Picoplankton Diel Variability and Estimated Growth Rates in Epipelagic and Mesopelagic Waters of the Central Red Sea

Abstract

The diel variability of the abundance and cell size of picoplanktonic groups in the central Red Sea was monitored every 2 h in situ on 4 occasions (once per season) from 2015 to 2016. We distinguished Prochlorococcus, low (LF-Syn) and high (HF-Syn) fluorescence Synechococcus, small (Speuk) and large (Lpeuk) picoeukaryotes and two groups of heterotrophic prokaryotes of low (LNA) and high (HNA) nucleic acid content. The diel variability in abundance was less marked than in cell size and more apparent in autotrophs than heterotrophs. Specific growth rates were estimated by an empirical relationship from measurements obtained in bottle incubations of surface and deep samples collected in the winter compared with in situ variations in cell size over 24 h. Autotrophic picoplankton groups generally grew faster (0.23–0.77 d–1) than heterotrophic prokaryotes (0.12–0.50 d–1). Surface to 100 m depth-weighted specific growth rates displayed a clear seasonal pattern for Prochlorococcus, with maxima in winter (0.77 ± 0.07 d–1) and minima in fall (0.52 ± 0.07 d–1). The two groups of Synechococcus peaked in spring, with slightly higher growth rates of LF-Syn (0.57 ± 0.04 d–1) than HF-Syn (0.43 ± 0.04 d–1). Speuk and Lpeuk showed different seasonal patterns, with lower values of the former (0.27 ± 0.02 and 0.37 ± 0.04 d–1, respectively). HNA consistently outgrew LNA heterotrophic prokaryotes, with a higher growth in the epipelagic (0–200 m, 0.36 ± 0.03 d–1) than in the mesopelagic (200–700 m, 0.26 ± 0.03 d–1), while no differences were found for LNA cells (0.19 ± 0.03 d–1 and 0.17 ± 0.02 d–1, respectively). With all data pooled, the mean diel abundances of autotrophic picoplankton in the upper epipelagic and of HNA cells in the epipelagic and mesopelagic layers were significantly correlated with the specific growth rates estimated from cell size variations. Our high-resolution sampling dataset suggests that changes in growth rates underlie the noticeable seasonality of picoplankton recently described in these tropical waters.

Full text

fmars-08-752910 November 5, 2021 Time: 14:18 # 1
ORIGINAL RESEARCH
published: 08 November 2021
doi: 10.3389/fmars.2021.752910
Edited by:
Ryan Paerl,
North Carolina State University,
United States
Reviewed by:
Raffaella Casotti,
Stazione Zoologica Anton Dohrn
Napoli, Italy
Pradeep Ram Angia Sriram,
UMR 6023 Laboratoire
Microorganismes Génome Et
Environnement (LMGE), France
Olga Mangoni,
University of Naples Federico II, Italy
*Correspondence:
Najwa Al-Otaibi
naotaibi@tu.edu.sa;
najwa.otaibi@kaust.edu.sa
Xosé Anxelu G. Morán
xelu.moran@kaust.edu.sa;
xelu.moran@ieo.es
Specialty section:
This article was submitted to
Aquatic Microbiology,
a section of the journal
Frontiers in Marine Science
Received: 03 August 2021
Accepted: 18 October 2021
Published: 08 November 2021
Citation:
Al-Otaibi N, García FC and
Morán XAG (2021) Picoplankton Diel
Variability and Estimated Growth
Rates in Epipelagic and Mesopelagic
Waters of the Central Red Sea.
Front. Mar. Sci. 8:752910.
doi: 10.3389/fmars.2021.752910
Picoplankton Diel Variability and
Estimated Growth Rates in
Epipelagic and Mesopelagic Waters
of the Central Red Sea
Najwa Al-Otaibi1,2*, Francisca C. García3and Xosé Anxelu G. Morán2,4*
1Department of Biology, College of Science, Taif University, Al-Hawiya, Saudi Arabia, 2Division of Biological
and Environmental Sciences and Engineering, Red Sea Research Center (RSRC), King Abdullah University of Science
and Technology (KAUST), Thuwal, Saudi Arabia, 3Environment and Sustainability Institute, University of Exeter, Exeter,
United Kingdom, 4Centro Oceanográfico de Gijón (IEO, CSIC), Gijón, Spain
The diel variability of the abundance and cell size of picoplanktonic groups in the
central Red Sea was monitored every 2 h in situ on 4 occasions (once per season)
from 2015 to 2016. We distinguished Prochlorococcus, low (LF-Syn) and high (HF-
Syn) fluorescence Synechococcus, small (Speuk) and large (Lpeuk) picoeukaryotes and
two groups of heterotrophic prokaryotes of low (LNA) and high (HNA) nucleic acid
content. The diel variability in abundance was less marked than in cell size and more
apparent in autotrophs than heterotrophs. Specific growth rates were estimated by
an empirical relationship from measurements obtained in bottle incubations of surface
and deep samples collected in the winter compared with in situ variations in cell size
over 24 h. Autotrophic picoplankton groups generally grew faster (0.23–0.77 d−1)
than heterotrophic prokaryotes (0.12–0.50 d−1). Surface to 100 m depth-weighted
specific growth rates displayed a clear seasonal pattern for Prochlorococcus, with
maxima in winter (0.77 ±0.07 d−1) and minima in fall (0.52 ±0.07 d−1). The two
groups of Synechococcus peaked in spring, with slightly higher growth rates of LF-Syn
(0.57 ±0.04 d−1) than HF-Syn (0.43 ±0.04 d−1). Speuk and Lpeuk showed different
seasonal patterns, with lower values of the former (0.27 ±0.02 and 0.37 ±0.04 d−1,
respectively). HNA consistently outgrew LNA heterotrophic prokaryotes, with a higher
growth in the epipelagic (0–200 m, 0.36 ±0.03 d−1) than in the mesopelagic (200–700
m, 0.26 ±0.03 d−1), while no differences were found for LNA cells (0.19 ±0.03 d−1
and 0.17 ±0.02 d−1, respectively). With all data pooled, the mean diel abundances of
autotrophic picoplankton in the upper epipelagic and of HNA cells in the epipelagic and
mesopelagic layers were significantly correlated with the specific growth rates estimated
from cell size variations. Our high-resolution sampling dataset suggests that changes in
growth rates underlie the noticeable seasonality of picoplankton recently described in
these tropical waters.
Keywords: Red Sea, picoplankton, Prochlorococcus,Synechococcus, picoeukaryotes, heterotrophic bacteria,
growth rate, diel variability
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Al-Otaibi et al. Diel Variability of Picoplankton
INTRODUCTION
Picoplankton (i.e., organisms with cell sizes ranging from 0.2 to 2
µm) play an essential role in the productivity and biogeochemical
cycles of the oceans and frequently dominate in oligotrophic
regions (Olson et al., 1990a;Zubkov et al., 2000;DuRand
et al., 2001). Flow cytometry has become the routine tool for
measuring their abundance and cellular characteristics. Using
this technique, two genera of picocyanobacteria (Prochlorococcus
and Synechococcus) and at least one group of picoeukaryotes are
easily distinguished by optical properties of light scatter (related
to cell size and complexity) and fluorescence (distinguished by
the red and orange fluorescence signals due to the presence
of chlorophyll aand phycoerythrin pigments). Heterotrophic
prokaryotes are in turn almost universally made up of two
clusters of differing relative nucleic acid content: low (LNA) and
high (HNA) after specific staining (Gasol et al., 1999;Bouvier
et al., 2007).
