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Aquaculture 245 (

The use of inert artificial commercial food sources as replacements

of traditional live food items in the culture of larval shrimp,

Farfantepenaeus aztecus

C.B. Robinsona,b, T.M. Samochaa,b,*, J.M. Foxb, R.L. Gandya, D.A. McKeeb

aTexas Agricultural Experiment Station–Shrimp Mariculture Research Facility, Texas A&M University System, 4301 Waldron Rd.,

Corpus Christi, TX 78418, USAbCenter for the Sciences, Room 251, Texas A&M University–Corpus Christi, 6300 Ocean Drive, Corpus Christi, TX 78412, USA

Received 10 May 2004; received in revised form 24 August 2004; accepted 30 November 2004

Abstract

Replacement of live feeds with alternate food sources is of major importance in commercial shrimp hatcheries. In this study,

partial and complete replacement of live microalgae and Artemia nauplii with microalgae pastes and inert feeds is reported in

the larval rearing of the brown shrimp Farfantepenaeus aztecus. Five different experimental feeds were used for each live food

replacement study. Partial replacement of live microalgae using Chaetoceros 1000 bPremium FreshQ Instant Algaek paste and

Liqualifek liquid larval feed showed survival rates similar to a control feed, however, postlarvae mean dry weight and length

were significantly less than the control. Use of other replacement feeds, Epifeedk liquid larval feed, Zeiglerk E-Z Larvae

liquid feed, and Zeiglerk Z-Plus feed, yielded inferior results compared to the control feed. Statistical analysis of results from

the live Artemia nauplii replacement study indicated that larval shrimp fed a control feed had significantly greater survival,

mean PL length, and mean PL dry weight. The only exception was the partial replacement of Artemia nauplii using Liqualifekin which there was no significant difference in survival between the control and this treatment. Results indicated that although

Liqualifek, Epifeedk, Zeiglerk E-Z Larvae, Zeiglerk Z-Plus and Zeiglerk E-Z Artemia feeds could serve as partial

replacement of newly hatched Artemia nauplii in production of F. aztecus postlarvae. Significantly improved results can be

expected when larvae are fed newly hatched Artemia nauplii (control diet) with no supplement.

D 2004 Elsevier B.V. All rights reserved.

Keywords: Shrimp; Larvae; Algae; Artemia

1. Introduction

0044-8486/$ - see front matter D 2004 Elsevier B.V. All rights reserved.

doi:10.1016/j.aquaculture.2004.11.051

* Corresponding author. Texas Agricultural Experiment

Station–Shrimp Mariculture Research Facility, 4301 Waldron

Rd., Corpus Christi, TX 78418 USA. Tel.: +1 361 937 22 68;

fax: +1 361 937 64 70.

E-mail address: samocha@falcon.tamucc.edu (T.M. Samocha).

Development of cost-effective, nutritionally com-

plete inert larval feeds would be of tremendous benefit

to commercial hatchery operations. Despite extensive

research, nutrient requirements of aquatic suspension

2005) 135–147

C.B. Robinson et al. / Aquaculture 245 (2005) 135–147136

feeders have not been well understood (Villamar and

Langdon, 1993). Sorgeloos and Leger (1992)

described finfish and crustacean larviculture diets for

the early first feeding stages as being the major

bottleneck for complete replacement of live larval

feeds.

In the wild, the diet of larval shrimp typically

consists of diverse phytoplankton and zooplankton

species of various sizes and biochemical composition.

For that reason, traditional feeding techniques used in

commercial shrimp hatcheries still rely on complex

regimes of live food items. Larval rearing diets are

typically composed of different species of microalgae

and newly hatched Artemia nauplii (New, 1976;

Kumlu and Jones, 1995) and, as such, are difficult

to maintain (Persoone and Claus, 1980). Assurance of

adequate concentrations of live food items requires

careful and frequent monitoring and can result in

increased operating costs, variable nutritional quality

(Watanabe et al., 1983) and could serve as vectors for

shrimp pathogens (Kurmaly et al., 1989). These

limitations underscore the importance of identifying

alternative inert feeds as substitutes for live food

items.

The determination of nutrient essentiality to larval

penaeid shrimp is also complicated by a life cycle

consisting of a series of planktotrophic stages result-

ing in substantial trophic level change (Jones et al.,

1993). Of the small amount of information available,

most has been derived from three species endemic to

the Gulf of Mexico: Litopenaeus setiferus, Farfante-

penaeus aztecus, Farfantepenaeus duorarum and

various Pacific species: Marsupenaeus japonicus,

Litopenaeus vannamei and Penaeus monodon.

Microparticulate feeds in microencapsulated,

microbound and microcoated forms have met with

various levels of success in commercial hatcheries as

partial replacements for live foods (Langdon et al.,

1985) and their use often results in deterioration of

water quality (Muir et al., 1991). Degradation and

leaching of nutrients from inert feeds not only

impairs shrimp growth and survival but also depre-

dates the quality of culture water via proliferation of

bacteria and general fouling (Jones et al., 1987; Muir

et al., 1991). Furthermore, the use of inert dry feeds

typically requires increased water exchange and

supplemental aeration to maintain food particles in

suspension. Constant monitoring of culture water is

also required to avoid significant bio-fouling or

decline in water quality due to residual, uneaten

feed (Muir and Sutton, 1994). Studies comparing

liquid to dry inert feeds have shown increased

benefits of liquid products, including reduced tank

fouling and increased suspension time in the water

column (Jones et al., 1987). Liquid feeds also appear

to allow for a reduction in both water exchange and

aeration (Jones et al., 1987). Ultimately, the develop-

ment of nutritionally stable inert feeds could reduce

operational costs for penaeid shrimp hatcheries by

minimizing reliance on live foods with variable

nutrient composition.

Our study addressed live feed replacement in the

larval rearing of F. aztecus, a species indigenous to the

Gulf of Mexico. F. aztecus was selected to increase

the knowledge and understanding of how various

formulated diets as substitutes for live feeds affect the

larviculture of this species as well as to provide

insight into its commercialization for the live bait-

shrimp industry.

The objectives of this study were to (1) evaluate

the partial and complete replacement of live micro-

algae and newly hatched Artemia nauplii by various

commercially available inert larval feeds and (2)

comment on their application to larviculture of F.

aztecus.

2. Materials and methods

2.1. Source of shrimp and stocking

This study was conducted at the Texas Agricultural

Experiment Station (TAES) Shrimp Mariculture

Research Facility (SMRF), Corpus Christi, TX.

