Aquaculture 468 (2017) 162–168

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Aquaculture

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Short Communication

The effect of different dietary microalgae on the fatty acid profile,

fecundity and population development of the calanoid copepod
Pseudodiaptomus hessei (Copepoda: Calanoida)
Oyama Siqwepu a, Nicole B Richoux b, Niall G Vine a,⁎
a
Department of Zoology and Entomology, University of Fort Hare, Alice 5700, South Africa
b
Department of Zoology and Entomology, Rhodes University, Grahamstown 6140, South Africa

a r t i c l e i n f o a b s t r a c t

Article history: We studied the effect of dietary microalgae on the fecundity, population development, and fatty acid profile [with
Received 3 June 2016 focus on the essential fatty acids docosahexaenoic acid (DHA; 22:6ω3) and eicosapentaenoic acid (EPA; 20:5ω3)]
Received in revised form 29 September 2016 of the calanoid copepod Pseudodiaptomus hessei, a potential live food for finfish larvae in aquaculture. Two mono-
Accepted 4 October 2016
algal diets, the Tahitian strain of Isochrysis galbana and Rhodomonas salina, and a 50:50 binary diet of the two
Available online 06 October 2016
were fed to copepods. Wild caught copepods were used as a baseline reference point. Copepods fed I. galbana
Keywords:
and the 50:50 binary diet had high DHA:EPA ratios and DHA content relative to those fed R. salina. The EPA con-
DHA tent of the copepods was similar for all three diets. Copepods fed R. salina had the highest fecundity relative to
EPA those fed I. galbana and the 50:50 binary diet. The largest population was obtained when the copepods were
First feeding fed I. galbana, which differed from those fed R. salina and the 50:50 binary diet. The results of this study showed
Live food that the fatty acid composition of P. hessei can be altered by changing its dietary microalgae, and that the copepod
Larviculture can accumulate fatty acids from its diet, especially DHA and EPA. Diet also affected fecundity and population de-
velopment of P. hessei, so this species represents a potential live food candidate for marine finfish larvae as its nu-
tritional composition and productivity can be manipulated to suit the needs of marine finfish larvae.
© 2016 Published by Elsevier B.V.

1. Introduction 1997). These three components are considered essential fatty acids
(EFAs) because they cannot be synthesized by fish de novo in quantities
Copepod nauplii are the main food source for fish larvae in the wild necessary for their normal metabolic needs (Watanabe, 1982, 1993).
(McKinnon et al., 2003; Chen et al., 2006). The small body size, high nu- Deficiencies in ω3 highly unsaturated fatty acids (HUFA, which includes
tritional content, and variable life stages make copepods important prey DHA and EPA) can lead to retarded fish growth, high mortality rates,
for fish larvae and other life cycle stages of fish. The mouth size or gape and a poor ability to handle stress (Izquierdo, 1996). The requirements
of the larvae limit what they can feed on, so copepods represent a better of these HUFA differ from species to species (Watanabe, 1993), but gen-
alternative as prey than brine shrimp (Artemia spp.), which may be too erally higher ratios of DHA:EPA increase the survival and growth of ma-
large to capture, and rotifers (Branchionus plicatilis), which may be nu- rine fish larvae (Nanton and Castell, 1998). For example, Bell et al.
tritionally inadequate (Lavens and Sorgeloos, 1996). Lack of suitable nu- (1985) found that the survival of juvenile turbot significantly increased
tritional live food or the inability to culture balanced nutritional live when the DHA:EPA ratio of the diet was increased from 0.1 to 0.5. The
food prey for first feeding larvae of marine finfish is a limiting factor in recommended ratio for marine finfish larvae feeds is 2:1 (Sargent et
aquaculture (Rippingale and Payne, 2001). There is a clear need to iden- al., 1997).
tify and culture suitable live prey that will allow suitable survival, Different species of algae have divergent nutritional qualities, one
growth, and eventual reproduction of the larvae (Rippingale and component being the relative proportions of EFAs, which in turn affect
Payne, 2001). the growth and productivity of the copepods (Brown et al., 1997; Tang
Fatty acids are important nutrients, particularly the polyunsaturated and Taal, 2005; Ohs et al., 2010a). To increase aquaculture-based pro-
fatty acids docosahexaenoic acid (DHA; 22:6ω3), eicosapentaenoic acid ductivity, the nutritional quality of the microalgae food must match
(EPA; 20:5ω3) and arachidonic acid (ARA; 20:4ω6), for the growth of the nutritional needs of the cultured copepods (Ohs et al., 2010a).
marine fish larvae (Watanabe, 1982; Watanabe, 1993; Brown et al., One of the key factors affecting population growth in copepods is fe-
cundity, and fecundity is, in turn, significantly affected by the extent to
⁎ Corresponding author. which the nutritional requirements of the copepod are met by its diet
E-mail address: [email protected] (N.G. Vine). (Kleppel and Burkart, 1995; Milione and Zeng, 2007). In addition to