The temporal dynamics of these picoplanktonic groups have
been mostly described at fairly broad time-scales, predominantly
seasonal studies based on monthly (e.g., Campbell et al., 1997;
DuRand et al., 2001;Malmstrom et al., 2010) or, less frequently,
weekly (e.g., Worden et al., 2004;Huete-Stauffer and Morán,
2012;Sabbagh et al., 2020) sampling intervals, typically as
part of long-term monitoring efforts. Understanding the key
biological factors (i.e., growth and mortality rates) driving the
changes in picoplankton populations abundance on a time scale
of hours requires a high-frequency sampling. So far, only a
few studies have investigated picoplankton communities with
a higher resolution of multiple samples per day, including
the equatorial Pacific (Durand and Olson, 1996;Landry et al.,
1996;Blanchot et al., 1997;Vaulot and Marie, 1999), the
north Atlantic (Olson et al., 1990b;Sosik et al., 2003;Hunter-
Cevera et al., 2014) and the Mediterranean Sea (Jacquet et al.,
1998, 2002). Most of them have revealed clear diel changes
in Prochlorococcus and Synechococcus abundance, with typically
maxima in the late afternoon or early night accompanied by an
inverse pattern in cell size (Binder and DuRand, 2002;Lefort
and Gasol, 2014), despite considerable variability exists among
depths and sampling locations. Although these two variables
are undoubtedly connected through synchronized cell division
(Durand and Olson, 1996;Vaulot and Marie, 1999), and would be
useful for calculating growth rates in the absence of loss processes,
in situ changes in cell abundance alone do not allow for an
accurate estimation.
Microbial plankton growth rates are commonly measured
from short-term production experiments involving the use of
radiolabeled substrates and standing stocks (Ducklow, 2000).
Another approach is to measure the changes in abundance using
experimental setups, either in the presence of the entire microbial
community (i.e., including protistan grazers and viruses),
yielding estimates of the net growth rate, or after diluting with
organism-free water and/or pre-filtering through appropriate
pore-size filters for minimizing grazing pressure in order to
estimate the specific growth rate (Landry and Hassett, 1982;
Kirchman, 2016). Recently, a model-based approach has been
used to obtain independent, high-resolution estimates of in situ
growth rates from continuous measurements of Synechococcus
cell size over the diel cycle (Sosik et al., 2003;Hunter-Cevera
et al., 2014). In open ocean tropical and subtropical waters,
picoplankton growth rates vary widely depending on the taxa
and environmental conditions, but also the approaches used.
For instance, Prochlorococcus specific growth rates ranged from
0.32 to 0.76 d−1in the tropical NE Atlantic Ocean (Partensky
et al., 1996;Quevedo and Anadón, 2001;Worden and Binder,
2003), but increased to 1.58 d−1in the tropical central Atlantic
(Agawin and Agustí, 2005). In contrast, Prochlorococcus growth
rates were reportedly lower in the north Pacific Ocean with an
average of 0.29 ±0.18 d−1(Selph et al., 2005) and increasing
up to 1.27 d−1in the upwelling region (Taniguchi et al.,
2014;Selph et al., 2016). The growth rate of Synechococcus
populations were within the same range of Prochlorococcus
in the NE Atlantic Ocean (Quevedo and Anadón, 2001), but
higher growth rates have been reported in the tropical Pacific
Ocean (1.52 d−1;Taniguchi et al., 2014;Selph et al., 2016).
Picoeukaryotes specific growth rates usually vary more than
those of cyanobacteria, with an overall range of 0.1 to >1.6 d−1
(Landry et al., 2003;Selph et al., 2005;Taniguchi et al., 2014).
Compared with picophytoplankton groups, the specific growth
rates of heterotrophic prokaryotes in the upper layers of low
latitude waters are usually lower (Kirchman, 2016). Very low
values have occasionally been measured (ca. 0.03 to 0.08 d−1) in
the western North Atlantic Subtropical waters (Steinberg et al.,
2001), but in the absence of predators they may reach up to 1.0
d−1(e.g., in the Atlantic Ocean, Gasol et al., 2002). The review
by Kirchman (2016) provides a mean growth rate of 1.10 ±0.83
d−1for heterotrophic prokaryotes, derived from seawater culture
experiments conducted over a large geographical scale. Although
the above-mentioned values refer to the average heterotrophic
bacteria, the LNA and HNA flow cytometric groups typically
present different growth rates, with higher values of the former
(Lebaron et al., 2001;Longnecker et al., 2005;Morán et al., 2007;
Wang et al., 2009).
The Red Sea offers a good opportunity to explore the fine-scale
dynamics of picoplankton communities in tropical waters. Its
highly saline oligotrophic waters are characterized by the highest
surface temperatures of any deep marine basin of the world
(even exceeding 35◦C in summer, Rasul et al., 2015;Chaidez
et al., 2017). The overall low primary productivity becomes more
marked in the central region (Raitsos et al., 2013). The abundance
of auto- and heterotrophic picoplankton are seemingly lower
than in other tropical ecosystems (Al-Otaibi et al., 2020), even
in shallow waters subject to human influence (Silva et al., 2019;
Sabbagh et al., 2020), likely caused by strong top-down control
(Sabbagh et al., 2020). However, our knowledge about the Red
Sea microbial plankton in situ diel dynamics is limited and the
growth rates of picoplankton remain largely unknown for this
marine basin, except only a few recent studies targeting the
specific growth rates of heterotrophic bacteria and archaea from
incubation bottles (Calleja et al., 2018;Silva et al., 2019) and the
net growth rates of phytoplankton along the eastern Red Sea from
isotopic labeling method (López-Sandoval et al., 2021).
Here, we provide the first high-resolution study of the diel
abundance and cell size of auto- and heterotrophic picoplankton
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Al-Otaibi et al. Diel Variability of Picoplankton
in the central Red Sea covering each of the four seasons. We
performed flow cytometric analysis of samples taken at 2 h
intervals during 24 h throughout the water column of a 700 m
deep station. The objectives were: (i) to examine any systematic
diel changes in picoplankton cell abundance and size in order to
allow for in situ growth rates estimation in the epipelagic (0–200
m) and mesopelagic layers (200–700 m), and (ii) to assess the
relationship between picoplankton growth rates and the observed
abundances over the seasonal cycle.
