Search PubMed⌕ Search

SEARCH · Search PubMed

Results for “EGGS”

Search indexed PubMed citations on genomics, clinical trials, systematic reviews and public health. Explore titles, authors and supplied subject terms, then open the PubMed record.

Quote a phrase for an exact phrase match. Source license links do not imply unrestricted reuse.

At least 919 records · Page 51Linked to original sources

The frequency of salmonellae on duck eggs.

Total plate counts on washed duck eggs from a breeder farm on Long Island were less than 30/shell during the winter (January to February) of 1982. Clean unwashed eggs had counts less than 9 X 10(1)/shell, whereas dirty unwashed eggs had counts as high as 9 X 10(5)/shell. Our results showed that washing with a chlorine sanitizer (under commercial conditions) was highly effective in reducing surface bacterial counts on egg shells. Prolonged storage reduced bacterial counts on clean eggs, but it did not significantly affect loads on dirty eggs. No salmonellae could be detected on shells or in the magma of all eggs examined. In a second trial (March 1982) bacterial loads on washed and clean duck eggs from six different breeder farms were low, ranging from too few to count to 10(2)/shell. A higher proportion of dirty eggs were heavily contaminated with counts ranging from 10(5) to 10(6)/shell, but no salmonellae were detected either on shells or in magma. In the third trial (May 1982) bacterial determinations on eggs from breeder ducks that were not confined followed the pattern of the second trial. However, in this trial Salmonella enteritidis was detected on dirty egg shells in four of six farms. In a fourth trial (May 1983), bacterial loads on washed and nest-clean eggs from the same breeder farms (not confined) ranged between 10(2) to 10(3)/shell and 10(2) to 10(4)/shell, respectively. S. enteritidis and S. badar were recovered from washed, nest clean, and dirty eggs in two of six farms. We conclude that proper egg washing and confinement of duck breeders should minimize the problem of salmonellosis in ducklings.

Animals↗

Effect of ascorbic acid and cooling during egg incubation on hatchability, culling, mortality, and the body weights of broiler chickens.

Five experiments involving 3,100 settable eggs with living embryos of commercial broiler parent stock were conducted to determine the effect of ascorbic acid (AA) and cooling during egg incubation on embryonic weight, hatchability, percentage of cull chicks, embryonic mortality, and body weights of the hatched chicks. The treatments were carried out at 15, 15, 17, 11 and 19 d of incubation for Experiments 1, 2, 3, 4, and 5, respectively. The treatments for the experiments were as follows: 1) eggs injected with 0.1 mL of sterile saline solution; 2) eggs injected with saline containing 0.5, 1, 3, and 12 mg of AA per egg; and 3) control for Experiment 1; 1) eggs injected with 3 mg of AA per egg; 2) eggs cooled at 22 C for 24 h; and 3) control for Experiments 2 and 3; 1) eggs injected with 3 mg AA per egg; and 2) control for Experiments 4 and 5. Ascorbic acid at a dose of 3 mg during egg incubation improved hatchability and body weight, and decreased number of cull chicks, whereas a dose of 12 mg decreased embryo weight and hatchability and increased the number of cull chicks and embryonic mortality. Injection of AA hatchability improved when injected on Days 11 and 15 of incubation; effects diminished if AA was injected on Day 19 of incubation. Cooling eggs at 22 C for 24 h had no significant effect on any of the variables studied when compared with controls. Comparison of AA injection at a dose of 3 mg per egg and egg cooling to 22 C for 24 h at 15 to 17 d of incubation led to the conclusion that AA had two advantages over cooling. These advantages were 1) low embryonic death and 2) greater chick weight at hatch and on Days 15 and 17 of incubation.

Animals↗

Persistence of eggs and hepatic fibrosis after treatment of Schistosoma mansoni-infected mice.

