The invasive behaviour of shape Cryptococcus neoformans: a possibility of direct access to the central nervous system?

Cryptococcocal meningoencephalitis is always considered secondary to initial lung infection. Because of the unquestionable evidence of haematogenious spread from the lungs, few publications have reported about other possible primary sites of infection or other routes to the central nervous system. This study was designed to investigate the infiltrative pattern of C. neoformans in immunocompromised mice by treatment with dexamethasone. The infection was performed by nasal instillation (30 mice) or injection into the retro-orbital space (12 mice). From the group infected intranasally, 3 mice presented diffuse invasive fungal colonisation of the mucosa and submucosa. The histologic findings showed infiltrative growth along the periosteum, sometimes surrounded nervous endings of submucosa, invasion along the olfactory nerve and simultaneous meningeal involvement in 2 mice on the 6th and 8th day of infection. All mice infected into the retro-ocular space developed lesions containing numerous cryptococci in the local of the inoculum. Out of the main lesion we observed preferential growth along the perineural spaces with adherence to the perineurium, perivascular spaces and sometimes along aponeurosis. Simultaneous invasion of trigeminal ganglio and trigeminal branches was observed in 4 mice. These morphologic patterns suggest the hypothesis of direct infiltrative invasion of the central nervous system. This revised version was published online in June 2006 with corrections to the Cover Date.     

Modulation of nuclear rotation in neuronal interphase nuclei by nerve growth factor, by gamma-aminobutyric acid, and by changes in intracellular calcium

Nuclear rotation (NR) is typically measured as motion of nucleoli within nuclei of cells in vitro. This occurs in cycling cells. However, its observation in neurons arrested in interphase indicates that mechanisms related to mitosis are not a prerequisite. We have recently shown that NR occurs in three dimensions within the nuclear space, that it occurs within the space delineated by the outer nuclear membrane and that it includes chromatin domains in addition to nucleoli and have postulated that this motion of chromatin domains is related to changes in gene expression. We now show that exposure of dorsal root, sensory neurons in vitro to nerve growth factor (NGF) or to gamma-aminobutyric acid (GABA), agents which alter gene expression, and to agents causing redistribution of calcium, such as EGTA and the calcium ionophore A23187, significantly alters NR. The NGF increased the mean rate of NR and did so at a time after exposure when activity of RNA polymerases have been shown to rise. Exposure to GABA resulted, within minutes, in shifts of the nucleolus within the three-dimensional space of the nucleus, associated in some neurons with significant, sigmoidal increases in the rate of NR. The calcium ionophore A23187 as well as chelation of extracellular calcium with EGTA similarly increased rates. Importantly, excess calcium, with EGTA remaining present, returned NR of all nucleoli to rates not different from controls. This indicates that the increase in NR seen with EGTA is specific to the chelation of calcium and not an nonspecific response to EGTA. It is difficult to link the action of agents which alter gene expression or transmembrane ion balance with changes in NR. Nevertheless, in support of our hypothesis, the results presented here show that agents known to alter gene expression, alter NR in a temporally coincident manner and that they do so, possibly, by calcium-dependent mechanisms.     

Neuron culture from adult goldfish

Neurons and gla from the central nervous system of the adult teleost Carassius auratus have been grown as explant cultures of minced brain tissue and as trypsin dissociated cells. These cultures exhibit extensive neurite growth from two neuronal types, have organotypic ultrastructure, and contain electrically active cells. Autoradiographic data indicate that these neurons do not divide in culture, and histological evidence suggests that some mature neurons survive explantation and regenerate processes. However, explantation of brain fragments not containing undifferentiated cells, localized in the ventricular and subventricular zones in the brains of fish, resulted in mesenchymal and glial cell cultures only. Therefore, a contribution to the population of cells in culture by undifferentiated cells must be considered. The cultured neurons remained viable for at least 19 weeks and ultrastructural and electrophysiological data indicate synaptic interaction between cells in explant cultures.     

Species-specific probe, based on 18S rDNA sequence, could be used for identification of the mucilage producer microalga Gonyaulax fragilis (Dinophyta)

In the Adriatic Sea, the correlation between mucilage phenomena and the presence of Gonyaulax fragilis (Schütt) Kofoid (Dinophyta) has been recently demonstrated. The application of PCR-based methods and the development of species-specific molecular probes might represent powerful technologies for rapid and specific monitoring of microalgal species in seawater samples. Here, we report sequencing of the small subunit (SSU) ribosomal RNA gene (18S rDNA) of G. fragilis and its comparative analysis within the Dinophyta. Total DNAs were extracted and amplified from cultured cells of G. fragilis, which were isolated from natural phytoplanktonic association in the northern Adriatic Sea. Total 18S rDNA gene was amplified using 16S1N and 16S2N primers and sequenced using ad hoc designed internal primers. The primers amplified a product of expected size (length 1700/1800 bp). The phylogenetic analysis carried out by comparing G. fragilis sequence to homologous sequences of Lingulodinium polyedrum (Stein) Dodge, Gonyaulax spinifera (Claparède et Lachmann) Diesing, Protoceratium reticulatum (Claparède et Lachmann) Bütschli revealed a great nucleotide divergence of G. fragilis SSU sequence. Therefore, the SSU sequence could be used as species-specific marker for the identification of this mucilage producer microalga. In addition, such sequence could be used as target to design oligonucleotide probes for the construction of DNA microchips as diagnostic tool for the routine monitoring of harmful algae in seawater. An erratum to this article is available at .