Antifungal Resistance and Virulence Among <Emphasis Type="Italic">Candida</Emphasis> spp. from Captive <Emphasis Type="Italic">Amazonian manatees</Emphasis> and West Indian Manatees: Potential Impacts on Animal and Environmental Health

This work aimed at evaluating the antifungal susceptibility and production of virulence factors by Candida spp. isolated from sirenians in Brazil. The isolates (n = 105) were recovered from the natural cavities of Amazonian and West Indian manatees and were tested for the susceptibility to amphotericin B, itraconazole, and fluconazole and for the production of phospholipases, proteases, and biofilm. The minimum inhibitory concentrations (MICs) for amphotericin B ranged from 0.03 to 1 µg/mL, and no resistant isolates were detected. Itraconazole and fluconazole MICs ranged from 0.03 to 16 µg/mL and from 0.125 to 64 µg/mL, respectively, and 35.2% (37/105) of the isolates were resistant to at least one of these azole drugs. Concerning the production of virulence factors, phospholipase activity was observed in 67.6% (71/105) of the isolates, while protease activity and biofilm production were detected in 50.5% (53/105) and 32.4% (34/105) of the isolates, respectively. Since the natural cavities of manatees are colonized by resistant and virulent strains of Candida spp., these animals can act as sources of resistance and virulence genes for the environment, conspecifics and other animal species, demonstrating the potential environmental impacts associated with their release back into their natural habitat.     

Cell-bound and extracellular carboxylesterases from Streptomyces: hydrolytic and synthetic activities

AIM: The distribution of cell-bound and extracellular carboxylesterases was investigated among the genus Streptomyces using 420 strains. METHODS AND RESULTS: Primary screening was carried out on solid media using tributyrin, triolein and Tween 60 as current substrates. Eleven representative strains were selected and grown in submerged cultures for evaluating their cell-bound and extracellular hydrolytic activity independently on various naphthyl and aliphatic esters. The best lipolytic strain was lyophilized and used as dry mycelium for catalysing the synthesis of various aliphatic esters in heptane, with molar conversions ranging from 28 to 78% after 3 days. CONCLUSIONS: Carboxylesterase activities can easily be found among the Streptomyces, often being cell-bound and also employable for catalysing esterification in organic solvent. SIGNIFICANCE AND IMPACT OF THE STUDY: A wide screening among Streptomyces, a genus poorly studied for the production of carboxylesterases, has allowed the selection of several strains with interesting enzymatic activities to be used in commercially valuable biotransformation.     

CNS myelin and sertoli cell tight junction strands are absent in Osp/claudin-11 null mice

Oligodendrocyte-specific protein (OSP)/claudin-11 is a recently identified transmembrane protein found in CNS myelin and testis with unknown function. Herein we demonstrate that Osp null mice exhibit both neurological and reproductive deficits: CNS nerve conduction is slowed, hindlimb weakness is conspicuous, and males are sterile. Freeze fracture reveals that tight junction intramembranous strands are absent in CNS myelin and between Sertoli cells of mutant mice. Our results demonstrate that OSP is the mediator of parallel-array tight junction strands and distinguishes this protein from other intrinsic membrane proteins in tight junctions. These novel results provide direct evidence of the pivotal role of the claudin family in generating the paracellular physical barrier of tight junctions necessary for spermatogenesis and normal CNS function.     

An attempt to apply ligand field theory to positronium reactions with 3d complexes

Positronium, Ps, the bound state between an electron, e, and a positron, c⁺, may have two spin states: theortho,o-Ps, and thepara,p-Ps, states, which differ for the spin orientation of the two particles. The two types of Ps atoms may be inter-converted by collision with paramagnetic compounds, such as several3d complexes. By investigating about 90 complexes of V^II, Cr^II, Cr^III, Mn^II, Fe^II, Co^II and Ni^II as a function of temperature, it was found that the rate constants k_CR of theo-Ps intop-Ps conversion reactions, CR, are linearly correlated to the delocalization β of metal electrons caused by the ligands. Therefore, if β known, k_CR may be estimated andvice versa. This suggests a new method for the experimental determination of β. The rate constants of the Ps oxidation reactions by Co^III complexes were also investigated and discussed. The paper is preceded by four sections dealing with: 1) the positron and positronium formation; 2) the positron annihilation modes; 3) the methods for measuring the Ps rate constants and establishing the Ps reaction types; 4) the application of the Smoluchowski equation to Ps reactions. Moreover, an attempt is made to ascertain the standard electrochemical potential of Ps atoms. The positron reactions and the formation of positronides are also taken into consideration. Presentata nella seduta del 13 dicembre 2002 dal Socio F. Calderazzo.     

