Detection of aromatic catabolic gene expression in heterogeneous organic matter used for reduction of volatile organic compounds (VOC) by biofiltration

A qualitative procedure of purified DNA/RNA co-extraction from complex organic matter, used as biofilter support for removing volatile organic compounds, was set up and applied to detect xylene monooxygenase gene expression by RT-PCR. A DNA/RNA extraction protocol based on a combination of sample lyophilization pre-treatment and CTAB––phenol/chloroform extraction procedure was optimized for the recovery of purified nucleic acids [100–500 ng DNA (10 kb) and 0.5–2 μg of rRNA 16S from 100 mg matrix]. PCR and RT-PCR protocols were established to detect xylene monooxygenase gene expression starting from differentially induced organic matrices obtained by biofiltration technology. This work allowed the microbial degradation activities in heterogeneous organic solid media to be studied and suggests a rapid method to follow specific biological activities during solid and/or semisolid organic substrates biotransformation.     

A new flounder,monolene helenensis (pleuronectiformes: Bothidae) from the eastern tropical atlantic

Monolene helenensis sp. nov. is described on the basis of seven specimens collected from the eastern tropical Atlantic Ocean. This new bothid flounder differs from all known congeners by the following combination of characters: 108–116 dorsal and 89–92 anal fin rays, 101–111 lateral line scales, 7–10+10–15=18–25 gill rakers, 10–12+37–38=48–50 vertebrae, body depth 2.58–2.77 in standard length, ocular side pectoral fin length 1.34–1.59, in head length and upper half of pectoral fin blackish.     

Use of an immobilized cell bioreactor for the continuous inoculation of milk in fresh cheese manufacturing

A system was developed to continuously acidify and inoculate skim milk for the production of fresh cheese. Four strains of mesophilic lactic acid bacteria were entrapped separately in κ -carrageenan/locust bean gum gel beads and used in a stirred bioreactor operated at 26°C with a 25% (v/v) gel load. The pH in the reactor was controlled at 6.0 by adding fresh milk using proportional integrated derived regulation. The bioreactor was operated during 8-h daily cycles for up to 7 weeks with different milks (heat treatment, dry matter content) and differing starting procedures. The heat treatment of the milk was an important factor for process performance: a dilution rate increase of 57% and an inoculation level decrease of 63% were observed with sterilized UHT skim milk (142°C – 7.5 s) compared with pasteurized skim milk (72°C – 15 s). The dry matter content of the milk (8–13% w/w) had no detectable effect on these parameters. A convenient starting procedure of the system was tested; steady-state was reached in less than 40 min following an interruption period of 16–60 h. These results combined with our published data on process performance show the feasibility of using an integrated immobilized cell bioreactor for milk prefermentation in cheese manufacture. Received 10 June 1996/ Accepted in revised form 15 October 1996     

Possible dynamic anchor points in a benzoxazinone derivative–human oxytocin receptor system — a molecular docking and dynamics calculation

In this study, we performed a molecular docking and dynamics simulation for a benzoxazinone–human oxytocin receptor system to determine the possible hydrophobic and electrostatic interaction points in the dynamic complex. After the homology modeling, the ligand was docked into the putative active using AutoDock 3.05. After the application of energetic and structural filters, the complexes obtained were further refined with a simulated annealing protocol (AMBER8) to remove steric clashes. Three complexes were selected for subjection to the molecular dynamics simulation (5 ns), and the results on the occurrence of average anchor points showed a stable complex between the benzoxazinone derivative and the receptor. The complex could be used as a good starting point for further analysis with site-directed mutagenesis, or further computational research. Figure The location of the ligands (complex B – blue; complex E – red; and complex F – green) in the transmembrane regions (TM1 – red; TM2 – blue; TM3 – yellow; TM4 – purple; TM5 – orange; TM6 – cyan; TM7 – pink) of the hOTR. For clarity, the EC and IC loops are not shown Electronic Supplementary Material Supplementary material is available at     

