José Roberto Machado-Silva

A new description of the reproductive system of Schistosoma mansoni (Trematoda: Schistosomatidae) analyzed by confocal laser scanning microscopy

Classical schemes of the adult Schistosoma mansoni reproductive system have been described. In our study, whole adult worms derived from unisexual or mixed infections and stained with carmine chlorine were virtually and tomographically analyzed under confocal laser scanning microscopy. We found that: (1) there were morphological differences in the ovary, vitteline glands and testicular lobes between specimens derived from unisexual or mixed infections; (2) there was always a single lobed ovary (three or four lobes), presenting differentiation from the anterior to the posterior lobes, where the most mature oocytes were located; (3) the proximal segment of oviduct was connected to an ampullary dilatation, full of tailed spermatozoa, characterizing a seminal receptacle; (4) there was no long vitelline duct, but a short one that begins at the end of the proximal region of the vitelline gland; (5) long cells of Mehlis’ gland placed radially around the ootype were not observed. Otherwise, the ootype was only lined by thick cuboidal epithelial cells with plaited bases and nuclei with flabby chromatin, making a clear distinction from the uterine epithelium. This morphological feature suggests that each cell represents a gland. (6) In coupled males, the specimens located inside the gynaecophoric canal had smaller testicular lobes, suckers, and body length and width when compared to their partners. Our results show that the reproductive system does not follow a unique pattern within flatworms. Due to its better resolution, confocal laser scanning microscopy, using a reflected mode with tomographic sections, allows new interpretations, modifying the adopted and current descriptions of the internal morphological structures of S. mansoni adult worms.

Morphological study of adult male worms of Schistosoma mansoni Sambon, 1907 by confocal laser scanning microscopy

Aiming to detail data obtained through brightfield microscopy (BM) on reproductive, excretory and digestive system, specimens of Schistosoma mansoni eight weeks old, were recovered from SW mice, stained with Langerons carmine and analyzed under a confocal laser scanning microscope CLSM 410 (Carl Zeiss). The reproductive system presented a single and lobate testis, with intercommunications between the lobes without efferent duct. Supernumerary testicular lobe was amorphous and isolated from the normal ones. Collecting tubules (excretory ducts), followed by the excretory bladder, opening to the external media through the excretory pore, were observed at the posterior extremity of the body. In the digestive tract, a cecal swelling was noted at the junction that originates the single cecum. It was concluded that through confocal laser scanning microscopy, new interpretations of morphological structures of S. mansoni worms could be achieved, modifying adopted and current descriptions. The gonad consists of a single lobed testis, similar to that observed in some trematode species. Moreover, the same specimens can be observed either by BM or CLSM, considering that the latter causes only focal and limited damage in tissue structures.

Morphological changes in the reproductive organs of male and female Schistosoma mansoni worms caused by streptozotocin, a drug used to induce diabetes mellitus

Host metabolic changes have been observed to affect Schistosoma mansoni egg production, worm survival and morphology. We examined worms recovered from streptozotocin-induced diabetic mice by morphometric and morphological analysis through brightfield and confocal laser scanning microscopy. Tegument thickness was slightly smaller and changes in the reproductive organs were observed in 23-30% and consecutively 88-100% of the worms. The testicular lobes had a large diminution of cells in one or more of the lobes, which was associated with a lack of spermatozoids in the seminal vesicle. Ovaries were atrophied, manifested by a complete or large reduction in oocytes but other parts of the reproductive system like the vitelline glands were mainly unaffected. Streptozotocin (STZ) instead of hyperglycaemia caused the degeneration since worms from mice injected with a non-diabetogenic dose, or with nicotinamide to prevent diabetes showed the same alterations. The drug did not affect worm survival or pairing. We conclude that STZ, an alkylating agent that provokes chromosome and DNA damage, changes the morphology of ovaries and testicular lobes in S. mansoni worms in vivo. This is the first report of STZ action in helminths and we suggest that STZ affects oogenesis and spermatogenesis and might cause sterilization of schistosomes.

