Abnormal Salmonid Otolith Mineralogy and its Implication for Hatchery Fish Health

Introduction

Over 100 million hatchery fish, in the US alone, are used to compensate for dwindling wild salmon populations [1]. Despite the reliance on hatchery fish to maintain worldwide fish stocks, hatchery-rearing techniques result in physical abnormalities that impact salmon health and survivability [2-5]. One such deformity is the abnormal mineralogy of otoliths, also known as ear stones. Otoliths form from layered deposits of calcium carbonate (CaCO3) in the inner ears of teleost fish [6]. These bony structures aid in fish balance, orientation and hearing, and as such are crucial structures for salmon survival [3,4]. Along with their importance to the fish, otoliths also provide critical life history data that informs salmon conservation and management. Recently, an endemic salmonid otolith mineralogy abnormality has been impairing salmonid survival rates as well as salmonid research efforts [4,5,7-9]. This abnormality is increasingly observed in hatchery salmonid otoliths, indicating a problem with current hatchery management techniques [3,5]. This review will assess the proposed causes of the aforementioned otolith abnormality. It will also discuss the impact of this abnormality on hatchery salmonid health and on current salmonid research. 

Background

Though otoliths exist in over 23,000 fish species, this paper will focus on salmonids since they are commonly reared in hatcheries [10]. According to the process governed by the U.S. Fish and Wildlife Service (FWS), hatcheries breed and raise salmonids while they are in the egg and fry phases of their life cycle. Just before reaching their smolt phase, they are released from the hatchery to spend 1-2 years in freshwater streams to prepare for their oceanic migration. The salmonids will spend most of their life at sea until they are ready to breed, at which point they will return to their natal streams [1]. Researchers can learn about the various aquatic conditions a salmon encounters throughout their migratory life cycle by analyzing the trace elements captured in each layer of the otolith [9,11].

Despite the presence of other otoliths in the fish’s inner ear, current research focuses on the sagittal otolith as it is the largest in size and therefore most easily observed. The sagittal otolith typically consists of layers of aragonite, a CaCO3 crystal formation [8]. Aragonite layers are deposited as the fish grows, accumulating material from the fish’s environment in each layer–making the otolith a useful indicator of fish age and environmental conditions [6]. Vaterite, another polymorph of CaCO3, is also known to form on the otolith. Since vaterite is a less dense polymorph than aragonite, vateritic otoliths impair fish sensory functions [3]. The less dense otolith cannot stimulate the proper nerve endings in the fish’s inner ear. Once vaterite forms, aragonite will no longer be deposited, affecting the severity of the damage to fish sensory functions [12]. While in the wild this abnormal form appears sporadically, vaterite is 10 times more likely to occur in hatchery-reared salmonids [3]. The hatchery related cause of this otolith deformity in conjunction with the salmonid fitness are current points of interest in aquaculture research.

Cause of Vaterite Formation 

Researchers know that vaterite occurs more frequently in hatchery salmonids than wild populations, but the cause of vaterite formation is still unknown [3,13]. Vaterite deposits are possibly linked to thermal conditions, increased growth rates, or genetics [5,13,14]. Researchers have tested the hypothesis of the effect varying thermal conditions has on vaterite formation. They tested their hypothesis by raising hatchery cohorts in tanks at a constant temperature of 11.5℃, as opposed to the 2 - 20℃ temperature fluctuations experienced in streams. Researchers then measured the resulting percentage of each otolith composed of vaterite and compared them to vaterite percentages for fish who experienced temperature fluctuations [5,13]. Ultimately, they found no correlation between temperature and vaterite occurrence because vateritic frequencies were inconsistent though thermal conditions were constant [5,13]. Their findings suggest another factor of hatchery cultivation results in the observed vaterite frequencies. 

