Toxin Gene Contents and Activity of <Emphasis Type="Italic">Bacillus thuringiensis</Emphasis> Strains Against Two Sugarcane Borer Species, <Emphasis Type="Italic">Diatraea saccharalis</Emphasis> (F.) and <Emphasis Type="Italic">D. flavipennella</Emphasis> (Box)

Bacillus thuringiensis (Berliner) bears essential characteristics in the control of insect pests, such as its unique mode of action, which confers specificity and selectivity. This study assessed cry gene contents from Bt strains and their entomotoxicity against Diatraea saccharalis (F.) and Diatraea flavipennella (Box) (Lepidoptera: Crambidae). Bioassays with Bt strains were performed against neonates to evaluate their lethal and sublethal activities and were further analyzed by PCR, using primers to identify toxin genes. For D. saccharalis and D. flavipennella, 16 and 18 strains showed over 30% larval mortality in the 7th day, respectively. The LC₅₀ values of strains for D. saccharalis varied from 0.08 × 10⁵ (LIIT-0105) to 4104 × 10⁵ (LIIT-2707) spores + crystals mL^?1. For D. flavipennella, the LC₅₀ values of strains varied from 0.40 × 10⁵ (LIIT-2707) to 542 × 10⁵ (LIIT-2109) spores + crystals mL^?1. For the LIIT-0105 strain, which was the most toxic to D. saccharalis, the genes cry1Aa, cry1Ab, cry1Ac, cry1B, cry1C, cry1D, cry1F, cry1I, cry2Aa, cry2Ab, cry8, and cry9C were detected, whereas for the strain LIIT-2707, which was the most toxic to D. flavipennella, detected genes were cry1Aa, cry1Ab, cry1Ac, cry1B, cry1D, cry1F, cry1I, cry2Aa, cry2Ab, and cry9. The toxicity data and toxin gene content in these strains of Bt suggest a great variability of activity with potential to be used in the development of novel biopesticides or as source of resistance genes that can be expressed in plants to control pests.     

Natural succession of grassland on the Loess Plateau of China affects multifractal characteristics of soil particle-size distribution and soil nutrients

Knowledge of the changes of soil properties and soil particle-size distribution (PSD) is important for quantifying soil quality and dynamics during natural succession. We applied multifractal theory to characterize the PSDs in five soil layers of nine croplands abandoned for less than 1 year, 2, 5, 7, 10, 15, 20, 30 and 35 years and evaluated the changes of soil organic carbon (SOC), total nitrogen (total N), total phosphorus (total P), soil texture and multifractal parameters [capacity dimension (D ₀), entropy dimension (D ₁), correlation dimension (D ₂) and Hölder exponent of order zero (α ₀)] during natural succession on the Loess Plateau of China. Rényi spectra (D _q ) and singularity spectra f(α) characterized the PSDs well and sensitively reflected the changes of heterogeneity of the surface soil (0–20 cm) during natural succession. SOC, total N and clay contents and the multifractal parameters increased significantly by the late stage of succession, mostly in the topsoil (0–10 cm). Natural succession thus effectively improved the soil physicochemical properties on the Loess Plateau of China, even though it was time-consuming. SOC and total N contents decreased with depth throughout the natural succession, but the multifractal parameters were higher in the topsoil only in the late stage. D ₁ and D ₂ were strongly and positively correlated with SOC and total N contents in the surface layers and with fine particles in all layers, suggesting that D ₁ and D ₂ may be sensitive and practical indices for quantifying changes in soil properties and erosion.     