Host-induced morphological changes of Schistosoma mansoni Sambon, 1907 male worms

In order to evaluate the permissiveness of Nectomys squamipes to Schistosoma mansoni and the influence of the albino mice on the morphological aspects of adult worms derived from a population isolated from N. squamipes, the morphology of adult S. mansoni Sambon, 1907 male worms was studied using a digital image analyser (MOP VIDEOPLAN) and light microscopy. Their sources were as follows: (1) recovered from the wild rodent N. squamipes Brants naturally infected from Sumidouro, RJ, Brazil; (2) recovered from albino mice experimentally infected with the strain derived from N. squamipes; (3) recovered after the isolation of a strain derived from aboriginal human infections in Sumidouro. Worms recovered from N. squamipes (group 1) showed body length and distance between suckers significantly bigger than those of the specimens maintained in mice (groups 2 and 3). The number of testes in group 1 was statistically less than that of groups 2 and 3. Group 2 strains which were maintained in mice, presented the length of the worms as the only significant different character. Data show that: (1) N. squamipes is a more suitable host for the development of S. mansoni when compared to the albino mice; (2) a strain of S. mansoni isolated from a natural host undergoes morphological changes after its passage in the white mouse.

Schistosoma mansoni Sambon, 1907: morphometric differences between adult worms from sympatric rodent and human isolates

A computer software for image analysis (IMAGE PRO PLUS, MEDIA CYBERNETICS) was utilized in male and females adult worms, aiming the morphological characterization of Schistosoma mansoni samples isolated from a slyvatic rodent, Nectomys squamipes, and humans in Sumidouro, Rio de Janeiro, Brazil and recovered from Mus musculus C3H/He. The following characters for maless testicular lobes were analyzed: number, area, density, larger and smaller diameter, longer and shorter axis and perimeter and extension; for females: area, longer and shorter axis, larger and smaller diameter and perimeter of the eggs and spine; oral and ventral suckers area and distance between them in both sex were determined. By the analysis of variance (one way ANOVA) significant differences (p < 0.05) were observed in all studied characters, except for the density of testicular lobes. Significant differences (p < 0.05) were detected for all characters in the female worms. Data ratify that sympatric isolates present phenotypic differences and the adult female characters are useful for the proper identification of S. mansoni isolates.

Enteroparasitoses em pré-escolares de comunidades favelizadas da cidade do Rio de Janeiro, Brasil

Prevalence of intestinal parasites was investigated in 1381 low-income children under five years of age from March 1990 to October 1991 in the city of Rio de Janeiro, Brazil. Stool tests were run using the Blagg et al. method. Observed prevalence of infection was 54.5%. The most frequent parasites were Giardia lamblia and Ascaris lumbricoides (25.0% of the children). A significant statistical difference (p<0.05) was demonstrated between age and infection according to the species of parasite studied. Infection rates demonstrated the need for control of parasites in these child.Prevalence of intestinal parasites was investigated in 1381 low-income children under five years of age from March 1990 to October 1991 in the city of Rio de Janeiro, Brazil. Stool tests were run using the Blagg et al method. Observed prevalence of infection was 54.5%. The most frequent parasites were Giardia lamblia and Ascaris lumbricoides (25.0% of the children). A significant statistical difference (p<0.05) was demonstrated between age and infection according to the species of parasite studied. Infection rates demonstrated the need for control of parasites in these children.A prevalencia das enteroparasitoses foi verificada atraves de inquerito coproparasitologico realizado de marco de 1990 a outubro de 1991, em 1.381 pre-escolares provenientes de quatro comunidades faveladas da cidade do Rio de Janeiro, Brasil. Por meio da tecnica de Blagg et al. (1955), foi encontrada uma positividade geral de 54,5. Os parasitos mais frequentes foram Giardia lamblia e Ascaris lumbricoides, em cerca de 25,0% das criancas investigadas. Correlacao estatisticamente significativa foi observada entre idade e positividade nas criancas menores de tres anos. Sugerimos que tratamento antiparasitario e modificacoes ambientais sejam implementados para o controle das infeccoes parasitarias nesta populacao.

Morphological aspects of Schistosoma mansoni adult worms isolated from nourished and undernourished mice: a comparative analysis by confocal laser scanning microscopy

Malnutrition hampers the course of schistosomiasis mansoni infection just as normal growth of adult worms. A comparative morphometric study on adult specimens (male and female) recovered from undernourished (fed with a low protein diet - regional basic diet) and nourished (rodent commercial laboratory food, NUVILAB) white mice was performed. Tomographic images and morphometric analysis of the oral and ventral suckers, reproductive system and tegument were obtained by means of confocal laser scanning microscopy. Undernourished male specimens presented smaller morphometric values (length and width) of the reproductive system (first, third and last testicular lobes) and thickness of the tegument than controls. Besides that, it was demonstrated that the dorsal surface of the male worms bears large tubercles unevenly distributed, but kept grouped and flat. At the subtegumental region, vacuolated areas were detected. It was concluded that the inadequate nutritional status of the vertebrate host has a negative influence mainly in the reproductive system and topographical somatic development of male adult Schistosoma mansoni, inducing some alterations on the structure of the parasite.

