ALBERT

Notes: Chemically defined minimal media for the cultivation of high temperature tolerant and pathogenic Naegleria spp. have been developed. A defined minimal medium, identical for N. fowleri and N. lovaniensis, consists of eleven amino acids (arginine, glycine, histidine, isoleucine, leucine, methionine, phenylalanine, proline, threonine, tryptophan, and valine), six vitamins (biotin, folic acid, hemin, pyridoxal, riboflavin, and thiamine), guanosine, glucose, salts, and metals. Three of the four strains of Naegleria fowleri tested (ATCCr̀30100, ATCCr̀30863, and ATCCr̀30896) and two strains of N. lovaniensis (ATCCr̀30467 and ATCCr̀30569) could be cultured beyond ten subcultures on this medium. For N. fowleri ATCCr̀30894 diaminopimelic acid, or lysine, or glutamic acid was also required. Mean generation time was reduced and population density increased for all strains with the introduction of glutamic acid. Glucose could be eliminated from the minimal medium only if glutamic acid was present. Without glucose, mean generation time increased and population density decreased. Diaminopimelic acid could substitute for lysine for ATCCr̀30894, indicating that Naegleria species may synthesize their lysine via the DAP pathway. Naegleria fowleri ATCCr̀30100 could be adapted to grow without serine or glycine in the minimal medium with glutamic acid added, but with mean generation time increased and population density decreased. The strain could be grown in the minimal medium in the absence of metals. For growth of N. australiensis ATCCr̀30958, modification of the medium by increasing metals ten-fold, substituting guanine for guanosine and adding lysine, glutamic acid, and six vitamins (p-aminobenzoic acid, choline chloride, inositol, vitamin B12, nicotinamide, and Ca pantothenate) was required.

Notes: From several surveys of environmental sites, the virulent human pathogen, Naegleria fowleri, was isolated from a pond in Georgia, a sewage treatment plant in Missouri, and from the Potomac and Anacostia rivers near and in Washington, D.C. Widely scattered, sparse populations seemed only a potential threat to human health at the time of sampling. The data support an estimate that the sites sampled contain 10,000 typical, low temperature, bactivorous amoebae for each heat tolerant amoeba able to grow at 45° C. Heat tolerant competitors were much more common than N. fowleri. Naegleria lovaniensis, which is heat tolerant but nonpathogenic, was isolated from and downstream from an open air thermal pollution temperature gradient. Hot piles of composting sewage sludge yielded no amoeboflagellates, many heat tolerant (45–49° C) amoebae, and one thermophilic (52° C) Acanthamoeba. Features of the methods used include two-stage incubation to increase isolation of sparse organisms and distinction of N. fowleri from almost all other amoebae on agar plates. The flagellate-empty habitat hypothesis postulates a general model in which human intervention and/ or natural events remove usual competitors and the ability to transform to a motile flagellate confers an advantage in recolonizing.

Notes: The ultrastructure of the bloodstream form of Cryptobia salmositica in rainbow trout was examined during the acute phase of experimental infection. The arrangement of the major groupings of cytoplasmic microtubules originating near the basal bodies is similar to that in other bodonids. The cytostome is reinforced both by pellicular microtubules and an electron-dense plaque. Certain microtubules associated with the flagellar pocket serve as nucleating sites for pellicular microtubules. A flagellar rootlet, consisting of two parallel fibers which are bound together intermittently by electron-dense plaques, curves posteriorly from the basal body of the recurrent flagellum towards the kinetoplast. The basal body associated plaque on the kinetoplast membranes is duplicated at the same time as the basal bodies. Cytoplasmic microtubules are found in association with the plaque and the outer kinetoplastic membrane. A pulsatile vacuole, described for the first time in a hemoparasitic cryptobiid, lies adjacent to the post-flagellar pit. Smaller, interconnected vesicles of the spongiome are continuous with the pulsatile vacuole. Since a pulsatile vacuole occurs not only in free-living and ectoparasitic cryptobiids but in the hemoparasitic (=trypanoplasm) forms as well, this is no longer a character by which the genus Trypanoplasma may be separated from the genus Cryptobia. Possession of this osmoregulatory complex may allow the organism to survive outside of a host and fulfill a monoxenous life cycle, in addition to the usual heteroxenous cycle involving a leech as vector.

Notes: The life-cycle of the amoeboflagellate Tetramitus rostratus includes amoeboid, cyst, and flagellate stages. The ultrastructure of these three stages is illustrated, with particular emphasis on flagellate morphology. Amoeba morphology is typical of that of limax amoebas. Cysts, forming from trophic amoebas, are enclosed by a wall made up of two layers: ectocyst (ca. 70 nm), and endocyst (200 nm). The wall apparently forms from precursor material present in vesicles in the pre-cyst stage cytoplasm. Flagellate morphology is characterized by a well-defined top-shaped profile, maintained by microtubules under the plasma membrane. The flagellar apparatus or mastigont consists of four flagella, their basal bodies, sheaves of microtubules associated with two of the basal bodies, and several rhizoplasts (periodicity 20 nm). A deep, microtubule-supported, ventral invagination appears to function as a gullet. A small number of mitotic stages observed in amoeboid and flagellate individuals suggests similarity in the division process in both stages: intranuclear mitotic apparatus, nucleolus persisting through mitosis, no centrioles or basal bodies functioning as centrioles, difficulty in resolving chromosomes. The text compares ultrastructures of several amoeboflagellate organisms and evaluates the phylogenetic significance of those features common to different species. On the basis of this study, Tetramitus most closely resembles Naegleria spp.

Notes: The hypothesis is advanced that all freshwater Euplotes species with a 9 type 1 fronto-ventral cirrus pattern (E. patella type) depend upon bacteria-like endosymbionts. Aposymbiotic cells of these species are unable to divide. The hypothesis is based on the investigation of 40 different freshwater Euplotes stocks collected in Germany, France, the USA, and Japan. No symbionts were found in E. crenosus and E. palustris, freshwater species with 10 fronto-ventral cirri, nor in E. muscicola, a representative of the freshwater Euplotes group with a 9 type 2 fronto-ventral cirrus pattern (E. affinis type). Characteristic for the essential endosymbionts are multiple nucleoids, a feature described earlier for omikron, an indispensable symbiont of E. aediculatus. Although the symbionts differ from omikron and among each other in size, shape, and their average number per host, they are believed to be related to omikron. In two stocks a different type of bacterium was found in which no defined nucleoids can be detected. Transfer of this symbiont into aposymbiotic cells, originally carrying omikron, revealed that it can restore the ability to multiply. Similarly, omikron was also able to restore the ability to divide in cells freed of this symbiont. It is assumed that this different type of symbiont is a secondary invader of Euplotes which displaced the original omikron-like endosymbiont. Some of the stocks were found to carry, in addition to omikron-like symbionts, other symbiotic bacteria; E. daidaleos carries in addition an alga. The findings suggest that the freshwater Euplotes species with a 9 type 1 cirrus pattern are closely related to each other and evolved from an ancestor (probably of cirrotype 10) which already was dependent upon endosymbionts of the omikron type. It supports the view that the two subgroups of freshwater Euplotes forms with a cirrotype of 9 have evolved independently from each other from species with 10 fronto-ventral cirri by losing a cirrus at different positions.

Notes: Promastigotes of Leishmania mexicana mexicana attach to mouse macrophages in vitro in the absence of serum by a wheat germ agglutinin-like ligand on the surface of the promastigote that binds to the N-acetyl glucosamine moiety of a receptor on the surface of the macrophage. The binding is temperature dependent, and the macrophage receptor is trypsin, cytochalasin B, and glutaraldehyde sensitive. The promastigote ligand is proteolytic enzyme and glutaraldehyde insensitive. Uptake follows attachment and is assisted or inhibited as for attachment. Treatment of promastigotes with proteolytic enzymes uncovers a receptor for a serum component that binds strongly to a mouse macrophage receptor in vitro. The strain of mice donating the macrophages had little effect upon attachment and uptake except that A strain mouse macrophages attached fewer promastigotes in 10 min than those of outbred mice, but took up as many promastigotes over 90 min as those of outbred mice. Low responder Biozzi mouse macrophages took up more promastigotes than high responder Biozzi mouse macrophages. Normal unheated human, rabbit, and guinea pig sera lysed promastigotes and so inhibited their attachment to macrophages in vitro. Unheated immune serum showed an enhanced inhibition of attachment. Heated normal serum allowed attachment and uptake, while promastigotes treated with heated immune serum showed enhanced attachment to and uptake by macrophages. Treatment of macrophages in vitro with immune serum enhanced their ability to attach promastigotes and to engulf them. Repeated 90-min exposures of a population of promastigotes to uptake by mouse macrophages in vitro did not deplete the population of any sub-population more likely to be taken by macrophages. The first sub-population to be taken up survived better in macrophages over 24 h than subsequently engulfed sub-populations.

Notes: BALB/c mice were hyperimmunized with non-infectious extracts of either Leishmania braziliensis promastigotes or Trypanosoma cruzi epimastigotes. When spleen cells from these mice were fused with P3X63Ag8 plasmacytoma cells, the resultant hybridomas synthesized monoclonal antibodies which displayed specific reactivity by indirect immunofluorescence with distinct subcellular components of the parasites. These studies revealed that antigens associated with the flagellum and with a nongranular component of the cytoplasm would account for much of the serologic cross-reactivity observed between the two species. Conversely, antigens associated with surface and/or cytoplasmic granules and with an intracellular organelle believed to be the kinetoplast appeared to be species-specific.

Notes: The invasion of liver parenchymal cells by sporozoites of Plasmodium berghei Vincke & Lips, 1948, was studied in vivo using transmission electron microscopy. Livers of Brown Norway rats were examined 30 and 60 min after intraportal injection of 15 million sporozoites each. Sporozoites found after incorporation into vacuoles in hepatocytes were often located near a bile canaliculus at the lateral cell surface, surrounded by hepatocyte lysosomal structures; however, degradation of sporozoites caused by lysosomal digestion inside hepatocytes was never observed. Due to the crescent shape of sporozoites, serial sections were necessary to demonstrate the actual process of invasion of the hepatocyte. The hepatocyte's plasmalemma appeared to invaginate due to the sporozoite's action, thereby creating a parasitophorous vacuole. It was suggested that the sporozoite actively penetrated the hepatocyte; however, no visible depletion of rhoptries and micronemes was observed.