In 1971 we estimated that Schistosoma mansoni eggs in the tissues of mice were destroyed with an approximate half-life of four weeks. Our present results of five experiments suggest that egg destruction is not as rapid, and no significant destruction of eggs was detected for up to 26 weeks after treatment. However, in these experiments, a mean of 60% of the eggs in intestinal tissues were found in the feces at the time of treatment. In previously reported experiments, only 15% of gut eggs were passed in the feces. We now believe that underestimation of the number of eggs passed in the feces led to an overestimation of the number of eggs destroyed in the tissues. We analyzed liver eggs separately because eggs lost from this site are unaffected by eggs passed in the feces. No significant decrease in liver eggs occurred in the present experiments, but reanalysis of previously published data showed significant egg destruction in the liver in several experiments, although at a much slower rate than previously estimated. However, inspection of the data in the previously published and present experiments does not show a convincing difference in the number of eggs in the liver after treatment. The persistence of egg shells is probably not important in the pathogenesis of disease, but is of concern in calculating worm fecundity. Hepatic collagen levels increased markedly two weeks after treatment and subsequently decreased significantly in some, but not all, experiments.

Analysis of Variance↗

Egg consumption patterns and Salmonella risk in Finland.

To estimate the consumer risk of contracting Salmonella infection via shell eggs and to evaluate the effect of possible preventative measures, quantitative microbiological risk assessment is being developed in Finland. As a part of the risk assessment, a survey of 918 respondents was conducted to study how households purchase, store, handle, and use eggs. In addition, suitability of the Internet as a survey method was compared with a postal survey. Shell eggs were usually purchased once every 2 weeks (41% of all the respondents). Ninety-one percent of the respondents bought eggs in groceries and 93% stored eggs at chilled temperatures. The majority of the respondents (80%) only had eggs in their home for which the best-before date had not expired. Only 34% of the respondents said that they always washed their hands after breaking eggs. Consumption of well-cooked eggs accounted for 84%, consumption of soft-boiled eggs for 12%, and consumption of raw eggs for 4% of the total amount of eggs consumed. The elderly used eggs more frequently than the whole population, but the consumption of raw egg dishes decreased with age. The Internet survey was a rapid method for transmitting information, but its response rate was low (9%), and it did not appear to be a suitable tool for data collection in a general population. The results indicate that although the majority of the respondents had safe egg-handling practices, a substantial minority of the consumers had risk-prone behavior.

Adult↗

Electron miscroscopy and histochemical studies on four Egyptian helminthes eggs of medical importance.

By SEM the Fasciola gigantica egg is ovoid with a small knob like operculum, while the egg of Heterophyes heterophytes is broad oval with the operculum more tapering. The egg shell of fertilized Ascaris lumbricoides has interconnected ridges and peak-like projections, while the egg of Enterobius vermicularis is flattened with a thicker margin at the curved side. By TEM, Fasciola egg shell consists of fine reticulum fibrils of three layers. The outer lipoprotein of perivitelline membrane beneath which 2 membranes separated by inclusions, middle of protein globules and inner lipoprotein layer with minute electron-dense granules of melanin or polymer origin, in some parts of the shell giving the egg its brown coloration. The Heterophyes egg shell is more or less similar to that of Fasciola but lacking the minute electron-dense granules. The egg shell of Ascaris has outer ulterine layer with three consecutive layers, basal lipoprotein layer and the inner lipid or ascaroside layer which is the most resistant layer. The Enterobius egg shell consists of five layers, external uterine, internal uterine, vitelline, chitinous and lipid layer. Histochemically, Fasciola egg shell consists of nine amino-acids, and that of Heterophyes consists of ten amino acids. In Ascaris, the lipid layer characteristically consists 25% protein and 75% lipid. The histochemical examination of Enterobius as a detailed example, showed different degrees of reactions with mercuric bromophenol blue, diazotization coupling, Sakaguchi reaction, Sudan black and Mallory's triple stain. Temperature showed marked effect on eggs survival. Eggs of Fasciola and Heterophyes withstand more low temperatures but those of Ascaris and Enterobius withstand more high ones. There are marked correlations between the egg shell constitution, histochemical compositions on one hand and water permeability and egg dryness on the other hand. The results were photographed and discussed.

Animals↗

Determination of egg proteins in snack food and noodles.