Shrimp larvae (F. aztecus) were provided by Lone

Star Farm (Austin, TX) from certified viral-pathogen-

free captive broodstock. Nauplii from a single spawn

were first acclimated to artificial seawater (35 ppt;

Instant Ocean Artificial Salt Mix, Aquarium Systems,

Mentor, OH) for a period of 60 min and then

transferred to a common holding tank and maintained

at a density of 100 nauplii L�1 until the initiation of

the study. Larvae (N5) from this holding tank were

used to stock experimental vessels for a microalgae

replacement study (Study 1). For the Artemia replace-

ment study (Study 2), the larvae were maintained in

C.B. Robinson et al. / Aquaculture 245 (2005) 135–147 137

this tank until Z3 and then stocked into cones. Larvae

in substages Z1 and Z2 were fed live diatoms

(Chaetoceros muelleri) at a density of 2.0�105 cells

mL�1. Each experimental tank (1 L Imhoff cone) was

stocked with 100 hand-counted larvae and filled with

1 L distilled artificial seawater.

2.2. Water source and monitoring

Artificial seawater was prepared daily to a salinity

of 35 ppt, 24 h in advance of use and completely

replaced in all culture vessels for both studies. In order

to chelate trace metals, EDTA was added to culture

water at 10 mg L�1. Dissolved oxygen (DO), pH,

salinity, temperature and ammonium–nitrogen con-

centrations were determined on a daily basis accord-

ing to Standard Methods (Clesceri et al., 1998).

Ammonium–nitrogen levels were subsequently used

to evaluate feed water stability.

2.3. Experimental system

The culture system used in Study 1 and Study 2

was a modification of the one developed by Wilken-

feld et al. (1984) and consisted of clear, 1-L plastic

Imhoff settling cones inserted into a common frame

and partially submerged in a thermally regulated

(29F1 C) water bath. Water circulation within the

bath was provided by a small submersible pump.

Aeration was supplied via a manifold system with

individual airlines connected to plastic pipettes

secured to the bottom of each cone. Rate of aeration

was limited to a slow trickle of bubbles which

maintained uniform distribution of larvae and food

particles within cones. A wooden-frame lid with a

clear plastic polyethylene cover was positioned on top

of the water bath to minimize evaporation and

introduction of airborne contaminants into the culture

medium. Continuous overhead illumination was

provided via two 60-W fluorescent light fixtures

suspended 1.5 m above the bath.

2.4. Daily monitoring and maintenance of experimen-

tal system

Survival and stage of metamorphosis were deter-

mined on a daily basis for all cones in both studies.

Larvae were also examined daily for extent of bio-

fouling and fullness of gut. Daily seawater replace-

ment in each cone was accomplished by first pouring

seawater through a sieve to concentrate larvae. Larvae

retained on sieves remained submerged for the

duration of the process. Filtrate from each exper-

imental cone was collected for subsequent ammo-

nium–nitrogen determination. Cones were then

refilled with newly cured artificial seawater. At this

point, microalgae, Artemia nauplii or inert feeds were

added to cones according to assigned treatments.

Larvae were then backwashed from collection sieves

into receiving cones.

2.5. Experimental feeds and feeding

Five experimental feeds (microalgal pastes, liquid

suspensions and dry powders) were used to evaluate

both 50% replacement and 100% replacement of

both, live microalga (C. muelleri; Study 1) and

Artemia nauplii (Study 2). A sevenfold level of

replication was used to evaluate a total of 11 dietary

treatments, including live control treatments of

microalgae and Artemia nauplii. Replacement feeds

for live microalgae included Chaetoceros 1000

bPremium FreshQ Instant Algaek (Reed Mariculture,

San Jose, CA), Liqualifek (Cargill, Inc., Minneap-

olis, MN), Epifeedk Liquid Larval Hatchery Feed

(Epicore Bionetworks, East Hampton, NJ), ZeiglerkZ-Plus (Zeigler Bros, Gardners, PA) and ZeiglerkE-Z Larvae (Zeigler Bros, Gardners, PA). Table 1

provides particle sizes and proximate analysis of inert

feeds. For Study 2, replacement of Artemia nauplii

was accomplished using the aforementioned inert

feeds. The only exception was replacement of Instant

Algaek, with E-Z Artemia (Zeigler Bros, Gardners,

PA). Preparation of inert larval feeds and final feed

concentrations for Study 1 and Study 2 followed

manufacturers’ instructions (Tables 2a and 2b,

respectively).

2.6. Termination of study and data collection

All cones within a dietary treatment were har-

vested when a mean of N80% of the larvae

metamorphosed to postlarva. Cones were harvested

by gently pouring contents onto a 500-Am sieve and

rinsing with fresh filtered artificial seawater. At this

point, 10 postlarvae were gently removed and

Table 1

Particle size, proximate analysis and marine fatty acid content of experimental feeds used in larval feeding trials with Farfantepenaeus aztecus

Experimental

feed

Particle

sizea (Am)

Crude

protein (%)bCrude

fat (%)bMoisture

(%)

Ash

(%)bDHAc

(%)

EPAd

(%)

E-Z Artemia 250–600 36.0 6.0 75.0 12.0

E-Z Larvae 10–100, 100–250 6.5 3.5 80.0 3.0 16.00 8.48

Z-Plus b100 50.0

100–150 12.0 8.0 12.0 6.07 7.55

150–250

Liqualifek 13–150 3.0

25–300 2.0 66.0 N/A 10.80 9.50

75–600

Epifeedk 4–44 14.0 15.0 43.0 5.0

4–124 14.0 15.0 39.0 5.0 8.32 8.65

30–400 19.0 16.0 36.0 6.0

Live microalgae 4–6 30.0 16.0 12.0 3.0 3.70 20.80

Live Artemia nauplii 400–500 50.0 19.0 6.1 8.0 TRe 1.0

a Particle size shown is that stated by manufacturer.b Dry matter concentration.c Docosohexaenoic acid; values are for one observation only.d Eicosohexaenoic acid; values are for one observation only.e Trace.