MATERIALS AND METHODS
Sampling
Sampling took place at a station located 6 km offshore to
the northwest of King Abdullah Economic City (KAEC) in
Saudi Arabia (latitude 22.46◦N, longitude 39.02◦E), on board of
RV Thuwal. Four high-frequency diel surveys were performed
covering the different seasons on a purely astronomical basis:
Winter (March 6t–7th, 2016), Spring (March 24–25th, 2015),
Summer (September 5–6th, 2015) and Fall (October 26–27th,
2016). Although the lag between the winter and spring samplings
(in different years) was of only 3 weeks due to logistic constraints,
we preferred to keep the seasonal references since the temporal
variations in oceanographic and planktonic variables over the
annual cycle at the study site were remarkable (Al-Otaibi et al.,
2020). For instance, although our samplings could also be
grouped as Winter-Spring and Summer-Fall, neither temperature
nor salinity did overlap in Winter and Spring (Figure 1).
Each time, sampling started at noon and 13 CTD casts were
made in total with a 2 h window. At each cast, seawater was
collected from 12 to 16 depths ranging from 5 to 700 m
with Niskin bottles attached to a rosette system equipped with
either a SeaBird SB9 Plus or an Idronaut 305 CTD for the
in situ measurement of temperature and salinity (Supplementary
Figure 1A). Chlorophyll aconcentration (Chl a) was obtained
after filtering 200 ml samples through Whatman GF/F glass fiber
filters (25 mm diameter, 0.7 µm nominal pore-size) in the 2015
cruises and sequentially through membrane filters of 20, 2, and
0.2 µm pore-size (IsoporeTM, RTTP, 47 mm diameter) in 2016.
The 3 size-fractions (micro-, nano- and picophytoplankton) were
summed to obtain total Chl aconcentration in the Summer and
Fall cruises. Filters were frozen at –80◦C until analysis in the
laboratory as described in detail in Al-Otaibi et al. (2020).
Flow Cytometric Analyses
Seawater samples (1.8 mL) were fixed with 1% paraformaldehyde
and 0.05% glutaraldehyde for estimating the abundance and
cellular characteristics of picoplankton communities, placed in
liquid nitrogen and stored at –80◦C until analysis. After thawing,
aliquots of 600 µL for picophytoplankton and 400 µL for
heterotrophic prokaryotes were usually run in a BD FACSCanto
II flow cytometer. Before analysis, heterotrophic prokaryotes
were stained with 2.5 µmol L−1of the DNA fluorochrome SYBR
Green II (Gasol and Moraìn, 2015). Picoplankton abundances
were estimated based on analysis time and the actual flow
rates, which were measured daily. Molecular Probes fluorescent
latex beads of 1 µm were added to each sample as an internal
standard for size and fluorescence measurements, according
to Calvo-Díaz and Morán (2006). Full details about the
FACSCanto settings can be found in Al-Otaibi et al. (2020).
The cytograms were analyzed using FCSExpress 5 software
in order to distinguish picocyanobacteria (Prochlorococcus and
Synechococcus) and picoeukaryotes based on their orange and
red fluorescence and light scatter at 90◦or side scatter (SSC)
signals. Two groups of heterotrophic prokaryotes were separated
based on their relative green fluorescence signal into low (LNA)
and high (HNA) nucleic acid content. An empirical calibration
between relative SSC and cell diameter was used to determine
the cell size of picoplankton groups, as previously described in
Calvo-Díaz and Morán (2006). Spherical shape was assumed for
estimating the biovolume for all the groups.
Experimental Incubations for Estimating
Specific Growth Rates From Cell Size
Changes
During the winter sampling, 10 L seawater was collected from
the surface (5 m) and from 550 m depth (i.e., the depth with the
strongest acoustic signal of the deep scattering layer formed by
mesopelagic fish during light hours, Calleja et al., 2018;Morán
et al., 2022) at midnight (Supplementary Figure 1B). Water
was pre-filtered through pre-combusted Whatman GF/C filters
(1.2 µm) in order to remove protistan grazers. A total of 3
bottles with 2 L each were incubated for 8 days in Percival
Twin Chamber I-22LLVL vertical incubators, which mimicked
the in situ temperature (±0.1◦C, as continuously recorded
by iBWetLand 22L, AlphaMach temperature loggers) and light
regimes (white fluorescent lights providing ca. 115 µmol photons
m−2s−1measured by a LI-COR Biosciences photosynthetically
active radiation sensor, in a 11:13 h light:dark for the surface
sample and in darkness for the 550 m depth one). Two replicates
of 1.8 mL were sampled 1–2 times per day and preserved in
1% paraformaldehyde and 0.05% glutaraldehyde until analysis
by flow cytometry as previously described. After plotting the
dynamics of each group during the incubation, the specific
growth rate was calculated as the slope of the ln-transformed
abundance vs. time for the first period of exponential growth,
which averaged 2.7 days (from 1.5 days for Prochlorococcus
and the two groups of Synechococcus to 6.3 days for LNA
prokaryotes from 550 m). Although viruses were not removed
in this experimental setup, these relatively short periods likely
precluded a major role of viral infections in the different groups’
growth dynamics. Our specific growth rate estimates should be
considered conservative in that case. The coefficients of variation
of the cell size (CV, %) measured over the full 24 h in situ
were then compared with the estimated specific growth rates of
Prochlorococcus, low (LF-Syn) and high (HF-Syn) phycoerythrin
fluorescence of Synechococcus, small picoeukaryotes, LNA and
HNA prokaryotes derived from the incubations.
Statistical Analysis
Mean values of cell abundance and size of each picoplankton
group were calculated for the first 12 sampling points along each
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FIGURE 1 | Environmental properties measured every 2 h at the surface of the study station in the four diel cycles. (A) temperature (◦C), (B) salinity, and (C)
chlorophyll aconcentration (µg L−1). The gray shading represents nighttime for each sampling.
diel cycle (the 13th, corresponding again to noon, was excluded).
The coefficient of variation of the mean (CV, %) was used to
assess the diel variability of both variables (abundance and size)
for each depth. Depth-weighted averages (from 0 to 100 m for
autotrophs and from 0 to 200 m and from 200 to ca. 700 m for
heterotrophs) were calculated by the trapezoidal method. Since
normal data distribution and homoscedasticity were usually not
met, a Kruskal-Wallis analysis of variance was employed to detect
significant differences in abundance and cell size between the
different seasons. The multiple comparison Dunn’s test was used
to determine the cases with significant differences. Wilcoxon
signed rank tests were used to determine whether mean day and
night values were significantly different in the vertical profiles
of abundance and cell size. All tests were performed with the
OriginPro software. The linear regression between the changes
in cell size along the 24 h and the estimated specific growth
rates from the winter experiment was performed according to the
ordinary least-squares (OLS) or Model I method, after checking
for normality of both datasets (Shapiro-Wilk goodness of fit,
p= 0.21 for cell size and p= 0.14 for specific growth rates).
