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.

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.

An effective and simple technique for seagrass bed restoration is required since the area of seagrass bed, one of the most productive marine ecosystems in Japan, is decreasing. The physiology of the plant Zostera marina, in relation to seed germination, was investigated in detail. Optimal seed storage condition and pretreatment for breaking seed dormancy increased the seed germination rate. Planting of biodegradable gel-coated seeds for Zostera marina seagrass bed restoration was proposed.

Although the status of microalgae as a primary producer in the aquatic environment is well-known, some algae are also an important biomass supporting our varied fishery resources as prey in fisheries industries. Algal blooms occur due to environmental changes in factors, such as temperature, salinity, nutrient concentrations, and light availability. The blooms occasionally result in serious damage to human activities including fisheries. Starting with information regarding several algal toxins from harmful algae, this chapter discusses the common characteristics of an algal bloom and several case studies of algal blooms causing fish-kill in coastal waters and aquaculture farms in Japan. In addition to low dissolved oxygen concentration induced by excessive proliferation of algae, some harmful algae produce distinctive toxins which contaminate marine foods like shellfish. Therefore, this chapter also outlines general features of algal toxins. The properties of bloom-forming algae described in this chapter require the fisheries industry and their related sectors to constantly monitor harmful algal blooms and verify whether the shellfish accumulates harmful algae.

Many studies have reported the changes in environmental conditions by using various water analysis (and environmental analysis) techniques. In these reports, “the necessity for water analysis,” “the basic knowledge for water analysis,” “the requirements, conditions, and methods for water analysis,” a “plan for water analysis,” and whether the entities to be measured satisfy the “Environmental Quality Standards” and the “Effluent Standards” are summarized.

Marine environmental assessments are needed to understand and conserve the marine ecosystem. In particular, the primary production of phytoplankton, which forms the basis of ecosystem functioning, is influenced by chemical species in seawater, including nitrogen (N), phosphorus (P), and iron (Fe). Thus, previous research by the author has focused on material dynamics in rivers and coastal areas to develop marine environmental technology. The investigations in the Chikugo River, Kesennuma Bay, and Kamaishi Bay are introduced in this chapter. Fe dynamics in the Chikugo River was investigated from July 2011 to May 2012. The trend in Fe concentration was different from that of N and P. Fe distribution in the river was specific and correlated to turbidity in the estuary. Thus, Fe dynamics might have contributed to the formation of the ecosystem in the Chikugo River and the Ariake Sea. In Kesennuma Bay, Fe, N, and P concentrations were monitored to gain insights on Fe dynamics after the 2011 tsunami disaster. The dynamics of Fe concentrations was found to be different before and after one year of the disaster (March 2012). Freshwater from terrestrial areas primarily influenced Fe concentrations close to the river mouths after one year of the disaster. In Kamaishi Bay, the concentrations of nutrients (N, P, and Si) and heavy metals (Cd, Pb, Cr, Zn, As, and Se) in seawater, as well as the concentrations of radioactive materials (134Cs and 137Cs) in sediments, were monitored from March 2012 to November 2013. Nutrient concentrations varied differently with seasons before and after the tsunami disaster. Heavy metal concentrations were below or equal to the accepted environmental standards in Japan after the disaster. Although radioactive materials produced by the Fukushima nuclear accident were detected in the sediments, they were only found in low concentrations. These results show that the coastal environment in Kamaishi Bay has recovered from the disaster.

Biofouling is described in this chapter. Biofouling is the overall phenomenon and processes related to the attachment of organisms to surfaces. It is classified into two main categories: microfouling and macrofouling. Microfouling is the phenomenon induced by bacterial activity. Bacteria, microalgae, etc. attach to materials’ surfaces, This induces the phenomenon. On the other hand, macrofouling occurs by the attachment of larger organisms such as barnacles, oysters, etc. Both types have a close relationship to each other. Actually, it occurs in a series. In this chapter, we provide an overview of each type and discuss how to effectively control them.

