Immature Eggs Research Articles

Using Xenopus laevis Oocytes to Functionally Characterize Plant Transporters.

Functionally characterizing plant membrane transport proteins is challenging. Typically, heterologous systems are used to study them. Immature eggs (oocytes) of the South African clawed frog Xenopus laevis are considered an ideal expression system for such studies. These large oocytes have a low number of endogenous transport systems in their plasma membranes and highly express foreign mRNA; the oocyte plasma membrane is the default destination of integral membrane proteins that lack recognized organellar sorting signals. These features facilitate almost background-free characterization of putative plant membrane transporters. Here we describe how to isolate Xenopus laevis oocytes, prepare capped sense RNA (cRNA) of the maize boron importer TASSEL-LESS1 (TLS1) as an example, microinject the cRNA into the isolated oocytes, and functionally assess the boron import capabilities of TLS1 in an oocyte swelling assay. These protocols can be easily adapted to study other plant and non-plant transporters with putative import function. © 2019 by John Wiley & Sons, Inc.

Screening of a PDE-focused library identifies imidazoles with in vitro and in vivo antischistosomal activity

We report the evaluation of 265 compounds from a PDE-focused library for their antischistosomal activity, assessed in vitro using Schistosoma mansoni. Of the tested compounds, 171 (64%) displayed selective in vitro activity, with 16 causing worm hypermotility/spastic contractions and 41 inducing various degrees of worm killing at 100 μM, with the surviving worms displaying sluggish movement, worm unpairing and complete absence of eggs. The compounds that did not affect worm viability (n = 72) induced a complete cessation of ovipositing. 82% of the compounds had an impact on male worms whereas female worms were barely affected. In vivo evaluation in S. mansoni-infected mice with the in vitro ‘hit’ NPD-0274 at 20 mg/kg/day orally for 5 days resulted in worm burden reductions of 29% and intestinal tissue egg load reduction of 35% at 10 days post-treatment. Combination of praziquantel (PZQ) at 10 mg/kg/day for 5 days with NPD-0274 or NPD-0298 resulted in significantly higher worm killing than PZQ alone, as well as a reduction in intestinal tissue egg load, disappearance of immature eggs and an increase in the number of dead eggs.

In vitro and in vivo evaluation of six artemisinin derivatives against Schistosoma mansoni.

Schistosomiasis is a tropical neglected disease whose socioeconomic impact is surpassed only by malaria. Until recently, praziquantel (PZQ) has been the only available drug, raising concerns that tolerant/resistant strains may appear. Since the discovery of the schistosomicidal potential of artemisinin (ART), new derivatives have been produced and evaluated. In this work, we evaluated the activity of ART derivatives against Schistosoma mansoni, both in vitro and in vivo. In the in vitro assay, worm survival, oviposition, and morphological alterations were evaluated. Further analysis of morphological alterations and membrane integrity was conducted using scanning electron microscopy and a cell-permeable, benzimidazole dye (Hoescht 33258) that binds to the minor groove of double stranded DNA. For the in vivo assay, artesunic acid (AcART) and dihydroartemisinin acetate (AcDQHS) were selected, since they showed the best in vitro results. Infected mice treated 21, 45, or 60days post-infection (dpi), with a concentration of 100mg/kg of either AcART or AcDQHS, showed a significant worm reduction (particularly in females), fewer eggs eliminated in feces, and a decrease of immature eggs in the intestinal tissues. Our results indicate that AcART and AcDQHS have some schistosomicidal activity against juvenile and adult stages of S. mansoni.