Hardness maximization or equalization? New insights and quantitative relations between hardness increase and bond dissociation energy

It has been overlooked that the change of hardness, η, upon bonding is intimately connected to thermochemical cycles, which determine whether hardness is increased according to Pearson’s “maximum hardness principle” (MHP) or equalized, as expected by Datta’s “hardness equalization principle” (HEP). So far the performances of these likely incompatible “structural principles” have not been compared. Computational validations have been inconclusive because the hardness values and even their qualitative trends change drastically and unsystematically at different levels of theory. Here I elucidate the physical basis of both rules, and shed new light on them from an elementary experimental source. The difference, Δη = η _mol – <η _at>, of the molecular hardness, η _mol, and the averaged atomic hardness, <η _at>, is determined by thermochemical cycles involving the bond dissociation energies D of the molecule, D ⁺ of its cation, and D ^? of its anion. Whether the hardness is increased, equalized or even reduced is strongly influenced by ΔD = 2D – D ⁺  ? D ^?. Quantitative expressions for Δη are obtained, and the principles are tested on 90 molecules and the association reactions forming them. The Wigner-Witmer symmetry constraints on bonding require the valence state (VS) hardness, η _VS, instead of the conventional ground state (GS) hardness, η _GS. Many intriguingly “unpredictable” failures and systematic shortcomings of said “principles” are understood and overcome for the first time, including failures involving exotic and/or challenging molecules, such as Be_2, B₂, O₃, and transition metal compounds. New linear relationships are discovered between the MHP hardness increase Δη _VS and the intrinsic bond dissociation energy D _i . For bond formations, MHP and HEP are not compatible, and HEP does not qualify as an ordering rule.     

Bacterial metabolites from intra- and inter-species influencing thermotolerance: the case of <Emphasis Type="Italic">Bacillus cereus</Emphasis> and <Emphasis Type="Italic">Geobacillus stearothermophilus</Emphasis>

Bacterial metabolites with communicative functions could provide protection against stress conditions to members of the same species. Yet, information remains limited about protection provided by metabolites in Bacillus cereus and inter-species. This study investigated the effect of extracellular compounds derived from heat shocked (HS) and non-HS cultures of B. cereus and Geobacillus stearothermophilus on the thermotolerance of non-HS vegetative and sporulating B. cereus. Cultures of B. cereus and G. stearothermophilus were subjected to HS (42 or 65 °C respectively for 30 min) or non-HS treatments. Cells and supernatants were separated, mixed in a combined array, and then exposed to 50 °C for 60 min and viable cells determined. For spores, D values (85 and 95 °C) were evaluated after 120 h. In most cases, supernatants from HS B. cereus cultures added to non-HS B. cereus cells caused their thermotolerance to increase (D ₅₀ 12.2–51.9) when compared to supernatants from non-HS cultures (D ₅₀ 7.4–21.7). While the addition of supernatants from HS and non-HS G. stearothermophilus cultures caused the thermotolerance of non-HS cells from B. cereus to decrease initially (D ₅₀ 3.7–7.1), a subsequent increase was detected in most cases (D ₅₀ 18–97.7). In most cases, supernatants from sporulating G. stearothermophilus added to sporulating cells of B. cereus caused the thermotolerance of B. cereus 4810 spores to decline, whereas that of B. cereus 14579 increased. This study clearly shows that metabolites in supernatants from either the same or different species (such as G. stearothermophilus) influence the thermotolerance of B. cereus.     

Specific light uptake rates can enhance astaxanthin productivity in <Emphasis Type="Italic">Haematococcus lacustris</Emphasis>

Lumostatic operation was applied for efficient astaxanthin production in autotrophic Haematococcus lacustris cultures using 0.4-L bubble column photobioreactors. The lumostatic operation in this study was performed with three different specific light uptake rates (q _e) based on cell concentration, cell projection area, and fresh weight as one-, two- and three-dimensional characteristics values, respectively. The q _e value from the cell concentration (q _e1D) obtained was 13.5 × 10^?8 μE cell^?1 s^?1, and the maximum astaxanthin concentration was increased to 150 % compared to that of a control with constant light intensity. The other optimum q _e values by cell projection area (q _e2D) and fresh weight (q _e3D) were determined to be 195 μE m^?2 s^?1 and 10.5 μE g^?1 s^?1 for astaxanthin production, respectively. The maximum astaxanthin production from the lumostatic cultures using the parameters controlled by cell projection area (2D) and fresh weight (3D) also increased by 36 and 22 % over that of the controls, respectively. When comparing the optimal q _e values among the three different types, the lumostatic cultures using q _e based on fresh weight showed the highest astaxanthin productivity (22.8 mg L^?1 day^?1), which was a higher level than previously reported. The lumostatic operations reported here demonstrated that more efficient and effective astaxanthin production was obtained by H. lacustris than providing a constant light intensity, regardless of which parameter is used to calculate the specific light uptake rate.     

