Calcined oyster is a commonly used shellfish traditional Chinese medicine in clinical practice in China. During the processing of oysters, their microscopic characteristics are destroyed, and open-fire calcination can damage the DNA of oysters, making it difficult to identify the primary source. The establishment of a specific polymerase chain reaction(PCR) method for the identification of calcined oysters can provide a guarantee for the safety and clinical efficacy of the medicine and its processed products. With Ostrea gigas as an example, the DNA extraction method of decoction pieces and formula particles of calcined oysters was improved, and high-quality DNA was obtained. Based on the specific single nucleotide polymorphism(SNP) sites of O. gigas and the other two species, the specific identification primers were designed, and the site-specific identification method of formula granules of calcined oyster(O. gigas) was established. The specificity and applicability of the method were investigated. The results showed that when the annealing temperature was 54 ℃, and the cycle was 44 times, the PCR amplified products of calcined oyster(O. gigas) and its formula granules produced a single bright identification band at 102 bp, while the other two species of oysters, O. talienwhanensis Crosse and O. rivularis Gould, had no band. In this study, DNA extraction and PCR identification of animal medicinal materials by calcination were established for the first time, which provided a tool for solving the difficult identification of calcined decoction pieces and ensuring drug safety.
Animals ; Ostrea/classification* ; Polymerase Chain Reaction/methods* ; Polymorphism, Single Nucleotide ; DNA/genetics*

The N⁶-methylation of RNA adenosines (N⁶-methyladenosine, m⁶A) is an important regulator of gene expression with critical implications in vertebrate and insect development. However, the developmental significance of epitranscriptomes in lophotrochozoan organisms remains unknown. Using methylated RNA immunoprecipitation sequencing (MeRIP-seq), we generated transcriptome-wide m⁶A-RNA methylomes covering the entire development of the oyster from oocytes to juveniles. Oyster RNA classes display specific m⁶A signatures, with messenger RNAs (mRNAs) and long non-coding RNAs (lncRNAs) exhibiting distinct profiles and being highly methylated compared to transposable element (TE) transcripts. Epitranscriptomes are dynamic and correspond to the chronological steps of development (cleavage, gastrulation, organogenesis, and metamorphosis), with minimal mRNA and lncRNA methylation at the morula stage followed by a global increase. mRNA m⁶A levels are correlated with transcript levels, and shifts in methylation profiles correspond to expression kinetics. Differentially methylated transcripts cluster according to embryo-larval stages and bear the corresponding developmental functions (cell division, signal transduction, morphogenesis, and cell differentiation). The m⁶A level of TE transcripts is also regulated and peaks during the gastrulation. We demonstrate that m⁶A-RNA methylomes are dynamic and associated with gene expression regulation during oyster development. The putative epitranscriptome implication in the cleavage, maternal-to-zygotic transition, and cell differentiation in a lophotrochozoan model brings new insights into the control and evolution of developmental processes.
Animals ; RNA/metabolism* ; Methylation ; RNA, Messenger/metabolism* ; Transcriptome ; Ostreidae/metabolism* ; RNA, Long Noncoding/metabolism*

Objective: To evaluate the application of real-time RT-PCR and semi-nested RT-PCR in the detection of norovirus in oysters and analyzing the genetic characteristics of the isolates. Methods: Real-time fluorescent RT-PCR and semi-nested RT-PCR were used to detect norovirus GⅠ/GⅡ in fresh oysters collected from the markets in Beijing from November 2014 to October 2015. The detection rate of the parallel test was also analyzed. In addition, the reliability of semi-nested RT-PCR was evaluated by agreement rate and consistency test (Kappa value). The positive products of norovirus GⅠ/GⅡ capsid protein region gene by semi-nested RT-PCR were sequenced. Software BioEdit 7.0.9.0 was used for sequence alignment, and software Mega 6.0 was used to construct the evolutionary tree. Results: In 72 samples, the detection rate of norovirus was 31.94% (23/72) by real-time RT-PCR, 38.89% (28/72) by semi-nested RT-PCR and 48.61% (35/72) by parallel test. The coincidence rate of the two methods was 73.61%, a moderate degree (Kappa value =0.43). A total of 13 norovirus strains were successfully sequenced, and 11 strains (7 GⅡ.17 strains, 2 GⅡ. 4 Sydney_ 2012 strains, 1 GⅡ. 1 strain and 1 GⅡ. 21 strain) were obtained from norovirus positive samples by two RT-PCR methods, two strains (1 GⅡ. 17 strain and 1 GⅡ. 3 strain) were obtained from real-time RT-PCR negative samples which were positive for norovirus by semi-nested RT-PCR. The similarity between these strains and reference strains from diarrhea patients, environmental sewage, and shellfish products were 84.4% - 100.0%. Conclusions: The parallel test of norovirus in oysters by two RT-PCR methods can improve the detection rate and detect more genotypes. Norovirus strains in oysters were highly homologous with reference strains from diarrheal patients, environmental sewage, and shellfish products. Therefore, surveillance, prevention and control for norovirus should be carried out in people who have frequent contacts with oysters and related environments.
Animals ; Beijing ; Humans ; Norovirus/genetics* ; Ostreidae ; RNA, Viral/genetics* ; Real-Time Polymerase Chain Reaction ; Reproducibility of Results ; Reverse Transcriptase Polymerase Chain Reaction