RESULTS
Diel Variations in Environmental
Variables, Picoplankton Abundance, and
Cell Size
Diel variations in temperature, salinity and chlorophyll a(Chl
a) concentration at the surface of the study site during the four
periods are shown in Figure 1. Seasonal differences were more
conspicuous for temperature, which ranged from 26.3 ±0.1◦C
(SD) in spring to 31.5 ±0.2◦C in summer, while low variability
was found between the day and night values (CVs = 0.3–
0.7%, Figure 1A). Surface salinity displayed lower values in
winter (38.4 ±0.4), with a noticeable diel pattern, but was
more similar in the remaining periods (ca. 39.5, Figure 1B).
Chl ashowed higher variability at the diel scale (CVs = 11.0 –
49.4%), with the highest values found in summer during the
day (0.17 ±0.01 µg L−1) and mean values in the other periods
were more similar (ca. 0.10 µg L−1,Figure 1C). Seasonal
differences in these environmental variables were significant
(Supplementary Table 1,p<0.001), but differences between day
and night time were not.
Diel variations in autotrophic and heterotrophic picoplankton
abundance at the surface were not consistent (Figure 2), similarly
to the rest of the water column (see below). Prochlorococcus
mean abundance ranged from 2.34 ±0.76 ×104cells mL−1
in summer to 6.00 ±1.13 ×104cells mL−1in spring, but
showed high diel variations (CVs = 18.9–32.5%, Supplementary
Table 1 and Figures 2A–D). Synechococcus cells dominated
autotrophic picoplankton in the four cycles (56–70% of total
picophytoplankton), but a seasonal shift was found between the
two groups of differing phycoerythrin content, with dominance
of LF-Syn in winter (5.36 ±0.82 ×104cells mL−1) and
spring (4.03 ±1.63 ×104cells mL−1), while HF-Syn clearly
dominated in summer (4.26 ±1.16 ×104cells mL−1) and
fall (5.14 ±1.13 ×104cells mL−1,Figures 2E–H). The
two populations of Synechococcus tended to show similar diel
dynamics, with abundance peaking at night, but LF-Syn was
more variable than HF-Syn (CVs = 15.3–40.6% and 15.6–27.3%,
respectively) (Supplementary Table 1 and Figures 2E–H). The
abundances of the two groups of picoeukaryotes were as expected
consistently lower, with values ranging from 1.40 to 2.13 ×103
cells mL−1for Speuk and 2.24 to 3.17 ×102cells mL−1for Lpeuk,
respectively, (Figures 1I–L). Picoeukaryotes abundances showed
higher diel variations than picocyanobacteria, especially Lpeuk
(CVs 32.7–55.9%, Supplementary Table 1 and Figures 1A–L).
LNA and HNA heterotrophic prokaryotes were one order of
magnitude more abundant than cyanobacteria. LNA cells (0.65–
2.26 ×105cells mL−1) outnumbered HNA cells (0.50–1.92 ×105
cells mL−1) at the surface, but the variability of HNA cells
(25.0–36.0%) was slightly higher than LNA cells (18.9–31.8%,
Figures 2M–P).
Contrary to abundance, the cell size of surface
picocyanobacteria presented clear and consistent diel
periodicities, with biovolume increasing during the day and
decreasing early at night at each sampling period (Figures 3A–
H). The precise timing of the decrease in cell size differed slightly
between Prochlorococcus and Synechococcus (Figures 3A–
H). Overall, cell size ranged from 0.04 to 0.11 µm3for
Prochlorococcus, from 0.08 to 0.21 µm3for LF-Syn and from
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FIGURE 2 | Diel variation in picoplankton abundance at the surface of the study site in winter, spring, summer, and fall. Prochlorococcus (A–D) low (LF-Syn, open
circles) and high (HF-Syn, filled circles) fluorescence Synechococcus (E–H), small (Speuk, filled circles) and large (Lpeuk, open circles) picoeukaryotes (I–L), low
(LNA, open circles), and high (HNA, filled circles) nucleic acid content heterotrophic prokaryotes (M–P). The gray shading represents nighttime.
0.10 to 0.28 µm3for HF-Syn and (Figures 3A–H). Diel
patterns were less marked in Speuk (1.19–1.59 µm3) and Lpeuk
(3.04–4.06 µm3,Figures 3I–M) and no obvious trends were
observed either for LNA (0.02–0.03 µm3;Figures 3M–P) or
HNA heterotrophic prokaryotes (0.03–0.05 µm3;Figures 3M–
P). Considering the cell size CVs of each group and sampling
period, there was a clear difference between the low variability of
heterotrophic prokaryotes (LNA, 3.3–11.2%; HNA, 11.4–19.2%)
and the more variable picocyanobacteria (Prochlorococcus,
24.6–36.9%; LF-Syn, 18.3–36.9%; HF-Syn, 16.9–26.0%). Surface
picoeukaryotes cell size CVs were similar to those of HNA cells
(Speuk, 7.03–14.7%; Lpeuk, 9.6–18.9%, Supplementary Table 1).
Similar to the surface values presented above, the abundance
and cell size of the different groups varied considerably over
24 h throughout the different sampling depths. The vertical
distributions of mean abundance and cell size during daytime
and nighttime are presented in Supplementary Figures 2, 3,
respectively. There were no consistently, significant differences
between the two periods for either abundance or cell size,
in spite of the conspicuous diel pattern for the latter. Since
picophytoplankton virtually disappeared below 100 m, Table 1
summarizes the 0–100 m depth-weighted abundances and cell
sizes of the autotrophic groups. Even with only four samples,
autotrophic picoplankton presented clear seasonal differences as
shown in detail in Al-Otaibi et al. (2020) and (Table 1). The
coefficient of variation of cell size was systematically lower than
that of abundance for all groups (Supplementary Figures 2, 3
and Table 1). Prochlorococcus showed the highest variation in
cell size regardless of the season (CVs = 22.3–33.7%), followed
by LF-Syn (16.1–23.3%) and the lowest was 4.2–10.4% for
LNA prokaryotes (Table 1). Table 2 shows the depth-weighted
abundances and cell sizes of heterotrophic prokaryotes in the
two zones, epipelagic (0–200 m) and mesopelagic (200–700 m).
The mean abundance of LNA and HNA prokaryotes presented
different seasonality in the epipelagic, with maximum values
recorded in winter and fall for LNA and in summer and fall
for HNA (Table 2). Seasonal patterns of LNA and HNA cells
were similar in the mesopelagic layer, with lower abundances in
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FIGURE 3 | Diel variation in picoplankton cell size at the surface of the study site in winter, spring, summer, and fall. Prochlorococcus (A–D) low (LF-Syn, open
circles) and high (HF-Syn, filled circles) fluorescence Synechococcus (E–H), small (Speuk, filled circles) and large (Lpeuk, open circles) picoeukaryotes (I–L), low
(LNA, open circles), and high (HNA, filled circles) nucleic acid content heterotrophic prokaryotes (M–P). The gray shading represents nighttime.
spring and 1.4- to 2.0-fold higher in winter and fall (Table 2).
The variability of LNA and HNA cell abundances was higher
in the epipelagic than in the mesopelagic layer (Table 2). The
variations of LNA and HNA mean cell size were less marked than
those of abundance (Table 2) and did not show any conspicuous
seasonal pattern.
