Algal endosymbiosis in brown hydra: host/symbiont specificity

Host/symbiont specificity has been investigated in non-symbiotic and aposymbiotic brown and green hydra infected with various free-living and symbiotic species and strains of Chlorella and Chlorococcum. Morphology and ultrastructure of the symbioses obtained have been compared. Aposymbiotic Swiss Hydra viridis and Japanese H. magnipapillata served as controls. In two strains of H. attenuata stable hereditary symbioses were obtained with Chlorococcum isolated from H. magnipapillata. In one strain of H. vulgaris, in H. oligactis and in aposymbiotic H. viridis chlorococci persisted for more than a week. Eight species of free-living Chlorococcum, 10 symbiotic and 10 free-living strains of Chlorella disappeared from the brown hydra within 1–2 days. In H. magnipapillata there was a graded distribution of chlorococci along the polyps. In hypostomal cells there were greater than 30 algae/cell while in endodermal cells of the mid-section or peduncle less than 10 algae/cell were found. In H. attenuata the algal distribution was irregular, there were up to five chlorocci/cell, and up to 20 cells/hydra hosted algae. In the dark most cells of Chlorococcum disappeared from H. magnipapillata and aposymbiotic hydra were obtained. Chlorococcum is thus an obligate phototroph, and host-dependent heterotrophy is not required for the preservation of a symbiosis. The few chlorococci that survived in the dark seem to belong to a less-demanding physiological strain. In variance with known Chlorella/H. viridis endosymbioses the chlorococci in H. magnipapillata and H. attenuata were tightly enveloped in the vacuolar membrane of the hosting cells with no visible perialgal space. Chlorococcum reproduced in these vacuoles and up to eight daughter cells were found within the same vacuole. We suggest that the graded or scant distribution of chlorococci in the various brown hydra, their inability to live in H. viridis and the inability of the various chlorellae to live in brown hydra are the result of differences in nutrients available to the algae in the respective hosting cells. We conclude that host/symbiont specificity and the various forms of interrelations we show in green and brown hydra with chlorococci and chlorellae are based on nutritional-ecological factors. These interrelations demonstrate successive stages in the evolution of stable obligatoric symbioses from chance encounters of preadapted potential cosymbionts.

Competition between Chlorellae in chimeric infections of Hydra viridis: the evolution of a stable symbiosis

Aposymbiotic polyps of Hydra viridis were infected with one or two of the following strains of Chlorella: the native strain obtained from green H, viridis, and the originally non-symbiotic strains Fs and 211/8p cultured in vitro. Larvae of Artemia served as infecting vectors. Chimeric infections were obtained with two different Chlorella strains cohabiting in the same cells and polyps. In time, the chimeric infections disappeared and mixed populations of Hydra were formed with different strains of Chlorella in different polyps. We suggest that the Chlorella/Hydra symbiosis was initiated originally by an infection of preadapted hydra by preadapted chlorellae. Through intracellular interalgal competition and competition between dissimilar infected cells and polyps, the present-day stable symbiosis has evolved.

Correlations between characteristics of some free-living Chlorella sp. and their ability to form stable symbioses with Hydra viridis

Aposymbiotic polyps of Hydra viridis were infected with 17 strains of in vitro cultured Chlorella sp. Larvae of Artemia fed with the chlorellae were used as an infecting vector. Of the 17 strains, seven formed stable symbioses and one formed a transient infection that disappeared within several weeks. Chlorellae of the nine other strains were cleared out of the infected hydra within 2–3 days. There was a distinct correlation between the ability of the chlorellae to form stable symbioses and their ability to adapt and grow in media enriched with 0.5% proteose peptone. Only strains that grew in the latter medium formed symbioses with the hydra. The symbioses formed with the different strains of chlorellae differed from one another. Hydra infected with some strains greened completely while those infected with other strains greened only partially. The degree of infection varied also within each population, and there were differences in the distribution of the various chlorellae along the stalk and inside the digestive cells of the hydra. Growth rates of the infected hydra were all less that those of aposymbiotic hydra or of hydra hosting native zoochlorellae. We conclude that adaptability to a nutrient-rich environment inside the perialgal vacuole of the digestive cell and a sufficient growth rate therein are crucial to the ability of chlorellae to form stable symbioses with H. viridis. In time, co-adaptation of hydra and chlorellae would restore the normal growth rate of the former and bring about regularity to the form and extent of infection by the latter.

Intracellular infection of aposymbiotic Hydra viridis by a foreign free-living Chlorella sp.: initiation of a stable symbiosis

An aposymbiotic strain of Hydra viridis became infected with free-living Chlorella sp. A stable symbiosis formed that differed in its characteristics from other known Chlorella/Hydra symbioses. The algae reproduced and formed clusters in host endodermal cells, inside large vacuoles filled with an electron-dense substance. A few algae were found to be digested by the hydra, but the apparently uncontrolled reproduction rate of the algae more than compensated for this loss. Surplus algae were expelled into the coelenteron and eventually into the surrounding medium. The expelled algae were repeatedly re-engulfed by the hydra during its feeding, forming a process of continuous reinfection. We suggest that such repeated reinfection of the hydra by the expelled algae provides the host with an endless number of Chlorella from which it might in time select a suitable adapted, controllable symbiont. The present newly formed symbiosis might serve as a model for the study of evolution of algal endosymbioses.