DEVELOPMENTAL BIOLOGY OF THE SCHISTOSOMA MANSONI IN THE INTERMEDIATE HOST SNAIL

Abstract

Snails from Biomplzalaria glahrata stock, unselected ordinary (Albino) susceptible, selected pigmented resistant and giant intermediate hosts of Schistosoma mansoni were studied for the developmental biology and histopathology. Histological serial sections of 128 snails of the three groups mass exposed to miracidia (MI) of S. mansoni were carried out in order to analyze the determination, location, migration and development of the various stages of the parasite and host responses. In chapter one comparative histology of above snails (50 susceptible, 55 resistant and 3 giant) of B. glahrata have been studied for development of the mother sporocyst eMS) after 1-10, 30 and 40 DPE. After penetration by S. mansoni MI detailed histological examination of various organs of the susceptible snails showed normal development of MS from 1-10 DPE. MS were visible in the foot, head, lips, tentacles, mantle, anus and buccal mass of the susceptible snails after 1-4 DPE. The beginning of the migration of the MS was observed at 3 and 4 DPE in the mantle, muscles of the anus and buccal mass of the susceptible snails. At 5 DPE some broad, few irregular, thin, elongated MS were observed in the above mentioned organs, neck and reached in the kidney of the susceptible snails. The next step of the migration of the MS was observed in the kidney at 5 DPE. From 6 DPE and onwards within the body of the mature MS embryos of the daughter sporocysts (DS) were found in the foot, head, lip, tentacles, mantle, kidney and esophagus of the susceptible snails. After 8 DPE in the foot, head, and lip of the susceptible snails within the body of the mature MS developed few embryos of the DS ruptured tegument and migrating towards deeper organs. After 9 and 10 DPE MS are visible in the pericardial cavity of the heart and respiratory epithelium of the lung of the susceptible snails. In resistant snails MS were located in the foot, head, lip, tentacles, mantle, anus, buccal mass, neck and columellar muscles from 1-10 DPE. Single MS were observed in the foot, head, lip, tentacles, mantle, neck, anus and buccal mass of these snails throughout infection period. While mUltiple MS were only visible in the tentacle and columellar muscles of the resistant snails after 9 DPE. No any further migration or mature MS was observed in these snails. During infections four types of the MS (single, multiple, mature and migratory) were recognized in the susceptible snails. Among all infected organs the highest total number of the 24 single and 189 multiple MS found in the head and foot of the susceptible snails respectively. In the resistant snails few number of single and multiple MS were found. Chi-square test results shows highly significant difference in between the overall MS in various infected organs of the susceptible and resistant snails O:::X2 (1) 21.58, P < 0.001). The body of the MS was characterized by thin tegument with cells, some germinal cells contained scanty cytoplasm, nuclei with prominent nucleolus, muscles, vacant spaces, apical gland and penetration glands at 1-2 DPE. At 2 DPE single MS further grow and become multiple at 3 DPE and onwards in the susceptible snails. After 5 DPE great proliferation of the germinal cells increased their number and size so that they occupy practically entire body of broad MS. The germinal cells as they increase in size have increased amount of cytoplasm and their nuclei are with prorriinent nucleoli. The germinal cells spread or distribute to form few clusters and masses of large numbers of these cells inside the body of the mature MS from 6 DPE and onwards. In the susceptible snails few embryos of the DS ruptured tegument and migrated towards deeper organs after 8 DPE. In the resistant snails development of the MS is characterized by tegument, vacant spaces, prominent penetration glands, few germinal cells had nuclei with nucleoli and little cytoplasm. Very few MS exhibits thin tegument, dark nuclei of germinal celis, increased vacant spaces, spread penetration gland and surrounded by herilocytes reaction. The highest mean diameter of the single MS found in the head susceptible snails ranges between 37.65 ± 5.55 )..tffi and 99.75 ± 9.38 )..tm from 1-10 DPE. In the foot of the resistant snails mean diameter (± SEM) of the single MS ranges between 48.08 ± 2.15 and 92.91 ± 1~.53 )..tffi during the infection. The giant snails had white patches on the regions of the ventral surface of the foot and selected for gigantism. In the posterior surface of the foot multiple MS were surrounded by strong hemocytes and mature MS containing embryos of DS after 30 DPE. In later infection at 40 DPE MS were visible in the most of the above organs, mantle rim, mantle cavity, radula, kidney, respiratory epithelium of the lung, shell epithelium, salivary glands, hemocytes producing organ and albumin glands of the resistant snails. MS was single in the foot, head, tentacles, buccal mass, neck, radula, and hemocytes producing organ. While increasing numbers of the multiple and mature MS were visible in the anus, mantle, kidney, respiratory epithelium of the lungs and albumin glands of the animals. In chapter two histopathological studies of the susceptible, resistant and giant snails have been done during the development of the MS. The histological examination of the various organs shows infiltration of hemocytes, encapsulations and cysts are developed in some of the susceptible snails. Whereas, in addition to these host responses early tissue reaction was visible in the resistant snails from 1-10 DPE. These responses were found in the foot, head, lip, tentacles, mantie, gut, mantle cavity and kidney of the susceptible snails. However, in the resistant snails these responses were also noticed in the first six organs mentioned earlier in the susceptible snails, pharynx, mantle rim, anus and respiratory epithelium of the lung during the