http://dx.doi.org/10.1016/j.aquaculture.2016.10.008
0044-8486/© 2016 Published by Elsevier B.V.
 O. Siqwepu et al. / Aquaculture 468 (2017) 162–168 163

fecundity, the productivity of copepods is related to hatching rate, sur- 2.3. Fatty acid experiment
vival and development of nauplii and copepodites to adults (Milione
and Zeng, 2007), with a diverse diet generally being associated with in- After 21 days, 50 adult copepods were collected from each container
creased consumer productivity (Kleppel and Burkart, 1995). Mixed using a 550 μm sieve and rinsed with clean seawater to remove any
diets tend to be superior, as individual food sources usually lack specific algae. Fatty acids of wild copepods collected from the Kariega Estuary
nutritional compounds (Koski et al., 2006). during June 2014 were used to compare with fatty acids of the copepods
Pseudodiaptomus hessei is a small (adults b 1.88 mm; copepodites fed the different experimental treatments. Fifty copepods in one filter
b0.508 mm; early nauplii b0.168 mm), dominant calanoid copepod in represented one sample. Adults were used in the analysis instead of
the zooplankton biomass of estuaries and lagoons along the west and nauplii because they are larger and easier to collect and clean. The sam-
south coasts of South Africa (Grindley, 1963; Jerling and Wooldridge, ples were freeze dried for 15 h and then were processed using a one-
1991; Isla and Perissinotto, 2004). It is able to tolerate a wide range of step method adapted from Indarti et al. (2005). Aliquots (1 μL) of the
salinities, and its optimum temperature for development is 26 °C (life fatty acid methyl esters (FAME) suspensions were injected at 260 °C
cycle takes ± 16 days; Jerling and Wooldridge, 1991; Isla and into an Agilent 7890 A/7000 gas chromatograph/mass spectrometer
Perissinotto, 2004). Members of the family Pseudodiaptomidae have (GC/MS) equipped with a Zebron ZB-WAX plus capillary column with
been cultured worldwide for use as live food in aquaculture, and several helium as a carrier gas. FAME peaks were created using a flame ioniza-
aspects of their cultivability have been investigated (Chen et al., 2006; tion or MS detector and visualized using Chemstation (B04.02) or
Puello-Cruz et al., 2009; Rhyne et al., 2009; Ohs et al., 2010a; Ohs et Masshunter (B.05.00) software, and peak identities were confirmed
al., 2010b). In South Africa, there is no copepod species identified as a using a NIST 08 MS library. Fatty acids were reported as the percentage
potential live food candidate for use in marine larviculture, therefore of total fatty acids (%TFAs, mean ± standard deviation) or μg/copepod
the aim of this study was to investigate the effects of different dietary and grouped into monounsaturated fatty acids (MUFAs), saturated
microalgae on the fatty acid profile, fecundity and population develop- fatty acids (SFAs), HUFA (includes DHA and EPA) and EFA (includes
ment of P. hessei under culture conditions. A suitable algal diet for the DHA, EPA and ARA). Quantitative data (μg/copepod) were used to per-
culture of P. hessei would be one that provides a DHA:EPA ratio in excess form a one-way ANOVA to investigate differences in the fatty acid con-
of 2:1, as recommended for marine finfish larvae (Sargent et al., 1997). tent of copepods and microalgae.
The diet should also provide optimum fecundity and increased popula-
tion growth and productivity. 2.3.1. Fecundity and population development experiment
After 21 days, twenty ovigerous females from each container were
collected and preserved in 10% buffered formalin. Each copepod was
2. Materials and methods placed in a petri dish, where the copepod was gently separated from
the egg sac membrane. The egg sac was placed in a 5% solution of sodi-
2.1. Algal and copepod culture um hypochlorite and gently agitated to dissolve the egg sac membrane.
The total number of eggs in each sac was counted under a dissecting ste-
Stock cultures of the algae Isochrysis galbana (Tahitian strain; CS-22) reo microscope (BestScope BS3040). Apart from those removed for the
and Tetraselmis suecica (CS-187) were sourced from the Department of fatty acid analyses, all the remaining copepods from each container
Agriculture, Forestry and Fisheries in Cape Town, and Rhodomonas sali- were preserved in 10% formalin, counted and each stage of develop-
na (CS − 201) from the Commonwealth Scientific and Industrial Re- ment identified according to Jerling and Wooldridge (1989). The final
search Organisation, Australia. The cultures were maintained at Pure population count per diet treatment was estimated as the average of
Ocean Aquaculture in East London at 22 ± 1 °C and salinity of 28‰. three replicates. The intrinsic rate of population increase r was calculat-
The photoperiod was set at 24 h of light with a fluorescent source. All ed for each treatment using the formula:
microalgae were fertilized using f/2 (Varicon aqua solutions, Cell-Hi
F2P) medium. r ¼ 1nðN1 =N0 Þ=t;
Copepods were collected at dusk during May 2013 from the Kariega
River in the Eastern Cape, South Africa. P. hessei was isolated from the where N0 = the population size at the beginning of the experiment,
rest of the zooplankton and fed a mixture of the microalgae T. suecica, N1 = the population size at the end of the experiment, while t (days)
I. galbana and R. salina; the increasing copepod population was scaled is the duration of the experiment.
up from tanks at Pure Ocean Aquaculture in East London.
2.4. Data analysis

