Paeniglutamicibacter terrestris sp. nov., isolated from phenanthrene-degrading consortium enriched from Antarctic soil.

A novel bacterium, designated strain ANT13_2T, was isolated from a phenanthrene-degrading consortium enriched from a soil sample collected near the Great Wall Station located in the southwestern area of King George Island, Antarctica. Following a polyphasic taxonomic study, a novel species belonging to the genus Paeniglutamicibacter was described. The strain was a Gram-stain-positive bacterium that exhibited a rod-coccus growth cycle. Strain ANT13_2T grew aerobically at an optimum temperature of 20-25 °C and at pH 7.0-8.0. Ribose, arabinose and glucose were detected as whole-cell sugars. The predominant menaquinone was MK-9. The diagnostic phospholipids were diphosphatidylglycerol, phosphatidylglycerol and an unidentified phospholipid. The predominant cellular fatty acids were anteiso-C15 : 0 (67.7 %) and anteiso-C17 : 0 (11.2 %). The DNA G+C content of the genomic DNA was 60.6 mol%. Based on 16S rRNA gene sequence analysis, strain ANT13_2T showed the highest similarities to Paeniglutamicibacter antarcticus SPC26T (98.9 %) followed by Paeniglutamicibacter gangotriensis Lz1yT (98.4 %), Paeniglutamicibacter sulfureus DSM 20167T (98.3%) and Paeniglutamicibacter kerguelensis KGN15T (97.9 %). The average nucleotide identity values between strain ANT13_2T and the type strains of P. antarcticus SPC26T and P. gangotriensis Lz1yT were 73.8 and 77.5 %, respectively, which are well below the 95-96 % species circumscription threshold. On the basis of this polyphasic taxonomic study, strain ANT13_2T is proposed to represent a novel species to be named Paeniglutamicibacter terrestris sp. nov. The type strain is ANT13_2T (=TBRC 11756T=NBRC 114615T).

Cryopreservation of sperm from the coral Acropora humilis.

Corals are sensitive to minute changes in their environments, and their continued existence is substantially threatened by the increasing number of destructive anthropogenic activities and unprecedented rates of climate change. Although cryopreservation has been successfully to preserve mammalian gametes for decades, coral cryopreservation was attempted for the first time less than 15 years ago, and freezing protocols exist for only a handful of coral species. The present study developed a cryopreservation protocol for the sperm of the common Indo-Pacific reef-builder Acropora humilis. Colonies of reefs of Sattahip Bay, Chonburi Province, Thailand were collected from 3 m depth with a mesh net during a spawning event. Immediately after collection, the sperm were isolated and subjected to a two-step freezing method featuring DMSO, polyethylene glycol, or methanol as the cryoprotectant. Viability and motility were assessed via a bioluminescence technique and a "computer-assisted semen analysis, and it was found that a 15-min equilibration with 2 M DMSO followed by cooling at 41.7 °C was the optimum cryopreservation protocol for A. humilis sperm. The post-thaw sperm achieved 45% fertilization success, and 35% of the fertilized eggs developed into blastopore larvae. The present optimized protocol can therefore facilitate the preservation of sperm for future propagation efforts of this species and provide an experimental platform for optimizing cryopreservation protocols for gametes of other scleractinian coral species.

Small microplastic ingestion by the calanoid Centropages furcatus in the Gulf of Thailand.