Evolutionary and dynamic stability in continuous population games

 Asymptotic stability under the replicator dynamics over a continuum of pure strategies is shown to crucially depend on the choice of topology over the space of mixed population strategies, namely probability measures over the real line. Thus, Strong Uninvadability, proved by Bomze (1990) to be a sufficient condition for asymptotic stability under the topology of variational distance between probability measures, implies convergence to fixation over a pure strategy x ^* only when starting from a population strategy which assigns to x ^* a probability sufficiently close to one. It does not imply convergence to x ^* when starting from a distribution of small deviations from x ^* , regardless of how small these deviations are. It is, therefore, suggested that when a metric space of pure strategies is involved, another topology, hence another stability condition, may prove more relevant to the process of natural selection. Concentrating on the case of a one dimensional continuous quantitative trait, we resort to the natural Maximum Shift Topology in which an ɛ-vicinity of the fixation on a pure strategy x ^* consists of all mixed population strategies with support which includes x ^* and is in the ɛ-neighborhood of x ^* . Under this topology, a relatively simple necessary and sufficient condition for replicator asymptotic stability, namely Continuous Replicator Stability (CRSS), is demonstrated. This condition is closely related to the static stability condition of Neighbor Invadability (Apaloo 1997), and slightly stronger than the condition of Continuous Stability (Eshel and Motro 1981). Received: 9 July 2002 / Revised version: 12 November 2002 / Published online: 19 March 2003 Key words or phrases: CSS – ESS – NIS – Strong uninvadability – Weak topology – Replicator dynamics – Long-term evolution – Continuous population games – Continuous replicator stability – Asymptotic stability     

Diastereoselective functionalisation of Baylis–Hillman adducts: a convenient approach to α-methyl-α-amino acids

The N-tosyl carbamates 4a–e, easily prepared starting from the Baylis–Hillman adducts 3a–e, underwent cyclization carried out with I₂/NIS in the presence of NaH, to give the corresponding 2-oxo-1,3-oxazolidines 5a–e in good yield and total stereoselection when the substituent at C-5 is Ar. After the removal of tosyl group, followed by the cleavage of the heterocyclic ring, the α-methyl-α-amino acids 8a,b and 10 were obtained in good yield as hydrochlorides.     

A new righteye flounder, Poecilopsetta multiradiata (Teleostei: Pleuronectiformes: Poecilopsettidae), from New Zealand and New Caledonia (South-West Pacific)

A new righteye flounder, Poecilopsetta multiradiata, is described from eight specimens (two males and six females) collected from deep waters (336–408 m) around New Zealand and New Caledonia (South-West Pacific). This new species is distinguished from its 14 congeners by the following combination of characters: high numbers of dorsal (70–73) and anal (58–62) fin rays, ca. 85–99 lateral-line scales, 31–32 caudal vertebrae, and a relatively shallow body depth of 36.9–41.9% SL.     

A new sea catfish,Netuma patriciae (Siluriformes: Ariidae), from the Philippines

A new sea catfish (Ariidae), Netuma patriciae, is described based on the holotype and nine paratypes, 127.3–303.4 mm in standard length, collected from Panay and Luzon islands, Philippines. The new species most resembles N. thalassina (Rüppell 1837), in sharing counts of anal-fin rays (17–19 in N. patriciae, 15–17 in N. thalassina) and a distinct dorsomedian head groove (longitudinal length of the groove, 33.7–40.1% of head length, 22.3–31.0%). However, the new species can be easily distinguished from the latter in having fused vomerine tooth patches, and more numerous free vertebrae (43–44 vs. 41–42 in N. thalassina). Although N. bilineata (Valenciennes in Cuvier and Valenciennes 1840a) has also fused vomerine tooth patches, the new species is distinguished by having fewer free vertebrae (43–44 vs. 47–51 in N. bilineata) and longer dorsomedian head groove (33.7–40.1% of head length vs. 0–9.7%). The new species can also be easily distinguished from both N. thalassina and N. bilineata by having a filamentous dorsal-fin ray (longest dorsal-fin ray 26.9–35.9% of standard length vs 20.3–23.3% in N. thalassina, 20.9–25.3% in N. bilineata, standard length > 127 mm) and a U-shaped junction of dorsomedian head ridge (vs. V-shaped in both species).     