Improving the acidic stability of <Emphasis Type="Italic">Staphylococcus aureus</Emphasis> α-acetolactate decarboxylase in <Emphasis Type="Italic">Bacillus subtilis</Emphasis> by changing basic residues to acidic residues

The α-acetolactate decarboxylase (ALDC) can reduce diacetyl fleetly to promote mature beer. A safe strain Bacillus subtilis WB600 for high-yield production of ALDC was constructed with the ALDC gene saald from Staphylococcus aureus L3-15. SDS-PAGE analysis revealed that S. aureus α-acetolactate decarboxylase (SaALDC) was successfully expressed in recombinant B. siutilis strain. The enzyme SaALDC was purified using Ni-affinity chromatography and showed a maximum activity at 45 °C and pH 6.0. The values of K _m and V _max were 17.7 μM and 2.06 mM min^?1, respectively. Due to the unstable property of SaALDC at low pH conditions that needed in brewing process, site-directed mutagenesis was proposed for improving the acidic stability of SaALDC. Homology comparative modeling analysis showed that the mutation (K52D) gave rise to the negative-electrostatic potential on the surface of protein while the numbers of hydrogen bonds between the mutation site (N43D) and the around residues increased. Taken together the effect of mutation N43D-K52D, recombinant SaALDC_N43D-K52D showed dramatically improved acidic stability with prolonged half-life of 3.5 h (compared to the WT of 1.5 h) at pH 4.0. In a 5-L fermenter, the recombinant B. subtilis strain that could over-express SaALDC_N43D-K52D exhibited a high yield of 135.8 U mL^?1 of SaALDC activity, about 320 times higher comparing to 0.42 U mL^?1 of S. aureus L3-15. This work proposed a  strategy for improving the acidic stability of SaALDC in the  B. subtilis host.     

Probiotic Effect of <Emphasis Type="Italic">Bacillus licheniformis fb11</Emphasis> on the Digestive Efficiency and Growth Performance in Juvenile <Emphasis Type="Italic">Chitala chitala</Emphasis> (Hamilton, 1822)

Five isocaloric (430 kcal 100 g^?1), isonitrogenous (40% CP) experimental diets were formulated with different concentrations of Bacillus licheniformis fb11 probionts (isolated from the gut of Chitala chitala) viz. Control (without probionts), 5 × 10⁴ CFU g^?1 (D₁), 5 × 10⁵ CFU g^?1 (D₂), 5 × 10⁶ CFU g^?1 (D₃), 5 × 10⁷ CFU g^?1 (D₄), 5 × 10⁸ CFU g^?1 (D₅) to evaluate its efficiency in C. chitala juvenile. The best growth performance, feed utilisation, specific α-amylase, total protease and lipase activity were observed with the diet D₃ (P < 0.05). The lowest Presumptive Pseudomonas Count, Motile Aeromonad Count, Total Coliform Count was observed for D₃ (P < 0.05) on 90th day of trial. Two uppermost values were achieved in case of crude protein for D₃ and D₂ (P > 0.05). The highest lipid content (12.12 ± 0.4 g 100 g^?1) was found for D₅ (P < 0.05). The highest gross energy (18.75 ± 0.21 MJ 100 g^?1) of carcass was recorded for D₃. Thus B. licheniformis fb11 at the concentration 5 × 10⁶ CFU g^?1 as probiotic supplement promoted growth, digestion in C. chitala juvenile significantly by modulating intestinal microflora.     

