ALBERT

Notes: . Actin, the major protein of amebae of Naegleria gruberi, proved to be strongly immunogenic in rabbits. The resulting precipitating antibodies are specific to actin of Naegleria. In a competitive solid-phase radioimmunoassay, these antibodies bound similarly to Naegleria G- and F-actin. Actins from amebae of Acanthamoeba and Dictyostelium, plasmodia of Physarum, sea urchin eggs, and vertebrate muscles gave no competition in the radioimmunoassay. Estimates of the amount of actin in Naegleria amebae ranged from a minimum of 5% of the total cell protein by radioimmunoassay to a maximum of 16% by electrophoresis. The unusual species specificity of these antibodies indicates that Naegleria actin, although conserved in many properties, is different enough to have unique antigenic determinants.

Notes: . Extracts of the pathogenic ameba Naegleria fowleri, prepared by freeze-thawing and sonication, were analyzed for their content of various hydrolytic enzymes that have acid pH optima. The organism is rich in acid phosphatase activity as well as a variety of glycosidases which include β-glucosidase, β-galactosidase, β-fucosidase, α-mannosidase, hexosaminidase, arylsulfatase A, and β-glucuronidase. The crude extract contained only negligible levels of sphingomyelinase, neuraminidase, or arylsulfatase B. All of the hydrolases exhibited higher activity at pH 5.5 than at 7.0, indicating that they are truly “acid” hydrolases. In general, after centrifugation (100,000 g, 1 h), except for arylsulfatase B, more than half of the activity of each of the various hydrolases was recovered in the supernatant fraction. The acid phosphatase in the high-speed supernatant was purified 45-fold (32% yield) by chromatography on QAE-Sephadex and Sephadex G-200 and shown to have the following properties: 1) pH optima, 5.5; 2) Km (4-methylumbelliferyl phosphate), 0.60 mM; 3) molecular weight (estimated by gel filtration chromatography), 92,000; 4) inhibited by heteropolymolybdate complexes but not by L(+) sodium tartrate (0.5 mM) or sodium fluoride (0.5 mM). In addition, unlike the tartrate-resistant acid phosphatase of Leishmania donovani, the major acid phosphatase of N. fowleri is less than 5% as effective in inhibiting superoxide anion production by f-Met-Leu-Phe-stimulated human neutrophils. The finding of high levels of a number of acid hydrolases in Naegleria fowleri raises several questions that merit further study: Do the hydrolases perform a housekeeping function in this single cell eukaryote or do they play some role in the pathogenic process that ensues when the organism infects a suitable host?

Notes: . We have shown that bacteria-fed Tetrahymena express at their surface and excrete into the medium a glycoconjugate absent in axenically grown cells. A preliminary analysis of the purified molecule is given. Immunolabeling of blotted surface extracts and fixed cells shows that bacteria-fed Tetrahymena build up a surface coat whose material originates totally or in part in the mucocysts. The glycoconjugate is located externally on the coat and mediates cell immobilization and immunolabeling by the serum. The results also indicate that axenic cells are probably devoid of surface coat.

Notes: . Axenic cultures of Entamoeba histolytica strains HK-9, HM-1, and Rahman were fractionated to provide plasma membranes, internal, vesiculated membranes, and a soluble cytosol. Each particulate fraction was solubilized and all fractions were analyzed by techniques designed to demonstrate molecular complexity and serologic reactivity. The cytosol contained more antigenic moieties than either membrane fraction; however, the antigens associated with the membranes had very high reactivity and lower nonspecific activity than the cytosol.

Notes: . Zygote development and oocyst wall formation of Eimeria truncata occurred in epithelial cells in renal tubules and ducts of experimentally infected lesser snow geese (Anser c. caerulescens). Post-fertilization stages were present throughout the kidneys beginning nine days post-inoculation. Initially, a single plasmalemma enclosed the zygote, and type 1 wall-forming bodies (WF1) became labyrinthine and moved toward the surface. There, WF1 degranulated and formed the outer layer of the oocyst wall between the plasmalemma and a newly formed second subpellicular membrane. Several WF2 fused and formed the inner layer, of the oocyst wall between the third and fourth subpellicular membranes. Six subpellicular membranes were observed during wall formation. Other features of oocyst development were similar to those of other eimerian species.

Notes: . Filaments in the oral apparatus of Tetrahymena appear similar, but not identical, to the intermediate filaments of multicellular organisms. The mean diameter of filaments measured in the present study was 16.4 nm, but the range of variation was much greater than has been reported for intermediate filaments. The organization of filaments within the oral apparatus has been studied by indirect immunofluorescence microscopy and immunogold localization at the electron microscopical level using antiserum raised in rabbits against the major subunit protein of the oral filaments (87K). The filaments were found to be organized into cables, networks, and chambers or cages which encase the basal bodies. At the highest level of organization, the filaments connect into a rigid framework capable of maintaining the overall architecture in the absence of microtubules. Like intermediate filaments, the oral filaments are insoluble at high ionic strength, have evolutionarily non-conservative subunit proteins, are probably non-contractile, and serve to stabilize persistent cellular architecture.

Notes: . Microgamonts and macrogamonts of Eimeria truncata were observed in renal epithelial cells of collecting tubules and ducts and occasionally in macrophages of experimentally infected lesser snow geese (Anser c. caerulescens) beginning 8.5 days post inoculation. Intraparasitophorous vesicles in parasitophorous vacuoles of both types of gamonts appeared to originate in host cell cytoplasm and enter gamonts through micropores by budding of plasmalemma or by pinocytosis. Within the parasite's cytoplasm, the vesicles were broken down in Golgi-associated vacuoles. The surfaces of microgamonts were highly invaginated to facilitate extrusion of numerous microgametes. Formation and maturation of microgametes were similar to those of other eimerian species. Each microgamete had two flagella, a mitochondrion, and a peculiarly shaped electron-dense nucleus that was oval anteriorly in cross section and somewhat dumbbell-shaped posteriorly. A longitudinally arranged inner membrane complex lay between a portion of the mitochondrion and the plasmalemma. About five subpellicular microtubules extended the length of the microgamete body. Macrogametogony differed little from that described in other eimerian species. Type 1 wall-forming bodies (WFB) formed in Golgi complexes early in macrogametogony, and type 2 WFB formed in cisternae of endoplasmic reticulum in intermediate stages of macrogamont development.

Notes: . The intraerythrocytic development and ultrastructure of Babesiosoma stableri Schmittner & McGhee, 1961 are described from Rana catesbeiana and Rana septentrionalis from Algonquin Park, Ontario. Morphometric and chronological observations on B. stableri in an experimentally infected Rana pipiens support the hypothesis that two successive types of merogonic cycles occur within the erythrocytes of infected frogs; the first cycle gives rise to the second and the second cycle produces merozoites destined to become gamonts. Merozoites, meronts, and gamonts are described by light and electron microscopy. Merozoites are typically coccidian and have a trilaminate pellicle with micropores, approximately 40 sub-pellicular microtubules, an apical and posterior polar ring, a conoid with two accessory rings and a pair of intra-conoidal microtubules, three rhoptries and numerous micronemes, and a nucleus with a nucleolus and a paranuclear Golgi body. The gamonts are larger than merogonic stages and are isogamous. They have approximately 55 sub-pellicular microtubules and large stores of amylopectin. These observations indicate that the genus Babesiosoma should be transferred from the Family Haemohormidiidae (Piroplasmida, Piroplasmia) to the Family Dactylosomatidae (Eucoccidiida, Coccidia).

Notes: . The development of Ichthyophthirius multifiliis trophonts in gill epithelium of channel catfish was studied on the first five days post-exposure (PE) by light and electron microscopy. Trophonts increased in average diameter from 48 μm at one day PE to 248 μm at five days PE. Although theronts invaded any part of the gill epithelium, at three days PE most parasites were found adjacent to major blood vessels, particularly the afferent vessel. From one to three days PE, 70–90% of trophonts were immediately adjacent to gill epithelium, but at four and five days PE only 50% of trophonts were closely apposed by host epithelial cells. Notable ultrastructural changes in the trophonts over the five-day period occurred in the mucocysts and lipid inclusions, both of which increased markedly in number. The few crystalline mucocysts present at one day PE were near the cell periphery but rarely attached to the plasma membrane. At three days PE, crystalline mucocysts were significantly more abundant, and at five days PE, they occurred in large numbers throughout the cytoplasm as well as at the periphery. At three days PE, secretory mucocysts were first observed. Contractile vacuoles were more prominent at five days PE than earlier in development. Development of mucocysts and lipid reserves is probably essential to survival and reproduction of the ciliate after it leaves the host.

Notes: . Two generations of pre-erythrocytic schizogony occurred in skeletal and cardiac muscle of domestic turkeys infected with sporozoites of Haemoproteus meleagridis. First generation schizonts reached maturity approximately five days post-inoculation (DPI) and developed in capillary endothelial cells and myofibroblasts. The schizonts ranged from 12 to 20 μm in diameter and produced long (5–6 μm), slender merozoites. Early second generation schizonts were first detected in capillary endothelial cells between 5 and 8 DPI. They were cylindrical and ranged in size from 5 to 8 μm in diameter and up to 28 μm in length. Second generation schizonts which reached maturity by 17 DPI were surrounded by a thick, hyaline wall and were packed with numerous spherical merozoites less than 1 μm in diameter. Mature megaloschizonts were fusiform, ranged from 30 to 113 μm in diameter, and extended as much as 465 μm along the long axis of muscle fibers. Merozoites developed as buds from cytomeres that formed between 8 and 14 DPI. Infected turkeys developed a moderate to severe myositis within 5 DPI and were lame in one or both legs. The myositis was associated with the necrosis of scattered groups of muscle fibers. Muscle fibers surrounding mature megaloschizonts were swollen and hyaline. Megaloschizonts were surrounded occasionally by fibroblasts and infiltrates of mononuclear cells. The morphology and site of development of mature megaloschizonts of Haemoproteus meleagridis are contrasted with those of other avian haemosporidians.

Notes: . Periodically, stocks of Tetrahymena vorax, which normally yield 70–90% macrostomes when subjected to heat shock or other induction methods, become low-transformers and yield ≥30% macrostomes. The addition to the post-heat-shock wash buffer (pH 6.8) of 2.7 × 10-4 M Fe3+, 1.6 × 10-5 M Cu2+, 1 × 10-4 M retinol palmitate or the adjustment of the buffer to a pH of 4 to 5 boosts transformation significantly over controls in inorganic medium alone. The addition of Fe2+ or Cu1+ has a similar, but less pronounced effect on transformation. Ferric ion (2.7 × 10-4 M) will significantly increase transformation in starved non-heat-shocked cells, whereas Fe2+, copper, retinol palmitate, and hydrogen ion concentration have no effect. The agents that boost transformation appear to act by delaying cell division in pre-transformants. Membrane fluidity, as inferred by fluorescence polarization measurements of 1,6-diphenyl-1,3,5-hexatriene, is altered in a consistent manner by starvation and heat shock. Enhancing agents, including compounds previously shown to boost heat-shock-induced macrostome formation, produce diverse shifts in membrane fluidity. Their effect on transformation of these low-transforming cells therefore appears to be attributable to some mechanism or mechanisms other than a direct alteration of membrane physical properties.

