Transactions of the American Microscopical Society

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The uptake and storage of two chlorinated hydrocarbon insecticides, aldrin and dieldrin, in freshwater ostracods (Chlamydotheca arcuata (Sars)) was studied using autoradiographic technique. Although aldrin and dieldrin uptake occurred over a wide body surface area, absorption was primarily through the body integument and the gut; lesser amounts of either insecticide were taken up through appendages. The shell absorbed or adsorbed only small quantities. The storage of these insecticides was largely in body lipids.
 
Specimens of Porrocaecum sulcatum (Rudolphi, 1819) from the sea turtle Chelone mydas collected off the coast of North Carolina are redescribed. General morphological features that are observed by light microscopy and scanning electron microscopy confirm the retention of this species in the genus Porrocaecum. The genus Sulcascaris is considered synonymous with Porrocaecum.
 
Electron microscopic techniques were employed to investigate the fine structure and function of the intestinal system of the oxyurid Leidynema appendiculata. The intestine is bounded at its lumenal aspect by a microvillous border and at its pseudocoelomic aspect by a filamentous basal lamina. The ultrastructural architecture of the caecum suggests that it is a specialized region concerned primarily with secretory activity. The cells of this structure contain a pervasive system of endoplasmic reticulum, multiple Golgi complexes, enlarged nuclei, pleomorphic mitochondria, lysosome-like granules, lipid droplets, and alpha glycogen deposits. The anterior intestine possesses both secretory and storage capabilities. Two patterns of cellular discharge are apparent in this region. Progressing from the caecum and anterior intestine to the middle and posterior intestinal regions, there exists a gradual alteration in fine structure which indicates a concomitant gradation in function. The latter areas appear to possess previously undescribed structures which appear to be accessory elements of support. Nuclear inclusions, which also have not been reported in nematodes, exist within the posterior intestinal cells. The results of the present study suggest a possible mechanism of food utilization.
 
With the importation of living materials from Taiwan, Republic of China, the life cycle of Fasciolopsis buski (Lankester, 1857), the giant intestinal fluke of Asia, is currently being maintained in the United States. The snail host, Segmentina hemisphaerula (Benson, 1842) may be cultured with moderate ease and pigs may be employed as definitive hosts. Preliminary susceptibility studies have indicated that most commonly used laboratory animals cannot be infected but the parasite may establish itself temporarily in rabbits or squirrel monkeys (Saimiri sciureus petrinus Thomas, 1927).
 
Although the crab, Eriocheir japonicus, is the principal host for Paragonimus westermani on Taiwan, the parasite was found in four species. Random samplings of E. japonicus from different parts of Taiwan indicated low overall metacercarial infections with the majority of positive sites being in the northern part of the island. Monthly collections in a highly endemic area, however, showed rates of infection to range from 30 to 83% with a tendency toward higher infections in the fall and winter months, the time at which these crabs are consumed by man. In several collections, 100% of crabs were infected. Metacercariae per host ranged from 4.2 parasites in summer months to 8.6 in the winter. Cysts of P. westermani apparently accumulate in the crab until time of death, with as many as 1,043 cysts recorded from one host.
 
Lecithaster helodes n. sp. is described from Mugil curema and M. cephalus. It differs from the closely related L. mugilis Yamaguti, 1970 by having four rather than three ovarian lobes and other minor differences. Discussions and measurements are presented for L. confusus Odhner, 1905, L. leiostomi Overstreet, 1970, Hysterolecitha elongata Manter, 1931, Aponurus pyriformis (Linton, 1910) n. comb., and A. elongatus Siddiqi & Cable, 1960. Branchadena Linton, 1910 is considered a synonym of Aponurus Looss, 1907. New host records are listed for some of the above hemiurid trematodes. Leurodera ocyri Travassos, Frietas & Bührnheim, 1965 and L. inaequalis Travassos, Freitas & Bührnheim, 1965 are considered synonyms of A. pyriformis.
 
The genus Rugipes Schaeffer, 1926 was established to accommodate two new species of amoebae, R. bilzi and R. vivax, the first a fresh-water species, the second marine. The type species, R. bilzi, may well fall as a member of an older genus; at any rate, the second species is no longer considered congeneric with it. Clydonella n. g. is proposed to provide a new generic home for certain marine amoebae with characters similar to, yet distinctly different from, Vannella Bovee, 1965 and Platyamoeba Page, 1969. At the same time, the new genus is considered appropriate for the former R. vivax, as its type species, although this organism had been removed to Vannella by Bovee and to Platyamoeba by Page. Comparative data on the three genera and specific characters for C. vivax (Schaeffer, 1926) n. comb. are presented briefly.
 
