Bridging the gap between chemistry, physiology, and evolution: quantifying the functionality of sperm whale myoglobin mutants

This work merges a large set of previously reported thermochemical data for myoglobin (Mb) mutants with a physiological model of O₂-transport and -storage. The model allows a quantification of the functional proficiency of myoglobin (Mb) mutants under various physiological conditions, i.e. O₂-consumption rate resembling workload, O₂ partial pressure resembling hypoxic stress, muscle cell size, and Mb concentration, resembling different organism-specific and compensatory variables. We find that O₂-storage and -transport are distinct functions that rank mutants and wild type differently depending on O₂ partial pressure. Specifically, the wild type is near-optimal for storage at all conditions, but for transport only at severely hypoxic conditions. At normoxic conditions, low-affinity mutants are in fact better O₂-transporters because they still have empty sites for O₂, giving rise to a larger [MbO₂] gradient (more varying saturation curve). The distributions of functionality reveal that many mutants are near-neutral with respect to function, whereas only a few are strongly affected, and the variation in functionality increases dramatically at lower O₂ pressure. These results together show that conserved residues in wild type (WT) Mb were fixated under a selection pressure of low P_O2.     

Proteome comparison of hypopharyngeal gland development between Italian and royal jelly producing worker honeybees (Apis mellifera L.)

The hypopharyngeal gland (HG) of the honeybee (Apis mellifera L.) produces royal jelly (RJ) that is essential to feed and raise broods and queens. A strain of bees (high royal jelly producing bee, RJb) has been selected for its high RJ production, but the mechanisms of its higher yield are not understood. In this study, we compared HG acini size, RJ production, and protein differential expressions between the RJb and nonselected honeybee (Italian bee, ITb) using proteomics in combination with an electron microscopy, Western blot, and quantitative real-time PCR (qRT-PCR). Generally, the HG of both bees showed age-dependent changes in acini sizes and protein expression as worker behaviors changed from brood nursing to nectar ripening, foraging, and storage activities. The electron microscopic analysis revealed that the HG acini diameter of the RJb strain was large and produced 5 times more RJ than the ITb, demonstrating a positive correlation between the yield and HG acini size. In addition, the proteomic analysis showed that RJb significantly upregulated a large group of proteins involved in carbohydrate metabolism and energy production, those involved in protein biosynthesis, development, amino acid metabolism, nucleotide and fatty acid, transporter, protein folding, cytoskeleton, and antioxidation, which coincides with the fact that the HGs of the RJb strain produce more RJ than the ITb strain that is owing to selection pressure. We also observed age-dependent major royal jelly proteins (MRJPs) changing both in form and expressional intensity concurrent with task-switching. In addition to MRJPs, the RJb overexpressed proteins such as enolase and transitional endoplasmic reticulum ATPase, protein biosynthesis, and development proteins compared to the ITb strain to support its large HG growth and RJ secretion. Because of selection pressure, RJb pursued a different strategy of increased RJ production by involving additional proteins compared to its original counterpart ITb. To our knowledge, this morphological and proteomic comparison study on the HG of the two strains of worker honeybees associated with their age-dependent division of labor is the first of its kind. The study provided not only the quantity and quality differences in the HG from the RJb and the ITb, but also addressed the cellular and behavioral biology development question of how the RJb strain can produce RJ more efficiently than its wild type strain (ITb).     

Lipid peroxidation in buffalo (Bubalus bubalis L.) spermatozoa: effect of added vitamin C and glucose

Addition of 2.5 mM vitamin C or 40 mM of glucose to washed buffalo spermatozoan suspensions in Ca2(+)-free Kreb's Ringer Hanseliet saline buffer (pH 7.0) resulted in significant lower malonaldehyde concentration and higher spermatozoan motility and liver spermatozoa compared to control levels after 45 min of aerobic incubation at 37 degrees C or pre-incubation levels.     

