Physiological effects of hypoxia in developing and adult honeybees <Emphasis Type=Italic>Apis mellifera</Emphasis> L.

The effect of hypoxia on activity of metabolism was studied in developing and adult honeybees. The feedback was established between the O₂ deficit in the gas medium and its consumption by honeybees. An elevation of temperature within the limits of vital diapason for development of honeybees activated the O₂ consumption regardless of the hypoxia level. A prolonged action of hypoxia on the honeybee physiological state was revealed. There was shown the existence of convergent similarity between the effect of O₂ deficit on honeybees and on homoiothermal animals.     

Exo- and Endogenous Water in the Honeybee Body (<Emphasis Type=Italic>Apis mellifera</Emphasis>, Hymenoptera,Apidae)

The exo- and endogenous water content in the honeybee body was studied in relation to age, season, and the physiological state of the bee colony. The water content in the carbohydrate food consumed by the bees was shown to affect brood production in the colony. The behavior of the bees supplying the colony with water was investigated and the amount of water used for larval food dilution and nest cooling was estimated. Seasonal interruption and resumption of the reproductive functions of the queen were shown to depend on the dynamics of the water content in the digestive tract of worker bees.     

Intraspecies Differences of Humoral Protective Response in the Honeybee <Emphasis Type="Italic">Apis mellifera</Emphasis>

Results of experiments on detection of differences in realization of a protective response to introduced pathogen (bacterial preparation, bitoxybacillin) at the molecular level between bees of the Central Russian and Caucasian strains as well as between their hybrids are presented. Interstrain differences in activity of enzymes and in ratio of the nucleic acid content in bees at the initial stage of development of protective reactions are studied. It is shown that bees of the Central Russian strain are capable for faster realization of responses after immunization and that bees of initial strains are advantageous over their hybrids in development of the protective response.     

Investigation of polyandry in honey bees (<Emphasis Type=Italic>Apis mellifera</Emphasis>) using microsatellites

Polyandry is a specific phenomenon increasing genetic diversity. To analyze the level of polyandry and to assess the contribution of drones to the genetic diversity of honey bee colonies, three microsatellite loci (A008, Ap049, and AC117) were studied in honey bees from colonies of different origin (Middle Russian race, Carpathian race, and hybrids) in Tomsk Province. The share of the introduced paternal alleles varied between colonies from 6.67 to 28.00%. The highest genetic diversity was observed in hybrid colonies (25–28% of introduced paternal alleles).     

Individual recruitment in honeybees, <Emphasis Type="Italic">Apis mellifera</Emphasis> L.

The recruitment of honeybee foragers individually exploiting a low-flow rate-feeder that presented different temporal reward programs was experimentally analyzed. By capturing hive bees that landed at the feeder in a 2-h period, the arrival rate of incoming bees could be obtained. With this procedure we quantitatively analyzed the maximum number of hive bees that can be brought to the feeding station by single foragers. Test bees collected sucrose solution during 12 visits to a rate-feeder located 160 m from the hive. The constant programs offered 0.6, 1.2, or 2.4 M sugar for all 12 visits, while the variable programs delivered either 0.6, 1.2, or 0.6 M or 0.6, 2.4, or 0.6 M, with four visits for each molarity. Results showed that the sucrose concentration exploited by single foragers increased the arrival rate. Moreover, there was a linear relationship within this range of sucrose concentrations that presented a slope of 1.58. Since the sugar solutions were provided at the same flow rate (5 μl/min) in all the programs, the arrival rate expressed in terms of sucrose flow rate (milligrams of sucrose/minute) shows that one additional incoming bee per hour arrived when the single forager assessed an increase in the sucrose flow rate of 0.75 mg sucrose/min at the rate-feeder. The absence of differences in the frequency of visits of the single foragers during the constant programs suggests that the differences observed in the arrival rate can mainly be explained by a more intensive display of the recruitment mechanisms performed per foraging trip instead of by their iterativeness throughout different foraging cycles. Variable reward programs showed that arrival rate is rapidly adjusted according to the reward change and is independent of its magnitude. Received in revised form: 17 August 2001 Electronic Publication     

New allele of the <Emphasis Type=Italic>COCHLEATA</Emphasis> gene in pea <Emphasis Type=Italic>Pisum sativum</Emphasis> L.