Animals ; Cadmium/toxicity* ; Dysbiosis/metabolism* ; Gastrointestinal Microbiome ; Glutamine/pharmacology* ; Methionine ; Mice ; Mice, Inbred C57BL ; Ostreidae ; Protein Hydrolysates ; Tyrosine

Background: One of the eight Ramsar sites in the Philippines is the Las Piňas – Parañaque Critical Habitat and Ecotourism Area(LPPCHEA), and it plays a significant role in the East Asian-Australian Flyway as a stopover site. The migratory birds coming from the north of Asia and Alaska feed on the molluscs in this area. However, there is paucity of literature on the species composition of molluscs found in this critical habitat. Baseline information on these organisms is essential as they are subject to the effects of anthropogenic activities close to and in the wetland, which in turn can have an impact on the ecosystem, particularly the birds foraging in this location. Methodology: The Natural Geography of in-Shore areas (NaGISA) protocol was used for the study. Transects were laid in three sampling sites in Freedom Island and Long Island. The sampling sites were GPS-referenced. A cylinder corer was used to collect mudflat soil, with the corer pushed into the sediment. Soil samples were sieved using a 0.5mm stainless mesh sieve pan, leaving shells and larger sand grains. The molluscs were sorted and identified through taxonomic keys. Sampling was done once for each site in November 2012. Results and Discussion: A total of 61 molluscan species belonging to two classes, 14 orders, and 33 molluscan families were identified and recorded. There were 34 species under the Class Gastropoda that belong to 5 orders and 18 families. For Class Bivalvia, there were 27 species belonging to 8 orders and 15 families. Among the molluscs recorded, 10 species were identified as non-indigenous. It is important to monitor molluscan species as anthropogenic activities may affect these organisms, and in turn, affect the wetland's function for migratory birds. The presence of non-indigenous species may be a potential threat to the ecosystem. Conclusion Baseline information of the molluscan community in the LPPCHEA was provided by the study. These species provide diet to the endemic and migratory birds in the area. There is a need to monitor these molluscs due to the effects of the man-caused activities close to the area. Also, the non-indigenous species should be studied for their potential to be invasive.
Mollusca

Aims: Bivalve molluscs are filter feeders that tend to accumulate and concentrate any suspended particles or bacteria from the water environment. Although they have been proven to accumulate heavy metals, investigation on the presence of antibiotic-resistant bacteria is still lacking. Therefore, this study examines the occurrence of antibiotic-resistant bacteria isolated from bivalve shellfish, namely, blood cockles (Anadara granosa) and green-lipped mussel (Perna viridis) purchased from markets in Johor, Malaysia. Methodology and results: Samples purchased were homogenized and then diluted. Viable cell count and bacterial isolation were performed using diluted samples followed by an antibiotic susceptibility test (ampicillin, ciprofloxacin and tetracycline) conducted on the pure isolates. The total viable count of bacterial colonies for cockle and mussel samples ranged from 1.1 × 107 to 4.4 × 109 CFU/mL and 1.2 × 109 to 4.3 × 109 CFU/mL. The numbers of colonies isolated from respective bivalves were 11 and 6. Generally, cockles isolates revealed higher resistance towards all three antibiotics at or above the Clinical and Laboratory Standard Institute (CLSI) threshold value. Meanwhile, mussel isolates showed full susceptibility to any ciprofloxacin concentration and tetracycline but exhibited resistance to ampicillin at a concentration exceeding the CLSI value. The number of drug resistance isolates in cockle and mussel samples decreased with increasing antibiotic concentration. The maximum number of antibiotics the mussel isolates were resistant to was two, whereas cockle isolates achieved three. Conclusion, significance and impact of study The outcome of this study concludes that some isolates from cockle and mussel samples can resist antibiotic concentration above the CLSI threshold value. Resistance of more than the CLSI threshold level revealed that these isolates could pose significant health risks especially when the bivalves are ingested raw or undercooked.
Drug Resistance, Bacterial ; Bivalvia