C.B. Robinson et al. / Aquaculture 245 (2005) 135–147138

subjected to a salinity stress-test similar to the one

described by Samocha et al. (1998). Postlarvae (PL)

were transferred into a 100-mL beaker filled with

fresh 35 ppt artificial seawater using dissecting scope

and a large-mouth eyedropper. From this beaker PL

were transferred into a separate cone filled with 17

ppt artificial seawater supplemented with gentle

aeration. After a 2-h exposure, PL were concentrated

on a sieve and survival was determined using a

dissecting scope. Following separation of the 10 PL

Table 2a

Feeding regimes and feed concentrations used in a larval feed study with

Treatment feed

(replacement level)

Feed concentrationa

Instant Algaek (50%) 0.5–1.0�105 cells mL�1

Instant Algaek (100%) 1.0–2.0�105 cells mL�1

Liqualifek (50%) 0.005–0.025 mL L�1 day�1

Liqualifek (100%) 0.010–0.050 mL L�1 day�1

Epifeedk (50%) 0.005–0.038 mL L�1 day�1

Epifeedk (100%) 0.010–0.075 mL L�1 day�1

Z-Plusk (50%) 0.005–0.080 mL L�1 day�1

Z-Plusk (100%) 0.010–0.160 mL L�1 day�1

E-Z Larvaek (50%) 0.009–0.043 mL L�1 day�1

E-Z Larvaek (100%) 0.018–0.086 mL L�1 day�1

Live microalgae Na

a Offered from Z1-PL1.b Offered from Z3-PL1.

for the stress-test, remaining animals on the sieve

were transferred into a labeled petri dish in 25 mL of

35 ppt fresh artificial seawater and placed in a

refrigerator for several hours to immobilize the

animals. At that point a dissecting scope was used

for counting, determination of larval developmental

stage and length measurement of 20 randomly

selected PL. Length measurements were taken from

the tip of the telson to the tip of the rostrum.

Harvested animals from each cone were rinsed with

Farfantepenaeus aztecus (Study 1)

Live algae concentration

(cells mL�1)aArtemia concentration

(nauplii mL�1)b

0.5–1.0�105 0.25–8.0

0 0.25–8.0

0.5–1.0�105 0.25–8.0

0 0.25–8.0

0.5–1.0�105 0.25–8.0

0 0.25–8.0

0.5–1.0�105 0.25–8.0

0 0.25–8.0

0.5–1.0�105 0.25–8.0

0 0.25–8.0

1.0–2.0�105 0.25–8.0

Table 2b

Feeding regimes and feed concentrations used in a larval feed study

with Farfantepenaeus aztecus (Study 2)

Treatment feed

(replacement

level)

Feed

concentration

(mL L�1 day�1)a

Artemia

concentration

(nauplii mL�1)a

Live

microalgae

(cells mL�1)b

E-Z Artemiak(50%)

0.016–0.050 0.13–4.0 0.25–8.0

E-Z Artemiak(100%)

0.032–0.092 0 0.25–8.0

Liqualifek(50%)

0.017–0.027 0.13–4.0 0.25–8.0

Liqualifek(100%)

0.034–0.054 0 0.25–8.0

Epifeedk(50%)

0.010–0.040 0.13–4.0 0.25–8.0

Epifeedk(100%)

0.020–0.080 0 0.25–8.0

Z-Plusk(50%)

0.012–0.090 0.13–4.0 0.25–8.0

Z-Plusk(100%)

0.024–0.150 0 0.25–8.0

E-Z Larvaek(50%)

0.016–0.050 0.13–4.0 0.25–8.0

E-Z Larvaek(100%)

0.032–0.092 0 0.25–8.0

Live Artemia

nauplii

na 0.25–8.0 0.25–8.0

a Offered from Z3-PL1.b Offered from Z1-PL1.

Table 3

Survival, final length and final weight of Farfantepenaeus aztecus

postlarvae in a study where larvae were offered inert feeds at the

50% and 100% replacement levels (Study 1)*

Treatment feed Survival (%) Mean length

(mm)

Mean weight

(mg)

50% replacement

Instant Algaek 85.00F4.55a 5.68F0.06c 0.239F0.01b

Liqualifek 88.14F2.04a 5.37F0.10b 0.201F0.01c

Epifeedk 79.43F8.99a 5.21F0.13b 0.197F0.01c

Z-Plus 71.00F20.53b 5.42F0.13b 0.212F0.02c

E-Z Larvae 69.14F23.79b 5.39F0.28b 0.208F0.02c

Live microalage 90.86F3.19a 6.55F0.24a 0.259F0.01a

100% replacement

Instant Algaek 0.00 (Day 4) na** na**

Liqualifek 10.00F6.16b 3.93F0.25b 0.153F0.02b

Epifeedk 2.00 2.58c 3.53F2.41b 0.139F0.10b

Z-Plus 10.14F9.23b 3.83F1.71b 0.147F0.07b

E-Z Larvae 14.86F14.35b 4.13F0.31b 0.160F0.03b

Live microalgae 90.86F3.19a 6.55F0.24a 0.259F0.01a

* Values are meansFS.D. Means within columns and having a

similar superscript are similar.

** na=not applicable; due to total mortality of larvae.

C.B. Robinson et al. / Aquaculture 245 (2005) 135–147 139

deionized water, blotted dry on absorbent paper and

placed on a marked pre-weighed microscope slide.

Slides were placed in an oven at 60 8C for 12 h.

Dried animals were group weighed, and individual

weight (reported in micrograms) was determined

based on the total number of animals on each slide.

2.7. Statistical analyses

Paired-samples Student’s t-test (SPSS; Chicago, IL)

was used to determine statistical differences in per-

formance factors (mean survival, final length, final dry

weight and stress-test survival) between level of

replacement of microalgae/Artemia nauplii and con-

trols for each experimental feed. Least Significant

Difference (LSD) and one-way ANOVAwere used at a

significance level of a=0.05 to identify statistical

differences among treatment means (SPSS; Chicago,

IL). All data were evaluated for normality and

homogeneity prior to analysis. Survival and stress-test

data were arcsine transformed prior to analysis.

3. Results

3.1. Replacement of live microalgae (Study 1)

3.1.1. Water quality analyses

Mean water temperature, salinity, dissolved oxygen

(DO) and pH in treatment cones was 29.1–29.9 C, 35

ppt, 5.15–6.08 mg L�1 and 7.93–8.40, respectively.

Ammonium–nitrogen ranged from 0.00 to 0.25 mg

L�1, except in experimental cones receiving ZieglerkZ-Plus at both the partial and complete-replacement

levels.

3.1.2. Growth and survival

Survival and final length and weight of shrimp

offered feeds as either partial or complete replacement

of live microalgae are shown in Table 3. Larvae offered

inert feeds at the 50% replacement level showed

increased survival, final length and final weight

compared to those fed at the 100% level (Pb0.05).

