Species composition and distribution of common Crassostrea and Saccostrea oysters along the coast of Hainan Island

Oyster cultivation is a pivotal economic industry in Hainan Island,South China,where a high oyster species diversity is shown.However,the specific distribution and biodiversity of oyster resources in the island have remained unclear.To elucidate the diversity of oyster species and their distribution there,307 oyster samples were collected from 29 sites in the intertidal zone around the island,and were identified using both morphological and molecular approaches.A minimum of 12 oyster species were identified in taxonomy,including Crassostrea species(C.gigas angulata,C.sikamea,C.iredalei,C.dianbaiensis,C.talonata,C.ariakensis,and C.hongkongensis),and Saccostrea species(S.malabonensis,S.mordax,S.echinata,S.circumsuta,and S.mordoides).The results revealed a remarkable diversity of oyster species along the coast of the island.Particularly noteworthy are that S.malabonensis and S.mordax constituted 36%and 22%of the collected specimens,respectively.This study provided a comprehensive overview on current state of oyster biodiversity in Hainan,serving as a valuable reference for conservation and research on species distribution and resource dynamics in oyster populations.

Characterisation of the Bacteria and Archaea Community Associated with Wild Oysters, At Three Possible Restoration Sites in the North Sea

With 85% of the global oyster reefs destroyed, there is an urgent need for large scale restoration to benefit from the ecosystem services provided by biogenic oyster reefs and their associated biodiversity, including microorganisms that drive marine biogeochemical cycles. This experiment established a baseline for the monitoring of the bacterial and archaeal community associated with wild oysters, using samples from their immediate environment of the Voordelta, with cohabiting Crassostrea gigas and Ostrea edulis, Duikplaats with only C. gigas attached to rocks, and the Dansk Skaldyrcentre, with no onsite oysters. The microbial profiling was carried out through DNA analysis of samples collected from the surfaces of oyster shells and their substrate, the sediment and seawater. Following 16S rRNA amplicon sequencing and bioinformatics, alpha indices implied high species abundance and diversity in sediment but low abundance in seawater. As expected, Proteobacteria, Bacteroidetes, Firmicutes and Thaumarchaeota dominated the top 20 OTUs. In the Voordelta, OTUs related to Colwellia, Shewanella and Psychrobium differentiated the oysters collected from a reef with those attached to rocks. Duikplaats were distinct for sulfur-oxidizers Sulfurimonas and sulfate-reducers from the Sva 0081 sediment group. Archaea were found mainly in sediments and the oyster associated microbiome, with greater abundance at the reef site, consisting mostly of Thaumarchaeota from the family Nitrosopumilaceae. The oyster free site displayed archaea in sediments only, and algal bloom indicator microorganisms from the Rhodobacteraceae, Flavobacteriaceae family and genus [Polaribacter] huanghezhanensis, in addition to the ascidian symbiotic partner, Synechococcus. This study suggests site specific microbiome shifts, influenced by the presence of oysters and the type of substrate.

Detection and quantification of Pb and Cr in oysters using laser-induced breakdown spectroscopy

The quantitative determination of heavy metals in aquatic products is of great importance for food security issues.Laser-induced breakdown spectroscopy(LIBS)has been used in a variety of foodstuff analysis,but is still limited by its low sensitivity when targeting trace heavy metals.In this work,we compare three sample enrichment methods,namely drying,carbonization,and ashing,for increasing detection sensitivity by LIBS analysis for Pb and Cr in oyster samples.The results demonstrate that carbonization can remove a significant amount of the contributions of organic elements C,H,N and O;meanwhile,the signals of the metallic elements such as Cu,Pb,Sr,Ca,Cr and Mg are enhanced by3–6 times after carbonization,and further enhanced by 5–9 times after ashing.Such enhancement is not only due to the more concentrated metallic elements in the sample compared to the dried ones,but also the unifying of the matter in carbonized and ashed samples from which higher plasma temperature and electron density are observed.This condition favors the detection of trace elements.According to the calibration curves with univariate and multivariate analysis,the ashing method is considered to be the best choice.The limits of detection of the ashing method are 0.52 mg kg-1 for Pb and0.08 mg kg-1 for Cr,which can detect the presence of heavy metals in the oysters exceeding the maximum limits of Pb and Cr required by the Chinese national standard.This method provides a promising application for the heavy metal contamination monitoring in the aquatic product industry.

