Facultative autogamy in Cyrtopodium polyphyllum (Orchidaceae) through a rain-assisted pollination mechanism

Cyrtopodium includes similar to 42 species, among which is Cyrtopodium polyphyllum (Vell.) Pabst ex F. Barros that occurs in a rainforest in south-eastern Brazil. Its non-rewarding flowers, which attract Centridini bees by deceit, are rain-assisted self-pollinated, a phenomenon rarely found in orchids and other plant families. In addition, self-pollination has never been reported in Cyrtopodiinae and data on the pollination of South American orchids are scarce. Flowers were observed at different times of the day, on both sunny and rainy days, to record floral morphology, visitors and the effects of rainfall on flowers. On rainy days, water accumulates on the stigma and dissolves the adhesive substance of the stigmatic surface. A viscous drop thus forms, which contacts the pollinarium. When evaporation makes the viscous drop shrink, the drop moves the pollinarium with the anther onto the stigmatic surface and promotes self-pollination. Fruit set in natural habitat was low, with 2.4% at one study site, where a similar value (2.2%) was recorded in flowers self-pollinated by rain. In C. polyphyllum, facultative self-pollination assisted by rain is thus an important strategy that guarantees fruit set when pollinator`s visits are scarce, which is common in species pollinated by deceit.

The benefits of polyandry in the free-spawning polychaete Galeolaria caespitosa

In many species, females are thought to benefit from polyandry due to the reduced risks of fertilization by genetically incompatible sperm. However, few studies that have reported such benefits have directly attributed variation in female reproductive success to the interacting effects of males and females at fertilization. In this paper, we determine whether male x female interactions influence fertilization in vitro in the free-spawning, sessile polychaete Galeolaria caespitosa. Furthermore, we determined whether polyandry results in direct fertilization benefits for females by experimentally manipulating the number of males contributing towards staged spawning events. To test for male x female interaction effects we performed an initial experiment that crossed seven males with six females (in all 42 combinations), enabling us to assess fertilization rates for each specific male-female pairing and attribute variation in fertilization success to males, females and their interaction. This initial experiment revealed a strong interaction between males and females at fertilization, confirming that certain male-female combinations were more compatible than others. A second experiment tested the hypothesis that polyandry enhances female reproductive success by exposing each female's eggs to either a single male's sperm (monandry) or the sperm from three males simultaneously (polyandry). We performed this second experiment at two ecologically relevant sperm concentrations. This latter experiment revealed a strong fertilization benefit of polyandry, independent of the effects of sperm concentration (which were also significant). We suggest that these direct fertilization gains arising from polyandry will constitute an important source of selection on females to mate multiply in nature.

Offspring size effects in the marine environment: A field test for a colonial invertebrate

A central tenet of life-history theory is the presence of a trade-off between the size and number of offspring that a female can produce for a given clutch. A crucial assumption of this trade-off is that larger offspring perform better than smaller offspring. Despite the importance of this assumption empirical, field-based tests are rare, especially for marine organisms. We tested this assumption for the marine invertebrate, Diplosoma listerianum, a colonial ascidian that commonly occurs in temperate marine communities. Colonies that came from larger larvae had larger feeding structures than colonies that came from smaller larvae. Colonies that came from larger larvae also had higher survival and growth after 2 weeks in the field than colonies that came from smaller larvae. However, after 3 weeks in the field the colonies began to fragment and we could not detect an effect of larval size. We suggest that offspring size can have strong effects on the initial recruitment of D. listerianum but because of the tendency of this species to fragment, offspring size effects are less persistent in this species than in others.

Does egg competition occur in marine broadcast-spawners?

When the availability of sperm limits female reproductive success, competition for sperm, may be an important broker of sexual selection. This is because sperm limitation can increase the variance in female reproductive success, resulting in strong selection on females to compete for limited fertilization opportunities. Sperm limitation is probably common in broadcast-spawning marine invertebrates, making these excellent candidates for investigating scramble competition between broods of eggs and its consequences for female reproductive success. Here, we report our findings from a series of experiments that investigate egg competition in the sessile, broadcast-spawning polychaete Galeolaria caespitosa. We initially tested whether the order in which eggs encounter sperm affects their fertilization success at two ecologically relevant current regimes. We used a split-clutch-split-ejaculate technique to compare the fertilization success of eggs from individual females that had either first access (competition-free treatment) or second access (egg competition treatment) to a batch of sperm. We found that fertilization success depended on the order in which eggs accessed sperm; eggs that were assigned to the competition-free treatment exhibited significantly higher fertilization rates than those assigned to the egg competition treatment at both current speeds. In subsequent experiments we found that prior exposure of sperm to eggs significantly reduced both the quantity and quality of sperm available to fertilize a second clutch of eggs, resulting in reductions in fertilization success at high and low sperm concentrations. These findings suggest that female traits that increase the likelihood of sperm-egg interactions (e.g. egg size) will respond to selection imposed by egg competition.

