Protein Synthesis in Isolated Symbionts from the Flagellate Protozoon Crithidia deanei1

Here we describe a method that allows the isolation of intact trypanosomatid symbionts in amounts sufficient for biochemical analysis. The isolated symbionts retain their characteristic morphological features and are reasonably free of subcellular debris. They actively incorporate [³H]leucine and [³⁵S]methionine into proteins. Chloramphenicol and rifampicin at 50 μg/ml almost completely inhibit the incorporation of protein precursors. The inhibition of protein synthesis by the antibiotics provides direct evidence for the existence of a prokaryotic protein-synthesizing system in this unusual intracellular structure. The pattern of protein synthesis of the isolated symbionts is complex. Several symbiont polypeptides are absent or poorly represented in the flagellate.     

Phosphorus transformations along a soil/vegetation series of fire-prone, dolomitic, semi-arid shrublands of southern Spain Soil P and Mediterranean shrubland dynamic

Models of P transformations duringpedogenesis and with succession have developed fromstudies in temperate humid regions with neutral toacidic soils. Little is known about P biogeochemistryand P availability in semi-arid Mediterranean-typeshrublands with alkaline soils. We studied Ptransformations in a series of semi-arid, dolomiticshrublands in southeastern Spain, ranging from afrequently-burned, open gorse-scrubland on erodedTypic Xerorthents to a long-unburned, mature garrigueon Entic Haploxerolls. In contrast to the commonpattern of decreasing total P concentrations in thesoil profile with soil development due to leaching,total P increased markedly in this system. This is dueto concentration increases of relatively insolubleelements (P, Al, Fe, Ti) as karstification of parentmaterial (dolomitic marbles with up to 94% Ca-Mgcarbonate) during pedogenesis released bicarbonateand, subsequently, Ca and Mg leached from the profileat a higher rate. The total element to Ti ratiosindicated that the relative ion weathering losssequence, from easily weathered to resistant ions, wasCa>Mg > > > Fe>=Al>Ti, showing that P islost from these shrublands at an intermediate rate. Inone extreme of the series (the open gorse-scrubland),most soil P was Ca bound and organic P concentrations,organic matter content and phosphatase activity werevery low, as predicted by the model of Walker andSyers for the initial stages of soil development.However, this site showed the highest inorganic soilsolution P concentration, low soil P fixation capacityand the lowest foliar N:P ratios. Soils from theintermediate stages of the series showed the highestlabile inorganic and labile organic P concentrations.At the other extreme of the series (the maturegarrigue), a high proportion of soil P was in occludedinorganic and organic forms as predicted by the Walkerand Syers'; model. However, Ca bound P still accountedfor the largest single P fraction. Soils showed veryhigh sorption capacity (and high extractable Fe and Alconcentrations) and released very little P tosolution. Increasing values for NaOH extractable Po,organic matter and phosphatase activity indicate thatcycling of P through organic matter is increasinglyimportant with ecosystem development through theseries.     

INFLUENCE OF ENVIRONMENTAL PARAMETERS ON REPRODUCTION OF THE EUROPEAN FLAT OYSTER (OSTREA EDULIS L.) IN A COASTAL LAGOON (MAR MENOR, SOUTHEASTERN SPAIN)

The influence of environmental parameters (temperature, dissolvedoxygen, suspended matter, chlorophyll a) on the condition indicesand gametogenic cycle of the European flat oyster (Ostrea edulisL.) was analyzed in the Mar Menor (Murcia, Spain) between January1990 and December 1992. The highest condition index values wererecorded during the prespawning season at temperatures between11–12°C. Condition index values gradually decreasedfrom 14°C, coinciding with the appearance of the first larvaein the plankton. Condition index values were lowest in summer.Multiple regression analysis revealed that the condition indiceswere correlated mainly with temperature and to a lesser extent,with dissolved oxygen, chlorophyll a and suspended matter. Absolutechlorophyll concentrations were low and presented a negativecorrelation with the condition index values, though this correlationwas less pronounced than that of temperature. Gametogenesiswas continuous all year round, but spawning took place onlyat temperatures of 14°C or higher, and larvae were presentin the plankton to 28°C. The low absolute values of chlorophylla and suspended matter found in the oligotrophic Mar Menor didnot seem to affect the gametogenesis, spawning or the larvaldevelopment of the oysters, which feed on picoplankton, principallycomposed of dinoflagellates and bacteria. (Received 29 April 1996; accepted 18 October 1996)     

