Monitoring Artificial Materials and Microbes in Marine Ecosystems: Interactions and Assessment Methods

Part 1: Marine Ecosystem and Environmental Monitoring

List of Contributors

Coral Reef Ecosystems in Marine Environments

Naoko Isomura*

Department of Bioresources Engineering, National Institute of Technology (KOSEN), Okinawa College, Nago, Japan

Abstract

Japan has different climatic zones running from the south to the north of the country, which include subtropical to cool continental zones. Within these zones, 78 genera and 415 species of corals are found. Coral reefs are continuing to decline worldwide owing to large-scale bleaching events that have accompanied climate change. In Japan, the decline of reef-building corals occurred from 1998 to 2007, with many corals also dying during a large-scale bleaching event in 2016. In this chapter, I introduce our studies regarding coral reproduction, "Hybridization and speciation in Acropora and Elucidation of synchronous spawning mechanism in Acropora." The importance and contribution of coral reproduction to the restoration of coral reefs are discussed because the influence of bleaching on reproduction is not limited only to the existing coral colonies, but it might also affect next generation colonies and the maintenance of coral populations.

Keywords: Bleaching, Corals, Multi-specific synchronous spawning, Reproduction.

* Corresponding author Naoko Isomura: Department of Bioresources Engineering, National Institute of Technology (KOSEN), Okinawa College, Nago, Japan; Tel: +81-980-55-4135; Fax: +81-980-55-4012; E-mail: [email protected]

INTRODUCTION

Hermatypic corals that form coral reefs are mostly species belonging to Scleractinia, Hexacorallia, Anthozoa, and Cnidaria, although some belong to Octocorallia and Hydrozoa [1]. Many corals inhabit the shallow sea, and they show intracellular symbiosis with dinoflagellate algae.

Japan has different climatic zones running from the south to the north of the country, which include subtropical to cool continental zones. Within these zones, 78 genera and 415 species have been identified to date [1-4]. In our previous study [5], we divided the area inhabited by corals into three parts. The coral reef region (24–30° N) has high coral species diversity and includes the Yaeyama Archipelago, Okinawa Islands, Amami Archipelago, Tokara Archipelago, and

Ogasawara Islands. The non-coral reef region (30–33° N) shows moderate coral species diversity and includes Kyushu, Shikoku, and the Kii Peninsula. In this region, corals assemble without making a reef structure. The peripheral region (33–35°N) shows undeveloped coral assemblages with low coral species diversity, and includes the Izu Peninsula, Boso Peninsula, and Izu Islands (Fig. 1). The number of species decreases with increasing latitude owing to lower temperatures, with the number of species in each region estimated at 415 in the coral reef region, 200 in the non-coral reef region, and 55 in the peripheral region.

Fig. (1))

Map of Japan showing the three coral regions (coral reef region, non-coral reef region, and peripheral region). Squares surrounded by a black line with white color show the coral reef region; squares with pale gray color show the non-coral reef region and the square with dark gray color shows the peripheral region (modified from Isomura and Fukami 2018 [5]).

In recent years, coral reefs have continued to decline worldwide owing to large-scale bleaching events that have accompanied climate change or by becoming prey for the crown-of-thorns starfish (Acanthaster spp.). In Japan, the decline of reef-building corals has continued after the large-scale bleaching in 1998, and Kawagoe [6] reported that approximately 70% of corals in Sekisei Lagoon (a barrier reef lying between the coasts of Ishigaki, Taketomi, and Iriomote Island) died owing to a large-scale bleaching event in 2016. Corals that previously dominated the subtropical zone have extended their habitat northwards in recent years, and the seaweed beds have been increasingly replaced by corals in fishing grounds [7]. Meanwhile, the area that the coral has joined by moving north has become a shelter of coral species that are reduced in the temperate areas [8]. The northern part of the peripheral region is a marginal zone, which never existed previously or had an extremely small quantity of coral populations, and is now where coral communities are expected to behave differently from those in subtropical areas.

Currently, we are mainly conducting research on "Hybridization and speciation in Acropora and Elucidation of synchronous spawning mechanism in Acropora." The genus Acropora is the largest group of reef corals and consists of approximately 150 species worldwide. This group of corals consists of very important organisms that are the foundation for the construction of coral reef ecosystems. The response and/or behavior of the genus Acropora under climate change conditions is expected to affect not only corals but also the organisms that use coral reefs as their habitats as well as the environment surrounding them. In this chapter, I introduce our studies of Acropora and discuss the future of corals including Acropora.