Features of embryo and morphogenesis trichurise eggs of cattle in vitro

One of the main factors hampering the development of livestock is the proliferation of a large number of diseases, both contagious and non-contagious etiology. Parasitic diseases of cattle and, in the first place, helminthiasis occupy a significant proportion of the diseases of contagious etiology. The dominant position among the helminthiasis of the digestive organs in ruminants is nematodosis of the gastrointestinal tract, in particular trichyrosis, the economic losses of which consist of a decrease in the productivity of animals due to the development of inflammatory processes in the thick intestine. Trihuris are geoellimits, which are localized predominantly in the blind and colon of the gastrointestinal tract. Females of the parasite lay eggs, which, with fecal animals, are released into the environment, where they are formed and reaches the invasive stage of the larva. Terms of preinvasive development depend on temperature. At a temperature of 30 ° in the eggs of T. ovis, the larva is formed after 17 days, and at 13–17 ° – after 118 days. At 10 º and below, the development of eggs is suspended, but they remain viable for several months. At temperatures above 38 ° the eggs die. According to U. A. Magometbekova (1953), in eggs of T. skrjabini, which parasitized in sheep, at 25–30 ° an invasive larva can be detected in 44–46 days after the eggs enter the environment. Eggs are bad for drying. The results of cultivation of eggs of T. skrjabini, the females of which parasitized in cattle, in vitro at a temperature of 27 ° for 51 days are given. The features of embryo and morphogenesis of eggs of pathogens during cultivation were studied. From the thick intestine of young cattle, by the method of a complete helminthological section by K.I. Scriabin, sexually mature nematodes were isolated, from which female gonads received immature eggs that were cultivated. It was found that at temperatures of 27 °C eggs T. skrjabini, parasitized in the thick intestine of cattle, pass into the stage of invasive larvae from the 27th day and completely stop their development to 51 days. During the period of cultivation, the length of eggs and caps is increased by 2.0 and 20.0%, the width of eggs and caps is reduced by 2.3 and 3.9%, compared to the cultivars. Reduced egg shell thickness by the 27th day of cultivation (by 2.1%) with a subsequent cessation of changes in this index.

ANTISCHISTOSOMAL ACTIVITY OF BALANITIES AEGYPTIACA FRUIT AQUEOUS EXTRACT AGAINST SCHISTOSOMA MANSONI INFECTED MICE

This study aims to evaluate the anthelminthic effect of Balanities aegyptiaca fruit aqueous extract against Schistosoma mansoniin vivo. Parasitological aspects were determined to evaluate the antischistosomal activity of B. aegyptiaca. In addition, tegumental surface topography alterations of S. mansoni adult male worms were noticed via scanning electron microscopic (SEM) investigation after treatment the infected mice orally with 200 mg/kg body weight (BW) B. aegyptiaca alone or combined with 300 mg/kg BW Praziquantel (PZQ). Results showed that treatment of S. mansoni infected mice with B. aegyptiaca, PZQ, or in combination led to a significant decrease (P<0.05) in the weight of liver and intestine as compared with those of S. mansoni-infected mice. Meanwhile, administration of the B. aegyptiaca, PZQ, or in combination decreased significantly (P<0.05) the total worm burden and ova count/g in hepatic and intestinal tissue. B. aegyptiaca and/or PZQ showed a remarkable effect on the oogram pattern in liver and intestine, where the immature and mature eggs were significantly decreased (P<0.05); whereas, dead eggs were significantly increased (P<0.05). The electron microscopy observation exhibited swelling, vacuolization, fusion of the tegumental folds, and loss or shortening of the spines on the tubercles, erosion, cracks and peeling of S. mansoni adult male worm after treatment with B. aegyptiaca and/or PZQ. This study revealed that B. aegyptiaca has a remarkable antischistosomal activity through deteriorating action on the adult worm evidenced by scanning electron microscopy.

ANTISCHISTOSOMAL ACTIVITY OF BALANITIES AEGYPTIACA FRUIT AQUEOUS EXTRACT AGAINST SCHISTOSOMA MANSONI INFECTED MICE

This study aims to evaluate the anthelminthic effect of Balanities aegyptiaca fruit aqueous extract against Schistosoma mansoniin vivo. Parasitological aspects were determined to evaluate the antischistosomal activity of B. aegyptiaca. In addition, tegumental surface topography alterations of S. mansoni adult male worms were noticed via scanning electron microscopic (SEM) investigation after treatment the infected mice orally with 200 mg/kg body weight (BW) B. aegyptiaca alone or combined with 300 mg/kg BW Praziquantel (PZQ). Results showed that treatment of S. mansoni infected mice with B. aegyptiaca, PZQ, or in combination led to a significant decrease (P<0.05) in the weight of liver and intestine as compared with those of S. mansoni-infected mice. Meanwhile, administration of the B. aegyptiaca, PZQ, or in combination decreased significantly (P<0.05) the total worm burden and ova count/g in hepatic and intestinal tissue. B. aegyptiaca and/or PZQ showed a remarkable effect on the oogram pattern in liver and intestine, where the immature and mature eggs were significantly decreased (P<0.05); whereas, dead eggs were significantly increased (P<0.05). The electron microscopy observation exhibited swelling, vacuolization, fusion of the tegumental folds, and loss or shortening of the spines on the tubercles, erosion, cracks and peeling of S. mansoni adult male worm after treatment with B. aegyptiaca and/or PZQ. This study revealed that B. aegyptiaca has a remarkable antischistosomal activity through deteriorating action on the adult worm evidenced by scanning electron microscopy.