Egg is one of the 5 major allergenic foods that are responsible for more than 3/4 of food allergies in children. Food-allergic responses can be controlled by avoidance of the offending foods. The applicability of a commercial enzyme-linked immunosorbent assay (ELISA) kit for the detection of egg in food products such as cookies, crackers, pretzels, salad dressings, and raw and cooked noodles was evaluated. A preliminary evaluation of an antibody-based biosensor was also performed. A National Institute of Standards and Technology (NIST) whole dried egg powder reference material, SRM 8415, was used as a standard. A homogeneous and stable aqueous egg suspension was prepared for the evaluation of the performance of the Veratox for Egg Allergen Test (Neogen Corp., Lansing, MI). This test does not detect egg yolk proteins. Each gram of the aqueous dried egg suspension contained 643 microg whole dried egg, 0.5 mg thimerosal, and 2.5 mg bovine serum albumin. When cookies, crackers, salad dressings, noodles, and ice cream were spiked at a level of 24 mg/kg SRM 8415, recoveries for whole egg averaged about 28%. All foods containing egg as indicated on the ingredient label were found positive by the Veratox test. No false positives occurred in samples that did not contain eggs. Similar results were obtained using the Naval Research Laboratory (NRL) array biosensor, an evanescent wave fluoroimmunosensor. Results for cooked noodles showed that they contained <1% of the egg found in uncooked noodles. A comparison of extracts from cooked and uncooked noodles by sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) revealed differences in protein profiles. The boiling of the noodles could have reduced the immunoreactivity of the egg proteins to the antibodies used in the kit or rendered the egg proteins nonextractable.

Allergens↗

Optimum site for in ovo amino acid injection in broiler breeder eggs.

Three experiments were conducted to evaluate the effect of differences in in ovo amino acid (AA) injection sites in broiler breeder eggs on subsequent hatchability and BW of chicks. In Experiment 1, an AA solution was injected into eggs with 13-mm or 19-mm, 27-ga needles. Uninjected eggs served as controls. Hatchability was decreased (P < 0.05) in eggs receiving AA injections with the 19-mm needle in comparison to the control and 13-mm-injected groups. However, BW of chicks increased (P < 0.05) relative to pre-incubational egg weight by AA injection with the 13-mm needle. In order to evaluate the in ovo location of AA injections from Experiment 1, India ink was injected into eggs in Experiment 2 with a 13-or 19-mm needle. Immediately after injection, the air cell end of the egg was windowed in order to observe effects of injection site. Windowing of eggs was accomplished by removing a piece of the eggshell over the air cell and the underlying membrane at Day 7 of incubation. The amount of injected India ink was higher in the extra-embryonic coelom in eggs treated by both needles. However, the occurrence of India ink in the extra embryonic coelom was higher (P < 0.05) in the group injected with AA solution using a 13-mm needle as compared to that after injection using a 19-mm needle. The observation of India ink in the amniotic cavity was higher (P < 0.05) in the group injected with AA solution using a 19-mm needle rather than that using a 13-mm needle. In Experiment 3, treatments consisted of control (uninjected eggs) or windowed eggs. Windowed eggs received AA to the chorioallantoic membrane, the yolk, extra-embryonic coelom, or amniotic cavity at Day 7 of incubation. Hatchability was reduced, but chicks hatched when eggs were windowed and when AA were injected into the yolk sac or extra-embryonic coelom. However, chicks did not hatch when AA were administered to the chorioallantoic membrane or into the amniotic cavity. These results suggest that the best AA injection sites in ovo may be the yolk and extra-embryonic coelom.

Amino Acids↗

Ecdysteroid and free amino acid content of eggs of the Colorado potato beetle, Leptinotarsa decemlineata.

