ALBERT

Notes: . Two gametocyte-forming clones, HB-3 and 3D7, were used. Concentrates of late stage parasites were mixed with bloods containing different proportions of young erythrocytes, and the parasitemia and proportion of gametocytes determined after 2, 3 or 4 days of culture. Significantly more gametocytes were formed in light cells than in heavy cells separated from the same normal blood samples. Up to seven times more gametocytes were formed in reticulocyte-rich bloods from patients with sickle cell anemia than in normal control blood.

Notes: Book reviewed in this article: Grimes, G. W. & Aufderheide, K. J. 1991. Cellular Aspects of Pattern Formation: The Problem of Assembly Foissner, W., Blatterer, H., Berger, H. & Kohmann, F. 1991. Taxonomische und okologische Revision der Ciliaten des Saprobiensystems. Band I: Cyrtophorida, Oligotrichida, Hypotrichida, Colpodea. Bryant, C. (ed.) 1991. Metazoan Lde Without Oxygen.

Notes: . Pentrich ciliates attached to small stones from the beds of two streams, one large with hard water, the other small with soft water, were enumerated throughout an annual cycle. Throughout the year, Platycola was the dominant peritrich in both streams, except for a brief period during the spring when Vorticella and Carchesium predominated. Vorticella reached peak levels of 89 ciliates cm2 of stone surface, and up to 102 Platycola per cm2 of stone surface were found. Mean volumes of samples of the main species were calculated, and used to estimate the standing stock biomasses. using a standard value of dry weight per unit volume. Published values of the growth rates of representatives of the main genera were used to estimate production values, which totalled about 6.5 g dry weight of peritrich cytoplasm/m2 of stream bed per annum in the large stream (mean annual density = 8.3 peritrichs/cm2 of stone surface), and 33 g dry weight/m2 of stream bed per annum in the small stream (mean annual density = 47 peritrichs/cm2 of stone surface). Food supply, temperature and predation were the primary factors determining peritrich abundance

Notes: . A method is described for obtaining large numbers of telotrochs from mass cultures of Vorticella convallaria. These free-swimming cells contract slightly along the aboral-oral plane when extracted with Triton X-100, and thus appear more similar in shape to zooids than unextracted telotrochs. Cytoskeletal structures associated with feeding, such as the infudibulum and the cytopharynx, are visible in cytoskeletal preparations of these non-feeding telotrochs and thus appear not to be disassembled during telotroch formation. The telotroch to stalked-zooid transition proceeds rapidly through a set series of morphogenetic stages. After telotroch attachment, the aboral cilia cease beating and are resorbed. Within 7 min the cell is inverted bell-shaped and the zooid begins feeding. Stalk elongation begins about 15–20 min after attachment, lengthening at the rate of 0.5 μ/min for the first hour and more slowly (0.1 μm/min) after that. Interestingly, these developmental stages are essentially the same as those described for the telotroch to zooid transition in the colonial peritrich Zoothamnium. This evolutionary conservation suggests that the precise sequence and timing of these events are critical for their successful completion. Furthermore, the facts that the telotroch to zooid transition occurs very rapidly, that the feeding structures are maintained throughout the transformation, and that basic cytoskeletal architecture is relatively unchanged is consistent with the hypothesis that the transformation occurs through controlled cytoskeletal rearrangements rather than by changes in gene expression.

Notes: . A new foraminiferan species, Rotaliella elatiana n. sp., was isolated in the Gulf of Elat, where it lives in association with a macrophytic green alga, Enteromorpha. The agamont of this tiny new species has a transparent test composed of a bilocular embryonic chamber followed by six to seven trochospirally coiled inflated chambers. The spiral sutures are undulated. The umbilical side has numerous denticules and has radial grooves. The gamont has only one inflated chamber. Rotaliella elatiana has a classical, heterophasic life cycle, with a regular alternation of diploid agamontic phase and haploid gamontic phase. The gamontic phase of the life cycle is exceptionally reduced and the uninucleated gamonts pair immediately after they build their first chamber. A few cases of autogamic reproduction were observed. R. elatiana is a heterocaryotic species; agamonts have one somatic and two to three generative nuclei.

Notes: . In Plasmodium falciparum. the rhoptries involved in the invasion process are a pair of flask-shaped organelles located at the apical tip of invading stages. They, along with the more numerous micronemes and dense granules, constitute the apical complex in Plasmodium and other members of the phylum Apicomplexa. Several proteins of varying molecular weight have been identified in P. falciparum rhoptries. These include the 225-, 140/130/110-, 80/60/40-, RAP-1 80-, AMA-1 80-, QF3 80-, and 55-kDa proteins. Some of these proteins are lost during schizont rupture and release of merozoites. Others such as the 140/130/110-kDa complex are transferred to the erythrocyte membrane during invasion. The ring-infected surface antigen (RESA). a 155-kDa polypeptide located in dense granules also associates with the erythrocyte membrane during invasion. Erythrocyte-binding studies have demonstrated that both the 140/130/110-kDa rhoptry complex and RESA bind to inside-out-vesicles (IOVs) prepared from human erythrocytes. The 140/130/110-kDa complex also binds to erythrocyte membranes prepared by hypotonic lysis. These proteins, however, do not bind to intact human erythrocytes. In a heterologous erythrocyte model, both the 140/130/110-kDa complex and RESA are shown to bind directly to mouse erythrocytes. Other studies have shown that RESA associates with spectrin in the erythrocyte cytoskeleton. We have recently developed a liposome-binding assay to demonstrate the lipophilic binding properties of the P. falciparum rhoptry complex of 140/130/110 kDa. The rhoptry complex binds to liposomes containing neutrally, positively, and negatively charged phospholipids. However, liposomes containing phosphatidylethanolamine compete effectively for rhoptry protein binding to mouse erythrocytes. The rhoptry complex also binds to membrane and inside-out-vesicles prepared from human erythrocytes and erythrocytes from other species. The rhoptry complex associated with the erythrocyte membrane in ring-infected erythrocytes is accessible to cleavage by phospholipase A. Studies are in progress to identify the molecular epitopes on the individual proteins within the complex responsible for lipid interaction in the erythrocyte bilayer and to determine the specificity of the phospholipid interaction using erythrocyte phospholipids.

Notes: . Trypanosoma congolense bloodstream forms preincubated with a high titer of anti-variant surface antigen (VSG)-specific antibody, a low amount of anti-VSG plus complement-active mouse serum (MS), MS alone, and trypsin were cocultivated with mouse peritoneal macrophages in vitro. Immunofluorescence as well as transmission and scanning electron microscopy revealed that upon attachment to the macrophages' surface, trypanosomes opsonized with anti-VSG/MS formed opsonized filopodia, which were rapidly internalized by the phagocytes. Although these cells attached as frequently as anti-VSG or trypsin-pretreated parasites, the rate of phagocytosis of anti-VSG/MS pretreated trypanosomes was reduced significantly. Trypanosomes pretreated with high antibody titers alone were lysed on the surface of the macrophages before phagocytosis was completed. Parasites opsonized with complement alone adhered only occasionally and were rarely phagocytosed. Trypsin-treated trypanosomes, which served as positive control cells, rapidly attached and remained intact until ingulfment by the macrophages was completed. Untreated control parasites did not attach to the macrophages and were not phagocytosed. Cocultivation of macrophages with anti-VSG/MS-opsonized trypanosomes caused internalization of the flagellum by membrane fusion. Filopodia formation by T. congolense is thus correlated with a marked reduction in phagocytosis even in the presence of only a sublytic antibody titer.

Notes: . Species of Tetrahymena, including T. vorax, T. thermophila, T. pyriformis, and T. pigmentosa, were tested for cloning efficiency in proteose peptone and in synthetic nutrient media to which were added hemin, protoporphyrin IX, chlorophyllin, or asolectin, an impure mixture of phospholipids. All species could be cloned with high efficiency in the crude media. In unsupplemented synthetic medium the cloning efficiencies were 0–10%, around 50%, around 50%, and 90–100% for T. thermophila, T. vorax, T. pyriformis, and T. pigmentosa, respectively. The first three were all stimulated to 90–100% by addition of the porphyrin or phospholipid compounds mentioned above. Uroporphyrin III and coproporphyrin I and III had no effect. We suggest that cells unable to form clones suffer from a lack of cellular energy. This situation may be alleviated by our additions: certain porphyrin rings may be built into cytochromes and phospholipids may be used as fuel. Thus, the synthetic media used so far for these ciliates have not been optimal.

Notes: . The first ultrastructural study of the actinosporean genus Triactinomyxon was carried out on Triactinomyxon legeri from the intestinal epithelium of Tubifex tubifex. The developmental cycle starts with bi- and uninucleate cells. We propose that these cells may be an early proliferative phase of the cycle and may unite to give rise to the four-cell stage, initiating pansporoblast formation. Valvogenic cells transform in the long stylus and anchor-like projections of the spore. In the capsulogenic cells, the primordium of the polar capsules originates as a simple, dense, club-shaped structure not observed in other actinosporeans. In all other respects, actinosporean ultrastructure follows more or less similar patterns. Comparison of actinosporean and myxosporean species gives evidence of considerable structural similarity, exemplified in both classes by the occurrence of cell junctions in their multicellular spores, identical polar capsules and their morphogenesis, cell-in-cell condition, pansporoblast formation, and presence of dense bodies (sporoplasmosomes) primarily in the sporoplasm. This unity of patterns speaks in favor of the postulated actinosporean-myxosporean transformation, which warrants further study.

Notes: . Immunoblotting tests involving cytoskeletal protein arrays and fluorescence microscopical examinations of whole cells using monoclonal antibody 424A8 gave substantially different results in three evolutionary subgroups within the genus Tetrahymena. These responses are described and some implications of the evolutionary divergence indicated in this ciliated protozoan are discussed.

Notes: . The infraciliatures of three Chesapeake Bay species of Eutintinnus conforming in lorica morphology to E. angustatus, E. pectinis, and E. tenuis were compared following Protargol silver impregnation. The kinetome of these species had a number of shared attributes including: 1) a right and left field that were of similar length and kinetal spacing; 2) two, rarely three, long dorsal kineties composed of monokinetids; 3) a ventral kinety; and 4) the absence of a true posterior kinety. Nonetheless, the specific organization of infraciliary components varied considerably among species. For example, the organization of the ventral kinety and several kineties along the border of the left field was different in all three species. In addition, the dorsal kineties of E. angustatus differed positionally from those of E. pectinis and E. tenuis. Some divergence in oral infraciliature was also noted, with the arrangement of infundibular oral polykinetids in E. pectinis and E. tenuis being distinct from that of E. angustatus. The amount of variation in the infraciliature of Eutintinnus spp. was much greater than previously noted for congeneric species of tintinnines and supports the separation E. angustatus, E. pectinis, and E. tenuis at the generic or subgeneric level.