Application of ICP-AES with Microwave Digestion to Detect Trace Elements in Oysters from Jiaozhou Bay,China

Using ICP-AES with microwave digestion, we determined the concentrations of 16 trace elements in oysters from six sampling points in Jiaozhou Bay.The distributions of the wholesome elements Zn, Fe and Mn, and heavy metals such as As,Cd,Hg and Pb were studied.The oysters collected are all rich in the wholesome trace elements,and the oysters from Licun River have the highest concentrations of the three wholesome trace elements.The concentrations of heavy metals in oysters from Licun River are the highest and those from Hongdao are the lowest. Compared with the domestic and foreign sea-areas,the heavy metal contents in the oysters from Jiaozhou Bay are less in amount than those from some developed countries, and more than those in Southeast China.This implies that the Jiaozhou Bay's oysters have been polluted by the environment to some extent.

论Oysters

林奈于1758年建立了Ostrea (s.l.)属。此后的两百多年中经拉马克(Lamark)、杜威尔(Douville)、乌雅洛夫(Vyalov)、斯坦兹尔(Stenzel)等许多学者的深入研究,不断修正、补充与完善,原来的Oatrea (s.l.)属现已扩展为牡蛎超科(SuperfamilyOstreacea),人们通常称之为Oysters(牡蛎类)。目前对Oysters的特征、分类、生态、起源、演化等方面的认识正不断深化,但仍存在不少分歧。

Study on Elemental Fingerprint of Traditional Marine Chinese Medicine Oysters from Jiaozhou Bay,China

In order to investigate the relationship between the trace elements and the characteristics of the oysters, we analyzed the trace elements present in the germplasm of oysters from different producing areas in the Jiaozhou Bay. The element fingerprints were established to reflect the elemental characteristics of the oysters. Concentration patterns of the elements were deciphered by principle component analysis (PCA) and hierarchical cluster analysis (HCA). The six regions were discriminated with accuracy using HCA and PCA based on the concentration of 16 trace elements. The elements were viewed as characteristic elements of the oysters and the fingerprints of these elements could be used to distinguish the quality of the oysters.

Sequence Analysis and Quantitative Detection of Norwalk-like Viruses in Cultured Oysters of China

We isolated 4 Norwalk-like viruses （NLVs） contaminated oysters from 33 Chinese oysters collected from local commercial sources of Shandong Province. After amplification of the RNA-dependent RNA polymerase （RdRp） region of NLVs genomes with RT-PCR, the open reading frame 1 （ORF 1） of the RdRp was sequenced and subjected to multiple-sequence alignment. The results showed that NLVs in the four isolates belong to genogroup Ⅱ. The sequence comparison showed that the similarity between four Chinese oyster isolates were higher than 99.0%, which indicated that NLVs prevalent in close areas have high homogeneity in genome sequences. In addition, the most conserved sequences between diverse NLVs were used to design primers and TaqMan probes, then the real-time quantitative PCR assay was performed. According to the standard curve of GII NLVs, the original amounts （copies） of NLVs in positive patient＇s fecal isolate, positive Japanese oyster isolate, and the Chinese oyster isolate were 8.9× 10^8, 1.25× 10^8 and 4.7× 10^1 respectively. The detecting limit of NLVs was 1 × 10^1 copies. This study will be helpful for routine diagnosis of NLVs pathogens in foods and thus for avoiding food poisoning in the future.

