• Parasites, Hosts and Diseases
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• v.61 no.1
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• pp.15-23
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• 2023

Concerns about foodborne illnesses caused by Kudoa septempunctata are steadily growing, but reports of K. septempunctata in clinical and food specimens related to food poisoning in Korea are limited. This study aimed to genetically identify K. septempunctata in patients with acute diarrhea and in clinical and food samples related to food poisoning caused by sashimi consumption. Both real-time and nested polymerase chain reaction assays were performed to detect K. septempunctata 18S and 28S rDNA genes in the stools of 348 patients with acute diarrhea, 11 samples (6 stool and 5 rectal swab samples) from patients with food poisoning, and 2 raw Paralichthys olivaceus samples collected from a restaurant where a food poisoning incident occurred. K. septempunctata was identified in 5 clinical specimens (4 stools and 1 rectal swab) and 1 P. olivaceus sashimi sample. All detected K. septempunctata were of genotype ST3. This is the first study to identify K. septempunctata in both patients and food samples with epidemiological relevance in Korea, providing evidence that it is a pathogen that causes food poisoning. Also, this is the first study to confirm the presence of K. septempunctata genes in rectal swabs. Despite continuing suspected occurrences of Kudoa foodborne outbreaks, the rate of identification of K. septempunctata is very low. One reason for this is the limitation in obtaining stool and vomit samples for the diagnosis of Kudoa infection. We strongly suggest the inclusion of rectal swabs among the diagnostic specimens for Kudoa food poisoning.

• Fisheries and Aquatic Sciences
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• v.26 no.2
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• pp.97-104
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• 2023

CD96 is a membrane-bound receptor discovered in humans in 1992 that is mainly present in natural killer cells and T cells derived from haematopoietic cells and performs immune functions. Based on the sequence of CD96 obtained from red seabream (Pagrus major), phylogenetic analysis with other species, infections of normal fish, Streptococcus iniae and red sea bream iridovirus (RSIV), and expression analysis was conducted using real-time polymerase chain reaction. Phylogenetic analysis showed the highest homology with Sparus aurata, and multiple sequence analysis confirmed the conservation of major domains between different fish species. Normal fish high expression results were confirmed in the head kidney, and spleen, which are the haematopoietic organs of the fish. High expression levels were confirmed in the gills, liver, spleen, and kidney on day three after RSIV infection. After S. iniae infection, high expression was confirmed in the gills and liver on day one, and high expression was confirmed in the spleen from 12 hours. These results show that PmCD96 functions as an immune gene in P. major and is considered a basic research case for CD96 in fish's hematopoietic organ immune system.

• Fisheries and Aquatic Sciences
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• v.26 no.3
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• pp.188-203
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• 2023

Lysozymes are well-known antibacterial enzymes that mainly target the peptidoglycan layer of the bacterial cell wall. Animal lysozymes are mainly categorized as g-type, c-type, and i-type based on protein sequence and structural differences. In this study, c-type (AcLysC) and g-like-type (AcLysG-like) lysozymes from Amphiprion clarkii were characterized in silico via expressional and functional approaches. According to in silico analysis, open reading frames of AcLysC and AcLysG-like were 429 bp and 570 bp, respectively, encoding the corresponding polypeptide chains with 142 and 189 amino acids. Elevated expression levels of AcLysC and AcLysG-like were observed in the liver and the heart tissues, respectively, as evidenced by quantitative real-time polymerase chain reaction assays. AcLysC and AcLysG-like transcript levels were upregulated in gills, head kidney, and blood cells following experimental immune stimulation. Recombinant AcLysC exhibited potent lytic activity against Vibrio anguillarum, whereas recombinant AcLysG-like showed remarkable antibacterial activity against Vibrio harveyi and Streptococcus parauberis, which was further evidenced by scanning electron microscopic imaging of destructed bacterial cell walls. The findings of this study collectively suggest the potential roles of AcLysC and AcLysG-like in host immune defense.

• Journal of Veterinary Science
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• v.24 no.3
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• pp.37.1-37.14
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• 2023

Background: Toll-like receptor (TLR) agonists have been used as adjuvants to modulate immune responses in both animals and humans. Objectives: The objective of this study was to evaluate the combined effects of the TLR 4 agonist monophosphoryl lipid A (MPL) and the TLR 3 agonist polyinosinic:polycytidylic acid (Poly I:C) on equine peripheral blood mononuclear cells (PBMCs), monocyte-derived dendritic cells (MoDCs), and bone marrow-derived mesenchymal stromal cells (BM-MSCs). Methods: The PBMCs, MoDCs, and BM-MSCs collected from three mixed breed horses were treated with MPL, Poly I:C, and their combination. The mRNA expression of interferon gamma (IFN-γ), interleukin (IL)-1β, IL-4, IL-6, IL-8, IL-12p40, tumor necrosis factor alpha (TNF-α), vascular endothelial growth factor (VEGF), and monocyte chemoattractant protein-1 (MCP-1) was determined using real-time polymerase chain reaction. Results: The combination of MPL and Poly I:C significantly upregulated immunomodulatory responses in equine cells/ without cytotoxicity. The combination induced greater mRNA expression of pro-inflammatory cytokines IFN-γ and IL-6 than MPL or Poly I:C stimulation alone in PBMCs. In addition, the combination induced significantly higher mRNA expression of IL-1β, IL-6, and IL-12p40 in MoDCs, and IL-8, MCP-1, and VEGF in BM-MSCs compared to stimulation with a single TLR agonist. Conclusions: The combination of MPL and Poly I:C can be used as a potential adjuvant candidate for vaccines to aid in preventing infectious diseases in horses.

