In Vitro Culture of Phytomonas Sp. Isolated from Euphorbia characias. Metabolic Studies by 1H NMR

ABSTRACT. We describe the in vitro culture of Phytomonas species isolated from Euphorbia characias . The best choice between tested media was SDM-79, in which promastigotes, after 6 days of culture, reached cell densities as high as 4 × 10⁷ cells/ml. Cells growing in LIT or MTL medium showed longer division times and lower cell densities. We succeeded in obtaining Phytomonas sp. amastigote and spheromastigote forms in modified GRACE's medium, yielding transformation rates of up to 70%. Electron microscopy studies were performed in order to characterize the ultrastructural features of these forms obtained in vitro. On the other hand, metabolic studies based on qualitative (nuclear magnetic resonance spectroscopy) and quantitative metabolic methods (enzymatic assays) showed that promastigote forms secreted mainly ethanol, acetate, glycine, glycerol, piruvate and succinate in SDM-79 medium, whereas the major metabolites found after transformation in modified Grace's medium were ethanol, acetate, glycine, piruvate and smaller amounts of glycerol.     

Isolation and identification of (2S)-2-amino-5-chloro-4-hydroxy-5-hexenoic acid from anAmanita of the sectionRoanokenses (Amanitaceae)

A chlorine-containing non-protein amino acid which was recently discovered from the fruit bodies ofAmanita gymnopus (2S)-2-amino-5-chloro-4-hydroxy-5-hexenoic acid, was isolated and crystallized for the first time from the fruit bodies of an unknown member ofAmanita belonging to the sectionRoanokenses, subsectionSolitariae. The results of elementary analyses, determination of optical rotations,¹H- and¹³C-NMR-spectra, and some chemical reactions supported an earlier proposed structure.Part 24 in the series Biochemical studies of nitrogen compounds in fungi. for Part 23, see Hatanaka, S. I. et al. 1994. this journal35: 391–394.     

Discrepancies between MIC and MLC values of amphotericin B against isolates ofAspergillus species

There is little information addressing the phenomena of discrepancy between minimal inhibitory concentrations (MIC) and minimal lethal concentrations (MLC) values of amphotericin B (AMB) to clinical isolates of fungi. This study assessed in vitro activity of AMB against 70 clinical isolates of aspergilli: 30 strains ofAspergillus fumigatus, 20 strains ofAspergillus flavus and 20 strains ofAspergillus niger. Susceptibility tests were accomplished using a macro broth dilution procedure, with special emphasis on ascertainment of MLCs. AMB exhibited low MIC values against all clinical isolates. While we did not identify any AMB resistant isolates among 70Aspergillus spp. studied as judged by MIC levels, analysis of the data demonstrated a clear discrepancy between the MIC and MLC levels of AMB obtained against clinical isolates ofAspergillus spp. The MLC values of AMB were significantly higher than the MIC values with MIC 50 and MIC 90 of 0.29 and 0.5 µg/ml, respectively, at the second reading time, and MLC 50 and MLC 90 of 2.31 and 9.24 µg/ml, respectively (p<0.001). Additionally, minimal lethal concentrations in 36/70 (51%) of aspergillal isolates studied produced drug concentrations above those which can usually be sustained in patient plasma or tissue.     

诺卡氏菌属GS-17耐热茁霉多糖酶的提纯和性质

诺卡氏菌属GS-17(Nocardia sp.GS-17)的耐热茁霉多糖酶(Pullulanase EC.3.2.1.41)的粗酶液经中空纤维柱超滤浓缩、羟基磷灰石柱层析和Pullulan-Sepharose 6B亲和层析,得到凝胶电泳均一的纯酶,比活提高264倍.酶作用最适温度为55℃,最适PH6.2,分子量140000,等电点pI为6.0.该酶水解茁霉多糖、支链淀粉和可溶性淀粉,但不水解糖原.酶在50℃作用于茁霉多糖的米氏常数K_m为0.90mg/ml,最大反应速度V_(max)为57μmol·min~(-1)·mg~(-1).Zn~(2 )、Fe~(3 )、Hg~(2 )、Cu~(2 )、Pb~(2 )和环状糊精对酶有抑制作用,Ca~(2 )对酶有激活作用.经蛋白质侧链化学修饰研究表明,色氨酸残基位于酶的活性位区.该酶是由1129个氨基酸残基组成的单肽链,酶的N末端序列经测定为:Ala-Gly-His-Gly-Pro-Asp-Val-Gln-Asp-Gly-     

Keraladiaptomus rangareddyi a new genus and new species of Diaptominae (Copepoda,Calanoida, Diaptomidae) from a temporary pond in Mattam,Kerala State,India

Calanoid copepods, belonging to the new genus and species Keraladiaptomus rangareddyi, were collected from temporary ponds in Kerala State, India. The new genus belongs to the family Diaptomidae, subfamily Diaptominae. It is described in detail and its affinity to the related genera, Arctodiaptomus Kiefer, 1932 and Eodiaptomus Kiefer, 1932, discussed.     

