Aurigamonas

Aurigamonas Vickerman et al., 2005 (ref. ID; 6794 original paper)

[ref. ID; 6794]
Diagnosis; Predatory flagellates with numerous stiff, radiating haptopodia and two flagella, one long and propulsive, the other short, crook-like and nonmotile. Two contractile vacuoles adjacent to flagellar bases and to single vesicular nucleus. Each retractible haptopodium bears a terminal haptosome and its base is anchored in the cell cortex. Eukaryotic prey captured by adhesion to haptopodia and either enveloped by pseudopods for ingestion into food vacuoles, or part of their cytoplasm is ingested by myzocytosis. One species. (ref. ID; 6794)
Remarks; On the strength of its unique characters it is suggested that the genus Aurigamonas deserves to be placed in a separate Order apart from the Sarcomonadea/Cercomonadida, along with Cercobodo agilis (Moroff, 1904) Sandon, 1927. We follow Sandon (1927) in distinguishing the genus Cercobodo Krassilstchik, 1886 from Cercomonas Dujardin, 1840, on the presence of an unattached posterior flagellum and a sedentary, non-progressing amoeboid phase in the former. (ref. ID; 6794)
Etymology; Latin Auriga charioteer, from rein-like motion of flagellum during locomotion. (ref. ID; 6794)

Aurigamonas solis Vickerman et al., 2005 (ref. ID; 6794 original paper, 7130)

Aurigamonas solis Vickerman et al., 2005 (ref. ID; 6794 original paper, 7130)

Diagnosis

Slow-moving spherical to ovoid flagellates, 3-18 um diameter, with 30-50 haptopodia, each ~6 um. Long trailing flagellum 9-52 um, beats in sinuous waves from base during translation along substratum producing characteristic oscillatory movement of body; short flagellum 4-8 um. (ref. ID; 6794)

Descriptions

In life, the flagellate's body is spherical to ovoid with a slightly flattened anterior end from which the two flagella emerge. Its size, 3-18 um diameter, depends on what prey it has been feeding on, and how recently it has fed. The moving flagellate has radiating stiff haptopodia of near uniform length, each ~6 um long, radiating from all parts of its surface. The two flagella differ markedly in size and motility. One is long, over three times the body diameter, 12-52 um, and beats slowly to provide the propulsive force for the organism. It emerges vertical to the body surface and trails behind in locomotion, its distal portion in direct contact with the substratum; undulations of the flagellum from its base do not progress into the adherent region. The second flagellum, which arises alongside that first, is short, 6-8 um long, and difficult to see in the moving organism, as it curves sideways and backwards and becomes concealed among the haptopodia. It is only visible in organisms that are highly compressed and it appears to lack motility. Substratum contact by the long flagellum appears to be essential for directional locomotion. The organism wanders erratically over the substratum but occasionally in a straight line at a speed of 0.3-0.5 mm/min, and with a back-and-forth or side-to-side oscillation of the body (angle of oscillation ~30 degrees, rate 150-200/min when the greater part of the flagellum is in contact with the substratum. This distinctive "nodding" or "bobbing" movement of the body plus its investment of haptopodia make the flagellate readily identifiable at all times. If dislodged from the substratum, the flagellate flounders without progression, the long flagellum thrashing vigorously, maintaining the bobbing body motion. Such thrashing results in the accumulation bacteria near the tip of the flagellum, giving it a "waving flag" appearance. Whether these particles are eventually cast or ingested could not be determined. Occasionally, non-motile organisms are encountered that lack a long flagellum, presumably as a result of autotomy. In addition to haptopodia, the flagellate produces prominent pseudopodia; both play a part in feeding. Pseudopodia range in form from broad lobopodia to fine filopodia. They may be very long in relation to the body size, thus a 6 um diameter flagellate may produce, within the space of minutes, three pseudopodia each up to 18 um long from anywhere on its surface. This response is seen when helioflagellates moving along the substratum are dislodged with gentle pippetting. Such pseudopodia serve to reanchor the flagellate to the substratum and after anchorage the flagellate may adpot an amoeboid form with a single spreading lamellipodium or with several highly branched pseudopodia with parsley leaf-terminations. Flagellar beating ceases in such amoeboid forms. On withdrawal of the pseudopodia, flagellum-based propulsion is resumed. Prominent among the cell contents are the nucleus (3-8 um diameter) with central nucleolus and several vacuoles. The nucleus is associated with the flagellar bases, but noticeably separated from them. Two contractile vacuoles situated alongside the flagellar bases, in front of the nucleus, discharge alternately either side of the flagella. The most striking inclusions under differential interference contrast microscopy in starved flagellates are highly refractile spicule-like bodies in vacuoles packing the cytoplasm or, in compressed specimens, garlandng the nucleus; many these spicules can be seen to be free in the vacuole and subject to Brownian motion. Orange lipid globules are occasionally visible in the cytoplasm. Bacteria appear to adhere in a could around the tips of the hatopodia of smaller individuals, sometimes forming a complete investment. (ref. ID; 6794)