Notes: . A new amoeba, isolated from well water in Gambia, West Africa, is described and named Phreatamoeba balamuthi n. g., n. sp. Requiring anaerobic conditions for growth, it is easily cultured monoxenically with Escherichia coli or axenically in complex, undefined organic media. Three phenotypes have been observed in the life cycle: an amoeba, a flagellate, and a cyst. The amoeba moves by monopodia, is predominantly multinucleate, and varies from 11 to 160 μm in length. The flagellate has a single flagellum and is from 6 to 50 μm long. The cyst is surrounded by a resistant wall that lacks pores and ranges from 9 to 18 μm in diameter. The transformation from amoeba to flagellate can be induced nutritionally, the exact inducing factor(s) being unknown. Sexual reproduction has not been observed.

Notes: . Three periods in development that strongly influence population dynamics of Ichthyophthirius multifiliis were identified in experimental infections of channel catfish. The first occurred upon establishment within the host, 0 to 10 min postexposure (PE), when the parasite population that gained entrance declined 50%. Survival from 10 to 45 min PE, however, was constant. The second period identified came after I. multifiliis left the host and the free-living tomont encysted. The third occurred during reproduction. Although survival of encysting tomonts approached 100% among individuals departing after three to five days residence in the host, theront production varied significantly with parasite size, culture temperature during development, and length of residence by the trophont in the host. Theront production per tomont increased daily and on days three, four, and five PE was significantly higher for parasites developing at 24°C than for those at 21°C. At five days PE, mean production was 562 theronts/tomont and 240 theronts/tomont, respectively, and production by tomonts of equal size was greater for parasites maintained at 24°C.

Notes: . Electron microscope observations of the trophic and cyst stages of the protostelid Protostelium pyriformis Olive & Stoianovitch, 1969 indicate a perinuclear golgi-vesicular region in the amoeboid cell. Within this region is a microtubule organizing center (MTOC) and associated microtubules which radiate into the cytoplasm. The cyst wall is revealed as having three distinct regions, certain areas of the wall possibly corresponding to exit pores. Protostelium pyriformis shares many similarities in trophic cell structure with other eumycetozoans and members of the non-mycetozoan family Acanthamoebidae.

Notes: . The feeding behavior and morphology of Nuclearia delicatula, the type species within the genus, was studied. Feeding was by penetration of damaged or old cells of Spirogyra or by actively ingesting cyanobacterial filaments (of Phormidium), a form of behavior placing it in the family Nucleariidae. This confirms Cienkowski's (1865) observations that N. delicatula penetrates directly into damaged algal cells. This contrasts with the vampyrellids which penetrate healthy cells of Spirogyra. Characteristic behavior of the contractile vacuole complex complied with previous observations on Nuclearia. Mitochondria contain tubular, non-anastomosing cristae.

Notes: . A new genus of rigid, colorless, phagotrophic euglenoid is described from a standing pond on a salt marsh. The cell body measures 21–25 μm long, is about 18 μm wide, has a slight dorso-ventral flattening, and is marked by distinct pellicular ridges. The organism, described as Serpenomonas costata, has two unequal, antapically inserted, heterodynamic flagella. The shorter flagellum is anteriorly directed during swimming and the longer one trails posteriorly. Cells move along the substrate with a creeping motion. The ingestion apparatus is composed of separate ribs extending the length of the cell and is incorporated into the ventral pellicular ridge. The apparatus is independent of the canal and reservoir and is not protrusible. The taxonomic affinities of Serpenomonas with other euglenoids are discussed.

Notes: . Paramecium caudatum, reared on bacterized hay infusions at pH 6.5 to 6.9, were washed into various buffered solutions containing 0.016 mM CaCl2 and a pH of 3.5 to 10.4. Solutions of pH 4.5 to 9.5 support strong swimming of the cells for at least 24 h. At pH values acid to the culture medium, cells show an increasing frequency of spontaneous ciliary reversal episodes (“avoiding reactions”). Uninterrupted forward swimming is usually observed over the pH range of 7.1 to 8.5, and above pH 8.5, forward motion is interrupted by circular swimming. For all pH values tested, transfer of cells to a more acidic test solution than the solutions into which they were washed (adaptation solution) usually induced short duration, periodic ciliary reversal behavior. With transfer to a more alkaline test solution than the adaptation solution, the cells shift from forward left spiralling motion to forward right spiralling motion. With decreasing pH, the cells show progressively less sensitivity to KC1 stimulation, and at pH values of less than 5.0, cells fail to show significant ciliary reversal response to any KC1 concentration tested (1 - 128 mM). At alkaline pH values and higher KC1 concentrations, the cells show very pronounced ciliary reversal behaviors but usually fail to regain forward swimming behavior.

Notes: . Parentodinium africanum was observed in rumen contents of four Brazilian cattle and constituted 4.4, 0.9, 〉 0.1, and 10.0% of the total ciliates. This appears to be the first observation of the family Cycloposthiidae in the rumen habitat. Blepharoconus krugerensis, previously observed in the intestines of the elephant, and an unknown species of ciliate with many characteristics similar to the family Paraisotrichidae were each observed in a single animal. A new subspecies, Diplodinium flabellum laterospinatum n. subsp., is also described. Total number of ciliates per ml of rumen contents ranged from 9.0 to 51.2 × 104 and included an overall total of 55 species and 4 subspecies.

Notes: . A quantitative study of the seasonal distribution of thermotolerant (37°C and 45°C), small free-living amoebae (FLA) was conducted in Willard's Pond, a warm, monomictic lake in the Piedmont region of South Carolina. Correlation of physical and chemical parameters with the seasonal distribution was facilitated by partitioning the aquatic ecosystem into benthic, planktonic, and neustonic habitats. Population densities of FLA peaked in late summer in each habitat; however, species composition varied between habitats. Littoral sediment appeared to be the major habitat for FLA, with peaks in populations of Acanthamoeba and Naegleria in August, Hartmannella in July, and Vahlkampfia in May. Populations in profundal sediment underwent dramatic seasonal shifts, apparently in response to the seasonal chemical changes in the hypolimnion. Acanthamoeba was most prevalent in late summer, representing as much as 82% of the FLA in profundal sediment. Distribution patterns and species composition of FLA from surface water were similar to those from littoral sediment; however, a greater percentage of Naegleria was found in surface water. Numerous FLA were isolated from the neustonic community (surface film), and the number of FLA isolated in the surface film at the deep water station was found to be significantly (P 〉 0.05) greater than the number from subsurface (5–10 cm) samples. In the water column, FLA populations consistently were highest in the detrital layer, which persisted at a depth of 3.0–3.4 m throughout the summer period. The large percentage of Naegleria contributing to FLA in the detrital layer suggests that Naegleria amoeboflagellates sink through the layer, flagellate, and swim back up, such migrations possibly being triggered by a reduction of nutrients below the layer or by the presence of anoxic, reducing conditions in the hypolimnion. In addition, weather events were found to play a major role in the redistribution of FLA between various habitats in the aquatic ecosystem, with such changes probably due to resuspension of FLA from littoral sediment by wind action and input from the watershed via runoff.

Notes: . There has been a distinct lack of morphological, physiological, and taxonomic studies on the marine psammophilic ciliates despite their importance in the benthic biotope. These fragile ciliates are considered a problematical group, most research being attempted under inconvenient conditions on the seashore or at adjacent sites. To alleviate this problem, an apparatus has been designed to maintain psammophilic ciliate communities for periods of up to six months in the laboratory, making them freely available for detailed cytological study.

Notes: Book reviewed in this article:Martinez, A. Julio. 1985. Free-Living Amebas: Natural History, Prevention, Diagnosis, Pathology, and Treatment of Disease.Albert, Adrien. 1985. Selective Toxicity, The Physico-Chemical Basis of Therapy.

Notes: . Behavioral responses to light at different oxygen tensions were studied in the ciliate Loxodes striatus. In the absence of O2 it does not react to light. In the presence of O2 it reacts to light as if the pO2 had been further increased, with the induction of positive geotaxis, a transient phobic response, and finally with a permanent kinetic response (increased swimming velocity and a decreased rate of tumbling). Cells treated with cyanide behave as cells in an anoxic environment and do not react to light. It is concluded that the light response is due to the photochemical production of oxygen radicals and that the sensory receptors for O2 and for light are identical. The three types of behavioral response (geotaxis, transient, and kinetic responses) are discussed in terms of their adaptive significance for the orientation of Loxodes in the natural environment.

Notes: Book reviewed in this article:Beaver, P. C. & Jung, R. C. (with contributions by A. D'Alessandro-Bacigalupo, J. H. Esslinger, S. P. Katz, M. D. Little, E. A. Malek, T. C. Orihel, and R. G. Yaeger).

Notes: . This study compared the response of protozoan communities colonizing artificial substrates to complex effluents in laboratory microecosystems and validated laboratory predictions using similar sampling methods in the field. Sampling stations were established on a small stream receiving heavy metal and sewage treatment plant effluents. Water from each station was used as a test medium to test the colonization of polyurethane foam artificial substrates in laboratory microecosystems. The heterotrophic index (HI), the ratio of total community biomass to chlorophyll biomass, was also measured. Field colonization trials and HI determinations were carried out at each sampling station. There was a strong negative relationship between the number of species colonizing artificial substrates and the total metal concentration of the water at each station; protozoan colonization was severely depressed at the highest metal concentrations (〉400 μg/liter) and recovered at downstream stations where dilution had taken place. The HI differentiated stations receiving high heavy metal concentrations in the field from those recovering from heavy metal influences. Effect concentrations based on laboratory experiments predicted that the concentration of total heavy metal that would produce a 5% decrease in species number (EC5) was 18 μg/liter, approximately the background concentration found in the stream studied. These experiments demonstrated that laboratory microecosystems or microcosms can be used to predict effects in the field accurately.

Notes: . The pellicular ultrastructure of Euglena ehrenbergii and Euglena oxyuris is examined for differences that might account for their very different motile activity. Their general ultrastructure is closely similar, but the continuity between adjacent pellicular strips found in E. oxyuris would severely restrict strip sliding and contribute to this species’very limited cell-shape changes. The strips of E. ehrenbergii have plate-like projections, whose constant orientation is consistent with strips sliding without deforming obliquely on cell rounding. Some variations and constant features in pellicular ultrastructure are noted and discussed in regard to the sliding strip mechanism for euglenoid shape changes.