The nitrite reductase activity of horse heart carboxymethylated-cytochrome <Emphasis Type="Italic">c</Emphasis> is modulated by cardiolipin

Horse heart carboxymethylated cytc (CM-cytc) displays myoglobin-like properties. Here, the effect of cardiolipin (CL) liposomes on the nitrite reductase activity of ferrous CM-cytc [CM-cytc-Fe(II)], in the presence of sodium dithionite, is reported between pH 5.5 and 7.6, at 20.0 °C. Cytc-Fe(II) displays a very low value of the apparent second-order rate constant for the NO₂ ^?-mediated conversion of cytc-Fe(II) to cytc-Fe(II)-NO [k _on = (7.3 ± 0.7) × 10^?2 M^?1 s^?1; at pH 7.4], whereas the value of k _on for NO₂ ^? reduction by CM-cytc-Fe(II) is 1.1 ± 0.2 M^?1 s^?1 (at pH 7.4). CL facilitates the NO₂ ^?-mediated nitrosylation of CM-cytc-Fe(II) in a dose-dependent manner, the value of k _on for the NO₂ ^?-mediated conversion of CL–CM-cytc-Fe(II) to CL–CM-cytc-Fe(II)-NO (5.6 ± 0.6 M^?1 s^?1; at pH 7.4) being slightly higher than that for the NO₂ ^?-mediated conversion of CL–cytc-Fe(II) to CL–cytc-Fe(II)-NO (2.6 ± 0.3 M^?1 s^?1; at pH 7.4). The apparent affinity of CL for CM-cytc-Fe(II) is essentially pH independent, the average value of B being (1.3 ± 0.3) × 10^?6 M. In the absence and presence of CL liposomes, the nitrite reductase activity of CM-cytc-Fe(II) increases linearly on lowering pH and the values of the slope of the linear fittings of Log k _on versus pH are ?1.05 ± 0.07 and ?1.03 ± 0.03, respectively, reflecting the involvement of one proton for the formation of the transient ferric form, NO, and OH^?. These results indicate that Met80 carboxymethylation and CL binding cooperate in the stabilization of the highly reactive heme-Fe atom of CL–CM-cytc.     

Parasitism Rate of <Emphasis Type="Italic">Habrobracon hebetor</Emphasis> to <Emphasis Type="Italic">Ephestia kuehniella</Emphasis> Larvae: Residual Effect of Deltamethrin,Fenvalerate and Azadirachtin

Sublethal concentrations of chemical insecticides may cause changes in some behavioral characteristics of natural enemies such as functional responses. The residual effect of three synthetic insecticides including deltamethrin, fenvalerate and azadirachtin were studied on functional response of Habrobracon hebetor Say to Ephestia kuehniella Zeller larvae. Seven host densities (2, 4, 8, 16, 32, 64 and 96) were used during a 24 h period. The resulting data were appropriately fit to Type II functional response models in all treatments: (1) control (0.0916 h^?1; and T _h  = 0.2011 h); (2) deltamethrin (a = 0.0839 h^?1; and T _h  = 0.3560 h); (3) fenvalerate (a = 0.0808 h^?1 and T _h  = 0.3623 h); and (4) azadirachtin (a = 0.0900 h^?1 and T _h  = 0.2042 h). Maximum theoretical parasitism rate (T/T _h ) was 119.34 estimated for control wasps. There was no significant difference between the values of attack rates (a and a + D _a ) in all treatments while the handling time was statistically affected in female wasps treated with fenvalerate. Our findings will be useful in safe application of these insecticides in pest management programmes.     

Biological Data on <Emphasis Type="Italic">Anovia punica</Emphasis> Gordon (Coleoptera: Coccinellidae), a Predator of <Emphasis Type="Italic">Crypticerya multicicatrices</Emphasis> Kondo &amp; Unruh (Hemiptera: Monophlebidae)