Microplastics could be ingested by many organisms, including zooplankton, involving bioaccumulation and biomagnification mechanisms a cross food webs. The information about microplastic ingestion by zooplankton keeps increasing worldwide. However, it is still limited for particle sizes under 300 µm (small microplastics, SMPs) and in areas such as Southeast Asia, which is considered one of the hotspots for plastic debris. This study aimed to characterize the size, shape, and polymer types of the SMPs ingested by the copepod Centropages furcatus in Si Chang Island (upper Gulf of Thailand). The study spans offshore and coastal waters, with data collected across wet, intermediate, and dry seasons. Using a semi-automated technique for micro-FTIR (Fourier-transform infrared) scanning spectroscopy for particle analysis, we found ingested SMPs in all samples. A total of 750 individuals of the calanoid Centropages furcatus were analyzed, finding 309 plastic particles and an average ingestion value of 0.41 ± 0.13 particles ind-1, one of the highest recorded values. All the particles were fragments, with a predominant size under 50 µm, and polymer types as Polypropylene (PP, 71 %), followed by Ethylene-Propylene-Diene-Monomer (EPDM, 16 %) and Polyethylene (PE, 7 %). Up to 470.2 particles m-3 were estimated to be retained by this calanoid species and potentially available for trophic transfer. The effect of rainfall on SMPs ingestion was inconclusive, with a non-significant observed tendency to higher ingestion values near the coastal area than offshore area, suggesting a decrease in particle exposure due to the runoff effect. Nevertheless, future studies should increase the frequency of surveys to arrive at better conclusions.

16S rRNA amplicon sequencing of seawater microbiota from Rayong Province, Thailand, affected by oil spills.

We reported on microbial communities isolated from 18 seawater samples affected by oil spills in Rayong province, Thailand, using the V3-V4 region of 16S rRNA gene sequencing.

Influence of monsoon seasonality and tidal cycle on microplastics presence and distribution in the Upper Gulf of Thailand.

Southeast Asian countries are recognized as significant contributors to the discharge of abundant plastic waste into the ocean. In this study, we conducted neuston net surveys on Si Chang Island of the Gulf of Thailand, a coral reef conservation area, to determine the presence of microplastic (MP) pollution. The survey, conducted during the wet (southwesterly monsoon), transition, and dry seasons (northeasterly monsoon), revealed that the MP abundance was in the range of 0.02-42.46 particles m-3. The precipitation, wind, and current direction induced by monsoons influenced the abundance and distribution of MP, presenting a significant seasonality. The cluster analysis for colors and polymer types of MPs suggested that the origin of plastic particles is diverse. Based on our results, a proposal for the generation, sources, and pathways for MPs in the Gulf of Thailand is presented: 1) plastic wastes exposed to strong UV light during the dry season get fragmented around the river, and 2) heavy rains wash away the particles during the wet season. This proposal is applicable to tropical regions, including the Gulf of Thailand. Therefore, this paper concluded that ocean currents induced by monsoons and the unique climate, resulting in the generation of MPs on land, increase MP presence and distribution in the ocean surrounding Southeast Asia countries. Furthermore, coral reef ecosystems can be particularly threatened by MPs in these areas. So, an increase in MP monitoring on coral ecosystems from Thailand and the world is highly recommended.

Pseudomonas and Pseudarthrobacter are the key players in synergistic phenanthrene biodegradation at low temperatures.

Hydrocarbon contamination, including contamination with polycyclic aromatic hydrocarbons (PAHs), is a major concern in Antarctica due to the toxicity, recalcitrance and persistence of these compounds. Under the Antarctic Treaty, nonindigenous species are not permitted for use in bioremediation at polluted sites in the Antarctic region. In this study, three bacterial consortia (C13, C15, and C23) were isolated from Antarctic soils for phenanthrene degradation. All isolated bacterial consortia demonstrated phenanthrene degradation percentages ranging from 45 to 85% for 50 mg/L phenanthrene at 15 â„ƒ within 5 days. Furthermore, consortium C13 exhibited efficient phenanthrene degradation potential across a wide range of environmental conditions, including different temperature (4-30 â„ƒ) and water availability (without polyethylene glycol (PEG) 6000 or 30% PEG 6000 (w/v)) conditions. Sequencing analysis of 16S rRNA genes revealed that Pseudomonas and Pseudarthrobacter were the dominant genera in the phenanthrene-degrading consortia. Moreover, six cultivable strains were isolated from these consortia, comprising four strains of Pseudomonas, one strain of Pseudarthrobacter, and one strain of Paeniglutamicibacter. These isolated strains exhibited the ability to degrade 50 mg/L phenanthrene, with degradation percentages ranging from 4 to 22% at 15 â„ƒ within 15 days. Additionally, the constructed consortia containing Pseudomonas spp. and Pseudarthrobacter sp. exhibited more effective phenanthrene degradation (43-52%) than did the individual strains. These results provide evidence that Pseudomonas and Pseudarthrobacter can be potential candidates for synergistic phenanthrene degradation at low temperatures. Overall, our study offers valuable information for the bioremediation of PAH contamination in Antarctic environments.