PCR-mediated detection of acidophilic, bioleaching-associated bacteria

The detection of acidophilic microorganisms from mining environments by culture methods is time consuming and unreliable. Several PCR approaches were developed to amplify small-subunit rRNA sequences from the DNA of six bacterial phylotypes associated with acidic mining environments, permitting the detection of the target DNA at concentrations as low as 10 fg.

The chemistry of 5-oxodihydroisoxazoles .19. The synthesis and photolysis of N-thioacylisoxazol-5(2H)-ones

5-Oxodihydroisoxazoles react with thiocarbonyl chlorides to afford N-thioacylisoxazol-5(2H)-ones which lose carbon dioxide under photochemical conditions and undergo intramolecular cyclisation of the iminocarbene to afford thiazoles, However, in some cases loss of carbon dioxide is accompanied by loss of sulfur, giving 1,3-oxazin-6-ones.

A comparison of the expressive communication skills of triplet, twin and singleton children

Few studies have focused on the language acquisition of higher multiple birth sets. In this study, the communication skills of 51 triplet children are described. The measures used were: mean length of utterance; type-token ratio; conversational nets; phoneme repertoire; and number of different types of phonological processes used. The data gained were used to compare the communication skills of triplets with those of twins, singletons and normative data available in the literature. Siblings within triplet sets were also compared using language samples obtained from adult-child interactions and when the three children were playing together. The results indicated that the triplets' early communication skills were different from those of both singletons and twins. The triplets' difficulties included delayed syntactic development, limited use of different language functions and delayed phonological development. In contrast, twins' communication profile is characterised by disordered phonological development.

Chemical characterization of soils of a tropical humid forest zone: A methodology

A methodology, based on a combination of routinely performed analyses and investigation of fundamental charge and anion sorption properties, was used to characterize the soils of the humid forest zone of Cameroon, In general, the soils have about 2 cmol kg(-1) permanent negative charge, with about 1 cmol kg(-1) from variable-charge sources at current soil pH values, Furthermore, they are impoverished with respect to Ca, Mg, and K, while Al frequently dominates the exchange complex. Thus, the ability of these soils to retain base cations is more limited than is suggested by the cation-exchange capacity (CEC), Therefore we propose the concept of a degradation index (DI) defined as: DI = 100(CEC5.5 - sum of basic cations)/CEC5.5, where CEC5.5 is the CEC measured at pH 5.5, This index encompasses degradation a soil may have experienced from natural or man-made causes, Extractable PO4 concentrations are considered very low and the soils have a moderate to high capacity to fix added PO4. Surface soil SO4 concentrations are considered marginal to deficient for plant growth, though adequate reserves of SO4 are held in the subsoil by SO4 sorption, The approach used demonstrated that the five morphologically different soil profile classes identified in the zone have similar chemical characteristics. Thus, the results of experimentation conducted on one of the soil profile classes will be applicable throughout the zone, Furthermore, the approach has provided a means of identifying comparable soil types in other parts of the world and will guide technology transfer, The analytical methods used in this study are relatively simple and require no specialized equipment, and are therefore within the capabilities of many laboratories in the developing world.

Influence of vitrification on mouse metaphase II oocyte spindle dynamics and chromatin alignment

Objective: To evaluate influences of vitrification and warming of metaphase II (MII) mouse oocytes on survival, spindle dynamics. spindle morphology, and chromatin alignment on metaphase plates. Design: Experimental animal Study. Setting: University animal laboratory. Animal(s): Eight-week-old B6D2F1 mice. Intervention(s): Denuded MII oocytes were used fresh (control), exposed to vitrification/warming solutions (Sol Expos), or vitrified and warmed (Vitr). Main Outcome Measure(s): Oocyte recovery and survival after warming and the influence of solution exposure and cryopreservation on spindle dynamics and chromatin alignment. Result(s): Cryopreservation of two or 10 oocytes per straw resulted in recovery (100% +/- 0% and 95% +/- 4%, respectively; mean SE) and survival (95% 2% and 98% 2%, respectively). Immediately after warming (Vitr), significantly fewer oocytes assessed with immunocytochemistry contained spindles, compared with control and Sol Expos. When oocytes were placed into a 3 degrees 7C environment for 2 hours after exposure or warming, the ability to recognize spindles by immunocytochemistry was not significantly different between groups. Using live-cell time-lapse imaging with LC-Polscope, similar time-dependent spindle formation dynamics were observed. At 2 hours after collection or treatment, spindle morphology and length were not significantly different between the groups, nor was the incidence of aberrant alignment of chromatin on metaphase plates. Conclusion(s): Immediately after warming of vitrified MII oocytes, beta-tubulin is depolymerized and chromatin remains condensed on the metaphase plate. Within a 2-hour period, beta-tubulin repolymerizes, forming morphologically normal metaphase spindles with properly aligned chromatin.

