Cephalopods include octopus, squid, cuttlefish, and nautilus, all of which are found across the world oceans from intertidal areas to deep-sea regions (Boletzky and Villanueva, 2014). They have separate sexes and lack a true larval stage, which means that they develop directly (Boletzky and Villanueva, 2014). Most cephalopods have short life spans (1–2 years) and high rates of growth and feed conversion; all cephalopods are carnivores (Lee et al., 1994; Domingues et al., 2004; Boletzky and Villanueva, 2014; Jiang et al., 2018). The global fishery industry has been increasing its production of cephalopods in the past 20 years and has sustainably met the demand for marine products (Boletzky and Villanueva, 2014; FAO, 2017). Among the various cephalopod species, the cuttlefish is regarded as having considerable potential for aquaculture because of its high nutritional profile and high market value (Gao et al., 2014). The natural distribution of cuttlefish can be divided into two major areas, namely, the Atlantic-Mediterranean and Indo-Pacific seas (Boletzky and Villanueva, 2014). Among the various cuttlefish species, Sepia officinalis is the predominant cuttlefish reared for research in European countries, including Spain, Portugal, and France (Boletzky, 1983; Domingues et al., 2001, 2005; Baeza-Rojano et al., 2010). In East and Southeast Asia (e.g., Thailand, China, Taiwan, and Japan), Sepia pharaonis, Sepia esculenta, and Sepiella japonica are the major species that are cultured (Nabhitabhata, 1978; Hao et al., 2007; Fan et al., 2011; Zheng et al., 2014).

In the early 1960s, Choe and Ohshima (1963) first used live shrimp as feed for rearing cuttlefish and squid and successfully bred Sepioteuthis lessoniana, S. esculenta, Sepia lycidas, and S. japonica. Since then, researchers have mainly used live prey and fresh diets (e.g., shrimp and fish) to rear cuttlefish and squid (Boletzky, 1974; Pascual, 1978; Minton et al., 2001; Villanueva et al., 2017; Jiang et al., 2018). However, the training of non-domesticated animals to accept artificial feed and further development of commercial feed are key factors that influence commercial aquaculture development; thus, minimizing or changing the practice of using live prey is a major challenge in the rearing of cuttlefish (Sorgeloos et al., 2001; Zohar and Mylonas, 2001; Nguyen et al., 2011; Ouraji et al., 2011; Takeuchi and Haga, 2015). Various studies have experimented with the use of artificial feed. Lee et al. (1991) were the first researchers to successfully feed S. officinalis using an artificial diet containing non-marine sources that included hot dogs, chicken puree, and raw chicken. Koueta et al. (2002) and Perrin et al. (2004) used enriched natural and frozen diets to enhance the growth of S. officinalis. Studies have reported positive results after feeding surimi to S. officinalis and highlighted that the protein content of the mantle muscle is a valuable indicator in long-term rearing (Castro et al., 1993; Castro and Lee, 1994). Numerous studies have reported that the higher the percentage of protein (35–50%) in an artificial diet, the better the growth performance of the fed cuttlefish (Domingues et al., 2005, 2008; Ferreira et al., 2010; Gao et al., 2014; Han et al., 2017; Jiang et al., 2021). Some studies have highlighted the crucial role of fatty acids for cuttlefish, particularly eicosapentaenoic acid and docosahexaenoic acid, with cuttlefish requiring high amounts of these nutrients (Perrin et al., 2004; Ferreira et al., 2010; Gao et al., 2014; Wen et al., 2014; Jiang et al., 2018).

Pharaoh cuttlefish (S. pharaonis), which have a unique tiger stripe pattern on their skin, are distributed in the Indo-Pacific sea, the waters north of Japan, and the waters west of the Red Sea (Nabhitabhata and Nilaphat, 1999). The pharaoh cuttlefish is a tropical and large (350–420 mm in length and 4,200–5,000 g in weight) species and a crucial fishery product in Southeast Asia (Nabhitabhata, 1978). Minton et al. (2001) cultured multiple generations of pharaoh cuttlefish under laboratory conditions. Although pharaoh cuttlefish can be reared for multiple generations, its mass production remains difficult. Jiang et al. (2021) proposed an optimum weaning method for pharaoh cuttlefish in mass culture and a specific culture protocol. However, the high cost and unstable quality of the live or fresh diet required for rearing pharaoh cuttlefish limit their mass production (Han et al., 2017; Jiang et al., 2018). Before mass production can be achieved, further information on the nutritional compositions of cuttlefish and their diets are required. Peng et al. (2015) and Jiang et al. (2018) analyzed the crude composition of the muscles of juvenile pharaoh cuttlefish in culture and compared it with the diets that they were fed; their results revealed that cuttlefish muscle and shrimp shared a similar composition. Wen et al. (2014) compared the nutritional quality of wild and cultured pharaoh cuttlefish and reported that the nutritional quality of both wild and cultured cuttlefish was similar to that of the shrimp that were fed to them. Because the nutritional quality of pharaoh cuttlefish is influenced by the prey that they eat, an enriched diet may increase their nutritional quality (Han et al., 2017).