In order to identify components of the Colorado potato beetle (CPB) egg that may be required by Edovum puttleri, a parasitic wasp that parasitizes the CPB egg, to complete development, ecdysteroid and free amino acid content of CPB eggs were analyzed by reversed phase high pressure liquid chromatography followed by radioimmunoassay to identify ecdysteroids. Ecdysteroid titers were relatively low (<300 pg/egg) through day 2 post-oviposition and then increased sharply, reaching concentrations >2,500 pg/egg on day 3 post-oviposition. Ecdysone (E), 20-hydroxyecdysone (20E), and polar conjugates of E were prominent ecdysteroids present in eggs sampled on days 0 and 1 post-ecdysis, and E, 20E, three peaks containing more polar ecdysteroids (metabolic inactivation products), and polar conjugates of E were present in eggs sampled on day 2. Thus, at a time when parasitization of CPB eggs by E. puttleri is relatively high (0-48 h), physiologically-active ecdysteroids (20E and perhaps E are physiologically active) are present at concentrations between 50 and 200 pg/egg. Ecdysone and 20E reached their highest levels in day-3 eggs, indicating that ecdysteroid may direct physiological processes associated with the completion of CPB embryonic development. In day-4 eggs, the concentration of E and 20E fall dramatically and polar metabolites of E and/or 20E are now responsible for the high ecdysteroid content of the eggs. Interestingly, conjugates of E decrease to relatively low levels in day-3 eggs and are absent in day-4 eggs. Therefore, it is likely that the increase in E in day-3 eggs is due, in part, to the breakdown of polar conjugates of E. Nine amino acids were present in significant quantities in eggs sampled at various times between 0 and 48 h post-oviposition. These include histidine, glutamine, proline, asparagine, serine, glutamic acid, threonine, lysine, and tyrosine. The first three amino acids were present at concentrations that were approximately 2 to 6 times greater than the concentrations of the last six amino acids. Amounts of most of the free amino acids varied with the age of the eggs from which the extract was prepared, but in general, there was no correlation between the levels at times of maximum parasitization (0 and 30 h) and the levels at the less favored times of parasitization (16 and 48 h). This information should facilitate the development of diets for both parasites and predators of pest species of beetles. Arch. Insect Biochem. Physiol. 44:172-182, 2000. Published 2000 Wiley-Liss, Inc.

Amino Acids↗

A study of hetero-specific sperm-egg interactions in the rat, mouse, and deer mouse using in vitro fertilization and sperm injection.

Hetero-specific fertilization of zone-free eggs is used in these experiments as a tool to analyze the barriers to hybridization and to gain insight into the mechanisms of normal fertilization. When the zonae of rat eggs, which are a barrier to hetero-specific fertilization, are removed with pronase, the eggs can be fertilized by mouse sperm and the zygotes start to develop normally. A rat egg fertilized with mouse sperm completes meiosis and forms both male and female pronuclei. Chromosomes from both parents are found on he spindle at the metaphase stage of the first cleavage division. Under present culture conditions, embryos develop only to the two-cell stage, but this initial development of the hybrid is apparently normal. The question of whether sperm and egg membrane fusion is requisite for normal development is addressed by injecting sperm directly into the cytoplasm of unfertilized eggs. The injection of mouse sperm into rat eggs frequently leads to activation and formation of male and female pronuclei. The first cleavage division is indistinguishable from that following hetero-specific fertilization. Capacitated and uncapacitated sperm react alike when injected into eggs. Egg activation, however, is necessary for male pronucleus formation. Sperm from the deer mouse Peromyscus maniculatus bairdii, which are incapable of fertilizing even zonea-free eggs, respond like mouse sperm when injected into rat eggs. These data indicate that sperm interactions with the egg cytoplasm are less species-specific than interactions at the egg surface. Furthermore, the normal surface interactions of sperm and eggs are not essential for the start of development.

Animals↗

The distribution of polymerized actin in the rat egg and its sensitivity to cytochalasin B during fertilization.