The spermatodesm of Cloeon dipterum (L.): fine structure and sperm movement

Sperm bundles of the ephemeropteran Cloeon dipterum (L.) were found in the seminal vesicle. They are apically protected by cyst cells and sperm anterior regions are embedded in a secretion produced by these cells. Cyst cells further degenerate making sperm cells free. The sperm bundles are able to perform a forward progression only when sperm are stick together with their tail posterior tips. On the contrary, when sperm have their tail ends free, they beat but are unable to progress. The functional significance of the sperm bundles is discussed.     

A comparison of ventilation rates and haemoglobin concentrations between two species of loach,Noemacheilus barbatulus (L.) and Cobitis taenia (L.)

The mean haemoglobin concentration of Noemacheilus barbatulus was found to be significantly higher than that of Cobitis taenia. Under conditions of reduced oxygen saturation, both species achieved maximum rates of ventilation at 10% 0₂. An alteration in opercular beat pattern accounted for the increased ventilation rates between 20% and 30%. A slightly higher tolerance for lowered oxygen concentrations was demonstrated by Cobitis taenia with disorientation and aerial respiration occurring at lower oxygen tensions than in Noemacheilus barbatulus.     

Fragmentation of condensed chromatin in nuclei of a plant sperm,pteridium aquilinum (L.) Kuhn

Nuclei of pteridium sperm have been dispersed by turbulence in natural or slightly alkaline buffer after stripping off the cytoplasm with nonionic detergent. The nuclei tended to break up into fragments arranged in a linear order. These fragments fluoresced brightly with acridine orange as did intact nuclei. Grounds are given for identifying the smaller fragments with chromosomes. It is proposed that the sperm nucleus of British Pteridium, possibly an autotetrapolid, consists of a sequence of paired homologues.     

Cryopreservation of common carp (Cyprinus carpio L.) sperm I. The importance of oxygen supply

Experiments were carried out on the cryopreservation of common carp ( Cyprinus carpio L.) sperm. The effects of pre-freezing oxygen supply on post-thaw motility and the efficacy of different extenders were studied. Sperm diluents contained DMSO as a cryoprotectant in 10% final concentration. The dilution rate was 1:9 (sperm:diluent). Sperm was diluted and equilibrated (10 min) at 0°C. Sperm was then frozen in plastic straws (0.5 ml) at the following rate: 0°C–4°C: 4°C min⁻¹−4°C–80°C: 11°C min⁻¹ from −80°C, straws were plunged directly into liquid nitrogen (−196°C) for further storage. Frozen samples were thawed in a water bath at 40°C.
The freezability of common carp sperm showing reduced motility (due to suboptimal oxygen supply) after transportation could be restored when 30 min of oxygenation was applied prior to freezing. Highest post-thaw motility (57%, percent of control) was achieved when sperm was diluted with modified Kurokura's 'Extender 2'.     

The effect of methanol on photosynthetic parameters of bean (Phaseolus vulgaris L.) under water deficit

Water availability is the main factor limiting crop growth and productivity in dry regions. This study was carried out in order to determine the effect of spraying methanol solution on the photosynthetic characteristics of bean plants. The main aim of our experiment was to improve plant performance under stress caused by water shortage. Two factors were involved: water-deficit stress, such as severe stress (25% of field capacity), mild stress (75% of field capacity), and no stress (100% of field capacity), and application of methanol solution spray at four concentrations: control (without spraying), 10, 20, and 30%. Methanol was applied three times at different growth stages (seedling, flowering, and podding stage) in 10-d intervals. The treatment with 20% methanol at the seedling stage resulted in increased net photosynthesis (P_N), intercellular CO₂ concentration (C_i), and decreased transpiration rate (E) under no stress and mild stress conditions. Under severe stress, 10 and 20%-methanol treatments resulted in increased C_i, maximal quantum yield of PSII photochemistry, and decreased E. At the flowering stage, methanol treatments resulted in decreased E and increased C_i under mild and severe stress. At the podding stage, 10 and 20%-methanol treatments resulted in increased P_N, C_i, and total chlorophyll content under mild stress. In conclusion, we suggested that foliar application of methanol had a positive role in enhancing photosynthetic performance.     