Notes: . Loxodes reached peak abundance close to the oxic-anoxic boundary (O2 5% atm) in two lakes, in test tube cultures, and in glass chambers with horizontal O2 gradients. Vertical profiles of CO2, pH, sulfide, and Fe2+ in a lake were not closely related to Loxodes abundance. In a laboratory experiment, Loxodes followed a retreating source of O2 and was repelled by a high pO2. This behavior was sustained when cells simultaneously swam up or down gradients of both CO2 and pH. Aggregation of cells was abolished by KCN (10-4-10-6 M). Sodium azide (10-1-10-4 M) had no effect and 2,4-DNP sharpened the aggregation. Rotenone, Antimycin A, and HOQNO had no obvious effect. Cytochrome oxidase is probably the oxygen receptor. Loxodes striatus contained low activities of superoxide dismutase and catalase. Extracellular production of superoxide (O-2) and hydrogen peroxide (H2O2) were probably not responsible for the exclusion of Loxodes from water with a high pO2. Continuous exposure of Loxodes to oxygen at normal atmospheric pressure at 10°C led to 50% mortality in 10 days. Cells left free to swim in an oxygen gradient doubled their number in the same period. Light exacerbated the toxic effects of O2. Behavioral responses to the dissolved oxygen tension probably controlled the spatial distribution of Loxodes.

Notes: . After formation, food vacuoles in the microstomal cell type of Tetrahymena vorax labeled with either India ink or cationized ferritin (CF) undergo three morphological stages during processing: an initial condensation with a size decrease, the appearance of a clear halo around the condensed marker that apparently results from expansion of the vacuole, and the disappearance of the halo region. These stages correspond generally to three of the four stages described previously for Tetrahymena. Fusion of older vacuoles prior to egestion may represent a fourth stage in vacuole processing. The developing vacuole has small coated regions of the membrane; two populations of vesicles, small coated vesicles and larger vesicles with electron-dense cores, are adjacent to it. During the initial condensation, tubular invaginations with CF project from the vacuole. After the disappearance of the halo, much of the vacuolar surface appears to be multilayered with extensive folds containing CF. Labeled flattened or curved disks and closed rings occur in the cytoplasm often adjacent to labeled vacuoles and in the vicinity of the cytoproct. The appearance of labeled disks and rings is identical to both labeled and unlabeled vesicles adjacent to the cytopharynx, adding additional support to the hypothesis that, like Paramecium, vesicles retrieved from food vacuoles during processing as well as after fusion with the cytoproct may be reused for vacuole formation.

Notes: . The purpose of this research was to examine the roles of physico-chemical parameters in structuring protist communities that colonize artificial substrates. Polyurethane foam (PF) substrates were placed in five lentic systems in northern lower Michigan during summer 1983. These lentic habitats represented a range of trophic states and included three lakes, a bog, and a marsh. Triplicate PF substrates were sampled after 1, 3, 7, 14, 21, and 42 days of exposure. During this study, 90 living protist samples were examined for the number and kinds of species. Water samples were analyzed for pH, conductivity, alkalinity, chloride, silica, temperature, dissolved oxygen, ammonia, and total and ortho-phosphate concurrent with artificial substrate collections. A total of 546 protist species was recorded. Only seven species were found in over 50% of the samples and 121 species were found in only one sample. The 96 most common species were examined in relation to environmental parameters using several multivariate statistical procedures. Factor analysis (principal components with varimax rotation) performed on the total environmental data set showed that three composite factors explained 85% of the variability of the data set. A reciprocal averaging ordination (RAO) was used to reduce species presence/absence data and to separate samples graphically by their species composition. Significant correlations, with RAO-generated axes from all five systems, were found for pH, oxygen, and a nutrient factor to axis 1. Factor analysis on the physico-chemical parameters of the three lakes showed that three factors explained 71% of the environmental data set variability. The RAO-generated axis (axis 1) was correlated with silica, ortho-phosphate, and Factor 2, which was primarily comprised of loadings from ortho-phosphate. These techniques support the hypothesis that a limited number of environmental parameters strongly affect protist community composition.

Notes: Book reviewed in this article:Poinar, G. O., Jr. & Thomas, G. M. 1984. Laboratory Guide to Insect Pathogens and Parasites.James, D. M. & Gilles, H. M. 1985. Human Antiparasitic Drugs: Pharmacology and Usage.

Notes: . The fine structure of cells and isolated pellicular sheets of Astasia longa is examined in relation to the sliding between neighboring pellicular strips believed to accompany euglenoid movement. Sonication breaks the repeating structure of the pellicle in the groove region where tannic acid fixation resolves separate but overlapping pellicular strips. All microtubules (including the two lying between the overlapping strips) remain with the inner strip. We therefore suggest that this fracture plane defines the boundary where sliding occurs between adjacent pellicular units. Consistent with this, traversing filaments running from groove to groove to connect the pellicular strips of adjacent units are displaced from transverse to oblique when elongated cells round up. In contrast, no changes suggestive of distortion within each unit are detected in the particle arrays of the ridge plasma membrane that overlies a single unit. The results are discussed in relation to previous ultrastructural studies, the site at which the force to cause sliding is generated, and the role of the traversing filaments.

Notes: . Cross-reactivity between fungal and Trypanosoma cruzi polysaccharides, owing to common residues of β-D-galactofuranose, β-D-galactopyranose, and α-D-mannopyranose, was demonstrated by using a) rabbit immune sera against T. cruzi epimastigotes and b) sera from patients with Chagas’disease. Several chagasic (Ch) sera precipitated partly purified galactomannans from Aspergillus fumigatus and from T. cruzi epimastigotes and also the galactoglucomannan from Dactylium dendroides. Reaction of one Ch serum with T. cruzi galactomannan (GM) was completely inhibited by synthetic β-D-Galf-(1 → 3)-Me α-D-Manp, and that of another Ch serum with a purified D. dendroides galactoglucomannan (GGM) was partly inhibited by (1 → 6)-linked (81%) or by (1 - 3)-linked (33%) β-D-Galf-Me α-D-Manp. The β-D-Galf-(1 → 3)-α-D-Manp epitope was present in both T. cruzi and D. dendroides polysaccharides. Rabbit anti-T. cruzi antisera precipitated A. fumigatus GM, T. cruzi antigenic extracts containing the lipopeptidophosphoglycan (LPPG), T. cruzi alkali-extracted GM, a synthetic GM, and D. dendroides GGM. Weak reactivities were obtained for a Torulopsis lactis-condensi GM containing β-D-Galp terminal residues and for baker's yeast mannan with α-D-Manp-(1 - 3)-α-D-Manp-(1- → 2)-α-D-Manp-(1 → 2) side chains. An anti-LPPG rabbit serum precipitated D. dendroides GGM—a reaction inhibited (82%) by β-D-Galf-(1 → 3)-Me α-D-Manp and, less efficiently, by a (1 → 5)-linked β-D-Galf-tetrasaccharide. Sera from mice immunized with D. dendroides whole cells reacted with CL-strain trypomastigotes as shown a) by indirect immunofluorescence, b) by a Staphylococcus adherence test, but were not lytic. Mice immunized with D. dendroides were not protected against a challenge with virulent T. cruzi trypomastigotes.

Notes: . The H immobilization antigens specified by the SerH locus of Tetrahymena thermophila have been purified by a procedure utilizing acid fractionation, ammonium sulfate precipitation, gel filtration, and ion exchange chromatography. Purified antigen migrates as a single band on SDS-PAGE and IEF. Molecular weights of the four allelic H antigens range from 44,000 to 52,000, and isoelectric points range from 4.1 to 4.5. No carbohydrate was detected.

Notes: . We have reinvestigated whether surface immunoglobulin (sIg) on Trypanosoma lewisi is antibody directed toward parasite antigen by using flow cytometry to analyze parasites stained with fluoresceinated F(ab′)2 fragments of antibodies to rat IgG and IgM. We have confirmed that IgG antibody to the parasites is present both in the serum of rats and on the surface of parasites between the fourth and twentieth days of infection, that the amount of sIg per cell increases as the infection progresses, and that considerably more IgG is present on parasites harvested from intact rats than on those from rats that had been immunosuppressed by whole body γ-irradiation. In addition sIgM was detected on trypanosomes from intact, but not on parasites from irradiated rats. We have also made two observations suggesting that not all sIg is specific antibody made in response to T. lewisi. First, a low but significant amount of sIgG was detected on parasites throughout infection in irradiated rats; no sIgM was detected on these parasites. Second, when parasites harvested from immunosuppressed rats were incubated in normal rat serum, the amount of both sIgG and sIgM detected by flow immunofluorescence increased. Parasites harvested from intact animals bound IgM but not IgG from normal rat serum. These results suggest either that natural antibody to the trypanosomes is present in the serum of uninfected rats or that some rat immunoglobulins bind to structures on the trypanosome surface in ways that do not depend on usual antigen-antibody interactions. Finally, flow immunofluorescence was also used to detect complement component C3 on the surface of both intact and trypsinized bloodstream forms harvested from intact or immunosuppressed rats. The amount of sC3 per cell did not increase until late in the infection and consequently did not correlate with the increase of sIgG. Therefore, T. lewisi avoids destruction by the immune system although immune effector molecules, IgG, IgM, and C3, are on its surface.

Notes: . A glycosphingolipid fraction from Trypanosoma mega was isolated after acetylation and was further purified on a silicic acid column. Final purification was by preparative thin-layer chromatography. The carbohydrate components of the glycolipid were fucose and galactose in approximately equimolar amounts. The neutral glycolipid of T. mega has a sphingosine base composition that consists of sphingosine and traces of dihydrosphingosine. Fatty acids forming amide groups with the sphingosine bases were analyzed by gas-liquid chromatography-mass spectrometry and are a mixture of normal and α-hydroxy fatty acids. Normal C16:0, C18:0, and 2-hydroxy C18:0 are the predominant fatty acids.

Notes: . The antigenic relationships between Leishmania mexicana pifanoi promastigotes, axenically grown amastigotes, and amastigotes isolated from the footpads of infected hamsters or from a J774 macrophage cell line were studied by three serologic methods. Amastigote and promastigote antigens were disrupted by freeze-thawing of intact cells in a lysis buffer. Antisera were prepared in rabbits by repeated subcutaneous inoculations of the parasite antigens in complete Freund's adjuvant and were tested against the homologous and heterologous antigens in a series of gel diffusion experiments. Negative results were obtained in all control experiments. In each instance, the homologous antigen-antiserum reactions yielded the largest numbers of precipitin lines. A pattern of cross-reactivity was also observed in the heterologous systems. Results indicated that the amastigote and promastigote forms had unique and common antigens. The two parasite antigen-antiserum systems were also examined by immunoelectrophoresis. Qualitative and quantitative differences between the promastigote and amastigote antigens were readily demonstrable by this technique. Results indicated that each parasite form had specific and many common antigens. In the homologous system, major proteins, with molecular weights (MW) of 23, 52, and 68 kd, were demonstrated in the promastigotes by immunoprecipitation of lactoperoxidase-catalyzed radioiodinated cells. In a similar (homologous) system, axenically grown amastigotes were found to contain three proteins with MW of 38, 70, and 74 kd and, therefore, different from those demonstrated for the promastigotes. All the results suggested that the three amastigote stages of different origins are antigenically similar to one another, but differ from the promastigote forms.