Notes: The human pathogenic amoeboflagellate Naegleria fowleri and the nonpathogenic species N. gruberi can be cultivated axenically but usually in different media. Naegleria fowleri 6088 has been adapted to grow in Balamuth H-4 medium, usually used to propagate N. gruberi nB81. and nB81 has been adapted to grow in supplemented Nelson's medium, usually used to propagate N. fowleri. N. gruberi nB81. grown in either medium, enflagellated 135 to 150 min after subculture to non-nutrient amoeba saline, whereas 6088 required 225 min. Naegleria gruberi nB81 grown in either medium was agglutinated by 100 ug concanavalin A/ml, whereas N. fowleri 6088 was not. Naegleria fowleri and N. gruberi grown in Nelson's medium became rounded to a greater extent upon chilling at 5° C and remained rounded longer than Naegleria grown in Balamuth medium. The specificity of the surface antigens was an inherent characteristic of each species and not dependent upon the propagating medium. but Naegleria grown in Nelson's medium was agglutinated more reproducibly and more effectively by antiserum. N. gruberi was somewhat more resistant to acriflavine, actinomycin D, cycloheximide, or tetracycline than N. fowleri, regardless of the culture medium. Naegleria fowleri 6088 grown in Nelson's medium, however, was more resistant to actinomycin D, daunomycin. mithramycin. sulfamethoxazole, or tyrocidine than 6088 grown in Balamuth medium. There are limitations on the validity of comparisons of N. fowleri and N. gruberi based upon cultures grown in different media.

Notes: Fine structural studies of the hydrogenosomes of Tritrichomonas foetus using an improved fixative reveal that they are enclosed by two closely apposed 6 nm membranes, which separate at some regions forming a large intramembranous vacuole where Ca++-binding sites are located. Fixation of the cells in a glutaraldehyde solution containing 5 mM CaCl2 and postfixation in an osmium tetroxide-potassium ferrocyanide solution led to the appearance of a reaction product associated with certain regions of the membrane of the hydrogenosomes and in the cisternae of the endoplasmic reticulum, in the recurrent flagellum, and in the plasma membrane. Treatment of ultrathin sections with EGTA removed the reaction product. These results, in association with others previously described, indicate the existence of several similarities between the hydrogenosomes and the mitochondria.

Notes: . One hundred eighty-eight stocks of Paramecium primaurelia. P. biaurelia, P. tetraurelia. and P. octaurelia were grown axenically and screened for variation in four different esterases and acid phosphatase using starch gel electrophoresis. Major observations: frequency of intraspecies variation for these enzymes is much lower in these four species than in other organisms; hypervariability for two esterases occurs in P. biaurelia both in isolates from worldwide locales and in a restricted locale; clustering of variations occurs in a high proportion of variant stocks in all four species; frequency of intraspecies variation is highest in Central and South America for all four species; and geographical differentiation is lacking between stocks in the same species both for common as well as variant phenotypes despite the cosmopolitan distribution of these species. These results are not correlated with adaptations that favor inbreeding over outbreeding. nor is the possession of bacterial endosymbionts strongly correlated with enzyme variation. When the riequency of intraspecies variation was examined for the aurelia complex of species as a whole for 13 enzymes, mitochondrial DNA, and ribosomal DNA, differences between enzymes in frequency of variation could be seen, ranging from less than 2% for seven enzymes to 12.4% for glucosephosphate isomerase, a value similar to that observed for malic dehydrogenase, mitochondrial DNA, and ribosomal DNA in P. tetraurelia. The percentage of polymorphic enzyme loci in the complex as a whole was found to be much lower than that observed for other organisms. For the species more intensely studied in this paper the level of genetic polymorphism was also much lower, although P. biaurelia showed greater variability for two of the enzymes.

Notes: Precursors of 2-aminobutanoic acid (2-ABA), found in the incubation medium of mixed rumen ciliate protozoa, were examined with washed or starved bacteria-free ciliates. Threonine and methionine strongly stimulated the formation of 2-ABA. Formation of 2-ABA by direct conversion of threonine and dethiomethylation of methionine was confirmed by radiotracer experiments with [U-14C]L-threonine and [carboxyl-14C] and [methyl-14C]L-methionine.

Notes: The effects of irreversible inhibition of protein synthesis by pactamycin in either infective forms of Trypanosoma cruzi or mammalian host cells on cellular invasion by this human pathogen were investigated. Treatment of bloodstream forms of T. cruzi with pactamycin markedly reduced their ability to bind either fibroblast-like cells of monkey origin or myoblasts of rat origin. The number of amastigote forms that could be established intracellularly was also significantly decreased with respect to control values obtained when mock-treated (medium alone) trypomastigotes were incubated with the cells. Pactamycin treatment also reduced the infectivity of T. cruzi trypomastigotes for mice as evidenced by both significantly reduced parasitemia levels and mortality rates when compared with those of control mice infected with mock-treated parasites. Inhibition of protein synthesis in the host cells neither prevented cell infection by untreated trypomastigotes nor altered the percentages of infected cells or the magnitude of the infection in vitro. These results indicate that protein synthesis is a requirement for cell invasion by T. cruzi and that the parasite can establish itself and replicate within cells relying on its own protein synthesis ability.

Notes: One hundred eighty-eight stocks of Paramecium primaurelia, P. biaurelia, P. tetraurelia, and P. octaurelia were grown axenically and tested for five esterases, visualized by starch gel electrophoresis, in a search for variant stocks. The five esterases can be distinguished on the bases of their substrate specificity, sensitivity to an inhibitor, and response to different growth conditions. This paper addresses the nature of the electrophoretic change in mobility of the variant stocks in order that species relationships can be more accurately assessed. Crosses carried out in all four species show that single genes determine the differences in mobility between variant and common subtypes. Extracts of variant stocks that gave similar patterns were run against each other, tested for their sensitivity to the inhibitor, and the pattern was compared to that found in extracts of stocks with variant and common subtypes in other species. The majority of the variants in P. primaurelia, P. tetraurelia, and P. octaurelia show an electrophoretic mobility characteristic of a common subtype, or a variant, in another species. The same proportion of variant subtypes as common subtypes have mobilities similar to esterase subtypes found in other species. Of the four species examined in this paper, P. tetraurelia and P. octaurelia appear to be most closely related on the basis of shared esterase subtypes. In P. biaurelia the mobilities of most of the variants are unique, as are the common esterase subtypes in this species. P. biaurelia stands out as having the greatest number of esterase subtypes, with very few of them homologous to subtypes found in other species. This observation supports the idea of greater diversification of stocks within P. biaurelia than for the other three species.

Notes: Glycogen phosphorylase and synthase activities were detected in the sonic lysate of rumen ciliates of the genus Entodinium. The ciliate phosphorylase had the following properties. The pH optimum was narrow and centered at pH 5.9. The activity was maximum at 30°C; above 40°C a rapid inactivation occurred. The Km value for glucose-1-phosphate (G-1-P) and for glycogen was 15 mM and 0.069% (w/v), respectively. NaF and ethylenediamine tetraacetic acid had no stimulative effect on the enzyme activity, though adenosine 3′,5′-monophosphate and theophylline activated it. NaHSO3 inhibited the enzyme activity at a concentration of 1 mM. The inhibition of glucose was noncompetitive for G-1-P. Glycolytic intermediates and nucleotides had a minor effect on phosphorylase activity. Glycogen synthase existed in two forms, glucose-6-phosphate dependent and independent forms: the proportion of the latter form increased with the decrease of reserve polysaccharide levels in the ciliates. Correlations between glycolytic enzyme activities included phosphorylase and synthase activities and reserve polysaccharide contents in the ciliates were determined, and a possible regulatory mechanism of polysaccharide synthesis and degradation was discussed.

Notes: The structure of the major protein of the pellicular membrane of Leishmania tropica was investigated. This protein is composed of two polypeptides, of ca. 50,000 d molecular weight, that were found to cross-react immunologically with the α and β subunits of pig brain tubulin. The polypeptides and pig brain tubulin subunits were partially digested with S. aureus V8 protease, and the peptides obtained analysed by SDS-polyacrylamide gel electrophoresis. A comparison of the patterns showed that the β subunits of Leishmania and pig tubulin have very similar primary structures, while the α subunits have evolved divergently. These experiments demonstrate that the major polypeptides found in the pellicular membrane of L. tropica are α and β subunits of tubulin. Immuno-electron microscopy indicates that the tubulin is located in the microtubules associated with the pellicular membrane of Leishmania. Arrays of microtubules were prepared by nonionic detergent treatment of the cells and observed by electron microscopy after negative staining. Optical diffraction reveals a 5 nm spacing between protofilaments in the microtubule and a 4 nm axial periodicity corresponding to the tubulin subunits. The pitch of the shallow left-hand three-start helix is 12°. A distance of 47 nm separates each microtubule from the next. These data show that the dimensions and supramolecular organization of the tubulin subunits in the microtubules are identical in the pellicular membrane of L. tropica and in mammalian brain.

Notes: Haemogregarina bigemina is redescribed from the blood of the marine fish Blennius pholis, and stages presumed to be those of the haemogregarine are recorded from the hematophagous praniza larva of the isopod Gnathia maxillaris. At College Rocks, Aberystwyth, Wales, the main study area, a high incidence of infection occurred in B. pholis. No exoerythrocytic stages were observed in these fish, nor was sexual dimorphism of the gametocyte evident. As in an earlier study, ecological evidence favored transmission by G. maxillaris rather than by leeches. Gametocytes, syzygy, oocysts, sporoblasts, and sporozoites were identified in the anterior hindgut of the isopod. The stages observed in G. maxillaris are compared with those of other haemogregarines described from the digestive tract of leeches. Mention is made of an intraleucocytic haemogregarine of another fish, Crenilabrus melops.

Notes: The nomenclature of three genera in the family Haemogregarinidae (Haemogregarina, Karyolysus, and Hepatozoon) has been reviewed and the following new names are introduced to replace homonyms or for previously unnamed species: Haemogregarina carlosi n. nom., in the erythrocytes of the lizard Lacerta ocellata; Haemogregarina tincae n. nom., in the stomach and intestine of the tench Tinca tinca; Hepatozoon insectivorae n. sp., in the leucocytes of the shrews Sorex araneus and Crocidura leucodon; Hepatozoon krampitzi n. sp., in the leucocytes of the vole Microtus oeconomus; Hepatozoon peromysci n. sp., in the leucocytes of the deermice Peromyscus boylii and P. truei gilberti; and Hepatozoon pallida (Pessoa et al., 1971) n. comb., in the erythrocytes of the snake Thamnodynastes pallidus nattereri.