Heavy metals in oysters, mussels and clams collected from coastal sites along the Pearl River Delta,South China

Concentrations of 8 heavy metals: cadmium (Cd), copper (Cu), zinc (Zn), lead (Pb), nickel (Ni), chromium (Cr), antimony (Sb) and tin (Sn) were examined in 3 species of bivalves (Perna viridis, Crassostrea rivularis and Ruditapes philippinarum) collected from 25 sites along the Pearl River Delta coastal waters in the South China Sea from July to August 1996. In general, Cd, Cu, Zn and Sn concentrations in the three bivalve species collected from the Estuarine Zone were significantly higher than those collected from the Western and Eastern Zones of the Pearl River Delta, which are related to the existence of various anthropogenic activities in the catchment of the Pearl River Delta. The Western Estuarine Zone is mainly impacted by Cr, Ni and Cu contamination. In Victoria Harbor, heavy metal contamination is mainly due to Cu and Pb. Cd, Cu and Zn concentrations in oysters were significantly higher than those in mussels and clams. This could be explained by the fact that oysters live mainly in the Estuarine Zone of the Pearl River Delta which receives most of the polluting discharges from the catchment of the Delta. During turbid condition, heavy metals(soluble or adsorbed on suspended particulates) discharged from the Delta are filtered from the water column and subsequently accumulated into the soft body tissues of oysters. Heavy metal concentrations in the three bivalve species were compared with the maximum permissible levels of heavy metals in seafood regulated by the Public Health and Municipal Services Ordinance, Laws of Hong Kong, and it was revealed that Cd and Cr concentrations in the three bivalve species exceeded the upper limits. At certain hotspots in the Delta, the maximum acceptable daily load for Cd was also exceeded.

A new identification method for five species of oysters in genus Crassostrea from China based on high-resolution melting analysis

The high phenotypic plasticity in the shell of oysters presents a challenge during taxonomic and phylogenetie studies of these economically important bivalves. However, because DNA can exhibit marked differences among morphologically similar species, DNA barcoding offers a potential means for oyster identification. We analyzed the complete sequences of the cytochrome oxidase subunit I （COI） of five common Crassostrea species in China （including Hong Kong oyster C. hongkongensis, Jinjiang oyster C. ariakensis, Portuguese oyster C. angulata, Kumamoto oyster C. sikamea, and Pacific oyster C. gigas） and screened for distinct fragments. Using these distinct fragments on a high-resolution melting analysis platform, we developed an identification method that does not rely on species-specific PCR or fragment length polymorphism and is efficient, reliable, and easy to visualize. Using a single pair of primers （Oyster- COI-1）, we were able to successfully distinguish among the five oyster species. This new method provides a simple and powerful tool for the identification of oyster species.

High Pressure Treatment Changes Spoilage Characteristics and Shelf Life of Pacific Oysters(Crassostrea gigas)During Refrigerated Storage

The effects of high pressure(HP) treatment on spoilage characteristic and shelf life extension of Pacific oysters(Crassostrea gigas) during refrigerated storage were studied.Results showed that HP treatment of 275 MPa for 3 min or 300 MPa for 2 min could achieve 100% full release of oyster adductor muscle,pressures higher than 350 MPa caused excessive release as the shells of oysters were broken,thus use of higher pressures should be cautious in oyster processing industry because of its adverse impact on the appearance of shells.HP treatment(300 MPa,2 min) was proper for the shucking of Pacific oyster(Crassostrea gigas) in China.This treatment caused no organoleptic disadvantage.Moreover,HP treatment resulted in obvious differences in biochemical spoilage indicators(p H,TVB-N and TBARS) changes and volatile compounds profile determined by electronic nose during storage.HP treatment(300 MPa,2 min) also led to a reduction of aerobic bacterial count(APC) by 1.27 log cycles.Furthermore,the APC values of oysters treated by HP were always lower than those of the control samples during storage.Based on the organoleptic,biochemical and microbiological indicators,shelf life of 6-8 d for control and 12 d for HP-treated oysters could be expected.HP treatment showed great potential in oyster processing and preservation.

An Efficient Method of Noroviruses Recovery from Oysters and Clams

Noroviruses (NoVs) are widespread causes of nonbacterial gastroenteritis. Outbreaks of NoVs caused diseases are commonly ascribed to the consumption of contaminated shellfish. The concentration and RNA extraction of NoVs are crucial steps of detecting NoVs in shellfish. This study aimed to select a simple, rapid and highly efficient recovery method of NoVs detection with real-time RT-PCR. Four methods of recovering GI.3 and GII.4 NoVs from spiked digestive tissues of oysters and clams, respectively, were compared, of them, the method involving proteinase K and PEG 8000 was found the most efficient. With this method, 9.3% and 13.1% of GI.3 and GII.4 NoVs were recovered from oysters and 9.6% and 12.3% of GI.3 and GII.4 NoVs were recovered from clams, respectively. This method was further used to detect NoVs in 84 oysters (Crassostrea gigas) and 86 clams (Ruditapes philippinarum) collected from 10 coastal cities in China from Jan. 2011 to Feb. 2012. The NoVs isolation rates were 10.47% of clams (9/86) and 7.14% of oysters (6/84). All the detected NoVs belonged to genotype GII. The NoVs recovery method selected is efficient for NoVs detection in oysters and clams.