Effect of Temperature and Humidity on the Seasonal Phenology of Drosophila suzukii (Diptera: Drosophilidae) in Wisconsin.

Drosophila suzukii (Matsumura) is an invasive pest of soft-skinned fruits that has caused significant economic damage worldwide. In this study, we focused on the seasonal abundance of D. suzukii during the early years of establishment in Wisconsin. We explored the seasonal patterns of summer and winter morphs, their reproductive output, and the effect temperature and humidity may have on their seasonal phenology. The seasonal abundance of D. suzukii during 2 yr (2014-2015) revealed that flies were detected in Wisconsin from early July to late December, with winter morphs being trapped from August through December. The adult populations trapped spanned 1 mo longer in 2015 than in 2014. The peak proportional abundance of D. suzukii in 2015 was recorded in August which was about 2 mo earlier than that in 2014. The combined factor [maximum temperature and maximum humidity] explained the most amount of variation in D. suzukii abundance consistently across the 2 yr in Wisconsin. We did not find significant differences in the fat content, number of mature eggs, proportion of females with immature eggs, or proportion of mated females between summer morph females at the beginning, summer and winter morph females during the middle, or winter morph females at the end of the collecting season in 2015. Our results build on the body of work providing a better understanding of the D. suzukii-overwintering abilities and strengthen the importance of early crop risk assessment and targeted control strategies.

Deficiency of Brummer Impaires Lipid Mobilization and JH-Mediated Vitellogenesis in the Brown Planthopper, Nilaparvata lugens.

Provisioning of sufficient lipids and vitellogenin to the oocytes is an indispensable process for fecundity of oviparous insects. Acute mobilization of lipid reserves in insects is controlled by the Brummer (Bmm), an orthologous of human adipose triglyceride lipase (ATGL). To investigate the functional roles of brummer-mediated lipolysis in the fecundity of the brown planthopper, Nilaparvata lugens, RNA interference (RNAi) analyses were performed with double-stranded RNA (dsRNA) against NlBmm in adult females. Knockdown of NlBmm expression resulted in obesity and blocked lipid mobilization in the fat body. In addition, NlBmm silencing led to retarded ovarian development with immature eggs and less ovarioles, decreased number of laid eggs, prolonged preoviposition period and egg duration. Furthermore, severe reductions of vitellogenin and its receptor abundance were observed upon NlBmm knockdown. The transcript levels of NlJHE (juvenile hormone esterase) which degrades JH were up-regulated, whereas the expression levels of JH receptors NlMet (Methoprene-tolerant) and NlTai (Taiman) and their downstream transcription factors NlKr-h1 (Krüppel-homolog 1) and NlBr (Broad-Complex) were down-regulated after suppression of NlBmm. JH-deficient females exhibited impaired vitellogenin expression, whereas JH exposure stimulated vitellogenin biosynthesis. Moreover, JH topical application partially rescued the decrease in vitellogenin expression in the NlBmm-deficient females. These results demonstrate that brummer-mediated lipolytic system is essential for lipid mobilization and energy homeostasis during reproduction in N. lugens. In addition to the classical view of brummer as a direct lipase with lipolysis activity, we propose here that brummer-mediated lipolysis works through JH signaling pathway to activate vitellogenesis and oocyte maturation that in turn regulates female fecundity.