Polymer types and additive concentrations in single-use plastic products collected from Indonesia, Japan, Myanmar, and Thailand.

Marine debris comprising single-use plastic products (SUPs) is ubiquitous in Asian coastal waters, but there is little information on the types of polymers and the concentrations of plastic additives such waste products contain. In this study, 413 SUPs randomly collected from 4 Asian countries between 2020 and 2021 were analyzed to obtain specific polymer and organic additive profiles. Polyethylene (PE), coupled with external polymers, was prominent in the inside of the SUPs, whereas polypropylene (PP) and polyethylene terephthalate (PET) were prevalent in both the insides and outsides of the SUPs. The use of different polymers in the insides and outsides of PE SUPs implies specific and complicated recycling systems are required to maintain the purity of the products. Phthalate plasticizers including dimethyl phthalate (DMP), diethyl phthalate (DEP), diisobutyl phthalate (DiBP), dibutyl phthalate (DBP), and di(2-ethylhexyl) phthalate (DEHP), and the antioxidant butylated hydroxytoluene (BHT) were prevalent in the SUPs (n = 68). High concentrations of DEHP were detected in PE bags from Myanmar (820,000 ng/g) and Indonesia (420,000 ng/g), which were an order of magnitude greater than the concentrations in PE bags collected in Japan. SUPs containing high concentrations of organic additives may be the primary source of harmful chemicals in the environment, and should be responsible for their ubiquitous distribution in ecosystems.

The seasonal investigation of Symbiodiniaceae in broadcast spawning, Acropora humilis and brooding, Pocillopora cf. damicornis corals.

The density and diversity of Symbiodiniaceae associated with corals can be influenced by seasonal changes . This study provided the first annual investigation of Symbiodiniaceae density and diversity associated with Acropora humilis and Pocillopora cf. damicornis corals in the Gulf of Thailand using both zooxanthellae cell count and next-generation sequencing (ITS-1, ITS-2 regions) techniques, respectively. The results from this study indicated that zooxanthellae cell densities in both coral species differ significantly. The number of zooxanthellae was negatively correlated with the physical environment variable (light intensity). The diversity within A. humilis consisted of two genera, Cladocopium (Cspc_C3: 56.39%, C3w: 33.62%, C93type1: 4.42% and Cspf: 3.59%) and a small amount of Durusdinium (D1: 1.03%) whereas P. cf. damicornis was found to be 100% associated with Durusdinium (D1: 95.58%, D6: 1.01% and D10: 2.7%) suggesting that each coral species may select their appropriate genus/species of Symbiodiniaceae in response to local environmental stressors. The results of this study provided some information on the coral-Symbiodiniaceae relationship between seasons, which may be applied to predict the potential adaptation of corals in localized reef environments.

First dynamics of bacterial community during development of Acropora humilis larvae in aquaculture.