Notes: . The cilia of Didinium nasutum are restricted to two girdles encircling the cell. Each row of cilia in both girdles is made up of two to three anterior pairs of kinetosomes followed by several single kinetosomes. Each single kinetosome has two sets of transverse microtubules, an overlapping postciliary microtubular ribbon, and a laterally directed kinetodesmal fiber. The pairs of kinetosomes are homologous to the oral dikinetids of other haptorians: the nonciliated kinetosome of the pair has a transverse microtubular ribbon that extends to line the membrane of the proboscis, a single short postciliary microtubule, and a nematodesma; the ciliated kinetosome has a ribbon of postciliary microtubules and two sets of transverse microtubules. The presence of these characters in Didinium invalidates Leipe & Hausmann's conclusion that the Didiniidae should be removed from the subclass that contains the other haptorians (Leipe, D. D. & Hausumann, K. 1989. Somatic infraciliature of the haptorid ciliate Homalozoon vermiculare (Kinetofragminophora, Gymnostomata) Ditransversalia n. subcl. and phylogenetic implications. J. Protozool., 36:280–289). In light of this, the justification for a subclass Ditransversalia is challenged and shown to be unnecessary.

Notes: . The morphology and morphogenesis of some oligotrichs were investigated using protargol impregnation, silver carbonate impregnation and scanning electron microscopy. The somatic kineties of Strobilidium caudatum form a spiral at the posterior pole. Strobilidiids without such a spiral are transferred to the genus Rimostrombidium. Fourteen new combinations and a nomen novum, Strobilidium kahli, are necessary, Meseres corlissi n. sp. is characterized by eight somatic kineties composed of long cilia which are not fused to “bristles” as they are in Halteria. Strombidium oblongum shows similar characteristics and is thus combined with Meseres. Strombidium rehwaldi n. sp. has an anterior and an equatorial girdle of extrusomes. The morphogenesis of Meseres and Halteria is very similar, i.e. the entire somatic ciliature and the oral primordium originate apokinetally on the cell surface; the parental somatic ciliature is resorbed. In strobilidiids and tintinnids, the oral anlagen develop in a subsurface pouch and the parental somatic kineties, which are not resorbed, elongate by intrakinetal proliferation of basal bodies. In strombidiids, the oral primordium develops in an intracellular sac or tube. These morphogenetic peculiarities and distinct morphologic characters (e.g. arrangement of adoral membranelles) were applied in constructing a phylogenetic system for oligotrichs using hypotrichs as outgroup. This shows that halteriids are more closely related to hypotrichs than they are to other oligotrichs. The Halteriidae are thus raised to ordinal and subclass ranks, Halteriida n. ord., Halteriia n. subcl.

Notes: A method for axenic cultivation of epimastigote and metacyclic forms of Trypanosoma (Duttonella) vivax at 27°C in vitro is described. Iscove's medium was supplemented with specific concentrations of foetal bovine serum, L-proline, L-glutamine, hypoxanthine, adenosine, pyruvate, and 2-mercaptoethanol. Bloodstream form parasites rapidly transformed into epimastigote forms that grew as surface-adherent colonies in plastic culture flasks. Transformation of epimastigotes to metacyclic forms was first observed 9–12 days after initiation of cultures. Percentages of metacyclics varied: East African T. vivax ranged up to 40% and West African T. vivax ranged up to 24%. Subcultures were made at two-week intervals and maintained for several months. Transformation of bloodstream forms to epimastigotes depended on initial attachment to the bottom of culture flasks and the presence of L-proline. The number and maturity of metacyclic forms was influenced by the concentrations of foetal bovine serum, L-proline, L-glutamine, and 2-mercaptoethanol. Trypanosomes from cultures were cryopreserved, revived, and used to re-establish fresh axenic cultures. These results represent a significant advance in cultivation of T. vivax insect forms that should enable studies to be accomplished on metabolism, differentiation, and pharmacology of this parasitic protozoan, free from the influence of extraneous cells.

Notes: . Macrophage-conditioned medium (MøCM) prepared from mouse peritoneal macrophages activated in vivo with bacillus Calmette-Guérin (BCG) or Propionibacterium acnes and triggered with lipopolysaccharide in vitro contained tumoricidal and amoebicidal activity. The murine fibroblast cell line L929 was used as the indicator of tumoricidal activity and Naegleria fowleri amoeba was used to detect amoebicidal activity in MøCM. The protease inhibitor, soybean trypsin inhibitor, decreased tumoricidal activity but had little effect on amoebicidal activity in MøCM. Anti-TNF antiserum inhibited tumoricidal activity in MøCM. The antiserum reduced amoebicidal activity in BCG-activated MøCM but had no effect on amoebicidal activity in P. acnes-activated MøCM. Recombinant TNF, rIL-1, or rIL-1 independently did not affect cytolysis of amoebae. Also, rTNF had no effect on the growth of amoebae. Preparative flat-bed electrofocusing of BCG-activated MøCM yielded fractions that exhibited different amoebicidal and tumoricidal activity profiles. Three domains of activity were analyzed (acidic, neutral, and basic). Anti-TNF antiserum eliminated tumoricidal activity, but not amoebicidal activity, in fractions from the acidic domain. A combination of anti-TNF and anti-IL-1 antisera failed to eliminate amoebicidal activity in fractions from the basic domain. These results indicate that different factors are responsible for macrophage amoebicidal and tumoricidal activity. The amoebicidal factors in MøCM affected cytolysis of several species of amoebae.

Notes: We previously reported that phospholipase increases host cell penetration by Toxoplasma gondii. Here we show that calcium-dependent phospholipase A (PLA) activity is found in the supernatant of sonically disrupted T. gondii. When fractions of disrupted T. gondii were incubated with host cells, the release of fatty acids and lysolipids was detected. Fractions of sonically disrupted T. gondii with PLA activity increased T. gondii host cell penetration in a bioassay. In addition, a protein of approximately 20 kDa was detected by immunoblot of T. gondii antigens with horse antiserum to snake venom, the major antibody of which recognizes PLA2. Incubation of T. gondii with exogenous PLA2 resulted in increased solubility of a rhoptry protein. This protein, which we previously characterized as involved with enhanced parasite invasion of host cells and which is recognized by monoclonal antibody Tg49, was detected in increased amounts in supernatant fractions of extracellular parasites treated with PLA2. Whereas without PLA2 treatment, it is only slightly soluble under physiological conditions. This raises the possibility that PLA may be implicated in the release of rhoptry proteins.

Notes: Riboflavin deficiency inhibits the growth of malaria parasites both in vitro and in vivo in infected animals and humans. Although the precise mechanisms underlying this inhibition are unknown, they may involve enhanced requirements for riboflavin by parasites. To investigate this possibility, the rate of uptake of [14C]riboflavin and the biosynthesis of FMN and FAD from riboflavin were studied in infected (5–8% parasitemia) and uninfected human erythrocytes. All cells were incubated for 0–3 h at 37° C in phosphate buffered saline containing MgCl2, glucose, and [14C]riboflavin (2.5–7.5 μM). At hourly intervals, samples were removed, centrifuged, washed twice with cold buffer, and lysed before counting the radioactivity. The rate of in vitro biosynthesis of FMN and FAD from riboflavin in erythrocytes was measured by ion exchange chromatography and reverse isotope dilution techniques. Results showed that the rate of riboflavin uptake and the biosynthesis of FMN and FAD were enhanced in erythrocytes with parasitemia as compared with results in unparasitized erythrocytes. Riboflavin uptake in erythrocytes was proportional to the extent of parasitemia and especially to percent of schizonts present in erythrocytes. These studies indicate that the requirement for riboflavin may be greater in the parasite than in the host erythrocyte. This increased riboflavin requirement may be due to rapid multiplication, higher metabolic rate, and extreme vulnerability to oxidative stress of malaria parasites compared with that of host erythrocytes. The differential requirement of riboflavin by the host and the malaria parasite may hold important potential for developing new strategies for malaria chemotherapy.

Notes: The complete small subunit rRNA gene sequences of the heterotrich Blepharisma americanum and the colpodid Colpoda inflata were determined to be 1719 and 1786 nucleotides respectively. the phylogeny produced by comparisons with other ciliates indicated that C. inflata is allied more closely with the nassophoreans and oligohymenophoreans than the spirotrichs. This is consistent with the placement of the colpodids in the Class Copodea. Blepharisma americanum was not grouped with the hypotrichs but instead was placed as the earliest branching ciliate. the distinct separation of B. americanum supports the elevation to class status given the heterotrichs based on morphological characters.

Notes: Endosymbiotic methanogenic bacteria of three species of anaerobic ciliates (Plagiopyla frontata, Metopus conforms, and M. palaeformis) were inactivated with the specific methanogen inhibitor 2-bromoethanesulfonic acid. the absence of endosymbiont methanogens reduced growth rate and growth yield by about 30% in P. frontata and M. contortus, while no significant change in fitness was observed in M. palaeformis. In Plagiopyla the growth rate constant is not affected by an artificially increased pH2 neither in normal nor in methanogen-free ciliates. the energetic advantage conferred by endosymbiont methanogens in Plagiopyla and in Metopus contortus probably is due to excretion of organic material from the bacteria at the expense of bacterial reproduction. It is unlikely that the maintenance of a low pH2 within the cells due to H2-consumption by the bacteria is important to the ciliates.

Notes: Feces from a juvenile specimen of the anteater Tamandua tetradactyla from Ponta de Pedras, Marajó, Pará, northern Brazil, contained three different coccidial oocysts: Eimeria tamanduae Lainson, 1968; E. corticulata Lainson & Shaw, 1990; and a third species previously unrecorded and described here as Eimeria marajoensis n. sp. Oocysts of the latter parasite are spherical to subspherical, 13.9 ± 1.5 times 13.4 ± 1.4 (11.1-16.5 times 11.1-16.5) μm, shape index (length/width) 1.0 (1.0-1.2). the oocyst wall is a single, Colorless layer about 0.6-1.0 μm thick with no striations or micropyle. There is no oocyst residuum, but a single, round, oval or irregularly shaped polar granule of about 0.75-2.5 μm is consistently present. the sporocysts are broadly ellipsoidal, 7.1 ± 0.7 ± 5.3 ± 0.6 (6.0-8.8 times 4.0-5.7) μm, shape index 1.3 (1.2-1.5), with a delicate wall bearing a minute stieda body. No sub-stieda body was visible. the sporocyst residuum consists of some 10-20 rounded granules, lying between the two slightly curved sporozoites which measure approximately 6.5 times 2.0 μm. Sporocyst refractile bodies were not discernablc.