Genetic diversity of Asian water buffalo (Bubalus bubalis): microsatellite variation and a comparison with protein-coding loci

Twenty-one microsatellite loci in 11 populations of Asian water buffalo (eight swamp, three river type) were analysed and, within and among populations, genetic variability was compared with results from 25 polymorphic protein-coding loci. Within-population mean heterozygosity ranged from 0·380–0·615, approximately twice that estimated from the protein-coding loci (0·184– 0·346). Only eight significant departures from Hardy–Weinberg equilibrium (involving four loci) were detected; global tests showed significant heterozygote deficiencies for these four loci. Non-amplifying alleles are likely to be segregating in some or all populations for one of these loci, and probably for the other three. There was significant differentiation between the swamp and river types of water buffalo, and among populations within each buffalo type. Estimates of θ (measure of population differentiation) for each locus for the eight swamp populations were all highly significant (mean θ = 0·168 ± 0·018). Mean θ for protein-coding loci was not significantly different (0·182 ± 0·041). The variance among protein-coding loci was significantly higher than among microsatellite loci, suggesting balancing selection affecting allele frequencies at some protein-coding loci. Genetic distances show clear separation of the swamp and river types, which were estimated to have diverged at least 10 000–15 000 years ago. The topology of the swamp populations’ microsatellite tree is consistent with their geographical distribution and their presumed spread through south-east Asia. By contrast, the tree based on the protein-coding loci distances is quite different, being clearly distorted by a bottleneck effect in one population, and possibly in at least two others. As many domestic livestock breeds are possibly descended from small numbers of founders, microsatellite-based trees are to be preferred in assessing breed genetic relationships.     

Genetic diversity, structure and marker-trait associations in a collection of Italian tomato (Solanum lycopersicum L.) landraces

The study of phenotypic and genetic diversity in landrace collections is important for germplasm conservation. In addition, the characterisation of very diversified materials with molecular markers offers a unique opportunity to define significant marker-trait associations of biological and agronomic interest. Here, 50 tomato landraces (mainly collected in central Italy), nine vintage and modern cultivars, and two wild outgroups were grown at two locations in central Italy and characterised for 15 morpho-physiological traits and 29 simple sequence repeat (SSR) loci. The markers were selected to include a group of loci in regions harbouring reported quantitative trait loci (QTLs) that affect fruit size and/or shape (Q-SSRs) and a group of markers that have not been mapped or shown to have a priori known linkage (NQ-SSRs). As revealed by univariate and multivariate analyses of morphological data, the landraces grouped according to vegetative and reproductive traits, with emphasis on fruit size, shape and final destination of the product. Compared to the low molecular polymorphism reported in tomato modern cultivars, our data reveal a high level of molecular diversity in landraces. Such diversity has allowed the inference of the existence of a genetic structure that was factored into the association analysis. As the proportion of significant associations is higher between the Q-SSR subset of markers and the subset of traits related to fruit size and shape than for all of the other combinations, we conclude that this approach is valid for establishing true-positive marker-trait relationships in tomato. Electronic supplementary material The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

The life cycles of Leptophlebia vespertina (L.) and L. marginata (L.) (Ephemeroptera) in Llyn Dinas, North Wales

Data is presented on the life cycles of Leptopblehia vespertina (L.) and L. marginata (L.) in Llyn Dinas, an oligotrophic North Wales lake. Both species are univoltine and growth continues throughout the winter although somewhat retarded. Except during July and August there is a clear size separation of the two species, L. marginata being larger and emerging earlier. The egg incubation period was determined directly from laboratory studies. The results are discussed in relation to previous research in the English Lake District.     