2.2. Experimental set-up and design Non-metric multidimensional scaling (nMDS) of the untransformed
fatty acid data (%TFA) was used to examine differences among the cope-
The experiment was run at Pure Ocean Aquaculture in East Lon- pods fed with the three microalgae treatments, the wild copepods, and
don for 21 days. The photoperiod was set at 24 h of light with a fluo- the microalgae. Analyses of similarity (ANOSIM) were performed to
rescent light. The temperature was 26 ± 1 °C and salinity was kept at identify whether any of these groups were statistically similar to one
28 ± 1. For the experiment, the copepods required a daily algal feed another based on their fatty acid profiles. The ANOSIM test statistic R
volume. The carbon concentrations (μg C/L) were calculated from was reported as a value between −1 and +1 (1 indicates complete dis-
cell volumes (μm3 ) (Strathmann, 1967). Carbon equivalents used similarity, 0 indicates complete similarity, with negative values rarely
were 7 pg C/cell for I. galbana and 40.7 pg C/cell for R. salina. The occurring), together with a corresponding p value. Additional R and p
daily carbon concentration used for feeding was 2000 μg C/L/day, a values were calculated using pair-wise comparisons as part of the
concentration above saturation (Kiørbe et al., 1985), which equated ANOSIM procedure. Similarity percentage (SIMPER) and principal com-
to 1.65 × 10 5 cells/mL for R. salina and 6.27 × 10 5 cells/mL for I. ponent analysis (PCA) were used to evaluate which fatty acids were the
galbana. Nine aerated 20 L containers were arranged as three repli- most influential on any differences between the copepods fed different
cate containers of the three feeding treatments: 100% I. galbana, dietary microalgae, wild copepods and the microalgae. Analyses were
100% R. salina and a 50%:50% mixture of I. galbana and R. salina. The conducted using PAST 3.00. The fecundity and population size data
water and the microalgae were exchanged every two days. Twenty were analysed using one-way ANOVA. When significant differences
two ovigerous female copepods were added into each 20 L container (p b 0.05) were found, a Tukey's multiple comparison test was used to
at the beginning of the experiment. determine specific differences among treatments (p b 0.05). Analyses
164 O. Siqwepu et al. / Aquaculture 468 (2017) 162–168

Table 1
Fatty acid composition of Pseudodiaptomus hessei fed different dietary microalgae and wild copepods and different dietary microalgae fed to copepods. All values are presented as mean %
total fatty acid ± standard deviation.