A new armored searobin <Emphasis Type="Italic">Paraheminodus longirostralis</Emphasis> (Teleostei: Peristediidae) from New Caledonia

A new species of armored searobin, Paraheminodus longirostralis, is described from five specimens collected from New Caledonia at depths of 412–467 m. It is distinguishable from its three known congeners in having 34 bony plates in the upper lateral row, a forward-directed spine on each plate between the 23rd–26th and 31st–32nd plates in the upper lateral row, 6–7 + 1 + 20–21 = 27–28 gill rakers, an elongate body posterior to the anus (49.9–52.1% standard length), an elongate rostral projection (53.0–59.3% head length), short upper jaw (42.1–43.4% head length), an elongate pectoral fin (70.6–79.4% head length), and long preopercular spine (39.2–57.7% head length).     

Larval salinity tolerance of the South American salt-marsh crab, Neohelice (Chasmagnathus) granulata: physiological constraints to estuarine retention, export and reimmigration

The semiterrestrial crab Neohelice (=Chasmagnathus) granulata (Dana 1851) is a predominant species in brackish salt marshes, mangroves and estuaries. Its larvae are exported towards coastal marine waters. In order to estimate the limits of salinity tolerance constraining larval retention in estuarine habitats, we exposed in laboratory experiments freshly hatched zoeae to six different salinities (5–32‰). At 5‰, the larvae survived for a maximum of 2 weeks, reaching only exceptionally the second zoeal stage, while 38% survived to the megalopa stage at 10‰. Shortest development and negligible mortality occurred at all higher salt concentrations. These observations show that the larvae of N. granulata can tolerate a retention in the mesohaline reaches of estuaries, with a lower limit of ca. 10–15‰. Maximum survival at 25‰ suggests that polyhaline conditions rather than an export to oceanic waters are optimal for successful larval development of this species. In another experiment, we tested the capability of the last zoeal stage (IV) for reimmigration from coastal marine into brackish waters. Stepwise reductions of salinity during this stage allowed for moulting to the megalopa at 4–10‰. Although survival was at these conditions reduced and development delayed, these results suggest that already the zoea-IV stage is able to initiate the reimmigration into estuaries. After further salinity reduction, megalopae survived in this experiment for up to >3 weeks in freshwater, without moulting to juvenile crabs. In a similar experiment starting from the megalopa stage, successful metamorphosis occurred at 4–10‰, and juvenile growth continued in freshwater. Although these juvenile crabs showed significantly enhanced mortality and smaller carapace width compared to a seawater control, our results show that the late larval and early juvenile stages of N. granulata are well adapted for successful recruitment in brackish and even limnetic habitats.     

Earthworm secondary production and N flux in agroecosystems: a comparison of two approaches

Production was estimated for Aporrectodea spp. and Lumbricus spp. populations in corn agroecosystems with a 5-year history of manure or inorganic fertilizer applications during 1994–1995 and 1995–1996. Earthworm biomass and production were greater in manure than inorganic fertilizer plots, although biomass and production declined by about 50% between 1994–1995 and 1995–1996 due to unfavorable climatic conditions. Production was highest during the spring and autumn when soil temperatures were between 4 and 22°C. Production was higher in Lumbricus spp. than Aporrectodea spp. populations due to greater Lumbricus spp. biomass. Aporrectodea spp. production was 3.47–16.14 g ash-free dry weight (AFDW) m^–2 year^–1, while Lumbricus spp. production was 6.09–18.11 g AFDW m^–2 year^–1, depending on the fertilizer treatment and the method used to estimate production. However, production estimates from the instantaneous growth rate method were within 27% of the values calculated using the size-frequency method. Nitrogen flux through earthworms was used to estimate efficiency quotients. Net production efficiency (P/A) ranged from 0.64 to 0.76, assimilation efficiency (A/C) ranged from 0.1 to 0.3, and gross production efficiency (P/C) ranged from 0.06 to 0.22. Annual N flux through earthworm populations was higher in manure than inorganic fertilizer plots, and ranged from 2.95 to 5.47 g N m^–2 year^–1 in 1994–1995 and 1.76 to 2.92 g N m^–2 year^–1 in 1995–1996. The N flux through earthworms represented an amount equivalent to 16–30% of crop N uptake during 1994–1995 and 11–18% of crop N uptake during 1995–1996. We concluded that the effects of earthworms on N cycling in corn agroecosystems were substantial, and that N flux through earthworms was influenced significantly by fertilizer amendments. Received: 20 September 1999 / Accepted: 24 March 2000     