Analysis with the polymerase chain reaction showed that the Khlorofill-4 pea Pisum sativum chlorophyll-deficient mutant with reduced stipules has an altered structure of the COCHLEATA (COCH) gene, carrying a new mutant COCH allele. The phenotype of the mutant was described in comparison with another form having reduced stipules (stipules reduced) and the control. Leaves of the coch mutant are smaller and have other proportions than in the control; stipules are absent from leaves of the first nodes and are narrow, bandlike, or spoonlike at later ontogenetic stages. It was concluded that the cell number in the stipule epidermis is reduced in the st and coch mutants compared to the wild type.     

Attenuation of fibrosis <Emphasis Type=Italic>in vitro</Emphasis> and <Emphasis Type=Italic>in vivo</Emphasis> with <Emphasis Type=Italic>SPARC</Emphasis> siRNA

Introduction  

SPARC is a matricellular protein, which, along with other extracellular matrix components including collagens, is commonly over-expressed in fibrotic diseases. The purpose of this study was to examine whether inhibition of SPARC can regulate collagen expression in vitro and in vivo, and subsequently attenuate fibrotic stimulation by bleomycin in mouse skin and lungs.     

Photosynthetic and transpiration responses of <Emphasis Type=Italic>in vitro</Emphasis>-regenerated <Emphasis Type=Italic>Solanum nigrum</Emphasis> L. plants to <Emphasis Type=Italic>ex vitro</Emphasis> adaptation

In vitro regeneration of black nightshade (Solanum nigrum L.) plants was achieved through callus-mediated shoot organogenesis followed by 30 d indoor ex vitro adaptation to nutritional stress under environmental ambience and thereafter 6-d outdoor acclimatization in pots prior to field establishment. Relevant physiological parameters including pigment content, chlorophyll a fluorescence, net photosynthetic rate (P _N), transpiration rate (E), and stomatal conductance (g _s) of in vitro-regenerated plants were investigated during the course of ex vitro adaptation. During the first 4 d of indoor transplantation to potting substrate, there was a marginal reduction in the leaf chlorophyll and carotenoid contents but P _N and E were strongly reduced. The stomatal conductance and E/P _N ratio were significantly higher in plants up to 20 d of indoor adaptation than those of comparable age grown naturally from seeds. The shape of the OJIP fluorescence transient varied significantly with acclimatization, and the maximum change was observed at 2.0 ms. The 2.0 ms variable fluorescence (V _j), 30 ms relative fluorescence (M ₀), photon trapping probability (TR₀/Abs), and photosystem II (PSII) trapping rate (TR₀/RC) showed initial disturbance and subsequent stabilization during 30 d of indoor acclimatization. Energy dissipation (DI₀/RC) and electron transport probability (ET₀/TR₀) showed an initial phase of increase during the 4 d after plants were transplanted outdoors. During the 6-d outdoor acclimatization after transfer of plants to soil, no significant change in total chlorophylls and carotenoids, E, and g _s were observed, but P _N improved after reduction on the first d. The OJIP-derived parameters experienced change on the first d but were stabilized quickly thereafter. There was no significant difference between outdoor acclimatized plants and those of the seed-grown plants of comparable age with respect to photosynthetic and fluorescence parameters. Direct transfer of plants without indoor acclimatization, however, showed a completely different trend with respect to P _N, E, and OJIP fluorescence transients. The bearing of this study on optimizing micropropagation is discussed.     

Regulation of the <Emphasis Type=Italic>ALBINO3</Emphasis>-mediated transition to flowering in <Emphasis Type=Italic>Arabidopsis</Emphasis> depends on the expression of <Emphasis Type=Italic>CO</Emphasis> and <Emphasis Type=Italic>GA1</Emphasis>

ALBINO3, a homologue of PPF1 in Arabidopsis, encodes a chloroplast protein, and is essential for chloroplast differentiation. In the present study, ALBINO3(−) transgenic plants exhibited a significant decrease in both the number of rosette leaves at bolting and the days before bolting, suggesting the important roles of ALBINO3 in regulating flowering during non-inductive short-day photoperiods. ALBINO3 mRNA was apparently accumulated in shoot apical meristem and floral meristems around the shoot apical meristem in wild-type plants. ALBINO3 might be predominantly involved in inducing the floral repression pathway by activating the expression of TFL1, and by suppressing the expression of LFY, respectively, in the shoot apical meristem. Moreover, the function of ALBINO3 in regulating flowering transition depended on the expression of CO and GA1, because ALBINO3 might function in the downstream integration of the photoperiod-dependent and the photoperiod-independent pathways. These results suggest that ALBINO3 may have an important integrative function in the flowering process in Arabidopsis.     