Dietary microalgae Copepods

Fatty acids Rhodomonas salina Isochrysis galbana 50:50 binary diet (calculated) Rhodomonas salina Isochrysis galbana 50:50 binary diet Wild

20:4ω6 (ARA) nd nd nd nd nd nd 0.12 ± 0.12

20:5ω3 (EPA) 0.64 ± 2.87 0.22 ± 0.31 0.91 ± 1.04 0.31 ± 0.67 0.19 ± 0.60 0.74 ± 4.71 3.09 ± 0.57
22:6ω3 (DHA) 0.67 ± 1.50 1.08 ± 3.56 2.02 ± 1.92 0.57 ± 0.99 0.69 ± 1.57 0.84 ± 2.84 4.85 ± 1.41
Sum PUFA 81.12 62.75 32.83 180.15 186.57 156.00 183.58
Sum MUFA 59.37 90.77 32.99 20.18 16.78 24.28 40.70
Sum SFA 205.92 346.48 120.78 599.67 896.65 519.73 175.72
Sum EFA 20.92 32.41 11.71 55.93 106.51 77.38 129.05
DHA: EPA 0.65 ± 2.25 0.97 ± 2.71 1.87 ± 1.61 0.24 ± 0.34 0.34 ± 0.96 0.32 ± 1.82 0.39 ± 0.10

nd indicates the fatty acid was not detected.

were performed using SPSS version 23, and summarized data are pre- p = 0.004). The DHA:EPA ratio of the copepods fed the binary diet
sented as means ± standard deviation. was similar to that of the copepods fed I. galbana (one-way ANOVA:
F(14, 26) = 2, p = 0.168); however, there was a significant difference be-
3. Results tween the ratios of copepods fed R. salina and I. galbana (one-way
ANOVA: F(1, 10) = 35, p = 0.001) and those fed I. galbana and the wild
3.1. Fatty acid composition of copepods copepods (one-way ANOVA: F(1, 14) = 71, p = 0.001). The DHA:EPA
ratio of copepods fed R. salina was significantly different from that of
A total of 31 fatty acids were detected in the calanoid copepod P. the wild copepods (one-way ANOVA: F(1, 10) = 5, p = 0.041).
hessei (Supplemental Table S1, summary in Table 1). The nMDS plots
and ANOSIM showed differences in fatty acid profiles among the differ- 3.2. Fecundity
ent food treatments (R = 0.60, p = 0.000) and the wild copepods (R =
0.58, p = 0.0001) (Fig. 1). The copepods fed on the mixed diet generally The experimental treatment had an effect on the fecundity of P.
occupied the space intermediate between the copepods fed with the hessei when compared to the wild caught copepods (one-way
single algal diets. SIMPER and PCA showed which fatty acids were influ- ANOVA: F(3, 3) = 96, p = 0.001). All three experimental groups were
ential in separating the copepod groups, with HUFA particularly domi- significantly different from each other (p = 0.001) except for the cope-
nant and distinctive in the wild copepods, whereas the cultured pods fed I. galbana and those fed the 50:50 binary diet (p = 0.981). Fig. 4
copepods tended to be dominated by various SFA such as 16:0, 17:0 shows that the wild copepods had the highest number of eggs, followed
and 18:0 (Fig. 1). by the copepods fed the mono-algal R. salina food, and those fed the
There were similarities in the EPA levels of wild copepods and those 50:50 binary food. Copepods fed I. galbana had the fewest number of
fed the 50:50 binary diet (one-way ANOVA: F(1, 10) = 0.288, p = 0.60), eggs.
but those fed R. salina and I. galbana were different from the wild cope-
pods (Fig. 2). The DHA levels of the copepods fed different dietary 3.3. Population growth and development
microalgae were similar between I. galbana and the 50:50 binary diet
(one-way ANOVA: F(3, 26) = 1, p = 0.735), but they were different for After 21 days of culture, the average final population numbers of P.
the 50:50 binary diet and R. salina (one-way ANOVA: F(1, 10) = 10, hessei were collated into four categories, namely ‘All Stages Included’