The coccinellid beetle Anovia punica Gordon (Coleoptera: Coccinellidae: Noviini) is an important predator of the Colombian fluted scale, Crypticerya multicicatrices Kondo & Unruh (Hemiptera: Monophlebidae). In order to gather information on the biological traits of A. punica, we conducted a series of studies, including of the developmental time of each life history stage, estimation of life table parameters, and predation rates under laboratory conditions [25.1 ± 1.6°C, with 70.5 ± 7.3% RH, and natural light regime, approx. 12:12 (L:D) h]. Developmental stages of A. punica were categorized as follows: egg stage, four larval instars, prepupal instar, pupal instar, and adult. Developmental time from egg to adult emergence averaged 29.41 ± 1.85 days, and 47.6% of the eggs developed to adulthood. Female and male survival was 94.42 and 90 days, respectively. Life table parameters show that one female of A. punica is replaced by 86 females (R ₀), the intrinsic growth rate (r _m ) was 0.1115, the average generation time (T) was 40 days, and the doubling time (D _t ) was 6.2 days. The life table parameters suggest that A. punica can be used as a potential predator of C. multicicatrices and, more importantly, provided baseline information for a mass-rearing protocol. This is the first detailed study on the biology of A. punica that reports the potential of this predator as a biological control agent for scale insects of the tribe Iceryini.     

Spectroscopic evidence for an all-ferrous [4Fe–4S]<Superscript>0</Superscript> cluster in the superreduced activator of 2-hydroxyglutaryl-CoA dehydratase from <Emphasis Type="Italic">Acidaminococcus </Emphasis><Emphasis Type="Italic">fermentans</Emphasis>

The key enzyme of the fermentation of glutamate by Acidaminococcus fermentans, 2-hydroxyglutarylcoenzyme A dehydratase, catalyzes the reversible syn-elimination of water from (R)-2-hydroxyglutaryl-coenzyme A, resulting in (E)-glutaconylcoenzyme A. The dehydratase system consists of two oxygen-sensitive protein components, the activator (HgdC) and the actual dehydratase (HgdAB). Previous biochemical and spectroscopic studies revealed that the reduced [4Fe–4S]⁺ cluster containing activator transfers one electron to the dehydratase driven by ATP hydrolysis, which activates the enzyme. With a tenfold excess of titanium(III) citrate at pH 8.0 the activator can be further reduced, yielding about 50% of a superreduced [4Fe–4S]⁰ cluster in the all-ferrous state. This is inferred from the appearance of a new Mössbauer spectrum with parameters δ = 0.65 mm/s and ΔE _Q = 1.51–2.19 mm/s at 140 K, which are typical of Fe(II)S₄ sites. Parallel-mode electron paramagnetic resonance (EPR) spectroscopy performed at temperatures between 3 and 20 K showed two sharp signals at g = 16 and 12, indicating an integer-spin system. The X-band EPR spectra and magnetic Mössbauer spectra could be consistently simulated by adopting a total spin S _t = 4 for the all-ferrous cluster with weak zero-field splitting parameters D = ?0.66 cm^?1 and E/D = 0.17. The superreduced cluster has apparent spectroscopic similarities with the corresponding [4Fe–4S]⁰ cluster described for the nitrogenase Fe-protein, but in detail their properties differ. While the all-ferrous Fe-protein is capable of transferring electrons to the MoFe-protein for dinitrogen reduction, a similar physiological role is elusive for the superreduced activator. This finding supports our model that only one-electron transfer steps are involved in dehydratase catalysis. Nevertheless we discuss a common basic mechanism of the two diverse systems, which are so far the only described examples of the all-ferrous [4Fe–4S]⁰ cluster found in biology.     

Ammonium,nitrate, and urea play different roles for lipid accumulation in the nervonic acid—producing microalgae <Emphasis Type="Italic">Mychonastes afer</Emphasis> HSO-3-1