Phenotypic plasticity in adult worms of Schistosoma mansoni (Trematoda:Schistosomatidae) evidenced by brightfield and confocal laser scanning microscopies

A comparative morphometric study was performed to identify host-induced morphological alterations in Schistosoma mansoni adult worms. A wild parasite population was obtained from a naturally infected rodent (Nectomys squamipes) and then recovered from laboratory infected C3H/He mice. Furthermore, allopatric worm populations maintained for long-term under laboratory conditions in Swiss Webster mice were passed on to N. squamipes. Suckers and genital system (testicular lobes, uterine egg, and egg spine) were analyzed by a digital system for image analysis. Confocal laser scanning microscopy (CLSM) showed details of the genital system (testicular lobes, vitelline glands, and ovary) and the tegument just below the ventral sucker. Significant morphological changes (p < 0.05) were detected in male worms in all experimental conditions, with no significant variability as assessed by CLSM. Significant changes (p < 0.05) were evident in females from the wild population related to their ovaries and vitelline glands, whereas allopatric females presented differences only in this last character. We conclude that S. mansoni worms present the phenotypic plasticity induced by modifications in the parasites microenvironment, mainly during the first passage under laboratory conditions.

Hepatic stereology of schistosomiasis mansoni infected-mice fed a high-fat diet

High-fat diets induce weight gain and fatty liver in wild-type mice. Schistosomiasis mansoni infection also promotes hepatic injury. This study was designed to quantify hepatic alterations in schistosomiasis mansoni-infected mice fed a high fat-rich chow compared to mice fed a standard rodent chow, using stereology. Female SW mice fed each either high-fat diet (29% lipids) or standard chow (12% lipids) over 8 months, and then were infected with Schistosoma mansoni cercariae. Four experimental groups were studied: infected mice fed a high-fat diet (IHFC) or standard chow (ISC), uninfected mice fed a high-fat diet (HFC) or standard chow (SC). Mice were sacrificed during early infection (9 weeks from exposure). The following hepatic biometry and the stereology parameters were determined: volume density (hepatocytes [h], sinusoids [s], steatosis [st] and hepatic fibrosis [hf]); numerical density (hepatocyte nuclei - Nv[h]); absolute number of total hepatocyte N[h], normal hepatocyte N[nh], and binucleated hepatocyte N[bh], percentage of normal hepatocyte P[nh] and binucleated hepatocyte P[bh]. IHFC and HFC groups exhibited TC, HDL-C, LDL-C, and body mass significantly greater (p < 0.05) than control group. No significant differences were found regards liver volume (p = 0.07). Significant differences were observed regards P[nh] (p = 0.0045), P[bh] (p = 0.0045), Nv[h] (p = 0.0006), N[h] (p = 0.0125), N[bh] (p = 0.0164) and N[nh] (p = 0.0078). IHFC mice group presented 29% of binucleated hepatocytes compared to HFC group (19%), ISC group (17%) and SC (6%). Volume density was significantly different between groups: Vv[h] (p = 0.0052), Vv[s] (p = 0.0025), Vv[st] (p = 0.0004), and Vv[hf] (p = 0.0007). In conclusion, schistosomiasis mansoni infection with concurrent high-fat diet promotes intensive quantitative changes in hepatic structure, contributing to an increasing on hepatic regeneration.

Schistosoma mansoni Sambon, 1907: comparative morphologica studies of some Brazilian strains

The morphology of Schistosoma mansoni adult male worms from three strains which have been maintained in albino mice for several generations, was compared to a strain that has been isolated from the natural host Nectomys squamipes (Rodentia:Muridae) captured in Sumidouro (Rio de Janeiro State) and have been maintained in the same sylvatic rodent under laboratory conditions. Total length of specimens, distance between suckers, the number of testes and extention of testes grouping were the taxonomic characters analysed. The worms recovered from N. squamipes showed expressive differences (p < 0.01) compared to the other strains regarding the considered morphological characters. The strains that were maintained in mice presented statistical differences (p < 0.01) in several characters. Some adult worms besides the normal position of the testes also showed an atypical arrangement of these glands. It can be concluded that the morphology of adult worms may be used to distinguish S. mansoni strains and that morphological changes in adult worms are not induced by successive inoculations of a strain in mice.