Analysis of survival, mean length and weight among

shrimp offered feeds at the 50% replacement level

showed a significant difference among treatments

(Pb0.02). Survival of larvae ranged from 69.1% to

88.1% (Table 3). Larvae fed Instant Algaek,

Liqualifek and Epifeedk demonstrated similar sur-

Table 4

Percentage survival by developmental substages of Farfantepe-

naeus aztecus larvae fed partial and complete replacement of live

microalgae (Study 1)*

Treatment Feed Z3 (%) M3 (%) PL1 (%)

50% replacement

Instant Algaek 97.86F0.69a 93.86F0.69b 85.00F4.55c

Liqualifek 98.00F0.82a 93.14F0.69b 88.14F2.04c

Epifeedk 97.71F0.76a 89.71F0.95b 79.43F8.99c

Z-Plus 97.43F0.98 a 95.14F1.22b 71.00F20.53c

E-Z Larvae 93.71F1.38 a 78.43F14.73b 69.14F23.79c

Live microalgae 97.86F0.69 a 96.29F1.11b 90.86F3.19c

100% replacement

Instant Algaek 88.71F1.11b 0.00 na**

Liqualifek 95.71F0.76b 35.43F3.55b 10.00F6.16b

Epifeedk 95.57F0.79b 26.29F2.81b 2.00F2.58b

Z-Plus 94.43F0.98b 31.00F3.92b 10.14F9.22b

E-Z Larvae 92.00F1.29b 28.43F3.10b 14.86F14.35b

Live microalgae 97.86F0.69a 96.29F1.11a 90.86F3.19a

* Values are meansFS.D. Means within columns and having a

similar superscript are similar.

** na=not applicable; due to total mortality of larvae at the Z3-

M1 molt.

C.B. Robinson et al. / Aquaculture 245 (2005) 135–147140

vival to those offered live microalgae (PN0.05). Other

inert feeds (Zeigler Z-Plusk and Ziegler E-Z

Larvaek) showed greatly reduced survival (Pb0.01).

Mean final length and weight of shrimp offered inert

feeds at the 50% replacement level ranged from 5.2 to

5.7 mm and 0.20 to 0.24 mg, respectively, with

Chaetoceros 1000 bPremium FreshQ Instant Algaekproducing the best performance (Pb0.01).

Significantly lower survival (Pb0.00) and final

mean length and weight (Pb0.02) were shown by

larvae offered inert feeds at the 100% replacement

level than those fed live microalgae. Survival of

shrimp offered inert feeds at the 100% replacement

level ranged from 0.00% to 14.9%, with the highest

numerical survival observed by larvae fed the Zeigler

E-Z Larvaek (Table 3). Shrimp fed the Zeigler E-Z

Larvaek, Liqualifek and Zeigler Z-Plusk feeds had

no significant differences in survival rates. However,

survival in these treatments was significantly better

than the shrimp fed the Epifeedk. Complete mortality

occurred within 4 days post-stocking for larvae

offered Instant Algaek. Final length and weight of

shrimp offered inert feeds at the 100% replacement

level ranged from 3.5–4.1 mm and 0.14–0.16 mg,

respectively, with Zeiglerk E-Z Larvae exhibiting the

best numeric performance, though not statistically

different from Liqualifek and Z-Plusk feeds.

In conclusion, except for similar survivals of the

Instant Algaek, Liqualifek and Epifeedk treat-

ments at the 50% replacement level of live algae, final

mean length and dry weight of larvae offered the

control feed were significantly higher than animals in

all other inert feed treatments (Pb0.009).

3.1.3. Survival, by developmental stage

Mean percent survival of larvae offered inert feeds

by developmental stage is presented in Table 4.

Survival through Zoea was better than through Mysis

at both levels of microalgae replacement (P=0.009). A

similar relationship was shown in terms of survival

through M3 compared to PL1 (P=0.030). The same

relationship was shown for shrimp fed the live micro-

algae control. Highest mortality of larvae offered inert

feeds (50% replacement) and live algae occurred

during metamorphosis from M3 to PL1 (Table 4).

Survival through M3 decreased as much as 24% for

shrimp offered inert feeds at the 50% replacement level

versus 5.4% for shrimp offered live microalgae. Shrimp

offered inert feeds at the 100% replacement level

experienced the greatest mortality during metamor-

phosis from Zoea to Mysis, at which point survival

decreased as much as 64%.

3.1.4. Marine fatty acid content and survival

Concentrations of DHA among the inert feeds and

the microalga control ranged from 3.7% to 16.0% and

showed no apparent trends for the 50% and the 100%

replacement levels (Table 5). Live microalgae, receiv-

ing the lowest concentration of DHA (3.7%), elicited

highest survival (90.9%).

Concentrations of EPA in inert feeds and the

microalga control ranged from 7.6% to 20.8% and

appeared to be positively correlated with survival

(Table 5). Shrimp offered live microalgae (highest

EPA concentration, 20.8%) had the highest percentage

survival (90.9%) among both treatments. These

results suggest that this level of EPA concentration

does not adversely affect survival.

3.1.5. Ammonium–nitrogen

Day 10 ammonium–nitrogen concentrations in

experimental cones receiving 50% replacement of live

microalgae were similar to those at the 100% level

(PN0.05). Analysis of day 10 ammonium–nitrogen in

Table 6

Day 10 ammonium–nitrogen concentration of treatment cones at the

50% and 100% replacement levels of live microalgae (Study 1)

Treatment feed Mean ammonium–nitrogen

concentration (mg L�1)*

50 % replacement

Instant Algaek 0.125F0.10a

Liqualifek 0.161F0.04a

Epifeedk 0.241F0.07a

Z-Plus 1.889F0.44b

E-Z Larvae 0.215F0.12a

Live microalgae 0.154F0.09a

100% replacement

Instant Algaek 0.197F0.02ac

Liqualifek 0.088F0.06a

Epifeedk 0.021F0.03ad

Z-Plus 2.139F0.34b

E-Z Larvae 0.139F0.05a

Live microalgae 0.154F0.09a

* Means with similar superscripts are not significantly different

( PN0.05).

Table 5

Comparison of Farfantepenaeus aztecus postlarvae survival and

fatty acid content of inert feeds offered at two different rates

Treatment feed Survival (%) % EPAa % DHAb

50% replacement

Liqualifek 88.14 9.50 10.80

Epifeedk 79.43 8.65 8.32

Z-Plus 71.00 7.55 6.97

E-Z Larvae 69.14 8.48 16.00

Live microalgae 90.86 20.80 3.70

100% replacement

Liqualifek 10.00 9.50 10.80

Epifeedk 2.00 8.65 8.32

Z-Plus 10.14 7.55 6.97

E-Z Larvae 14.86 8.48 16.00

Live microalgae 90.86 20.80 3.70

a Eicosopenaenoic acid.b Docosohexaenoic acid.

C.B. Robinson et al. / Aquaculture 245 (2005) 135–147 141

experimental cones receiving 50% replacement of live

microalgae showed a significant difference among

treatments (Pb0.00), with concentrations ranging from

0.125 to 1.889 mg L�1 (Table 6). Cones receiving

Zeiglerk Z-Plus elicited a significantly higher ammo-

nium–nitrogen at day 10 than all other algal replace-

ment feeds (Pb0.00), including live microalgae.

