Ultrastructural aspects of the hyphal tip ofSclerotium rolfsii preserved by freeze substitution

Summary The hyphal tip ofSclerotium rolfsii was examined after fixation by freeze substitution. The Spitzenkörper consisted of a dense mass of apical vesicles and microvesicles surrounding a vesicle-free zone. Linear arrangements of microvesicles were occasionally observed within the Spitzenkörper. Abundant microfilaments were seen within the Spitzenkörper region, often in close association with apical vesicles and microvesicles. Microtubules passed through the Spitzenkörper and terminated at the plasmalemma at the extreme hyphal apex. Filasomes were mostly observed within the apical region and were in close proximity to the plasmalemma. Rough ER, mitochondria, microtubules, and vacuoles were abundant in the subapical region and were usually oriented parallel to the long axis of the hypha. Ribosomes were aligned on the outer surfaces of mitochondria. Golgi body equivalents were observed throughout the subapical region and appeared as inflated cisternae of varying shapes and electron opacities. Relationships to other basidiomycetous hyphal tip cells are discussed.Abbreviations AV apical vesicle - C Celsius - diam diameter - f filasome - G Golgi body equivalent - h hour - nm nanometer - M mitochondria - ME membranous elements; min minute - MV microvesicle - MVB multivesicular body - N nucleus - OsO₄ osmium tetroxide - R ribosome - ER endoplasmic reticulum - S Spitzenkörper - Va vacuole - m micrometer     

Prenatal and lactational transmission of Toxocara canis and Ancylostoma caninum: experimental infection of the bitch before pregnancy

Breeding of five parasite-free and five experimentally infected (6000 Toxocara canis eggs orally and 2500 Ancylostoma caninum larvae subcutaneously) beagle bitches was done so that pairs of bitches (1 uninfected, 1 infected) whelped simultaneously. Pups born to an infected bitch were removed at birth and nursed by the paired uninfected bitch until 4 weeks of age when pups were necropsied to determine the number of parasites they had acquired prenatally from their infected mother. Pups born to the parasite-free bitch were nursed by the infected bitch until necropsied at 4 weeks of age to determine the number of parasites passed via the lactational route. Of 680 ascarids transmitted to pups by either route, 98.5% were transmitted prenatally and 1.5% lactationally. Transmission of 2746 hookworms to 22 pups occurred solely by the lactational route; prenatal transmission of this parasite did not occur in any of the 25 pups born to infected bitches.     

Xenopus laevis lectin is localized at several sites in Xenopus oocytes, eggs, and embryos

The endogenous lectin of Xenopus laevis oocytes, unfertilized eggs, and blastula-stage embryos was immunohistochemically localized using a highly specific antiserum. Each tissue was examined with several techniques, including paraformaldehyde or glutaraldehyde fixation, frozen or plastic sections, and immunofluorescence or immunoperoxidase staining. In oocytes and unfertilized eggs, lectin was detected in association with yolk platelets, cortical granules, and the vitelline envelope. In embryos, cortical granules had disappeared and lectin was found in the cleavage furrows between the embryonic cells. The distribution of the lectin suggests that it plays more than one role in this developing system.     

Reversal of Fv-1 host range by in vitro restriction endonuclease fragment exchange between molecular clones of N-tropic and B-tropic murine leukemia virus genomes.

We molecularly cloned unintegrated viral DNA of the BALB/c endogenous N-tropic and B-tropic murine leukemia retroviruses and in vitro passaged N-tropic Gross (passage A) murine leukemia retroviruses. Recombinant genomes were constructed in vitro by exchanging homologous restriction enzyme fragments from N- or B-tropic parents and subsequent recloning. Infectious virus was recovered after transfection of these recombinant genomes into NIH-3T3 cells and cocultivation with the Fv-1 nonrestrictive SC-1 cells. XC plaque assays of recombinant virus progeny on Fv-ln and Fv-lb cells indicated that the Fv-l host range was determined by sequences located between the BamHI site in the p30 region of the gag gene (1.6 kilobase pairs from the left end of the map) and the HindIII site located in the pol gene (2.9 kilobase pairs from the left end of the map).     

Serological response of non-human primates to human melanoma disialoganglioside GD3

Summary The immunogenicity of the disialoganglioside, GD3, a melanoma-tumor-associated antigen, has been evaluated in non-human primates. Sera from four chimpanzees and two monkeys were evaluated for anti-GD3 antibody activity by solid-phase radioimmunoassay using GD3 and control gangliosides as targets. Serum from one monkey, immunized with cells from a melanoma cell line, was strongly reactive with GD3, having a titer of >2500. In contrast, serum from this animal was non-reactive with several other gangliosides including the structurally similar GM3. Anti-GD3 reactivity was also demonstrable, albeit in low titer, in the sera of an additional monkey and a chimpanzee. Each of these animals had likewise been immunized using cells from melanoma cell lines. On the basis of these observations, suggestive of a primate anti-GD3 antibody response, we initiated a series of immunizations of chimpanzee using purified GD3 bound to Salmonella minnesota, R595. IgG reactive with melanoma cells in the cell-binding assay was first detected in sera collected after 4 immunizations and increased in titer against each reactive melanoma cell line during the immunizations. Reactivity of this serum with melanoma cell lines demonstrated a direct correlation with the expression of GD3 by the respective cell line. Anti-GD3 reactivity was evident in solid-phase radioimmunoassay against purified GD3 beginning with serum collected after 11 immunizations. By comparison with its binding to the control ganglioside panel, this serum demonstrated strong specificity for GD3 (titer=640) while having only marginal reactivity with GM3 (titer=40). Immune serum from this animal was also able specifically to block subsequent binding of a murine IgM anti-GD3 antibody (DMab7) to target GD3 in solid-phase radioimmunoassay. Together, these observations suggest that GD3, in the form of a purified molecule bound to a bacterial matrix or as part of the intact melanoma cell membrane, can be immunogenic in non-human primates, and is able to elicit an antibody response of appropriate specificity.Supported in part by grant CA32672 from the National Cancer Institute, Veterans Administration Program 821 and by the Yerkes Regional Primate Center, Atlanta, Georgia. The Yerkes Center is fully accredited by the American Association for Accreditation of Laboratory Animal Care     