Notes: . We used high-resolution polyacrylamide gradient gel electrophoresis (PGGE) to separate four babesial enzymes to aid in the identification of two Babesia microti isolates established in hamsters. The isolates were compared to two different hamster passages of the “Gray” strain. All isoenzyme patterns from the two isolates and the “Gray” strain were similar except glucose phosphate isomerase (GPI) from one of the “Gray” strain passages. It showed a polymorphic GPI pattern as opposed to a monomorphic GPI pattern seen in the other “Gray” strain passage and the two isolates. The observed differences suggested (i) that some populations of B. microti are capable of having polymorphic GPI, (ii) that the “Gray” strain originally contained (and may still contain) a heterogenous population of B. microti, and (iii) that the population possessing polymorphic GPI was selected over that with monomorphic GPI. This information was obtained by a PGGE method that eliminated hemoglobin from gels and allowed, for the first time, detection of babesial leucine amino peptidase (LAP) and isocitrate dehydrogenase (IDH). In addition, this method provided molecular weight estimations on babesial GPI, LAP, IDH, and glutamate dehydrogenase (GDH), and it proved useful in the identification and characterization of the B. microti isolates.

Notes: . Crude homogenates of the ciliate protozoon, Tetrahymena thermophila, can hydrolyze the potent acetylcholinesterase inhibitors O, O-diisopropylphosphorofluoridate (DFP) and O-1,2,2-trimethylpropylmethylphosphonofluoride (soman). Characterization of the enzymatic activity of the homogenate has been performed. The DFPase operates over a pH range of 4 to 10 and an ionic range of 0–500 mM NaCl. Rate of reaction increases three-to four-fold from 25°C to 40°C and is still present at 55°C. These results indicate that the enzymatic activity operates over a broad range of environmental conditions, making it an attractive material for use in the detoxification and detection of organofluorophosphates. DFPases may be important in the metabolism of naturally occurring organophosphates.

Notes: . Twenty-six species of bacteria and seven species of yeasts were aseptically presented separately as potential food sources to the estuarine heterotrich ciliate, Fabrea salina, under standardized conditions of cell number, medium, and temperature. The bacteria and yeasts were classified according to their effect on the intrinsic growth rate of the ciliate: Nutritious bacteria: Photobacterium fisherii, Vibrio neresis, V. natriegens, Flavobacterium sp. strain ASN16, V. harveyi, Xanthomonas sp. strain ASN22; Maintainer bacteria: V. vulnificus, Vibrio sp. strain V344, V. alginolyticus, V. anguillarum, Bacillus sp. strain ST332B2, Vibrio sp. strain V415, Acinetobacter sp. strain BHT8, Flectobacillus marinus, and Enterobacter aerogenes; Maintainer yeasts: Candida albicans and Cryptococcus marcerans strain 2; Nonnutriuous bacteria: V. parahaemolyticus, Planococcus sp. strain ASN13, P. mandapapanensis, Hyphomicrobium vulgari, Pseudomonas sp. strain CNS1, an unidentified gram-negative, yellow-pigmented, motile bacillus strain IG9A2, Thiobacillus thioparus, Escherichia coli, Corynebacterium sp. strain BR 17, Achromobacter sp. strain 23030, and Oceanospirillum beijerinckii; Nonnutriuous yeasts: C. marcerans strain 1, Saccharomyces cerevisiae, Debaryomyces hansenii, an unidentified black yeast, and C. laurentii. Under the conditions specified, bacteria appear to have a certain minimal value for the growth of Fabrea while yeast have little to none.

Notes: . A new and inexpensive medium is described for axenic mass cultivation of Paramecium tetraurelia stock 51s and the double mutant pawn A/pawn B. Skim milk powder is the major carbon and nitrogen source in this medium. Growth characteristics (proliferation rate, final cell density, and cell size) are similar to those observed with other axenic culture media. Cultures were run in one-liter erlenmeyer flasks, a 20-liter, and a 250-liter airlift bioreactor. The yield of a large bioreactor is 750 g (wet wt.) Paramecium or 5 × 109 cells. This easy, economical culture technique will greatly facilitate the use of Paramecium as a model organism for extensive biochemical studies.

Notes: . Metacyclic trypanosomes developed in populations of procyclic forms of four stocks of Trypanosoma brucei rhodesiense cultivated at 28°C in a liquid medium containing explants of tsetse fly head-salivary glands, alimentary tract, abdominal body wall, or thoracic muscle. The cultures became infective for mice 7–16 days after they were prepared, and infective trypanosomes were present for prolonged periods. In the culture series of stock TRUM 545, infectivity persisted for 138 days when the cultures were terminated. Only one explant of thoracic muscle tissue was required for the production of metacyclic stages in stock TRUM 497 cultures. Infectivity titrations on trypanosome suspensions from cultures of stocks TRUM 497, TRUM 545, and TRUM 567 revealed that only a small proportion of the culture population was infective. Using stock TRUM 530, mice were infected consistently from inoculations of trypanosomes grown in the presence of explants; infectivity of the trypanosomes ceased when the explants were removed from the flasks, but reappeared when they were returned to the cultures. Parasites grown in medium “conditioned” by explants produced sporadic infections in mice. The control cultures of trypanosomes grown in medium alone were generally not infective, but two of the stocks produced occasional parasitemias. Stained samples of infective inocula contained a few epimastigote-like and metacyclic-like trypanosomes.

Notes: . The Taiwanese strain of Leucocytozoon caulleryi was isolated from an infected chicken in Taipei, Taiwan, and established in chickens and biting midges Culicoides arakawae from Japan. Sporogony of the strain in C. arakawae was completed on day 3 after the infective blood meals at 25°C. Sporozoites isolated from the salivary glands of C. arakawae on days 3 or 4 after feeding caused infection in all the chickens inoculated. The strain showed high pathogenicity for chickens. Mortality of chickens rose with an increase in the number of sporozoites inoculated. The prepatent period for chickens inoculated with sporozoites was 14 days. Parasites appeared in the peripheral blood of chickens on day 15 and disappeared on day 26 after sporozoite inoculation. Soluble antigens were found in the sera of chickens infected with the strain between 10 and 17 days after inoculation, and homologous antibodies appeared after 17 days. Antigens prepared from sera, schizonts, merozoites, and gametocytes of the Taiwanese strain reacted with the sera of chickens infected witt the same strain or the strain isolated in Japan. The chickens that recovered from a primary infection with the Taiwanese strain demonstrated complete resistance to reinfection with the same strain or the strain isolated in Japan.

Notes: . Cells of Euplotes patella syngen 2 have been found to excrete mating-type-specific gamones (mating-inducing factors) into the surrounding medium. In strains of E. patella syngen 2, intraclonal conjugation (selfing) sometimes occurred spontaneously. Cell-free fluids of clonal cultures in which spontaneous selfing occurred were not effective in inducing homotypic pair formation in other cultures of the same mating type as the culture from which the cell-free fluids were derived. This indicates that selfing cultures produce only those gamones expected of their own mating types. This result suggests that spontaneous selling in E. patella syngen 2 is not due to change of mating type expression. The possible mechanisms for spontaneous selfing in E. patella syngen 2 are discussed.

Notes: . A trichomonad flagellate, Tritrichomonas mobilensis n. sp., is described from the large intestine of the squirrel monkey, Saimiri boliviensis boliviensis. The organism has a lanceolate body 7–10.5 μm in length; a well developed undulating membrane; a stout, tubular axostyle with periaxostylar rings that terminate in a cone-shaped segment projecting from the posterior end of the cell; and a moderately wide costa. The anterior flagella are about as long as the body, and the recurrent flagellum is of the acroneme type. All its characteristics suggest that the new species belongs in the Tritrichomonas augusta type of the subfamily Tritrichomonadinae.

Notes: Book reviewed in this article:Lee, J. J., Hutner, S. H. & Bovee, E. C., eds. 1985. An Illustrated Guide to the Protozoa.

Notes: . Thirty-seven strains of Acanthamoeba are described using 69 physiological characters. The characterizations of the strains are based on quantitative estimates of growth responses which were standardized to eliminate any effects of differences in growth rates of the strains. Thirty-six distinct patterns are generated using numerical profiles, but it is possible to distinguish within this total diversity 20 sub-generic groups which, although not completely congruent with the morphological species, are consistent with other non-morphological characters. Evidence is presented that Acanthamoeba is a coherent, well-founded genus, but it is not desirable to recognize formal taxa at the sub-generic level.

Notes: . This paper examines the biology, morphology, and pathogenicity for mice of an amoeboflagellate isolated from human nasal mucosa. The biological and morphological relationships of this isolate with the amoebae (Lobosea) and the true slime molds (Eumycetoza) are discussed though the taxonomic affinities of the organism have not been determined.

Notes: . A quantitative technique for the assessment of sporozoite infectivity in vivo, using intra-cecal inoculation of Eimeria tenella sporozoites, has been developed. Evaluation of the infection using cecal lesion scores and oocyst counts showed that this technique should be useful for the quantitation of sporozoite viability and thus for the anti-sporozoite activity of different treatments prior to inoculation. Pre-treatment of sporozoites with heat-inactivated hyperimmune antisera neutralized sporozoite infectivity in vivo and indicated that antibodies in the absence of complement inhibited sporozoite infectivity in vivo.

Notes: . An assay has been developed using parasite-specific incorporation of 3H-uracil to assess the intracellular growth of Eimeria tenella in vitro. As shown by both scintillation counts and autoradiography, 3H-uracil was incorporated specifically into intracellular parasites from the onset of infection and continued throughout development of the first generation schizonts. Mature schizonts and first generation merozoites did not continue to incorporate additional 3H-uracil, indicating that RNA synthesis had halted in these stages. Based on these findings, a semi-automated microscale uracil incorporation assay was developed to determine parasite viability. This method should be useful for biochemical studies with intracellular parasites and for screening compounds for anticoccidial activity. The ease, rapidity, and quantitative nature of this assay contrasts favorably with standard morphometric approaches of determining parasite development. In addition, parallel studies using host cell incorporation of 3H-uridine have been introduced as a method of determining whether antiparasitic activity is direct or indirect in relation to effects on the host cell.