Cosmocercoides dukae (Holl, 1928) Travassos, 1931 was recovered from 122 (7.7%) of 1,578 terrestrial molluses in Nebraska. Stenotrema leai (Binney), Succinea ovalis Say, Triodopsis albolabris (Say), T. multilineata (Say), and Cionella lubrica (Müller) are reported as new hosts for C. dukae.
 
The integument of critical-point dried adult S. mattheei was observed by scanning electron microscopy at 38-6,240 magnifications. There are marked differences in surface features of male and female schistosomes as well as different regions on the same parasite. SEM reveals surface structures which suggest basic adaptations for clasping of male and female schistosomes as well as for parasite attachment or anchorage in blood vessels of definitive hosts. Surface features are similar to those of S. haematobium, a morphologically related species.
 
The transformation of the heterotrich ciliate, Blepharisma americanum (Suzuki, 1954), from a bactivore to a carnivore, or cannibal giant is described. This transformation process is induced by exogenous d-alpha-tocopheryl acid succinate present at 10-6 to 10-4 M. In the presence of such concentrations, B. americanum undergoes a sequential alteration of morphology, especially with respect to the oral ciliature and buccal cavity. The length and volume of the organism also are increased greatly. Over a 6-10-h period following the addition of the vitamin, Blepharisma americanum develops an enlarged undulating membrane and adoral zone of membranelles due to the elongation of the component cilia. At 10-4 M tocopheryl, some monsterism is observed in the form of L-shaped or greatly elongated organisms after 3.5 h. At lower concentrations, transformation takes 8-15 h, and fewer individuals attain giant size. When B. americanum is removed from the vitamin solution, washed, and placed in sterile Chalkley's solution at pH 6.8, the ciliates revert to a normal bactivorous form by means of numerous cell divisions over a 3-day period. The results obtained in this study are essentially the same as the results of similar research on forms found in declining cultures of B. americanum, and bear considerable resemblance to the stomatin-induced transformations of Tetrahymena vorax. However, in the present work, a defined substance was used to induce transformation.
 
Raillietina (R.) loeweni had a minimal prepatent period of 14 days, and a patent period of 11 to 27 days in the hare and seven to 259 days in the laboratory rabbit. Numbers of cysticercoids given to experimental rabbits and the numbers of tapeworms recovered were directly related. Discharge of proglottids was greatest in afternoons and peaked three to eight days following patency. Proglottids discharged per day decreased as the tapeworm burden increased in Dutch Rabbits, which precludes using proglottid discharge rate to determine cestode burden in this host. Based upon limited data, resistance to reinfection appeared not to be marked following elimination of R. loeweni from an initial infection. Sodium chloride concentration, temperature, and interaction of salt and temperature conditioned evagination of cysticercoids in vitro.
 
In experimentally infected male albino rats, aberrant pre-adults of Trichinella spiralis are reported for the first time outside of the gastrointestinal tract.
 
Knobs projecting from membranes were never demonstrable by electron microscopy of negatively stained mitochondrial fractions isolated from a respiratory-deficient strain (cytoplasmic petite) of baker's yeast grown into stationary phase. Knobs projecting from some membranes were always demonstrable by electron microscopy of negatively stained mitochondrial fractions isolated from a respiratory-competent strain (grande) of baker's yeast grown into stationary phase, provided grande mitochondrial fractions were aged or osmotically shocked. Electron microscopy of sectioned petite cells showed there were fewer cristae per mitochondrion, fewer mitochondria per cell, and decreased total cristal area per cell.
 