period of the infection. The number of the infected organs of the snails was counted. Chi-square test result shows highly significant difference between the infected organs of the susceptible and resistant snails (LX2(1) 33.47, P < 0.001). The histopathological studies of an infiltration of hemocytes exhibited accumulation of the several layers of the cells in the various organs of the snails. Similarly, an encapsulation of the MS was also surrounded by few to several layers of the hemocytes in the resistant snails. In case of the susceptible snails an encapSUlation exhibited complete destruction of the multiple MS. The cyst was also surrounded by thin layers of hemocytes and characterized by complete destruction of the MS, few remains of the nuclei of the germinal cells, and increase vacuolated necrotic spaces. After 30 DPE in the posterior surface of the foot of the giant snails few multiple MS were surrounded by strong hemocytes reaction. An infiltration of the hemocytes was observed in the foot and tentacle of these snails. An encapSUlation of the MS and granulomas were also visible in the foot of the giant snails. In later infection after 40 DPE an infiltration of the hemocytes was visible in the respiratory epithelium of lung, kidney, gut, and heart of resistant snails. Inflammation and thin outer layer of respiratory epithelium of lung of resistant snail was also observed. An encapsulation of the MS was visible in the mantle, mantle cavity, and respiratory epithelium of lung of these snails. A cyst formation was also seen in the foot, head, tentacle, mantle, mantle cavity, mantle collar, buccal mass columellar muscles, lung, kidney, and gut of the resistant snails. In chapter three development of the DS and cercariae (C) plus host responses in the various organs of the susceptible, resistant, and giant snails have been studied. At 9 and 10 DPE few DS were observed in the posterior viscera which include gut, digestive gland, and ovotestis of the susceptible snails. The DS increased in number, multiplied and migrated in the deeper tissues of above organs at 15 DPE. In the digestive glands greater number of the DS was observed on 30 and 40 DPE respectively. Whereas, in resistant snails sections were made only on 9, 10 and 40 DPE. No DS was observed on 9 and 10 DPE, however, in later infection at 40 DPE the increased number of the DS visible in the gut and digestive glands of the resistant snail. Chi-square test results shows highly significant difference bet\\reen the infected susceptible and resistant snails and DS found in the gut, digestive glands and ovotestis of the snails (LX2 (3) 511.9, P < 0.001). The convoluted, longitudinal and broad shaped DS exhibits thick red tegument with spines at 9 DPE. Initially germinal cells were smaller then they increased in size, contained round or oval nucleus with prominent dark nucleoli in the center. At 20 DPE mature DS contained several germballs of the cercaria, tegument with prominent nuclei of cells, increase vacant spaces and in few early developments of the cercaria was observed. The head region, pre and post-acetabular and penetration glands and short tail region of the cercariae were also visible within the body of the DS. This stage further developed in digestive gland and ovotestis, matured and free cercariae were visible in the susceptible snails at 30 and 40 DPE respectively. During these developments few areas of an infiltration of hemocytes was visible in these organs. In the digestive glands thin walls and overgrowth was also prominent. Atrophy of the acini of the ovotestis was observed in these snails. At 30 DPE in the anterior surface of the foot of giant snails few mature DS containing few germballs and some encapsulations of the cercariae were visible. Enormous numbers of the DS and normal cercariae were observed in the digestive glands and ovotestis of these snails. Whereas, in the · gut of the resistant snails at 40 DPE some mature DS contained several germballs of the cercaria. The range mean diameter of the DS found in the gut, digestive gland and ovotestis of the susceptible snails was 33.01 ± 4.41 )..Lm to 238.79 ± 19.08 )..Lm. In the chapter four histopathology of ovotestis of the unselected susceptible snails of B. glahrata the intermediate host of S. manson; were studied after 9, 10, 15, 20, 30 and 40 DPE. The ovotestis was composed of transverse, longitudinal and irregular shapes and sizes of acini in the sectioning material and collecting canal. In S. manson; infection the highest mean diameter of the acini was 312 ± 28.76 )..Lm at 20 DPE. Ai 30 and 40 DPE infection of the mature DS and cercariae were progressing in the ovotestis of the snails. During these days the mean diameter of the acini was reduced. After detailed histological examination four types of female germinal cells were recognized. They are early oogonia, previtellogenic, stage I and II vitellogenic oocytes. Few atretic oogonia were also found during the infections. Early oogonia are very thin elongated, few in number, mostly located in the sidewall of the acinus, surrounded by dark thin outer membrane and cytoplasm at 9 DPE. They possess dark scanty cytoplasm. Atretic oogonia are surrounded by prominent outer membrane, filled with strongly dark cytoplasm with few vacuol~s. Normal previtellogenic oocytes are spindle, elongated and oval shaped with darkly stained homogenous cytoplasm and spherical nucleus with one or two nucleolus. The stage I oocytes are mostly quadrilateral and triangular in shape. They are located close or below the stage II vitello genic oocyte at the dorsal regions and sidewalls of the acini. The stage I vitellogenic oocyte is surrounded by few nurse cells with thin nuclei. These oocytes show prominent plasma membrane, dark vacuolated cytoplasm with yolk deposition and round nucleus. The early oogonia and previtellogenic oocytes have thin cell membrane, dark homogenous cytoplasm and nucleus with prominent nucleolus. The stage I and II