose sequence is identical to a natural hirudin variant, displayed enhanced anti-thrombotic activity in vivo, having a 100-fold lower ED50 compared to HV2 in the rabbit Wessler venous thrombosis model. These results support a role for the prothrombin-like domain in thrombin binding and, moreover, demonstrate that in vivo antithrombotic efficiency correlates with the dissociation constant of the inhibition reaction.     

Degradation of underlying extracellular matrix by sensory neurons during neurite outgrowth

The ability of differentiating sensory neurons to remodel a fibronectin substratum was examined. During the early stages of neurite outgrowth, fibronectin was cleared from areas beneath the neuronal soma and processes. The removal of fibronectin occurred in the presence and absence of plasminogen and was associated with the release of fibronectin fragments into the culture medium. The degradation of fibronectin was dependent upon neuronal contact with the substratum. Extraction of cells with the nonionic detergent Triton X-114 identified plasminogen activator and plasmin associated with the cell surface. These findings suggest that the plasminogen activator/plasmin system may play an important role in the interaction of differentiating sensory neurons with the extracellular matrix during axonal outgrowth.