Status of research during April 1 through June 30, 1999, on the effect of PUFAs on the growth and development of the Tautog (Tautoga onitis) and proposed studies for remainder of 1999

I.  Progress

A) Morphometric analysis

1) The techniques for the computer based measurement of morphological characteristics have been successfully applied to initial fixed samples of larval tautog. The identification of useful landmarks allows for a rapid measurement of growth and development. This approach also provides the ability to monitor for deformities as might occur (particularly in the cranial region) with nutritional deficiencies. At present, a comprehensive analysis is is limited by sample size and the use of fixed samples for this purpose for working out the methods.

B) Lipid analysis

1) Analysis of lipids, both gravimetric and gas chromatographic, has been carried out on three algae, three adult tautog foods and individual larval (n=2) and juvenile tautog (n=14).

2) The lowest quantity of tissue used in the analysis above was 26 mg. The capability to perform analysis on quantities as low a 5 mg has been developed making analysis of individual larval tautog possible.

3) The use of 5 commercially available fatty acid standards has allowed for the identification of some 45 fatty acids; greater than 80 % of observed peaks are identified.
 
 

C) Lipid data analysis

1) a comprehensive table of fatty acid compositions can be found on http://www.bio.umass.edu/biology/kunkel/fish/tautog/. Below are condensed figures of the fatty acid composition data collected from the three algae used in 1998 (below). Percent composition is shown on the y axes. Each of these three algae have distict PUFA profiles obviated by the categories of carbon chain length and by double bond pattern (Figure 1). This includes a pair of rare C:16 PUFA’s found in the Ply429 culture.

Figure 1. Algal strain differences in PUFA content.

The relative differences occur in these three algae while the total fraction of fatty acids is similar in the three groups (Figure 2).

Figure 2. Algal strain similarity in FA content.

It is our intent, based on the above data and capabilities, to carry out the studies outlined below.
 
 

II.  Proposed study goals and designs

A.  Experiment 1

Objective: Determine if different FA’s, including PUFA’s are accumulated in rotifers

Rationale: For tautog, survival and growth up to the time when young will take commercial feed has been a crucial hurdle in the development of a culture protocol. Current literature suggests that fatty acids, and in particular polyunsaturated fatty acids (PUFAs) are crucial during the early growth of fish and potentially tautog. Prior to commercially prepared feed, tautog are fed rotifers. The rotifers in turn are fed with algae from a varied sources. The pertinent question is whether or not the use of any given source of algae to raise the forage food, rotifers, would benefit the growth and development of tautog larvae. If this is indeed the case, (composition of the algae having an effect on tautog development) then these compounds must be accumulated in the intermediate, rotifers. If the important compounds are the PUFAs, then these FAs would be quantifiably different in rotifers under different feeding regimes.

The fatty acid composition of three types of algae currently in use (PLY 249, UTEX 2341, and T150) has been determined. These algae have differing FA compositions as demonstrated above. It is necessary to compare the fatty acid profiles of rotifers reared on each of the measure fatty acids reared with each of the algal types.

Design:

1) Four rotifer cultures will be maintained, of each being fed each of the three algae types or combinations of them (e.g. PLY 249, UTEX 2341, T150, or all three algae).

2) From each culture, approximately 0.15 g of rotifers must be thoroughly separated from the algae being used to feed them.

3) Each of the cultures will be filtered and a wet weight determined; a fraction of the purified rotifers will be used to determine the dry weight/wet weight ratio.

4) The remaining sample will be used to a) determine protein content b) determine lipid content c) determine fatty acid profiles
 

B.  Experiment 2

Objective: Determine if rotifers raised on different algae result in different FA accumulation, and growth of tautog.

Rationale: At the heart of this grant is the is the determination whether rotifers fed a given algae diet can positively affect the initial growth and and development of tautog larvae up until they can take commercial feed. The most important parameters to be measured in an aquaculture setting are survival and growth. Survival in tautog is varied and problematic, therefore growth is the best quantifiable measure. Morphological measurements through the growth period can be used to compare allometric processes.

Design: Tautog from an individual spawning event will be reared until hatching. Approximately 20 individuals will be removed and fixed at this time for morphometric study.  Ideally, four groups of tautog will be reared on rotifers fed PLY 249, UTEX 2341, T150, or all three algae. If numbers are sufficient, duplicates could be used to test for tank effects and fish will be sampled at 3 wks and 5 wks. The most important time for sampling is when the fish are likely to begin to take commercial feed. This time marks the end of potential algal influence on growth in culture.

At each point, a minimum of 10 fish from each group will be removed, anesthetized in 100 mg/L MS-222 (methane tricaine sulfonate, pH = 7.0) weighed, a digital image taken, and the fish snap frozen for analysis of protein and lipids. If possible, 30 fish will be sampled.

C.  Experiment 3

Objective: To determine if the morphology and composition of wild reared tautog differ from hatchery reared fish

Rationale: Wild fish will be used as an external validation of the morphological and compositional approaches. Analysis of wild fish can also serve as an exploratory exercise in finding potential differences in the allometry of growth during the early life history of these fish.

Design: Wild fish will be captured, positively identified and analyzed as described above.
 

III.  Personnel:

1. Joe Kunkel, PI.  Morphometric analysis and protein purification and analysis.
2. Joe Zydlewski, Postdoctoral Associate.  Lipid analysis and immunologic assay.
3. Ray Moniz, Undergraduate.  Lab technician projects assisting Joe Zydlewski.
4. Rahul Sharma, Undergraduate.  Lab technician projects assisting Joe Kunkel.