Notes: Transmission and scanning electron microscopy of specimens of Paramecium multimicronucleatum treated with the Rio-Hortega silver-impregnation method as modified by Fernández-Galiano demonstrate that considerable deposition of silver occurs around the kinetosomes, especially at the level of the basal plate and also at the proximal end of the kinetosome. In addition, silver is heavily deposited within the kinetodesmal fibers, in the fibrous matrix that surrounds the postciliary and transverse microtubules, in the connective structures observed between the two kinetosomes of a pair and between the kinetodesmal fiber and the anterior kinetosome, and in the trichocysts. Differences and similarities in sites of deposit when other methods of silver impregnation are employed are discussed and the particular value of the present technique in studies of ciliate systematics and phytogeny is stressed.

Notes: . Various ions and treatments known to alter the availability of free Ca2+ were examined with respect to their effects on the cytopharyngeal pouch, a large prey receptacle found in the potentially carnivorous macrostomal form of Tetrahymena vorax. Addition of Ca2+, Ba2+, or Sr2+ induced the pouch to separate from the region of the cytostome, forming a large empty vacuole. Na+, alone, had no effect, but lowered the concentration of Ca2+ required to produce maximum vacuolar formation in populations of cells. Vacuolar induction was also initiated by the cation ionophore A23187 or by subjecting macrostomal cells to an electric current. In the presence of divalent cation chelators EDTA and EGTA, the cytopharyngeal pouch collapsed and was resorbed. Taken together, these results suggest that Ca2+ plays an important role during phagocytosis in this cell type.

Notes: Oocysts of Eimeria morainensis n. sp. are described from the golden-mantled ground squirrel, Spermophilus lateralis. in Northern Colorado. The oocysts of E. morainensis are double-walled and subspherical, 20.3 × 19.8 (18.7–26.2 × 17.5–21.2) μm; and the sporocysts are ellipsoid, 12.1 × 6.9 (8.7–13.7 × 6.2–8.7) μm. Oocyst residuum and micropyle are absent, but a polar granule is present. Sporocyst residuum and Stieda body are present. Differences in oocyst characteristics provide the basis for recognition of this new species of Eimeria.

Notes: Trichoduboscqia epeori Léger was found to parasitize nymphs of the mayfly Rhithrogena iridina Kolenati in southwest Germany for a new host record. It was studied by light and electron microscopy. The pansporoblast membrane is evaginated at several points, usually four, to produce long needle-like appendages 〉20 μm in length with a resilient inner core superficially resembling collagen, which is thought to maintain their orientation. It is suggested that the pansporoblast appendages may play a role in host infection. The structure and ultrastructure of developmental stages are recorded for the first time. Apart from the pansporoblast appendages, the ultrastructure of T. epeori conforms to the general pattern seen in many other pansporoblastic Microspora. Typically 16 spores are produced per pansporoblast but 32-spore pansporoblasts were also found, and the taxonomic significance of this is discussed.

Notes: Electron microscopy of the tomite of Conidophrys pitelkae confirms that Jankowski was correct in including the pilisuctorians in the Apostomatida. Like other apostome tomites, the tomite of Conidophrys possesses a rosette opening to the exterior, kinetodesmata made up of stacks of individual kinetodesmal fibrils, and canaliculi that are surrounded by dense inclusion bodies and open on the ventral surface. The fine structure of the trophont of Conidophrys, however, is quite unlike that of other apostome trophonts. The elaborate infraciliature of the tomite disappears immediately after it settles and reappears de novo on the trophont just before tomitogenesis. The cyst wall, which completely encloses the trophont and grows with it, attaches the ciliate to a seta on its, host, the shrimp Crangon crangon. The setae on which tomites settle vary greatly in size and shape, but each appears to have at its tip some digitiform cuticular projections that surmount a pore, which opens into the lumen of the seta. The trophont's only direct connection to its host is at the cytostome, a unique structure formed of delicate tubules that pass through the pore into the lumen of the seta. Ingestion is by micropinocytosis, and there are no visible food reserves.

Notes: Les colorations au protargol ainsi que la microscopie électronique á transmission et á balayage permettent de distinguer quatre parties dans I'organisation de Spirochona gemmipara: la collerette formée d'une entonnoir et d'une volute abritant la ciliature, le pseudatrium, et le cytostome; le cou contenant le cytopharynx, le systéme excréteur. et I'appareil cytoproctal; le corps renfle par les noyaux et les vacuoles digestives: et le pseudostyle allonge assurant la fixation au substrat. En majeure partie, le spirochone est limité par une pellicule non ciliée et dépourvue de cils; la pellicule comprend la membrane cellulaire, un épiplasme épais percé de nombreux pores et des triplets de microtubules (MT) sous-pelliculaires. Principalement située dans I'entonnoir, la ciliature somatiquc du spirochone est répartie en deux ensembles inégaux, le champ gauche et le champ droit. Les cinéties sont séparées par des crétes contenant les MT post-ciliaires disposés en une couche verticale; les MT sous-cinétiens sont nombreux, arrangés parallélement á la base des cinétosomes; ceux-lá présentent également une lame dense et des MT transverses, et une fibre cinétodesmale discrete. Un important réseau de faisceaux fibrillaires est disposé orthogonalement par rapport aux cinéties. La base de I'entonnoir est déprimée en une petite cavitéévasée, le pseudatrium; celui-lá conduit à un cytostome ouvert en permanence. Dépourvu de némadesmes, le cytopharynx est un tube cylindrique formé par une dizaine de rideaux microtubulaires; prés du cytostome, chaque rideau porte en plus quelques MT radiaires assimilés aux lamelles Z des Nassulida. Le phagoplasme est riche en tubules complexes et en vésicules de taille moyenne à contenu contrasté. Le champ X, peu organisé, comprend 10–30 cinétosomes situés à gauche du cytostome; il ne correspond certainement pas á la ciliature périorale droite de Chilodochona. Si cette difference se retrouve chez d'autres Chonotriches, il sera nécessaire de séparer taxonomiquement les espéces possédant une ciliature périorale de celles qui en sont dépourvues.〈section xml:id="abs1-2"〉〈title type="main"〉ABSTRACTIn the organization of Spirochona gemmipara, four parts can be demonstrated by protargol staining and also by scanning and transmission electron microscopy: the collar, composed of a funnel and a volute, which shelters the cilia, the pseudatrium, and the cytostome: the neck, which contains the cytopharynx, the excretory system, and the cytoproctal apparatus; the body, enlarged by the nuclei and the digestive vacuoles; and the elongated pseudostyle, which ensures attachment to the substrate. Most of the surface of the spirochone is covered by the pellicula devoid of cilia and alveoli; the pellicula comprises the cell membrane, a thick epiplasm perforated with numerous pores and subpellicular triplets of microtubules (MT). The somatic ciliature of the spirochone is located principally in the funnel and is divided into two unequal parts, the left and right fields. The kineties are separated from one another by ridges, each containing one layer of postciliary MT: numerous subkinetal MT run in a parallel direction under the kinetics; moreover, the kinetosomes show a transverse dense spur and MT, and a modest kinetodesmal fiber. A conspicuous net of fibrillar bundles runs orthogonally to the kineties. The base of the funnel forms a small splayed depression, the pseudatrium; the latter leads to a permanently open cytostome. The cytopharynx is a cylindrical tube devoid of nematodesmata but containing ca. 10 microtubular curtains, each bearing also some radial MT resembling the Z lamellae of the Nassulida. The phagoplasm contains many complex tubules and numerous middle-sized vesicles with an electron-dense content. The X field, which is not well organized and comprises 10–30 kinetosomes, lies on the left of the cytostome; it certainly does not correspond to the right perioral ciliature of Chilodochona. If this disparity is found again in other chonotrichs, it will be necessary to separate taxonomically the species that possess a perioral ciliature from those that do not.

Notes: Cells of the ciliate Tetrahymena pyriformis GL overproduce and accumulate massive quantities of the heme intermediate, protoporphyrin IX. Protoporphyrin is localized intracellularly in discrete membranous compartments. The amount of porphyrin stored in the cell changes dramatically as cells progress through the growth cycle. Porphyrin overproduction is stimulated by δ-aminolevulinic acid, but only during the mid-stationary phase. Overproduction of protoporphyrin IX apparently results from an increase, late in the growth cycle, of activities subsequent to δ-aminolevulinic acid synthetase. Feedback inhibition in the pathway by accumulated protoporphyrin IX does not occur. The presence of Co2+ completely inhibits accumulation of protoporphyrin IX in a manner reversed by δ-aminolevulinic acid. Sn4+ stimulates protoporphyrin IX accumulation in the culture.

Notes: The occurrence of amoebae in the rhizosphere of a beach grass (Panicum sp.) collected at the Hempstead Lake State Park, Long Island, New York, was investigated throughout the growing season of 198 1. Amoebae achieved increased population density in the root system when compared with that in the surrounding bare sand; moreover, numbers of amoebae were higher only during the period of active plant growth and up to flowering. Following flowering, the numbers of amoebae in the root system fell to the level found in bare sand. The species diversity of amoebae in this system was compared with that in the carposphere of the mushroom Laccaria trullisata which appears at the same site but at a different time of year.

Notes: Studies of the life cycle of Myxosoma cerebralis showed that development of infectivity did not occur endogenously but that the spore “aging” process required participation of an aquatic tubificid oligochaete. Data suggestive of such involvement were derived from trials in which spores were “aged” in an array of inert, sterilized, pasteurized, or natural aquatic substrates and from examination of aquatic soils from trout hatcheries in which whirling disease was epizootic. The role of the aquatic oligochaete was confirmed two ways. First, signs of whirling disease developed, and M. cerebralis spores were produced in young rainbow trout (Salmo gairdneri) that had been fed oligochaetes harvested from pond soil taken from two hatcheries where whirling disease was epizootic. Second, when containers of pasteurized soil were populated with four genera of oligochaetes–Aeolosoma, Dero, Stylaria, or Tubifex– from a biological supply house, or with tubificid worms from trout hatcheries free of whirling disease, and then seeded with M. cerebralis spores and “aged” for 4 months, whirling disease occurred only in trout held with Tubifex and with hatchery tubificids.

Notes: During the cellular differentiation induced by starvation of Acanthamoeba castellanii, the expression of a number of genes is regulated. Evidence is reviewed that at least one of these, the precursor ribosomal RNA transcription unit, is regulated at the level of transcription. The structure of the rRNA transcription unit and of the RNA polymerases responsible for transcription in Acanthamoeba are reviewed. Utilizing an in vitro transcription system constructed from these components, preliminary evidence has been obtained that pre-rRNA gene expression is regulated by a modification of RNA polymerase I that affects the enzyme's ability to participate efficiently in the initiation of transcription. These results are reviewed in relation to other known mechanisms of transcriptional regulation in eukaryotes.