Arbutin production in <Emphasis Type=Italic>Ruta graveolens</Emphasis> L. and <Emphasis Type=Italic>Hypericum perforatum</Emphasis> L. in vitro cultures

Optimization of conditions for hydroquinone biotransformation into its β-d-glucoside, arbutin, in agitated shoot cultures of Ruta graveolens L. and Hypericum perforatum L. allowed us to obtain a maximum content of this important therapeutic and cosmetic product of 7.8 and 7.2% (dry weight), respectively. These contents are higher than respective values required for standardization of known arbutin-containing plant raw materials according to the European Pharmacopoeia and national pharmacopoeias of European countries.     

Presence of <Emphasis Type=Italic>C. albidus</Emphasis>, <Emphasis Type=Italic>C. laurentii</Emphasis> and <Emphasis Type=Italic>C. uniguttulatus</Emphasis> in Crop and Droppings of Pigeon Lofts (<Emphasis Type=Italic>Columba livia</Emphasis>)

Columba livia is an important reservoir and carrier of Cryptococcus neoformans, Cryptococcus uniguttulatus, Cryptococcus laurentii and Cryptococcus albidus. Upper digestive tract of this species is also known as a habitat for Cryptococcus neoformans. Given the increasing clinical interest of this microorganism, 331 swabs from crop and 174 dropping samples from pigeon lofts in Grand Canary Island have been studied. The obtained results show an extensive presence samples 81 positive (24.47%) of Cryptococcus spp. in analysed crops: 32 (9.66%) for C. neoformans, 24 (7.2%) for C. uniguttulatus, 23 (6.9%) for C. albidus and 2 (0.6%) for C. laurentii. In the same way, Cryptococcus spp was also isolated in 82 (47.13%), dropping samples: C. neoformans in 59 (33.9%), C. uniguttulatus, in 9 (5.17%), C. laurentii in 8 (4.59%) and C. albidus in 6 (3.44%) of the investigated samples, respectively. The cryptococcosis produced by species of cryptococci other than C. neoformans has become more important during the last decade, supporting the study on the role of pigeon in the epidemiology of this disease.     

Probiotic <Emphasis Type=Italic>Lactobacillus</Emphasis> strains: <Emphasis Type=Italic>in vitro</Emphasis> and <Emphasis Type=Italic>in vivo</Emphasis> studies

Three Lactobacillus strains (LOCK 0900, LOCK 0908, LOCK 0919) out of twenty-four isolates were selected according to their antagonistic activity against pathogenic bacteria, resistance to low pH and milieu of bile salts. Intragastric administration of a mixture of these strains to Balb/c mice affected cytokine T_H1-T_H2 balance toward nonallergic T_H1 response. Spleen cells, isolated from lactobacilli-treated mice and re-stimulated in vitro with the mixture of heat-inactivated tested strains, produced significantly higher amounts of anti-allergic tumor necrosis factor- and interferon-γ than control animals whereas the level of pro-allergic interleukin-5 was significantly lower. Lactobacillus cells did not translocate through the intestinal barrier into blood, liver and spleen; a few Lactobacillus cells found in mesenteric lymph nodes could create antigenic reservoir activating the immune system. The mixture of Lactobacillus LOCK 0900, LOCK 0908 and LOCK 0919 strains represents a probiotic bacterial preparation with possible use in prophylaxis and/or therapy of allergic diseases.     

Nestmate Recognition Differences between Honeybee Colonies of <Emphasis Type="BoldItalic">Apis cerana</Emphasis> and <Emphasis Type="BoldItalic">Apis mellifera</Emphasis>

Nestmate recognition in Apis cerana and Apis mellifera was studied by introducing sealed queen cells heterospecifically between queenless colonies. No A. cerana queens were accepted by queenless A. mellifera; but A. mellifera queens were accepted in queenless A. cerana colonies. A. mellifera queens oviposited in queenless A. cerana colonies, but A. cerana workers removed most eggs. In time, egg removals declined, and some A. mellifera larvae that hatched from these eggs reached adulthood, and eventually about half of the workers were newly emerged A. mellifera. Eventually, the colonies consisted only of A. mellifera after A. cerana workers died by attrition. A. mellifera workers are more sensitive to nestmate recognition and killed the A. cerana virgin queens. In mixed-species colonies, after newly emerged A. mellifera workers matured, they removed eggs laid by the A. cerana queens until there were no workers to replace the old ones.     

The order of the <Emphasis Type=Italic>bs,Skdh</Emphasis>, and <Emphasis Type=Italic>Aadh1</Emphasis> genes in chromosome 5R of rye <Emphasis Type=Italic>Secale cereale</Emphasis> L.