Fig. 1. Non-metric multidimensional scaling output using fatty acid profiles of Pseudodiaptomus hessei fed microalgal diets Isochrysis galbana, Rhodomonas salina, a 50:50 binary diet of the
two microalgae and wild copepods. Polygons representing the fatty acid groups of different dietary microalgae and wild copepods. Fatty acids influential in separating the 3 diets and wild
copepods (derived from SIMPER and PCA) are superimposed in the plot. Squares = 50:50 binary diet, hollow circles = I. galbana, triangles = R. salina and filled circles = wild copepods.
 O. Siqwepu et al. / Aquaculture 468 (2017) 162–168 165

treatment was significantly greater than all other treatments (p =

DHA:EPA ratio
0.009). The treatment producing the second highest population was
the 50:50 binary treatment, which was not different from R. salina.
The number of nauplii was significantly different between the three
treatments (ANOVA: F(2, 8) = 14, p = 0.006). The intrinsic rate of pop-
ulation increase r ranged from 0.14 ± 0.005 in the 50:50 binary treat-
% total fatty acid

ment to 0.20 ± 0.06 for those in the I. galbana treatment (Table 2)

(Fig. 3). I. galbana produced the highest number of copepods across all
life stages.

4. Discussion

Dietary microalgae had an effect on the fatty acid profile of P. hessei.

These results are consistent with studies on other copepod species fed
(A) different diets that changed the copepod fatty acid profiles (Graeve et
al., 1994; Rasdi et al., 2015). Rasdi et al. (2015) found that the fatty
acid profile of Cyclopina kasignete was altered when it was fed dry
Nannochloropsis oculata, dry Melosira sp., Schroederiella apiculata,
Scenedesmus pectinatus and Tetraedrom minimum. The fatty acid profiles
of the copepods were altered in that the levels of PUFA, including EPA,
DHA and ARA, were dependent on the type of algae they were fed.
Graeve et al. (1994) also found similar results, in that the copepods
Calanus finmarchicus, Calanus hyperboreus and Calanus glacialis fed
Thalassiosira antarctica and Amphidinium cartera changed their fatty
acid profile depending on which food was provided. The fatty acid pro-
file of the copepods in our study was dominated by SFA and MUFA, with
lower concentrations of PUFA. Similarly, McKinnon et al. (2003) studied
the calanoid copepods Bestiolina similis and Parvocalanus crassirostris, to
(B) which they fed different dietary microalgae. They found that the profiles
of the copepods were dominated by SFA (16:0, 18:0, 14:0) and MUFA
(22:1, 20:1) and that the EFA, DHA and EPA were present in smaller pro-
portions (McKinnon et al., 2003).
We used wild copepods as a baseline for comparison with copepods
fed the three dietary treatments. The higher number of fatty acids de-
tected in the wild copepods was likely due to the fact that copepods in
the wild do not feed exclusively on microalgae but also on heterotrophic
protists and dinoflagellates, giving them a more diverse fatty acid profile
(Tang and Taal, 2005). In our study, the wild zooplankton had signifi-
cantly higher concentrations of long chain EFA, DHA and EPA, and low
levels of ARA were detected, as opposed to copepods fed the dietary
microalgae, which had no detectable ARA content. These results were
similar to findings by Nanton and Castell (1999) working on the effect
of temperature and dietary fatty acids on the composition of the
(C) harpacticoid copepods Tisbe sp. and Amonardia sp. They found that
wild zooplankton contained large amounts of both long chain EFA, par-
Fig. 2. A) Docosahexaenoic acid (DHA), B) eicosapentaenoic acid (EPA) and C) DHA:EPA ticularly EPA and DHA (Nanton and Castell, 1999).
ratio levels in Pseudodiaptomus hessei corresponding to three dietary treatments. Values P. hessei fed I. galbana met the required DHA:EPA ratio of 2:1 recom-
are means ± standard errors.
mended by Sargent et al. (1997) for marine finfish larvae. Other cope-
pods such as Acartia sinjiensis, Bestiolina similis and Parvocalanus
(adults, copepodites and nauplii), ‘Adults Only’ (only stage C6), crassirostris also reached the required DHA:EPA ratio of 2:1 when fed
‘Copepodites Only’ (C1–C5), and ‘Nauplii Only’ (N2–N6, Table 2). The the dietary microalgae Isochrysis sp. Tetraselmis chuii, Rhodomonas sp.
final population size (including all stages) for copepods in the I. galbana and Heterocapsa niei (McKinnon et al., 2003).