Modelling the impact of insecticide-based control interventions on the evolution of insecticide resistance and disease transmission

BackgroundCurrent strategies to control mosquito-transmitted infections use insecticides targeted at various stages of the mosquito life-cycle. Control is increasingly compromised by the evolution of insecticide resistance but there is little quantitative understanding of its impact on control effectiveness. We developed a computational approach that incorporates the stage-structured mosquito life-cycle and allows tracking of insecticide resistant genotypes. This approach makes it possible to simultaneously investigate: (i) the population dynamics of mosquitoes throughout their whole life-cycle; (ii) the impact of common vector control interventions on disease transmission; (iii) how these interventions drive the spread of insecticide resistance; and (iv) the impact of resistance once it has arisen and, in particular, whether it is sufficient for malaria transmission to resume. The model consists of a system of difference equations that tracks the immature (eggs, larvae and pupae) and adult stages, for males and females separately, and incorporates density-dependent regulation of mosquito larvae in breeding sites.ResultsWe determined a threshold level of mosquitoes below which transmission of malaria is interrupted. It is based on a classic Ross-Macdonald derivation of the malaria basic reproductive number (R0) and may be used to assess the effectiveness of different control strategies in terms of whether they are likely to interrupt disease transmission. We simulated different scenarios of insecticide deployment by changing key parameters in the model to explore the comparative impact of insecticide treated nets, indoor residual spraying and larvicides.ConclusionsOur simulated results suggest that relatively low degrees of resistance (in terms of reduced mortality following insecticide contact) can induce failure of interventions, and the rate of spread of resistance is faster when insecticides target the larval stages. The optimal disease control strategy depends on vector species demography and local environmental conditions but, in our illustrative parametrisation, targeting larval stages achieved the greatest reduction of the adult population, followed by targeting of non-host-seeking females, as provided by indoor residual spraying. Our approach is designed to be flexible and easily generalizable to many scenarios using different calibrations and to diseases other than malaria.

Suppression of Schistosoma japonicum Acetylcholinesterase Affects Parasite Growth and Development.

To further investigate the importance of Schistosoma japonicum acetylcholinesterase (SjAChE) in cholinergic signaling for parasite growth and development, we used RNA interference (RNAi) to knock-down its expression in adults and eggs in vitro. This resulted in its reduced transcription but also expression of other important genes involved both in cholinergic signaling and glucose uptake were impacted substantially. Significant decreases in AChE protein expression, AChE enzymatic activity, and glucose uptake were observed in the SjAChE-knockdown parasites compared with luciferase controls. In vaccine/challenge experiments, we found that immunization of mice with recombinant SjAChE (rSjAChE) expressed in Escherichia coli elicited reductions in male worm numbers (33%), liver granuloma density (41%), and reduced numbers of mature intestinal eggs (73%) in the vaccinated group compared with the control group. These results indicate AChE plays an important role in the metabolism of male worms, and impacts indirectly on female fecundity leading to increased numbers of immature eggs being released and reduced sizes of liver granulomas. Furthermore, cytokine analysis showed that immunization of mice with rSjAChE elicited a predominantly Th1-type immune response characterized by increased production of IFNγ in splenic CD4+ T cells of vaccinated mice. The study confirms the potential of SjAChE as a vaccine/drug candidate against zoonotic schistosomiasis japonica.

Biocontrol of Meloidogyne incognita by Metarhizium guizhouense and its protease

Crude extracts of the fungus Metarhizium guizhouense PSUM02 grown on protein- enriched medium showed high proteolytic activity and produced one main protein indicated as fungal protease with 33 kDa molecular mass. The protease activity had high stability when incubated at 10 or −20 °C for 6 months. The use of Metarhizium spores (1 × 104 spore mL−1) in combination with its protease (354.1 ± 2.2 U mL−1) completely inhibited hatching of the root-knot nematode Meloidogyne incognita within 24 h. Moreover, the mortality of juveniles increased to 100% at 48 h post treatment. The results were significantly different (p < 0.05) from Metarhizium spores or protease individually, as well as from extraction buffer and distilled water controls. Metarhizium spores suspended in crude protease rapidly germinated in 24 h and then mycelia invaded the immature eggs of M. incognita, while the eggs containing juveniles and the hatched juveniles were resistant. SEM observations showed extensive damage to eggshell and cuticle when treated with Metarhizium spore and protease, either individually or in combination. This is the first report on biocontrol potential of M. guizhouense against M. incognita. The combination of Metarhizium spores and protease has enhanced potential to develop into a safe and reliable method for plant parasitic nematode control.

Chorionic sculpture of eggs in the subfamily Dismorphiinae (Lepidoptera: Papilionoidea: Pieridae).

