Polyphyly of Asian Tree Toads,Genus Pedostibes Günther, 1876 (Anura: Bufonidae), and the Description of a New Genus from Southeast Asia

The Asian Tree Toad genus Pedostibes, as currently understood, exhibits a conspicuously disjunct distribution, posing several immediate questions relating to the biogeography and taxonomy of this poorly known group. The type species, P. tuberculosus and P. kempi, are known only from India, whereas P. hosii, P. rugosus, and P. everetti are restricted to Southeast Asia. Several studies have shown that these allopatric groups are polyphyletic, with the Indian Pedostibes embedded within a primarily South Asian clade of toads, containing the genera Adenomus, Xanthophryne, and Duttaphrynus. Southeast Asian Pedostibes on the other hand, are nested within a Southeast Asian clade, which is the sister lineage to the Southeast Asian river toad genus Phrynoidis. We demonstrate that Indian and Southeast Asian Pedostibes are not only allopatric and polyphyletic, but also exhibit significant differences in morphology and reproductive mode, indicating that the Southeast Asian species’ are not congeneric with the true Pedostibes of India. As a taxonomic solution, we describe a new genus, Rentapia gen. nov. to accommodate the Southeast Asian species.     

Leaching of concanavalin A during affinity chromatographic isolation of cell surface glycoproteins from human fetal neurons and glial cells.

Preparation of surface glycoproteins from human fetal brain cells by affinity chromatography on Con A-Sepharose 4B was a problematic endeavor due to leaching of Con A from the matrix. Dissociation of Con A from the matrix took place irrespective of the presence of lipid and/or detergent and the buffer composition during chromatography and was apparently related to the nature of the protein under study. Pretreatment of Con A-Sepharose with 6 M guanidine or 8 M urea reduced Con A leaching. The Con A eluate also contained noncovalently associated glycolipid. Elution at 25 degrees C rendered fractions containing a higher degree of Con A and glycolipid contamination compared to the negligible contamination by these two components when elution was carried out at 4 degrees C. This phenomenon was attributed to the formation of heterogeneous mixed micelles of glycoprotein.     

Characterization of the oriC region of Mycobacterium smegmatis.

A 3.5-kb DNA fragment containing the dnaA region of Mycobacterium smegmatis has been hypothesized to be the chromosomal origin of replication or oriC (M. Rajagopalan et al., J. Bacteriol. 177:6527-6535, 1995). This region included the rpmH gene, the dnaA gene, and a major portion of the dnaN gene as well as the rpmH-dnaA and dnaA-dnaN intergenic regions. Deletion analyses of this region revealed that a 531-bp DNA fragment from the dnaA-dnaN intergenic region was sufficient to exhibit oriC activity, while a 495-bp fragment from the same region failed to exhibit oriC activity. The oriC activities of plasmids containing the 531-bp sequence was less than the activities of those containing the entire dnaA region, suggesting that the regions flanking the 531-bp sequence stimulated oriC activity. The 531-bp region contained several putative nine-nucleotide DnaA-protein recognition sequences [TT(G/C)TCCACA] and a single 11-nucleotide AT-rich cluster. Replacement of adenine with guanine at position 9 in five of the putative DnaA boxes decreased oriC activity. Mutations at other positions in two of the DnaA boxes also decreased oriC activity. Deletion of the 11-nucleotide AT-rich cluster completely abolished oriC activity. These data indicate that the designated DnaA boxes and the AT-rich cluster of the M. smegmatis dnaA-dnaN intergenic region are essential for oriC activity. We suggest that M. smegmatis oriC replication could involve interactions of the DnaA protein with the putative DnaA boxes as well as with the AT-rich cluster.     

The effects of antioxidants on the metabolism and mutagenicity of benzo[a]pyrene in vitro.

Antioxidants inhibit the rat liver microsomal mixed-function-oxidase-catalysed hydroxylation of benzo[a]pyrene. These antioxidants also decrease the formation of mutagenic products from benzo[a]pyrene as judged by the Ames bacterial-mutagenicity assay [B.N. Ames, J. McCann & E. Yamazaki (1975) Mutat. Res. 31, 347-364]. It is suggested that antioxidants exert their protective effect against cancer by inhibiting the formation of carcinogenic metabolites.     