Notes: . Binary fission of the sessiline loricate peritrich Thuricola folliculata was examined by light microscopy. Cytokinesis, which occurs in the oral-aboral median plane, is 35-40 min in duration. Cytochalasin B (CB) was used at final concentrations of 50–100 μg/ml in dimethyl sulfoxide (DMSO). A transition point occurs at about 20 min after the beginning of binary fission and about 10 min before cytokinesis. In most cases, 50 μg/ml and 100 μg/ml CB added to cells before the transition point resulted in delays of cytokinesis which were significant compared to DMSO controls; sometimes, cytokinesis was blocked completely. If added after the transition point, CB had no effect on cells.

Notes: . Water-soluble and 0.6 M KCl-soluble protein fractions prepared from Tetrahymena pyriformis, when inoculated into mice, could effectively induce activated macrophages having the ability to kill Toxoplasma gondii in vitro. This effect was not induced by other proteins tested, such as bovine serum albumin, pepsin from porcine stomach mucosa and chicken egg-white lysozyme, nor by muramyl dipeptide (MDP), a potent immunoadjuvant. Five fractions obtained by DEAE-Sephadex chromatography of the water-soluble protein fraction were compared with regard to induction of toxoplasmacidal activity in macrophages. The first peak was most effective for activation of macrophages. Five fractions obtained by chromatography of the 0.6 M KCl-soluble protein fraction were also examined and it was found that the first peak had the activity. No marked difference in activity was observed between the active fractions of water-soluble and 0.6 M KCl-soluble protein fractions. For practical use, we focused on the water-soluble active fraction. The minimum effective dose of the active fraction was 100 μg and the fraction could activate macrophages directly in vitro. Four fractions obtained by gel filtration of the active fraction on Sephadex G-200 were compared and the first peak had the activity. The first peak contained a single protein, revealed by SDS-polyacrylamide gel electrophoresis; its apparent molecular weight was 64,000.

Notes: . We have determined the complete nucleotide sequence of the coding region of the small subunit rRNA gene of the hypotrichous ciliate Euplotes aediculatus. It is 1882 nucleotides long and contains several inserts not present in the small subunit rRNA genes of the hypotrichs Oxytricha nova and Stylonychia pustulata. A comparison of the sequences suggests that E. aediculatus is much less closely related to these other two hypotrichs than they are to each other. Although the gene sequence of E. aediculatus is drifting more rapidly than those of these other two species, its faster evolutionary clock is not enough to account for the degree of difference between them.

Notes: . DNA is eliminated during development of the somatic MACronucleus from the germinal MICronucleus in the ciliated protozoan, Tetrahymena thermophila. Facultatively persistent sequences are a class of sequences that persist in the MAC DNA of some cell lines but are eliminated from the MAC DNA of other cell lines. One cloned MAC fragment contains a persistent sequence as well as sequences normally retained in the MAC. When this cloned fragment was used to construct MAC restriction maps of this region in cell lines whose MAC DNAs do, or do not, contain the persistent sequence, extensive variation in the map flanking this region was observed. The different DNA rearrangements of this MIC segment are epigenetically determined during or soon after MAC development. Moreover, different rearrangements may occur among the 45 copies of this MIC segment as a MAC is formed, resulting in polymorphisms that are later resolved by phenotypic assortment.

Notes: . One stock, GBC, has a maximum temperature of growth about 5°C lower than other recently collected stocks of Paramecium primaurelia. The resistant stocks (R) are able to grow continuously at 35°C while the sensitive stock (S) cells die within 48 h. The F1s of R x S crosses exhibited a cytoplasmic pattern of inheritance and all F2-by-autogamy lines derived from the S cytoplasmic parent are sensitive. The F2-by-autogamy lines cytoplasmically descended from the R parent were predominantly (93%) R in the initial assay. Upon reinvestigation one year later, only 64% of these lines were R, 9% were S, and 27% had a new phenotype, weak (W), intermediate between R and S. Backcrosses of W lines to both R and S strongly suggest that the W lines have normal cytoplasm (i.e. R) but also have nuclear gene(s) for temperature sensitivity that are derived from the original S stock. The delayed manifestation of the W phenotype is not understood.

Notes: . The combined action of cadmium and selenium was investigated in two unicellular marine phytoplankton organisms, Prorocentrum micans Ehrbg. and Crypthecodinium cohnii Biecheler, in culture. When present simultaneously in low concentrations, the toxicity of Cd and Se is decreased (antagonism) whereas if one of these elements is present at high concentration, especially for a long period, their toxicity is increased (synergism) more than if they were separate. This interaction affected cell organelles in the following ways: vacuolation, development of chloroplast pyrenoids, and elicitation of lipid vesicles, starch vesicles, giant mitochondria, and lysosomes. Possible competition between these two elements for binding sites is discussed.

Notes: Book reviewed in this article:Müller, H. J., ed. 1985. Bestimmung wirbelloser Tiere im Gelände. Bildtafeln für zoologische Bestimmungsübungen und Exkursionen. G. Fischer

Notes: . The yield of the bacterium Enterobacter aerogenes and the ciliate Colpidium colpoda was dependent on initial phosphorus concentrations in batch cultures containing 125 or 250 mg/liter glutamate and 50–1000 μg/liter phosphorus. For both, yield per unit phosphorus declined at higher phosphorus concentrations. A marked decline in growth rate in bacterial cultures was coincident with the depletion of dissolved phosphorus and the development of rapid orthophosphate turnover times. Colpidium introduced to these cultures consumed about 16,000 bacteria/h/ciliate while multiplying exponentially and relieved phosphorus limitation, as indicated by a longer turnover-time for phosphate. The longer turnover-time was due to the reduction of bacterial numbers; in cultures with ciliates, bacteria appear to be more active in taking up phosphate, and much of the total phosphorus accumulates in ciliates. Ciliates released both inorganic and organic phosphorus, but the organic phosphorus did not accumulate to excess in the cultures to an extent that would indicate that it is less used by bacteria. Although ciliates release enough phosphorus to account for ca. 20% of the bacterial uptake, ciliates appear to behave as phosphorus sinks as much as phosphorus recyclers in these closed systems.

Notes: . Cells of Paramecium bursaria, which harbor a symbiotic alga of the genus Chlorella, reverse the effective beat of their cilia and swim backward when stimulated in either of two ways. Ionic stimulation is introduction of cells into a solution high in K+ while step-down photostimulation is a sharp reduction in the intensity of light falling on the culture. Much is known about the mechanisms of ionic stimulation of ciliary reversal, but little is known about step-down photobehavior. Inhibitors of ionically stimulated ciliary reversal were applied to cells undergoing step-down photobehavior; Ca-channel inhibitors, neomycin and W-7, inhibit both behaviors. Activation of Ca-channels in the ciliary membrane is involved in step-down photobehavior, suggesting that the algae may alter the Paramecium membrane to make it more excitable.

Notes: . Isolated mitochondria of the anaerobic protozoan Blastocystis hominis were subjected to spectral analysis, color, catalase, and peroxidase tests and found to be completely negative for cytochrome enzymes, catalase, and peroxide. Based on the absence of cytochrome enzymes, the possible evolution of B. hominis mitochondria from anaerobic bacteria is postulated.

Notes: . New information on the life cycle and fine structure of Pilosporella chapmani, a microsporidium of the mosquito Aedes triseriatus, is presented. Pilosporella chapmani is shown to have two sporulation sequences, one of them being involved in transovarial transmission. One sequence, involving meiosis and production of a moniliform sporogonial plasmodium, occurs in the larval fat body, resulting in eight uninucleate, spherical, and fully developed spores. The other occurs in oenocytes of adult mosquitoes and results in isolated, binucleate, elongate, and thin-walled spores. Also, for the first time, metabolic products are shown to be expelled into the surrounding host tissues through the wall of the sporocyst.

Notes: . The flagellated protozoon Trichomonas vaginalis, parasite of the human urogenital tract, possesses a well developed microtubule system organized in highly differentiated structures. We have shown by immunoblotting that monospecific anti-sheep brain tubulin antibodies are able to react with the microtubular tubulin of T. vaginalis. These antibodies were used to study the microtubular system of T. vaginalis both in interphase and mitosis by indirect immunofluorescence. The interphase microtubular pattern, characterized by an axostyle, a pelta, four anterior flagella, and a recurrent flagellum, displayed remarkable changes at the onset of mitosis: the axostyle disappeared, and two pole bodies connected by a short spindle became evident; chromosomal fibers arose while pole-to-pole fibers elongated. The last phases of mitosis were marked by the disappearance of chromosomal fibers, the appearance of two small axostyles, and the depolymerization of the pole-to-pole bundle. At the end of mitosis, the normal interphase microtubule pattern was observed.

Notes: . The Müller vesicle is a characteristic organelle of loxodid ciliates. Its structure and development have been investigated using light microscopy and TEM. The organelle consists of a membrane-covered mineral body (the statolith), a vacuole, and various structures derived from the overlying kinety. There is strong evidence that the vesicle functions as a gravity sensor: a) its structure and relative dimensions fulfil the minimum requirements of a functional statocyst; b) its structure bears a close resemblance to the statocysts of some higher animals; c) re-orientation of the cell with respect to gravity produces a gravity-induced displacement of the mineral body, and d) geotaxis in Loxodes can be demonstrated experimentally. The transduction of the signal probably takes place at the level of the two kinetosomes of the organelle, one of which is in close contact with the cell membrane, while the other is connected to the statolith by a fairly rigid stalk containing a bundle of microtubules.

Notes: . Mouse peritoneal macrophages exposed to the RNA from the spleens of mice infected with Trypanosoma cruzi (iRNA) exhibit enhanced resistance to this parasite. The poly(A)-containing iRNA was found to be the active fraction. No such activity was observed in macrophages incubated with RNA from normal mice (nRNA) or with synthetic poly A.

Notes: . A large number of trypanosomatids was seen in the latex of the milkweed plant Euphorbia hyssopifolia found in Rio de Janeiro, Brazil. The parasites, which belong to the genus Phytomonas, were isolated and axenically cultivated in a biphasic culture medium. The form and the structural organization of the parasites as found in the intact plant, in the isolated latex, and in the culture medium was studied by scanning and transmission electron microscopy.

Notes: . A method to determine the volume of individual live Tetrahymena with improved precision has been developed. Cells are compressed between a microslide and coverslip and their volume calculated from their thickness and area determined by interference microscopy and from a photomicrograph, respectively. A formula has been derived to correct for the “edge error” which arises because the rounded edges of the cells are projected on the film as if they were rectangular. The minimal surface area of compressed cells, i.e. that necessary to surround the cells smoothly, can also be calculated. Cells may be recovered after measurements. The method may also be applied to other cells.