Biological Effects and Dose-Response Assessment of Diesel Exhaust Particles on In Vitro Early Embryo Development in Mice

An increased risk of early pregnancy loss in women briefly exposed to high levels of ambient particulate matter during the preconceptional period was recently observed. The effects of this exposure on early embryo development are unknown. This study was designed to assess the dose-response and biological effects of diesel exhaust particles (DEP) on in vitro embryo development using the in vitro fertilization (IVF) mouse model. Zygotes obtained from superovulated mice after IVF were randomly cultured in different DEP concentrations (0, 0.2, 2, and 20 mu g/cm(2)) for 5 days and observed for their capacity to attach and develop on a fibronectin matrix until day 8. Main outcome measures included blastocyst rates 96 and 120 h after insemination, hatching discriminatory score, total cell count, proportion of cell allocation to inner cell mass (ICM) and trophectoderm (TE), ICM morphology, attachment rate and outgrowth area, apoptosis and necrosis rates, and Oct-4 and Cdx-2 expression. Multivariate analysis showed a negative dose-dependent effect on early embryo development and hatching process, blastocyst cell allocation, and ICM morphology. Although blastocyst attachment and outgrowth were not affected by DEP, a significant impairment of ICM integrity was observed in day 8 blastocysts. Cell death through apoptosis was significantly higher after DEP exposure. Oct-4 expression and the Oct-4/Cdx-2 ratio were significantly decreased in day 5 blastocysts irrespective of DEP concentration. Results suggest that DEP appear to play an important role in disrupting cell lineage segregation and ICM morphological integrity even at lower concentrations, compromising future growth and viability of the blastocyst.

Prospective randomized comparison of human oocyte cryopreservation with slow-rate freezing or vitrification

Objective: To compare cryopreservation of mature human oocytes with slow-rate freezing and vitrification and determine which is most efficient at establishing a pregnancy. Design: Prospective randomized. Setting: Academically affiliated, private fertility center. Patient(s): Consenting patients with concerns about embryo cryopreservation and more than nine mature oocytes at retrieval were randomized to slow-rate freezing or vitrification of supernumerary (more than nine) oocytes. Intervention(s): Oocytes were frozen or vitrified, and upon request oocytes were thawed or warmed, respectively. Main Outcome Measure(s): Oocyte survival, fertilization, embryo development, and clinical pregnancy. Result(s): Patient use has resulted in 30 thaws and 48 warmings. Women`s age at time of cryopreservation was similar. Oocyte survival was significantly higher following vitrification/warming (81%) compared with freezing/thawing (67%). Fertilization was more successful in oocytes vitrified/warmed compared with frozen/thawed. Fertilized oocytes from vitrification/warming had significantly better cleavage rates (84%) compared with freezing/thawing (71%) and resulted in embryos with significantly better morphology. Although similar numbers of embryos were transferred, embryos resulting from vitrified oocytes had significantly enhanced clinical (38%) pregnancy rates compared with embryos resulting from frozen oocyte (13%). Miscarriage and/or spontaneous abortion rates were similar. Conclusion(s): Our results suggest that vitrification/warming is currently the most efficient means of oocyte cryopreservation in relation to subsequent success in establishing pregnancy. (Fertil Steril (R) 2010; 94: 2088-95. (C) 2010 by American Society for Reproductive Medicine.)

Tests for ovarian reserve: reliability and utility

Purpose of review This review discusses ovarian reserve tests for ovulation induction and their application in determining fertility capacity, and their current applications to assess risk of natural ovarian failure and to estimate ovarian function after cancer treatment. Recent findings The current arsenal of ovarian reserve tests comprises hormonal markers [basal follicle stimulating hormone, estradiol, inhibin-B, antimullerian hormone (AMH)] and ultrasonographic markers [ovarian volume, antral follicle counts (AFCs)]. These markers have limitations in terms of which test(s) should be used to reliably predict ovarian reserve with regard to accuracy, invasiveness, cost, convenience, and utility. Several studies have correlated sonographic AFCs with serum AMH levels for predicting the ovarian response to ovulation induction protocols during assisted reproduction treatments. Summary Serum AMH levels and AFC are reliable tests for predicting the ovarian response to ovulation induction. However, none of the currently employed tests of ovarian reserve can reliably predict pregnancy after assisted conception. Further, ovarian reserve tests cannot predict the onset of reproductive and hormonal menopause; thus, they should be used with caution for reproductive life-programming counseling. Moreover, there is no evidence to support the use of ovarian reserve tests to estimate the risk of ovarian sufficiency after cancer treatments.