Hierarchic organization of globular proteins: Experimental studies on thermolysin

Previous studies from this laboratory have shown that the thermolysin fragment 121–316, comprising entirely the“all-α” COOH-terminal structural domain 158–316, as well as fragment 206–316 (fragment FII) are able to refold into a native-like, stable structure independently from the rest of the protein molecule. The present report describes conformational properties of fragments 228–316 and 255–316 obtained by chemical and enzymatic cleavage of fragment FII, respectively. These subfragments are able to acquire a stable conformation of native-like characteristics, as judged by quantitative analysis of secondary structure from far-ultra-violet circular dichroism spectra and immunochemical properties using rabbit anti-thermolysin antibodies. Melting curves of the secondary structure of the fragments show cooperativity with a temperature of half-denaturationT _mof 65–66°C. The results of this study provide evidence that it is possible to isolate stable supersecondary structures (folding units) of globular proteins and correlate well with predictions of subdomains of the COOH-terminal structural domain 158–316 of thermolysin.     

Two new soldierfishes (Beryciformes: Holocentridae:Myripristis) from Japan

Two species of the holocentrid fish genusMyripristis are described as new from Japan, both with a single pair of tooth patches outside the gape at tip of lower jaw, the third anal spine longer than the fourth, and 28–29 lateral-line scales:M. kochiensis from Kashiwa-jima, Kochi Prefecture, previously misidentified asM. murdjan, is distinct in having small scales in axil of pectoral fins, 32–36 gill rakers, interorbital space 4.45–5.1 in head, and lower jaw not strongly projecting when mouth closed;M. greenfieldi, from the Ryukyu Islands and Ogasawara Islands to Kochi Prefecture, previously misidentified asM. woodsi andM. randalli, is distinct in lacking scales in the pectoral axil, having 35–52 cteni on largest body scales, 43–47 gill rakers, and longest dorsal spine 2.0–2.25 in head.     

A new righteye flounder, Poecilopsetta pectoralis (Pleuronectiformes: Poecilopsettidae), from New Caledonia

A new righteye flounder, Poecilopsetta pectoralis, is described from 4 specimens (2 males, 2 females) collected from deep waters (510–580 m) around New Caledonia. The species is easily distinguished from 13 congeners by having 99–105 lateral line scales, a narrow interorbit [male: 37.00–40.25 in head length (HL); female: 47.33–50.71], long pectoral fin on ocular side (male: 0.89–0.91 in HL; female: 1.04–1.20), no scales from snout to anterior interorbital region, and 4 spots on caudal fin. Sexual dimorphism is evident in this species, showing males bearing the ocular-side pectoral fin longer than females, and it is the first example in Poecilopsetta.     

Intense cold and mortality in Castile-La Mancha (Spain): study of mortality trigger thresholds from 1975 to 2003

Studies on temperature–mortality time trends especially address heat, so that any contribution on the subject of cold is necessarily of interest. This study describes the modification of the lagged effects of cold on mortality in Castile-La Mancha from 1975 to 2003, with the novelty of also approaching this aspect in terms of mortality trigger thresholds. Cross-correlation functions (CCFs) were thus established with 15 lags, after application of ARIMA models to the mortality data and minimum daily temperatures (from November to March), and the results for the periods 1975–1984, 1985–1994 and 1995–2003 were then compared. In addition, daily mortality residuals for the periods 1975–1989 and 1990–2003 were related to minimum temperatures grouped in 2°C intervals, with a cold threshold temperature being obtained in cases where such residuals increased significantly (p < 0.05) with respect to the mean for the study period. A cold-related mortality trigger threshold of −3°C was obtained for Ciudad Real for the period 1990–2003. The significant number of lags (p < 0.05) in the CCFs declined every 10 years in Toledo (5–2–0), Cuenca (4–2–0), Albacete (4–3–0) and Ciudad Real (3–2–1). This meant that, while the trend in cold-related mortality trigger thresholds in the region could not be ascertained, it was possible to establish a reduction in the lagged effects of cold on mortality, attributable to the improvement in socio-economic conditions over the study period. Evidence was shown of the effects of cold on mortality, a finding that renders the adoption of preventive measures advisable in any case where intense cold is forecast.     