Notes: Exoerythrocytic forms of Plasmodium gallinaceum were cultured in vitro using salivary gland sporozoites extracted from experimentally infected Aedes fluviatilis mosquitoes. the host cells were macrophage precursors from chicken bone marrow. At various times after introduction of Sporozoites, the cultures were stained by Giemsa or by immunofluorescence assay (IFA) using anti-sporozoite-specific monoclonal antibodies (MAb). the time to complete parasite development in vitro was 50-70 h. By 70 h, ruptured segmenters and free merozoites were visible within the cells. Inoculation of normal chickens with infected cultures induced parasitemia after a pre-patent period of 10-11 days. In vitro young exoerythrocytic forms, late schizonts that include the matured segmenters, and free merozoites shared common antigens with the sporozoites as revealed by IFA using anti-sporozoite-specific MAbs. Our data indicate that macrophages support development of P. gallinaceum sporozoites and that the circumsporozoite proteins are present until Ac end of the primary exoerythrocytic schizogony.

Notes: . Ditrichomonas honigbergii n. g., n. sp. is a small trichomonad flagellate that has three emergent flagella arising from four basal bodies, a parabasal apparatus (single dictyosome with associated striated flagellar rootlets), a microtubular axostyle, a short undulating membrane, and hydrogenosomes. Cultures of D. honigbergii were isolated from the sediments of a freshwater lake and there is no known metazoan host. Cells form walled cysts with internalized flagella and go through all phases of the life cycle (excystment, binary division, encystment) without any perturbations to the culture medium. Ditrichornonas honigbergii is capable of ingesting and digesting bacteria by phagocytosis. These facts suggest that D. honigbergii may be a free-living inhabitant of oxygen-reduced environments. The structure of D. honigbergii is similar to that of retortamonads and the relationship of trichomonads to other amitochondrial protists is discussed.

Notes: This past-presidential address considers the relationship between small-scale assembly and large-scale patterning in ciliates. Certain model examples of assembly-based patterning are reviewed, followed by the introduction of Williams and Honts' concept of “meta-assembly” as applied to the development of the oral apparatus. The major part of this review then explores two topics: the nature and origin of large-scale circumferential order, and the manner by which the large-scale order of cell directions influences the organization of membranelles of the oral apparatus. In this review, I summarize an existing formal description of large-scale positional order, allude to a more precise abstract theoretical model, and end with a brief discussion of the problem of searching for molecular mechanisms.

Notes: Ellobiophrya conviva clasps tentacles of the bryozoan Bugula neritina with a ring-like structure formed from aboral extensions of its body that taper into two slender arms. The tips of the arms overlap and join to form a unique organelle, the bouton. Each arm contains a massive myoneme that splays out at the bouton. The bouton consists of the cupped tips of the arms and a cavity, which is filled with dense homogeneous material. Long digitations containing longitudinal microtubules at their periphery project from the inner surface of the tip of each arm into the cavity. Deep folds of pellicle with pores opening into their depths line the wall of the cavity. Conventional kinetosomes are not visible in the bouton, but circular or elliptical arrays of microtubules are found at the bases of digitations. The nonfunctional scopula of the adult is in a depression enclosed by pellicular folds. The bouton is distant from the scopula, but its fine structure somewhat resembles it, supporting Chatton and Lwoff's hypothesis that the cinctal arms carry parts of the scopula at their tips. The fine structure of the cinctum supports their suggestion that the cinctal arms are homologous to the spasmonemes of vorticellid peritrichs.

Notes: Twenty monoclonal antibodies were produced against trophozoites of Entamoeba histolytica strains HK-9 and HM-1: IMSS. When reactivity to various enteric protozoa was examined by an indirect fluorescence antibody test, 15 of the monoclonal antibodies were strongly reactive with E. histolytica trophozoites. Species-specific antigens recognized by these monoclonal antibodies were located on the plasma membrane, nucleus, cytoplasm, and cytoskeletal structures of the trophozoites. Two of the remaining five monoclonals reacted strongly with trophozoites of the E. histolytica-like Laredo strain. The determinant antigen was located in the cytoplasm. The three remaining monoclonal antibodies were found to recognize cross-reactive antigens between E. histolytica and E. histolytica-like Laredo, E. hartmanni, E. coli, Dientamoeba fragilis, Giardia lamblia, and Trichomonas hominis. These three antibodies were also reactive with T. vaginalis

Notes: . Variants of a cloned laboratory stock of the trypanosomatid parasite Crithidia luciliae have been distinguished from “parental type” organisms. These variants accumulated spontaneously over time as the protozoan was maintained by continuous passage in a chemically defined medium. Cloned lines of these variants have been isolated by plating on nutrient agar and partially characterized on the basis of their growth characteristics in culture, their colony and cellular morphology as well as their surface protein expression. One cloned line consisted of motile, flagellated forms which, unlike “parental type” organisms, did not adhere to the surface of culture flasks. Another cloned line was composed of non-adherent, nonmotile, amastigote-like forms which were further distinguished from “parental type” cells by virtue of their constitutive expression, in nutrient-replete medium, of high levels of a surface membrane associated 3′-nucleotidase/nuclease (3′-N'ase) activity. Both the motile, flagellated and amastigote-like variants, like the “parental type” organisms, exhibited elevated levels of the 3′-N'ase activity upon exposure to purine starvation conditions. The variants described are of potential importance in elucidating the mechanism of induction of the highly regulated 3′-N'ase activity as well as for understanding the cytoskeletal systems and the surface properties of these protozoa.

Notes: . A marine kinetoplastid flagellate, Cryptobia eilatica n. sp., is described from the gills of cultured gilt-head sea bream Sparus aurata L. and wild black-spot sea bream Diplodus noct (Valenciennes) in the Red Sea. The trophozoite is elongated and lacks a contractile vacuole and undulating membrane. The body averages 13.5 × 4.1 μm, anterior flagellum 9.7 μm, and free portion of recurrent flagellum 15.2 μm. The ultrastructural features of the species exhibit great similarity to various previously studied Cryptobiids. Cryptobia eilatica trophozoites feed on bacteria, show a preference for the branc hial interlamellar crypts, and attach to the host epithelium by means of the recurrent flagellum. Neither penetration into the epithelial cells, nor any direct damage to host tissue was observed. Cryptobia eilatica inhabits a purely marine habitat, but its trophozoite tolerates salinities as low as 10 ppt.

Notes: . Macrostomal cell formation is blocked by the antibiotic cerulenin at levels of 15 μg/ml or higher. Inhibition can be reversed up to 4 h following cerulenin addition by washing and resuspending cells in new, noncerulenin-treated transforming principle. In these latter cases, additional time equal to the time spent in the inhibitor, is needed for cells to reach control values of transformation. Neither the addition of saturated or unsaturated fatty acids, cholesterol added alone or in combination with stearic acid, nor a mixture of lipids extracted from Tetrahymena vorax reversed the cerulenin effect. Radioisotope incorporation data showed while protein synthesis was reduced by the end of 1 h and tetrahymanol synthesis by the end of 2 h, little or no effect of this inhibitor occurred on RNA or fatty acid synthesis during these times. One interpretation of these results is that cerulenin, by preventing first protein synthesis and later tetrahymanol synthesis, interferes with synthesis and formation of membranes required for the microstome to macrostome transition.

Notes: Microfilum lutjani n. g., n. sp. (Microsporida) was found on the gill filaments of Lutjanus fulgens (Teleost) inhabiting the coasts of Senegal. This microsporidium forms xenomas distinguished by the microvilli covering the plasma membrane. At all stages of development individuals have isolated nuclei and are in direct contact with the host cytoplasm. Merogony is binary and sporogony is tetrasporoblastic. the spore (4.75 times 2.60 μm)) is characterized by a manubrium inserted on a laterally offset anchoring disc and extending into a very short, noncoiled polar filament (no longer than 500 nm) in the form of a hook. This type of polar filament has not been described previously in the Microsporida.

Notes: Geosim was identified as the cause of a distinct earthy/grassy odour detected in cultures of a free-living amoeba, Vannella species. Volatile components of cell lysates were isolated and concentrated by the Closed Loop Stripping method. Capillary, gas chromatography/mass spectrometry was used to identify odorous compounds. Bacterial symbionts observed in the cytoplasm of the amoebae may be responsible for production of the geosmin. This appears to be the first report of odorous compounds associated with a free-living protozoan and suggests that in some circumstances, Vannella sp. may contribute to taste and odour problems in drinking water.

Notes: . Tetrahymena thermophila mutants homozygous for the oad mutation become nonmotile when grown at the restrictive temperature, and axonemes isolated from nonmotile mutants lack approximately 90% of their outer dynein arms. Electrophoretic analyses of axonemes isolated from nonmotile mutants (oad axonemes) indicate they contain significantly fewer of the 22 S dynein heavy chains that axonemes isolated from wild-type cells (wild-type axonemes) contain. The 22 S dynein heavy chains that remain in axonemes isolated from nonmotile, oad mutants are assembled into 22 S dynein particles that exhibit wild-type levels of ATPase activity. Two-dimensional gel electrophoresis of oad axonemes show that they are deficient in no proteins other than those proteins thought to be components of 22 S dynein. This report is the first formal proof that outer dynein arms in Tetrahymena cilia are composed of 22 S dynein.

Notes: The literature on discriminant feeding by planktonic protozoans using geometric and nongeometric criteria is reviewed with emphasis on recent studies that indicate phagotrophic protists can use information other than particle size or shape to sort among potential prey. Sufficient data are available for ciliates, aplastidic microflagellates, and phagotrophic dinoflagellates. Numerous representative taxa of all three groups have chemosensory capabilities, either to specific chemicals or to prey exudates, that modify their motility patterns resulting in aggregation or dispersal. Representatives of all three groups also have specific prey preferences. These considerations imply, but do not prove, selectivity in feeding through use of chemical cues. Although prey geometry is clearly a first-order determinant of ingestion through passive mechanical selection, recent studies illustrate that planktonic ciliates and flagellates can use other criteria to discriminate among prey. the evidence clearly implicates use of chemical cues, most likely perceived through contact chemoreception. Filter feeders as well as raptors have such abilities indicating that feeding mechanisms per se do not imply limitations on feeding behavior. Evidence of considerable flexibility and complexity in chemoperceptive feeding suggests that we have only glimpsed the more detailed features of feeding behavior in aquatic protozoans.