Microbial Groups Dynamics in the
Experimental Incubations
All autotrophic picoplankton groups were able to grow in
the winter incubation experiments. Initial abundances were
2.85 ×103cells mL−1for Prochlorococcus, 4.33 ×103cells mL−1
for LF-Syn, 3.47 ×103cells mL−1for HF-Syn and 2.85 ×102
cells mL−1for Speuk (Supplementary Figures 4A–C). Although
the initial cell concentrations were similar for most groups, their
growth curves were quite different, as well as the duration of
the exponential growth phases (Supplementary Figures 4A–
C). Prochlorococcus and LF-Syn reached a comparatively
higher carrying capacity (8.33 ×103±2.69 ×102and
1.02×104±2.19 ×103cells mL−1, respectively) than HF-Syn
(6.10 ±2.29 ×103cells mL−1), while Speuk were able to grow for
a longer period (until 4.25 days), reaching a maximum abundance
of 1.57 ±0.16 ×103cells mL−1(Supplementary Figures 4A–C).
The dynamics of the LNA and HNA heterotrophic prokaryotes
differed clearly between the surface and 550 m (Supplementary
Figures 4D,E).
Autotrophic picoplankton always grew faster than
heterotrophs, with the highest specific growth rates
corresponding to LF-Syn (0.86 ±0.03 d−1) and the lowest
to HF-Syn (0.19 ±0.03 d−1) (Supplementary Figure 5A). HNA
and LNA showed consistent differences in their specific growth
rates at both depths, with higher growth rates of the former both
at the surface (0.25 ±0.02 and 0.11 ±0.04 d−1, respectively)
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TABLE 1 | Depth-weighted (0–100 m) averages of autotrophic picoplankton abundances and cell sizes with the corresponding coefficients of variation (%) between
parentheses in winter, spring, summer, and fall.
Winter Spring Summer Fall
Abundance Prochlorococcus (104cell ml−1) 3.39 ±0.83 (27.0%) 6.35 ±1.26 (31.0%) 7.40 ±2.34 (27.3%) 5.03 ±1.17 (40.1%)
LF-Syn (104cell ml−1) 2.90 ±0.77 (61.7%) 2.34 ±1.18 (54.8%) 1.08 ±0.21 (52.7%) 1.08 ±0.30 (39.4%)
HF-Syn (104cell ml−1) 2.38 ±0.57 (33.1%) 1.74 ±0.51 (36.8%) 2.50 ±1.29 (49.6%) 2.75 ±0.97 (87.3%)
Speuk (104cell ml−1) 0.29 ±0.12 (41.1%) 0.22 ±0.77 (34.6%) 0.42 ±0.19 (38.5%) 0.35 ±0.12 (62.0%)
Lpeuk (104cell ml−1) 0.04 ±0.00 (53.7%) 0.02 ±0.00 (63.0%) 0.03 ±0.00 (47.9%) 0.06 ±0.00 (68.6%)
Cell size Prochlorococcus (µm3) 0.21 ±0.06 (33.7%) 0.21 ±0.03 (26.1%) 0.26 ±0.04 (25.1%) 0.23 ±0.03 (22.3%)
LF-Syn (µm3) 0.16 ±0.03 (23.3%) 0.10 ±0.03 (24.9%) 0.15 ±0.03 (19.6%) 0.12 ±0.02 (16.1%)
HF-Syn (µm3) 0.28 ±0.04 (15.4%) 0.36 ±0.07 (18.2%) 0.34 ±0.05 (16.7%) 0.29 ±0.04 (16.0%)
Speuk (µm3) 1.50 ±0.6 (12.4%) 1.53 ±0.2 (10.3%) 1.67 ±0.14 (9.2%) 1.37 ±0.16 (11.8%)
Lpeuk (µm3) 3.3 ±0.2 (19.0%) 3.1 ±0.5 (13.3%) 4.1 ±0.7 (19.0%) 2.7 ±0.3 (11.4%)
TABLE 2 | Depth-weighted averages of heterotrophic picoplankton abundances and cell sizes in the epipelagic (0–200 m) and mesopelagic (200–700 m) with the
corresponding coefficients of variation (%) between parentheses in winter, spring summer, and fall.
Winter Spring Summer Fall
Abundance Epipelagic (0–200 m) LNA (105cell ml−1) 1.39 ±0.27 (19.1%) 0.87 ±0.29 (44.0%) 1.03 ±0.29 (37.8%) 1.33 ±0.34 (45.0%)
HNA (105cell ml−1) 0.87 ±0.20 (20.7%) 0.72 ±0.27 (46.6%) 1.30 ±0.29 (34.0%) 1.25 ±0.29 (40.8%)
Mesopelagic (200–700 m) LNA* (105cell ml−1) 0.46 ±0.09a(14.8%) 0.25 ±0.07b(28.5%) 0.34 ±0.06a(17.3%) 0.45 ±0.06a(10.8%)
HNA* (105cell ml−1) 0.41 ±0.08a,b(14.5%) 0.29 ±0.09a(29.1%) 0.47 ±0.09a,b(16.0%) 0.57 ±0.08b(11.5%)
Cell size Epipelagic (0–200 m) LNA (µm3) 0.02 ±0.00 (4.9%) 0.02 ±0.00 (9.7%) 0.02 ±0.00 (5.9%) 0.04 ±0.00 (9.0%)
HNA (µm3) 0.05 ±0.01 (17.6%) 0.04 ±0.01 (14.4%) 0.04 ±0.01 (17.5%) 0.05 ±0.01 (11.5%)
Mesopelagic (200–700 m) LNA (µm3) 0.02 ±0.00 (4.1%) 0.02 ±0.00 (8.0%) 0.02 ±0.00 (4.3%) 0.02 ±0.00 (8.1%)
HNA (µm3) 0.04 ±0.01 (12.2%) 0.03 ±0.00 (7.6%) 0.04 ±0.01 (8.5%) 0.03 ±0.00 (8.8%)
Stars and superscript letters indicate significant differences between seasons (Kruskal-Wallis analysis and Dunn’s test; *p < 0.05).
and 550 m depth (0.17 ±0.03 and 0.05 ±0.01 d−1, respectively)
(Supplementary Figure 5B).
Specific Growth Rate Estimates
The changes in cell size of the epipelagic and mesopelagic
autotrophic and heterotrophic groups monitored in situ for 24 h
reflected their specific growth rates estimated in the concurrent
laboratory incubations of the winter sampling (Figure 4). CVs
of cell size ranged from 2.5 (for LNA cells at 550 m) to 37.0%
(for LF-Syn). The fitted linear regression between both variables
was able to explain 85% of the variance in specific growth rates.
This model was subsequently applied to estimate the specific
growth rates from the cell size CVs for each of the sampling
periods and depths.
The vertical distribution of specific growth rates resembled
that of abundance for Prochlorococcus, systematically peaking at
the deep chlorophyll maximum (DCM) with a rather invariable
value of ca. 0.80 d−1except in winter (Figure 5A). The vertical
distribution of the LF-Syn and HF-Syn specific growth rates
were less coupled to their abundance distribution (Figures 5B,C).