Tracing paternal ancestry in mice, using the Y-linked, sex-determining locus, Sry

The molecular evolution of mammalian Y-linked DNA sequences is of special interest because of their unique mode of inheritance: most Y- linked sequences are clonally inherited from father to son. Here we investigate the use of Y-linked sequences for phylogenetic inference. We describe a comparative analysis of a 515-bp region from the male sex- determining locus, Sry, in 22 murine rodents (subfamily Murinae, family Muridae), including representatives from nine species of Mus, and from two additional murine genera--Mastomys and Hylomyscus. Percent sequence divergence was < 0.01% for comparisons between populations within a species and was 0.19%-8.16% for comparisons between species. Our phylogenetic analysis of 12 murine taxa resulted in a single most- parsimonius tree that is highly concordant with phylogenies based on mitochondrial DNA and allozymes. A total evidence tree based on the combined data from Sry, mitochondrial DNA, and allozymes supports (1) the monophyly of the subgenus Mus, (2) its division into a Palearctic group (M. musculus, M. domesticus, M. spicilegus, M. Macedonicus, and M. spretus) and an Oriental group (M. cookii++, M. cervicolor, and M. caroli), and (3) sister-group relationships between M. spicilegus and M. macedonicus and between M. cookii and M. cervicolor. We argue that Y- chromosome DNA sequences represent a valuable new source of characters for phylogenetic inference.      

New chromosomal translocations correlate with specific immunophenotypes of childhood acute lymphoblastic leukemia

Cytogenetic analysis of leukemic cells obtained at diagnosis from 122 patients with childhood acute lymphoblastic leukemia (ALL) disclosed chromosomal translocations in 36 cases. Two new nonrandom translocations were identified and found to be associated with specific immunophenotypes of the disease. The first, identified in 4 of 16 cases of T-cell ALL positive for sheep erythrocyte receptors (E+), involved the short arm (p) of chromosome 11 and the long arm (q) of chromosome 14 and was designated t(11;14) (p13;q13). The second, found in 7 of 23 cases with a pre-B-cell phenotype, involved the long arm of chromosome 1 and the short arm of chromosome 19; it was designated t(1;19) (q23;p13.3). A third abnormality involving a common breakpoint on chromosome 12 (band p 12) was also identified. These two new differentiation-specific translocations suggest a mechanism for aberrant expression of genes that influence lymphoid cell growth and development, as well as leukemogenesis.     

Active and passive cation transport and L antigen hertogeneity in low potassium sheep red cells

Several lines of experimental evidence are presented suggesting that the L antigens in low potassium (LK) sheep red cells are associated with separate Na(+)K(+) pump flux is distinct from the action of anti-L(l) on K(+) leak flux, implying that K(+) leak transport sites may not be converted into active pumps by the L antiserum. Treatment of LK red cells with trypsin completely abolished both the stimulation of K(+) pump flux and the enhancement of the rate of ouabain binding brought about by anti- L. That this effect is due to a total destruction of the L(p) determinant associated with the LK pump was evident from the complete failure of anti-L(p) to bind to trypsinized LK red cells. The L(p) antigen can be effectively protected against the trypsin attack by prior incubation with anti-L, indicating that the sites for antibody binding and trypsin action may be closely adjacent at the structural level. Trypsin treatment, however, did not interfere with anti-L(l) reducing ouabain insensitive K(+) leak influx, nor did it prevent binding of anti-L(ly), the hemolytically active L antibody which is probably identical with anti-L(l). The functional independence of the L(p) and L(l) sites was documented by the observation that anti-L(l) still reduced K(+) leak influx in LK cells with experimentally induced high potassium concentrations, at which K(+) pump flux is fully suppressed, whether or not anti-L(p) was binding to the L(p) antigen associated with the LK pump.     

Caprine Arthritis-Encephalitis Virus Is Distinct from Visna and Progressive Pneumonia Viruses as Measured by Genome Sequence Homology

Competitive inhibition of hybridization between ¹²⁵I-labeled caprine arthritis-encephalitis viral RNA and homologous cDNA by heterologous viral RNA shows that the caprine retrovirus shares <20% genome sequence homology with visna and progressive pneumonia viruses. These viruses, however, are indistinguishable in immunodiffusion reactions involving the major structural protein (p28).     

Heparin-inhibitable lectins: Marked similarities in chicken and rat

Extracts of young rat lung contain a heparin-inhibitable lectin that closely resembles one recently purified from chicken liver. Both lectins interact with heparin and N-acetyl-D-galactosamine, and were purified by gel filtration on Sepharose CL-2B followed by affinity chromatography on heparin-Sepharose. They both behave as high molecular weight aggregates that can be dissociated into two peptides with apparent molecular weights of 13,000 and 16,000 by gel electrophoresis in SDS. Samples of purified lectin contained up to 20% DNA by weight, and the degree of lectin aggregation and hemagglutination activity was greatly reduced by treatment with micrococcal nuclease without inhibiting heparin-binding activity. Association of lectin with DNA is an artifact of homogenization in high salt, since only 2% of the lectin is found associated with a purified nuclear fraction.