Species Diversity of Acropora via Hybridization

Among scleractinian corals (hereinafter referred to as corals), the genus Acropora is an important component in tropical, subtropical, and temperate areas, and contains approximately 150 species. Due to its large number of species, hybrid speciation is believed to have occurred within this genus [9]. Evidence of hybrid speciation in Acropora has been shown as follows: (1) many colonies of intermediate forms have been observed and (2) many species showed inter-specific synchronized spawning, which causes multi-specfic crossing. Examples of. (1) are the three coral species in the Caribbean Sea, A. cervicornis, A. palmata, and A. prolifera, withA. prolifera shown to be a hybrid of A. cervicornis and A. palmata [10]. Although genetic information confirmed these hybrids, it was predicted that this species was a hybrid because it contained two other intermediate forms. Regarding point (2), multi-specfic synchronous spawning has been reported on the Great Barrier Reef [11] and inter-specific hybridization experiments have been conducted at various locations, which have revealed that they can cross between genetically similar species [12, 13]. Although these studies reveal the ability of crossbreeding and hybrid formation in Acropora, actual hybrid speciation has not been confirmed to date. In addition to the above conditions (1) and (2), two more conditions may be required as follows: (3) hybrids can survive at any stage of life history as well as the parent species, and (4) hybrids are fertile, which is required for the success of the next generation. Moreover, two sub conditions of point (4) exist as follows: (4a) mating between hybrids is established more frequently than crossing with other species or parent species, and (4b) under inter-specific synchronous spawning conditions, inter-specific breeding is equivalent to or more effective than intra-specific crossing.

We focused on two species, A. florida (Fig. 2A) and A. intermedia (Fig. 2B) in Akajima Island, Okinawa, to study the hybrid speciation of Acropora. These two species can hybridize artificially and are distinguished morphologically because A. florida has a bottle-brushed morphology (Fig. 2A), in which, a number of short branches protrude from the main thick branches, whereas A. intermedia has a long branching morphology (Fig. 2B). Previous studies in the Caribbean suggest that hybrids are likely to exhibit an intermediate form of the parent species; however, it is expected that hybrids can be easily discriminated, such as in these two species with distinctly different colony shapes. In fact, putative hybrids showing intermediate forms of both species were confirmed from Akajima Island (Fig. 2C). In 2007, two types of cross breeding were performed. The obtained larvae (confirmed hybrids) were cultivated for 7 years and it was confirmed for the first time globally that these confirmed hybrids spawned gametes and were fertile (Fig. 2D). Furthermore, from the results of cross breeding experiments over some years, it was possible to verify the above four conditions that are required for hybrid speciation to occur in A. florida and A. intermedia [14-16].

From 2007, the authors bred hybrids of artificially produced A. florida and A. intermedia outdoors, and measured their morphological traits in 2011 and 2014. The morphological traits of the parent species colonies were also measured during the same period, and morphological analysis using the multinomial logit model was performed. The hybrids showed an intermediate morph between the two parents and the traits were similar to the mother species [17]. In addition, the mother species was estimated from the morphology of the putative hybrids that were found in the field. Therefore, the parent species of the hybrids could be estimated more accurately together with the results from the gene analyses.

Furthermore, it was confirmed that there was an intermediate form of colonies that did not have a clear branching structure similar to A. intermedia, although it had a bottle-brushed form that is a feature of A. florida (Fig. 2E). This colony had a larger axial corallite and possessed the characteristics of A. gemmifera (Fig. 2F), which inhabited the same area as the other two species. We are now examining the ability of hybrid formation in A. gemmifera using A. florida and A. intermedia to verify how many species of Acropora can form hybrids under multi-specific synchronous spawning in Okinawa.

Fig. (2))

Photographs of the Acropora species that were used in our study. A: A. florida, B: A. intermedia, C: Putative hybrid considered to originate from A. florida and A. intermedia, D: Spawning of real hybrid by A. florida and A. intermedia, E: Putative hybrid considered to be related to A. gemmifera, F: A. gemmifera. Scale bars = 10 cm. This figure is modified from Isomura 2018 (Umiushi Tsushin, 98, 8-10, written in Japanese).