A symbiosis of bacterial community (sometimes called microbiota) play essential roles in developmental life cycle and health of coral, starting since a larva. For examples, coral bacterial holobionts function nitrogen fixation, carbon supply, sulfur cycling and antibiotic production. Yet, a study of the dynamic of bacteria associated coral larvae development is complicated owning to a vast diversity and culturable difficulty of bacteria; hence this type of study remains unexplored for Acropora humilis larvae in Thai sea. This study represented the first to utilize 16S rRNA gene sequencing to describe the timely bacterial compositions during successfully cultured and reared A. humilis larval transformation in aquaculture (gametes were collected from Sattahip Bay, Chonburi province, Thailand), from gamete spawning (0 h) and fertilization stage (1 h), to embryonic cleavage (8 h), round cell development (28, 39 and 41 h), and planula formation (48 h). The sequencing results as estimated by Good's coverage at genus level covered 99.65 ± 0.24% of total bacteria. While core phyla of bacteria were observed (Proteobacteria, Actinobacteria, Firmicutes and Bacteroidetes), changes in bacterial population structures and differential predominant core bacterial orders were denoted for each larval developmental stage, from fertilization to embryonic cleavage and subsequently from the embryonic cleavage to round cell development (P = 0.007). For instances, Pseudoalteromonas and Oceanospirillales were found prevalent at 8 h, and Rhizobiales were at 48 h. The bacterial population structures from the round cell stage, particularly at 41 h, showed gradual drift towards those of the planula formation stage, suggesting microbial selection. Overall, this study provides preliminary insights into the dynamics of bacterial community and their potentially functional association (estimated from the bacterial compositions) during the developmental embryonic A. humilis in a cultivation system in Southeast Asia region.

An updated inventory of sea slugs from Koh Tao, Thailand, with notes on their ecology and a dramatic biodiversity increase for Thai waters.

Improved access to field survey infrastructure throughout South-East Asia has allowed for a greater intensity of biodiversity surveys than ever before. The rocky bottoms and coral reef habitats across the region have been shown to support some of the highest sea slug biodiversity on the planet, with ever increasing records. During the past ten years, intensive SCUBA surveys have been carried out at Koh Tao, in the Gulf of Thailand, which have yielded remarkable findings in sea slug biology and ecology. In this work a brief history of sea slug biodiversity research from Thailand is covered and a complete inventory of sea slugs from Koh Tao, Gulf of Thailand is provided. This inventory is based on surveys from 2012 to 2020, with previously unreported findings since 2016. Habitat specificity and species-specific ecology are reported where available with a focused comparison of coral reef habitats and deeper soft-sediment habitats. The findings contribute 90 new species records for Thai waters (92 for the Gulf of Thailand) and report a remarkable consistency in the proportional diversity found to be exclusive to one habitat type or another. Additionally, taxonomic remarks are provided for species documented from Koh Tao that have not been discussed in past literature from Thailand, and a summary of previous records in the Indo-West Pacific is given.

Two new species of the genus Chironephthya (Octocorallia, Alcyonacea, Nidaliidae, Siphonogorgiinae) from the Gulf of Thailand.

Two new species of the genus Chironephthya, C. sirindhornae sp. nov. and C. cornigera sp. nov., are described based on three specimens collected from the Gulf of Thailand. Both species are well distinguished from the previously described species of the genus Chironephthya by their colonies consisting of multiple stems that stand upright from a common base, and by a significantly thinner canal wall without large spindles. As a result of phylogenetic analyses using COI, mtMutS, and 28S rDNA sequences of these two species, these three specimens constituted an independent small clade within a large mixed clade of Siphonogorgia and Chironephthya, with the two species slightly different from each other. The discrepancy in the morphology suggested the erection of a new genus to accommodate these species, however, as the subclade was included in a large mixed clade of Siphonogorgia and Chironephthya, we place these species within genus Chironephthya. Our results further highlight the continuing confusion between Siphonogorgia and Chironephthya, and demonstrate the need for taxonomic revision of these genera.

An interlaboratory comparison exercise for the determination of microplastics in standard sample bottles.

An interlaboratory comparison exercise was conducted to assess the consistency of microplastic quantification across several laboratories. The test samples were prepared by mixing one liter seawater free of plastics, microplastics made from polypropylene, high- and low-density polyethylene, and artificial particles in two plastic bottles, and analyzed concurrently in 12 experienced laboratories around the world. The minimum requirements to quantify microplastics were examined by comparing actual numbers of microplastics in these sample bottles with numbers measured in each laboratory. The uncertainty was due to pervasive errors derived from inaccuracies in measuring sizes and/or misidentification of microplastics, including both false recognition and overlooking. The size distribution of microplastics should be smoothed using a running mean with a length of >0.5 mm to reduce uncertainty to less than ±20%. The number of microplastics <1 mm was underestimated by 20% even when using the best practice for measuring microplastics in laboratories.