Feeding: Aurigamonas preys on various flagellate protozoa and possibly on bacteria. Three modes of ingestion, related to the size of prey, have been observed.

(1) Phagocytosis, after capture and killing, of entire protists.
(2) Myzocytosis of some of the cytoplasm of large prey.
(3) Trawling of surface-attached bacteria by a large lamellipodium.

From a mixed population of soil protozoa taken from Sourhope soil, ingestion of entire Spumella sp. (4-9 um), Bodo saltans (5-9 um), Rhynchobodo sp. (12-15 um), Goniomonas truncata (8-12 um), Apusomonas proboscidea (9-14 um) and Ploeotia sp. (15-21 um) has been observed. Trophic forms only were eaten, encysted stages were ignored by the predator. Species of Heteromita (4-8 um), Cercomonas (12-21 um) and Thaumatomonas were not ingested; encounters with these flagellates led to temporary attachment only, followed by abandonment. Soil ciliate, such as Colpoda cucullus, and amoebae, such as Mayorella spp., were not attacked. Most observations on feeding have concerned predation on B. saltans and on a rigid soil euglenid, Ploeotia sp. The former was routinely supplied as convenient prey in the culture of isolates from all three sites as it does not encyst; the Ploeotia sp. (which does encyst) arounded in an early enrichment culture from Sourhope sample. The violent flicking movements of B. saltans while attached the substratum brought the prey into contact with the predator's haptopodial tips and a single pseudopodium was everted by the Aurigamonas to engluf it, the entire procedure being completed in 2 min, whereas the large Rhynchobodo sp. was ingested over 4 min. Ingested prey occasionally appeared to have in their own cytoplasm refractile spicules similar to those prominent in the cytoplasm of the predator, suggesting that these spicules may be passed from predator to prey during capture. An Aurigamonas on encoutering a creeping Ploeotia made initial contact via the haptopodial tips. One contact had been made, adherence followed with flagellar beating continuing in both predator and prey. Occasionally the prey managed to disengage itself and swim away unharmed, but usually adherence was followed by the production of a single large pseudopodium from the flagellate, its digit-like extensions enveloping the prey; contraction in length of haptopodia following prey seizure was not observed. Contact of pseudopodium and prey was followed by immobilisation of the latter, and from then onward locomotion of the pair was that typical of Aurigamonas alone, with its characteristic bobbing movement. The pseudopod eventually enveloped the entire prey into a food vacuole and the prey was digested. Complete ingestion of Ploeotia took over 20 min from initial contact. Often, a single Ploeotia was 'mobbed" by several Aurigamonas, as many as nine at once. After one predator had made haptopodial and pseudopodial contact, others began to put out cup-like pseudopodia in the absence of haptopodial contact, but only one or two succeeded in making pseudopodial contact and attempting ingestion. Such communal meals were completed with each predator removing its own share of the prey, and without the cytoplasmic fusion characteristic of communal meals in heliozoans. Alternatively simultaneous predation took the form of myzocytosis as described below for predation on Euglena gracilis, each predator removing some cytoplasm and leaving the prey as a pellicle "ghost" with a totally disintegrated interior. Similar feeding with ghost production was observed on Apusomonas proboscidea. The ability of Aurigamonas to feed on even larger, free-swimming prey was studied by offering it E. gracilis (37-45 um) from axenic culture; unlike the rigid Ploeotia, E. gracilis is capable of vigorous squirming movements ("metaboly"). Flagellar entaglement appeared to play an important part in the capture of Euglena. Aurigamonas readily attached to the much larger flagellate close to the point of emergence of the Euglena long flagellum from its pocket. Haptopodia then adhered to the prey's anterior end (which is less subject to compression during squirming activity) and tenaciously held on to it. Euglenas in the act of binary fission appeared particularly susceptible to attack, presumably because they were less motile. Flagellar movement of the Euglena soon ceased but squirming continued for several minutes. Captured euglenas could readily be identified as they too exhibited the characteristic oscillation movement engendered by the movement of Aurigamonas itself. The predator spread itself on the euglena's surface but released its hold after 5-10 min, having ingested some of the prey's cytoplasm through a tubular pseudopodium; often the pseudopodium was inserted into the flagellar pocket of the euglena. That the prey was killed at the pellicle ruptured was witnessed initially by local vacuolation of the prey's cytoplasm followed later by the emission of cell contents (chloroplast fragments, paramylon) after the predator's departure. Increase in size and number of the Aurigamonas were observed over the next 7 days, while the proportion of dead euglenas in the culture increased over the same period. Aurigamonas would also attack and greedily ingest motile forms of Ochromonas danica, as many as three Aurigamonas attacking one Ochromonas simultaneously. Here, however, predation did not always lead to an increase in number of the predator; following engulfment, the predator ejected the chloroplasts and subsequently underwent dissolution, the prey appearing to have killed its attacker. On several occasions Aurigamonas was seen to project a large, flattened lamellipodium over bacteria attached to a surface and ingest them, but attempts to grow the flagellate with only bacteria as food failed. Haptopodia appear to be stable structures, only rarely withdrawn. Their absence was observed, however, in flattened attached individuals which had produced extensive branched pseudopodia and in which flagellar activity had virtually ceased. (ref. ID; 6794)
Cultures of Aurigamonas from all three sites were readily maintained in perpetuity with B. saltans as food, and with mixed bacteria, employing cerophyl, tryptone soya broth or soil extract as the supporting medium. Transfer to fresh medium every 2-3 weeks was sufficient to initiate rapid growth of B. saltans followed by growth of the predator. Serial cultivation with E. gracilis was not achieved; Aurigamonas disappeared form the culture after 3 weeks. (ref. ID; 6794)