The distribution of polymerized actin in rat eggs fertilized in vitro was determined using NBD-phallacidin (NBD-ph). Unfertilized and fertilized eggs exhibited a 3-5-micron-thick band of fluorescence that encompassed the entire cortical cytoplasm. There was no dramatic increase in the staining of the cortex in association with any component of the fertilizing sperm during its incorporation into the egg. Unfertilized eggs and fertilized eggs obtained at intervals after sperm-egg fusion were treated with cytochalasin B (CB; 5 micrograms/ml) and subsequently stained with NBD-ph. Unfertilized eggs treated with CB exhibited a continuous ring of cortical staining identical to that seen in untreated eggs. Eggs treated with CB 15 min after sperm-egg fusion exhibited small gaps in the cortical staining pattern, whereas those exposed to CB 1 hr after fusion exhibited larger gaps and the staining pattern appeared punctate. This pattern could be seen throughout the remainder of the 7 hr period of sperm incorporation and for at least 13 hr thereafter. CB-treated fertilized eggs that were washed to remove the drug again exhibited uninterrupted cortical staining on treatment with NBD-ph. CB also induced the resorption of surface elevations that are normally seen on the eggs during sperm incorporation, but it did not affect the morphology of unfertilized eggs. The sensitivity to CB during fertilization coincides with the onset of a variety of egg shape changes that occur during the period of sperm incorporation (Battaglia and Gaddum-Rosse, Gamete Res., 10:107-118, 1984a).(ABSTRACT TRUNCATED AT 250 WORDS)

Actins↗

Embryonic osmoregulation: consequences of high and low water loss during incubation of the chicken egg.

The rates of water loss of domestic chicken eggs were varied during incubation to measure the osmoregulatory ability of the avian embryo. Egg water loss was increased by drilling holes in the eggshell over the airspace on day 13 (I = 21 days) and then placing these eggs in a low relative humidity (r.h.: 0-10%) incubator until hatch. Egg water loss was decreased by placing other eggs in a high-r.h. (85-90%) incubator on day 0. Eggs with low water loss (approximately 6% of initial fresh mass [IFM]) produced embryos and yolks that were not different in wet or dry mass when compared to control eggs that lost approximately 12% of IFM. However, 1-4 gm of excess albumen were left in low-water-loss eggs on day 21. Hatching success was 71% and 89% for low and control eggs, respectively. Low egg water loss did not appear to disturb embryonic growth. The allantoic fluid volume and millimolar allantoic Na+ and Cl- ions declined faster with high and slower with low rates of water loss. Thus, excess water was lost as a result of increased movement of water out of allantoic fluid, which was due to increased active transport of Na+ ions by the chorioallantoic membrane (CAM). Eggs with high water loss had elevated Cl- levels after day 17 in plasma and amniotic fluid, which indicated a period of osmotic stress after depletion of allantoic fluid between day 18 and hatch. The decrease in wet embryo mass measured in embryos from high-water-loss eggs was due principally to dehydration of skin. Embryonic skin may serve as an emergency water reservoir during osmotic stress. Dehydrated chicks produced from high-water-loss eggs were 6 gm less in wet mass at hatch compared to controls. However, these chicks regained the water deficit 7 days after hatch and grew at a rate not different from control chicks through 6 weeks of age. Total egg water loss of 12% of IFM results in highest hatching success. However, water losses between 6% and 20% of IFM do not appear to affect adversely the growth or water content of the chick. Water losses above 20% of IFM cause early depletion of allantoic fluid, prolong the period of osmotic stress, and result in subsequent dehydration of blood, amniotic fluid, and embryonic skin.(ABSTRACT TRUNCATED AT 400 WORDS)

Allantois↗

The surface events of fertilization: the movements of the spermatozoon through the sea urchin egg surface and the roles of the surface layers.

