Three new species of penicillium

Three new species of microfungi belonging to the genus Penicillium Link ex Fries are described and illustrated. All but one have been isolated from the atmosphere of las Palmas, capital city of the island of Gran Canaria (Canary Islands, Spain). They clearly differ from all species of the genus described so far and are, therefore, described and proposed as new species: Penicillium hispanicum sp. nov., Penicillium grancanariae sp. nov., and Penicillium palmensis sp. nov.     

Bullous pemphigoid antigen. II. Isolation from the urine of a patient.

This report concerns the isolation of bullous pemphigoid antigen from the nondialyzable urinary components of a patient with the disease. The isolation was accomplished by ion exchange chromatography and gel filtration. Pemphigoid antigen was found to be a basic glycoprotein that on SDS gel electrophoresis showed two major bands, one in the 18,000 m. w. region and the second with a m. w. of 74,000. Between these two bands, two additional bands appeared; one of 35,000 daltons and the other of 68,000 daltons. The 18,000 m. w. band was eluted from the gel and rerun on SDS gels. These gels showed the 18,000 m.w. band and also the appearance of the 35,000 and 74,000 m.w. bands. This finding indicates that urinary pemphigoid antigen may exist both as a single monomeric form and in polymeric aggregates.     

Attachment of Trypanosoma cruzi trypomastigotes to receptors at restricted cell surface domains

We have used glutaraldehyde-fixed target cells to study the attachment phase of cell invasion by live trypomastigotes of Trypanosoma cruzi, and determined that attachment is polarized and receptor-mediated. T. cruzi trypomastigotes bind much less efficiently to confluent epithelial cells, which are polarized, than to sparse epithelial cells. When the tight junctions of confluent epithelial cells are disrupted by removing Ca2+ from the incubation medium before glutaraldehyde fixation, binding of T. cruzi increases. T. cruzi also shows preference for attachment underneath cells or to the edges of cells. The binding occurs within a few minutes, is saturable, and is influenced by the parasite developmental stage. Fab fragment derived from monoclonal antibodies that immunoprecipitate a 160-kDa molecule present only on the surface of trypomastigotes inhibit adhesion to fixed and live cells. Future characterization of the target cell receptors for this molecule and the use of fixed target cells should facilitate studies of the mechanisms involved in the initial interaction of T. cruzi with its host cells.     

Streptococcus pneumoniae DNA polymerase I lacks 3''-to-5'' exonuclease activity: localization of the 5''-to-3'' exonucleolytic domain.

The Streptococcus pneumoniae polA gene was altered at various positions by deletions and insertions. The polypeptides encoded by these mutant polA genes were identified in S. pneumoniae. Three of them were enzymatically active. One was a fused protein containing the first 11 amino acid residues of gene 10 from coliphage T7 and the carboxyl-terminal two-thirds of pneumococcal DNA polymerase I; it possessed only polymerase activity. The other two enzymatically active proteins, which contained 620 and 351 amino acid residues from the amino terminus, respectively, lacked polymerase activity and showed only exonuclease activity. These two polymerase-deficient proteins and the wild-type protein were hyperproduced in Escherichia coli and purified. In contrast to the DNA polymerase I of Escherichia coli but similar to the corresponding enzyme of Thermus aquaticus, the pneumococcal enzyme appeared to lack 3'-to-5' exonuclease activity. The 5'-to-3' exonuclease domain was located in the amino-terminal region of the wild-type pneumococcal protein. This exonuclease activity excised deoxyribonucleoside 5'-monophosphate from both double- and single-stranded DNAs. It degraded oligonucleotide substrates to a decameric final product.     

Time-resolved fluorescence study of human recombinant interferon alpha 2. Association state of the protein, spatial proximity of the two tryptophan residues.

Human recombinant interferon alpha 2 belongs a to family of proteins active against a wide range of viruses. It contains two tryptophan residues located at positions 77 and 141 in the peptide sequence. The fluorescence emission spectrum of these tryptophan residues displays a maximum at 335 nm. The fluorescence intensity decay is described by one broad excited-state-lifetime population centered around a value of 1.7 ns (full width at half maximum, 1.5 ns). These observations suggest that in the native protein, both tryptophan residues emit from similar environments, not directly exposed to the surrounding solvent. The anisotropy decay is essentially biexponential. The correlation-time value characterizing the Brownian rotation of the protein varies linearly with the viscosity/temperature ratio. The calculated hydrodynamic volumes are compatible with the existence of a dimer and a tetramer, at pH 5.5 and 9.4, respectively. Addition of urea at pH 5.5 disrupts the dimer and modifies to some extent the excited-state-lifetime distribution which becomes more heterogeneous. Disulfide-bond reduction also dissociates the dimer and leads to a highly heterogeneous fluorescence-intensity decay with four excited-state-lifetime populations. An opening of the local structure in the Trp region of the protein is likely to occur in these conditions. The fast-anisotropy-decay components can be due to either fast rotation or energy transfer between the indoles. Close proximity of the two Trp residues (less than 1 nm) is suggested from steady-state and time-resolved fluorescence-anisotropy measurements in vitrified medium [95% (by mass) glycerol at -38 degrees C]. This suggestion is in agreement with the recently published three-dimensional structure of the homologous protein murine interferon beta [Senda, T., Shimazu, T., Matsuda, S. Kawano, G., Shimizu, H., Nakamura, K. T. & Mitsui, Y. (1992) EMBO J. 11, 3193-3201].     