Although the implementation of artificial diets for cuttlefish has been explored, most studies have done so by mixing various fresh diets (e.g., fish and shrimp) to produce moist feed (MF; Han et al., 2017; Jiang et al., 2018). Currently, dry feed (DF) for cuttlefish is unavailable. Our cultivation experience suggests that during the mass culture of cuttlefish, most cuttlefish (wild and artificially reared stocks) only accept bait with complete fish and shrimp shapes, and artificial diets cannot be used unless training is provided. Therefore, we developed a novel process comprising (1) feed preference tests and (2) progressive training programs. Subsequently, we assessed the growth performance achieved with the use of available artificial diets.

Cephalopods have complex brains and central nervous systems that provide them with excellent learning and memorization abilities (Cole and Adamo, 2005; Correia et al., 2005; Cartron et al., 2013; Yang and Chiao, 2016; Zepeda et al., 2017; Shinzato et al., 2018; Yasumuro and Ikeda, 2018). As a member of the cephalopod family, the pharaoh cuttlefish has been proven to have number sense and excellent learning ability (Yang and Chiao, 2016; Yasumuro and Ikeda, 2018; Huang et al., 2019). Studies have verified several methods of pharaoh cuttlefish weaning, providing us with reliable message training methods that enable cuttlefish to become accustomed to eating artificial feed (Jiang et al., 2018, 2020, 2021). Our cultural experience (we weaned more than 5000 juveniles per year between 2015 and 2021) suggests that the key challenge is not the weaning process but the reduction of rearing cost and preparation of nutritional diets. Thus, training cuttlefish to eat an artificial diet is crucial to future commercial culture and enhances its efficiency.

Studies have reported that for the purpose of training cuttlefish to eat an artificial diet, their feed should be shaped in a form that is similar to the shape of their natural prey. Lee et al. (1991) used various artificial baits as feed for cultured S. officinalis and reported that artificial pellets had the longest latency-to-grab time, whereas rod-shaped artificial feed (e.g., hot dog-shaped feed) exhibited a shorter latency-to-grab time. Castro et al. (1993) and Shinzato et al. (2018) reported that cuttlefish were more attracted to shrimp-shaped bait. Thus, artificial feed should be shaped like a rod or a flat rod. In our tests, juvenile cuttlefish also tended to grab rod- and flat rod-shaped artificial feed (Figure 2). However, the feed-movement test did not reveal any significant difference; that is, pharaoh cuttlefish did not exhibit a preference in this regard (Figure 3). This result is inconsistent with that of Cole and Adamo (2005), who reported that the feed given to S. officinalis must be slowly lowered. Although the fishing lines used in our tests were not used to control the sinking speed of bait, we came to the same conclusion regarding the necessity of allowing feed to sink.

While they are hunting, cuttlefish search, detect, select, grab, handle, and kill their prey, and these activities require experience and learning (Cole and Adamo, 2005; Villanueva et al., 2017). Changes in the movement of a bait may confuse a cuttlefish or cause it to ignore the bait. Our results revealed that pharaoh cuttlefish preferred MF to DF in general, even when they had the same shape (Figure 4). This could be because the sinking speed of MF is higher than that of DF. Moreover, cuttlefish do not necessarily consume the feed that they have detected, selected, grabbed, and handled. We discovered that they may discard a piece of feed and refrain from grabbing it again. We speculate that the similarity of the texture of MF to that of real prey (i.e., soft surface) was the reason why pharaoh cuttlefish preferred MF to DF. As was highlighted in an earlier section, we believe that the texture of artificial feed influences the selection behavior of cuttlefish; this inference is also supported by Lee et al. (1991), who reported that feed texture is essential for cuttlefish weaning. Our results indicate that MF and agarose-coated feed are more suitable feed options for cuttlefish (Figures 4, 5). The latency-to-grab time was shorter for agarose-coated feed and longer for gelatin-coated feed. We believe that these two types of coated feed achieved different results because of their texture. The texture of gelatin CDF was softer than that of agarose CDF. We speculate that the gelatin coating was too “liquid-like,” such that our cultured pharaoh cuttlefish could not identify it as food and refused to eat it as a result. This study is the first to employ coated feed for cuttlefish, and the experiment revealed that coating has a considerable effect on cuttlefish. However, the suitability of various types of coating warrants further research.

