Feed Fortification: Initial Efficacy Study Results

Summary

In the second half of 2024, we conducted a study to test whether a custom-formulated fish feed improves fish welfare compared to the feed commonly utilised by farmers of Indian major carp in Andhra Pradesh. This blog post summarises the key outcomes of the study. Analysis of the data revealed there is a statistical difference between the two feeds for weight/feed-related indicators, which indicates that our custom-formulated fish feed may be more nutritious.

However, there is no statistical difference between the two feeds for water quality indicators, suggesting that the custom-formulated fish feed may not indirectly improve welfare in the way we had hoped by improving the environment in which fishes live. Additionally, we couldn’t obtain the data for direct welfare improvements—physiological indicators of fish stress response—that we had hoped for.

Overall, the findings are encouraging, but we are being cautious about moving forward with a program built around an improved feed without stronger evidence. We are currently making plans for studies to provide the level of evidence we need to take a program forward or not. Of particular note, we are planning a field-based effectiveness study where the fortified feed will be tested in real-world conditions (i.e. tested in real farms by real farmers) as opposed to being tested by researchers in a test facility. This study will ultimately inform the decision about proceeding with a program.

Background

Feed quality and management are critical determinants of fish welfare. Most farmers of Indian major carp (IMC) in Andhra Pradesh use de-oiled rice bran (DORB) as the primary supplemental feed during the grow-out stage. DORB is a by-product of oil extraction from rice bran, which itself is a by-product of rice production. Unfortunately, DORB’s poor nutritional value—lacking essential proteins, amino acids, vitamins, and minerals—makes it unsuitable for fishes. Additionally, DORB contains mycotoxins and other anti-nutritional factors that reduce nutrient absorption and digestibility. FWI believes that the use of DORB likely causes suffering for the fishes and increases their susceptibility to other welfare issues, such as disease.

Apart from its poor nutritional value, feed management practices typically employed for DORB-based feeding contribute to poor water quality. Observations previously made by FWI suggest that many farmers commonly overfeed (i.e. provide more feed than is considered necessary for the fish biomass in the pond), and employ poor practices, such as dumping uneaten feed into the pond. Uneaten DORB can undergo decomposition, consuming oxygen, and can also settle at the bottom of the pond and contribute to excessive organic detritus, causing anaerobic conditions in the sediment. All together, the dynamics of dissolved oxygen (DO) levels in the pond can be greatly impacted by poor DORB-based feed management practices.

Throughout the second half of 2024 we conducted a study to compare DORB to a custom-formulated fish feed. We undertook this study as we believe that DORB’s poor nutritional value (e.g. protein, amino acid, vitamins, and mineral content that do not match the requirements of the fishes), coupled with the presence of mycotoxins and other “anti-nutritional factors” (components that reduce nutrient absorption and digestibility), likely impacts the fishes’ growth and development, increases their susceptibility to disease, and ultimately impacts their overall health and welfare. Apart from its poor nutritional value, feed management practices that farmers typically follow for DORB—which is formulated as a powder-like mash—contribute to poor water quality, compounding welfare issues for the fishes.

DORB is the feed type of choice for IMC farmers in Andhra Pradesh due to its low cost. Cost is a critical consideration for IMC farmers, as margins are so low. FWI was interested in testing whether a modified version of DORB, supplemented with a small number of nutrients and formulated as a pellet, can improve fish welfare while being a viable option for IMC grow-out farmers to utilize in place of conventional DORB. DORB was selected as the basis for our customized feed, as our ultimate aim is to present farmers with a cost-effective product that they are likely to accept; given the low-cost and widespread use of DORB, we reasoned that a fortified version of DORB—rather than formulating a whole new bespoke feed—was the best strategy to pursue. We produced this fortified feed in partnership with a local feed manufacturer.

Top: A farmworker preparing DORB to feed to the fishes. Bottom: A (separate) farmworker ties the DORB-filled bags to a rope in the pond. The bags have small holes cut in them throughout, which allows the fishes to nibble on them to access feed, though also allows the feed to leech into the water, degrading water quality.

What We Did

Our research question was as follows: Does a modified formulation of DORB—supplemented with selected nutrients and formulated as a pellet—improve the welfare of fish through improving their nutritional requirements and quality of the water in which they live, while resulting in fishes that are at least the same size as those fed on conventional DORB?

To answer our research question, we ran a study from the end of June to the middle of November 2024, during which we collected data to assess welfare (water quality indicators, as well as nutritional and physiological indicators) and the growth of fishes. Growth indicators are considered important for two reasons. Firstly, if the feed is suitably nutritious, the fishes should grow well. Secondly, a future program built around the provision of fortified feed needs to consider the economics from the perspective of the fish farmers.