Descriptions of Two New Species of Chronogaster Cobb, 1913 from India

Two new species of Chronogaster in India were described and illustrated, based on light and scanning electron microscopy. Chronogaster neotypica n. sp. collected from a sewage slurry was characterized by a medium-sized body, a ventral tail mucro without additional spines, absence of longitudinal incisures in lateral fields, and by the presence of crystalloids in the body. Diagnostic for C. spinicauda n. sp. collected from soil around roots of mango were a medium-sized body, a tail mucro with 10 spines, and absence of lateral lines and crystalloids. Males were not found.     

The influence of culture conditions on mycorrhiza formation between the ectomycorrhizal fungus Pisolithus sp. and Afzelia africana Sm. seedlings

Interactions between an isolate of the ectomycorrhizal fungus Pisolithus sp. and Afzelia africana Sm. seedlings were studied at the structural and ultrastructural levels. Several different conditions were tested with or without sugar and in a sterile or nonsterile medium. In the growth cabinet, the A. africana/Pisolithus sp. interactions did not produce ectomycorrhizas. A fungal sheath was formed but no Hartig net, and an unusual host epidermal cell wall was observed. Hyphae of Pisolithus sp. induced modifications of epidermal cells of 15-day-old A. africana seedlings indicative of non-mycorrhizal interactions, such as wall thickening, wall ingrowth, papillae formation, degraded host wall material and the presence of intracellular hyphae. Wall ingrowth consisted of depositions of host cell wall materials giving a positive reaction for polysaccharides; however, wall thickenings and papillae showed no homogeneous reactions for polysaccharides. In glasshouse conditions, inocula of Pisolithus sp. in the form of spores or mycelia entrapped in peat-vermiculite added to sterilized soil produced typical ectomycorrhizae only with 6-month-old A. africana seedlings. Under these conditions, no conspicuous cell wall reactions occurred on A. africana roots. The results demonstrate that the establishment of an association between an ectomycorrhizal fungus and a potential host plant is strongly influenced by seedling age and/or environmental conditions. Therefore, in vitro synthesis is not a conclusive demonstration of a symbiotic relationship.     

Natural levels of fungal infections in grasshoppers in Northern Benin

The infection of grasshoppers by naturally occurring, entomopathogenic fungi was monitored at two sites in Malanville, northern Benin, Africa. Grasshoppers were collected and recorded from the sites between June and December 1992 and all of them, barring the first instars, were incubated in ventilated cages. At the first site, 1343 individuals of 35 grasshopper species were incubated, and at the second site, 857 individuals of 36 grasshopper species were incubated. Three hyphomycete fungi (Deuteromycotina: Hyphomycetes), Metarhizium flavoviride Gams and Rozsypal, Beauveria bassiana (Bals.) Vuillemin and Sorosporella sp. were found infecting grasshoppers. The average incidence of M. flavoviride infection was 2.9% and 1.8% at the two sites for all host species. M. flavoviride sporulated on most grasshopper cadavers within 10 days of collection. B. bassiana and Sorosporella sp. were only collected from one and five grasshopper individuals respectively. A significant difference was noted in the time to death of small grasshopper species infected with M. flavoviride compared to larger species. At one site, M. flavoviride infection was positively correlated with rainfall during the 10‐day period in which samples were taken.     

H2 production by mixed cultures

Production of H₂ from glucose by an anoxygenic phototrophic bacterium (Rhodobacter sphaeroides), a cyanobacterium (Synechococcus cedrorum) and a heterotrophic bacterium (Pseudomonas fluorescens) was tested individually and in mixed cultures of various combinations in light. H₂ production was maximal with a mixed culture of R. sphaeroides and P. fluorescens, which could be further enhanced by immobilization of the bacteria in alginate gel. Inhibition of H₂ photoproduction was observed in a mixture of S. cedrorum and P. fluorescens and a co-culture of all the three organisms.Ch. Sasikala and Ch. V. Ramana are and G. S. Prasad was with the Microbial Biotechnology Laboratory, Department of Botany, Osmania University, Hyderabad-500 007, India. G. S. Prasad is now with the Microbial Type Culture Collection Centre (MTCC), IMTECH, Chandigar, India.     

植物细胞离析酶的制备和应用

用 Aspergillus sp．A-19菌经固体发酵研制成一种新的植物细胞离析酶(SeparatasezA—P)。其离析单细胞的酶活力平均为70 767u／g,有效作用的pH在3．0—7．0,温度为20—45℃。发酵培养基配方是麸皮：桔皮粉：(NH₄)₂SO₄(w／w)为100:100:O．63,料水比为1 :2．0,培养适宜条件为25℃、60小时。