Although this is only a preliminary analysis, we examined the multiple exon regions of A. florida, A. intermedia, A. gemmifera, and putative hybrids, and found the following: (1) the haplotype was shown to be shared among A. florida, A. intermedia, A. gemmifera, and the putative hybrids, (2) another haplotype was shared between A. intermedia and the putative hybrids, and (3) another haplotype was shared only among the A. florida colonies. These results indicate that A. gemmifera may be related to hybrid production and introgression. Further detailed genetic analysis will clarify how many hybrids exist and how much introgression is occurring among these species. Moreover, hybrid speciation will be elucidated, which has been a longstanding mystery of coral research, by integrating crossing experiments, distribution surveys, and genetic analyses.

Search for Endocrine System Regulating Multi-specific Synchronous Spawning in Acropora

The genus Acropora develops their gametes throughout the year, and they spawn eggs and sperm synchronously during the summer season. Synchronization of these reproductive activities was thought to be due to changes in the external environment such as the lunar age, water temperature, and the tide. Corals receive changes in the external environment as stimuli and regulate gametogenesis and spawning by hormones and/or neurotransmitters. However, the physiological mechanism for synchronous spawning is almost unknown. Sex steroid hormones are produced from cholesterol, and sex hormones containing male and female sex hormones are formed as metabolites from cholesterol. Among these hormones, estrogen is known as the female hormone and plays an important role in reproductive activities such as the formation of ovaries and uteri. Estrogens act via the estrogen receptor (hereinafter referred to as ER). After estrogen is secreted, it enters the nucleus, binds to the ER, and regulates the transcription of reproductive genes. Invertebrates already have the same ER gene as that of humans [18] and the association between sexual maturation and the ER has been reported in oysters [19]. In scleractinian corals, estrogen has been detected in Euphyllia ancora and the estrogen level increases during the spawning months [20].

Catecholamine acts as a neurotransmitter and regulates important functions such as locomotion, biological rhythm, blood pressure, and the reproductive and endocrine systems. Noradrenaline, a type of catecholamine, functions via a noradrenaline transporter (hereinafter referred to as NAT). Even in invertebrates such as bivalves, catecholamines have been reported to be related to reproduction and spawning [21]. It has been reported that dopamine, a catecholamine, inhibited the spawning of Acropora [22]. Due to next generating sequencing, the whole genome decoding of A. digitifera was performed [23], and this genomic information can be used in studies of reef corals. We found both the ER and NAT genes using the genome database of A. digitifera.

Recently, we began to examine the expression kinetics of the ER and NAT genes in gametogenesis and spawning in A. intermedia to elucidate a part of the endocrine mechanism concerning the reproductive activity of Acropora. Although only a preliminary result, it was suggested that A. intermedia secreted much estrogen during the initial formation of the oocyte and regulated the amount of secretion during the development of the oocyte. In contrast, the expression level of the NAT gene decreased toward spawning and recovered after spawning. Noradrenaline and dopamine, a precursor of noradrenaline, are known to inhibit spawning and reproductive activity in corals and invertebrates [22, 24]. It has also been suggested that noradrenaline suppresses spawning and that a reduction of noradrenaline secretion triggered spawning in A. intermedia. In previous studies, catecholamines containing noradrenaline have been detected as substances in Acropora spp. before and after spawning [25]. Moreover, it is necessary to detect the proteins and substances related to reproduction in coral tissues, and to the clarify changes that take place before and after spawning and between seasons.

Contribution of Coral Reproduction to the Restoration of Coral Reefs

The NOAA (2015) predicted that a global bleaching event, which was the third largest in recorded history, would occur from 2015 to 2016 [26], with this predicted global bleaching event beginning at the Mariana Islands in 2014 and expanding to the South Pacific and Indian Ocean in 2015, and the northern Great Barrier Reef in 2016 [27]. Subsequent to this, a mass coral-bleaching event occurred in Ryukyu Islands, Japan, during the summer of 2016. Severe damage was observed, especially around Miyako Island and in Sekisei Lagoon [6, 28].

The amount of energy resources available to the coral hosts may decrease by the loss or reduction of zooxanthellae during bleaching. Energy resources are allocated among the processes of growth, survival, and reproduction to maximize evolutionary fitness [29], and thus the effects of bleaching on reproduction may be expected. The effects of bleaching on reproduction have been reported in Japan [30, 31] as have the effects of climate change and/or ocean acidification on reproduction [32, 33].