• Clinical and Experimental Reproductive Medicine
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• v.49 no.4
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• pp.248-258
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• 2022

Objective: This research investigated the effects of human chorionic gonadotropin (HCG)-producing peripheral blood mononuclear cells (PBMCs) on the implantation rate and embryo attachment in mice. Methods: In this experimental study, a DNA fragment of the HCG gene was cloned into an expression vector, which was transfected into PBMCs. The concentration of the produced HCG was measured using enzyme-linked immunosorbent assay. Embryo attachment was investigated on the co-cultured endometrial cells and PBMCs in vitro. As an in vivo experiment, intrauterine administration of PBMCs was done in plaque-positive female mice. Studied mice were distributed into five groups: control, embryo implantation dysfunction (EID), EID with produced HCG, EID with PBMCs, and EID with HCG-producing PBMCs. Uterine horns were excised to characterize the number of implantation sites and pregnancy rate on day 7.5 post-coitum. During an implantation window, the mRNA expression of genes was evaluated using real-time polymerase chain reaction. Results: DNA fragments were cloned between the BamHI and EcoRI sites in the vector. About 465 pg/mL of HCG was produced in the transfected PBMCs. The attachment rate, pregnancy rate, and the number of implantation sites were substantially higher in the HCG-producing PBMCs group than in the other groups. Significantly elevated expression of the target genes was observed in the EID with HCG-producing PBMCs group. Conclusion: Alterations in gene expression following the intrauterine injection of HCG-producing PBMCs, could be considered a possible cause of increased embryo attachment rate, pregnancy rate, and the number of implantation sites.

• Clinical and Experimental Reproductive Medicine
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• v.49 no.4
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• pp.239-247
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• 2022

Objective: Asafoetida is a gum derived from Ferula assa-foetida, which is used in traditional Iranian medicine to treat some reproductive system disorders. The effects of asafoetida on ovarian tissue, expression of certain genes associated with polycystic ovary syndrome (PCOS), and levels of liver, kidney, and blood cell factors after treatment in a rat model were investigated. Methods: Thirty rats were divided into five groups: normal, polycystic, and treatment with three doses of asafoetida (12.5, 25, and 50 mg/kg for 3 weeks after PCOS induction). PCOS was induced by letrozole at a dose of 1 mg/kg administered orally for 3 weeks. Blood samples were taken, and the ovaries were removed and prepared for histomorphometric examination. Liver and kidney parameters were measured. The mRNA expression levels of luteinizing hormone receptor, CYP11A1, adenosine monophosphate-activated protein kinase, adiponectin, and adiponectin receptors 1 and 2 were also measured by real-time polymerase chain reaction. Results: The levels of liver, kidney, and blood parameters did not significantly differ between the treatment groups and the control group. At doses of 25 and 50 mg/kg, ovarian histopathology, especially the thicknesses of the theca and granulosa layers, was significantly improved relative to the PCOS group. The expression of target genes also improved in the 25 and 50 mg/kg treatment groups. Conclusion: Asafoetida can be used to treat PCOS as a complementary approach to conventional therapies. Asafoetida appears to act by regulating and activating metabolic and ovarian cycle enzymes.

• The Plant Pathology Journal
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• v.39 no.2
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• pp.191-206
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• 2023

Ground cherry (Physalis pubescens) is the most prominent species in the Solanaceae family due to its nutritional content, and prospective health advantages. It is grown all over the world, but notably in northern China. In 2019 firstly bacterial leaf spot (BLS) disease was identified on P. pubescens in China that caused by both BLS pathogens Xanthomonas euvesicatoria pv. euvesicatoria resulted in substantial monetary losses. Here, we compared whole genome sequences of X. euvesicatoria to other Xanthomonas species that caused BLS diseases for high similarities and dissimilarities in genomic sequences through average nucleotide identity (ANI) and BLAST comparison. Molecular techniques and phylogenetic trees were adopted to detect X. euvesicatoria on P. pubescens using recQ, hrpB1, and hrpB2 genes for efficient and precise identification. For rapid molecular detection of X. euvesicatoria, loop-mediated isothermal amplification, polymerase chain reaction (PCR), and real-time PCR techniques were used. Whole genome comparison results showed that the genome of X. euvesicatoria was more closely relative to X. perforans than X. vesicatoria, and X. gardneri with 98%, 84%, and 86% ANI, respectively. All infected leaves of P. pubescens found positive amplification, and negative controls did not show amplification. The findings of evolutionary history revealed that isolated strains XeC10RQ, XeH9RQ, XeA10RQ, and XeB10RQ that originated from China were closely relative and highly homologous to the X. euvesicatoria. This research provides information to researchers on genomic variation in BLS pathogens, and further molecular evolution and identification of X. euvesicatoria using the unique target recQ gene through advance molecular approaches.