LF-Syn specific growth rates peaked at the depth of maximum
detection (ca. 50–70 m), with values ranging from 0.41 to 1.12
d−1(Figure 5B). The specific growth rate of HF-Syn slightly
decreased from the surface to 100 m, with values ranging
from 0.40 to 0.60 d−1and from 0.22 to 0.50 d−1, respectively
(Figure 5C). The specific growth rate of Speuk and Lpeuk did not
show clear vertical patterns, with values ranging from 0.13 to 0.43
d−1and from 0.12 to 0.64 d−1, respectively (Figures 5D,E). The
specific growth rate of LNA cells exhibited a more homogeneous
vertical distribution, with values generally lower than 0.20 d−1,
while HNA prokaryotes slightly increased from the surface
(overall range 0.28–0.45 d−1) to the DCM (0.36–0.51 d−1) and
remained quite stable within the mesopelagic layer at values of
ca. 0.25 d−1except in winter, with a higher mean value of 0.41
d−1(Figures 6A,B).
Figure 7 shows the seasonal of picophytoplankton and
heterotrophic prokaryotes specific growth rates averaged for
the different layers of the study site, the upper epipelagic (0–
100 m) for picophytoplankton and the epipelagic (0–200 m)
and mesopelagic (200–700 m) for heterotrophic prokaryotes.
Prochlorococcus cyanobacteria consistently exhibited the highest
mean specific growth rates, ranging from 0.52 ±0.07 to
0.77 ±0.07 d−1(Figure 7A). The two groups of Synechococcus
showed similar patterns with consistently higher values of LF-
Syn (from 0.38 ±0.05 to 0.57 ±0.04 d−1) than HF-Syn (from
0.37 ±0.04 to 0.43 ±0.04 d−1) (Figure 7A). Remarkably, the
two groups of picoeukaryotes distinguished according to their
size showed an inverse pattern to that of the two Synechococcus
clusters with higher values of Lpeuk (0.25 ±0.03 – 0.44 ±0.07
d−1) than Speuk (0.23 ±0.04 – 0.30 ±0.03 d−1) except
in Fall (Figure 7A). The estimated specific growth rate of
HNA prokaryotes in the epipelagic was higher (0.28 ±0.01 –
0.54 ±0.04 d−1) than in the mesopelagic layer (0.20 ±0.02
– 0.34 ±0.02 d−1), while LNA mean seasonal values were
very similar in both layers of (0.14 ±0.01 – 0.25 ±0.02
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FIGURE 4 | Relationship between the diel coefficient of variation in cell size
measured over 24 h from in situ samples and the estimated specific growth
rate from the incubation experiments conducted in winter. Pro,
Prochlorococcus; LF-Syn, low phycoerythrin fluorescence Synechococcus,
HF-Syn, high phycoerythrin fluorescence Synechococcus; Speuk, small
picoeukaryotes; LNA, low nucleic acid content; HNA, high and low nucleic
heterotrophic prokaryotes. Samples were collected from the surface (S) and
550 m depth. See details in the main text.
in the epi- and 0.13 ±0.01 – 0.21 ±0.02 d−1in the
mesopelagic) (Figure 7B).
With all epipelagic data pooled, we found a positive
relationship between the estimated specific growth rates and
the mean diel abundances of picophytoplankton (Figure 8).
The correlation between specific growth rate and mean
diel abundance was also significant for HNA heterotrophic
prokaryotes when pooling data from the epi- and mesopelagic
layers, but not for LNA cells (Figures 8B,C).
DISCUSSION
The Red Sea belongs to the vast area occupied by tropical
and nutrient-poor waters, but stands out as having significantly
warmer (Raitsos et al., 2013;Chaidez et al., 2017) and saltier
(Tesfamichael and Pauly, 2016) surface waters. However, by
being more accessible than the open Atlantic, Indian or Pacific
oceans, the oligotrophic and quasi-permanently stratified Red
Sea is an appropriate model for studying the spatio-temporal
changes in picoplankton communities, the dominant planktonic
size class in low latitude waters (Campbell et al., 1997;DuRand
et al., 2001;Agawin and Agustí, 2005). Flow cytometry allowed
us to consistently identify seven groups of picoplanktonic
organisms: five autotrophs (Prochlorococcus, two populations of
Synechococcus characterized by differing phycoerythrin content
and two differently sized clusters of picoeukaryotes) plus the
two widespread clusters of heterotrophic prokaryotes, LNA, and
HNA. The two Synechococcus groups, named as low (LF-Syn)
and high (HF-Syn) fluorescence (Al-Otaibi et al., 2020), seem
to be a general characteristic of at least the central, both in
coastal (Sabbagh et al., 2020) and offshore waters (Al-Otaibi
et al., 2020), and southern regions of the Red Sea (Veldhuis
and Kraay, 1993). They had already been reported in other
oligotrophic sites such as an Indian estuary (Mitbavkar et al.,
2012) and the subtropical North Atlantic (Taucher et al., 2018).
Although not always consistently found (Grégori et al., 2001;Liu
et al., 2014;Thyssen et al., 2014), we were also able to discern
two groups of picoeukaryotes in the Red Sea, with the small
picoeukaryotes (Speuk) group also probably more homogeneous
taxonomically than the larger (Lpeuk) one (Not et al., 2009;
Cabello et al., 2016). While the numbers of picoeukaryotes were
comparable, the abundance of single-celled cyanobacteria was
generally lower than in other warm regions (Zubkov et al., 2000;
Worden et al., 2004). Lower picoplankton abundances were even
more noticeable in the case of LNA and HNA prokaryotes,
when compared to other low latitude waters (e.g., Carlson et al.,
1996;Gundersen et al., 2001;Hale et al., 2017). Sabbagh et al.
(2020) have recently suggested that top-down control by viruses
and heterotrophic nanoflagellates was fundamental in keeping
autotrophic and heterotrophic bacterial abundances low in the
nearby shallow waters of KAUST Harbor, a process probably also
behind the low numbers at our study site (Al-Otaibi et al., 2020).
Recent studies have shown that the seasonal variability of the
diverse picoplanktonic groups was higher than anticipated for
tropical oceans and comparable to higher latitude regions (Calleja
et al., 2019;Al-Otaibi et al., 2020). However, no information
with a frequency higher than once per day was available for Red
Sea microbial plankton. The high resolution (with a total of 52
vertical profiles from the surface down to the bottom equally
distributed in each season) reached in this study has allowed us to
even estimate in situ specific growth rates, providing a dynamical
and functional representation of picoplankton across the epi- and
mesopelagic layers of the central Red Sea.