The mechanism of the ring trap of the predacious hyphomyceteDactylella brochopaga Drechsler

Summary A mutant ofDactylella brochopaga which produces giant functional ring traps, was isolated and studied. From experiments in which such traps were compressed, punctured and caused to inflate and deflate in environments of varying humidity levels it was concluded that trap cells are inflated by absorbing water or water vapour directly through their cell walls from their surroundings. Water entry occurs by osmosis, is initiated by a sudden increase in plasma membrane permeability and the increased pressure causes the inner of the two cell wall layers either to expand elastically or to unfold from a prepackaged state. In the fully inflated cell this wall layer becomes plastic finally.     

A radioimmunoassay method for the rapid detection of Candida antibodies in experimental systemic candidiasis

Rabbits were employed as experimental models to evaluate a solid-phase radioimmunoassay (RIA) method for the diagnosis of systemic candidiasis. Ten rabbits were inoculated subcutaneously to mimic superficial candidiasis and were found to produce no antibodies to Candida as determined by both immunodiffusion and RIA procedures. However, 94 per cent of 18 rabbits systemically infected by intravenous injection of Candida cells were observed to produce antibody as assessed by the RIA technique. These data encourage further tests with human sera and the continued development of this RIA procedure as a useful tool in the early serodiagnosis of systemic candidiasis.     

Detection of virus-like particles in the coremia of Penicillium claviforme.

A double-stranded ribonucleic acid (dsRNA) was isolated from coremial extracts of wild type P. claviforme, by methylated-albumin kieselguhr chromatography. Differential centrifugations of the coremial extracts from WT and Sh mutant strains yielded two classes of virus-like particles (VLP), of dimensions 25-30 nm, and 50-70 nm. The possible ecological significance of fungal viruses is discussed.     

Interactions of steroids with human placental cytochrome P-450 in the presence of carbon monoxide.

Spectral analyses of the carbon monoxide (CO) complex of human placental microsomal cytochrome P-450 revealed absorption maxima at 426 and 450 nm when NADPH (2×10⁻⁴M) was utilized as a reducing agent. Additional NADPH or NADH did not produce any further increases in the absorption maximum at 450 nm. A period of 10–15 minutes was required for the complete reduction. Various steroids were added to both sample and reference cuvettes to examine their interactions with the CO-cytochrome P-450 complex. The resulting spectral changes indicated that low concentrations of steroids (≃10⁻⁷M) such as androstenedione, 19-hydroxyandrostenedione, 19-oxoandrostenedione and testosterone completely eliminated the absorbance maxima at 450 nm while 19-norandrostenedione, 19-nortestosterone, pregnenolone and benzo[a]-pyrene did not eliminate this peak. Since ample time was allowed to reduce the cytochrome P-450 with NADPH, the observed interaction of steroids with cytochrome P-450 in the presence of CO does not represent an effect on reductase activity, but on the formation of the CO-cytochrome P-450 complex.     

Spectral characteristics of human fetal adrenal microsomes.

Investigations of human fetal adrenal gland microsomes indicated that a carbon monoxide binding pigment had an absorption maximum of 446 to 448 nm. This pigment, upon heat treatment at 37°C was degraded to the form of cytochrome p-420. NADPH reduced cytochrome p-450 slowly and completely. Typical concentrations of 0.75 and 0.16 nmoles/mg protein cytochrome P-450 and b₅, respectively, were observed. Reduced ethylisocyanide spectra were similar to those of rat hepatic microsomes with absorption maxima at 430 as well as 454 nm. Typical type I spectral changes were observed with progesterone, 17-α-OH-progesterone, pregnenolone and androstenedione when these steroids were added to the sample cuvettes. Androstenedione exhibited an apparent spectral dissociation constant (K_S) of 5×10⁻⁶M pregnenolone and progesterone exhibited higher affinities with apparent dissociation constants of 1.1×10⁻⁷M and 1.8×10⁻⁷M, respectively. The maximal absorbance change induced by androstenedione was lower (Emax = 0.027 per mg protien) than the changes in absorbance maxima induced by pregnenolone or progesterone (Emax = 0.060 and 0.047 per mg protein, respectively) when saturating concentrations of these steroids were added to the sample cuvettes. Ethylmorphine and aminopyrine (10⁻³M final concentrations) did not exhibit observable spectral changes; however, type II spectra could be elicited with aniline and nicotinamide and apparent dissociation constants of 3.5×10⁻²M and 2.5×10⁻²M, respectively, were obtained.