Antibody responses against natural Taenia hydatigena infection in dogs in Kenya

Antibody responses (IgG) against Taenia hydatigena infection in dogs in Kenya were analysed in ELISA using excretory/secretory products of T. hydatigena scoleces derived from goat cysticercus cysts. Helminth infections of individual dogs were confirmed at autopsy. T. hydatigena worms were found in 89.5% of 143 dogs, and positive anti-T. hydatigena antibody levels were detected in 58.7% of infected dogs. Positive antiscolex antibody levels were detected in 40.0% of Turkana dogs uninfected with T. hydatigena, suggesting previous infection. Antibody was not detected in 34.4% of infected dogs. There was no relationship between individual T. hydatigena worm burdens and absorbance values for sera in ELISA. It was not possible to distinguish between sera from T. hydatigena-infected and uninfected dogs.     

The structure of human mitochondrial DNA variation

Summary Restriction analysis of mitochondrial DNA (mtDNA) of 3065 humans from 62 geographic samples identified 149 haplotypes and 81 polymorphic sites. These data were used to test several aspects of the evolutionary past of the human species. A dendrogram depicting the genetic relatedness of all haplotypes shows that the native African populations have the greatest diversity and, consistent with evidence from a variety of sources, suggests an African origin for our species. The data also indicate that two individuals drawn, at random from the entire sample will differ at approximately 0.4% of their mtDNA nucleotide sites, which is somewhat higher than previous estimates. Human mtDNA also exhibits more interpopulation heterogeneity (G_ST=0.351±0.025) than does nuclear DNA (G_ST=0.12). Moreover, the virtual absence of intermediate levels of linkage disequilibrium between pairs of sites is consistent with the absence of genetic recombination and places constraints on the rate of mutation. Tests of the selective neutrality of mtDNA variation, including the Ewens-Watterson and Tajima tests, indicate a departure in the direction consistent with purifying selection, but this departure is more likely due to the rapid growth of the human population and the geographic heterogeneity of the variation. The lack of a good fit to neutrality poses problems for the estimation of times of coalescence from human mtDNA data.     

Transformational exchanges in the dihydropteroate synthase gene of Neisseria meningitidis: a novel mechanism for acquisition of sulfonamide resistance.

The nucleotide sequences of the chromosomal dihydropteroate synthase (dhps) genes in sulfonamide-susceptible and sulfonamide-resistant strains of Neisseria meningitidis of serogroups A, B and C were determined. The molecular weights and the amino acid sequences showed similarity to those of all other known dihydropteroate synthase polypeptides. Sequence comparison of the N. meningitidis dhps genes indicated horizontal transfer of DNA segments rather than point mutations as the cause for resistance in meningococci. The dhps genes in three of four sulfonamide-resistant meningococci contained identical central regions of 424 bp. Compared with the corresponding genes in susceptible strains, each central region included an insert of 6 bp. In one of the sulfonamide-resistant strains, the dhps gene was similar to the corresponding genes in the sensitive strains in its NH2-terminal and C-terminal parts. Its central region, however, was identical to the corresponding regions of two of the other resistant genes, and thus it could be seen as a hybrid dhps gene. Transformation experiments and mapping of transformed dhps genes indicated the existence of a novel mechanism for the dissemination of sulfonamide resistance in N. meningitidis. The origin of the resistance-mediating segment of the gene is unknown, but hybridization results showed the presence of homologous dhps genes in Neisseria gonorrhoeae and N. lactamica but not in N. subflava or Branhamella catarrhalis.     

Radiolabeling of DNA can induce its fragmentation in HL-60 human promyelocytic leukemic cells.

Incorporation of radiolabeled thymidine is commonly used to investigate DNA damage. Using a filter-binding assay, we observed that the addition of various doses of [methyl-3H]thymidine (0.2 and 2 microCi/ml) or [2-14C]thymidine (0.02 and 0.2 microCi/ml) in the culture medium for 2 days, a standard method for cell-labeling, induces DNA fragmentation in HL-60 human promyelocytic cells. This effect was dose- and time-dependent and the DNA fragments were not protein-linked since the levels of DNA fragmentation were identical in the presence and in the absence of proteinase K (0.5 mg/ml). Radiolabeled thymidine-induced DNA fragmentation was associated with an inhibition of cell growth, but cells remained able to exclude trypan blue, suggesting that plasma membrane integrity was conserved, except at very high doses of [methyl-3H]thymidine (2 microCi/ml). By agarose-gel electrophoresis, the DNA-fragmentation was demonstrated to be internucleosomal with a typical ladder pattern. Addition of unlabeled thymidine to the culture medium prevented DNA fragmentation in a dose-dependent manner, indicating that radiolabeled thymidine incorporation in DNA was directly responsible for DNA fragmentation. We conclude that radiolabeling of DNA using thymidine incorporation can induce DNA fragmentation in some cell lines such as HL-60. This observation must be taken into account in methods using radiolabeling to study DNA damage in these cells.     

A rapid procedure for the construction of PCR cDNA libraries from small amounts of plant tissue

We describe a general method for the preparation of λZAP II cDNA libraries from very small amounts (<50 mg) of plant tissue. We have achieved this by combining an efficient method for RNA extraction with a modified PCR protocol for the synthesis and amplification of cDNA. Using this protocol we have found it possible to generate cDNA libraries containing more than 10⁶ clones from as little as 1 μg of total RNA.     

R-phycoerythrin from the macroalgaCorallina officinalis (Rhodophyceae) and application of a derived phycofluor probe for detecting sugar-binding sites on cell membranes

R-Phycoerythrin (absorption spectrum 280; 495;sh 535, 564 nm with A564/A280 ratio of 5.3) was purified from the red macroalgaCorallina officinalis. The relative molecular mass determined from PAGE was 240 000. SDS-PAGE demonstrated two major subunits ofM _r 20 000 and 21 000, respectively, and a minor subunit ofM _r 30 000. A fucospyranosyl phenylisothiocyanate conjugate was prepared and this novel fluorescent affinity reagent used in conjunction with a flow cytometer to probe fucose-binding sites on blood mononuclear cells. By varying the sugar and using other phycobiliproteins the approach has the potential for simultaneously monitoring different sugar binding sites on subsets of cells within populations.