Titanospirillum velox: a huge, speedy, sulfur-storing spirillum from Ebro Delta microbial mats

Abstract

A long (20-30 micrometer), wide (3-5 micrometer) microbial-mat bacterium from the Ebro Delta (Tarragona, Spain) was grown in mixed culture and videographed live. Intracellular elemental sulfur globules and unique cell termini were observed in scanning-electron-microprobe and transmission-electron micrographs. A polar organelle underlies bundles of greater than 60 flagella at each indented terminus. These Gram-negative bacteria bend, flex, and swim in a spiral fashion; they translate at speeds greater than 10 body lengths per second. The large size of the spirillum permits direct observation of cell motility in single individual bacteria. After desiccation (i.e., absence of standing water for at least 24 h), large populations developed in mat samples remoistened with sea water. Ultrastructural observations reveal abundant large sulfur globules irregularly distributed in the cytoplasm. A multilayered cell wall, pliable and elastic yet rigid, distends around the sulfur globules. Details of the wall, multiflagellated termini, and large cytoplasmic sulfur globules indicate that these fast-moving spirilla are distinctive enough to warrant a genus and species designation: Titanospirillum velox genus nov., sp. nov. The same collection techniques at a similar habitat in the United States (Plum Island, northeast Essex County, Massachusetts) also yielded large populations of the bacterium among purple phototrophic and other inhabitants of sulfurous microbial-mat muds. The months-long survival of T. velox from Spain and from the United States in closed jars filled with mud taken from both localities leads us to infer that this large spirillum has a cosmopolitan distribution.

Figure 1

(a) Mat surface at the Ebro Delta field site (3) showing lack of standing water. (Bar = 10 cm.) (b) Two spirilla cells (S, sulfur globule) shown by differential interference contrast (Nomarski). (Bar = 5 μm.) (c) Phase contrast microscopy of live spirillum cells. (Bar = 5 μm.) (d) Bipolar lophotrichous large spirillum in which only one pole has retained flagella. Sulfur globules are visible through the cell wall (scanning electron micrograph). (Bar = 5 μm.) (e) Negative-stain transmission electron micrograph of an entire bipolar lophotrichous large spirillum showing flagella “braids” (double arrowheads) compared with standard-sized spirilla (single arrowhead). (Bar = 5 μm.) (f) This scanning-electron micrograph of a cell terminus shows one vaulted end with residual flagella. The indentation coated by the polar organelle (P; see Fig. 2) is implied. (Bar = 0.5 μm.) (g) This Gram-stain brightfield preparation compares the two size classes, huge and standard, of Gram-negative spirilla. (Bar = 5 μm.) (h) Standard-sized spirillum Gram stain. The lighter spots are probably sulfur globules. (Bar = 5 μm.)

Figure 2

Transmission electron micrographs of Titanospirillum velox. (a) The polar organelle (po) underlies the indented terminus. (Bar = 1 μm.) (b) Complex cell wall (cw), flagella (f), and clear zone (cz) at the cell terminus subtended by thickening (t) and empty sulfur-globule (s) vacuoles. (Bar = 0.5 μm.) (c) Distended cell wall around a peripheral membrane-bound sulfur-globule vacuole (arrow). (Bar = 1 μm.) (d) The left (l) and right (r) polar organelles lie proximal to at least nine layers of wall material at the cell termini. (Bar = 0.25 μm.) (e) Sulfur-globule vacuoles distributed irregularly in the cytoplasm are especially abundant at the cell periphery distal to the nucleoid (n). (Bar = 1 μm.)

Figure 3

Huge spirilla rendered from both life and micrographs.