Objective: Owing to antibiotic abuse and the subsequent development of antibiotic resistance, bacterial infection has become one of the most persistent unresolved problems. New antibacterial agents, especially those that are environmental-friendly, are urgently needed. Methods: Melanin extracted by filtration centrifugation and acid and proteolytic hydrolysis was characterized using UV, FTIR, TEM, and XPS. Photothermal conversion was calculated, and the bacteriostatic effects, and , were assessed by plate counting and ratios (%) of wound areas. Results: Natural melanin hydrolyzed by trypsin had good photothermal conversion effects, which resulted in superior bacteriostatic activities. The extracted melanin along with laser NIR irradiation at 808 nm promoted the healing of wounds infected by drug-resistant bacteria and was biocompatible according to toxicity tests and . Conclusion The present findings indicated a safe and efficient method of developing natural antibacterial agents.
Animal Shells ; chemistry ; Animals ; Anti-Bacterial Agents ; pharmacology ; Escherichia coli ; drug effects ; radiation effects ; Escherichia coli Infections ; drug therapy ; Melanins ; pharmacology ; Mytilus edulis ; chemistry ; Photochemical Processes ; Rats ; Rats, Sprague-Dawley ; Staphylococcal Infections ; drug therapy ; Staphylococcus aureus ; drug effects ; radiation effects ; Wound Healing

The 2019 hepatitis A outbreak has become increasingly prevalent among adults in Korea and is the largest outbreak since that in 2009–2010. The incidence in the current outbreak is highest among adults aged 35–44 years, corresponding to the peak incidence among those aged 25–34 years 10 years ago. This may indicate a cohort effect in the corresponding age group. Causes of these repeated outbreaks of hepatitis A in Korea are low level of immunity among adults, Korean food culture that consumes raw seafood such as salted clam and inadequate public health system. Among countermeasures, along with general infectious disease control measures including control of the infectious agent, infection spread, and host, urgent actions are needed to review the vaccination policy and establish an adequate public health system.
Adult ; Bivalvia ; Cohort Effect ; Communicable Diseases ; Disease Outbreaks ; Epidemiology ; Hepatitis A virus ; Hepatitis A ; Hepatitis ; Humans ; Immunization ; Incidence ; Korea ; Public Health ; Seafood ; Vaccination

Gymnophalloides seoi (Digenea: Gymnophallidae) is a human intestinal trematode contracted by eating raw oysters (Crassostrea gigas) in the Republic of Korea (=Korea). It has been known to be highly endemic in Aphae Island, Shinan-gun, Jeollanam-do (Province). However, recent epidemiological status of G. seoi has not been reported since the 1990s. In this study, we investigated the prevalence of G. seoi metacercariae in natural and cultured oysters collected from 3 islands and 2 coastal areas in western parts of Korea. The oysters were examined using the artificial digestion method followed by stereomicroscopy. The overall positive rate of G. seoi metacercariae in natural oysters was 66.0% (99/150), and the oysters collected from Yubu Island showed the highest infection rate (74.0%). However, the metacercarial density per oyster was relatively low (1.5–2.4 per oyster). By contrast, no metacercaria was found in cultured oysters purchased from 2 coastal areas in Chungcheongnam-do. Thus, we could confirm that natural oysters produced from 3 western coastal islands are infected with G. seoi metacercariae, whereas cultured oysters purchased from 2 coastal areas were free from infection.
Chungcheongnam-do ; Digestion ; Eating ; Humans ; Islands ; Jeollanam-do ; Korea ; Metacercariae ; Methods ; Ostreidae ; Prevalence ; Republic of Korea

Anisakiasis (anisakidosis) refers to a foodborne zoonosis caused by ingesting raw or undercooked marine fish or cephalopods infected with anisakid larvae. The present study was performed to investigate the prevalence of anisakid larvae in anchovies (Engraulis japonica) purchased from 2 local markets in Gyeongsangnam-do, the Republic of Korea (=Korea), during 2018–2019. Anchovies were transported to our laboratory and examined by pepsin-HCl artificial digestion technique followed by microscopic observations and molecular analyses. The overall prevalence of anisakid larvae was 19.5% (39/200), from which a total of 51 larvae (av. 1.3 larvae/infected anchovy) were recovered. Sequencing of the larvae targeting the ITS region, including ITS1, 5.8S rRNA, and ITS2 genes confirmed the species of larvae as Anisakis pegreffii (54.9%; 28/51), Hysterothylacium sinense (23.5%; 12/51), and Hysterothylacium aduncum (21.5%; 11/51). The results suggested that anchovies could be a potential source of human anisakiasis in Korea.
Anisakiasis ; Anisakis ; Cephalopoda ; Digestion ; Gyeongsangnam-do ; Humans ; Korea ; Larva ; Prevalence ; Republic of Korea