The properties of the tegumental hexose transport systems of the intestinal helminths Hymenolepis diminuta (Cestoda: Cyclophyllidea) and Moniliformis dubius (Acanthocephala) are described and compared to those of the glucose transport systems of other tapeworm species and of the mucosal brush border of the vertebrate intestine. Glucose transport and accumulation in H. diminuta and in other cestodes are Na+-dependent and K+- and phlorizin-sensitive, and the tegumental glucose transport loci have substrate specificities very similar to those of the vertebrate intestinal mucosa. In contrast, glucose transport across the tegumental surface of M. dubius is neither Na+-dependent nor phlorizin-sensitive, and the substrate specificities of the facilitated diffusion systems for hexose transport in the tegument of this acanthocephalan are markedly different from those observed for the concentrative transport systems of the H. diminuta tegument and of the host intestinal mucosa. The accumulation of free glucose pools within the body wall of M. dubius appears to involve the "shuttling" of absorbed glucose through the nonreducing disaccharide trehalose. It is suggested that the differences in mechanisms of glucose transport and accumulation between M. dubius, on the one hand, and the tapeworm tegument and the vertebrate intestinal mucosa, on the other, derive from differences in the cytoarchitecture of these absorptive surfaces which share a common biochemical environment. It is further suggested that Na+-coupled carbohydrate transport against a concentration difference may be a unique feature of tissues whose absorptive surfaces comprise a brush border membrane.
 
The concept of the brush border as a digestive-absorptive surface is reviewed and brought up to date in terms of brush border membrane enzymes, brush border membrane transport systems, and those brush border membrane diseases which result from deletion of enzymic or transport activities. The molecular basis of brush border membrane disease, in the case of sucrase-isomaltase deletion, is indicated to be an absence of enzyme protein from the membrane rather than its presence with altered properties.
 
Detailed descriptions of the karyotypes of 47 species in the tribes Sciomyzini and Tetanocerini demonstrate that different genera of Sciomyzidae are evolving karyologically at different rates. All have 2n = 12 chromosomes, except two species of Pteromicra (2n = 10) and Sepedon spinipes americana (2n = 11 in $\male$ and 12 in $\female$). Various sex-chromosome systems, including XO/XX, XY/XX, and X1X2Y/X1X1X2X2, have been found, and the sex chromosomes are heterochromatic in about one-fourth of the species studied. The autosomes are less variable than the sex chromosomes. There is considerable karyological variation in the Sciomyzini, especially in Pherbellia. In the Tetanocerini, species of genera centered around Dictya have retained similar and apparently conservative karyotypes; those of Tetanocera are relatively variable, and those of Sepedon are even more variable.
 
Recently-hatched oncospheres in the invasive condition, obtained from washed centrifuged stools of infected rats, were prepared as fixed whole mounts on slides, treated with saliva or ribonuclease, then stained with acetic acid-orcein. The general morphology is described by means of a series of photomicrographs from dorsal to ventral levels through the body. The protoplasm of the penetration gland cells, following the dorsal body contour beneath the subcuticular musculature, had an extensive meshwork. The nucleus of each gland cell contained a small nucleolus and moderately evident chromosomes. The region behind the hooks contained the majority of cells, particularly five large embryonic pairs separable into two distinct classes. Class I cells included three pairs with very distinct, thickened chromosomes, arranged as a spireme, an irregular nucleolus, and dense, compact cytoplasm. The class II embryonic cells had the largest nuclei, indistinct chromosomes, but massive nucleoli rich in nucleoprotein.
 
Acini (alveoli) from mammary tissue of lactating rats were isolated using collagenase and examined in the transmission electron microscope to determine if changes occur in cellular structure and cellular relationships as a result of the isolation procedure. The secretory cells within isolated acini maintain their secretory apparatus and polarity following the collagenase isolation procedure. Tight junctions (zonula occludens) were present between secretory cells. Myoepithelial cells and their contacts between one another, as well as their contacts with secretory cells, were present in isolated acini. Preparations of isolated acini were nearly free of basal laminae, adipocytes, ductal elements, and connective tissue components. Thus, basolateral membranes of secretory cells within isolated acini are in direct contact with the extracellular medium. Lanthanum stained the plasma membrane of secretory cells on either side of the tight junction, but was absent within the tight junction itself. The presence of lanthanum within the lumina of acini, coupled with the absence of cascin micelles within lumina, indicates that lumina are in direct contact with the extracellular medium, probably by way of the primary ductal opening. Below the tight junction, intermittent gaps were present between lateral membranes of adjoining secretory cells. The gaps may be the result of osmotic changes and mechanical forces generated during the isolation procedure.
 