Analysis of day 10 ammonium–nitrogen in exper-

imental cones receiving 100% replacement of live

microalgae showed a significant difference among

treatments (Pb0.00), with concentrations ranging from

0.088 to 2.139 mg L�1 (Table 6). Cones receiving

Zeiglerk Z-Plus had a significantly higher day 10

ammonium–nitrogen concentration than all other rep-

lacement feeds (Pb0.00), including live microalgae.

Furthermore, with the exception of Instant Algaekand Z-Plus, it appears that ammonium–nitrogen levels

were lower in the 100% replacement in terms of

reduced water quality degradation. Nevertheless, even

if the stability of some of the inert feeds means a slight

improvement in water quality, the dominating factor is

the poor survival and the growth of the larvae on these

feeds.

3.2. Replacement of live Artemia nauplii (Study 2)

3.2.1. Water quality analyses

Mean water temperature, salinity, dissolved oxygen

and pH in treatment cones was 28.8–29.5 8C, 35 ppt,

5.4–6.1 mg L�1 and 8.1–8.3, respectively. Ammo-

nium–nitrogen ranged from 0.00 to 0.55 mg L�1,

except in cones fed Zieglerk Z-Plus at both the 50%

and 100% replacement levels where maximum con-

centrations of 1.94 and 3.56 mg L�1, respectively.

3.2.2. Growth and survival

Survival, final length, final weight and stress-test

survival of shrimp offered inert feeds at the 50%

replacement level of live Artemia (Table 7) were

greater than corresponding values for the 100%

replacement level (Pb0.04, Table 7). There was a

significant difference in survival, final length and final

weight of shrimp fed the 50% replacement feeds

(Pb0.01). Shrimp offered live Artemia exhibited

significantly higher survival rates than those fed inert

replacement feeds (Pb0.05), with the exception of

those fed Liqualifek (P=0.22). Overall survival of

shrimp fed 50% replacement feeds ranged from 87.1%

to 92.1%, with the highest numeric survival seen in

those offered the Liqualifek. Shrimp offered

Liqualifek exhibited significantly higher survival than

all others fed 50% replacement diets. Stress-test

survival was similar among all shrimp fed the 50%

replacement feeds (P=0.01). Stress-test survival of

shrimp offered 50% replacement feeds ranged from

7.1% to 12.9%, with the highest numeric survival

shown by those fed Zeiglerk E-Z Larvae. Shrimp

Table 7

Survival, final length, final weight and stress-test survival of postlarvae when larvae were offered inert feeds at the 50% and 100% live Artemia

nauplii replacement levels (Study 2)*

Treatment feed Survival (%) Mean length

(mm)

Mean weight

(mg)

Stress-test

survival (%)

50% replacement

E-Z Artemia 89.71F4.39bc 5.55F0.08cd 0.287F0.01c 7.14F7.56b

Liqualifek 92.14F1.95a 5.91F0.16b 0.304F0.01b 10.00F8.17b

Epifeedk 90.43F1.72bc 5.71F0.18bc 0.294F0.01bc 10.00F8.17b

Z-Plus 87.14F3.02c 5.87F0.12b 0.300F0.01b 8.57F6.90b

E-Z Larvae 90.86F1.77b 5.51F0.12de 0.281F0.01cd 12.86F7.56b

Live Artemia nauplii 93.43F4.32a 6.95F0.22a 0.356F0.01a 18.57F6.90a

100% replacement

E-Z Artemia 81.14F1.95bc 4.26F0.13e 0.217F0.01e 1.43F3.78b

Liqualifek 80.14F1.35c 5.40F0.15b 0.278F0.01b 1.43F3.78b

Epifeedk 83.14F1.86b 5.18F0.21c 0.263F0.02c 1.43F3.78b

Z-Plus 81.14F2.27b 4.57F0.20d 0.235F0.01d 2.86F4.88b

E-Z Larvae 84.29F3.90b 5.32F0.09b 0.273F0.01b 2.86F4.88b

Live Artemia nauplii 93.43F4.32a 6.95F0.22a 0.356F0.01a 18.57F6.90a

* Values are meansFS.D. Means within columns and having a common superscript are similar.

C.B. Robinson et al. / Aquaculture 245 (2005) 135–147142

offered the live Artemia diet exhibited significantly

higher survival in the stress-test than seen in with all

other Artemia replacement feeds. Shrimp offered inert

replacement feeds showed significantly lower final

length and weight than those fed live Artemia

(Pb0.000). Final length and weight of shrimp fed

50% replacement feeds ranged from 5.5 to 5.9 mm and

0.281 to 0.304 mg, respectively. Shrimp fed

Liqualifek exhibited significantly greater final length

and weight compared to those fed Zeiglerk E-Z

Artemia ( P=0.001) or Zeiglerk E-Z Larvae

(Pb0.000). Shrimp fed Liqualifek exhibited similar

final length and weight gains compared to those fed

Epifeedk (P=0.06) and Zeiglerk Z-Plus (P=0.44).

Shrimp offered the live Artemia control had

significantly higher survival, mean length, mean

weight and survival in stress-test than those offered

inert feeds at the 100% replacement level (Pb0.00). A

significant difference in survival, final length, final

weight and stress-test survival was also shown among

shrimp offered inert feeds (Pb0.000). Survival of

shrimp fed at the 100% replacement level ranged from

80.1% to 84.3%, with the highest numeric survival

shown by those offered Zeiglerk E-Z Larvae. Shrimp

fed this feed had significantly greater survival than

those fed Liqualifek (P=0.04), but not those fed

Zeiglerk E-Z Artemia (P=0.10), Epifeedk (P=0.5)

or Zeiglerk Z-Plus (P=0.10).

Shrimp fed inert feeds at the 100% replacement

level showed significantly lower final length and

weight than those offered live Artemia nauplii

(Pb0.00). Mean final length and weight of shrimp

fed 100% replacement feeds ranged from 4.3 to 5.4

mm and 0.217 to 0.278 mg, respectively, with

Liqualifek showing the best results. Shrimp fed

Liqualifek had significantly greater final length and

weight than those fed Zeiglerk E-Z Artemia ,

Epifeedk and Zeiglerk Z-plus (Pb0.004), but not

those fed Zeiglerk E-Z Larvae.

Shrimp fed live Artemia nauplii had significantly

higher stress-test survival than those fed any of the

replacement feeds (Pb0.00). Stress-test survival of

shrimp fed inert feeds at the 100% replacement level

ranged from 1.4% to 2.9%, with the highest numeric

survival seen in those fed Zeiglerk Z-Plus and E-Z

Larvae. No significant difference in stress-test survival

was shown among shrimp fed inert feeds (P=0.50).

