Media engineering in marine diatom Phaeodactylum tricornutum employing cost-effective substrates for sustainable production of high value renewables

Phaeodactylum tricornutum is a marine diatom, and well-studied model of unicellular microalga. This diatom contains a wide range of high-value renewables (HVRs) with high commercial relevance owing to their importance in human nutrition and health. In this study, we screened P. tricornutum for biomass, eicosapentaenoic acid (EPA) and fucoxanthin production under photoautotrophic and mixotrophic condition with various substrate combinations. Results highlights that culture supplemented with glycerol and urea lead to enhanced biomass, biochemical and HVR production. Further continuous feeding of urea in glycerol supplemented medium results in an increase in biomass yield (0.77 g L-1) by ~ 2-fold. Additionally, continuous feeding of urea channelizes the carbon flux towards biosynthesis of fatty acids increasing FAME content by ~2-fold as compared to the control conditions. Overall EPA and fucoxanthin production was 27 mg L-1 and 11 mg L-1 (~2 & 4 fold) in urea fed cultures respectively. Present study demonstrates efficient valorization of cost-effective substrates such as glycerol and urea for the production of high-value renewables in P. tricornutum.

Comparative Study Highlights the Potential of Spectral Deconvolution for Fucoxanthin Screening in Live Phaeodactylum tricornutum Cultures

Microalgal biotechnology shows considerable promise as a sustainable contributor to a broad range of industrial avenues. The field is however limited by processing methods that have commonly hindered the progress of high throughput screening, and consequently development of improved microalgal strains. We tested various microplate reader and flow cytometer methods for monitoring the commercially relevant pigment fucoxanthin in the marine diatom Phaeodactylum tricornutum. Based on accuracy and flexibility, we chose one described previously to adapt to live culture samples using a microplate reader and achieved a high correlation to HPLC (R2 = 0.849), effectively removing the need for solvent extraction. This was achieved by using new absorbance spectra inputs, reducing the detectable pigment library and changing pathlength values for the spectral deconvolution method in microplate reader format. Adaptation to 384-well microplates and removal of the need to equalize cultures by density further increased the screening rate. This work is of primary interest to projects requiring detection of biological pigments, and could theoretically be extended to other organisms and pigments of interest, improving the viability of microalgae biotechnology as a contributor to sustainable industry.

Response of Chlorophyll Fluorescence Characteristics and Dissolved Organic Matter for Marine Diatom Skeletonema dohrnii under Stress from Penicillin and Zn2+

Skeletonema dohrnii is a good model diatom for studying environmental stress and has promising applications and prospects in various fields. Antibiotics and heavy metals are commonly exceeded in the nearshore marine habitats. In this work, we investigated the effects of an antibiotic (penicillin, 2 µg/L) and a heavy metal ion (Zn2+, 10 µmol/L) stress on marine diatom S. dohrnii, mainly using excitation-emission matrices (EEMs) fluorescence methods and OJIP test. Results indicated that algal cells grown with the antibiotic showed higher biomass, specific growth rate, doubling time, chlorophyll a, and chlorophyll fluorescence variables. Moreover, excess zinc had negative effects on S. dohrnii. We found that zinc not only inhibited the relative photosynthetic electron transfer efficiency but also reduced the Chl a content, which ultimately affected algal growth and organic matter production. In addition, the combined effect of penicillin and Zn2+ further affected the physiological state of S. dohrnii. The dissolved organic matter (DOM) characteristics of the four cultures were also different, including fluorescence indices (fluorescence index, biological index, β/α, and humification index) and fluorescence peaks (peaks A, C, M and T). In brief, characterization of chlorophyll fluorescence characteristics and DOM-related variables are important for understanding the effects of environmental stress on microalgae.

Quorum Sensing Regulation of Virulence Gene Expression in Vibrio harveyi during its Interaction with Marine Diatom Skeletonema marinoi

Communication between species from different kingdoms may be as important as intra-kingdom communication. It has recently been confirmed that co-existing bacteria and phytoplankton in aquatic ecosystems do cross-talk. This study examined the signs of possible cross signalling between V. harveyi, one of the predominant bacterial species of the marine ecosystem and a dominant diatom species, S.marinoi, to understand communication over species borders. It is known that V.harveyi employ quorum sensing for cell-to-cell communication, bioluminescence (luxR), and the regulation of the virulence gene (vhp, chiA). Former studies have also shown, this kind of interactions being disrupted by compounds secreted by a few algal species existing in the aquatic ecosystem. We investigated the QS communication by quantifying the expression levels of virulence regulator luxR and virulence factors metalloprotease (vhp) and chitinase (chiA) in four different V. harveyi strains grown in the presence of S. marinoi strain. Results obtained in this study indicate that quorum sensing was activated in strains of V. harveyi analysed but did not regulate the expressions of vhp and chiA virulence factors. This observation suggests that the existence of S. marinoi did not interfere with the QS behaviour of V. harveyi and its interaction with marine diatom; it may be due to the commensalism relationship.

Chlorophyll fluorescence as a light signal enhances iron uptake by the marine diatom Phaeodactylum tricornutum under high-cell density conditions

Background Diatoms usually dominate phytoplankton blooms in open oceans, exhibiting extremely high population densities. Although the iron uptake rate of diatoms largely determines the magnitude and longevity of diatom blooms, the underlying mechanisms regulating iron uptake remain unclear. Results The transcription of two iron uptake proteins, ISIP2a and ISIP1, in the marine diatom Phaeodactylum tricornutum was enhanced with increasing cell density, whereas the cellular iron content showed the opposite trend. When compared with the wild-type strain, knockdown of ISIP2a resulted in 43% decrease in cellular iron content, implying the involvement of ISIP2a in iron uptake under high-cell density conditions. Incubation of the diatom cells with sonicated cell lysate conditioned by different cell densities did not affect ISIP2a and ISIP1 expression, ruling out regulation via chemical cues. In contrast, ISIP2a and ISIP1 transcription were strongly induced by red light. Besides, chlorophyll fluorescence excited from the blue light was also positively correlated with population density. Subsequently, a “sandwich” illumination incubator was designed to filter out stray light and ensure that the inner layer cells only receive the emitted chlorophyll fluorescence from outer layers, and the results showed that the increase in outer cell density significantly elevated ISIP2a and ISIP1 transcription in inner layer cells. In situ evidence from Tara oceans also showed positively correlated between diatom ISIP transcripts and chlorophyll content. Conclusions This study shows that chlorophyll fluorescence derived from neighboring cells is able to upregulate ISIP2a and ISIP1 expression to facilitate iron assimilation under high-cell density. These results provide novel insights into biotic signal sensing in phytoplankton, which can help to elucidate the underlying mechanisms of marine diatom blooms.