BIOGEOGRAPHY OF CLADOPHOROPSIS MEMBRANACEA (CHLOROPHYTA) BASED ON COMPARISONS OF NUCLEAR rDNA ITS SEQUENCES1

Nucleotides were compared at 988 sites, spanning both internal transcribed spacers (ITS1 and ITS2) of the nuclear ribosomal DNA, among 17 isolates of the green alga Cladophoropsis membranacea (Hofman Bang ex C. Agardh) Boergesen and two isolates of Struvea anastomosans (Harvey) Piccone and Grunow. Collections were made from Bonaire, Curaçao, St. Croix, the Canary Islands, the Cape Verde Islands, Mauritania, Syria, the Red Sea, Okinawa, and Hawaii. Two nucleotide substitutions were found between the sequenced coding regions of C. membranacea and S. anastomosans. Of the 720 nucleotides compared in ITS1 and ITS2, an average of 6.7% sequence divergence was found within C. membranacea and 17.4% between C. membranacea and S. anastomosans. Sequences were analyzed using maximum parsimony. Phylogenetic hypotheses were compared with the biogeographic hypothesis of an east-west Tethyan vicariance. Results of the study allow the identification of widely dispersed biogeographic populations, the identification of an underlying Tethyan imprint, and support for the hypothesis that dispersal has occurred between the Caribbean and the tropical eastern Atlantic. These results demonstrate the usefulness of ITS sequences at the sub-specific level in C. membranacea.     

MOLECULAR PHYLOGENY OF THE GENUS CAULERPA (CAULERPALES,CHLOROPHYTA) INFERRED FROM CHLOROPLAST tufA GENE1

The genus Caulerpa consists of about 75 species of tropical to subtropical siphonous green algae. To better understand the evolutionary history of the genus, a molecular phylogeny was inferred from chloroplast tufA sequences of 23 taxa. A sequence of Caulerpella ambigua was included as a potential outgroup. Results reveal that the latter taxon is, indeed, sister to all ingroup sequences. Caulerpa itself consists of a series of relatively ancient and species‐poor lineages and a relatively modern and rapidly diversifying clade, containing most of the diversity. The molecular phylogeny conflicts with the intrageneric sectional classification based on morphological characters and an evolutionary scheme based on chloroplast ultrastructure. High bootstrap values support monophyly of C. mexicana, C. sertularioides, C. taxifolia, C. webbiana, and C. prolifera, whereas most other Caulerpa species show para‐ or polyphyly.     

PHYLOGENETIC POSITION OF CRUSTOMASTIX STIGMATICA SP. NOV. AND DOLICHOMASTIX TENUILEPIS IN RELATION TO THE MAMIELLALES (PRASINOPHYCEAE,CHLOROPHYTA)1

A new marine microalga from the Mediterranean Sea, Crustomastix stigmatica Zingone, is investigated by means of LM, SEM, TEM, and pigment and molecular analyses (nuclear‐encoded small subunit [SSU] rDNA and plastid‐encoded rbcL). Pigment and molecular information is also provided for the related species Dolichomastix tenuilepis Throndsen et Zingone. Crustomastix stigmatica has a bean‐shaped cell body 3–5 μm long and 1.5–2.8 μm wide, with two flagella four to five times the body length. The single chloroplast is pale yellow‐green, cup‐shaped, and lacks a pyrenoid. A small bright yellow stigma is located in the mid‐dorsal part of the cell under the chloroplast membrane. An additional accumulation of osmiophilic globules is at times seen in a chloroplast lobe. Cells lack flat scales, whereas three different types of hair‐like scales are present on the flagella. The main pigments of C. stigmatica are those typical of Mamiellales, though siphonein/siphonaxanthin replaces prasinoxanthin and uriolide is absent. The pigment pool of D. tenuilepis is more similar to that of Micromonas pusilla (Butcher) Manton et Parke and of other Mamiellales. The nuclear SSU rDNA phylogeny shows that the inclusion of C. stigmatica and D. tenuilepis in the Mamiellales retains monophyly for the order. The two species form a distinct clade, which is sister to a clade including all the other Mamiellales. Results of rbcL analyses failed to provide phylogenetic information at both the order and species level. No unique morphological or pigment characteristics circumscribe the mamiellalean clade as a whole nor its two daughter clades.     

De novo synthesis of steroids (from acetate) by isolated rat Sertoli cells.

Sertoli cells from 17 day old rats were shown to convert [¹⁴C]acetate to [¹⁴C]-labelled cholesterol, pregnenolone and 17α-hydroxypregnenolone$\text{in}$$\text{vitro}$. Identification was by several systems of thin layer and gas chromatography of the extracted steroids and their sylil and acetyl derivatives and by recrystallizations with authentic and acetylated unlabelled steroids. Several other steroids formed from acetate were tentatively identified. No androstenedione or testosterone were formed. That the Sertoli cell cultures were free of Leydig cells was established by the absence of histochemically detectable 3β-hydroxysteroid dehydrogenase activity and the inability of the cultures to oxidize the 3β-hydroxyl group of [¹⁴C]pregnenolone. This is the first direct evidence that Sertoli cells have the capacity to synthesize steroids $\text{de}$$\text{novo}$ from acetate.     