Abnormal methylation at the KvDMR1 imprinting control region in clinically normal children conceived by assisted reproductive technologies

Genomic imprinting alterations have been shown to be associated with assisted reproductive technologies (ARTs) in animals. At present, data obtained in humans are inconclusive; however, some epidemiological studies have demonstrated an increased incidence of imprinting disorders in children conceived by ARTs. In the present study, we focused on the effect of ARTs [IVF and intracytoplasmic sperm injection (ICSI)] on the epigenetic reprogramming of the maternally methylated imprinting control region KvDMR1 in clinically normal children. Qualitative and quantitative methylation at KvDMR1 were assessed by the methylation-specific PCR approach and by the methylation-sensitive enzymatic digestion associated with real-time PCR method, respectively. DNA was obtained from peripheral blood of 12/18 and umbilical cord blood and placenta of 6/18 children conceived by IVF or ICSI. The methylation patterns observed in this group were compared with the patterns observed in 30 clinically normal naturally conceived children (negative controls) and in 3 naturally conceived Beckwith-Wiedemann syndrome patients (positive controls). Hypomethylation at KvDMR1 was observed in 3/18 clinically normal children conceived by ARTs (2 conceived by IVF and 1 by ICSI). A discordant methylation pattern was observed in the three corresponding dizygotic twins. Our findings corroborate the hypothesis of vulnerability of maternal imprinting to ARTs. Furthermore, the discordant methylation at KvDMR1 observed between dizygotic twins could be consequent to one of the following possibilities: (i) a differential vulnerability of maternal imprints among different embryos; or (ii) epimutations that occurred during gametogenesis resulting in the production of oocytes without the correct primary imprint at KvDMR1.

Effects of biomass burning on nasal mucociliary clearance and mucus properties after sugarcane harvesting

Objective: Biofuel from sugarcane is widely produced in developing countries and is a clean and renewable alternative source of energy. However, sugarcane harvesting is mostly performed after biomass burning. The aim of this study was to evaluate the effects of harvesting after biomass burning on nasal mucociliary clearance and the nasal mucus properties of farm workers. Methods: Twenty seven sugarcane workers (21-45 years old) were evaluated at the end of two successive time-periods: first at the end of a 6-month harvesting period (harvesting), and then at the end of a 3-month period without harvesting (non-harvesting). Nasal mucociliary clearance was evaluated by the saccharine transit test, and mucus properties were analyzed using in vitro mucus contact angle and mucus transportability by sneeze. Arterial blood pressure, heart rate, respiratory rate, pulse oximetry, body temperature, associated illness, and exhaled carbon monoxide were registered. Results: Data are presented as mean values (95% confidence interval). The multivariate model analysis adjusted for age, body-mass index, smoking status and years of working with this agricultural practice showed that harvesting yielded prolonged saccharine transit test in 7.83 min (1.88-13.78), increased mucus contact angle in 8.68 degrees (3.18-14.17) and decreased transportability by sneeze in 32.12 mm (-44.83 to -19.42) compared with the non-harvesting period. No significant differences were detected in any of the clinical parameter at either time-period. Conclusion: Sugarcane harvesting after biomass burning negatively affects the first barrier of the respiratory system in farm workers by impairing nasal mucociliary clearance and inducing abnormal mucus properties. (C) 2011 Elsevier Inc. All rights reserved.

Age-related increase of reactive oxygen species in neat semen in healthy fertile men

OBJECTIVES The effects of advancing paternal age on the male reproductive system are well known, but its effects on fecundity remain controversial. Although oxidative stress is associated with poor semen quality and function, a relationship with advancing male age has not been established. The objective of this study was to analyze the relationship between male age and seminal reactive oxygen species (ROS) levels in men presenting for voluntary sterilization. METHODS We prospectively evaluated 98 fertile men who were candidates for vasectomy. These were divided into 2 age groups: less than 40 years (n = 78) and 40 or more years (n = 20). We used 46 infertile patients as positive controls. Standard semen analysis, seminal leukocyte count and ROS levels were measured in all samples. Fertile men with leukocytospermia were excluded. RESULTS The mean age of the men was 35.1 +/- 5.6 years. Men 40 years and older had significantly higher ROS levels compared with younger men (P < 0.001). We observed a positive correlation between seminal ROS levels and age (r = 0.20; P = 0.040). In addition, ROS was negatively correlated with sperm concentration (r = -0-48; P < 0.001) and motility (r = -0.21; P = 0.030). CONCLUSIONS Reactive oxygen species levels are significant higher in seminal ejaculates of healthy fertile men older than 40 years. ROS levels in whole ejaculate are significantly correlated to age among fertile men. Because ROS are clearly implicated in the pathogenesis of male infertility, these data suggest that delayed fatherhood may reduce the chances of pregnancy as men become progressively less fertile with age.