Occurrence of Toxigenic Clostridium difficile in Louisiana Oysters (<i>Crassostrea virginica</i>) and Environmental Waters

Clostridium difficile infection (CDI) was generally considered as a hospital-associated disease;however, recent community-acquired CDI has raised the concerns regarding the transmission of the pathogen through environmental sources. Limited data are available regarding the presence of C. difficile in food and water. In this study, oysters (Crassostrea virginica) and the harvest water collected from the commercial harvesting areas along the Louisiana Gulf Coast as well as the influent and effluent of a municipal treatment plant in New Orleans, LA were analyzed for toxigenic C. difficile. The bacterium was isolated from 47.37% (9/19) of oysters and 37.5% (3/8) of harvest water samples. Toxigenic C. difficile were also detected in all the wastewater influent and effluent samples. All the isolates harbored the gene tcdB encoding the virulence factor toxin B. Further PCR-ribotyping showed that the C. difficile isolated from the oysters and harvest water differed from the wastewater isolates. However, similar ribotypes were found in oysters and the surrounding harvest water. We found that oysters growing in contaminated water could bioaccumulate toxigenic C. difficile and pose a health risks by serving as a vehicle for the transmission of the pathogen to humans.

Selection of Reference Genes for Expression Analysis of Kumamoto and Portuguese Oysters and Their Hybrid

Quantitative real-time polymerase chain reaction(q RT-PCR) is a rapid and reliable technique which has been widely used to quantifying gene transcripts(expression analysis). It is also employed for studying heterosis, hybridization breeding and hybrid tolerability of oysters, an ecologically and economically important taxonomic group. For these studies, selection of a suitable set of housekeeping genes as references is crucial for correct interpretation of q RT-PCR data. To identify suitable reference genes for oysters during low temperature and low salinity stresses, we analyzed twelve genes from the gill tissue of Crassostrea sikamea(SS), Crassostrea angulata(AA) and their hybrid(SA), which included three ribosomal genes, 28 S ribosomal protein S5(RPS5), ribosomal protein L35(RPL35), and 60 S ribosomal protein L29(RPL29); three structural genes, tubulin gamma(TUBγ), annexin A6 and A7(AA6 and AA7); three metabolic pathway genes, ornithine decarboxylase(OD), glyceraldehyde-3-phosphate dehydrogenase(GAPDH) and glutathione S-transferase P1(GSP); two transcription factors, elongation factor 1 alpha and beta(EF1α and EF1β); and one protein synthesis gene(ubiquitin(UBQ). Primers specific for these genes were successfully developed for the three groups of oysters. Three different algorithms, ge Norm, Norm Finder and Best Keeper, were used to evaluate the expression stability of these candidate genes. Best Keeper program was found to be the most reliable. Based on our analysis, we found that the expression of RPL35 and EF1α was stable under low salinity stress, and the expression of OD, GAPDH and EF1α was stable under low temperature stress in hybrid(SA) oyster; the expression of RPS5 and GAPDH was stable under low salinity stress, and the expression of RPS5, UBQ, GAPDH was stable under low temperature stress in SS oyster; the expression of RPS5, GAPDH, EF1β and AA7 was stable under low salinity stress, and the expression of RPL35, EF1α, GAPDH and EF1β was stable under low temperature stress in AA oyster. Furthermore, to evaluate their suitability, the reference genes were used to quantify six target genes. In conclusion, we have successfully developed primers appropriate for the expression analysis in SS, SA and AA.