Total direct counts of aquatic bacteria from natural samples were estimated by epifluorescence microscopy after acridine orange staining. Cells were concentrated by membrane filtration on black membrane filters and counted directly. Using this method, bacterial counts of pure cultures corresponded to those obtained by the Petroff-Hauser and Coulter counter methods. Epifluorescence counting is the method of choice for ecological studies of the natural distribution of bacteria in aquatic environments since it permits ready discrimination of bacteria from detritus and does not rely on the adequacy of culture methods to elicit growth of all viable organisms. Total counts obtained in this way were related to glucose and acetate uptake by resident microbial populations of lake water sample. Viable counts of the same samples were not related.
 
The complete life cycle of two odoriferous aquatic actinomycetes was observed in slide cultures. Branching of the vegetative mycelium yielded sporogenous hyphae. Frequent intimate contacts were observed between vegetative hyphae produced by the same or by different spores. No "initial cells" were detected. Contrary to previous reports, these aquatic actinomycetes appeared to be true streptomycetes and lacked an isogamous reproductive pattern. Vegetative mycelium developed pigments, sporogenous hyphae and odor under aerobic conditions, but not when grown under an atmosphere of inert gases or in the presence of 300 ppm malonate. Only small amounts of sporogenous hyphae could be produced by washed vegetative mycelium suspended in a balanced salt solution.
 
The fine structure and the cytochemical distribution of acid phosphatase and aryl sulfatase in the intermediate cell layer of cysticercoids of Hymenolepis diminuta are reported. The cells comprising this layer are of two types in the developing cysticercoid and of a single type in the mature cysticercoid. In the former stage, one cell type appears to be responsible for the elaboration of the peripheral layer with its radiating fibers (= "hairy processes"). The second type appears to be involved in the formation of the circular fibers and, in part, the fibers of the intermediate layer. The latter type cell displays acid phosphatase and aryl sulfatase activity. Much of the activity is cytoplasmic in distribution and possibly is responsible for the ultimate degeneration of the cell.
 
The normal pharate adult development of the abdominal epidermis of the silkmoth Hyalophora cecropia was analyzed at the ultrastructural level and was compared with changes in cell morphology resulting from the application of the insect growth hormones, β-ecdysone and juvenile hormone. The onset of adult development elicited by endogenous β-ecdysone is accompanied by a change in cell ultrastructure from pupal cells having few mitochondria, few Golgi, numerous free ribosomes to epidermis containing rough endoplasmic reticulum and numerous Golgi. The microvilli of the apical plasma membrane appear to secrete pupal endocuticle during early pharate adult development and adult endocuticle a little later. Giant autophagic vacuoles, appearing early, may be requisite for cellular remodeling that necessarily precedes differentiation into adult cell types. Glycogen is accumulated in the epidermal cells during pharate adult development, and masses of glycogen rosettes are still evident in the newly eclosed moth. Adult endocuticle continues to be secreted after eclosion. Within an hour after injection of β-ecdysone into pupae, blebbing of the apical plasma membrane occurs; this is followed by the premature accumulation of giant autophagic vacuoles and the precocious deposition of cuticle. Similar responses occurred after juvenile hormone injection, except that giant autophagic vacuole development was completely suppressed. The ecdysone-like effects of juvenile hormone treatment are probably a result of juvenile hormone stimulation of endogenous ecdysone secretion. In response to juvenile hormone, spaces appear between the epidermal cells in the basal region, suggesting that juvenile hormone may modulate the permeability of the epidermal monolayer.
 
Sambonia parapodum n. sp. is described from Varanus salvator as the fourth member of the genus after Megadrepanoides, Self & Kuntz, 1957, with its two species, is relegated to synonymy with Sambonia based on Fain's (1961) redescription of the latter. Armillifer agkistrodontis n. sp. is described from Agkistrodon acutus.
 
The secretory bodies observed in the tegument of Haematoloechus medioplexus were examined for nucleoside phosphatase activity and for the presence of macromolecular diglycols. Reaction products resulting from the hydrolysis of ADP and ATP were found in the Golgi cisternae of the perinuclear cytoplasm and in the homogeneous portion of the secretory bodies. The same organelles were reactive after the test for macromolecular diglycols by the periodic acid-chromic acid-silver methenamine technique. It is hypothesized, from these observations, that the secretory bodies originate from the Golgi complex in the perinuclear cytoplasm and pass into the distal cytoplasm (tegument). The presence of sensory bulbs in the distal cytoplasm (tegument) and junctional complexes associated with the perinuclear cytoplasm are noted for the first time in this organism.
 