The sea urchin egg surface at fertilization has been examined with the scanning electron microscope to reveal the movements of the spermatozoon from the exterior, through the surface layers, and into the egg cytoplasm. The layers that the spermatozoon encounter have been studied to determine their physical and chemical natures and their role in early development. By studying the outside of whole eggs and the inner face of surfaces isolated shortly after fertilization, it has been possible to compile data on the movements of the spermatozoon through the egg surface. The spermatozoon initially contacts the egg with the elongated acrosomal process. The vitelline sheet, the outermost layer of the egg, separates slightly next to the attached spermatozoon. As membrane fusion between the gametes occurs, the plasma membrane from the egg engulfs the spermhead, the cortical granules start to discharge their contents, and a spreading surface deformation, concommitant with a distortion of the fibrous cortex, is initiated. A cluster of elongate microville surround the perpendicularly fusing spermatozoon. These microvilli interidigitate as the spermatozoon is forced to lie upon the egg surface between the plasma membrane and the matrix of cortical fibers. The spermatozoon then rotates additionally to enter the egg cytoplasm with the posterior end first; it has rotated 180 degrees through the cell surface. Finally, it detaches into the egg cytoplasm, leaving a scar in the cortex through which it penetrated. The egg cortex, previously unobserved by electron microscopy, is revealed to be composed of 50-200 nm fibers. At fertilization they are uniformly organized but during later development this order is lost. The cortex is from 0.2-0.5 micronm thick and is a contractile structure. The role of the outer surface in releasing the cell from the metabolic constraints of the unfertilized egg is shown, and the apparent differences in the mobilities of the membranes derived from the sperm and from the egg are demonstrated. The relation of these layers to the movements of the spermatozoon, to the activation of the egg, to the block to polyspermy, and to each other are discussed.

Acrosome↗

Observations of hamster sperm-egg fusion in freeze-fracture replicas including the use of filipin as a sterol marker.

We have extended the observations of previous transmission electron microscopy studies of sperm-egg fusion to include those of freeze-fracture replicas showing sperm-egg interactions before, during, and following sperm head fusion with the egg membrane. Hamster eggs were incubated with hamster sperm under polyspermic conditions and were observed after a period of 5-30 minutes. After fixation, the eggs and sperm were exposed to filipin, which binds beta-OH-sterols to form visible complexes in freeze-fracture replicas. Filipin can act as a marker for egg plasma membrane wherein it is abundant, while filipin is relatively scarce in the acrosome-reacted hamster sperm membrane, found only in the plasma membrane of the equatorial segment. The earliest sperm-egg interactions are observed between the egg microvilli and the perforatorium and the equatorial segment of the sperm, and the initial fusion between egg and sperm occurs in the vicinity of the equatorial segment. At later stages of fusion involving the postacrosomal segment, a clear line of demarcation is observed between the filipin-rich egg membrane and the filipin-poor sperm postacrosomal segment, suggesting that filipin binding lipids from the egg intercalate into the sperm membrane following membrane fusion. The anterior segment of the sperm does not fuse with the egg but is instead incorporated into a cytoplasmic vesicle derived from both sperm and egg membranes. In this latter step, filipin-sterol complexes are not found in sperm-derived membranes suggesting that there may be barriers to the movement of filipin binding lipids from the egg into these sperm membranes.

Animals↗

Successive electrical responses to insemination and concurrent sperm entries in the polyspermic egg of the ctenophore Beroe ovata.

In the ctenophore Beroe ovata, action potentials and sperm-induced egg electrical responses were recorded, in unfertilized and in vitro inseminated eggs, respectively. The unfertilized egg had a resting membrane potential of -65.6 +/- 1.3 mV and a membrane resistance of 1.45 +/- 0.09 M omega. When depolarized beyond a threshold of about -20 mV, the membrane displayed an action potential with a fast phase and then a peak reaching 38 +/- 0.6 mV, followed by a positive declining plateau which lasted about 20 sec and ended by a biphasic repolarization of the membrane. The action potential was mainly Ca(2+)-dependent and to a lesser extent, Na(+)-dependent. Upon one or several inseminations, monospermic eggs generated one fertilization potential and polyspermic eggs generated several sperm induced egg electrical responses. These sperm-induced egg electrical responses were composed of a voltage-gated action potential elicited by sperm-egg contact, resembling the action potential triggered by a depolarizing current, followed by a true sperm-gated depolarizing plateau which lasted about 60 sec and was mainly Na(+)-dependent. The number of electrical responses displayed by each egg corresponded to the number of male pronuclei detected in its cortex. An electrical response occurred for all egg membrane potentials ranging from -78 to +16 mV. In addition, the histogram giving the number of eggs in each class of egg characterized by a number of sperm entries indicated that each sperm entry constituted an independent process. However, when we compared the effectiveness of the in vitro inseminations during electrophysiological measurements and the in vivo inseminations under simulated natural conditions, the results showed two predominant classes of fertilized eggs, with one and two male pronuclei in the cortex respectively. Consequently, although our overall results exhibit features characteristic of polyspermy, they do not yet preclude the possibility that the eggs of the ctenophore B. ovata might be monospermic, or might, under natural planktonic conditions, display a small number of sperm entries.

