Toxic Characteristics and Action Mode of the Mixotrophic Dinoflagellate Akashiwo sanguinea on Co-Occurring Phytoplankton and Zooplankton

The mixotrophic dinoflagellate Akashiwo sanguinea frequently forms harmful algal blooms around the world and has caused massive deaths of shellfish, finfish and birds, yet its toxic mechanism is still unclear. In this study, toxic effects of A. sanguinea on co-culturing phytoplankton and zooplankton were investigated. The results showed that sonicated cultures of A. sanguinea JX13 and JX14, isolated from the Pearl River Estuary, had a significant lethal effect on the rotifer Brachionus plicatilis, with the highest mortality rate of 80%. The highest inhibition rates of A. sanguinea cultures JX13 (90%) and JX14 (80%) on R. salina were much higher than that of AS2 (20%). Toxicity varied with the growth stage, during which A. sanguinea cells in the exponential stage showed the highest toxicity (40%), while A. sanguinea filtrate had the highest toxicity (10%) in the decline stage. The action mode of A. sanguinea toxicity on plankton was explored through an osmotic membrane culture device. It was found that A. sanguinea JX13 displayed an inhibitory effect on coexisting phytoplankton, whether they had contact or not, but the inhibition rate increased by 25% with contact. A lethal effect of A. sanguinea JX13 on rotifer Brachionus plicatilis was observed only in contact treatment. This study suggests that direct contact is the key action mode to trigger the release of toxins and induce toxic effects of A. sanguinea on co-occurring plankton.

Integrative Taxonomy of Two Peruvian Strains of Brachionus plicatilis Complex with Potential in Aquaculture

Two Peruvian strains of the genus Brachionus were isolated from impacted coastal wetlands. With an integrative taxonomic view, we described their taxonomic status, morphological characters, productive parameters, and phylogenetic position. In the case of both strains, the relationship between biometrics and productive parameters obtained with Principal Components Analysis indicated that the lorica length was associated with longevity, progeny, egg production, and reproductive age, while the lorica width and aperture were associated with the maximum number of eggs carried. Maximum Likelihood and Bayesian Inference analysis carried out with mtDNA COI gene and rDNA ITS1 region showed that both strains were clustered in two clades with distinct phylogenetic positioning from what is currently known for Brachionus plicatilis s.l. One of the strains, Z010-VL, is proposed to be a subspecies of L4 (B. paranguensis), and the other strain, Z018-SD, is proposed as a sub species of SM2 (B. koreanus). In addition, 33 and 31 aquaculture production lineages are proposed, delimited by COI and concatenated COI+ITS1 sequences, respectively. Finally, this study provides new tools that enhance the traceability of the origin of each sub-species throughout the world.

Novel Non-paralytic Shellfish Toxin and Non-spirolide Toxicity to Finfish, Brine Shrimp, and Rotifer Observed in a Culture of the Dinoflagellate Alexandrium insuetum Isolated From the Coastal Water of China

The genus Alexandrium is one of the major harmful algal blooms (HABs)-forming dinoflagellate group and at least half of ~40 described species have been reported to produce paralytic shellfish toxins (PSTs). The potentially harmful species Alexandrium insuetum has been reported from many countries of Asia and Europe, and to have paralytic shellfish poisoning toxicity, but no mortality of marine animals was observed during its bloom. Therefore, it is ecologically important to characterize the possible toxicity and toxins of this organism. In this study, based on the establishment of two clonal cultures through cyst germination collected from the Yellow Sea, we identified A. insuetum from China as the first record via light microscopy (LM) and scanning electron microscopy (SEM) observations and phylogenetic analyses. The cultures of A. insuetum were further observed to be toxic to finfish and zooplankton and deleterious to rotifer eggs via laboratory bioassays. The exposure bioassays using rotifer (Brachionus plicatilis), brine shrimp (Artemia salina), and larval finfish (Oryzias melastigma) demonstrated that A. insuetum caused significant lethal effects on finfish and zooplankton species. Rotifer bioassays using cell-free culture medium, heat-treated cultures, and water, methanol, and trichloromethane extracts of algal cells revealed that A. insuetum produced heat-labile, water-soluble toxin(s) that could be excreted from A. insuetum cells and steadily accumulated in the medium during the growth phases. Hatching success of rotifer eggs was also found to be seriously affected by the exposure to A. insuetum. Importantly, ultra-high performance liquid chromatography-tandem mass spectrometry [UPLC (or LC)-MS-MS] analyses suggest the above-described toxicity of A. insuetum was caused by neither PSTs nor spiroimines (13-desmethyl spirolide C and gymnodimine). Collectively, our findings demonstrated the novel toxicity to finfish and zooplankton in A. insuetum, which is ecologically important in not only possibly contributing to population dynamics and even the formation of HABs of the species, but also affecting the on-the-spot survival and the reproduction potency of marine animals. The present work is believed to set a cornerstone for the monitoring and risk assessment of the species along the coastal waters of China and for understanding the general ecology of A. insuetum.

Ammonia Bioremediation from Aquaculture Wastewater Effluents Using Arthrospira platensis NIOF17/003: Impact of Biodiesel Residue and Potential of Ammonia-Loaded Biomass as Rotifer Feed

The present work evaluated the capability of Arthrospira platensis complete biomass (ACDW) and the lipid-free biomass (LFB) to remove ammonium ions (NH4+) from aquaculture wastewater discharge. Under controlled conditions in flasks filled with 100 mL of distilled water (synthetic aqueous solution), a batch process ion-exchange was conducted by changing the main parameters including contact times (15, 30, 45, 60, 120, and 180 min), initial ammonium ion concentrations (10, 20, 30, 40, 50, and 100 mg·L−1), and initial pH levels (2, 4, 6, 8, and 10) at various dosages of ACDW and LFB as adsorbents (0.02, 0.04, 0.06, 0.08, and 0.1 g). After lab optimization, ammonia removal from real aquaculture wastewater was also examined. The removal of ammonium using ACDW and LFB in the synthetic aqueous solution (64.24% and 89.68%, respectively) was higher than that of the real aquaculture effluents (25.70% and 37.80%, respectively). The data of IR and Raman spectroscopy confirmed the existence of various functional groups in the biomass of ACDW and LFB. The adsorption equilibrium isotherms were estimated using Freundlich, Langmuir, and Halsey models, providing an initial description of the ammonia elimination capacity of A. platensis. The experimental kinetic study was suitably fit by a pseudo-second-order equation. On the other hand, as a result of the treatment of real aquaculture wastewater (RAW) using LFB and ACDW, the bacterial counts of the LFB, ACDW, ACDW-RAW, and RAW groups were high (higher than 300 CFU), while the LFB-RAW group showed lower than 100 CFU. The current study is the first work reporting the potential of ammonia-loaded microalgae biomass as a feed source for the rotifer (Brachionus plicatilis). In general, our findings concluded that B. plicatilis was sensitive to A. platensis biomass loaded with ammonia concentrations. Overall, the results in this work showed that the biomass of A. platensis is a promising candidate for removing ammonia from aquaculture wastewater.