The influence of bleaching on reproduction is not limited to the present coral colonies; it may also affect next generation colonies and population maintenance. The recovery of Acropora depends on the density of nearby adult colonies [34]; therefore, adequate fecundity of adult colonies that are sexually mature will be important for both population fitness and recruitment to self and/or other populations. However, we have only limited information regarding coral reproduction, even for the well-studied genus Acropora, although there are more than 400 corals species around Japan. Further studies on coral ecology and reproduction are required to understand how corals are affected by ongoing climate change and how to minimize the influence of the declining coral cover in the future.

CONSENT FOR PUBLICATION

Not applicable.

CONFLICT OF INTEREST

The author confirms that this chapter contents have no conflict of interest.

ACKNOWLEDGEMENTS

Declare none.

REFFERENCES

Marine Polymers as Ecofriendly Alternatives to Petroleum-Based Plastics

Minato Wakisaka

Graduate School of Life Science and Systems Engineering, Kyushu Institute of Technology, Kitakyushu, Japan

Abstract

Nanofibers with diameters between 100 nm to 200 nm were easily prepared from various water-soluble marine polysaccharides by combining ultrasonic atomization with freeze casting. Scanning electron microscopy demonstrated considerable differences in fiber diameter and morphology between the types as well as the concentrations of polymers. Nanofibers with uniform orientation were obtained by rapid freezing. The anti-bacterial activity of chitosan nanofibers is suitable for their use as food packaging material. Biodegradable composites of chitosan nanofibers and cellulose paper could be a solution to the problem of ocean pollution from single-use petroleum-based plastics.

Keywords: Freeze casting, Nanofiber, Polysaccharides.

* Corresponding author Minato Wakisaka: Graduate School of Life Science and Systems Engineering, Kyushu Institute of Technology, Kitakyushu, Japan; Tel/Fax: +81-093-695-6066; E-mail: [email protected]

INTRODUCTION

Nanofiber production from polysaccharides has attracted a great deal of research attention because of properties of the nanofibers, including biodegradability and biocompatibility [1].

Marine polysaccharides, which are present in many marine organisms, are very important biological macromolecules. Chitin is a long-chain polymer of N-acetylglucosamine (Fig. 1A). It is one of the most abundant polysaccharides isolated from the shells of crabs and shrimps. Chitosan is a natural cationic polysaccharide (Fig. 1B) produced commercially by deacetylation of chitin. Other types of polysaccharides are rich in seaweed, such as carrageenan from red sea-weeds and alginates from brown seaweeds. Carrageenan consists of linear sulfated polysaccharides. There are three main varieties that differ based on their degree of sulfation. κ-carrageenan has one sulfate group per disaccharide (Fig. 1C). Alginate is a natural anionic polysaccharide with a linear structure (Fig. 1D) composed of β-(1–4) linked D-mannuronic acid and its epimer comprised of α-guluronic acid units. Along its polymer chain, alginate has regions rich in sequential mannuronic acid units, guluronic acid units, and regions in which both monomers are equally prevalent.

Fig. (1))

Chemical structure of marine polysaccharides.

Spinning methods, such as electrospinning [2] or wet spinning, are commonly used for nanofiber fabrication. Wet spinning requires complex processes and is expensive. In addition to the risk of electric shock, electrospinning using high-voltage electrical fields is limited by the difficulty in controlling fiber orientation and is sensitive to the conductivity of the spinning dope solution.

A novel and unique technique of fabricating nanofibers with a uniform orientation combines ultrasonic atomization and freeze casting. In this study, the applicability of this technique to various marine polysaccharides was investigated. Furthermore, biodegradable composites of chitosan nanofiber with cellulose paper were developed. The potential of these composites as food packaging material was evaluated. Such a biodegradable packaging material could provide a solution to the problem of ocean pollution with single-use petroleum-based plastics.

MATERIALS AND METHODS

Chitosan 10 powder (deacetylation degree: 80%, MW = approximately 22 kDa), sodium alginate, and κ-carrageenan were all purchased from Wako Pure Chemical Industries, Ltd. (Osaka, Japan). All the chemicals were of analytical grade, for laboratory research and investigational use only, and were used as received.