The M.ADP.P(i) state is required for helical order in the thick filaments of skeletal muscle

The thick filaments of mammalian and avian skeletal muscle fibers are disordered at low temperature, but become increasingly ordered into an helical structure as the temperature is raised. Wray and colleagues (Schlichting, I., and J. Wray. 1986. J. Muscle Res. Cell Motil. 7:79; Wray, J., R. S. Goody, and K. Holmes. 1986. Adv. Exp. Med. Biol. 226:49-59) interpreted the transition as reflecting a coupling between nucleotide state and global conformation with M.ATP (disordered) being favored at 0 degrees C and M.ADP.P(i) (ordered) at 20 degrees C. However, hitherto this has been limited to a qualitative correlation and the biochemical state of the myosin heads required to obtain the helical array has not been unequivocally identified. In the present study we have critically tested whether the helical arrangement of the myosin heads requires the M.ADP.P(i) state. X-ray diffraction patterns were recorded from skinned rabbit psoas muscle fiber bundles stretched to non-overlap to avoid complications due to interaction with actin. The effect of temperature on the intensities of the myosin-based layer lines and on the phosphate burst of myosin hydrolyzing ATP in solution were examined under closely matched conditions. The results showed that the fraction of myosin mass in the helix closely followed that of the fraction of myosin in the M.ADP.P(i) state. Similar results were found by using a series of nucleoside triphosphates, including CTP and GTP. In addition, fibers treated by N-phenylmaleimide (Barnett, V. A., A. Ehrlich, and M. Schoenberg. 1992. Biophys. J. 61:358-367) so that the myosin was exclusively in the M.ATP state revealed no helical order. Diffraction patterns from muscle fibers in nucleotide-free and in ADP-containing solutions did not show helical structure. All these confirmed that in the presence of nucleotides, the M.NDP.P(i) state is required for helical order. We also found that the spacing of the third meridional reflection of the thick filament is linked to the helical order. The spacing in the ordered M.NDP.P(i) state is 143.4 A, but in the disordered state, it is 144. 2 A. This may be explained by the different interference functions for the myosin heads and the thick filament backbone.     

Association of Ribosomes with Intracellular Vesicular Stomatitis Virus Particles

Ribosomes are observed intimately associated with nucleocapsids of vesicular, stomatitis virus, especially those that line structures that are either cytoplasmic vesicles or invaginations of the plasma membrane.     

Immunocytochemical localization of vesicular stomatitis virus proteins N and NS with monoclonal antibodies

The purpose of this paper is to describe the immunocytochemical-localization of N and NS nucleocapsid proteins of vesicular stomatitis virus in the cells throughout the infectious cycle. N protein was detected in the cytoplasm at 2 h after infection and formed small cytoplasmic clusters which progressively increased in size and number. At 5-6 h, it formed large cytoplasmic inclusions. NS protein was detected in the cytoplasm a little later than N protein and showed almost the same immunostaining pattern. However, diffuse background staining of NS protein was identified throughout the cytoplasm by double immunostaining methods. At electron microscopic level, N protein was mostly granular and occasionally organized in strands at 2-3 h. At 5-6 h, numerous immunostained reaction products were organized in strands. The reaction products of NS protein were almost the same as those of N protein with the exception that diffuse background staining was observed. Cos cells, transfected with SV40 vector containing N gene obtained by recombinant DNA technique, showed clusters of N protein, but virtually no strand at electron microscopic levels. The rapid-freezing and deep-etching replica method demonstrated that loosely coiled VSV genome coated with N protein was localized on cytoplasmic sides of cell membranes in the infected cells. These results showed that complete virus genome replication was needed for strand formation of N and NS proteins and suggested that they were bound to VSV genomes in the infected cells.