Foodborne Pathogens in Retail Oysters in South China

Objective To investigate the occurrence of important foodbome pathogens in shellstock Pacific oysters in the food markets in South China. Methods From July 2007 to June 2008, retail oysters were collected in different seasons from South China and analyzed for the prevalence and levels of Listeria monocytogenes, Vibrio vulnificus and Vibrio parahaemolyticus. Results None of L. monocytogenes could be detected in any of the 202 oyster samples tested, while E vulnificus and E parahaemolyticus could be detected in 67 （54.9%） and 109 （89.3%） of the 122 oyster samples analyzed, respectively, with an MPN （most probable number） value greater than or equal to 3. V. wdnificus and Eparahaemolyticus with a more than 102 MPN/g were found in 36 （29.5%） and 59 （48.4%） of the 122 oyster samples, respectively. The tdh and trh genes were detected in 4 （0.3%） and 8 （0.6%） of the 1 349 V. parahaemolyticus isolates, respectively. Of the 122 samples, 4 （3.3%） was positive for either tdh or trh. The levels of E vulnificus and total V. parahaemolyticus in oysters in South China varied in different seasons. Conclusion V. vulnificus and pathogenic V. parahaemolyticus are frequently found in oysters in south China, which may pose a potential threat to public health. Data presented here will be useful for the microbiological risk assessment in oysters in China.

Acid Sulfate Soil Induced Acidification of Estuarine Areas Used for the Production of Sydney Rock Oysters, <i>Saccostrea glomerata</i>

This study investigated estuarine acidification, associated with drainage and excavation of acid sulfate soils, in areas used for commercial cultivation of Sydneyrock oysters (Saccostrea glomerata). Regular measurements of pH and electrical conductivity were collected in oyster cultivation areas and acidified reaches of the Hastings River estuary and Port Stephens estuary located on the mid north coast of New South Wales, Australia. Water quality information from acidified floodplain drains was also collected in theHastingsRiverfollowing heavy rainfall. Both estuaries experienced acidification of tributaries following periods of heavy rainfall. Drain outflow waters were acidic

Factors Affecting Dermo Disease(Perkinsus marinus)in Eastern Oysters(Crassostrea virginica)in Galveston Bay,Texas

The oyster disease Dermo(Perkinsus marinus)affects the viability of oyster reefs of Galveston Bay,Texas.Documenting the relationships between distribution and prevalence of Dermo disease in the eastern oyster(Crassostrea virginica)and environmental conditions is beneficial to management of the eastern oyster in Galveston Bay.We sampled four sites located in Galveston Bay every other month from November 2014 through September 2015.The focus of the study was the relationship of water quality parameters(i.e.,fresh-water flow,salinity,water temperature,and water turbidity)to prevalence and parasite concentration of Dermo disease in oysters.Dermo was present in oysters at all reefs sampled,and Dermo prevalence was greatest at April Fool and Confederate reefs,but declined after heavy rainfall.Linear regression analysis indicated water variables such as temperature,salinity,turbidity,and fresh water inflow explained different amounts of the variability in the Mackin Dermo Intensity Scale among sampled reefs.We found combinations of low fresh-water inflow,high salinity,and high temperatures accounted for majority of the variance of Dermo in oysters located in Galveston Bay.

Annual Component Analysis of Pacific Oysters （Crassostrea gigas） and Mussels （Mytilus gallopro vincialis）

To study the change in components of Pacific oysters （Crassostrea gigas） and mussels （Mytilus galloprovincialis） over a year, their annual obesity ratio, umami components （glutamic acid and glycogen）, functional components （Zn, Mn, Cu, and vitamin B12）, and Cd and noroviruses （NoVs） contents were quantified. In oysters, the obesity ratio and chemical components showed seasonal changes depending on the spawning season. Glutamic acid content increased from November to March. In contrast, glycogen content peaked in May and November. Mineral content markedly increased during spawning. From the comparison of NoVs GI and GII strains, oyster predominately infected by the GI. A remarkable difference in NoVs contents was observed depending on the oyster farms. In mussels, glutamic acid, Zn and Cu contents were stable throughout the year. Mussels＇ Cd content was about one-fifths times lower than that found in oysters. Although the NoVs strain was affected by the collection area, mussels from both areas showed decreased NoVs contents in summer, corresponding with the consumption season of mussels in Japan. Our comparative study on the seasonal variability in the chemical components and NoVs contents in pacific oysters （C. gigas） and mussels （M. galloprovincialis） clearly showed an association with physiological and environmental factors.