Notes: The study of microbial food webs is dominated by field measurements of microbial standing stocks and rate processes and to a lesser extent by laboratory studies. These approaches reflect the concerns of microbial ecologists to assess accurately the capabilities of microorganisms and to compare microbial processes to other ecosystem parameters. These approaches have led to enormous advances in understanding microbial food webs. Reconciling our expanding knowledge with general questions about the significance and representation of microbial food webs in ecosystem studies requires additional approaches including comparative studies and field experiments. Comparative studies, analyses of microbial stocks or rates across a wide range of ecosystems, lead to quantitative models of microbial processes. These models facilitate testing of hypotheses at a very general level, allow the comparison of different stocks or rate processes across a gradient of systems, and detect unusual situations or outlier systems. Field experimental manipulations offer the advantages of working with intact natural communities, of direct evaluation of results with statistical methods, and of testing important qualitative hypotheses. Both comparative and field manipulation studies have led to important advances in the study of microbial food webs and should be expanded.

Notes: . We have used ultrastructural techniques in different malarial species to demonstrate a lysosomal system. First, we have tried to localize acid phosphatase, a typical lysosomal label. Its activity was localized in the endoplasmic reticulum and in endocytic vesicles, and in dense-cored vesicles near the digestive vacuoles, especially in Plasmodium falciparum (FCR3 strain). Then, we have studied the different cellular compartments of the malarial parasite by the zinc iodide-osmium tetroxide technique that heavily contrasted the cellular compartments of the parasite. This experiment led to the observation of a profound rearrangement of the endoplasmic reticulum, especially in P. berghei. A very atypical but functional Golgi apparatus was demonstrated in all the growing stages of the parasite and lysosome-like vesicles were observed, showing a structure very similar to those of the coated vesicles of a true Golgi complex. The presence of these organelles are in favor of the existence of a lysosomal system and of the endogenicity of some enzymes involved hemoglobin degradation.

Notes: . This paper reports on a new phenomenon in the ciliated protists: cytoplasmically determined early sexual maturity. Stock MN1 of the marine hypotrich Euplotes crassus matures immediately after conjugation. We analyzed the respective contribuboas of the nucleus and the cytoplasm to the inheritance of this stable condition. A genetic marker, and new methods in E. crassus for cytoplasmic labeling, production of amicronucleates, and induction of selfing were used. Crosses within and among the early mature (EM) variants and late mature (LM) “wild type” lines were done in ovarious combinations. Descendants of EM conjugants continued to be EM, and descendants of LM continued to be LM, regardless of the different experimental approaches used. The results of the crosses clearly show that the clonally stable, variant EM phenotype is transmitted at conjugation in a non-Mendelian manner through the cytoplasmic lineage. The expression of the trait is independent of the micronuclear genome, but the precise site and nature of the hereditary basis is unknown.

Notes: . Mutant strain d48 and d12 cannot express serotype A. In d48, the A i-antigen gene is present in the micronucleus, but not in the macronucleus. It has recently been shown that d12 contains the A gene in its micronucleus, but its macronucleus lacks the gene. Micronuclear transplantations into enucleated cells were performed to analyze those mutants. Reciprocal transplantation between wild type and d48 confirmed that d48 contains the A gene in the micronucleus and its cytoplasm is defective. Wild type 51 enucleated cells into which were transplanted d12 micronuclei could not express A. Amiccronucleate d12 cells into which were transplanted normal micronuclei from 51 or d48 showed no expression of A. These results show that even if the micronucleus of d12 contains the A gene, it must be abnormal, and its cytoplasm is also defective the same as d48. Genetic analysis showed that heterozygote of d12 and wild type 51 or d48 caused a cure of the cytoplasmic defect of d48 and d12 during the development of macronuclei.

Notes: . The effect of culture age on the rate of oxidation of short-, medium-, and long-chain fatty acids by Leishmania major promastigotes was investigated. Promastigotes from 5-day stationary phase cultures oxidized several saturated fatty acids about 3-to-4-fold faster than cells from late log phase cultures, but [10−14C]oleate was oxidized 9-fold faster. The increase in rate of oxidation was partially reversed within 5 h and almost completely reversed within 30 h after resuspending cells from a 5-day stationary culture in fresh medium. Addition of acetate, leucine, or alanine caused moderate inhibitions of [1-14C]palmitate oxidation, while glycerol had little effect. Glucose, however, was a powerful inhibitor of the oxidation of [1-14C]palmitate and of [1-14C]octanoate. Mannose and fructose were also strong inhibitors of palmitate oxidation, but neither galactose, 2-deoxyglucose or 6-deoxyglucose caused appreciable inhibition. The extent of inhibition by acetate increased with increasing culture age, whereas inhibition by glucose decreased. In addition to demonstrating a reversible rise in β-oxidation capacity with culture age, these data also demonstrate a hitherto unrecognized strong and culture age-dependent inhibition of fatty acid oxidation by glucose.

Notes: . Studies of in vitro interactions between Plasmodium berghei sporozoites and peritoneal macrophages from mice and rats were performed. A videomicroscopic analysis was made of interactions observed by phase-contrast microscopy. Our results showed a diversity of dynamic interactions between sporozoites and macrophages that included no interaction, surface interaction without sporozoite interiorization, active sporozoite penetration, active penetration with subsequent sporozoite escape, macrophage destruction, and the formation of “tethers” or web-like structures by sporozoites that had actively invaded macrophages. Sporozoites are thus clearly capable of actively invading host macrophages and are not restricted to being phagocytosed for interiorization. The formation of “tethers” by the moving sporozoite might function in vivo by anchoring the sporozoite to the cells lining the lumen of the liver sinusoid. Active sporozoite motility appears to be a functional phenomenon involved in sporozoite invasion of host liver cells.

Notes: Book reviewed in this article:Gardiner, C. H., Fayer, R. & Dubey, J. P. 1988. An Atlas of Protozoan Parasites and Animal Tissues.Bryant, C. & Behm, C. 1990. Biochemical Adaptation in Parasites.Dubey, J. P. & Beattie, C. P. 1988. Toxoplasmosis of Animals and Man.Margulis, L., Corliss, J. O., Melkonian, M. & Chapmann, D. J. (ed.) (with 60 contributors). 1990. Handbook of Protoctistia.Esch, G., Bush, A. & Aho, J. (ed.) 1990. Parasite Communities: Patterns and Processes.

Notes: The major manifestations of amoeboid locomotion in Naegleria—cytoplasmic streaming, pseudopod production, cell polarity and focal contact production—require that the actin-based cytoskeleton be extremely dynamic. Whether these features are causally linked is unclear. In an attempt to answer this question we have used the fungal product cytochalasin B (cyt B) to dissect the motility process. This drug can perturb the organisation of actin filaments both in vivo and in vitro. Essentially cyt B acts as a molecule which can cap the barbed ends of actin filaments. Not surprisingly therefore cyt B has an effect on rates of actin polymerization and the dynamic state of actin in the cytoplasm.We have found that cyt B has a profound effect on focal contact production and breakdown. Within minutes of addition of cyt B focal contact production ceases, existing focal contacts are stabilised but cytoplasmic streaming and pseudopod production are not blocked. In conclusion it is now clear that the state of actin required for focal contact production is different from that required for pseudopod extension and cytoplasmic streaming.

Notes: . A cell culture of Paramecium with a precise negative gravitaxis was exposed to 4 times 10-6g during a parabolic flight of a sounding rocket for 6 min. Computer image analysis revealed that without gravity stimulus the individual swimming paths remained straight. In addition, three reactions could be distinguished. For about 30 s, paramecia maintained the swimming direction they had before onset of low gravity. During the next 20 s, an approximate reversal of the swimming direction occurred. This period was followed by the expected random swimming pattern. Similar behavior was observed under the condition of simulated weightlessness on a fast-rotating clinostat. Control experiments on the ground under hyper-gravity on a low-speed centrifuge microscope and on a vibration test facility proved that the observed effects were caused exclusively by the reduction of gravity.

Notes: . The nuclear apparatus of H. vermiculare consists of a single moniliform macronucleus and about 25 micronuclei. the micronuclei are about 3 μm in diameter and characterized by a meshwork of thick condensed chromatin. Mitosis is intranuclear and acentric as in all other ciliates. In metaphase, interpolar and chromosomal microtubules are abundant and the length of the micronuclei increases to about 5 μm. In late anaphase, interzonal microtubules become prominent and the spindle elongates to about 50 μ. In meta- and anaphase, the microtubules of the spindle are attached to the polar vesicles, and in anaphase, chromosomes become attached to it. In contrast to most other eukaryotes, micronuclear mitosis is not strictly bound to cell division in H. vermiculare. While most of the micronuclei divide prior to cytokinesis, others retain their interphasic shape or degenerate. In addition, some micronuclei divide in the interdivision period, i.e. between two successive divisions of the cell and macronucleus. Mating cells of H. vermiculare become joined to each other in the cilia-free region covering the cytostome. In the course of conjugation, the cell membranes and the underlying oral filamentous sheaths of both cells fuse, thus uniting the endoplasm of both cells in the mouth region. Synaptonemal complexes in the meiotic chromosomes are more distinct in H. vermiculare than in most other dilates. the micrographs presented here depict dearly the central filament, transverse elements, and other substructures.

Notes: A chemically defined medium using commercially available α-MEM supplemented with HEPES, L-glutamine, D-glucose, folic acid, D-biotin and adenine supports the luxuriant growth and propagation of Leishmania donovani promastigotes. A peak parasite population of about 7.0 × 107/ml at stationary phase and a population doubling time of 11.4 h for high-subpassage promastigotes were obtained. The medium was suitable for transformation of isolated amastigotes from infected hamster spleen. Promastigotes could be detected by culturing kala-azar patients’bone-marrow aspirate or spleen puncture material in this medium. Four out of six freshly transformed isolates gradually adapted and grew well in this medium. Macroscopic colonies appeared on agar plates prepared with the medium within 16–20 days after inoculation. The cloning efficiency was increased about five-fold by glycerol supplementation.

Notes: Strombidium sulcatum is the type species for the genus Strombidium and has been repeatedly referred to over the last 130 yr. However, there are several taxonomic problems associated with it. We discuss why the original description of S. sulcatum lacks resolution to describe a single species. We conclude that: (1) the description of S. sulcatum sensu Fauré-Fremiet, 1912 be used to diagnose the species; (2) there are ambiguities in several redescriptions of S. sulcatum; and (3) S. sulcatum sensu Lynn et al., 1988 is Strombidium emergens (Leegaard, 1915) Kahl, 1932. From this analysis we present a description for Strombidium inclinatum n. sp. (previously S. sulcatum sensu Fenchel and Jonsson, 1988).

Notes: The microsporidium Nudispora biformis n. g., n. sp., a parasite of a larva of the damsel fly Coenagrion hastulatum in Sweden, is described based on light microscopic and ultrastructural characteristics. Merogonial stages and sporonts are diplokaryotic. Sporogony comprises meiotic and mitotic divisions, and finally eight monokaryotic sporoblasts are released from a lobed plasmodium. Sporophorous vesicles are not formed. The monokaryotic spores are oval, measuring 1.4–1.8 × 2.8–3.4 μm in living condition. The thick spore wall has a layered exospore, with a median double-layer. The polaroplast has two lamellar parts, with the closest packed lamellae anteriorly. The isofilar polar filament is arranged in 6 (to 7) coils in the posterior half of the spore. Laminar and tubular extracellular material of exospore construction is present in the proximity of sporogonial stages. In addition to normal spores teratological spores are produced. The microsporidium is compared to the microsporidia of the Odonata; its possible relations to the genus Pseudothelohania and to the Thelohania-like microsporidia are discussed. The new genus is provisionally included in the family Thelohaniidae.