Feed represents the single most expensive component for fish farming, and even though our fortified feed is designed with low cost in mind, it would still be more expensive than regular DORB. While improved welfare is FWI’s primary concern, farmers will be very mindful of operational costs. In order to encourage farmers to switch to a modified formulation, we have to show that the higher unit cost of feed is met—or exceeded—by the increased margins that farmers gain from (i) higher prices received for their fish and/or (ii) lower costs due to utilizing less feed. 

The key metrics for assessing the performance of fortified feed—and by extension, the success of the study—were:

• Water quality indicators: Does the use of fortified feed result in improved water quality of ponds compared to ponds where conventional DORB is used?

  
  

• Weight and feed-related indicators: Do fishes eat the fortified feed, and if yes, how does their weight/size gain compare to fishes fed on conventional DORB? (Note: While growth is not necessarily an indicator of welfare, if the feed is sufficiently nutritious, fishes should grow well.)

  
  

• Physiological indicators: Is the welfare of fishes, as measured by physiological indicators of the fish stress response, improved when fortified feed is used?

We conducted this study at our former test facility at Adikavi Nannaya University (AKNU) in Rajamahendravaram, Andhra Pradesh, between late June and mid-November last year. Across six test ponds, we tested three different feeds (two ponds each): (1) our fortified feed; (2) conventional DORB; and (3) a commercially-available pelleted feed (this was included as a form of "positive control" and it's considered nutritionally superior to both conventional DORB and our fortified feed, with the rationale being that our fortified feed would produce results somewhere in-between; the high cost of this feed limits its use by farmers).

Preliminary analysis of results extended into the second quarter of this year, with further time taken to explore certain components (notably, assessing physiological indicators of stress).

The six small, custom-built ponds used for this study.

Issues Encountered During Study

It’s important to note that during the study we encountered various issues at the research ponds—a lice outbreak across all ponds, a significant invasion of tilapia in one pond, and a significant overgrowth of vegetation in the pond bottom of another—which likely led to (or, at least, contributed to) the highly variable data, thereby complicating comparisons between the feeds. For example, while we successfully overcame a lice outbreak, the treatments provided to combat lice—along with the direct impact of lice on the fishes—may have impacted benefits (water quality and/or nutritional) from fortified feed.

Summary of Key Findings

Below are the key findings from each of the metrics:

Water Quality Indicators

• The overall finding was that fortified feed did not result in improved water quality. There were no significant differences between ponds where fortified feed was used compared to ponds where conventional DORB was used.

Weight and Feed-related Indicators

• Fortified feed was consumed by fishes, with no obvious signs of fishes rejecting or reacting strangely to a new feed type.

  
  

• Statistical analysis revealed that fishes fed with the fortified feed achieved higher final body weight (p<0.05) compared to fishes fed with either conventional DORB or the commercial feed control.

  
  

• Fortified feed demonstrated the best feed conversion ratio (FCR) and feed efficiency (FE) among all diets tested, meaning fishes required less feed to achieve superior growth. The Daily Growth Index (DGI) and Specific Growth Rate (SGR) were also highest for fortified DORB, indicating faster and more efficient growth relative to body size over time. These findings indicate that fortified feed may be more nutritious for IMC than conventional DORB, and is more efficiently metabolised.

• We compared the total protein in plasma samples from fishes fed the two diets to assess if the fortified feed led to higher total protein levels, a direct nutritional indicator. However, there was no difference in total protein in plasma samples. This suggests that our fortified feed did not have the intended nutritional impact that we were hoping for.

Growth performance of fishes fed the three diets over the course of four months.

Physiological Indicators

• The four selected indicators of stress response (cortisol, lactate, glutathione, and triglycerides) did not provide the information we were seeking. We contracted an external laboratory to conduct the laboratory tests for us, but the results were unusable as data from technical replicates were highly variable (we believe the lab that conducted the assays did not do a good job, either due to poor equipment, poor lab procedures, a combination of both, or some other reason). We have struggled to find a laboratory in India that can conduct the tests we require. In effect, this means that we couldn’t use most of the physiological data. 

  
  

• Analysis of triglyceride levels—a tertiary stress response indicator—suggest that conventional DORB may impose greater metabolic stress compared to fortified feed and commercial pellet feed, although these results should be interpreted cautiously given sample variability.

Another intriguing finding from our study is that, while fortified feed has a higher per-unit cost than conventional DORB, the superior feed efficiency means fishes consume less feed overall to achieve greater growth. Cost analysis suggests that, with optimal feeding ratios, farmers could save significantly on feeding costs—the most expensive aspect of IMC farming—while achieving better harvest weights. While this finding does not contribute directly to improving fish welfare, this could act as an important “pull” to convince farmers to use fortified feed if we ultimately proceed with a program.

You may view the raw data and our internal analysis here:

Raw Data

Internal Analysis

In summary, analysis of the data revealed that there is a statistical difference between fortified feed and conventional DORB for weight/feed-related indicators, but not for water quality indicators. We couldn’t obtain the data for physiological indicators of fish stress response that we had hoped for.