Notes: . Pyrotheca hydropsycheae n. sp. is described from caddis fly larvae, Hydropsyche siltalai Döhler, 1963. All stages were found in oenocytes and fat body cells. Meronts were uni- or binucleate with simple surface membranes. The sporogonic stages were recognized ultrastructurally by the separation of an envelope, the sporophorous vesicle, from their surfaces. Mature sporogonial plasmodia were tetranucleate and gave rise by longitudinal fission to four uninucleate elongate sporoblasts with polar nuclei. Spores were lageniform with an inflated posterior end, containing the polar tube coils and the nucleus, and a narrow anterior section comprising two-thirds of the length, containing the polaroplast and straight part of the polar tube. The polaroplast consisted of an anterior region of loosely packed membranes arranged as partitions at angles to one another and a posterior region of increasingly closely packed parallel membranes. The spore wall consisted of an electron-dense exospore with a fuzzy coat and a thin electron-lucent endospore. All four spores derived from a sporont faced in the same direction in the sporophorous vesicle. Spores measured 8.7 μm long and extruded polar filaments were about 20 μm.

Notes: . The interaction between the predator, Discophrya collini, and the prey, Tetrahymena pyriformis, was investigated in order to determine what prey factors(s) may be recognized, facilitating the feeding response of the suctorian predator. When viable, deciliated T. pyriformis were offered to starved D. collini, only 12% of the suctoria captured a ciliate compared to 98% when presented with ciliated T. pyriformis. Starved suctoria were also offered three types of tanned sheep erythrocytes used as artificial prey. After exposure to erythrocytes coated with sonicates of T. pyriformis, 16% of the D. collini displayed one or more attached erythrocytes but after incubation with erythrocytes coated with T. pyriformis cilia, 72% of the suctoria had tentacles with attached cells. No untreated, tanned erythrocytes were captured. When T. pyriformis were treated with ciliary antiserum and then offered to starved D. collini, the resulting capture rate was low (13%), but T. pyriformis treated with normal rabbit serum were captured at a rate comparable to controls (92%). These results suggest that the prey's cilia and/or their surface macromolecules may facilitate prey capture.

Notes: . Entamoeba histolytica kills mammalian target cells in a multi-step sequential process with separate adherence, cytolytic, and phagocytic events. In the studies reported here, we used fluorescein isothiocyanate linked to dextran to label the endocytic vesicles of the HM1 strain of E. histolytica and measure vesicle pH (5.1 ± 0.2 by spectrofluorimetry). Concentrations of NH4Cl (1.0–10.0 mM) sufficient to increase vesicle pH to °5.7 inhibited amebic killing of target Chinese hamster ovary (CHO) cells as assayed by trypan blue staining, by the release of 3H-thymidine previously incorporated into CHO cell monolayers, and by the release of 111indium oxine from radiolabeled CHO cells. Similar effects were also observed with two other weak bases, primaquine and chloroquine (both 50 μM). In contrast, NH4Cl (10 mM) did not affect either the adherence or phagocytic events, as measured by amebic adherence to CHO cells at 4°C and by the binding and ingestion of 3H-leucine-labeled bacteria. In the presence of NH4Cl and and the carbohydrate ligand asialofetuin, there was no evidence of intracellular trapping of the amebic galactose-inhibitable lectin; inhibition of adherence by cycloheximide (10 μg/ml for 3 h) suggested rapid turnover of the surface lectin. Prolonged exposure to NH4Cl for 48 h (which had no effect on amebic protein synthesis) or shorter exposure to cycloheximide (10 μg for 3 h) produced persistent inhibition of cytolysis. These results indicate that an uninterrupted acid pH in intracellular endocytic vesicles is necessary for the cytolysis of target cells by E. histolytica trophozoites.

Notes: . The cytotoxic activity of a cell-free extract of Naegleria fowleri amebae on B103 rat nerve cells in culture was investigated. The cell-free extract was prepared by subjecting lysed amebae to centrifugation at 100,000 g for 1 h, precipitation of the supernatant fluid with 30–60% saturated ammonium sulfate, and desalting by group exclusion chromatography utilizing Sephadex G-25. The supernatant fluid recovered from this procedure was termed the soluble fraction. The Naegleria cytotoxic activity present in the soluble fraction was assayed by 51Cr released from labeled B103 cells. The Naegleria soluble fraction, when added to nerve cells, elicited blebs on the B103 target cell surface within 5 min after exposure to the fraction. Later, holes were observed in the B103 cell plasma membrane. These alterations were never observed on untreated B103 cells. Phospholipase A, phospholipase C, and protease activities were associated with the desalted ammonium sulfate-precipitable cytotoxic activity of N. fowleri cell-free lysate. The cytotoxic activity was impaired by ethylenediamine-tetraacetate (EDTA), phospholipase A inhibitor (Rosenthal's reagent), heating at 50°C for 15 min, or incubation at pH 10 for 60 min. Repeated freeze-thawing and inhibitors of proteolytic enzymes had no effect on the cytotoxic activity. Small amounts of ethanol (5% v/v) enhanced cytotoxic activity of the fraction. Phospholipases A and C, as well as other as yet unidentified cytolytic factors may be responsible for producing 51Cr release from target cells by the soluble fraction of N. fowleri extracts.

Notes: . Restriction endonuclease mapping of the Trypanosoma cruzi kinetoplast DNA maxicircle was performed in nine cloned stocks using maxicircle probes from T. brucei. Analysis of the maxicircle 13–15-kbp encoding region allowed cloned stocks to be divided into three groups: A, B, and C. Parasites from groups A and B had 3% sequence divergence while parasites from group C showed 16–17% sequence divergence with regard to parasites from groups A and B. Cross-hybridization experiments demonstrated that the 23–25-kbp maxicircle divergent region was similar in sequence in group A and B, but different in group C parasites. The high homology of the T. cruzi and T. brucei encoding regions allowed the use of T. brucei probes to detect T. cruzi transcripts, whose overall picture did not differ for parasites from any of the nine assayed clones.

Notes: . Agarose gel electrophoresis was used to identify metabolic enzymes in Babesia bovis and B. bigemina. Glutamate dehydrogenase, lactate dehydrogenase, glucose phosphate isomerase, and hexokinase were identified in B. bovis- and B. bigemina-infected erythrocytes and B. bovis merozoite preparations. A specific electrophoretic mobility was observed for each enzyme. Malate dehydrogenase, glucose 6-phosphate dehydrogenase, 6-phosphogluconate dehydrogenase, and adenylate kinase were only detected in normal erythrocyte preparations. Inter-species, but not intra-species, variation was noted when comparing electrophoretograms of both species. Kinin-activating activity was not detected in B. bovis-infected erythrocyte or merozoite preparations at pH 4.2 or 7.6.

Notes: . In the state of Espirito Santo, Brazil, a cassava disease was recently described, and subsequently a high density of trypanosomatids was revealed in the latex of unhealthy plants. To better characterize this flagellate, Phytomonas françai, we tried to grow it axenically. Successful results were obtained using a biphasic medium containing rabbit blood in the solid phase and a defined medium as overlay.

Notes: . Clones of in vitro-cultured Babesia bigemina-infected erythrocytes were concentrated by several density gradient procedures. The density range of infected erythrocytes containing pairs of parasites was 1.077 to 1.089 g/ml, whereas the density range of infected erythrocytes containing single parasites was 1.092 to 1.100 g/ml. Three enzymes-lactate dehydrogenase, glucose-phosphate isomerase, and glutamate dehydrogenase-were found associated with infected erythrocytes. The parasite-specific enzyme and/or isoenzymes were shown to have different mobility patterns in starch gel electrophoresis from those found in the normal bovine erythrocytes. The enzyme 6-phosphogluconate dehydrogenase was not detected as a parasite-specific enzyme in B. bigemina-infected erythrocytes.

Notes: . Alpha-difluoromethylornithine (DFMO), an irreversible inhibitor of ornithine decarboxylase, has been demonstrated to be an effective agent against a variety of parasitic protozoa but not against Leishmania spp. In this report, we show that Leishmania donovani promastigotes in continuous culture are sensitive to the growth inhibitory and cytotoxic effects of DFMO. Incubation of the promastigotes with DFMO obliterates intracellular putrescine pools and depletes spermidine concentrations, which correlates with the onset of growth inhibition. The effects of DFMO on the growth and the intracellular polyamine pools can be reversed completely by the addition of 10 μM putrescine to the culture medium. These results suggest that the treatment of leishmaniasis may be amenable to chemotherapeutic manipulation by DFMO.

Notes: . Trophozoites of Entamoeba invadens IP-1 can be induced to encyst in simple solutions composed of semipermeable constituents (buffer, salts, or sugars) provided that their osmotic pressure is in the range of 60–160 mosmol/kg. Optimal yield of mature cysts was obtained when the osmotic pressure of the medium was 110 mosmol/kg. Encystation could be obtained in the absence of serum although higher yields were obtained in its presence. No difference in the yield of mature cysts was found when either dialyzed or full serum was used. High yields of encystation were obtained (〉70%) in the presence of 5% serum in solutions of NaCl, KCl, or MgSO4, suggesting that the mechanism of encystation is not induced via sodium or potassium channels. Cysts were obtained in the presence of 72 mM glucose, indicating that depletion of a carbon source is not the only requirement for encystation. A rapid change in the density of the Entamoeba cells was observed upon transfer of trophozoites (density 1.061–1.073 g/ml) from growth medium to the low osmotic pressure encystation solutions. Within the first 2 min their density decreased (to 1.050 g/ml), but it soon increased, reaching within 30 min a density higher than 1.120 g/ml. As the encystation process continued to completion, the density of the cells gradually decreased, the mature cysts reaching a density of 1.049–1.061 g/ml.

Notes: . DL-α-difluoromethylornithine (DFMO), a specific irreversible inhibitor of ornithine decarboxylase (ODC), rapidly depletes cells of intracellular putrescine. When administered to animals and humans, DFMO cures acute infections of trypanosomiasis. In order to determine if the mechanism of drug action is related to initiation of transformation and biochemical alterations subsequent to polyamine depletion, trypanosome morphology and mitochondrial activation were studied in a monomorphic strain of Trypanosoma brucei brucei. Exposure of trypanosomes to DFMO in vivo in infected rodents or in vitro in culture resulted in a depletion of intracellular putrescine and a cessation of cell division without specific cytotoxicity. These events were followed by a transformation of the long slender bloodstream form to a short stumpy form via an intermediate morphology. Putrescine, the product of the ODC reaction, abrogates this effect. When introduced into SDM-79 medium, the intermediate form is capable of further transformation to an “insect” procyclic trypomastigote whereas the long slender form and short stumpy form are not. Short stumpy forms are incapable of binary fission and have lost their infectivity for the vertebrate host. In addition, the mitochondrial marker enzyme, NAD diaphorase, was found only in the short stumpy and intermediate forms. We hypothesize that the short stumpy phenotype may not be a viable stage in the natural transformation of the trypanosome from its mammalian host to the insect vector.