Notes: The course of malarial infection was compared in pregnant mice inoculated with Plasmodium berghei at different stages of gestation. When 12–14 wk old, pregnant BALB/c mice were inoculated with 1 × 106 of P. berghei NK65-infected red cells at gestation day 0, 2, 4, 6, 8, 10, 12, 14 or 16, the mice inoculated on gestation days 6–12 expired 6.5 days after inoculation compared to 9.5 days in non-pregnant mice. Parasitemia in these pregnant mice increased rapidly on day 4 after inoculation and anemia also developed earlier on day 5. However, the degree of parasitemia and anemia in the terminal stage of infection in these pregnant mice was milder than that of non-pregnant controls. Blood urea nitrogen increased at the terminal stage although the degree of increase in mice inoculated on gestation days 6–10 was comparatively small. Pregnant malarial mice died earlier with less physiological changes than non-pregnant controls. It was concluded that pregnancy makes the host susceptible to physiological changes caused by malaria.

Notes: The pattern of cytospindle assembly and the modifications of the microtubular cytoplasmic network during division of Paramecium are studied by means of indirect immunofluorescence. The assembly of cytospindle starts at two independent areas placed respectively around the proter's and opisthe's buccal overture. The moment of the microtubule bundles’appearance depends on their distance from the buccal opening, with those closest appearing 1st. The existence of microtubule organizing centers that act transiently during division of Paramecium is discussed.

Notes: The fatty acid profiles and contents of protozoa from the rumen fluid of cattle varied according to the type of diet consumed by their host. Changing from a high-quality hay diet to a low-quality hay diet (DA) decreased the proportions of saturated acids and increased the proportions of the unsaturated acids 18:1 cis-9, 18:2 and 18:3 in neutral lipids (NL) and phospholipids (PL). Adding sucrose, urea and sulphur (SUS) to DA increased the proportions of branched chain acids in PL while addition of safflower oil increased polyunsaturated acids in PL and 18:1 trans-11 in NL. Diet did not alter the PL fatty acid content of protozoa but oil supplement of DA resulted in a 10-fold increase in the content of free fatty acids. The defaunating effect of oil supplement was partly reversed by SUS suggesting that factors other than the fatty acid content of cells are important in determining the toxicity of oil to rumen protozoa. The results indicate that the amounts of individual long-chain fatty acids taken up by rumen ciliates are largely determined by their concentrations in rumen digesta.

Notes: In the cell cortex of the parasitic ciliate Ichthyophthirius multifiliis different kinds of cisternae were observed: the alveolar sacs, thick membrane cisternae and the endoplasmic reticulum. The thick membrane cisternae possess coated dilated rims and sometimes could be observed close to the endoplasmic reticulum. Using cytochemical techniques acid phosphatase, thiamine pyrophosphatase and nucleoside diphosphatase activities were detected in the thick membrane cisternae and in the alveolar sacs of trosphozoites. In the endoplasmic reticulum acid phosphatase activity was not detected and only very small amounts of thiamine pyrophosphatase and nucleoside diphosphatase reaction product were observed. After exit from the host, a reduction in acid phosphatase activity was evident in the alveolar sacs. At theront stage acid phosphatase activity is absent from these structures. However, high thiamine pyrophosphatase and nucleoside diphosphatase activities remain in the alveolar sacs during the whole life cycle. On the other hand, acid phosphatase, thiamine pyrophosphatase and nucleoside diphosphatase activities were detected in thick membrane cisternae of theronts. Based on the morphological aspects and enzymatic content the thick membrane cisternae of the cell cortex are designated as golgian-like cisternae. The cytochemical results point out a relationship between the alveolar sacs and the Golgi complex.

Notes: Electron microscopy of salivary glands of the phytophagous hemipteran Phihia picta infected with Phytomonas serpens revealed the presence of flagellates in the gland periphery beneath the gland envelope, in the gland central lumen, between gland cells in the intercellular space and inside the gland cells. In the latter case, flagellates were found in the cytoplasm whether or not it was surrounded by a vacuolar membrane. Flagellates were always of the promastigote type, sometimes displaying a large twisted body. Morphological peculiarities of flagellates in different gland locations are recorded.

Notes: Freeze fracturing of Myxosporidian spores reveals the occurrence of a continuous layer of transmembrane particles all over the surface area of the valve cells which form the spore envelope. These particles are densely packed all over the P face membrane. Due to their polygonal outline, their diameter (6-7 nm) and their central core, they resemble the particles forming the connections of gap junctions which metabolically couple the neighboring cells in animal tissues. In the present report, the role of the transmembrane particles is still hypothetical. However, they might represent a membrane structural specialization of the spores which are submitted to osmotic variations of the fluid external medium. Furthermore similar transmembrane particles are observed at the level of the septate junction which seals the valve cells. In this occurrence, they are arranged in a series of 40 double rows parallel to the suture of the spore envelope. These findings support the view that Myxosporidia are Metazoa and raise the problem of their origin.

Notes: Coleman, A. W., Goff, L. J. & Stein-Taylor, J. R. (ed.). 1989. Algae as Experimental Systems. Plant Biology.

Notes: A rare phenomenon can occur in ciliated protists of the genus Euplotes, which can undergo genetic recombination by the normal outbreeding process of conjugation following mild starvation. Occasionally, the dominant mutation for the autogamy trait arises. Individuals possessing the trait show obligate self-fertilization upon mild starvation. This yields, after normal asexual division, a population of individuals that are reproductively isolated from the parental outbreeding strain. A morphometric analysis of sympatric autogamous and non-autogamous populations of Euplotes vannus from Somalia demonstrates that there has been morphological drift in gross body proportions in the autogamous populations. However, the positional patterns of the locomotory organelles on the ventral surface remain unchanged. The changes in body proportions in the autogamous populations are relevant to the mechanics of the conjugation process, which involves fusion of the oral regions of paired cells belonging to complementary mating types.

Notes: A new species of microsporidium, Nolleria pulicis, is described and named here from the cat flea, Ctenocephalides felis. The genus Nolleria is created and placed within the family Chytridiopsidae. The family is slightly modified to accommodate certain features of intracellular development seen in N. pulicis, which is otherwise very similar to other species in the family Chytridiopsidae. Sporulation is described from ultrastructural analysis of infected midgut epithelial cells of adult C. felis. The term “multiple division by vacuolation” is proposed for describing sporogony as it occurs in this species and certain related species of microsporidia. The probable mode of transmission and apparent absence of merogony are discussed.

Notes: Sphaerospores were found in the kidneys of alevin channel catfish (Ictalurus punctatus) from a farm in Central California. MulticelluUr developmental stages, similar to C-blood protozoans described for Sphaerospora spp. from cyprinid fishes, were observed in circulating blood and numerous tissues. Upon a 2nd examination of the same population offish 10 days later, sporogonic stages were seen developing into mature Sphaerospores in the lumina of the kidney tubules. Sporogeoesis was asynchronous with simple unicellular stages adjacent to more complex forms with developing polar capsules and valves. Only one elliptical spore (5.6 μm in width, 6.5 μm in thickness by 5.8 μm in length) developed within the surrounding pscudoplasmodium. Thin valves surrounded two sporoplasm cells and two subspherical polar capsules (1.7 × 1.9 μm) which contained a polar filament with four to five turns. The blood stages of the Sphaerospora sp. described here are similar to the trophozoites seen in channel catfish with proliferativc gill disease (PGD). Early stages of PGD also observed in the same population of channel catfish containing developmental and sporogonic stages of this newly recognized Sphaerospora sp. may suggest a causal relationship between this new myxosporean and the gill disease.

Notes: Chromatin spreads made from isolated nuclei of the unicellular green alga Chlamydomonas reinhardtii show the beaded fibers typical of eukaryotic polynucleosomes. Micrococcal nuclease digestions confirmed the presence of nucleosomes with a repeat length of 189 base pairs, essentially the same as typical mammalian cells. Basic nuclear proteins extracted from isolated nuclei or chromatin with 1 M calcium chloride and 0.3 M hydrochloric acid are resolved into seven major components by electrophoresis in the presence of sodium dodecyl sulfate (SDS). These seven components were subjected to qualitative peptide mapping with V8 protease on SDS gels for comparison with the major histone components of calf thymus. Finally, the C. reinhardtii basic nuclear proteins were fractionated by reversed phase high performance liquid chromatography and their amino acid composition determined. From these studies, we conclude that C. reinhardtii has a full complement of the five histones with properties very similar to those of both higher animals and higher plants.

Notes: Toxoplasma gondii tachyzoites were quiescent in mouse peritoneal fluid or in K2SO4 buffer at pH 8.2. They became consistently motile when K+ was replaced by other monovalent or divalent cations at a constant pH (pH = 8.2). They also became motile when Cl− was substituted for SO42-. Nitrate or SCN−, can also be substituted for Cl− to a certain extent. Tachyzoites showed independent movement for more than 15 min in KCl, and for about 5 min in the other buffers at pH 8.2 after which they were exhausted and stopped. These tachyzoites could not then be further stimulated to motility by renewal of the suspension buffer. Infection of monolayer cells was demonstrated only with parasites which were motile during inoculation. The highest infectivity was thus obtained either with freshly collected tachyzoites or with those preincubated in K2SO4 buffer for 30 min at 37° C at alkaline pH and thus not yet exhausted for motility. Approximately 34 to 38% of these latter organisms were seen to enter cells when they were inoculated into cultures immediately after being resuspended in MEM for 30 min at 37° C. Conversely, those whose motility had been exhausted by the preincubation in buffers other than K2SO4, pH 8.2 could not enter monolayer cells. Additionally, parasites were unable to enter cells when inoculated into cultures in K2SO4 buffer at alkaline pH; instead they remained quiescent on the surface of the monolayer cells, suggesting that Toxoplasma enters the host cells by active invasion.