3.2.3. Percentage survival by developmental stage

Survival of shrimp through metamorphosis to

Postlarva is presented in Table 8. Survival through

M3 exceeded that through PL1 for all types of inert

feeds at all levels of replacement (P=0.003). Survival

of shrimp fed live Artemia nauplii through M3

exceeded that through PL1 (P=0.038). Survival from

M3 through PL1 decreased as much as 15% for shrimp

Table 8

Percentage survival of Farfantepenaeus aztecus developmental

substages when inert feeds were fed at the 50% and 100%

replacement levels of live Artemia nauplii (Study 2)*

Treatment feed M3 PL1

50% replacement

E-Z Artemia 96.43F1.13 89.71F4.39

Liqualifek 97.14F1.22 92.14F1.95

Epifeedk 96.00F1.16 90.43F1.72

Z-Plus 95.86F1.07 87.14F3.02

E-Z Larvae 95.86F0.90 90.86F1.77

Live Artemia nauplii 97.00F1.00 93.43F4.32

100% replacement

E-Z Artemia 94.86F1.35 81.14F1.95

Liqualifek 95.00F1.41 80.14F1.35

Epifeedk 94.86F0.90 83.14F1.86

Z-Plus 95.71F1.38 81.14F2.27

E-Z Larvae 94.57F0.98 84.29F3.90

Live Artemia nauplii 97.00F1.00 93.43F4.32

* Values are meansFS.D. None of the values shown, by

substage of metamorphosis, were significantly different ( Pb0.050).

Table 9

Survival, stress-test survival of Farfantepenaeus aztecus postlarvae

and fatty acid concentrations of experimental feeds offered at the

50% and 100% replacement levels (Study 2)

Treatment feed Survival

(%)

Stress-test

survival (%)

% EPAa % DHAb

50% replacement

E-Z Artemiak 89.71 7.14 5.21 4.84

Liqualifek 92.14 10.00 9.50 10.80

Epifeedk 90.43 10.00 8.65 8.32

Z-Plus 87.14 8.57 7.55 6.97

E-Z Larvae 90.86 12.86 8.48 16.00

Live Artemia nauplii 93.43 18.57 1.00 TRc

100% replacement

E-Z Artemiak 81.14 1.43 5.21 4.84

Liqualifek 80.14 1.43 9.50 10.80

Epifeedk 83.14 1.43 8.65 8.32

Z-Plus 81.14 2.86 7.55 6.97

E-Z Larvae 84.29 2.86 8.48 16.00

Live Artemia nauplii 93.43 18.57 1.00 TR

a Eicosopentaenoic acid; values shown constitute one observa-

tion only.b Docosohexaenoic acid; values shown constitute one observa-

tion only.c Trace.

C.B. Robinson et al. / Aquaculture 245 (2005) 135–147 143

offered inert feeds at the 100% replacement level. On

the other hand, this decrease did not exceed 8% for

shrimp fed at the 50% replacement level. Shrimp fed

live Artemia nauplii experienced a decrease in

survival of only 4% for the same period (e.g.,

metamorphosis from M3 to PL1).

3.2.4. Marine fatty acid content and survival

Comparison of concentrations of EPA and DHA in

inert feeds and live Artemia nauplii to overall survival

and stress-test survival suggest some trends (Table 9).

At the 50% replacement level, shrimp offered live

Artemia nauplii (lowest EPA concentration) exhibited

the highest overall survival (93.4%) and stress-test

survival (18.6%). Concentration of EPA among inert

feeds ranged from 5.21% to 9.50% and showed no

apparent influence on overall survival at the 50%

replacement level. Among shrimp offered inert feeds,

those offered Liqualifek exhibited the highest EPA

concentration (9.50%) and percent survival (92.1%).

Shrimp offered E-Z Larvae (EPA=8.5%) demonstrated

the highest stress-test survival (12.9%). There were no

apparent trends in EPA concentration and overall

survival or stress-test survival at the 100% replacement

level.

Shrimp offered live Artemia nauplii (trace amounts

of DHA) exhibited the highest overall survival

(93.43%) and stress-test survival (18.6; Table 9).

Among the inert feeds, shrimp offered E-Z Larvae

(highest DHA concentration of 16.00%) at the 50%

replacement level had the highest stress-test survival

(12.9%). Shrimp offered E-Z Larvae (DHA=16.00%)

at the 100% replacement level showed the highest

overall survival (84.3%) and stress-test survival

(2.9%).

3.2.5. Ammonium–nitrogen

Day 7 ammonium–nitrogen concentrations in

100% Artemia replacement cones were significantly

different from those offered inert feeds at the 50%

replacement level (Pb0.04), with the exception of the

Zeiglerk E-Z Larvae treatment (P=0.32). A signifi-

cant difference (Pb0.00) was shown in day 7

ammonium–nitrogen concentrations, which ranged

from 0.001 to 1.938 mg L�1 among inert feeds

offered at the 50% replacement level (Table 10). The

highest concentration was shown in cones containing

Zeiglerk Z-Plus (1.938 mg L�1). This feed yielded a

significantly higher ammonium–nitrogen concentra-

tion (Pb0.00) than all other replacement feeds,

including live Artemia nauplii.

Table 10

Day 7 ammonium–nitrogen concentration of treatment cones at the

50% and 100% replacement levels of live Artemia nauplii (Study 2)

Treatment feed Mean ammonium–nitrogen

concentration (mg L�1)*

50% replacement

E-Z Artemia 0.039F0.05b

Liqualifek 0.001F0.02b

Epifeedk 0.424F0.07b

Z-Plus 1.938F0.13a

E-Z Larvae 0.087F0.05b

Live Artemia nauplii 0.127F0.07b

100% replacement

E-Z Artemia 0.356F0.08b

Liqualifek 0.113F0.12b

Epifeedk 0.548F0.18b

Z-Plus 3.563F0.12a

E-Z Larvae 0.061F0.04b

Live Artemia nauplii 0.127F0.07b

* Means with similar superscripts are not significantly different

( PN0.05).

C.B. Robinson et al. / Aquaculture 245 (2005) 135–147144

A significant difference (Pb0.00) was seen in day

7 ammonium–nitrogen concentration of cones receiv-

ing inert feeds at the 100% replacement level (Table

10): concentrations ranged from 0.061 to 3.563 mg

L�1, with the highest occurring in the Zeiglerk Z-

Plus cones. This feed elicited a significantly higher

ammonium–nitrogen concentration than all other

replacement feeds (Pb0.00), including live Artemia

nauplii (Pb0.00).