Notes: . The heat-shock method for induction of the macrostomal form of Tetrahymena vorax involves the transfer of cells to reduced nutrient medium and the application of a series of elevated temperature shocks followed by washing the protozoa into inorganic medium. The component of the procedure that had the greatest effect on food vacuoles was the heat shocks. At the end of the heat shocks, cells formed vacuoles at a lower rate than non-heat-shocked cells, but the size of the vacuoles formed was larger and the protozoa contained an increased number of vacuoles and total vacuolar membrane. The rate was further reduced by washing cells into nonnutrient medium. In the absence of the heat shocks, the medium had little effect on the capacity of the cells to form vacuoles although after 7.5 h in inorganic medium, the vacuoles formed were smaller and the protozoa possessed fewer vacuoles and therefore less vacuolar membrane. The amount of membrane required to form the cytopharyngeal pouch of the macrostomal cell type was equivalent to the surface area of food vacuoles present in cells prior to the onset of the heat shocks, but the number and surface area of vacuoles decline between the time of oral resorption and pouch development.

Notes: . The morphology of Blastocrithidia triatomae, when colonizing the rectum of Triatoma infestans, was studied by scanning electron microscopy. The posterior end of the epimastigote's body is tapered and often rolled up. The “straphangers” (cyst stages) are connected with the flagellum by “cytoplasmic bridges”; after “straphanger” detachment these bridges persist as remnants. A comparison of colonization densities of B. triatomae and Trypanosoma cruzi shows that both prefer to colonize the rectal gland, but minor density differences are apparent for B. triatomae in other rectal regions. The colonization density in established infections of B. triatomae is always greater than that of T. cruzi, especially near the midgut exit.

Notes: . Electron microscopy of schizont development in erythrocytic Plasmodium knowlesi has revealed that spheroidal vacuoles 250 nm in diameter with semi-dense contents appear at the periphery of the parasite prior to the budding of merozoites. When treated with non-polar solvents, their contents are completely extracted, and after fixation in tannic-glutaraldehyde they contain regular lamellae with a periodicity of 5.5 nm. Both of these reactions are typical of lipids. Some of these structures are associated with phagosomal vacuoles which may contribute to their lamellae. They disappear at the onset of merozoite formation, but membranous whorls of various sizes continue to be associated with the schizont surface during budding of merozoites. It is suggested that the lipidic vacuoles are a source of preformed lipid which can be utilized rapidly during the generation of merozoites.

Notes: . Thirty-eight of 51 (74.5%) shrew moles collected in Japan were infected with from one to four species of Eimeria and/or Isospora including six of six Dymecodon pilirostris and 32 of 45 (71.1%) Urotrichus talpoides. Four eimerians and two isosporans were identified and all are described as new species. Sporulated oocysts of Eimeria amorphospora n. sp. were subspheroid/ellipsoid, 21.1 × 17.9(18-25 × 16-21) μm. Sporocysts were amorphous, gelatinoid envelopes 20.3 × 7.5 (17–24 × 7–9) μm. Sporozoites were enclosed together within a membrane in each sporocyst. This species was found in 9 of 45 (20%) U. talpoides. Sporulated oocysts of Eimeria gonocilia n. sp. were subspheroid/ellipsoid, 28.8 × 24.4 (25–30 × 21–28) μm; a highly ornate outer oocyst wall gave the appearance of a ciliated ball. Sporocysts ovoid, pointed at both ends, were 17.0 × 9.0 (15–19 × 7–11) μm; this species was found in 4 of 45 (8.9%) U. talpoides. Sporulated oocysts of Eimeria talpoidei n. sp. were asymmetrical ovoid, 20.6 × 13.3 (18–23 × 12–15) μm, with sporocysts lacrimiform, 12.0 × 5.8 (10–14 × 5–7) μm. This species was found in 7 of 45 (15.6%) U. talpoides. Sporulated oocysts of Eimeria honshuensis n. sp. were ellipsoid, 15.5 × 11.4 (13–18 × 10–13) μm, with sporocysts ovoid, 9.1 × 5.2 (8–10 × 4–6) μm. This species was found in 10 of 45 (22.2%) U. talpoides and in 5 of 6 (83.3%) D. pilirostris. Sporulated oocysts of Isospora dymecodi n. sp. were subspheroid/ellipsoid, 15.8 × 12.6 (13–17 × 11–13) μm, with sporocysts ellipsoid, 10.9 × 6.9 (10–13 × 6–8). This species was found in six of six D. pilirostris. Sporulated oocysts of Isospora urotrichi n. sp. were spheroid/subspheroid, 13.4 × 12.4 (11–16 × 9–14) μm, with sporocysts ovoid, 9.2 × 6.3 (8–11 × 5–7) μm. This species was found in 27 of 45 (60%) U. talpoides. Only 14 of 38 (36.8%) infected hosts (one D. pilirostris, 13 U. talpoides) were seen to be naturally infected with only one coccidian species when sampled.

Notes: . An undescribed species of aseptate gregarine, for which the name Pterospora schizosoma is proposed, occurs in the coelom of the maldanid polychaete Axiothella rubrocincta (Johnson). The body of the mature gamont is divided into two cylindrical trunks, which are joined anteriorly. Posteriorly, each trunk bifurcates twice, thereby producing four terminal branches. The length of the entire organism varies from 230–600 μm. The organism actively pumps endoplasm from one trunk to the other, and the nucleus (average = 56.2 × 25.9 μm) moves with the flowing endoplasm. In all four of the previously described species, the gamont's main trunks arise from a substantial body mass whereas in P. schizosoma the trunks make up most of the body mass. Pterospora schizosoma's oocysts possess a distinct outer and inner capsule, which contains eight tightly appressed sporozoites.

Notes: . Since May 1979, 190 rodents in the family Sciuridae, representing three genera and nine species, have been collected in the western United States and northern Mexico and examined for coccidia; 71 (37%) had coccidian oocysts in their feces. These included 2 of 12 (17%) Eutamias canipes; 7 of 12 (58%) E. dorsalis; 18 of 50 (36%) E. merriami; 33 of 96 (34%) E. obscurus; 3 of 4 (75%) E. townsendii; 3 of 9 (33%) Sciurus aberti; 1 of 1 S. griseus; 1 of 1 Tamiasciurus hudsonicus mogollonensis; and 3 of 5 (60%) T. mearnsi. The following coccidians were identified from infected rodents: Eimeria cochisensis n. sp. and Eimeria dorsalis n. sp. from E. canipes; E. cochisensis, E. dorsalis, and E. tamiasciuri from E. dorsalis; E. dorsalis and E. tamiasciuri from E. merriami; E. cochisensis, E. dorsalis, E. tamiasciuri, and E. wisconsinensis from E. obscurus; E. cochisensis and E. dorsalis from E. townsendii; E. ontarioensis and E. tamiasciuri from S. aberti; E. tamiasciuri from S. griseus; E. tamiasciuri and E. toddi from T. h. mogollonensis; and E. tamiasciuri from T. mearnsi. Sporulated oocysts of Eimeria dorsalis n. sp. were ovoid, 21.9 × 16.8 (17–24 × 14–20) μm with sporocysts ovoid, 11.5 × 6.9 (10–14 × 6–8) μm. Sporulated oocysts of Eimeria cochisensis n. sp. were spheroid to subspheroid, 16.7 × 15.3 (15–18 × 14–17) μm, with sporocysts ovoid, 8.4 × 5.6 (6–11 × 4–7) μm. Fifty-five of 71 (77%) infected hosts had oocysts of only one eimerian species in their feces at the time they were examined. One eimerian, E. tamiasciuri, was found in seven of nine host species in three genera. A list is provided of all eimerians (22, including the species described here) that have been described in the literature from Eutamias, Sciurus, and Tamiasciurus spp.

Notes: . Gibbsia archiuli n. g., n. sp. (Eucoccidiorida, Adeleidae) is named for the apicomplexan protozoon described by Gibbs (1952) from the blood cells of the South African garden millipede Archiulus moreleti. Merogony, gamogony, and sporogony all take place in the host's blood cells. The oocysts contain four sporocysts, each with one sporozoite. The microgametes are not flagellated.

Notes: . The life cycle and morphology of a previously undescribed species of Cryptosporidium isolated from commercial broiler chickens is described. The prepatent period for Cryptosporidium baileyi n. sp. was three days post oral inoculation (PI) of oocysts, and the patent period was days 4–24 PI for chickens inoculated at two days of age and days 4–14 for chickens inoculated at one and six months of age. During the first three days PI, most developmental stages of C. baileyi were found in the microvillous region of enterocytes of the ileum and large intestine. By day 4 PI, most parasites occurred in enterocytes of the cloaca and bursa of Fabricius (BF). Mature Type I meronts with eight merozoites first appeared 12 h PI and measured 5.0 × 4.9 μm. Mature Type II meronts with four merozoites and a large granular residuum first appeared 48 h PI and measured 5.1 × 5.1 μm. Type I meronts with eight short merozoites and a large homogeneous residuum first appeared 72 h PI and measured 5.2 × 5.1 μm. Microgamonts (4.0 × 4.0 μm) produced 16 micro-gametes that penetrated into macrogametes (4.7 × 4.7 μm). Macrogametes gave rise to two types of oocysts that sporulated within the host cells. Most were thick-walled oocysts (6.3 × 5.2 μm), the resistant forms that passed unaltered in the feces. Some were thin-walled oocysts whose wall (membrane) readily ruptured upon release from the host cell. Sporozoites from thin-walled oocysts were observed penetrating enterocytes in mucosal smears. The presence of thin-walled, autoinfective oocysts and the recycling of Type I meronts may explain why chickens develop heavy intestinal infections lasting up to 21 days. Oocysts of C. baileyi were inoculated orally into several animals to determine its host specificity. Cryptosporidium baileyi did not produce infections in suckling mice and goats or in two-dayold or two-week-old quail. One of six 10-day-old turkeys had small numbers of asexual stages only in the BF. Four of six one-day-old turkeys developed mild infections only in the BF, and sexual stages of the parasite were observed in only one of the four. All seven one-day-old ducks and seven two-day-old geese developed heavy infections only in the BF with all known developmental stages present.

Notes: . A new species of Eimeria from the Moorish gecko, Tarentola mauritanica, of Minorca, Spain, is described. Oocysts of Eimeria tarentolae are ellipsoidal and measure 17.8 (17.6–18.7) μm by 13.5 (12.9–14.0) μm. Micropyle, oocyst residuum, and polar granule are absent. Each oocyst contains four spherical to slightly subspherical sporocysts 6.8 (6.4–7.0) μm in diameter. A sporocyst residuum is present, while a Stieda body is lacking. Sporulation is completed within 24–32 h at 21 ± 2°C.

