Population Growth in Stationary and Suspension Culture of Paramoeba pemaquidensis Page (Amoebida,Paramoebidae)1

Paramoeba pemaquidensis Page requires attachment to a substrate in order for population growth to occur. Populations of amoebae suspended in bacterized culture media on a roller tube apparatus did not increase in size whereas amoebae maintained in stationary control tubes multiplied in number. Amoebae maintained in suspension for as long as three weeks attached and multiplied in culture when allowed to settle. This suggests that Paramoeba found suspended in the surface microlayer of the ocean may be in a “dormant” state. Implications for the survival and geographic distribution of the genus are discussed.     

Paramoeba invadens n. sp. (Amoebida,Paramoebidae), a Pathogenic Amoeba from the Sea Urchin,Strongylocentrotus droebachiensis,in Eastern Canada1

Paramoeba invadens n. sp. is described from sea urchin tissues and from culture. Amoebae are 20–40 μm in length with an irregular hyaline region producing short subpseudopodia. One parasome per cell is present. The parasome is bipolar with basophilic, Feulgen-positive poles and a Feulgen-negative median segment. The fine structure of the parasome resembles those in other species of Paramoeba and the surface of the amoeba is plain with no hairs or scales. Amoebae dislodged from tissues often adopt a semifloating form; floating forms have been seen in culture. Amoebae circulate within lumina of the water vascular system and can migrate readily into or out of tissues.     

Effect of Temperature and Prey Availability on Growth of Paramoeba invadens in Monoxenic Culture

Paramoeba invadens Jones 1985 is a pathogenic marine amoeba responsible for mass mortalities of sea urchins (Strongylocentrotus droebachiensis) of Nova Scotia between 1980 and 1983. A direct relationship between temperature and sea urchin paramoebiasis has been shown in previous laboratory and field studies. This study examined the effect of prey availability and temperature on the growth of P. invadens in monoxenic culture (with the marine bacterium Pseudomonas nautica). At 15°C, the specific growth rate of P. invadens increased with bacterial prey concentration and was highest at 10⁸ bacterial cells ml⁻¹. Growth rate of P. invadens was maximal at 15 to 20°C (which corresponds to annual sea temperature maxima in the natural environment) and the minimum generation time was 19.41 h at 20°C. At 10 and 12°C, generation times were 91.18 and 73.39 h, respectively; at 2 and 5°C, there was no growth. P. invadens did not survive in monoxenic culture at 27°C. Growth rates of P. invadens in vitro were positively correlated with time to morbidity of infected S. droebachiensis.     

Adhesion,Morphology, and Locomotion of Paramoeba pemaquidensis Page (Amoebida,Paramoebidae): Effects of Substrate Charge Density and External Cations1

Morphology and locomotive behavior in the marine amoeba, Paramoeba pemaquidensis Page, was examined under different environmental conditions. Paramoeba requires a minimum surface negative charge density for adhesion of amoebae to substrata. Once adhesion to the substratum has been attained, however, surface negative charge density has no effect on morphology or locomotive rate. Divalent cations are not required for adhesion, but external calcium is required for normal locomotion. In the presence of calcium, Paramoeba often assumes a locomotive form with a broad, well-developed anterior hyaline region and truncate posterior region. Locomotive forms vary from those with only a well-developed hyaline region (Flabellula-like) to forms with long digitiform sub-pseudopodia (Vexillifera-like), with intermediate morphotypes. Locomotive rates decrease and anteroposterior polarity disappears in the presence of living or heat-killed bacteria, indicating that phagocytosis temporarily interferes with locomotion and alters form.     

Paramoeba sp. (Amoebida, Paramoebidae) as the possible causative agent of sea urchin mass mortality in Nova Scotia

An amoeba resembling Paramoeba has been cultured from tissues of diseased sea urchins. Cultures containing the amoeba produced signs of the disease when injected into the coelom of healthy urchins. Control treatments lacking the amoeba did not cause the disease. The amoeba was cultured from radial nerve fragments and seen in tissue sections from experimentally infected urchins. Cultures of the amoeba from these experimentally infected urchins reproduced the disease in healthy urchins by both injection and water-borne routes. These observations suggest involvement of Paramoeba in recent mass mortalities of sea urchins in Nova Scotia.     