Mechanisms towards compensation of long-term haemopoietic injury in mice after 5 Gy irradiation:In vivo andin vitro enhancement of superoxide anion production by granulocytes

This paper analyzes the long-term (6 and 12 months) function of mouse granulocytes after total body irradiation with a single dose (5 Gy) of X-rays. Superoxide anion production has been investigated in granulocytes from peripheral blood, and also in those harvested from long term bone marrow cultures, with the aim of correlating the environmental damage induced by radiation with the functional properties of granulocytes. Anin vivo andin vitro enhancement of superoxide anion production and protein levels in granulocytes from irradiated mice is described. The presence of some colony stimulating factor in the supernatant of cultures from irradiated mice could play an important role in the priming of granulocytes.     

Alteration of the Magnetic Properties of Aquaspirillum magnetotacticum by a Pulse Magnetization Technique

The presence of a narrow shape and size distribution for magnetite crystals within magnetotactic organisms suggests strongly that there are species-specific mechanisms that control the process of biomineralization. In order to explore the extent of this control, cultures of Aquaspirillum magnetotacticum in the exponential growth phase were exposed to increasing magnetic pulses with the aim of separating cell populations on the basis of their magnetic coercivities. Isothermal remanent magnetization and anhysteretic remanent magnetization studies were performed with freeze-dried magnetic cells after the remagnetization treatment. Subpopulations of A. magnetotacticum that showed an increase in coercivity correlated with the intensity of the magnetic pulses were isolated. After successive subcultures of the remaining north-seeking cells, a maximum bulk coercivity (H_b^max) of 40 mT was obtained after treatment with a 55-mT pulse. Although we obtained A. magnetotacticum variants displaying higher coercivities than the wild-type strain, changes in crystal size or shape of the magnetite crystals were below reliable detection limits with transmission electron microscopy. Attempts to shift the coercivity towards higher values caused it to decrease, a change which was accompanied by an increase in magnetostatic interactions of the magnetosome chains as well as an increase in the cell population displaying an abnormal distribution of the magnetosome chains. Ultrastructural analyses of cells and magnetosomes revealed the appearance of cystlike bodies which occasionally contained magnetosomes. The increase in cystlike cells and abnormal magnetosome chains when higher magnetic pulses were used suggested that magnetosomes were collapsing because of stronger interparticle magnetostatic forces.     

Naturally occurring benzodiazepines in human milk

The presence of benzodiazepine-like molecules was detected radioimmunologically in the plasma and milk of 12 women and in the plasma of 9 men. All subjects were non-users of benzodiazepines. The concentration of these biological materials expressed as diazepam equivalents per mL amounted to 2.54 +/- 0.74 ng in male plasma; to 2.20 +/- 0.35 ng in female plasma and to 1.91 +/- 0.54 ng in milk. Further investigation of the active compounds in milk permitted the unequivocal identification of diazepam, both free and bound to a presumably protein carrier and, at least, three more benzodiazepine-like molecules. Their origin either from dietary sources or as a result of endogenous biosynthesis is still unclear.     

Biological characterization of Trypanosoma cruzi zymodemes: in vitro differentiation of epimastigotes and infectivity of culture metacyclic trypomastigotes to mice

Thirty-one Trypanosoma cruzi isolates from Chile, Peru, and Bolivia were studied in their capacity to differentiate in vitro from epimastigotes to metacyclic trypomastigotes on TAU-3AAG medium. Zymodeme 1 parasites displayed the best level of differentiation, which ranges from 60 to 90% depending on the isolate. Zymodeme 2 parasites exhibited highly heterogenous differentiation rates. This differentiation method permits the obtention of large amounts of metacyclic trypomastigotes from zymodeme 1 parasites. Metacyclic trypomastigotes obtained in vitro were infective to nude Balb/c hybrid mice. Zymodeme 1 parasites produced high parasitemias in this murine model; in contrast, zymodeme 2 parasites displayed lower parasitemias. Of a total of 27 T. cruzi isolates, 20 proved to be infective to mice, 12 gave enough parasites for further studies, and 8 of these were used for biological characterization. Results are compared with the infective clone Dm28 and Tulahuén strains maintained since 1954 in mice.