Diel changes in the abundance of picoplankton groups
are driven by the periodicity in physical (e.g., light and
temperature) and biological factors (e.g., cell division, grazing,
and viral lysis, Ribalet et al., 2015;Tsai et al., 2018). The diel
dynamics of autotrophic picoplankton abundance in oligotrophic
waters have shown typically an increase in the afternoon or
early evening and a decrease through the night until dawn,
following the typically synchronized cell division (Landry et al.,
1996;Blanchot et al., 1997;Vaulot and Marie, 1999). While
increases in autotrophic picoplankton abundance can only be
explained by cell division in the absence of strong advection,
decreases can be caused by cell death, grazing mostly caused by
heterotrophic nanoflagellates (Dolan and Šimek, 1999;Christaki
et al., 2001;Guillou et al., 2001) and/or viral infection (Suttle
and Chan, 1994;Dolan and Šimek, 1999;Sullivan et al.,
2003). The fact that the surface diel variations of picoplankton
abundance did not show any consistent pattern in this dataset
(Figure 2), contrary to other studies (André et al., 1999;
Vaulot and Marie, 1999;Lefort and Gasol, 2014), can be
explained by the tight coupling between growth and loss
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FIGURE 5 | Vertical profile of the estimated specific growth rates of Prochlorococcus (Pro, A), low (LF-Syn, B) and high (HF-Syn, C) fluorescence Synechococcus,
small (Speuk, D), and large (Lpeuk, E) picoeukaryotes in the four sampling cycles. Note that different scales were used.
FIGURE 6 | Vertical profile of the estimated specific growth rates of low (LNA, A) and high (HNA, B) nucleic acid heterotrophic prokaryotes in the four sampling
cycles.
processes (Lefort and Gasol, 2014), preventing standing stocks
from sustained, marked increases or decreases on a diel basis. The
diel variations in the abundance of heterotrophic picoplankton
could also be explained by the numerous species likely present
within the flow cytometric categories of LNA (for instance the
Alphaproteobacteria SAR11 or the Gammaproteobacteria SAR86
clades) and HNA cells (frequently members of Roseobacter,
Gammaproteobacteria and Bacteroidetes,Schattenhofer et al.,
2011;Morris et al., 2012). Although lacking any clear pattern,
the diel variations in the abundance of picophytoplankton
were usually higher than those of heterotrophic prokaryotes
(Tables 1,2,Figure 2, and Supplementary Figure 2).
In contrast to variations in abundance, highly consistent
periodicities in cell size were apparent for picocyanobacteria
regardless of the sampled period (Figure 3). The diel increase in
surface LF- and HF-Synechococcus cell size ceased approximately
2 h earlier than that of Prochlorococcus, indicating that the time
of cell division differed between these picocyanobacteria genera
(Figures 3A–H). Although diel changes in cell size have been
used to estimate in situ growth rate in some studies (Sosik
et al., 2003;Hunter-Cevera et al., 2014;Ribalet et al., 2015), the
straightforward application of relationships obtained in regions
with different environmental characteristics to the Red Sea is not
feasible. In an early study, Moigis (1999) showed high growth
rates values (up to 3 d−1) of marine cyanobacteria along the
latitudinal axis of the Red Sea, especially in the territorial waters
of Sudan and Yemen. With regard to heterotrophic prokaryotes,
Silva et al. (2019) recently provided estimates of the specific
growth rates of LNA and HNA groups at the surface of the
shallow waters of KAUST Harbor, but there is virtually no
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FIGURE 7 | Depth-weighted averages of the estimated specific growth rates of autotrophic picoplankton groups for the upper 100 m (A) and heterotrophic
prokaryotes for the epi- (0–200 m) and mesopelagic (200–700 m) layers (B).
information about their vertical variability in Red Sea open
waters. At the study site off KAEC, we only have direct growth
measurements of autotrophic and heterotrophic picoplankton
groups in predator-free incubations of samples collected in
the epipelagic and mesopelagic zone in the winter 24 h cycle
(Figure 4 and Supplementary Figure 5). In situ changes in cell
size reflected remarkably well the measured specific growth rates
of all picoplankton groups, except Lpeuk which were removed
by the pre-filtration step, in the experimental incubations.
Unfortunately, this was the only concurrent autotrophic and
heterotrophic picoplankton growth rate experiment conducted at
the study site but given the high percentage of variance explained
we felt it was safe to extrapolate this relationship to the remaining
periods. Two additional experiments focused on heterotrophic
prokaryotes and how migrating mesopelagic fishes were able to
supply labile DOC to sustain their growth rate (Calleja et al., 2018;
Morán et al., 2022) were not included because no information
about picophytoplankton was available. Previous studies have
estimated in situ division rates of picoplankton populations from
the ratio of the maximum to the minimum flow cytometric right
angle light scatter value, a common proxy for picoplankton cell
size (Vaulot and Marie, 1999;Jacquet et al., 2002;Lefort and
Gasol, 2014). The higher values of this ratio were indicative of
higher division rates. In this study, SSC values were converted
to biovolume assuming a spherical shape. However, rather than
using only two values of cell size, the minimum and maximum,
we decided to use the coefficient of variation (CV) of all data of
cell size available over 24 h. Although the cell size CVs and the
maximum to minimum SSC ratios were as expected correlated
(r= 0.41, p= 0.0002, n= 91), by choosing the CVs we minimized
the possible errors of anomalously low or high values in the ratio.
The vertical distributions of Prochlorococcus abundance and
estimated growth rates showed similar trends with depth
(Figure 5A and Supplementary Figure 2), while the match
was less marked for the other groups of picophytoplankton
(Figures 5B–E). Our maximum estimated specific growth rates
of Prochlorococcus in the central Red Sea were slightly lower than
the values measured in the tropical Pacific (ca. 1 d−1,Vaulot
et al., 1995;Selph et al., 2005) and central Atlantic (ca. 1.5 d−1,
Quevedo and Anadón, 2001;Worden and Binder, 2003;Agawin
and Agustí, 2005;Figure 7A). The maximum specific growth
rates of the two groups of Synechococcus were slightly higher
than in tropical Pacific waters (0.50 d−1,Landry et al., 2003;
Selph et al., 2005), but notably lower than in the central Atlantic
(1.85 d−1,Agawin and Agustí, 2005) and the NW Mediterranean
(1.79 d−1,Agawin et al., 1998;Figure 7A). However, none of
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FIGURE 8 | Relationships between the mean diel abundance of (A)
autotrophic picoplankton and (B,C) HNA and LNA heterotrophic prokaryotes
in the epipelagic and mesopelagic layers, excluding the second-noon to avoid
duplicate data, and the estimated specific growth rate. Group abbreviations
as in Figure 4.
these studies distinguished between low and high fluorescence
groups as we did here. It is remarkable that the difference between
the LF-Syn and HF-Syn groups usually held year-round, so that
the similar seasonal variability in the upper epipelagic suggests
a common response of both types to environmental factors. In
shallow waters, however, the HF-Syn group is virtually absent
from April to September (Sabbagh et al., 2020), suggesting that
some requirements for HF-Syn growth are absent in shallow
waters for a substantial period of the year. Not many studies
have assessed the specific growth rates of picoeukaryotes in
oligotrophic tropical waters. Those we are aware of report values
for the central (Landry et al., 1995), northwest (Selph et al., 2005)
and eastern tropical Pacific (Taniguchi et al., 2014). Although the
large picoeukaryotes group typically grew faster than the small
one, our values (Figure 7A) were uniformly lower than in the
tropical Pacific (0.77 d−1,Selph et al., 2005).