Encystation: In old cultures (3 weeks after initiation) the Aurigamonas all became very small and acquired a uniform layer of adherent bacteria. They eventually lost their motility having withdrawn their flagella and their haptopodia. These non-motile small (~3.7-4.8 um) forms were deemed to be encysted, as actively motile Aurigamonas could be recovered from old cultures showing neither trophic predator or prey after storage at 4-8 degrees C for several months, on the addition of fresh medium and B. saltans. (ref. ID; 6794)

Electron Microscopy: Chromium-shadowed whole mounts of Aurigamonas show that, unlike axopodia, haptopodia exhibit a more or less uniform thickness (0.3 um) along their length and possess clubbed termini. Such preparations also show the long flagellum to have fine lateral non-tubular hairs with a distinctive organization, and an acroneme tip. The hairs are ~690 nm long, relatively stiff and of uniform thickness. They are seen emerge in bunches spaced at ~70 nm along almost the length of the axonemal shaft. Their tips appear to mesh to form the felted boundary layer to an extraflagellar sheath composed of the hairs. In some profiles the hairs appear on one side of the flagellum only, in others along both sides. In the latter, the sheath is usually narrower along one side and may reflect bending of unilateral hairs under the flagellar shaft during drying, of fan-shaped bunches of hairs emerging over a broad arc of the flagellar membrane. The short second flagellum also carries hairs but, being bent back against the body, it is difficult to visualise these surface adornments in the same detail. In scanning electron micrographs the number of haptopodia per flagellate varies from 15 to 50, their length being ~5-7 um. The reflexed short flagellum is readily visible in scanning micrographs. During flagellate division it abandons its reflexed posture and grows out to become the long flagellum of the daughter cell. Transmission electron microscopy of thin (~60-70 nm) and thick (~150 nm) sections shows details of the structure of flagella, cytoplasm and haptopodia. The two flagella arise from basal bodies, ~1 um in length, lying close to the nucleus but not abutting on the nuclear envelope. A cartwheel structure is evident in the proximal portion of each basal body. The basal body-axoneme transition region in short with the transition plate lying above the level of the surface membrane with thickened rim next to the flagellar membrane; beyond this plate, the flagellum shaft increases in thickness. Two electron-dense plates flank each basal body and appear to be nucleating centres for a labile system of microtubules that radiate outward in the cortex of the flagellate. Organised flagellar roots have not been observed. The flagellar axonemes have the conventional '9+2' structure; a paraxonemal rod is not evident in transverse sections. The basal bodies are flanked by the two contractile vacuoles, one of which lies alongside a conventional Golgi apparatus that sits in the prenuclear cytoplasm in a shallow cavity of the nucleus. Cisternae of rough endoplasmic reticulum emanate from the nuclear envelope and some appear to contain filamentous material in the lumen; this may represent flagella hairs. A remarkable feature of the cytoplasm is that it is packed with small smooth cisternae bounded by thickened (10 nm) membrane: these are presumed to be in waiting for the moment when the helioflagellate has to synthesise vast quantities of plasma membrane to cover the pseudopodium and lining of the food vacuole. Other inclusions are mitochondria, lipid globules, undischarged haptosomes