Action Potentials↗

The sperm-induced Ca2+ wave following fertilization of the Xenopus egg requires the production of Ins(1, 4, 5)P3.

We used fluorescence ratio imaging of fura-2 in the egg of Xenopus laevis to study the initiation and propagation of the wave of increased free Ca2+ that is normally triggered at fertilization. Naturally matured, jellied eggs were injected with fura-2 and ratio-imaged with fluorescence excitation at 350 and 385 nm while sperm were added. The [Ca2+]i rise normally begins as a small spot near the surface of the egg, remains fairly localized for 20-60 sec, and then spreads more rapidly across the egg at 7.5 +/- 0.05 microns/sec to reach the antipode about 5 min after fertilization. The [Ca2+]i wave velocity is slowed by increasing the concentration of fura-2 in the cytoplasm to 100 microM, and 250 microM fura-2 blocks wave propagation. The peak [Ca2+]i in a sperm-activated wave is 2.2 +/- 0.1 microM and [Ca2+]i returns to preactivation levels within 21 +/- 0.7 min after fertilization. We further studied the mechanism by which sperm trigger the Ca2+ wave by injecting substances that interfere with Ins(1,4,5)P3-induced Ca2+ release or Ca(2+)-induced Ca2+ release (CICR). Heparin (3 kDa) inhibits sperm-induced egg activation in a manner that is linearly proportional to its cytoplasmic concentration. At 130 microM (390 micrograms/ml), sperm-induced activation is completely blocked and at 75 microM (225 micrograms/ml) activation of half of the eggs is inhibited. All eggs injected with 130 microM heparin are polyspermic as verified using Hoechst dye to label nuclear DNA. Imaging eggs injected with 75 microM heparin revealed multiple, transient "spots" of increased [Ca2+]i that failed to spread across the egg. Injection of a monoclonal antibody to PIP2 (0.2 microM), kt3g, blocked sperm-induced egg activation in 73% of the 30 eggs injected, suggesting that activation requires the hydrolysis of PIP2 in the membranes of the egg rather than simply the introduction of Ins(1,4,5)P3 from the sperm. This sperm-induced egg activation is not blocked by either of two CICR inhibitors, procaine (10 mM) or ruthenium red (30 microM), and egg activation is not triggered by either of two stimulators of CICR, caffeine (10 mM) or ryanodine (50 microM).(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗

Fertilization stimulates an increase in inositol trisphosphate and inositol lipid levels in Xenopus eggs.

Previous experiments from our lab have suggested that the hydrolysis of phosphatidylinositol 4,5-bisphosphate (PIP2) is required for sperm-induced egg activation in Xenopus laevis. Here we measure the endogenous production of both Ins(1,4,5)P3 and PIP2 during the sperm-induced and ionomycin-induced calcium wave in the egg and find that both increase following fertilization. Ins(1,4,5)P3 increases 3.2-fold from an unfertilized egg level of 0.13 pmole per egg (0.29 microM) to a peak of 0.42 pmole per egg (0.93 microM) as the calcium wave reaches the antipode in the fertilized egg. This continuous production of Ins(1,4,5)P3 during the time that the Ca2+ wave is propagating across the egg suggests the involvement of Ins(1,4,5)P3 in wave propagation. This increase in Ins(1,4,5)P3 is smaller in ionomycin-activated eggs than in sperm-activated eggs, suggesting that the sperm-induced production of Ins(1,4,5)P3 involves a PIP2 hydrolysis pathway that is not simply raising intracellular Ca2+. While one might expect PIP2 levels to fall as a result of hydrolysis, we find that PIP2 actually increases 2-fold. The total lipid fraction in unfertilized egg exhibits 0.8 pmole PIP2 per egg and this increases to 1.5 pmole as the calcium wave reaches the antipode. The PIP2 concentration peaks 2 min after the completion of the calcium wave at 1.8 pmole per egg. The amount of PIP2 in the animal and vegetal hemispheres of the egg was also measured by cutting frozen eggs in half. The vegetal hemisphere contained twice the amount of PIP2 as the animal hemisphere but it also contained twice the amount of lipid. Thus, there was an equivalent amount of PIP2 normalized to lipid in each hemisphere. Isolated animal and vegetal hemisphere cortices exhibit similar PIP2 concentrations, suggesting that the 2-fold higher total PIP2 in the vegetal half is not due to a gradient of PIP2 in the plasma membrane, but rather implies that cytoplasmic organelle membranes also contain PIP2.