Producing valuable coproducts from oleaginous microalgae is an option to reduce the total cost of biofuel production. Here, the influence of nitrogen sources on biomass yield and lipid accumulation of a newly identified oleaginous green microalgal strain, Mychonastes afer HSO-3-1, was evaluated. Carbon assimilation and the following lipid biosynthesis of M. afer were inhibited to some extent under weak acidic conditions (6 < pH < 7) and any of the tested nitrogen source. The highest lipid productivity of 50.7 mg L^?1 day^?1 was achieved with a 17.6 mM nitrogen supplement in the form of urea. The cell polar lipid content was significantly higher than triacylglycerol (TAG), and saturated palmitic acid (C16:0) occupied a dominant position in the fatty acid profiles while culturing M. afer in acidic medium with NH₄ ⁺ as the nitrogen source. Under neutral conditions, the lipid productivities of M. afer cultivated in media containing 17.6 mM of NaNO₃, NH₄Cl, and NH₄NO₃ were 76.2, 77.5, and 79.0 mg L^?1 day^?1, respectively. The greatest TAG content (58.56%) of total lipids was obtained when NaNO₃ was used as the nitrogen source. There was no significant difference in the fatty acid composition of M. afer cells when they were cultivated in neutral media supplemented with NaNO₃, urea, NH₄Cl, and NH₄NO₃. Therefore, NH₄ ⁺ was not a suitable nitrogen source for M. afer cultivation due to the additional labor, working procedures, and alkali required to adjust the medium pH. Considering that using urea as nitrogen source could reduce the cost of nutrient salts substantially and urea can be taken up and utilized by most microalgae, it is a preferred nitrogen source. The major properties of biodiesel derived from M. afer HSO-3-1 met biodiesel quality, and nervonic acid concentrations remained at approximately 3.0% of total fatty acids.     

<Emphasis Type="Italic">In vitro</Emphasis> propagation of endangered <Emphasis Type="Italic">Mammillaria</Emphasis> genus (Cactaceae) species and genetic stability assessment using SSR markers

In vitro propagation protocols were established for endangered species of cacti Mammillaria hernandezii, M. dixanthocentron, and M. lanata. In vitro-germinated seedlings were used as the explant source. Three explant types were evaluated as apical, basal, and lateral stem sections. Shoot multiplication was achieved using Murashige and Skoog (MS) medium supplemented with benzyladenine, kinetin, meta-topolin, and thidiazuron in equimolar concentrations (0.0, 0.4, 1.1, 2.2, 4.4, and 8.9 μM). Shoot regeneration was obtained primarily in the lateral stem section explants. In M. hernandezii, an average of 7.4 shoots was regenerated in MS medium with 2.2 μM meta-topolin. M. dixanthocentron and M. lanata averaged 16.7 and 17.9 shoots/explant, respectively, in MS medium supplemented with 1.1 μM meta-topolin. Rooting occurred in MS medium without growth regulators. Three in vitro culture cycles were performed to validate the propagation protocols and to verify genetic stability. Shoots were collected in each cycle and genomic DNA was extracted. Amplified microsatellites were used to compare each genotype with its respective donor plant. Polymorphic information content analysis showed low levels of intra-clonal polymorphisms—M. hernandezii 0.04 and M. dixanthocentron and M. lanata both 0.12. More than 95% of the plants were successfully acclimatized in the greenhouse. After 12 months, plants of M. hernandezii reached the flowering stage; M. dixanthocentron and M. lanata flowered at 24 mo.     

Characterization of the interactions between coumarin-derivatives and acetylcholinesterase: Examination by NMR and docking simulations

Alzheimer’s disease (AD) is one of the most common forms of dementia and a significant threat to the elderly populations, especially in the Western world. The rapid hydrolysis of the principal neurotransmitter into choline and acetate by acetylcholinesterase (AChE) at synapses causes the loss of cognitive response that becomes the real cause of AD. Therefore, inhibition of AChE is the most fundamental therapy among currently available treatments for AD. In this context, we designed and performed molecular recognitions studies of coumarin-based inhibitors towards AChE. STD NMR and Tr-NOESY applications were utilized to evaluate the binding epitope, the dissociation constant (K_D) and bound conformations of these inhibitors within this inhibitor-AChE complex. Compound 1, which has a similar inhibition activity to tacrine (a current drug) led in this study as a stronger binder with K_D?=?30 μM ,even greater than tacrine (K_D?=?140 μM). Moreover, docking simulations mimic NMR results and provided evidence of synchronizing binding of compound 1 with three sites; the peripheral anionic site, the bottom of the gorge, and the catalytic site. Therefore, we envisioned from our experimental and theoretical results that coumarin-based inhibitors containing a piperidinyl scaffold might be a potential drug candidates for AD in the future.     