Notes: Enterocytozoon was 1st described in 1985, in an AIDS patient with intestinal malabsorption and diarrhea. Since then, additional cases of infection with this organism have been observed, but only in individuals with AIDS and malabsorption. Intestinal tissue biopsies were obtained from a 45-year-old man prior to AIDS diagnosis, again nine months later and then at autopsy two months later. When the biopsies were examined electron microscopically, both sets contained the microsporidian parasite. However, the 2nd intestinal biopsy, when wasting was much more severe, contained infection in almost every small intestinal enterocyte examined. The parasite was actively developing, allowing us to detail its life cycle. The parasite is apansporoblastic, polysporous and has characteristics not previously reported in the Microsporida: (1) an electron lucent inclusion not usually seen in Microsporida is prominent and always present; (2) extremely elongated sausage-shaped nuclei occur in the proliferative phase of parasite development; (3) the polar tube development uniquely involves the production of electron dense discs, yet results in the formation of a typical spore; and (4) polar tube development occurs prior to the final division of the multi-nucleate sporont. On the basis of these characteristics, we are placing this genus in a new family, Enterocytozoonidae, n. fam.

Notes: Gametogenesis in the foraminifer Cribrothalammina alba involves changes in both the gamontic test and cytoplasm. As gametes begin to differentiate and gametic flagella emerge, pores form in a regular array over the gamontic test, constituting the only avenue for gamete release. The spherical, biflagellated gametes average 1.5μ in diameter and are released in rapidly moving swarms along with flagellated “spherical masses” that probably result from incomplete gametic differentiation. Gametogenesis occurs entirely within the test and utilizes the entire cytoplast. After gamete release is complete, the agglutinated test collapses and disaggregates within a fairly short time. Similar modifications of the gamontic test occur during gametogenesis in Ovammina opaca Dahlgren, but are otherwise unknown among monothalamous agglutinated foraminifera at present.

Notes: Methods for inducing selfing, and the relation between selfing and the life cycle of Euplotes woodruffi syngen 3 are reported. Three intercrossing stocks were used in this experiment. Selfing was induced with several treatments as follows: cell-free fluid from the cultures of complementary mating types; intact cells of GI or S phase in the cell cycle; heat-killed cells, and lysed cells of GI-, S-, and D-phase cells which were prepared by freeze-thawing. Stock SJ-27 was used as a parental stock from which Fl clones were originated through selfing. The other two stocks, SJ-8 and SJ-19, were used as testers. The period of immaturity varied from clone to clone. The heterotypic conjugation of clones with cells of stock SJ-8 seems to occur earlier in the life cycle than with cells of stock SJ-19. This result shows that this syngen has an adolescent period in the life cycle. The length of selfing immaturity seems to be different from that of crossing immaturity, and selfing appeared slightly later than crossing with testers. But the clones in which selfing 1st occurred are considered to be in adolescence or maturity, not in senility. Once selfing appeared in any clone, the clone continued to produce selfing pairs till just before clonal death. The viability of selfing and of outcrossing were compared and found not significantly different. Inbreeding depression took place in some of the F2 clones by successive selfing. The viability of F2 clones from young parents was significantly higher than that from old parents (220 to 230 fissions) both in selfing and outcrossing. The total life spans which were studied in three F1 clones were 168 to 264 fissions.

Notes: Dietary riboflavin deficiency is known to diminish malarial parasitemia. In this study, we determined whether imipramine and amitriptyline, drugs which inhibit riboflavin metabolism, have antimalarial efficacy. In addition, we evaluated whether these drugs, like other antimalarial agents, increase the hemolytic response to ferriprotoporphyrin IX (FP). The growth of Plasmodium falciparum (FCR3) in the absence and presence of these drugs (10 to 75 μM) was measured by determining (3H)hypoxanthine uptake by intraerythrocytic parasites for 48 h in RPMI 1640 medium. The uptake of (3H)hypoxanthine was significantly reduced in a dose-dependent manner by both imipramine and amitriptyline. The IC50 values of imipramine and amitriptyline at 48 h were 56 and 45 μM, respectively. Both drugs enhanced hemolysis induced by FP (10 or 20 μM). No hemolysis by these drugs was detected in the absence of FP. It is concluded that the tricyclic antidepressants, imipramine and amitriptyline, possess substantial antimalarial properties.

Notes: Localization of the S-antigen of Plasmodium falciparum isolate FCQ27/PNG, from Papua New Guinea, was studied by post-embedding immunoelectron microscopy using affinity-purified rabbit antibodies raised against the repeat region of the antigen. Labelling was found in the parasitophorous vacuole (PV) space of early to late schizonts and in PV-related vesicles within the erythrocyte cytoplasm of schizont-infected cells. Other subcellular structures within the erythrocyte cytoplasm were not labelled. After breakdown of the PV membrane, label was observed around the merozoites, consistent with mixing of the PV contents and erythrocyte cytoplasm. The antigen was not found in uninfected cells, ring stages, trophozoites or associated with free merozoites. Antibodies to FCQ27/PNG S-antigen did not react with other isolates tested, whereas rabbit antibodies to the Palo Alto/Wellcome S-antigen repeat region reacted with isolates FCR3 and ItG2F6 but not with FCQ27/PNG.

Notes: . The cell volume provided by electronic particle counters may be incorrect. As a particle, or cell, passes the counting device, its volume is calculated as a sphere. The electronically derived, mean cell volume (electronic MCV) of a population of Tetrahymena (prolate spheroid) is smaller than the volume (morphometric MCV) calculated from measured cell length and width. This discrepancy was studied using a Coulter Multisizer particle counter and cell morphometry. The electronic MCV averaged 0.70 of the morphometric MCV (1.00) but changed from 0.72 (fast growth) to 0.63 and 0.76 (slow or no growth) for cells having a mean length/width of 2.05, 2.33, and 1.61, respectively. The measured diameter of latex particles (used for calibration) was identical to that stated, but the diameter of the electronic MCV was larger than the width of the cells which related to wehther the length/width of the cells was above, or below, 2.00. Hence, electron particle counters register primarily the width of a prolate-spheroidal cell, oriented with its long axis in the direction of flow, and uses this value as diameter for the calculated sphere, whereas for more spherical cells, tumbling without any orientation, a mean of the axes is used. Factors for correction of the electronic MCV of Tetrahymena are provided.

Notes: . Three strains of Phytomonas serpens two from tomatoes, Lycopersicon esculentum one from the insect Phtia picta (Hemiptera, Coreidae), were cultivated in a chemically defined medium developed from a defined medium for cultivating insect flagellates. Besides organic growth factors required by other insect trypanosomatids this flagellate requires, serine and inositol. Glutamine stimulates growth, and, surprisingly, does not require heme.

Notes: . Cricetid rodents, Peromyscus truei and P. boylii, were inoculated with sporulated oocysts of Eimeria arizonensis collected from wild P. truei maintained in the lab. In P. truei the prepatent period was 4–5 days, the patent period was 9–11 days, and sporulated oocysts were 21.5 × 25.0 (20–23 × 24–26) μm with sporocysts 7.7 × 12.0 (6–8 × 10–13) pm. In P. boylii the prepatent period was 6–7 days, the patent period was 8–9 days, and sporulated oocysts were 20.1 × 23.2 (18–22 × 21–24) pm with sporocysts 6.8 × 10.0 (5–8 × 9–12) pm. Sporulated oocysts from both host species were used in direct side-by-side comparison of isozyme banding patterns using protein electrophoresis. The parasite has polytypic loci for leucine aminopeptidase (LAP), lactate dehydrogenase (LDH), and 6-phosphogluconate dehydrogenase (6-PGD). In oocysts from P. truei, LAP showed one band with fast migration and LDH and 6-PGD each showed two bands, one with fast and one with slow migration. In oocysts from P. boylii, LAP and LDH each had one band with slow migration and 6-PGD had one band with moderate migration. Oocysts of E. arizonensis collected from P. boylii were used to inoculate P. truei. The prepatent and patent periods, structural measurements, and isozyrne banding patterns of the resultant oocysts were the same as those from P. truei when inoculated with oocysts from P. truei.

Notes: . The morphology and morphogenesis of two species of the genus Lembadion, L. lucens and L. bullinum, are described. In both species, left and right ventral kineties converge behind the mouth forming a postoral suture. Buccal infraciliature is formed by one polykinety and two very close paroral kineties (inner and outer). During stomatogenesis, the new oral structures originate from the paroral kineties. The inner paroral kinety forms the new adoral polykinety and regenerates the outer paroral kinety of the proter, while the paroral kineties of the opisthe originate from the outer paroral kinety of the parental cell. Somatic proliferation starts before the stomatogenesis at the equatorial level of the cell, and extends towards the poles forming an equatorial band. Two large invariant zones, anterior and posterior, remain in the dividing cell. Moreover, the kinetodesmal fibers disappear in the proliferation band during the bipartition (fission) process.

Notes: . Both the lag period and the time required for the filament and sporoplasm to emerge from Nosema algerae spores were prolonged when germination occurred under hyperosmotic conditions. Polyethylene glycol (PEG) and sucrose inhibited germination, first by preventing eversion of the filament, and then at higher concentrations by preventing stimulation. The size of the spore cases decreased by about 21% following germination, indicating an elastic spore wall and turgor pressure in the dormant spores. Increased pressure during germination was indicated by less osmotically-induced shrinkage in stimulated than in dormant spores and by higher concentration of solutes in the homogenates of germinated than ungerminated spores. These results are consistent with the hypothesis of a pressure increase during germination that is caused by an endogenous increase in solute concentration.

Notes: . Miamiensis avidus, a marine scuticociliate, undergoes microstome to macrostome transformation. This process is induced by prey ciliates. the morphogenesis of this process was examined using scanning electron microscopy and light microscopy of quantitative protargol-stained specimens. Several stages are identified and described.

Notes: Amoebae were isolated from a natural thermal water source in Michoacaan, Mexico, in September 1986. Two 500-ml samples were taken from pools with water at 45°C and 46°C and concentrated at 2,000 g for 15 min. The sediment was seeded on nonnutritive agar plates and incubated at 42°C. The isolates were axenized in bactocasitone-serum medium. The identification of the isolates was based on their morphology, total protein and isoenzyme patterns by agarose isoelectric focusing, serology, fine structure, agglutination with Concanavalin A, sensitivity to trimethoprim, capacity to kill mice, and their cytopathic effect in Vero cells. The results showed several morphophysiological, biochemical and serological differences between the isolates and the type strain Aq/9/1/ 45D of Naegleria lovaniensis. These remarkable differences provide sufficient evidence to consider one of the isolates a new subspecies, and the other one a morphological variant of N. l. lovaniensis, which can be differentiated from other Naegleriae by their morphology, biochemistry, serology and physiology. The authors propose the name tarasca for the subspecies and purepecha for the morphological variant.