Three novel umami peptides derived from the alcohol extract of the Pacific oyster(Crassostrea gigas):identification,characterizations and interactions with T1R1/T1R3 taste receptors

Oyster(Crassostrea gigas),the main ingredient of oyster sauce,has a strong umami taste.In this study,three potential umami peptides,FLNQDEEAR(FR-9),FNKEE(FE-5),and EEFLK(EK-5),were identified and screened from the alcoholic extracts of the oyster using nano-HPLC-MS/MS analysis,i Umami-Scoring Card Method(i Umami-SCM)database and molecular docking(MD).Sensory evaluation and electronic tongue analysis were further used to confirm their tastes.The threshold of the three peptides ranged from 0.38 to 0.55 mg/m L.MD with umami receptors T1R1/T1R3 indicated that the electrostatic interaction and hydrogen bond interaction were the main forces involved.Besides,the Phe592 and Gln853 of T1R3 were the primary docking site for MD and played an important role in umami intensity.Peptides with two Glu residues at the terminus had stronger umami,especially at the C-terminus.These results contribute to the understanding of umami peptides in oysters and the interaction mechanism between umami peptides and umami receptors.

Use of natural trophic resources by Eastern oysters and Pacific oysters of different ploidy

Ploidy manipulation,such as triploidy,in farmed oysters has been used as a tool to enhance oyster quality throughout all seasons because triploid oysters allocate less energy to gametogenesis,and therefore are deemed better performers than fertile diploids.Nevertheless,scientific reports describing no differences between ploidies and,in certain conditions,disadvantages of triploids are not uncommon.As the use of triploid oyster seed increases culture cost for growers,consistency in performance of triploids is considered to be an important goal.Thus,research to assess how ploidy affects physiological processes underlying oyster performance is fundamental for the aquaculture industry.This work was undertaken to assess if ploidy-based differences in performance in the two most commonly cultivated commercial species of oyster in the United States,Crassostrea virginica and Crassostrea gigas,are associated with filtration,feeding,and metabolism.To test this hypothesis,biodeposition measurements were made with oysters exposed to ambient water conditions at locations and seasons providing a variety of environmental conditions.Oysters did not show differences in filtration and feeding associated with ploidy,but physiological feeding variables fluctuated with environmental characteristics associated with spatial and seasonal differences.A preliminarily test of the hypothesis that differences in energy metabolism may account for differences in performance among ploidy levels indicates that basal metabolic rates of diploid,triploid,and tetraploid Eastern oysters are not different.

Spatial compartmentalization and temporal stability of associated microbiota in Pacific oyster Crassostrea gigas

The Pacific oyster Crassostrea gigas,one of the most exploited molluscs in the world,has suffered from massive mortality in recent decades,and the occurrence mechanisms have not been well characterized.In this study,to reveal the relationship of associated microbiota to the fitness of oysters,temporal dynamics of microbiota in the gill,hemolymph,and hepatopancreas of C.gigas during April 2018-January 2019 were investigated by 16 S rRNA gene sequencing.The microbiota in C.gigas exhibited tissue heterogeneity,of which Spirochaetaceae was dominant in the gill and hemolymph while Mycoplasmataceae enriched in the hepatopancreas.Co-occurrence network demonstrated that the gill microbiota exhibited higher inter-taxon connectivity while the hemolymph microbiota had more modules.The richness(Chao 1 index)and diversity(Shannon index)of microbial community in each tissue showed no significant seasonal variations,except for the hepatopancreas having a higher richness in the autumn.Similarly,beta diversity analysis indicated a relatively stable microbiota in each tissue during the sampling period,showing relative abundance of the dominant taxa exhibiting temporal dynamics.Results indicate that the microbial community in C.gigas showed a tissue-specific stability with temporal dynamics in the composition,which might be essential for the tissue functioning and environmental adaption in oysters.This work provides a baseline microbiota in C.gigas and is helpful for the understanding of host-microbiota interaction in oysters.