Criteria for identification of Schistosoma mansoni eggs in faecal sediments prepared with the Helmintex method and stained by ninhydrin

Helmintex is a sensitive method used for detecting Schistosoma mansoni eggs. Here, we describe the observed frequency of six proposed criteria associated with the identification of S. mansoni eggs prepared with the Helmintex method and stained with ninhydrin. The efficacy of these criteria in classifying S. mansoni eggs when applied in various combinations was also examined. Nine observers registered the presence or absence of 6 different criteria in 100 eggs using a microscope at 100x magnification. Ninhydrin purple, which was frequently observed, was the criterion associated with the lowest inter-observer variability. At least three criteria were associated with a significantly better performance in egg identification. In conclusion, ninhydrin staining and a combination of criteria are recommended for microscope examination of faecal sediments.

Morphology of eggs of Dioctophyme renale Goeze, 1782 (Nematoda: Dioctophymatidae) and influences of temperature on development of first-stage larvae in the eggs / Morfologia de ovos de Dioctophyme renale Goeze, 1782 (Nematoda: Dioctophymatidae) e influência da temperatura no desenvolvimento de larvas de primeiro estágio nos ovos

This study aims to provide information on morphological data of D. renale eggs, as well as on first-stage larvae development into eggs kept at different temperatures. Eggs were obtained by centrifugation of infected dog urine, placed in Petri dishes, and stored in BOD chamber for a 90-day period. Each treatment (GI - 15 ºC, GII - 20 ºC, and GIII - 26 ºC) was repeated five times. Eggs showed average measures of 67.23 x 42.78 µm, and the mean incubation time was inversely proportional to the incubation temperature. Larvae motility was observed one week after being observed in eggs.

Este estudo teve a finalidade de fornecer dados morfológicos de ovos de D. renale e do desenvolvimento de larvas de primeiro estádio em ovos mantidos em diferentes temperaturas. Os ovos foram obtidos por centrífugação da urina de cães parasitados e colocados em placas de Petri em estufa BOD, durante 90 dias. O experimento consistiu de três tratamentos (GI - 15 ºC, GII - 20 ºC e GIII - 26 ºC) com cinco repetições cada. Os ovos apresentaram tamanho médio de 67,23 x 42,78 µm, e o tempo médio de incubação foi inversamente proporcional à temperatura de incubação e as larvas apresentaram motilidade por aproximadamente uma semana após sua formação.

Identification of differentially expressed genes in developing cotton fibers (Gossypium hirsutum L) through differential display

Cotton fibers are differentiated, non-dividing cells that originate from the epidermal layer of developing ovules. To identify genes involved in cotton fiber development, we performed non-radioactive differential display reverse transcriptase PCR (DDRT-PCR) on the purified mRNA. This technique was tested on mRNA isolated from five different developmental stages of cotton fiber including 0, 5, 10, 15 and 20 DPA (days after pollination). The mRNA purified from total RNA was reversibly transcribed using three anchored oligo-dT primers. Polymerase chain reaction (PCR) amplification of each cDNA preparation was carried out in combination with seven arbitrary primers. The amplified products were resolved on 1 percent agarose gel containing ethidium bromide. DNA was extracted from seventeen differentially expressed bands and cloned in pTZ57R/T vector. The sequencing and BLAST search analysis indicated that 12 of the differentially expressed genes matched the previously characterized genes, while 3 of them matched the uncharacterized sequences of cotton fiber expressed sequence tags (ESTs) reported previously to be associated with cotton fiber and 2 of the clones had homology with putative proteins. The technique can be used to efficiently identify differentially expressed genes and can be expanded to large scale studies by increasing the number of random decamers.