Notes: The somatic and buccal infraciliature of Lagynus elegans are described, and aspects of its division and conjugation are reported. Its somatic infraciliature is made up of 37–46 meridianal kineties composed of isolated kinetosomes that have thick and long kinetodesmal fibers. In the anterior zone of the cell, the circumoral infraciliature can be observed: it is composed of short, slightly oblique kinetal segments, which are formed of three kinetosomes each. The brosse of this species consists of 3 or 4 groups that possess 4 to 6 ciliated kinetosomes each; these kinetosomes lack kinetodesmal fibers. On the apical pole of the cell, surrounding the oral opening, a crown of nematodesmata is observed; these nematodesmata are connected to each other by a fibrillar structure. Taking into account these features, we propose that this genus be transferred from the order Prostomatida to a new family, Lagynidae, of the order Prorodontida.

Notes: The present study was undertaken to determine whether murine macrophage cell lines exhibited in vitro amoebicidal activity comparable to that elicited by activated murine peritoneal macrophages. Peritoneal macrophages activated in vivo by bacillus Calmette-Guérin or Propionibacterium acnes demonstrated significant cytolysis of Naegleria fowleri amoebae. The macrophage cell line RAW264.7 also effected cytolysis of amoebae, but to a lesser extent than that elicited by activated peritoneal macrophages. However, the macrophage cell lines, J774A.1 and P388D1, did not exhibit amoebicidal activity. Macrophage conditioned medium prepared from RAW264.7 macrophages mediated cytolysis of L929 tumor cells but had no effect on N. fowleri amoebae. In addition, neither recombinant tumor necrosis factor nor recombinant interleukin-1 exhibited amoebicidal activity. Scanning electron microscopy of co-cultures revealed that N. fowler bound to activated peritoneal macrophages and RAW264.7 macrophages. These results suggest that RAW264.7 macrophages treated in vitro with lipopolysaccharide are similar to macrophages activated in vivo in that they effect contact-dependent cytolysis of Naegleria fowleri amoebae. The RAW264.7 macrophages are unlike primary macrophage cultures in that they either do not release soluble amoebicidal factors into the conditioned medium or they release insufficient quantities.

Notes: The natural ecology of a heterosporous microsporidium, Amblyospora connecticus was investigated at three different salt marsh habitats during 1986–1989. The parasite has a well-defined seasonal transmission cycle that occurs regularly each year and intimately involves the primary mosquito host, Aedes cantator, and the intermediate copepod host, Acanthocyclops vernalis. In the spring, the microsporidium is horizontally transmitted from the copepod, where it appears to overwinter, to the mosquito via the ingestion of haploid spores produced in the copepod. Mosquitoes develop a benign infection, and females transmit the microsporidium transovarially to their progeny via infected eggs. Oviposition occurs during the summer and infected eggs hatch synchronously in the fall causing widespread epizootics. Infected larvae die, and the cycle is completed when meiospores are released into the pool and subsequently are eaten by A. vernalis, which reappears in the fall and early winter. Amblyospora connecticus thereby persists by surviving in one of two living hosts throughout most of its life cycle rather than in the extra-corporeal environment. This represents an important survival strategy for A. connecticus as results show the salt marsh habitat to be a relatively unstable environment that is subject to periodic flooding and drying. The adaptive significance of utilizing an intermediate host in the life cycle is discussed as it directly facilitates transmission and enhances survival of the microsporidium.

Notes: Morphogenesis of cell division was investigated in Uronychia transfuga utilizing both light microscopy of living and stained specimens and SEM of preserved specimens. The cortical morphogenetic pattern of Uronychia is similar in several respects to that of the members of the family Euplotidae. These features include: the de novo development of the opisthe oral primordium in a subcortical pouch; the development of frontoventral and transverse cirri for both the proter and opisthe from 5 cirral primordia that form de novo within a single latitudinal developmental zone; and the absence of right marginal cirri. The members of the genus Uronychia also show a number of unique characteristics: development of a proter oral primordium that causes partial replacement of the parental adoral zone of oral polykinetids during development of the proter; a large oral membrane that is divided into a right and left component; large caudal cirri that bend to the left; and dorsal kineties comprised of closely set paired-kinetosome kinetids. When compared to the other euplotid-like ciliates, these unique features support the placement of the genus Uronychia in a separate family, Uronychiidae.

Notes: To identify the surface features of Holospora obtusa during its differentiation from the reproductive short form to the infectious long form, bacteria of four different buoyant densities were isolated by Percoll density gradient centrifugation of homogenates of host cells or isolated macronuclei, and examined with a scanning electron microscope. Bacteria of buoyant density 1.09 g/ml were reproductive short forms as well as cells at various stages in the elongation process including fully elongated ones. Bacteria of buoyant densities 1.11 g/ml and 1.13 g/ml were premature long forms and those of 1.16 g/ml were mature infectious long forms. Bacteria of buoyant density 1.09 g/ml had an entirely rough surface while those of buoyant densities 1.11 g/ml and 1.13 g/ml were smooth and had wale-like stripes on their surface. A small tapered tip was observed at one end of the bacteria of buoyant density 1.13 g/ml. Bacteria of buoyant density 1.16 g/ml had an entirely smooth surface, but one end always showed a rough surface; this locally differentiated surface of the special tip of the infectious long form may be responsible for both the nuclear and species specificities of the infectivity of H. obtusa. These observations indicate that the surface of H. obtusa changes during differentiation and the special tip develops in bacteria of buoyant density 1.13 g/ml.

Notes: The axenically cultured, weakly pathogenic Naegleria fowleri LEE and the highly pathogenic, mouse passaged N. fowleri LEEmp are cytopathic for B103 rat nerve cells in culture. Cytopathogenicity was measured by release of radiolabeled rubidium or radiolabeled chromium from B103 target cells. Cytopathogenicity was time-dependent for up to 18 h and dependent upon amoebae effector to nerve cell target ratios of less than 1:1. Release of51 Cr from B103 cells by either LEE or LEEmp amoebae was enhanced by addition of calcium or magnesium to medium free of these divalent cations but the ion-channel inhibitor, verapamil, or the ionophore A23187 and phorbol myristate acetate did not alter release of 51 Cr from B103 cells cocultured with the amoebae. Cycloheximide or actinomycin D impaired release of 51 Cr from B103 target cells injured by either LEE or LEEmp amoebae. Both strains of amoebae were fractionated by glass bead disruption and high speed centrifugation into membrane and soluble fractions. Each fraction was incubated with either 86Rb or 51 Cr labeled nerve cells. The membrane fraction from LEEmp was more active than the soluble fraction in facilitating rubidium and chromium release. In contrast, the soluble fraction from LEE was more active than the membrane fraction in facilitating rubidium release from radiolabeled target cells. The sequential release of 86Rb and 51 Cr from target cells rather than the simultaneous release of the two isotopes indicates that target cell death is due to the release of ions followed later by the release of large macromolecules. The results indicate that N. fowleri amoebae injure nerve cells by two alternate mechanisms, trogocytosis or contact-dependent lysis.

Notes: This paper describes the cortical anatomy and development of mirror-image doublets of Stylonychia mytilus, analyzed using the protargol technique. The reversed, or “left-handed” (LH) component of these doublets is a mirror image of the normal or “right handed” (RH) component with regard to the arrangement of cortical structures. The mirror-image patterning is imperfect, however, as the individual ciliary structures of the LH component all are of normal internal asymmetry, and the orientation of membranelles is inverted. Certain structures that would be expected to form near the line of symmetry are absent. During cell division and cortical reorganization, ciliary primordia arise and become arranged in a mirror-image pattern that is more perfect than that exhibited by the mature structures. Deviations from a mirror-image pattern appear at late stages when organelle sets differentiate within ciliary primordia: for example, the membranelle set differentiates within the oral primordium of the LH component in a sequence that is an inversion rather than a mirror image of the corresponding sequence of the RH component. This mixed control of oral development by different cortical “informational systems” accounts for some of the characteristic abnormalities of the mature oral structures of the LH component.

Notes: . Scanning electron microscopic observations of feeding plasmodia show three characteristic features: 1) extension of multilobed pseudopodia protruding from the leading edge of the plasmodium as it advances onto the surface of a food particle, 2) confluence of the lobes to form a sheath-like pseudopodium attached to the surface of the food particle, and 3) protrusion of small nodules with thin lamellar projections from the leading edge of the plasmodium. Sections through freeze-dried preparations of the feeding plasmodium exhibit a highly convoluted under surface in contact with loosened starch grains that appear to be released by extracellular digestion. the cytoplasm, viewed by transmission electron microscopy, contains branched, internally penetrating canals (ca. 2 μm wide) enclosing engulfed starch grains. Starch grains in the deeper part of the canals are more electron dense and appear to be digested. Micropseudopodia (70-80 nm dia.), projecting from the surface of the canals, protrude toward and into the ingested starch grains. Digestive marker enzyme (acid phosphatase) activity was detected cytochemically in food particles penetrated by micropseu-dopodia indicating a digestive role for these structures not reported previously.

Notes: . Intestinal microsporidiosis in patients diagnosed with acquired immunodeficiency syndrome (AIDS) and having chronic diarrhea was first reported in 1985 and the associated microsporidian was named Enterocytozoon bieneusi. the intracellular developmental cycle of E. bieneusi in enterocytes has been demonstrated and many cases have been reported worldwide. This report presents the life cycle of a second intestinal microsporidian, associated with the same symptoms, in five AIDS patients. This new microsporidian also infects enterocytes but its pathology and morphology differ from that of E. bieneusi. It involves lamina propria macrophages, fibroblasts, and endothelial cells and can disseminate to infect other parts of the body, e.g. the kidney and gall bladder. the parasite cycle includes development of rounded uninucleate and elongated bi- or tetranucleate cells without the formation of plasmodial stages. Sporogony is similar to the more typical development of microsporidia with sporoblast morphogenesis occurring after the last cell division. the development of cells within chambers of a septate, honeycomb-like, parasite-secreted fibrillar network and surrounded by a parasitophorous vacuole, however, is unique to this microsporidian, justifying the establishment of a new genus and species, Septata intestinalis n. g., n. sp.

Notes: . Metabolism of tryptophan by promastigotes of Leishmania donovani donovani was investigated in cells suspended in a simple buffer solution supplemented with glucose. Metabolites from supernatant and lysed cell pellets were analyzed by capillary gas liquid chromatography and 13C nuclear magnetic resonance spectroscopy, with structural confirmation by gas liquid chromatography-mass spectrometry. Tryptophan does not appear to serve as a carbon energy source for L. d. donovani promastigotes since parasites could survive for only short periods in buffer containing tryptophan without glucose, levels of tricarboxylic acid cycle intermediates remained unchanged in the presence of added tryptophan and label from [13C]tryptophan was not detected in any of the intermediates. Leishmania d. donovani catabolized l-tryptophan via aminotransferase and aromatic lactate dehydrogenase reactions to form one major end product, indole-3-lactic acid. The activity of aromatic lactate dehydrogenase required manganese and was NADH-dependent in these organisms that lack lactate dehydrogenase. Promastigotes taken from the mid-log stage of growth produced higher concentrations of indole-3-lactic acid than those from the stationary stage. Conservation of a similar tryptophan catabolic pathway among four Leishmania species suggests the pathway is physiologically important to the parasites themselves.