Microbes, including bacteria and plankton, play important roles in aquatic ecosystems, particularly marine ecosystems. Techniques to detect and evaluate marine microbes are necessary for the understanding and conservation of the ecosystems, and to better understand microbial corrosion of artificial structures built in the marine environment. Fundamental techniques to detect and evaluate microbes have been developed from laboratory experiments using cultured model microorganisms, such as Escherichia coli. Such laboratory-based techniques are often insufficient in the marine environment because of interference from organisms other than target microbes of each research, and from organic and inorganic compounds in the environment. Precise evaluation of microbial communities under such circumstances demands techniques with greater sensitivity and specificity than those of the laboratory-based approaches. This chapter considers more precise and sensitive techniques than the conventional techniques for the collection, detection, and evaluation of microbes. These techniques can also help experts and professional readers achieve practical evaluation and control of microbes in complex conditions such as marine environments.

The collection and isolation of microorganisms from seawater in aquaculture systems, aquariums, fishing port facilities, and recreation areas is the most fundamental means of studying microbiological, physiological, and pathogenic properties of water to improve public health and the environment. However, there are few methods for concentration of living microorganisms. Thus, it is necessary to develop a technology for collection and concentration of microorganisms from seawater. In this study, we examined the removal and concentration of the bacterium Vibrio and fungus Fusarium from seawater by foam separation using dispersed bubbles and surfactants. After batch processing with only 1 mg/L milk casein added as a surfactant and after injection of bubbles, Vibrio and Fusarium were isolated at removal efficiency rates of more than 80% and 99.9%, respectively, and most of the microbial cells were concentrated alive in the foam water within 5 min. When the continuous foam separation unit was installed at the actual site of a fishery harbor, though the removal efficiency for viable bacteria was 49.2%, the bacteria were isolated at a huge concentration in the foam water, and the concentration factor was 18.5. Foam separation is a feasible convenient technology for not only seawater purification but also membrane-free concentration of live microorganisms.

We developed an electrical retrieval method for living environmental microorganisms using a weak electrical potential applied to an optically transparent electrode. Living microorganisms resuspended in non-nutritive solutions such as calcium- and magnesium-free phosphate-buffered saline [PBS(-)] and artificial seawater were attracted by and selectively attached to an indium tin oxide (ITO) electrode surface to which a negative potential between −0.2 and −0.4 V vs. Ag/AgCl was applied. The electrically attached microorganisms could be cultured on the agar medium after detachment from the ITO electrode by application of a ±20 mV vs. Ag/AgCl, 9 MHz sine wave potential. When a ±0.2 V vs. Ag/AgCl, 12 MHz sine wave potential was applied to the attached microorganisms on the electrode with an applied negative potential, actinomycetes were selectively cultured and were expanded 60- to 4000-fold in terms of colony-forming units (CFUs) as compared with application of a ±20 mV vs. Ag/AgCl, 9 MHz sine wave potential. Using the electrical retrieval method, we identified and isolated Nocardiopsis sp., Dietzia sp., Pseudonocardia sp., Brachybacterium sp., Nesterenkonia sp., Microcella sp., Microbacterium sp., and Streptomyces sp. from deep-sea sediment core samples in Suruga Bay, Japan. Phylogenetic analyses based on 16S ribosomal RNA gene sequences indicated that 59% of the obtained strains are novel species or are highly likely to be novel species. The electrical retrieval method for living microorganisms holds promise as a novel screening strategy for hardly cultivable microorganisms.

Ascidians are known to accumulate extremely high levels of vanadium in their blood cells. The concentration of vanadium in seawater is 35 nM, while the concentration of vanadium reaches 350 mM in blood cells, which corresponds to 107 times that of seawater. Ascidians can be regarded as a natural ecosystem that harbors vanadium-related symbiotic bacteria and serves as a useful bacterial resource. Since the 1990s, vanadium-accumulating or -reducing bacteria have been isolated from vanadium-rich ascidians. Recent functional screening and comprehensive molecular studies also identified symbiotic bacteria in the branchial sac and the intestine. These bacteria could contribute to the high accumulation of vanadium by ascidians. In this chapter, the authors overview the vanadium accumulation and reduction in ascidians, review the studies on the isolation and functional analyses of vanadium-accumulating or -reducing bacteria, and provide perspectives on utilization of these bacteria for bioremediation of heavy metals.