Oschmarinella albamarina (Treshchev, 1968) n. comb., a liver fluke from the killer whale Orcinus orca (L.) off the British coast

Oschmarinella albamarina n. comb. is described from the bile-ducts of a killer whale Orcinus orca stranded on the British coast. The systematic position of the species is discussed in relation to the genera Campula Cobbold, 1858, Orthosplanchnus Odhner, 1905 and Oschmarinella Skrjabin, 1947, and the species is transferred from Orthosplanchnus to Oschmarinella. Campula rochebruni (Poirrier, 1886) is transferred to Oschmarinella, as O. rochebruni n. comb., and Oschmarinella mascomai Raga, 1986 is considered its synonym.     

The evolution of gonadotropins: some molecular data concerning a non-mammalian pituitary gonadotropin, the hormone from a teleost fish (Cyprinus carpio L.)

The amino acid and sugar compositions as well as long N-terminal sequences and the C-terminal amino acids of the two subunits of carp gonadotropin, SU I and SU II, were determined. An important homology was demonstrated between SU I and alpha-subunits and between SU II and beta-subunits of mammalian gonadotropins. Moreover SU II was more closely related to the beta-subunit of LH than to the beta-subunit of FSH.     

The sterols of Balanus balanoides (L.): The effect of controlled diets

The effect of a range of diets with varying cholesterol content on the sterol composition of the barnacle Balanus balanoides (L.) has been studied.The animals appear able to maintain their sterol composition for considerable periods (4 months), both quantitatively and qualitatively, in the absence of dietary sterols. This suggests either an extremely slow metabolic turnover or some capacity for de novo synthesis of sterols from other biochemical components.     

The spiracle of Ixodes ricinus (L.) (Ixodidae: Metastigmata: Acarina): a passive diffusion barrier for water vapour

The structure of the spiracle in the adult female of Ixodes ricinus (L.) is assessed. Gaseous exchange between the tracheal system and the external atmosphere is shown to occur only via the aeropyles in the surface of the sieve plate and is regulated by the atrial valve. The elaborate structure is shown to be an effective passive mechanism to retard the transpiration of water vapour from the tracheal system. The pedicels reduce airflow within the labyrinth to a minimum, while the size and arrangement of the aeropyles allow the formation of a cloud of water vapour above the surface of the spiracle. It is from this cloud which diffusion of water vapour takes place, with no significant changes in water vapour pressure occurring within the tracheal system or the spiracle itself. The mechanics of the system are analysed.     

Comparative effect of water, heat and light stresses on photosynthetic reactions in Sorghum bicolor(L.) Moench

Five varieties of Sorghum bicolor (L.) Moench., differing in their drought tolerance under field conditions have been used to study the effect of individual components of drought stress, namely high light intensity stress, heat stress and water stress, on their photosynthetic performance. Chlorophyll content, chlorophyll fluorescence, ribulose-1,5-bisphosphate carboxylase (Rubisco, EC 4.1.1.39) content, phosphoenolpyruvate carboxylase (PEPcase, EC 4.1.1.31) activity and photo-synthetic oxygen evolution were used as key parameters to assess photosynthetic performance. The results indicated that photochemical efficiency of photosystem II (PSII) was severely reduced by all three stress components, whereas PEPcase activity was more specifically reduced by water stress. Degradation of Rubisco and chlorophyll loss occurred under high light and water stress conditions. Of the four drought-tolerant varieties, E 36-1 showed higher PEPcase activity, Rubisco content and photochemical efficiency of PSII, and was able to sustain a higher maximal rate of photosynthetic oxygen evolution under each stress condition as compared to the other varieties. A high stability to stress-induced damage, or acclimation of photosynthesis to the individual components of drought stress may contribute to the high yields of E 36-1 under drought conditions. In the E 36-1 variety markedly higher levels of the chloroplastic chaperonin 60 (cpn 60) were observed under all stress conditions than in the susceptible variety CSV 5.Key words: Chlorophyll fluorescence, drought stress, oxygen evolution, phosphoenopyruvate carboxylase, Sorghum.