Notes: . A ciliate isolated from a pond in Brazil, transformed to a giant form when its food was shifted from a bacterial prey to a ciliate prey. This polymorphism is immediately reversible when the prey ciliates, either Tetrahymena or Colpidium, disappear from the culture medium. By its life cycle, morphology, and ultrastructure, this ciliate belongs to the Class Colpodea. it could belong to the genus Platyophryides Foissner, 1987, except that its micronucleus is not enveloped by the macronuclear membrane. The systematic position of the genus Platyophryides, the validity of the three species in this genus, and the characteristics of the Cyrtolophosidida are discussed.

Notes: . Dramatic and consistent changes of mitochondrial or kinetoplast DNA (kDNA) were observed in certain variants of Leishmania amazonensis (A variants) selected in vitro for arsenite-resistance. This was found initially by comparing different lots of wild-type cells and their respective A variants resistant to 30 μM arsenite. The kDNAs isolated from these two groups had different restriction patterns and hybridized poorly to each other, whereas those from different lots within each of the two groups were identical. Hybridization data showed an overall identity of less than 10−3 between total kDNAs of the two groups. This difference was further examined in three independent series of variants, which were selected from three different clones for resistance to graded concentrations of arsenite (5–30 μM). In all three series, their kDNAs were found to change abruptly in an identical pattern at a late step of the selection process, i.e. A variants resistant to 15 μM or 30 μM arsenite. There was no apparent loss of kDNA in the process. Most of the changes observed appear to involve a shift in either the dominance or the copy number of different minicircle subclasses. Surprisingly, the kDNAs of tunicamycin-resistant variants (T variants) were also found to undergo similar changes. Genetic changes previously described in both A and T variants are limited to their nuclei. Namely, different chromosomal regions are amplified to produce large DNA circles which are responsible for the drug-resistant phenotypes. Interestingly, other arsenite-resistant clones without such chromosomal DNA amplification (A'variants) had kDNA of the wild-type pattern. The profound changes of kDNA observed are unprecedented. We propose the term “transkinetoplastidy” for this phenomenon to distinguish it from dyskinetoplastidy or the loss of kDNA described previously in trypanosomatid protozoa. This phenomenon is discussed with respect to the possible mechanisms of its generation, regulation and relation to the drug-resistant phenotypes.

Notes: . The marine ciliate Parauronema acutum harbors a group of symbiotic bacteria, termed xenosomes, which reside exclusively in the cytoplasm where they grow and divide in a remarkable synchronism with the host. When released into the ambient culture medium by gentle rupture of the protozoans, the symbionts can infect homologous as well as heterologous Parauronema stocks and, oddly, Miamiensis avidus, a distantly related marine ciliate. Xenosomes from certain Parauronema stocks can kill other marine ciliates, particularly those of the genus Uronema. Our principal aims with this host-symbiont system have been to study, at the molecular level, the nature of the interaction of the xenosome with the host, infection and the killer effect. Our most recent investigations have been directed toward establishing the phylogenetic origins of the symbionts using molecular approaches. This paper summarizes our previous work and updates our more recent studies on the association of a bacterial symbiont with its protozoan host. The reader is referred to previous reviews for more details on the subject [4, 5].

Notes: Repeated exposure of trypanosomes in vitro or in vivo to low concentrations of the methylating agent 1,2-bis(methylsulfonyl)-1-methylhydrazine induces a series of moderately synchronous morphological and biochemical changes. Cell division halts and the long-slender bloodstream forms transform to short-stumpy forms via larger intermediate-stage cells which contain approximately double the normal G2 content of DNA. In common with naturally occurring short-stumpy trypanosomes, drug-induced short-stumpy forms do not infect rodents and when transferred to Cunningham's medium, transform to and replicate as procylics. Furthermore, these short-stumpy forms exhibit α-ketoglutarate supported motility and oxygen consumption, acquire the ability to reduce nitroblue tetrazolium (NADH diaphorase positivity) and appear to be in the G1 or G0 stage of the cell cycle based upon DNA content.

Notes: Tomonts and their theront offspring of the hymenostomatid fish parasite Ichthyophthirius multifiliis were exposed to calcium levels from 0 to 0.8 mM Ca2+. The survival and reproductive rates of tomonts in the absence of extracellular calcium were not significantly different from rates of tomonts provided calcium. Theronts that developed in the absence of calcium, however, were not infective for Ictalurus punctatus even when the extracellular magnesium concentration was doubled. Theronts that developed in 0.10 mM Ca2+ were infective (0.77 trophonts/mm2 of pectoral fin) to essentially the same extent as theronts provided 0.33 mM Ca2+. Infectivity of those provided 0.8 mM Ca2+ was 1.79 trophonts/mm2 of fin, similar to that of theront controls. Theronts deprived of extracellular calcium as they developed contained significantly fewer secretory mucocysts than did theronts provided 0.1 to 0.8 mM Ca2+ although no significant differences among groups occurred with respect to abundance of crystalline or differentiating mucocysts. Theronts deprived of extracellular calcium also had swollen or enlarged mitochondria and abnormal crystalline mucocysts.

Notes: Four types of anomalous conjugation were documented in Stylonychia mytilus. Type I pairs were formed between mates of different sizes. These pairs exhibited an abnormal site of fusion in at least one of the mates, and the mates might face each other ventrally throughout conjugation instead of the normal side-by-side position. Type I pairs underwent sexual nuclear development and proceeded with the first cortical reorganization as in normal conjugants. Type II involved pairing at the anterior ends of mates with ventral surfaces facing the same direction. These pairs also underwent sexual nuclear development. Hence, aberrant orientation of the mates, and also ectopic sites of cytoplasmic fusion, if extensive, would permit sexual development. Type III pairs were united ventral-to-ventral with their anterior-left sides at the adoral zone of membranelles, and remained as such throughout conjugation. In these pairs, nuclear and cortical events were typical of the asexual development of physiological reorganization. In Type IV pairs, one mate of the pair possessed a fission furrow and developed two sets of ciliature typical of binary fission, while the other mate might undergo physiological reorganization or binary fission. Type III and Type IV pairs thus reveal the asexual state of early conjugants, which can pursue either one of the two modes of asexual cortical reorganization; these cases reinforce the notion of overlap of asexual and sexual cycles during conjugation of hypotrichs. Spontaneous autogamy was documented for the first time for this genus. The autogamonts proceeded with nuclear development and with the first cortical reorganization. Some probably underwent second and third reorganizations, as in conjugants, but accompanied by abnormalities, particularly in the stages beyond fertilization. Post-autogamous clones were nonviable except for one dubious case.

Notes: Leishmania major promastigotes were washed and resuspended in an iso-osmotic buffer. The rate of oxidation of 14C-labeled substrates was then measured as a function of osmolality. An acute decrease in osmolality (achieved by adding H2O to the cell suspension) caused an increase in the rates of 14CO2 production from [6-14C]glucose and, to a lesser extent, from [1, (3)-14C]glycerol. An acute increase in osmolality (achieved by adding NaCl, KCl, or mannitol) strongly inhibited the rates of 14CO2 production from [1-: 14C]alanine, [1-14C]glutamate, and [1, (3)-14C]glycerol. The rates of 14CO2 formation from [1-14C]laurate, [1-14C]acetate, and [2-14C]glucose (all of which form [1-14C]acetyl CoA prior to oxidation) were also inhibited, but less strongly, by increasing osmolality. These data suggest that with increasing osmolality there is an inhibition of mitochondrial oxidative capacity, which could facilitate the increase in alanine pool size that occurs in response to hyper-osmotic stress. Similarly, an increase in oxidative capacity would help prevent a rebuild up of the alanine pool after its rapid loss to the medium in response to hypo-osmotic stress.

Notes: The developmental stages of a recently described microsporidian from the nucleus of hematopoietic cells of salmonid fish were found to be unique among the Microsporida. All observed stages, including meronts, sporonts, and spores were in direct contact with the host cell nucleus (principally hematopoietic cells) of chinook salmon (Oncorhynchus tshawytscha). There is no par-asitophorous vacuole and sporogony does not involve formation of a pansporoblastic membrane as with other members of the suborder Apansporoblastina. The extrusion apparatus differentiates prior to division of sporogonial plasmodia. The spores are ovoid (1 times 2 μm) and uninucleate, and possess a coiled polar tube with 8-12 turns. Developmental stages of the salmonid microsporidian are similar to those described for Enterocytozoon bieneusi as found in the intestinal mucosa of human AIDS patients. However, the intranuclear development, different cell types, and host infected clearly separate the salmonid and human parasites. Accordingly, the intranuclear parasite of salmonids is given the name Enterocytozoon salmonis n. sp. within the suborder Apansporoblastina.

Notes: . Mirror-image symmetry doublets of the ciliate Stylonychia pustulata were obtained from the progenies of dividing cells in which cell division was inhibited by heat-shocks. In two components consisting of the doublet, the left (cell's) component possessed ciliary organelles arranged in almost the same pattern as in normal singlets, while the right one had surface organelles located in a mirror-image symmetry of those of the left component. In cell division of the doublet, two sets of ciliary primordia that were arranged in a mirror-image symmetry developed synchronously in both components. In about 80% of oral primordia (OP) of the right components, the arrangement of the membranellar bands became abnormal. In some cases, OP of the right component were occasionally separated into two longitudinal halves, each consisting of normal membranelles and inverted membranelles. A set of primordia of the paroral membranelles and fronto-ventro-transverse cirri was rarely derived from the basal bodies of the right half with a band of normal membranelles. As a result, a third component with the ciliary organelles normally arranged emerged on the right side of the original right component. The differentiation of membranelles and segmentation of the primordial streaks into cim proceeded from anterior to posterior. A cytoplasmic bulge with multiple right marginal cirral rows was frequently formed at the right margin of the doublet. The behavior in the separation of third and fourth streaks from a primordial streak of dorsal cirri was not mirror-image symmetrical in each component.

Notes: . Norlevinea n. g. is established for microsporidia in which a uninucleate meront changes into a sporont by secreting a thin, membranous, sporontogcnetic and fragile sporophorous vesicle (pansporoblast membrane) in which four uninucleate sporoblasts are formed. In contrast to the genus Gurleya, the sporoblasts and later the spores are permanently joined into doublets, being laterally cemented by an electron-dense substance structurally identical to and continuous with the exospore layer. The polar filament is of the anisofilar type. The type species is Norlevinea daphniae (Weiser, 1947) n. comb., a parasite of the ovaries of Daphnia longispina occurring in several carp ponds in Czechoslovakia.