This is the second exploration, comparison, and analysis of the chorion of species (45 sspp.) of the subfamily Dismorphiinae (Pieridae). This study includes nearly 50% of the species of the subfamily, including six of the seven genera in its two subtribes: Leptidea (Leptideini), Enantia, Pseudopieris, Lieinix, Moschoneura, and Dismorphia (Dismorphiini). The material studied originates from more than three dozen localities in six different countries on three continents (America, Asia, and Europe) and two biogeographical regions, the Palearctic and Neotropical, over the last 20 years. We have corrected and added information regarding several morphological aspects of the chorion. The precision of the citriform configuration and the elimination of the meloniform shape in the egg of Dismorphiini were determined with detailed observations on the maturation of the chorion in the ovarioles where each stage appears in a linear sequence. We discerned that the meloniform states correspond to incompletely differentiated or immature eggs. This was confirmed by the study of new samples of Dismorphia amphione, D. eunoe, and D. lewyi. The chorion of Dismorphiinae is basically plesiomorphic with respect to those of Coliadinae and Pierinae because it lacks several typical synapomorphies of these subfamilies, such as the presence of micro-grid and/or perimicropylar and apex differentiation, respectively. The eggs of each Dismorphiinae genus can be diagnosed by a combination of chorionic features, although sometimes by one or more plesiomorphies or apomorphies in each genus, with respect to the form or character states in axes, ribs, and poles in the grid of the three regions of the egg - two polar regions and one equatorial (basal, medial, and apical). Leptidea and Enantia show the most generalized grid pattern; however, two genera retain several plesiomorphies with respect to the undifferentiated axes or a small number of short axes (Pseudopieris), as well as many equidistant ribs (Lieinix). The chorionic grid of Moschoneura, although practically lacking short axes, shows the fewest number of axes in the entire subfamily (eight aligns it with Pseudopieris). The chorionic grid in Dismorphia is highly diverse, as it shows the most derived states; however, it comprises symplesiomorphies or atavisms in two groups of species, which aligns them closer to Lieinix or Pseudopieris, but we do not take them into account in some cases where they are convergences or structural parallelisms. It seems that the combination of the shape and its length:width ratio is correlated with the alar configuration (design, sexual dimorphism, and coloring patterns) and separates three groups of species in Dismorphia, and often correlates with the number of ribs. This also coincides with the Batesian participation in the number of mimetic complexes in which a subgroup of species and their stenoecy are integrated within the primary forests. Finally, two schemes are presented that synthesize and illustrate the changes or progression of the form and chorionic grid in the genera of the subfamily.

Trapping system comparisons for and factors affecting populations of Drosophila suzukii and Zaprionus indianus in winter-grown strawberry.

Drosophila suzukii (Matsumura) is a major fruit pest in temperate regions worldwide, but in subtropical Florida, winter-grown strawberries have not been severely affected. Zaprionus indianus Gupta is another invasive drosophilid species and a pest of some tropical fruits. To improve monitoring, trapping systems for D. suzukii and Z. indianus were tested. Morphology, ovarian status and the suitability and availability of non-crop hosts as possible D. suzukii population-limiting factors were assessed. Traps with commercial attractants captured more D. suzukii but fewer Z. indianus than those with a homemade mixture. In central and northern Florida, < 10% and 30-80% of D. suzukii, respectively, exhibited darker, winter morph coloration, and 55-75% of females from central Florida were carrying mature and/or immature eggs. Adult D. suzukii were reared from fruits of two of 28 potential hosts: elderberry (Sambucus nigra) and nightshade (Solanum americanum). Nightshade, but not elderberry, was common on field perimeters (21 and six of 36 fields, respectively). Traps placed in wooded or partially wooded field edges yielded the most D. suzukii. Florida strawberry is at risk of D. suzukii infestation, as flies were captured throughout the growing season. However, fly captures remained relatively low, peaking at 1.5 flies per trap per day. In central Florida, the low availability and suitability of non-crop hosts likely limit population growth. The finding of few flies in northern Florida may additionally be attributable to a greater proportion of flies displaying winter morph coloration than in central Florida. © 2018 Society of Chemical Industry.