Characterization of a common wheat (<Emphasis Type="Italic">Triticum aestivum</Emphasis> L.) high-tillering dwarf mutant

Key message

A novel high-tillering dwarf mutant in common wheat Wangshuibai was characterized and mapped to facilitate breeding for plant height and tiller and the future cloning of the causal gene.

Abstract

Tiller number and plant height are two major agronomic traits in cereal crops affecting plant architecture and grain yield. NAUH167, a mutant of common wheat landrace Wangshuibai induced by ethylmethyl sulfide (EMS) treatment, exhibits higher tiller number and reduced plant height. Microscope observation showed that the dwarf phenotype was attributed to the decrease in the number of cells and their length. The same as the wild type, the mutant was sensitive to exogenous gibberellins. Genetic analysis showed that the high-tillering number and dwarf phenotype were related and controlled by a partial recessive gene. Using a RIL_2:6 population derived from the cross NAUH167/Sumai3, a molecular marker-based genetic map was constructed. The map consisted of 283 loci, spanning a total length of 1007.98 cM with an average markers interval of 3.56 cM. By composite interval mapping, a stable major QTL designated QHt.nau-2D controlling both traits, was mapped to the short arm of chromosome 2D flanked by markers Xcfd11 and Xgpw361. To further map the QHt.nau-2D loci, another population consisted of 180 F₂ progeny from a cross 2011I-78/NAUH167 was constructed. Finally, QHt.nau-2D was located within a genetic region of 0.8 cM between markers QHT239 and QHT187 covering a predicted physical distance of 6.77 Mb. This research laid the foundation for map-based cloning of QHt.nau-2D and would facilitate the characterization of plant height and tiller number in wheat.

     

Cloning and characterization of two distinct water-forming NADH oxidases from <Emphasis Type="Italic">Lactobacillus pentosus</Emphasis> for the regeneration of NAD

Two uncharacterized nicotinamide adenine dinucleotide (NADH) oxidases (named as LpNox1, LpNox2) from Lactobacillus pentosus ATCC 8041 were cloned and overexpressed in Escherichia coli BL21 (DE3). The sequence analysis revealed that the two enzymes are water-forming Noxs with 64 % and 52 % identity to LbNox from Lactobacillus brevis DSM 20054. The optimal pH and temperature of the purified LpNox1 and LpNox2 were 7.0 and 8.0 and 35 and 40 °C, respectively, with K _M of 99.0 μM (LpNox1) and 27.6 μM (LpNox2), and yielding catalytic efficiency k _cat/K _M of 1.0 and 0.2 μM^?1 s^?1, respectively. Heat inactivation studies revealed that the two enzymes are relatively instable. The application of LpNox1 for the regeneration of NAD⁺ was demonstrated by coupling with a glycerol dehydrogenase-catalyzed oxidation of glycerol to 1,3-dihydroxyacetone. The characteristics of the LpNox1 could prove to be of interest in industrial application such as NAD⁺ regeneration in dehydrogenase-catalyzed oxidations.     

Feed-backs between genetic structure and perturbation-driven decline in seagrass (<Emphasis Type="Italic">Posidonia oceanica</Emphasis>) meadows