Notes: All Stylonychia mytilus-like ciliates which were collected and sent to us during the last 20 years belonged either to S. mytilus or to a new species, S. lemnae, which is described here. The only morphological differences are the shape and the size. Stylonychia mytilus that have been starved for one day average 300 μm in length, and S. lemnae starved for a day are 230 μm long. The occurrence of mating types is described.

Notes: The ultrastructural organization of Nuclearia moebiusi is described. The organism has filose pseudopodia without supportive microtubules; it has mitochondria with flattened cristae, lacks extrusomes, microtubule-organizing centers, and cytoplasmic microtubules. During cell division, microtubules appear only within the nucleus, and reorganization of the nuclear envelope occurs late in the nuclear division cycle. Ultrastructural studies reveal that Nuclearia spp. have a pattern of cellular organization distinct from that of other amoebae. Comparison of ultrastructural features suggests that this organism is only distantly related to other rhizopod amoebae (including other filose amoebae) and to the heliozoa.

Notes: Metacyclic forms of Trypanosoma cruzi isolated from the hindgut of infected insect vectors (Rhodnius prolixus) were found to be immunologically cross-reactive with cultured epimastigote, amastigote, and metacyclic stages of the parasite as well as with bloodstream trypomastigote forms by direct agglutination and indirect immunofluorescence techniques. Sera specific for each of these forms of the parasite systematically yielded maximal antibody titers when measured against the homologous antigen, indicating that antigenic determinants are shared by all of the developmental forms used in this work. Supporting this conclusion were the significant reductions in anti-insect-derived metacyclic antibody titer caused by absorption with any of the other life stages of T. cruzi. These results are relevant to the potential use of laboratory-grown forms of T. cruzi in vaccination against a natural infection with this parasite.

Notes: Core-protected DNA can drive only 60% of the Tetrahymena thermophila macronuclear genome into duplexes in hybridization experiments. This core-protected DNA therefore contains only a subset of the genome complexity. We interpret this to mean that a large fraction, if not all, of the genome is phased with respect to nucleosome placement. Among the sequences present in total DNA and absent from core-protected DNA are most of the sequences containing N6-methyladenine (MeAde) residues, consistent with our previous demonstration that most of these residues lie in linker DNA. We show that these results are not due to artifacts resulting from the small size of the DNA driver, nor are they due to any sequence preferences exhibited by staphylococcal (staph) nuclease. This is the first evidence that nucleosome phasing may be a bulk genome characteristic.

Notes: Book review in this Article Cox, F. E. G., ed. 1982. Modern Parasitology. A Textbook of Parasitology

Notes: Column chromatography with Biogel P2 (molecular exclusion of 1800 daltons) indicates that the transforming principle causing microstomes to become macrostomes is a small molecule. Absorbance tests show that only those fractions with high absorbance at 260 nm have biological activity, indicating that the active principle is a component of nucleic acids. Tests of purines and pyrimidines show that purines are active, with hypoxanthine having the highest activity. The combination of hypoxanthine with uridine shows a synergistic reaction. As these two compounds are the natural catabolic excretory products from nucleic acids in Tetrahymena, the fact that they induce transformation in concentrated, starving cells may be a survival mechanism allowing cannibalism to be induced when nutrients are depleted, thereby allowing the survival of the transformed cells until such time as adequate nutritional conditions are restored.

Notes: RESUME.Après résorption des structures buccales du protomonte, la stomatogenèse ne commence que sur les produits de l'avant-dernière division du tomonte. Elle se poursuit et se complète sur les tomites individualisés de la dernière division. Au niveau des extrémités antérieures des cinéties somatiques intercalaires et des extrémités rompues de cinéties bipolaires, une lère vague de proliferation des cinétosomes produit de courtes lignes, obliques, de cinétosomes isolés. Elles se transforment en lignes d'une double rangèe de cinétosomes, à la suite d'une 2ème vague de multiplication de cinétosomes. Un alignement de ces segments, procédant de gauche à droite et d'avant en arrière, constitue successivement les 3 membranelles longitudinales en doublets (M1 puis M2 et M3) ainsi que la membrane parorale, également en doublet. Une 3ème vague de proliferation cinétosomienne juxtapose une rangée de cinétosomes à droite des doublets (sauf à l'extrémité postérieure de M1 et au niveau de la parorale) transformant les promembranelles en triplets. Cette proliferation cinétosomienne se prolonge par addition d'une 4ème rangée de cinétosomes au trajet médian et postérieur de M2 et peut-ětre à M3 et par juxtapositions successives de nouvelles rangées supplémentaires à l'extrémité postérieure de M2 (flamme). Les cinétosomes des rangées droites de promembranelles portent de larges rideaux de nombreuses fibres postciliaires. Les cinétosomes des autres rangées de M2, au moins, ont également des fibres postciliaires. Entre les cils de promembranelles il n'y a pas de couche alvéolaire, ni d'épiplasme. Une résorption des cinétosomes commence à se manifester par disparition des cinétosomes de la rangée gauche de la parorale dont subsistent les cinétosomes droits porteurs de fibres postciliaires. A vec le raccourcissement de l'aire buccale la résorption s'étend aux cinétosomes postérieurs des 2 rangées droites de M1, et des extrémités antérieures des 2 (ou 3?) rangées droites dc M3. Une invagination de la dépression buccale entraǐne vers la gauche les organelles buccaux et enfonce les cinéties vestibulaires en remontant en avant et à gauche leurs extrémités postérieures tronquées. Il y a régression postéro-antérieure totale des cinétosomes de la parorale. M1 reste constituée au départ de 3 rangées ciliaires; M2 est également formée de 3 rangées ciliaires doublées postérieurement de nombreuses rangées constituant la flamme; M3 n'est finalement constituée que d'une seule rangée ciliaire. Une ultime proliferation cinétosomienne aux extrémités antérieures de cinéties vestibulaires serait peut-ětre à l'origine d'un champ allongé de nouveaux cinétosomes vestibulaires.〈section xml:id="abs1-1"〉〈title type="main"〉ABSTRACTAfter resorption of the buccal structures of the protomont, stomatogenesis begins only in the products of the penultimate division of the tomont. It continues to completion in the individualized tomites of the last division. At the anterior ends of the intercalary somatic kineties and the broken ends of bipolar kineties, a first wave of kinetosome proliferation produces short streak lines of isolated kinetosomes. These develop into lines formed of double rows of kinetosomes following a second wave of kinetosome multiplication. An alignment of these segments, proceeding from left to right and from front to rear, constitutes successively the three longitudinal membranelles in doublets (M1 then M2 and M3), and the paroral membrane, also a doublet. A third wave of kinetosome proliferation juxtaposes a row of kinetosomes to the right of the doublets (except at the posterior end of M1 and at the level of the paroral membrane) to give triplets. This proliferation is extended by addition of a fourth row of kinetosomes on the median and posterior path of M2 and perhaps M3, and by successive juxtaposition of further rows at the posterior end of M2 (flare). The kinetosomes of the right hand rows of promembranelles bear wide ribbons of numerous postciliary fibers. There is no alveolar layer nor epiplasm between the cilia of the promembranelles. Resorption of kinetosomes begins by disappearance of the kinetosomes of the left hand row of the paroral membrane; the right hand kinetosomes carrying postciliary fibers remain. With shortening of the buccal zone, resorption extends to the kinetosomes of the two posterior rows of M1 and the anterior ends of the two (or three?) left hand rows of M3. An invagination of the buccal cavity draws the buccal organelles to the left and pushes in the vestibular kineties while raising forward and to the left their truncated posterior ends. Total postero-anterior regression of the kinetosomes of the paroral membrane occurs. Membranelle 1 remains composed of three rows of cilia; M2 is also composed of three rows of cilia edged posteriorly by numerous rows constituting the flare; M3 is composed of a single row of cilia. A final kinetosome proliferation at the anterior ends of the vestibular kineties might be responsible for the extended field of new vestibular kinetosomes.

Notes: . A new procedure is described that utilizes Percoll gradients for purifying micronuclei (MIC) and macronuclei (MAC) from Tetrahymena thermophila. Separation of MIC from MAC during certrifugation in Percoll gradients occurs as a result of their difference in size rather than density. Three kinds of tests were used to evaluate the purity of the nuclei: visualization of the nuclei by light microscopy; examination of the nuclei by electron microscopy; and Southern blots of MIC and MAC DNA probed with the 5s rRNA genes or a fragment from the MAC extrachromosomal rDNA molecule. When examined under the light microscope, the isolated MIC and MAC have much lower nuclear cross contamination levels than previous methods have reported. MIC's contaminated with less than 1 MAC in 1000 MIC and MAC's contaminated with less than 1 MIC in 500 MAC can be routinely prepared. Quantitative analyses of electron micrographs gave higher estimates of cross contamination in our purified nuclei, which may, in part, be explained by the difficulty in identifying small MIC or MAC fragments. Southern blots of MIC and MAC DNA probed with 5s rDNA confirmed the level of MAC contamination in the MIC estimated by light microscopy during purification of the nuclei. The level of nucleolar contamination in the MIC was estimated at 10% by Southern blots of MIC and MAC DNA, derived from a heterokaryon with distinctive MIC and MAC Bam HI sites, using an rDNA probe.

Notes: . Polyurethane substrates were anchored near the surface of 14 lakes in the northern tip of the lower peninsula of Michigan. I wo substrates were removed from each take and taken to the laboratory after 1, 3, 6, 15, and 21 days of exposure. At the laboratory, one substrate was used for determining the number of species of diatoms and the other for protozoa. A cluster analysis of the matrix of Jaccard's coefficients for all diatom samples from all lakes showed that virtually all samples from any given lake consistently clustered together. This indicates that, with respect to species occurrence, distinct and compositionally stable diatom assemblages formed on the substrates in fewer than 21 days. Analysis of all protozoan samples from all lakes did not show such clustering, however, and the correspondence of clusters for protozoan and diatom communities for the 14 lakes was not particularly good. This suggests that the link between the two groups at the species level is not particularly strong during the early phases of artificial substrate colonization.