Heterotrophic bacteria and archaea consistently showed lower
variations in cell size than autotrophic picoplankton (Table 2
and Figures 3M–P), resulting in lower specific growth rate
estimates according to our model. The vertical distribution of
LNA prokaryotes growth was as expected more uniform than
their HNA counterparts, with no significant differences between
the epipelagic and mesopelagic layers (Figures 6A,7B). Similar
to Prochlorococcus, we typically observed higher specific growth
rates of HNA prokaryotes at the DCM than at the surface
(Figure 6B), indicating that copiotroph taxa were able to respond
to higher substrate inputs as suggested elsewhere (Morán et al.,
2011). In contrast to our findings, Scharek and Latasa (2007)
showed higher specific growth rates of LNA cells (ca. 0.90 d−1)
at the DCM of the NW Mediterranean, doubling the values of
the HNA group (ca. 0.40 d−1), while HNA grew clearly faster
at the surface (1.18 for HNA and 0.50 d−1for LNA). Also,
seasonality was much more marked for HNA than for LNA
cells in the epipelagic layer (Figure 7B). Our estimated specific
growth rates of LNA and HNA heterotrophic prokaryotes were
lower than the maxima reported for the NW Mediterranean
Sea (ca. 1.0 d−1,Vaqué et al., 2001;Sala et al., 2002) and the
tropical Atlantic Ocean (ca. 0.8 d−1,Steinberg et al., 2001).
Compared with the few studies reporting specific growth rates
of heterotrophic prokaryotes in the Red Sea, our values were
higher than in the surface northern Red Sea (0.15 d−1;Grossart
and Simon, 2002), but lower than in the central (ca. 1.5 d−1)
and southern regions (ca. 0.7 d−1;Weisse, 1989). A recent study
reported notably higher values (up to 1.1 d−1for LNA cells
and 2.3 d−1for HNA cells) in the nearby shallow waters of
KAUST Harbor (Silva et al., 2019). The low mean values of
dissolved inorganic nitrogen (nitrite plus nitrate, 0.17 ±0.11
µmol L−1) and dissolved organic carbon (76 ±8µmol C L−1)
concentrations characterizing this study site (Al-Otaibi et al.,
2020) could partially explain our lower values compared with the
shallower environment (occasionally exceeding 20 µmol L−1of
DIN and up to 98 µmol L−1of DOC, Silva et al., 2019). Another
important process sustaining heterotrophic prokaryotes growth
in the Red Sea is the diel vertical migration of mesopelagic fish
(Calleja et al., 2018;García et al., 2018). The deep scattering
layer (DSL), located between 400 and 600 m at our study site
(Klevjer et al., 2016), is dominated by Benthosema pterotum
lanternfishes. The labile dissolved organic matter released by the
fishes fuels an active community of heterotrophic prokaryotes at
the DSL (Calleja et al., 2018;García et al., 2018), with significant
differences between day and night linked to the presence and
absence, respectively, of mesopelagic fish (Morán et al., 2022).
Our LNA and HNA prokaryotes specific growth rate estimates
did not show any consistent increase at the DSL depth (Figure 6).
Although in direct incubations Calleja et al. (2018) had reported
higher specific growth rates of heterotrophic prokaryotes from
mesopelagic depths than those at the surface, a more recent study
using the same approach found the opposite (Morán et al., 2022),
indicating that vertical variations in their growth rates can also
vary temporally, especially for the HNA group (see for instance
Winter values in Figure 6B).
Differences in the various picoplankton groups’ specific
growth rates can, as mentioned above, be partially due to
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Al-Otaibi et al. Diel Variability of Picoplankton
the different approaches being used, but we would argue that
offshore Red Sea microbial plankton grow at slower rates than
other tropical sites. However, an interesting finding is that
higher specific growth rates of picophytoplankton and HNA
heterotrophic prokaryotes were mirrored by higher abundances
averaged for the different 24 h periods (Figures 8A,C), suggesting
a tight coupling between the growth and loss rates over the
annual cycle. Although we lacked concomitant measurements of
grazing and viral lysis, strong top-down control by heterotrophic
nanoflagellates and viruses was the plausible cause for the low
abundances of autotrophic and heterotrophic bacterioplankton
found in shallower waters of the central Red Sea (Silva et al.,
2019;Sabbagh et al., 2020). Future detailed studies about grazing
and viral mortality in the whole water column are needed to
fully understand the diel variability of planktonic microbes in
this tropical basin. By filling the gap of diel scale studies of
picoplankton communities in low latitude regions, our results
also add to the currently limited database on their growth
rates in the Red Sea.
CONCLUSION
Our findings confirm the importance of the diel scale for
variations in the abundance and cell size of the major groups of
picoplankton dominating plankton communities in the central
Red Sea. The diel coefficient of variation in cell size allowed
us to estimate the specific growth rates of autotrophic and
heterotrophic picoplankton in offshore Red Sea waters after
confirming a strongly positive association between in situ
variations in cell size and the specific growth rates obtained
in concurrent experimental incubations. The vertical profiles
of the estimated specific growth rates of Prochlorococcus and
HNA heterotrophic prokaryotes were more similar to their
corresponding in situ abundances than the other groups. The
estimated specific growth rates were generally lower than in other
tropical environments. Prochlorococcus showed the highest values
year-round, followed by the low phycoerythrin fluorescence
group of Synechococcus and the lowest corresponded to small
picoeukaryotes. HNA consistently outgrew LNA heterotrophic
prokaryotes, which failed to show marked vertical or seasonal
variations. These results suggest a fine coupling between
picophytoplankton and HNA prokaryotes standing stocks and
specific growth rates over the seasonal cycle in Red Sea waters.
DATA AVAILABILITY STATEMENT
The original contributions presented in the study are included
in the article/Supplementary Material, further inquiries can be
directed to the corresponding authors.
AUTHOR CONTRIBUTIONS
NA-O analyzed the data, prepared figures and/or tables, authored
or reviewed drafts of the manuscript, and approved the final draft.
FCG analyzed the data, reviewed drafts of the manuscript, and
approved the final draft. XAGM conceived the research, analyzed
the data, contributed to the interpretation of results, reviewed
drafts of the manuscript, and approved the final draft.
ACKNOWLEDGMENTS
We gratefully acknowledge the crew of the RVs Thuwal and
KAUST Explorer and all the personnel from the Coastal
and Marine Resources Core Lab for their diligent field-
work assistance. Luis Silva designed and performed the
winter experiment for estimating specific growth rates.
Miguel Viegas, Eman I. Sabbagh, Abbrar Labban, Maria Ll.
Calleja, Katherine Rowe, Anders Røstad, and Luis Silva aided
enormously with fieldwork.
SUPPLEMENTARY MATERIAL
The Supplementary Material for this article can be found
online at: https://www.frontiersin.org/articles/10.3389/fmars.
2021.752910/full#supplementary-material
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