and the spicule vacuoles that are so prominent a feature by light microscopy. The mitochondria have short tubular or ampulliform cristae with constricted bases. In sections the haptopodia have capitate tips, but no granules are observed along their shafts; the shafts is seen to contain not microtubules but a single hollow cylindrical core, while the tip contains a single haptosome. Inside the dense bounding membrane of the haptosome lies a hyaline shell enclosing in electron-dense core. The hollow cylinder supporting the haptopodial shaft is composed of closely packed parallel microfilaments, each ~7 nm diameter. The proximal end of the cylinder is anchored in a inclusion-free papilla (0.7 um high, 0.85 um diameter) at the base of the haptopodium. In this base the microfilamentous cylinder has a more marked electron density and, in transverse sections of the papilla, radiating filaments are seen to be associated with the anchor region. The haptosomes that cap the haptopodia are found in abundance in the cytoplasm and beneath the surface membrane. Sections of short haptopodia (presumed to be in formation) show filamentous structures associated with the haptosome membrane, but not yet in the papilla at the haptopodium base, suggesting that the haptopodial anchor differentiates after the shaft formation. The spicule-like, striated structures visible in sections of the large vacuoles are believed to correspond to the prominent refractile cytoplasmic inclusions seen by light microscopy. Each spicule is inserted into a short flat-ended diverticulum of the vacuolar membrane, sometimes at both ends. The thickened membrane surrounding the vacuole is taken to be of Golgi origin. The last cytoplasmic inclusion of note is a single branched organelle with thin (6 nm) bounding membrane and occasional tubular structures faintly visible in its electron-dense matrix. It appears to be loosely wrapped around the nucleus and will be referred to as the paranuclear organelle. Sections of candidate cysts (not shown) reveal a thin (~100 nm) homogeneous wall without pores. Around the nucleus lie persistent basal bodies, mitochondria enveloped by granular endoplasmic reticulum, Golgi and paranuclear organelle; remnants of haptopodia and spicules are not evident. (ref. ID; 6794)

We isolated one culutre from grassland at the edge of Listvyanka village, 0.5 km from the Lake Baikal shore (Siberia), and another from volcanic soil from Costa Rica, with an extremely similar 18S rDNA sequence to the three A. soils strains of Vickerman et al. (2005). Our strain AurBaikal (GenBank HQ176323) is available as CCAP 1909/1. Our observations confirm and supplement those of Vickerman et al. (2005): cell size 8.2 um (6-10 um); shorter AF ca. 4 um difficult observe, often curved; PF 38.2 um (32-48 um). PF flickers rapidly along its length occasionally, while tethering cell body to substratum by its tip. Cell spherical or slightly elongated; radiating filose spine-like haptopodia all over periphery, 1.5-8.5 um long (mean 4.1 um), usually slightly bulbous at distal tips. Much variation in cell and haptopodia length. Pseudopodia not observed. Cell and PF often unattached to substratum, PF movement makes cell rapidly yo-yo up and down in medium; occasionally its distal end tethers cell to substratum. Culture medium Volvic and grain; grows best in company of other eukaryotes (probably prey; feeding not observed). Cv at anterior end of cell where flagella emerge. Globular cysts readily formed. (ref. ID; 7130)

Etymology

Latin solis, of the sun; in Greek legend the sun was drawn across the heavens by a charioteer. (ref. ID; 6794)