Notes: . Herpetomonas megaseliae, Crithidia fasciculata, and Leptomonas collosoma from culture survived gut passage in Anolis carolinensis following their ingestion by this lizard. Maximum persistence of H. megaseliae in lizards, as detected by fecal culture, was seven days. No invasion of tissues by H. megaseliae could be detected by means of sectioned material, stained impression slides, or cultures inoculated with material from organs. Crithidia fasciculata was evident in cloacal fluid for up to three days in wet mount preparations. Leptomonas collosoma was observed in feces 24 h after the organisms were fed to lizards. Both C. fasciculata and L. collosoma were cultured from feces of lizards fed the parasites 24 h earlier. Herpetomonas megaseliae was differentiated in lizard feces, with greater than 40% of the forms observed being paramastigotes or opisthomastigotes. Truncate, semispherical forms resembling choanomastigotes were seen, but the kinetoplast was posterior to the nucleus in some of these. Many forms showed extensive coiling of the axoneme within the body of the flagellate. Choanomastigotes and spheromastigotes of C. fasciculata and promastigotes, sphero-mastigotes and amastigotes of L. collosoma were also observed in the feces.

Notes: . To answer whether Blepharisma hyalinum is truly unpigmented, the organism must be established in culture as pointed out by Giese in 1973. Accordingly, the present study deals with B. hyalinum kept in culture since its isolation in 1975. The organism still remains colorless after growth in the dark; however, it contains cortical granules resembling pigment granules in colored species. A comparative study was therefore undertaken of B. hyalinum and B. steini; both species have a compact macronucleus, though of different shape. Crude pigment was extracted with acetone from organisms grown in the dark for three weeks and the maxima were measured by absorption. Purified pigment was obtained from TLC-plate preparations and the absorption maxima were measured after removal of lipids with chloroform. No maxima characteristic of blepharismin were found in extracts of B. hyalinum, but these were present in extracts of B. steini. Electron microscopy of the cortical region revealed membrane-bound granules in both species; these granules differed in content but not in their capacity to extrude. In B. hyalinum all granules had a homogenous electron-dense substructure; in B. steini the granules had a net-like granulated substructure of varying electron density. This difference corresponds to that published on “pigment” granules in albino and pigmented strains of B. undulans. Our conclusions are that B. hyalinum is unpigmented (and a valid separate species) and that the cortical granules may serve other functions than that of storing blepharismin.

Notes: . Sporozoites of Eimeria vermiformis from the mouse were first seen in the epithelial cells of villus tips and the crypts of Lieberkühn four hours after inoculation (HAI). They were always within a parasitophorous vacuole. By 12 HAI, most were in crypt epithelial cells between the basement membrane and host cell nucleus. The sporozoites in the villus tips had 26 subpellicular microtubules, two polar rings, two preconoidal rings, two refractile bodies surrounded by amylopectin-like granules, a lamellar Golgi apparatus, numerous micronemes, and rhoptries. The sporozoites in the crypt cells had fewer amylopectin-like granules, micronemes, and rhoptries. A nucleolus was visible, as were pieces broken off from the posterior refractile body. Later, the sporozoites folded over to become U-shaped; the infolded membranes fused; and then the inner membranes disappeared so that spherical meronts were formed. Folding sporozoites were first seen 16 HAI and persisted until 52 HAI.

Notes: . From an intermittent stream in College Station, Texas, a Paramecium was isolated that did not appear to belong to any recognized species. On the basis of nuclear and whole-body morphology, it can be assigned to the Paramecium aurelia species-complex, and it can be distinguished from other members of that complex on the basis of mating-type reactivity and isoenzyme patterns. These characteristics are felt sufficient to justify a new species assignment. The new species has been named Paramecium sonneborni n. sp. in honor of the late Dr. Tracy M. Sonneborn of Indiana University.

Notes: . Enzyme electrophoresis was exploited to identify stocks of paramecia previously not identified to particular species. Stocks collected in India and one from Panama belong to Paramecium jenningsi, while others collected in Panama or in Brazil are assignable to syngen 2 of P. multimicronucleatum on the basis of similarity of their esterase and acid phosphatase phenotypes. Inclusion of these doubled the numbers of stocks available in the two species, thereby facilitating examination of intraspecies variation and comparison of particular features of intraspecies variation found for the P. aurelia complex. Variant stocks were observed in P. jenningsi and in syngens 2, 3, and 4 of P. multimicronucleatum. In some cases the variant lacked the enzyme; in others, a change in mobility of the enzyme occurred that resulted in an electrophoretic form similar to one common in another species. Unique phenotypes were displayed by the variants of syngen 2 in P. multimicronucleatum. Hypervariability for Esterase B was observed in this syngen, where, in addition, several subtypes were seen for three other esterases. Unique phenotypes and hypervariability were also noted in P. biaurelia. Clustered variations were observed in these species and in the P. aurelia species. Unlike the situation for members of the aurelia complex, where lack of geographical differentiation between stocks in the same species is a unique feature, some such differentiation does occur in P. multimicronucleatum-2. The frequency of variant stocks in P. jenningsi was similar to that observed in the aurelia sibling species. In contrast, a significantly higher frequency of variant stocks was found in syngens 2, 3, and 4 of P. multimicronucleatum.

Notes: The surface charge of Tritrichomonas foetus was evaluated by means of the binding of colloidal iron hydroxide particles at pH 1.8 and cationized ferritin particles at pH 7.2 to the cell surface, as visualized by electron microscopy and by direct measurements of the electrophoretic mobility (EPM), of cells suspended in solutions of different ionic strength and pH. At pH 7.2, T. foetus has a negative surface charge with a mean EPM of −1.03 μmμs−1μV−1μcm. At lower pH, there is a decrease in the negative surface charge with an isoelectric point at pH 1.2. At higher pH (〉 9.0), there is an increase in the surface charge reaching an EPM of −2.5 μmμs−1μV−1μcm. These results indicate that the surface of T. foetus contains both negatively and positively charged dissociating groups. Binding of colloidal iron hydroxide and cationized ferritin particles throughout the cell surface of the protozoon was observed. Treatment of T. foetus with neuraminidase or trypsin reduced significantly the EPM of the cells. Enzyme-treated cells recovered their normal EPM when incubated for 6 h in fresh culture medium by a process that is inhibited by puromycin.

Notes: Amastigotes of Trypanosoma cruzi were purified from overlays of infected Vero cell cultures by centrifugation over a discontinuous gradient of metrizamide. Pure amastigote preparations were usually recovered from the pellet under the layer of specific gravity 1.086. The isolated amastigotes grew in cell-free ML-15HA medium. Growth rate for the different strains of T. cruzi were in the order Y 〉 Tulahuan 〉 CL. The generation time of amastigotes in ML-15HA medium was 16.8, 18.0, and 26.4 h for the Y, Tulahuen, and CL strains, respectively, in the presence of 5% CO2) and 16.8, 31.2, and 36.4 h, respectively, in the absence of CO2. Intracellular amastigotes did not differ ultrastructurally from amastigotes from either the density-gradient fractionation or culture in cell-free medium.

Notes: Unicapsula seriolae n. sp. forms long plasmodia within the striated muscle fibers of Seriola lalandi off eastern Australia. Its spores are composed of three valves, only one of which contains a developed polar capsule. Within the capsule, the filament makes 2 1/2–3 turns, and this criterion can be used to separate U. seriolae from the two other members of the genus. The flesh of infected fish disintegrates during slow cooking.

Notes: Twenty-nine (64.4%) of 45 reindeer, Rangifer tarandus, examined over a two-year period were infected with trypanosomes. Trypomastigotes and dividing epimastigotes were found in the blood of fawns, cows, and bulls. Morphometric analysis of bloodstream trypomastigotes from reindeer and comparison of these parasites with similar stages of trypanosomes from elk, mule deer, and white-tailed deer from the contiguous United States proved them conspecific; the trypanosomes from these members of the Cervidae are identified as Trypanosoma cervi Kingston & Morton, 1975. This is the first report of trypanosomes from reindeer. No pathogenic effects are known to be caused by these parasites.

Notes: We investigated the growth requirements of symbiont-free and symbiont lambda-bearing Paramecium octaurelia stock 299 for folic acid and biopterin in chemically defined culture medium. Symbiont-free P. octaurelia required both folic acid and biopterin for growth. In the absence of these substances growth of symbiont-free P. octaurelia failed after the first transfer, whereas symbiont lambda-bearing P. octaurelia could be maintained indefinitely in serial subculture. In the absence of folic acid and biopterin, sulfanil-amide inhibited growth of the symbiont lambda-beating protozoa. In the presence of folic acid and biopterin, the antiobiotic selectively inhibited growth of lambda symbionts but did not affect growth of the protozoa. In both cases, inhibition by sulfanilamide was reversed by addition of p-aminobenzoic acid to the medium. These results support our earlier finding that folic acid is required for growth of symbiont-free P. octaurelia 299 and that growth of the lambda-bearing strain without exogenous folate denoted synthesis of folic acid by the symbionts. In addition, it appears that the symbionts produce sufficient biopterin to meet the needs of the host protozoon for growth.

Notes: Book Reviewed in this article:Canning, Elizabeth U., ed. 1981. Parasitological Topics: a Presentation Volume to P. C. C. Garnham F.R.S. on the Occasion of His 80th Birthday 1981Krylov, M. V. 1981. Piroplazmidy. [Piroplasms.]Baker, J. R. 1982. The Biology of Parasitic ProtozoaBarriga, Omar O. 1981. The Immunology of Parasitic Infections: a Handbook for Physicians, Veterinarians, and BiologistsBeyer, T. V., Bezukladnikova, N. A., Galuzo, I. G., Konovalova, S. I. & Pak, S. M., eds. 1979. Toksoplasmidy. [The ToxoplasmidsGeltzer, Ya. G., Korganova, G. A., Mavlyanova, M. I. & Nikolyuk, V. I., eds. 1980. Pochvennye Prosteyshie. [The Soil Protozoa.] (ProtozoologiyaBeyer, T. V., Kazakova, I. I., Lakhonina, G. M., Roigas, E. M. & Teras, J. H., eds. 1981. Vzaimootnosheniya Prosteyshikh s Virusami. [The Interaction between Protozoa and Viruses.] (ProtozoologiyaOgimoto, Keiji & Imai, Soichi 1981 Allas of Rumen MicrobiologyLong, Peter L., ed. 1982. The Biology of the CoccidiaLloyd, David, Poole, Robert & Edwards, Steven W. 1982. The Cell Division Cycle: Temporal Organization and Control of Cellular Growth and ReproductionFrederick, J. F., ed. 1981. Origins and Evolution of Eukaryotic Intracellular Organelles. [Ann. N.Y. Acad. Sci.Hayat, M. A. 1981. Fixation for Electron MicroscopyBuetow, D. E., ed. 1982. The Biology of Euglena.Ogden, C. G. & Hedley, R. H. 1980. An Atlas of Freshwater Testate AmoebaeParker, S. P., ed. 1982. Synopsis and Classification of Living OrganismsMargulis, L. & Schwartz, K. V. 1982. Five Kingdoms: an Illustrated Guide to the Phyla of Life on EarthCairns, J., Jr., ed. 1982. Artificial SubstratesCurds, C. R. 1982. British and Other Freshwater Ciliated Protozoa

Notes: . Freeze-fracture techniques reveal differences in fine structure between the anterior three flagella of Tritrichomonas foetus and its recurrent flagellum. The anterior flagella have rosettes of 9–12 intramembranous particles on both the P and E faces. The recurrent flagellum lacks rosettes but has ribbon-like arrays of particles along the length of the flagellum, which may be involved in the flagellum's attachment to the cell body. This flagellum is attached to the membrane of the cell body along a distinct groove that contains few discernible particles. Some large intramembranous particles are visible on the P face of the cell body membrane at the point where the flagellum emerges from the cell body. The randomly distributed particles on the P and E faces of the plasma membrane have a particle density of 919/μm2 and 468/μm2 respectively, and there are areas on both faces that are devoid of particles. Freeze-fracture techniques also reveal numerous fenestrations in the membrane of the Golgi complex and about 24 pores per μm2 in the nuclear. membrane.