Segregation analysis was performed in the progenies obtained in analyzing crosses of hybrids of spring and winter accessions of rye Secale cereale L. and wild S. montanum subsp. anatolicum (Grossh.) Tzvel. (syn. S. strictum (J. Presl) J. Presl). The test genes controlled the brittle stem (bs), the allelic variants of aromatic alcohol dehydrogenase (Aadh1) and shikimate dehydrogenase (Skdh), and the growth habit (Vrn1). A linkage was observed in the inheritance of the brittle stem and the aromatic alcohol dehydrogenase and shikimate dehydrogenase alloenzymes. The order of genes was established to be bs-Skdh-Aadh1, and the genetic distances were estimated to be bs-(9.0%)-Skdh, bs-(10.8%)-Aadh1, and Skdh-(5.3%)-Aadh1. The recombination coefficient between the Skdh and Aadh1 genes varied from 2.2 to 18.2%, averaging 5.3%. The growth habit was inherited independently of the bs-Skdh-Aadh1 linkage group.     

Efficient separation and purification of allophycocyanin from <Emphasis Type=Italic>Spirulina</Emphasis> (<Emphasis Type=Italic>Arthrospira</Emphasis>) <Emphasis Type=Italic>platensis</Emphasis>

Allophycocyanin (APC) is a minor component of phycobiliproteins in cyanobacteria and red algae. This paper describes a simple and inexpensive extracting method for isolating APC from Spirulina (Arthrospira) platensis with high efficiency. The crude phycobiliprotein extract was pretreated by ammonium sulfate fractionation. Then, by adding hydroxylapatite into crude phycobiliprotein extract dissolved in 20 mM phosphate buffer (pH 7.0), APC was selectively adsorbed by hydroxylapatite but C-phycocyanin (C-PC) was not. The hydroxylapatite was collected and APC was extracted from the crude phycobiliprotein extract. Then, the enriched APC was washed off from the hydroxylapatite using 100 mM phosphate buffer (pH 7.0). In this simple extracting method it was easy to remove C-PC and isolate APC in large amounts. The absorbance ratio A ₆₅₀/A ₂₈₀ of extracted APC reached 2.0. The recovery yield was 70%, representing 4.61 mg · g⁻¹ wet weight. The extracted APC could be further purified by a simple anion-exchange chromatography with a pH gradient from 5.6 to 4.0. The absorbance ratio A ₆₅₀/A ₂₈₀ of the purified APC reached 5.0, and the overall recovery yield was 43%, representing 2.83 mg · g⁻¹ wet weight. Its purity was confirmed by native polyacrylamide gel electrophoresis (PAGE) and sodium dodecyl sulfate-PAGE.     

Interaction of <Emphasis Type=Italic>Bdellovibrio bacteriovorus</Emphasis> with bacteria <Emphasis Type=Italic>Campylobacter jejuni</Emphasis> and <Emphasis Type=Italic>Helicobacter pylori</Emphasis>

Interaction of Bdellovibrio bacteriovorus 100NCJB with bacteria Campylobacter jejuni (strains 1, 2, 3, 4, and 5) and Helicobacter pylori, strain TX30a, was confirmed. The results indicate that lytic activity of bdellovibrios both in liquid media and cells attached to a surface was observed. The potential use of the antimicrobial activity of predatory bacteria for environmental bioprotection and public health is discussed.     

<Emphasis Type=Italic>In vitro</Emphasis> organogenesis of <Emphasis Type=Italic>Passiflora alata</Emphasis>

Passiflora alata in vitro organogenesis was studied based on explant type, culture medium composition, and incubation conditions. The results indicated that the morphogenic process occurred more efficiently when hypocotyl segment-derived explants were cultured in media supplemented with cytokinin and AgNO₃ incubated under a 16-h photoperiod. The shoot bud elongation and plant development were obtained by transferring the material to MSM culture medium supplemented with GA₃ and incubated in flasks with vented lids. Histological analyses of the process revealed that the difficulties in obtaining plants could be related to the development of protuberances and leaf primordia structures, which did not contain shoot apical meristem. Roots developed easily by transferring elongated shoots to 1/2 MSM culture medium. Plant acclimatization occurred successfully, and somaclonal variation was not visually detected. The efficiency of this organogenesis protocol will be evaluated for genetic transformation of this species to obtain transgenic plants expressing genes that can influence the resistance to Cowpea aphid borne mosaic virus.