Huevo, larva en primer estadio y aparato genital femenino de la mariposa Chabuata castanea (Lepidoptera: Noctuidae)

Egg, first larval stage and female genitalia of the moth Chabuata castanea (Lepidoptera: Noctuidae). Egg, first larval stage, and female genitalia of the widely distributed moth Chabuata castanea are described, based on material from Talcahuano, VIII region, Chile. Egg microestructures are illustrated with scannig electron microscope images which show that egg morphology allows identification to species level. Rev. Biol. Trop. 55 (2): 659-664. Epub 2007 June, 29.

Se describe el huevo, larva de primer estadio y aparato genital de la hembra de Chabuata castanea con material proveniente de Talcahuano, VIII región, Chile y de huevos obtenidos en laboratorio. Se fotografiaron los huevos con microscopia electrónica de barrido para analizar las variaciones entre micropila, celdas primarias y secundarias, concluyéndose que las diferencias permiten una identificación al nivel de especie.

Comparison between morphological and staining characteristics of live and dead eggs of Schistosoma mansoni

Schistosoma mansoni eggs are classified, according to morphological characteristics, as follows: viable mature and immature eggs; dead mature and immature eggs, shells and granulomas. The scope of this study was to compare the staining characteristics of different morphological types of eggs in the presence of fluorescent labels and vital dyes, aiming at differentiating live and dead eggs. The eggs were obtained from the intestines of infected mice, and put into saline 0.85 percent. The fluorescent labels were Hoechst 33258 and Acridine Orange + Ethidium Bromide and vital dyes (Trypan Blue 0.4 percent and Neutral Red 1 percent). When labelled with the probe Hoechst 33258, some immature eggs, morphologically considered viable, presented fluorescence (a staining characteristic detected only in dead eggs); mature eggs did not present fluorescence, and the other types of dead eggs, morphologically defined, showed fluorescence. As far as Acridine Orange + Ethidium Bromide are concerned, either the eggs considered to be live, or the dead ones, presented staining with green color, and only the hatched and motionless miracidium was stained with an orange color. Trypan Blue was not able to stain the eggs, considered to be dead but only dead miracidia which had emerged out of the shell. Neutral Red stained both live and dead eggs. Only the fluorescent Hoechst 33258 can be considered a useful tool for differentiation between dead and live eggs.

Methylene blue staining method for identification of Opisthorchis viverrini egg.

Methylene blue staining method was used to distinguish O. viverrini eggs from Haplorchis taichui and Prosthodendrium molenkampi eggs. All eggs were obtained from dissected adult worms, fixed in 10% formalin, and stained with methylene blue prior to light microscopy observation. The distinct musk-melon-like texture of the O. viverini eggshell surface and the thread-like texture of H. taichui eggshell surface were recognized, while P. molenkampi eggs showed a smooth eggshell. We also evaluated the sensitivity and specificity of the method by training investigators to differentiate surface textures. After training, the investigators were randomly tested with 10 slides containing fluke eggs. The sensitivity and specificity were 95% and 95%, respectively.

Superovulation in vesper mouse Calomys musculinus (Rodentia-Sigmodontinae)

Optimal conditions for pharmacological induction of ovulation of vesper mouse, Calomys musculinus, were analyzed. The best superovulation (a mean of about 21 eggs per female, range 12-45) was induced by the administration of 12 IU of PMSG followed 48 hr later by injection of 15 IU of hCG. Ovulation started about 10 hr after administration of hCG and was completed during the next 4-5 hr. The induction of ovulation was achieved irrespective of the stage of the oestrus cycle at the moment of PMSG administration. The majority of females (105, 82.7) responded to the treatment with either an ovulatory (53.4) or superovulatory (49.7) response. Oocyte recovery and egg quality were clearly influenced by the age of females, 30 days to more than 120 days old. The majority (90.3) of superovulated eggs was morphologically normal, and only a small proportion of eggs showed morphological abnormalities (7.4) or were spontaneously activated (2.3). Superovulated oocytes under these conditions, were able to undergo normal fertilization in vitro. After 6 hr of sperm-egg interaction in vitro, 87.5 of the oocytes had extruded the second polar body and/or developed pronuclei