Animals↗

Transmission of injected DNA sequences to multiple eggs of Metaseiulus occidentalis and Amblyseius finlandicus (Acari: Phytoseiidae) following maternal microinjection.

The persistence of DNA injected into two species of adult female phytoseiids and its transmission to serial eggs deposited by them was assessed by the polymerase chain reaction (PCR). The effect of DNA concentration on persistence and transmission was examined in Metaseiulus occidentalis. M. occidentalis females were microinjected with plasmid DNA at three different concentrations (250, 500, 750 ng microL-1) and allowed to deposit one to five eggs before the females and their last eggs were analyzed. Plasmid DNA was found in 82% of the females assayed and in 70% of all the eggs analyzed (including the fifth eggs produced after microinjection). Transmission of DNA to multiple eggs was also examined in Amblyseius finlandicus. Females of this species are less traumatized by microinjection allowing analysis of transmission over a more extended number of eggs. Females were microinjected and allowed to deposit eggs until their death. DNA from every fifth egg was analyzed by the PCR. PCR products were amplified from 51% of the eggs and from all egg classes except the 30th egg. The persistence and presence of plasmid DNA in both eggs and females suggests that (1) maternal microinjection is a more efficient method for DNA delivery than traditional egg microinjection, (2) it may be possible to isolate transformants from fewer maternally-microinjected females than originally expected, and (3) maternal microinjection could be useful as a DNA delivery system in other phytoseiids.

Animals↗

Culturable and VBNC Vibrio cholerae: interactions with chironomid egg masses and their bacterial population.

Vibrio cholerae, the etiologic agent of cholera, is autochthonous to various aquatic environments. Recently, it was found that chironomid (nonbiting midges) egg masses serve as a reservoir for the cholera bacterium and that flying chironomid adults are possible windborne carriers of V. cholerae non-O1 non-O139. Chironomids are the most widely distributed insect in freshwater. Females deposit egg masses at the water's edge, and each egg mass contains eggs embedded in a gelatinous matrix. Hemagglutinin/protease, an extracellular enzyme of V. cholerae, was found to degrade chironomid egg masses and to prevent them from hatching. In a yearly survey, chironomid populations and the V. cholerae in their egg masses followed phenological succession and interaction of host-pathogen population dynamics. In this report, it is shown via FISH technique that most of the V. cholerae inhabiting the egg mass are in the viable but nonculturable (VBNC) state. The diversity of culturable bacteria from chironomid egg masses collected from two freshwater habitats was determined. In addition to V. cholerae, representatives of the following genera were isolated: Acinetobacter, Aeromonas, Klebsiella, Shewanella, Pseudomonas, Paracoccus, Exiguobacterium, and unidentified bacteria. Three important human pathogens, Aeromonas veronii, A. caviae, and A. hydrophila, were isolated from chironomid egg masses, indicating that chironomid egg masses may be a natural reservoir for pathogenic Aeromonas species in addition to V. cholerae. All isolates of V. cholerae were capable of degrading chironomid egg masses. This may help explain their host-pathogen relationship with chironomids. In contrast, almost none of the other bacteria that were isolated from the egg masses possessed this ability. Studying the interaction between chironomid egg masses, the bacteria inhabiting them, and V. cholerae could contribute to our understanding of the nature of the V. cholerae-egg mass interactions.

Animals↗