| Karlodinium veneficum,a cosmopolitan,toxic,and harmful algal bloom-forming dinoflagellate,has caused numerous blooms that killed massive fish by production of karlotoxins in many places of the world.The adaption to various environments and rapid reproduction of K.veneficum are attributed to its multiple survival strategies.This dissertation investigated three important survival strategies of K.veneficum through multiple approaches:toxicity to animals,allelopathy to co-cultured phytoplankton,and phagotrophy,to explore their mechanism and ecological implication.We examined the toxicity and the allelopathy of four strains of K.veneficum that were isolated from coastal waters of China(Beidaihe,Ningde,and the Pearl River Estuary)and the USA(Chesapeake Bay)and explored the ecological implications of toxicity and allelopathy.The toxicity bioassays were carried out by adding three species of test animals,namely,the finfish Oryzias melastigma,the brine shrimp Artemia salina,and the rotifer Brachionus plicatilis,into“whole live culture”and“sonicated culture”of the four strains of K.veneficum,respectively.The same numbers of three species of animals maintained in f/2 and Isochrysis galbana culture were set as controls,respectively.Half lethal concentrations(LC50)were calculated to compare the toxicity among strains.The 24-h ingest rates of Artemia salina and Brachionus plicatilis on K.veneficum were calculated to evaluate the effects of toxicity on the animals'ingest rates.The experiments for allelopathy were conducted by co-culturing four strains of K.veneficum with a selected target species(the naked dinoflagellate Akashiwo sanguinea,a species demonstrated being sensitive to allelopathy)in a series of density gradient.All monocultures in the same conditions and initial densities were used as controls.The parameter“Growth relative to control%(Grc)”was quantified to compare the allelopathic effects among strains and different cell densities.We also launched an investigation on the phagotrophy of K.veneficum using the clonal culture isolated from the coastal water of East China Sea(Ningde,Fujian Province).We observed the feeding behavior of K.veneficum by adding different species or types of preys,and compared the ingestion ability of K.veneficum under different preys,growth stages,and nutrient levels by staining lysosomes or/and phagolysosomes with fluorescence probes.The major findings of our study are summarized as follows:(1)Regarding the toxicity of K.veneficum,the four strains significantly varied and appeared a decreasing order as KVND-1(from Ningde,isolated in 2016)>KVBDH-1(from Beidaihe,isolated in 2014)>KVPRE-1(from the Pearl River Estuary,isolated in 2011)>KV7+8(from Chesapeake Bay,isolated in 2006),which coincided with their culturing histories(i.e.duration of maintenance as clonal cultures)in the laboratory,suggesting K.veneficum may have gradually lost toxicity during laboratory culturing.This result further indicates that karlotoxins production may be a response of K.veneficum to grazing pressure or other stressors.Moreover,we found that the ingest rates of A.salina on different strains of K.veneficum were corresponded to the strength of toxicity and that A.salina avoided feeding on more toxic strains.This phenomenon suggests that karlotoxin(s)may help K.veneficum to avoid predation by zooplankton,which consequently provides an advantage for K.veneficum in terms of the population survival.(2)As for the allelopathy of K.veneficum,all four strains of K.veneficum at high cell densities(>100,000 cells m L-1)significantly inhibited the growth of A.sanguinea to different extents,and the inhibited ability from strong to weak was in the following order,KVBDH-1>KV7+8>KVND-1,KVPRE-1.However,due to the difference of strains,co-culturing time,and the initial cell density ratios of tested species and target species,they exhibited either positive(stimulative)or negative(inhibited)allelopathy at lower densities.These results suggest that allelopathy may not play an important role in initiating blooms of K.veneficum.The order of allelopathy among K.veneficum strains were not perfectly parallel to that for the toxic potency,indicating that either the allelochemicals and toxins of K.veneficum may be different chemicals,or the same chemicals functioned in different modes.(3)As for the mixotrophy of K.veneficum,we found K.veneficum is an omnivorous phagotroph which can feed on all kinds of organisms tested,including both live and dead cells/bodies of co-cultivated microalgae Akashiwo sanguinea,Margalefidinium polykrikoides,Alexandrium leei,Rhodomonas salina,Isochrysis galbana,rotifer(Brachionus plicatilis),brine shrimp(Artemia salina),fish(Oryzias melastigma),and even its own dead species.Especially,the feeding behavior of K.veneficum on animals with sizes much larger than itself was called micropredation.Karlodinium veneficum exhibited flexible feeding modes,it can either engulf the whole cell of small cells directly(i.e.phagotrophy sensu stricto)or by sucking the cell contents of all species provided through a peduncle(i.e.myzocytosis).Karlodinium veneficum preferred to feed on preys which were non-motile or newly dead,no matter whether they were phytoplankton,zooplankton,or fish.This result suggested that K.veneficum may find and capture preys by chemoattraction.Cannibalism in K.veneficum,i.e.a cell feeds on other unhealthy or dead cells of the same species,was observed for the first time,which can contribute to the growth and long-term maintaining of the population under limited nutrient condition.The growth rate of K.veneficum exhibited significant positive correlation with the ingestion rate of K.veneficum,which varied with prey species,and the highest growth rate was observed when adding R.salina as prey.The ingest capacity of K.veneficum was stimulated by nutrient deficiency.In summary,the omnivorous mixotrophy is proposed to be a key autecological mechanism for K.veneficum to widen its ecological niche and succeed in forming a cosmopolitan distribution and frequent blooms. |
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