The charge-transfer complex between protochlorophyllide and NADPH: an intermediate in protochlorophyllide photoreduction

A hypothesis describing the mechanism of photoactive protochlorophyllide (P) photoreduction in vivo, relating mainly to the molecular nature of the intermediates, is proposed. The hypothesis is compatible with currently published experimental data. After illumination of etiolated barley leaves at 143 to 153 K, the absorption of P remains essentially unchanged, but a new absorption band at 690 nm is observed. Appearance of this new intermediate enables to distinguish between light and dark stages of the photoconversion reaction. When returned to the higher temperature in the dark, the treated leaves begin accumulating chlorophyllide (Chlide), concomitant with the disappearance of the 690-nm band. The decay time of the excited P (P^*) is estimated at 300 ps, which approximates the time constant of photoinduced electron transfer (ET). It is suggested that the charge-transfer complex (CTC) in its ground state (GS) (ground state of CTC formed by the partial (δ) electron transfer), i.e. (P^δ−•••H–D^δ+), between P and NADPH – the electron and proton donor (H–D) – accumulates in the following sequence: P^* + H–D → (P^*•••H–D)→[(P^*•••H–D)←(P⁻•••H–D⁺)] → ¹(P⁻•••H–D⁺)] → ³(P⁻•••H–D⁺) → (P^δ−•••H–D ^δ+), where an equilibrium state (ES) – [(P^*•••H–D)←(P⁻•••H–D⁺)] – with a lifetime of about 1 to 2 ns, exists between the local excited (LE) and ET states. The existence of a triplet ET state – ³(P⁻•••H–D⁺) – is proposed because the time interval between recording of the ES and appearance of the CTC GS (35–250 ns) does not fit the lifetime of the singlet excited complex (exciplex). It is feasible that apart from NADPH, other intermediate proton carriers are contemporaneously involved in the dark reaction (P^δ−•••H–D^δ+) → Chlide, because proton binding to the C₇–C₈ bond in vivo takes place in the trans-configuration. The hydride ion may approach the C₇–C₈ bond from one side by heterolytic fission and an additional proton, donated by the protein group, may be simultaneously added to this bond from the opposite side of the porphyrin nucleus surface. This revised version was published online in June 2006 with corrections to the Cover Date.     

<Emphasis Type="Italic">Epigonus cavaticus</Emphasis> (Teleostei: Perciformes), a new epigonid fish from Palau,western Central Pacific

A new epigonid fish, Epigonus cavaticus, is described on the basis of eight specimens (59.2–69.5 in standard length: SL) collected from a cave at depth 20 m, southern fringing reef of Ngemelis Island, Palau. The species differs from other congeners by having minute teeth on both jaws, no opercular spine, pyloric caeca 7–8, gill rakers 25–27, total pored lateral line scales 48–50, dorsal fin rays VII-I, 10–11 (mode VII-I, 10), pectoral fin rays 16, vertebrae 10 + 15, body depth 21.4–25.0% SL, pectoral fin length 22.7–24.6% SL, eye diameter 44.4–47.5% head length: HL, upper jaw length 40.2–42.5% HL. Four paratypes (63.7–66.8 mm SL) of the new species are female with mature gonads, it is the smallest in size at sexual maturity among the congeners.     

Chemical synthesis of15N-labeled analogues of zervamicin IIB

Analogues of 16-membered peptide antibiotic zervamicin IIB with the Gln³ and Gln¹¹ residues¹⁵N-labeled at the Cα-atoms were synthesized by coupling the antibiotic segments (1–4), (5–9), and (10–16). In turn, these were prepared by a stepwise chain elongation in solution starting from theirC-termini using benzotriazol-1-yloxy-tris(dimethylamino)phosphonium hexafluorophosphate (BOP) as an activating agent. The sterically hindered 2-aminoisobutyric acid was introduced by the BOP-dimethylaminopyridine system with the preactivation of the carboxyl component. The segment condensation was performed with the use of the 6-trifluoromethylbenzotriazol-1-yloxy-tris(pyrrolidino)phosphonium hexafluorophosphate activating reagent. The homogeneity of the resulting zervamicin analogues was confirmed by HPLC, and their structures were proved by NMR spectroscopy and FAB mass spectrometry.