Ultrastructural observation of oocytes in six types of stony corals.

In this study, the ultrastructure of the oocytes of 6 types of scleractinian corals was observed by transmission electron microscopy (TEM). Moreover, histological and ultrastructural analyses were performed to improve our understanding of the organelles involved in coral oocyte formation. In all 6 stony coral species, the microvilli were tubular and directly grew from the surface of the oocyte membrane; yolk bodies, lipid granules, and cortical alveoli accounted for most of the volume inside the oocytes, suggesting that they are associated with energy storage and buoyancy. Clear differences were observed in the size of yolk bodies and lipid granules in the oocytes of the 6 stony coral species, which occupied approximately 55%-80% of the inner space of the oocytes. Galaxea fascicularis exhibited the largest lipid granule volume, but the oocytes contained only an average number of 12.45 lipid granules per unit area. Only Montipora incrassata oocytes contained symbiotic algae. The smallest size and proportion of lipid granules in M. incrassata oocytes may be attributed to the presence of symbiotic algae and large yolk bodies, which may help oocytes produce energy and function as a nutritional source. This study is crucial for improving the understanding of the basic biology of coral reproduction, and the ensuing datasets is critical for conservation-oriented studies seeking to cryopreserve corals during these times of dramatic global climate change.

Development of Cryopreservation Techniques for Gorgonian (Junceella juncea) Oocytes through Vitrification.

Gorgonian corals are slowly declining due to human interaction and environmental impacts. Cryopreservation of gorgonian corals is an ex-situ method of conservation, ensuring future reproduction. The present study assessed the vitrification properties of cryoprotectant (CPT) mixtures using the cryotop, cryoloop and open pulled straw (OPS) cryopereservation methods prior to experimentation on gorgonian (Junceella juncea) oocytes. Investigations of the equilibration and vitrification solutions' (ES and VS) effect on oocytes throughout different incubation periods were conducted. The cryotop method was found to be the most successful in ensuring vitrification. The most favourable VS was composed of propylene glycol (PG), ethylene glycol (EG) and methanol with concentrations of 3.5 M, 1.5 M and 2 M respectively. Experiments were performed using the cryotop method to cryopreserve Junceella juncea oocytes using VS2, the solution had the least impact on oocytes at 5°C rather than at 26°C. The success of the vitrification procedures was determined by adenosine triphosphate (ATP) levels in cooled-thaw oocytes and the highest viability obtained from the present study was 76.6 ± 6.2%. This study provides information regarding gorgonian corals' tolerance and viability throughout vitrification to further advance the vitrification protocol on whip corals.

Membrane lipid phase transition behavior of oocytes from three gorgonian corals in relation to chilling injury.

The lipid phase transition (LPT) from the fluid liquid crystalline phase to the more rigid gel structure phase that occurs upon exposure to low temperatures can affect physical structure and function of cellular membranes. This study set out to investigate the membrane phase behavior of oocytes of three gorgonian corals; Junceela fragilis, J. juncea and Ellisella robusta,at different developmental stages after exposure to reduced temperatures. Oocytes were chilled to 5°C for 48, 96 or 144 h, and the LPT temperature (LPTT) was determined with Fourier Transform Infrared (FTIR) spectroscopy. The J. fragilis oocytes had a higher LPTT (∼23.0-23.7°C) than those of J. juncea and E. robusta oocytes (approximately 18.3-20.3°C). Upon chilling for 96 h at 5°C, the LPTTs of J. juncea and E. robusta oocytes in the early (18.0±1.0 and 18.3±0.6°C, respectively) and late (17.3±0.6 and 17.7±1.2°C, respectively) stages were significantly lower than those of J. fragilis oocytes (20.3±2.1 and 19.3±1.5°C for the early and late stages, respectively). The LPTTs of early stage gorgonian oocytes was significantly lower than those of late stage oocytes. These results suggest that the LPT of three gorgonian oocytes at different developmental stages may have been influenced by the phospholipid composition of their plasma membranes, which could have implications for their low temperature resistance.