4. Discussion

Replacement of live microalgae with inert feeds

appeared to be marginally successful at the 50%

replacement level; however, complete replacement

could not be achieved without a significant negative

effect on final weight and length and survival of

shrimp. Complete mortality of shrimp fed Instant

Algaek (a paste) could have been associated with

nutritional deficiencies associated with inappropriate

preparation procedures, as recommended by the

manufacturer. Microscopic evaluation of this paste-

like product showed substantial lysed and small-

diameter cells compared to that of the live microalga

control. This observation suggests that shrimp offered

this feed may have not received adequate nutrition.

Overall performance of shrimp offered inert feeds

at the 100% replacement level was poor with respect

to survival and final length and weight. Other studies

with different inert feeds have reported good survival

but only moderate growth of larval shrimp offered

these feeds (Kanazawa et al., 1982; Jones et al., 1987;

Smith and Lawrence, 1987; Kurmaly et al., 1989;

Kumlu and Jones, 1995). Although no explanation for

their results was provided, these studies suggested

further investigation to determine whether inert feeds

were nutritionally imbalanced, less digestible or of a

non-suitable particle size. It was further suggested that

it may be necessary to evaluate and characterize the

potentially toxic conditions caused by the increased

release of toxic nitrogenous compounds via ammoni-

fication and/or nitrification.

Jones et al. (1979) demonstrated that the use of

non-encapsulated microparticulate feeds warranted

strict monitoring of feed levels and quality to avoid

deterioration of water quality and associated adverse

effects on growth and survival of M. japonicus.

Nitrogenous products including ammonium, nitrite

and nitrate were reported to be toxic to several marine

organisms at extremely low concentrations (Spotte,

1979; Muir et al., 1991). Muir et al. (1991) reported

that degradation of microencapsulated feeds yielded

an increase in dissolved ammonium–nitrogen within

1.5 h of addition to culture water, which surpassed a

two- to threefold increase within 48 h of the addition

of microcapsules. These concentrations were not

considered as toxic until the final 2 days of the trial.

In the present studies, sub-lethal ammonium–nitrogen

concentrations were detected within a short time after-

feeding (24 h) and steadily increased until the end of

the trial. Cones receiving the Zeiglerk Z-Plus had

ammonium–nitrogen concentrations as high as 2.14

mg L�1 in the live algae study and 3.56 mg L�1 in the

live Artemia study. This level is substantially higher

than the 0.13 mg L�1 considered toxic to shrimp

(Chin and Chen, 1987). Chin and Chen (1987)

reported ammonium–nitrogen 24- and 96-h LC50s

of 5.71 and 1.26 mg L�1, respectively, for postlarval

P. monodon. The present studies failed to elucidate a

direct relationship between sub-lethal concentrations

of ammonium–nitrogen and reduced growth and/or

survival.

Shrimp excretion of NH3–N probably contributed

to overall ammonium–nitrogen concentrations in the

C.B. Robinson et al. / Aquaculture 245 (2005) 135–147 145

present studies, but was tempered by survival.

Wajsbrot et al. (1989) reported excretion rates for

various sizes of Penaeus semisulcatus were highest

between 4 and 8 h post-feeding. The dominant waste

product in their study was NH3–N (61–83%).

Although larval shrimp were not included in that

study, a trend of decreasing NH3–N excretion per unit

body weight was seen as shrimp weight increased. In

the present studies, higher levels of ammonium–

nitrogen were observed with the Zieglerk Z-Plus

treatments in both the live algae and live Artemia

replacement studies. In the live algae study, with the

exception of Instant Algaek and Z-Plus, it appears

that ammonium–nitrogen levels were lower in the

100% replacement treatments in terms of reduced

water quality degradation. These results are not what

would be expected when increasing substitution levels

from 50% to 100%. A high level of shrimp mortality

was recorded early on in the study with the 100%

replacement treatments. The fall in larval density in

these cones means a decrease in excretion rates. It also

means fewer animals are feeding on and breaking

down the particles. The stability of the test feeds

would be intake for a longer period of time. Break-

down of the feed particles would take longer as would

the rise in ammonium–nitrogen levels. The result

would probably be a decrease in ammonium–nitrogen

concentrations. The live Artemia study supports these

findings. Ammonium–nitrogen levels were higher in

the 100% replacement treatments. Shrimp survival

throughout the study was higher which means an

increase in larval excretion rates and a more rapid

breakdown of feed particle stability due to feeding.

Even if the stability of some of the inert feeds means a

slight improvement in water quality, the dominating

factor is the poor survival and the growth of the larvae

on these feeds. Nevertheless, no clear trend of

decreased survival was seen as a result of these

higher ammonium–nitrogen levels. Although the 50%

replacement of live algae and live Artemia out-

performed the 100% replacement levels, feeding the

larvae live algae and live Artemia provided signifi-

cantly improved survival, growth and performance in

stress-tests than all inert feeds tested in the present

study.

Feed particle diameter could have reduced growth

and survival of larval shrimp. Feed particles must be

of an appropriate size in order to assure maximum

rate of ingestion. Jones et al. (1979) reported that the

optimum particle size for Zoea stage M. japonicus

was approximately 10 Am. Those same authors

observed a trend towards selection of larger particles

at M2–M3 and suggested an optimum particle

diameter approximating 28 Am. They measured the

median particle diameter (Md) of capsules prior to

and post-feedings. In the majority of replicates, Md

increased for subsequent feedings, suggesting larvae

were selecting for more suitable-diameter particles

and rejecting larger diameter particles. In the present

study, which focused on larval feeding of F. aztecus,

it is possible that feed particle size, especially during

early Zoea stage, did not possess the proper Md.

Guts of shrimp offered inert feeds at the 100%

replacement level during Zoea and early Mysis

stages were seldom observed to be full, whereas

larvae offered inert feeds at the 50% replacement

level typically revealed full guts throughout the

duration of the study. As shown in Table 1, inert

feeds had minimum particle sizes which were of

similar, smaller or slightly larger diameter than that

recommended by Jones et al. (1979). The only

exception was Ziegler Z-Plus, which had a minimum

particle size of bb100 Am.Q The maximum particle

size for all inert feeds evaluate was substantially

outside this optimal range, especially with respect to

the Zoea stage. Frequency distribution of particles

sizes of inert feeds was not evaluated; however, it is

postulated that a high proportion were inappropriate

and thus contributed to higher overall survival in

Study 2. Most mortality was experienced during the

metamorphosis from Zoea to Mysis stages, especially

for shrimp fed the 100% replacement treatments

(Table 4).

According to Jones et al. (1979), to ensure high

growth and survival rates, particle size and feed rates

must be closely controlled, implying a need for

increased frequency of feeding of an appropriate

particle size. Unfortunately, with the inert feeds tested

in Study 1 (microalgae replacement) this recommen-

dation would have ultimately resulted in substantial

waste of expensive feed and a reduced cost/benefit to

commercial hatcheries. Probably the most efficient

means of improving microparticle feed efficiency lies

in reducing variance in particle Md and could

ultimately be derived from improvements in feed

particle manufacturing technology.