Notes: . Extraction of the ciliated protozoon Tetrahymena with nonionic detergents produces a surface-related cytoskeleton that consists of a basic lamina of whole-cell dimensions together with associated microtubule and microfilament systems, including all ciliary basal bodies. The organization of the isolated cytoskeleton has been studied using scanning electron microscopy, and several new features are described in the oral region. Here the ciliary basal bodies are arranged in a very stable and highly complex pattern. This pattern was found to be identical in the four species of Tetrahymena we examined. In addition, various microtubular bundles and two separate systems of filaments were observed in scanning electron micrographs of isolated oral skeletons. The appearance of the deep fiber bundle in preparations of this type suggests that it arises, at least in part, as an extension of the ribbed wall microtubules. On the basis of its distribution within the oral skeleton, one of the filament systems described is suggested to be a contractile system responsible for pinching off food vacuoles.

Notes: . Dehydrogenase activity with hydroxysteroids has been observed in Tetrahymena furgasoni (formerly T. pyriformis strain W), and the enzyme responsible has been isolated from this organism. The purified dehydrogenase is active with a variety of steroid alcohols at apparent Km values ranging from 0.2 to 4.0 mM. The C-3 hydroxyl of ring A of the steroid nucleus is the preferred position of oxidation. However, a variety of other secondary alcohols are also substrates, with apparent Km values for 2-butanol, 2-pentanol, and cyclohexanol of 880, 1000. and 150 mM, respectively. With both steroidal and nonsteroidal alcohols. NAD is the preferred co-substrate, although low activity with NADP is observed. Evidence is presented that the activity with secondary alcohols, whether steroidal or not, is the property of a single protein species.

Notes: . Domestic turkeys naturally infected with Leucocytozoon smithi were blinded by bilateral ocular enucleation, pinealectomized, sham-pinealectomized, pinealectomized plus enucleated, or maintained as controls. Groups of turkeys were acclimated to either light-dark periods of 14L:10D or “darkness” with intermittent periods (10–20 min) of red light at irregular hours approximately every three days as required for maintenance of turkeys. Peripheral gametocyte numbers of L. smithi in all groups were determined every 2 h over a 36 h period. Under 14L: 10D photoperiod, no observable difference in the pattern of gametocyte circadian rhythmicity between pinealectomized, enucleated, pinealectomized plus enucleated, and control turkeys was noted. Although mean parasitemias differed among groups, peak gametocyte numbers occurred between 1000 and 1800 h; how parasitemias occurred between 2000 and 0400 h. However, the phase of gametocyte rhythmicity in pinealectomized plus enucleated turkey hosts did exhibit a lag with reference to other hosts when examined by least squares fits of simple harmonics. Under conditions of “darkness” with intermittent, irregular periods of red light, L. smithi gametocyte numbers of individual turkeys, pinealectomized, sham-pinealectomized, or maintained as controls, exhibited a circadian periodicity though parasite cycles were out of phase with the natural photoperiod to which the turkeys previously had been exposed. A slight drift out of phase of L. smithi gametocyte periodicity occurred among turkeys in the sham-pinealectomized and the control groups while a considerably more prominent drift out of phase was seen among the parasite rhythmicity patterns of the pinealectomized birds. Data indicate that the pineal gland of the turkey did not directly mediate L. smithi gametocyte circadian periodicity, although an indirect involvement in regulating the timing of parasite rhythmicity is suggested.

Notes: . The surface of merozoites and sporozoites of Eimeria tenella was affected by incubation with E. tenella-immune chicken serum (ICS). Normal chicken serum (NCS) and heat-inactivated ICS had no effect on the pellicular surface of either developmental stage. Sporozoites formed surface bulges or swellings after 10 min of incubation with ICS, and by 15 min postincubation, the morphology of the sporozoites was distorted by a surface coating of fibrinous material. Merozoites exposed to ICS were similarly coated, but surface swelling was not as severe. The coating formed rapidly and was seen as early as 5 min postincubation. Sporozoites incubated with heat-inactivated ICS supplemented with normal chicken serum were coated with a fibrinous material and in some cases lysed. These data indicated that complement must be present for the surface interaction to occur.

Notes: . The fine structure of the trophozoite, encysting cells, and the cyst of Acanthamoeba astronyxis has been examined. In the trophic form a microtubule organizing center was associated with a well developed Golgi complex. During encystment the organelles of the amoeba changed considerably. The profiles of rough endoplasmic reticulum elongated and were often arranged in circles of multilayered concentric systems, enclosing mitochondria, the nucleus, or other inclusions. The mitochondria showed a tendency toward elongation and constriction. One or two nucleolus-like bodies appeared in the nucleus. Lipid droplets increased considerably in amount and were distributed individually or as aggregates. The mature cyst was star-shaped and surrounded by an almost circular exocyst and an endocyst that was closely apposed to the cell membrane. Both walls differed in their thickness and granulation. The exocyst was continuous over the entire cyst, while the endocyst was interrupted by gaps, ostioles. in the region of the rays. Within the ostioles was a bell-shaped structure, the operculum. The latter was composed of a granular material comparable in electron density to that of the endocyst.

Notes: Ambiphrya ameiuri is an ectocommensal peritrich that attaches to the gills of warm-water fishes and filters bacteria from the water. The ultrastructure of this protozoon, its attachment to the fish gills, and its effect on the gill tissue were investigated by scanning and transmission electron microscopy. The peritrich attached to the gills by fibers extending from the scopula. A microtubular array, apparently a barren kinetosome, was present in each lobular projection, but no scopular cilia were observed. At low densities Ambiphrya had no apparent harmful effects on the fish; however, at high densities respiration may be impeded. Ultrastructural studies indicate that this organism receives no nourishment from the host tissue.

Notes: Une méthode d'obtention et de maintien des Ciliés du rumen (genre Polyplastron) en culture axénique, est décrite. Elle comporte deux périodes d'incubation en anaérobiose avec différentes associations d'antibiotiques, séparées par un lavage et un renouvellement du milieu nutritif. Dans ces conditions. Polyplastron multivesiculatum est obtenu à l'état axénique et maintenu en survie pendant cinq jours.〈section xml:id="abs1-2"〉〈title type="main"〉ABSTRACTA method to obtain and keep rumen ciliates (genus Polyplastron) in an axenic condition is described. It consists of two incubating periods with different antibiotic mixtures, separated by washing and renewing of nutrient medium. Under these conditions, Polyplastron multivesiculatum is established in an axenic state and can survive thusly for five days.

Notes: Proteins of whole cell extracts of Naegleria fowleri, precipitated with acetone, have been resolved by two-dimensional polyacrylamide gel electrophoresis. Autoradiograms of the [35S]-methionine-labeled polypeptides were scanned and analyzed by a computer-assisted program in order to determine whether there were correlations between selected attributes of proteins (e.g., subunit size and charge). The majority of the polypeptides had molecular sizes within the range of 20–60 kilodaltons. The mean amount of polypeptide was less for those with molecular sizes between 20 and 45 kilodaltons than for those larger than 45 kilodaltons. The mean amount of polypeptide was greater in the isoelectric focusing range of pH 5–6 than in the range of pH 6–7. Polypeptides in the size range of 20–40 kilodaltons had a median isoelectric point of 6.1, whereas polypeptides in the size range of 40–80 kilodaltons had a median pI of 5.6. Our data indicated that molecular size and charge were not entirely independent variables, and that the composition of a polypeptide might have an important influence on its steady state level in N. fowleri.

Notes: During a survey of the coccidian parasites of reptiles from Iowa, three specimens of Crotalus horridus L., the Timber Rattlesnake, and one of Sistrurus catenatus (Rafinesque), the Massasauga Rattlesnake, were found to be passing oocysts of a Caryospora, here described as C. bigenetica n. sp. Since these snakes (family Crotalidae) are known to subsist mainly on small mammals, oocysts from one of the Timber Rattlesnakes were fed to laboratory white mice (Mus musculus L.) to determine if mammals might be involved as alternate hosts in the life cycle. At necropsy, tissues of the tongue and dermis of the mice revealed a sequence of stages which included mature male and female gamonts, fully sporulated sporocysts, “excysted” sporozoites, and “resting” sporozoites that lay individually in solitary, cyst-like host cells termed “caryocysts.” A coccidia-free Massasauga that was fed an infected mouse, at a time when caryocysts in the mouse would have been present, later passed oocysts similar to those of the original inoculum. These results, along with the discovery of endogenous stages (asexual and sexual) in the intestine of the Timber Rattlesnake and the experimentally infected Massasauga, suggest that this parasite has a heteroxenous life cycle pattern, with sexual stages occurring both in the ophidian and the mammalian hosts.

Notes: Ciliophrys marina is a small marine helioflagellate, with a central nucleus, which is capable of reversibly transforming from a rapidly swimming flagellate cell with no axopodia to the structure of a heliozoan with a flagellum that beats only a few times a minute. When in the flagellate form, the flagellum acts as a tractellum due to the tubular mastigonemes found along its length. When the rapidly swimming flagellate strikes a piece of debris, the flagellum goes through a very characteristic shock-induced avoidance reaction. Similarly, when a mechanical shock is delivered to the cell in its heliozoan form, the axopodia are contracted in less than 20 msec. Both reactions are inhibited in low calcium seawater. Transformation from the heliozoan to the flagellate form is accomplished by slow retraction and absorbance of the axopodia and activation of the flagellum. Ultrastructurally, each axopodium is found to contain three microtubules which attach to the outer nuclear membrane of the central nucleus at sites that this study characterizes by electron microscopy of thin sections and freeze fracture preparations. The mitochondria have tubular cristae, each containing an intracristal filament. Finally, a taxonomic review of the helioflagellates is presented, and it is suggested that C. marina is derived from the chrysomonads. An argument is also made for classifying C. marina with the heliozoan order Actinophryida, as a recently published classification of the protozoa does.