Involvement of blood serum in amphibian fertilization

A proteolytic enzyme secreted by the first portion of amphibian oviduct, pars recta, called oviductin in Xenopus laevis, causes ultrastructural alterations on the extracellular matrix of coelomic eggs, turning them susceptible to fertilization. Although great advances have been made in the field of reproduction, the molecular mechanisms responsible for the fusion between the egg and the sperm are yet to be understood. We have recently demonstrated the presence of proteins from pars recta fluid in blood serum and extracellular matrix of coelomic eggs in Bufo arenarum. Here we show, using immunofluorescence procedures, that blood serum components are present in the extracellular matrix of coelomic and pars recta fluid-conditioned eggs in Bufo arenarum. Furthermore, by assessing the neutralizing effect on the conditioning activity of pars recta fluid on coelomic eggs we found that antibodies against pars recta secretions and blood serum inhibited the effect of sperm-lysin on the vitelline envelope of conditioned oocytes and impaired fertilization by sperm. Thus, serum proteins appear to be implicated in the molecular events that lead to amphibian fertilization

Vitelline envelope formation during oogenesis in Bufo arenarum

The formation of vitelline envelope (VE) during the oogenesis of Bufo arenarum (Amphibia Anura) is described. At the stage of early vitellogenesis, the first structures appear: the number of oocyte microvilli increases, and many cross sections of them are observed between the follicle cells and the oocyte. A filamentous material is observed inside the follicle cells and between the follicle cells and the oocyte. Multivesicular bodies are also found in the follicle cells, and in the perivitelline space. The micrographs also suggest the participation of the oocyte in the process of VE formation: large vesicles are present in the cortex of the oocyte, filled with an amorphous material of low and uniform electron density. Some of them are in the process of releasing their content to the perivitelline space. Many vesicles (probably resulting from microvilli fragmentation) are also observed in the perivitelline space. During late vitellogenesis the VE is a continuous structure between the layer of follicle cells and the oocyte. The filamentous material is aggregated in bundles, forming a net, and the spherical components are now either included in the orifices of the net, or free near the oocyte's surface. At the end of oogenesis, when the VE is completely formed, it is difficult to distinguish its components independently. Immunolocalization with antibodies against VE, show a positive reaction in follicle cells and oocytes in previtellogenic and full grown ovarian follicles. This analysis suggests that both the oocyte and the follicle cells are directly involved in the synthesis and secretion of the components of the vitelline envelope in Bufo arenarum.

Desenvolvimento embrionário do Curimbatá Prochilodus lineatus (Valenciennes, 1836) (Cypriniformes, Prochilodontidae) / Embryonal development of the curimbatá Prochilodus lineatus (Valenciennes, 1836) (Cypriniformes, Prochilodontidae)

Através da análise da morfologia externa dos ovos e dos aspectos microscópicos de seus cortes histológicos, foram caracterizados 18 estádios durante o desenvolvimento embrionário do curimbatá Prochilodus lineatus, que ocorreu a uma temperatura média de 26 + 0,5 C e no período de 16 horas, desde a fertilizaçao até a eclosao das larvas.

Study on helminth parasites of man in Sohag governorate, Upper Egypt II- Biological studies on encountered parasites

Detailed morphological studies on the eggs of parasitic infection in Sohag Governorate were done. For this purpose, collected and intra-uterine eggs of helminth parasites were studied, different sizes and shapes of eggs were noticed. Examination of intrauterine eggs of worms which were obtained form treated patients and kept in formol saline revealed different types of eggs in the same worm. Knowledge of such differences is of very important aid in identification of eggs and diagnosis of parasitic infections

Schistosoma japonicum: the modulation of lung granuloma and inhibition of egg maturation in mice by human sera.