Overall, the findings are encouraging, particularly the superior growth performance displayed by fishes fed with fortified feed, which indicates that fortified feed may be more nutritious for the fishes. However, while differences in weight/feed-related indicators were significantly different between fortified feed and conventional DORB, it’s important to note that there was a high degree of variability in the data across the ponds/diets. This variable data is likely due to—fully or partially—the issues we encountered during the study (see above). This high degree of variability, coupled with the lack of data to indicate improved welfare—either indirect (water quality of the environment in which fishes live) or direct (physiological indicators of welfare/stress)—means that we are being cautious about moving forward without stronger evidence.

Our team at our test ponds, measuring and weighing fishes as part of the monthly data collection exercise.

Next Steps

This study was conducted to provide a proof of concept that our fortified feed is better for fishes compared to conventional DORB. The intention was to use the findings from this pond-based “efficacy study” to inform next steps. Pending positive outcomes, the plan was to proceed to a field-based “effectiveness study”, where the fortified feed would be tested in real-world conditions (i.e. tested in real farms by real farmers) as opposed to being tested by researchers in a test facility; this, in-turn, would inform decisions about proceeding with a program built around encouraging farmers to utilize the fortified feed. Unfortunately, the study at our test ponds did not provide the evidence we were hoping for to inform moving to the field study phase.

In essence, the issues we encountered, and the high variability across replicates, gives rise to a lot of “what ifs”. We want to be confident that the general concept works ahead of committing the resources to a larger study. However, terminating the project based on the inconclusive results—especially knowing that there were issues during the study that likely impacted how we can compare the different feeds—would be disappointing, considering there is a general opinion that our fortified feed is likely to be a welfare improvement given how poor DORB is.

Despite the inconclusive findings from our pond-based study, we remain excited about the potential for a program built around an improved feed. We have decided to follow three parallel tracks:

1. Tank-based study to assess the impact of fortified feed on welfare and growth

This will be a laboratory-based version of the study we conducted last year at our AKNU test facility. Instead of conducting the study in test ponds, we will utilize tanks, similar to how we conducted our DO Tolerance study. This tank-based study is more “controlled” than the previous pond-based efficacy study, so the issues that plagued us during that previous study are less likely to be an issue. This increases confidence that conclusive results—either positive or negative—will be obtained. 

2. Tank-based study to assess the impact of fortified feed on water quality

To provide a proof-of-concept that our bespoke fortified feed is better for water quality than mash DORB, we will conduct a simple tank-based experiment. This will focus on nutrient leaching from the two feeds, and how water quality is impacted (this study will not involve any fishes). Assessing any positive impact on water quality from fortified feed in the tank-based study described above to assess the impact of fortified feed on welfare and growth would be difficult—if not impossible—because we’ll be constantly conducting water exchange to prevent buildup of toxic levels of ammonia and to maintain favourable levels of DO.

3. Field-based study to compare fortified feed to DORB in a real-world setting

We plan to recruit 12 farmers, half of whom will be provided with fortified feed and the others provided with regular DORB. We will study fishes and water quality at these farms for a period of four months.

We are currently making plans for all three of these studies, and expect to initiate them in January, potentially earlier. The original intention was to only proceed to the field-based efficacy study (which are typically large and resource-intensive studies to conduct) only if we had promising findings from an efficacy study (which are typically smaller and less resource-intensive). Ultimately, conducting a field-based effectiveness study is needed to inform decisions around scaling-up a program built around fortified feed. Proceeding with a program based merely on proof-of-concept results is fraught with danger, and is not good practice. However, with our 2026 Goal in mind, we have decided to take a calculated risk, guided by promising trends from our pond-based efficacy study and proceed with the field-based study before the findings from the tank-based efficacy study are in.

Loss of Fishes During the Study 

Unfortunately, during the course of the study, we lost a number of fishes. In total, 11 fishes died. Six fishes died as a result of bird predation. This prompted us to install wires across the ponds to deter bird further attacks. Four fishes died as a result of the lice outbreak. The team identified this outbreak early and worked hard to minimise its impact. One fish died during the transportation process (i.e. transferring of fishes from the supplier to our facility). While losing fish is always upsetting, any study involving animals always comes with the risk of losing some of them. We mitigate that risk as much as we can.

Release of Study Fishes

The study we conducted at our test facility was the first time we procured fishes for a study that we have conducted. An important consideration in our decision to proceed with this study in the first place was our commitment to releasing the study fishes into a natural water body at the end of the study, as it is against FWI’s ethical values to sell fishes. Over the course of two days at the end of November 2024, we released the fishes into a river in Andhra Pradesh, with the approval of relevant authorities. Everyone involved in the study was happy to see the fishes being released into their natural environment.