Table 2
Final population and intrinsic rate (r) of population increase of Pseudodiaptomus hessei cultures after a 21 day period fed on three microalgal diets.

FNP — all stages r (all stages Copepodite r (Copepodite r (Nauplii

Microalgal diet included included) Adults only r (adults only) only only) Nauplii only only)

Rhodomonas 144.3 ± 45.08a 0.05 ± 0.01 65.1 ± 8.84a 0.03 ± 1.7 × 25.9 ± 20.88a 0.02 ± 0.02 26.7 ± 9.54a 0.02 ± 0.004
salina 10−17
Isochrysis galbana 236.4 ± 53.25b 0.07 ± 0.03 93.8 ± 0.04 ± 0.003 43.3 ± 12.42a 0.03 ± 0.01 49.7 ± 11.59b 0.03 ± 0.003
18.11ba
ac a ac
50:50 binary diet 145.8 ± 23.10 0.05 ± 0.002 56.1 ± 0.03 ± 0.01 32.6 ± 8.84 0.02 ± 0.003 28.6 ± 9.68 0.02 ± 0.01
34.83ba

Data are presented as mean ± standard error; different superscript letters in the same column indicate significant differences (p b 0.05).
FNP = final population number.
166 O. Siqwepu et al. / Aquaculture 468 (2017) 162–168

400

Mean population composition of P.hessei

350

300

250

200

150

100

50

0
Rhodomonas salina Isochrysis galbana 50:50 Binary diet
Dietary Microalgae

N2-N6 C1-C5 Adult (C6) Total

Fig. 3. Effect of three microalgal diets on the average population composition of Pseudodiaptomus hessei over a 21 day culture period. The experiment started with 21 ovigerous females.
Data are presented as means ± standard errors. N2–N6 = naupliar stages 2 to 6; C1–C5 = copepodite stages 1 to 5.

50

45
Mean egg production of P. hessei

40

35

30

25

20

15

10

0
Rhodomonas salina Isochrysis galbana 50:50 binary diet Wild caught copepods

Copepods fed dietary microalgae and wild copepods

Fig. 4. Mean P. hessei egg production of wild caught and copepods fed three different dietary microalgae over 21 days. Data are presented as mean ± standard error.

The different food treatments in our study also had an effect on the species specific differences between P. hessei and A. tonsa. Camus et al.
fecundity of P. hessei, evident in that the number of eggs produced sig- (2009) found that when B. similis was fed a tri-algal diet, it showed op-
nificantly differed between the treatments (except for copepods fed I. timum egg production, hatching success and population numbers. Sim-
galbana and the 50:50 binary diet). Wild copepods had the highest ilarly, Ohs et al. (2010a) and Camus and Zeng (2012) found that the
number of eggs, followed by copepods fed R. salina and the 50:50 binary productivity of copepods increased when they were fed a mixed diet
diet. These results were contrary to those of Støttrup and Jensen (1990), rather than a mono-algal diet. In our study, however, when the mono-
who found that Acartia tonsa had higher egg production when fed the algal diet R. salina was fed to P. hessei it yielded higher egg production
mono-algal diet I. galbana than Rhodomonas sp. Støttrup and Jensen than the 50:50 binary diet and the mono-algal diet I. galbana. Consump-
(1990) attributed this outcome to a high DHA:EPA ratio in I. galbana. tion rate in copepods is influenced by size, quality and abundance of the
In our study, the results differed in that I. galbana-fed copepods were food item (Kiørbe et al., 1985), therefore it is possible that the two die-
not as fecund as copepods fed R. salina or the 50:50 binary diet (as tary algal species were not ingested at equal rates. Copepods may have
when A. tonsa was fed I. galbana); this difference was probably due to selected I. galbana, whose cell size is 4–6 μm and was provided at 6.27 ×

