Utilization of urea and expression profiles of related genes in the dinoflagellate <i>Prorocentrum donghaiense</i>

<div><p>Urea has been shown to contribute more than half of total nitrogen (N) required by phytoplankton in some estuaries and coastal waters and to provide a substantial portion of the N demand for many harmful algal blooms (HABs) of dinoflagellates. In this study, we investigated the physiological and transcriptional responses in <i>Prorocentrum donghaiense</i> to changes in nitrate and urea availability. We found that this species could efficiently utilize urea as sole N source and achieve comparable growth rate and photosynthesis capability as it did under nitrate. These physiological parameters were markedly lower in cultures grown under nitrate- or urea-limited conditions. <i>P</i>. <i>donghaiense</i> N content was similarly low under nitrate- or urea-limited culture condition, but was markedly higher under urea-replete condition than under nitrate-replete condition. Carbon (C) content was consistently elevated under N-limited condition. Consequently, the C:N ratio was as high as 21:1 under nitrate- or urea-limitation, but 7:1 under urea-replete condition and 9:1 to 10:1 under nitrate-replete condition. Using quantitative reverse transcription PCR, we investigated the expression pattern for four genes involved in N transport and assimilation. The results indicated that genes encoding nitrate transport, urea hydrolysis, and nickel transporter gene were sensitive to changes in general N nutrient availability whereas the urea transporter gene responded much more strongly to changes in urea concentration. Taken together, our study shows the high bioavailability of urea, its impact on C:N stoichiometry, and the sensitivity of urea transporter gene expression to urea availability.</p></div

Information of primers and thermal cycling conditions used in RT-qPCRs.

<p>Information of primers and thermal cycling conditions used in RT-qPCRs.</p

Cellular nitrogen and carbon contents of <i>P</i>. <i>donghaiense</i> grown under the N-replete and N-deprived conditions.

<p>Cellular nitrogen and carbon contents of <i>P</i>. <i>donghaiense</i> grown under the N-replete and N-deprived conditions.</p

N-nutrient concentrations in the media in the first 6 days of experimental period.

<p>N-nutrient concentrations in the media in the first 6 days of experimental period.</p

Transcriptional levels of N transporter and assimilation genes normalized to <i>calmodulin</i> (<i>Pdcalm</i>) in <i>Prorocentrum donghaiense</i> grown under nitrogen replete and deprived conditions.

<p>(A) Nitrate transporter (<i>PdNRT</i>). (B) Urea transporter (<i>PdUT</i>). (C) Urease (<i>PdURE</i>). (D) High-affinity nickel transporter (<i>PdNiT</i>). Error bars indicate ± SD of biological triplicates. Significant differences (<i>p</i> < 0.05) between experiment groups and control (nitrate-replete) are indicated by an asterisk (*).</p

Seasonal nitrogen concentrations and nitrate isotopes of a New England River from 2018 to 2019

Data collection occurred in four parts during 2018 to 2019 for the Pawcatuck River: weekly collection from the Stillman and Westerly Bridges in Westerly, RI; collections were also taken seasonally from various bridges in a transect from head of the Pawcatuck River at Warden Pond to Westerly, RI; rain water was collected at UConn Avery Point - Groton, CT; and wastewater data reported by Westerly Wastewater Facility (which was corroborated in house at UConn Avery Point). Standard data collected was nutrient concentrations of nitrate, nitrite, ammonium, and phosphate. Total dissolved nitrogen, particulate nitrogen, and chlorophyll-a were also collected and measured. Our study utilized stable isotopes of nitrate and particulate nitrogen with the intent of tracking sources, cycling, and loading along the river. We focused on δ15N-NO3, δ18O-NO3, δ17O-NO3, and δ15N-PN. Through collection of rainwater at UConn Avery Point, percent atmospheric deposition of river samples based on the mass independent fractionation between δ17O and δ18O was calculated. Loading was calculated for each nutrient source based on collected data and river discharge reported from the USGS