Notes: Laboratory-reared Fundulus grandis and F. heteroclitus were experimentally infected with Eimeria funduli by being fed Palaemonetes pugio (grass shrimp) collected from endemic areas. Histological sections were made of heart, liver, hepatopancreas, spleen, gall bladder, kidney, intestine, peri-intestinal fat, reproductive organs, and brain from F. grandis sacrificed at 1, 2, 6, 12, 18, and 24 h and from F. heteroclitus at 5, 6, 9, 14, 19, 24, 29, 34, 39, and 44 days after consuming naturally infected shrimp. We first found merogonous stages at day 9 postinfection (p.i.). No developmental stages of the parasite could be positively identified in the tissues of experimentally infected fish prior to day 9 p.i. Mature meronts were found 14 days p.i. The majority contained 8–16 (mean, 13) merozoites, but a few meronts had 18–26 (22) merozoites. Gamonts first appeared on day 14, were mature by day 19, and fertilization was completed by day 24 p.i. After sporoblast formation, sporopodia appeared during sporocyst wall formation, between days 24 and 29 p.i. Sporozoite formation was completed by day 44 p.i. in most sporocysts. Most endogenous stages occurred in hepatocytes; however, pancreatic and spleen cells were sometimes infected with gamonts.

Notes: A recent isolate of Eimeria praecox, strain G, was obtained from Georgia and purified. Studies of the life history, pathogenicity, and cross-immunity of the isolate were conducted to verify its identity. In inoculated three-week-old chickens, the occurrence of merogony and gametogony was limited to the superficial epithelium of the upper intestine. Oocysts, 23 × 19.5 äm, with a shape index of 1.17 were first observed 83 h after inoculation. Mortality and morbidity were not observed in any of the experimental birds. However, there was a positive correlation between dose of oocysts, reduced weight gain, and the incidence of exudative diathesis. These studies showed that E. praecox depresses weight gains in chickens and may be of economic importance. Although complete immunity to avian coccidiosis is believed to be species specific, chickens immune to E. praecox (G) or E. acervulina had a degree of cross-immunity to a heterologous challenge. Electrophoretic analysis of glucose phosphate isomerase and lactate dehydrogenase prepared from the European strain of E. praecox and E. praecox (G) showed no differences, confirming the identity of the isolate as E. praecox.

Notes: The antiviral agent phosphonoacetic acid inhibits growth of Tetrahymena thermophila at concentrations comparable to those inhibiting growth of other eukaryotic cells, with 50% inhibition at 0.5 mM phosphonoacetic acid. The compound is cytotoxk to Tetrahymena at concentrations greater than 2.0 mM. When a culture of Tetrahymena the growth of which was totally inhibited by 2.0 mM phosphonoacetic acid was diluted with fresh medium, growth resumed in an exponential, rather than synchronous, fashion. [2–14C]phosphonoacetic acid is not metabolized by Tetrahymena.

Notes: Agglutinins were not detected in sera from mice given one, two, or three intranasal (i.n.) inoculations or a single intravenous (i.v.) inoculation of trophozoites of Naegleria fowleri. However, agglutinins were produced following second and third i.v. inoculations. Serum immunoglobulin levels increased in both i.n.- and i.v.-inoculated mice. IgG and IgM increased substantially more for i.v.-inoculated mice. IgA levels increased more consistently for i.n.-inoculated mice.

Notes: The sensitivity and specificity of the indirect fluorescent antibody (IFA) test for the detection of serum antibodies were examined in mice that were infected with Eimeria falciformis, E. ferrisi, E. papillata, or E. vermiformis. For the study of each species, five groups of mice were given graded inoculation doses of 10, 102, 103, 104, or 105 sporulated oocysts in a primary infection. The sixth group was infected with three sequential doses of 1.5 times 103, 1.5 times 104, and 1.5 times 105 sporulated oocysts per mouse at two- to three-week intervals. All groups of infected mice developed serum antibodies. Sera were titrated by the IFA test with purified sporozoites. Strong fluorescence and high IFA titers were observed with homologous reactions mainly with the sera from mice infected with the higher inoculation dose levels in primary infections and from those given three sequential inoculation doses. Immunological cross reaction among the four species of Eimeria occurred at dilutions of 1:10 to 1:160. Very weak or no fluorescence of free sporozoites was observed with sera from noninfected mice, and there was no fluorescence of sporozoites contained in intact sporocysts.

Notes: The structure of both the host and parasite membranes during stages in the asexual development of Plasmodium chabaudi in mouse red blood cells is examined by transmission electron microscopy of thin sections and freeze-fracture preparations. The erythrocyte's plasma membrane, the membrane of the parasitophorous vacuole, and the plasma membrane of the parasite exhibit different structural properties in terms of membrane width and the frequency and diameter of the typical intramembrane-particles (IMP) populating the membrane's fracture faces. The difference between the parasitophorous vacuolar membrane and host cell's plasma membrane is remarkable because the vacuolar membrane is formed from an invagination of the erythrocyte's plasma membrane. The vacuolar membrane has significantly reduced frequencies and diameters of IMP's on both faces and reveals a marked temperature response manifesting itself as large IMP-devoid domains emerging on both faces on chilling to 4°C. In contrast, cooling induces only some very small IMP-devoid patches on both faces of the host plasma membrane. Neither of these membranes changes significantly as parasite development progresses. In contrast, the parasite's plasma membrane shows local alterations during its development, forming compaction domains with the nuclear envelope in ca. 30% of the ring-stages and trophozoites. These compaction domains disappear in late uninuclear trophozoites and schizonts. Furthermore, the plasma membrane of the host cell, the vacuolar membrane, and the parasite's plasma membrane do not respond to externally applied Ca2+, and their temperature-response remains unaltered during the infection cycle. Thus, modification of these three membranes as a consequence of invasion and development of the parasites, as recently found in the primate malaria caused by P. knowlesi, can be detected neither directly nor indirectly via temperature- and/or Ca2+-response in the rodent malaria caused by P. chabaudi.

Notes: A system of prescreens and screen has been developed to select chelators as potential drugs against Trypanosoma brucei brucei EATRO 110. The chelators tested were nearly all commercially available, low molecular, and having moderate to high affinity for Fe(III). We prescreened 70 compounds showing heme-sparing or inhibitory activity in a Crithidia fasciculata growth system having excess Fe and minimal hemin. Of these, 45 were highly trypanocidal for suspensions of bloodstream T. b. brucei; criteria of activity here were immobilization, lysis, and loss of infectivity. Eighteen of the chelators highly active in the suspension prescreen were tried in T. b. brucei-infected mice. Thirteen of these chelators were curative in mice with 24-h infections, that is, they allowed survival 〉30 days beyond the untreated controls. 3,4-Dihydroxycinnamic acid (caffeic acid). 2,9-dimethyl-1, 10 phenanthroline (neocuproine), and 2-pyridinecarboxaldehyde-2-pyridyl-hydrazone cured five out of five mice after an i.v. dose of 100 mg/kg. Salicylaldehyde thiosemicarbazone cured five out of five mice at an i.p. dose of 500 mg/kg. Lesser activity was shown by several other chelators.

Notes: The size and fatty acid composition of Tetrahymena pyriformis W cells were influenced by the provision of a nutritional supplement of ergosterol, cholesterol, or tetrahymanol, but not of 20-isocholesterol. Ergosterol and cholesterol addition led to a reduction in cellular volume, an increase in glycerophospholipid saturated fatty acid content, and an increase in palmitoleic acid and its metabolic products when compared to unsupplemented controls. Tetrahymanol supplementation resulted in an increase in cellular volume, a decrease in saturated fatty acid content, and a reduction in palmitoleic acid and derivatives. 20-Isocholesterol was accumulated by the cells; however, this compound had no effect on any of the parameters followed in this investigation and had only a small depressant effect on tetrahymanol biosynthesis. Ergosterol and cholesterol had the same impact on the ciliates, even though the ergosterol-supplemented cells contained approximately three times as much free sterol as did cholesterol-grown cells. The amount of the free cholesterol and metabolic products in supplemented cultures was similar to the amount of tetrahymanol present in control cultures. This observation suggests that the cells recognize qualitative differences among the various polycyclic alcohols rather than responding to the amount of sterol present. Increased cellular levels of tetrahymanol led to a response unlike that of the true sterols, which again suggests that the high degree of specificity depends on the structure of the added polycyclic alcohol. The changes in fatty acid composition may be required to maintain proper interaction of the polar lipids and the polycyclic alcohols to give an appropriate degree of membrane fluidity.

Notes: An unknown, ninhydrin-positive substance detected on paper chromatograms of the endogenous metabolites of mixed rumen ciliate protozoa was isolated and purified by column chromatography with ‘Dowex’ 50-X8 resin and identified as 2-aminobutanoic acid (α-amino-n-butyric acid) on the basis of elementary analysis, mass spectrometry, paper chromatography, infrared spectrometry, and melting point.

Notes: Book reviews in this article: Dillon, Lawrence S. 1981. Ultrastructure, Macromolecules and Evolution Schwemmler, Werner & Schenk, Hainfried E. A., eds. 1980. Endocytobiology: Endosymbiosis and Cell Biology, a Synthesis of Recent Research Krylov, M. V. & Starobogatov, Y. I., eds. 1980. Principles of the Construction of the Macrosystem of the Unicellular Animals Palmer, W. J., ed. 1981. Rapid & Automated Methods in Microbiology & Immunology: A Bibliography, 1976–1980 de Harven, E. & Nemanic, M. K., organizers. 1981. Cell Surface Labeling Inoki, Shozo, ed. 1981. Atlas of Protozoa Czapik, Anna 1980. Pdostawy Protozoologii. [Introduction to Protozoology.] Levine, Norman D. & Ivens, Virginia 1981. The Coccidian Parasites (Protozoa, Apicomplexa) of Carnivores Page, Frederick C. 1981. The Culture and Use of Free-Living Protozoa in Teaching

Notes: The trophont stage in the life cycle of Ichthyophthirius multifiliis was studied in the electron microscope. Surface ridges contain up to 24 ridge microtubules, disposed as a ribbon. Kinetosomes show the classic morphology of 9 triplets of microtubules. Associated with each kinetosome is a kinetodesmal fibril, originating in proximity to triplets 5, 6, and 7, and having a 30 nm periodicity; 3 to 5 postciliary microtubules, originating between triplets 8 and 9; and up to 3 transverse microtubules, originating at triplet 4, as well as a parasomal sac. Each cell is partially enclosed by a system of 3 “unit” membranes: the outer limiting membrane, and the outer and inner alveolar membranes. The last two membranes define the alveolar sac. Mucocysts, each with a dense core, are present in large numbers. The contractile vacuole system includes the contractile vacuole, associated tubules and vesicles, injection canals, a discharge canal, and a pore. Microtubules abound in the walls of the contractile vacuole, injection and discharge canals, and in the region of the pores, where both ring and radial microtubular arrangements are noted. The ultrastructure suggests that I. multifiliis is more closely related to Tetrahymena pyriformis than to Paramecium aurelia.