Human sera taken from patients with chronic schistosomiasis japonica have been demonstrated to have two effects on mice. Sera from those patients reduced the size of granuloma in mice sensitised for accelerated granuloma formation to eggs entrapped in the lungs of mice injected with the sera shortly before and at day 2 after intravenous egg challenge. The sera with this effect on the mouse lung granuloma models caused large segmented precipitates in the optimised circumoval precipitin test (COPT). Such sera also reduced the rate at which eggs matured in the liver and intestines of mice infected with S. japonicum. The results strongly support our postulate that a major cause of granuloma modulation in cases of chronic schistosomiasis japonica is antiembryonation immunity and that mice provide useful models for the analysis of our postulate. Identification of egg antigens responsible for the anti-embryonation effect should facilitate progress towards the development of a vaccine against granulomatous disease.

Haplorchis pumilio (Looss) infection in man in northeastern Thailand.

During a clinical trial of praziquantel for human opisthorchiasis, Haplorchis pumilio Looss were recovered from the stools of 12 patients. This is the third species of Haplorchis spp. reported from man in Thailand.

The Effect of Lecucocytes on the Maturation of Mouse Oocytes in vitro

This experiment was undertaken in order to know the effect of leucocytes on the maturation of mouse oocytes in vitro. Leucocytes obtained from heart puncture of mouse (3 X 10(4) cells/mm3) inhibited the maturation of mouse oocytes. The egg toxic activity declined with decreasing leucocyte concentration. It was found that egg toxic effect of leucocytes is not species specific. The activity of intact leucocytes or equal numbers of leucocytes that were destroyed was similar and which seems not to be influenced by the physiological stats of leucocytes.

Volume Changes During the Preimplantation Stages of Mouse Egg Development

The mouse eggs in the various stages, of the development prior to implantation were collected and measurements were made on both the largest and smallest diameters of the vitellus, inner and outer surface of the zona pellucida. The various stages of development used were ovarian oocytes (germinal vesiA®e stage), ovulated but unfertilized egg, ovulated and fertilized egg, the 2-cell embryo on the second day of pregnancy, 4-8-cell embryo on the third day of pregnancy and morulablastocyst on the fourth day of pregnancy, A further comparative study on unfertilized and fertilized tubal eggs was made, The time of l2 hours after H.C.G. injection was chosen as the starting point from which to follow the collection of eggs every 3 hours for 24 hours. Since the volume gives a better comparison of size than diameter, the volume of the total eggs, intrazonal cavity, perivitelline space and the various were calculated for the various preimplantation stages of mouse egg. The volume of zona pellucida was also calsulated by subtraction of the volume of the inner zonal cavity from the volume of total egg and compared with the zona pellucida thickness. All calculations were made by computor(CEIR Time-sharing Computor). The diameter and volume of the vitellus in the ovarian oocyte is the largest one of any stage during the preimplantation stages of development, while the total volume of the entire egg as determined from the diameter of outer surface of the zona pellucida of the ovarian cocyte is the smallest one of any stage during development. The diameter and total volume of the entire egg increases from the ovarian oocytes to the first day of pregnancy and then gradually decreases until the third day of pregnancy. An increase in these parameters again takes place on the fourth day of pregnancy. The zona pellucida of the tubal ova is thicker than that of the oocyte, with the zona pellucida of the fertilized egg being definitely thinner when compared with unfertilized eggs. This phenomenon of decreased thickness in fertilized egg may be associated with zona reaction. The perivitelline space between the vitellus and zona pellucida thus formed following ovulation occupied approximately 40 percent of the total volume enclosed by the inner surface of the zona pellucida (intrazonal cavity) in the 1-cell tubal ova. Neither the cause of the rapid accumulation of fluid after ovulation which resulted in the production of the perivitelline space nor the actual time of the formation of the perivitelline space are known. Some possible reasons for the formation or origin of the perivitelline space are discussed. The size and shape of the vitellus undergo compartive reduction during preimplantation stages of development. The possible reason for the reduction of vitelline volume are discussed.