Since hatchery-raised fish tend to display faster growth rates than wild fish, it has been suggested that increased growth rate leads to vaterite formation [3]. Though studies have been done to test if faster growing fish showed increased vaterite occurrence, inconsistent vaterite frequencies between hatchery fish of the same size suggest there is no correlation between growth rate and vaterite formation [5,14]. However, Chalan et al. found decreased vateritic frequency in coho salmon treated with the growth hormone transgene OnMTGH1. The decrease in vaterite deposits indicates a growth related biological mechanism may be contributing to salmon otolith abnormality. Current research supports this hypothesis, as varying hatchery methods of increasing salmonid growth rates results in different amounts of vaterite occurrence [5,14]. Chalan et al. proposes the protein matrices, that result from the growth hormone used in their experiment, prevent vaterite formation despite the increased growth rate. Considering that protein matrices are a result of genetic expression, the genetic makeup of hatchery populations may be a key factor in vaterite occurrence. The protein expression hypothesis suggested by Chalan et al., aligns with the findings of Austad et al. where vateritic formation appeared to be most reliant on the specific hatchery of origin. Since hatchery fish tend to become more inbred with each subsequent population, there may be a genetic rationale for the lack of protein matrices and therefore vaterite formation [2]. Further research should focus on the effect of hatchery population genetics on protein expression and therefore vaterite formation. 

Vaterite Prevention Treatments 

Despite not knowing the cause of otolith polymorphisms, multiple hormones and protein matrices have been studied for their effect on otolith formation or mineralization. Treating hatchery salmon with proteins such as OnMTGH1, has been proposed as a potential method of preventing vaterite formation [14]. The presence of other proteins like OMM-64 have been suggested as potential causes for increased vaterite deposits [15]. While attempting to determine if increased growth rate causes vaterite occurrence, Chalan et al. discovered that fish treated with the OnMTGH1 growth hormone produced less vateritic otoliths. Though the mechanisms behind this hormone’s ability to prevent vaterite is still unknown, researchers propose it be used as a vaterite preventing agent in hatchery fish. The effects of other macromolecules on CaCO3 polymorphs have also been examined for their use in vaterite prevention. Organic matrix macromolecule-64 (OMM-64) was found to inhibit aragonite mineralization on otoliths, meaning other polymorphic crystal forms, like vaterite, are then more likely to form [15]. Since aragonite crystals were seldom found in the otoliths of fish treated with OMM-64, it is suspected that the OMM-64 deficiency leads to vateritic mineralization of the otoliths [15]. If the presence of this protein matrix inhibits aragonitic formation, then a treatment that suppresses OMM-64 expression could be effective at preventing abnormal otolith polymorphisms. Since the presence or deficiency of specific protein matrices results in the prevention of vateritic deposits, more research should be done into the protein products that are, or are not, being created in hatchery fish genomes.  

Implications on Fish Welfare 

Since otoliths are pivotal sensory structures, affectation of otoliths is speculated to reduce salmonid survival rates by increasing their predation risk [4]. To test the effect of vateritic otoliths on salmonid survival rates, Austad et al. determined a baseline frequency of vateritic occurrence in Atlantic salmon hatchery smolt cohorts. This baseline was then compared to the vateritic frequency present in the salmon groups who returned from their ocean migration. Due to the observed lower frequency of vaterite in the returning group, researchers concluded there is a correlation between vaterite occurrence and marine survival rates [5]. Delaval et al., in contrast, found more vateritic fish after marine migration, when conducting a similar experiment assessing vaterite frequency. Since Delaval et al. observed larger hatchery fish were more likely to have vateritic otoliths, they propose the increased body size counteracts the predation risk caused by vaterite otolith hearing impairments. In the study done by Delaval et al., abnormal otoliths were negatively correlated with salmonid survival rates, while Austad et al. found a positive correlation. This conflict of research results implies a more complex relationship between abnormal otoliths and salmonid survival rates during oceanic migration. The most significant difference between the study done by Delaval et al. and the one by Austad et al. is the hatchery facility where the experiments were conducted. Different hatcheries will have various rearing conditions resulting in salmon with diverse physiological features. While all hatcheries report high occurrence of vaterite, not all hatchery populations will produce the same physiological traits to counteract the negative consequences of vaterite. 