Notes: Experiments with Tetrahymena thermophila using ferritin probes revealed that these cells take up ferritin conjugated to antibodies (not directed against Tetrahymena) much more readily than they do ferritin or cationized ferritin. The massive and rapid uptake of antibody-ferritin offers certain advantages for studies of endocytosis and membrane flow in cells of this type, and the method may be applicable to other types of protozoa as well.

Notes: The mechanism of action of antileishmanial compounds is poorly understood. Ultrastructural changes in Leishmania tropica within human macrophages exposed in vitro to Pentostam, pentamidine, amphotericin B, WR 6026, ketoconazole, and Formycin B were examined in these experiments. In Pentostam-treated cultures, some organisms exhibited diminished definition of mitochondrial and other membranes, while other organisms had completely disintegrated. Pentostam-exposed macrophages demonstrated loss of membrane definition in the absence of further alterations; it is therefore hypothesized that impaired macrophage membrane function may contribute towards the effect of this drug against macrophage-contained organisms. Leishmania parasites in pentamidine-treated cultures initially demonstrated swollen kinetoplasts and fragmentation of the kinetoplast DNA core. The initial observed effect of the other four drugs on the parasites was cytoplasmic condensation. These ultrastructural studies suggest that all five non-antimonial drugs may have different mechanisms of action than antimony (Pentostam) against Leishmania.

Notes: Immobilization antigens from 12 serotypes of three stocks of Paramecium tetraurelia and from one serotype of one stock of P. primaurelia were isolated and purified. Purified proteins were cleaved with cyanogen bromide, and the patterns of the fragment peptides were determined by electrophoresis on SDS-polyacrylamide gels. It was shown that each of the serotypes of stock 51 of P. tetraurelia has an antigen that produces a characteristic and unique pattern. Consequently, the antigens can be identified by their patterns. Antigens from the allelic serotypes tested had identical patterns. The method is sensitive enough for the investigation of small sample volumes, and useful as a simple biochemical technique for the identification of serotypes.

Notes: Nutritional requirements of Acanthamoeba polyphaga (strain PD) were compared to those reported for A. castellanii. Although A. polyphaga and A. castellanii have essentially the same minimal amino acid requirements–arginine, methionine, leucine, isoleucine, and valine–A. polyphaga cannot utilize acetate as sole carbon source, but A. castellanii can if the medium is supplemented with glycine.

Notes: Classification at the species level has been difficult in the genus Acanthamoeba. The taxonomic designations of a number of strains are in doubt and new approaches to classification are needed. We describe the use of electrophoretic patterns obtained with restriction enzyme digests of mitochondrial DNA as a basis for one new approach. Results from analysis of ten strains of A. castellanii, two of A. polyphaga and one of A. astronyxis are discussed. Examples both of nucleotide sequence diversity and of sequence conservation have been found among strains with the same species designation. Five strains from Europe, North America and New Zealand had identical digestion phenotypes with five enzymes; consequently, very similar nucleotide sequences are predicted. All are pathogenic to humans or mice. The mtDNA sequences of eight remaining strains are predicted to differ from this cluster and, in most cases, from each other at least as much as in sibling species of Paramecium aurelia.

Notes: When Tetrahymena furgasoni (W) are stained by the silver impregnation technic. a relatively small percentage of them show cortical argentophilic bands or stripes that are intermeridional in position. The number and arrangement of the bands as well as their distinctive appearance and abrupt daily peaks of occurrence render them distinct from other recognized components of the argyrome. The term “virgulene” is proposed as a name for this feature.

Notes: The differentiation of Trypanosoma cruzi epimastigotes into trypomastigotes was studied in diffusion chambers sub-cutaneously implanted in mice. Using epimastigotes of the Tulahuén strain, transformation was first evident at 16 h after implantation and reached its maximum (92% trypomastigotes) by 24 h. Shortly before their differentiation into trypomastigotes, epimastigotes were found to develop resistance to lysis by the alternative pathway of complement. Furthermore, implantation of stationary-phase (as opposed to log-phase) parasites resulted in the accumulation of large numbers of complement-resistant epimastigotes in the chambers. These observations suggest that epimastigotes pass through a complement-resistant transitional stage before differentiating into trypomastigotes and that transformation may require cell division. In a further series of experiments, epimastigotes recovered 7 h after implantation in mice were found to differentiate into trypomastigotes when cultured in vitro for an additional 17 h at 37°C. This observation indicates that the events which trigger the morphologic transformation of epimastigotes into trypomastigotes can be dissociated operationally from the differentiation process itself.

Notes: Morphological, cultural, and biochemical criteria that have been used in describing lower trypanosomatids, genera Blastocrithidia, Crithidia, Leptomonas, Herpetomonas, Rhynchoidomonas, and Phytomonas are reviewed. Kinetoplast structure, carbohydrate utilization, electrophoretic mobilities of isoenzymes, and kDNA fingerprinting are among the recommended criteria for species differentiation. Temperature, pH, and osmolarity tolerance are useful growth parameters. Generic placement may be assisted by the determination of nitrogenous excretion products and ornithine-arginine cycle enzymes.

Notes: Ten antiprotozoal drugs were tested in vitro against four axenic strains of the intestinal parasite Blastocystis hominis. Inhibitory drugs in order of effectiveness were emetine, metronidazole, furazolidone, trimethoprim sulfamethoxazole, 5-chloro-8-hydroxy-7-iodo-quinoline (Entero-Vioform), and pentamidine. Moderately inhibitory were two quinolines other than iodochlorhydroxquin. These were chloroquine and 5, 7-diiodo-8-hydroxy-quinoline(Floraquin). Diloxanide furoate was not inhibitory. Paromomycin and other antibiotics were not inhibitory. Entero-Vioform and metronidazole have been effective in human and higher primate diarrhea caused by B. hominis.

Notes: Groups of mice were given an intraperitoneal injection of one of six monoclonal antibodies to Toxoplasma gondii, a mixture of equal amounts of five monoclonal antibodies to T. gondii, or the murine myeloma protein MOPC 21, and challenged with either a highly virulent or moderately virulent parasite strain. Two monoclonal antibodies (FMC 19 and FMC 22) conferred total protection against the moderately virulent challenge, with all mice surviving, whereas 90% of control mice died. FMC 19 and FMC 22 also conferred significant protection against the highly virulent challenge as indicated by a prolonged mean time to death (MTD) of immunized compared with control groups of mice. One monoclonal antibody (FMC 23) and the mixture of five antibodies gave significant protection against the moderately virulent challenge only. Passive immunization with dilutions of FMC 22 antibody indicated that the lowest serum titer needed to confer significant protection to mice against a moderately virulent Toxoplasma challenge was 1/640. Mice challenged with highly virulent tachyzoites that had been preincubated with FMC 22 had a significantly longer MTD than mice challenged with highly virulent tachyzoites that had been preincubated with MOPC 21 or phosphate buffered saline, pH 7.2 (PBS). Immunoprecipitation and autoradiography of radiolabeled tachyzoites confirmed that FMC 19 was directed against a 35,000 molecular weight (mol. wt.) antigen and FMC 22 was directed against a 14,000 mol. wt. fraction. The potential for use of single antigens as protective immunogens in preventing toxoplasmosis is raised.

Notes: The heliozoan Echinosphaerium nucleofilum produced about 75 times 103 floating cells per 19-cm culture dish per day when fed the green flagellate Chlorogonium elongatum. This method yields enough cells for usable quantities of subcellular fractions. Heliozoa were lysed in a detergent mixture containing stabilizing reagents, and axonemal bundles of axopodial microtubules were isolated from the lysate by differential centrifugation. Polyacrylamide gel electrophoresis in sodium dodecylsulfate showed two prominent bands tentatively designated alpha- and beta-tubulin. Apparent molecular weights were 51.8 times 103 and 48.1 times 103, respectively. As assayed by electron microscopy of negatively stained whole mounts, the microtubule bundles splintered readily, although glycerol tended to inhibit this fraying. Intermicrotubule bridges could be observed in some axonemal splinters.

Notes: The antipsychotic drugs chlorpromazine, pimozide, and penfluridol caused a 50% inhibition of growth of Tetrahymena at concentrations of 4.5, 5.5, and 1.5 μM, respectively. The degree of growth inhibition was dependent on the concentration of cells; higher drug concentrations were needed to produce inhibition of denser cell cultures. Binding studies with penfluridol showed that 50% growth inhibition resulted when approximately 50 μmoles of drug were bound per 106 cells. A 20-min preincubation of cells with chlorpromazine (14.7 μM) inhibited DNA synthesis by 46%, and with penfluridol (4 μM) DNA synthesis was inhibited by 27%. The incorporation of labeled thymidine into the thymidine triphosphate pool was inhibited by chlorpromazine but not by penfluridol, indicating that the drugs produce their growth inhibitory effects by different mechanisms. TDP kinase activity was demonstrated in a particle-free fraction of the cells. Its enzymatic activity was not affected by added chlorpromazine, penfluridol, or calmodulin, suggesting that inhibition of DNA synthesis by these drugs may be a consequence of growth inhibition.

Notes: The protozoan Blepharisma japonicum showed negative phototaxis caused by transient reversal of the direction of ciliary beat and changes of swimming velocity induced with varying intensities of light. The ciliary reversal occurred at 1–2 sec after a sudden increase in light intensity. When light intensity was decreased, no response was observed. Moreover, the ciliates swam fast in light areas but slowly in dark areas; the mean velocity of swimming was 80 μ m/sec at 5 × 102 lux but reached about 400 μMm/sec at 5 × 103 lux. In addition, the cell body elongated in response to light application; the mean length of the body was 308 μm at 5 × 102 lux, which increased to 397 μ m at 104 lux. Such body elongation seems to contribute to rapid swimming. Negative phototaxis may be an important behavior in B. japonicum because the organisms are killed by exposure to strong light.