The mixture of polyvinyl alcohol (PVA) and Bouin's solution was found to be a satisfactory fixative and adhesive for smears of parasitic protozoa. PVA keeps material firmly attached to slides but does not interfere with staining or washing of specimens, while Bouin's fixation enables one to stain for flagella using the Bodian silver technique. Smears prepared with this mixture may be stored dry prior to staining with little apparent damage to the protozoa.
 
Twenty-eight golden hamsters (Mesocricetus auratus Waterhouse) representing days 6-15 of pregnancy, the postpartum condition, as well as the non-pregnant state, were used to study, following intraperitoneal injection of tritiated thymidine, the pattern of DNA synthesis in the uterus, yolk sac and allantoic placentae, and adnexa, during the course of development and differentiation, by means of autoradiography. Tables and graphs of the mitotic indices obtained demonstrate the varying cell proliferation activity from day to day in six uterine and 11 placental (and adnexa) tissues, and the difference among these several tissues. Uterine tissues have an overall range of mitotic index from a low of 0.26% in the myometrium, to a high of 10.23% in the myometrial fibroblasts. Fetal membranes have an overall range from a low of 1.63% in the visceral yolk sac mesoderm, to a high of 22.23% in the Träger. The labyrinth undergoes a remarkable increase in volume, and irrespective of how much conversion there may be of trophospongium into labyrinth, evidence points to an important participation of interstitial growth within the labyrinth.
 
An ATP phosphohydrolase was localized in the tegumental invagination of adult Schistosoma mansoni using a modification of the Wachstein and Meisel lead-ATP protocol. This reaction is not affected by deletion of Mg+- or K+, but is sensitive to cysteine. ATP phosphohydrolase activity is also observed in the tegumental basal plasma membrane of females, and in the nuclear envelopes and plasma membranes of parenchymal cells. These latter reactions are ouabainsensitive, Mg++-dependent, inhibited by Ca++, but are not affected by deletion of K+ from the medium. From these data, it is hypothesized that the reaction in the tegumental invaginations is due to non-specific alkaline phosphohydrolase activity, while the reaction observed elsewhere is indicative of a Mg++-dependent transport ATPase. Limited use of the strontium-nitrophenol phosphate protocol indicates a dependency on Na+ in these latter sites.
 
Snails containing encysted and free metacercariae and adult worms of a new species of trematode were collected in from 2-3 m of water 20-30 m off Entebbe, Uganda in Lake Victoria during a study of snails for schistosomes. This report is the first to note the occurrence of adult trematodes in fresh-water pulmonates. Whether the complete life cycle occurs in the snail is unknown.
 
Extensive feeding experiments involving 96 hosts of 19 species were undertaken to investigate the host-specificity of Cotylurus flabelliformis among domestic and wild avian hosts. Four new experimental hosts, representing three new orders were found: Pekin duck (Anseriformes); pied-billed grebe (Podicipediformes); American coot (Gruiformes); and song sparrow (Passeriformes). Although development was delayed in the pied-billed grebe, no host-induced variations of enough significance to question previously described species of Cotylurus were noted. Temperature of the fixative (AFA) greatly influenced the size of the worms in permanent preparations. Experimental studies on the development of C. flabelliformis in the domestic mallard indicate that adults are capable of producing eggs within 48 hr following their ingestion as metacercariae. Greatest changes in the parasite body occurred in the development of the hindbody and reproductive system during the first 42 hr. Untanned eggs were present in the uterus after 42 hr and eggs were present in the feces in less than 51 hr.
 
The visceral musculature of Canton S strain of Drosophila melanogaster was studied at the ultrastructural level to investigate the fine structural alterations that occur in this organism with age. Although drastic age-related alterations have been reported in the fat body, brain, midgut epithelium, cardiac and flight muscles of various insects, only limited and minor changes were noted in the midgut musculature for the first 90 days of life. Additional ultrastructural alterations, such as disarray of myofilaments, were noted in Drosophila over 100 days of age.
 
Top-cited authors
Robert P. Higgins
  • Museum of Natural History, Smithsonian Institution
Hjalmar Thiel
  • University of Hamburg
Anil Kumar Mandal
  • University of Florida
Timothy K Maugel
  • University of Maryland, College Park
John J. Gilbert
  • Dartmouth College