Table 3
Summary of data on the use of Rhodomonas salina, Isochrysis galbana and a 50:50 binary diet in the culture of Pseudodiaptomus hessei.

Parameter R. salina I. galbana 50:50 binary diet Wild Optimal diet

a b b c
DHA content (μg/copepod) 0.57 ± 0.99 0.69 ± 1.57 0.84 ± 2.84 4.85 ± 1.41 I. galbana and 50:50 binary diet
DHA:EPA ratio 0.24 ± 0.34m 0.34 ± 0.96n 0.32 ± 1.82n 0.39 ± 0.10m I. galbana and 50:50 binary diet
Fecundity (no. of eggs/female) 34.60 ± 0.77x 22.88 ± 0.70y 23.3 ± 0.87y 40.70 ± 0.91z R. salina
Population development (no. of copepods/20 L) 144.3 ± 45.08p 236 ± 53.25q 145.8 ± 23.10p Not tested I. galbana

Different superscripts in each row denote significant difference (p b 0.05).

 O. Siqwepu et al. / Aquaculture 468 (2017) 162–168 167

105 cells/mL, over R. salina, whose cell size is 5–12 μm fed at 1.65 × 105 In conclusion, P. hessei has a DHA:EPA ratio suited to meet the needs
cells/mL. Støttrup and Jensen (1990) found that A. tonsa ingested I. of marine finfish larvae. Moreover, when fed I. galbana, high yields of P.
galbana at a higher rate than Rhodomonas sp. This may have been the hessei could be obtained while providing the copepods with a HUFA rich
case in our study, as copepods may have selected I. galbana over R. sali- diet and maintaining a high DHA:EPA ratio. Further research is neces-
na, leading to lower fecundity in copepods fed the 50:50 binary diet. sary to investigate the effect of other diets on the fatty acid and amino
Based on the finding that the 50:50 binary diet did not improve the fe- acid profiles of P. hessei.
cundity of P. hessei, a 50:50 combination of I. galbana and R. salina may
not be as nutritionally adequate as R. salina for enhancing the fecundity
Acknowledgements
of P. hessei.
The different algal treatments fed to the copepods affected the pop-
We thank Pure Ocean for infrastructural support, the South African
ulation growth of the copepods. I. galbana had the highest population
Institute of Aquatic Biodiversity (SAIAB) for assisting with copepod col-
size, followed by the 50:50 binary diet, which was similar to the cope-
lections, and the African Coelacanth Ecosystem Programme (ACEP)
pods fed R. salina (Table 2). This effect was similar in Milione and
Phuhlisa project, the National Research Foundation (NRF) of South Afri-
Zeng (2007), who observed that microalgal diet affected population
ca and the University of Fort Hare (GMRDC) for funding the study.
growth of the copepod A. sinjiensis, with a binary diet of Tetraselmis
chuii and I. galbana resulting in more successful growth than any
Appendix A. Supplementary data
mono or binary diet. However, in our study, the mono-algal diet of I.
galbana was more successful than the 50:50 binary diet for population
Supplementary data to this article can be found online at http://dx.
growth [although fecundity was lower in copepods fed I. galbana (all
doi.org/10.1016/j.aquaculture.2016.10.008.
stages included)]. Koski et al. (2006) observed a similar trend with
Temora longicornis, which produced a high number of eggs followed
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