Insights into the mode of action of 1,2,6,7-tetraoxaspiro [7.11] nonadecane (N-89) against adult Schistosoma mansoni worms

Control of morbidity associated with schistosomiasis via chemotherapy largely relies on the drug praziquantel. Repeated therapy with praziquantel has created concerns about the possible selection of resistant worms and necessitated the search for novel drugs to treat schistosomiasis. Here, a murine model was infected with Schistosoma mansoni and treated with oral 1,2,6,7-tetraoxaspiro [7.11] nonadecane (N-89), which caused a significant reduction in fecundity and egg burden and reduced morbidity when administered at 5-weeks post-infection.The analysis showed that the mode of action occurred through the ingestion of activated N-89 by the worms, and that there was no direct external effect on the S. mansoni worms. Ultrastructural analysis of the treated worms showed disruptions in the gut lumen and the presence of large volumes of material, suggestive of undigested blood meals or red blood cells. In addition, there were reduced vitelline cells in female worms and damage to sub-tegmental musculature in male worms. Eggs recovered from the treated mice showed both damage to the eggs and the production of immature eggs. Expression of mRNA responsible for gut and digestive function and egg production was also significantly affected by N-89 treatment, whereas control genes for musculature showed no significant changes.Thus, N-89 drastically affected the total digestive function and egg production of S. mansoni worms. Physiological processes requiring heme uptake such as egg production and eggshell formation were subsequently affected, suggesting that the compound could be a possible therapeutic drug candidate for schistosomiasis control.

Assessment of Echinacea purpurea and mefloquine in treatment of Schistosoma mansoni

Objective To assess the effect of combined Echinacea purpurea (EP) and mefloquine (MQ) in the treatment of experimental Schistosoma mansoni infection. Background Schistosomiasis is a prevalent parasitic disease in tropical and subtropical areas caused by the flatworm S. mansoni. Several medications are used in the treatment of schistosomiasis. Praziquantel (PZQ) is the drug of choice because of its safety and broad-spectrum activity. Materials and methods The current study was carried out on 120 laboratory-bred Swiss albino female mice. They were classified into six groups: group I (control), group II (PZQ treated), group III (MQ treated), group IV (EP treated), group V (PZQ and EP), and group VI (MQ and EP). Evaluation was done parasitologically using the worm count technique, ova count technique in intestine and liver, and oogram pattern and histopathological examination of liver sections for assessment of granuloma size and diameter. Moreover, electron microscopic changes of the adult worms were evaluated. Results The highest worm burden reduction of male (79.41%), female (82.28%), couples (86.6%), and total (92%) worms was obtained in the combined MQ and EP treated group. The highest reduction in the mean number of ova count/g tissue in either the intestine (93%) or the liver (89%) was detected in MQ and EP treated group. In combined treated groups, the highest percentage of reduction in the immature and mature eggs was detected in the group MQ and EP; it was 9.16 ± 4.11 and 5.25 ± 2.56, respectively. Moreover, there was an increase in the percentage of dead eggs (85.58 ± 6.43). The highest reductions in hepatic granuloma diameter and number were seen in the combination of MQ and EP at 67 and 85%, respectively (P Conclusion MQ has effective schistosomicidal properties through reduction of worm burden and tissue egg load. Moreover, EP extract showed promising antischistosomal activity on S. mansoni-infected mice. It is recommended to study the exact mechanism of action of EP on S. mansoni and its possible effects on other parasitic infections.

Resource Storage in the Neotropical Social Wasp Mischocyttarus socialis (Saussure, 1854) (Vespidae: Polistini)

Like other Hymenoptera, the social wasps can store liquid and solid resources inside the cells of their nests as reserve for periods of foof shortage. This study describes the occurrence of honey storage in colonies of the neotropical independently-founding wasp Mischocyttarus socialis. During August 2016, in the dry season, 15 colonies in the post-emergence phase were recorded at the botanical garden of Universidade Federal de Juiz de Fora in southwestern of Brazil. Five of the colonies showed honey stored inside empty and immature (eggs) cells. The presence of these two patterns for this species suggests that the storage behavior can be a short-term strategy to feed larvae as soon as they emerge and also a long-term storage to complement population diet during food shortages.