Biologic,Immunocytochemical, and Cytogenetic Characterization of Two New Human Melanoma Cell Lines: IIB-MEL-LES and IIB-MEL-IAN

Two human melanoma cell lines, derived from metastases of two patients with epithelioid malignant amelanotic melanomas, and designated IIB-MEL-LES and IIB-MEL-IAN, have been established. Both cell lines have been in continuous culture over 2 years and were propagated continuously for 85 and 75 serial passages, respectively. Morphologically, IIB-MEL-LES is composed predominantly of spindle shaped cells, whereas IIB-MEL-IAN grows as a monolayer of cuboid and stellate shaped cells with many rounded cells in suspension. Immunocytochemical studies revealed that both cell lines express S-100 protein, vimentin, and GD₃ and GD₂ gangliosides but are negative for keratin and collagen. Both cell lines express HLA class I and HLA-DR antigens in variable proportions. The MAGE-1 gene is expressed only by the IIB-MEL-IAN cell line, as revealed by PCR analysis. Cytogenetic analysis of both cell lines revealed abnormal karyotypes; the modal chromosome numbers of IIB-MEL-LES and IIB-MEL-IAN were 48 and 81, respectively. IIB-MEL-LES cells presented rearrangements in chromosomes 1, 14 and X, gains in chromosomes 10,20, and 21 losses in chromosomes 15 and Y. The most frequent markers observed in IIB-MEL-IAN cells were 7q+, 10p+, 2p+, i(6p), 2q+, and 10q-. Clonal gains were observed in chromosomes 12 and 21, whereas losses were seen in chromosomes 1, 2, 3, 4, 6, 7, 11, and 17. Both cell lines were capable of forming colonies in soft agar and developed tumors when transplanted into nude mice, reproducing and maintaining the characteristics of the original tumors. These cell lines and their xenografts appear to provide useful systems for studying the biology, genetics and histogenesis of human malignant melanoma and could be utilized for the development of melanoma vaccines.     

Measured and Calculated Transpiration in Trifolium repens under Different Water Potentials

The relationship between transpiration measured gravimetrically,a generalized transpiration equation, and the ratio VPD/r_leafwas investigated in Trifolium repens plants subjected to varyingwater potentials. Dawn leaf water potential was measured witha pressure chamber, leaf diffusion resistance with a diffusionporometer, leaf temperatures with a thermistor, and relativehumidity with an aspirated psychrometer. During drought transpirationrates determined by both methods were quite similar particularlyat the lowest water potentials. After rewatering calculatedrates were somewhat higher than measured ones. It is concludedthat transpiration calculated by the indirect method is a usefuland reasonable estimate of transpiration for single plants undervarying water potentials.     

Comparative Study of Experimental Ocular Melanoma Using Two Implantation Techniques of B16-F10 Melanocytes

Objective: The aim of this work was to evaluate the behaviour of B16-F10 melanoma cell cultures implanted in the anterior chamber of the eye of New Zealand white rabbits by studying the clinical-pathological and ultrastructural characteristics of the lesions. Methods: One group (A) (consisting of 30 rabbits) was transclerally inoculated (1 mm from sclero-corneal limbus) with 4×10⁶ melanocytes and another group (B) (also 30 animals) was inoculated once per week for 3 consecutive weeks with 5×10⁶ cells (total 15×10⁶); 30 animals acted as the control group (C). All the lesions were processed for optic and electronic microscopy. Results: Tumoral growth in group A was 43% (13/30) and in group B 80% (24/30). All lesions were pigmented and none perforated the eyeball. Microscopically, they were a mixture of epithelioid and fusiform cells disposed around the blood vessels. Ultrastructurally, the presence of melanosomes in different stages of maturation and aberrant melanosomes were characteristic. Conclusion: We suggest that the transcleral inoculation of 15×10⁶ B16-F10 melanocytes into the anterior chamber of the eye of New Zealand white rabbits may be a valid and reproducible method for obtaining an experimental ocular melanoma model.