We attempted evaluation of biofilm formation by the electrochemical quartz crystal microbalance (EQCM) method. The EQCM method is applicable to in situ settings and may be able to determine corrosion and a chemical reaction on materials under the influence of biofilm formation. We used Au having high chemical stability to measure frequency and changes of the potential with biofilm formation alone. Additionally, sample surfaces were examined by a conventional biofilm formation assay such as fluorescent X-ray analysis and Raman spectroscopy. We conducted three types of experiments, 1st: rest-potential measurement, 2nd: spontaneous-potential measurement, and 3rd: constant-potential measurement. From the results of these measurements, it was concluded that frequency changes depend on changes in the potential. We investigated biofilm formation by conventional methods of evaluation and found that the above phenomenon was caused by concentration of ions such as Ca2+ and Cl− from tap water in biofilm. In the positive voltage range, the magnitude of biofilm formation was inhibited by a halogen ion such as Cl−, whereas in the negative range, cations such as Ca2+ were concentrated at an accelerated rate. Therefore, electrochemical evaluation of biofilm formation was carried out successfully by means of Au, with high chemical stability. In conclusion, the EQCM method appears to be able to measure biofilm formation.

Biofilms formed on glass plates or polystyrene petri dishes were investigated in-situ in physiological salt water by scanning ion conductance microscopy (SICM). Environmental biofilms formed in a laboratory biofilm reactor (LBR) as well as biofilms formed by a single bacterium, Aliivibrio fischeri, could be observed by SICM. Maintaining consistency of salt concentration in the water used for SICM observation and that used in the LBR and the addition of glutaraldehyde to water before observation might be effective for clear and stable observation of biofilms by SICM.

Fluorescence-activated cell sorting (FACS) that is often interchangeably termed as flow cytometry allows for rapid multiparametric analysis of the physical characteristics of individual particles or cells as they flow in a fluid stream one by one through a fixed laser beam. FACS also implies physical sorting of particles with selected optical properties out of the flow stream and collecting them for subsequent studies. Unicellular microalgae of freshwater and seawater habitats represent a very convenient object for FACS analysis. The current chapter summarizes major achievements of FACS in the study of morpho-functional peculiarities and taxonomic affiliation of microalgae representing eukaryotic organisms. Life cycle analysis of green microalgae using a combination of key life cycle parameters such as cell size/volume, DNA and chlorophyll content distributions is also considered here

Fluorescence in situ hybridization (FISH) has become a standard technique in the visual detection and phylogenetic identification of environmental microorganisms in marine microbiology. Thirty years have passed since the appearance of FISH, and now FISH can detect not only ribosomal RNA but also mRNA and functional genes with high sensitivity. This chapter describes the principles, drawbacks, and applications of FISH and highly sensitive FISH. In particular, we introduce two high-sensitivity FISH methods developed by the authors. The first is in situ DNA hybridization chain reaction (HCR), a highly sensitive and highly penetrating detection technique for ribosomal-RNA–targeting FISH, and the second is two-pass tyramide signal amplification (TSA)-FISH that can detect single-copy genes. Finally, we discuss how the FISH technique has contributed to the field of marine microbiology.

Biofilm is inhomogeneous, thin film-like matter formed on materials’ surfaces. It is actually a hydrogel and composed of water (about 80% depending on the formation stages), external polysaccharides (EPS) and bacteria themselves. When materials are immersed into marine environments, biofilms form by bacterial activity. However, the process contains many steps. The formation and growth capability of biofilms are determined by the combination of environmental factors (bacteria, biota, temperatures, etc.) and materials. In this chapter, the authors describe the general biofilm formation process and discuss how various materials affect biofilm formation and growth.

Regulating biofouling is important for the maintenance of marine vessels, bridges, and other constructions. Marine organisms must also be protected. Here we propose a silane-based polymer coating that has features of both polysiloxanes and silicates: nontoxicity, weatherability, and a self-cleaning effect as an anti-biofouling method. We focus on the antibacterial activity of some metals and metallic nanoparticles applied to the silane-based coating that can remove biofouling at the late stage of biofilm formation and may be effective at the early stage. Silver, nickel, or copper nanoparticles dispersed to a silane-based polymer coating exhibit antibiofouling behavior, whereas tungsten or molybdenum nanoparticles enhance biofouling.