• Journal of Oral Medicine and Pain
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• v.47 no.1
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• pp.10-26
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• 2022

Purpose: The purpose of this study was to investigate whether various saliva collection methods affect the observed salivary microbiome and whether microbiomes of stimulated and unstimulated saliva and plaque differ in richness and diversity. Methods: Seven sampling methods for unstimulated saliva, stimulated saliva, and plaque samples were applied to six orally and systemically healthy participants. Bacterial 16S ribosomal RNA genes of 10 major oral bacterial species, namely, Aggregatibacter actinomycetemcomitans, Porphyromonas gingivalis, Tannerella forsythia, Treponema denticola, Fusobacterium nucleatum, Prevotella intermedia, Prevotella nigrescens, Streptococcus mitis, Streptococcus sobrinus, and Lactobacillus casei, were analyzed by real-time polymerase chain reaction. We comprehensively examined the dependence of the amount of bacterial ribosomal DNA (rDNA), bacterial-community composition, and relative abundance of each species on sample collection methods. Results: There were significant differences in the bacterial rDNA copy number depending on the collection method in three species: F. nucleatum, P. nigrescens, and S. mitis. The species with the highest richness was S. mitis, with the range from 89.31% to 100.00%, followed by F. nucleatum, P. nigrescens, T. denticola, T. forsythia, and P. intermedia, and the sum of the proportions of the remaining five species was less than 1%. The species with the lowest observed richness was P. gingivalis (<0.1%). The Shannon diversity index was the highest in unstimulated saliva collected with a funnel (4.449). The Shannon diversity index was higher in plaque samples (3.623) than in unstimulated (3.171) and stimulated (3.129) saliva and in mouthwash saliva samples (2.061). Conclusions: The oral microbial profile of saliva samples can be affected by sample collection methods, and saliva differs from plaque in the microbiome. An easy and rapid technique for saliva collection is desirable; however, observed microbial-community composition may more accurately reflect the actual microbiome when unstimulated saliva is assayed.

• Journal of Korean Neurosurgical Society
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• v.66 no.4
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• pp.400-408
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• 2023

Objective : Nicotinamide mononucleotide adenylyl transferase 2 (NMNAT2) is a crucial factor for the survival of neuron. The role of NMNAT2 in damage following traumatic brain injury (TBI) remains unknown. This study was designed to investigate the role of NMNAT2 in TBI-induced neuronal degeneration and neurological deficits in rats. Methods : The TBI model was established in Sprague-Dawley rats by a weight-dropping method. Real-time polymerase chain reaction, western blot, immunofluorescence, Fluoro-Jade C staining, and neurological score analyses were carried out. Results : NMNAT2 mRNA and protein levels were increased in the injured-side cortex at 6 hours and peaked 12 hours after TBI. Knocking down NMNAT2 with an injection of small interfering RNA in lateral ventricle significantly exacerbated neuronal degeneration and neurological deficits after TBI, which were accompanied by increased expression of BCL-2-associated X protein (Bax). Conclusion : NMNAT2 expression is increased and NMNAT2 exhibits neuroprotective activity in the early stages after TBI, and Bax signaling pathway may be involved in the process. Thus, NMNAT2 is likely to be an important target to prevent secondary damage following TBI.

• Journal of Marine Life Science
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• v.1 no.2
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• pp.88-94
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• 2016

Determining the infection history of living organisms is essential for understanding the evolution of infection agents with their host, particularly for key aspects such as immunity. Viruses, which can spread between individuals and often cause disease, have been widely examined. The increasing availability of fish genome sequences has provided specific insights into the diversity and host distribution of retroviruses in fish. The shortspine spurdog (Squalus mitsukurii) is an important elasmobranch species; this medium-sized dogfish typically lives at depths of 100~500 m. However, the retroviral envelope polyprotein in dogfish has not been examined. Thus, the aim of the present study was to identify and analyze the retroviral envelope polyprotein in various tissues of dogfish. The 1334-base pair full-length novel cDNA of dogfish envelope polyprotein (dEnv) was obtained by 3' and 5'-rapid amplification of cDNA end analysis from S. mitsukurii. The open reading frame showed a complete coding sequence of 815 base pairs with a deduced peptide sequence of 183 amino acids that exhibited 34~50% identity with other fish and bird species. It was also expressed according to reverse transcription and real-time polymerase chain reaction in the kidney, liver, intestine, and lung, but not in the gill. This distribution can be assessed by identifying and analyzing endogenous retroviruses in fish, which consists of three main genes: gag, pol and env. Dogfish envelope polyprotein sequence is likely important in evolution and induces rearrangements, altering the regulatory and coding sequences. This is the first report of the identification and molecular characterization of retroviral envelope polyprotein in various tissues of S. mitsukurii.