Histopathology of Dryas octopetala leaves co-infected by Subanguina radicicola and Aphelenchoides sp. and molecular caracterization of the nematodes

During a survey, in the Perthshire hills, Scotland, altitude 800–900 m a.s.l., samples of Dryas octopetala leaves showing concomitantly symptoms of galls and discolored spots, were collected. Several nematodes, juveniles and adults, were isolated by dissecting foliar tissues. Two species of nematodes, recovered and identified at morphological and molecular level as Aphelenchoides sp. and Subanguina radicicola, are herein reported. The most significant diagnostic characters for species identification of nematode specimens extracted from Dryas leaf galls fitted well with those previously reported for S. radicicola. Concerning the Aphelenchoides sp., the diagnostic characters were very similar to those reported for A. ritzemabosi.The ITS and the D2-D3 expansion domain analyses confirmed the presence of S. radicicola from galls, while Aphelenchoides specimens from discoloured leaf areas), despite the morphological similarity with A. ritzemabosi, were genetically distinct from A. ritzemabosi based on two genomic regions (LSU and ITS). The 18S rRNA gene (SSU) was only amplified in S. radicicola. Phylogenetic analyses using ITS and LSU sequences placed S. radicicola with other populations of the same species and Aphelenchoides sp. with species of the same genus. The histopathology, caused by both endoparasites in naturally infected leaves, was examined via serial sections mounted on glass slides. Green Dryas leaves infected by Aphelenchoides sp. appeared discolored (yellowish spots), slightly deformed with a population density of four specimens/cm2 per single leaf blade. Dryas octopetala leaves infected by S. radicicola showed an average of three galls/leaf, located randomly, mainly at the upper leaf edge. Dissected galls contained a central cavity with a variable number of adult females (3–6)/gall and males, together with several immature females, juvenile stages and eggs. The present study reports on a new host plant and an undescribed type of sedentary parasitism of the root-gall nematode, S. radicicola on D. octopetala leaves with the concomitant infection of specimens of Aphelenchoides sp.

Age at Menopause: Do Chemical Exposures Play a Role?

Age at Menopause: Do Chemical Exposures Play a Role?

A Parental Care-Mating Dilemma? Potential Risks for Offspring in the Pholcid Spider When Egg-Carrying Females Accept Mating

Many species face a trade-off between additional mating opportunities and the offspring benefits (viability, quality) provided by parental care. Female Holocnemus pluchei spiders must abandon their egg-sac, which they otherwise carry with their chelicerae, to copulate. This may involve risks for the offspring, such as predation and fungal infection. We assessed whether (1) males discriminate between egg-carrying females according to the egg development stage, (2) females are influenced by the egg development stage in regard to their proneness to mate, and (3) offspring are less likely to survive, due to high humidity and/or predation, in the absence of egg-carrying females. Apparently, males did not distinguish between females according to the developmental stage of the eggs they carried. However, females were more likely to mate when carrying relatively mature versus immature eggs (14–18 days and 3–6 days post-oviposition, respectively). All egg-sacs hatched successfully when guarded by egg-carrying females because this avoided both fungal infections (at high humidity) and cannibalism by conspecific spiders. Thus, H. pluchei females may face a trade-off between mating and parental care. Further research should clarify why egg-carrying females mate and how females prevent their egg-sacs from being infected by fungi.

Characterization of the injection funnel during intracytoplasmic sperm injection reflects cytoplasmic maturity of the oocyte

To quantify cytoplasmic maturity on the basis of intracytoplasmic sperm injection (ICSI) injection funnel manifestation and to evaluate influence factors of the latter. Prospective study. Private fertility center. A total of 31 patients with good ovarian response. Mature and immature oocytes were injected intracytoplasmatically. Formation and persistence of an injection funnel was documented and measured. ICSI funnel size, persistence of injection funnel, rates of degeneration and fertilization, embryo quality. Funnel volume in germinal vesicle stage oocytes (prophase I [PI]) was significantly smaller than that of metaphase I (MI) and MII oocytes. Immature eggs (PI, MI) almost never showed a persistent funnel 2-4minutes after ICSI, whereas in MII eggs the funnel was still observable in 35% (117/334) of the cases. Uni- and multivariate analysis revealed that pipette type and stimulation protocol significantly influenced appearance of injection funnel. Funnel volume in oocytes that fertilized regularly was significantly higher compared with three-polar body and degenerated oocytes. Oocyte maturation within the follicle is closely associated with a remarkable change in cytoplasm viscosity from an aqueous to a more viscous subtype. Precise evaluation of the injection funnel may help to explain deviations from expected ICSI outcome and could also assist in optimizing controlled ovarian hyperstimulation.