A Comparative Characterization of the Mitochondrial Genomes of Paramoeba aparasomata and Neoparamoeba pemaquidensis (Amoebozoa,Paramoebidae)

Marine amebae of the genus Paramoeba (Amoebozoa, Dactylopodida) normally contain a eukaryotic endosymbiont known as Perkinsela‐like organism (PLO). This is one of the characters to distinguish the genera Neoparamoeba and Paramoeba from other Dactylopodida. It is known that the PLO may be lost, but PLO‐free strains of paramoebians were never available for molecular studies. Recently, we have described the first species of the genus Paramoeba which has no parasome—Paramoeba aparasomata. In this study, we present a mitochondrial genome of this species, compare it with that of Neoparamoeba pemaquidensis, and analyze the evolutionary dynamics of gene sequences and gene order rearrangements between these species. The mitochondrial genome of P. aparasomata is 46,254 bp long and contains a set of 31 protein‐coding genes, 19 tRNAs, two rRNA genes, and 7 open reading frames. Our results suggest that these two mitochondrial genomes within the genus Paramoeba have rather similar organization and gene order, base composition, codon usage, the composition and structure of noncoding, and overlapping regions.     

Isolation and identification of pathogenic Naegleria australiensis (Amoebida, Vahlkampfiidae) from a spa in northern Italy

Samples from therapeutic swimming pools and mud basins were cultured for free-living amoebae. Seven strains of pathogenic Naegleria species were isolated. Although some of the strains were as virulent as Naegleria fowleri, the etiological agent of primary amoebic meningoencephalitis, they were identified as Naegleria australiensis with the indirect fluorescent-antibody technique. The virulence of the isolates for mice corresponded with the cytopathic effect for Vero cells. The N. australiensis strains were isolated from swimming pools with water temperatures ranging from 32 to 35 degrees C and from mud with temperatures from 25 to 43 degrees C. The presence of pathogenic N. australiensis in the swimming pools did not correlate with bacterial indicators.     

Paramoeba aparasomata n. sp., a symbiont-free species,and its relative Paramoeba karteshi n. sp. (Amoebozoa,Dactylopodida)

Two brackish water amoebae have been isolated and studied from the benthic biotopes of the Chupa Inlet (Kandalaksha Bay, northwestern Russia). Both strains can be identified as new species of the genus Paramoeba (Amoebozoa, Dactylopodida, Paramoebidae) based on light microscopical characters, structure of microscales on the cell surface and molecular evidence based on the analyses of two genes, nuclear SSU rRNA and mitochondrial cytochrome c oxidase subunit 1 (COI). Paramoeba aparasomata n. sp. is of particular interest because this amoeba is permanently lacking a symbiotic Perkinsela-like organism (PLO) present in other species of Paramoeba and Neoparamoeba. The results obtained show that scaly dactylopodial amoebae lacking PLO are not necessarily members of Korotnevella. In particular, we suggest that Korotnevella nivo Smirnov, 1997, with microscales very similar to those of Paramoeba eilhardi and the species studied here in structure, may be in fact a member of Paramoeba. Molecular data on K. nivo have to be obtained and analysed to test this hypothesis. Based on our new results we emend the diagnosis of the genus Paramoeba to make it more fit to the current phylogenetic conception.     

Isolation and identification of pathogenic Naegleria australiensis (Amoebida, Vahlkampfiidae) from a spa in northern Italy.

Samples from therapeutic swimming pools and mud basins were cultured for free-living amoebae. Seven strains of pathogenic Naegleria species were isolated. Although some of the strains were as virulent as Naegleria fowleri, the etiological agent of primary amoebic meningoencephalitis, they were identified as Naegleria australiensis with the indirect fluorescent-antibody technique. The virulence of the isolates for mice corresponded with the cytopathic effect for Vero cells. The N. australiensis strains were isolated from swimming pools with water temperatures ranging from 32 to 35 degrees C and from mud with temperatures from 25 to 43 degrees C. The presence of pathogenic N. australiensis in the swimming pools did not correlate with bacterial indicators.     

Growth of Paramecium tetraurelia in Bacterized,Monoxenic Cultures1

Wild type and mutant Paramecium tetraurelia were grown in monoxenic cultures by first growing Enterobacter aerogenes on a defined medium and then adding the Paramecium to the stationary phase bacterial culture. The bacterial growth was proportional to the concentration of the carbon source (citrate), and the Paramecium growth was dependent upon both the bacterial density and the starting density of Paramecium. The behavior, electrophysiological properties, ciliary lipid composition, and growth characteristics were similar to the commonly used bacterized medium (Cerophyl) except that 5–10 times greater Paramecium yields were reliably obtained.     

Two Marine Species of Flabellula (Amoebida,Mayorellidae)*

SYNOPSIS. Two species of amoebae identified as Flabellula citata Schaeffer, 1926, and F. calkinsi (Hogue, 1914) were isolated from saltwater habitats on the Maine coast. Both species showed the locomotive form and behavior of the genus Flabellula emend. Bovee, 1965. Both showed a mitotic pattern distinct from that of the genus Vannella Bovee, 1965, supporting Bovee's subdivision of Schaeffer's genus Flabellula. Preparations of F. calkinsi had an unusually high proportion of cells with 2 or more nuclei. F. calkinsi is a voracious cannibal. Neither F. citata nor F. calkinsi formed cysts in culture. Both species could be cultivated on media made with highly dilute sea water, but neither survived in a medium equivalent to a freshwater medium.     