The oxidizing slag discarded from the oxidation process in an electric arc furnace was analyzed using the tank leaching test based on the Japanese industrial standards. The dissolved concentrations of environmentally regulated substances from the oxidizing slag were lower than those stipulated in the environmental quality standards. Therefore, this slag could be used as soil. Ca, K, P, Mg, S, Fe, Mn, Zn, Cu, B and Mo, which are essential elements for plants, were dissolved in the slag. Si was also dissolved in the slag. The slag-containing aqueous solution exhibited buffering action, and its pH was approximately 8.5. This value was smaller than those of mortar and limestone solutions. The slag-free eluate also exhibited buffering action. The Fe(II) ion was dissolved in the slag, and therefore, it could reduce the Cr(VI) ion.

In recent years, many studies have been conducted on the use of steelmaking slag to improve marine environments. By burying steelmaking slag alone or in a mixture of steelmaking slag and soil (or sand) containing organic materials in marine environments, the growth of marine plants can be promoted and rocky-shore denudation can be repaired. Furthermore, it is known that the occurrence of blue tide can be prevented. These effects are understood to be the result of a steady supply of iron, which is a major component of steelmaking slag. This chapter outlines the generation and conventional applications of ironmaking and steelmaking slags, which are by-products of the steel industry. Author then highlight previous studies on the environmental protection of sea areas using steelmaking slag and discuss novel applications of steelmaking slag.

Sea desertification has become a serious problem in sea coasts worldwide. Owing to Japan’s long sea coast, fishery is one of its key industries; thus, sea desertification is a crucial problem for the Japanese society. The positive effect of steelmaking slag containing humic substances or soil in sea coast on the rehabilitation of damaged coastal environments has been phenomenologically confirmed. This study has focused on elucidating this phenomenon, and it has been clarified that the coexistence of organic substances and steelmaking slag supported the stable dissolution of nutrients from steelmaking slag by chelating the inorganic ions.

This chapter describes effects of an eluate from steelmaking slag on phytoplankton, particularly Chlorella species as model phytoplankton. Eluates used to assess toxicity in this study were derived from slag using a leaching test condition based on JIS K 0058-1. Electric arc furnace slag in particular was used as a slag. In this study, we also developed methods to estimate not only the number of algal cells by hemocytometry but also their physiological activities via flow cytometry. After a leaching test, the concentrations of eluates from slag were almost lower than those of environmental quality standards. In addition to an elemental analysis, a Chlorellabased bioassay was performed on the eluates. After the treatment of algae with eluates, algae were analyzed by microscopy and flow cytometry. As a result, treatment with eluates induced neither death nor growth inhibition. This treatment was not extremely toxic but rather induced algal growth and a vigorous physiological state.

Reduction of seaweed beds in coastal areas, called isoyake in Japanese, is a serious problem in Japan, which results in the disappearance of habitats for many fishes and the decline of coastal fisheries. Many factors may contribute to the reduction of seaweed beds, including seawater temperature rising, grazing by herbivores, deficiencies of nutrients (nitrogen, phosphorus) and iron, among others. We examined fertilization by selected some nutrients and minerals, with a particular focus on iron, and developed a fertilizer, VivaryTM Unit. VivaryTM Unit is a 1:1 (v:v) mixture of the steelmaking slag and humus soil. Steelmaking slag, which is a by-product of the steelmaking process, is a source of iron, and artificial humus soil contains chelators such as humic acid. To establish a regeneration technology for seaweed beds, we verified the role of iron in seaweed growth and the bioavailability of VivaryTM Unit with several experiments in various scales. In vitro culture was used to demonstrate the synthesis of chlorophyll a in Pyropia yezoensis thalli and the requirement of iron in gametophyte maturation in Saccharina japonica var. religiosa. The effects of VivaryTM Unit were demonstrated through two experiments. (1) a cultivation test of P. yezoensis in a mesocosm facility, (2) a field test in Hokkaido, Japan. In the cultivation of P. yezoensis the elution of nitrogen, phosphorus, silica and iron from VivaryTM Unit were revealed, and P. yezoensis grew only in the mesocosm with VivaryTM Unit. In the field test in Hokkaido, seaweed beds, especially S. japonica var. religiosa, were restored following fertilization via a buried VivaryTM Unit. We indicated the effect of dissolved iron supplied by the buried VivaryTM Unit by examining the correlation between EC (electrical conductivity) and dissolved iron compared with the correlation between EC and dissolved silica.