The effect of feed odor on cuttlefish was difficult to evaluate. The flavor of surimi-supplemented feed was more similar to that of fresh shrimp, but the latency-to-grab times of surimi-supplemented and normal feed did not differ significantly (Figure 6). This means that cuttlefish rely more on their vision than their sense of smell to locate prey (Correia et al., 2005). However, olfactory effects still merit discussion. Guibé et al. (2010) reported that the visual prey preference of cuttlefish can be changed by odor experiences, and Lee et al. (2020) highlighted that pharaoh cuttlefish embryo can detect environmental cues and change their behavior accordingly after hatching. In our preliminary test, we added a large amount of shrimp surimi to our artificial feed, and some cuttlefish refused to eat such feed and consequently died. Therefore, we refrained from training cuttlefish to accept artificial feed directly. Instead, we changed the odor of fresh shrimp. Accordingly, the first step of the progressive training program involved the soaking of oriental shrimp into artificial feed liquid to change the odor of fresh shrimp. This enabled us to train our cuttlefish to accept artificial feed; some groups even maintained a 100% survival rate throughout the tests (Table 4). These results indicate that the role of olfactory stimulation in training should be considered, even though vision is the primary factor. The combined results of the odor test and training programs revealed that visual training yielded short-term results, whereas olfactory training can be beneficial in the long term. Olfactory training is necessary to enable cuttlefish to eat artificial feed in the long term; this position is supported by Guibé et al. (2010) and Lee et al. (2020).

The growth performance of cuttlefish that were fed artificial feed was less favorable than that of cuttlefish that were fed fresh shrimp (Table 5). Domingues et al. (2008) examined the growth performance of cuttlefish that were fed frozen baits and artificial diets. They reported that despite the addition of fresh fish and squid in the artificial diets, the feeding and growth rates were lower among the cuttlefish that were fed artificial diets than among those that were fed frozen baits. Ferreira et al. (2010) balanced the nutritional composition of frozen baits and artificial diets; however, they observed that the growth rate of cuttlefish that were fed artificial diets was still lower than that of cuttlefish that were fed frozen baits. Most studies have reported unfavorable results for artificial diets (Lee et al., 1991; Castro et al., 1993; Castro and Lee, 1994; Domingues et al., 2005, 2008; Ferreira et al., 2010; Han et al., 2017); however, Domingues et al. (2005) and Han et al. (2017) have suggested that a high crude protein content increases the weight of the cuttlefish. Although our artificial feed contained approximately 53% of crude protein, PWG, and FCR of the cuttlefish that were fed artificial feed were still less than those of the cuttlefish that were fed fresh shrimp. We discovered that the artificial feed was not always consumed completely by the cuttlefish, whereas the fresh shrimp was almost always consumed completely by the cuttlefish. The preference of cuttlefish for fresh shrimp might explain why they disliked the artificial feed and engaged in foraging behavior only when they were starving. Although our study revealed that the growth performance of cuttlefish that were fed artificial feed was less favorable than that of cuttlefish fed with fresh shrimp, no cuttlefish died during our tests. This indicates that the artificial feed is suitable for cuttlefish.

In this study, we observed that pharaoh cuttlefish preferred rod-shaped, moist artificial feed to other types of feed that were shaped differently or dry. We also discovered that the texture of feed should be similar to that of the prey of cuttlefish. Pharaoh cuttlefish can be trained to eat DF through odor stimulation, and this training process should be longer than 3 weeks (for 8-week-old juveniles). The use of compound feed for long-term cuttlefish rearing is possible, but the nutritional composition of the compound feed that is used should be studied further.

The original contributions presented in the study are included in the article/Supplementary Material, further inquiries can be directed to the corresponding author.

The animal study was reviewed and approved by Institutional Animal Care and Use Committee of National Taiwan Ocean University (NTOU).

C-HM, C-HLn, Y-CC, C–HLo, and T-HH contributed to conception and design of the study. C-HM, C-HLn, and Y-CC collected the data. C-HM, Y-CC, and H-TL performed the statistical analysis. C-HM and T-HH wrote the first draft of the manuscript. C-HLn, Y-CC, H-TL, and C-HLo wrote sections of the manuscript. All authors contributed to manuscript revision, read, and approved the submitted version.

The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.

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