Influence of temperature gradients on leaf water potential

Water potential was monitored at nine locations along single maize (Zea mays L.) leaf blades with aluminum block in situ thermocouple hygrometers. Water potential showed a continuous decrease toward the tip, with a 2- to 4-bar difference between leaf base and tip under both moist and dry soil conditions. The water potential difference between the soil and the leaf base was about 4 bars. Water potentials decreased during the day and during a drying cycle, and increased at night and after irrigation. Heating a band of a leaf to 40 C or cooling it to 7 C had no influence on the water potential of the affected portion when this was corrected for hygrometer output over standard calibrating solutions at the respective temperatures. Heating or cooling a portion of a leaf had neither short nor long term effects on water potential of more distal leaf portions continuously monitored by hygrometers in dew point readout. Water potential fluctuated with an amplitude of about 1.5 bars and an irregular period of 10 to 30 minutes. Measurements with silver foil in situ psychrometers gave similar results.     

A simple procedure for solubilizing 3beta-hydroxysteroid dehydrogenase from microsomes of rat adrenals and testis interstitial cells

A simple procedure is described for solubilizing microsomal 3beta-hydroxysteroid dehydrogenase (3beta-HSD). Microsomes from rat adrenals or from testicular interstitial cells were incubated for 1 or 2 h at 0 C in a buffer containing NaCl followed by overnight storage at -20 C. Maximum solubilization of 3beta-hydroxy-5beta-androstan-17-one-HSD (androstane-3beta-HSD) was obtained by incubating adrenal microsomes with 1 M NaCl and interstitial cell microsomes with 2 M NaCl. Incubation with NaCl for 1 or 2 h resulted in maximum solubilization; incubation with NaCl for 4, 8 or 24 h did not change the amount of enzyme solubilized. From adrenal microsomes incubated with 1 M NaCl, up to 80% (105.7 millimicron/mg microsomes) of the total androstane-3beta-HSD activity was recovered in the supernatant following centrifugation at 130,000 x g for 1 h. The maximum amount of androstane-3beta-HSD solubilized from interstitial cell microsomes was 56% (29.5 millimicron/mg microsomes) at 2 M NaCl. The "solubilized" androstane-3beta-HSD was retarded when chromatographed on a Sephadex G-200 column and it did not pellet out when centrifuged at 130,000 x g for 15 h. KCL appeared to be equally effective in solubilizing androstane-3beta-HSD from microsomes. Other steroid dehydrogenase activities such as pregnanolone-HSD and 3beta-hydroxy-5alpha-androstan-17-one-HSD were also found in the 130,000 x g supernatant.     

MOLECULAR EVIDENCE CONFIRMS SISTER RELATIONSHIP OF ARDISSONEA,CLIMACOSPHENIA, AND TOXARIUM WITHIN THE BIPOLAR CENTRIC DIATOMS (BACILLARIOPHYTA,MEDIOPHYCEAE), AND CLADISTIC ANALYSES CONFIRM THAT EXTREMELY ELONGATED SHAPE HAS ARISEN TWICE IN THE DIATOMS1

With the use of a new kit from Qiagen to amplify total genome quantity, DNA was bulked up from two diatoms that are difficult to grow (Ardissonea and Climacosphenia), and the nuclear SSU rRNA gene was successfully amplified. Results of Bayesian analyses showed that these diatoms are sister to Toxarium and belong to the bi‐ and multipolar centric diatoms. The results indicate that extremely elongate shape has arisen at least twice in diatoms, in the true pennates, and in the bipolar centrics. The two lateral pattern centers of Ardissonea and Climacosphenia likely represent a modified annulus that subtends ribs internally as well as externally. Studies of sexual reproduction are needed to determine whether Ardissonea, Climacosphenia, and Toxarium achieve their elongate shape by similar means to each other and to true pennates, that is, by controlling the expansion of the auxospores by sequential addition of silicified bands (to form a properizonium or perizonium).     

At-line determination of octanoic acid in cultivation broth--an electronic tongue (ET) feasibility study

Production monitoring of "natural" 2-heptanone from octanoic acid in an industrial fed-batch cultivation based on Penicillium roqueforti requires development of a method for determination of octanoic acid dissolved in the water phase. An electronic tongue array using six non-specific potentiometric sensors with solid inner contact, and a pH electrode, has been introduced by spiking octanoic acid to a substrate obtained from four different cultivations, representing variations in the relevant industrial matrix. Multivariate calibration was performed on acid concentrations spanning 0.65-20 mmol l(-1). Excluding the lowest concentration a global Partial Least Square regression model with a predicted versus measured correlation of 0.98 and a relative root mean square error of prediction of 5.1% (ln units) (RPD=5.5) signifies a highly acceptable prediction facility. This model was further tested by subjecting it to undiluted as well as diluted samples obtained from a cultivation process in which octanoic acid was catabolized; this led to acceptable prediction errors within the same range as for the global model. It is concluded that the ET sensor array can be applied for determination of octanoic acid in cultivation systems of the general P. roqueforti type.