Effects on Trace Element Readings 

Aside from their sensory functions, otoliths are also useful resources for assessing salmon ambient water conditions through trace element analysis. Although the presence of vaterite often obscures the clarity of trace element data [8,9]. Typically, aragonite deposits form  distinct bands of material that can be counted like tree rings. The material precipitated in each band of otolith is then analyzed for trace elements such as strontium (Sr), carbon isotopes or nitrogen isotopes [11]. In otoliths, the amounts and ratios of trace elements inform researchers about the fish’s environmental conditions, migration patterns, and physiological activity [7]. The presence of vaterite means different ratios of trace elements would be found, skewing data and resulting in incorrect ecological conclusions [9]. Since different polymorphs have varying affinities for trace elements, an otolith’s mineralogy can influence trace element concentrations [9]. Without proper identification of the polymorphic content, trace element data can be incorrectly attributed to external factors when it is actually due to an otolith’s mineralogy [9].

Various methods have been proposed to deal with this experimental flaw when studying otoliths. Wood et al. determined visual identification is not nearly accurate enough for polymorphic detection. They highly recommend using Wide-Angle Neutron Diffraction (WAND), but this technique can be expensive and inaccessible. The least time consuming and costly method of polymorphic identification that Wood et al. identified was the use of polarized light spectroscopy, which is used in most current studies. Regardless of the chosen method of identification, a comprehensive assessment of polymorphic content is essential for ensuring accurate conclusions are made from trace element readings [9] Once vaterite presence is confirmed, otoliths can still be used for trace element data. In order to use vateritic otoliths in trace element studies, new baselines for trace element concentrations must be established for that polymorph [8]. This method requires excess otolith samples to establish a baseline for the element of interest. Using the new vaterite specific threshold, environmental conditions for the fish’s lifespan can still be determined [8]. Vaterite poses new challenges for studying otolith mineralogy and microchemistry, though with proper identification and polymorph specific scales, vateritic otoliths can still be useful sources of fish age related data. 

Conclusion 

This review aimed to evaluate current research on the causes and consequences of abnormal otoliths on fish welfare, as well as determine a direction for future otolith research. While current research methods have found a way to account for vateritic content, vateritic otoliths still pose potential threats to salmon fitness. Researchers have reached a consensus that vaterite occurs much more frequently in hatchery reared fish. Considering hatchery fish make up a significant portion of current worldwide salmonid populations, it is crucial we determine solutions for preventing this otolith deformity from occurring. Currently, research into the cause of otolith polymorphic abnormalities are all conflicting and variable. However, the most promising factor for predicting vaterite occurrence is determining the hatchery of origin for a population. Since protein matrices are proving to be effective measures at treating hatchery fish for vaterite, hatchery specific population genetics should be the focus of future salmonid aquaculture research.

About the Author: Adele Ferrer

Adele Ferrer is a fourth year Biological Sciences major at UC Davis. She currently interns at the UC Davis Center for Watershed Sciences in the Johnson and Jeffres lab. It is here that she learned all about salmon and their otoliths. While preparing otoliths for spectroscopy, she became curious about the different crystal structures she was seeing in each otolith. When tasked with choosing a research topic for her UWP 102B class, she decided this was the perfect opportunity to delve into our current understanding of this abnormal otolith crystal formation. She hopes readers will gain a new appreciation for otoliths and their utility for aquaculture research, or at the very least learn what otoliths are.

Author’s Note

I wrote this literature review for my UWP102B class with Professor Amy Goodman-Bide. When it came time to choose a topic, I had just started working with otoliths in the lab I’m a part of at the Center for Watershed Sciences. As I was being trained on how to prep the otoliths for spectroscopy, I was taught about this abnormal otolith crystallization. When I asked my supervisor why this abnormal form occurs, I was told that we still aren’t sure of the cause. This piqued my interest, so I used this literature review assignment to attempt to figure out why we know so little about how this abnormality occurs. My search generated some conflicting results, and left me with even more questions about otolith mineralogy. I wrote this literature review with those unanswered questions in mind. I hope my literature review points towards those gaps in our current knowledge of the subject, potentially leading to new otolith research.  

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