Monoxenic nodulation process of Acacia mangium (Mimosoideae, Phyllodineae) by Bradyrhizobium sp

Acacia mangium Willd., a native tree in Australia and Papua New Guinea, has been introduced to countries in Asia and South America where plantations have been established that cover several hundred thousand ha. The present study investigated the early stages of the nodulation process in A. mangium using an homologous Australian Bradyrhizobium strain. After optimizing the axenic nodulation, histological and cytological studies were conducted using light and electron microscopy. These documented the proliferation of Bradyrhizobium, the lysis of mucilage at the root surface, root hair deformation and initiation, as well as the development and growth of multiple infection threads. A belt of tannin-filled cells was shown to surround the central nodular fixation zone. The nodules were of the indeterminate type and the bacteroids had a rod shape, without size modification and with few polyhydroxybutyrate (PHB) granules. Several bacteroids can share the same symbiosome. A. mangium exhibits both classical and novel features in its nodulation.     

Acanthamoeba pearcei N. Sp. (Protozoa: Amoebida) from Sewage Contaminated Sediments

ABSTRACT. Seabottom sediments from a discontinued Philadelphia-Camden 40-Mile ocean sewage disposal site were cultured for cyst-forming free-living amoebae. Barge delivered wastes were discharged at the site from 1973 until 1980 when the site was closed. One station at the southeast margin of the site was sampled at a depth of approximately 50 m, twice in 1978 and once in 1982, 1983 and 1984. Sediment from the 1978 collection yielded Acanthamoeba polyphaga, Vahlkampfia sp., and an unknown amoeba with stellate endocysts similar to those of A. astronyxis. Trophozoites and cysts of the isolate were typical of those described for the genus Acanthamoeba. Biochemical tests employing enzyme electrophoresis and morphological studies on live and stained specimens showed that the isolate was distinct from other well-described species within the family Acanthamoebidae Sawyer & Griffin, 1975.     

Acanthamoeba stevensoni N. Sp. (Protozoa: Amoebida) from Sewage-Contaminated Shellfish Beds in Raritan Bay, New York

Marine sediments from 12 shallow water stations in Raritan Bay, New York were tested for the presence of Acanthamoeba. Eight stations were positive for one or more species of Acanthamoeba, A. castellanii, A. comandoni, A. hatchetti, A. lenticulata, A. polyphaga, A. rhysodes, and Acanthamoeba spp. Isolates that grew at 38–40° C were found at four stations (A. comandoni, A. lenticulata, and two unidentified strains). The two unknown strains were characterized on the basis of morphological features, isoenzyme profiles, and mouse pathogenicity tests. One of the two strains was determined to be a new species and is designated herein as Acanthamoeba stevensoni n. sp., ATCC 50388. Mature cysts were most similar to those of morphological Group II of Pussard & Pons (1977). Acanthamoeba stevensoni n. sp. was isolated from inshore coastal sediments where seawater ranged from 20–30%‰ (ppt.). The sediments supported commercially valuable populations of hard clams, Mercenaria mercenaria, that required depuration prior to sale because of contamination by sewage-associated bacteria.     

Spawning and larval development of Tegula euryomphala (Jones, 1844) (Trochoidea: Tegulidae) from La Herradura Bay,Chile

The spawning and larval development of Tegula euryomphala (Jones, 1844) were studied under laboratory conditions. The release of gametes was stimulated by raising temperature from 16 to 19?°C, and could be induced during several months of the year. Males always spawned first. Eggs were 155.6?μm in diameter and remained unattached to each other. Development was similar to that described for other Trochoidea and lasted 3–5?days at 19?°C. Larvae hatched approximately 24 h post-fertilization at the pre-torsional veliger stage. The larval protoconch of the metamorphic veliger measured 247?μm and was ornamented with six spiral threads and weakly ordered granulation. Larvae metamorphose without any natural substratum.     

A Light- and Electron-Microscopical Study of Protacanthamoeba caledonica n. sp., Type-Species of Protacanthamoeba n. g. (Amoebida,Acanthamoebidae)1

In its amoeboid stage, Protacanthamoeba caledonica n. g., n. sp. closely resembles the genus Acanthamoeba, on both light- and electron-microscopical levels, including possession of a centrosphere with a plaque-shaped centriole-like body. The cyst wall differs from that of Acanthamoeba in lack of preformed exit pores and in fine structure; the occasional apparent division into exocyst and endocyst is due to irregular splitting. The strain isolated from a Scottish estuary did not grow at 37°C and did not grow normally on agar made with 25% sea water, but cysts remained viable after a week in full-strength sea water. Protacanthamoeba n. g. is distinguished from Acanthamoeba on the basis of cyst structure, but it is assigned to the family Acanthamoebidae.