C.B. Robinson et al. / Aquaculture 245 (2005) 135–147146

The replacement of Artemia nauplii with inert test

feeds appeared marginally successful. Larvae offered

the live control diet in this study performed

significantly better in terms of final length, and

weight and performance in stress-tests than those fed

any of the inert feeds. The only exception was that

of similar survival associated with replacement of

Artemia nauplii with Liqualifek at the 50% level.

The significantly better performance of the control

diet suggests that survival by itself should not be

used as the sole criteria to select inert diets for larval

shrimp. Potential for replacement of live feed at

lower rates than those tested in the present study

(e.g., 10–25%) should be evaluated to determine

whether reduction in live feed consumption is

feasible without compromising shrimp growth and

performance in stress-tests. The results from Study 1

suggest that further investigation is necessary to

determine whether inert feeds offers a less nutrition-

ally balanced diet, a less digestible diet, or inad-

equate particle sizes that are rejected by larval

shrimp. Additional research is also required to study

the long-term effects of inert feeds on growth and

survival of the shrimp in the nursery and grow-out

phases.

5. Conclusions

Partial and complete replacement of live micro-

algae at the 50% and 100% level showed inferior

performance to the live microalgae control diet.

Similarly, Study 2 suggests that significantly better

results were obtained when the larvae were fed live

Artemia (control diet) than with any of the inert feeds.

No clear conclusion can be made concerning the

correlation between the concentrations of EPA and

DHA in the inert feeds and the performance of the

larvae. More in-depth studies are needed to clarify this

issue.

Acknowledgments

Funding for this research was provided by Lone

Star Farm Inc. and Texas Agricultural Experiment

Station-Shrimp Mariculture Research Facility. Spe-

cial thanks to the Lone Star Farm staff for the help

provided. Special thanks also to Reed Mariculture,

Inc., Cargill, Inc., Epicore Bionetworks, Inc. and

Zeigler Bros. Inc. for donating the feeds used in

these studies. Finally, we thank Mrs. Linda Smith

Lemmon for her help in improving the quality of this

manuscript.

References

Chin, T.S., Chen, J.C., 1987. Acute toxicity of ammonia to larvae of

the tiger prawn, Penaeus monodon. Aquaculture 66, 247–253.

Clesceri, L.S., Greenberg, A.E., Eaton, A.D., 1998. Standard

Methods for the Examination of Water and Wastewater, 20th

ed. American Public Health Association, Washington, DC,

USA.

Jones, D.A., Kanazawa, A., Abdel Rahman, S., 1979. Studies on the

presentation of artificial diets for rearing the larvae of Penaeus

japonicus (Bate). Aquaculture 17, 33–43.

Jones, D.A., Kurmaly, K., Arshard, A., 1987. Penaeid shrimp

hatchery trials using microencapsulated diets. Aquaculture 64,

133–146.

Jones, D.A., Kamarudin, M.S., LeVay, L., 1993. The potential

replacement of live feed in larval culture. Journal of the World

Aquaculture Society 24 (2), 199–210.

Kanazawa, A., Teshima, S., Sasada, H., Rahman, S.A., 1982.

Culture of prawn larvae with micro-particulate diets. Bulletin of

the Japanese Society of Scientific Fisheries 48 (2), 195–199.

Kumlu,M., Jones, D.A., 1995. The effect of live and artificial diets on

growth, survival and trypsin activity in larvae ofPenaeus indicus.

Journal of the World Aquaculture Society 26 (4), 406–415.

Kurmaly, K., Jones, D.A., Yule, A.B., East, J., 1989. Comparative

analysis of the growth and survival of Penaeus monodon

(Fabricius) larvae, from protozoea I to postlarvae I, on live

feeds, artificial diets and on a combination of both. Aquaculture

81, 25–27.

Langdon, C.J., Levine, D.M., Jones, D.A., 1985. Microparticulate

feeds for marine suspension-feeders. Journal of Microencapsu-

lation 2 (11), 1–11.

Muir, P.R., Sutton, D.C., 1994. Bacterial degradation of micro-

encapsulated foods used in larval culture. Journal of the World

Aquaculture Society 25 (3), 371–378.

Muir, P.R., Sutton, D.C., Owens, L., 1991. Nitrate toxicity to

Penaeus monodon protozoea. Marine Biology 108, 67–71.

New, M.B., 1976. A review of dietary studies with shrimp and

prawns. Aquaculture 9, 101–144.

Persoone, G., Claus, C., 1980. Mass culture of algae: a bottleneck in

the nursery culture of mollusks. In: Shelef, G., Sveder, C.J.

(Eds.), Algal Biomass. Elsevier North Holland Biomedical

Press, Amsterdam, pp. 265–285.

Samocha, T.M., Guajardo, H., Lawrence, A.L., Castille, F.L., Speed,

M., Mckee, D.A., Page, K.I., 1998. A simple stress test for

Penaeus vannamei postlarvae. Aquaculture 165, 233–242.

Smith, L.L., Lawrence, A.L., 1987. Experimental use of dry diets in

penaeid larviculture as replacements for Artemia. Journal of the

World Aquaculture Society 18, 17A.

C.B. Robinson et al. / Aquaculture 245 (2005) 135–147 147

Sorgeloos, P., Leger, P., 1992. Improved larviculture outputs of

marine fish, shrimp and prawn. Journal of the World Aqua-

culture Society 23 (4), 251–264.

Spotte, S., 1979. Seawater Aquariums. John Wiley and Sons, New

York, NY, USA.

Villamar, D.F., Langdon, C.J., 1993. Delivery of dietary compo-

nents to larval shrimp (Penaeus vannamei) by means of

complex microcapsules. Marine Biology 115, 635–642.

Wajsbrot, N., Krom, M.D., Gasith, A., Samocha, T., 1989.

Ammonia excretion of green tiger prawn Penaeus semisulcatus

as a possible limit on the biomass density in shrimp ponds. The

Israeli Journal of Aquaculture-Bamidgeh 41 (4), 159–164.

Watanabe, T., Kitajima, C., Fujita, S., 1983. Nutritional value of live

organisms used in Japan for mass propagation of fish: a review.

Aquaculture 34, 115–143.

Wilkenfeld, J.S., Lawrence, A.L., Kuban, F.D., 1984. Survival,

metamorphosis and growth of penaeid shrimp larvae reared on a

variety of algal and animal food. Journal of the World

Mariculture Society 15, 31–49.