We explored the relationships between perturbation-driven population decline and genetic/genotypic structure in the clonal seagrass Posidonia oceanica, subject to intensive meadow regression around four Mediterranean fish-farms, using seven specific microsatellites. Two meadows were randomly sampled (40 shoots) within 1,600 m² at each site: the “impacted” station, 5–200 m from fish cages, and the “control” station, around 1,000 m downstream further away (considered a proxy of the pre-impact genetic structure at the site). Clonal richness (R), Simpson genotypic diversity (D*) and clonal sub-range (CR) were highly variable among sites. Nevertheless, the maximum distance at which clonal dispersal was detected, indicated by CR, was higher at impacted stations than at the respective control station (paired t-test: P < 0.05, N = 4). The mean number of alleles (Â) and the presence of rare alleles ( _r) decreased at impacted stations (paired t-test: P < 0.05, and P < 0.02, respectively, N = 4). At a given perturbation level (quantified by the organic and nutrient loads), shoot mortality at the impacted stations significantly decreased with CR at control stations (R ² = 0.86, P < 0.05). Seagrass mortality also increased with  (R ² = 0.81, P < 0.10), R (R ² = 0.96, P < 0.05) and D* (R ² = 0.99, P < 0.01) at the control stations, probably because of the negative correlation between those parameters and CR. Therefore, the effects of clonal size structure on meadow resistance could play an important role on meadow survival. Large genotypes of P. oceanica meadows thus seem to resist better to fish farm-derived impacts than little ones. Clonal integration, foraging advantage or other size-related fitness traits could account for this effect.     

Unspecific histological and hematological alterations in anadromous and resident <Emphasis Type="Italic">Salvelinus malma</Emphasis> induced by volcanogenic pollution

Knowledge of larval dispersal and connectivity in coral reef species is crucial for understanding population dynamics, resilience, and evolution of species. Here, we use ten microsatellites and one mitochondrial marker (cytochrome b) to investigate the genetic population structure, genetic diversity, and historical demography of the powder-blue tang Acanthurus leucosternon across more than 1000 km of the scarcely studied Eastern African region. The global AMOVA results based on microsatellites reveal a low but significant F _ST value (F _ST = 0.00252 P < 0.001; D _EST = 0.025 P = 0.0018) for the 336 specimens sampled at ten sample sites, while no significant differentiation could be found in the mitochondrial cytochrome b dataset. On the other hand, pairwise F _ST, PCOA, and hierarchical analysis failed to identify any genetic breaks among the Eastern African populations, supporting the hypothesis of genetic homogeneity. The observed genetic homogeneity among Eastern African sample sites can be explained by the lengthy post-larval stage of A. leucosternon, which can potentiate long-distance dispersal. Tests of neutrality and mismatch distribution signal a population expansion during the mid-Pleistocene period.     

Genetic population structure and low genetic diversity in the over-exploited sea cucumber <Emphasis Type="Italic">Holothuria edulis</Emphasis> Lesson, 1830 (Echinodermata: Holothuroidea) in Okinawa Island

Understanding genetic connectivity is fundamental for ecosystem-based management of marine resources. Here we investigate the metapopulation structure of the edible sea cucumber Holothuria edulis Lesson, 1830 across Okinawa Island, Japan. This species is of economic and ecological importance and is distributed from the Red Sea to Hawai‘i. We examined sequence variation in fragments of mitochondrial cytochrome oxidase subunit I (COI) and 16S ribosomal RNA (16S), and nuclear histone (H3) at six locations across Okinawa Island. We found higher haplotype diversity for mtDNA (COI: Hd = 0.69 and 16S: Hd = 0.67) and higher heterozygosity of nDNA (H3: H _E = 0.39) in populations from the west coast of Okinawa compared to individuals from populations on the east coast (COI: Hd = 0.40; 16S: Hd = 0.21; H3: H _E = 0.14). Overall population structure was significant (AMOVA results for COI: Φ _ST = 0.49, P < 0.0001; 16S: Φ _ST = 0.34, P < 0.0001; H3: Φ _ST = 0.12, P < 0.0001). One population in the east, Uruma, showed elevated pairwise Φ _ST values in comparisons with all other sites and a marked reduction of genetic diversity (COI: Hd = 0.25 and 16S: Hd = 0.24), possibly as a consequence of a shift to a more dominant asexual reproduction mode. Recent reports have indicated that coastal development in this area influences many marine organisms, and ecosystem degradation in this location could cause the observed decrease of genetic diversity and isolation of H. edulis in Uruma. Our study should provide valuable data to help with the urgently needed management of sea cucumber populations in Okinawa, and indicates particular attention needs to be paid to vulnerable locations.