Notes: . Naegleria fowleri amebae, but not those of N. australiensis, N. gruberi, or N. lovaniensis, demonstrated enhanced motility when placed in proximity to mammalian cells. Amebae of nonpathogenic species of Naegleria, however, were more motile in cell culture medium than the amebae of N. fowleri. The locomotory response of highly pathogenic mouse-passaged N. fowleri amebae to nerve cells was greater than axenically cultured amebae. The enhanced mobility elicited by whole nerve cells or disrupted nerve cells was not directed migration but chemokinetic. Naegleria fowleri responded to disrupted neuroblastoma cells more vigorously than to disrupted African green monkey kidney (Vero) cells.

Notes: . Immunological relationships between genera and species of microsporidia were examined by immunoblot analysis. Exospore polypeptides from two Nosema spp., three Vairimorpha spp., and two undescribed Vairimorpha-like isolates were analyzed. Gel electrophoresis and immunoblot analysis revealed that a variety of polypeptides, mostly between 15,000 and 90,000 molecular weight, are present on the exospore. The three Vairimorpha spp. are closely related immunologically to each other, but less so to the two undescribed Vairimorpha-like isolates. The two Nosema spp. are immunologically distant from each other and from the Vairimorpha spp. Indirect evidence, however, indicated that many internal spore polypeptides present in both genera are similar. Cross-reactivity between exospore polypeptides from entomophilic microsporidia and antisera to a mammalian microsporidium, Encephalitozoon cuniculi, was very limited. These results indicate that immunoblot analysis of exospore polypeptides may be employed to investigate the interrelatedness of microsporidian species, and that exospore polypeptides of some microsporidia are sufficiently diverse to be of immunodiagnostic value.

Notes: . Drugs that interact with microtubules (colchicine and vinblastine) and microfilaments (cytochalasin B) partially inhibited cell growth and motility of Tritrichomonas foetus. Parasites incubated with these substances became rounded and cell division was blocked. Neither colchicine nor vinblastine disrupted the microtubules that form the peltar-axostylar system. Any one of these drugs interfered with the net negative surface charge of T. foetus as evaluated by determination of the cellular electrophoretic mobility (EPM). The decrease in the EPM of cytochalasin B-treated cells was caused by dimethylsulfoxide, which was used as solvent. Untreated cells as well as cytochalasin B-treated cells showed a uniform distribution of anionic sites on the plasma membrane as seen with cationized ferritin particles. In cells treated with colchicine or vinblastine the anionic sites were distributed in patches. These results are discussed in terms of participation of labile cytoplasmic microtubules and microfilaments in the control of the distribution of anionic sitecontaining macromolecules located on the cell surface of T. foetus.

Notes: . Bloodstream forms of Trypanosoma musculi, cultured in Schneider's drosophila medium at room temperature, multiply and differentiate through a series of developmental stages into infective metacyclic trypomastigotes in 10 days. Oral inoculation with these culture forms into CBA mice produced a parasitemia similar to that produced by intraperitoneal infection with bloodstream forms except for a three-day longer prepatent period. Attempts to induce parasitemia with bloodstream forms given orally were unsuccessful.

Notes: . Critical use of Nomarski DIC optics and a rotocompressor permits basal bodies and kinetodesmal fibers to be visualized in the cortices of living Paramecium tetraurelia and Paramecium sonneborni. The identification of these structures is confirmed by the correspondence of the images obtained by DIC optics of living cells and by brightfield optics of fixed cells stained by the Fernández-Galiano silver technique.Examination of cells carrying cortical inversions (portions of the cortex rotated 180°) shows that inverted regions may be identified and distinguished from normal regions by the orientation of the kinetodesmal fibers of the kinetids (cortical units) within the kineties (ciliary rows). This demonstrates that both the asymmetry and the polarity of each cortical unit may be assessed in the living cell. This technique has useful applications in the study of morphogenesis and patterning in living cells and for the screening of mutations and variants conferring altered cortical phenotypes.

Notes: . Suckling mice were used as a model host to compare the endogenous development of three different isolates of Cryptosporidium: one from a naturally infected calf, one from an immunocompetent human with a short-term diarrheal illness, and one from a patient with acquired immune deficiency syndrome (AIDS) and persistent, life-threatening, gastrointestinal cryptosporidiosis. After oral inoculation of mice with oocysts, no differences were noted among developmental stages of the three isolates in their sites of infection, times of appearance, and duration, morphology, and fine structure. Sporozoites excysted within the lumen of the duodenum and ileum, penetrated into the microvillous region of villous enterocytes, and developed into type I meronts with six or eight merozoites. Type I merozoites penetrated enterocytes and underwent cyclic development as type I meronts or they became type II meronts with four merozoites. Type II merozoites did not exhibit cyclic development but developed directly into sexual forms. Microgamonts produced £16 small, bullet-shaped microgametes, which were observed attaching to and penetrating macrogametes. Approximately 80% of the oocysts observed in enterocytes had a thick, two-layered wall. After sporulating within the parasitophorous vacuole, these thick-walled oocysts passed through the gut unaltered and were the resistant forms that transmitted the infection to a new host. Approximately 20% of the oocysts in enterocytes consisted of four sporozoites and a residuum surrounded only by a single oocyst membrane that ruptured soon after the parasite was released from the host cell. The presence of thin-walled, autoinfective oocysts and recycling of type I meronts may explain why a small oral inoculum can produce an overwhelming infection in a suitable host and why immune deficient persons can have persistent, life-threatening cryptosporidiosis in the absence of repeated oral exposure to thick-walled oocysts.

Notes: . Malarial parasites infecting mammalian hosts are considered to be homolactate fermentors at their asexual intraerythrocytic developmental stage; however, existing ultrastructural and biochemical evidence suggest that their acristate mitochondria could be involved in energy metabolism. In the present study, inhibitors of mitochondrial function including compounds which act on NADH and succinate dehydrogenases, electron transport and mitochondrial ATPase, as well as uncouplers, were found to inhibit the growth and propagation of the human parasite Plasmodium falciparum in in vitro cultures at concentrations that specifically affect mitochondrial functions. Direct measurement of parasite protein and nucleic acid synthesis in synchronized cultures showed that throughout the parasite life cycle both processes were inhibited, the latter process being more sensitive. These results strongly suggest that intraerythrocytic malarial parasites require mitochondrial energy production.

Notes: . Isoenzyme electrophoretic techniques were applied to the characterization of seven Sarcocystis spp. that had been identified by conventional morphological studies. Cystozoites were harvested from macroscopic cysts from sheep, cattle, and mice and from microscopic cysts from sheep, cattle, and goats. Soluble cystozoite extracts were subjected to cellulose acetate gel electrophoresis and characterized at 15 of the 39 enzyme loci examined. Genetic relationships among isolates were examined by simple phenetic clustering. Two different morphological types of macroscopic cysts from sheep, identified as S. gigantea (syn. S. ovifelis) and S. medusiformis, consistently differed at 40% of the loci examined. Such genetic divergence confirms their separate morphotypic classification. Both differed from microscopic cyst isolates from sheep at 87% of the loci examined; however, two different morphotypes of microscopic cysts were found in the sheep sampled (thick-walled and thin-walled cysts). Until sufficient numbers of each type can be isolated and examined separately, both were regarded as belonging to the species S. tenella (syn. S. ovicanis). Macroscopic and microscopic cysts from cattle consistently differed at 80% of the loci thereby supporting their separate classification as S. hirsuta (syn. S. bovifelis) and S. cruzi (syn. S. bovicanis), respectively. Isolates from goats (microscopic cysts identified as S. capracanis) differed from S. tenella and S. cruzi at 20% and 47% of the loci, respectively. All macroscopic cyst isolates from the various host animal species (including S. muris from mice) differed from each other at nearly all loci. Isoenzyme electrophoretic techniques therefore provided genetic evidence supporting the classification of these various Sarcocystis spp. by their morphological characteristics.

Notes: . Plasmids containing inserts of Babesia bovis DNA were prepared and clones suitable for use in the diagnosis of B. bovis infections were isolated. Dot blot hybridization with DNA from these plasmids, which probably contain repetitive sequences, can detect after an overnight exposure 100 pg of B. bovis DNA, which corresponds to the amount of DNA present in 50 μl of 0.01% parasitemic erythrocytes. No detectable cross-hybridization was observed with Babesia microti, Plasmodium falciparum, Plasmodium vivax, Boophilus, or cow DNA. A small amount of cross-hybridization was observed with 10 ng Babesia bigemina DNA. Use of these probes in a hybridization assay may be helpful in the diagnosis of babesiosis in cattle and ticks, in the confirmation of strain identities, and in correlating virulence with particular strains of Babesia.

Notes: . Disc-shaped plaques were found in the primary cyst wall of sarcocysts of Sarcocystis hemionilatrantis in mule deer in Montana. Ultrastructurally, the plaques were 190.5 nm in diameter, 161.6 nm thick and consisted of six distinct layers with microfilaments arising from the innermost layer. These unusual plaques have not been reported previously for any species of Sarcocystis.

Notes: . We have developed an improved procedure for isolating and purifying the metacyclic trypomastigote form of Trypanosoma cruzi from infected Triatoma infestans. The procedure was simple, did not require time-consuming removal of the insect gut, and gave a good recovery of metacyclics. Purification involved centrifugal flotation of the parasites in Percoll followed by diethylaminoethyl cellulose column chromatography. The resulting purified metacyclics exhibited no loss of infectivity when assayed in mice as compared to metacyclics taken directly from the insects.

Notes: . Four littermate, laboratory-reared Palestine vipers, Vipera xanthina palestinae (#149, #150, #151, #152) (Serpentes: Viperidae) were used to determine modes by which Caryospora simplex (Apicomplexa: Eimeriidae) could be transmitted to snakes. Viper #149 was inoculated orally by stomach tube with 5.0 x 104 sporulated oocysts of C. simplex obtained from the feces of an Ottoman viper, V. x. xanthina and began passing unsporulated oocysts of C. simplex 121 days post-inoculation (DPI). Viper #150 was fed five mice that had been inoculated orally £25 days previously with 5.0 x 104 sporulated oocysts of C. simplex and it began passing unsporulated oocysts of C. simplex 33 days after being fed the first two of five mice. Viper #151 was inoculated orally with sporulated oocysts of C. simplex obtained from viper #150 and began passing oocysts 52 DPI. Viper #152 served as an uninoculated control and did not pass oocysts of any species of coccidian. This study demonstrates that snake/snake and mouse/snake transmission of C. simplex readily occurs.